CN105085714A - Process for extracting chondroitin sulfate in cartilage - Google Patents
Process for extracting chondroitin sulfate in cartilage Download PDFInfo
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- CN105085714A CN105085714A CN201510557844.0A CN201510557844A CN105085714A CN 105085714 A CN105085714 A CN 105085714A CN 201510557844 A CN201510557844 A CN 201510557844A CN 105085714 A CN105085714 A CN 105085714A
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Abstract
The invention belongs to the field of biochemical pharmacy, and provides a process for extracting chondroitin sulfate in cartilage. The process is characterized by comprising the following steps: (1) cleaning; (2) material boiling; (3) enzymolysis; (4) filter pressing; (5) centrifugal separation: conducting centrifugal separation on filtrate obtained in the step (4) to obtain a separation liquid, residue and grease; (6) backflow: pumping grease obtained in the step (5) into a grease separating tank, cooling, solidifying and separating grease through the grease separating tank, and flowing a separated enzymatic hydrolysate back to a stewing tank in the step (2); (7) ion exchange; (8) elution; (9) chondroitin sulfate extraction. Through improvement of the process, double separation of grease and the enzymatic hydrolysate as well as enzymatic hydrolysate backflow recycle is adopted, and the viscosity of chondroitin sulfate solution is reduced, so that the yield and purity of the product are effectively improved when the production cost of the process is reduced.
Description
Technical field
The invention belongs to biochemical pharmacy field, be specifically related to a kind of technique extracting chondroitin sulfate in cartilage.
Background technology
Chondroitin sulfate (chondroitinsulfate, CS) is a kind of natural acid mucopolysaccharide, and be mainly present in animal cartilage tissue, its molecular weight is generally 10 × 10
3~ 50 × 10
3, there is the important biomolecule functions such as anti-inflammatory, anticancer, AntiHIV1 RT activity, reducing blood-fat, anticoagulant active, prevention sacroiliitis.In China, it is mainly used in treatment neurodynia, sacroiliitis, omarthralgia and abdominal postoperative pain etc., the dysacousis that prevention and therapy Streptomycin sulphate causes and hypoacusia, tinnitus disease etc. that various noise causes.In Australia, chondroitin sulfate generally exists as dietary supplements, for preventing and treating osteoarthritis, Japan its as nonprescription drugs, be applied to eye drops, arthralgia relief medicine, in more American-European developed countries, CS as dietary supplements prolonged application in diseases such as control coronary heart diseases and angina pectoris, myocardial infarction, coronary insufficiency, myocardial ischemias.On the other hand, chondroitin sulfate, not only as medicine, also as important additive, is widely used in various high-grade healthcare products, food, feed, the cosmetics of super quality.Along with the continuous expansion of chondroitin sulfate novelty teabag, world market also presents quick growth to the demand of chondroitin sulfate.
Along with the continuous expansion of bioactive constantly revealed and its Application Areas of chondroitin sulfate, will be more and more higher to the requirement of its degree that becomes more meticulous.At present, about the extraction of chondroitin sulfate, purification and separation technology achieve certain achievement in research, but in the production and research of chondroitin sulfate, existing production technique to be improved further to improve productive rate and the purity of product, reduce production cost, adapt to the new demand both at home and abroad to raw material and preparation.
In prior art, how unprocessed through isolated grease, the present invention is not only by the separation and purification again of the grease through three phase separation, namely cooling solidification separation is carried out with grease separating tank, effectively enzymolysis solution contained by grease and its is carried out secondary separation, improve grease purity, be conducive to further utilization, improve economic worth; On the other hand the enzymolysis solution containing chondroitin sulfate and other materials be separated is refluxed in digester, improving the utilization ratio of raw material, while reducing costs, adding the productive rate of product, decreasing the discharge of refuse, killing three birds with one stone.
Preferably, described a kind of technique extracting chondroitin sulfate in cartilage, is characterized in that in described step (6), separation temperature is 6 ~ 9 DEG C.This preferred version can effectively by enzymolysis solution and grease secondary separation.
