CN103088006A - Method for extracting protease from squeezed juice of sisal - Google Patents
Method for extracting protease from squeezed juice of sisal Download PDFInfo
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- CN103088006A CN103088006A CN2013100077846A CN201310007784A CN103088006A CN 103088006 A CN103088006 A CN 103088006A CN 2013100077846 A CN2013100077846 A CN 2013100077846A CN 201310007784 A CN201310007784 A CN 201310007784A CN 103088006 A CN103088006 A CN 103088006A
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Abstract
The invention discloses a method for extracting protease from squeezed juice of sisal. The method for extracting protease from squeezed juice of sisal disclosed by the invention comprises the following steps: squeezing juice by squeezing fresh sisal residue to obtain squeezed juice of sisal; filtering by filtering the squeezed juice of sisal and removing coarse residue to obtain a filter liquor without the coarse residue; centrifuging by centrifuging a crude enzyme liquor without coarse residue to obtain a transparent crude enzyme liquor; ultra-filtering and concentrating by ultra-filtering and concentrating the crude enzyme liquor to obtain a concentrated enzyme liquor; precipitating via ammonium sulfate by adding the ammonium sulfate into the concentrated enzyme liquor, salting out the enzyme and precipitating to obtain an enzyme precipitate; centrifuging by centrifuging the enzyme precipitate and collecting a pasty enzyme paste; drying and crushing by drying the enzyme paste and crushing to obtain powdery finished products of protease. The sisal protease extracted by the method for extracting protease from squeezed juice of sisal disclosed by the invention has the advantages of good quality and high purity. The method for extracting protease from squeezed juice of sisal disclosed by the invention has the advantages of simple process, low production cost, high recovery rate, energy conservation, environmental friendliness and the like.
Description
Technical field
The present invention relates to the proteins extraction purification technique, is that a kind of squeezing the juice from sisal hemp extracted the method for proteolytic enzyme specifically.
Background technology
Agavain is a kind of thiol proteinase, enzyme activity is high, solvability is good, thermostability is high, the zymin finished product is white or milk yellow powder, under the deepfreeze condition, and good stability, inner enzyme vigor reduction in 1 year is less than 10%, is widely used in the industries such as food, medicine, makeup, weaving, feed, leather.
The methods such as the extracting and purifying method of protein mainly contains and saltouts, organic solvent deposit, isoelectric precipitation, ion exchange chromatography, tannin precipitation, affinity chromatography, ultrafiltration and concentration.But, use separately the wherein a kind of technology in aforesaid method, the effect of extracting the purifying agavain is also bad.At first the present invention will remove the sedimentary sisal hemp ultrafiltration and concentration of squeezing the juice, and then add ammonium sulfate to precipitate wherein agavain, can remove a part of micromolecular protein on the one hand in ultra-filtration process, can reduce in a large number on the other hand the consumption of ammonium sulfate, reduced production cost, also reduced environmental pollution, the vigor of agavain finished product is high, quality good.
China sisal hemp main producing region in Guangxi, the areas such as Guangdong and Hainan, the sisal hemp cultivated area in Guangxi is maximum at present.Sisal hemp has a large amount of sisal dregs after sisal fibers is extracted in processing, also be not developed at present.Abundant sisal hemp raw material resources are for industrialization of the present invention provides solid raw material guarantee.
Summary of the invention
At first the present invention will remove the sedimentary sisal hemp ultrafiltration and concentration of squeezing the juice, and then add ammonium sulfate to precipitate wherein agavain, can remove the little protein of molecular weight when ultrafiltration, can also reduce in a large number the consumptions of ammonium sulfate, reduced cost, reduced environmental pollution, the vigor of agavain finished product is high, quality good.
The present invention is achieved by the following technical programs:
A kind of squeezing the juice from sisal hemp extracted the method for agavain, comprises the steps:
(1) squeeze the juice, with fresh sisal dregs squeezing, obtain sisal hemp and squeeze the juice;
(2) filter, with the sisal hemp filtration of squeezing the juice, remove thick slag, obtain removing the filtrate of thick slag;
(3) centrifugal, the crude enzyme liquid of thick slag is centrifugal with going, and obtains transparent crude enzyme liquid;
(4) ultrafiltration and concentration with transparent crude enzyme liquid ultrafiltration and concentration, obtains concentrated enzyme liquid;
(5) ammonium sulfate precipitation adds ammonium sulfate in concentrated enzyme liquid, make enzyme saltout, precipitate and obtain the enzyme throw out;
(6) centrifugal, the enzyme throw out is carried out the centrifugal enzyme cream of collecting paste;
(7) dry and pulverizing, dry with enzyme cream, pulverizes, and gets the proteolytic enzyme finished product of powdery.
Preferably, described fresh sisal dregs is the fresh sisal dregs after fiber.
Preferably, the filtration that is filtered into employing 200 order filter clothes of described step (2).
Preferably, the centrifugal of described step (3) is centrifugal with the high-speed dish piece whizzer, and rotating speed is 5000 rev/mins～8000 rev/mins.
