food for thoughts and plastic for dinner: the latest ......©2015 waters corporation 1 food for...
TRANSCRIPT
©2015 Waters Corporation 1
Food for Thoughts and Plastic for Dinner: the
Latest Analytical Landmarks in the Field of
Contact Material Contamination
©2015 Waters Corporation 2
Overview
Extractables & Leachables Analysis
– Definition – Workflows – Directives – Simulants
– Incidents
Example #1: Analysis of Primary Aromatic Amines (PAA)
using the ACQUITY H-Class - Xevo TQ-S micro
Example #2: Analysis of N-nitrosamines in Saliva
using the ACQUITY UPLC I-Class - Xevo TQ-S micro
Example #3: Analysis of Polymer Additives in Simulated Migration Solvents
using the ACQUITY UPLC – SQD2
Example #4: Analysis of Polymer Additives in Polymer matrix using the ACQUITY UPLC – QDa
Example #5: Analysis of Phthalates in Beverages using the ACQUITY H-Class - Xevo TQD
Example #6: Analysis of Bisphenols A, B & E in Baby Food and Infant Formula
using the ACQUITY UPLC – Xevo TQD
©2015 Waters Corporation 3
Definitions
Leachables are chemical entities, both organic and inorganic, that migrate from
components of a container closure system or device into a drug/food product over the
course of its shelf-life. In other words, leachables are chemical species that make their way
into the product under normal application conditions.
Extractables are chemical entities, both organic and inorganic, that will migrate from
packaging or container materials into the contents when exposed to certain solvents under
exaggerated temperature and time conditions. They are used to identify and quantify
potential leachables.
Typical extractables include monomers and oligomers from incomplete polymerization
reactions, plasticizers, stabilizers, fillers, coloring agents, antioxidants, and antistatic
agents, as well as their degradants. Additionally, residues from detergents and mold
release agents that can be present on the resin after the molding process.
©2015 Waters Corporation 4
Food/Pharmaceutical manufacturing equipment
– Belts, gaskets, lubricants, etc.
Food/Pharmaceutical packaging
– Paper, plastic, carton board, glass, etc.
Dining wares
– Cutlery, bowl, plate, etc.
Toys
Balloons
Cosmetics
Contact Materials/ Substances
©2015 Waters Corporation 5
Workflows
Identification of extractables from packaging
material
– Sample extraction
– Targeted screening for known or expected
components and their degradants (IAS)
using HR-MS
– Unknown screening of unidentified
components (NIAS) using HR-MS
Targeted analysis of extractables from
packaging material
– Migration experiments using simulated
migration solvents or beads (to mimic a
worst case scenario)
– Targeted analysis using TQ-MS
Compound name: Aniline
Correlation coefficient: r = 0.999930, r^2 = 0.999859
Calibration curve: 11380.5 * x + -898.041
Response type: External Std, Area
Curve type: Linear, Origin: Exclude, Weighting: 1/x, Axis trans: None
ppb-0 5 10 15 20 25 30 35 40 45 50 55 60 65 70 75 80 85 90 95 100
Re
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200000
400000
600000
800000
ppb
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-6.0
-4.0
-2.0
0.0
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4.0
©2015 Waters Corporation 6
NIAS
Non-intentionally added substances (NIAS) are chemical compounds that are present in a
material but have not been added for a technical reason during the production process.
Their presence in food/pharma contact materials is generally not known by the consumer
and often is a challenge for the producer.
NIAS originate from break-down products of food/pharma contact materials, impurities of
starting materials, unwanted side-products and various contaminants from recycling
processes.
