CN104789545A - Bromelain purification method - Google Patents
Bromelain purification method Download PDFInfo
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- CN104789545A CN104789545A CN201410020389.6A CN201410020389A CN104789545A CN 104789545 A CN104789545 A CN 104789545A CN 201410020389 A CN201410020389 A CN 201410020389A CN 104789545 A CN104789545 A CN 104789545A
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- bromeline
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/48—Hydrolases (3) acting on peptide bonds (3.4)
- C12N9/50—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25)
- C12N9/63—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25) derived from plants
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y304/00—Hydrolases acting on peptide bonds, i.e. peptidases (3.4)
- C12Y304/22—Cysteine endopeptidases (3.4.22)
- C12Y304/22004—Bromelain (3.4.22.4)
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Abstract
The present invention relates to a bromelain purification method, which comprises (1) crude bromelain pre-treatment, (2) refining, and (3) freeze-drying. According to the present invention, the process steps are reasonable and simple, the method is suitable for large scale industrial production, the freezing is accurate and is easy to control, and the finished product enzyme activity is up to 8 million units/g; and the process has characteristics of simpleness, low energy consumption, and low cost.
Description
Technical field
The present invention relates to a kind of method of purification of pharmaceuticals, particularly a kind of method of purification of bromeline.
Background technology
Bromeline (English: Bromelain, also referred to as Traumanase) two kinds of materials can be meant, one of them is the proteolytic enzyme that narrow sense refers to squeeze out from pineapple section plant, and another is then the binding substances that broad sense refers to from other compositions extracts and these proteolytic enzyme.
Traumanase can be used in the field of many medical aspects.It is used in field of medicaments is for the first time nineteen fifty-seven, and now it is successfully used in and intercepts using of some inflammation metabolite (metabolite), and these metabolites originally can accelerate and worsen inflammatory response.Traumanase is a kind of medicine of anti-inflammatory, so be also used in sport injury, wound, sacroiliitis, with the redness of other kinds.Its main purposes be used in sport injury, intestines and stomach problem, phlebitis (phlebitis), sinusitis paranasal sinusitis and postoperative recovery treatment help above.The Traumanase of 200mg dosage has proved effective substitute of non-steroidal anti-inflammatory anodyne.Traumanase is also used in the treatment of the disease symptomses such as sacroiliitis, chronic venous varicose (venous insufficiency), slight ecchymosis, gout, hemorrhoid, dysmenorrhoea, autoimmune disorders (autoimmunedisease) and ulcerative colitis (ulcerative colitis).Research display Traumanase is also effective above thrombus in blood reducing hematoblastic excess buildup and reducing, the particularly blood of artery.Traumanase may for treatment human immunodeficiency virus disease effect to some extent.The Traumanase mixture of volume production can be used in the treatment of third stage burn, and the burn treatment of more high-level is then audited.
In existing bromeline production technique, have multiple to the purification process of bromeline, wherein vacuum-freezing process can reduce the enzyme loss alive of bromeline, but refrigerating process difficulty controls at present, and enzyme activity is not high.
Summary of the invention
The object of the invention is to solve above-mentioned deficiency, a kind of method of purification of bromeline is provided.
The technical scheme realizing the object of the invention is: a kind of method of purification of bromeline, comprises the pre-treatment of (1) bromeline crude product; (2) refining; (3) step of freeze-drying.
The method of purification of above-mentioned bromeline, described step (1) is specially: the bromeline crude product of recipe quantity is crossed 14 mesh sieves, obtains bromeline crude product particle; Take the purified water of 1/4 recipe quantity, add Sodium Benzoate and the EDTA-2Na of recipe quantity, stirring and dissolving, add above-mentioned bromeline crude product particle, stir to obtain mixture; Mixture colloidal mill is ground, obtains suspension; The purified water that takes a morsel washing plant and container, washings is incorporated in suspension, proceeds in treatment tank.
The method of purification of above-mentioned bromeline, described step (2) is refining to be specially: in treatment tank, add residue recipe quantity purified water, stirring reaction 1 hour; Added by the ammonium sulfate of recipe quantity in above-mentioned suspension, stir and saltout completely, liquid centrifuge of saltouing, obtains bromeline wet product.
