CN101948399A - Method for continuously separating and purifying valine in fermentation liquor by using simulated moving bed chromatography - Google Patents
Method for continuously separating and purifying valine in fermentation liquor by using simulated moving bed chromatography Download PDFInfo
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- CN101948399A CN101948399A CN2010105028128A CN201010502812A CN101948399A CN 101948399 A CN101948399 A CN 101948399A CN 2010105028128 A CN2010105028128 A CN 2010105028128A CN 201010502812 A CN201010502812 A CN 201010502812A CN 101948399 A CN101948399 A CN 101948399A
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Abstract
The invention discloses a method for continuously separating and purifying valine in fermentation liquor by using simulated moving bed chromatography, and belongs to the technical field of biological product processing. In the method, a simulated moving bed chromatography separating device suitable for the composition characteristic of valine fermentation liquor is designed, water is taken as a mobile phase, and the previously invented resin special for valine separation (disclosed in the Chinese patent 200910231735.4) is taken as a stationary phase. The method can completely remove impurities of alanine and leucine from the valine fermentation liquor, achieves product purity of over 99.7 percent to meet the medicinal standard, is suitable for continuous production, has higher recovery rate and production strength, and overcomes the defects of low purity, low efficiency, high pollution and the like in the conventional valine separation and purification process.
Description
Technical field
A kind of method of using Xie Ansuan in the continuous separation and purification fermented liquid of simulated moving bed chromatography belongs to the processing technique field of biological products.
Background technology
The L-Xie Ansuan belongs to branched chain amino acid (branched chain amino acid, BCAA), it is one of essential amino acid, has different physiological roles, be widely used in the manufacturing of medicine, food and seasonings, animal-feed and makeup, the effect in medical research and treatment also comes into one's own day by day.
Under China scientific and technical personnel's effort, the upstream technology of China's fermentative Production Xie Ansuan also makes great progress, and has reached industrialized level substantially in recent years.But about 0.5% L-Ala and the leucine that produce in the Xie Ansuan fermenting process and its chemical property is close, separation difficulty.Valine content is low in the finished product that traditional separation methods such as ion-exchange are produced, and impurity is many, and major part only meets the quality standard of foodstuff additive, has restricted its price.
Wuxi Jinghai Amino Acid Co., Ltd. rather is good for and flies to wait the people is that the method for using membrane sepn to separate coupling extraction separation L-Xie Ansuan with industrial chromatography in CN101503366 " membrane sepn separates the method for coupling extraction separation L-Xie Ansuan with industrial chromatography " has higher yield and product purity in the patent No., but is to use shortcomings such as having efficient is low, time length, cost height.The Peng Qi equalization is in " a kind of method of separating the purification Xie Ansuan from Xie Ansuan liquid " of patent No. CN101168512, designed a kind of method of five-region simulated moving bed separation Xie Ansuan, but it is about 93% Xie Ansuan that this method only can obtain purity, can't obtain more highly purified Xie Ansuan, its metal-chelate resin that adopts is relatively poor to the specific adsorption of Xie Ansuan in addition, has caused the decline of separation purity.(simulated moving bed technology separates Xie Ansuan and L-Ala to people such as Wan Guihong, food and fermentation industries, 2005,31 (12), 50-53) reported that also a kind of simulated moving bed chromatography separates the method for Xie Ansuan, but need to adopt reagent such as ammoniacal liquor, can produce very big pollution to environment, and product purity is also lower.
Summary of the invention
The objective of the invention is to: at the problem that exists in the current Xie Ansuan sepn process, the present invention is moving phase with water, Xie Ansuan separate, dedicated resin with previous invention is a stationary phase, and designed the moving bed imitation chromatogram separation facility that is applicable to the valine fermentation liquid component characteristic, can remove impurity L-Ala and leucine in the valine fermentation liquid fully, product purity is reached more than 99.7%, reach medicinal standard, can realize serialization production, the higher rate of recovery and production intensity are arranged.
