CN1594559A - Process for extracting actinidiaceae proteinase - Google Patents
Process for extracting actinidiaceae proteinase Download PDFInfo
- Publication number
- CN1594559A CN1594559A CN 200410026244 CN200410026244A CN1594559A CN 1594559 A CN1594559 A CN 1594559A CN 200410026244 CN200410026244 CN 200410026244 CN 200410026244 A CN200410026244 A CN 200410026244A CN 1594559 A CN1594559 A CN 1594559A
- Authority
- CN
- China
- Prior art keywords
- polyphenol
- tea
- fruit juice
- aqueous solution
- precipitation
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Landscapes
- Coloring Foods And Improving Nutritive Qualities (AREA)
- Medicines Containing Plant Substances (AREA)
Abstract
A method of extracting kiwi fruit proteinase is disclosed. The invention employs 0.1-0.5% green tea polyphenols solution as precipitator, the volume ratio of green tea polyphenols solution with kiwi fruit juice is 0.8-1.1:1. Compared with the prior arts, the invention has merit of high yield, simple process, short period, low cost, innocuity etc, which is suitable for industrialized production of kiwi fruit proteinase.
Description
Technical field
The invention belongs to the improvement of separation and Extraction Actinidin method from kiwifruit fruit.
Background technology
Kiwifruit is the very high fruit of a kind of nutritive value, is called as " king of Vc ", health fruit and space food.Except that containing a large amount of Vc and several kinds of mineral elements, also contain a large amount of Actinidins in the kiwifruit fruit.Actinidin is the plant protease that sulfydryl is contained in a kind of its active centre, has been widely used in fields such as medicine, food, beer production, make-up and beauty, biochemical reagents.
China is Kiwifruit big producing country, existing bimillennial cultivation history.Over nearly 20 years, manual inseminating's area of Kiwifruit enlarges rapidly, and national total area is 870,000 mu at present, produces 330000 tons in fresh fruit per year, and the Kiwifruit resource is very abundant.Kiwifruit fruit belongs to the berry of climacteric type, and storage tolerance not if the fresh fruit dull sale causes overstockedly, will cause waste on the resource and loss economically.Annual in the Kiwifruit production simultaneously have 20% cull fruit also to await processing and utilization approximately.In order to make full use of these Kiwifruit resources, can adopt biochemical method, separation and Extraction goes out Actinidin wherein.By the mode of deep processing, not only can solve the problem of outlet of Kiwifruit cull fruit effectively, for the orchard worker increases income, extend the industrial chain that Kiwifruit is produced, simultaneously also can be for the product of Actinidin comparatively in short supply is provided on the market.
The separation and extraction technology of relevant Actinidin, present disclosed report is also fewer.Xiamen University woods in 1986 the human ammonium sulfate precipitation method such as beautiful jade that oozes separates purification Actinidin (Lin Qinying etc., A.chinensis Planch. proteolytic enzyme separates purifies and property research, " Xiamen University's journal (nature version) ", 1986,25 (3): 347～353), this method belongs to breadboard preparation method, used plant and instrument costliness, the enzyme sample amount of preparation is less, only for the usefulness of scientific research analysis, can't carry out suitability for industrialized production.Extract used ammonium sulfate simultaneously and remain in the enzyme product, also limited its enzyme in medicine, Application in Food.The inventor has submitted the patent application (application number: CN 20,041,002 5932.8) of using sodium-chlor salt precipitator method separation and Extraction Actinidin in a few days ago, though this method has overcome the defective of prior art, reduced extraction cost, can carry out suitability for industrialized production, enzyme preparation also meets the requirement of purposes such as food medicine, but as the industrialized production process technology, the equipment such as salt large usage quantity, process cycle are grown, need refrigerator freezer that still have carry out productivity problems such as low-temperature sludge.
In the method for common separation and Extraction plant protease, mainly be to adopt the precipitator method, and the technology of most critical is the selection of precipitation agent in the precipitator method.In the biochemical method of classics, precipitation agent commonly used is a neutral salt, aforesaid ammonium sulfate and sodium-chlor etc., and these methods were continued to use nearly 200 years always.In the process of modern age the natural plant activeconstituents being furtherd investigate, it is found that the main active ingredient tea-polyphenol in the tealeaves also is a kind of good protein precipitant, can be used for precipitate and separate protein or enzyme.
