溶 Lysozyme, also known as muramidase or N-acetyl glutathione hydrolase, is a very biochemical substance at home and abroad, widely used in clinical medicine. Has a variety of pharmacological effects, anti-infection, anti-inflammatory, detumescence, enhance the immune response in vivo, etc., have antibacterial effects, often used in the treatment of a variety of mucous membrane inflammation, dermatology, herpes zoster and other diseases. Is an excellent natural preservative, can be used for food preservation and preservation. It is particularly important that lysozyme has become an indispensable tool for genetic engineering and cell engineering in recent years. With the development of bioengineering, extraction of lysozyme has important strategic significance. Currently, the lysozyme used in the pharmaceutical industry is mainly extracted from egg white as the raw material. This article describes a new method for the simple extraction of lysozyme. First, the pharmaceutical raw materials: sodium chloride, acetic acid, sodium hydroxide. The above materials are all chemically pure reagents. Second, equipment: glass or ceramic containers, pH meter, fine gauze, glass stir bar, plastic dropper, Buchner funnel and so on. Third, the production methods:? (1) collection of egg: a knock out of a small hole in each of the fresh eggs, so that the egg white out (preferably fresh eggs, pH value of not less than 8, otherwise can not be used), according to its Add water twice the volume and gently stir for 5 minutes to make the consistency of the egg white solution uniform. Note that no bubbling can occur during the stirring process. The stirring should not be too fast. The stirring rod should be smooth to prevent protein denaturation and affect the lysozyme product. The yield and quality of the final, with a double gauze filter egg white solution in the umbilical cord block and broken eggshell and so on. (2) Add sodium chloride: add 2 g of sodium chloride per 100 ml of egg white solution, slowly add sodium chloride powder to the egg white solution, stir it while stirring to promote the timely dissolution of sodium chloride powder. In order to avoid excessive local concentration or precipitation in the bottom of the container, otherwise it will cause protein denaturation and produce a large number of white precipitate. (3) Crude lysozyme: After addition of sodium chloride fine powder, carefully adjust the pH of the egg white solution to 10.8 with 1 mol/l sodium hydroxide solution. Adjust the egg white with sodium hydroxide solution. The pH of the solution was dropped dropwise with a plastic dropper and stirring constantly to avoid local over-alkalization and protein denaturation, thereby affecting the yield and quality of lysozyme. In order to accelerate the crystallization of lysozyme, an appropriate amount of lysozyme crystals can be added as a seed crystal. After standing at low temperature for several days, the lysozyme crystals will slowly precipitate and reach a maximum yield in 72-96 hours. After the crystallization is complete, the supernatant is decanted and the crystals are filtered off using a Buchner funnel to obtain crude lysozyme crystals. (4) Purified lysozyme: The prepared crude crystals were dissolved with acetic acid of pH 4.6, and then the enzyme solution was allowed to stand for 2 hours, followed by filtration to remove insoluble matter. The filtrate was collected and the volume was measured, and the filtrate was added per 100 ml of filtrate. The proportion of 5 g of sodium chloride is added (the method of addition is the same as the method of adding sodium chloride in the second step), and then the pH is slowly adjusted to 10.8 with 1 mol/l sodium hydroxide solution, and the solution is allowed to stand at a low temperature. crystallization. In order to accelerate the crystallization process, lysozyme seed crystals can be added to the enzyme solution, and after the crystallization is completed, the supernatant liquid is decanted, and the lysozyme crystals are obtained by filtration using a Buchner funnel. If the purity is not high, the purification can be repeated until it reaches a certain level. With the required purity, the crystallization product is obtained after drying at 30-40 degrees. Fourth, packaging and storage: products should be installed in glass or ceramic containers, placed in a cool place, so as not to lose activity after lysozyme heat. V. Notes: (1) The entire process can only be carried out in glass or ceramic containers. Metal containers are not available to avoid inactivation of enzymes and affect the yield and quality of products. (2) The whole process of operation should be carried out at a low temperature (below 10°C) to prevent the deterioration of raw materials and inactivation of enzymes. (3) The specific operation of adding the lysozyme seed in the third step is to uniformly suspend the lysozyme crystal in a small amount of pH value of 10.8 and a concentration of 5% sodium chloride solution, and take a few drops to adjust the pH value. In the egg white of sodium chloride concentration, crystallize at a low temperature. (541002 Guilin Gaotouling Air Compressor Factory Gao Guijun)
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