CN104357507A - Fermentation production process of high-concentration L-sorbose - Google Patents
Fermentation production process of high-concentration L-sorbose Download PDFInfo
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- CN104357507A CN104357507A CN201410561050.7A CN201410561050A CN104357507A CN 104357507 A CN104357507 A CN 104357507A CN 201410561050 A CN201410561050 A CN 201410561050A CN 104357507 A CN104357507 A CN 104357507A
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- fermentation
- sorbose
- citric acid
- high density
- manufacturing technique
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- 230000004151 fermentation Effects 0.000 title claims abstract description 57
- 238000000855 fermentation Methods 0.000 title claims abstract description 56
- LKDRXBCSQODPBY-AMVSKUEXSA-N L-(-)-Sorbose Chemical compound OCC1(O)OC[C@H](O)[C@@H](O)[C@@H]1O LKDRXBCSQODPBY-AMVSKUEXSA-N 0.000 title claims abstract description 48
- 238000004519 manufacturing process Methods 0.000 title claims abstract description 18
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 claims abstract description 36
- LVHBHZANLOWSRM-UHFFFAOYSA-N methylenebutanedioic acid Natural products OC(=O)CC(=C)C(O)=O LVHBHZANLOWSRM-UHFFFAOYSA-N 0.000 claims abstract description 21
- 238000000034 method Methods 0.000 claims abstract description 19
- 239000001963 growth medium Substances 0.000 claims abstract description 10
- 108010010803 Gelatin Proteins 0.000 claims abstract description 9
- 229920000159 gelatin Polymers 0.000 claims abstract description 9
- 239000008273 gelatin Substances 0.000 claims abstract description 9
- 235000019322 gelatine Nutrition 0.000 claims abstract description 9
- 235000011852 gelatine desserts Nutrition 0.000 claims abstract description 9
- 238000009630 liquid culture Methods 0.000 claims abstract description 9
- FBPFZTCFMRRESA-FSIIMWSLSA-N D-Glucitol Natural products OC[C@H](O)[C@H](O)[C@@H](O)[C@H](O)CO FBPFZTCFMRRESA-FSIIMWSLSA-N 0.000 claims abstract description 7
- FBPFZTCFMRRESA-JGWLITMVSA-N D-glucitol Chemical compound OC[C@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-JGWLITMVSA-N 0.000 claims abstract description 7
- 229960002920 sorbitol Drugs 0.000 claims abstract description 7
- 239000000758 substrate Substances 0.000 claims abstract description 5
- NAOLWIGVYRIGTP-UHFFFAOYSA-N 1,3,5-trihydroxyanthracene-9,10-dione Chemical compound C1=CC(O)=C2C(=O)C3=CC(O)=CC(O)=C3C(=O)C2=C1 NAOLWIGVYRIGTP-UHFFFAOYSA-N 0.000 claims description 11
- 241000193830 Bacillus <bacterium> Species 0.000 claims description 9
- 235000010356 sorbitol Nutrition 0.000 claims description 6
- 239000007788 liquid Substances 0.000 abstract description 26
- 239000000463 material Substances 0.000 abstract description 3
- 241000589232 Gluconobacter oxydans Species 0.000 abstract 3
- JAHNSTQSQJOJLO-UHFFFAOYSA-N 2-(3-fluorophenyl)-1h-imidazole Chemical compound FC1=CC=CC(C=2NC=CN=2)=C1 JAHNSTQSQJOJLO-UHFFFAOYSA-N 0.000 abstract 1
- 230000015572 biosynthetic process Effects 0.000 abstract 1
- 230000019522 cellular metabolic process Effects 0.000 abstract 1
- 238000004904 shortening Methods 0.000 abstract 1
- 239000002609 medium Substances 0.000 description 9
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 6
- VTYYLEPIZMXCLO-UHFFFAOYSA-L Calcium carbonate Chemical compound [Ca+2].[O-]C([O-])=O VTYYLEPIZMXCLO-UHFFFAOYSA-L 0.000 description 6
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 description 5
- 238000003756 stirring Methods 0.000 description 5
- 239000001888 Peptone Substances 0.000 description 3
- 108010080698 Peptones Proteins 0.000 description 3
- 229960000583 acetic acid Drugs 0.000 description 3
- 229910000019 calcium carbonate Inorganic materials 0.000 description 3
- 229940041514 candida albicans extract Drugs 0.000 description 3
- 238000012258 culturing Methods 0.000 description 3
- 239000012362 glacial acetic acid Substances 0.000 description 3
- 235000019319 peptone Nutrition 0.000 description 3
- 235000019154 vitamin C Nutrition 0.000 description 3
- 239000011718 vitamin C Substances 0.000 description 3
- 239000012138 yeast extract Substances 0.000 description 3
