CN101899484A - Preparation method of genipin - Google Patents
Preparation method of genipin Download PDFInfo
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- CN101899484A CN101899484A CN2009101868133A CN200910186813A CN101899484A CN 101899484 A CN101899484 A CN 101899484A CN 2009101868133 A CN2009101868133 A CN 2009101868133A CN 200910186813 A CN200910186813 A CN 200910186813A CN 101899484 A CN101899484 A CN 101899484A
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Abstract
The invention belongs to the biotechnology field and relates to a preparation method of genipin. The method comprises a preparation process of immobilized enzyme, a transformation process of gardenoside and a separation and purification process of genipin and is characterized by fermenting the strains for producing beta-glucosidase, collecting the fermentation liquor after reaching the enzyme producing peak, co-immobilizing enzyme and cells or enzyme and hyphae by combining embedding with crosslinking to prepare the immobilized enzyme, utilizing a packed-bed or stirred reactor to carry out catalyzed hydrolysis on the gardenoside and obtaining genipin after purification. The invention dispenses with separation and purification of the fermentation liquor, the immobilized enzyme obtained by preparation has high activity still keeping over 50% after operation for 960h and produces less pollution to the environment, genipin has high yield and the method has obvious advantages compared with other methods.
Description
Technical field
The invention belongs to biological technical field, relate to a kind of preparation method of genipin, particularly a kind of co-immobilization enzyme and the hydrolysis of cell biocatalysis jasminoidin prepare the method for genipin.
Background technology
Genipin is by Geniposide, also claims jasminoidin, the iridoid that obtains after the beta-glucoside enzymic hydrolysis.Genipin has multiple use, and the one, can generate cyanine with the amino acid reaction, this natural blue pigment can be used as foodstuff additive, and its nontoxic, stable in properties is generally used abroad; The 2nd, genipin medically can be used as the natural biological linking agent, in the modern biomedical practice in order to reduce organism to implanting the rejection of heterogeneity biological organization or artificial surrogate, increase the biocompatibility and the mechanical property of biomaterial, usually use crosslinked way to achieve the goal, traditional linking agent major part is the chemosynthesis linking agent, as glutaraldehyde, carbodiimide, tripoly phosphate sodium STPP, epoxy compounds etc., these synthetic cross-linking agents all have bigger cytotoxicity, can influence the healthy tissues growth behind the implantable bioartificial body, and genipin nontoxicity and good biocompatibility can form stable crosslinked; The 3rd, genipin can be used for the treatment of type ii diabetes, studies show that a kind of plastosome uncoupling protein 2 (Uncoupled Protein 2 that cries, UCP2) albumen discharges Regular Insulin restraining effect, if therefore remove the inhibition that UCP2 discharges Regular Insulin, may impel more Regular Insulin to discharge, improve part type ii diabetes symptom, therefore this just effective constituent that can remove UCP2 to suppression of insulin release of genipin be expected to open new epoch for treating type ii diabetes clinically.In addition, genipin can also be used for the treatment of hepatic diseases, hypotensive, and fingerprint collecting reagent, staining agent etc.In a word, the genipin applied range, potentiality to be exploited is big, has vast market prospect.
At present the preparation method of genipin generally adopts beta-glucosidase after the purification to transform jasminoidin as CN 101104745A; CN 101396455A; CN 101029066A passes through extracted with diethyl ether again, the preparation of acetone crystalline method, and the genipin productive rate that this method obtains is low, purity is not high, and the enzyme of input can not reclaim, so cost is higher, so the genipin of Xiao Shouing costs an arm and a leg in the market.In addition, the preparation of genipin need be used a large amount of poisonous organic solvents, and environment is easily polluted; Also have and utilize microbe fermentation method to transform the report that jasminoidin prepares genipin, as Huo Lei, Soviet Union builds, Shen Jing, etc., research and development of natural products 2008,20:70-73; Wang Yonghong, Soviet Union builds, Wang Jianfang, etc. the time precious traditional Chinese medical science traditional Chinese medicines, 2007,18 (12): 3003-3004; Meng Meng XU, Qun SUN, Jian SU, et al.Enzyme and MicrobialTechnology, 2008,42:440-444, but this method exists a bigger defective to be, more amino acid and protein matter are arranged in the fermented liquid during microbial fermentation, after the jasminoidin hydrolysis generates genipin, genipin can with the amino acid generation cyanine that reacts, reduce the yield of genipin greatly; There is report to utilize the immobilization beta-glucosidase to transform the technology that jasminoidin prepares genipin abroad, as ShigeakiFUJIKAWA, Tomoko YOKOTA, Kunimasa KOGA, et al.Biotechnology Letters, 1987,9 (10): 697-702, transformation efficiency can reach more than 95%, and the transformation period of immobilized enzyme reaches 600 hours, but this method need obtain a large amount of pure beta-glucosidases, so cost is also higher.
