The Study Of New Methods For Isolation And Quantification Of Seven Major Proteins From Dairy

The quality of food products has become more and more attention as the social and economic importance of the dairy products market. The analytical methods of milk quality show especially significance in the field of food quality. It requests the development of the analytical methods to suit the market for controlling the origin and quality of produts. Meanwhile, some fraudulent manipulations appear frequently, for instance, Fraud may add some amino acids and vegetable proteins into the products for cutting down the cost. And with the improvement of living standards, dairy products will become functional foods when we add some proteins or peptides into formula milk. GB (GB/T 5413.2- 1997) has been unable to meet the increasingly stringent requirements of analysis and has been unable to accurately reflect the composition of proteins in dairy products. Therefore, we are eager to need a stable and universal dairy analysis method to quantitate each component of dairy products. In this paper, two kinds of quantitative analysis methods, RP-HPLC and SEC are successfully established. We compare characteristics of the three analysis methods (RP-HPLC, SEC, SDS-PAGE) in protein identification and quantitation. In the liquid chromatography method, we use the SAS statistical software to design experiment and optimize experimental conditions. With the traditional single-factor test compared, SAS can also consider a variety of factors. SAS software program can establish the data model based on various changed factors. The interaction relationship among various factors were quantified. All information due to changing various factors can be obtained by carrying out a small number of experiments. We identify each protein by using the retention time and peak area ratio and quantitate protein according to the standard curve. Then, we quantitated a variety of samples from market, such as milk powder, liquid milk and unprocessed raw milk. In our study, the puzzle that denaturation proteins are not accurately quantitated was resolved. In the established method, the samples don’t need to centrifuge comparing to the traditional experimental methods. In the determination of albumin components, the predominant casein proteins are not removed too. We simplify the test steps to avoid the cumbersome process as a result of the loss components, improving the recovery rate of the proteins. The seven major proteins (κ-casein,α-casein,α-casein,β-casein,α-lactalbumin,β-lactoglobulin B andβ-lactoglobulin A) may be separated and quantified in one run by the RP-HPLC method. The hard issue separatingκ-CN fromαs2-CN is resolved, as well asα-La ,β-LgB andβ-LgA. By now, these methods we established are unique and not only have a significant value in the field of protein variant separation and accurate quantitation but also have a very broad application prospect. The RP-HPLC can test the content of natural albumin active components. While, the SEC can quantitate the content of the most variations including natural and nonnatural things. The SEC method resolve the puzzle whichα-La andβ-LgB of the deep-processing products can be precisely quantitated. As same as the RP-HPLC, the SEC has good stability and reproducibility as well as higher recovery rate. As reference standard, GB (GB/T 5413.2-1997) contrast with the RP-HPLC and SEC methods in the area of identification and quantitation.