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Background: A method for the quantification of total amino acids (including taurine and excluding tryptophan) using ultra-HPLC separation coupled to UV detection (UHPLC-UV) was granted First Action status (AOAC 2018.06) by the AOAC INTERNATIONAL Stakeholder Program for Infant Formula and Adult Nutritionals (SPIFAN) in 2018. Objective: An interlaboratory study was conducted to further assess method performance against the AOAC Standard Method Performance Requirements (AOAC SMPR® 2014.013). Dairy and cereal matrixes were added to expand the scope of the method in collaboration with IDF (International Dairy Federation), ISO (International Organization for Standardization), and AACCI (American Association of Cereal Chemists International, now Cereals & Grains Association). Methods: Sixteen different matrixes were chosen to cover the requirements of AOAC, IDF/ISO, and AACCI. Blind duplicate samples were organized into specific series to ensure that each pair was analyzed on the same day. Fifteen laboratories returned results. Data fromfour laboratories were considered invalid and removed fromthe dataset. Remaining data were assessed according to the Appendix D of the AOAC Official Methods of AnalysisSM (guidelines for collaborative study procedures). Results: This method generally met the requirements listed in the SMPR for infant formulas and adult nutritionals, except for taurine. Method performance was comparable in dairy and cereal matrixes. Five different UHPLC instruments were used with either commercial or in-house reagents, demonstrating that the method is not limited to a single supplier. Conclusion: This method was recommended for Final Action in infant and adult/pediatric nutritional formulas by the AOAC SPIFAN Nutrients Expert Review Panel in April 2021, with the exception of taurine. The corresponding IDF/ISO Draft International Standard (DIS) was approved by national bodies in May 2022, and comments collected during the ballot were incorporated into this manuscript. Highlights: AOAC Official Method 2018.06 for the determination of total amino acids in infant formulas, adult nutritionals, dairy, and cereal matrixes was successfully validated in an interlaboratory study. [ABSTRACT FROM AUTHOR]

A β-casein enriched whey produced from skim milk using a 2-step microfiltration process was used to manufacture a β-casein enriched infant formula (β-CN+ IF) with about 60% of β-CN in its casein fraction. Bringing the casein composition closer to that of human milk affected gelation properties and in vitro proteolysis in comparison with a control IF with the casein profile of cow milk. β-CN+ IF had a higher buffering capacity and produced a weaker gel during acidification but β-CN+ IF exhibited a lower degree of protein hydrolysis (DH) and slower β-CN proteolysis during in vitro gastric digestion. No marked differences were observed during the intestinal phase. β-CN bioactive peptides were more abundant in the β-CN+ IF digesta. Further investigations are needed to assess the impact of these differences on amino acid absorption, gastrointestinal hormone release and gut microbiota in babies fed with this formula composition. [ABSTRACT FROM AUTHOR]

A multilaboratory testing study was conducted on AOAC First Action Official MethodSM 2016.15: Quantification of Whey Protein Content in Infant Formula Powders by Sodium Dodecyl Sulfate-Capillary Gel Electrophoresis (SDS-CGE). Nineteen laboratories participated in the analysis of duplicate blind-coded samples of 15 formula powder products for infants and young children. Electrophoregrams were recorded at UV220 nm and integrated. The normalized peak areas of whey and casein proteins were summed separately to calculate total whey protein content. Apart from one sample [NIST Standard Reference Material (SRM) 1849a], relative standard deviation of repeatability (RSDr) and reproducibility (RSDR) ranged from 0.83 to 2.11% and from 2.18 to 4.22%, respectively, and Horwitz ratios ranged from 1.02 to 1.85, meeting the precision limits specified in the whey protein Standard Method Performance Requirements and in the guidelines recommended for the Horwitz ratio. In these samples, the measured whey protein content was between 98 and 108% of the declared value. NIST SRM 1849a showed atypical results, with elevated RSDr (3.51%), RSDR (5.94%), Horwitz ratio (2.62), and recovery (134%). There is no clear reason for this. The percent whey protein value for NIST is calculated from the formulation and is not a reference or certified value. Multiple instrument models and makes, as well as capillary sources, were used in this collaborative study, demonstrating the robustness of the method. The method is fit-for-purpose for the quantification of whey protein content in milk-based formula powder products for infants and young children. It is not applicable to the analysis of hydrolyzed or plant protein-based infant formulas. [ABSTRACT FROM AUTHOR]

The article focuses on a study reagrding the quantification of whey protein content in infant formulas by sodium dodecyl sulfate-capillary gel electrophoresis (SDS-CGE). It mentions that proteins in infant formula samples are denatured by anionic surfactant SDS and states that infant formula electrophoregrams are optimized by batch analysis. It also mentions the use of acidic wash solution and SDS gel buffer for quantification of whey protein.