©2015 Waters Corporation 7
Quantitative Analyses
Photo-initiators in ink
– In-situ, after migration
– 2005: Ink curing agent Isopropylthioxanthone (ITX) detected in cardboard packaged milk
Primary Aromatic Amines in textile, printed ink, kitchenware
– In-situ, after migration
– 2007: Poorly manufactured polyamide utensils leach carcinogenic amines
Polymer additives
– In-situ, after migration from packaging
N-nitrosamines
– After migration from paint, cosmetics, balloons
Bisphenol A
– Recent concern involving infant feeding bottles
2-ethylhexyl-phthalate
– 2011: Clouding agent in probiotics substituted by probable carcinogenic compound in order to cut costs
Epoxydised soy bean oil (ESBO)
– 2005: Swiss survey of jarred foods identified concentrations exceeding the TDI allowance
©2015 Waters Corporation 8
Directive No 10/2011
Commission Regulation (Eu) No 10/2011 of 14 January 2011 on Plastic Materials
and Articles Intended to Come into Contact with Food
This Directive concerns packaging materials which are in direct contact with foodstuffs. This
packaging must be such that it does not transfer a hazardous part of its constituents
into/onto the food thus avoiding contamination, which could lead to adverse effects on both
human health and/or the quality of the food.
This transfer, known as migration, indicates to what extent the material is inert and
therefore a safe product. A specific migration limit (SML) is applicable for each substance. It
is expressed in mg substance per kg food. It is indicated ND if the substance shall not
migrate in detectable quantities.
In many cases a SML of 0.05 mg/kg applies. For substances for which no specific migration
limit is provided, a generic specific migration limit of 60 mg/kg shall apply.
©2015 Waters Corporation 9
Food Simulants
Food simulants A, B and C are assigned for foods that have a hydrophilic character and are able to extract hydrophilic substances. Food simulant B shall be used for those foods which have a pH below 4.5. Food simulant C shall be used for alcoholic foods with an alcohol content of up to 20 % and those foods which contain a relevant amount of organic ingredients that render the food more lipophilic.
Food simulants D1 and D2 are assigned for foods that have a lipophilic character and are able to extract lipophilic substances. Food simulant D1 shall be used for alcoholic foods with an alcohol content of above 20 % and for oil in water emulsions. Food simulant D2 shall be used for foods which contain free fats at the surface.
Food simulant E (Tenax) is assigned for testing specific migration into dry foods.
©2015 Waters Corporation 10
Xevo TQ & SQ Portfolio
Xevo TQD
Xevo TQ-S
Xevo TQ-S micro
SQD2
ACQUITY QDa
©2015 Waters Corporation 11
Example #1: Analysis of Primary Aromatic Amines (PAA)
using the ACQUITY H-Class - Xevo TQ-S micro
©2015 Waters Corporation 12
Background
The term “primary aromatic amines” (PAA) denotes a group of chemical compounds whose
simplest version is Aniline.
PAA are substances that are used, for example, in the production of certain colorants, so-
called azo pigments, notably in the color range yellow - orange - red.
Whereas a large number of PAA are safe in this respect, some PAA are known human
carcinogens. On the basis of studies involving animal experiments, others are seen as
potentially carcinogenic for humans.
For kitchenware, paper napkins and baker’s bags with colorful print and other printed food
contact items, some PAA may pose a health risk, if they are transferred to food.
©2015 Waters Corporation 13
Guidelines - Simulant
As regards polyamide kitchenware, the specific migration limit (SML) for PAAs is set as non-
detectable except for those on the positive list of the relevant legislation. They should not
release into foods or food simulants PAAs in a detectable quantity.
In accordance with Regulation (EU) No. 10/2011 on plastic materials and articles intended
to come into contact with food, the transfer of PAA that have not been assessed specifically,
must not be detectable in the sum.
Therefore a group SML of 0.01 mg/kg has been assigned.
As of 01/01/2013 the rules under Regulation (EU) No 10/2011 will apply. These provisions
include that for primary aromatic amine migration from polyamide kitchenware only one
migration test will be carried out.
The test is conducted with simulant B: 3% (w/v) acetic acid, as it has been demonstrated
that this simulant represents the worst case for the migration of PAAs from polyamide
kitchenware.
