Studies On Extract And Dring Purify Of Immunoglobulin G In Colostrum

Immunoglobulin G is an important functional component of colostrum, and it's comprehensive research is an important issue for the dairy industry. The purpose of this task is to study the method of extracting high purity immunoglobulin G from colostrum, and through the research on vacuum freeze-drying technology to get high activity immunoglobulin G. The achievement of this task will help comprehensive utilization of colostrum resources, will fully reflect the potential value of the available of colostrum resources, broaden the application areas of immunoglobulin G and have greater economic and social benefits.In this task, the determination of total protein content of colostrum, the extraction of immunoglobulin G and the process of vacuum freeze-drying for purificated eluate were studied. This study provided the theoretical and technical support for comprehensive utilization of IgG in colostrum. The main results of the study are as follows:1. Determine the determination of total protein content of colostrum through studies. The absorbance decreases with time in half-hour when use bradford G-250 method and di-wavelength ultraviolet spectrophotometry method to determine the total protein content of colostrum, but the degree of change of di-wavelength ultraviolet spectrophotometry method is smaller than bradford G-250 method, and it proves that the stability of di-wavelength ultraviolet spectrophotometry method is better than bradford G-250 method. The result of 6 parallel determinations for the same sample is that the RSD of bradford G-250 method is 2.041%, and the RSD of di-wavelength ultraviolet spectrophotometry method is 0.082%, and it proves that the accuracy stability of di-wavelength ultraviolet spectrophotometry method is better than bradford G-250 method. It can be defined that using di-wavelength ultraviolet spectrophotometry to determine total protein content of colostrum is a good method.2. Determine the best technology of using ammonium sulfate-extraction to extract the immunoglobulin G in colostrum through studies. The RSD is 0.430% and the recovery rate is 99.58% when using SRID method to determine the content of IgG. The optimal parameters of ammonium sulfate-extraction to extract the immunoglobulin G are as follows in colostrum:the centrifugal speed is 4000r/min, the temperature is 40℃, the concentration of ammonium sulfate is 35%, the pH-number is 7.4. Under these conditions to do verifying test, the result is the purity of IgG is 61.72%, and the content of IgG is 28.41mg/mL. The conditions of gel chromatography are as follows:the afiller is Sephadex G-75, the specification of chromatography column is 1.6×70cm, the elution is phosphate buffer whose concentration is 0.01mol/L and pH is 7.4, the flow rate is 0.5mL/min, the sample volume is 5mL. The content of IgG is 8.26mg/mL, the content of total protein is 8.87 mg/mL and the purity of IgG is 93.12% after purification with gel chromatography.3. Determine the best technology of vacuum freeze-drying to immunoglobulin G. The Best parameters of vacuum freeze-drying are as follows:the glycine and sucrose ratio is 1:1 by 10% to add, the install column height is 8mm, pre-freezing temperature is-20℃and the pre-freeze time is 2h. Under these conditions to drying the purified eluate with vacuum freeze-drying method, the result is the rate of residual.activity of IgG is 92.64%.