Tea-polyphenol (being called for short TP, tea polypHenol) is the general name of polyphenols in the tealeaves, is the complex body that contains a plurality of free phenolic hydroxyl groups in the molecule structure.The content of tea-polyphenol in tealeaves is 18%～36%.Because tea-polyphenol has many important purposes in fields such as medicine, food, in recent years the research of tea-polyphenol chemical property is also goed deep into gradually.It has been recognized that at first tea-polyphenol has tannin and the peculiar receipts sword of tannin, be in the tea-polyphenol free phenolic hydroxyl group can with virus, protein bound on the bacterium, bacterioprotein is precipitated, thereby have an anti-virus sterilizing effect (Yang Xianqiang etc., " tea-polyphenol chemistry " Shanghai science tech publishing house, 2003,44-46).Along with going deep into of research, and then find that tea-polyphenol has common chemical property of polyphenolic compound: i.e. a large amount of free phenolic hydroxyl group in the tea-polyphenol, can with protein generation complexing action, when iso-electric point, just can be with protein or enzyme precipitate and separate from solution.
Research for the proteinic chemical property of tea-polyphenol complex-precipitation, people (Haslam E.Plat Polyenols Vegetable Tannis Revisited such as Haslam E., Cambridge Universitypress, 1989,10,170) proposed the theoretical model and the action condition of tea-polyphenol and protein bound, robot systems such as Yang Xianqiang have been described the binding mode of tea-polyphenol precipitating proteins.Under these guide of theory, people had begun tea-polyphenol is used for the real work of separation and Extraction protein in recent years.But because tea-polyphenol still is in the starting stage in this applied research on the one hand, example that can reference is less, and range of application also only is confined in a few protein.And,, still find no the research report that closes this respect through data-searching for utilizing tea-polyphenol separation and Extraction Actinidin.
Summary of the invention
The purpose of this invention is to provide a kind of extracting method that can be used in industrial Actinidin, make it further reduce extraction cost, and operation is more simplified, process cycle shortens, improve the yield of enzyme, make whole technological process be fit to suitability for industrialized production more.
The Actinidin separation and Extraction that the present invention proposes is to adopt the complex-precipitation method, and method steps includes: fruit juice mixes with precipitation agent, staticly settles solid-liquid separation, drying and crushing; Characteristics of the present invention are that employed precipitation agent is the aqueous solution that contains tea-polyphenol, and the weight ratio of its polyphenol content is 0.1%～0.5%, and itself and carambola juice blended volume ratio are 0.8～1.1: 1.The described preparation that contains the tea-polyphenol aqueous solution can be adopted powdered commodity tea-polyphenol, and water is mixed with the aqueous solution of 0.2%～0.5% (w/w), and itself and carambola juice blended volume ratio are 0.8～1: 1; Contain the Aqua Folium Camelliae sinensis that the aqueous solution of tea-polyphenol also can adopt tealeaves to make, wherein the content of tea-polyphenol (w/w) is 0.1%～0.4%, and itself and carambola juice blended volume ratio are 0.9～1.1: 1.
Complex-precipitation of the present invention is, after adding contains the aqueous solution of tea-polyphenol in carambola juice, tea-polyphenol just can with Actinidin generation complex reaction, form tea-polyphenol-proteinic mixture, and then aggegation generation flocks, thereby being precipitated from solution, Actinidin separates out.
Normal temperature of the present invention staticly settles and is, the precipitation precipitation process of proteolytic enzyme is to carry out at ambient temperature, the aqueous solution that is about to contain tea-polyphenol be placed under the room temperature condition after carambola juice mixes, can finish precipitation after 2～4 hours and separate out.Generally speaking, if the precipitation agent that uses is the commodity tea-polyphenol, required sedimentation time is longer, and minimum sedimentation time is 4 hours; If the precipitation agent that uses is a fresh tea leaf in its water, required sedimentation time is shorter, and minimum sedimentation time is 2 hours.
Solid-liquid separation of the present invention, drying and crushing and ethanol are purified and are all adopted prior art.
When precipitating, the present invention there is no strict demand for the iso-electric point in the mixed solution.Because the proteinic ability of tea-polyphenol complex-precipitation is stronger, iso-electric point is faint for the effect of assisting precipitating proteins, so the pH value a wider range that requires, generally speaking, all can in pH value to 3.0～5.5 of adjusting mixed solution.Be standard operation, can be advisable by adjust pH to 5.0.
