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Symmetry®
ColumnsApplications
Notebook
Symmetry®
SymmetryShield™
Symmetry300™
SymmetryPrep™
Intelligent Speed (IS™)
1Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Column: Symmetry® C18,4.6 x 150 mm, 5 µm
Mobile Phase A: H2OMobile Phase B: MeCNMobile Phase C: pH 3.75; 100 mM
NH4COOH in H2OFlow Rate: 1.4 mL/minIsocratic: 30% A; 60% B; 10% CInjection Volume: 5.0 µLTemperature: 30 °CDetection: UV @ 233 nm
Instrumentation: Waters Alliance® HT 2795Waters 996 PDA detectorSampling Rate: 5 pts./sec
Setting the Standard for Reproducibility
As today’s chemists establish new analytical methods for the latest pharmaceu-tical and biopharmaceutical products, theselection of a reproducible HPLC column isessential. The selected column needs to provide the same chromatographic resultsover the life of the new drug product.Symmetry® columns demonstrate superiorreproducibility over many years. Batches randomly selected over the past 5 yearsdemonstrate excellent reproducibility in theexample shown.
Reproducibility is our number one priority insupplying Symmetry® columns. This excellentreproducibility is a result of our commitmentto maintaining the tightest specifications in theHPLC column industry. Symmetry® columnsare engineered with high purity raw materials, tightly controlled manufacturingprocesses and column packing proceduresthat provide today’s scientists with the best,most reproducible HPLC column available.
Our goal is to supply a family of HPLCcolumns that you can rely on for rugged androbust methods. Symmetry® columns let youincrease your laboratorys' productivity, andallow easier method transfer between labsand around the globe.
A three-page Certificate of Analysis is included with every Symmetry® column. Wereport both our specifications, which are thetightest in the industry, and the results of the22 critical tests each column must passbefore it carries the Symmetry® brand.
RSD’s for Retention Times
1. Terbinafine HCl 0.7%
2. Ibuprofen 0.8%
3. Lovastatin 0.6%
4. Simvastatin 0.7%
Batch 136 (1998)
Batch 142 (1998)
Batch 149 (1999)
Batch 151 (2000)
Batch 156 (2001)
Unmatched Year-to-year Reproducibility
Comprehensive Certificate of Analysis
Minutes
Symmetry® Columns
2 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Table of Contents
Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1
Compound Index . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3-4
List of Applications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5-8
Applications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9-182
Ordering Information . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .183-186
Notes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 187-192
3Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Compound Index
Acephate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5, 9
Acetaminophen . . . . . . . . . . . . . . . . . . . . . . . 5, 11-13
Acetanilide . . . . . . . . . . . . . . . . . . . 122-123, 147-148
Acetophenone . . . . . . . . . . . . . . . . . . . . . . . . 147-148
2-Acetylfuran . . . . . . . . . . . . . . . . . . . . . . . . . 147-148
Acifluorfen . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14
Acyclovir . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5, 15-17
Alachlor . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10
Albuterol . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5, 18-19
Allobarbital . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 33
Alpha-Tocopherol (Vitamin E) . . . . . . . . . . . . . . . . 179
6a-Methyl-17l-hydroxyprogesterone . . . . . . . . 122-123
2-Amino-7-chloro-5-oxo-5H-[1]
Benzopyrano[2,3-b]pyridine-3-carbonitrile . . . 122-123
7-Amino Clonazepam . . . . . . . . . . . . . . . . . . . . . . . 86
3-Aminofluoranthene . . . . . . . . . . . . . . . . . . . 122-123
7-Amino Flunitrazepam . . . . . . . . . . . . . . . . . . . . . . 86
7-Amino Nitrazepam . . . . . . . . . . . . . . . . . . . . . . . 86
Amitriptyline . . . . . . . . . . . . . . . . . . . . . . 173-175, 29
Amobarbital . . . . . . . . . . . . . . . . . . . . . . . . . . . 33-34
Amoxapine. . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5, 21
Amoxicillin . . . . . . . . . . . . . . . . . . . . . . . . . . 5, 22-24
Amphetamine . . . . . . . . . . . . . . . . . . . . . . . . 5, 26-27
Ampicillin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5, 25
Angiotensin I . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 115
Angiotensin II . . . . . . . . . . . . . . . . . . . . . . . . . . . . 115
Antipyrine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5, 31
Atrazine . . . . . . . . . . . . . . . . . . . . . 5, 32, 36, 81, 172
AZT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 182
β-hydroxyethyltheophylline (I.S.) . . . . . . . . . . . 45, 140
Bamethan (I.S.) . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19
Barbital . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 33
Beclomethasone . . . . . . . . . . . . . . . . . . . . . . 5, 35, 59
Benomyl . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5, 36, 81
Bentazon . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14
Benzoic acid . . . . . . . . . . . . . . . . . . . . . . . . . 145-146
Benzophenone. . . . . . . . . . . . . . . . . . . . . . . . 147-148
Benzoylecgonine . . . . . . . . . . . . . . . . . . . . . . . . . . . 62
Betamethasone. . . . . . . . . . . . . . . . 5, 37-38, 149-150
Betamethasone Valerate . . . . . . . . . . . . . . . . . 149-150
Berberine. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20
Bisphenol A . . . . . . . . . . . . . . . . . . . . . . . . . 5, 36, 81
2-Bromofluorene . . . . . . . . . . . . . . . . . . . . . . 122-123
8-Bromoguanosine . . . . . . . . . . . . . . . . . . . . . 122-123
BSA . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 39-42
Bupropion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5, 43
Busiperone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5, 44
Butabarbital . . . . . . . . . . . . . . . . . . . . . . . . . . . . 33-34
Butalbarbital . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 33
Butalbital . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 34
Butyl paraben . . . . . . . . . . . . . . . . . . . . 79, 135, 150
Butyrophenone . . . . . . . . . . . . . . . . . . . . . . . 147-148
Caffeine . . . . . . . . . . . . . . . . . . . . 5, 45, 48, 140, 170
Cafteric acid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 125
Calcium Pantothenate . . . . . . . . . . . . . . . . . . . . . . 180
Captopril . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5, 46
Captopril Disulfide . . . . . . . . . . . . . . . . . . . . . . . . . 46
Carbadox. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5, 47
Catechins. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 48
Carbamezapine . . . . . . . . . . . . . . . . . . . . . . . . . . . 30
Carbamezapine-10,11-epoxide . . . . . . . . . . . . . . . . 30
Carbaryl . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 36, 81
Cefixime . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5, 49
Cefuroxime . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5, 50
Cephalexin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5, 51
Chloramben . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14
Chloramphenicol . . . . . . . . . . . . . . . . . . . . . . . . . 5, 52
Chlorcyclizine HCL . . . . . . . . . . . . . . . . . . . . . . . . . 65
Chlordiazepoxide . . . . . . . . . . . . . . . . . . . . . 6, 53-54
2-Chloro-2,6-diethylacetanilide . . . . . . . . . . . . . . . . 10
Chlorogenic acid . . . . . . . . . . . . . . . . . . . . . . . . . . 125
Chlorpheniramine . . . . . . . . . . . . . . 6-7, 55, 137, 160
1-(3-Chlorophenyl)peperazine . . . . . . . . . . . . . . . . 171
Cholecalciferol (Vitamin D3) . . . . . . . . . . . . . . . . . 179
Cichoric acid . . . . . . . . . . . . . . . . . . . . . . . . . . . . 125
Cimetidine . . . . . . . . . . . . . . . . . . . . . . . . . . . 6, 56-58
Clavulanate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23
Clobetasol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6, 59
Clonazepam . . . . . . . . . . . . . . . . . . . . . . . . . 6, 60, 86
Cocaethylene . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 62
Cocaine . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6, 61-62
Codeine . . . . . . . . . . . . . . . . . . . . . 6, 11, 63-64, 139
Codeine-6-Glucuronide (C-6-G) . . . . . . . . . . . . . 63-64
Corticosterone . . . . . . . . . . . . . . . . . . . . . . . . 151-152
Cortisone . . . . . . . . . . . . . . . . . . . . . . . . . . . . 151-152
Cyanazine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 172
Cyclizine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6, 65
Cytochrome C. . . . . . . . . . . . . . . . . . . . . . . . . 6, 66-69
2,4-D . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14
2,4-DB . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14
Demoxepam . . . . . . . . . . . . . . . . . . . . . . . . . . . 53-54
Deoxidized (Insulin). . . . . . . . . . . . . . . . . . . . . . . . . 91
Desamino (Insulin) . . . . . . . . . . . . . . . . . . . . . . . . . . 91
Desethylatrazine . . . . . . . . . . . . . . . . . . . . . . . 32, 172
Desethyldesisopropylatrazine . . . . . . . . . . . . . . . . . . 32
Desisopropylatrazine . . . . . . . . . . . . . . . . . . . . 32, 172
Dextromethorphan . . . . . . . . . . . . . . . . . . 6, 70-71, 87
Diazepam . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6, 72
Dicamba. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14
3,5-Dichlorobenzoic Acid. . . . . . . . . . . . . . . . . . . . . 14
Dichlorprop . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14
Diclofenac Sodium . . . . . . . . . . . . . . . . . . . . . . . 6, 73
2,6-Diethylaniline . . . . . . . . . . . . . . . . . . . . . . . . . . 10
10,11-Dihydrocarbamezapine . . . . . . . . . . . . . . . . . 30
Dihydroergotamine mesylate . . . . . . . . . . . . . . . . 6, 74
Dilitiazem HCL . . . . . . . . . . . . . . . . . . . . . . . . . . 75-76
Dinoseb . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14
Diphenoxylate HCL . . . . . . . . . . . . . . . . . . . . . . . 6, 77
Diquat . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7, 120
Disopyramide . . . . . . . . . . . . . . . . . . . . . . . . . . . 6, 78
Doxepin . . . . . . . . . . . . . . . . . . . . 6, 29, 79, 173-175
Doxycycline . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 167
EDDP (Methadone Metabolite) . . . . . . . . . . . . . . . . . 99
Enalapril . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 80
Ephedrine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 26-27
Epicatechin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 48
Epigallocatechin Gallate . . . . . . . . . . . . . . . . . . . . . 48
Estazolam . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 99
Estradiol . . . . . . . . . . . . . . . . . . . . . . . . . . . . 151-152
Estrone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 151-152
Ethosuximide . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 153
Etodolac . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6, 83
Fenoprofen . . . . . . . . . . . . . . . . . . . . . . . . . . 131-132
Flunitrazepam . . . . . . . . . . . . . . . . . . . . . . . . 6, 85-86
Flurbiprofen . . . . . . . . . . . . . . . . . . . . . . . . . . 131-132
Folic Acid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8, 181
Furazolidone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 161
Furosemide . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6, 88
Guanosine hydrate . . . . . . . . . . . . . . . . . . . . . . . . . 17
Heptanophenone . . . . . . . . . . . . . . . . . . . . . . 147-148
Hexanophenone . . . . . . . . . . . . . . . . . . . . . . 147-148
Hydrastine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20
Hydrocodone Bitartrate . . . . . . . . . . . . . . . . . . . . 5, 12
Hydrocortisone . . . . . . . . . . . . . . . . . . 6, 89, 122-123
Hydrocortisone-21-Acetate . . . . . . . . . . . . . . . . . . . 89
4-Hydroxyantipyrine . . . . . . . . . . . . . . . . . . . . . . . . 31
Hydroxyatrazine . . . . . . . . . . . . . . . . . . . . . . . 32, 172
1-Hydroxybenzotriazole . . . . . . . . . . . . . . . . . . . . . 51
Hydroxydesethylatrazine . . . . . . . . . . . . . . . . . . . . . 32
Hydroxydesisopropylatrazine . . . . . . . . . . . . . . . . . . 32
7-(2-Hydroxyethyl) Theophylline . . . . . . . . . . . . . . 169
Ibuprofen . . . . . . . . . . . . . . . . . . . . . . 6, 90, 131-132
Imipramine . . . . . . . . . . . . . . . . . . . . . . . . . . 173, 175
Insulin(s) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 91-93
Isorhamnetin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 84
Kaempferol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 84
Ketamine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 124
Ketoconazole . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 94
Ketoprofen . . . . . . . . . . . . . . . . . . . . . . . . . . 131-132
Lasix (furosemide) . . . . . . . . . . . . . . . . . . . . . . . . . . 88
Click in the box, Type desired page number, Then click OK
4 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Compound Index
Leu enkephalin . . . . . . . . . . . . . . . . . . . . . . . . . . . . 82
Lidocaine HCL . . . . . . . . . . . . . . . . . . . . . . . . . . . 6, 95
Maleate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 160
Maleic Acid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 80
Maprotiline HCL . . . . . . . . . . . . . . . . . . . . . 6, 97, 171
MCPA . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14
MCPP . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14
Meclizine HCL . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 98
Mephenytoin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 28
Mephobarbital . . . . . . . . . . . . . . . . . . . . . . . . . 33-34
Met enkephalin . . . . . . . . . . . . . . . . . . . . . . . . . . . . 82
Methadone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7, 99
Methamphetamine . . . . . . . . . . . . . . . . . . . . . . . 26-27
Methoxyverapamil . . . . . . . . . . . . . . . . . . . . . 29, 178
Methsuximide . . . . . . . . . . . . . . . . . . . . . . . . . . . . 153
Methylparaben . . . . . . . . . . . . . . . . . . . . . . . . . . . . 95
Methylphenidate HCL . . . . . . . . . . . . . . . . . . . . 7, 100
Metolachlor. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10
Metolachlor-metabolite . . . . . . . . . . . . . . . . . . . . . . 10
Metoprolol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 134
Metoprolol tartrate . . . . . . . . . . . . . . . . . . . . . . 7, 101
Metronidazole . . . . . . . . . . . . . . . . . . . . . . 7, 102-103
Minocycline . . . . . . . . . . . . . . . . . . . . . . . . . . 167-168
Morphine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7, 104
Morphine-D3-glucuronide . . . . . . . . . . . . . . . . . . . 104
Morphine-D6-glucuronide . . . . . . . . . . . . . . . . . . . 104
N-Acetylprocainamide . . . . . . . . . . . . . . . . . . 128-129
N-Propionylprocainamide . . . . . . . . . . . . . . . 128-129
Nadolol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7, 105
Nalidixic acid. . . . . . . . . . . . . . . . . . . . . . . . 145, 161
Naloxone . . . . . . . . . . . . . . . . . . . . . . . . . . . . 106-109
Naltrexone . . . . . . . . . . . . . . . . . . . . 7, 106-109, 114
Naphtho(2,3-a)pyrene . . . . . . . . . . . . . . . . . . 122-123
Naproxen . . . . . . . . . . . . . . . . . . . . . . 7, 90, 110-111
Niacinamide . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 180
Nicergoline . . . . . . . . . . . . . . . . . . . . . . . 7, 112, 181
Nifedipine . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7, 113
Nitrazepam . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 86
4-Nitrophenol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14
Norchlordiazepoxide . . . . . . . . . . . . . . . . . . . . . 53-54
Nordiazepam . . . . . . . . . . . . . . . . . . . . . . . . . . 53-54
Nordoxepin . . . . . . . . . . . . . . . . . . . . . . . 29, 174-175
Nortriptyline . . . . . . . . . . . . . . . . . . . . . . 29, 173-175
Norverapamil. . . . . . . . . . . . . . . . . . . . . . . 8, 29, 178
Octanophenone . . . . . . . . . . . . . . . . . . . . . . . 147-148
O-dichlorobenzene . . . . . . . . . . . . . . . . . . . . . . . . . 60
Oxazepam . . . . . . . . . . . . . . . . . . . . . . . . . . . . 53-54
Oxolinic Acid . . . . . . . . . . . . . . . . . . . . . . . . . . . . 161
Oxprenolol . . . . . . . . . . . . . . . . . . . . . . . . . . 101, 134
Oxycodone . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7, 114
Oxytocin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7, 115
Paclitaxel (Taxol) . . . . . . . . . . . . . . . . . . . . 7, 116-117
Papaverine HCL . . . . . . . . . . . . . . . . . . . . . . . . . 7, 118
Paracetamol . . . . . . . . . . . . . . . . . . . . . . . . . . . 7, 119
Paraquat . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7, 120
Paraxanthine . . . . . . . . . . . . . . . . . . . . . . . . . 45, 140
Peptides. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7, 121
Perylene . . . . . . . . . . . . . . . . . . . . . . . . . . . . 122-123
Phencyclidine (PCP) . . . . . . . . . . . . . . . . . . . . . . . . 124
Phenobarbital . . . . . . . . . . . . . . . . . . . . 28, 30, 33-34
Phensuximide . . . . . . . . . . . . . . . . . . . . . . . . . . . . 153
2-Phenyl-2ethylmalonamide . . . . . . . . . . . . . . . . . . . 30
Phenytoin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 30
Phthalic Acid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 146
p-Hydroxyphenobarbital . . . . . . . . . . . . . . . . . . . . . 30
Picloram . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14
Prednisolone . . . . . . . . . . . . . . . . . . . . . . . . . 149-152
Prednisolone Acetate . . . . . . . . . . . . . . . . . . . . . . . 149
Prednisone . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7, 126
Primidone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 28, 30
Procainamide . . . . . . . . . . . . . . . . . . . . . . 7, 127-129
Prochlorperazine Edisylate . . . . . . . . . . . . . . . . . . . 130
Progesterone . . . . . . . . . . . . . . . . . . . . . . . . . 151-152
Promethazine HCL . . . . . . . . . . . . . . . . . . . . . . . . . 133
Propiophenone . . . . . . . . . . . . . . . . . . . . . . . 147-148
Propranolol . . . . . . . . . . . . . . . . . . . . 7, 134-135, 160
Propylparaben . . . . . . . . . . . . . . . . . . . . . . . . . . 37-38
Protriptyline HCL . . . . . . . . . . . . . . . . . . . . . . . . . . 173
Pseudoephedrine HCL . . . . . . . . . . . . . . . . 7, 136-139
Pyridoxal . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 180
Pyridoxamine Dihydrochloride . . . . . . . . . . . . . . . . 180
Pyrodoxine HCL . . . . . . . . . . . . . . . . . . . . . . . . . . 180
Quercetin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 84
Quinidine sulfate . . . . . . . . . . . . . . . . . . . . . . . . 8, 141
Ranitidine . . . . . . . . . . . . . . . . . . . 8, 63-64, 142-144
Riboflavin . . . . . . . . . . . . . . . . . . . . . . . . . . . 180-181
Salbutamol . . . . . . . . . . . . . . . . . . . . . . . . . . 5, 18-19
Salicylic acid . . . . . . . . . . . . . . . . . . . . . . . 8, 145-146
Secobarbital . . . . . . . . . . . . . . . . . . . . . . . . . . . 33-34
Simazine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 172
Succinimide . . . . . . . . . . . . . . . . . . . . . . . . 8, 28, 153
Succinylsulfathiazole . . . . . . . . . . . . . . . . . . . . 96, 154
Sulfadiazine . . . . . . . . . . . . . . . . . . . . . . 97, 154-157
Sulfadimethoxine . . . . . . . . . . . . . . . . . . . . . . 156-157
Sulfadimidine . . . . . . . . . . . . . . . . . . . . . . . . 156-157
Sulfamerazine . . . . . . . . . . . . . . . . . 97, 154-157, 159
Sulfamethazine . . . . . . . . . . . . . . . . . . . . . . . . 96, 154
Sulfamethoxazole . . . . . . . . . . . . . . . . . . . 8, 156-157
Sulfanilamide . . . . . . . . . . . . . . . . . 97, 127, 144, 154
Sulfathiazole . . . . . . . . . . . . . . . . . . . . . . 97, 154-157
Sulfisoxazole . . . . . . . . . . . . . . . . . . . . . . . . . 156-157
2,4,5-T . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14
Tamoxifen citrate . . . . . . . . . . . . . . . . . . . . . . 162-163
Taxol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8, 116-117
Terconazole . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 94
Terfenadine . . . . . . . . . . . . . . . . . . . . . . . . 8, 164-165
Testosterone acetate . . . . . . . . . . . . . . . . . . . . . . . . 166
Testosterone benzoate . . . . . . . . . . . . . . . . . . . . . . 166
Testosterone enanthate (I.S.) . . . . . . . . . . . . . . . . . . 166
Testosterone Propionate (I.S.) . . . . . . . . . . . . . . 35, 166
Tetracaine. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6, 87
Tetracycline. . . . . . . . . . . . . . . . . . . . . . . . . . . 167-168
Theobromine . . . . . . . . . . . . . . . . 8, 45, 140, 169-170
Theophylline . . . . . . . . 8, 45, 140, 147-148, 169-170
Thiamine Hydrochloride . . . . . . . . . . . . . . . . . 180-181
Toluamide . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 160
2,4,5-TP . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14
Trans-Retinol (Vitamin A) . . . . . . . . . . . . . . . . . . . . 179
Trazodone . . . . . . . . . . . . . . . . . . . . . . . 5, 8, 21, 171
Triamcinolone . . . . . . . . . . . . . . . . . . . . . . . . 122-123
Trimethoprim . . . . . . . . . . . . . . . . . . . . . . . 8, 158-159
Trimipramine . . . . . . . . . . . . . . . . . . . . . . . . 173, 175
Tripelennamine . . . . . . . . . . . . . . . . . . . . . . . . . 8, 176
Triprolidine HCL . . . . . . . . . . . . . . . . . . . . . . . 87, 138
Uracil . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 149, 179
Valerophenone . . . . . . . . . . . . . . . . . . . . . . . 147-148
Verapamil . . . . . . . . . . . . . . . . . . . . . . 8, 29, 177-178
Vitamins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 179-181
Warfarin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6, 87
Zidovudine . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8, 182
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List of ApplicationsAcephate in Drinking Water . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9
Acetamide Herbicides and Metabolites in Drinking Water . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10
Acetaminophen and Codeine. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
Acetaminophen and Hydrocodone Bitartrate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12
Acetaminophen in Serum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13
Acidic Herbicides in Drinking Water . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14
Acyclovir . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 15,16
Acyclovir in Plasma. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17
Albuterol (Salbutamol) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18
Albuterol (Salbutamol) in Serum. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19
Alkaloids in Golden Seal . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20
Amoxapine & Trazodone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21
Amoxicillin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 22
Amoxicillin & Clavulanate Potassium Augmentin Tablet . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23
Amoxicillin in an Oral Suspension Formulation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 24
Ampicillin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 25
Amphetamines . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 26
Amphetamines in Human Urine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 27
Anticonvulsants. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 28
Antidepressants in Plasma . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 29
Antiepileptics & Metabolites . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 30
Antipyrine & Its Metabolites . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 31
Atrazine & Metabolites in Drinking Water . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 32
Barbiturates . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 33
Barbiturates in Human Urine. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 34
Beclomethasone. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35
Benomyl & Bisphenol A. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 36
Betamethasone. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 37
Betamethasone in Syrup Formulation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 38
BSA Tryptic Digest . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 39
BSA Tryptic Digest - Column Length Effects . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 40
BSA Tryptic Digest - Flow Rate Effects . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 41
BSA Tryptic Digest - Gradient Duration Effects . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 42
Bupropion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 43
Busiperone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 44
Caffeine and Metabolites in Serum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 45
Captopril . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 46
Carbadox. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 47
Catechins - 4.6 x 20 mm Intelligent Speed™ Separation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 48
Cefixime. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 49
Cefuroxime . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 50
Cephalexin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 51
Chloramphenicol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 52
5Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
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List of ApplicationsChlordiazepoxide & Metabolites. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 53