Protein separation by sodium dodecyl sulfate-capillary gel electrophoresis, followed by UV absorption at 220 nm, allows for the quantification of major proteins in raw milk. In processed dairy samples such as skim milk powder (SMP) and infant formulas, signals from individual proteins are less resolved, but caseins still migrate as one family between two groups of whey proteins. In the first group, α-lactalbumin and β-lactoglobulin migrate as two distinct peaks. Lactosylated adducts show delayed migration times and interfere with peak separation, but both native and modified forms as well as other low-MW whey proteins still elute before the caseins. The second group contains high-MW whey proteins (including bovine serum albumin, lactoferrin, and immunoglobulins) and elutes after the caseins. Caseins and whey proteins can thus be considered two distinct nonoverlapping families whose ratio can be established based on integrated areas without the need for a calibration curve. Because mass-to-area response factors for whey proteins and caseins are different, an area correction factor was determined from experimental measurement using SMP. Method performance assessed on five infant formulas showed RSDs of 0.2-1.2% (within day) and 0.5-1.1% (multiple days), with average recoveries between 97.4 and 106.4% of added whey protein. Forty-three different infant formulas and milk powders were analyzed. Of the 41 samples with manufacturer claims, the measured whey protein content was in close agreement with declared values, falling within 5% of the declared value in 76% of samples and within 10% in 95% of samples. [ABSTRACT FROM AUTHOR]

Filtration is frequently employed in the dairy industry to tailor protein profiles and manipulate the nutri- and techno-functionality of ingredients. One milk protein of particular interest in infant formula is β-casein. However, during the enrichment of β-casein, co-fractionation of κ-casein, as 2–10% of the total protein, is common. In this study, isolation of β-casein from micellar casein using filtration, incorporating chymosin-treatment, was investigated. At 5 °C, casein curd formation can be avoided, allowing the fractionation of liquid streams. Three treatments with chymosin were applied: feed without treatment and feed or permeate treated with chymosin. An average β-casein enrichment of ∼90% (w/w) total protein was achieved. Treatment of feed with chymosin resulted in caseinomacropeptide (CMP)-enrichment. CMP-enriched β-casein showed a wider particle size distribution at 50 °C, and a reduced level of solubility. The novel materials produced may be relevant in applications such as encapsulation, emulsification, infant formula and cheese manufacture. [ABSTRACT FROM AUTHOR]

Background: An acid hydrolysis ultrahighperformance LC-UV method was evaluated for the determination of total amino acids in infant formula and adult/pediatric nutritional formula. Objective: It was assessed for compliance against AOAC INTERNATIONAL Standard Method Performance Requirements (SMPR®) established by the Stakeholder Panel for Infant Formula and Adult Nutritionals (SPIFAN). Methods: A single-laboratory validation (SLV) study was conducted as a first step in the process to validate the method. In this SLV, 17 SPIFAN matrices representing a range of infant formula and adult nutritional products were evaluated for their amino acid content. Results: The analytical range was found to be within the needs for all products; some may require a dilution. Evaluation of trueness performed on Standard Reference Material 1849a (Infant/Adult Nutritional Formula) showed all compounds met the SMPR theoretical value, with exceptions for threonine and tyrosine. These may have a bias for the National Institute of Standards and Technology (NIST) data, depending on hydrolysis used in the determination of the NIST certificate of analysis. Conclusions: Based on the results of this SLV, this method met the SMPR and was approved as a First Action method by the AOAC Expert Review Panel on Nutrient Methods on August 28, 2018. [ABSTRACT FROM AUTHOR]