Capuano E, Alewijn M, van Ruth SM, Verkaik-Kloosterman J
RIVM Rapport 350124003
Adequate methods with known performance characteristics used for the assessment of the amount of nutrients in formula are essential in the evaluation whether the composition deviates from the legal amounts. Many standardised analytical methods for the analysis of nutrients in formula are internationally available. Currently, EU regulation regarding formula (commission directive 2006/141/EC) does not provide any indication on reference methods to be used in official controls for the analysis of vitamins, choline and inositol in formula. Therefore, an overview of the available standardised methods of analysis for the measurement of vitamins, inositol and choline in infant and follow-on formula is provided. 89 standardised analytical methods from both national and international organizations developing standard analytical methods have been collected and their performance characteristics (i.e. analytical technique, target compound, matrixes validated, LOD, LOQ, analytical range, repeatability, reproducibility, accuracy/spike recovery) are described. For most nutrients studied, several standardised analytical methods are available, with a maximum of 12 for vitamin A. Especially for water-soluble vitamins some current standardised methods are more than 25 years old. However, methods of analysis based on more modern analytical technologies have been developed in the recent decades. 11 new official AOAC methods for the analysis of Vitamin A, D, E, B12, Folic acid and based on more advanced techniques have been adopted as First Action method in 2011 and new AOAC official methods of analysis are expected to be approved in the near future for other nutrients. For several nutrients the overall activity is the sum of the activities of several forms of the compound. However, it is important to notice that the analytical methods available may not always measure the overall activity but only a limited proportion based on specific forms of the compound. In the EU legislation the compound forms that may be used in the manufacturing of formula are listed, however, it does not provide unambiguous specification about the forms of the vitamin to be assessed. Moreover, the minimum and maximum values are provided per energy content, logic from a nutritional point of view. This means that both the uncertainty in the energy calculation and the uncertainty in the measurement of the target nutrient contribute to the total uncertainty in the assessment of the nutrient content when this is provided per energy content. The existence of many different available methods with different performance characteristics and possibly different target forms of the nutrients is identified as a serious gap in the current harmonisation status. In absence of relevant European Union rules, the choice of the best method of analysis for each of the target nutrients in formula should be based on several factors, such as the capacity of the method to assess all the forms of the vitamin that are nutritionally relevant in formula and the analytical performances of the method (i.e. accuracy/spike recovery, reproducibility, lower and upper limit of quantification). As CODEX is the most internationally recognized organization publishing standardised analytical methods, the methods of analysis recommended by CODEX might, although arbitrarily chosen, be taken as guidance for the choice.