Chlordiazepoxide & Metabolites in Serum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 54
Chlorpheniramine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 55
Cimetidine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 56-58
Clobetasol and Related Compound . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 59
Clonazepam . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 60
Cocaine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 61
Cocaine & Its Metabolites in Serum. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 62
Codeine and Its Glucuronide Metabolite in Human Urine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 63
Codeine and Its Glucuronide Metabolite in Porcine Plasma . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 64
Cyclizine & Chlorcyclizine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 65
Cytochrome c Tryptic Digest - Column Lifetime Study, Critical Pair Resolution . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 66
Cytochrome c Tryptic Digest - Pore Size Effects . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 67
Cytochrome c Tryptic Digest - Recovery Study . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 68
Cytochrome c Tryptic Digest - Reproducibility Study . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 69
Dextromethorphan . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 70
Dextromethorphan in a Cough Syrup . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 71
Diazepam in Serum - LC/MS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 72
Diclofenac Sodium . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 73
Dihydroergotamine Mesylate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 74
Diltiazem HCL . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 75
Diltiazem HCL -Isolation of Impurities, Transfer from Analytical to Prep . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 76
Diphenoxylate HCL. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 77
Disopyramide . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 78
Doxepin in Serum. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 79
Enalapril . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 80
Endocrine Disruptors in Soil . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 81
Enkephalins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 82
Etodolac . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 83
Flavanoids in Ginko . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 84
Flunitrazepam . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 85
Flunitrazepam and Related Compounds . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 86
Four Drug Mixture: Warfarin, Dextromethorphan, Triprolidine, and Tetracaine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 87
Furosemide . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 88
Hydrocortisone and Related Compunds. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 89
Ibuprofen and Naproxen in Serum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 90
Insulin Chain - QC Batch Test for Symmetry 300™. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 91
Insulin- Impurity Isolation, Transfer from Analytical to Prep . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 92
Insulin- Isophase Suspension . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 93
Ketoconazole. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 94
Lidocaine HCL . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 95
Lifetime of 3.5 µm Symmetry® Column: 10,000 Injections of Sulfa Drugs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 96
Maprotiline HCL. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 97
Click on the application titles below to go directly to the desired applicationFor a new search, click on the "List of Applications" bookmarks to the left
List of ApplicationsMeclizine HCL. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 98
Methadone in Human Urine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 99
Methylphenidate HCL . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 100
Metoprolol Tartrate and Oxyprenolol. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 101
Metronidazole . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 102,103
Morphine & Its Glucuronide Metabolites in Serum. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 104
Nadolol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 105
Naltrexone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 106
Naltrexone in Plasma . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 107
Naltrexone in Plasma - LC/MS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 108
Naltrexone in Serum. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 109
Naproxen . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 110,111
Nicergoline . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 112
Nifedipine. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 113
Oxycodone in Serum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 114
Oxytocin & Angiotensins in Serum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 115
Paclitaxel (Taxol) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 116
Paclitaxel (Taxol)- Resolution of Impurities. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 117
Papaverine HCL. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 118
Paracetamol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 119
Paraquat & Diquat in Tap Water . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 120
Peptides- Alberta Peptide Mixture . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 121
Pharmaceutical Test Mix - 2.1 x 20 mm Intelligent Speed™ Separation. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 122
Pharmaceutical Test Mix - 4.6 x 20 mm Intelligent Speed™ Separation. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 123
Phencyclidine in Human Urine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 124
Phenolics in Echinacea . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 125
Prednisone. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 126
Procainamide in Serum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 127
Procainamides at pH 3.0 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 128
Procainamides at pH 6.0 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 129
Prochlorperazine- Isolation of Degradation Products, Transfer from Analytical to Prep . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 130
Profens (Gradient) - 4.6 x 20 mm Intelligent Speed™ Separation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 131
Profens (Isocratic) - 4.6 x 20 mm Intelligent Speed™ Separation. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 132
Promethazine- Impurities Profile . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 133
Propranolol in Human Urine. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 134
Propranolol in Serum. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 135
Pseudoephedrine HCL. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 136
Pseudoephedrine HCL & Chlorpheniramine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 137
Pseudoephedrine HCL & Triprolidine HCL . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 138
Pseudoephedrine HCL & Codeine. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 139
Purine Alkaloids . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 140
Quinidine Sulfate and Related Substance. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 141
Ranitidine. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 142,143
7Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Click on the application titles below to go directly to the desired applicationFor a new search, click on the "List of Applications" bookmarks to the left
8 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
List of ApplicationsRanitidine in Serum. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 144
Salicyclic Acid & Related Compounds in Serum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 145
Salicyclic Acid & Related Compounds. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 146
Speed Mix - 2.1 x 20 mm Intelligent Speed™ Separation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 147
Speed Mix - 4.6 x 20 mm Intelligent Speed™ Separation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 148
Steroids. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 149
Steroids in Serum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 150
Steroids - 2.1 x 20 mm Intelligent Speed™ Separation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 151
Steroids - 4.6 x 20 mm Intelligent Speed™ Separation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 152
Succinimides . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 153
Sulfa Drugs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 154,155
Sulfa Drugs - 2.1 x 20 mm Intelligent Speed™ Separation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 156
Sulfa Drugs - 4.6 x 20 mm Intelligent Speed™ Separation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 157
Sulfamethoxazole Oral Suspension . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 158
Sulfamethoxazole and Trimethorprim in Serum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 159
Symmetry® C18 pH 3 Batch Test. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 160
Synthetic Antibiotics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 161
Tamoxifen . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 162
Tamoxifen, Isolation of Impurities, Transfer from Analytical to Prep Columns . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 163
Terfenadine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 164
Terfenadine and Related Compound. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 165
Testosterone Esters in Serum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 166
Tetracyclines . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 167
Tetracyclines in Serum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 168
Theophylline & Related Compounds . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 169
Theophylline & Theobromine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 170
Trazodone. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 171
Triazine Herbacides in Drinking Water . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 172
Trycyclic Antidepressants . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 173
Trycyclic Antidepressants & Metabolites . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 174
Trycyclic Antidepressants in Plasma . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 175
Tripelennamine. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 176
Verapamil . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 177
Verapamil and Norverapamil. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 178
Vitamins: Fat-Soluble . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 179
Vitamins: Water-Soluble. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 180
Vitamins: Water-Soluble Folic Acid Mixture. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 181
Zidovudine (AZT) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 182
Click on the application titles below to go directly to the desired applicationFor a new search, click on the "List of Applications" bookmarks to the left
Acephate in Drinking Water
acephate
Minutes0 10 155
blank cartridge
nonspiked sample
spiked tap water
Oasis® HLB Extraction MethodConditions for Oasis® HLB Cartridge, 3 cc, 60 mg
Part Number WAT094226
% RecoverySpike Level Spike Level10 µg/L 5 µg/L5 replicates 7 replicates91.0% (16.3% RSD) 83.6% (15.0% RSD)
(Source water: chlorinated municipal tap water)
HPLC Method
Column: SymmetryShield™ RP8, 3.9 x 150 mm, 5 µmPart Number: WAT200655Mobile Phase: H2O/ACN 96:4Flow Rate: 1.0 mL/minInjection volume: 75 µLDetection: UV @ 200 nm (0.02 AUFS)
SP
NC
O
CH3
OH3C
HOH3C
- contact and systematic organophosphorous insecticide- highly polar, water soluble
Prepare Sample:To 20 mL sample add 5 g NaCl
Adjust to pH 3 with H3PO4
Condition:3 mL methylene chloride
Rinse: 3 mL methanolRinse: 3 mL H2O
Load:20 mL sample @ 4 mL/min
Elute:2 mL methylene chloride
Evaporate and Reconstitute:with 250 µL mobile phase
Acephate
9Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Minutes
10 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Acetamide Herbicides and Metabolitesin Drinking Water
Minutes0 10 20
1
2 3 4 5
spiked well water(200 ng/L)
blank cartridge
nonspiked sample
Oasis® HLB Extraction MethodConditions for Oasis® HLB Cartridge, 3 cc, 60 mg
Part Number WAT094226
Compounds % RecoveryTap Water Tap Water Well Water2.0 µg/L 200 ng/L 200 ng/L5 replicates 5 replicates 5 replicates
1. Metolachlor-metabolite* 99.1 (10) 78.9 (22) 88.3 (3.3)2. 2-Chloro-2,6-diethylacetanilide 98.5 (3.8) 109 (34) 105 (24)3. 2,6-Diethylaniline 93.3 (11) 80.3 (23) 89.2 (7.8)4. Alachlor 93.9 (3.2) 90.1 (21) 89.9 (1.2)5. Metolachlor 97.1 (6.6) 93.0 (16) 112 (3.3)
* 2-[(2-ethyl-6-methylphenyl)amino]1-propanol
HPLC Method
Column: Symmetry® C8 3.9 x 150mm, 5µm Part Number: WAT046970Mobile Phase A: 10mm phosphate buffer pH 6.8,/acetonitrile 70:30Mobile Phase B: AcetonitrileGradient: Time Profile
(min) %A %B0 100 020 40 60
Flow Rate: 1.2 mL/minInjection volume: 80 µLDetection: UV @ 214 nm (0.02 AUFS)
CH2CH3
CH2CH3
N
COCH2Cl
CH2OCH3
Condition:3 mL methanol
Rinse:2 mL H2O
Load:150 mL sample @ 4 mL/min
Wash:1 mL H2O
Elute:1.2 mL methanol
Evaporate to 0.2 mLReconstitute to 0.5 ml with reagent H2O
Alachlor
Minutes
11Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Acetaminophen and Codeine
HPLC Method
Column: Symmetry® C8, 3.9 x 150mm, 5 µmPart Number: WAT046970Mobile phase: 10mM docusate sodium in methanol/water/THF/phosphoric acid 60:36:4:1Flow rate: 0.75mL/minInjection volume: 15 µL of 55µg/mL acetaminophen and 35 µg of codeine sampleDetection: UV @ 254 nm
USP Tailing Factors
1. 1.092. 1.04
1
2
0 2 4 6 8 10 12
Minutes
1. Acetaminophen
2. Codeine
Minutes
12 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Acetaminophen and Hydrocodone Bitartrate
HPLC Method
Column: Symmetry® C18, 3.9 x 150mm, 5µmPart Number: WAT046980Mobile phase: 50 mM Ammonium phosphate, pH 5.8/acetonitrile 85:15 Flow rate: 1.0 mL/minInjection volume: 20 µL of 775µg/mL acetaminophen and 8 µg hydrocodone bitartrate sampleDetection: UV @ 214 nm
1. Acetaminophen
2. Hydrocodone Bitartrate
Minutes
13Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Acetaminophen in Serum
0.02 AU
1
2
B
A
0 4 8 12Minutes
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30 mg Extraction Cartridge
Part Number WAT094225
Compound Concentration % Recovery %RSD µg/mL (n=6)
Acetaminophen 0.100 107% 1.5%
0.500 101% 0.40%
HPLC Method
Column: SymmetryShield™ RP8, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ guard column, 3.9 x 20 mm, 5 µmPart Numbers: Column - WAT200655, Guard - WAT200675Mobile phase: 20 mM potassium phosphate, pH 3.0/methanol 97:3)Flow rate: 1.0 mL/minInjection volume: 50 µL of reconstituted porcine serum extractTemperature: 30˚ CDetection: UV @ 220 nm
Compounds:1: Sulfanilamide (I.S.)2: Acetaminophen
HO
O
2CHHN
Condition1 mL methanol/1 mL water
Load1 mL spiked porcine serum
Wash1 mL 5% methanol in water
Elute1 mL methanol
Evaporate and Reconstitute40 ˚C under nitrogen stream
200 µL of 50 µg/mL sulfanilamide(I.S.) in water
2. Acetaminophen
Chromatogram of Serum Extracts: A) Blank B) Spiked Sample
Minutes
1. Sulfanilamide (I.S)
14 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Acidic Herbicides in Drinking Water
15
Minutes0 10 20 30
1
23
4 5
67 8
91011 12
1314
spiked tap water(400 ng/L)
nonspiked sample
Compounds
1. Picloram2. Dicamba3. Chloramben4. 4-Nitrophenol5. Bentazon6. 2,4-D
7. MCPA8. Dichlorprop9. 2,4,5-T
10. MCPP11. 3,5-Dichlorobenzoic12. 2,4-DB
13. 2,4,5-TP
14. Acifluorfen15. Dinoseb
HPLC Method
Column: SymmetryShield™ RP8 3.9 x 150mm, 5µmPart Number: WAT200655Mobile Phase A: 13 mM phosphate buffer, pH 3.4Mobile Phase B: AcetonitrileGradient: Time Profile
(min) %A %B0 85 158 70 3015 70 3030 40 6035 10 90
Flow Rate: 1.0 mL/minInjection volume: 75 µLDetection: UV @ 230 nm (0.015 AUFS)
Oasis® HLB Extraction MethodConditions for Oasis® HLB Cartridge, 3 cc, 60 mg
Part Number WAT094226
Prepare Sample: 75 mL sample,adjust to pH 2 with H3PO4
Condition:3 mL 10:90 methanol/MTBE*
Risnse: 2 mL methanolRinse: 2 mL H2O
Load:75 mL sample @ 4 mL/min
Wash:1 mL H2O
Elute:2 mL 10:90 methanol/MTBE*
Evaporate to 0.2 mLReconstitute to 0.5 mL with H2O
* methyl t-butyl etherdiethyl ether can be used as an alternative to MTBE
N
Cl
OH
O
N
Cl
Cl
1. Picloram
Cl
OH
O
N
Cl
3. Chloramben
N+
O O
OH
4. 4-Nitrophenol
N
S
N
O
O
O
9. 2,4,5-T5. Bentazon
O
Cl
Cl
OH
O
6. 2,4-D
O
Cl
OH
O
7. MCPA
Cl
OHO
Cl
11. 3,5-Dichlorobenzoic acid
O
Cl
Cl
OH
O
8. Dichloroprop
O
Cl
OH
O
10. MCPP
OO
N+
O
O
O
F
F
F
Cl
O OO
N
14. Acifluorfen
OH
N+
N+
O
O
O O
15. Dinoseb
O
Cl
Cl
Cl
OH
O
13. 2,4,5-TP
O
Cl
Cl
OH
O
12. 2,4-DB
2. Dicamba
Minutes
15Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µmPart number: WAT046970Mobile phase: 0.1% glacial acetic acid (aq), pH 3.3Flow rate: 1.0 mL/minInjection volume: 5 µL of 100 µg/mL sampleDetection: UV @ 254 nm
USP Tailing Factors
1. 1.1
Minutes
Acyclovir
1. Acyclovir
16 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
HPLC Method
Column: SymmetryShield™ RP18, 4.6 x 150 mm, 5 µmPart number: 186000109Mobile phase: 20 mM, pH 7.0 phosphate buffer/acetonitrile 95:5Flow rate: 1.0 mL/minInjection volume: 5 µLDetection: UV @ 254 nm
USP Tailing Factors
1. 1.1
Acyclovir
1. Acyclovir
Minutes
17Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Acyclovir in Plasma
Compound Concentration % Recovery %RSD µg/mL (n=6)
Acyclovir 1.0 95.1% 3.4%0.20 98.3% 5.0%
HPLC Method
Column: SymmetryShield™ RP8, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ guard column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT200655, Guard - WAT200675Mobile phase: 20 mM ammonium acetate, pH 5Flow rate: 1.0 mL/minInjection volume: 50 µLDetection: UV @ 250 nm
Compounds:1. Acyclovir2. Guanosine hydrate
Chromatogram of Plasma Extracts: A) Blank B) Spiked Sample
2. Guanosine hydrate (I.S.)
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30 mg Extraction Cartridge
Part Number WAT094225
Condition
1 mL methanol/1 mL water
Load
1 mL spiked plasma
Wash
300 µL water
Elute
500 µL 1% glacial acetic acid
with 10% methanol
Evaporate and Reconstitute
Dilute and add internal standard:
480 µL mobile phase and
20 µL internal standard
0.015AU
Minutes
0 5 15
1
2
B
A
1. Acyclovir
18 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Albuterol (Salbutamol)
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: Water/methanol/glacial acetic acid 69:30:1 with 5 mM sodium 1-hexanesulfonateFlow rate: 1.0 mL/minInjection volume: 10 µL of 200 µg/mL salbutamolDetection: UV @ 276 nm
USP Tailing Factors
1. 1.2
1. Salbutamol
Minutes
1
0 2 4 6 8
HN
CH3
CH3
HO
HO
HO
19Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Albuterol (Salbutamol) in Serum
A
B
2015105
12
Minutes
0
100 mV
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30 mg Extraction Cartridge
Part Number WAT094225
Compound Concentration % Recovery %RSD µg/mL (n=6)
Salbutamol 0.400 98.8% 3.4%2.00 98.0% 3.5%
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ guard column, 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046980, Guard - WAT054250Mobile phase: 5 mM hexanesulfonate and 1% glacial acetic acid in
water/1% glacial acetic acid in methanol, 75:25Flow rate: 1.0 mL/minInjection volume: 20 µL of reconstituted porcine serum extractDetection: Fluorescence, Ex = 275 nm, Em = 310 nm
Compounds:1: Salbutamol2: Bamethan (I.S.)
Condition:1 mL methanol/1 mL water
Load:1 mL spiked porcine serum
with 1.0 µg/mL bamethan (I.S.)
Wash:1 mL 5% methanol in water
Elute:1 mL methanol
Evaporate and Reconstitute:
40 ˚C under nitrogen stream200 µL mobile phase
1. Salbutamol
Chromatogram of Serum Extracts: A) Blank B) Spiked Sample
2. Bamethan (I.S.)
HN
CH3
CH3
HO
HO
HO
Minutes
20 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Alkaloids in Goldenseal
Oasis® HLB method for alkaloids in GoldensealConditions for Oasis® HLB cartridge, 3 cc, 60 mg
Part Number WAT094226
Condition/Equilibrate:
1 mL methanol/1 mL 150 mM K2HPO4
Load:
3 mL of diluted sample
Wash:
1 mL 30% methanol/ 150 mM K2HPO4
Elute:
3 mL methanol
0 5 10
Sample pre-preparation100 mg sample is extracted with 25 mL of ethanol:water (70:30)1 mL of the ethanolic extract is diluted 1:3 with water (~ pH 7)
• Recovery, measured with certified standards at 100 ppm in reagent water,was greater than 85 % for all compounds
• All analyses gave results within ± 35 % of the expected values with theexception of the liquid supplement (see goldenseal analysis)
• The selective SPE extraction and cleanup procedure provided a convenientanalysis of echinacea phenolics in complex matrix such as in herbal tea
HPLC Method
Column: Symmetry® C18, 4.6 x 250 mm, 5 µmPart number: WAT054275Mobile Phase A: 0.1% phosphoric acidMobile Phase B: AcetonitrileGradient: Time Profile
(min) %A %B0 90 1013 78 2240 60 40
Flow Rate: 1.5 mL/minInjection volume: 10 µLTemperature: 35° CDetection: UV @ 330 nm
Herbal tea extractbefore SPE cleanup
Herbal tea extract after SPE cleanupusing Oasis® MAX
Minutes
hydrastineberberine
21Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: 20 mM potassium phosphate pH 7.0/ methanol 35:65Flow rate: 1.0 mL/minInjection volume: 10 µL of 100 µg/mL amoxapine and 100 µg/mL trazodoneDetection: UV @ 254 nm
USP Tailing Factors
1. 1.12. 1.4
1. Trazodone
2. Amoxapine
Amoxapine and Trazodone
Minutes
22 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Amoxicillin
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: 10 mM sodium acetate, pH 4.0/acetonitrile 96:4Flow rate: 1.0 mL/minInjection volume: 10 µL of 100 µg/mL sampleDetection: UV @ 230 nm
USP Tailing Factors
1. 1.15
1. Amoxicillin
Minutes
23Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
HPLC Method
Column: Symmetry® C8 ,3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: 56 mM sodium phosphate, pH 4.4/methanol 95:5Flow rate: 1.0 mL/minInjection volume: 10 µL of 100 µg/mL amoxicillin and 25 µg/mL clavulante extracted tablet sample at 10 to 39 µg/mLDetection: UV @ 220 nm
USP Tailing Factors
1. 1.22. 1.1
1. Clavulanate
2. Amoxicillin
Amoxicillin and Clavulanate Potassium Augmentin Tablet
Minutes
24 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Amoxicillinin in an Oral Suspension Formulation
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: 10 mM sodium acetate, pH 4.0/acetonitrile 96:4Flow rate: 1.0 mL/minInjection volume: 10 µL of 100 µg/mL sampleDetection: UV @ 230 nm
USP Tailing Factors
1. 1.13
1. Amoxicillin
Minutes
25Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Ampicillin
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: 10 mM sodium acetate, pH 4.0/acetonitrile 92:8Flow rate: 1.0 mL/minInjection volume: 10 µL of 100 µg/mL sampleDetection: UV @ 230 nm
USP Tailing Factors
1. 1.07
1. Ampicillin
Minutes
26 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Amphetamines
HPLC Method
Column: SymmetryShield™ RP18, 3.9 x 150 mm, 5 µm, Part Number: 186000108Mobile phase: 0.1% TFA in Water/acetontrile/methanol 91:7:2Flow rate: 1.0 mL/minInjection volume: 50 µLDetection: UV @ 210
USP Tailing Factors
1. 1.282. 1.323. 1.30
1. Ephedrine (I.S.)
2. Amphetamine
3. Methamphetamine
Minutes
27Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Amphetamines in Human Urine
HPLC Method
Column: SymmetryShield™ RP18, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ Guard Column, 3.9 x 20 mm, 5 µmPart Numbers: Column - 186000108, Guard - 186000107Mobile phase: 0.1% TFA in Water/Acetontrile 90:10Flow rate: 1.0 mL/minInjection volume: 50 µL Temperature: 30˚ CDetection: UV @ 214 nm (0.14 AUFS)
Compounds:1. Ephedrine2. Amphetamine3. Methamphetamine
%Recovery (%RSD) Spike Levels: 0.7 µg/mL 3.5 µg/mL
Amphetamine (n=8) 99.3 (4.9) 104.7 (1.8)
Methamphetamine (n=8) 105.5 (5.8) 100.9 (4.4)
Amphetamine Interday (n=16) 102.8 (2.7)
Amphetamine Interperson (n=16) 98.8 (6.2) 101.8 (3.3)
Methamphetamine Interday (n=16) 98.5 (5.4)
Methamphetamine Interperson (n=16) 96.4 (11.5) 99.4 (6.3)
Ephedrine (I.S.) (n=8) 89.5 (1.4)
2 4 6 8
Minutes
0.14
AU
FS
12
3
B
A
Chromatogram of A) Spiked Urine, B) Blank Urine
1. Ephedrine (I.S.)
Oasis® HLB Extraction MethodOasis® HLB Extraction Plate, 30 mg/96-well
Part Number WAT058951
Conditioning:1 mL methanol
1 mL water
Load:2 mL of acidfied spiked human urine
Wash 1:1 mL 5% methanol with
2% ammonium hydroxide
2. Amphetamine 3. Methamphetamine
Wash 2:1 mL 20% methanol with 2% ammonium hydroxide
Elute:0.5 mL 20% methanol with
2% acetic acid
Minutes
28 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Anticonvulsants
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µmPart number: WAT046970Mobile phase: 100 mM potassium phosphate, pH 6.9/acetonitrile/water 20:30:50Flow rate: 1.0 mL/minInjection volume: 10 µL of11-160 µg/mL sampleTemperature: 30˚ CDetection: UV @ 214 nm
USP Tailing Factors
1. 1.412. 1.033. 1.114. 1.025. 1.07
1. Succinimide
2. Primidone
3. Phenobarbital
4. Mephenytoin
5. Mephobarbital
Minutes
29Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
HPLC Method
Column: SymmetryShield™ RP8, 3.9 x 150 mm, 5 µmPart number: WAT200655Mobile phase: 100 mM phosphate pH 7.0/acetonitrile/methanol 55:35:10Flow rate: 1.0 mL/minInjection volume: 100 µL acidified plasma sample solutionTemperature: 30˚ CDetection: UV @ 230 nm
Compounds:1. Nordoxepin2. Nortriptyline3. Norverapamil4. Doxepin5. Amitriptyline6. Verapamil7. Methoxyverapamil (I.S.)
Antidepressants in Plasma
Compound Concentration % Recovery %RSD
µg/mL (n=6)
Amitriptyline 1.46 99.9% 1.10%0.29 94.9% 0.60%
Doxepin 1.00 98.4% 0.69%0.20 98.0% 2.30%
Nordoxepin 0.50 92.8% 0.91%0.10 88.0% 1.33%
Nortriptyline 0.62 95.3% 1.29%0.12 91.1% 1.03%
Verapamil 2.50 96.7% 0.63%0.50 98.8% 0.70%
Norverapamil 1.00 95.9% 0.59%0.20 98.5% 1.04%
Chromatogram of Plasma Extracts: A) Blank B) Spiked Sample
B.
A.