The method of purification of above-mentioned bromeline, described step (3) freeze-drying is specially: bromeline wet product uniform particles loaded in material disc, every disc thickness is no more than 15mm; Material disc is placed on the shelf successively; By the Freeze Drying Equipment freeze-drying of bromeline wet product particle; Start freeze-drying after material inlet: goods pre-freezing temperature all below-30 DEG C after, condenser temperature is down to less than-50 DEG C, start vacuum pump evacuation, treat front case vacuum be evacuated to below 60Pa start heat up, enter sublimation stage; In sublimation process, the setting of thermal oil temperature is to heat up 5 DEG C per half an hour, and evenly rise to 22 DEG C, regulate vacuum simultaneously by aeration, control between 20Pa-60Pa, condenser temperature controls all the time below-50 DEG C; Close aeration when two products temperatures all reach 22 DEG C, by vacuum degree control at below 10Pa, make two products temperatures within 30 minutes ~ 60 minutes, can shut down discharging 18 DEG C ~ 22 DEG C insulations, product dried terminates, discharging, obtains bromeline finished product.
The method of purification of above-mentioned bromeline, described step (3) is if close aeration in freeze-drying operation, when products temperature is all lower than 18 DEG C, by heat conductive oil inlet temperature adjusting, make products temperature 18 DEG C ~ 22 DEG C insulations, control single goods top temperature in process all below 25 DEG C, thermal oil top temperature controls below 35 DEG C.
The method of purification of above-mentioned bromeline; aeration is closed when two products temperatures all reach 22 DEG C in described step (3) freeze-drying operation; by vacuum degree control at below 10Pa; two products temperatures are made within 45 minutes ~ 60 minutes, can discharging to be shut down 20 DEG C ~ 22 DEG C insulations; product dried terminates; discharging, obtains bromeline finished product.
The method of purification of above-mentioned bromeline; preferably aeration is closed when two products temperatures all reach 22 DEG C in described step (3) freeze-drying operation; by vacuum degree control at below 10Pa; two products temperatures are made within 60 minutes, can discharging to be shut down 22 DEG C of insulations; product dried terminates; discharging, obtains bromeline finished product.
Beneficial effect of the present invention: present invention process step is rationally simple, is applicable to large-scale commercial production, freezing being accurately easy to controls, finished product enzyme activity Da Gaoda 8,000,000 units/gram made; (2) technique is simple and easy, and energy consumption is little, and cost is low.
Embodiment
(embodiment 1)
The bromeline crude product of recipe quantity is crossed 14 mesh sieves, obtains bromeline crude product particle; Take the purified water of 1/4 recipe quantity, add Sodium Benzoate and the EDTA-2Na of recipe quantity, stirring and dissolving, add above-mentioned bromeline crude product particle, stir to obtain mixture; Mixture colloidal mill is ground, obtains suspension; The purified water that takes a morsel washing plant and container, washings is incorporated in suspension, proceeds in treatment tank.Then in treatment tank, residue recipe quantity purified water is added, stirring reaction 1 hour; Added by the ammonium sulfate of recipe quantity in above-mentioned suspension, stir and saltout completely, liquid centrifuge of saltouing, obtains bromeline wet product.Bromeline wet product uniform particles loaded in material disc, every disc thickness is no more than 15mm; Material disc is placed on the shelf successively; By the Freeze Drying Equipment freeze-drying of bromeline wet product particle; Start freeze-drying after material inlet: goods pre-freezing temperature all below-30 DEG C after, condenser temperature is down to less than-50 DEG C, start vacuum pump evacuation, treat front case vacuum be evacuated to below 60Pa start heat up, enter sublimation stage; In sublimation process, the setting of thermal oil temperature is to heat up 5 DEG C per half an hour, and evenly rise to 22 DEG C, regulate vacuum simultaneously by aeration, control at 20Pa, condenser temperature controls all the time below-50 DEG C; Aeration is closed when two products temperatures all reach 22 DEG C; by vacuum degree control at below 10Pa; two products temperatures are made within 60 minutes, can discharging to be shut down 18 DEG C of insulations; if closedown aeration; when products temperature is all lower than 18 DEG C, by heat conductive oil inlet temperature adjusting, make products temperature 18 DEG C ~ 22 DEG C insulations; control single goods top temperature in process all below 25 DEG C, thermal oil top temperature controls below 35 DEG C.Product dried terminates, discharging, obtains bromeline finished product.