Preferably, the ultrafiltration and concentration of described step (4) uses 6000～20000 daltonian ultra-filtration membranes, and operating pressure is positive 0.03MPa～0.1MPa, and temperature is 15 ℃～30 ℃, and concentration ratio is 10 times.
Preferably, the ammonium sulfate concentrations of described step (5) is 20%～50%.
Preferably, described step (6) centrifugal, rotating speed is 3000 rev/mins～7000 rev/mins.
Preferably, the drying of described step (7) is lyophilize, and temperature is-10 ℃～-50 ℃.
Beneficial effect of the present invention is: at the bottom of less energy consumption, cost, technique is simple, and the rate of recovery is high, energy-conserving and environment-protective.The purity of the agavain finished product that extracts is high, and quality is good.
Embodiment
The present invention is described in further detail below in conjunction with embodiment, but embodiments of the present invention are not limited to the scope that embodiment represents.
Embodiment 1
Fresh sisal dregs is squeezed, obtain the sisal hemp that proteinase activity is about 3000 unit of activity/milliliters and squeeze the juice; With 200
The order filter cloth is removed thick impurity wherein, the filtrate that obtains removing thick slag with the sisal hemp filtration of squeezing the juice; With disk plate centrifuge, that filtrate is centrifugal, remove the trickle impurity of particle wherein, obtain transparent crude enzyme liquid; Ultrafiltration and concentration uses 10000 daltonian ultra-filtration membranes, is 0.05MPa in operating pressure, and temperature is under 15 ℃ of conditions, and to 10% of original volume, obtaining proteinase activity is the concentrated enzyme liquid of 22000 unit of activity/milliliters with transparent crude enzyme liquid ultrafiltration and concentration; Add ammonium sulfate to final concentration 50% in concentrated enzyme liquid, make enzyme saltout, precipitate; Enzyme cream with the centrifugal collection paste of atresia tripod pendulum type batch centrifugal; With the lyophilize of enzyme cream, then the dry enzyme piece of gained is pulverized, get the proteolytic enzyme finished product of powdery, enzyme activity reaches 1,510,000 units/gram.
Embodiment 2
Fresh sisal dregs is squeezed, obtain the sisal hemp that proteinase activity is about 3500 unit of activity/milliliters and squeeze the juice; With the sisal hemp filtration of squeezing the juice, remove thick impurity wherein, the filtrate that obtains removing thick slag with 200 order filter clothes; With disk plate centrifuge, that filtrate is centrifugal, remove the trickle impurity of particle wherein, obtain transparent crude enzyme liquid; Ultrafiltration and concentration uses 20000 daltonian ultra-filtration membranes, is 0.08MPa in operating pressure, and temperature is under 17 ℃ of conditions, and to 8% of original volume, obtaining proteinase activity is the concentrated enzyme liquid of 31000 unit of activity/milliliters with transparent crude enzyme liquid ultrafiltration and concentration; Add ammonium sulfate to final concentration 20% in concentrated enzyme liquid, centrifugal with disk plate centrifuge, remove precipitation (impurity); Add ammonium sulfate to final concentration 46% in its enzyme liquid, make enzyme saltout, precipitate; Enzyme cream with the centrifugal collection paste of atresia tripod pendulum type batch centrifugal; With the lyophilize of enzyme cream, then the dry enzyme piece of gained is pulverized, get the proteolytic enzyme finished product of powdery, enzyme activity reaches 1,830,000 units/gram.
Embodiment 3
Fresh sisal dregs is squeezed, obtain the sisal hemp that proteinase activity is about 3100 unit of activity/milliliters and squeeze the juice; With the sisal hemp filtration of squeezing the juice, remove thick impurity wherein, the filtrate that obtains removing thick slag with 200 order filter clothes; With disk plate centrifuge, that filtrate is centrifugal, remove the trickle impurity of particle wherein, obtain transparent crude enzyme liquid; Ultrafiltration and concentration uses 20000 daltonian ultra-filtration membranes, is being 0.10MPa as pressure mutually, and temperature is under 22 ℃ of conditions, and to 9% of original volume, obtaining proteinase activity is the concentrated enzyme liquid of 26000 unit of activity/milliliters with transparent crude enzyme liquid ultrafiltration and concentration; Add ammonium sulfate to final concentration 25% in concentrated enzyme liquid, centrifugal with disk plate centrifuge, remove precipitation (impurity); Add ammonium sulfate to final concentration 45% in its enzyme liquid, make enzyme saltout, precipitate; Enzyme cream with the centrifugal collection paste of atresia tripod pendulum type batch centrifugal; With the lyophilize of enzyme cream, then the dry enzyme piece of gained is pulverized, get the proteolytic enzyme finished product of powdery, enzyme activity reaches 1,680,000 units/gram.
Although embodiment of the present invention are open as above, but it is not restricted to listed utilization in specification sheets and embodiment, it can be applied to various suitable the field of the invention fully, for those skilled in the art, can easily realize other modification, therefore do not deviating under the universal that claim and equivalency range limit, the present invention is not limited to specific details and illustrates here and the embodiment that describes.
