Separation and quantification of major milk proteins in different species by reversed phase high performance liquid chromatography
In order to detect milk proteins, reversed phase high performance liquid chromatography (RP-HPLC) (Waters 2695 Series chromatograph, USA) was successfully performed to separate and quantify the major proteins from cow (n=20), goat (n=20), buffalo (n=20) and yak (n=20) milk which were collected around the fourth month of lactation for each species from February to April, 2013 within 30 min. Descriptive statistics and Duncan’s multiple comparison of protein contents were obtained by SAS 9.1 via GLM model (SAS Institute Inc., Cary, NC). Main protein data sets were analyzed by PCA using the Unscrambler 9.8 (CAMO software AS, Oslo, Norway). Results showed that the contents (g/L) of milk total protein were significantly different in milk from yak (62.87), buffalo (56.09), goat (42.83) and cow (37.35), respectively. Average content of major protein fractions obtained were all converted by purity of individual protein standard (Sigma-Aldrich, St. Louis, MO, USA) to assess the real individual protein content. Composition (g/L) of yak milk contained the highest κ-CN content (9.80), and this was significantly higher than in buffalo, cow, and goat milk (7.35, 6.24 and 5.56). Content (g/L) of α-CN in yak milk (16.14) was significantly higher than those for buffalo, cow, and goat milk (11.53, 11.81 and 9.48). The β-CN content (g/L) was similar in yak and buffalo milk (23.34, 22.68), but higher than in goat and cow milk (20.53, 13.31). The β-LgB quantification (g/L) in goat and buffalo milk was similar (3.69, 3.88), significantly lower than in yak milk (9.46), but higher than in cow milk (1.19). The α-La content (g/L) was highest in buffalo milk (7.96), while significantly lower in cow, goat and yak milk (1.45, 1.28 and 1.47). Cow milk contained highest content (g/L) of β-LgA (2.15), and this was significantly higher than that in buffalo, yak and goat milk (0.13, 0.12 and 0.14). And different chromatographic profiles were obtained for them. The data of milk protein can be differentiated according to animal species by principal component analysis (PCA). It was concluded that major milk proteins from different animals had special profiles.
Keywords: milk protein, reversed phase high performance liquid chromatography, species