Acrylamide ERP Resubmission Review

(d) 2,3,3-d 3 -acrylamide (d 3 -AA) working standard solution 4 µg/mL in water (IS) .—Add 2 200 mL of stock standard solution (E.c) into a 100-mL volumetric flask, and complete to the mark 201 with water. Store at - 20 °C for maximum 9 months. 202 (e) 13 C 3 -2,3,3-d 3 -acrylamide ( 13 C 3 -d 3 -AA) ca 20 µg/mL in water (IS) .— 13 C 3 -d 3 -AA is 203 supplied in small amount (ca. 1 mg). To record the weighed mass as precisely as possible, 204 weigh the original commercial bottle (m1). Transfer the content into a 50-mL volumetric flask. 205 Rinse the bottle with water and complete the flask to the mark with water. Weight again empty 206 dried commercial bottle (m2) to record weighed amount of 13 C 3- d 3 -AA by difference (m IS = 207 m1- m2). Store at - 20°C for maximum 9 months. 208 (f) 13 C 3 -2,3,3- d 3 -acrylamide ( 13 C 3 -d 3 -AA) working standard standard solution 2 µg/mL in 209 water (IS) .—Into a 100-mL volumetric flask, dilute an adequate volume of stock solution (E.e) 210 to obtain a concentration of 2 µg/mL (± 0.3 µg/mL) in water. Store at - 20 °C for maximum 9 211 months. 212 (g) Calibration solutions .—Into separate 50-mL volumetric flasks (n=7), transfer 0 / 0.15 213 / 0.5 / 1.5 / 2.5 / 5 and 10 mL of the AA 0.5 µg/mL working standard solution (E.b), 214 corresponding to 0 / 1.5 / 5 / 15 / 25 / 50 and 100 ng/mL concentration. Add in each flask 0.1 215 mL of the IS working standard solution. Complete to the mark with water. Calibration solutions 216 are stable at - 20°C for at least 6 months. 217 (h) Rinsing solution during SPE cleanu-up Methanol-water (60+40) .— Into a 200-mL 218 glass bottle, transfer 120 mL of methanol and 80 mL of water (measured separately in 200 and 219 100-mL volumetric cylinders) and mix. Store at ambient temperature for maximum one month. 220 (i) Enzymatic solution for cereals samples preparation.— Prepare the enzymatic solution 221 by weighing Fungamyl® 2500 SG, applying a 1/400 (w/v) dilution factor in water. As an 222 example, for 24 samples preparation, dilute 1.20 g ± 0.05 g of enzyme into 480 mL of water. 223 Homogenize using magnetic stirrer plate and let it rest for 10 min before using. This solution 224 can’t be stored, it must be freshly prepared. 225

F.

Laboratory test sample preparation

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(a) Laboratory sample.— A representative sample should be sent to the laboratory, ideally 227 with a minimum of 100 g. Sample must not be damaged or changed during transport and 228 storage. It should be transported in air-tightness container, protected from light, and stored at 229

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