©2015 Waters Corporation 14
Compounds - Samples
A mix of 24 PAA in methanol was provided at a concentration of 100 µg/mL. Structures for a
small selection of PAA is given below.
23 out of 24 compounds could be detected at the desired levels. 3-chloro-o-toluidine was not
detected. No information is available in literature suggesting sensitivity is a common issue for
this compound.
9 extracts of kitchenware in 3% acetic acid were provided for analysis
Compound Mass Structure
Aniline 93
o-Toluidine 107
2,4-Diaminotoluene
122
©2015 Waters Corporation 15
Additional Sample Preparation
Some of the compounds are small, basic compounds, which are ionized with low pH. As a
result of their basic properties and the acidic sample solvent, some PAA don’t focus well on
the head of the column, resulting in poor peak shape and/or loss of retention.
Using Ammonium Hydroxide added to the 3% Acetic Acid samples prior to injection, the pH
of the sample is increased and the polar and weakly basic PAA such as Aniline will be in its
neutral form.
We believe this approach is preferred over the use of ion-pair reagent, currently described
in literature.
Sample X Sample X neutralized with NH4OH
©2015 Waters Corporation 16
A generic gradient was applied to separate the 23 PAAs
Adding Formic Acid to the mobile phase is not advised as it leads to loss
of retention of basic compounds on a reversed phase column. However,
using post-column addition of Formic Acid, a significant increase in
sensitivity could be obtained. The Xevo TQ-S micro integrated
fluidics simplify and fully automate this post-column addition.
UPLC Method
ACQUITY UPLC HSS T3, 1.8 µm, 2.1 x 100 mm
Mobile Phase A Water
Mobile Phase B Methanol
Column Temp 45ºC
Sample Temp 10ºC
Flow Rate 0.4 ml/min
Run Time 10 min
Injection volume 20 µL
Gradient
0 min 5% B
10 min 100% B
12 min 100% B
12.01 min 5% B
15 min 5% B
©2015 Waters Corporation 17
Typical Chromatogram
Overview of the 23 primary aromatic amines which could be quantified
The concentration of the compounds depicted equals 6.2 ppb (20 µL injection)
©2015 Waters Corporation 18
Typical Calibration Curve
Calibration curve from 0.8 to 100 ppb
Example of Aniline
Compound name: Aniline
Correlation coefficient: r = 0.999930, r^2 = 0.999859
Calibration curve: 11380.5 * x + -898.041
Response type: External Std, Area
Curve type: Linear, Origin: Exclude, Weighting: 1/x, Axis trans: None
ppb-0 5 10 15 20 25 30 35 40 45 50 55 60 65 70 75 80 85 90 95 100
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400000
600000
800000
ppb
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-2.0
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4.0
©2015 Waters Corporation 19
LOQ
Signal to Noise was calculated on 0.8 ppb standard (20 µL injection)
Compound S/N ratio
LOQ (ppb)
Aniline 377.4 0.021
o-Toluidine 768.1 0.010
2,4-Diaminotoluene 52.4 0.149
o-Anisidine 89.2 0.087
4-Chloroaniline 322.5 0.024
3-Chloro-o-toluidine ND ND
2,4,5-Trimethyl aniline 692.7 0.011
2-Methoxy-5-methylaniline
1444.3 0.005
4-Chloro-2-methylaniline 3503.1 0.002
2-Amino naphthalene 1858.2 0.004
2-Methyl-5-nitroaniline 26.6 0.293
4-Aminobiphenyl 225.5 0.035
2-Aminobiphenyl 271.6 0.029
Benzidine 559.3 0.014
4-Phenyl azoaniline 1930.7 0.004
©2015 Waters Corporation 20
LOQ
23 out of 24 compounds could be quantified well below the requested levels, with
quantitation limits ranging from 3 ng/L to 300 ng/L, taking into account an injection
volume of 20 µL. 19 out of the 23 compounds have detection limits below 30 ng/L. 3-
chloro-o-toluidine could not be detected, and this is confirmed by literature references.
