CN105820214A - Method for separating polymyxin E fermentation liquid by multistage ceramic membrane microfiltration - Google Patents
Method for separating polymyxin E fermentation liquid by multistage ceramic membrane microfiltration Download PDFInfo
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- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
- C07K7/50—Cyclic peptides containing at least one abnormal peptide link
- C07K7/54—Cyclic peptides containing at least one abnormal peptide link with at least one abnormal peptide link in the ring
- C07K7/60—Cyclic peptides containing at least one abnormal peptide link with at least one abnormal peptide link in the ring the cyclisation occurring through the 4-amino group of 2,4-diamino-butanoic acid
- C07K7/62—Polymyxins; Related peptides
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Abstract
The invention discloses a method for separating a polymyxin E fermentation liquid by multistage ceramic membrane microfiltration. The method comprises the following steps: 1) acidizing the polymyxin E fermentation liquid; and 2) sequentially passing the acidized polymyxin E fermentation liquid through the multistage ceramic membrane microfiltration system to perform microfiltration separation, wherein the pore sizes of the ceramic membranes in the ceramic membrane microfiltration system gradually decrease along the polymyxin E fermentation liquid microfiltration separation direction. The method has the advantages of mild conditions and low worker labor intensity, reduces the separation process steps and enhances the separation recovery yield.
Description
Technical field
The present invention relates to the separation of polymyxin E fermentation liquid, in particular it relates to use the method that multistage microfiltration of ceramic membrane separates polymyxin E fermentation liquid.
Background technology
Polymyxin E, has another name called colistin sulfate (colistin, polymyxinsulfate), is by alkalescence ring type polypeptide class antibiotic produced by aerobacillus polymyxa Donker (Bacilluspolymyxa).Polymyxin E is amorphous white powder shape, high temperature resistant, soluble in water, stable in properties, can effectively suppress the propagation of all kinds of gram negative bacteria, notable to causing the effects such as the bacillus pyocyaneus of animal feeding disease, Salmonella, dysentery bacterium, aerobacteria and pneumobacillus especially.Polymyxin E can be attached to bacterium surface, hydrophobic functional groups therein can be combined with the lipoprotein phosphate radical in bacterial cell membrane, destroy the integrity of adventitia, whole membrane passage is made to increase, causing the little molecular chaperones such as intracytoplasmic phosphoric acid, nucleoside to escape, bacterial growth is suppressed the most lethal dying.Polymyxin E all has good effect to the antibacterial of trophophase and resting stage, and is difficult to cause antibacterial to produce drug resistance.It can be effectively prevented the infectious intestinal tract disease caused by gram negative bacteria, and the domestic animal such as chicken and ducks in poultry, cattle pig are had obvious prophylactic treatment effect.Meanwhile, this product is little to domestic animal avian physiological toxic and side effects, is difficult to digested road and absorbs, and can be obviously promoted poultry growth and improve efficiency of feed utilization, being current safest poultry growth promotion antibiotic.
The mode of industrial usual employing separation and Extraction bacillus polymyxa fermentation liquid prepares polymyxin E, and typical process flow is: fermentation liquid acidifying, plate-and-frame filtration remove impurity, ion exchange resin extraction, nanofiltration concentrate, are spray-dried finished product.So that the plate-and-frame filtration impurity removal process cycle is tediously long, production efficiency is low, labor intensity is big in above-mentioned steps, and impurity removal is incomplete, and product separation yield is low, has a strong impact on the efficiency of subsequent process steps.
The patent of publication number CN101974075 discloses a kind of with extracting polymyxin B, the method for E from fermentation technique culture, under the conditions of 25 DEG C, fermentation technique culture is stirred 30 minutes after alkaline matter regulation pH to 10.0-14.0,3000 rotating speed centrifugations are collected centrifugal thing and are obtained polymyxin E free alkali precipitation, add acid solution regulation pH value and make polymyxin E resolution of precipitate to 3.0-4.0, acquire the concentrated solution containing polymyxin E by sucking filtration lysate.The method remove impurity is incomplete, and product purity is low, and polymyxin E precipitation in different pH values solution and dissolving cause product to lose in a large number, and yield is relatively low, and consumes a large amount of acidic and alkaline waste water in technique, and treatment cost is high, is unfavorable for industrialization promotion.