Technical scheme of the present invention: a kind of method of using Xie Ansuan in the continuous separation and purification fermented liquid of simulated moving bed chromatography, designed the moving bed imitation chromatogram separation facility that is applicable to the valine fermentation liquid component characteristic, with water is moving phase, with Xie Ansuan separate, dedicated resin is stationary phase, Xie Ansuan in the valine fermentation liquid is carried out continuous separation and purification, and step is as follows:
(1) designed open five-region simulated moving bed chromatographic separation device, this device has 5 districts, and each district is made up of the 1-3 root chromatogram column, and parallel-series connects, and the specification of all chromatographic columns is identical, and column diameter is 1-50cm, chromatographic column adopting chuck heat tracing; In the middle of per two pillars 3 outlets are arranged, 3 inlets, sepn process except distinguish 5 and distinguish 1 between have two entry and exit can use, has only entry and exit to use in the middle of other district/post, no entry and exit all are in closing condition; The direction in 4-district, 3-district, 2-district, 1-district 5, district is the liquid flow path direction, and 1-district, 2-district, 3-district, 4-district, 5-district 5, district is the post switching direction; Xie Ansuan dedicated separation resin is housed in the post; District 1 and the open architecture of distinguishing between 5 have guaranteed that filler is fully cleaned, thereby can prepare highly purified Xie Ansuan.Solid line is the liquid flow path direction in the accompanying drawing, and dotted line is the post switching direction, and the article that the ultimate principle of equipment operation can have been delivered referring to contriver before (simulated moving bed chromatography technology and application thereof, chromatogram, 2004,22 (2), 111-115).The Xie Ansuan dedicated separation resin of independent research is housed in the post, and Chinese patent 200910231735.4 is open, and this resin has very strong specific adsorption ability to Xie Ansuan.
(2) after the Xie Ansuan fermenting process finishes, with hydrochloric acid with the fermented liquid pH regulator to 2-5, centrifugal removal thalline, protein and polysaccharide impurity are then with the clear liquid activated carbon decolorizing that obtains;
(3) valine fermentation liquid after will decolouring concentrates at 60 ℃ of vacuum decompressions, and the Xie Ansuan mass concentration is 70-80 g/L in the control clear liquid; Perhaps fermented liquid directly separates without concentrating;
(4) clear liquid is put into the described moving bed imitation chromatogram separation facility of step (1) and separated, obtain highly purified Xie Ansuan solution 1, obtain solid Xie Ansuan finished product through concentrated, crystallization again from exporting; Moving phase is water, and water temperature is controlled at 20-80 ℃; Liquid linear velocity in 5 districts is controlled at 2-100cm/min; Outlet obtains weak absorption impurity in 2, is adsorbed impurity by force in the outlet 3, and these impurity comprise L-Ala, the Isoleucine that produces in the glucose that ferment, the fermenting process, heteroproteins on a small quantity.
The preparation method of the disclosed macroporous adsorbent resin special for separating valine of described Chinese patent 200910231735.4, step is:
(1) prepares aqueous phase substance with distilled water: wherein polyvinyl alcohol mass concentration 0.1%-2%, and NH
4SO
4Mass concentration 0.5%-6%;
(2) preparation organic phase material: get linking agent and mix with polar monomer, linking agent and polar monomer weight ratio are 10:1-2.5:1, to the pore-creating agent that wherein adds mixture 1-3 times volume, make polar monomer and linking agent carry out suspending copolymerization, obtain the organic phase material;
(3) resins: aqueous phase substance is warming up to 30-40 ℃, while stirring organic phase material and tert-Butyl dicarbonate are poured in the aqueous phase substance, be warming up to 70-80 ℃ after finishing mixing, polyreaction 4-5 hour, the mass ratio of water and organic phase is 20:1-5:1, and tert-Butyl dicarbonate accounts for the 0.1%-1% of reaction system gross weight;
(4) aftertreatment: leach resin, water cleans, vacuum-drying, and vacuum tightness is 100Pa, 40 ℃ of drying temperatures obtain 120-150 μ m particle diameter finished product macroporous adsorbent resin;
Described linking agent is selected for use: one or more in vinylbenzene, divinylbenzene, triethylene benzene, divinyl toluene, Vinylstyrene, divinyl dimethylbenzene, the divinyl ethylbenzene;
Described polar monomer is selected for use: ethyl propenoate, allyl acetate, allyl acetate or methyl ethylene pyrrolidone;
Described pore-creating agent is selected for use: toluene, dimethylbenzene, 200# gasoline, heptane, octane-iso, propyl carbinol or tertiary amyl alcohol.
Described NH
4SO
4Can use NaSO
4, NaCl, MgCl
2Or MgSO
4Substitute.
Described linking agent selects for use divinylbenzene and vinylbenzene to use with the ratio of mass ratio 1 ︰ 1.
The ratio of described polar monomer and linking agent is 0.18 ︰ 1.
Described pore-creating agent is toluene or 200# gasoline, and the dosage of pore-creating agent is 2 times of volumes of monomer mixture total amount.
Beneficial effect of the present invention: 1, obtain highly purified Xie Ansuan, also can obtain the L-Ala and the leucine of higher degree simultaneously; 2, adopt continuous chromatography to separate, resin is effectively utilized; 3, production process fully automated, production intensity is low; 4, the production process water is cooked moving phase, and is with low cost, do not have pollutent to produce substantially, helps protecting environment.
