CN101407804A - Preparation of mixed enzyme of molecular sieve immobilized barley beta-amylase and Pullulanase - Google Patents
Preparation of mixed enzyme of molecular sieve immobilized barley beta-amylase and Pullulanase Download PDFInfo
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- CN101407804A CN101407804A CNA2008101415641A CN200810141564A CN101407804A CN 101407804 A CN101407804 A CN 101407804A CN A2008101415641 A CNA2008101415641 A CN A2008101415641A CN 200810141564 A CN200810141564 A CN 200810141564A CN 101407804 A CN101407804 A CN 101407804A
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- molecular sieve
- pullulanase
- amylase
- immobilized
- enzyme
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Abstract
The invention discloses a preparation method for two mixed enzymes of a molecular sieve immobilized barley Beta-amylase and a Pullulanase. The existing immobilized enzyme technology has the defects of complexity, higher cost, low yield and short service life. In the invention, a macroporous molecular sieve is used as a vector and the methods of absorbing the enzymes for a plurality of times as well as crosslinking glutaric dialdehyde for a plurality of times are adopted for preparing the immobilized barley Beta-amylase and the Pullulanase; after the prepared immobilized barley Beta-amylase and the Pullulanase are packaged in column, a cornstarch liquefied liquid of 20 percent is continuously supplied for 18 days under the temperature of 55 DEG C, and then a saccharified liquid with a DE value of more than 58 percent and a maltose content of more than 72.5 percent can be generated; and after 25 days, the DE value can be still as high as 52 percent and the maltose content is about 60 percent. The vector used by the invention is easy to be shaped; and the preparation method has the advantages of simple preparing technique of the immobilized enzyme, gentle condition, a plurality of times of absorbing and crosslinking the enzyme, better mechanical property, stable operation, being capable of repeated use and being suitable for a filling bed reactor. The method can also immobilize other matters with catalyzing activity.
Description
Technical field
The present invention relates to two kinds of mixed enzyme preparation methods of a kind of molecular sieve immobilized barley beta-amylase and Pullulanase, especially relate to a kind of molded molecular sieve immobilized barley beta-amylase and two kinds of mixed enzyme preparation methods of Pullulanase of utilizing.
Background technology
China is from the nineties in last century, because production technology is improved, scale enlarges, and cost descends, Dian Fentang production develop rapidly, and along with China's living standards of the people improve, the demand of Dian Fentang also can increase year by year from now on, and quality standard requires higher.Dian Fentang is mainly used in foodstuffs industry, medicine and chemical industry etc.A lot of to the research of fixation glucose amylase both at home and abroad.The report of external existing scale operation.At solid, the particularly existing a large amount of report (I.CHIBATA of the method for immobilized enzyme on solids, immobilized enzyme, chapter 2, p.9-107, Wiley and Sons, 1978) .Tate and Lyle company as carrier, adopt acetone precipitation with bone black, the mode of glutaraldehyde cross-linking prepares immobilized enzyme, has been applied in the american corn source mill.Be reported in the method for immobilized enzyme on the polymeric film according to patent of invention 01139257.6, this method technical sophistication, cost is higher, the immobilization difficulty.Zhao's honor etc. has been studied the zeolite immobilized enzyme that amination is handled, 45 ℃ of reactions behind the dress post, and calculating the transformation period is 38 days.But these research enzyme immobilization technology complexity, cost is higher, and yield is low, and the life-span is short, does not fit into continuous production, and suitability for industrialized production is restricted.
Summary of the invention
The objective of the invention is to adopt the molecular sieve of moulding is carrier, and it is carrier immobilized barley beta-amylase and two kinds of mixed enzyme preparation methods of Pullulanase that a kind of molded molecular sieve that utilizes is provided.It can make the carrier moulding easy, and immobilized enzyme preparation technology is simple, mild condition, and better mechanical property, stable operation, and be applicable to packed bed reactor and other material that fixedly has catalytic activity.
In order to achieve the above object, technical scheme of the present invention is: a kind of molecular sieve is carrier immobilized barley beta-amylase and two kinds of mixed enzyme of Pullulanase (Novi's letter) and preparation method thereof, it is characterized in that: specifically may further comprise the steps:
(1) Al of adding about 20% in the molecular sieve carrier
2O
3Binding agent, the mixed pressure moulding, drying at room temperature, roasting is driven away and is broken into required granular size;
(2) molecular sieve bore diameter scope 1-25nm;
(3) two kinds of enzyme ratios of barley beta-amylase and Pullulanase are mixed at 1: 2;
(4) carrier is adorned post, put into some broken porcelain at the bottom of the post, put some glass yarn above, post is thermal insulation layer in addition, 25-55 ℃ fed barley beta-amylase and two kinds of mixed enzyme solution absorbs of Pullulanase 5-10 hour down, 25-30 ℃, with the crosslinked 3-5 of glutaraldehyde solution hour of 1%-4%, hang repeatedly that enzyme is repeatedly crosslinked to get final product under the pH4.5-7 condition.
5) immobilized enzyme that will repeatedly use inactivation adsorbs through 500-650 ℃ of roasting, and crosslinked the continuation used.
Described molecular sieve is with USY, and DAY and beta-zeolite are raw material, and molecular sieve is a multi-hole type, and the molecular sieve bore diameter scope is 1-25nm, drives broken even particle size away before the molecular sieve dress post.
Two kinds of enzyme ratios of described barley beta-amylase and Pullulanase are mixed at 1: 2, and mix.