(embodiment 2)
The bromeline crude product of recipe quantity is crossed 14 mesh sieves, obtains bromeline crude product particle; Take the purified water of 1/4 recipe quantity, add Sodium Benzoate and the EDTA-2Na of recipe quantity, stirring and dissolving, add above-mentioned bromeline crude product particle, stir to obtain mixture; Mixture colloidal mill is ground, obtains suspension; The purified water that takes a morsel washing plant and container, washings is incorporated in suspension, proceeds in treatment tank.Then in treatment tank, residue recipe quantity purified water is added, stirring reaction 1 hour; Added by the ammonium sulfate of recipe quantity in above-mentioned suspension, stir and saltout completely, liquid centrifuge of saltouing, obtains bromeline wet product.Bromeline wet product uniform particles loaded in material disc, every disc thickness is no more than 15mm; Material disc is placed on the shelf successively; By the Freeze Drying Equipment freeze-drying of bromeline wet product particle; Start freeze-drying after material inlet: goods pre-freezing temperature all below-30 DEG C after, condenser temperature is down to less than-50 DEG C, start vacuum pump evacuation, treat front case vacuum be evacuated to below 60Pa start heat up, enter sublimation stage; In sublimation process, the setting of thermal oil temperature is to heat up 5 DEG C per half an hour, and evenly rise to 22 DEG C, regulate vacuum simultaneously by aeration, control between 40Pa, condenser temperature controls all the time below-50 DEG C; Aeration is closed when two products temperatures all reach 22 DEG C; by vacuum degree control at below 10Pa; two products temperatures are made within 45 minutes, can discharging to be shut down 20 DEG C of insulations; if closedown aeration; when products temperature is all lower than 18 DEG C, by heat conductive oil inlet temperature adjusting, make products temperature 22 DEG C of insulations; control single goods top temperature in process all below 25 DEG C, thermal oil top temperature controls below 35 DEG C.Product dried terminates, discharging, obtains bromeline finished product.
(embodiment 3)
The bromeline crude product of recipe quantity is crossed 14 mesh sieves, obtains bromeline crude product particle; Take the purified water of 1/4 recipe quantity, add Sodium Benzoate and the EDTA-2Na of recipe quantity, stirring and dissolving, add above-mentioned bromeline crude product particle, stir to obtain mixture; Mixture colloidal mill is ground, obtains suspension; The purified water that takes a morsel washing plant and container, washings is incorporated in suspension, proceeds in treatment tank.Then in treatment tank, residue recipe quantity purified water is added, stirring reaction 1 hour; Added by the ammonium sulfate of recipe quantity in above-mentioned suspension, stir and saltout completely, liquid centrifuge of saltouing, obtains bromeline wet product.Bromeline wet product uniform particles loaded in material disc, every disc thickness is no more than 15mm; Material disc is placed on the shelf successively; By the Freeze Drying Equipment freeze-drying of bromeline wet product particle; Start freeze-drying after material inlet: goods pre-freezing temperature all below-30 DEG C after, condenser temperature is down to less than-50 DEG C, start vacuum pump evacuation, treat front case vacuum be evacuated to below 60Pa start heat up, enter sublimation stage; In sublimation process, the setting of thermal oil temperature is to heat up 5 DEG C per half an hour, and evenly rise to 22 DEG C, regulate vacuum simultaneously by aeration, control between 20Pa-60Pa, condenser temperature controls all the time below-50 DEG C;
Aeration is closed when two products temperatures all reach 22 DEG C; by vacuum degree control at below 10Pa; two products temperatures are made within 50 minutes, can discharging to be shut down 2022 DEG C of insulations; if closedown aeration; when products temperature is all lower than 18 DEG C, by heat conductive oil inlet temperature adjusting, make products temperature 18 DEG C ~ 22 DEG C insulations; control single goods top temperature in process all below 25 DEG C, thermal oil top temperature controls below 35 DEG C.Product dried terminates, discharging, obtains bromeline finished product.