These sensitivity levels meet the requirements for this assay, i.e. individual quantification
limits of 3 ng/mL per PAA and a total LOQ of 10 ng/mL for total PAA content.
Compound S/N ratio LOQ (ppb)
4,4’-Diamino diphenylmethane
1353 0.006
4,4’-Oxydianiline 311.6 0.025
3,3’-Dimethyl benzidine 164.5 0.047
4,4’-Thiodianiline 2582.3 0.003
o-Amino azotoluene 1745.5 0.004
3,3’-Dimethyl-4,4’-diaminodiphenylmethane
1818.2 0.004
3,3’-Dimethoxy benzidine 528.1 0.015
3,3’-Dichloro benzidine 925.9 0.008
4,4’-Methylene bis (2-chloroaniline)
1522.3 0.005
©2015 Waters Corporation 21
Repeatability
Repeatability (n=6) was assessed at different concentration levels: 0.4 – 0.8 – 3.1 – 12.5 –
100 ppb
The assay was found to be repeatable with RSD values (n = 6) generally lower than 5%.
©2015 Waters Corporation 22
Conclusion
23 out of 24 compounds could be detected at the desired levels. 3-chloro-o-toluidine was not
detected.
Using:
– Ammonium Hydroxide to neutralize the acidic samples
– post-column addition of Formic Acid to increase the signal intensity for some critical cpds
a sensitive assay was developed that can reach low ng/L concentrations
The assay was found to be repeatable with %CV values in general lower than 5%.
The samples were all below detection limits except for Aniline which was detected at 0.3 – 0.5
ng/mL.
©2015 Waters Corporation 23
Example #2: Analysis of Nitrosamines in Saliva
using the ACQUITY UPLC I-Class - Xevo TQ-S micro
©2015 Waters Corporation 24
Background
N-Nitroso compounds (NOC) are amongst the most potent carcinogens.
N-nitrosamines are used in the manufacturing of cosmetics, pesticides, and in most rubber
products. N-nitrosamines occur in latex products such as balloons, and in many foods and
other consumables.
Primary, secondary and tertiary amines can all be nitrosated to generate nitrosamines. The
secondary amines in general are the most reactive compounds towards nitrosating agents,
generating nitrosamines.
Other sources of N-nitrosamines are
– Elastomers (e.g. rubbers, silicones and thermoplastic elastomers)
– N-nitrosamines may be formed in finger paints under certain acidic conditions if they
include N-nitrosatable substances
©2015 Waters Corporation 25
Guidelines
Elastomers are extracted with a saliva solution containing nitrite, as ingestion will be
primarily via the mouth.
Finger paints are extracted with
– Water, simulating skin absorption
– Saliva solution containing nitrite, simulating ingestion
SML are in place for individual and the sum of all N-nitrosamines
©2015 Waters Corporation 26
Sample Analysis
5 nitrosamines and 1 internal standard were used for setting up a quantitative assay.
7 saliva samples were provided by the customer
ESI /EScI / APcI were compared
Linearity, sensitivity and repeatability were investigated
©2015 Waters Corporation 27
UPLC Method
ACQUITY UPLC I-Class system (Sample manager FTN)
Mobile phases
– Solvent A: water + 2 mM CH3COONH4
– Solvent B: methanol
– Sample manager wash solvent : Solvent B
Flow rate: 0.4 mL/min
Column temperature: 55°C
Injection volume: 30 µL
Column: ACQUITY UPLC CSH C18, 2.1 x 100 mm, 1.7 µm
Analysis time: 6 min
Gradient: from 5%B to 100%B in 4 minutes, after an isocratic hold of 30s.
