Extraction of acrylamide from food for analysis following EN16618:2015

Figure 1. Structure of Acrylamide.

Introduction 

Acrylamide is a small polar highly water soluble molecule formed when carbohydrate rich foodstuffs are cooked at high temperatures.

Acrylamide was first discovered in foodstuffs in 2002 causing widespread alarm due to its classification as a probable carcinogen. By 2017, regulatory authorities established bench- mark levels and recommended that companies continue to monitor and minimise the acrylamide levels in heat processed foods. There has since been an increased demand for a reliable and accurate method for acrylamide detection.

The sample preparation methodology outlined in this application note reflects the European standard methodology documented in EN16618:2015 ‘Food analysis — Determination of acrylamide in food by liquid chromatography tandem mass spectrometry (LC-ESI-MS/MS)’. For additional information and analytical details, refer to the EN documentation.

The sample preparation process is based on a two-part solid phase extraction procedure. Part 1 utilizes ISOLUTE® Multimode SPE cartridges to trap unwanted interference compounds from the sample matrix. Part 2 utilizes ISOLUTE® ENV+ cartridges to retain and concentrate the acrylamide from aqueous solution.

Analyte

Acrylamide

Matrix

Baked products such as bread, toasted bread, crisp bread, butter cookies, and biscuits, as well as potato products such as potato chips, potato crisps, and potato pan cake and roasted coffee.

Sample preparation procedure

Sample pre-treatment

Baked products and potato products:

Weigh 2 g +/-0.1 g of test material and add 40 mL water, along with appropriate internal standard solution. Shake vigorously. Centrifuge and transfer a 10 mL aliquot into a clean test tube. Avoid transfer of any fat layer.

Coffee samples:

Weigh 2 g +/-0.1 g of test material and add 5 mL n-hexane or cyclohexane. Add 40 mL water, along with appropriate internal standard solution. Shake vigorously and centrifuge.
Remove and discard the organic (hexane) layer and transfer a 10 mL aliquot of the aqueous sample into a clean test tube.

Step 1: SPE clean-up

Format

ISOLUTE® Multimode 1 g/6 mL (p/n 904-0100-C)

Condition

Condition the cartridge with methanol (3 mL).

Equilibrate

Equilibrate the cartridge with water (2 x 6 mL).

Sample loading

Load 10 mL of the aqueous sample and collect the eluate.

Step 2: SPE concentration

Format

ISOLUTE® ENV+ 500 mg/6 mL (p/n 915-0050-C)

Condition

Condition the cartridge with methanol (5 mL).

Equilibrate

Equilibrate the cartridge with water (5 mL).

Sample loading

Load the extract (approximately 10 mL) from the previous step and discard the eluate.

Wash

Rinse the cartridge with water (4 mL).

Elute

Elute acrylamide with 60/40 methanol/water (v/v, 2 mL).

Post extraction

Evaporate the methanol from the extract at <4o oC. Do not reduce the volume to less than 500 µL. Transfer the extract to a suitable vial for LC-MS/MS analysis.

Analytical conditions

Please refer to EN16618:2015 for analytical LC-MS/MS conditions.

Additional information

If required, this methodology can be scaled down to utilize smaller SPE cartridges or plates.

Reduced bed masses bring the advantages of lower sample volumes, and subsequently reduced final extract volumes. This can lead to higher throughput due to reduced processing time and reduced post extraction evaporation.

The table below suggests appropriate volumes for use with reduced bed masses.

*Note these volumes are based on theoretical considerations and should be optimized by users.

Ordering information

Literature Number: AN966