Description of drawings
Fig. 1 simulated moving bed chromatography post zone chart.
Embodiment
Embodiment 1:
Xie Ansuan concentration is about 30g/L after the fermentation ends, is adjusted to pH2 with hydrochloric acid, obtains supernatant liquor after 3000 commentaries on classics/min are centrifugal, adds the decolorizing with activated carbon of clear liquid weight 5%, filters and removes activated carbon, and Xie Ansuan concentration is 80g/L behind the clear liquid vacuum concentration.The diameter of simulation moving-bed inner prop is 5cm, and the number of the post in 1-5 district is: 2,2,2,2,2, and the flow velocity degree in 1-5 district is respectively 25,15,20,16,30 cm/min, and system temperature is controlled at 60 ℃, and the treatment capacity of fermentation clear liquid is 98 mL/min.Final is 32g/L from exporting the 1 Xie Ansuan product concentration that obtains, and purity reaches 99.5%, and the rate of recovery is 80%.
Embodiment 2:
Xie Ansuan concentration is about 35 g/L after the fermentation ends, is adjusted to pH5 with hydrochloric acid, obtains supernatant liquor after 8000 commentaries on classics/min are centrifugal, adds the decolorizing with activated carbon of clear liquid weight 10%, filters and removes activated carbon, and Xie Ansuan concentration is 70g/L behind the clear liquid vacuum concentration.The diameter of simulation moving-bed inner prop is 1cm, and the number of the post in 1-5 district is: 1,1,1,1,1, and the flow velocity degree in 1-5 district is respectively 30,22,27,18,50 cm/min, and system temperature is controlled at 80 ℃, and the treatment capacity of fermentation clear liquid is 3.9mL/min.Final is 32.8 g/L from exporting the 1 Xie Ansuan product concentration that obtains, and purity reaches 99.1%, and the rate of recovery is 75 %.
Embodiment 3:
Xie Ansuan concentration is about 20g/L after the fermentation ends, is adjusted to pH3 with hydrochloric acid, obtains supernatant liquor after 10000 commentaries on classics/min are centrifugal, adds the decolorizing with activated carbon of clear liquid weight 4%, filters and removes activated carbon, and Xie Ansuan concentration is 75g/L behind the clear liquid vacuum concentration.The diameter of simulation moving-bed inner prop is 20cm, and the number of the post in 1-5 district is: 3,3,3,3,3, and the flow velocity degree in 1-5 district is respectively 35,25,32,16,40 cm/min, and system temperature is controlled at 20 ℃, and the treatment capacity of fermentation clear liquid is 2.2 L/min.Final is 37.8 g/L from exporting the 1 Xie Ansuan product concentration that obtains, and purity reaches 99.0%, and the rate of recovery is 72 %.
Embodiment 4:
Xie Ansuan concentration is about 30g/L after the fermentation ends, is adjusted to pH4 with hydrochloric acid, obtains supernatant liquor after 5000 commentaries on classics/min are centrifugal, adds the decolorizing with activated carbon of clear liquid weight 10%, filters and removes activated carbon, and Xie Ansuan concentration is 80g/L behind the clear liquid vacuum concentration.The diameter of simulation moving-bed inner prop is 50cm, and the number of the post in 1-5 district is: 2,3,3,1,2, and the flow velocity degree in 1-5 district is respectively 37,20,30,15,35 cm/min, and system temperature is controlled at 70 ℃, and the treatment capacity of fermentation clear liquid is 19.6 L/min.Final is 35.8 g/L from exporting the 1 Xie Ansuan product concentration that obtains, and purity reaches 99.3%, and the rate of recovery is 76 %.
Embodiment 5:
Xie Ansuan concentration is about 30g/L after the fermentation ends, is adjusted to pH3 with hydrochloric acid, obtains supernatant liquor after 7000 commentaries on classics/min are centrifugal, adds the decolorizing with activated carbon of clear liquid weight 6%, filters and removes activated carbon, and Xie Ansuan concentration is 80g/L behind the clear liquid vacuum concentration.The diameter of simulation moving-bed inner prop is 30cm, and the number of the post in 1-5 district is: 2,2,3,1,3, and the flow velocity degree in 1-5 district is respectively 28,19,26,17,32 cm/min, and system temperature is controlled at 50 ℃, and the treatment capacity of fermentation clear liquid is 4.9 L/min.Final is 46.6 g/L from exporting the 1 Xie Ansuan product concentration that obtains, and purity reaches 99.6%, and the rate of recovery is 75 %.