©2015 Waters Corporation 28
Typical Chromatogram
Chromatogram of a 10 ng/mL standard. Compound names are depicted in the top
right corner of each trace. MRM transitions have been scheduled to maximize dwell time (15 points across a peak)
©2015 Waters Corporation 29
Ionization Source
NDBzA and NDBA are better in ESI,
IonSabre2 APcI is better for the small compounds like NDMA, NDEA and NePhA
IonSabre2 APcI is better than the combined EScI mode for NDMA, NDPhA and NDEA.
Because NDMA and NDEA are the least sensitive compounds, IonSabre2 APcI is the
preferred ionization mode.
The IonSabre2 APcI probe can be mounted by simply screwing the probe on the source
housing. No tools are needed.
©2015 Waters Corporation 30
Linearity
Calibration curve (0.1 – 10 ng/mL) for NDBA
Excellent linear correlation with r2 value of 0.9997 and % residuals of maximum 7.5%
Compound name: NDBA
Correlation coefficient: r = 0.999863, r^2 = 0.999726
Calibration curve: 13869.8 * x + -238.344
Response type: External Std, Area
Curve type: Linear, Origin: Exclude, Weighting: 1/x, Axis trans: None
ng/mL-0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5 6.0 6.5 7.0 7.5 8.0 8.5 9.0 9.5 10.0
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25000
50000
75000
100000
ng/mL
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0.0
5.0
©2015 Waters Corporation 31
S/N at the 0.1 ng/mL level
The S/N for a 30 µL injection of the 5 nitrosamines
S/N based on pk-to-pk noise, no extra processing
Sensitivity
©2015 Waters Corporation 32
Repeatability
The repeatability (n = 6) of the quantitative analysis was determined at the level of 0.5 ng/mL
and 10 ng/mL. The % RSD’s are:
©2015 Waters Corporation 33
Conclusions
The nitrosamine assay on the Xevo TQ-S micro is sensitive with detection limits at and below the
desired 0.1 ng/mL level.
The assay is both linear and repeatable.
IonSabre2 APcI is the best ionization source for this multi-residue application.
©2015 Waters Corporation 34
Example #3: Analysis of Polymer Additives in Simulated Migration
Solvents using the ACQUITY UPLC – SQD2
©2015 Waters Corporation 35
Background
Plastics are made of monomers and other starting substances which are chemically reacted
to a macromolecular structure, the polymer, which forms the main structural component of
the plastics.
To the polymer, additives are added to achieve defined technological effects. The polymer
as such is an inert high molecular weight structure, and usually cannot be absorbed in the
body.
Potential health risk may occur from non- or incompletely reacted monomers or other
starting substances or from low molecular weight additives which are transferred into food
via migration from the plastic food contact material.
Therefore monomers, other starting substances and additives should be risk assessed and
authorized before their use in the manufacture of plastic materials and articles.
©2015 Waters Corporation 36
Sample Analysis: Objectives
Demonstrate the performance characteristics to quantify polymer additives
At levels of 10 ng/mL in 3% acetic acid and 10% alcohol
At levels of 6 µg/mL in olive oil
This report focuses on the results of the analysis of 10 polymer additives in terms of sensitivity,
linearity and repeatability.