0 5 10 15Minutes
0.002 AU
1
6. Verapamil
3. Norverapamil
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30 mg Extraction Cartridge
Part Number WAT094225
Condition:1 mL methanol/1 mL water
Load:1 mL of acidified plasma
sample solution
Wash:1 mL of 70% methanol,
2% ammonium hydroxide
Elute:0.5 mL of 70% methanol,
2% acetic acid
Evaporate and Reconstitute:Not required
7. Methoxyverapamil (I.S.)
1. Nordoxepin 2. Nortriptyline
5. Amitriptyline
2 34 5
6
7
4. Doxepin
30 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250 Mobile phase: 100 mM potassium phosphate, pH 6.9/acetonitrile/methanol/water 17:25:5:53Flow rate: 1.0 mL/minInjection volume: 5 µL of 25µg/mL sampleDetection: UV @ 214 nm
USP Tailing Factors
1. NA 5. 1.102. NA 6. 1.173. 1.13 7. NA4. 1.17 8. NA 1. 2-Phenyl-2ethylmalonamide
2. p-Hydroxyphenobarbital
3. Primidone
4. Phenobarbital
5. Carbamezapine-10,11-epoxide
6. Carbamezapine
Antiepileptics and Metabolites
Minutes
7. 10,11-Dihydrocarbamezapine 8. Phenytoin
31Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Antipyrine and Its Metabolite
HPLC Method
Column: SymmetryShield™ RP8, 4.6 x 50 mm, 5µmPart Number: 186000224Mobile phase: 0.1% glacial acetic acid/methanol 85:15Flow rate: 1.0 mL/minInjection volume: 5 µL solution with antipyrine (240 ng) and 4-hydroxyantipyrine (29 ng)Detection: UV @ 254 nm
1. Antipyrine
2. 4-Hydroxyantipyrine
1
2
2 4
Minutes
32 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Atrazine and Metabolites in Drinking Water
1
23
4 5
6
7
0 2010
Minutes
PDA
(0.
08 A
UFS
)
Spiked Sample
Blank Sample
Oasis® MCX SPE MethodConditions for Oasis® MCX Cartridge, 6 cc, 150 mg
Part Number 186000256
Compounds (% Recovery - % RSD, n=5)0.2 µg/L 1.0 µg/L
1. Hydroxydesisopropylatrazine 94 (3) 85 (3)2. Desethyldesisopropylatrazine 75 (8) 76 (5)3. Hydroxydesethylatrazine 89 (6) 76 (7)4. Desisopropylatrazine 79 (4) 83 (2)5. Hydroxyatrazine 107 (7) 101 (2)6. Desethylatrazine 79 (5) 83 (3)7. Atrazine 89 (5) 77 (3)
HPLC Method
Column: SymmetryShield™ RP18, 3.9 x 150 mm, 5 µmPart number: 186000108Mobile Phase A: 20 mM Phosphate buffer, pH 6.8Mobile Phase B: AcetonitrileGradient: Time Profile
(min) %A %B0 95 52 95 520 25 75
Flow Rate: 0.8 mL/minInjection volume: 80 µLDetection: PDA (215 nm)
Prepare Sample:adjust to pH 1.5 with HCI
Condition: 2 mL methanolRinse with 2 mL H2O
Load Sample
Wash #1:2 mL 0.1 N HCI
Wash #2:2 mL methanol
Elute:3 mL methanol (4% NH4OH)
Evaporate and Reconstitute
Drinking water samples (100 mL) were spikedwith the herbicides and adjusted to pH 1.5. Thesamples were then analyzed using 6 ccOasis® MCX cartridges using the protocol forbasic compounds
Minutes
33Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: 100 mM Potassium phosphate, pH 6.9/acetonitrile/water 20:30:50Flow rate: 1.0 mL/minInjection volume: 10 µL of 25µg/mL sampleDetection: UV @ 214 nm
USP Tailing Factors1. 1.17 5. 1.112. 1.12 6. 1.113. 1.14 7. 1.094. 1.10 8. 1.09
1. Barbital 2. Allobarbital
3. Phenobarbital 4. Butabarbital
5. Butalbital 6. Amobarbital
Barbiturates
8. Secobarbital7. Mephobarbital
34 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Barbiturates in Human Urine
%Recovery (n=8) (%RSD) (n=8)Compound 0.2 µg/mL 1.0 µg/mL
1. Phenobarbital 114.3 (1.7) 106.5 (0.5)
2. Butabarbital 95.7 (1.3) 105.5 (0.7)
3. Butalbital 109.5 (0.9) 104.2 (0.9)
4. Amobarbital (I.S.) 86.3 (1.7)
5. Mephobarbital 92.5 (3.6) 92.4 (1.7)
6. Secobarbital 101.5 (5.2) 94.8 (2.2)
1. Phenobarbital
HN
NH
OO
O
H3C
H C3
H3C
4. Amobarbital (I.S.)2. Butabarbital
HPLC Method
Column: SymmetryShield™ RP18, 2.1 x 150 mm, 5 µmGuard Column: Sentry™ Guard Column RP18, 3.9 x 20 mm, 5 µmPart numbers: Column - 186000111, Guard - 186000107Mobile phase: 50 mM potassium phosphate, pH 7.0/acetonitrile 71: 29Flow rate: 1 mL/minInjection volume: 80 µL urine extractTemperature: 30 ˚CDetection: UV @ 214 nm (0.350 AUFS)
Oasis® HLB Extraction MethodOasis® HLB Extraction Plate, 30 mg/96-well
Part Number WAT058951
Condition:1 mL of methanol
Equilibrate:1 mL water
Wash 1:1 mL of 5% methanol
Wash 2:1 mL 25% methanol with 2% acetic acid
Elute:1 mL 35% methanol with 2% ammonium hydroxide
Evaporated:To dryness under N2 at 40˚C
Load:2 mL water of spiked human urine
Reconstituted:300 µL of water
5. Mephobarbital3. Butalbital6. Secobarbital
0 8
BA
16
1
23
45
6
0.35
0 A
UFS
Minutes
Chromatogram of A) Blank Urine, B) Spiked Urine
Minutes
35Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Beclomethasone
HPLC Method
Column: Symmetry® C18, 4.6 x 100 mm, 3.5 µmPart number: WAT066220Mobile phase: Acetonitrile/water 65:35Flow rate: 1.4 mL/minInjection volume: 15 µL of 0.1 mg/mL of each beclomethasone-dipropionate and testosterone propionate (internal standard)Detection: UV @ 254 nm
USP Tailing Factors1. 1.12. 1.0
1. Beclomethasone Dipropionate
2. Testosterone Propionate (I.S.)
Minutes
0 105
1
2
36 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Benomyl and Bisphenol A
1
234
Minutes
0 5 10
sample
blank
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile Phase A: 10 mM phosphate, pH 6.8Mobile Phase B: MethanolGradient: Time Profile
(min) %A %B0 60 4020 0 100
Flow Rate: 1.0 mL/minInjection volume: 100 µLDetection: PDA (283 nm extracted)
Compounds % RECOVERY ± % RSD1. Benomyl 65 ± 102. Carbaryl N/A3. Atrazine N/A4. Bisphenol A 83 ± 2
100 ppb spike level in pear
Oasis® SPE Method for Endocrine DisruptorsConditions for Oasis® HLB Cartridge, 6 cc, 200 mg
Part Number WAT106202
Prepare Sample:adjust to pH 3
Condition:3 mL MTBE*
Rinse: 3 mL methanolRinse: 3 mL H2O
Load:up to 500 mL sample
Wash:3 mL 5% methanol in H2O
Elute:6 mL 10% methanol/MTBEt*
* methyl t-butyl etherdiethyl ether can be used as an alternative to MTBE
For GC analysis, dryextract over Na2SO4,
then adjust to 1 mL
For LC analysis, exchangeto acetonitrile, then
adjust to 1 mL
1. Benomyl 2. Carbaryl
3. Atrazine 4. Bisphenol A
Minutes
37Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 mm x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: Water/acetonitrile/phosphoric acid 63:37:0.1Flow rate: 1.0 mL/minInjection volume: 10 µL of 50 µg/mL sampleDetection: UV @ 254 nm
USP Tailing Factors1. 1.122. 1.09
2. Propylparaben(internal standard)
Betamethasone
1. Betamethasone
Minutes
38 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: Water/acetonitrile/phosphoric acid 63:37:0:1Flow rate: 1.0 mL/minInjection volume: 10 µL of 150 µg/mL sampleDetection: UV @ 254 nm
USP Tailing Factors1. 1.142. 1.12
1. Betamethasone
2. Propylparaben (I.S.)
Betamethasone in Syrup Formulation
Minutes
Celestone is a registered trademark of Schering Corporation.
39Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
HPLC Method
Column: Symmetry300™ C18, 4.6 x 50 mm, 5 µm Part number: WAT106209Mobile phase A: 0.1% TFA in waterMobile phase B: 0.1% TFA in acetonitrileGradient: Time Profile
(min) %A %B0 100 045 70 30
Flow rate: 1.0 mL/minInjection volume: 20 µLDetection: UV @ 214 nm
BSA Tryptic Digest
0.00 20.00 40.00
Minutes
40 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
HPLC Method
Column A: Symmetry300™ C18, 4.6 x 50 mm, 5 µm Column B: Symmetry300™ C18, 4.6 x 150 mm, 5 µmColumn C: Symmetry300™ C18, 4.6 x 250 mm, 5 µmPart numbers: A: WAT106209
B: WAT106157C: WAT106151
Mobile phase A: 0.1% TFA in waterMobile phase B: 0.1% TFA in acetonitrileGradient: Time Profile
(min) %A %B0 100 045 70 30
Flow rate: 0.75 mL/minInjection volume: 20 µL (7 µL for 4.6 x 50 mm)Temperature: 35˚CDetection: UV @ 214 nm
BSA Tryptic Digest - Column Length Effects
0.00 20.00 40.00
Minutes
0.00 20.00 40.00
Minutes
0.00 20.00 40.00
Minutes
A
B
C
41Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
HPLC Method
Column: Symmetry300™ C18, 4.6 x 50 mm, 5 µm Part number: WAT106209Mobile phase A: 0.1% TFA in waterMobile phase B: 0.1% TFA in acetonitrileGradient: Time Profile
(min) %A %B0 100 045 70 30
Injection volume: 20 µLTemperature: 35˚CDetection: UV @ 214 nm
BSA Tryptic Digest - Flow Rate Effects
0.00 20.00 40.00
Minutes
0.00 20.00 40.00
Minutes
0.00 20.00 40.00
Minutes
0.00 20.00 40.00
Minutes
0.3 mL/min
0.5 mL/min
1.0 mL/min
2.0 mL/min
42 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
HPLC Method
Column: Symmetry300™ C18, 4.6 x 50 mm, 5 µm Part number: WAT106157Mobile phase A: 0.1% TFA in waterMobile phase B: 0.1% TFA in acetonitrileGradient: Time Profile
(min) %A %B0 100 045 70 30
Flow rate: 0.75 mL/minInjection volume: 20 µLTemperature: 35˚CDetection: UV @ 214 nm
BSA Tryptic Digest - Gradient Duration Effects
0.00 20.00 40.00
Minutes
0.00 20.00 40.00
Minutes
0.00 20.00 40.00
Minutes
0.00 20.00 40.00
Minutes
0.00 20.00 40.00
Minutes
Gradient time: 15 min.
Gradient time: 25 min.
Gradient time: 50 min.
Gradient time: 150 min.
Gradient time: 75 min.
43Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Bupropion
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: 20 mM potassium phosphate, pH 7.0/methanol 35:65Flow rate: 1.0 mL/minInjection volume: 10 µL of 20 µg/mL sampleDetection: UV @ 254 nm
USP Tailing Factors1. 105
1. Bupropion
Minutes
44 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Busiperone
HPLC Method
Column: SymmetryShield™ RP18, 4.6 x 150 mm, 5 µm Part number: 186000109Mobile phase: 20 mM phosphate buffer, pH 7.0/acetonitrile 55:45Flow rate: 1.0 mL/minInjection volume: 5 µLDetection: UV @ 240 nm
USP Tailing Factors1. 1.08
1. Busiperone
Minutes
45Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Caffeine and Metabolites in Serum
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30 mg Extraction Cartridge
Part Number WAT094225
Compound Concentration % Recovery %RSD µg/mL (n=6)
Theobromine 0.100 103.0% 3.4%0.500 104.0% 2.4%
Paraxanthine 0.100 101.0% 6.2%0.500 98.9% 2.3%
Theophylline 0.100 100.0% 5.2%0.500 101.0% 2.1%
Caffeine 0.100 92.6% 4.4%0.500 97.5% 3.2%
HPLC Method
Column: SymmetryShield™ RP8, 3.0 x 150 mm, 5 µmGuard Column: Sentry™ guard column, 3.9 x 20 mm, 5 µmPart numbers: Column - WAT094243, Guard - WAT200675Mobile phase: 20 mM ammonium acetate, pH 5/acetonitrile, 95:5Flow rate: 0.6 mL/minInjection volume: 20 µL of reconstituted porcine serum extract Temperature: 25˚ CDetection: UV @ 273 nm
Compounds:1: Theobromine2: Paraxanthine3: Theophylline4: β-hydroxyethyltheophylline (I.S.)5: Caffeine
Condition1 mL methanol/1 mL water
Load1 mL porcine serum with 0.2 µgß-hydroxyethyltheophylline (I.S.)
Wash1 mL water
Elute1 mL methanol
Evaporate and Reconstitute40 ˚C under nitrogen stream
200 µL mobile phase
1. Theobromine
Chromatogram of Serum Extracts: A) Blank B) Spiked Sample
3. Theophylline2. Paraxanthine
4. ß-hydroxyethyltheophylline (I.S.)
N
NN
N
O
O CH2CH2OH
H3C
CH3
5. Caffeine
N
NN
N
O
O CH2
H3C
CH3
B
A
0.03 AU
0 5 10 15
1
2
3
4
5
Minutes
46 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Captopril
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: 0.1% phosphoric acid in water/methanol 60:40Flow rate: 1.0 mL/minInjection volume: 10 µL of 200 µg/mL captopril and 20 µg/mL of captopril disulfideTemperature: 40˚ CDetection: UV @ 220 nm
USP Tailing Factors:1. 1.02. 1.2
1. Captopril
Minutes
47Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Carbadox
HPLC Method
Column: Symmetry® C18, 4.6 x 100 mm, 3.5 µmPart number: WAT066220Mobile phase: 20 mM ammonuim acetate, pH 5.0/Acetonitrile 85:15Flow rate: 1.4 mL/minInjection volume: 15 µL of 0.025 mg/mLTemperature: 30˚ CDetection: UV @ 254 nm
USP Tailing Factors1. 1.1
1. Carbadox
Minutes
48 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Catechins - 4.6 x 20 mm Intelligent Speed™ Separation
HPLC Method
Columns: Symmetry® C18, 4.6 x 20 mm, 3.5 µm IS™
SymmetryShield™ RP18, 4.6 x 20 mm, 3.5 µm IS™
Part numbers: Symmetry - 186002090, Shield - 186002092Mobile phase A: 0.1% HCOOH in WaterMobile phase B: 0.1% HCOOH in MeOHGradient: Time Profile
(min) %A %B0.0 100 04.0 60 40
Flow rate: 3.0 mL/minInjection volume: 10 µLTemperature: 30˚CDetection: UV @ 280 nm
Compounds:1. Caffeine2. Epicatechin3. Epigallocatechin Gallate
Minutes
t0= 0.13 minMax psi= 2225
1
2 3
0.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.000.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00
3
1
2
0.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.000.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00
Minutes
t0= 0.13 minMax psi= 2111
SymmetryShield™ RP18
Symmetry® C18
OOH
OH
O
OH
OH
OH
OOH
OH
OH
OOH
OH
OH
OH
OH
N
NN
N
O
O CH2
H3C
CH3
3. Epigallocatechin Gallate2. Epicatechin1. Caffeine
49Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Cefixime
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: 10 mM sodium acetate, pH 3.8/acetonitrile 80:20Flow rate: 1.0 mL/minInjection volume: 10 µL of 100 µg/mL extracted suspension sampleDetection: UV @ 254 nm
USP Tailing Factors1. 1.27
1. Cefixime
Minutes
50 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Cefuroxime
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: 5 mM sodium acetate, pH 3.8/acetonitrile 80:20Flow rate: 1.0 mL/minInjection volume: 10 µL of 100 µg/mL sampleDetection: UV @ 254 nm
USP Tailing Factors1. 1.44
1. Cefuroxime
Minutes
51Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Cephalexin
HPLC Method
Column: Symmetry® C18, 3.0 x 150 mm, 5µmPart number: WAT054200Mobile phase: 1.25% glacial acetic acid in water/methanol 88:12Flow rate: 0.6 mL/minInjection volume: 400 µL of cephalexin (1 mg/mL) and 200 µL of 1-hydroxybenzotriazole (Internal standard, 1 mg/mL)Detection: UV @ 254 nm
USP Tailing Factors1. 1.42. 1.2
1. 1-hydroxybenzotriazole (IS)
2. Cephalexin
1
100
2
Minutes
52 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
HPLC Method
Column: Symmetry® C8, 3.9 x 150mm, 5 µmGuard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: Water/methanol/glacial acetic acid 55:45:0.1Flow rate: 1 mL/minInjection volume: 10 µL of 100 µg/mLDetection: UV @ 280 nm
USP Tailing Factors1. 1.1
1. Chloramphenicol
Chloramphenicol
Minutes
53Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Chlordiazepoxide and Metabolites
HPLC Method
Column: Symmetry® C18, 3.0 x 150 mm, 5 µmPart number: WAT054200Mobile phase: 20 mM Potassium phosphate pH 7.0/methanol/acetonitrile 49:28:23Flow rate: 0.59 mL/minInjection volume: 5 µL of 40 74 µg/mLDetection: UV @ 240 nm
USP Tailing Factors1. 1.12. 1.23. 1.14. 1.25. 1.1
1. Norchlordiazepoxide
2. Oxazepam
3. Noradiazepam
4. Chlordiazepoxide
5. Demoxepam
impurity
5 10
1
2
3
45
Minutes
54 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Chlordiazepoxide and Metabolites in Serum
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30 mg Extraction Cartridge
Part Number WAT094225
Compound Concentration % Recovery %RSD µg/mL (n=6)
1. Norchlordiazepoxide 0.200 97.8% 4.3%0.040 96.5% 5.5%
2. Oxazepam 0.200 104.0% 2.6%0.040 90.0% 2.5%
3. Nordazepam 0.200 101.0% 3.0%0.040 98.9% 1.2%
4. Chlordiazepoxide 0.200 90.0% 2.4%0.040 100.0% 4.1%
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ guard column, 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046980, Guard - WAT054225Mobile phase: 20 mM Potassium phosphate, pH 7.0/acetonitrile/methanol 56:21:23Flow rate: 1.0 mL/minInjection volume: 20 µL of reconstituted porcine serum extractDetection: UV @ 240 nm
Compounds:1: Norchlordiazepoxide2: Oxazepam3: Nordazepam4: Chlordiazepoxide5: Demoxepam (I.S.)
Condition:1 mL methanol/1 mL water
Load:1 mL spiked porcine serum
with 1.5 µg/mL demoxepam (I.S.)