Embodiment 5:
Xie Ansuan concentration is about 32g/L after the fermentation ends, is adjusted to pH3 with hydrochloric acid, obtains supernatant liquor after 8000 commentaries on classics/min are centrifugal, adds the decolorizing with activated carbon of clear liquid weight 4%, filters and removes activated carbon, and Xie Ansuan concentration is 70 g/L behind the clear liquid vacuum concentration.The diameter of simulation moving-bed inner prop is 10cm, and the number of the post in 1-5 district is: 2,1,2,1,3, and the flow velocity degree in 1-5 district is respectively 8,2,7,3,10cm/min, and system temperature is controlled at 30 ℃, and the treatment capacity of fermentation clear liquid is 392mL/min.Final is 42 g/L from exporting the 1 Xie Ansuan product concentration that obtains, and purity reaches 99.1%, and the rate of recovery is 72 %.
Embodiment 6:
Xie Ansuan concentration is about 35g/L after the fermentation ends, is adjusted to pH3 with hydrochloric acid, obtains supernatant liquor after 5000 commentaries on classics/min are centrifugal, adds the decolorizing with activated carbon of clear liquid weight 4%, filters and removes activated carbon, and Xie Ansuan concentration is 80 g/L behind the clear liquid vacuum concentration.The diameter of simulation moving-bed inner prop is 50cm, and the number of the post in 1-5 district is: 2,1,2,1,3, and the flow velocity degree in 1-5 district is respectively 100,60,90,70,100 cm/min, and system temperature is controlled at 30 ℃, and the treatment capacity of fermentation clear liquid is 58.9L/min.Final is 17.9 g/L from exporting the 1 Xie Ansuan product concentration that obtains, and purity reaches 99.0%, and the rate of recovery is 68 %.
Claims (1)
1. method of using Xie Ansuan in the continuous separation and purification fermented liquid of simulated moving bed chromatography, it is characterized in that designing the moving bed imitation chromatogram separation facility that is applicable to the valine fermentation liquid component characteristic, with water is moving phase, with Xie Ansuan separate, dedicated resin is stationary phase, Xie Ansuan in the valine fermentation liquid is carried out continuous separation and purification, and step is as follows:
(1) designed open five-region simulated moving bed chromatographic separation device, this device has 5 districts, and each district is made up of the 1-3 root chromatogram column, and parallel-series connects, and the specification of all chromatographic columns is identical, and column diameter is 1-50cm, chromatographic column adopting chuck heat tracing; In the middle of per two pillars 3 outlets are arranged, 3 inlets, sepn process except distinguish 5 and distinguish 1 between have two entry and exit can use, has only entry and exit to use in the middle of other district/post, no entry and exit all are in closing condition; The direction in 4-district, 3-district, 2-district, 1-district 5, district is the liquid flow path direction, and 1-district, 2-district, 3-district, 4-district, 5-district 5, district is the post switching direction; Xie Ansuan dedicated separation resin is housed in the post;
(2) after the Xie Ansuan fermenting process finishes, with hydrochloric acid with the fermented liquid pH regulator to 2-5, centrifugal removal thalline, protein and polysaccharide impurity are then with the clear liquid activated carbon decolorizing that obtains;
(3) valine fermentation liquid after will decolouring concentrates at 60 ℃ of vacuum decompressions, and the Xie Ansuan mass concentration is 70-80 g/L in the control clear liquid; Perhaps fermented liquid directly separates without concentrating;
(4) clear liquid is put into the described moving bed imitation chromatogram separation facility of step (1) and separated, obtain highly purified Xie Ansuan solution 1, obtain solid Xie Ansuan finished product through concentrated, crystallization again from exporting; Moving phase is water, and water temperature is controlled at 20-80 ℃; Liquid linear velocity in 5 districts is controlled at 2-100 cm/min; Outlet obtains weak absorption impurity in 2, is adsorbed impurity by force in the outlet 3, and these impurity comprise L-Ala, the Isoleucine that produces in the glucose that ferment, the fermenting process, heteroproteins on a small quantity.