Summary of the invention
The present invention utilizes microbial fermentation technology, preparation contains beta-glucosidase and cell, or and the fermented liquid of mycelia, again fermented liquid is directly utilized immobilization technology, with β glucuroide and cell, or and the mycelia co-immobilization prepare immobilized enzyme, then jasminoidin is changed into genipin, thereby has finished the present invention.This method need not be carried out separation and purification to fermented liquid, and the immobilized enzyme vigor height for preparing, reach 960 hours working time after enzyme live and still to keep more than 50%, environmental pollution is little, genipin productive rate height is compared with additive method and to be had significant advantage.
The object of the present invention is to provide a kind of co-immobilization technology that adopted, preparation immobilized enzyme and cell or enzyme and mycelia bio-transformation jasminoidin prepare the preparation method of genipin.
The object of the present invention is achieved like this: it comprises the preparation technology of immobilized enzyme, the conversion process of jasminoidin and three parts of separation purifying technique of genipin, the strain fermentation of beta-glucosidase will be produced, after waiting to reach product enzyme peak, fermented liquid is collected, with embedded-cross-linked bonded method co-immobilization enzyme and cell or enzyme and mycelia, the preparation immobilized enzyme adopts packed bed or stirring reactor that jasminoidin is carried out catalytic hydrolysis reaction, obtains genipin after purified.
Genipin is the iridoid that obtains after utilizing beta-glucosidase that Geniposide (also claiming jasminoidin) is hydrolyzed, and its structural formula and hydrolytic process are as shown in Figure 1.With fermentations such as the superior strain that produces beta-glucosidase such as aspergillus niger, Rhizopus oryzae, Aspergillus usamiis, make fermented liquid and mycelia, adopt the co-immobilization method, by crosslinked-embedding in conjunction with the preparation immobilized enzyme, this method need not be carried out separation and purification to enzyme, has reduced production cost.We have used stirring-type and filling bed type immobilized enzyme that jasminoidin is transformed respectively, the result is that transformation efficiency is all more than 95%, but shorten the work-ing life of stirring-type immobilized enzyme greatly, and reason is that stirring reactor has considerable influence to the activity of immobilized enzyme.
Each several part technology is:
(1) preparation technology of immobilized enzyme: the superior strain fermentation that will produce beta-glucosidase, the bacterium liquid that ferments adopts the co-immobilization method to fix together with cell or mycelia, promptly adding final concentration in fermented liquid is the cross-linked material of 0.05%-2%, be that the linking agent of 0.05%-2% carries out crosslinked with final concentration then, carry out embedding with sodium alginate again, dropwise join in the calcium chloride solution then and harden, make globular immobilized enzyme particle;
(2) conversion process of jasminoidin: utilize immobilized enzyme particle, adopt stirring-type or filling bed type to carry out the conversion of jasminoidin, in packed bed jasminoidin solution with the flow velocity of 0.1-3 column volume per hour by the immobilized enzyme bed, invert point is 45-55 ℃, promptly obtain genipin solution, the transformation efficiency of jasminoidin is more than 95%; If adopt stirring-type to transform, then jasminoidin solution is mixed with immobilized enzyme, under 45-55 ℃ of temperature, transform with certain mixing speed, mixing speed is best with 50-200r/min, and transformation time was the best with 2-6 hour, and transformation efficiency reaches more than 95%.
