CN105296316A - Complex enzyme preparation as well as preparation method and application thereof - Google Patents
Complex enzyme preparation as well as preparation method and application thereof Download PDFInfo
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- CN105296316A CN105296316A CN201510737914.0A CN201510737914A CN105296316A CN 105296316 A CN105296316 A CN 105296316A CN 201510737914 A CN201510737914 A CN 201510737914A CN 105296316 A CN105296316 A CN 105296316A
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Abstract
The invention provides a complex enzyme preparation which comprises the following components by weight percent: 1-2.5*10<4> U/L papain, 1000-3000U/L bromelain, 0.1-10PPU/L proline incision enzyme, 0.1-1% of sodium benzoate, 0.1-1% of ascorbic acid, 0.1-1% of sodium ethylene diamine tetracetate, 20-50% of glycerinum and the balance of solvent deionized water. The complex enzyme preparation has the advantages of high stability, long guarantee period, long-lasting obvious effect of eliminating cold muddy effect, capability of greatly reducing the dosage of the enzyme preparation and ultrahigh commercial and popularizing values.
Description
Technical field
The present invention relates to compound enzymic preparation technical field, particularly relate to a kind of albumen compound enzymic preparation eliminating cold muddiness, the preparation method of this compound enzymic preparation, and this compound enzymic preparation eliminates the application in cold turbid phenomenon in the manufacture such as beer, nectar and storage process.
Background technology
Beer and fruit juice, as the unstable colloid of one, wherein contain a large amount of albumen and polyphenols.And the albumen of polyphenol and proline rich can form insoluble mixture in long term storage, thus there is cold concrete phenomenon, affect the quality of beer and fruit juice.In order to solve the cold concrete problem of beer, current main technique is in beer, add a certain amount of formaldehyde, eliminates most of polyphenol substance, thus solves this problem.But this technique one to cause beer flavor to decline, two is to make final product for containing formaldehyde beer.The beer that this technique is produced not only affects the quality of beer, the more important thing is and also can be detrimental to health.And common process adopts the Filtration Adsorption media such as silica gel, wilkinite or kaolin in the production of fruit juice, repeatedly filter and remove albumen and polyphenols, thus improve the stability of fruit juice.But this technique can produce a large amount of Filtration Adsorption medium wastes, effective nutritive substances such as a large amount of albumen polyphenol in fruit juice also can be caused to be removed, thus to make taste and the quality degradation of fruit juice.Therefore develop a kind of green non-pollution, and the technique at utmost can preserving the nutrition of beer, fruit juice etc. is very necessary.
Bio protease, especially conjugated protein zymin, can eliminate cold concrete phenomenon as one, and the method that can improve again beer good flavor becomes study hotspot.In patent CN201410493227, develop a kind of composite pawpaw glycoprotein proteolytic enzyme as Beer Clarifier, achieve good effect, but in clarity, still have huge raising space.Experimental study also finds that simple can not solve the cold concrete problem in appearance after long-term storage such as beer, fruit juice with papoid and bromeline simultaneously.This mainly because papoid and bromeline are prozyme, almost all has certain hydrolysis effect to all albumen.But, can not directly excise albumen with proline residue, thus cause the albumen that these can not be eliminated proline residue in the process of long storage periods still can form insoluble mixture with polyphenol, occur cold concrete phenomenon.Therefore, exploitation one can remove most of protein efficiently, again can specificity eliminate albumen compound enzymic preparation with proline residue there is huge marketable value.
Summary of the invention
Technical problem to be solved by this invention is to provide a kind of compound enzymic preparation, this compound enzymic preparation good stability, long quality-guarantee period, eliminates cold muddy effect obvious lastingly, and can significantly reduce zymin dosage, have high business and promotional value.
The technical solution adopted in the present invention is:
A kind of compound enzymic preparation, said preparation comprises each component of following proportioning: papoid 1-2.5 × 10
4u/ liter, bromeline 1000-3000U/ liter, proline(Pro) restriction endonuclease 0.1-10PPU/ liter, Sodium Benzoate 0.1-1% (percent weight in volume), xitix 0.1-1% (percent weight in volume), sodium ethylene diamine tetracetate 0.1-1% (percent weight in volume), glycerine 20-50% (percent weight in volume, final concentration), solvent is deionized water.