The following Polymer Additives were analyzed: Irgafos 168, Irgafos 168 phosphate, Irganox
1010, Irganox 3114, Irganox 1076, Irganox 1330, Oleamide, Erucamide, PS800, PS802
©2015 Waters Corporation 37
Instrument Settings
UPLC Column: ACQUITY UPLC BEH Phenyl 1.7 µm, 2.1 mm x 100 mm
UPLC Parameters
MS settings:
– Auto dwell times (15 points across peak) and pos/neg switching in APCI mode
– All compounds in APCI positive ion mode except for Irganox 1330 (APCI-)
– RT windows according to elution time
– One m/z for quantitation and one m/z (in-source fragment) for confirmation (where applicable)
Mobile Phase A 0.1% formic acid in water
Mobile Phase B Methanol
Column Temp 40ºC
Sample Temp 15ºC
Flow Rate 0.4 mL/min
Run Time 8.5 min
Injection V 2 or 20 µL
Gradient 0 min 80% B
6 min 100% B (6)
7 min 100% B (6)
8.5 min 80% B (1)
[MH]+
N2 molecules
[Fragment]+
Collision Area
10-4
mbar
~1 mbar
©2015 Waters Corporation 38
Typical Chromatogram
100 ng/mL - Compound names are denoted on the right side
©2015 Waters Corporation 39
Acetic Acid & Alcohol
Excellent linearity for all compounds, in both 3% HOAc and 10% alcohol. The residual error
on the calibration plot below 10% for virtually all measured points.
The repeatability of the 6 QC’s of both 10 ng/mL and 50 g/mL standard levels was very good
on all measured concentrations (RSD < 10%).
The instrument sensitivity was enough to provide a S/N of min. 10 (required for LOQ) for a
standard at 5 ng/mL. This is half the required concentration.
IonSabre2 APcI is the best ionization source for this multi-residue application.
©2015 Waters Corporation 40
Typical Calibration Curve
Calibration curve and residuals for Irganox 1330
Excellent linearity (r2 = 0.999) and % residuals (all below 5.3%)
Compound name: 1330
Correlation coefficient: r = 0.999547, r̂ 2 = 0.999093
Calibration curve: 332.424 * x + 299.498
Response type: External Std, Area
Curve type: Linear, Origin: Exclude, Weighting: 1/x, Axis trans: None
ppb-0 5 10 15 20 25 30 35 40 45 50 55 60 65 70 75 80 85 90 95 100
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10000
20000
30000
ppb
Res
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4.00
©2015 Waters Corporation 41
Repeatability at 10 ng/mL
Most % RSD values below 5%
©2015 Waters Corporation 42
Olive Oil
Polymer Additives analysis in diluted olive oil is possible at the requested level of 6 µg/mL.
The confirmation trace is a good way to increase the selectivity for some compounds in this
complex matrix (see chromatograms for PS800).
min3.450 3.500 3.550 3.600 3.650 3.700 3.750 3.800 3.850 3.900 3.950 4.000 4.050 4.100 4.150 4.200 4.250
%
18
F2:SIR of 2 channels,AP+
329.2
20150327 008
IonSabre2 6 ug/g in olie > 60 ppb in vial
1.452e+0054.05
PS800
3.94
5023.55
3.46 3.723.53 3.593.55 3.63
4.17
min
%
1
F2:SIR of 2 channels,AP+
515.35
20150327 008
IonSabre2 6 ug/g in olie > 60 ppb in vial
3.033e+005
PS800
3.94
13865.31*
4.17
©2015 Waters Corporation 43
Olive Oil Spiked at 6 µg/mL
Some peak detoriation occurs (for 1076 eg)
©2015 Waters Corporation 44
Example #4: Analysis of Polymer Additives in
Polymer matrix using the ACQUITY UPLC - QDa
LC/MS System
©2015 Waters Corporation 45
Background
Customer has several HPLC as well as TLC methods for the detection of Polymer Additives
in extracts of the polymer itself.
The aim of this analysis was to find an alternative for the TLC analyses of the UV
transparent polymer additives:
– PS800
– PS802
– Oleamide
– Erucamide
– Stearic Acid
– Weston 618
– Hostanox SE 10
Required sensitivity levels: 10-100 ppm in AcN extract
– After polymer extraction & precipitation
©2015 Waters Corporation 46
Polymer Additives
Most common polymer additives are analyzed in APCI mode
– Low ppb levels can be obtained (SQD2 fit for purpose for migration purposes)
– See example #4
QDa only available with ESI probe
– Low ppm levels possible with ESI?