Wash:1 mL 5% methanol in water
Elute:1 mL methanol
Evaporate and Reconstitute:40 ˚C under nitrogen stream
200 µL of 20 mM potassium phosphate pH7/methanol 80:20 (v/v)
5. Demoxepam
Chromatogram of Serum Extracts: A) Blank B) Spiked Sample
4. Chlordiazepoxide3. Nordazepam1. Norchlordiazepoxide 2. Oxazepam
N
N
Cl
OH
2 3
4
1
5
5 10 15 20
0.002 AU
Minutes
B
A
Minutes
55Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µmPart number: WAT046970Mobile phase: 50 mM potassium phosphate, pH 3.0/ acetonitrile 80:20Flow rate: 1.0 mL/minInjection volume: 5 µL of 120 µg/mL sampleDetection: UV @ 261 nm
USP Tailing Factors1. 1.33
1. Chlorpheniramine Maleate
Chlorpheniramine
Minutes
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Cimetidine- (SymmetryShield™ RP18)
HPLC Method
Column: SymmetryShield™ RP18, 4.6 x 250 mm, 5 µmPart number: 18600012Mobile phase: 0.3 mL phosphoric acid, methanol/water 18:82Flow rate: 1.8 mL/minInjection volume: 50 µL Detection: UV @ 220 nm
USP Tailing Factors1. 1.22
1. Cimetidine
0 1 2 3Minutes
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Cimetidine - Ion-Pair Separation
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: 6.6 mM sodium 1-hexanesulfanate, 0.04% phosphoric acid in water/methanol 76:24Flow rate: 1.0 mL/minInjection volume: 10 µL of 100 µg/mLDetection: UV @ 220 nm
USP Tailing Factors1. 1.2
1. Cimetidine
Minutes
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Cimetidine- (Symmetry® C18 )
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: 0.03% Phosphoric acid in water/methanol 90:10Flow rate: 1.0 mL/minInjection volume: 10 µL injection of 10 µg/mLDetection: UV @ 220 nm
USP Tailing Factors1. 1.2
1. Cimetidine
1
1 432
Minutes
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Clobetasol and Related Compound
HPLC Method
Column: SymmetryShield™ RP18, 4.6 x 150 mm, 5 µmPart number: 186000109Mobile phase: Acetonitrile/water/methanol 48:42:10Flow rate: 1.0 mL/minInjection volume: 10 µLDetection: UV @ 240 nm
USP Tailing Factors1. 1.072. 1.05
1. Clobetasol
2. Beclomethasone
Minutes
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Clonazepam
HPLC Method
Column: Symmetry® C18, 4.6 x 250 mm, 5 µmPart number: WAT054275Mobile phase: Water/methanol/acetonitrile 40:30:30Flow rate: 1.5 mL/minInjection volume: 25 µL of sample with 40 µg/mL of clonazepam and 80 µL/mL of o-dichlorobenzene (internal standard)Detection: UV @ 254 nm
USP Tailing Factors1. 1.082. 1.04
1. Clonazepam
2. O-dichlorobenzene (IS)
Minutes
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HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046980, Guard - WAT054225Mobile phase: 20 mM potassium phosphate, pH 2.18/methanol 75:25Flow rate: 1.0 mL/minInjection volume: 5 µL of 200 µg/mL sampleDetection: UV @ 254 nm
USP Tailing Factors1. 1.4
Cocaine
N
O
Me
OMeO
O1. Cocaine
Minutes
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Cocaine and its Metabolites in Serum
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30 mg Extraction Cartridge
Part Number WAT094225
Compound Concentration % Recovery %RSD µg/mL (n=6)
1. Benzoylecgonine 2.00 93.7% 1.7%0.40 93.6% 2.2%
2. Cocaine 2.00 98.0% 1.1%0.40 93.4% 2.9%
3. Cocaethylene 2.50 97.7% 2.8%0.50 98.2% 5.1%
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ guard column, 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250 Mobile phase A: 20 mM potassium phosphate, pH 7 Mobile phase B: MethanolGradient: Time Profile
(min) %A %B0 78 228.5 78 228.6 42 5830 42 58
Flow rate: 1.0 mL/minInjection volume: 1 mL of reconstituted porcine serum extract Detection: UV @ 235 nm
Condition1 mL methanol/1 mL water
Load1 mL serum spiked porcine sample
with 20 µL phosphoric acid
Wash1 mL 5% methanol in water
Elute1 mL methanol
Evaporate and Reconstitute
40 ˚C under nitrogen stream250 µL water:methanol 80:20 (v/v)
1. Benzoylecgonine
Chromatogram of Serum Extracts: A) Blank B) Spiked Sample
N
O
Me
OMeO
O
2. Cocaine 3. Cocaethylene
N
O
Me
OO
O
2
3
1
AB
5
0.01. AU
20 251510MinutesMinutes
63Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Codeine and Its Glucuronide Metabolite in Human Urine
Compound % Recovery %RSD 0.12 µg/mL 0.6 µg/mL
Codeine (n=3) 88.5 (2.7) 99.5 (1.2)
Codeine-6-Glucuronide (n=3) 99.3 (5.4) 98.7 (0.4)
Codeine Interday (n=6) 102.5 (3.5) 105.4 (4.0)
Codeine Interperson (n=9) 91.6 (7.1) 104.1 (5.4)
C-6-G Interday (n=6) 116.1 (9.4) 111.5 (3.3)
C-6-G Interperson (n=9) 102.4 (7.7) 107.4 (7.3)
HPLC Method
Column: Symmetry® C18, 2.1 x 150 mm, 3.5 µmGuard Column: Symmetry® C18, 2.1 x 10 mm, 3.5 µmPart numbers: Column - 186000174, Guard - 186000169Mobile phase: Methanol/Acetonitrile/0.05% TFA in water 5:5:90Flow rate: 0.3 mL/minInjection volume: 80 µL urine extractTemperature: 30˚ CDetection: UV @ 220 nm (0.04 AUFS)
Compounds:1. Ranitidine (I.S.)2. Codeine-6-Glucuronide (C-6-G)3. Codeine
Chromatogram of Urine Extracts: A) Blank B) Spiked Sample
CH3O
O H
NCH3
HO
3. Codeine2. Codeine-6-Glucuronide(C-6-G)
O
OH OH
OH
OHOC
O
O
H3CO
N-
H
CH3
Oasis® MCX Extraction MethodOasis® MCX Extraction Cartridge, 3 cc/60 mg
Part Number 186000254
Load3 mL acidified spiked human urine
Wash 12 mL 0.1N hydrochloric acid
Wash 22 mL methanol
Elute2 mL 5% ammonium hydroxide
with 95% methanol
After elution, the extract is diluted 1:3 with water for HPLC analysis
No Conditioning
Minutes0 105
1
2
3
nonspiked sample
spiked sample0.04
AU
FS
1. Ranitidine
Minutes
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Codeine and Its Glucuronide Metabolite in Porcine Plasma
HPLC Method
Column: Symmetry® C18, 2.1 x 150 mm, 3.5 µmGuard Column: Symmetry® C18, 2.1 x 10 mm, 3.5 µmPart numbers: Column - 186000174, Guard - 186000169Mobile phase: Methanol/Acetonitrile/0.05% TFA in water 5:5:90Flow rate: 0.3 mL/minInjection volume: 80 µL urine extractTemperature: 30˚ CDetection: UV @ 220 nm (0.04 AUFS)
Compounds:1. Ranitidine (I.S.)2. Codeine-6-Glucuronide (C-6-G)3. Codeine
Chromatogram of Plasma Extracts: A) Blank B) Spiked Sample
Oasis® MCX Extraction MethodOasis® MCX Extraction Cartridge, 3 cc/60 mg
Part Number 186000254
No Conditioning
Wash 1
2 mL 0.1N hydrochloric acid
Wash 2
2 mL methanol
Elute
2 mL 5% ammonium hydroxide
with 95% methanol
After elution, the extract is diluted 1:3 with water for HPLC analysis
Load
3 mL acidified plasma sample
CH3O
O H
NCH3
HO
3. Codeine2. Codeine-6-Glucuronide(C-6-G)
O
OH OH
OH
OHOC
O
O
H3CO
N-
H
CH3
1. Ranitidine
% Recovery(n=3) %RSD(n=3) Spike Level: 0.12 µg/mL 0.6 µg/mL
Codeine 95.4 (5.0) 103.4 (0.9)
Codeine-6-Glucuronide 106.8 (1.9)1 04.6 (1.3)
Minutes
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HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: 7.4 mM sodium 1-heptanesulfonate in water/acetonitrile 65:35Flow rate: 1.0 mL/minInjection volume: 8 µL of 100 µg/mL sampleDetection: UV @ 230 nm
USP Tailing Factors1. 1.282. 1.28
2. Chlorcyclizine HCL
Cyclizine and Chlorcyclizine
1. Cyclizine
Minutes
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Cytochrome c Tryptic Digest - Column Lifetime Study, Critical Pair Resolution
HPLC Method
Column: Symmetry300™ C18, 3.9 x 150 mm, 5 µmPart number: WAT106154Mobile phase A: 0.1% TFA water/acetonitrile 90:10Mobile phase B: 0.1% TFA water/acetonitrile 40:60Gradient: Time Profile
(min) %A %B0 100 045 34 66
Flow rate: 0.75 mL/minInjection volume: 20 µLTemperature: 35˚CDetection: UV @ 214 nm
Minutes Minutes
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Cytochrome c Tryptic Digest - Pore Size Effects
HPLC Method
Column: Symmetry® C18, 4.6 x 150 mm, 5 µm, 100ÅGuard Column: Symmetry300™ C18, 4.6 x 150 mm, 5 µm, 300ÅPart numbers: Column - WAT045905, Guard - WAT106157Mobile phase A: 0.1% TFA in WaterMobile phase B: 0.1% TFA in AcetonitrileGradient: Time Profile
(min) %A %B0 100 050 70 30
Flow rate: 0.75 mL/minInjection volume: 20 µLTemperature: 35˚CDetection: UV @ 214 nm
Symmetry® C18, 4.6 x 150 mm, 5 µm, 100Å
0.00 20.00 40.00
Minutes
0.00 20.00 40.00
Minutes
Symmetry300™ C18, 4.6 x 150 mm, 5 µm, 300Å
68 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Cytochrome c Tryptic Digest -Recovery Study
HPLC Method
Column: Symmetry300™ C18, 4.6 x 50 mm, 5 µmPart number: WAT106209Mobile phase A: 0.1% TFA in WaterMobile phase B: 0.1% TFA in AcetonitrileGradient: Time Profile
(min) %A %B0 100 045 70 30
Flow rate: 1.0 mL/minInjection volume: 20 µLTemperature: 35˚CDetection: UV @ 214 nm
FirstRun
SecondRun
TFA Blank InjectionMinutes
69Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Cytochrome c Tryptic Digest - Reproducibility Study
HPLC Method
Column: Symmetry300™ C18, 3.9 x 150 mm, 5 µmPart number: WAT106154Mobile phase A: 0.05% TFA in WaterMobile phase B: 0.1% TFA in AcetonitrileGradient: Time Profile
(min) %A %B0 100 045 72 28
Flow rate: 1.0 mL/minInjection volume: 20 µLTemperature: 35˚CDetection: UV @ 220 nm
Batch 107
Batch 106
Batch 104
Batch 103
Minutes
70 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: 23 mM ammonium nitrate, pH 3.4/acetonitrile 30:70Flow rate: 1.0 mL/minInjection volume: 20 µL of 100 µg/mL sampleDetection: UV @ 254 nm
USP Tailing Factors1. 0.98
1. Dextromethorphan
Dextromethorphan
1
Minutes
71Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: 23 mM ammonium nitrate, pH 3.4/acetonitrile 30:70Flow rate: 1.0 mL/minInjection volume: 20 µL of 100 µg/mL sampleDetection: UV @ 254 nm
USP Tailing Factors1. 0.95
1. Dextromethorphan
Dextromethorphan in a Cough Syrup
Minutes
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Diazepam in Serum - LC/MS
HPLC Method
Column: Symmetry® C18, 2.1 x 100 mm, 3.5 µmPart number: WAT058965Mobile phase: 2 mM Ammonium Acetate/Acetonitrile/Formic Acid 65:35:0.1Flow rate: 200 µL/minInjection volume: 10 µLMS: Micromass Quattro LCIon Mode: ES+Cone Voltage: 45 VCollision Energy: 25 eV
Diazepam
60 80 100 120 140 160 180 200 220 240 260 280 300 320 340m/z0
100
%
285.1
287.1
60 80 100 120 140 160 180 200 220 240 260 280 300 320 340m/z0
100
%
154.1
285.4
222.4193.4
182.0167.0
228.2 257.3
0.0 0.5 1.0 1.5 2.0 2.5 3.0
3.5
4.0 4.5 5.0ng/mL-0.399
771
Response
0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0ng/mL-0.399
165
Response
1.00 2.00 3.00 4.00Time0
100
%
2.26
1.00 2.00 3.00Time0
100
%
2.27
1.00 2.00 3.00 4.00Time0
100
%
2.2839
A B C
N
N
Cl
O3CH
Precursor(m/z)285
Product(m/z)154
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30mg Extraction Cartridge
Part Number WAT04225
Condition:1 mL methanol/1 mL water
Wash: 1 mL 5% methanol in water
Elute:1 mL methanol
Load:1 mL porcine serum
Evaporate and Reconstitute:40˚ C under nitrogen stream,
200 µL mobile phase
Figure 1: Background-subtracted electrospray mass spectrum of pure diazepam standard (5µg), under optimum conditions. Chromatographed as described above to remove contaminants.
Figure 2: Background-subtracted electrospray product tion spectrum of pure diazepam standard (5µg), under optimum conditions. Chromatographed as described above to remove contaminants.
Figure 4: MRM Chromatograms under optimum conditions of pure diazepam standard at (A) 0.2ng/mL (LOD) and (B) 5.0ng/mL and (C) a processed human plasma sample with
a low concentration of diazepam (calculated as 0.75ng/mL)
Compound 3 name: DiazepamCoefficent of Determination: 0.998439Calibration curve: 150.210* x + -0.398957Response type: External Std., AreaCurve type: Linear, Origin: Exclude, Weighting: 1/x Axis trans: None
Figure 3: Calibration curve with triplicate injections for each point and demonstrating LOQ at 0.2ng/mL
73Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Diclofenac Sodium
HPLC Method
Column: SymmetryShield™ RP18, 4.6 x 150 mm, 5 µmPart number: 186000109Mobile phase: 20 mM phosphate buffer, pH 7.0/acetonitrile 65:35Flow rate: 1.0 mL/minInjection volume: 5 µLDetection: UV @ 220 nm
USP Tailing Factor1. 1.18
1. Diclofenac
1
0 1 632 4 5
Minutes
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Dihydroergotamine Mesylate
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: 50 mM ammonium phosphate, pH 5.8/Acetonitrile 35:65Flow rate: 1.0 mL/minInjection volume: 10 µL of 100 µg/mL dihydroergotamine mesylateDetection: UV @ 281 nm
USP Tailing Factors1. 1.1
1. Dihydroergotamine mesylate
Minutes
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HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: 100 mM sodium acetate, 5mM camphosulfonic acid, pH 6.2/acetonitrile/methanol 40:30:30Flow rate: 1.0 mL/minInjection volume: 10 µL of 50 µg/mL sampleDetection: UV @ 254 nm
USP Tailing Factors1. 1.08
1. Dilitiazem HCL
Diltiazem HCL
Minutes
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Diltiazem HCL- Isolation of Impurities, Transfer fromAnalytical to Prep Columns
HPLC Method
Column: A. Symmetry® C18, 3.9 x 150 mm, 5 µmB. SymmetryPrep™ C18, 7.8 x 150 mm, 7 µmC. SymmetryPrep™ C18, 19 x 150 mm, 7 µm
Part numbers: A. WAT046980B. WAT066288C. WAT066240
Mobile phase: Acetonitrile/0.1% TFA in water 30:70Flow rates: A. 0.7 mL/min B. 2.8 mL/min C. 16.6 mL/minInjection volume: Diltiazem: A. 0.5 mg B. 2.0 mg C. 11.9 mgDetection: UV @ 280 nm
1. Diltiazem Hydrochloride
B. SymmetryPrep™ C18 ,7 µm, 7.8 mm x 150 mm
A. Symmetry® C18 ,5 µm, 3.9 mm x 150 mm
C. SymmetryPrep™ C18 ,7 µm, 19 mm x 150 mm
Minutes
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Diphenoxylate HCL
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: 50 mM potassium phosphate, pH 6.0/acetonitrile 30:70Flow rate: 1.0 mL/minInjection volume: 10 µL of 100 µg/mL Diphenoxylate hydrochlorideDetection: UV @ 210 nm
USP Tailing Factors1. 1.3
1. Diphenoxylate HCL
Minutes
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Disopyramide
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: 50 mM potassium phosphate, pH 7.0/methanol 70:30Flow rate: 1.0 mL/minInjection volume: 20 µL of 100 µg/mL sampleDetection: UV @ 254 nm
USP Tailing Factors1. 1.3
1. Disopyramide
Minutes
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Doxepin in Serum
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30 mg Extraction Cartridge
Part Number WAT094225
Compound Concentration % Recovery %RSD µg/mL (n=6)
Doxepin 0.200 95.8% 1.4%4.00 96.6% 1.9%
HPLC Method
Column: SymmetryShield™ RP8, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ Guard Column, 3.9 x 20 mm, 5 µmPart numbers: Column - WAT200655, Guard - WAT200675Mobile phase: 20 mM potassium phosphate, pH 7/methanol 38:62Flow rate: 1.0 mL/minInjection volume: 20 µL of reconstituted porcine serum extractTemperature: 30˚ CDetection: UV @ 254 nm
Compounds:1: Butyl paraben (I.S.)2: Doxepin
Condition1 mL methanol/1 mL water
Load1 mL spiked porcine serum
Wash1 mL 5% methanol in water
Elute1 mL methanol with 5 µg
butyl paraben (I.S.)
Evaporate and Reconstitute40 ˚C under nitrogen stream
200 µL methanol
2. Doxepin
Chromatogram of Serum Extracts: A) Blank B) Spiked Sample
1
0.001 AU
2
0 4 8 12Minutes
AB
1. Butyl paraben
Minutes
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HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µmPart number: WAT046970Mobile phase: 10 mM potassium phosphate, pH 2/acetonitrile 75:25Flow rate: 2 mL/minInjection volume: 2 µL of 1.67 mg/mLTemperature: 80˚ CDetection: UV @ 215 nm
Compounds1. Maleic Acid2. Enalapril
Enalapril
2. Enalapril1. Maleic Acid
Minutes
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Endocrine Disruptors in Soil
1
2
3
4
Minutes
0 5 10
sample
blank
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046970Mobile Phase A: 10 mM phosphate pH 6.8Mobile Phase B: MethanolGradient: Time Profile
(min) %A %B0 60 4020 0 100
Flow Rate: 1.0 mL/minInjection volume: 100 µLSample: 10 g potting soil extracted with 25 mL
acetonitrile; then SPE on Oasis® HLBDetection: PDA (225 nm extracted, 0.04 AUFS)
50 ppb spike levelCompounds % Recovery ± % RSD1. benomyl 62 ± 62. carbaryl 91 ± 43. atrazine 84 ± 54. bisphenol A 78 ± 6
Soil samples (5 g) were spiked with the appropriate compounds and extracted with 25 mL of acetonitrile (30 minutes on shaker). A 5 mL aliquotof the acetonitrile extract was diluted to 100 mL with reagent water (MilliQ)and then processed by SPE.
Oasis® SPE Method for Endocrine DisruptorsConditions for Oasis® HLB Cartridge, 6 cc, 200 mg
Part Number WAT106202
Prepare Sample:adjust to pH 3
Condition:3 mL MTBE*
Rinse: 3 mL methanolRinse: 3 mL H2O
Load:up to 500 mL sample
Wash:3 mL 5% methanol in H2O
Elute:6 mL 10% methanol/MTBEt*
* methyl t-butyl etherdiethyl ether can be used as an alternative to MTBE
For GC analysis, dryextract over Na2SO4,
then adjust to 1 mL
For LC analysis, exchangeto acetonitrile, then
adjust to 1 mL
1. Benomyl 2. Carbaryl 3. Atrazine 4. Bisphenol A
Minutes
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Enkephalins
Peptides
1: Met enkephalin: Tyr-Gly-Phe-Met
2: Leu enkephalin: Tyr-Gly-Gly-Phe-Leu
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase A: 0.1% TFAMobile phase B: 0.1% TFA in acetonitrile/water 90:10Gradient: Time Profile
(min) %A %B0 80 2030 40 60
Flow rate: 1.0 mL/minInjection volume: 5 µL of mixture with 500 µg/mL of each peptideDetection: UV @ 214 nm
USP Tailing Factors1. 0.992. 0.97
Minutes
83Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Etodolac
HPLC Method
Column: SymmetryShield™ RP18, 4.6 x 150 mm, 5 µm Part number: 186000109Mobile phase: 20 mM phosphate buffer, pH 7.0/acetonitrile 65:35Flow rate: 1.0 mL/minInjection volume: 5 µLDetection: UV @ 220 nm
USP Tailing Factors1. 1.16
1. Etodolac
Minutes
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Flavonoids in Ginkgo
1
2
3
Minutes10 2010
HPLC Method
Column: Symmetry® C18, 4.6 x 250 mm, 5 µmPart number: WAT054275Mobile Phase: 0.5% Phosphoric acid/Methanol 50:50Flow Rate: 1.5 mL/minInjection volume: 10 µLTemperature: 25° CDetection: UV @ 270nm 0.02 AUFS
Sample pre-preparation1 g sample is refluxed in 50 mL of ethanol:3M HCl (70:30) for 2.5 hrThe cooled sample is adjusted to exactly 100 mL0.3 mL of the ethanolic extract is diluted 1:10 with water
• Recovery, measured with certified standards at 100 ppm in reagent water, was 82 % for quercitin
and > 90 % for the other compounds
• All analyses gave results within ± 40 % of the expected values with the exception of the
capsule (+ 60%)
• The selective SPE extraction and cleanup procedure provided a convenient analysis of ginkgo
flavonoids in a complex matrix (Herbal One with 16 herbal ingredients)
Oasis® HLB method for flavonoids in ginkgo
Conditions for Oasis® HLB cartridge, 3 cc, 60 mgPart Number WAT094226
Condition/Equilibrate:1 mL methanol/1 mL water
Load:3 mL of diluted sample
Wash 1:2 mL 5% methanol/water
Wash 2: 2 ml methanol/50mM NH4OAc (30:70) pH 5
Dry Cartridge:air dry for 15 min with vacuum
Elute:3 ml MTBE/methanol (80:20)
Evaporate and Reconstitute:1 mL mobile phase
Compounds % RECOVERY1. quercetin 82%2. kaempferol > 90%3. isorhamnetin > 90%total: 11 mg / tablet
1. Quercetin 2. Kaempferol
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Flunitrazepam
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: 50 mM potassium phosphate pH 6.0/acetonitrile 60:40Flow rate: 1.0 mL/minInjection volume: 10 µL of 30 µg/mL flunitrazepamDetection: UV @ 219 nm
USP Tailing Factors1. 1.2
1. Flunitrazepam
N
NO
FO2N
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Flunitrazepam and Related Compounds
Condition:
1 mL methanol
Equilibrate:
1 mL water
Load:
250 µL
Wash:
1 mL, 10% methanol with acetic acid
Elute 1 mL, 40% methanol with
ammonium hydroxide
Compound % Recovery
7-Amino Nitrazepam 118
7-Amino Clonazepam 100
7-Amino Flunitrazepam 95.9
Nitrazepam 93.7
Clonazepam 101
Flunitrazepam 85.9
Flunitrazepam
N
NO
FO2N
Minutes
1
1
2
2
3
3
4
4
5
5
6
6
7 8 9 10
HPLC Method
Column: SymmetryShield™ RP8, 3.9 x 150 mm, 5 µmPart number: WAT200655Mobile phase: Water/Acetonitrile 70:30Flow rate: 1.5 mL/minInjection volume: 100 µLTemperature: AmbientDetector: UV @ 220 nm
Compounds: 1. 7-Amino Nitrazepam2. 7-Amino Clonazepam3. 7-Amino Flunitrazepam4. Nitrazepam5. Clonazepam6. Flunitrazepam
Data supplied by J. Vukovik, L. SchofieldWaters Ltd., Canada
Oasis® HLB Extraction MethodOasis® HLB Extraction Plate,30 mg 96-well
Part Number WAT058951
Minutes
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Four Drug Mixture-Warfarin, Dextromethorphan, Triprolidine, andTetracaineHPLC Method
Column: SymmetryShield™ RP18, 3.9 x 150 mm, 5 µmPart number: 186000108Mobile phase A: 25 mM Potassium phosphate, pH 7Mobile phase B: MethanolGradient: Time Profile
(min) %A %B0 85 1526 20 80
Flow rate: 1.0 mL/minInjection volume: 10 µL of mixture with 0.3 mg/mL triprolidine, 0.4 mg/mL dextromethorphan,
0.15 mg/mL tetracaine, and 0.05 mg/mL of warfarinDetection: UV @ 280 nm
USP Tailing Factors1. 1.32. 1.63. 1.14. 1.2
1. Warfarin
2. Dextromethorphan HCL
3. Triprolidine HCL
4. TetracaineMinutes
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Furosemide
HPLC Method
Column: SymmetryShield™ RP18, 4.6 x 150 mm, 5 µmPart number: 186000109Mobile phase: 20 mM phosphate buffer, pH 7.0/acetonitrile 75:25Flow rate: 1.0 mL/minInjection volume: 5 µLDetection: UV @ 220 nm
USP Tailing Factors1. 1.30
1. Furosemide
Minutes
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Hydrocortisone and Relate Compound
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: Acetonitrile/water 35:65Flow rate: 1.0 mL/minInjection volume: 10 µL of 10 µg/mL hydrocortisone and 30 µg/mL hydrocortisone-21-acetateDetection: UV @ 254 nm
USP Tailing Factors1. 1.12. 1.1
2. Hydrocortisone-21-Acetate1. Hydrocortisone
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Ibuprofen and Naproxen in Serum
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30 mg Extraction Cartridge
Part Number WAT094425
Compound Concentration % Recovery %RSD µg/mL (n=6)
Naproxen 0.100 90.0% 4.0%0.500 98.7% 1.9%2.00 99.2% 0.85%
Ibuprofen 0.500 86.8% 4.6%2.50 95.0% 3.7%
Condition1 mL methanol/1 mL water
Load
1 mL spiked porcine serum acidified with
20 µL of phosphoric acid
Wash
1 mL 5% methanol in water
Elute
1 mL methanol
Evaporate and Reconstitute
40 ˚C under nitrogen stream
200 µL of mobile phase
1. Naproxen
O
OH
CH3
CH3
O
OH
CH3
CH3
2. Ibuprofen
Chromatogram of Serum Extracts: A) Blank B) Spiked Sample
0 5 10 15
Minutes
BA
HPLC Method
Column: SymmetryShield™ RP8, 3.0 x 150 mm, 5 µmGuard Column: Sentry™ Guard column, 3.9 x 20 mm, 5 µmPart Numbers: Column - WAT094243, Guard - WAT200675Mobile phase: 1.2% glacial acetic acid in water, Acetonitrile 50:50Flow rate: 0.6 mL/minInjection: 20 µL of reconstituted porcine serum extract Temperature: 25 ˚CDetection: UV @ 254 nm
Compounds:
1: Naproxen2: Ibuprofen
91Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Insulin Chain - QC Batch Test for Symmetry300™ HPLC Columns
HPLC Method
Column: Symmetry300™ C4 , 3.9 x 150 mm, 5 µmPart number: WAT106154Mobile phases: A. 25% of 0.05% TFA in Acetonitrile / 75% of 0.05% TFA in Water
B. 32% of 0.05% TFA in Acetonitrile/ 68% of 0.05% TFA in Water (Linear Gradient from 100% A to 100% B in 14 Min.) Flow rate: 1.0 mL/minInjection volume: 4 µg, 10 µLDetection: UV @ 220 nm
Compounds:1. Insulin2. Desamino3. Deoxidized
Batch 3
Batch 2
Batch 1
