CN103709091A - Method for extracting L-tryptophan - Google Patents
Method for extracting L-tryptophan Download PDFInfo
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- CN103709091A CN103709091A CN201310626054.4A CN201310626054A CN103709091A CN 103709091 A CN103709091 A CN 103709091A CN 201310626054 A CN201310626054 A CN 201310626054A CN 103709091 A CN103709091 A CN 103709091A
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D209/00—Heterocyclic compounds containing five-membered rings, condensed with other rings, with one nitrogen atom as the only ring hetero atom
- C07D209/02—Heterocyclic compounds containing five-membered rings, condensed with other rings, with one nitrogen atom as the only ring hetero atom condensed with one carbocyclic ring
- C07D209/04—Indoles; Hydrogenated indoles
- C07D209/10—Indoles; Hydrogenated indoles with substituted hydrocarbon radicals attached to carbon atoms of the hetero ring
- C07D209/18—Radicals substituted by carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals
- C07D209/20—Radicals substituted by carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals substituted additionally by nitrogen atoms, e.g. tryptophane
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Abstract
The invention relates to a method for extracting L-tryptophan. The method comprises the following steps of 1, concentrating an L-tryptophan-containing solution by evaporation so that L-tryptophan content is 30-95g/L, 2, adding an organic acid into the concentrated L-tryptophan solution according to a mole ratio of L-tryptophan to the organic acid of 1: 4-12, and carrying out continued evaporation until the L-tryptophan content is 200-400g/L, 3, carrying out crystallization to obtain an L-tryptophan crude product, and 4, adding water into the L-tryptophan crude product, carrying out high-temperature treatment and carrying out crystallization to obtain an L-tryptophan pure product. The method reduces a production cost and sewage discharge and improves a yield and content of L-tryptophan.
Description
Technical field
The invention belongs to biological chemical field, be specifically related to a kind of method of extracting L-Trp.
Background technology
L-Trp is one of biogenic 8 seed amino acids of humans and animals, is important nutrition agent.In recent years, along with the development of domestic and international fodder industry and medicine industry, L-Trp becomes a kind of world market development prospect well and the larger product of Chinese market demand.At present, the production method of L-Trp has chemical synthesis, proteolysis method, and microbe fermentation method.The production method of L-Trp mainly relies on chemical synthesis and proteolysis method the earliest, and along with microbial method being produced to deepening continuously of L-Trp research, it is practical and in dominant position that the method has been moved towards.The extraction purifying of L-Trp mainly adopts absorption and ion exchange method.In absorption method, due to L-Trp light current from, cause resin low to the adsorption rate of L-Trp, product yield is low.In ion exchange method, washing can produce a large amount of trade effluents.The energy consumption of the production process of these two kinds of methods is higher, thereby causes production cost higher.
Summary of the invention
For the shortcoming of above-mentioned prior art, the invention provides a kind of method of extracting L-Trp, it comprises the following steps:
1) solution evaporation containing L-Trp is concentrated into every liter containing 30-95 gram of L-Trp;
2) with every mole of L-Trp 4-12 mole organic acid consumption, add organic acid, continue to be evaporated to every liter containing 200-400 gram of L-Trp;
3) crystallization obtains L-Trp crude product;
4) to described L-Trp crude product, add water, after pyroprocessing, crystallization obtains L-Trp sterling.
Described organic acid is selected from: formic acid, acetic acid, propionic acid, butyric acid, phenylformic acid, lactic acid, fumaric acid, toxilic acid, propanedioic acid, citric acid, oxysuccinic acid, Whitfield's ointment, oxalic acid and Sorbic Acid.Preferable formic acid.
In step 2) in, every mole of organic acid consumption corresponding to L-Trp can be 4-5,5-6,6-7,7-8,8-9,9-10,10-11 or 11-12 mole.
In preferred embodiments, in step 1), the solution evaporation containing L-Trp is concentrated into every liter containing 50-75 gram of L-Trp.
In preferred embodiments, in step 2) in, with every mole of L-Trp 4-12 mole organic acid consumption, add organic acid, continue to be evaporated to every L containing 260-330 gram of L-Trp.
Preferably, in step 3) and step 4), the low temperature slow crystallization of crystallization for carrying out at 15-25 ℃ preferably carried out at 20 ℃; The time of carrying out is 8 hours-16 hours, is preferably 12 hours.
In preferred embodiments, the described pH value containing L-Trp solution is 6.8-7.2.
