Determination of Caseinomacropeptide by an RP‐HPLC Method and Monitoring of the Addition of Rennet Whey to Powdered Milk

Abstract

A precise, sensitive, and reliable RP‐HPLC method was developed and validated to enable the separation and quantification of caseinomacro peptide. The optimized method used a polystyrene‐divinylbenzene column and gradient elution with two solvents. Solvent A was 0.1% trifluoroacetic acid in water and solvent B was 95% acetonitrile‐5% water‐0.1% trifluoroacetic acid. The flow rate used was 1 mL/min. The effluent was monitored by a UV detector at 214 nm and enabled the separation of two peaks corresponding to the aglyco components of the two principal genetic variants (A and B), eluted after the less well resolved glyco caseinomacropeptide components. The determinations were performed in the linear range of 15–200 µg/mL. Detection limit was 2 µg/mL. The validity of the method was verified. The recoveries were 93 and 98%. The precision of the method was also evaluated being the %CV less than 4.87%. The method was applied to the analysis of rennet and acid wheys and whey protein concentrates produced by the dairy industry. It was also applied to the detection of rennet whey in powdered milks.