92 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Insulin- Impurity Isolation, Transfer from Analytical to Prep
HPLC Method
Columns: A. Symmetry® C18, 4.6 x 150 mm, 5 µmB. SymmetryPrep™ C18, 7.8 x 300 mm, 7 µmC. SymmetryPrep™ C18, 19 x 150 mm, 7 µm
Part numbers: A. WAT045905B. WAT066235C. WAT066240
Mobile phases: A. 0.1% TFA in Water B. 0.1% TFA/acetonitrile 26% B to 33% B in 14 minutes adjusted for gradient delay on prep system
Flow rates: A. 1.0 mL/min b. 5.76 mL/min c. 17mL/min Detection: UV @ 280 nmSample Load: Bovine pancreas insulin, 10 mg/mL
A. 0.07 mg b. 3 mg c. 8.5mgHPLC Systems: A. analytical system B. and C. prep system with 0.04 in. I.D. tubing
Compounds:
1. Insulin A and B chains with 51 residues, 5733 daltons2. Impurity
A. Symmetry® C18 , 5 µm, 4.6 mm x 150 mm
B. SymmetryPrep™ C18 , 7 µm, 7.8 mm x 300 mm
C. SymmetryPrep™ C18 , 7 µm, 19 mm x 150 mm
Minutes
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Insulin- Isophase Suspension
HPLC Method
Column: Symmetry® C18, 4.6 x 150 mm, 5 µmPart number: WAT045905Mobile phase A: 0.1% TFA in Water Mobile phase B: Acetonitrile with 0.1% TFAGradient: Time Profile
(min) %A %B0 100 020 40 60
Flow rate: 1.0 mL/minInjection volume: 5 µL of sample with 150 µL of insulin suspension and 300 µL of 0.01N HCL
Suspension contains zinc-insulin crystals and protamine sulfate in buffered watersDetection: UV @ 280 nm
USP Tailing Factors1. 1.12. 1.23. 1.2
Compounds: 1. Unknown2. Insulin A and B chains with 51 residues,
5733 daltons3. Unknown
Minutes
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Ketoconazole
HPLC Method
Column: SymmetryShield™ RP18, 4.6 x 250 mm, 5 µmPart number: 186000112Mobile phase: 70% diisopropylamine solution/30% ammonium acetate solution
Diisopropylamine solution: 2 mL diisopropylamine in 1 L methanolAmmonium acetate solution: 1 g in 200 mL of water
Flow rate: 1.5 mL/minInjection volume: 20 µLDetection: UV @ 225 nm
USP Tailing Factors1. 1.032. 1.06
1. Ketoconazole
2. Terconazole
Minutes
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Lidocaine HCL
HPLC Method
Column: SymmetryShield™ RP18, 4.6 x 250 mm, 5 µmPart number: 186000112Mobile phase: 50 mL glacial acetic acid in 930 mL of water, pH adjusted
to 3.4 with 0.1 N sodium hydroxide/70% buffer/30% acetonitrileFlow rate: 1.5 mL/minInjection volume: 20 µLDetection: UV @ 254 nm
USP Tailing Factors1. 2.492. 0.99
1. Lidocaine HCL
2. Methylparaben
Minutes
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Lifetime of Symmetry® 3.5 µm Column: 10,000Injections of Sulfa Drugs
HPLC Method
Column: Symmetry® 3.5µm C18, 4.6 x 100 mm Guard Column: Sentry™ Guard column, 3.9 x 20 mm, 5 µmPart numbers: Column - WAT066220, Guard - WAT054225Mobile phase: Water/methanol/glacial acetic acid 79:20:1Flow rate: 1.5 mL/minInjection volume: 10 µL of mixture of six sulfa drugs at 10 to 39 µg/mLDetection: UV @ 254 nm
SNH
O
O
N
N
NH2SNH
O
O
N
N
NH2
S
NHN
S
O
O
NH2
S
NHN
S
O
O
NH2
S
NHN
O
O
N
NH2
CH3
S
NHN
O
O
N
NH2
CH3
1. Sulfanilamide
2. Sulfadiazine
3. Sulfathiazole
4. Sulfamerazine
5. Sulfamethazine
6. Succinylsulfathiazole
EndStart
Minutes
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Maprotiline HCL
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: 50 mM potassium phosphate pH 3.0/acetonitrile 65:35Flow rate: 1.0 mL/minInjection volume: 5 µL of 100 µg/mL maprotiline hydrochlorideDetection: UV @ 254 nm
USP Tailing Factors1. 1.4
1. Maprotiline HCL
Minutes
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Meclizine HCL
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: 7.4mM sodium 1-heptanesulfonate in water/acetonitrile 58:42Flow rate: 1.0 mL/minInjection volume: 20 µL of 25 µg /mL sampleDetection: UV @ 230 nm
USP Tailing Factors1. 1.36
1. Meclizine HCL
Minutes
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Methadone in Human Urine
%Recovery (%RSD) Compound n=3 n=9
(1 Subject) (3 Subject)(3 SPE Washes) (2 SPE Washes)
Methadone (0.5 µg/mL) 95.2 (1.0) 97.4 (3.0)
Methadone (0.1 µg/mL) 98.0 (4.8) 114.7 (8.2)
EDDP (0.2 µg/mL) 93.3 (1.0) 92.9 (2.4)
EDDP (0.4 µg/mL) 98.4 (2.9) 100.5 (5.8)
Estazolam (IS) 94.4 (3.3) 91.2 (91.2)
Chromatogram of Human Urine: A) Spiked Sample, B) Blank
Oasis® MCX Extraction MethodOasis® MCX Extraction Cartridge, 3 cc/60 mg
Part Number 186000254
Load3 mL acidified spiked human urine
Wash 12 mL 0.1M hydrochloric acid
Wash 22 mL methanol
Each elution diluted 1:1 with water for HPLC analysis
Elute2 mL 5% ammonium hydroxide
with 95% methanol
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ Guard Column, 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046980, Guard - WAT054225Mobile phase: 0.1% TFA in water/Methanol 60:40Flow rate: 1 mL/minInjection volume: 100 µL urine extractTemperature: 30˚ CDetection: UV @ 210 nm
Compounds:1. EDDP (0.2 µg/mL)2. Estazolam (I.S.) (0.2 µg/mL)3. Methadone (0.2 µg/mL)
No Conditioning
Optional Wash 32 mL of 5% ammonium hydroxide
with 60% methanol
AB
5 10 15
3
Minutes
1
2
0
0.01 AU
1. EDDP (Methadone Metabolite)
N+
CH3
CH3
CH3
3. Methadone
CH3
O
N
CH3
CH3
CH3
2. Estazolam
N
N
NN
Cl
Minutes
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HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: 20 mM potassium phosphate, pH 2.18/methanol 80:20Flow rate: 1.0 mL/minInjection volume: 15 µL of 200 µg/mLDetection: UV @ 254 nm
USP Tailing Factors1. 1.5
1. Methylphenidate HCL
Methylphenidate HCL
Minutes
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HPLC Method
Column: SymmetryShield™ RP18, 4.6 x 250 mm, 5 µmPart number: 186000112Mobile phase: 1.92 g of 1-pentansulfonic acid sodium salt monohydrate,
0.27 g of sodium acetate trihydrate and 1.14 mL glacial acetic acid in 940 mL of water; 54% methanol/46% buffer
Flow rate: 1.0 mL/minInjection volume: 30 µLDetection: UV @ 254 nm
USP Tailing Factors1. 1.292. 1.98
Metoprolol Tartrate and Oxprenolol
1. Metoprolol tartrate
2. Oxprenolol
Minutes
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HPLC Method
Column: SymmetryShield™ RP18, 4.6 x 150 mm, 5 µmPart number: 186000109Mobile phase: 20 mM phosphate buffer, pH 7.0/acetonitrile 95:5Flow rate: 1.0 mL/minInjection volume: 5 µLDetection: UV @ 254 nm
USP Tailing Factors1. 1.03
Metronidazole (SymmetryShield™ RP18)
1. Metronidazole
Minutes
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Metronidazole (Symmetry C18)
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: Water/Methanol 90:10Flow rate: 1.0 mL/minInjection volume: 10 µL of 100 µg/mL metronidazoleDetection: UV @ 254 nm
USP Tailing Factors:1. 1.2
1. Metronidazole
Minutes
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Morphine and Its Glucuronide Metabolites in Serum
6
05
1
2
3
10
Minutes
15
mV
B
A
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30 mg Extraction Cartridge
Part Number WAT094225
Compound Concentration % Recovery %RSD µg/mL (n=6)
Morphine-D3-glucuronide 0.48 90.7 2.00.097 100 3.1
Morphine-D6-glucuronide 2.4 92.2 2.80.49 93.1 2.5
Morphine 3.6 102 3.60.73 102 3.2
HPLC Method
Column: SymmetryShield™ RP8, 3.9 x 150 mm, 5 µmPart number: WAT200655Mobile phase: 20 mM potassium phosphate, pH 6.4Flow rate: 1.0 mL/minInjection volume: 100 µL of porcine serum extractDetection: Fluorescence; ex 280 nm, em 355 nm
Compounds:1: Morphine-D3-glucuronide2: Morphine-D6-glucuronide3: Morphine
Condition1 mL methanol
Equilibrate1 mL water
Load1 mL spiked porcine serum
Elute0.5 mL 3% triethylamine
No Evaporation
Sample injected directly
Chromatogram of Serum Extracts: A) Blank B) Spiked Sample
Wash1 mL water
3. Morphine
O
HO
ONCH3
H
OCOOH
OH
OHHO
1. Morphine-D3-glucuronide
HO
O H
NCH3
HO
2. Morphine-D6-glucuronide
OH
OH
COOHO O
ONCH3
H
HO
HO
Minutes
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HPLC Method
Column: SymmetryShield™ RP18, 4.6 x 250 mm, 5 µmPart number: 186000112Mobile phase: Methanol/water 35:65, 2.93 g of sodium chloride,
1.0 mL of 0.1 N hydrochloric acidFlow rate: 1.0 mL/minInjection volume: 20 µLDetection: UV @ 220 nm
USP Tailing Factors1. 1.19
Nadolol
1. Nadolol
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HO
O
NOHO
HPLC Method
Column: SymmetryShield™ RP8, 3.9 x 150 mm, 5 µmPart number: WAT200655Mobile phase: 100 mM Ammonium acetate, pH 5.0/acetonitrile 88:12Flow rate: 1.0 mL/minInjection volume: 35 µL (500 µL eluate)Detection: UV @ 281 nm
USP Tailing Factors1. 1.052. 1.04
Naltrexone
1. Naloxone (I.S.)
2. Naltrexone
Minutes
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Naltrexone in Plasma
Compound Concentration % Recovery %RSD µg/mL (n=6)
Naltrexone 3.3 108% 4.2%
HPLC Method
Column: SymmetryShield™ RP8, 3.9 x 150 mm, 5 µmPart number: WAT200655Mobile phase: 100 mM Ammonium acetate pH 5/acetonitrile 88:12Flow rate: 1.0 mL/minInjection volume: 1 mL spiked plasma with naloxone (I.S.)Temperature: 8˚ CDetection: UV @ 281 nm
Compounds:1. Naloxone (I.S.)2. Naltrexone
Chromatogram of Plasma Extracts: A) Blank B) Spiked Sample
BA
0 2 4 6 8 10
0.001 AU
1
2
HO
O
NOHO
1. Naloxone (I.S.) 2. Naltrexone
Oasis® HLB Extraction MethodOasis® HLB Extraction Plate, 30 mg/96-well
Part Number WAT058951
Condition
1 mL methanol/1 mL water
Load
1 mL spiked plasma
with naloxone (I.S.)
Wash
1) 1 mL 5% methanol containing
2% ammonium hydroxide
2) 1 mL 20% methanol containing
2% ammonium hydroxide
Elute
0.5 mL of 25% methanol
containing 2% acetic acid
Evaporate and Reconstitute
Not required
Minutes
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Naltrexone in Plasma - LC/MS
CID mass spectra
Rela
tive
Abu
ndan
ce (%
)
Mass/Charge (m/z)
100 200 300 4000
0
324
324
100
A -H2 O
0
0
328
310100
B
-H2O
HO
O
NOHO
HO
O
NOHO
Oasis® HLB Extraction MethodOasis® HLB Extraction Plate, 30 mg/96-well
Part Number WAT058951
Loading1 mL of spiked plasma
Wash 11 mL of 5% methanol/
2% ammonium hydroxide
Wash 21 mL of 20% methanol/2% ammonium hydroxide
Elution0.5 mL 25% methanol/
1% acetic acid
Dilution0.5 mL water
HPLC Method
Column: Symmetry® C8, 2.1 x 30 mm, 5 µm Part number: WAT058977Mobile phase: 10 mM NH4OH, pH 5/Acetonitrile 88:12Flow rate: 200 µL/minInjection volume: 50 µL plasma extractMS: Micromass Quattro™ IIIon Source Temp: 120˚ CCID: 1.8e-3 mBar, 20 eVDesolvation Temp: 400 ˚CNebulizer: 40 L/hrDrying gas: 400 L/hrCone volt: 56 V
Conc Average % Std. Dev (RSD%) (rel. error)ng/mL n=8
0.5 0.6 0.09 16 15
1 0.9 0.06 6 -14
5 5.1 0.2 4 3
10 10.1 0.4 4 1
20 19.7 0.8 4 -2
50 49.1 3.9 8 -2
100 101.1 3.7 4 1
Coefficient of Determination: 0.996457Calibration Curve: 0.0508208* x + 0.00306776Response type: Internal Std (20 ng/mL Naloxone), Area * ( IS Conc. / IS Area)Curve type: Linear, Origin: Exclude, Weighting: 1x, Axis trans: None
Conc. (ng/mL)
0.0 50.0 100.00
5.35
Resp
onse
0 100
0.6HPLC-MS/MS (MRM Analysis at 0.5 ng/mL and 100 ng/mL
Naltrexone m/z 342 > 324Naloxone m/z 328 > 310
Analyte:Naltrexone
Int. Std.:Naloxone
0.20 0.40 0.60 0.80 1.00 1.20 1.40 1.60 1.80Time0
100
%
0
100
%
1.38
0.380.12 0.58 1.12 1.80
1.19
342.1>324.0
779
328.0>310.02.14e4
1.38
1.19
0.20 0.40 0.60 0.80 1.00 1.20 1.40 1.60 1.80Time0
100
%
0
100
%
1.34
1.17
100 ng/mL 342.1>324.08.90e4
328.0>310.02.42e4
1.34
1.17
0.5 ng/mL
0.5 1.0 1.5 2.00
1000
100
0
1000
100
Time (min)
A
B
C
D
Condition1 mL of methanol/1mL water
109Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Naltrexone in Serum
1
2
B
A
0
0.05 AU
5 10 15
Minutes
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30 mg Extraction Cartridge
Part Number WAT094225
Compound Concentration % Recovery %RSD µg/mL (n=6)
Naltrexone 0.200 98% 3.2%
1.00 100% 3.5%
HPLC Method
Column: SymmetryShield™ RP8, 3.0 x 150 mm, 5 µmGuard Column: Sentry™ guard column, 3.9 x 20 mm, 5 µmPart numbers: Column - WAT094243, Guard - WAT200675Mobile phase: 20 mM ammonium acetate, pH 5/acetonitrile, 90:10Flow rate: 0.6 mL/minInjection volume: 20 µL of reconstituted porcine serum extractTemperature: 25˚ CDetection: UV @ 215 nm
Compounds:1: Naloxone (I.S.)2: Naltrexone
Condition1 mL methanol/1 mL water
Load1 mL spiked porcine serumwith 0.2 µg naloxone (I.S.)
Wash1 mL 5% methanol in water
Elute1 mL methanol
Evaporate and Reconstitute40 ˚C under nitrogen stream
200 µL mobile phase
1. Naloxone (I.S.)
Chromatogram of Serum Extracts: A) Blank B) Spiked Sample
HO
O
NOHO
2. Naltrexone
Minutes
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Naproxen (Symmetry® C8)
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 mm x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: Acetonitrile/water/glacial acetic acid 50:49:1Flow rate: 1.0 mL/minInjection volume: 5 µL of 100 µg/mL sampleDetection: UV @ 254 nm
USP Tailing Factors1. 1.15
1. Naproxen
111Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Naproxen (SymmetryShield™ RP18)
HPLC Method
Column: SymmetryShield™ RP18, 4.6 x 250 mm, 5 µm Part number: 186000112Mobile phase: Acetonitrile/water 46:54, 2.0 mL of glacial acetic acid in 1 L of mobile phaseFlow rate: 1.0 mL/minInjection volume: 20 µLDetection: UV @ 254 nm
USP Tailing Factors1. 1.092. 1.13
1. Naproxen
Minutes
2. Butyrophenone
OO
112 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Nicergoline
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: Acetonitrile/100 mM sodium acetate, pH 5.0 40:60Flow rate: 1.0 mL/minInjection volume: 10 µL of 30 µg/mL nicergolineDetection: UV @ 227 nm
USP Tailing Factor1. 1.2
1. Nicergoline
Minutes
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Nifedipine
HPLC Method
Column: SymmetryShield™ RP18, 4.6 x 150 mm, 5 µm Part number: 186000109Mobile phase: 20 mM phosphate buffer, pH 7.0/acetonitrile 40:60Flow rate: 1.0 mL/minInjection volume: 5 µLDetection: UV @ 240 nm
USP Tailing Factors1. 1.03
1. Nifedipine
Minutes
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Oxycodone in Serum
Compound Concentration % Recovery %RSD µg/mL (n=6)
Oxycodone 0.200 96.0% 6.4%
1.00 97.9% 1.0%
HPLC Method
Column: SymmetryShield™ RP8, 3 x 150 mm, 5 µmGuard Column: Sentry™ RP8, guard column, 3.9 x 20 mm, 5 µmPart Numbers: Column - WAT094243, Guard - WAT200675Mobile phase: 20 mM ammonium acetate, pH 5/acetonitrile, 90:10Flow rate: 0.6 mL/minInjection volume: 20 µL of reconstituted porcine serum extractTemperature: 25˚ CDetection: UV @ 215 nm
Compounds:1: Oxycodone2: Naltrexone (I.S.)
1. Oxycodone
Chromatogram of Serum Extracts: A) Blank B) Spiked Sample
H3CO
O
NCH3
OHO
2. Naltrexone (I.S.)
HO
O
NOHO
A
B
2
1
100 155Minutes
0.05 AU
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30mg Extraction Cartridge
Part Number WAT094225
Condition:1 mL methanol/1 mL water
Wash: 1 mL 5% methanol in water
Elute:1 mL methanol
Load:1 mL spiked porcine serum with
0.2 µg naloxone (I.S.)
Evaporate and Reconstitute:40˚ C under nitrogen stream,
200 µL mobile phase
Minutes
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Oxytocin and Angiotensins in Serum
Compound Concentration % Recovery %RSD µg/mL (n=6)
Oxytocin 5.0 103.7% 1.9%
Angiotensin I 5.0 97.8% 2.0%
Angiotensin II 5.0 99.2% 4.3%
HPLC Method
Column: SymmetryShield™ RP8, 4.6 x 100 mm, 3.5 µmGuard Column: Sentry™ guard column 3.9 x 20 mm, 3.5 µmPart numbers: Column - WAT094226, Guard - 186000704Mobile phase A: 0.1% TFA in water Mobile phase B: Acetonitrile/water 90:10 with 0.1% TFA
15 to 60% B in 22 minutesGradient: Time Profile
(min) %A %B0 85 1522 40 60
Flow rate: 1.4 mL/minInjection volume: 30 µL of reconstituted porcine extractTemperature: 30˚ CDetection: UV @ 214 nm
Compounds:1. Oxytocin2. Angiotensin II3. Angiotensin I (I.S.)
1
4 6 Minutes 8
0.05
2
3
B.
A.
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30 mg Extraction Cartridge
Part Number WAT094225
Condition1 mL methanol/1 mL water
Load1 mL spiked porcine serum with
20 µL of phosphoric acid
Wash1 mL 10% acetonitrile in 0.1% TFA
Elute1 mL acetonitrile/water 80:20
(with or without 0.1% TFA-no impact)
Evaporate and Reconstitute
40 ˚C under nitrogen stream1 mL 0.1% aqueous TFA (for greater
sensitivity use less volume for reconstituting sample)
1. Oxytocin 2. Angiotensin II 3. Angiotensin I
Chromatogram of Serum Extracts: A) Blank B) Spiked Sample
Minutes
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HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: 20 mM ammonium acetate, pH 5.0/acetonitrile/methanol 50:40:10Flow rate: 1 mL/minInjection volume: 5 µL of 100 µg/mLDetection: UV @ 227 nm
USP Tailing Factors1. 1.0
1. Paclitaxel (Taxol)
Paclitaxel (Taxol)
Minutes
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Paclitaxel (Taxol)- Resolution of Impurities
1. Paclitaxel (Taxol)
HPLC Method
Column: Symmetry® C18, 4.6 x 100 mm, 3.5 µmPart number: WAT066220Mobile phase: Water/acetonitrile/methanol 57:34:9Flow rate: 1.4 mL/minInjection volume: 25 µL of paclitaxel, 1 mg/mL in methanolDetection: UV @ 230 nm
Minutes
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Papaverine HCL
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: 10 mM sodium phosphate pH 3.0, 2.88 g/L sodium lauryl sulfate/acetonitrile 50:50Flow rate: 1.0 mL/minInjection volume: 10 µL of 20 µg/mL papaverine hydrochlorideDetection: UV @ 254 nm
USP Tailing Factor1. 1.1
1. Papaverine HCL
Minutes
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Paracetamol
HPLC Method
Column: SymmetryShield™ RP18, 4.6 x 150 mm, 5 µmPart number: 186000109Mobile phase: 20 mM phosphate buffer, pH 7.0/acetonitrile 85:15Flow rate: 1.0 mL/minInjection volume: 5 µLDetection: UV @ 254 nm
USP Tailing Factor1. 1.06
1. Paracetamol
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Paraquat and Diquat in Tap Water
2 64
1
2
PDA
(0.
08 A
UFS
)
257 nm
308 nm
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile Phase: 25 mM Octanesulfonate, pH 3.0/methanol 45:55Flow Rate: 0.75 mL/minInjection volume: 80 µLSample: 200 mL spiked tap water processed
on Oasis® MCX cartridgeDetection: PDA, 257 nm for Paraquat, 308 nm for Diquat
Oasis® MCX Method for QuaternaryAmines in Drinking Water
Conditions for Oasis® MCX Cartridge, 3 cc, 60 mgPart Number 186000254
Condition: 3 mL methanol
Rinse with 3 mL H2O
Load:
250 mL sample
Wash:
3 mL 50% methanol/ H2O
Elute: 2 mL of 1 M NH4CI
in 50% methanol/ H2O
For LC analysis dilute the eluent to 5 mL
with 50 mM sodium octanesulfonate
NN CH3CH3++
Compounds 2 mg/L Spike Level % Recovery, n=50.5 µg/L 2.0 µg/L
1. Paraquat 111 + 7 110 + 52. Diquat 91 + 4 96 + 4
1. Paraquat 2. Diquat
+
+N
N
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Peptides- Alberta Peptide Mixture
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase A: 0.1% TFA in water Mobile phase B: 0.1% TFA in acetonitrileGradient: Time Profile
(min) %A %B0 90 1030 60 4035 40 60
Flow rate: 0.8 mL/minInjection volume: 9 µL of peptide mixture; lyophilized sample dissolved in 1 mL mobile phaseTemperature: 35˚ CDetection: UV @ 214 nm
USP Tailing Factors1. 1.32. 1.23. 1.24. 1.25. 1.3
Peptides:Ac-Arg-Gly-x-x-Gly-Leu-Gly-Leu-Gly-Lys-amidewhere x-x=S1 Ala=Gly with free N alpha amino groupS2 Gly-GlyS3 Ala-GlyS4 Val-GlyS5 Val-Val
Minutes
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Pharmaceutical Test Mix - 2.1 x 20 mm Intelligent Speed™ Separation
HPLC Method
Columns: Symmetry® C18, 2.1 x 20 mm, 3.5 µm IS™
SymmetryShield™ RP18, 2.1 x 20 mm, 3.5 µm IS™
Part numbers: Symmetry - 186002066SymmetryShield - 186002068
Mobile phase A: 0.1% HCOOH in Water Mobile phase B: 0.1% HCOOH in ACNGradient: Time Profile
(min) %A %B0 100 04 0 100
Flow rate: 0.6 mL/minInjection volume: 5 µLTemperature: 30˚ CDetection: UV @ 254 nm
5
81
2
3 710
9
4
6
0.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00
Minutes
0.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00
109
12
3
4
75 6
0.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00
Minutes
0.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00
8
SymmetryShield™ RP18, 3.5µm
Symmetry® C18, 3.5µm
t0 = 0.20 min
Max PSI = 1424
t0 = 0.20 min
Max PSI = 1224
NH
N
N
O
NH2N
O
HOH
HH
H
HO
H
Br
NH
N
N
O
NH2N
O
HOH
HH
H
HO
H
NH
N
N
O
NH2N
O
HOH
HH
H
HO
H
BrBr
NH
O
H3C O
F
HO
O
OH
OH
OH
H
H
O
F
HO
O
OH
OH
OH
H
H
O
HO
O
OH
OH
H H
H
O
HO
O
OH
OH
H H
H
NH2NH2
O
OH
O
H H
H
O
OH
O
H H
H
BrBr
5. 2-Amino-7-chlro-5-oxo-5H-[1]benzopyrano [2,3-b]pyridine-3 carbonitrile
8. 2-Bromofluorene
6. 6a-Methyl-17a-hydroxyprogesterone
2. Acetanilide
7. 3-Aminofluorathene
3. Triamcinolone
4. Hydrocortisone
1. 8-Bromoguanosine
10. Naptho(2,3-a)pyrene9. Perylene
Compounds:1. 8-Bromoguanosine2. Acetanilide3. Triamcinolone4. Hydrocortisone5. 2-Amino-7-chloro-5-oxo-5H-[1]
benzopyrano[2,3-b]pyridine-3-carbonitrile6. 6λMethyl-17λ-hydroxyprogesterone7. 3-Aminofluoranthene8. 2-Bromofluorene9. Perylene10. Naphtho(2,3-a)pyrene
Minutes
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Pharmaceutical Test Mix - 4.6 x 20 mm Intelligent Speed™ Separation
HPLC Method
Columns: Symmetry® C18, 4.6 x 20 mm, 3.5 µm IS™
SymmetryShield™ RP18, 4.6 x 20 mm, 3.5 µm IS™
Part numbers: Symmetry - 186002090SymmetryShield - 186002092
Mobile phase A: 0.1% HCOOH in Water Mobile phase B: 0.1% HCOOH in ACNGradient: Time Profile
(min) %A %B0 100 04 0 100
Flow rate: 3.0 mL/minInjection volume: 10 µLTemperature: 30˚ CDetection: UV @ 254 nm
SymmetryShield™ RP18, 3.5µm
Symmetry® C18, 3.5µm
t0 = 0.17 min
Max PSI = 1122
t0 = 0.14 min
Max PSI = 1169
Minutes
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50
1
7
2
3
4
5
6 89
10
Minutes
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50
Minutes
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50
Minutes
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50
7
1
2
35
68
910
4
Minutes
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50
Minutes
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50
Compounds:1. 8-Bromoguanosine2. Acetanilide3. Triamcinolone4. Hydrocortisone5. 2-Amino-7-chloro-5-oxo-5H-[1]
benzopyrano[2,3-b]pyridine-3-carbonitrile6. 6λMethyl-17λ-hydroxyprogesterone7. 3-Aminofluoranthene8. 2-Bromofluorene9. Perylene10. Naphtho(2,3-a)pyrene
NH
N
N
O
NH2N
O
HOH
HH
H
HO
H
Br
NH
N
N
O
NH2N
O
HOH
HH
H
HO
H
NH
N
N
O
NH2N
O
HOH
HH
H
HO
H
BrBr
NH
O
H3C O
F
HO
O
OH
OH
OH
H
H
O
F
HO
O
OH
OH
OH
H
H
O
HO
O
OH
OH
H H
H
O
HO
O
OH
OH
H H
H
NH2NH2
O
OH
O
H H
H
O
OH
O
H H
H
BrBr
5. 2-Amino-7-chlro-5-oxo-5H-[1]benzopyrano [2,3-b]pyridine-3 carbonitrile
8. 2-Bromofluorene
6. 6a-Methyl-17a-hydroxyprogesterone
2. Acetanilide
7. 3-Aminofluorathene
3. Triamcinolone
4. Hydrocortisone
1. 8-Bromoguanosine
10. Naptho(2,3-a)pyrene9. Perylene
Minutes
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Phencyclidine in Human Urine
Compound % Recovery (n=6) %RSD (n=6) 0.3 µg/mL 1.5 µg/mL
Phencyclidine (PCP) 101.7 (2.3) 100.2 (0.8)
Ketamine (I.S.) 86.5 (1.0)
HPLC Method
Column: SymmetryShield™ RP18, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ Guard Column RP18, 3.9 x 20 mm, 5 µmPart numbers: Column - 186000108, Guard - 186000107Mobile phase: 50 mM Phosphate buffer, pH 7/Acetonitrile 70:30 Flow rate: 1 mL/minInjection volume: 20 µL urine extractTemperature: 30˚ CDetection: UV @ 210 nm (0.01 AUFS)
Compounds:1. Phencyclidine2. Ketamine HCI
Chromatogram of A) Spiked Urine, B) Blank Urine
Oasis® MCX Extraction MethodOasis® MCX Extraction Plate, 30 mg/96-well
Part Number 186000248
Load
1 mL x 4 of acidified spiked human
urine (4 mL Total Sample)