In actual production operation, for complete operation at ambient temperature better, can not in time handle and room temperature when too high (more than 20 ℃) when the carambola juice that squeezes, the salt that can add 8%～12% (w/v) in fruit juice is beneficial to fruit juice and deposits.The purpose that adds sodium-chlor can play preservative activity on the one hand, and sodium-chlor also can be used as precipitation agent and plays and help heavy effect simultaneously, makes the precipitation of formation more thorough, to improve the yield of enzyme.
The present invention adopts the principle of complex-precipitation method institute foundation to be: tea-polyphenol is a kind of novel precipitation agent that developed recently gets up, and is strong to proteinic precipitation ability, the deposition efficiency height.Tea-polyphenol is a polyphenolic compound, contained a large amount of free phenolic hydroxyl group can with protein generation complexing action, tea-polyphenol and the formed mixture of protein are gone out at iso-electric point sinking precipitation, just can reach the purpose of separation and Extraction protein or enzyme.About the theoretical pattern of tea-polyphenol and protein bound, the pattern that Haslam E. proposes is: the principal mode of Polyphenols and protein bound is a hydrogen bond, promptly on phenolic hydroxyl group-OH in the polyphenol and the protein peptide chain-NH-CO-and with side chain on-OH ,-NH
2,-COOH etc. are with the combination of hydrogen bond form.People such as Yang Xianqiang have explained that further Polyphenols and binding pattern between the protein can be the multiple spot hydrophobic bonds and the combining of hydrogen bond, be that hydrophobic grouping in the polyphenol molecule enters the hydrophobic position in the protein molecule, multidigit point hydrogen bonded more than 2 takes place in phenolic hydroxyl group on the polyphenol and the polar group in the protein molecule (as peptidyl-NH-CO, hydroxyl-OH, carboxyl-COOH etc.) then, forms a kind of stable bond between polyphenol and the protein thereby make.
The present invention adopts tea-polyphenol as protein precipitant, has following clear superiority: 1. tea-polyphenol is the secondary metabolite of tea tree, the content in tealeaves quite high (18%～36%).Belong to the natural plant activeconstituents, food grade materials, nontoxic, meet the special safety and health request of medicine food, therefore be suitable as the extraction Actinidin, the tealeaves among the present invention can be low-grade tea, tea stalk, tea dust etc.2. the effectiveness height of tea-polyphenol precipitate and separate proteolytic enzyme, and consumption is few, and the throw out of formation is intensive thoroughly, be easy to filter, and it is short to form the required time of precipitation, and whole process cycle is shortened.3. tea-polyphenol has anti-oxidant and provide protection for biomacromolecule, can the stabilized enzyme activity; Because Actinidin belongs to the sulfydryl plant protease, the easily oxidized and inactivation of its sulfydryl, the present invention is used for the separation and Extraction of enzyme with tea-polyphenol, the sulfydryl of enzyme active center is protected, thereby has special meaning.4. tealeaves is the potability plant of extensively planting, and China is Tea Production big country, a large amount of low and middle-grade tea and tea stalk, tea is all arranged not every year, and raw material resources are very sufficient, and it is used as precipitation agent, can improve the overall economic efficiency of tealeaves industry.
Find out that by above the extracting method of Actinidin proposed by the invention has following characteristics:
1) uses tea-polyphenol as precipitation agent separation and Extraction Actinidin, both can reduce the salt consumption or, can omit the desalination step again, reduce the technological operation link without salt;
2) because tea-polyphenol is stronger to proteinic precipitation ability, required sedimentation time is short, is reduced to 2～4 hours by 12 hours of salting-out process; And the filtration velocity quickening, whole process cycle is shortened;
3) precipitation process among the present invention is to carry out (salting-out process is to carry out at low temperatures) at normal temperatures, therefore, needn't drop into main equipments such as refrigerator, freezer in the production;
4) reduced extraction cost, compared with the salt precipitator method, the commodity in use tea-polyphenol can reduce fixed cost more than 10%, uses Aqua Folium Camelliae sinensis can reduce fixed cost more than 20%;
5) improved the yield of enzyme, in following several extracting method, the thick enzyme yield of Actinidin is respectively: the salt precipitator method are 1.22%, and the commodity in use tea-polyphenol is 1.46%, and using Aqua Folium Camelliae sinensis is 1.52%, and using Aqua Folium Camelliae sinensis and adding salt is 1.76%;
6) the present invention is directly with the on-the-spot Aqua Folium Camelliae sinensis precipitation separation Actinidin that boils, and does like this that not only raw material is easy to get, low price and tea-polyphenol be without grading purification, and molecular weight ranges is wide, the better effects if of its precipitating proteins; Simultaneously, directly use Aqua Folium Camelliae sinensis can avoid of the influence of the contained dispersion agent of commodity tea-polyphenol to enzyme preparation purity.
