AOAC SPIFAN Nutrients ERP (December 7, 2022)

Amino-07 (Method) 10-2022 FOR ERP USE ONLY DO NOT DISTRIBUTE

F.

Sample Preparation (a) Slurry preparation .—Make fresh prior to analysis (i) Accurately weigh ~5 g powder into a 50 mL disposable centrifuge tube. Record weight. (ii) Accurately weigh ~40 g water to powder. Record weight. (iii) Cap firmly and shake to mix. (iv) Shake each slurry immediately prior to weighing to ensure homogeneity. (v) Accurately weigh ~1.0 mL slurry into a 15 mL centrifuge tube. Record weight. (vi) Add 0.3 mL of SILSS to each sample centrifuge tube. (vii) Add 5 mL water. Cap and shake to mix. (b) Slurry extraction (i) Add 0.1 mL Carrez I solution and 0.1 mL Carrez II solution. Shake to mix. (ii) Stand for 20 min. Shake to mix every five minutes. (iii) Centrifuge at 2000 × g for 10 min. (iv) Syringe filter supernatant into a microcentrifuge vial. (v) Transfer 0.1 mL extract in the microcentrifuge vial into HPLC vial, add 0.9 mL acetonitrile. Cap and vortex mix.

G.

Chromatography (a) Set-up the UHPLC system with the following configuration: Table XXXX.XXB. Chromatographic instrument settings Parameter Value Mobile phase

acetonitrile:water:formic acid 900:100:1 v/v/v

Column

Kinetex HILIC 2.6 μm, 4.6 × 100 mm

Oven temperature Chiller temperature Injection volume

30 °C 10 °C

5 μL

0.8 mL min − 1 (isocratic)

Flow rate:

H.

Mass Spectrometry (a) Set up the mass spectrometer with the following instrument settings