We compare co-immobilization and simple immobilized enzyme, promptly adopt two kinds of enzymes after the different fixationsization with the filling bed type reactor jasminoidin to be transformed respectively, jasminoidin concentration is 1%, with acetate-sodium acetate buffer regulator solution pH value is 4.5, pass through the immobilized enzyme bed with 1BV/hr (BV refers to column volume) flow velocity, respectively take out the 1g immobilized enzyme behind the certain hour and carry out enzyme activity determination, the enzyme activity determination method adopts pNPG method (Liang Huazheng, Liu Fuliang, Peng Lingxi, Deng. Food science, 2006,4:182-185), the result shows, co-immobilization enzyme and cell, or enzyme and mycelia enzyme transformation period of living obviously be longer than the ordinary fixed enzyme, concrete outcome sees the following form 1:
Table 1 co-immobilization takes out the enzyme slip-knot fruit of measuring with simple immobilized enzyme after transforming certain hour
As shown in Table 1, the co-immobilization half life of enzyme reaches about 960hr, and the enzyme that is higher than the ordinary fixed method is lived the transformation period.And the co-immobilization enzyme is lived at preceding 120hr enzyme and is changed all lessly, and reason is to include in the immobilized enzyme that the co-immobilization method makes produces enzyme viable cell or mycelia, and these cells also are in active condition after by embedding, still have enzymatic productivity.
(3) separation purifying technique of genipin: with the genipin conversion fluid through macroporous resin adsorption, wash-out, reconcentration, drying, crystallization, get final product the genipin product, its high-efficient liquid phase chromatogram is as shown in Figure 2.With the high performance liquid chromatography peak area production standard curve of genipin reference substance (purchase in Japanese Wako company), calculation result shows, reaches 98.65% with the genipin content of present method preparation.
(4) yield of genipin: get content and be 95% jasminoidin 100 grams, with pH is that to be dissolved into concentration be 1.5% solution for acetate-sodium acetate buffer of 4.5, other adopts the inventive method co-immobilization to prepare immobilized enzyme and cell, or enzyme and mycelia 500 grams, enzyme is adorned post be placed on 50 ℃ of preheatings, (detect to can not detecting amino acid with the damping fluid flushing then with the genipin development process, with reference to Liang Huazheng, Liao Fusheng, happy growing tall, Deng. research and development of natural products, 2006 (3): 467-470), again with jasminoidin solution with the flow velocity of 1BV/hr by the immobilized enzyme bed, with conversion fluid HPD100 absorption with macroporous adsorbent resin, through wash-out, concentrate, crystallization, obtain Powdered genipin 46.82 grams after the drying, detecting genipin content with the HPLC method is 96.26%, and genipin yield calculation formula is: yield=[(genipin quality * genipin content)/(jasminoidin quality * jasminoidin content)]/(226.23/388.36), calculating yield is 81.45%.
Comprise the steps: during the invention process
(1) will produce superior strain fermentation 48-150 hour of beta-glucosidase, fermented liquid is taken out together with mycelia, directly adding final concentration is the 0.1%-2% cross-linked material, with final concentration be carry out in the linking agent of 0.1%-2% crosslinked, crosslinking time is 0.5-4 hour, crosslinking temperature is 4-20 ℃, or crosslinked again after fermented liquid concentrated together with mycelia;
The superior strain of described product beta-glucosidase is aspergillus niger, Rhizopus oryzae or Aspergillus usamii;
Described cross-linked material is chitosan or gelatin;
Described linking agent is glutaraldehyde, carbodiimide, tripoly phosphate sodium STPP or epoxy compounds;
(2) enzyme after crosslinked and mycelia liquid are carried out embedding with sodium alginate, sodium alginate concentration is 1%-6%;
(3) embedded mixed solution is dropwise added in the calcium chloride solution fix, calcium chloride concentration 0.5%-8%, preparation immobilized enzyme bead;
(4) the immobilized enzyme bead with preparation places room temperature to harden at the calcium chloride solution of 0.1%-1%, and setting time is 1-3 hour;
(5) filter the good immobilized enzyme of sclerosis, place 4 ℃ to preserve standby down;
(6) with immobilized enzyme wet method dress post, prepare the jasminoidin solution of 0.1%-5% again with deionized water, with acetate-sodium acetate buffer regulator solution pH is 4.5, with the flow velocity of 0.1-3 column volume per hour with jasminoidin solution by the immobilized enzyme column bed, hydrolysis temperature is 50 ℃;
(7) with the conversion fluid absorption with macroporous adsorbent resin, through wash-out, concentrate, cryodrying obtains Powdered genipin, purity can reach about 95%, can reach more than 98% through genipin purity behind the recrystallization again;
Described per-cent is weight percentage.