Embodiment
Example 1 is got fresh carambola juice 1L, the 0.3% tea-polyphenol aqueous solution 800ml (the powdered tea-polyphenol that the green precious company in Wuxi produces) of preparation is mixed with it, stir, adjust the pH value to 5.0 of mixed solution, at room temperature staticly settle more than 4 hours, flocculent aggregate is deposited in container bottom.Abandoning supernatant is stuck with paste the rare enzyme of lower floor to change over to and is carried out suction filtration in the B, and the throw out in the collection funnel carries out vacuum-drying, pulverizes and sieves through ball mill, obtains thick enzyme dry products 14.6 grams of Actinidin, and its thick enzyme yield is 1.46%.
Example 2 is got fresh carambola juice 1L, and (use shanqing tea 22g, add 1500ml water, boiled 3 minutes, 20 minutes after-filtration are soaked in insulation, obtain the Aqua Folium Camelliae sinensis of polyphenol content about 0.15% with self-control Aqua Folium Camelliae sinensis 1L.Usually the solubility rate of tea-polyphenol is about 10%) mix, stir, adjust the pH value to 4.5 of mixed solution, at room temperature staticly settle more than 2 hours, discard supernatant liquor, the rare enzyme of lower floor is stuck with paste carried out suction filtration, the collecting precipitation thing carries out vacuum-drying, pulverize and sieve through ball mill, obtain the thick enzyme dry products of Actinidin 15.2g, thick enzyme yield is 1.52%.
Example 3 is got fresh carambola juice 2L, adds salt (NaCl) 200g, after the stirring and dissolving, can at room temperature preserve 1-3 days.Get example 2 homemade Aqua Folium Camelliae sinensis 1.8L, mix with carambola juice, adjust pH value to 4.0, staticly settled under the room temperature 3 hours, abandoning supernatant is stuck with paste decompress filter with the rare enzyme in bottom, obtains the wet product 62.8g of the thick enzyme of Actinidin.Get wherein that 31.4g carries out vacuum lyophilization, carry out ball mill pulverizing then and sieve, obtain the thick enzyme dry products of Actinidin 17.6g, its thick enzyme yield is 1.76%; With the wet product of the thick enzyme of remaining 31.4g, carrying out ethanol purifies, the ethanolic soln that adds 800ml 80% (V/V) makes thick enzyme dissolving, stirs, and changes centrifuge tube under 4000r/min speed centrifugal 15 minutes, remove supernatant liquor, collect the throw out of bottom, carry out vacuum lyophilization, grind and sieve with mortar, obtain the pure enzyme dry products of Actinidin 10.7g, the yield of its pure enzyme is 1.07%.
Claims (5)
1, a kind of method of extracting Actinidin is to adopt the complex-precipitation method, and method steps includes: fruit juice mixes with precipitation agent, staticly settles solid-liquid separation, drying and crushing; It is characterized in that: employed precipitation agent is the aqueous solution that contains tea-polyphenol, and the weight ratio of its polyphenol content is 0.1～0.5%, and itself and fruit juice blended volume ratio are 0.8～1.1: 1.
2, method according to claim 1 is characterized in that: the described aqueous solution that contains tea-polyphenol is 0.2～0.5% the aqueous solution that adopts the preparation of commodity tea-polyphenol, and itself and fruit juice blended volume ratio are 0.8～1: 1; It staticly settles is to leave standstill at ambient temperature to be not less than 4 hours.
3, method according to claim 1 is characterized in that: the described aqueous solution that contains tea-polyphenol is that the polyphenol content that adopts tealeaves to make is 0.1～0.4% Aqua Folium Camelliae sinensis, and itself and fruit juice blended volume ratio are 0.9～1.1: 1; It staticly settles is to leave standstill at ambient temperature to be not less than 2 hours.
4, method according to claim 1 is characterized in that: pH to 3.0～5.5 of adjusting mixed solution before staticly settling.