The patent of publication number CN103059105B discloses the extracting method of a kind of colistin sulfate, polymyxin E fermentation liquid adds sulfur acid for adjusting pH to 3.9-4.1, the preliminary remove impurity of via hole diameter 500 nano ceramics membrane filtration, after filtrate adjusts pH4.5-5, then purified by further remove impurity of step such as Continuous Moving Bed absorption parsings.The method is tentatively filtered by large aperture ceramic membrane, and rapidly and efficiently large granular impurity in coarse filtration fermentation liquid, and yield is high.But 500 nano apertures cannot retain numerous impurity such as bacterial chip, macromole particle, has a strong impact on purity and the quality of polymyxin E product, it is therefore desirable to use complicated loaded down with trivial details Continuous Moving Bed absorption to resolve the further remove impurity of technique, purification.The subsequent handlings such as the Continuous Moving Bed absorption parsing in the method produce a large amount of spent acid, salkali waste, and power consumption water consumption.
The patent of publication number CN102718842B discloses the technique that a kind of sedimentation method extract colistin sulfate, fermentation liquid oxalic acid containing polymyxin E or concentrated sulphuric acid are regulated pH value to 2.0-5.0, filter through 50-200 nano ceramics membrane micropore, after filtrate is sequentially added into metal chelating agent and inorganic flocculating agent normal-temperature reaction, it is again heated to 70-95 DEG C and is incubated 0.5-2.0 hour, subsequently through cooling, sucking filtration, make impurity be precipitated out rapidly, thus reach the purpose of remove impurity.The method optimizes extraction process, it is thus achieved that product purity is high, quality is excellent, but adds the processing step that multiple tracks is numerous and diverse.This invention uses the ceramic membrane microporous filter retaining a diameter of 50-200 nanometer, it is possible to the dreg in removing fermentation liquid, bacterial chip effectively, but cannot remove the particle in filtrate and the organic polymer impurity such as albumen, pigment.In order to remove this kind of impurity further, in this invention, the interpolation of metal chelating agent and flocculant considerably increases production cost, and the follow-up still use alkalization sedimentation method extract polymyxin E, add the process discharge of alkaline waste water, are unfavorable for environmental conservation.At present separating technology is disadvantageous in that: lack the separation method of the polymyxin E fermentation liquid of a kind of processing step refine, easy and simple to handle, with low cost, Environmental Safety.
Summary of the invention
It is an object of the invention to provide a kind of method using multistage microfiltration of ceramic membrane to separate polymyxin E fermentation liquid, the method mild condition, staff labor intensity is low, decreases separate process steps, improves separation and recovery yield.
To achieve these goals, the invention provides a kind of method using multistage microfiltration of ceramic membrane to separate polymyxin E fermentation liquid, the method includes:
1) polymyxin E fermentation liquid is carried out acidification；
2) the polymyxin E fermentation liquid after acidification is sequentially passed through multistage microfiltration of ceramic membrane system and carry out microfiltration separation；
Wherein, the direction that the microfiltration along polymyxin E fermentation liquid separates, the aperture of the ceramic membrane in microfiltration of ceramic membrane system is gradually reduced.
Pass through technique scheme, the present invention use multistage ceramic membrane device work continuously, wherein, the bigger microfiltration of ceramic membrane system in aperture uses and retains more greatly pore size ceramic membrane element, it is mainly used in filtering the bulky grain thing such as dreg, bacterial chip, and polymyxin E product and albumen, pigment, all kinds of ion can freely pass through；The wide-aperture rate of filtration that can increase fermentation liquid that arranges, improves dust removal rate, shortens operation duration；And decrease the operation pressure of the microfiltration systems less to aperture.The microfiltration systems employing nanoscale pore size ceramic membrane that aperture is less, the filtrate of upper level is separated again, in filtrate, the organic polymer impurity such as high molecular weight protein of more than 60kDa molecular weight will not pass through ceramic membrane and is retained down, quickly it is effectively improved filtrate quality, the convenient step simplifying subsequent product Hydrolysis kinetics.
Present invention process and existing sedimentation method technique comparison have the advantage that the multistage microfiltration of ceramic membrane system of employing increases substantially process efficiency and the clarity of filtrate and quality；Multistage microfiltration of ceramic membrane system uses difference to retain pore size ceramic membrane unit, the big small impurities of Selective Separation different molecular weight, be conducive to reducing the impurity pressure to next stage level ceramic membrane nano aperture, increase the membrane flux of filtrate, shorten the cycle of separation of fermentative broth operation.The present invention, without using the additive such as alkali liquor, inorganic flocculating agent, decreases production cost and waste water and dregs pollution problem；Do without the multiple tracks separation circuits such as heating, cooling and sucking filtration, greatly reduced the loss of polymyxin E product, reduce manual labor's intensity, energy efficient, effectively shorten the production cycle.Ceramic membrane separation device operating automation degree is high, cleans simple efficient, and beneficially persistence produces.
