CN103589759B - A kind of waste molasses and the two carbon source through fermentation of sewer oil produce the method for sophorolipid - Google Patents
A kind of waste molasses and the two carbon source through fermentation of sewer oil produce the method for sophorolipid Download PDFInfo
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- CN103589759B CN103589759B CN201310590289.2A CN201310590289A CN103589759B CN 103589759 B CN103589759 B CN 103589759B CN 201310590289 A CN201310590289 A CN 201310590289A CN 103589759 B CN103589759 B CN 103589759B
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- sophorolipid
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Abstract
The invention discloses a kind of method of waste molasses and sewer oil two carbon source through fermentation production sophorolipid, it is characterized in that, described method comprises the steps: that torulopsis bacterium is inoculated into bacteria suspension processed (2) seed liquor processed (3) on YPD slant medium and seed liquor is inoculated in and extracts sophorolipid with fermentation (4) ethyl acetate in waste molasses and the sewer oil fermention medium that is substrate by (1).The present invention realizes the recycling of waste, greatly reduces the production cost of sophorolipid.Solve sewer oil exhaust emission greatly, again to enter edible field and endanger large problem, achieve the rational utilization of sewer oil resource.This technique does not need complicated equipment, and fermenting process is simple, is conducive to the exploitation of sophorolipid, application and popularization.
Description
Technical field
The invention belongs to the preparing technical field of bio-surfactant, be specifically related to a kind of method of waste molasses and sewer oil two carbon source through fermentation production sophorolipid.
Background technology
Tensio-active agent is except in daily life as washing composition, and other application almost can cover all field of fine chemical.Sophorolipid is a kind of environmentally friendly bio-surfactant produced by microorganism, compared with the tensio-active agent produced, except total characteristic, also has himself advantage with chemosynthesis or refining of petroleum method.Sophorolipid is glycolipid system bio-surfactant, has toxicity low, with human body and Environmental compatibility good, have the characteristics such as good emulsifying, dispersion, solubilising; There is in fields such as food, medicine, makeup, metallurgy, environment protection, waste oil recovery research and development be worth.The structural stability of sophorolipid is high, can obtain various derivative by it, and being has application future most in glycolipid system bio-surfactant.In foodstuffs industry, petroleum industry, environment protection, makeup, washing composition and pharmacy, obtained at present application in various degree, carried out for many years the theoretical investigation of sophorolipid and application and development in the world, China is also in the starting stage in this field at present.
Fermention medium component is the important factor determining sophorolipid bio-surfactant production cost.At present, sophorolipid fermentation mostly adopts oil soluble and water-soluble pair of carbon source through fermentation technology, and wherein, oil soluble carbon source mainly comprises oleic acid, rapeseed oil, peanut oil, soybean wet goods, and water-soluble carbon source is generally glucose.The price of above-mentioned carbon source own is higher, causes the production cost of sophorolipid can not to be in any more, is unfavorable for applying of sophorolipid.
Waste molasses is a kind of byproduct of sugar refinery, not only containing the fermentable sugars up to 40-55%, and containing Na
+, K
+ca
2+be conducive to the nutritive substance of microorganism growth Deng the necessary element of microorganism growth and VITAMIN, protein etc., can be used as the substitute of the water-soluble carbon sources such as glucose in sophorolipid fermentative production.Along with the development of China's sugar industry, the output of waste molasses increases day by day, abundance, is the very good material of large-scale industry fermentative production.
Sewer oil also claims waste cooking oil or recycled wood materials, main component is palmitinic acid, stearic acid, oleic acid and linolic acid, its lipid acid composition is substantially the same with other animal-plant oil, carbon chain lengths, at 14 ~ 18, can be used as the substitute of peanut wet goods oil soluble carbon source in sophorolipid fermentative production.At present, along with the raising that people's unboiled water is flat, the output of sewer oil is also increasing, and according to incompletely statistics, the annual production of China's sewer oil is at 400 ~ 8,000,000 tons; As recycled not in time, not only cause serious environmental pollution, and edible oil market can be entered by the processing backflow of some illegal enterprises, cause serious harm to people are healthy.
Bio-surfactant is as a kind of novel green tensio-active agent, have a good application prospect in fields such as daily-use chemical industry, oil production, environment remediation, at present, high production cost is the key of restriction sophorolipid bio-surfactant in each industrial trade widespread use.
the report that current employing sewer oil makes hydrophobic carbon source on the one hand concentrates in detection method, and in the esterification mainly concentrating on biofuel and lipid acid on the other hand and saponification, that carries out fermenting as substrate have not been reported.therefore, how developing with waste is the method that fermenting substrate prepares sophorolipid, and while turning waste into wealth, improving its utility value, greatly reduce the production cost of sophorolipid, is the technical problem to be solved in the present invention.
Summary of the invention
The object of the invention is to the weak point overcoming above-mentioned prior art, provide the two carbon source through fermentation of a kind of waste molasses and sewer oil to produce the method for sophorolipid, is a kind of effective and can reduce sophorolipid production cost, be easy to the method for production application.
