303580 Determination of acrylamide in food SPE clean-up procedure for LC-MS-MS

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Determination of acrylamide in food SPE clean-up procedure for LC-MS-MS

Summary

Type: SPE
Application #: 303580
Catalog#: 730533 - Chromab. columns ABC18, 6 ml, 500 mg
Detection: LC-MS-MS
Conditions: 1) Purpose and field of application
This method describes a clean-up procedure for the preparation of aqueous extracts for the determination of acrylamide by LC-MS-MS.
The method is suited for ultra heated starch-containing food, such as potato chips and other snacks, french fries, crispbread, cereals, etc.
The method is specially recommended for food with an acrylamide content > 100 μg/kg.

2) Summary
Food samples are superficially defatted with hexane/t-butyl methyl ether. Acrylamide is extracted from the food at 40 °C using water. For correction of recovery losses and quenching effects during the subsequent analysis, deuterated acrylamide is added as an internal standard. The crude extract is clarified by Carrez precipitation and further purified by filtrating solid phase extraction on a cartridge with a mixed phase of an ion exchanger and RP material.

3) Chemicals
Unless otherwise stated, chemicals of analytical purity must be used. Water must be either double-distilled or of comparable purity. The term "solution" is used for an aqueous solution.

Note:
Handling of the hazardous chemicals used in this method requires suitable care and protective means, such as avoiding skin contact and use of a fume hood. Local regulations concerning work with hazardous chemicals must be observed.

3.1) Isohexane / t-butyl methyl ether (95+5/vv)

3.2) Carrez solution I: 150 g K4[Fe(CN)6] x 3 H2O are dissolved in double-distilled water and filled up to 1 l with more bidist. water.

3.3) Carrez solution II: 300 g ZnSO4 x 7 H2O are dissolved in bidist. water and filled up to 1 l with more bidist. water.

3.4) Solid phase extraction cartridges with ion exchange and RP functions
CHROMABOND® ABC18, 6 ml, 500 mg, Cat. No. 730533
supplied e.g. by MACHEREY-NAGEL GmbH & Co. KG, Düren, Germany

3.5) Internal standard: triply deuterated acrylamide (CD2=CDCONH2)
concentration about 2 – 5 μg/ml in methanol or acetonitrile
Acrylamide and the isotopically labelled form, is carcinogenic, mutagenic and neurotoxic! Special precaution is required when weighing powdery standards (use rubber gloves and a respirator).

4) Equipment
4.1) Heatable ultrasonic bath
4.2) Vacuum manifold and vacuum pump for solid phase extraction
4.3) Centrifuge with centrifuge tubes for about 100 ml volume
4.4) Standard glassware, like e.g. Erlenmeyer flasks, glass funnels, pipettes
4.5) Folded filters, e.g. MACHEREY-NAGEL 615 1/4 , 15 cm diameter

5) Sampling
Prior to the analysis, samples should be stored, so that changes in composition are avoided. For dry products exclusion of atmospheric humidity is especially important.

6) Procedure
6.1) Sample pretreatment
For the analysis a representative amount of sample is homogenised, e.g. by grinding or crushing in a suitable mill.
The sample size for homogenisation should be at least 50 g. For smaller amounts of sample the whole sample is homogenised.
For verification of the analysis it is recommended, that for each series of samples a comparative sample is included, the content of which is known from at least a 5-fold determination.

6.2) Work-up procedure
About 5 g of the homogenised sample (for very high contents of acrylamide correspondingly smaller samples) are weighed in a folded filter and superficially defatted by slowly pouring 50 ml isohexane / butyl methyl ether (3.1) through the filter.
Alternative: pour about 50 ml isohexane / butyl methyl ether over 5 g sample into an Erlenmeyer flask, let stand for 10 minutes, shaking slightly now and then. Filter through a folded filter, rinse flask and filter with about 20 ml isohexane / butyl methyl ether.
The filter is dried in air, the contents are transferred into a 250 ml Erlenmeyer flask, the filter is cut in small pieces and also added to the flask.
The sample is spiked with about 2 – 5 μg of the internal standard (maximum 2 ml)
An aliquot of the used internal standard is filled into an autosampler vial and sent to the LC-MS lab together with the corresponding measuring series.
After addition of 100 ml of double-distilled water the flask is ultrasonically treated at 40 °C for 10 minutes (shake flask now and then).
The contents of the flask are mixed with 0.5 – 1 ml Carrez solution I and then with the same volume of Carrez solution II (mix well every time).
The aqueous phase is filtered through a paper filter, about the first 10 ml are discarded, the next 10 – 20 ml are collected separately for further investigation.
If necessary, a centrifugation step must precede the filtration.
A CHROMABOND® ABC18 cartridge is conditioned with about 10 ml methanol, followed by about 10 ml double-distilled water, then air is aspirated through the column for about 30 seconds. 2 ml of the sample solution are poured on to the cartridge and drop by drop aspirated through the packing. The eluate is discarded.
Then another 5 ml(*) of the sample solution are aspirated through the cartridge drop by drop, this eluate is collected.
1 ml of the eluate is transferred into an autosampler vial and used for LC-MS-MS, the rest is frozen till the end of the analysis.

(*) The detection limit can be decreased to 40μg/kg, using 25 ml instead of 5 ml in step 6.2. Even high contaminated samples show no break through of the matrix. After extraction with 10 ml ethyl acetate the extract is evaporated to a small volume and refilled to 1 ml. This sample can be analysed directly with GC/MS.
Authors: Rosen J Hellenäs K E Gutsche B Weißhaar R Buhlert J
Source: The Analyst 127 871-879 (2002)Deutsche Lebensmitte


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