In preferred embodiments, in preferred embodiments, in step 4), described pyroprocessing is carried out at the temperature of >=80 ℃, preferably at 80-100 ℃, carries out, and the time of carrying out is 5-15 minute, preferably 10 minutes.
In preferred embodiments, in step 4), the consumption of the water that every gram of L-Trp crude product is corresponding is 1.5-2.5 milliliter.
In preferred embodiments, the described solution containing L-Trp is L-Trp fermented liquid, preferred intestinal bacteria L-Trp fermented liquid, and before carrying out described step 1), described L-Trp fermented liquid is filtered and the processing of decolouring.Preferably, the L-Trp content of described L-Trp fermented liquid is 20-28 grams per liter.In preferred embodiments, with described organic acid, pH is adjusted to 4.5 described L-Trp fermented liquid being filtered and decolour before processing.In preferred embodiments, described decolouring is processed and is used Powdered Activated Carbon to carry out.In preferred embodiments, the consumption of described Powdered Activated Carbon is 3 grams per liters.In preferred embodiments, described decolouring is processed and is carried out at 60-80 ℃, and bleaching time is 20-40 minute, is preferably 30 minutes.
L-Trp and organic acid form the hydration L-Trp organic acid salt that L-Trp purity is higher under finite concentration in the method for the invention, resulting hydration L-Trp organic acid salt can make organic acid salt Crystal Breakup through pyroprocessing, thereby obtains L-Trp.
Method of the present invention has the following advantages:
1. no longer adopt traditional resin adsorption method, simple to operate, reduced the energy expenditure in producing, and raw material is cheap and organic acid consumption is less, has reduced cost;
2. in step 3), solution is concentrated to every liter containing 200-400 gram of L-Trp, make the mother liquor after step 3) crystallization less, and the L-Trp crude product producing in present method is the hydration tryptophane organic acid salt that tryptophane purity is higher, so the impurity that the mother liquor after the crystallization in step 4) contains is considerably less, can reuse or reclaim, environmentally friendly;
3. when described method is used formic acid extraction tryptophane, mother liquor in step 3) after crystallization can be used for preparing calcium formiate and (primary crystallization mother liquor is regulated to PH to 5.0 with calcium hydroxide, spraying is dried the feed grade calcium formiate of Dry Sack histidine content approximately 5%), thus make whole leaching process substantially not produce waste liquid.
4. improved yield and the content of L-Trp, tryptophane content in crude product is more than 90%, even can reach 95%, and yield can reach 80%; Tryptophane sterling content >=98.5, yield can reach more than 70%.
Embodiment
With specific embodiment, technical scheme of the present invention is done to further technical specification below; But the present invention is not limited to these embodiment." the L-Trp crude product " of the use in specification sheets of the present invention and claims refers to L-Trp organic acid salt crystal." every liter containing gram L-Trp " using in specification sheets of the present invention and claims refers to the amount of average every liter of L-Trp existing in any form containing in reaction system (such as the tryptophane in free tryptophane, tryptophane organic acid salt etc.), the volume (L) of total amount (the g)/reaction system of its L-Trp that equals to exist in any form in reaction system (such as the tryptophane in free tryptophane, tryptophane organic acid salt etc.).The L-Trp fermented liquid using in embodiment is herein prepared by Escherichia coli fermentation.Other preparations used herein and equipment are common commercially available product.
Embodiment 1
Formic acid adjust pH 4.5 for the L-Trp fermented liquid that is 23 grams per liters by 10 liters of concentration, be heated to 50 ℃, 0.1 micron of tubular fibre membrane filtration, obtain 10 liters of clear liquids, described clear liquid is heated to 70 ℃, add 30 grams of Powdered Activated Carbons, slowly stir 30 minutes, vacuum filtration obtains the clear liquid that decolours, and filter cake is obtained to washing water with 30 ml pure water washings, and the decolouring clear liquid obtaining is mixed stand-by with washing water.The mixing solutions of gained is evaporated to 4 liters (every liter containing 57.5 grams of L-Trps), add formic acid (0.4 liter), continue to be evaporated to 0.77 liter (every liter containing 298.7 grams of L-Trps), discharging is to crystallization cylinder, at 20 ℃, slow cooling crystallization is 12 hours, 3000rpm is centrifugal, pour out supernatant (, mother liquor after crystallization for the first time, volume is 0.57L), gained crystallization is washed with 40 ml pure waters, obtain after dry at 80 ℃ as 196.37 grams of the L-Trp crude products of rhabdolith, content 94.97%, yield 81.08%.L-Trp crude product is added in 400 ml pure waters, at 85 ℃, slowly stir 10 minutes, at 20 ℃, slow cooling crystallization is 12 hours, and 3000rpm is centrifugal, pours out supernatant (, mother liquor after crystallization for the second time, volume is 0.23L), 30 ml pure waters washings, after dry at 80 ℃ 165.55 grams of sheet L-Trp sterlings, content 98.72%, yield 71.06%.