- ZZZCUOFIHGPKAK-UHFFFAOYSA-N D-erythro-ascorbic acid Natural products OCC1OC(=O)C(O)=C1O ZZZCUOFIHGPKAK-UHFFFAOYSA-N 0.000 description 2
- 229930003268 Vitamin C Natural products 0.000 description 2
- 230000001580 bacterial effect Effects 0.000 description 2
- 230000003570 biosynthesizing effect Effects 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 241000894006 Bacteria Species 0.000 description 1
- 241000196324 Embryophyta Species 0.000 description 1
- 241000589236 Gluconobacter Species 0.000 description 1
- 241001052560 Thallis Species 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 229960005070 ascorbic acid Drugs 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000005265 energy consumption Methods 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- RWSXRVCMGQZWBV-WDSKDSINSA-N glutathione Chemical compound OC(=O)[C@@H](N)CCC(=O)N[C@@H](CS)C(=O)NCC(O)=O RWSXRVCMGQZWBV-WDSKDSINSA-N 0.000 description 1
- 235000003969 glutathione Nutrition 0.000 description 1
- 229960003180 glutathione Drugs 0.000 description 1
- 230000036039 immunity Effects 0.000 description 1
- 239000002054 inoculum Substances 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 238000004886 process control Methods 0.000 description 1
- 229930003231 vitamin Natural products 0.000 description 1
- 235000013343 vitamin Nutrition 0.000 description 1
- 239000011782 vitamin Substances 0.000 description 1
- 229940088594 vitamin Drugs 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/02—Monosaccharides
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- Organic Chemistry (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Zoology (AREA)
- Life Sciences & Earth Sciences (AREA)
- Wood Science & Technology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Microbiology (AREA)
- General Chemical & Material Sciences (AREA)
- Biotechnology (AREA)
- Health & Medical Sciences (AREA)
- Biochemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention discloses an efficient fermentation production process of L-sorbose under the condition of high concentration. In the process, gluconobacter melanogenus is taken as a production strain and D-sorbitol is taken as a substrate to ferment to produce the L-sorbose, 0.1-5 mg/ml of gelatin is added in a formula of a strain liquid culture medium, and 0.1-5 mg/ml of citric acid and 0.1-5 mg/ml of itaconic acid are added in a separated or combined manner into the fermentation liquor at any time of the 0-16 hours of fermentation. The fermentation production process improves the growth state of a gluconobacter melanogenus cell and the quality of the strain liquid, adding allogenic materials, namely the citric acid and the itaconic, in the fermentation process promotes the cellular metabolism of the gluconobacter melanogenus, so that the formation rate of the L-sorbose is improved in the fermentation process, the content of the L-sorbose after the fermentation is more than 420 mg/ml, and the purposes of improving the fermentation efficiency and shortening the fermentation cycle are realized.
Description
Technical field
The invention belongs to ascorbic preparing technical field, particularly a kind of high density L-sorbose fermentation manufacturing technique.
Background technology
Vitamins C is the necessary a kind of water-soluble vitamins of human body, has oxidation-resistance, improves the effects such as immunity of organisms, be widely used in the fields such as medicine, food, makeup.
In current industrial vitamin C production technique, generating L-sorbose by D-glucitol fermentation is an indispensable step in production of vitamin C operational path, raw Black Grape saccharic acid bacillus (Gluconobacter melagenus) of industrial general employing is as producing bacterium, produce in the process of L-sorbose at high concentration substrate situation bottom fermentation, still there is the technical problem that thalli growth is suppressed, fermentation period is long, the high restriction of energy consumption is produced, therefore, improve original fermentation manufacturing technique, improve L-sorbose fermentation efficiency and be very important.
