Active gel electrophoresis method for lactalbumin

A gel electrophoresis and whey protein technology, applied in the field of protein electrophoresis, can solve the problems of expensive equipment, sample absorption, and high analysis cost, and achieve the effects of less sample consumption, easy application, and high sensitivity

Inactive Publication Date: 2009-07-08
CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

These methods have their own advantages, but the required equipment is expensive, the analysis cost is high, the technology is strong, and there are many factors that affect the test results, and the immune method is usually difficult to achieve under ordinary conditions because it needs to obtain the antibody of the corresponding protein first. And the repeatability is not good; the capillary electrophoresis method also has the problem of sample absorption; in addition, each of the above methods cannot comprehensively summarize the changes in the whey protein of heat-treated dairy products, so it is necessary to find a simple, effective and comprehensive method for detecting whey Method of protein change to identify dairy products with different heat treatments

Method used

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  • Active gel electrophoresis method for lactalbumin
  • Active gel electrophoresis method for lactalbumin
  • Active gel electrophoresis method for lactalbumin

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0061] Example 1 Active Gel Electrophoresis Spectrum and SDS Gel Electrophoresis Electrophoresis Comparison

[0062] Take fresh cow's milk and commercially available UHT fresh milk, centrifuge at 8500g at 4°C for 15min to skim the fat, and divide them evenly into active gel electrophoresis group and SDS gel electrophoresis group. Fresh milk samples from the above two groups (commercially available fresh UHT milk without heat treatment) were heat treated according to the following processes: 1) without heat treatment; 2) 65°C / 30min; 3) 85°C / 20s; 4) frozen at -80°C for 24h, and the heat-treated samples were cooled to room temperature.

[0063] The pH of all samples was adjusted to 4.6 with 6% lactic acid, and after standing for 15 minutes, the casein was removed by centrifugation at 8500g at 4°C for 15 minutes to obtain whey protein, which was stored at -20°C for later use.

[0064] Prepare SDS polyacrylamide gel with 12% separating gel and 4% stacking gel. First prepare 5 mL ...

Embodiment 2

[0073] Example 2 Different heating temperatures, heating times, active gel electrophoresis patterns of heat-treated bovine milk whey protein at pH values

[0074] 1) Active gel electrophoresis patterns of heat-treated bovine whey protein with different heating temperatures and the same heating time:

[0075] Fresh milk was degreased by centrifugation at 8500g at 4°C for 15 minutes, distributed evenly, and heated at 65°C, 75°C, 85°C, 95°C, and 100°C for 10 minutes, and the samples were cooled rapidly after heating. After cooling to room temperature, the pH was adjusted to 4.6 with 2% hydrochloric acid, and after standing for 15 minutes, the casein was removed by centrifugation at 10,000 g at 4°C for 15 minutes, and the obtained supernatant was whey protein. The whey protein was aliquoted and stored at -20°C for later use.

[0076] Prepare an active acrylamide gel with 8% separating gel and 4% stacking gel. The whey protein sample was mixed with 2 times active protein loading ...

Embodiment 3

[0092] Example 3 Identification of Heat-treated Bovine Whey Whey Protein by Active Gel Electrophoresis at Different Heating Temperatures and Different Heating Times

[0093] Fresh milk is degreased by centrifugation at 8500g at 4°C for 15 minutes, evenly distributed and heated at 65°C, 75°C, 85°C, 95°C, and 100°C. Each temperature is divided into different heating time gradients, respectively 5, 10, 20, 30min. The samples were cooled rapidly after heating. After cooling to room temperature, the pH was adjusted to 4.6 with 6% lactic acid, and after standing for 15 minutes, the casein was removed by centrifugation at 8500g at 4°C for 15 minutes, and the obtained supernatant was whey protein. The whey protein was aliquoted and stored at -20°C for later use.

[0094] Prepare active acrylamide gel with 10% separating gel and 4% stacking gel. Mix the whey protein sample with the active protein loading buffer at a ratio of 1:3, centrifuge, and load 12 μL of the sample.

[0095] T...

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Abstract

The invention relates to an active gel electrophoresis method for lactalbumin. The method utilizes active acrylamide gel, mixes a lactalbumin sample with a buffer solution, and then directly performs electrophoretic analysis under the conditions as follows: an electrophoresis buffer solution is a Tris-glycine buffer solution with the pH of 8.3; the sample loading amount is that the ratio of lactalbumin to 2-times active protein sample buffer solution is 1:3-5, 7.5 mu L; the voltage is between 75 and 150 V; the time is between 25 and 75 minutes; and the temperature is between 2 and 4 DEG C. The active electrophoresis method has the advantages of small quantity of samples, high resolution, high sensitivity, good repeatability and capability of accurately analyzing the content of various types of lactalbumin and sensitively judging the processing techniques and quality situations of dairy products.

Description

technical field [0001] The invention relates to a protein electrophoresis method, in particular to an active gel electrophoresis method for whey protein. Background technique [0002] Milk protein is mainly composed of casein and whey protein, of which the casein content is about 83%, and the whey protein content is about 17%. Whey protein refers to the protein (casein precipitation) that is still soluble in dairy products under the acidic condition of pH 4.6, mainly composed of bovine serum albumin (BSA), α-lactalbumin (α-La), β-lactoglobulin (β-Lg), immunoglobulin, lactoferrin, peroxidase and lysozyme. In order to ensure the quality of dairy products, dairy products undergo different heat treatment processes. During the heat treatment process, various substances of dairy products undergo significant changes, among which the change of whey protein is particularly obvious. The content of whey protein will increase with the heating temperature and temperature. The increase ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N27/447
Inventor 姜微波林素菊
Owner CHINA AGRI UNIV
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