AOAC SPIFAN Nutrients ERP-Final Review (Jan. 2019)

2017.03 (JAN 2019) Amino-02-(ERP Review for Pubs) Tryptophan by enz hyd_Abbott Labs_Method_10172018

FOR ERP USE ONLY DO NOT DISTRIBUTE

HPLC Determination of Total Tryptophan in Infant Formula and Adult/Pediatric Nutritional Formula Following Enzymatic Hydrolysis Jonathan Draher and Norman White Abbott Nutrition, Division of Abbott Laboratories l) Stir bars m) Stir plates n) ACRODISC disposable filter assembly, 0.45 micron pore size From Gelman Scientific, cat. # 4217, or equivalent o) 3 cc or 5 cc disposable syringe (s) p) Weighing paper q) Weigh boats r) Funnel(s) s) Water bottle t) Membrane filter, 0.45 micron u) Autosampler vials and caps (see autosampler requirements). v) Analytical Column: Preferred column: YMC PACK C8, 3 um particle, 3.0 50 mm – YMC America, INC, Product No. OC12S03-0503WT, or alternate w) HPLC system with autosampler, column oven to maintain 30°C, Fluorescence Detector, binary or quaternary HPLC Pump with degasser, HPLC peak area integrator or chromatography data processor. 2. Instrument Parameters and Conditions a) Mobile Phase Flow Rate : 0.5 mL /minute b) Column Temperature: 30 degrees Centigrade c) Injection volume: 10 µ L. d) Detector settings: Excitation wavelength to 295 nm and emission wavelength to 345 nm e) Typical PMT setting: 7 for Agilent G1321A f) Run time: 8.5 minutes, or as needed to fully collect the 5-methyl-DL-tryptophan peak.

D. REAGENTS 1. Standards L-Tryptophan USP Standard Cat #1700501, or equivalent. 2. Chemicals

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