AOAC SPIFAN Stakeholder Panel Meeting Book (March 12, 2020)

Summary of Existing Methods in OMA, Chapter 50

Method

Status

Method Applicability

Comments

2011.11 “Vitamin D in Infant Formula

• AOAC First Action

• Vitamin D 2 and D 3 • Applicable to Infant Formula and Adult/Pediatric Nutritional Formula • Range, LOD, LOQ, RSD r pass stated SMPR • RSD R generally passes SMPR – failed for a few matrices • Vitamin D 2 and D 3 • Applicable to Infant Formula and Adult/Pediatric Nutritional Formula • Range, LOD, LOQ, RSD r (generally) pass SMPR • RSD R not determined

• Sap, Extraction • RP UHPLC; APcI-MS/MS Analysis • d3 -D 2 and d3 -D 3 Stable Isotope IS • pre-D 2 and pre-D 3 measured • SLV/MLT completed on SPIFAN kit • Selectivity via MS/MS is good • some chromatography challenges noted

and Adult/Pediatric Nutritional Formula”

2012.11 "Simultaneous Determination of Vitamins D 3 and D 2 by ESI LC-MS/MS"

• AOAC First Action

• Sap, Extraction • RP UHPLC; ESI-MS/MS Analysis • d3 -D 2 and d3 -D 3 Stable Isotope IS • pre-D 2 (D 2 ) and pre-D 3 (D 3 ) coelute • Selectivity via MS/MS • some chromatography challenges addressed by ESI • MLT not completed • Sap, Extraction; PTAD Adduct • RP UHPLC; APcI-MS/MS Analysis • d6 -D 2 and d6 -D 3 Stable Isotope IS • pre-D 2 and pre-D 3 included • Selectivity via PTAD adduct and LC-MS/MS is excellent

2016.05 “Analysis of Vitamin D 2 and Vitamin D 3 in Fortified Milk Powders, Infant Formulas, and Adult/Pediatric Nutritional Formulas”

• AOAC Final Action Method • ISO 20636 • Codex Type II (Infant Formula)

• Vitamin D 2 and D 3 • SPIFAN Matrices • SLV and MLT completed pass SMPR

LC-MS/MS Methods

Stable-Isotope Internal Standardization Improved Selectivity compared to LC-UV based methods Even so, some sample matrix challenges Selectivity of OMA 2016.05 is very good due to PTAD Adduct

Conclusions

Regulatory Landscape has changed (EU) Vitamin D SMPR amendment needed A single SMPR is adequate All methods reviewed are generally fit-for-purpose LC-UV methods have limited selectivity vs LC-MS/MS methods LC-UV methods have not been formally assessed using current product formulations UHPLC-MS/MS selectivity is generally improved; can utilize Stable Isotope IS Chromatography of direct-measure methods is matrix challenged Selectivity of OMA 2015.15 is very good due to PTAD Adduct with UHPLC-MS/ MS These methods still have utility (Codex Type III, QC) Other findings SPIFAN methods are applicable to Codex categories beyond Infant Formula Follow on Formula; Special Foods (FSDU) Not being captured in Codex Stan 234 (992.26 listed as Type II for FSDU Special Foods)