(embodiment 4)
The bromeline crude product of recipe quantity is crossed 14 mesh sieves, obtains bromeline crude product particle; Take the purified water of 1/4 recipe quantity, add Sodium Benzoate and the EDTA-2Na of recipe quantity, stirring and dissolving, add above-mentioned bromeline crude product particle, stir to obtain mixture; Mixture colloidal mill is ground, obtains suspension; The purified water that takes a morsel washing plant and container, washings is incorporated in suspension, proceeds in treatment tank.Then in treatment tank, residue recipe quantity purified water is added, stirring reaction 1 hour; Added by the ammonium sulfate of recipe quantity in above-mentioned suspension, stir and saltout completely, liquid centrifuge of saltouing, obtains bromeline wet product.Bromeline wet product uniform particles loaded in material disc, every disc thickness is no more than 15mm; Material disc is placed on the shelf successively; By the Freeze Drying Equipment freeze-drying of bromeline wet product particle; Start freeze-drying after material inlet: goods pre-freezing temperature all below-30 DEG C after, condenser temperature is down to less than-50 DEG C, start vacuum pump evacuation, treat front case vacuum be evacuated to below 60Pa start heat up, enter sublimation stage; In sublimation process, the setting of thermal oil temperature is to heat up 5 DEG C per half an hour, and evenly rise to 22 DEG C, regulate vacuum simultaneously by aeration, control between 20Pa-60Pa, condenser temperature controls all the time below-50 DEG C; Aeration is closed when two products temperatures all reach 22 DEG C; by vacuum degree control at below 10Pa; two products temperatures are made within 60 minutes, can discharging to be shut down 22 DEG C of insulations; if closedown aeration; when products temperature is all lower than 18 DEG C, by heat conductive oil inlet temperature adjusting, make products temperature 18 DEG C ~ 22 DEG C insulations; control single goods top temperature in process all below 25 DEG C, thermal oil top temperature controls below 35 DEG C.Product dried terminates, discharging, obtains bromeline finished product.
Above-described specific embodiment; object of the present invention, technical scheme and beneficial effect are further described; be understood that; the foregoing is only specific embodiments of the invention; be not limited to the present invention; within the spirit and principles in the present invention all, any amendment made, equivalent replacement, improvement etc., all should be included within protection scope of the present invention.
Claims (7)
1. a method of purification for bromeline, is characterized in that: comprise the pre-treatment of (1) bromeline crude product; (2) refining; (3) step of freeze-drying.
2. the method for purification of bromeline according to claim 1, is characterized in that: described step (1) is specially: the bromeline crude product of recipe quantity is crossed 14 mesh sieves, obtains bromeline crude product particle; Take the purified water of 1/4 recipe quantity, add Sodium Benzoate and the EDTA-2Na of recipe quantity, stirring and dissolving, add above-mentioned bromeline crude product particle, stir to obtain mixture; Mixture colloidal mill is ground, obtains suspension; The purified water that takes a morsel washing plant and container, washings is incorporated in suspension, proceeds in treatment tank.
3. the method for purification of bromeline according to claim 1, is characterized in that: described step (2) is refining to be specially: in treatment tank, add residue recipe quantity purified water, stirring reaction 1 hour; Added by the ammonium sulfate of recipe quantity in above-mentioned suspension, stir and saltout completely, liquid centrifuge of saltouing, obtains bromeline wet product.
