SPIFAN ERP Final Action Review (June 9, 2017)

Study Title

Analysis of Vitamin D

and Vitamin D

by LC MS/MS in Infant Formulas and Nutritionals Products, First

Action 2016.05: Multi-Laboratory Testing.

Method Principle

Samples are saponified at high temperature then lipid soluble components are extracted into isooctane. A

portion of the isooctane layer is transferred, washed, and an aliquot of 4-phenyl-1,2,4-triazoline-3,5-dione

(PTAD) is added to derivatise vitamin D to form a high molecular mass, easily ionisable adduct. 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

-vitamin D

and

-vitamin D

internal standards are used for quantitation to correct for

losses in extraction and any variation in derivatisation and ionisation efficiencies.

Study Author

Brendon Gill, PhD

Senior Research Scientist

Fonterra Co-operative Group Ltd

P.O. Box 7, Waitoa 3341

New Zealand

Objective

The objective of this study is to complete the validation of AOAC 2016.05 by determination of inter-

laboratory precision (reproducibility) with a multi-laboratory testing study as fulfilment of the requirements as

a SPIFAN Final Action method.

Acknowledgments

The Study Director would sincerely like to thank the following laboratories and individuals for their

participation in this study: Andrew Todd and Mark Reynolds (Fonterra, New Zealand), Carolyn Burdette and

Melissa Phillips (NIST, USA), Danny Samson and Maurice Seegers (Mead-Johnson, Netherlands), Emeline

Tissot, Frédéric Martin, and Esther Campos-Gimenez (Nestlé, Switzerland), Greg Hostetler and Steve

Tennyson (Perrigo Nutritionals, USA), Marlène Daminato and Isabelle Malaviole (Aquanal, France), Shane

Wei (Fonterra, New Zealand), Tiffany Gallegos-Peretz and Sneh Bhandari (Merieux Nutrisciences), Vicky

Manti (Danone Nutricia, Netherlands), Yang Zhou (Eurofins, USA).

The assistance of Melissa Phillips (NIST, USA) for the supply of NIST1869 included as part of this study and

Shellie Stassi (Covance, USA) for arranging the shipping of the SPIFAN kits to participating labs is greatly

appreciated.

2016.05 (FEBRUARY 2017) VITD-18

MLT REPORT

FOR ERP USE ONLY

DO NOT DISTRIBUTE