Claims (8)
1. squeeze the juice from sisal hemp and extract the method for agavain for one kind, it is characterized in that comprising the steps:
(1) squeeze the juice, with fresh sisal dregs squeezing, obtain sisal hemp and squeeze the juice;
(2) filter, with the sisal hemp filtration of squeezing the juice, remove thick slag, obtain removing the filtrate of thick slag;
(3) centrifugal, the crude enzyme liquid of thick slag is centrifugal with going, and obtains transparent crude enzyme liquid;
(4) ultrafiltration and concentration with transparent crude enzyme liquid ultrafiltration and concentration, obtains concentrated enzyme liquid;
(5) ammonium sulfate precipitation adds ammonium sulfate in concentrated enzyme liquid, make enzyme saltout, precipitate and obtain the enzyme throw out;
(6) centrifugal, the enzyme throw out is carried out the centrifugal enzyme cream of collecting paste;
(7) dry and pulverizing, dry with enzyme cream, pulverizes, and gets the proteolytic enzyme finished product of powdery.
2. according to claim 1 squeezing the juice from sisal hemp extracted the method for agavain, it is characterized in that, described fresh sisal dregs is the fresh sisal dregs after fiber.
3. according to claim 1 squeezing the juice from sisal hemp extracted the method for agavain, it is characterized in that the filtration that is filtered into employing 200 order filter clothes of described step (2).
4. according to claim 1 squeezing the juice from sisal hemp extracted the method for agavain, it is characterized in that, the centrifugal of described step (3) is centrifugal with the high-speed dish piece whizzer, and rotating speed is 5000 rev/mins～8000 rev/mins.
5. according to claim 1 squeezing the juice from sisal hemp extracted the method for agavain, it is characterized in that the ultrafiltration and concentration of described step (4), use 6000～20000 daltonian ultra-filtration membranes, operating pressure is positive 0.03MPa～0.1MPa, and temperature is 15 ℃～30 ℃, and concentration ratio is 10 times.
6. according to claim 1 squeezing the juice from sisal hemp extracted the method for agavain, it is characterized in that, the ammonium sulfate concentrations of described step (5) is 20%～50%.
7. according to claim 1 squeezing the juice from sisal hemp extracted the method for agavain, it is characterized in that, and described step (6) centrifugal, rotating speed is 3000 rev/mins～7000 rev/mins.
8. according to claim 1 squeezing the juice from sisal hemp extracted the method for agavain, it is characterized in that, the drying of described step (7) is lyophilize, and temperature is-10 ℃～-50 ℃.
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| CN2013100077846A CN103088006A (en) | 2013-01-09 | 2013-01-09 | Method for extracting protease from squeezed juice of sisal |
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| CN2013100077846A CN103088006A (en) | 2013-01-09 | 2013-01-09 | Method for extracting protease from squeezed juice of sisal |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109503120A (en) * | 2018-12-27 | 2019-03-22 | 广州华隆建筑材料有限公司 | A kind of acoustic brick and preparation method thereof of high intensity |
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| CN1766099A (en) * | 2004-10-31 | 2006-05-03 | 广西师范大学 | Grosvenor momordica proteinase and its extraction method |
| CN101319208A (en) * | 2008-04-08 | 2008-12-10 | 徐闻县美仑生物制品有限公司 | Method for preparation of bromelin |
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2013
- 2013-01-09 CN CN2013100077846A patent/CN103088006A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1766099A (en) * | 2004-10-31 | 2006-05-03 | 广西师范大学 | Grosvenor momordica proteinase and its extraction method |
| CN101319208A (en) * | 2008-04-08 | 2008-12-10 | 徐闻县美仑生物制品有限公司 | Method for preparation of bromelin |
Non-Patent Citations (8)
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| 《华南农业大学学报》 19930402 徐凤彩, 等 "东一号"剑麻蛋白酶分离纯化及特性研究 第14卷, 第1期 * |
| 《科学通报》 19800215 孙崇荣, 等 剑麻蛋白酶--一种新发现的巯基蛋白酶 , 第3期 * |
| 孙崇荣, 等: "剑麻蛋白酶——一种新发现的巯基蛋白酶", 《科学通报》, no. 3, 15 February 1980 (1980-02-15) * |
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| 徐凤彩, 等: ""东一号"剑麻蛋白酶分离纯化及特性研究", 《华南农业大学学报》, vol. 14, no. 1, 2 April 1993 (1993-04-02) * |
| 沈家柏, 等: "灰叶剑麻叶片中一种蛋白酶的分离纯化", 《热带作物学报》, vol. 4, no. 1, 2 July 1983 (1983-07-02) * |
| 苏小建, 等: "罗汉蛋白酶的分离纯化和特性研究", 《生物工程》, vol. 28, no. 5, 15 May 2007 (2007-05-15) * |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109503120A (en) * | 2018-12-27 | 2019-03-22 | 广州华隆建筑材料有限公司 | A kind of acoustic brick and preparation method thereof of high intensity |
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Application publication date: 20130508 |