Adopting molecular weight is that 6000 to 30000 hollow fiber membrane ultrafiltration device concentrates together with mycelia fermented liquid, is concentrated into sixth to two/one of original volume.
The present invention adopts the method for co-immobilization enzyme and cell or enzyme and mycelia, only need with the fermented liquid that ferments together with cell or mycelia crosslinked-embedding, not only prolonged the duration of service of immobilized enzyme, also greatly reduce cost, overcome and used in the past that pure enzymatic conversion jasminoidin cost height, efficient are low, the shortcoming of separation difficulty.Its practicality is in particular in following three aspects with superiority: the one, and fermented liquid need not separate, and preparation immobilized enzyme cost is low; The 2nd, the enzymic activity height that co-immobilization enzyme and cell or enzyme and mycelia obtain, long half time; The 3rd, immobilized enzyme washes reject amino acid, protein etc. effectively with damping fluid behind the dress post, and green change can not take place conversion fluid, has improved the yield of genipin.
Description of drawings
Fig. 1, jasminoidin of the present invention are hydrolyzed into the reaction formula of genipin through beta-glucoside.
The high-efficient liquid phase chromatogram of genipin behind Fig. 2, the purifying of the present invention.
Chromatographic condition among Fig. 2 is: Hypersil ODS C18 (4.6 * 250mm, 5 μ m) chromatographic column, moving phase is methyl alcohol: water (22: 78), flow velocity are 1.0ml/min, detect wavelength 238nm.
Embodiment
The present invention can specifically implement by the technology that illustrates in the summary of the invention, can further describe the present invention by the following examples, yet scope of the present invention is not limited to following embodiment, and described per-cent is weight percentage:
(1) will produce enzyme Aspergillus niger strain fermentation 96 hours, fermented liquid is taken out together with mycelia, adopting molecular weight is that 6000 hollow fiber membrane ultrafiltration device concentrates it, be concentrated into about 1/3rd of original volume, adding final concentration and be 1% gelatin and final concentration and be 0.25% glutaraldehyde carries out crosslinked, crosslinking time is 1 hour, and crosslinking temperature is 10 ℃;
(2) enzyme after crosslinked and mycelia liquid are carried out embedding with sodium alginate, sodium alginate concentration is 3%;
(3) embedded mixed solution is dropwise added in the calcium chloride solution fix, calcium chloride concentration 3%, preparation immobilized enzyme bead;
(4) the immobilized enzyme bead with preparation places room temperature to harden at 0.5% calcium chloride solution, and setting time is 2 hours;
(5) filter the good immobilized enzyme of sclerosis, place 4 ℃ to preserve standby down;
(6) with immobilized enzyme wet method dress post, preparing 1% jasminoidin solution again, is 4.5 with acetate-sodium acetate buffer regulator solution pH, with the flow velocity of 0.5 column volume per hour jasminoidin solution is passed through the immobilized enzyme column bed, and hydrolysis temperature is 50 ℃;
(7) conversion fluid is adsorbed with macroporous adsorbent resins such as model HPD-100; Flowing out with deionized water rinsing resin to colourless liquid, is 35% ethanolic soln wash-out again with concentration, collects elutriant; Elutriant is concentrated, obtain the genipin of paste, lyophilize obtains Powdered genipin again, and purity can reach about 95%.Can reach more than 98% through genipin purity behind the recrystallization.