In order to solve the problem, our company develops a kind of working method (publication number is CN103114111A) of the L-of raising sorbose fermentation productivity, the invention provides a kind of employing raw Black Grape saccharic acid bacillus for producing bacterial strain, be that fermenting substrate produces L-sorbose with D-glucitol, in the fermentation culture any time of 0 ~ 12 hour, in fermented liquid, add 0.01% ~ 0.05% reduced glutathion, the biosynthesizing speed ratio prior art of L-sorbose can improve 31.2% at most; In the fermentation culture any time of 0 ~ 12 hour, in fermented liquid, add 0.01% ~ 0.05% casamino acid, the biosynthesizing speed ratio prior art of L-sorbose can improve 27.9%.Effectively can shorten L-sorbose fermentation period by above technical scheme, improve vitamin c fermenting production efficiency.Also following data are listed: ferment in embodiment 1 24 hours sorbose content 287.2mg/ml in the embodiment scheme of this patent; Ferment in embodiment 2 26 hours sorbose content 307.8mg/ml; Ferment in embodiment 3 24 hours sorbose content 270.6mg/ml; In embodiment 4,25 hours fermentation record sorbose content is to terminal 315.4g/L.In actual use, the maximum sorbose content obtained after program fermentation is only 315.4g/L, still there is the problem that fermentation concentration is low.
Summary of the invention
Object of the present invention will solve the problems of the technologies described above, and provides a kind of generating rate comparatively to contrast and can improve at most, effectively shortens a kind of high density L-sorbose fermentation manufacturing technique of L-sorbose fermentation period.
In order to realize above-mentioned object, this invention takes following technical solution: the L-sorbose high-efficiency fermenting production technique under a kind of high concentration substrate condition, is characterized in that:
Its processing step is as follows:
1, bacterial classification: raw Black Grape saccharic acid bacillus;
2, culture medium prescription:
Shaking flask kind liquid culture medium (mg/ml): D-glucitol 150, yeast extract paste 5, peptone 5, calcium carbonate 1.5; Glacial acetic acid regulates pH to 4.8 ~ 5.4,121 DEG C of autoclaving 25min;
Plant liquid enlarged culturing base (mg/ml): D-glucitol 150, yeast extract paste 3, peptone 3, calcium carbonate 1.5; Glacial acetic acid regulates pH to 4.8 ~ 5.4,121 DEG C of autoclaving 25min;
Fermentation initial medium (mg/ml): D-glucitol 450, yeast extract paste 1.5, peptone 1.5, calcium carbonate 1.5; Glacial acetic acid regulates pH to 4.8 ~ 5.4,121 DEG C of autoclaving 25min;
3, shaking flask kind liquid preparation:
From the appropriate raw Black Grape saccharic acid bacillus of picking slant medium in kind of liquid culture medium, shaking flask volume 750ml, liquid amount 250ml, 32 ± 1 DEG C, shaking speed 230r/min, cultivates 14h for subsequent use as kind of liquid, controls kind of a liquid L-sorbose content 80mg/ml ~ 120mg/ml;
4, secondary kind liquid culturing process controls:
Inoculum size is 1%, culture temperature 33 ± 1 DEG C, cultivates 16h for subsequent use as kind of liquid, controls kind of a liquid L-sorbose content 80mg/ml ~ 120mg/ml;
5, fermentation processes:
Initial pH value 4.8 ~ 5.4, culture temperature 34 ± 1 DEG C, stirring velocity 500 ~ 800r/min, in any time of fermentation culture 0 ~ 16h, adds 0.1mg/ml ~ 5mg/ml citric acid, 0.1mg/ml ~ 5mg/ml methylene-succinic acid in fermented liquid, continues fermentation to terminal;
The present invention adds 0.1mg/ml ~ 5mg/ml gelatin in secondary kind liquid culture medium formula, during L-sorbose fermentation culture 0 ~ 16h, add 0.1mg/ml ~ 5mg/ml citric acid, in its fermented liquid, the generating rate of L-sorbose comparatively contrasts and can improve 19.82% at most; During L-sorbose fermentation culture 0 ~ 16h, add 0.1mg/ml ~ 5mg/ml methylene-succinic acid, in its fermented liquid, the generating rate of L-sorbose comparatively contrasts and can improve 24.56% at most; During L-sorbose fermentation culture 0 ~ 16h, add 0.1mg/ml ~ 5mg/ml citric acid, 0.1mg/ml ~ 5mg/ml methylene-succinic acid, in its fermented liquid, the generating rate of L-sorbose comparatively contrasts and can improve 26.32% at most.