5, method according to claim 1 is characterized in that: the salt that allows to contain in the fruit juice 8～12% weightmeasurement ratios before fruit juice and precipitant mix.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 200410026244 CN1273588C (en) | 2004-06-18 | 2004-06-18 | Process for extracting actinidiaceae proteinase |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 200410026244 CN1273588C (en) | 2004-06-18 | 2004-06-18 | Process for extracting actinidiaceae proteinase |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1594559A true CN1594559A (en) | 2005-03-16 |
| CN1273588C CN1273588C (en) | 2006-09-06 |
Family
ID=34663862
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN 200410026244 Expired - Fee Related CN1273588C (en) | 2004-06-18 | 2004-06-18 | Process for extracting actinidiaceae proteinase |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN1273588C (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102499351A (en) * | 2011-11-17 | 2012-06-20 | 西南科技大学 | Actinidia clarifying agent and process |
| CN102499352A (en) * | 2011-11-17 | 2012-06-20 | 西南科技大学 | Kiwi fruit meat tenderizer and process |
| CN108504646A (en) * | 2018-06-07 | 2018-09-07 | 贵州沣稷生物科技有限公司 | A kind of highly effective extraction method of bromelain |
-
2004
- 2004-06-18 CN CN 200410026244 patent/CN1273588C/en not_active Expired - Fee Related
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102499351A (en) * | 2011-11-17 | 2012-06-20 | 西南科技大学 | Actinidia clarifying agent and process |
| CN102499352A (en) * | 2011-11-17 | 2012-06-20 | 西南科技大学 | Kiwi fruit meat tenderizer and process |
| CN102499352B (en) * | 2011-11-17 | 2013-05-08 | 西南科技大学 | Kiwi fruit meat tenderizer and process |
| CN108504646A (en) * | 2018-06-07 | 2018-09-07 | 贵州沣稷生物科技有限公司 | A kind of highly effective extraction method of bromelain |
Also Published As
| Publication number | Publication date |
|---|---|
| CN1273588C (en) | 2006-09-06 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN101508690B (en) | Novel methods for simultaneously extracting tea polyphenol, tea polysaccharide and caffeinum from tea | |
| CN103238819B (en) | Plant salt and preparation method of plant salt | |
| CN104892785B (en) | A kind of extracting method of algal polysaccharides | |
| CN1566356A (en) | Method for preparing earthworm bioactive peptide and earthworm biological micro-fertilizer and use thereof | |
| CN101942014A (en) | Food grade phycocyanin and preparation method thereof | |
| CN1124784C (en) | Biological Pesticide prepared with the extraction from xianggu mushroom leftover and the preparation process | |
| CN101851300A (en) | Process for extracting chondroitin sulfate | |
| CN1091152C (en) | Physical extraction of bromelain | |
| CN1304488C (en) | Technique for extracting edible natural red color element from psammophyte of nitraria | |
| CN1273588C (en) | Process for extracting actinidiaceae proteinase | |
| CN1259335C (en) | Method for preparing phycocyanin from blue algae in water bloom | |
| CN109504573B (en) | Washing liquid composition prepared from fruit and vegetable garbage and preparation method thereof | |
| CN111471732A (en) | Novel selenium-rich tea source ACE inhibitory peptide and preparation method thereof | |
| CN102885056A (en) | Natamycin preparation and preparation method thereof | |
| CN1817822A (en) | Special fertilizer of sweet wormwood | |
| CN101880203B (en) | Vegetable foliar biological fertilizer and preparation method thereof | |
| CN104327175B (en) | A kind of method for separating antibacterial peptide | |
| CN1334258A (en) | Process for preparing nourishing agent of plant | |
| CN1243095C (en) | Extracting process of actinidia proteinase | |
| CN109111300A (en) | A kind of grape special nutritional bacteriostatic agent and the preparation method and application thereof | |
| CN1100563C (en) | Extraction of effective component in Northern cordyceps producing matrix | |
| CN1156002A (en) | Preparation method of barley green powder | |
| CN1566036A (en) | Production method of amino acid chelate compound environmental protection fertilizer | |
| CN108504646A (en) | A kind of highly effective extraction method of bromelain | |
| CN1743341A (en) | Method for preparing lycium barbarum polysaccharides |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| C19 | Lapse of patent right due to non-payment of the annual fee | ||
| CF01 | Termination of patent right due to non-payment of annual fee |