The preparation method of above-mentioned compound enzymic preparation, comprises the following steps:
(1) Accurate Determining bromeline, papoid and proline(Pro) restriction endonuclease enzyme are lived, and get bromeline, papoid, proline(Pro) restriction endonuclease respective amount, add the deionized water accounting for cumulative volume 30-40% and fully dissolve;
(2), after zymin is dissolved completely, add Sodium Benzoate, xitix, sodium ethylene diamine tetracetate, finally add glycerine to final concentration, and add deionized water constant volume to final volume;
(3) pasteurization is carried out to the above-mentioned zymin configured, and preserve in 2-8 DEG C.
The present invention further provides above-mentioned compound enzymic preparation and eliminate application in cold turbid phenomenon at beer, nectar.
Compared with prior art, the present invention has following remarkable advantage and beneficial effect:
(1) compound enzymic preparation of the present invention effectively can eliminate the cold muddiness in drinks and beverage storage process, and the pH scope of application is 2-9.5, and Optimal pH use range is 4-8; Temperature stability is high, and 60 degrees Celsius of constant temperature process 1 hour, enzyme lived loss less than 5%;
(2) compound enzymic preparation of the present invention can not only effectively to be degraded the protein fragments of molecular weight at about 40kD in beer, nectar storage, eliminate cold muddiness, can also efficient degradation high-molecular-weight protein (60-90kD), improve package stability, extend the shelf life, the enzyme amount needed for traditional enzyme method technique can be reduced simultaneously, thus reduce costs, increase economic efficiency;
(3) generally speaking, compound enzymic preparation of the present invention has good stability, long quality-guarantee period, eliminates the lasting significantly advantage of cold muddy effect, and significantly can reduce the advantage of zymin dosage, have high business and promotional value.
Accompanying drawing explanation
Shown in Fig. 1 is the pH stability test result of compound enzymic preparation of the present invention;
Shown in Fig. 2 is the temperature stability test result of compound enzymic preparation of the present invention;
Shown in Fig. 3 is that prozyme of the present invention is eliminating the effect in Formation of Beer Chill Haze;
Shown in Fig. 4 is reach the zymin consumption needed for same clarifying effect (turbidity is lower than 0.2) in beer long storage periods;
Shown in Fig. 5 is the effect of prozyme of the present invention after eliminating fruit juice in muddiness;
Shown in Fig. 6 is reach the required zymin consumption of same clarifying effect (turbidity is lower than 0.3) in fruit juice long storage periods.
Embodiment
Below in conjunction with embodiment, the present invention is further described in detail.Iting is noted that following illustrating is all exemplary, being intended to the invention provides further instruction.Except as otherwise noted, all Science and Technology terms that the present invention uses have the identical meanings usually understood with the technical field of the invention personnel.
In following examples compound enzymic preparation, the extraction of bromeline and papoid carries out according to the method described in patent 2013102350043 that (wherein the raw material of bromeline is pineapple, the raw material of papoid is papaya), the solid polypeptide formulation obtained measures according to GB " SBT10317-1999 protease activity amylograph ", final papoid zymin enzyme work is 40,000 units (U)/milligram, and the work of bromeline enzyme is 3020 units (U)/milligram.
Proline(Pro) restriction endonuclease (PESP) is purchased from DSM company limited.
Embodiment 1:
The present embodiment compound enzymic preparation comprises each component of following proportioning: papoid 15000U/ liter, bromeline 2000U/ liter, proline(Pro) restriction endonuclease 5PPU/ liter, Sodium Benzoate 0.5% (g/L), xitix 0.5% (g/L), sodium ethylene diamine tetracetate 0.5% (g/L), glycerine 35% (g/L, final concentration), solvent is deionized water.
The preparation steps of this compound enzymic preparation is as follows:
1. Accurate Determining bromeline, papoid and proline(Pro) restriction endonuclease enzyme are lived, and get bromeline, papoid, proline(Pro) restriction endonuclease respective amount, the deionized water adding cumulative volume about 30-40% fully dissolves;
2., after zymin is dissolved completely, add preservative sodium benzoate, antioxidants ascorbic acid, stablizer sodium ethylene diamine tetracetate, finally add glycerine (to final concentration 35%), and add deionized water constant volume to final volume;
3. pair above-mentioned zymin configured carries out pasteurization (be heated to 62-65 DEG C, keep 30 minutes), and preserves in 2-8 DEG C.