Experimental details
– 6 minute gradient from 80% B to 100%B
– Mobile phases:
o A = 0.02% formic acid in water
o B = methanol
– 1 µL injection volume
o Sample injected without any further sample preparation
©2015 Waters Corporation 47
Polymer Additives
A 10 ppm mixture of following polymer additives was analyzed (see next slide):
– Irganox PS 800
– Irganox PS 802
– Oleamide
– Erucamide
– Weston 618
– Hostanox SE10
– Stearic acid
©2015 Waters Corporation 48
Polymer Additives
Irganox PS 800
Irganox PS 802
Oleamide
Erucamide
Weston 618
Hostanox SE10
Stearic acid
10 ppm could clearly be detected for 6 out of 7 compounds. The observed intensities allow
for lower detection limits.
1 out of 7 compounds, Hostanox SE10, is not detected. Based on the structure ([H3C-
(CH2)17-S-]2 of the compound, it is likely that this compound is not MS sensitive at all.
©2015 Waters Corporation 49
Example #5: Analysis of Phthalates in Beverages
using the ACQUITY H-Class - Xevo TQD
©2015 Waters Corporation 50
Phthalates
Phthalates cover a large group of compounds, esters of phthalic acid Known toxic effects
− Considered endocrine disruptors, related to reproduction in animal studies Migration can occur during production and storage, from packaging materials,
coatings, equipment coatings, sealants, etc.
In 2013, China banned the importation of European spirits unaccompanied by CoA for phthalates
©2015 Waters Corporation 51
Challenges in phthalate analysis
Background ~7e4
DBP at 100 ppb
Traditionally, analysed by gas chromatography
– Derivatisation and/ or extraction required
– Non selective m/z 149 monitored for multiple compounds
Growing interest liquid chromatography method
– Reduced sample preparation: dilute and shoot
– Improved selectivity on m/z transitions
Ubiquitous contaminants
– Significant background impacts accurate
quantification by GC and/ or LC
©2015 Waters Corporation 52
Isolator column: Overview
Ubiquitous contaminants
AcQuity C18 isolator column
− Part number: 186004476
Isolator separates background
contaminants from analytes of
interest
Isolator column
Sample injector
Solvent mixer
©2015 Waters Corporation 53
Isolator column: Contaminant separation
BBP Background BBP contaminant
©2015 Waters Corporation 54
Example of spirit matrices spiked with BBP at 100 µg.l-1
Summary of Results
Sample A
Brandy
Sample B
Gin
Sample C
Whiskey
Compound name: BBPCorrelation coefficient: r = 0.997755, r^2 = 0.995515Calibration curve: 839.103 * x + 1264.79Response type: External Std, AreaCurve type: Linear, Origin: Exclude, Weighting: 1/x, Axis trans: None
ppb-0 10 20 30 40 50 60 70 80 90 100
Re
sp
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-0
20000
40000
60000
80000
BBP
Compound name: DNOPCorrelation coefficient: r = 0.998477, r^2 = 0.996957Calibration curve: 763.204 * x + 168.408Response type: External Std, AreaCurve type: Linear, Origin: Exclude, Weighting: 1/x, Axis trans: None
ppb-0 10 20 30 40 50 60 70 80 90 100
Re
sp
on
se
-0
20000
40000
60000
DNOP
Example of whiskey matrix matched calibration from 5 to 100 µg.l-1
©2015 Waters Corporation 55
Conclusions
Vast and varied area of analysis with international interest increasing
Extensive legislation in place, with some commonalities and solid base for safety and
analytical approach
Challenges faced by food industry for good communication with packaging manufacturers
Simple and quick “dilute and shoot” method for the accurate quantification of migration of
materials in contact with foodstuffs
Improves consumer confidence and ensures compliance for export with high throughput
©2015 Waters Corporation 56
Example #6: Analysis of Bisphenols A, B & E in Baby Food
and Infant Formula using the ACQUITY UPLC – Xevo TQD
©2015 Waters Corporation 57
BPA Background
What is BPA?