Embodiment 2
Formic acid adjust pH 4.5 for the L-Trp fermented liquid that is 22 grams per liters by 15 liters of concentration, be heated to 50 ℃, 0.1 micron of tubular fibre membrane filtration, obtain 15.1 liters of clear liquids, described clear liquid is heated to 70 ℃, add 45 grams of Powdered Activated Carbons, slowly stir 30 minutes, vacuum filtration obtains the clear liquid that decolours, and filter cake obtains washing water with 45 ml pure water washings, and the decolouring clear liquid obtaining is mixed stand-by with washing water.The mixing solutions of gained is evaporated to 5.5 liters (every liter containing 60 grams of L-Trps), add formic acid (0.7 liter), continue to be evaporated to 1.2 liters (every liter containing 275 grams of L-Trps), discharging is to crystallization cylinder, at 20 ℃, slow cooling crystallization is 12 hours, 3000rpm is centrifugal, pour out supernatant (, mother liquor after crystallization for the first time, volume is 0.92L), gained crystallization is washed with 58 ml pure waters, obtain after dry at 80 ℃ as 286.32 grams of the L-Trp crude products of rhabdolith, content 93.15%, yield 80.82%.L-Trp crude product is added in 550 ml pure waters, 90 ℃ are slowly stirred 10 minutes, at 20 ℃, slow cooling crystallization is 12 hours, and 3000rpm is centrifugal, pours out supernatant (, mother liquor after crystallization for the second time, volume is 0.28L), 42 ml pure waters washings, after dry at 80 ℃ 236.12 grams of sheet L-Trp sterlings, content 99.02%, yield 70.85%.
The mother liquor and the first subcrystalline wash water that in embodiment 1 and 2, carry out after crystallization are for the first time mixed, obtain 1.6L.Under the stirring (180rpm) of magnetic stirring apparatus, to described solution, slowly add calcium hydroxide, adjust pH to 5.0.Then use small spraying drying instrument (far away purchased from Shanghai generation) spraying dry, 140 degrees Celsius of inlet temperature, inlet amount 20 ml/min, obtain 102 grams of calcium formiates that contain 5.14% tryptophane.
Embodiment 3
Acetic acid adjust pH 4.5 for the L-Trp fermented liquid that is 22 grams per liters by 10 liters of concentration, be heated to 50 ℃, 0.1 micron of tubular fibre membrane filtration, obtain 10.1 liters of clear liquids, described clear liquid is heated to 70 ℃, add 30 grams of Powdered Activated Carbons, slowly stir 30 minutes, vacuum filtration obtains the clear liquid that decolours, and filter cake obtains washing water with 30 ml pure water washings, and the decolouring clear liquid obtaining is mixed stand-by with washing water.The mixing solutions of gained is evaporated to 3.7 liters (every liter containing 59.5 grams of L-Trps), adds acetic acid (0.5 liter), continue to be evaporated to 0.80 liter (every liter containing 275 grams of L-Trps), discharging is to crystallization cylinder, at 20 ℃, slow cooling crystallization is 12 hours, and 3000rpm is centrifugal, pours out supernatant (, mother liquor after crystallization for the first time, volume is 0.63L), 40 ml pure waters washings, obtain after dry at 80 ℃ as 102.21 grams of the L-Trp crude products of rhabdolith, content 89.54%, yield 41.60%.L-Trp crude product is added in 200ml pure water, 85 ℃ are slowly stirred 10 minutes, at 20 ℃, slow cooling crystallization is 12 hours, and 3000rpm is centrifugal, pours out supernatant (, mother liquor after crystallization for the second time, volume is 0.09L), 15ml pure water washing, after dry at 80 ℃ 75.21 grams of sheet L-Trp sterlings, content 98.61%, yield 33.71%
Embodiment 4
Lactic acid adjust pH 4.5 for the L-Trp fermented liquid that is 23 grams per liters by 10 liters of concentration, be heated to 50 ℃, 0.1 micron of tubular fibre membrane filtration, obtain 10 liters of clear liquids, described clear liquid is heated to 70 ℃, add 30 grams of Powdered Activated Carbons, slowly stir 30 minutes, vacuum filtration obtains the clear liquid that decolours, and filter cake obtains washing water with 30 ml pure water washings, and the decolouring clear liquid obtaining is mixed stand-by with washing water.The mixing solutions of gained is evaporated to 3.63 liters (every liter containing 63.4 grams of L-Trps), adds lactic acid (0.4 liter), continue to be evaporated to 0.76 liter (every liter containing 302.6 grams of L-Trps), discharging is to crystallization cylinder, at 20 ℃, slow cooling crystallization is 12 hours, and 3000rpm is centrifugal, pours out supernatant (, mother liquor after crystallization for the first time, volume is 0.6L), 39 ml pure waters washings, obtain after dry at 80 ℃ as 89.71 grams of the L-Trp crude products of rhabdolith, content 91.32%, yield 35.62%.L-Trp crude product is added in 180ml pure water, 85 ℃ are slowly stirred 10 minutes, at 20 ℃, slow cooling crystallization is 12 hours, and 3000rpm is centrifugal, pours out supernatant (, mother liquor after crystallization for the second time, volume is 0.08L), 11ml pure water washing, after dry at 80 ℃ 62.85 grams of sheet L-Trp sterlings, content 98.73%, yield 26.98%.