Description of drawings
Accompanying drawing 1 is a process flow sheet of the present invention
The invention has the beneficial effects as follows:
Immobilized enzyme preparation technology is simple, mild condition, and it is repeatedly crosslinked repeatedly to hang enzyme, better mechanical property, stable operation can use repeatedly, and is applicable to packed bed reactor.
Embodiment:
Embodiment: the present invention utilizes molded molecular sieve immobilized barley beta-amylase and two kinds of mixed enzyme preparation methods of Pullulanase, specifically may further comprise the steps:
(1) making of molecular sieve:
Molecular sieve is starting material with the beta-zeolite, adds about 20% Al
2O
3Binding agent, the mixed pressure moulding, drying at room temperature, roasting is driven away and is broken into particulate state, the 20-40 order that sieves, even particle size.
(2) preparation of saccharification post
Put into some broken porcelain in glass column, put some glass yarn above, in the post of then molecular sieve of moulding being packed into, the glass column outside adds a thermal insulation layer and gets final product.
(3) hang enzyme for the first time
Get two kinds of enzymes of barley beta-amylase and Pullulanase (Novi's letter), mix at 1: 2 by mass ratio, get quantitative mixed enzyme, under 25-30 ℃ with volume pump circulate 5-10 hour (pressure and flow are determined according to column volume and post are high, but are hung enzyme time proper extension).With distillation washing enzyme, wash water and remaining enzyme are merged, measure enzyme activity, detect and hang the enzyme effect.
(4) for the first time crosslinked
Get the glutaraldehyde solution of quantitative 1%-4%,, under the pH4.5-7 condition circulation 3-5 hour, unnecessary glutaraldehyde solution is washed with distilled water at 25-30 ℃.
(5) it is repeatedly crosslinked repeatedly to hang enzyme
It is for the second time crosslinked to carry out hanging the second time enzyme according to step (3) and (4), so repeats 3-5 time, can obtain comparatively stable immobilized enzyme.
Claims (3)
1, two kinds of mixed enzyme preparation methods of a kind of molecular sieve immobilized barley beta-amylase and Pullulanase is characterized in that: specifically may further comprise the steps:
(1) Al of adding about 20% in the molecular sieve carrier
2O
3Binding agent, the mixed pressure moulding, drying at room temperature, roasting is driven away and is broken into required granular size;
(2) molecular sieve bore diameter scope 1-25nm;
(3) two kinds of enzyme ratios of barley beta-amylase and Pullulanase are mixed at 1: 2;
(4) carrier is adorned post, put into some broken porcelain at the bottom of the post, put some glass yarn above, post is thermal insulation layer in addition, 25-55 ℃ fed barley beta-amylase and two kinds of mixed enzyme solution absorbs of Pullulanase 5-10 hour down, 25-30 ℃, with the crosslinked 3-5 of glutaraldehyde solution hour of 1%-4%, hang repeatedly that enzyme is repeatedly crosslinked to get final product under the pH4.5-7 condition.
(5) immobilized enzyme that will repeatedly use inactivation adsorbs through 500-650 ℃ of roasting, and crosslinked the continuation used.
2, a kind of molecular sieve immobilized barley beta-amylase according to claim 1 and two kinds of mixed enzyme preparation methods of Pullulanase, it is characterized in that: described molecular sieve is with USY, DAY and beta-zeolite are raw material, molecular sieve is a multi-hole type, the molecular sieve bore diameter scope is 1-25nm, drives broken even particle size away before the molecular sieve dress post.
3, a kind of molecular sieve immobilized barley beta-amylase according to claim 1 and two kinds of mixed enzyme preparation methods of Pullulanase is characterized in that: two kinds of enzyme ratios of described barley beta-amylase and Pullulanase are mixed at 1: 2, and mix.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNA2008101415641A CN101407804A (en) | 2008-10-08 | 2008-10-08 | Preparation of mixed enzyme of molecular sieve immobilized barley beta-amylase and Pullulanase |
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| CNA2008101415641A CN101407804A (en) | 2008-10-08 | 2008-10-08 | Preparation of mixed enzyme of molecular sieve immobilized barley beta-amylase and Pullulanase |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106520734A (en) * | 2016-11-28 | 2017-03-22 | 山东正德食品有限公司 | Method for improving heat resistance of pullulanase for catalyzing maltosyl-beta-cyclodextrin synthesis |
| CN107012138A (en) * | 2017-05-03 | 2017-08-04 | 福州晨翔环保工程有限公司 | Silicon/carbon-based complex solidifying enzyme environment-friendly materials and preparation method thereof |
| CN107574200A (en) * | 2017-10-31 | 2018-01-12 | 无锡甜丰食品有限公司 | A kind of production technology for preparing high maltose syrup |
-
2008
- 2008-10-08 CN CNA2008101415641A patent/CN101407804A/en active Pending
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106520734A (en) * | 2016-11-28 | 2017-03-22 | 山东正德食品有限公司 | Method for improving heat resistance of pullulanase for catalyzing maltosyl-beta-cyclodextrin synthesis |
| CN107012138A (en) * | 2017-05-03 | 2017-08-04 | 福州晨翔环保工程有限公司 | Silicon/carbon-based complex solidifying enzyme environment-friendly materials and preparation method thereof |
| CN107012138B (en) * | 2017-05-03 | 2020-04-10 | 福州晨翔环保工程有限公司 | Silicon/carbon-based composite immobilized enzyme environment-friendly material and preparation method thereof |
| CN107574200A (en) * | 2017-10-31 | 2018-01-12 | 无锡甜丰食品有限公司 | A kind of production technology for preparing high maltose syrup |
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Open date: 20090415 |