AOAC SPIFAN ERP Reviewer Forms (March 16, 2017)

FOR EXPERT REVIEW PANEL USE ONLY NOT FOR DISTRIBUTION

Expert Review Panel for Infant Formula and Adult Nutrition

Evaluation of Method AOAC 2016.05: Vitamin D in Nutritional Products by LC- MS/MS

Title: Analysis of Vitamin D2 and Vitamin D3 by LC MS/MS in Infant Formulas and Nutritionals Products, First Action 2016.05: Multi-Laboratory Testing

Author: Brendon Gill, Fonterra Co-operative Group

Summary of Method: Samples are saponified at high temperature, and undergo Liquid-liquid extraction into isooctane. Then the extract is derivatized with 4-phenyl-1,2,4-triazoline-3,5-dione (PTAD) with a 5 min reaction. The vitamin D- adduct is then back-extracted into a small volume of acetonitrile and analysed by reversed-phase liquid chromatography. Detection is by mass spectrometry using multiple reaction monitoring (MRM). Stable isotope labelled d6 -vitamin D2 and d6 -vitamin D3 internal standards are used as internal standards. Method Scope/Applicability: This method is applicable to powders, RTF liquids, and liquid concentrates of vitamin D fortified infant, adult, and pediatric nutritional formulas. The previtamin Ds are not determined or monitored by this method. General comments about the method: Excellent precision and reproducibility data. Good concentration coverage. The concentration range in the tested samples is from 0.7 to 10.2 ug/hg sample (powder), which corresponds to 0.08 – 1.1ug/hg of RTF products; The SMPR requirement is 0.12-5.1 ug/hg;

Method Clarity: Method is well written, and clear to follow. One typo in method section G (d), 70 o C should be 0 o C.

Method Safety Concerns: Generally addressed by Caution notice at the beginning of the method

Pros/Strengths: • Excellent precision and reproducibility results Cons/Weaknesses • Previtamin Ds are not measured or monitored.

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