Embodiment 2
(1) will produce enzyme Aspergillus usamii strain fermentation 120 hours, fermented liquid is taken out together with mycelia, adopting molecular weight is that 30000 hollow fiber membrane ultrafiltration device concentrates it, be concentrated into about 1/5th of original volume, adding final concentration and be 0.5% chitosan and final concentration and be 0.5% glutaraldehyde carries out crosslinked, crosslinking time is 2 hours, and crosslinking temperature is 20 ℃;
(2) enzyme after crosslinked and mycelia liquid are carried out embedding with sodium alginate, sodium alginate concentration is 3%;
(3) embedded mixed solution is dropwise added in the calcium chloride solution fix, calcium chloride concentration 3%, preparation immobilized enzyme bead;
(4) the immobilized enzyme bead with preparation places room temperature to harden at 0.5% calcium chloride solution, and setting time is 3 hours;
(5) filter the good immobilized enzyme of sclerosis, place 4 ℃ to preserve standby down;
(6) with immobilized enzyme wet method dress post, preparing 2% jasminoidin solution again, is 4.5 with acetate-sodium acetate buffer regulator solution pH, with the flow velocity of 0.3 column volume per hour jasminoidin solution is passed through the immobilized enzyme column bed, and hydrolysis temperature is 50 ℃;
(7) conversion fluid is adsorbed with model HPD-200 resin; Flowing out with deionized water rinsing resin to colourless liquid, is 35% ethanolic soln wash-out with concentration again, collects elutriant; Elutriant is concentrated, obtain the genipin of paste, lyophilize obtains Powdered genipin again, and purity can reach about 95%.Can reach more than 98% through genipin purity behind the recrystallization.
Embodiment 3
(1) will produce enzyme Rhizopus oryzae bacterial strain fermentation 96 hours, fermented liquid is taken out together with mycelia, and add final concentration and be 2% gelatin and final concentration and be 1% glutaraldehyde and carry out crosslinkedly, crosslinking time is 1 hour, and crosslinking temperature is 10 ℃;
(2) enzyme after crosslinked and mycelia liquid are carried out embedding with sodium alginate, sodium alginate concentration is 3%;
(3) embedded mixed solution is dropwise added in the calcium chloride solution fix, calcium chloride concentration 4%, preparation immobilized enzyme bead;
(4) the immobilized enzyme bead with preparation places room temperature to harden at 0.5% calcium chloride solution, and setting time is 2 hours;
(5) filter the good immobilized enzyme of sclerosis, place 4 ℃ to preserve standby down;
(6) with immobilized enzyme wet method dress post, preparing 2% jasminoidin solution again, is 4.5 with acetate-sodium acetate buffer regulator solution pH, with the flow velocity of 1.5 column volumes per hour jasminoidin solution is passed through the immobilized enzyme column bed, and hydrolysis temperature is 50 ℃;
(7) conversion fluid is adsorbed with model HPD-100 resin; Flowing out with deionized water rinsing resin to colourless liquid, is 35% ethanolic soln wash-out with concentration again, collects elutriant; Elutriant is concentrated, obtain the genipin of paste, lyophilize obtains Powdered genipin again, and purity can reach about 95%.Can reach more than 98% through genipin purity behind the recrystallization.
Embodiment 4
(1) will produce enzyme Aspergillus niger strain fermentation 96 hours, fermented liquid is taken out together with mycelia, and add final concentration and be 1% chitosan and final concentration and be 0.25% glutaraldehyde and carry out crosslinkedly, crosslinking time is 1 hour, and crosslinking temperature is 10 ℃;
(2) enzyme after crosslinked and mycelia liquid are carried out embedding with sodium alginate, sodium alginate concentration is 4%;
(3) embedded mixed solution is dropwise added in the calcium chloride solution fix, calcium chloride concentration 5%, preparation immobilized enzyme bead;
(4) the immobilized enzyme bead with preparation places room temperature to harden at 1% calcium chloride solution, and setting time is 3 hours;
(5) filter the good immobilized enzyme of sclerosis, place 4 ℃ to preserve standby down;
(6) the jasminoidin solution of preparation 0.5% is 5 liters, is 4.5 with acetate-sodium acetate buffer regulator solution pH, adds immobilized enzyme 1kg, and placing is hydrolyzed in 50 ℃ of thermostat water baths transforms 5 hours;
(7) conversion fluid is adsorbed with model HPD-200 resin; Flowing out with deionized water rinsing resin to colourless liquid, is 35% ethanolic soln wash-out with concentration again, collects elutriant; Elutriant is concentrated, obtain the genipin of paste, cryodrying obtains Powdered genipin again, and purity can reach about 95%.Can reach more than 98% through genipin purity behind the recrystallization.
