CN101709049A - Method for extracting L-tryptophan - Google Patents
Method for extracting L-tryptophan Download PDFInfo
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- CN101709049A CN101709049A CN200910224005A CN200910224005A CN101709049A CN 101709049 A CN101709049 A CN 101709049A CN 200910224005 A CN200910224005 A CN 200910224005A CN 200910224005 A CN200910224005 A CN 200910224005A CN 101709049 A CN101709049 A CN 101709049A
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Abstract
The invention relates to a method for extracting L-tryptophan. The method comprises the following steps: performing pressure filtration on L-tryptophan fermentation liquor flocculated through flocculant; decolorizing clean liquid obtained through pressure filtration through activated carbon; treating the obtained product through ion exchange resin; eluting the obtained product with ammonia water; and performing concentration and crystallization on eluent so as to obtain the L-tryptophan. The method for extracting L-tryptophan has the advantages of high extraction yield, high product purity, simple process, convenient operation, low labor intensity and little environmental pollution.
Description
Technical field
The invention belongs to biological chemical field, particularly relate to a kind of method of the L-of extraction tryptophane.
Background technology
The L-tryptophane is to contain the German-Chinese property of indyl die aromatischen Aminosaeuren, is white in color or slightly yellowy leaflet crystal or powder, and odorless, pleasantly sweet.Solubleness is 1.14g/l (25 ℃) in the water, is dissolved in diluted acid or diluted alkaline, and is more stable in alkali lye, decomposes in the strong acid.Be slightly soluble in ethanol, be insoluble to chloroform, ether.
The L-tryptophane is one of indispensable amino acid of humans and animals, the restrictive function certain to having grown of humans and animals.But in vivo from L-tryptophane synthetic hormone, pigment, alkaloid, coenzyme and several physiological active substances.Be mainly used in medicine clinically, as amino acid transfusion, anti-vexed dose, anti-smelly filter medicine is regulated effects such as brain metabolism.Recently research report, the L-tryptophane in vivo can be by vitamins B
6Effect generate neural hormonal precursor serotonine, thereby promote sleep and ataraxy have been used as protective foods and tranquilizer.The main application of tryptophane is to make the nutrition medicament at present, synthetic meals, indispensable amino acid sheet, and the additive of protein hydrolysate.Tryptophane and chalybeate, VITAMIN etc. are share and can be improved anti-anaemia curative effect, in addition, also can be used as the adjutant of peptic ulcer Histidine, are used for the treatment of insomnia, anxiety disorder, stress and craving for tobacco drug addiction etc.
In recent years, the L-tryptophane is as indispensable amino acid, and is increasingly extensive in the every field application, along with the extensive application of Methionin, methionine(Met), the higher always L-tryptophane of price more wishes to have the production and the extracting method of Cheap highly effective, so that advance the application of L one tryptophane.The same with the other biological engineering product, the tryptophane industrial production also usually can be subjected to the restriction of production cost, and in the formation of production cost, the cost of downstream engineerings such as separation and Extraction occupies suitable ratio.Therefore, the extracting method of research L one tryptophane has important significance for theories and practical value.When direct fermentation was produced L one tryptophane, by-product amino acid was more in the fermented liquid, has increased the difficulty of separation and Extraction L one tryptophane.
Summary of the invention
The method that the purpose of this invention is to provide a kind of L-of extraction tryptophane is to solve the problems referred to above that prior art exists.
The invention provides a kind of method of the L-of extraction tryptophane, comprise the steps: L-tryptophane fermented liquid by flocculation agent flocculation back press filtration, the clear liquid activated carbon decolorizing that press filtration obtains, then after ion exchange resin treatment, use the ammoniacal liquor wash-out again, the gained elutriant promptly gets the L-tryptophane through condensing crystal.
Described L-tryptophane fermented liquid is obtained by Escherichia coli fermentation, and wherein the content of L-tryptophane is 20～30g/L.
Described flocculation agent is selected from A645 or A856, and the flocculation agent addition is 10～30ppm/L, and flocculation temperature is 50～80 ℃, and flocculation time is 30～60min; Wherein said flocculation agent is available from flocculation agent company limited of Jiangsu Nan Tian, and model is A645 and A856, and the flocculating effect of A856 is better.
Described gac addition is 5～10g/L, preferred 8g/L, and bleaching temperature is 60～80 ℃, bleaching time 30～60min; The preferred Zhuxi Powdered Activated Carbon that adopts available from Zhuxi, Chinese Liyang gac company limited.
The clear liquid decolouring back that described press filtration obtains is earlier with 98% vitriol oil adjust pH to 2.0～4.0.
Described ion exchange resin type number is the WD-2 amino-acid resin, and ion exchange column is in 50～70 ℃ of insulations.
The ammonia concn that wash-out is used is 1.5～2.5% (V/V), is preferably 1.8%, and eluting temperature is 50～70 ℃.
The tryptophane elutriant is concentrated at 50～70 ℃ of following vacuum decompressions.
