Preparation method of catfish visceral protease
A protease and fish viscera technology is applied in the field of preparation of catfish viscera protease, which can solve the problems of inconvenient operation, low protease purity, low efficiency, etc., and achieves the effects of enhancing disease prevention, improving lymphocyte activity and high purity
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Embodiment 1
[0014] The preparation method of catfish viscera protease comprises leaching, salting out, dialysis and purification, and the specific steps are:
[0015] 1) Extraction: After homogenizing catfish viscera with a homogenizer, add buffer to extract for 3~5 hours, and centrifuge the extract at a speed of 8000~9000r / min for 10~15min, and the obtained supernatant is the crude catfish viscera extract Disperse and grind the catfish visceral tissue into a uniform paste through a homogenizer, so that it can fully contact with the buffer, thereby fully releasing the cell contents, leaching and centrifuging, so that the solid particles and The liquid containing the cell contents is separated to obtain the catfish crude extract;
[0016] 2) Salting out: After adding saturated ammonium sulfate to the crude catfish viscera extract for precipitation, the weight-to-volume ratio of the crude catfish viscera extract to saturated ammonium sulfate is 1g:1.3~1.6ml. Centrifuge at 10000r / min for 12...
Embodiment 2
[0021] The preparation method of catfish viscera protease comprises the following steps:
[0022] 1) Extraction: After homogenizing catfish viscera with a homogenizer, add buffer to extract for 3~5 hours, and centrifuge the extract at a speed of 8000~9000r / min for 10~15min, and the obtained supernatant is the crude catfish viscera extract ;
[0023] 2) Salting out: After adding saturated ammonium sulfate to the crude catfish viscera extract for precipitation, the weight-to-volume ratio of the crude catfish viscera extract to saturated ammonium sulfate is 1g:1.6ml, centrifuge at 5°C and 10,000 r / min 13min;
[0024] 3) Dialysis: Dissolve the precipitate in phosphate buffer, dialyze in distilled water at 5°C for 30 hours, and change the distilled water every 7 hours;
[0025] 4) Purification: The dialysate was prepared by hydroxyapatite adsorption chromatography to prepare catfish visceral pure protease. The column specifications were 2 cm in diameter and 52 cm in length. The m...
Embodiment 3
[0028] The preparation method of catfish viscera protease comprises the following steps:
[0029] 1) Extraction: After the homogenizer is homogenized, add the same amount of buffer as before to extract for 4 hours, and centrifuge the extract at a speed of 9000r / min for 13 minutes, and the obtained supernatant is the crude catfish viscera extract.
[0030] 2) Salting out: After adding saturated ammonium sulfate to the crude catfish viscera extract for precipitation, the weight-to-volume ratio of the crude catfish viscera extract to saturated ammonium sulfate is 1g:1.3ml, centrifuge at 4°C and 9000 r / min 12min.
[0031] 3) Dialysis: Dissolve the precipitate in phosphate buffer, dialyze in distilled water at 6°C for 32 hours, and change the distilled water every 8 hours.
[0032] 4) Purification: The dialysate was prepared by hydroxyapatite adsorption chromatography to prepare catfish visceral pure protease. The column specifications were 5 cm in diameter and 60 cm in length. Th...
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