AOAC SPIFAN Nutrients ERP (December 7, 2022)

Amino-06 (SLV) 10-2022 FOR ERP USE ONLY DO NOT DISTRIBUTE

m. Centrifuge tubes with Screw cap tube - 15 mL (Sarstedt) or equivalent n. Filter paper – Whatman Grade 595 (Cytivia) or equivalent o. Syringe filters – 0,20 µm Regenerated Cellulose (Macherey-Nagel) or equivalent p. HPLC vials – 2 mL with suitable screw cap or equivalent

D. Chemicals and reagents

Note: All reagents and water should be of analytical grade.

a. Taurine – Certified purity >99% (Sigma-Aldrich) or equivalent b. 2-Aminoethane-d 4 -sulfonic acid – (CDN Isotopes) or equivalent c. Acetonitrile - Hypergrade for LC-MS (Merck-Millipore) or equivalent d. Trichloroacetic acid – (J.T. Baker, Aventor) or equivalent e. Ammonium Formate – (Fluka) or equivalent f. Formic acid – UPLC-MS (Biosolve) or equivalent g. UHQ Water – ≥ 18,2M Ω ·cm (Merck Millipore Milli-Q) or equivalent

E. Preparation of reagents

a. Taurine standard stock solutions Weigh an amount of taurine according to table 1 accurately at 0,1 mg in seven volumetric flasks of 500 mL. Dissolve in approximately 20 mL UHQ water for one minute by ultra-sonification. Fill the volumetric flasks to the mark and homogenize the solutions. Stock solutions can stored in portions for at least 2 months at -20°C.

Table 1. Standard stock solution preparation

Standard Stock Solution

Taurine (mg)

Taurine Stock Concentration (ug/mL)

T1 T2 T3 T4 T5 T6 T7

7,5

15

62,5 125 250 625 1250 2500

125 250 500

1250 2500 5400

b. 2-Aminoethane-d 4 -sulfonic acid IS stock solution Weigh accurately 50 mg of 2-aminoethane-d 4 -sulfonic acid onto a weighing boat. Transfer quantitatively into a 50 mL volumetric flask. Fill the volumetric flask with UHQ water to the mark and homogenize. I.S. stock solutions can be stored at least for 2 months at -20°C. c. Trichloroacetic acid (TCA) solution (50%) Weigh with the help of a porcelain spatula 250 grams of trichloroacetic acid into a beaker glass. Add approximately 200 mL of UHQ water. Transfer in a 500 mL volumetric flask. Fill the flask to the mark and homogenize.