Slowly lower the temperature during described elutriant condensing crystal, keep 1～2 hour after temperature is reduced to 20～25 ℃, centrifugal, crystal washs with pure water, places 50～80 ℃ of oven dry again.
The method of extraction L-tryptophane of the present invention has following beneficial effect: the extract yield height, and the product purity height, the technology simple operations is easy, and labour intensity is little, and is little to the pollution of environment.
Embodiment
Following examples are used to illustrate the present invention, but are not used for limiting the scope of the invention.
L-tryptophane fermented liquid source: by intestinal bacteria (Anhui Fengyuan Fermentation Technology Engineering Research Co., Ltd.) fermentation gained, wherein the L-tryptophane in the fermented liquid is 20～30g/L;
Flocculating agent A 856 and A645 are available from flocculation agent company limited of Jiangsu Nan Tian;
Used ion exchange resin is that WD-2 amino acid is resin dedicated among the embodiment, available from Anhui Wan Dong chemical industry company limited;
The Zhuxi Powdered Activated Carbon is available from Zhuxi, Chinese Liyang gac company limited.
Embodiment 1
With 20L concentration is to add 200ml1g/L A856 flocculation agent after the L-tryptophane fermented liquid of 25g/L is heated to 60 ℃, use little filter press behind the insulation 30min, obtain the 22L clear liquid, add Zhuxi Powdered Activated Carbon 140g after clear liquid is heated to 65 ℃ and slowly stir decolouring 30min, vacuum filtration, filter cake washs with pure water, destainer and wash water mixed collection; The destainer of collecting is transferred the laggard ion exchange column of pH to 3.0 with the vitriol oil, treat the column outlet effluent liquid be subacidity when (being about pH5) with 1.8% (V/V) ammoniacal liquor wash-out, eluting temperature is 60 ℃, collects elutriant 40L altogether, and vacuum decompression is concentrated into behind the 5L slowly decrease temperature crystalline, mixing speed 150RPM, kept after temperature is reduced to 20 ℃ 1 hour, centrifugal, crystal washs, obtains 422gL-tryptophane product 50 ℃ of vacuum-dryings with the 500ml pure water, purity is 99.25%, and yield is 83.77%.
Embodiment 2
50L concentration is to add 550ml1g/L A856 flocculation agent after the L-tryptophane fermented liquid of 23.6g/L is heated to 60 ℃, use little filter press behind the insulation 30min, obtain the 58L clear liquid, add Zhuxi Powdered Activated Carbon 450g after clear liquid is heated to 65 ℃ and slowly stir decolouring 30min, vacuum filtration, filter cake washs with pure water, destainer and wash water mixed collection, the destainer of collecting is transferred the laggard ion exchange column of pH to 3.2 with the vitriol oil, treat the column outlet effluent liquid be subacidity when (being about pH5) with 1.8% (V/V) ammoniacal liquor wash-out, eluting temperature is 65 ℃, collect elutriant 110L altogether, vacuum decompression is concentrated into behind the 10L slowly decrease temperature crystalline, mixing speed 150RPM, after reducing to 20 ℃, temperature kept 1 hour, centrifugal, crystal obtains 1006g L-tryptophane product with the washing of 2000ml pure water 50 ℃ of vacuum-dryings, purity is 98.8%, and yield is 84.23%.
Embodiment 3
50L concentration is to add 600ml1g/L A856 flocculation agent after the L-tryptophane fermented liquid of 24g/L is heated to 65 ℃, use little filter press behind the insulation 30min, obtain the 60L clear liquid, add Zhuxi Powdered Activated Carbon 500g after clear liquid is heated to 65 ℃ and slowly stir decolouring 30min, vacuum filtration, filter cake washs with pure water, destainer and wash water mixed collection, the destainer of collection is transferred the laggard ion exchange column of pH to 3.0 with the vitriol oil, treat the column outlet effluent liquid be subacidity when (being about pH5) with 2.0% (V/V) ammoniacal liquor wash-out, eluting temperature is 60 ℃, collect elutriant 108L altogether, vacuum decompression is concentrated into behind the 10L slowly decrease temperature crystalline, mixing speed 150RPM, after reducing to 10 ℃, temperature kept 1 hour, centrifugal, crystal obtains 1050gL-tryptophane product with the washing of 2000ml frozen water 50 ℃ of vacuum-dryings, purity is 99.1%, and yield is 86.71%.
Embodiment 4
20L concentration is to add 200ml1g/L A856% flocculation agent after the L-tryptophane fermented liquid of 23.8g/L is heated to 60 ℃, use little filter press behind the insulation 30min, obtain the 23L clear liquid, add Zhuxi Powdered Activated Carbon 150g after clear liquid is heated to 65 ℃ and slowly stir decolouring 30min, vacuum filtration, filter cake washs with pure water, destainer and wash water mixed collection, the destainer of collecting is transferred the laggard ion exchange column of pH to 3.0 with the vitriol oil, treat the column outlet effluent liquid be subacidity when (being about pH5) with 2.0% (V/V) ammoniacal liquor wash-out, eluting temperature is 60 ℃, collects elutriant 38L altogether, vacuum decompression is concentrated into behind the 5L slowly decrease temperature crystalline, mixing speed 150RPM keeps 1 hour after temperature is reduced to 10 ℃, and is centrifugal, crystal obtains 422g L-tryptophane product with the washing of 500ml pure water 50 ℃ of vacuum-dryings.Purity is 97.8%, and yield is 84.51%.