4. the method for purification of bromeline according to claim 1, is characterized in that: described step (3) freeze-drying is specially: bromeline wet product uniform particles loaded in material disc, every disc thickness is no more than 15mm; Material disc is placed on the shelf successively; By the Freeze Drying Equipment freeze-drying of bromeline wet product particle; Start freeze-drying after material inlet: goods pre-freezing temperature all below-30 DEG C after, condenser temperature is down to less than-50 DEG C, start vacuum pump evacuation, treat front case vacuum be evacuated to below 60Pa start heat up, enter sublimation stage; In sublimation process, the setting of thermal oil temperature is to heat up 5 DEG C per half an hour, and evenly rise to 22 DEG C, regulate vacuum simultaneously by aeration, control between 20Pa-60Pa, condenser temperature controls all the time below-50 DEG C; Close aeration when two products temperatures all reach 22 DEG C, by vacuum degree control at below 10Pa, make two products temperatures within 30 minutes ~ 60 minutes, can shut down discharging 18 DEG C ~ 22 DEG C insulations, product dried terminates, discharging, obtains bromeline finished product.
5. the method for purification of bromeline according to claim 4, it is characterized in that: described step (3) is if close aeration in freeze-drying operation, when products temperature is all lower than 18 DEG C, by heat conductive oil inlet temperature adjusting, make products temperature 18 DEG C ~ 22 DEG C insulations, control single goods top temperature in process all below 25 DEG C, thermal oil top temperature controls below 35 DEG C.
6. the method for purification of bromeline according to claim 4; it is characterized in that: in described step (3) freeze-drying operation, close aeration when two products temperatures all reach 22 DEG C; by vacuum degree control at below 10Pa; two products temperatures are made within 45 minutes ~ 60 minutes, can discharging to be shut down 20 DEG C ~ 22 DEG C insulations; product dried terminates; discharging, obtains bromeline finished product.
7. the method for purification of bromeline according to claim 4; it is characterized in that: in described step (3) freeze-drying operation, close aeration when two products temperatures all reach 22 DEG C; by vacuum degree control at below 10Pa; two products temperatures are made within 60 minutes, can discharging to be shut down 22 DEG C of insulations; product dried terminates; discharging, obtains bromeline finished product.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410020389.6A CN104789545A (en) | 2014-01-16 | 2014-01-16 | Bromelain purification method |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410020389.6A CN104789545A (en) | 2014-01-16 | 2014-01-16 | Bromelain purification method |
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| CN104789545A true CN104789545A (en) | 2015-07-22 |
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| CN201410020389.6A Pending CN104789545A (en) | 2014-01-16 | 2014-01-16 | Bromelain purification method |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108741099A (en) * | 2018-06-11 | 2018-11-06 | 广州尚天诚品生物科技有限公司 | One kind contributing to comfortable alimentation composition in joint and its preparation method and application |
| CN115637211A (en) * | 2022-09-05 | 2023-01-24 | 祝雪花 | Ficin extraction equipment and use method thereof |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1131159A (en) * | 1994-09-30 | 1996-09-18 | 阎隆飞 | Bromelin decrustation preparation and its preparing method |
| CN101319208A (en) * | 2008-04-08 | 2008-12-10 | 徐闻县美仑生物制品有限公司 | Method for preparation of bromelin |
| CN103305492A (en) * | 2013-06-09 | 2013-09-18 | 诸辉 | Method for extracting high-purity pharmaceutical grade bromelain |
-
2014
- 2014-01-16 CN CN201410020389.6A patent/CN104789545A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1131159A (en) * | 1994-09-30 | 1996-09-18 | 阎隆飞 | Bromelin decrustation preparation and its preparing method |
| CN101319208A (en) * | 2008-04-08 | 2008-12-10 | 徐闻县美仑生物制品有限公司 | Method for preparation of bromelin |
| CN103305492A (en) * | 2013-06-09 | 2013-09-18 | 诸辉 | Method for extracting high-purity pharmaceutical grade bromelain |
Non-Patent Citations (1)
| Title |
|---|
| 王燕等: "菠萝蛋白酶工业化提取工艺的改良及酶学性质研究", 《食品工业科技》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108741099A (en) * | 2018-06-11 | 2018-11-06 | 广州尚天诚品生物科技有限公司 | One kind contributing to comfortable alimentation composition in joint and its preparation method and application |
| CN115637211A (en) * | 2022-09-05 | 2023-01-24 | 祝雪花 | Ficin extraction equipment and use method thereof |
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Application publication date: 20150722 |