Preferably, adopt ultrasonic cleaning in described step (1), scavenging solution is 0.01 ~ 0.06mol/LNaHCO
3solution.Many experiments shows, this preferred version is comparatively thorough to cartilage cleaning.
Preferably, adopt double enzymolysis technique in described step (3), wherein the add-on of 2709 proteolytic enzyme and the add-on of Pancreas Sus domestica are than being 1:10 ~ 15, are 0.1 ~ 0.9% of cartilage quality, hydrolysis temperature is 45 DEG C ~ 55 DEG C, pH8.5 ~ 9.0, and the treatment time is 4 ~ 8 hours.Under these processing condition, chondroitin sulfate extraction yield is higher, and protein digestion rate is higher.
Preferably, intensification described in described step (4) is gone out after enzyme step, has pH4.0 ~ 4.5 time process 0.5 ~ 3 hour, then adjusts the step of pH to 5.0 ~ 6.0.
Contain-S0 in chondroitin sulfate more
3h and the-group such as COOH, in the alkaline environment of enzymolysis process with the positively charged ion such as sodium, potassium in conjunction with salify, acidic treatment, by the positively charged ion such as sodium, potassium wash-out, makes it revert to-S0
3the form of H and-COOH, is conducive to the ion-exchange in step (6) like this; The solution viscosity of chondroitin sulfate is larger, and be now solid, liquid, oily three-phase mixture, chondroitin sulfate solution easily sticks in solid phase and oil phase, cause the loss of product, cause productive rate to decline, and due to viscosity large, also can adhere to many solid phases and oil phase substance, cause the difficulty in separation, reduce product purity.Research shows, pH value is lower, and chondroitin sulfate solution viscosity is lower, but pH value must not be too low, this instability under sour environment of chondroitin sulfate.It is high that pH regulator of the present invention efficiently solves chondroitin sulfate solution viscosity, easily loses, and the difficult feature be separated, effectively improves the productive rate of chondroitin sulfate.
Preferably, after enzyme is gone out in intensification in step (4), process 2 hours under also comprising pH4.3, then adjust pH to 5.4 to the treating processes of the obvious layering of enzymolysis solution.
Under this optimum condition, chondroitin sulfate solution viscosity effectively reduces, and three-phase layering is obvious, and chondroitin sulfate character controls also comparatively stable.
Preferably, described step (5) adopts disk centrifuge to be separated, and operating parameters is disengaging time 1 ~ 3 hour, rotary drum rotating speed 6000 ~ 8000rpm/min, temperature 45 ~ 50 DEG C.
Preferably, the mass ratio that it is characterized in that parting liquid and macroporous anion exchange resin in described step (6) according to a kind of technique extracting chondroitin sulfate co-producing amino acid powder in cartilage according to claim 1 is 1 ~ 1.5:1, adsorption time 1 ~ 2 hour.
Preferably, described cartilage is one or more in nose of an ox bone, ox larynx bone, crops bone, ox trunnion, ox cuirass bone.
Preferably, elutriant described in described step 9 also has centrifugal treating technique after oxidative decoloration.
Preferably, alcohol precipitation process described in described step 9 is secondary alcohol precipitation.
Preferably, drying process described in described step 9 is vacuum-drying.
The present invention is in the technical foundation of prior art Enzymatic Extraction chondroitin sulfate, by the improvement to operational path and processing parameter, especially adopt the secondary separation of grease and enzymolysis solution, enzymolysis solution refluxes and recycle, and by reducing the viscosity of chondroitin sulfate solution, while production cost of the present invention is reduced, effectively improve productive rate and the purity of product.