Wash 1
1 mL 0.1N hydrochloric acid
Wash 2
1 mL methanol
Elute
1 mL 5% ammonium hydroxide
with 95% methanol
14
Minutes
0 7
1
2
B
A
0.01
AU
FS
N
1. Phencyclidine (PCP) 2. Ketamine (I.S.)
CH
CIO
3
NH
Minutes
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Phenolics in Echinacea
HPLC Method
Column: Symmetry® C18, 4.6 x 250 mm, 5 µmPart number: WAT054275Mobile Phase A: 0.1% phosphoric acidMobile Phase B: AcetonitrileGradient: Time Profile
(min) %A %B0 90 1013 78 2240 60 40
Flow Rate: 1.5 mL/minInjection volume: 10 µLTemperature: 35° CDetection: UV @ 330 nm
Sample pre-preparation100 mg sample is extracted with 25 mL of ethanol:water (70:30)1 mL of the ethanolic extract is diluted 1:3 with water (~ pH 7)
• Recovery, measured with certified standards at 100 ppm in reagent water,was greater than 85 % for all compounds
• All analyses gave results within ± 35 % of the expected values with theexception of the liquid supplement (see goldenseal analysis)
• The selective SPE extraction and cleanup procedure provided a convenientanalysis of echinacea phenolics in complex matrix such as in herbal tea
Oasis® MAX method for phenolics in echinaceaConditions for Oasis® MAX Cartridge, 3 cc, 60 mg
Part Number 186000367
Condition/Equilibrate:1 mL methanol/1 mL water
Load:3 mL of diluted sample
Wash 1:2 mL of 50 mM sodium acetate
Wash 2:2 mL of methanol
Dry Cartridge:air dry for 15 min with vacuum
Elute:3 mL MTBE/methanol/TFA (49:49:2)
Evaporate and Reconstitute:1 mL mobile phase
1
2
3
Tea bagextract
Minutes10 200
1. cafteric acid2. chlorogenic acid3. cichoric acidtotal: 5 mg / teabag
Herbal tea extractbefore SPE cleanup
Herbal tea extract after SPE cleanupusing Oasis® MAX
Minutes
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Prednisone
HPLC Method
Column: SymmetryShield™ RP18, 4.6 x 150 mm, 5 µmPart number: 18600109Mobile phase: 20 mM Phosphate buffer, pH 7/Acetonitrile 65:35 Flow rate: 1.0 mL/minInjection volume: 5 µLDetection: UV @ 254 nm
USP Tailing Factor1. 1.02
1. Prednisone
Minutes
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Procainamide in Serum
Compound Concentration % Recovery %RSD µg/mL (n=6)
Procainamide 0.100 93.8% 1.8%0.500 98.1% 2.1%
HPLC Method
Column: SymmetryShield™ RP8, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ guard column, 3.9 x 20 mm, 5 µmPart numbers: Column - WAT200655, Guard - WAT200675Mobile phase: 20 mM potassium phosphate, pH 3.0/methanol 97:3Flow rate: 1.0 mL/minInjection volume: 50 µL of reconstituted porcine serum extractTemperature: 30˚ CDetection: UV @ 220 nm
Compounds:1: Procainamide2: Sulfanilamide (I.S.)
1. Procainamide
Chromatogram of Serum Extracts: A) Blank B) Spiked Sample
O
NH2 2 3
2 3
H NN
CH CH
CH CH
2. Sulfanilamide
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30 mg Extraction Cartridge
Part Number WAT094225
Condition1 mL methanol/1 mL water
Load1 mL spiked porcine serum
Wash1 mL 5% methanol in water
Elute1 mL methanol
Evaporate and Reconstitute
40 ˚C under nitrogen stream200 µL of 50 µg/mL sulfanilamide (I.S.)
in water
Minutes
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HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: 50 mM potassium phosphate, pH 3.0/methanol 85:15Flow rate: 1.0 mL/minInjection volume: 10 µL of 40 µg/mL sampleDetection: UV @ 254 nm
USP Tailing Factors1. 1.22. 1.03. 1.0
1. Procainamide
2. N-Acetylprocainamide
3. N-Propionylprocainamide
Procainamides at pH 3.0
Minutes
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HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µmPart number: WAT046970Mobile phase: 20 mM potassium phosphate, pH 6.0/acetonirtrile 90:10Flow rate: 1.0 mL/minInjection volume: 10 µL of 40 µg/mL each compoundDetection: UV @ 254 nm
USP Tailing Factors1. 1.42. 1.43. 1.4
1. Procainamide
2. N-Acetylprocainamide
3. N-Propionylprocainamide
Procainamides at pH 6.0
Minutes
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Prochlorperazine - Isolation of Degradation Products,Transfer from Analytical to Prep
HPLC Method
Columns: A. Symmetry® C18, 3.9 x 150 mm, 5 µmB. SymmetryPrep™ C18, 7.8 x 150 mm, 7 µm
Part numbers: A. WAT046980, B. WAT066288Mobile phase A: 0.1% TFA in Water Mobile phase B: AcetonitrileGradient: Time Profile
(min) %A %B0 90 1050 40 60
Flow rate: A. 0.7 mL/min B. 2.8 mL/minInjection volume: Prochlorperazine edisylate
A. 0.8 mg B. 3.2mgDetection: UV @ 280 nm
1. Prochlorperazine Edisylate
1. Symmetry® C18 , 5 µm, 3.9 mm x 150 mm
2. SymmetryPrep™ C18 , 7 µm, 7.8 mm x 150 mm
Minutes
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Profens (Gradient)-4.6x 20mm Intelligent Speed™ Separation
HPLC Method
Columns: Symmetry® C18, 4.6 x 20 mm, 3.5 µm IS™
SymmetryShield™ RP18, 4.6 x 20 mm, 3.5 µm IS™
Part numbers: Symmetry - 186002090SymmetryShield - 186002092
Mobile phase A: 0.1% HCOOH in Water Mobile phase B: 0.1% HCOOH in ACNGradient: Time Profile
(min) %A %B0 65 352 60 40
Flow rate: 3.0 mL/minInjection volume: 10 µLTemperature: 30˚ CDetection: UV @ 220 nm
Compounds:1. Ketoprofen2. Fenoprofen3. Flurbiprofen4. Ibuprofen
SymmetryShield™ RP18, 3.5µm
t0 = 0.19 min
Max PSI = 1090
13
2
4
0.40 0.60 0.80 1.00 1.20 1.40 1.60 1.80 2.000.40 0.60 0.80 1.00 1.20 1.40 1.60 1.80 2.00
2
31
4
0.40 0.60 0.80 1.00 1.20 1.40 1.60 1.80 2.000.40 0.60 0.80 1.00 1.20 1.40 1.60 1.80 2.00
Symmetry® C18, 3.5µm
t0 = 0.15 minMax PSI = 1111
Minutes
Minutes
O
O
OH
CH3O
O
OH
CH3
O
O OH
CH3
O
O OH
CH3
O
F
OH
CH3
O
F
OH
CH3
O
OH
CH3
CH3
CH3
O
OH
CH3
CH3
CH3
1. Ketoprofen
2. Fenoprofen
3. Flurbiprofen
4. Ibuprofen
132 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
3
1
2
4
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.000.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00
Profens (Isocratic) - 4.6 x 20 mm Intelligent Speed Separation
HPLC Method
Columns: Symmetry® C18, 4.6 x 20 mm, 3.5 µm IS™
SymmetryShield™ RP18, 4.6 x 20 mm, 3.5 µm IS™
Part numbers: Symmetry - 186002090SymmetryShield - 186002092
Mobile phase A: 0.1% HCOOH in Water Mobile phase B: 0.1% HCOOH in ACNIsocratic: 65% A, 35% BFlow rate: 3.0 mL/minInjection volume: 10 µLTemperature: 30˚ CDetection: UV @ 220 nm
Compounds:1. Ketoprofen2. Fenoprofen3. Flurbiprofen4. Ibuprofen
SymmetryShield™ RP18, 3.5µm
t0 = 0.18 min
Max PSI = 1082
Symmetry® C18, 3.5µm
t0 = 0.15 min
Max PSI = 1111
Minutes
1
2
43
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00
Minutes
O
O
OH
CH3O
O
OH
CH3
O
O OH
CH3
O
O OH
CH3
O
F
OH
CH3
O
F
OH
CH3
O
OH
CH3
CH3
CH3
O
OH
CH3
CH3
CH3
1. Ketoprofen
2. Fenoprofen
3. Flurbiprofen
4. Ibuprofen
133Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Promethazine- Impurities Profile
HPLC Method
Column: SymmetryShield™ RP18, 3.0 x 150 mm, 5 µmPart number: 186000692Mobile phase A: 20 mM potassium phosphate, pH 7.0Mobile phase B: AcetonitrileGradient: Time Profile
(min) %A %B0 95 530 20 80
Flow rate: 1.0 mL/minInjection volume: 10 µL of 0.5 mg/mLDetection: UV @ 254 nm
1. Promethazine HCL
Minutes
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Propranolol in Human Urine
Compound %Recovery (%RSD) 0.08 µg/mL 0.4 µg/mL
Propranolol (n=3) 105.8 (2.7) 98.5 (0.7)
Metoprolol (n=3) 101.0 (3.4) 99.6 (0.4)
Propranolol Interday (n=6) 99.0 (0.9)
Propranolol Interperson (n=9) 100.4 (6.4)
Metoprolol Interday (n=6) 100.3 (0.9)
Metoprolol Interperson (n=9) 97.3 (2.6)
Oxprenolol (I.S.) 91.1 (3.2)
Chromatogram of A) Spiked Urine, B) Blank Urine
HPLC Method
Column: SymmetryShield™ RP18, 3.9 x 150 mm, 5 µmGuard Column: SymmetryShield™ RP18, 3.9 x 20 mm, 5 µmPart numbers: Column - 186000108, Guard - 186000107 Mobile phase: 0.1% TFA in water/Acetonitrile 80:20Flow rate: 1 mL/minInjection volume: 50 µL Temperature: 30˚ CDetection: UV @ 275 nm
Compounds: 1. Metoprolol2. Oxprenolol (I.S.)3. Propranolol
Oasis® MCX Extraction MethodOasis® MCX Extraction Cartridge, 3 cc/60 mg
Part Number 186000254
Load3 mL acidified spiked human urine
Wash 12 mL 0.1M hydrochloric acid
Wash 22 mL of 100% methanol
Elute2 mL 95% methanol with 5% ammonium hydroxide
Neutralize each elution with 100 µL of acetic acid. Evaporate une N2 at 40 ˚C andreconstitute with 300 µL of water
Wash 32 mL of 50% methanol with 5% ammonium hydroxide
No Conditioning
1
23
4 8 12 160
Minutes
AB
0.00
80 A
UFS
2. Oxprenolol (I.S.)
O
OCH2
HN CH3
CH3
3. Propranolol
O
OH
NH
CH3
CH3
1. Metoprolol
O
OHHN CH3
CH3H3CO
Minutes
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Propranolol in Serum
1
2
A
B
0
Minutes
124 8
0.002 AU
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30mg Extraction
CartridgePart Number WAT094225
Compound Concentration % Recovery %RSD µg/mL (n=6)
Propranolol 0.800 85.9% 2.1%4.00 96.4% 2.4%
HPLC Method
Column: SymmetryShield™ RP8, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ guard column, 3.9 x 20 mm, 5 µmPart numbers: Column - WAT200655, Guard - WAT200675Mobile phase: 20 mM potassium phosphate, pH 7/methanol 38:62Flow rate: 1.0 mL/minInjection volume: 20 µL of reconstituted porcine serum extractTemperature: 30˚ CDetection: UV @ 254 nm
Compounds:1: Propranolol2: Butyl paraben (I.S.)
O
OHNH
Condition1 mL methanol/1 mL water
Load1 mL spiked porcine serum
Wash1 mL 5% methanol in water
Elute1 mL methanol with 5 µg
butyl paraben
Evaporate and Reconstitute
40 ˚C under nitrogen stream200 µL methanol
1. Propranolol
Chromatogram of Serum Extracts: A) Blank B) Spiked Sample
Minutes
2. Butyl paraben (I.S.)
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HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: 50mM potassium phosphate, pH 3.0/acetonitrile 85:15Flow rate: 1.0 mL/minInjection volume: 5 µL of 2.88 µg/mL sampleDetection: UV @ 261 nm
USP Tailing Factors1. 1.65
1. Pseudoephedrine HCL
Pseudoephedrine HCL
Minutes
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Pseudoephedrine HCL and Chlorpheniramine
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046970Mobile phase A: 50 mM potassium phosphate, pH 3.0Mobile phase B: AcetonitrileGradient: Time Profile
(min) %A %B0 85 151 85 1515 50 50
Flow rate: 1.0 mL/minInjection volume: 5 µL of 1.8 mg/mL pseudoephedrine and 120 µg/mL chlorpheniramine sampleDetection: UV @ 261 nm
USP Tailing Factors1. 1.72. 1.2
2. Chlorpheniramine Maleate
1. Pseudoephedrine HCL
Minutes
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HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase A: 50 mM potassium phosphate, pH 3.0Mobile phase B: AcetonitrileGradient: Time Profile
(min) %A %B0 85 151 85 1515 50 50
Flow rate: 1.0 mL/minInjection volume: 5 µL of 2.88 µg/mL pseudoephedrine and 120 µg/mL triprolidine extracted tablet sampleDetection: UV @ 261 nm
USP Tailing Factors1. 1.532. 1.16
1. Pseudoephedrine HCL
Pseudoephedrine HCL and Triprolidine HCL
2. Triprolidine HCL
Minutes
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Pseudoephedrine HCL and Codeine
HPLC Method
Column: SymmetryShield™ RP8, 3.9 x 150 mm, 5 µmPart Number: WAT200655Mobile phase A: 50 mM potassium phosphate, pH7Mobile phase B: Acetonitrile Gradient: Time Profile
(min) %A %B0 85 1515 50 50
Flow rate: 1 mL/minInjection Volume: 5 µL of 2 mg/mL pseudoephedrine HCL and 0.2 mg/mL codeineDetection: UV @ 254 nm
USP Tailing Factors1. 1.82. 1.1
1. Pseudoephedrine HCL
2. Codeine
Minutes
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Purine Alkaloids
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30 mg Extraction Cartridge
Part Number WAT094225
Compound Concentration % Recovery %RSD µg/mL (n=6)
Theobromine 0.100 103.0% 3.4%0.500 104.0% 2.4%
Paraxanthine 0.100 101.0% 6.2%0.500 98.9% 2.3%
Theophylline 0.100 100.0% 5.2%0.500 101.0% 2.1%
Caffeine 0.100 92.6% 4.4%0.500 97.5% 3.2%
HPLC Method
Column: SymmetryShield™ RP8, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ guard column, 3.9 x 20 mm, 5 µmPart Numbers: Column - WAT094243, Guard - WAT200675Mobile phase: 20 mM ammonium acetate, pH 5/acetonitrile, 95:5Flow rate: 0.6 mL/minTemperature: 25˚ CInjection volume: 20 µL of reconstituted porcine serum extract Detection: UV @ 273 nm
Compounds:1: Theobromine2: Paraxanthine3: Theophylline4: b-hydroxyethyltheophylline (I.S.)5: Caffeine
Condition1 mL methanol/1 mL water
Load1 mL porcine serum with 0.2 µgß-hydroxyethyltheophylline (I.S.)
Wash1 mL water
Elute1 mL methanol
Evaporate and Reconstitute40 ˚C under nitrogen stream
200 µL mobile phase
1. Theobromine
Chromatogram of Serum Extracts: A) Blank B) Spiked Sample
3. Theophylline2. Paraxanthine
4. ß-hydroxyethyltheophylline (I.S.)
N
N N
N
O
O CH2CH2OH
H3C
CH3
5. Caffeine
N
N N
N
O
O CH2
H3C
CH3
B
A
0.03 AU
0 5 10 15
1
2
3
4
5
141Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Quinidine Sulfate and Related Substance
HPLC Method
Column: SymmetryShield™ RP18, 4.6 x 250 mm, 5 µmPart Number: 186000112Mobile phase: 860 mL water, 20 mL methanesulfonic acid sloution, 20 mL diethylamine solution,
100 mL acetonitrile; pH adjusted to 2.6 with diethylamine solutionDiethylamine solution: 10 mL diethylamine in 100 mL water Methanesulfonic acid solution: 14 mL methanesulfonic acid and 8 mL glacial acetic acid in 200 mL water
Flow rate: 1.0 mL/minInjection volume: 50 µL Detection: UV @ 235 nm
USP Tailing Factor1. 1.762. 1.33
1. Quinidine sulfate
2. Dihydroquinidine sulfate
Minutes
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Ranitidine (SymmetryShield™ RP18)
HPLC Method
Column: SymmetryShield™ RP18, 4.6 x 150 mm, 5 µmPart Number: 18600109Mobile phase: 20 mM phosphate buffer, pH 7.0/acetonitrile, 85:15Flow rate: 1.0 mL/minInjection volume: 5 µLDetection: UV @ 254 nm
USP Tailing Factor1. 1.07
1. Ranitidine
1
Minutes
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HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: 100 mM sodium acetate, 5mM camphosulfonic acid, pH 6.2/ acetonitrile/methanol 80:10:10Flow rate: 1.0 mL/minInjection volume: 5 µL of 100 µg/ml sampleDetection: UV @ 254 nm
USP Tailing Factor1. 1.12
1. Ranitidine
Ranitidine (Symmetry® C8)
Minutes
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Ranitidine in Serum
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30mg Extraction Cartridge
Part Number WAT094225
Compound Concentration % Recovery %RSD µg/mL (n=6)
Ranitidine 0.100 104% 1.9%
0.500 102% 1.7%
HPLC Method
Column: SymmetryShield™ RP8, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ guard column, 3.9 x 20 mm, 5 µmPart numbers: Column - WAT200655, Guard - WAT200675Mobile phase: 20 mM potassium phosphate, pH 3.0/methanol 97:3Flow rate: 1.0 mL/minInjection volume: 50 µL of reconstituted porcine serum extractTemperature: 30˚ CDetection: UV @ 220 nm
Compounds:1: Sulfanilamide (I.S.)2: Ranitidine
Condition1 mL methanol/1 mL water
Load1 mL spiked porcine serum
Wash1 mL 5% methanol in water
Elute1 mL methanol
Evaporate and Reconstitute
40 ˚C under nitrogen stream200 µL of 50 µg/mL sulfanilamide (I.S.)
