ISO/TC 34/WG 14 - Vitamins, carotenoids and other nutrients

This document provides guidelines on: — how to express vitamin quantity, — the expression of different molecular forms in appropriate units, — and in some cases, vitamin activity, according to vitamers present or used in food products, in addition to the quantitative content determination obtained from ISO and CEN analytical standards. It provides information to be used as a basis for discussion between stakeholders and food control laboratories. It is not intended to be prescriptive or exhaustive.

This document specifies a method for the simultaneous quantitative determination of four water-soluble vitamins in infant formula and related nutritional products, including relevant forms of vitamins B1, B2, B3 and B6 by enzymatic digestion and UHPLC-MS/MS. This document is not intended to be used on products where vitamins have not been added.

This document specifies a method for the determination of total or free choline and carnitine in infant formula and adult nutritionals by liquid chromatography and tandem mass spectrometry (HPLC-MS/MS).

This document specifies a method for the quantitative determination of β-carotene and lycopene in infant formula and adult nutritionals in solid (i.e. powders) or liquid (i.e. ready-to-feed liquids and liquid concentrates) forms using reversed-phase ultra-high performance liquid chromatography (RP-UHPLC) and UV-visible detection. The application range runs from 1 μg/100 g to 1 500 μg/100 g for lycopene and from 1 μg/100 g to 2 250 μg/100 g for β-carotene. Based on the single-laboratory validation, the limit of detection (LOD) was 0,1 μg/100 g and the limit of quantification (LOQ) was 0,3 μg/100 g for each carotenoid. The method does not apply to materials that contain measurable levels of β-apo-8′-carotenal. The reproducibility data meets the requirements given in References [8] and [10]. Annex C specifies the determination of lutein. The reproducibility data does not meet the requirements given in Reference [9].

This document specifies a method for the quantitative determination of biotin and/or biocytin in fortified milk powders, infant formula and adult nutritionals in solid (i.e. powders) or liquid (i.e. ready-to-feed liquids and liquid concentrates) forms using liquid chromatography coupled with immunoaffinity column clean-up extraction. Precision data from an interlaboratory study is given in Annex B. A comparison between data obtained with the method in this document and EN 15607 is given in Annex C.

This document specifies a method for the quantitative determination of trans and total (cis + trans) vitamin K1 in infant, pediatric and adult nutritionals using normal phase (NP) high-performance liquid chromatography (HPLC) with post-column reduction and fluorescence detection. The method demonstrated good linearity over a standard range of ~2 μg/l to 80 μg/l trans vitamin K1, and the limit of quantification (LOQ) was estimated to be 0,4 μg/l for standards and 0,09 μg/100 g ready to feed (RTF) for samples assuming 4 grams of sample are diluted to 10 ml.

This document specifies a method for the quantitative determination of vitamin D2 and/or vitamin D3 in infant formula, and adult nutritionals in solid (i.e. powders) or liquid (i.e. ready-to-feed liquids and liquid concentrates) forms using liquid chromatography-mass spectrometry. The application range runs from 0,15 µg/100 g (limit of quantification) to 59 µg/100 g for vitamin D2 and from 0,25 µg/100 g to 65 µg/100 g for vitamin D3.

This document specifies a method for the determination of vitamin C (L-ascorbic acid) present in all forms of infant and adult formulas (powders, ready-to-feed liquids and liquid concentrates), using (ultra) high performance liquid chromatography with ultraviolet detection (U)HPLC-UV. The application range runs from 2,5 mg/100 g (limit of quantification) to 50 mg/100 g expressed in the product as consumed. The method is able to distinguish between D-ascorbic acid (isoascorbic- or erythorbic acid) and L-ascorbic acid.

ISO 20637:2015 specifies a method for the determination of myo-inositol (free or free plus bound as phosphatidylinositol) in infant formula and adult nutritionals using liquid chromatography and pulsed amperometry with column switching.

ISO 20639:2015 specifies a method for the quantitative determination of pantothenic acid, excluding bound forms, in infant formula and adult nutritionals (i.e. powders) using ultra high performance liquid chromatography and tandem mass spectrometry method (UHPLC-MS/MS).

ISO 20638:2015 specifies a method for the quantitative determination of 5′-mononucleotides in infant formula in solid (i.e. powders) or liquid (i.e. ready-to-feed liquids and liquid concentrates) forms using liquid chromatography.

ISO 20633:2015 specifies a method for the simultaneous quantitative determination of vitamin E (α-tocopherol and α-tocopheryl acetate) and vitamin A (13-cis and all-trans isomers of retinyl palmitate and retinyl acetate) present in all forms of infant and adult formulas (powders, ready-to-feed liquids and liquid concentrates). Retinol is not used for fortification purposes and therefore is not addressed in this method. The innate amount in products is insignificant. Stereoisomers of vitamin E, α-tocopherol and α-tocopheryl acetate, are not differentiated in this method.

ISO 20634:2015 specifies a method for the quantitative determination of vitamin B12 in infant and adult formula (powders, ready-to-feed liquids and liquid concentrates) by reversed phase high performance liquid chromatography.