Summary of the invention
In view of this, the object of this invention is to provide and a kind ofly extract in cartilage the technique extracting chondroitin sulfate, simple to solve China's existing crude product chondroitin sulfate production technology, product yield is low, the problems such as purity is low, the solution of the present invention is: a kind of technique extracting chondroitin sulfate in cartilage, comprises the steps:
(1) clean;
(2) material is boiled: boiling in digester, cooling;
(3) enzymolysis;
(4) press filtration: heat up the enzyme that goes out, and filters to get filtrate and filter residue;
(5) centrifugation: filtrate is through centrifugation in step (4), obtains parting liquid and residue and grease;
(6) reflux: the grease in step (5) is through pump in grease separating tank, and grease is separated through the cooling solidification of grease separating tank, the digester in the enzymolysis solution backflow progressive rapid (2) of separation.
(7) ion-exchange: the parting liquid in step (5) adsorbs through anionite-exchange resin, obtains penetrating liquid;
(8) wash-out: step (7) ion exchange resin, through eluent, obtains elutriant;
(9) extraction of chondroitin sulfate: the elutriant in step (8) is centrifugal through oxidative decoloration, ultrafiltration, alcohol precipitation, dehydration, drying obtains chondroitin sulfate.
Accompanying drawing explanation
Fig. 1 is the process flow sheet extracting chondroitin sulfate in cartilage
Embodiment
Below in conjunction with specific embodiment, the invention will be further described, and to help understanding the present invention better, but protection scope of the present invention is not limited in these embodiments.
Embodiment 1
Extract a technique for chondroitin sulfate in cartilage, comprise the steps:
(1) clean: use 0.05mol/LNaHCO
3solution ultrasonic cleaning cartilage;
(2) material is boiled: boiling 5 hours in digester, cooling;
(3) enzymolysis: add 2709 proteolytic enzyme and the broken liquid mixture of Pancreas Sus domestica in the boiling cooling fluid in step (2), 48 DEG C, pH8.8 process 6 hours, wherein the add-on of 2709 proteolytic enzyme and the add-on of Pancreas Sus domestica are than being 1:13, are 0.8% of cartilage quality;
(4) press filtration: enzymolysis solution heats up and goes out after enzyme, temperature 80 DEG C, 0.5 hour enzyme that goes out, is cooled to 62 DEG C, process 1 hour under pH4.2, then after adjusting pH to 5.4 to the obvious layering of enzymolysis solution, with 747 filter cloths filtrations, obtains filtrate and filter residue;
(5) centrifugation: filtrate is through centrifugation in step (4), obtains parting liquid and residue and grease; Realize solid, fat, liquid through disk centrifugal separator and be separated 1 hour, rotary drum rotating speed 6000rpm/min, temperature 45 C, obtain parting liquid and residue and grease.
(6) reflux: the grease in step (5) is through pump in grease separating tank, and grease is 7 DEG C through grease separating tank control temperature and solidifies separation, the digester in the enzymolysis solution backflow progressive rapid (2) of separation.
(7) ion-exchange: the parting liquid in step (5) adsorbs through anionite-exchange resin, the mass ratio of parting liquid and anionite-exchange resin is 1.4:1, and adsorption time 1.5 hours, obtains penetrating liquid;
(8) wash-out: step (7) ion exchange resin through the NaCl solution wash-out of concentration 2mol/L, elution time 2.5 hours, pH8.5.
(9) extraction of chondroitin sulfate: the elutriant in step (8) is after oxidative decoloration, inlet pipe formula centrifuge is separated, supernatant liquor pumps into ultra-filtration equipment, carry out desalination and concentration through ultrafiltration apparatus and obtain concentrated solution, ultra-filtration membrane molecular weight cut-off 1000 ~ 8000 dalton, working pressure is 0.4MPa, and temperature is at 50 DEG C, face speed 4m/s, trapped fluid obtains chondroitin sulfate through ethanol precipitation twice, centrifugal, the vacuum-drying of dewatering.
