final ms apps ppt
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GC MS
LC MS/MS
SELDI-TOF
CEMS
HPTLC MS
ICPMS
TGAMS2
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Niemann-Pick C1 Disease from Human Plasma
Adrenal Cortical Dysfunction by Liquid Chromatography-Tandem Mass
Spectrometry
Cancer using SELDI-TOF MS
Propionic Acedemia using GCMS
Diagnosing Diabetes
Diagnosis of vitamin B12 deficiency
Diagnosis of neuroblastomas etc.
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Phenyalanine to Tyrosine
PKU is a metabolic disorder caused by a deficiency of the liver
enzyme phenylalanine hydroxylase
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Fig. 1. A drop of blood is spotted onto a sample card, and an aliquot is extracted
for analysis
by MS or GC-MS. The peak profile is used to diagnose metabolic errors.
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Amino acids extraction
200 L methanol/HCl 0.1% in a screw cap vial, 1 h, at 4C or bysonication for 1 min.
100 L of the extract placed in a vial,
Addition of the internal standard and removing of the reagent inanitrogen stream
AA derivatization
Esterification: 500 Lbutanol: acetyl chloride, 4:1 (v/v), 30 min, at
100C. The excess reagent is removed with a stream of nitrogen7
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Analytical instrument :
Trace DSQ ThermoFinnigan quadruple mass spectrometer
Trace GC.
Column
capillary column Rtx-5ms
30 m 0.25 mm, 0.25 m
TC Prog. : : 50 C (1 min) to 310 C, at 20 C/min.
Carrier gas : Helium (99.9995%)
Flow rate : 1 mL/min
MS
Ion source : EI mode
SIM mode
Scan m/z 50 to 5008
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Fig. 2. The mass spectra of the phenylalanine and tyrosine as trifluoroacetic
butyl ester derivatives used for PKU diagnosis
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The fingerprint chromatograms in SIM-GC-MS for a PKU patient (our results) present
the differences between the first (top chromatogram) and the third month of treatment
(second chromatogram) . The retention times for the amino acids: 15N-Gly at 6.40
min, Val at 7.02 min, Leu at 7.50 min, Pro at 8.40 min, Phe at 9.70 min and Tyr at
11.28 min. 10
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PKU diagnosis could be tested by calculating the ratio Phe/Tyr using 20
L of blood spots.
A comparison of two chromatograms demonstrates the benefit of thetreatment in the case of a PKU child .
The first fingerprint chromatogram presents higher intensity of the
phenylalanine peak in the case of a patient presenting PKU, and after
three months of treatment the peak of Phe is significantly decreased.
The ratio of Phe/Tyr decreased from 2.30 at 1.02.
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What is Niemann-Pick Disease?
- Neurodegenerative disease
- metabolic dysfunction
Why LC-MS/MS?
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How is it Diagnosed?
Monitoring oxidation products of cholesterol in plasma
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Fig.2 Spectra for 7-ketocholesterol
dimethylglycinate (KC-DMG)
Fig.1 Spectra for cholestane-5-hydroxy-3,6-bis (dimethylglycinate)(CH-DMG2)
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Analysis:
- Sample preparation
Derivatization by N,N-Dimethyl glycine
-LC-MS/MS
APCI
MRM scan Mode
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What is SELDI-TOF MS ?
Difference between the mass spectrometric proteomic patterns ofserum
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Take 1-3 l serum sample
Spot sample on the activated protein chip array
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Also known as Kimmelstiel-Wilson syndrome
Renal disease
Caused by angiopathy of capillaries in the kidney glomeruli
Risk factors- elevated urinary albumin excretion, increased blood
pressure, and poor glycemic control
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To extract maximal information proteins present in smallquantities
CE-ESP-MS: Fast, sensitive, automated
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Detection of the current status of diabetic nephropathy of diabeticpatients.
Differentiation between patients with diabetes mellitus and
healthy controls in general.
Evaluation of CE-MS-defined patterns derived from urinarypolypeptides of patients with diabetic nephropathy differ fromthose of patients with other chronic renal diseases.
Making a statement of the progression of diabetic nephropathy.
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Validation of the treatment success of drugs, likecandesartan in context of diabetic nephropathy
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Analogue insulin.
Animal insulin :-bovine insulin and porcine insulin.
Human insulin
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Planar chromatography is well suited for peptide analysis.
Analysis of intact proteins.
Simple to medium complex mixtures.
Peptides from tryptic digest can be separated by HPTLC and
analyzed by MS.
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Human insulin
Bovine insulin
Porcine insulin
This were all diluted in 0.1% TFA (1mg/ml)
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HPTLC silica gel Si 60 F254 20 10 cm.
Automated TLC Sampler 4
Band length :- 5 mm
Track distance :-10 mm
Application volume :- 5 micro litre.
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Human insulin Porcine insulin Bovine insulin
968,7816[M + 6H]6+ 963,78[M+6H]6+ 956,4421 [M+6H]
1162,5367[M+5H] 1156,3347[M+5H] 1147,5288[M+H5H]
1452,6680[M+4H] 1445,1658[M+4H] 1434,1584[M+4H]
1936,5513[M+3H] 1926,5491[M+3H] 1912,2065[M+3H]
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Provide rapid, comprehensive, multi-componentanalyses
Maintain or improve sensitivity, selectivity, and
accuracy
Replacing many older traditional techniques
Is becoming an indispensable tool in clinical laboratory
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THANK YOU
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