Present invention optimizes separating technology, decrease that the step generally existed in existing separation of fermentative broth technique is lengthy and jumbled, operation duration, cannot continuously, hand labor intensity is big and consumes a large amount of soda acid and produces the problems such as contaminated wastewater environment, the present invention is greatly simplified separating step simultaneously, effectively reduce the product loss that separation process causes, improve the recovery yield of product.The present invention embodies environmental protection, safe efficient, the new concept of low energy consumption, has bigger application value.
Other features and advantages of the present invention will be described in detail in detailed description of the invention part subsequently.
Detailed description of the invention
Hereinafter the detailed description of the invention of the present invention is described in detail.It should be appreciated that detailed description of the invention described herein is merely to illustrate and explains the present invention, it is not limited to the present invention.
The invention provides a kind of method using multistage microfiltration of ceramic membrane to separate polymyxin E fermentation liquid, the method includes:
1) polymyxin E fermentation liquid is carried out acidification；
2) the polymyxin E fermentation liquid after acidification is sequentially passed through multistage microfiltration of ceramic membrane system and carry out microfiltration separation；
Wherein, the direction that the microfiltration along polymyxin E fermentation liquid separates, the aperture of the ceramic membrane in microfiltration of ceramic membrane system is gradually reduced.
In the present invention, the progression of multistage microfiltration of ceramic membrane system can select in wide scope, although in the case of progression is more, clarity and the quality of filtrate can be improved further, but consider from production cycle and production cost, preferably, the progression of multistage microfiltration of ceramic membrane system is 2-3 level.More preferably, the direction separated along the microfiltration of polymyxin E fermentation liquid, multistage microfiltration of ceramic membrane system is made up of first order microfiltration of ceramic membrane system and second level microfiltration of ceramic membrane system successively, wherein, containing the first ceramic membrane in first order microfiltration of ceramic membrane system, containing the second ceramic membrane in the microfiltration of ceramic membrane system of the second level.
In above-mentioned two-stage microfiltration of ceramic membrane system, the aperture of the first ceramic membrane and the second ceramic membrane can select in wide scope, but so that first order microfiltration of ceramic membrane system can effective filter out the bulky grain thing such as dreg, bacterial chip, and polymyxin E product and albumen, pigment, all kinds of ion can freely pass through；But simultaneously so that the filtrate of first order microfiltration of ceramic membrane system can the less operation pressure to second level microfiltration of ceramic membrane system, operation pressure has the most important impact to clarity and the quality of filtrate, this is that inventor is obtained by substantial amounts of experiment, therefore, preferably, the aperture of the first ceramic membrane is 40-60nm, and the aperture of the second ceramic membrane is 4-6nm.
Simultaneously, first ceramic membrane and/or the second ceramic membrane can be single layer structures in the present invention, it can also be multiple structure, in order to improve the microfiltration effect of microfiltration of ceramic membrane system further, preferably, the first ceramic membrane and/or the second ceramic membrane use multiple structure, more preferably, first ceramic membrane is made up of 5-7 ceramic membrane element, and the second ceramic membrane is made up of 3-5 ceramic membrane element.
In addition, the concrete material of ceramic membrane element can select in wide scope, but in order to improve clarity and the quality of the polymyxin E product after microfiltration separates further, it is preferable that the composition material of ceramic membrane element is one or more in aluminium oxide, titanium oxide and zirconium oxide.