Embodiment 5
With 20L concentration is to add 500ml1g/L A645 flocculation agent after the L-tryptophane fermented liquid of 25.8g/L is heated to 60 ℃, use little filter press behind the insulation 60min, obtain the 22L clear liquid, add Zhuxi Powdered Activated Carbon 200g after clear liquid is heated to 65 ℃ and slowly stir decolouring 30min, vacuum filtration, filter cake washs with pure water, destainer and wash water mixed collection; The destainer of collecting is transferred the laggard ion exchange column of pH to 4.0 with the vitriol oil, treat the column outlet effluent liquid be subacidity when (being about pH5) with 2.5% (V/V) ammoniacal liquor wash-out, eluting temperature is 70 ℃, collects elutriant 40L altogether, and vacuum decompression is concentrated into behind the 5L slowly decrease temperature crystalline, mixing speed 150RPM, kept after temperature is reduced to 20 ℃ 1 hour, centrifugal, crystal washs, obtains 422gL-tryptophane product 50 ℃ of vacuum-dryings with the 500ml pure water, purity is 99.15%, and yield is 81.78%.
Claims (9)
1. a method of extracting the L-tryptophane is characterized in that, press filtration after L-tryptophane fermented liquid is flocculated by flocculation agent, the clear liquid activated carbon decolorizing that press filtration obtains, after ion exchange resin treatment, use the ammoniacal liquor wash-out more then, the gained elutriant promptly gets the L-tryptophane through condensing crystal.
2. the method for extraction L-tryptophane according to claim 1 is characterized in that described L-tryptophane fermented liquid is obtained by Escherichia coli fermentation, and wherein the content of L-tryptophane is 20～30g/L.
3. the method for extraction L-tryptophane according to claim 1 is characterized in that described flocculation agent is selected from A645 or A856, and the flocculation agent addition is 10～30ppm/L, and flocculation temperature is 50～80 ℃, and flocculation time is 30～60min.
4. the method for extraction L-tryptophane according to claim 1 is characterized in that described gac addition is 5～10g/L, 60～80 ℃ of bleaching temperatures, bleaching time 30～60min.
5. the method for extraction L-tryptophane according to claim 1 is characterized in that, the clear liquid decolouring back that described press filtration obtains is earlier with 98% vitriol oil adjust pH to 2.0～4.0.
6. the method for extraction according to claim 1 L-tryptophane is characterized in that, described ion exchange resin type number is the WD-2 amino-acid resin, 50～70 ℃ of insulations of ion exchange column.
7. the method for extraction L-tryptophane according to claim 1 is characterized in that the ammonia concn that wash-out is used is 1.5～2.5% (V/V), 50～70 ℃ of eluting temperatures.
8. the method for extraction L-tryptophane according to claim 1 is characterized in that, the tryptophane elutriant is concentrated at 50～70 ℃ of following vacuum decompressions.
9. the method for extraction according to claim 1 L-tryptophane is characterized in that, slowly lowers the temperature during described elutriant condensing crystal, keeps 1～2 hour after temperature is reduced to 20～25 ℃, and centrifugal, crystal washs with pure water, places 50～80 ℃ of oven dry again.
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| CN2009102240051A CN101709049B (en) | 2009-11-17 | 2009-11-17 | Method for extracting L-tryptophan |
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| CN101709049B CN101709049B (en) | 2011-09-14 |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109369501A (en) * | 2018-11-28 | 2019-02-22 | 河南巨龙生物工程股份有限公司 | It is a kind of to pass through the technique for carrying out tryptophan purification of directly flocculating to fermentation liquid |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| GB0218012D0 (en) * | 2002-08-05 | 2002-09-11 | Ciba Spec Chem Water Treat Ltd | Production of a fermentation product |
| CN101376646A (en) * | 2007-08-29 | 2009-03-04 | 天津科技大学 | Novel method for extracting L-tryptophan from fermentation liquor |
| CN101565395A (en) * | 2009-05-25 | 2009-10-28 | 河南孟成生物药业股份有限公司 | Technique for extracting L-tryptophan in fermentation liquor |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109369501A (en) * | 2018-11-28 | 2019-02-22 | 河南巨龙生物工程股份有限公司 | It is a kind of to pass through the technique for carrying out tryptophan purification of directly flocculating to fermentation liquid |
| CN109369501B (en) * | 2018-11-28 | 2020-11-10 | 河南巨龙生物工程股份有限公司 | Process for purifying tryptophan by directly flocculating fermentation liquor |
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