in water
Chromatogram of Serum Extracts: A) Blank B) Spiked Sample
O
S
HN
HN
CH3
CHNO2
N
H3C
H3C
2. Ranitidine1. Sulfanilamide
Minutes
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Salicylic Acid and Related Compoundsin Serum
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30 mg Extraction Cartridge
Part Number WAT094225
Compound Concentration % Recovery %RSD µg/mL (n=6)
Benzoic Acid 0.100 84.8% 3.8%0.500 87.1% 3.0%
Salicylic Acid 0.100 92.3% 5.0%0.500 91.3% 3.3%
HPLC Method
Column: SymmetryShield™ RP8, 3.0 x 150 mm, 5 µmGuard Column: Sentry™ guard column, 3.9 x 20 mm, 5 µmPart numbers: Column - WAT094243, Guard - WAT200675Mobile phase: 1.2% glacial acetic acid/acetonitrile 75:25Flow rate: 0.6 mL/minInjection volume: 10 µL of reconstituted porcine serum extractTemperature: 40˚ CDetection: UV @ 236 nm
Compounds:1: Benzoic acid2: Nalidixic acid (I.S.)3: Salicylic acid
Condition1 mL methanol/1 mL water
Load1 mL spiked porcine serum
acidified with 20 µL phosphoric acid
Wash1 mL 5% methanol in water
Elute1 mL methanol
Evaporate and Reconstitute
40 ˚C under nitrogen stream1 mL 5 µg/mL nalidixic acid
in mobile phase
1. Benzoic acid
Chromatogram of Serum Extracts: A) Blank B) Spiked Sample
2. Nalidixic acid (I.S.)
1
2
3
B
A
0
0.035 AU
5 10 15Minutes
3. Salicylic acid
Minutes
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Salicylic Acid and Related Compounds
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: 50 mM potassium phosphate pH 3.0/acetonitrile 70:30Temperature: 40˚ CFlow rate: 0.7 mL/minInjection volume: 6 µL of phthalic acid (0.5 mg/mL), benzoic and salicylic acids (1 mg/mL)Detection: UV @ 254 nm
USP Tailing Factors1. 1.72. 1.13. 1.4
2. Salicylic Acid
1. Phthalic Acid
3. Benzoic Acid
Minutes
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Speed Mix - 2.1 x 20 mm Intelligent Speed™ Separation
HPLC Method
Columns: Symmetry® C18, 2.1 x 20 mm, 3.5 µm IS™
SymmetryShield™ RP18, 4.6 x 20 mm, 3.5 µm IS™
Part numbers: Symmetry - 186002066SymmetryShield - 186002068
Mobile phase A: 0.1% HCOOH in Water Mobile phase B: 0.1% HCOOH in ACNGradient: Time Profile
(min) %A %B0 100 04 0 100
Flow rate: 0.6 mL/minInjection volume: 5 µLTemperature: 30˚ CDetection: UV @ 254 nm
SymmetryShield™ RP18, 3.5µm
t0 = 0.17 minMax PSI = 1452
Symmetry® C18, 3.5µm
t0 = 0.17 minMax PSI = 1247
1
2
3
4
5
6
7
89
1110
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.000.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00
1
2
3
4
56
7
89 1110
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.000.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00
Compounds:1. Theophylline2. 2-Acetylfuran3. Acetanilide4. Acetophenone5. Propiophenone6. Butyrophenone7. Benzophenone8. Valerophenone9. Hexanophenone10. Heptanophenone11. Octanophenone
O
O
O
O
O
CH3
O
CH3
OO
OO
NH
O
H3C
OO
OO
1. Theophylline 2. 2-Acetylfuran
4. Aceophenone
3. Acetanilide
5. Propiophenone 6. Butyrophenone
8. Valerophenone7. Benzophenone
OO
9. Hexanophenone
OO
10. Heptanophenone
OO
11. Octanophenone
Minutes
Minutes
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Speed Mix - 4.6 x 20 mm Intelligent Speed™ Separation
HPLC Method
Columns: Symmetry® C18, 4.6 x 20 mm, 3.5 µm IS™
SymmetryShield™ RP18, 4.6 x 20 mm, 3.5 µm IS™
Part numbers: Symmetry - 186002090SymmetryShield - 186002092
Mobile phase A: 0.1% HCOOH in Water Mobile phase B: 0.1% HCOOH in ACNGradient: Time Profile
(min) %A %B0 100 04 0 100
Flow rate: 3.0 mL/minInjection volume: 10 µLTemperature: 30˚ CDetection: UV @ 254 nm
SymmetryShield™ RP18, 3.5µm
t0 = 0.16 min
Max PSI = 1124
Symmetry® C18, 3.5µm
t0 = 0.15 min
Max PSI = 1162
34
68
910
111
5
2
7
0.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.000.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00
7
1
2
3
4
56 8 9
1011
0.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.000.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00
Compounds:1. Theophylline2. 2-Acetylfuran3. Acetanilide4. Acetophenone5. Propiophenone6. Butyrophenone7. Benzophenone8. Valerophenone9. Hexanophenone10. Heptanophenone11. Octanophenone
O
O
O
O
O
CH3
O
CH3
OO
OO
NH
O
H3C
OO
OO
1. Theophylline 2. 2-Acetylfuran
4. Aceophenone
3. Acetanilide
5. Propiophenone 6. Butyrophenone
8. Valerophenone7. Benzophenone
OO
9. Hexanophenone
OO
10. Heptanophenone
OO
11. Octanophenone
Minutes
Minutes
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HPLC Method
Column: Symmetry® C8 ,3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: Acetonitrile/water 50:50Flow rate: 1.0mL/minInjection volume: 5µL of 10 µg/mL sampleDetection: UV @ 254nm
USP Tailing Factors1. 1.082. 1.183. 1.104. 1.31
1. Prednisolone
Steroids
Vo: Uracil
2. Betamethasone
3. Prednisolone Acetate
4. Betamethasone Valerate
Minutes
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Steroids in Serum
0 4 8 12 16Minutes
BA
12 3
4
0.05 AU
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30 mg Extraction Cartridge
Part Number WAT094225
Compound Concentration % Recovery %RSD µg/mL (n=6)
Prednisolone 1.00 93.8% 4.7%5.00 92.0% 1.4%
Betamethasone 1.00 85.7% 2.3%5.00 91.3% 1.5%
Betamethasone 4.00 95.4% 2.2%
Valerate 20.00 90.6% 1.3%
HPLC Method
Column: SymmetryShield™ RP8, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ guard column, 3.9 x 20 mm, 5 µmPart numbers: Column - WAT200655, Guard - WAT200675Mobile phase: 20 mM potassium phosphate, pH 7/methanol 41:59Flow rate: 1.0 mL/minInjection volume: 5 µL of reconstituted porcine serum extractDetection: UV @ 254 nm
Compounds:1: Prednisolone2: Betamethasone3: Butyl Paraben (I.S.)4: Betamethasone Valerate
Condition1 mL methanol/1 mL water
Load1 mL spiked porcine serum
Wash1 mL 5% methanol in water
Elute1 mL methanol, added 50 µL of 0.1 mg/mL butyl paraben (I.S.)
Evaporate and Reconstitute40 ˚C under nitrogen stream
200 µL acetonitrile
4. Betamethasone valerate
Chromatogram of Serum Extracts: A) Blank B) Spiked Sample
1. Prednisolone 2. Betamethasone 3. Butyl Paraben (I.S.)
Minutes
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Steroids - 2.1 x 20 mm Intelligent Speed™ Separation
HPLC Method
Column: Symmetry® C18, 2.1 x 20 mm, 3.5 µm IS™
Part number: 186002066Mobile phase A: Water Mobile phase B: ACNGradient: Time Profile
(min) %A %B0 80 204 36 64
Flow rate: 0.6 mL/minInjection volume: 5 µLTemperature: 30˚ CDetection: UV @ 220 nm
t0 = 0.33 min
Max PSI = 1280
6
2 34 5
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.000.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00
1
1. Prednisolone 2. Cortisone
O
O
O
OH
CH3
CH3
OH
H
H H
O
O
O
OH
CH3
CH3
OH
H
H H
3. Corticosterone
O
O
CH3OH
CH3
OH
H
H
H
O
O
CH3OH
CH3
OH
H
H
H
4. Estradiol
CH3OH
OH
H
H H
CH3OH
OH
H
H H
5. Estrone
OCH3
OH
H
H
H
OCH3
OH
H
H
H
6. Progesterone
O
O
CH3
CH3
CH3
H
H
H
O
O
CH3
CH3
CH3
H
H
H
Minutes
Symmetry® C18, 3.5µmCompounds:1. Prednisolone2. Cortisone3. Corticosterone4. Estradiol5. Estrone6. Progesterone
152 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Steroids - 4.6 x 20 mm Intelligent Speed™ Separation
HPLC Method
Columns: Symmetry® C18, 4.6 x 20 mm, 3.5 µm IS™
SymmetryShield™ RP18, 4.6 x 20 mm, 3.5 µm IS™
Part numbers: Symmetry - 186002090SymmetryShield - 186002092
Mobile phase A: Water Mobile phase B: ACNGradient: Time Profile
(min) %A %B0 80 204 36 64
Flow rate: 3.0 mL/minInjection volume: 10 µLTemperature: 30˚ CDetection: UV @ 220 nm
SymmetryShield™ RP18, 3.5µm
t0 = 0.14 min
Max PSI = 1190
Symmetry® C18, 3.5µm
t0 = 0.14 min
Max PSI = 1132
Minutes
Minutes
12 3
45
6
0.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.000.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00
1
4
2 35
6
0.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.000.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00
Compounds1. Prednisone2. Cortisone3. Corticosterone4. Estradiol5. Estrone6. Progesterone
1. Prednisolone 2. Cortisone
O
O
O
OH
CH3
CH3
OH
H
H H
O
O
O
OH
CH3
CH3
OH
H
H H
3. Corticosterone
O
O
CH3OH
CH3
OH
H
H
H
O
O
CH3OH
CH3
OH
H
H
H
4. Estradiol
CH3OH
OH
H
H H
CH3OH
OH
H
H H
5. Estrone
OCH3
OH
H
H
H
OCH3
OH
H
H
H
6. Progesterone
O
O
CH3
CH3
CH3
H
H
H
O
O
CH3
CH3
CH3
H
H
H
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Succinimides
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: 100 mM potassium phosphate, pH 6.9/methanol/water 20:30:50Flow rate: 1.0 mL/minInjection volume: 10 µL of 40-500µg/mL sampleDetection: UV @ 214 nm
USP Tailing Factors1. 0.962. 0.923. 0.91
1. Ethosuximide
2. Phensuximide
3. Methsuximide
Minutes
1
2
3
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Sulfa Drugs
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5µmGuard Column: Sentry™ Guard Column 3.9 x 20 mm, 5µmPart numbers: Column - WAt046970, Guard - WAT054250Mobile phase: Water/methanol/glacial acetic acid 79:20:1Flow rate: 1.0 mL/minInjection volume: 10 µL of sample with components at 10 to 39 µg/mLDetection: UV @ 254 nm
USP Tailing Factors1. 1.272. 1.193. 1.184. 1.155. 1.126. 1.11
1. Sulfanilamide
SNH
O
O
N
N
NH2SNH
O
O
N
N
NH2
2. Sulfadiazine
S
NHN
S
O
O
NH2
S
NHN
S
O
O
NH2
3. Sulfathiazole
S
NHN
O
O
N
NH2
CH3
S
NHN
O
O
N
NH2
CH3
4. Sulfamerazine
5. Sulfamethazine 6. Succinylsulfathiazole
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Sulfa Drugs in Serum
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30 mg Extraction Cartridge
Part Number WAT094225
Compound Concentration % Recovery %RSD µg/mL (n=6)
Sulfadiazine 2.00 98.4% 2.7%10.0 96.9% 0.65%
Sulfamerazine 2.00 93.5% 1.9%10.0 96.4% 0.74%
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: 1% glacial acetic acid and 4% methanol in waterFlow rate: 1.0 mL/minInjection volume: 10 µL of reconstituted porcine serum extractTemperature: 25˚ CDetection: UV @ 254 nm
Compounds:1: Sulfadiazine2: Sulfathiazole (I.S.)3. Sulfamerazine
SNH
O
O
N
N
NH2SNH
O
O
N
N
NH2
Condition
1 mL methanol/1 mL water
Load
1 mL spiked porcine serum
with 10 µg/mL sulfathiazole (I.S.),
20 µL phosphoric acid
Wash
1 mL 5% methanol in water
Elute
1 mL methanol
Evaporate and Reconstitute
40 ˚C under nitrogen stream
1 mL mobile phase
1. Sulfadiazine
Chromatogram of Serum Extracts: A) Blank B) Spiked Sample
S
NHN
S
O
O
NH2
S
NHN
S
O
O
NH2
2. Sulfathiazole (I.S.)
S
NHN
O
O
N
NH2
CH3
S
NHN
O
O
N
NH2
CH3
3. Sulfamerazine
0.004 AU
B
A
15105
23
1
MinutesMinutes
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Sulfa Drugs - 2.1 x 20 mm Intelligent Speed™ Separation
HPLC Method
Column: Symmetry® C18, 2.1 x 20 mm, 3.5 µm IS™
Part number: 186002066Mobile phase A: 0.1% HCOOH in Water Mobile phase B: 0.1% HCOOH in MeOHGradient: Time Profile
(min) %A %B0 100 04 50 50
Flow rate: 0.6 mL/minInjection volume: 10 µLTemperature: 30˚ CDetection: UV @ 270 nm
Compounds:1. Sulfadiazine2. Sulfathiazole3. Sulfamerazine4. Sulfadimidine5. Sulfamethoxazole6. Sulfisoxazole7. Sulfadimethoxine
Symmetry® C18, 3.5µm
t0 = 0.17 min
Max PSI = 2050
Minutes
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.000.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00
1
32
4
5
6 7
SNH
O
O
N
N
NH2SNH
O
O
N
N
NH2
1. Sulfadiazine
S
NHN
O
O
N
NH2
CH3
S
NHN
O
O
N
NH2
CH3
3. Sulfamerazine
S
NHN
S
O
O
NH2
S
NHN
S
O
O
NH2
2. Sulfathiazole
S
NHN
N
O
O
NH2
CH3
CH3
S
NHN
N
O
O
NH2
CH3
CH3
4. Sulfadimidine
S
NHN
OO
O
NH2CH3
S
NHN
OO
O
NH2CH3
5. Sulfamethoxazole
SNH
N
O
O
O
CH3 NH2
CH3
SNH
N
O
O
O
CH3 NH2
CH3
6. Sulfisoxazole
S
NH
NN
O
O
O
O
NH2
CH3
CH3 S
NH
NN
O
O
O
O
NH2
CH3
CH3
7. Sulfadimethoxine
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Sulfa Drugs -4.6 x 20 mm Intelligent Speed™ Separation
HPLC Method
Column: Symmetry® C18, 4.6 x 20 mm, 3.5 µm IS™
Part number: 186002090Mobile phase A: 0.1% HCOOH in Water Mobile phase B: 0.1% HCOOH in MeOHGradient: Time Profile
(min) %A %B0 100 04 50 50
Flow rate: 3.0 mL/minInjection volume: 10 µLTemperature: 30˚ CDetection: UV @ 270 nm
Compounds:1. Sulfadiazine2. Sulfathiazole3. Sulfamerazine4. Sulfadimidine5. Sulfamethoxazole6. Sulfisoxazole7. Sulfadimethoxine
Symmetry® C18, 3.5µm
t0 = 0.15 min
Max PSI = 2280
Minutes
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.000.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00
53
1
4
26 7
SNH
O
O
N
N
NH2SNH
O
O
N
N
NH2
1. Sulfadiazine
S
NHN
O
O
N
NH2
CH3
S
NHN
O
O
N
NH2
CH3
3. Sulfamerazine
S
NHN
S
O
O
NH2
S
NHN
S
O
O
NH2
2. Sulfathiazole
S
NHN
N
O
O
NH2
CH3
CH3
S
NHN
N
O
O
NH2
CH3
CH3
4. Sulfadimidine
S
NHN
OO
O
NH2CH3
S
NHN
OO
O
NH2CH3
5. Sulfamethoxazole
SNH
N
O
O
O
CH3 NH2
CH3
SNH
N
O
O
O
CH3 NH2
CH3
6. Sulfisoxazole
S
NH
NN
O
O
O
O
NH2
CH3
CH3 S
NH
NN
O
O
O
O
NH2
CH3
CH3
7. Sulfadimethoxine
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Sulfamethoxazole and Trimethoprim
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: Water/methanol/glacial acetic acid 79:20:1Flow rate: 1.0 mL/minInjection volume: 10 µL of 100 µg/mL sulfamethoxazole and 30 µg/mL trimethroprimDetection: UV @ 254 nm
USP Tailing Factors1. 1.52. 1.1
1. Trimethoprim
2. Sulfamethoxazole
Minutes
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Sulfamethoxazole and Trimethoprimin Serum
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30 mg Extraction Cartridge
Part Number WAT094225
Compound Concentration % Recovery %RSD µg/mL (n=6)
Sulfamethoxazole 1.20 90.4% 1.3%0.240 95.4% 3.4%
Trimethoprim 1.50 96.7% 1.1%0.300 99.3% 3.2%
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ guard column, 3.9 x 20 mm, 5 µm Part number: Column - WAT046980Mobile phase: 25 mM ammonium acetate, pH 5/methanol 82:18 (v/v)Flow rate: 1.0 mL/minInjection volume: 20 µL of reconstituted porcine serum extractDetection: UV @ 290 nm
Compounds:1: Sulfamerazine (I.S.)2: Trimethoprim3: Sulfamethoxazole
Condition1 mL methanol/1 mL water
Load1 mL spiked porcine serum
with 20 µL phosphoric acid and2.5 µg/mL sulfamerazine (I.S.)
Wash1 mL 5% methanol in water
Elute1 mL methanol
Evaporate and Reconstitute
40 ˚C under nitrogen stream200 µL mobile phase
1. Sulfamerazine (I.S.)
` 5 10 15 20Minutes
00.02 AU
Chromatogram of Serum Extracts: A) Blank B) Spiked Sample
B.
A.
1
2
2. Trimethoprim 3. Sulfamethoxazole
3
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Symmetry® C18 pH 3 Batch Test
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: 50 mM potassium phosphate pH 3.0/acetonitrile 80:20Flow rate: 1.0 mL/minInjection volume: 20 µL of mixture with chlorpheniramine, maleate, toluamide and propranololDetection: UV @ 230 nm
Compounds1. Maleate2. Toluamide3. Chlorpheniramine4. Propranolol
O
OH
NH
CH3
CH3
1. Maleate
2. Toluamide
3. Chlorpheniramine
4. Propranolol
Minutes
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Synthetic Antibiotics
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: 0.1% phosphoric acid in water/methanol 62:38Flow rate: 1.0 mL/minInjection volume: 10 µL of mixture with 30 µg/mL furazolidone and oxolinic acid and 100 µg/mL nolidixic acidDetection: UV @ 254 nm
USP Tailing Factors1. 1.22. 1.13. 1.2
1. Furazolidone
2. Oxolinic Acid
3. Nalidixic Acid
Minutes
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HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µmPart number: WAT046970Mobile phase: 50 mM potassium phosphate, pH 3/acetonitrile 55:45Flow rate: 1.0 mL/minInjection volume: 5 µL of 100 µg/mL Detection: UV @ 254 nm
USP Tailing Factors1. 1.3
Tamoxifen
1. Tamoxifen Citrate
Minutes
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Tamoxifen- Isolation of Impurities, Transfer from Analytical to Prep
HPLC Method
Columns: Symmetry® C18, 3.0 x 150 mm, 5 µmSymmetryPrep™ C18, 7.8 x 300 mm, 7 µm
Part numbers: Symmetry - WAT054200SymmetryPrep - WAT066235
Mobile phase: Acetonitrile/50 mM potassium phosphate buffer, pH 3.0 40:60Flow rate: A. 0.4 mL/min B. 5.6 mL/minInjection volume: 5 mg/mL tamoxifen citrate
A. 15 mg B. 101mgDetection: UV @ 254 nm
1. Tamoxifen Citrate
1. Symmetry® C18 , 5 µm, 3.0 mm x 150 mm
1
1. SymmetryPrep™ C18 , 7 µm, 7.8 mm x 300 mm
Minutes
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HPLC Method
Column: SymmetryShield™ RP18, 4.6 x 150 mm, 5 µmPart number: 186000109Mobile phase: 20 mM phosphate buffer, pH 7.0/acetonitrile 25:75Flow rate: 1.0 mL/minInjection volume: 5 µL Detection: UV @ 220 nm
USP Tailing Factor1. 1.04
Terfenadine
1. Terfenadine
Minutes
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HPLC Method
Column: Symmetry® C8, 4.6 x 250 mm, 5 µmPart number: WAT054270Mobile phase: 100 mM triethylammonium phosphate, pH 7.0/methanol 20:80Flow rate: 1.0 mL/minInjection volume: A. 15 µL of mixture with 0.6 mg/mL terfenadine and 0.15 mg/mL
of terfenadine related compound ARSB.15 µL of 8mg/mL crushed tablet ( equivalent to 0.6mg/mL terfenadine)
Detection: UV @ 235 nm
Compounds:1. Terfenadine2. Terfenadine Related Compound ARS
USP Tailing Factor1. 1.0
1. Terfenadine
Terfenadine and Related Compound
Tablet:
Standard:
Minutes
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Testosterone Esters in Serum
Oasis® HLB Extraction MethodOasis
®HLB 1 cc/30 mg Extraction Cartridge
Part Number WAT094225
Compound Concentration % Recovery %RSD µg/mL (n=6)
Testosterone 1.00 98% 1.2%Acetate 0.200 94% 3.7%
Testosterone 1.00 100% 1.9%Propionate 0.200 94% 4.3%
Testosterone 1.00 94% 1.6%Benzoate 0.200 87% 3.2%
HPLC Method
Column: Symmetry® C8, 4.6 x 75 mm, 3.5 µmGuard Column: Sentry™ guard column, 3.9 x 20 mm, 5 µmPart Numbers: Column - WAT066224, Guard - WAT064225Mobile phase: Acetonitrile/water 60:40Flow rate: 1.5 mL/minInjection Volume: 20 µL of reconstituted porcine serum extractDetection: UV @ 254 nm
Compounds:1: Testosterone acetate2: Testosterone propionate3: Testosterone benzoate4: Testosterone enanthate (I.S.)
Condition1 mL methanol/1 mL water
Load1 mL sample with 800 µL spiked porcine
serum and 200 µL of methanol
Wash1 mL 5% methanol in water
Elute1 mL methylene chloride: methanol 50:50 (v/v)
Evaporate and Reconstitute40 ˚C under nitrogen stream
500 µL methanol with 5 µg/mL of testosterone enanthate
3. Testosterone benzoate
Chromatogram of Serum Extracts: A) Blank B) Spiked Sample
1. Testosterone acetate 2. Testosterone propionate 4. Testosterone enanthate (I.S.)
Minutes0 10 20
1
2
34
B
A
0.008 AU
Minutes
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Tetracyclines
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: 10 mM sodium phosphate, pH 6.25/methanol 45:55Flow rate: 1.0 mL/minInjection volume: 20 µL of 500 µg/mL sampleDetection: UV @ 280 nm
USP Tailing Factors1. 1.052. 1.293. 1.14
1. Doxycycline
2. Minocycline
3. Tetracycline
Minutes
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Tetracyclines in Serum
10
0.006
3
2
1
20 30
B
A
Oasis® HLB Extraction MethodOasis® HLB 1 cc/30 mg Extraction Cartridge
Part Number WAT094225
Compound Concentration % Recovery %RSD µg/mL (n=6)
Minocycline 2.50 96.1% 1.3%0.500 88.0% 1.6%
Tetracycline 2.50 101.0% 0.5%0.500 97.8% 1.4%
HPLC Method
Column: SymmetryShield™ RP8, 3.0 x 150 mm, 5 µmGuard Column: Sentry™ guard column, 3.9 x 20 mm, 5µmPart numbers: Column - WAT094243, Guard - WAT200675Mobile phase: 0.1% TFA in Water, Methanol, Acetonitrile 91:2:7Flow rate: 0.90 mL/minInjection volume: 20 µL of reconstituted porcine serum extractDetection: UV @ 350 nm
Compounds:1: Minocycline2: Tetracycline3: Demeclocycline (I.S.)
Condition1 mL methanol/1 mL water
Load1 mL spiked porcine serum
with 20 µL phosphoric acid and5.0 µg/mL demeclocycline (I.S.)