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Cited By (8)
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|---|---|---|---|---|
| WO2013119034A1 (en) | 2012-02-06 | 2013-08-15 | Cj Cheiljedang Corporation | An apparatus for continuous separation of valine and a method for continuous separation of valine using the same |
| EP2656892A1 (en) * | 2012-04-23 | 2013-10-30 | Merck Patent GmbH | Chromatography method |
| CN104592047A (en) * | 2014-12-24 | 2015-05-06 | 三达膜科技（厦门）有限公司 | Separation and purification method of valine |
| CN104744277A (en) * | 2015-02-12 | 2015-07-01 | 新疆阜丰生物科技有限公司 | Simulated moving bed and method for extracting three-branch chain amino acid based on simulated moving bed |
| CN107185273A (en) * | 2017-06-15 | 2017-09-22 | 苏州麦可旺志生物技术有限公司 | The method and its equipment of a kind of continuous chromatography |
| CN109836346A (en) * | 2019-04-16 | 2019-06-04 | 同舟纵横（厦门）流体技术有限公司 | A kind of purifying technique of isoleucine fermentation liquid |
| CN111841073A (en) * | 2019-09-12 | 2020-10-30 | 浙江大学宁波理工学院 | Multi-column switching cycle chromatographic separation system and method for separating and concentrating target components from raw materials |
| CN116813492A (en) * | 2023-06-29 | 2023-09-29 | 山东兆光色谱分离技术有限公司 | Method for chromatographic separation of valine |
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| CN101168512A (en) * | 2007-11-30 | 2008-04-30 | 江南大学 | Method for separating and purifying valine from valine liquid |
| CN101721979A (en) * | 2009-12-01 | 2010-06-09 | 江南大学 | Method for preparing macroporous adsorbent resin special for separating valine |
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| CN101721979A (en) * | 2009-12-01 | 2010-06-09 | 江南大学 | Method for preparing macroporous adsorbent resin special for separating valine |
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| JP2018024679A (en) * | 2012-02-06 | 2018-02-15 | シージェイ チェイルジェダング コーポレイション | Apparatus for continuous separation of valine and method for continuous separation of valine using the same |
| KR101449808B1 (en) | 2012-02-06 | 2014-10-14 | 씨제이제일제당 (주) | An apparatus for continuous separation of valine and a method for continuous separation of valine using the same |
| CN104203903A (en) * | 2012-02-06 | 2014-12-10 | Cj第一制糖株式会社 | An apparatus for continuous separation of valine and a method for continuous separation of valine using the same |
| JP2015506958A (en) * | 2012-02-06 | 2015-03-05 | シージェイ チェイルジェダング コーポレイション | Apparatus for continuous separation of valine and method for continuous separation of valine using the same |
| CN104203903B (en) * | 2012-02-06 | 2016-09-28 | Cj第一制糖株式会社 | The continuous separation device of valine and the method being continuously separated valine with it |
| WO2013119034A1 (en) | 2012-02-06 | 2013-08-15 | Cj Cheiljedang Corporation | An apparatus for continuous separation of valine and a method for continuous separation of valine using the same |
| EP2656892A1 (en) * | 2012-04-23 | 2013-10-30 | Merck Patent GmbH | Chromatography method |
| JP2015520667A (en) * | 2012-04-23 | 2015-07-23 | メルク パテント ゲゼルシャフト ミット ベシュレンクテル ハフツングＭｅｒｃｋ Ｐａｔｅｎｔ Ｇｅｓｅｌｌｓｃｈａｆｔ ｍｉｔ ｂｅｓｃｈｒａｅｎｋｔｅｒ Ｈａｆｔｕｎｇ | Chromatographic method |
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| CN104592047A (en) * | 2014-12-24 | 2015-05-06 | 三达膜科技（厦门）有限公司 | Separation and purification method of valine |
| CN104744277A (en) * | 2015-02-12 | 2015-07-01 | 新疆阜丰生物科技有限公司 | Simulated moving bed and method for extracting three-branch chain amino acid based on simulated moving bed |
| CN107185273A (en) * | 2017-06-15 | 2017-09-22 | 苏州麦可旺志生物技术有限公司 | The method and its equipment of a kind of continuous chromatography |
| CN109836346A (en) * | 2019-04-16 | 2019-06-04 | 同舟纵横（厦门）流体技术有限公司 | A kind of purifying technique of isoleucine fermentation liquid |
| CN109836346B (en) * | 2019-04-16 | 2022-03-15 | 同舟纵横（厦门）流体技术有限公司 | Purification process of isoleucine fermentation liquor |
| CN111841073A (en) * | 2019-09-12 | 2020-10-30 | 浙江大学宁波理工学院 | Multi-column switching cycle chromatographic separation system and method for separating and concentrating target components from raw materials |
| CN111841073B (en) * | 2019-09-12 | 2022-02-22 | 浙江大学宁波理工学院 | Multi-column switching cycle chromatographic separation system and method for separating and concentrating target components from raw materials |
| CN116813492A (en) * | 2023-06-29 | 2023-09-29 | 山东兆光色谱分离技术有限公司 | Method for chromatographic separation of valine |
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Application publication date: 20110119 |