Embodiment 2:
The present embodiment compound enzymic preparation comprises each component of following proportioning: papoid 12000U/ liter, bromeline 1200U/ liter, proline(Pro) restriction endonuclease 3PPU/ liter, Sodium Benzoate 0.2% (g/L), xitix 0.2% (g/L), sodium ethylene diamine tetracetate 0.2% (g/L), glycerine 50% (g/L, final concentration), solvent is deionized water.
The preparation steps of this compound enzymic preparation is with embodiment 1.
Embodiment 3:
The present embodiment compound enzymic preparation comprises each component of following proportioning: papoid 25000U/ liter, bromeline 2500U/ liter, proline(Pro) restriction endonuclease 7PPU/ liter, Sodium Benzoate 0.8% (g/L), xitix 0.8% (g/L), sodium ethylene diamine tetracetate 0.8% (g/L), glycerine 50% (g/L, final concentration), solvent is deionized water.
The preparation steps of this compound enzymic preparation is with embodiment 1.
Embodiment 4: the pH stability of compound enzymic preparation
At 60 DEG C, this compound enzymic preparation is added 50mMpH be respectively in the potassium phosphate buffer of 2,4,6,8,10 measure enzyme live, carry out enzyme activity determination according to enzyme activity determination method in patent CN2007101502823.Having sphere of action through mensuration compound enzymic preparation of the present invention is pH2-9.5, and Optimal pH use range is 4-8, specifically as shown in Figure 1.
Embodiment 5: the temperature stability of compound enzymic preparation:
This compound enzymic preparation is added the potassium phosphate buffer of 50mMpH4.7, under condition of different temperatures, hatch 1h, measure residual enzyme and live, result is as shown in Figure 2.After hatching 1h under this compound enzymic preparation 60 degrees Celsius after measured, proteinase activity does not almost lose.Therefore this compound enzymic preparation can meet the practical application request in the fields such as beer, grape wine and nectar.
Embodiment 6: the application of compound enzymic preparation in beer storage
Compound enzymic preparation of the present invention is added in the beer sample tentatively filtered through PVPP and diatomite (purchased from China Resources brew-house), addition is 1-20mg/L, not add any zymin beer sample and to add equivalent PESP for contrast, turbidity is measured about zero degrees celsius, measure turbidity once weekly, METHOD FOR CONTINUOUS DETERMINATION 12 weeks, result as shown in Figure 3.Result shows, the beer of this compound enzymic preparation process, highly stable, and turbidity stable maintenance is about 0.1.Relative to blank combination, single proline(Pro) restriction endonuclease treatment group, compound enzymic preparation group can eliminate the cold muddiness in beer storage process better, reaches the effect improving beer shelf-stable.
Meanwhile, under the condition reaching same clarifying effect, investigate the consumption of papoid, bromeline, papain bromelain compound formulation, proline(Pro) restriction endonuclease and compound enzymic preparation involved in the present invention, experimental result as shown in Figure 4.Experiment found at long storage periods within 6 weeks, reach same clarifying effect (turbidity less than 0.2), the compound enzymic preparation amount used that the present invention develops is minimum, for 5mg/L, papoid is 30mg/L, and bromeline is 50mg/L, and proline(Pro) restriction endonuclease is 20mg/L, papoid and bromeline compound formulation (1:1, m/m) are 12mg/L.Therefore, relative to the single use of various zymin, it is less that the compound enzymic preparation that the present invention develops has consumption, the advantage that effect is more stable.
Embodiment 7: the application of compound enzymic preparation in nectar
Compound enzymic preparation of the present invention is added in the samples of juice (laboratory self-control) such as Fresh Juice (apple, orange), addition is 1-20mg/L, not add compound enzymic preparation sample and only to add the sample (addition 1-20mg/L) of proline(Pro) restriction endonuclease for contrast, turbidity is measured about zero degrees celsius, measure turbidity once weekly, METHOD FOR CONTINUOUS DETERMINATION 6 weeks, result as shown in Figure 5.Result shows, this compound enzymic preparation is after fruit juice stores six weeks, and turbidity still can lower than 0.5.Illustrate that this compound enzymic preparation can significantly improve the clarity of fruit juice, be beneficial to production and the storage of fruit juice, there is great application potential.