– BPA is the starting material for polycarbonate polymerization
– It has been an important industrial chemical not only to make polycarbonate plastic but
also epoxy resins; both of which are used in a wide variety of applications.
o Polycarbonate plastics are used in baby bottles, tableware and other food containers
• (Eyeglass lenses, medical equipment, digital media (e.g., CDs and DVDs), cell
phones, consumer electronics, computers and other electrical equipment, …)
o Epoxy resins are used in can coatings, industrial floorings, adhesives, industrial
protective coatings, powder coatings, and printed circuit boards
©2015 Waters Corporation 58
BPA Background
What are the potential health issues?
– Can leach into food from the epoxy resin lining of cans and from
consumer products such as polycarbonate tableware, food storage
containers, water bottles, and baby bottles.
– BPA is a weak "estrogen like" endocrine disruptor which can mimic
circulatory endogenous hormones and is suspected to be linked to a
higher incidence of certain cancers, reproductive disorders, diabetes
and heart disease in humans
©2015 Waters Corporation 59
International Regulations
Canada - First country to take action, listed it as a toxic substance
US-FDA - Released a report in 2010 expressing some concern regarding exposure of fetuses, infants and young children to BPA. Report encourages further studies into the safety of BPA
European Union - EU directive (2011/8/EU) has banned the use of BPA in infant feeding bottles
o France Bans Bisphenol A in All Food Contact Materials: French National Assembly has been voting to ban BPA from 2014 on. Concerning baby nutrition products the ban will step into force in 2013
Japan - Between 1998-2003, canning industry voluntarily replaced its BPA-containing epoxy resin can liners with BPA-free polyethylene terephthalate (PET) in many of its products
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Column: ACQUITY UPLC BEH C18 2.1 x 50 mm, 1.7 m
Column temp 40 C
Mobile phase A 0.5 % NH4OH in H2O
Mobile phase B 0.5 % NH4OH in MeOH
Elution 3 minute linear gradient from 5% (B) to 95% (B)
Flow Rate 0.5 mL/min
Injection volume
50 L
Experimental Conditions
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Part 1 – Protein precipitation
Part 2 - DisQUE
Part 3 – OASIS HLB
Optimised Sample Prep Protocol
Precipitation (10g sample, 10mL CH3CN) Centrifuge and collect supernatant
Add contents from DisQuE tube 1. Shake.
Centrifuge & collect 10 mL supernatant
Add contents from DisQuE tube 2. Shake.
Centrifuge & collect supernatant.
Dilute supernatant with 70 ml H2O. Load on Oasis HLB (3cc)
Wash – 2 mL 40 % MeOH Elute – 1 mL 100 % MeOH
Dilute eluate with 1 mL H2O Inject 50 µL for LC-MS/MS
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Spiked Compounds: 1 ng/mL in Samples
S/N > 3
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Correlation coef f icient: r = 0.998164, r^2 = 0.996332
Response type: Internal Std ( Ref 4 ), Area * ( IS Conc. / IS Area )
Curve type: Linear, Origin: Exclude, Weighting: 1/x^2,
Conc-0.0 2.5 5.0 7.5 10.0 12.5 15.0 17.5 20.0
Res
po
nse
-0.00
0.10
0.20
0.30
0.40
0.50
0.60
0.70
0.80
0.90
1.00
1.10 Bisphenol Infant Formula (10 pg/μL spike)
BPA 102% (3.2%)
BPB 95% (5.5%)
BPE 81% (4.6%)
Linearity and Recovery
Bisphenol Baby food
(20 pg/μL spike)
BPA 110% (7.8%)
BPB 112% (6.7%)
BPE 99% (6.1%)
©2015 Waters Corporation 64
Thank you for your attention
©2015 Waters Corporation 65
Additional Information about Legislation