Embodiment 5
(1) will produce enzyme Aspergillus niger strain fermentation 120 hours, fermented liquid is taken out together with mycelia, and add final concentration and be 1% gelatin and final concentration and be 0.25% glutaraldehyde and carry out crosslinkedly, crosslinking time is 1.5 hours, and crosslinking temperature is 15 ℃;
(2) enzyme after crosslinked and mycelia liquid are carried out embedding with sodium alginate, sodium alginate concentration is 3%;
(3) embedded mixed solution is dropwise added in the calcium chloride solution fix, calcium chloride concentration 4%, preparation immobilized enzyme bead;
(4) the immobilized enzyme bead with preparation places room temperature to harden at 0.5% calcium chloride solution, and setting time is 2 hours;
(5) filter the good immobilized enzyme of sclerosis, place 4 ℃ to preserve standby down;
(6) the jasminoidin solution of preparation 0.5% is 10 liters, is 4.5 with acetate-sodium acetate buffer regulator solution pH, adds immobilized enzyme 2kg, and placing is hydrolyzed in 50 ℃ of thermostat water baths transforms 6 hours;
(7) conversion fluid is adsorbed with model HPD-100 resin; Flowing out with deionized water rinsing resin to colourless liquid, is 35% ethanolic soln wash-out with concentration again, collects elutriant; Elutriant is concentrated, obtain the genipin of paste, lyophilize obtains Powdered genipin again, and purity can reach about 95%.Can reach more than 98% through genipin purity behind the recrystallization.
Claims (4)
1. the preparation method of a genipin, it comprises the preparation technology of immobilized enzyme, the conversion process of jasminoidin and three parts of separation purifying technique of genipin, it is characterized in that and to produce the strain fermentation of beta-glucosidase, after waiting to reach product enzyme peak, fermented liquid is collected, with embedded-cross-linked bonded method co-immobilization enzyme and cell or enzyme and mycelia, the preparation immobilized enzyme, adopt packed bed or stirring reactor that jasminoidin is carried out catalytic hydrolysis reaction, obtain genipin after purified.
2. the preparation method of a kind of genipin according to claim 1 is characterized in that each several part technology is:
(1) preparation technology of immobilized enzyme: the superior strain fermentation that will produce beta-glucosidase, the bacterium liquid that ferments adopts the co-immobilization method to fix together with cell or mycelia, promptly adding final concentration in fermented liquid is the cross-linked material of 0.05%-2%, be that the linking agent of 0.05%-2% carries out crosslinked with final concentration then, carry out embedding with sodium alginate again, dropwise join in the calcium chloride solution then and harden, make globular immobilized enzyme particle;
(2) conversion process of jasminoidin: utilize immobilized enzyme particle, adopt stirring-type or filling bed type to carry out the conversion of jasminoidin, in packed bed jasminoidin solution with the flow velocity of 0.1-3 column volume per hour by the immobilized enzyme bed, invert point is 45-55 ℃, promptly obtain genipin solution, the transformation efficiency of jasminoidin is more than 95%; If adopt stirring-type to transform, then jasminoidin solution is mixed with immobilized enzyme, under 45-55 ℃ of temperature, transform with certain mixing speed, mixing speed is best with 50-200r/min, and transformation time was the best with 2-6 hour, and transformation efficiency reaches more than 95%.
(3) separation purifying technique of genipin: with the genipin conversion fluid through macroporous resin adsorption, wash-out, reconcentration, drying, crystallization, get final product the genipin product;
Described per-cent is weight percentage.