In the first order microfiltration of ceramic membrane system of the present invention, although service condition can be configured by concrete needs, but inventor find service condition select have the most important impact for clarity and the quality of polymyxin E product, preferably, in first order microfiltration of ceramic membrane system, service condition at least meets following condition: running temperature is not higher than 35 DEG C, and operating pressure is 0.4-0.6MPa, and membrane flux is 50-150L/m2·h；It is highly preferred that operating pressure is 0.4-0.5MPa, membrane flux is 120-130L/m2·h。
In like manner, in the second level microfiltration of ceramic membrane system of the present invention, although service condition can also be configured by concrete needs, but inventor find service condition select also have the most important impact for clarity and the quality of polymyxin E product, preferably, in the microfiltration of ceramic membrane system of the second level, service condition at least meets following condition: running temperature is not higher than 35 DEG C, operating pressure is 0.5-1.5MPa, and membrane flux is 40-60L/m2·h；It is highly preferred that operating pressure is 0.8-0.9MPa, membrane flux is 40-60L/m2·h。
On the basis of the above, through step 1) pH of polymyxin E fermentation liquid after acidifying can select in wide scope, but so that polymyxin E can resolution of precipitate effectively, preferably, after acidified process, the pH of polymyxin E fermentation liquid is 2.0-5.5.Wherein, during acidification, the selection of acid can be the conventional selection of this area, but considers from acidizing effect, it is preferable that acidification is adjusted by concentrated sulphuric acid or oxalic acid.
Hereinafter will be described the present invention by embodiment.
Embodiment 1
1) with containing the fermentation liquid 2000L that polymyxin E concentration is 60.01 ten thousand U/mL as raw material, import souring tank and add sulfur acid for adjusting pH value to 2.7, and continuously stirred 50min (40-60min).
2) being separated by first order microfiltration of ceramic membrane system by the fermentation liquid being acidified, the ceramic membrane using 6 layers of aperture to be 50 nanometers, membrane flux is 120L/m2H, running temperature 30 DEG C, operating pressure is 0.4MPa, obtains first order filtrate 4800L, and polymyxin E concentration is 24.85 ten thousand U/mL.
4800L first order filtrate being passed through second level microfiltration of ceramic membrane system separate, the ceramic membrane using 4 layers of aperture to be 5 nanometers, membrane flux is 40L/m2H, running temperature 32 DEG C, operating pressure is 0.8MPa, obtains second level filtrate 10000L, and polymyxin E concentration is 11.9 ten thousand U/mL, and filtrate is clarified, and ceramic membrane yield is 99.1%.
Embodiment 2
1) with containing the fermentation liquid 2000L that polymyxin E concentration is 65.23 ten thousand U/mL as raw material, import souring tank and add sulfur acid for adjusting pH value to 2.7, and continuously stirred 50min (40-60min).
2) being separated by first order microfiltration of ceramic membrane system by the fermentation liquid being acidified, the ceramic membrane using 6 layers of aperture to be 50 nanometers, membrane flux is 130L/m2H, running temperature 31 DEG C, operating pressure is 0.4MPa, obtains first order filtrate 4600L, and polymyxin E concentration is 28.21 ten thousand U/mL.
4600L first order filtrate being passed through second level microfiltration of ceramic membrane system separate, the ceramic membrane using 4 layers of aperture to be 5 nanometers, membrane flux is 40L/m2H, running temperature 32 DEG C, operating pressure is 0.9MPa, obtains second level filtrate 9800L, and polymyxin E concentration is 13.1 ten thousand U/mL, and filtrate is clarified, and ceramic membrane yield is 99.2%.
Embodiment 3
1) with containing the fermentation liquid 2000L that polymyxin E concentration is 62.55 ten thousand U/mL as raw material, import souring tank and add sulfur acid for adjusting pH value to 2.7, and continuously stirred 50min (40-60min).
2) being separated by first order microfiltration of ceramic membrane system by the fermentation liquid being acidified, the ceramic membrane using 6 layers of aperture to be 50 nanometers, membrane flux is 120L/m2H, running temperature 32 DEG C, operating pressure is 0.5MPa, obtains first order filtrate 4600L, and polymyxin E concentration is 27.03 ten thousand U/mL.
4600L first order filtrate being passed through second level microfiltration of ceramic membrane system separate, the ceramic membrane using 4 layers of aperture to be 5 nanometers, membrane flux is 40L/m2H, running temperature 33 DEG C, operating pressure is 0.9MPa, obtains second level filtrate 10500L, and polymyxin E concentration is 11.80 ten thousand U/mL, and filtrate is clarified, and ceramic membrane yield is 99.1%.
Embodiment 4
Carrying out according to the method for embodiment 1, ceramic membrane yield is 98.5%, except that the service condition of first order microfiltration of ceramic membrane system is: membrane flux is 50L/m2·h。
Embodiment 5
Carrying out according to the method for embodiment 1, ceramic membrane yield is 99.2%, except that the service condition of first order microfiltration of ceramic membrane system is: membrane flux is 150L/m2H, operating pressure is 0.6MPa.
