Isolation of casein for stable isotope ratio analysis of butter, cheese, and milk powder

Abstract

Rationale: Stable isotope ratio analysis (SIRA) is commonly used for the authentication of dairy commodities, providing evidence to support the geographical origin and production background of products. We set out to optimise methods for the isolation of a common constituent (casein) from three dairy commodities, which would permit easier inter- and intra-commodity comparisons following SIRA.

Methods: Three published methods for isolation of protein (from cheese, milk, and butter) were adapted to yield protein (casein) fractions from commercial cheddar cheese, whole milk powder (WMP), and butter samples with a high degree of purity for subsequent SIRA. The casein fractions isolated underwent elemental analysis (H, C, and N), protein determination, and some also underwent SIRA of O and S. Two-way analysis of variance and Tukey post hoc comparisons tested differences between methods.

Results: For each product, an optimised casein isolation method was chosen based on the C/N ratio and protein content. An optimum solvent lipid extraction (petroleum spirit-diethyl ether (2:1)) and casein precipitation method was chosen for cheddar cheese casein. A final solvent lipid extraction (heptane-isopropanol (3:2)) was necessary for WMP and butter casein extraction. δ¹³ C and δ² H values validated the methods' abilities to remove contaminating lipid and isolate pure casein.

Conclusions: Casein of high purity, for subsequent SIRA, can be isolated from cheddar cheese, WMP, and butter following modifications of previously published methods.