Meanwhile, under the condition reaching the same clarifying effect of fruit juice, investigate the consumption of papoid, bromeline, papain bromelain compound formulation, proline(Pro) restriction endonuclease and compound enzymic preparation involved in the present invention, experimental result as shown in Figure 6.Experiment found at long storage periods within 6 weeks, reach same clarifying effect (turbidity less than 0.3), the compound enzymic preparation amount used that the present invention develops is minimum, for 20mg/L, papoid is 55mg/L, and bromeline is 80mg/L, and proline(Pro) restriction endonuclease is 35mg/L, papoid and bromeline compound formulation (1:1, m/m) are 50mg/L.Therefore, relative to the single use of various zymin, it is less that the compound enzymic preparation that the present invention develops has consumption, the advantage that effect is more stable.
The material that the embodiment of the present invention relates to, reagent and experimental installation, if no special instructions, be the commercially available prod meeting compound enzymic preparation technical field.
The above, be only the preferred embodiments of the present invention, should be understood that; for those skilled in the art; under the prerequisite not departing from core technology of the present invention, can also make improvements and modifications, these improvements and modifications also should belong to scope of patent protection of the present invention.Any change in the implication suitable with claims of the present invention and scope, all should think to be included in the scope of claims.
Claims (4)
1. a compound enzymic preparation, is characterized in that each component comprising following proportioning: papoid 1-2.5 × 10
4u/ liter, bromeline 1000-3000U/ liter, proline(Pro) restriction endonuclease 0.1-10PPU/ liter, Sodium Benzoate 0.1-1%, xitix 0.1-1%, sodium ethylene diamine tetracetate 0.1-1%, glycerine 20-50%, solvent is deionized water.
2. the preparation method of compound enzymic preparation according to claim 1, is characterized in that comprising the following steps:
(1) Accurate Determining bromeline, papoid and proline(Pro) restriction endonuclease enzyme are lived, and get bromeline, papoid, proline(Pro) restriction endonuclease respective amount, add the deionized water accounting for cumulative volume 30-40% and fully dissolve;
(2), after zymin is dissolved completely, add Sodium Benzoate, xitix, sodium ethylene diamine tetracetate, finally add glycerine to final concentration, and add deionized water constant volume to final volume;
(3) pasteurization is carried out to the above-mentioned zymin configured, and preserve in 2-8 DEG C.
3. compound enzymic preparation according to claim 1 eliminates the application in cold turbid phenomenon at beer.
4. compound enzymic preparation according to claim 1 eliminates the application in cold turbid phenomenon at nectar.
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Cited By (2)
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| CN108576547A (en) * | 2018-04-17 | 2018-09-28 | 王智辉 | A kind of complex enzyme formulation and preparation method thereof containing solid acid |
| CN109430683A (en) * | 2018-10-10 | 2019-03-08 | 宁波希诺亚海洋生物科技有限公司 | Remove the complex enzyme formulation of seitan |
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| EP1729593A1 (en) * | 2004-02-24 | 2006-12-13 | Natbio Pty Ltd. | Cysteine protease from ginger (zingiber) as a food improver and anti-inflammatory |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108576547A (en) * | 2018-04-17 | 2018-09-28 | 王智辉 | A kind of complex enzyme formulation and preparation method thereof containing solid acid |
| CN109430683A (en) * | 2018-10-10 | 2019-03-08 | 宁波希诺亚海洋生物科技有限公司 | Remove the complex enzyme formulation of seitan |
| CN109430683B (en) * | 2018-10-10 | 2022-05-27 | 宁波希诺亚海洋生物科技有限公司 | Complex enzyme preparation for removing gluten |
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Denomination of invention: Composite enzyme preparations and their preparation methods and applications Effective date of registration: 20230327 Granted publication date: 20180116 Pledgee: China Construction Bank Corporation Xiangshan Branch Pledgor: NINGBO XINUOYA MARINE BIOTECHNOLOGY Co.,Ltd. Registration number: Y2023330000594 |