The finished product yield is 17.5%, and after testing, every quality index of chondroitin sulfate is as shown in table 1:
Every quality index of table 1 embodiment 1 gained chondroitin sulfate
Embodiment 2
Extract a technique for chondroitin sulfate in cartilage, comprise the steps:
(1) clean: use 0.01mol/LNaHCO
3solution ultrasonic cleaning cartilage;
(2) material is boiled: boiling 4 hours in digester, cooling;
(3) enzymolysis: add 2709 proteolytic enzyme and the broken liquid mixture of Pancreas Sus domestica in the boiling cooling fluid in step (2), 45 DEG C, pH8.5 process 4 hours, wherein the add-on of 2709 proteolytic enzyme and the add-on of Pancreas Sus domestica are than being 1:10, are 0.1% of cartilage quality;
(4) press filtration: enzymolysis solution heats up and goes out after enzyme, temperature 85 DEG C, 1 hour enzyme that goes out, is cooled to 65 DEG C, process 3 hours under pH4.0, then after adjusting pH to 6.0 to the obvious layering of enzymolysis solution, with 747 filter cloths filtrations, obtains filtrate and filter residue;
(5) centrifugation: filtrate is through centrifugation in step (4), obtains parting liquid and residue and grease; Realize solid, fat, liquid through disk centrifugal separator and be separated 3 hours, rotary drum rotating speed 8000rpm/min, temperature 50 C, obtain parting liquid and residue and grease.
(6) reflux: the grease in step (5) is through pump in grease separating tank, and grease is 6 DEG C through grease separating tank control temperature and solidifies separation, the digester in the enzymolysis solution backflow progressive rapid (2) of separation.
(7) ion-exchange: the parting liquid in step (5) adsorbs through anionite-exchange resin, the mass ratio of parting liquid and anionite-exchange resin is 1:1, and adsorption time 1 hour, obtains penetrating liquid;
(8) wash-out: step (7) ion exchange resin through the NaCl solution wash-out of concentration 3mol/L, elution time 2 hours, pH8.0.
(9) extraction of chondroitin sulfate: the elutriant in step (8) is after oxidative decoloration, enter centrifuge to be separated, supernatant liquor pumps into ultra-filtration equipment, carry out desalination and concentration through ultrafiltration apparatus and obtain concentrated solution, ultra-filtration membrane molecular weight cut-off 1000 ~ 8000 dalton, working pressure is 0.4MPa, and temperature is at 50 DEG C, face speed 4m/s, trapped fluid obtains chondroitin sulfate through ethanol precipitation twice, centrifugal, the vacuum-drying of dewatering.
The finished product yield is 17.1%, and purity is 96.15%, and after testing, every quality index of chondroitin sulfate all reaches standard-required.
Embodiment 3
Extract a technique for chondroitin sulfate in cartilage, comprise the steps:
(1) clean: use 0.06mol/LNaHCO
3solution ultrasonic cleaning cartilage;
(2) material is boiled: boiling 5 hours in digester, cooling;
(3) enzymolysis: add 2709 proteolytic enzyme and the broken liquid mixture of Pancreas Sus domestica in the boiling cooling fluid in step (2), 55 DEG C, pH9.0 process 8 hours, wherein the add-on of 2709 proteolytic enzyme and the add-on of Pancreas Sus domestica are than being 1:15, are 0.9% of cartilage quality;
(4) press filtration: enzymolysis solution heats up and goes out after enzyme, temperature 83 DEG C, 0.8 hour enzyme that goes out, is cooled to 60 DEG C, process 0.5 hour under pH4.5, then after adjusting pH to 5.0 to the obvious layering of enzymolysis solution, with 747 filter cloths filtrations, obtains filtrate and filter residue;
(5) centrifugation: filtrate is through centrifugation in step (4), obtains parting liquid and residue and grease; Realize solid, fat, liquid through disk centrifugal separator and be separated 2 hours, rotary drum rotating speed 7000rpm/min, temperature 48 DEG C, obtain parting liquid and residue and grease.
(6) reflux: the grease in step (5) is through pump in grease separating tank, and grease is 9 DEG C through grease separating tank control temperature and solidifies separation, the digester in the enzymolysis solution backflow progressive rapid (2) of separation.
