Utilization Of The Pancreas: Single-component Separation And Purification Of Porcine Insulin

Insulin is one of the most important biochemical medicines extracted from porcine pancreas. When existing insulin, most of other enzymes would lose their activities, so the leavings are useless, which is a huge waste to industrial production. So what in study is to look for a condition which can extract insulin effectively and can extract other medicinal enzymes using the leavings, in order to give a base to the general utilize of pancreas.First of all, test the extracting terms, and make sure that under this condition other enzymes will keep their activities. It includes the following four aspects: Exaction Factor I, Exaction Factor II, hours of extracting and times of extracting. Through experiments, it is the best extracting conditions when the Factor I is C, Factor II is b, two hours extracting and extracted twice. Besides, temperature could also effects the extracting, so we choose temperature between 0-4℃.When purifying insulin, we do not use the traditional ways which are reduced pressure concentration and salt out, but use chromatogram I to separate insulin from ethanol solution. Chromatogram I can wipe off fat and pigment from insulin extractions, so the eluting solution is pellucid, which is a convenient for the next close purification.Use chromatogram II to get mono component insulin(MCI). After this process, insulin purity can reach to 95%; then use chromatogram III, which can make the insulin purity reach over 99%. Using SDS-PAGE and NATIVE-PAGE to detect the purity, the prepared sample reaches electrophoretic purity; using gel- filtration to detect large molecule protein impurity, the result shows impurity protein peak area is no more than 1% insulin peak area, which according to the ChP2005. Using protein content determination, every 1kg pancreas can obtain 7.46mg insulin, which is almost one time more than the traditional method. Using HPLC to detect insulin, the sample insulin has the same time of appearing peak with standard insulin; the sample insulin purity reaches more than 99%. Using Mouse convulsion way to detect insulin activity, under the set dose, mouce give the convulsion phenomenon, which also according to the ChP2005.