The present invention is by adding allogenic material gelatin in L-sorbose fermentation kind of liquid culturing process, improving raw Black Grape saccharic acid bacilli-cell growth conditions, improve kind of a liquid quality; Allogenic material citric acid, methylene-succinic acid is added in fermenting process, facilitate the metabolism of raw Black Grape saccharic acid bacilli-cell, thus improve L-sorbose generating rate in fermenting process, after fermentation, L-sorbose content is all greater than 420mg/ml, reach the object improving fermentation efficiency and shorten fermentation period, apply and there is good economic and social benefit.
Embodiment
The invention will be further described below, but not as limitation of the present invention:
Embodiment 1,1.5mg/ml gelatin is added in former secondary kind liquid culture medium, add the citric acid of 2.5mg/ml in the fermentation medium, 10L Fermentation is utilized to cultivate raw Black Grape saccharic acid bacillus secondary kind liquid, by cultured secondary kind liquid 7.5L, access is containing in the 100L Fermentation of 50L fermention medium, initial pH value 5.2, culture temperature 34 DEG C, stirring velocity 650r/min, ferments 39h to terminal, L-sorbose content 427.20mg/ml, produce sugared speed 10.95mg/ml/h, L-sorbose throughput rate comparatively to contrast and improve 17.87%.
Embodiment 2,2.5mg/ml gelatin is added in former secondary kind liquid culture medium, add the methylene-succinic acid of 2mg/ml in the fermentation medium, 10L Fermentation is utilized to cultivate raw Black Grape saccharic acid bacillus secondary kind liquid, by cultured secondary kind liquid 7.5L, access is containing in the 100L Fermentation of 50L fermention medium, initial pH value 5.3, culture temperature 34 DEG C, stirring velocity 650r/min, ferments 38h to terminal, L-sorbose content 423.90mg/ml, produce sugared speed 11.16mg/ml/h, L-sorbose throughput rate comparatively to contrast and improve 21.04%.
Embodiment 3, 4mg/ml gelatin is added in former secondary kind liquid culture medium, add the citric acid of 0.5mg/ml in the fermentation medium, the methylene-succinic acid of 2.5mg/ml, 10L Fermentation is utilized to cultivate raw Black Grape saccharic acid bacillus secondary kind liquid, by cultured secondary kind liquid 7.5L, access is containing in the 100L Fermentation of 50L fermention medium, initial pH value 5.2, culture temperature 34 DEG C, stirring velocity 650r/min, fermentation 37h to terminal, L-sorbose content 422.7mg/ml, produce sugared speed 11.42mg/ml/h, L-sorbose throughput rate comparatively contrasts and improves 24.40%.
Embodiment 4,2.5mg/ml gelatin is added in former secondary kind liquid culture medium, 10L Fermentation is utilized to cultivate raw Black Grape saccharic acid bacillus secondary kind liquid, by cultured secondary kind liquid 7.5L, access is containing in the 100L Fermentation of 50L fermention medium, initial pH value 5.3, culture temperature 34 DEG C, stirring velocity 650r/min; Cultivate after 4h, add the citric acid of 1mg/ml, the methylene-succinic acid of 0.5mg/ml in fermented liquid, to terminal, L-sorbose content 420.6mg/ml, produces sugared speed 11.07mg/ml/h, L-sorbose throughput rate and comparatively contrasts and improve 21.11% fermentation 38h.
Obviously, above-described embodiment is only for example of the present invention is clearly described, and the restriction not to embodiment.For those of ordinary skill in the field, can also make other changes in different forms on the basis of the above description.Here exhaustive without the need to also giving all embodiments.And thus the apparent change of amplifying out or variation be still among protection scope of the present invention.
Claims (5)
1. a high density L-sorbose fermentation manufacturing technique, it is characterized in that: utilize raw Black Grape saccharic acid bacillus to be fermentation strain, gelatin is added in kind of liquid culture medium formula, fermenting process D-glucitol is that substrate produces L-a kind of high density L-sorbose fermentation manufacturing technique according to claim 1, it is characterized in that: the addition of described gelatin is 0.1 mg/ml ~ 5mg/ml.