(7) ion-exchange: the parting liquid in step (5) adsorbs through anionite-exchange resin, the mass ratio of parting liquid and anionite-exchange resin is 1.5:1, and adsorption time 2 hours, obtains penetrating liquid;
(8) wash-out: step (7) ion exchange resin through the NaCl solution wash-out of concentration 1.5mol/L, elution time 3 hours, pH9.0.
(9) extraction of chondroitin sulfate: the elutriant in step (8) is after oxidative decoloration, carry out desalination and concentration through ultrafiltration apparatus and obtain concentrated solution, ultra-filtration membrane molecular weight cut-off 1000 ~ 8000 dalton, working pressure is 0.4MPa, temperature is at 50 DEG C, face speed 4m/s, trapped fluid obtains chondroitin sulfate through alcohol precipitation, centrifugal, the drying of dewatering.
The finished product yield is 16.7%, and purity is 95.45%, and after testing, every quality index of chondroitin sulfate all reaches standard-required.
Embodiment 4
Replace with except ox trunnion except by nose of an ox bone, other are with embodiment 1, and the yield of chondroitin sulfate is 14.8%, and purity is 94.8%.
Embodiment 5
Replace with except crops bone except by nose of an ox bone, other are with embodiment 2, and the yield of chondroitin sulfate is 14.2%, purity 93.6%.
Embodiment 6
Replace with except ox crescent moon except by nose of an ox bone, other are with embodiment 2, and the yield of chondroitin sulfate is 13.7%, purity 94.4%.
Above-described embodiment is the present invention's preferably embodiment; but embodiments of the present invention are not restricted to the described embodiments; change, the modification done under other any does not deviate from spirit of the present invention and principle, substitute, combine, simplify; all should be the substitute mode of equivalence, be included within protection scope of the present invention.
Claims (10)
1. extract a technique for chondroitin sulfate in cartilage, it is characterized in that comprising the steps:
(1) clean;
(2) material is boiled: boiling in digester, cooling;
(3) enzymolysis;
(4) press filtration: heat up the enzyme that goes out, and filters to get filtrate and filter residue;
(5) centrifugation: filtrate is through centrifugation in step (4), obtains parting liquid and residue and grease;
(6) reflux: the grease in step (5) is through pump in grease separating tank, and grease is separated through the cooling solidification of grease separating tank, the digester in the enzymolysis solution backflow progressive rapid (2) of separation;
(7) ion-exchange: the parting liquid in step (5) adsorbs through anionite-exchange resin, obtains penetrating liquid;
(8) wash-out: step (7) ion exchange resin, through eluent, obtains elutriant;
(9) extraction of chondroitin sulfate: the elutriant in step (8) is centrifugal through oxidative decoloration, ultrafiltration, alcohol precipitation, dehydration, drying obtains chondroitin sulfate.
2. a kind of technique extracting chondroitin sulfate in cartilage as claimed in claim 1, is characterized in that in described step (6), separation temperature is 6 ~ 9 DEG C.
3. a kind of technique extracting chondroitin sulfate in cartilage according to claim 1, it is characterized in that adopting ultrasonic cleaning in described step (1), scavenging solution is 0.01 ~ 0.06mol/LNaHCO
3solution.
4. a kind of technique extracting chondroitin sulfate in cartilage as claimed in claim 1, it is characterized in that adopting double enzymolysis technique in described step (3), wherein the add-on of 2709 proteolytic enzyme and the add-on of Pancreas Sus domestica are than being 1:10 ~ 15, are 0.1 ~ 0.9% of cartilage quality.
5. a kind of technique extracting chondroitin sulfate in cartilage as described in claim 1 or 4, it is characterized in that hydrolysis temperature in described step (3) is 45 DEG C ~ 55 DEG C, pH8.5 ~ 9.0, the treatment time is 4 ~ 8 hours.
6. a kind of technique extracting chondroitin sulfate in cartilage as claimed in claim 1, it is characterized in that heats up in described step (4) goes out after enzyme, also comprises pH4.0 ~ 4.5 time process 0.5 ~ 3 hour, then adjusts the treating processes of pH to 5.0 ~ 6.0.