3. the preparation method of a kind of genipin according to claim 1 and 2 is characterized in that it comprises the steps:
(1) will produce superior strain fermentation 48-150 hour of beta-glucosidase, fermented liquid is taken out together with mycelia, directly adding final concentration is the 0.1%-2% cross-linked material, with final concentration be carry out in the linking agent of 0.1%-2% crosslinked, crosslinking time is 0.5-4 hour, crosslinking temperature is 4-20 ℃, or crosslinked again after fermented liquid concentrated together with mycelia;
The superior strain of described product beta-glucosidase is aspergillus niger, Rhizopus oryzae or Aspergillus usamii;
Described cross-linked material is chitosan or gelatin;
Described linking agent is glutaraldehyde, carbodiimide, tripoly phosphate sodium STPP or epoxy compounds;
(2) enzyme after crosslinked and mycelia liquid are carried out embedding with sodium alginate, sodium alginate concentration is 1%-6%;
(3) embedded mixed solution is dropwise added in the calcium chloride solution fix, calcium chloride concentration 0.5%-8%, preparation immobilized enzyme bead;
(4) the immobilized enzyme bead with preparation places room temperature to harden at the calcium chloride solution of 0.1%-1%, and setting time is 1-3 hour;
(5) filter the good immobilized enzyme of sclerosis, place 4 ℃ to preserve standby down;
(6) with immobilized enzyme wet method dress post, prepare the jasminoidin solution of 0.1%-5% again with deionized water, with acetate-sodium acetate buffer regulator solution pH is 4.5, with the flow velocity of 0.1-3 column volume per hour with jasminoidin solution by the immobilized enzyme column bed, hydrolysis temperature is 50 ℃;
(7) with the conversion fluid absorption with macroporous adsorbent resin, through wash-out, concentrate, cryodrying obtains Powdered genipin, purity can reach about 95%, can reach more than 98% through genipin purity behind the recrystallization again;
Described per-cent is weight percentage.
4. the preparation method of a kind of genipin according to claim 2, it is characterized in that adopting molecular weight is that 6000 to 30000 hollow fiber membrane ultrafiltration device concentrates together with mycelia fermented liquid, is concentrated into sixth to two/one of original volume.
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| EP3666089A1 (en) | 2011-11-07 | 2020-06-17 | Wild Flavors, Inc. | Method for preparing a colorant from a genipin-rich extract of genipa americana |
| CN103211829A (en) * | 2012-01-13 | 2013-07-24 | 樊向德 | Compounds having virus resistance and composition thereof |
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| CN104313009A (en) * | 2014-10-21 | 2015-01-28 | 江南大学 | Method for immobilizing cellobiose epimerase whole cells |
| CN104498470A (en) * | 2015-01-16 | 2015-04-08 | 齐鲁工业大学 | New type co-immobilization white rot fungi gel particle |
| CN104498470B (en) * | 2015-01-16 | 2017-10-24 | 齐鲁工业大学 | A kind of co-immobilization whiterot fungi gel particle |
| CN105063119B (en) * | 2015-08-31 | 2018-11-30 | 广西红瑶药业有限公司 | The method of Geniposide is extracted from cape jasmine |
| CN105063119A (en) * | 2015-08-31 | 2015-11-18 | 桂林茗兴生物科技有限公司 | Method for extracting genipin from gardenias |
| CN106701858A (en) * | 2017-02-28 | 2017-05-24 | 南京理工大学 | Preparation method of genipin |
| CN106701858B (en) * | 2017-02-28 | 2020-08-11 | 南京理工大学 | Preparation method of genipin |
| CN108913734A (en) * | 2018-08-01 | 2018-11-30 | 河南农业大学 | A kind of optimization method of beta-glucosidase to aminoglycoside hydrolysising condition |
| CN115011506A (en) * | 2022-05-05 | 2022-09-06 | 浙江科技学院 | Method for synthesizing gardenia blue pigment and gardenia red pigment by high-yield beta-glucosidase lactic acid bacteria immobilized cells |
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