Embodiment 6
Carrying out according to the method for embodiment 1, ceramic membrane yield is 99.0%, except that the service condition of second level microfiltration of ceramic membrane system is: membrane flux is 60L/m2H, operating pressure is 1.5MPa.
Embodiment 7
Carrying out according to the method for embodiment 1, ceramic membrane yield is 99.0%, except that the service condition of second level microfiltration of ceramic membrane system is: membrane flux is 40L/m2H, operating pressure is 0.5MPa.
Embodiment 8
Carrying out according to the method for embodiment 1, ceramic membrane yield is 99.2%, except that the pH of the fermentation liquid being acidified is 2.0.
Embodiment 9
Carrying out according to the method for embodiment 1, ceramic membrane yield is 98.4%, except that the pH of the fermentation liquid being acidified is 5.5.
By above example it is known that the method mild condition of present invention offer, staff labor intensity is low, and separate process steps is few, has the separation and recovery rate of excellence simultaneously
The preferred embodiment of the present invention described in detail above; but, the present invention is not limited to the detail in above-mentioned embodiment, in the technology concept of the present invention; technical scheme can be carried out multiple simple variant, these simple variant belong to protection scope of the present invention.
It is further to note that, each concrete technical characteristic described in above-mentioned detailed description of the invention, in the case of reconcilable, can be combined by any suitable means, in order to avoid unnecessary repetition, various possible compound modes are illustrated by the present invention the most separately.
Additionally, can also carry out combination in any between the various different embodiment of the present invention, as long as it is without prejudice to the thought of the present invention, it should be considered as content disclosed in this invention equally.
Claims (10)
1. one kind uses the method that multistage microfiltration of ceramic membrane separates polymyxin E fermentation liquid, it is characterised in that described method includes:
1) polymyxin E fermentation liquid is carried out acidification；
2) the described polymyxin E fermentation liquid after acidification is sequentially passed through multistage microfiltration of ceramic membrane system and carry out microfiltration separation；
Wherein, the direction that the microfiltration along described polymyxin E fermentation liquid separates, the aperture of the ceramic membrane in described microfiltration of ceramic membrane system is gradually reduced.
Method the most according to claim 1, wherein, the progression of described multistage microfiltration of ceramic membrane system is 2-3 level.
Method the most according to claim 2, wherein, the direction separated along the microfiltration of described polymyxin E fermentation liquid, described multistage microfiltration of ceramic membrane system is made up of first order microfiltration of ceramic membrane system and second level microfiltration of ceramic membrane system successively, wherein, containing the first ceramic membrane in first order microfiltration of ceramic membrane system, containing the second ceramic membrane in the microfiltration of ceramic membrane system of the second level.
Method the most according to claim 3, wherein, the aperture of described first ceramic membrane is 40-60nm, and the aperture of described second ceramic membrane is 4-6nm.
Method the most according to claim 4, wherein, described first ceramic membrane is made up of 5-7 ceramic membrane element, and described second ceramic membrane is made up of 3-5 ceramic membrane element.
Method the most according to claim 5, wherein, the composition material of described ceramic membrane element is one or more in aluminium oxide, titanium oxide and zirconium oxide.
7. according to the method described in any one in claim 3-6, wherein, in described first order microfiltration of ceramic membrane system, service condition at least meets following condition: running temperature is not higher than 35 DEG C, and operating pressure is 0.4-0.6MPa, and membrane flux is 50-150L/m2·h。
Method the most according to claim 7, wherein, in the microfiltration of ceramic membrane system of the described second level, service condition at least meets following condition: running temperature is not higher than 35 DEG C, and operating pressure is 0.5-1.5MPa, and membrane flux is 40-60L/m2·h。
Method the most according to claim 8, wherein, after acidified process, the pH of described polymyxin E fermentation liquid is 2.0-5.5.
Method the most according to claim 9, wherein, described acidification is adjusted by concentrated sulphuric acid or oxalic acid.
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| CN102351930A (en) * | 2011-11-02 | 2012-02-15 | 福建和泉生物科技有限公司 | Method for separating spiramycin fermentation liquid by ceramic membrane microfiltration separation technique |
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| CN111100823A (en) * | 2020-01-17 | 2020-05-05 | 黄石曼菲特生物科技有限公司 | Polymyxin B sulfate production strain, preparation method and application of polymyxin B sulfate |
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