Claims (10)
1. a method of extracting L-Trp, it comprises the following steps:
1) solution evaporation containing L-Trp is concentrated into every liter containing 30-95 gram of L-Trp;
2) with every mole of L-Trp 4-12 mole organic acid consumption, add organic acid, continue to be evaporated to every liter containing 200-400 gram of L-Trp;
3) crystallization obtains L-Trp crude product,
4) to described L-Trp crude product, add water, after pyroprocessing, crystallization obtains L-Trp sterling.
2. the process of claim 1 wherein that described organic acid is selected from: formic acid, acetic acid, propionic acid, butyric acid, phenylformic acid, lactic acid, fumaric acid, toxilic acid, propanedioic acid, citric acid, oxysuccinic acid, Whitfield's ointment, oxalic acid and Sorbic Acid, preferable formic acid.
3. the method for claim 1, in step 3), the low temperature slow crystallization of described crystallization for carrying out at 15 ℃-25 ℃ preferably carried out at 20 ℃; The time of carrying out is 8 hours-16 hours, is preferably 12 hours.
4. the method for claim 1, described pyroprocessing is carried out at the temperature of >=80 ℃, preferably at 80-100 ℃, carries out; The time of carrying out is 5-15 minute, preferably 10 minutes.
5. the method for claim 1, in step 4), described crystallization is the slow crystallization of low temperature at 15 ℃-25 ℃, preferably at 20 ℃, carries out; The time of carrying out is 8 hours-16 hours, is preferably 12 hours.
6. the method for claim 1, in step 4), the consumption of the water that every gram of L-Trp crude product is corresponding is 1.5-2.5 milliliter.
7. the method for claim 1, comprises the following steps:
1) solution evaporation containing L-Trp is concentrated into every liter containing 50-75 gram of L-Trp;
2) with every mole of L-Trp 4-12 mole organic acid consumption, add organic acid, continue to be evaporated to every liter containing 260-330 gram of L-Trp;
3) crystallization obtains L-Trp crude product,
4) to described L-Trp crude product, add water, after pyroprocessing, crystallization obtains L-Trp sterling.
8. the method for claim 1-7 any one, the wherein said solution containing L-Trp is L-Trp fermented liquid, preferably intestinal bacteria L-Trp fermented liquid.
9. the method for claim 8, the L-Trp content 20-28 grams per liter of wherein said L-Trp fermented liquid.
10. the method for claim 8 was filtered and the processing of decolouring described L-Trp fermented liquid before carrying out described step 1), and described decolouring is processed and used Powdered Activated Carbon to carry out at 60-80 ℃; Bleaching time is 20-40 minute, is preferably 30 minutes, with described organic acid, pH is adjusted to 4.5 described L-Trp fermented liquid being filtered and decolour before processing.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105693592A (en) * | 2016-03-03 | 2016-06-22 | 天津科技大学 | Process method for efficiently extracting L-tryptophan from fermentation liquor through thallus carrying crystallization |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105693592A (en) * | 2016-03-03 | 2016-06-22 | 天津科技大学 | Process method for efficiently extracting L-tryptophan from fermentation liquor through thallus carrying crystallization |
| CN105693592B (en) * | 2016-03-03 | 2019-03-19 | 天津科技大学 | A kind of carry disease germs from fermentation liquid crystallizes the process of high efficiency extraction L-Trp |
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