HPLC Determination of Total Tryptophan in Infant Formula and Adult/Pediatric Nutritional Formula Following Enzymatic Hydrolysis: Single-Laboratory Validation, First Action 2017.03

Abstract

Abstract A method for tryptophan (Trp) analysis designed to comply with AOAC Standard Method Performance Requirement 2014.013 is described. Unlike AOAC 988.15, which uses alkaline hydrolysis, this method uses enzymatic hydrolysis to release the Trp from the intact protein. The method achieves an LOQ of 0.18 mg/100 g Trp on a ready-to-feed basis with mean recoveries ranging from 93.8 to 104.9%. Repeatability ranged from 0.2 to 5.0%. Intermediate precision ranged from 1.0 to 6.9%. The analytical range was determined to be 0.18–300 mg/100 g, with linearity over eight calibration standard levels giving an average deviation from theoretical levels of 0.3%. No single calibration point had a deviation of >5.0%. Two standard reference materials (SRMs 1849a and 927e) were analyzed, and the average deviation from the certified value was 98.5% for SRM 1849a and 101.2% for SRM 927e. Sample preparation is very similar to existing methods in terms of time and complexity. The use of an internal standard reduces laboratory error and allows for reproducible results.