Wash1 mL 5% methanol in water
Elute1 mL methanol
Evaporate and Reconstitute
40 ˚C under nitrogen stream200 µL mobile phase
2. Tetracycline
Chromatogram of Serum Extracts: A) Blank B) Spiked Sample
1. Minocycline 3. Demeclocycline (I.S.)
Minutes
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Theophylline and Related Compounds
HPLC Method
Column: SymmetryShield™ RP18, 3.0 x 150 mm, 5 µmPart number: 186000692Mobile phase: 5 mM potassium phosphate, pH 2.0/acetonitrile 95:5Flow rate: 0.6 mL/minInjection volume: 5 µL injection of solution containing theobromine (0.22 mg/mL),
theophylline (0.26 mg/mL), and 7-(2-hydroxyethyl) theophylline (0.13 mg/mL)Detection: UV @ 270 nm
1. Theobromine
2. Theophylline
3. 7-(2-hydroxyethyl) Theophylline
Minutes
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HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ Guard Column 3.9 x 20mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: 10 mM potassium phosphate, pH7.0/ methanol 76:24Flow rate: 1.0 mL/minInjection volume: 20 µL of 10 µg/mL sampleDetection: UV @ 280 nm
USP Tailing Factors1. 1.182. 1.033. 1.01
2. Theophylline
Theophylline and Theobromine
1. Theobromine
N
N N
N
O
O CH2
H3C
CH3
3. Caffeine
Minutes
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HPLC Method
Column: SymmetryShield™ RP8, 3.9 x 150 mm, 5 µmPart number: 186000108Mobile phase: 50 mM phosphate, pH7.0/methanol 36:64Flow rate: 1.0 mL/minInjection volume: 25 µLDetection: UV @ 250 nm
USP Tailing Factors1. 1.312. 1.213. 1.12
Trazodone
1. 1-(3-chlorophenyl)peperazine
3. Trazodone
2. Maprotiline (I.S.)
Minutes
Oasis® HLB Extraction MethodConditions for Oasis® HLB Cartridge, 3 cc, 60 mg
Part Number WAT094226
Condition: 3 mL methanolRinse: 3 mL H2O
Load:75 mL sample @ 4 mL/min
Wash:1 mL H2O
Elute:1.2 mL methanol
Evaporate to 0.2 mL
Reconstitute to exactly 1.0 mL with H2O
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Compounds % RecoveryTap Water Tap WaterSpike Level Spike Level500 µg/L 200 ng/L5 replicates 7 replicates
1. Desisopropylatrazine 98.4 (5.0) 95.6 (5.8)2. Hydroxyatrazine 132 (1.3) 109 (11)3. Desethylatrazine 106 (5.1) 104 (4.0)4. Simazine not determined 97.7 (3.9)5. Cyanazine not determined 93.1 (3.7)6. Atrazine 101 (5.0) 101 (4.4)
Triazine Herbicides in Drinking Water
HPLC Method
Column: SymmetryShield™ RP8, 3.9 x 150 mm, 5 µmPart number: WAT0200655Mobile Phase A: 5 mM phosphate buffer, pH 6.7/Acetonitrile 85:15 Mobile Phase B: AcetonitrileGradient: Time Profile
(min) %A %B0 100 02 100 025 30 70
Flow Rate: 1.0 mL/minInjection volume: 75 µLDetection: UV @ 214 nm (0.02 AUFS)
1
2
34
5 6
Minutes0 5 10 15
blank cartridge
spiked well water
nonspiked sample
6. Atrazine
3. Desethylatrazine2. Hydroxyatrazine
4. Simazine 5. Cyanazine
1. Desisopropylatrazine
Minutes
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HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: 20 mM potassium phosphate, pH 7/methanol 35:65Flow rate: 1.0 mL/minInjection volume: 20 µL of 100 µg/mL sampleDetection: UV @ 254 nm
USP Tailing Factors1. 1.72. 1.73. 1.84. 1.55. 1.46. 1.2
Tricyclic Antidepressants
1. Protriptyline HCL
2. Nortriptyline HCL
3. Doxepin HCL
4. Imipramine HCL
5. Amtriptyline HCL
6. TrimipramineMinutes
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HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart number: WAT046980Mobile phase: 20 mM potassium phosphate, pH 7.0/methanol 29:71Flow rate: 1.0 mL/minInjection volume: 5 µL of 1 to 1.16 mg/mLTemperature: 35˚ CDetection: UV @ 254 nm
USP Tailing Factors1. 1.52. 1.53. 1.34. 1.2
1. Nordoxepin HCL
2. Doxepin HCL
4. Amtriptyline HCL
3. Nortriptyline HCL
Tricyclic Antidepressants and Metabolites
Minutes
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Tricyclic Antidepressants in Plasma
500 ng/mL 100 ng/mL
Compound Recovery RSD Recovery RSD(n=96) (n=95)
Nortriptyline 92.3% 1.4% 90.8% 5.7%
Doxepin 90.6% 1.4% 90.4% 4.7%
Imipramine 92.2% 1.7% 86.4% 5.3%
Amitriptyline 90.2% 1.6% 85.3% 5.8%
Trimipramine 90.3% 1.9% 89.8% 6.1%
0 1 2 3 4
B
A
0.02 AU
1
2 34
56
Chromatogram of Plasma Extracts: A) Blank B) Spiked Sample
Oasis® HLB Extraction MethodOasis® HLB Extraction Plate, 30 mg/96-well
Part Number WAT058951
Condition1 mL methanol/1 mL water
Load1 mL spiked plasma plus
20 µL concentrated phosphoric acid
WashA. 1 mL 2% ammonium
hydroxide in 5% methanolB. 1 mL 2% ammonium
hydroxidein 65% methanolC. 1 mL 2% acetic acid in 5% methanol
Elute600 µL 65% methanol
Add internal standard (I.S.)60 µL 36 µg/mL nordoxepin in
10% ammonium hydroxide
HPLC Method
Column: SymmetryShield™ RP8, 4.6 x 75 mm, 3.5 µmPart number: WAT094263Mobile phase: 50 mM phosphate, pH 7/methanol 26:74Flow rate: 1.4 mL/minInjection volume: 1 mL spiked plasma plus 20 µL concentrated phosphoric acid, 100 µL injectionTemperature: 29˚ CDetection: UV @ 254 nm
Compounds:Spiked at 500 ng/mL versus Blank:1. Nordoxepin (I.S.)2. Nortriptyline3. Doxepin4. Imipramine5. Amitriptyline6. Trimipramine
3. Doxepin1. Nordoxepin 2. Nortriptyline
4. Imipramine 5. Amitriptyline 6. Trimipramine
Minutes
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HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20 mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: 50 mM potassium phosphate, pH 3.0/acetonitrile 85:15Flow rate: 1.0 mL/minInjection volume: 10 µL of 200 µg/mL sampleDetection: UV @ 261 nm
USP Tailing Factor1. 1.6
Tripelennamine
1. Tripelennamine
Minutes
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Verapamil
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µm Guard Column: Sentry™ Guard Column 3.9 x 20mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250 Mobile phase: 10 mM sodium acetate with 3.3% glacial acetic acid, pH 2.6/acetonitrile 70:30Flow rate: 1.0 mL/minInjection volume: 10 µL of 100 µg/mL sampleDetection: UV @ 278 nm
USP Tailing Factor1. 1.27
1. Verapamil
Minutes
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Verapamil and Norverapamil
HPLC Method
Column: SymmetryShield™ RP8, 3.9 x 150 mm, 5 µm Part number: WAT200655Mobile phase: 50 mM phosphate pH 7.0/acetonitrile/methanol 41:37:22Flow rate: 1.0 mL/minInjection volume: 40 µLDetection: UV @ 230 nm
USP Tailing Factors1. 1.102. 1.063. 1.07
1. Norverapamil
2. Verapamil
3. Methoxyverapamil (I.S.)
Minutes
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HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µmGuard Column: Sentry™ Guard Column 3.9 x 20mm, 5 µmPart numbers: Column - WAT046970, Guard - WAT054250Mobile phase: Methanol/acetonitrile/water 45:45:10Flow rate: 1.0 mL/minInjection volume: 10 µL of 25 µg/mL sampleDetection: UV @ 280 nm
USP Tailing Factors1. 1.332. 1.203. 1.16 Vo. Uracil
1. Trans-Retinol (Vitamin A)
2. Cholecalciferol (Vitamin D3)
3. Alpha-Tocopherol (Vitamin E)
Vitamins- Fat-Soluble
Minutes
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Vitamins- Water-Soluble
1. Niacinamide
6. Riboflavin
7. Thiamine Hydrochloride
2. Calcium Pantothenate
3. Pyridoxal
4. Pyrodoxine Hydrochloride
5. Pyridoxamine Dihydrochloride
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart numbers: Column - WAT046980, PIC B6 - WAT084199Mobile phase: 20 mM PIC B6 (hexane sulfonic acid) and 0.1% phosphoric acid,
pH 2.2/methanol 75:25Flow rate: 0.7 mL/minInjection volume: 10 µL of mixture with component 1 at 11 µg/mL, 2 at 133 µg/mL,
3 at 17 µg/mL, 4 at 6 µg/mL, 5 at 25 µg/mL, 6 at 34 µg/mL, and 7 at 67 µg/mL
Detection: UV @ 210 nm
USP Tailing Factors1. n.c.2. n.c.3. 1.44. 1.15. 1.26. 1.17. 1.2
Minutes
181Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
HPLC Method
Column: Symmetry® C18, 3.9 x 150 mm, 5 µmPart numbers: Column - WAT046980, PIC B6 - WAT085140Mobile phase: 20 mM PIC B6 (hexane sulfonic acid) and 0.1% phosphoric acid, pH 2.2/methanol 75:25Flow rate: 0.7 mL/minInjection volume: 10 µL of mixture with 0.02 to 0.17 mg/mLTemperature: 40˚ CDetection: UV @ 280 nm
USP Tailing Factors1. 1.62. 1.43. 1.24. 1.2
Vitamins- Water-Soluble Folic Acid Mixture
1. Nicergoline
2. Folic Acid
3. Riboflavin
4. Thiamine Hydrochloride
Minutes
182 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
HPLC Method
Column: Symmetry® C8, 3.9 x 150 mm, 5 µmPart number: WAT046970Mobile phase: 50 mM potassium phosphate, pH 4.6/methanol 85:15Flow rate: 1.0 mL/minInjection volume: 20 µL of 100 µg/mL sampleDetection: UV @ 254 nm
USP Tailing Factor1. 1.2
1. Zidovudine
Zidovudine (AZT)
Minutes
183Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Particle Part No. Part No.Dimension Size C18 C81.0 x 50 mm 3.5 µm WAT106056 WAT1060521.0 x 150 mm 3.5 µm WAT248059 WAT2480722.1 x 30 mm 3.5 µm WAT058973 WAT0589772.1 x 50 mm 3.5 µm WAT200650 WAT2006242.1 x 100 mm 3.5 µm WAT058965 WAT0589612.1 x 150 mm 3.5 µm WAT106005 WAT1060113.0 x 100 mm 3.5 µm 186000696 1860006983.0 x 150 mm 3.5 µm 186000695 1860006974.6 x 30 mm 3.5 µm 186000271 1860002704.6 x 50 mm 3.5 µm WAT200625 WAT2006204.6 x 75 mm 3.5 µm WAT066224 WAT0662004.6 x 100 mm 3.5 µm WAT066220 WAT0662044.6 x 150 mm 3.5 µm WAT200632 WAT200630
2.1 x 50 mm 5 µm 186000206 1860002122.1 x 150 mm 5 µm WAT056975 WAT0569553.0 x 150 mm 5 µm WAT054200 WAT0542303.0 x 250 mm 5 µm 186000690 1860006913.9 x 150 mm 5 µm WAT046980 WAT0469704.6 x 50 mm 5 µm 186000207 1860002134.6 x 150 mm 5 µm WAT045905 WAT0459954.6 x 250 mm 5 µm WAT054275 WAT054270
Symmetry® Analytical Columns
Particle Part No. Part No.Dimension Size C18 C82.1 x 20 mm 3.5 µm 186000269 1860002682.1 x 50 mm 3.5 µm 186000152 1860001492.1 x 100 mm 3.5 µm 186000151 1860001532.1 x 150 mm 3.5 µm 186000150 1860001484.6 x 75 mm 3.5 µm WAT066260 WAT0662104.6 x 100 mm 3.5 µm WAT066265 WAT066215
3.9 x 50 mm 5 µm WAT054220 WAT0542403.9 x 150 mm 5 µm WAT054205 WAT0542354.6 x 150 mm 5 µm WAT054210 WAT0542554.6 x 250 mm 5 µm WAT054215 WAT054245
Symmetry® Cartridge Columns (Requires endfittings, see Page 186)
Particle Part No. Part No.Dimension Size C18 C82.1 x 10 mm 3.5 µm WAT106127 WAT1061283.9 x 20 mm 5 µm WAT054225 WAT054250
Symmetry® Sentry™ Guard Columns (2 per pack)(Requires Sentry™ Guard Holders, see Page 186)
Particle Part No. Part No.Dimension Size C18 C80.32 x 50 mm 3.5 µm 186000945 1860009460.32 x 100 mm 3.5 µm 186000951 1860009520.32 x 150 mm 3.5 µm 186000957 186000958
0.32 x 50 mm 5 µm 186000929 1860009300.32 x 100 mm 5 µm 186000935 1860009360.32 x 150 mm 5 µm 186000347 186000941
Symmetry® Capillary Columns
Symmetry® Columnsand Cartridges
Dimension Particle Size Part Number
75 µm x 50 mm 3.5 µm 18600140775 µm x 100 mm 3.5 µm 18600140875 µm x 100 mm 5 µm 18600140975 µm x 150 mm 5 µm 186001410
Symmetry® C18 PicoFrit™ Columns
Dimension Particle Size Part Number
75 µm x 50 mm 3.5µm 18600140375 µm x 100 mm 3.5 µm 18600140475 µm x 100 mm 5 µm 18600140575 µm x 150 mm 5µm 186001406
Symmetry® C18 IntegraFrit™ Columns
Particle Part No. Part No.Dimension Type Size C18 C84.6 x 150 mm Column 3.5 µm WAT094240 WAT0942372.1 x 150 mm Column 5 µm WAT094234 WAT0942313.0 x 150 mm Column 5 µm WAT054446 WAT0544343.9 x 150 mm Column 5 µm WAT047210 WAT0469554.6 x 150 mm Column 5 µm WAT054448 WAT0544354.6 x 250 mm Column 5 µm WAT054450 WAT054438
3.9 x 150 mm Cartridge* 5 µm WAT054452 WAT0544404.6 x 150 mm Cartridge* 5 µm WAT054454 WAT0544424.6 x 250 mm Cartridge* 5 µm WAT054456 WAT054444
* Require Endfittings, see Page 13
Symmetry® Column and Cartridge ColumnMethod Validation Kits Three Columns from Three Different Batches to Test Reproducibility.
Particle Part No. Part No.Dimension Size C18 C87.8 x 10 mm 5 µm 186000711 1860007127.8 x 50 mm 5 µm 186000208 1860002147.8 x 100 mm 5 µm 186000209 18600021519 x 10 mm 5 µm 186000715 18600071619 x 50 mm 5 µm 186000210 18600021619 x 100 mm 5 µm 186000211 18600022930 x 50 mm 5 µm 186000235 18600023730 x 100 mm 5 µm 186000236 186000238
7.8 x 10 mm 7 µm 186000713 1860007147.8 x 150 mm 7 µm WAT066288 WAT0662857.8 x 300 mm 7 µm WAT066235 WAT06622519 x 10 mm 7 µm 186000717 18600071819 x 150 mm 7 µm WAT066240 WAT06622819 x 300 mm 7 µm WAT066245 WAT066230
Purification and Isolation Cartridge Column Holders
7.8 x 10 mm 5, 7 µm — 18600070819 x 10 mm 5, 7 µm — 186000709
Replacement o-ring 7.8 mm, 2 pkg 700001019Replacement o-ring 19 mm, 2 pkg 700001020
SymmetryPrep™ Columns for Purification and Isolation
184 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Particle Part No. Part No.Dimension Size RP18 RP80.32 x 50 mm 3.5 µm 186000947 1860009480.32 x 100 mm 3.5 µm 186000953 1860009540.32 x 150 mm 3.5 µm 186000959 186000960
0.32 x 50 mm 5 µm 186000931 1860009320.32 x 100 mm 5 µm 186000937 1860009380.32 x 150 mm 5 µm 186000942 186000943
SymmetryShield™ Capillary Columns
SymmetryShield™
Columns and Cartridges
Symmetry300™
Columns and Cartridges
Particle Part No. Part No.Dimension Size RP18 RP81.0 x 50 mm 3.5 µm 186000175 WAT1060601.0 x 150 mm 3.5 µm 186000176 WAT1060482.1 x 30 mm 3.5 µm 186000171 WAT1060012.1 x 50 mm 3.5 µm 186000172 WAT0942572.1 x 100 mm 3.5 µm 186000173 WAT0589692.1 x 150 mm 3.5 µm 186000174 WAT1060083.0 x 100 mm 3.5 µm 186000700 1860007033.0 x 150 mm 3.5 µm 186000699 1860007024.6 x 50 mm 3.5 µm 186000177 WAT0942604.6 x 75 mm 3.5 µm 186000178 WAT0942634.6 x 100 mm 3.5 µm 186000179 WAT0942664.6 x 150 mm 3.5 µm 186000180 WAT094269
2.1 x 50 mm 5 µm 186000217 1860002232.1 x 150 mm 5 µm 186000111 WAT0942453.0 x 150 mm 5 µm 186000692 WAT0942433.0 x 250 mm 5 µm 186000693 1860006943.9 x 150 mm 5 µm 186000108 WAT2006554.6 x 50 mm 5 µm 186000218 1860002244.6 x 150 mm 5 µm 186000109 WAT2006624.6 x 250 mm 5 µm 186000112 WAT200670
SymmetryShield™ Columns
Particle Part No. Part No.Dimension Size RP18 RP82.1 x 50 mm 3.5 µm 186000168 1860001472.1 x 100 mm 3.5 µm 186000167 1860001462.1 x 150 mm 3.5 µm 186000166 1860001454.6 x 75 mm 3.5 µm 186000183 WAT0942724.6 x 100 mm 3.5 µm 186000170 WAT094275
3.9 x 50 mm 5 µm — WAT0942483.9 x 150 mm 5 µm 186000106 WAT2006584.6 x 150 mm 5 µm 186000110 WAT2006654.6 x 250 mm 5 µm 186000113 WAT200661
SymmetryShield™ Cartridge Columns (Requires endfittings, see Page 186)
Particle Part No. Part No.Dimension Size RP18 RP82.1 x 10 mm 3.5 µm 186000169 WAT1061293.9 x 20 mm 3.5 µm 186000701 1860007043.9 x 20 mm 5 µm 186000107 WAT200675
SymmetryShield™ Sentry™ Guard Columns (2 per pack)(Requires Sentry™ Guard Holders, see Page 186)
Particle Part No. Part No.Dimension Size RP18 RP819 x 10 mm 5 µm 186001835 18600184119 x 50 mm 5 µm 186001836 18600184219 x 100 mm 5 µm 186001837 18600184319 x 150 mm 5 µm 186001838 186001844
19 x 150 mm 7 µm 186001839 18600184519 x 300 mm 7 µm 186001840 186001846
Purification and Isolation Cartridge Column Holders
7.8 x 10 mm 5, 7 µm — 18600070819 x 10 mm 5, 7 µm — 186000709
Replacement o-ring 7.8 mm, 2 pkg 700001019Replacement o-ring 19 mm, 2 pkg 700001020
SymmetryShield™ Columns for Purification and Isolation
Particle Part No. Part No.Dimension Size C18 C41.0 x 150 mm 3.5 µm 186000185 1860002762.1 x 50 mm 3.5 µm 186000187 1860002772.1 x 100 mm 3.5 µm 186000188 1860002782.1 x 150 mm 3.5 µm 186000200 1860002794.6 x 50 mm 3.5 µm 186000201 1860002804.6 x 75 mm 3.5 µm 186000189 1860002814.6 x 100 mm 3.5 µm 186000190 1860002824.6 x 150 mm 3.5 µm 186000197 186000283
2.1 x 150 mm 5 µm WAT106172 1860002853.9 x 150 mm 5 µm WAT106154 1860002864.6 x 50 mm 5 µm WAT106209 1860002874.6 x 150 mm 5 µm WAT106157 1860002884.6 x 250 mm 5 µm WAT106151 186000289
Symmetry300™ Analytical Columns
Particle Part No. Part No.Dimension Type Size RP18 RP82.1 x 150 mm Column 3.5 µm 186000182 —4.6 x 150 mm Column 3.5 µm 186000181 WAT0942782.1 x 150 mm Column 5 µm 186000100 WAT0942543.0 x 150 mm Column 5 µm - WAT0942513.9 x 150 mm Column 5 µm 186000104 WAT2105944.6 x 150 mm Column 5 µm 186000103 WAT2105884.6 x 250 mm Column 5 µm 186000102 WAT210591
3.9 x 150 mm Cartridge* 5 µm 186000105 WAT2105824.6 x 150 mm Cartridge* 5 µm 186000101 WAT2105854.6 x 250 mm Cartridge* 5 µm 186000114 WAT210579
* Require endfittings, see Page 186
SymmetryShield™ Column and Cartridge ColumnMethod Validation Kits Three Columns from Three Different Batches to Test Reproducibility.
185Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Particle Part No.Dimension Size C1819 x 10 mm 5 µm 18600184719 x 50 mm 5 µm 18600184819 x 100 mm 5 µm 18600184919 x 150 mm 5 µm 186001850
Purification and Isolation Cartridge Column Holders
7.8 x 10 mm 18600070819 x 10 mm 186000709
Replacement o-ring 7.8 mm, 2 pkg 700001019Replacement o-ring 19 mm, 2 pkg 700001020
Symmetry300™ Columns for Purification and Isolation
Particle Part No.Dimension Size C182.1 x 50 mm 3.5 µm 1860001992.1 x 100 mm 3.5 µm 1860001912.1 x 150 mm 3.5 µm 1860001964.6 x 75 mm 3.5 µm 1860001924.6 x 100 mm 3.5 µm 186000193
3.9 x 150 mm 5 µm WAT1061694.6 x 150 mm 5 µm WAT1061634.6 x 250 mm 5 µm WAT106160
Symmetry300™ Cartridge Columns (Requires endfittings, see Page 186)
Particle Part No. Part No.Dimension Size C18 C42.1 x 10 mm 3.5 µm 186000198 1860002753.9 x 20 mm 5 µm WAT106166 186000284
Symmetry300™ Sentry™ Guard Columns (2 per pack)(Requires Sentry™ Guard Holders, see Page 186)
Particle Part No. Part No.Dimension Size C18 C40.32 x 50 mm 3.5 µm 186000949 1860009500.32 x 100 mm 3.5 µm 186000955 1860009560.32 x 150 mm 3.5 µm 186000961 186000962
0.32 x 50 mm 5 µm 186000933 1860009340.32 x 100 mm 5 µm 186000939 1860009400.32 x 150 mm 5 µm 186000348 186000944
Symmetry300™ Capillary Columns
Particle Part No. Part No.Dimension Type Size C18 C42.1 x 150 mm Column 3.5 µm 186000194 1860002904.6 x 150 mm Column 3.5 µm 186000195 186000291
2.1 x 150 mm Column 5 µm WAT106193 1860002923.9 x 150 mm Column 5 µm WAT106187 1860002934.6 x 150 mm Column 5 µm WAT106190 1860002944.6 x 250 mm Column 5 µm WAT106184 186000295
3.9 x 150 mm Cartridge* 5 µm WAT106181 —4.6 x 150 mm Cartridge* 5 µm WAT106175 —4.6 x 250 mm Cartridge* 5 µm WAT106178 —
* Require endfittings, see Page 186
Symmetry300™ Column and Cartridge ColumnMethod Validation Kits Three Columns from Three Different Batches to Test Reproducibility.
Particle Symmetry® Symmetry®
Dimension Size C18 C82.1 x 20 mm 3.5 µm 186002066 1860020672.1 x 20 mm 3.5 µm 186002070 1860020713.0 x 20 mm 3.5 µm 186002074 186002075
3.0 x 20 mm 5 µm 186002078 1860020793.9 x 20 mm 5 µm 186002082 1860020833.9 x 20 mm 5 µm 186002086 1860020874.6 x 20 mm 5 µm 186002090 1860020914.6 x 20 mm 5 µm 186002094 186002095
Symmetry IS™ Columns
Particle Symmetry® Symmetry®
Dimension Size RP18 RP82.1 x 20 mm 3.5 µm 186002068 1860020692.1 x 20 mm 3.5 µm 186002072 1860020733.0 x 20 mm 3.5 µm 186002076 186002077
3.0 x 20 mm 5 µm 186002080 1860020813.9 x 20 mm 5 µm 186002084 1860020853.9 x 20 mm 5 µm 186002088 1860020894.6 x 20 mm 5 µm 186002092 1860020934.6 x 20 mm 5 µm 186002096 186002097
SymmetryShield IS™ Columns
Particle Symmetry300™
Dimension Size C180.35 x 5 mm 5 µm 186001492
Opti-Pak™ Symmetry300™ C18 trap columns (5 per pack)
186 Waters, Symmetry, SymmetryShield, Sentry, SymmetryPrep, Platform LC, Quattro Ultima, IS, Oasis and Alliance are trademarks of Waters Corporation. ©2003 Waters Corporation.
Cartridge and Guard Columns
Integrated Sentry™ Guard Holder to gowith cartridge end-fittings (see above)
3.0 x 20 mm, 3.9 x 20 mm – use Part Number WAT046905
Universal Sentry™ Guard Holder (Use on any column)
2.1 x 10 mm use Part Number WAT0979582.1 x 20 mm use Part Number 1860002623.0 x 20 mm use Part Number WAT0469103.9 x 20 mm use Part Number WAT0469104.6 x 20 mm use Part Number WAT046910
Waters Cartridge Column
Waters cartridge system requires re-useable endfittings. These cartridge endfit-tings need to be ordered when purchasing a Waters HPLC cartridge columnfor the first time. These cartridge columns should be connected to your HPLCinstrument using Waters standard connectors.
Disposable Cartridge
Reusable End Connectors
Waters Cartridge Column
ConnectorEndfitting
WAT037525
C-Clip
GrooveRetaining
Nut
Cartridge
Waters Sentry™ Guard Column andUniversal Guard Holder
Waters offers a line of Sentry™ Guard columns that contain the same high performance packing materials as all of the Waters analytical columns.Developed for customers needing increased capacity for biological fluid clean-up and/or temperature control, the Sentry™ Guard column is available in twoholder designs--one to be used as an integrated part of the Waters steel cartridge column with reusable endfitting, the other for use with any HPLC column. Both are installed without tools.
Sentry™ Guard Column
Integrated Guard Holder
Universal Guard Holder
Cartridge End-Fittings
2.1 x 50 mm, 2.1 x 100 mm, 2.1 x 150 mm, 2.1 x 250 mm use Part Number 7000001173.0 x 50 mm, 3.0 x 100 mm, 3.0 x 150 mm, 3.0 x 250 mm use Part Number WAT0375253.9 x 50 mm, 3.9 x 100 mm, 3.9 x 150 mm, 3.9 x 250 mm use Part Number WAT0375254.6 x 50 mm, 4.6 x 100 mm, 4.6 x 150 mm, 4.6 x 250 mm use Part Number WAT037525
Sentry™ Guard Column
©2003 Waters Corporation, 6/03 720000593EN
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