7. a kind of technique extracting chondroitin sulfate in cartilage as claimed in claim 1, it is characterized in that heats up in described step (4) goes out after enzyme, also to comprise under pH4.3 process 2 hours, then adjusts pH to 5.4 to the treating processes of the obvious layering of enzymolysis solution.
8. a kind of technique extracting chondroitin sulfate in cartilage as claimed in claim 1, it is characterized in that described step (5) adopts disk centrifuge to be separated, operating parameters is disengaging time 1 ~ 3 hour, rotary drum rotating speed 6000 ~ 8000rpm/min, temperature 45 ~ 50 DEG C.
9., according to a kind of technique extracting chondroitin sulfate in cartilage according to claim 1, it is characterized in that the mass ratio of parting liquid and macroporous anion exchange resin in described step (6) is 1 ~ 1.5:1, adsorption time 1 ~ 2 hour.
10. extract chondroitin sulfate technique in cartilage according to one according to claim 1, it is characterized in that described cartilage is one or more in nose of an ox bone, ox larynx bone, crops bone, ox trunnion, ox moon dentale; Or elutriant described in described step 9 also has centrifugal treating technique after oxidative decoloration; Or alcohol precipitation process described in described step 9 is secondary alcohol precipitation; Or drying process described in described step 9 is vacuum-drying.
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| CN201510557844.0A CN105085714A (en) | 2015-09-02 | 2015-09-02 | Process for extracting chondroitin sulfate in cartilage |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108043353A (en) * | 2018-01-29 | 2018-05-18 | 湖南伍星生物科技有限公司 | A kind of chondroitin sulfate extractor |
| CN108148150A (en) * | 2018-01-29 | 2018-06-12 | 湖南伍星生物科技有限公司 | A kind of technique for shortening chondroitin sulfate extraction time |
| CN109502922A (en) * | 2018-12-29 | 2019-03-22 | 四川瑞宝生物科技股份有限公司 | A kind of gelatine wastewater processing method |
| CN114805635A (en) * | 2022-05-10 | 2022-07-29 | 滨海宇美科技有限公司 | Desalination protein powder and sodium chondroitin sulfate co-production process |
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| JP2005113106A (en) * | 2003-10-07 | 2005-04-28 | Ryota Katahira | Method for producing crude chondroitin sulfates |
| CN101348815A (en) * | 2008-09-03 | 2009-01-21 | 南京工业大学 | Process for preparing chondroitin sulfate by natural enzyme and protease synergistic method |
| CN104418957A (en) * | 2013-08-22 | 2015-03-18 | 青岛蓝农谷农产品研究开发有限公司 | Method for comprehensively utilizing cartilage to realize clean extraction of chondroitin sulfate |
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2015
- 2015-09-02 CN CN201510557844.0A patent/CN105085714A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2005113106A (en) * | 2003-10-07 | 2005-04-28 | Ryota Katahira | Method for producing crude chondroitin sulfates |
| CN101348815A (en) * | 2008-09-03 | 2009-01-21 | 南京工业大学 | Process for preparing chondroitin sulfate by natural enzyme and protease synergistic method |
| CN104418957A (en) * | 2013-08-22 | 2015-03-18 | 青岛蓝农谷农产品研究开发有限公司 | Method for comprehensively utilizing cartilage to realize clean extraction of chondroitin sulfate |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108043353A (en) * | 2018-01-29 | 2018-05-18 | 湖南伍星生物科技有限公司 | A kind of chondroitin sulfate extractor |
| CN108148150A (en) * | 2018-01-29 | 2018-06-12 | 湖南伍星生物科技有限公司 | A kind of technique for shortening chondroitin sulfate extraction time |
| CN109502922A (en) * | 2018-12-29 | 2019-03-22 | 四川瑞宝生物科技股份有限公司 | A kind of gelatine wastewater processing method |
| CN114805635A (en) * | 2022-05-10 | 2022-07-29 | 滨海宇美科技有限公司 | Desalination protein powder and sodium chondroitin sulfate co-production process |
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