2. a kind of high density L-sorbose fermentation manufacturing technique according to claim 1, is characterized in that: the addition of described citric acid is 0.1mg/ml ~ 5mg/ml.
3. a kind of high density L-sorbose fermentation manufacturing technique according to claim 1, is characterized in that: the addition of described methylene-succinic acid is 0.1mg/ml ~ 5mg/ml.
4. a kind of high density L-sorbose fermentation manufacturing technique according to claim 1, is characterized in that: described citric acid, methylene-succinic acid are for adding separately.
5. a kind of high density L-sorbose fermentation manufacturing technique according to claim 1, is characterized in that: the combination of described citric acid, methylene-succinic acid is added.
Priority Applications (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410561050.7A CN104357507B (en) | 2014-10-21 | 2014-10-21 | A kind of high concentration L sorbose fermentation manufacturing techniques |
| PCT/CN2015/092248 WO2016062232A1 (en) | 2014-10-21 | 2015-10-20 | A fermentation production process of l-sorbose with high concentration |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410561050.7A CN104357507B (en) | 2014-10-21 | 2014-10-21 | A kind of high concentration L sorbose fermentation manufacturing techniques |
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| Publication Number | Publication Date |
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| CN104357507A true CN104357507A (en) | 2015-02-18 |
| CN104357507B CN104357507B (en) | 2017-08-22 |
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| Country | Link |
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| WO (1) | WO2016062232A1 (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2016062232A1 (en) * | 2014-10-21 | 2016-04-28 | Dsm Jiangshan Pharmaceutical (Jiangsu) Co., Ltd. | A fermentation production process of l-sorbose with high concentration |
| CN113913354A (en) * | 2021-12-03 | 2022-01-11 | 山东天力药业有限公司 | Method for promoting growth and metabolism of gluconobacter nigricans |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114045243B (en) * | 2021-12-03 | 2023-12-26 | 山东天力药业有限公司 | Method for shortening fermentation period of gluconobacter nigrum |
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| CN87100655A (en) * | 1986-02-11 | 1987-09-23 | 武田药品工业株式会社 | Method for producing L-sorbose |
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| CN104357507B (en) * | 2014-10-21 | 2017-08-22 | 帝斯曼江山制药（江苏）有限公司 | A kind of high concentration L sorbose fermentation manufacturing techniques |
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2014
- 2014-10-21 CN CN201410561050.7A patent/CN104357507B/en active Active
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2015
- 2015-10-20 WO PCT/CN2015/092248 patent/WO2016062232A1/en active Application Filing
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2016062232A1 (en) * | 2014-10-21 | 2016-04-28 | Dsm Jiangshan Pharmaceutical (Jiangsu) Co., Ltd. | A fermentation production process of l-sorbose with high concentration |
| CN113913354A (en) * | 2021-12-03 | 2022-01-11 | 山东天力药业有限公司 | Method for promoting growth and metabolism of gluconobacter nigricans |
| CN113913354B (en) * | 2021-12-03 | 2023-10-10 | 山东天力药业有限公司 | Method for promoting growth metabolism of gluconobacter nigrum |
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| Publication number | Publication date |
|---|---|
| CN104357507B (en) | 2017-08-22 |
| WO2016062232A1 (en) | 2016-04-28 |
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Address after: 214500, 61 Jiangshan Road, Taizhou, Jiangsu, Jingjiang Applicant after: DSM JIANGSHAN PHARMACEUTICAL (JIANGSU) CO.,LTD. Address before: 214500, 61 Jiangshan Road, Taizhou, Jiangsu, Jingjiang Applicant before: DSM JIANGSHAN PHARMACEUTICAL (JIANGSU) Co.,Ltd. |
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Address after: 214500 No.61 Jiangshan Road, Jingjiang City, Taizhou City, Jiangsu Province Patentee after: DSM JIANGSHAN PHARMACEUTICAL (JIANGSU) Co.,Ltd. Country or region after: China Address before: 214500 No.61 Jiangshan Road, Jingjiang City, Taizhou City, Jiangsu Province Patentee before: DSM JIANGSHAN PHARMACEUTICAL (JIANGSU) CO.,LTD. Country or region before: China |