Method for pectic enzyme separating purification by double aqueous phase extraction system

A technology for separation and purification, pectinase, applied in the field of separation and purification of protein biological activity target products, can solve the problems of enzyme inactivation, difficult to meet food-grade quality requirements, low activity, etc., to optimize the preparation process and shorten the operation period , the effect of improving yield

Inactive Publication Date: 2007-10-31
SHAANXI UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] At present, most of the enzyme industry adopts the salting-out method. This method can only obtain crude enzyme liquid containing bacteria, salts, miscellaneous proteins and other solids. This kind of enzyme is difficult to meet the quality requirements of food grade.
In addition, the direct contact between the organic solvent and the pectinase during the operation process of the current pectinase extraction method will easily lead to the inactivation of the enzyme, which is also one of the main reasons for the low activity of domestic pectinase

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0011] Example 1, pretreatment of fermented liquid: Since the pectinase produced by Aspergillus niger is an extracellular enzyme, the method of filtration is firstly used for rough extraction; at 20°C, polyethylene glycol, ammonium sulfate and water are mixed to prepare Mixed solution, wherein the mass percentage of polyethylene glycol is 27%, the mass percentage of ammonium sulfate is 19%, and the rest is water; the mixed solution is oscillated and shaken, and then 10% of the mixed solution is added after standing for phase separation Pectinase solution, shake well, stand at room temperature for 5-10 minutes, extract and separate the phases, measure the enzyme activity in the upper and lower phases respectively after the phase separation is complete, at this time, the distribution coefficient K of pectinase can reach the maximum Value K=11.42; Most of the pectinase enters the upper phase polyethylene glycol phase, and the upper phase liquid polyethylene glycol is separated for...

Embodiment 2

[0012] Embodiment 2, the pretreatment of fermented liquid: because the pectinase produced by Aspergillus niger is an extracellular enzyme, first adopt the method of filtration to carry out rough extraction; At 20 ℃, polyethylene glycol, ammonium sulfate and water are mixed to prepare Mixed solution, wherein the mass percentage of polyethylene glycol is 25%, the mass percentage of ammonium sulfate is 15%, and the rest is water; the mixed solution is oscillated and shaken, and then 10% of the mixed solution is added after standing for phase separation Pectinase solution, shake well, extract and phase-separate after standing at room temperature for 5-10 minutes, measure the enzyme activity in the upper and lower phases respectively after the phase separation is complete, at this time, the distribution coefficient of pectinase K=2.15, Most of the pectinase enters the upper phase polyethylene glycol phase, and the upper phase liquid polyethylene glycol is separated for later use; at...

Embodiment 3

[0013] Example 3, pretreatment of fermented liquid: since the pectinase produced by Aspergillus niger is an extracellular enzyme, the method of filtration is firstly used for crude extraction; at 20°C, polyethylene glycol, ammonium sulfate and water are mixed to prepare Mixed solution, wherein the mass percentage of polyethylene glycol is 27%, the mass percentage of ammonium sulfate is 13%, and the rest is water; the mixed solution is oscillated and shaken, and then 10% of the mixed solution is added after standing for phase separation Pectinase solution, shake well, stand at room temperature for 5-10 minutes and then carry out extraction and phase separation. After the phase separation is complete, respectively measure the enzyme activity in the upper and lower phases. At this time, the distribution coefficient of pectinase K=6.67, Most of the pectinase enters the upper phase polyethylene glycol phase, and the upper phase liquid polyethylene glycol is separated for later use; ...

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PUM

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Abstract

The invention discloses a method to separate and purify pectic enzyme with double water phase extractive system, which comprises the following steps: mixing carbowax, ammonia sulfate and water; producing mixed solution; shaking the mixed solution evenly; stewing; phase-splitting; adding into pectic enzyme liquid; shaking evenly; extracting and phase-splitting at room temperature; separating upper phase liquid carbowax to reserve; mixing with potassium sodium and water; back-extracting at room temperature; separating lower phase; filtering with film; getting pectic enzyme. This invention can optimize preparing craft, shorten operating period and increase receiving ratio of the product.

Description

technical field [0001] The invention relates to a technology for separating and purifying protein bioactive target products, in particular to a method for separating and purifying pectinase in a two-phase extraction system. Background technique [0002] The pharmacological effects of biological products, especially protein biomacromolecules, are closely related to their physiological activities. Therefore, in the separation and purification process, the extraction operation must be carried out under the premise of maintaining its biological activity according to the characteristics of the target product. The aqueous two-phase extraction system is a new type of separation technology developed based on the liquid-liquid extraction theory while considering the maintenance of biological activity in consideration of this situation. [0003] Aqueous two-phase extraction technology is an effective method for separating and purifying biological macromolecules such as protein mixtur...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/24
Inventor 杨辉张娟王旭
Owner SHAANXI UNIV OF SCI & TECH
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