Extraction method of soybean trypsin inhibitor
A technology of trypsin inhibition and extraction method, which is applied in the extraction field of soybean trypsin inhibitor, and achieves the effects of simple steps, improved quality and easy operation.
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Embodiment 1
[0033] A method for extracting soybean trypsin inhibitor, comprising the following steps:
[0034] (1) preparing and extracting soybean trypsin inhibitor primary supernatant;
[0035] The concrete preparation steps of this primary supernatant are:
[0036] (1.1) Take dry soybeans and add water to soak them overnight, then grind them and fully mix them into a slurry;
[0037] (1.2) After stirring the slurry obtained in step (1.1), add the solution and filter it with more than 3 layers of gauze to obtain filtrate and bean dregs;
[0038] The water in step (1.1) is deionized water at 50°C, and the weight ratio of deionized water to soybean is 5:3; the sodium citrate in the mixed solution of step (1.2) is 10mM, and trisodium phosphate is 10mM. And the weight ratio of sodium citrate and trisodium phosphate is 1:1.
[0039] (1.3) Sodium citrate and trisodium phosphate are added in the filtrate obtained by step (1.2) in a ratio of 2:1.5 until the pH value of the filtrate reaches 4...
Embodiment 2
[0056] This embodiment is basically the same as Embodiment 1, the difference is:
[0057] The weight ratio of deionized water to soybeans selected in step (1.1) is 5:3.
[0058] The sodium citrate in step (1.2) is 70mM, and trisodium phosphate is 70mM.
[0059] Step (2.1) When ammonium sulfate is added to the primary supernatant, the primary supernatant needs to be heated to 22° C. and stirred for 3 hours.
[0060] The stirring in step (2.3) needs to be stirred for 3 hours under the environment of 4°C, and then stirred at the speed of 3000r / min for 10min under the environment of 4°C.
[0061] The EDTA used in the step (3.1) is 0.006mM EDTA.
[0062] Step (3.2) When the dialysate is purified by affinity chromatography, first take 3g of the dialysate and dissolve it in 100ml, 3.5M, and pH value of 6.5 phosphate buffer, slowly absorb for 2 hours, and add a concentration of 5% 50ml of pentylene glycol was stirred evenly.
[0063] The elution buffer in step (3.3) is 100 ml of c...
Embodiment 3
[0065] This embodiment is basically the same as Embodiment 2, and its difference is:
[0066] The weight ratio of deionized water to soybeans selected in step (1.1) is 7:3.
[0067] The sodium citrate in the step (1.2) is 100 mM, and the trisodium phosphate is 100 mM.
[0068] Step (2.1) When ammonium sulfate is added to the primary supernatant, the primary supernatant needs to be heated to 25° C. and stirred for 4 hours.
[0069] The stirring in step (2.3) needs to be stirred for 4 hours under the environment of 4°C, and then stirred at the speed of 8000r / min for 10min under the environment of 4°C.
[0070] The EDTA used in the step (3.1) is 0.01 mM EDTA.
[0071] Step (3.2) When the dialysate is purified by affinity chromatography, first take 5 g of the dialysate and dissolve it in 100 ml of 0.5 M phosphate buffer with a pH value of 7.0, slowly absorb for 3 hours, and then add a concentration of 3% 30ml of pentylene glycol was stirred evenly.
[0072] The elution buffer ...
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