For achieving the above object, the invention provides a kind of method of waste molasses and sewer oil two carbon source through fermentation production sophorolipid, it is characterized in that, described method comprises the steps:
(1) preparation of bacteria suspension: by torulopsis bacterium (
torulopsisbombicola) ATCC22214 is inoculated on the YPD slant medium that is made up of glucose, peptone, yeast powder and agar, is settled to 1L, pH=6.0 with distilled water;
In 30 DEG C of constant temperature culture 2-3 days, be the physiological saline wash-out thalline of 0.85% by appropriate mass concentration, obtained bacteria suspension;
(2) seed liquor processed: the shaking flask seed culture medium 100ml be made up of glucose, yeast powder and urea being placed in 500ml, 121 DEG C of sterilizing 20min, above-mentioned bacteria suspension is accessed according to the inoculum size of volume percent 3-5% after cooling, then shaking flask is placed in 25-32 DEG C, 24h cultivated by the shaking table of 120-180rpm, obtain seed liquor;
(3) ferment: be used for above-mentioned seed liquor inoculating aseptic 5L fermentor tank with in waste molasses and the sewer oil fermention medium that is substrate, 3L is settled to tap water, pH=6.0, by fermentation culture based on 121 DEG C of sterilizing 20min, by in above-mentioned seed liquor access fermention medium after cooling, keep leavening temperature 25-32 DEG C, dissolved oxygen 20-60%, ferments;
(4) sophorolipid processed: after fermentation ends, extracts sophorolipid with the isopyknic organic solvent ethyl acetate of fermented liquid, and underpressure distillation removing organic solvent, obtains sophorolipid.
Slant medium composition (g/L) in above-mentioned steps (1): glucose 20, peptone 20, yeast powder 10, agar 15-20.
In bacteria suspension in above-mentioned steps (1), mycetocyte content is about 10
7-10
8individual/mL.
Seed culture medium composition (g/L) in above-mentioned steps (2): glucose 60-100, yeast powder 5-8, urea 0.5-1.
Fermention medium composition (g/L) in above-mentioned steps (3): sewer oil 60-120, waste molasses 60-180, yeast powder 3-5.
The inoculum size of seed liquor according to volume ratio 3-5% is accessed in fermention medium in above-mentioned steps (3).
Fermentation time in above-mentioned steps (3) is 72-120h.
In above-mentioned steps (4), the output of sophorolipid is 30-100g/L.
In above-mentioned steps (3)
it is refining sewer oil (buying on the market) that sewer oil have employed two kinds: one, and two is reclaim in the trench dining room, merely through simple centrifugal treating, gets upper strata oil reservoir and makes raw material.
Beneficial effect of the present invention:
1, the present invention adopts waste molasses and the two carbon source through fermentation of sewer oil to produce the Technology of sophorolipid, realizes the recycling of waste, reduces environmental pollution improvement's expense, greatly reduce the production cost of sophorolipid.
2, solve sewer oil exhaust emission greatly, again to enter edible field and endanger large problem, open " bottleneck " that sewer oil is recycled, effective guarantee people are healthy, achieve the rational utilization of sewer oil resource.
3, this technique does not need complicated equipment, and fermenting process is simple, is conducive to the exploitation of sophorolipid, application and popularization.
4, the method can also carry out the production of sophorolipid by continuous feeding fermentation, and actual application value is higher, wide market.
Embodiment
Embodiment 1
(1) by torulopsis (
torulopsisbombicola) ATCC22214 is inoculated on YPD slant medium, 30 DEG C of constant temperature culture 2 days, with for 0.85% physiological saline wash-out thalline, make cell content and be about 5.3 × 10
7the bacteria suspension of individual/mL;
(2) preparation of seed liquor: seed culture medium composition (g/L): glucose 60, yeast powder 5, urea 0.5, pH6.0; This seed culture medium of 100mL is placed in 500mL shaking flask, and 121 DEG C of sterilizing 20min, access above-mentioned bacteria suspension according to the inoculum size of volume percent 3% after cooling, shaking flask are placed in 25 DEG C, 24h cultivated by the shaking table of rotating speed 120rpm, obtains seed liquor;
(3) above-mentioned seed liquor is for inoculating aseptic 5L fermentor tank with in waste molasses and the sewer oil fermention medium that is substrate.Fermention medium composition (g/L): sewer oil 60, waste molasses 60, yeast powder 3, pH=6.0, tap water is settled to 3L, 121 DEG C of sterilizing 20min, according to the inoculum size of volume ratio 3%, above-mentioned seed liquor is accessed in fermention medium after cooling, leavening temperature 30 DEG C, dissolved oxygen is 20%, fermentation time 72h;
(4) after fermentation ends, extract sophorolipid with the isopyknic ethyl acetate of fermented liquid, underpressure distillation removing organic solvent, obtaining sophorolipid output is 33.5g/L.
Embodiment 2
(1) by torulopsis (
torulopsisbombicola) ATCC22214 is inoculated on YPD slant medium, 30 DEG C of constant temperature culture 2 days, with for 0.85% physiological saline wash-out thalline, make cell content and be about 6.2 × 10
7the bacteria suspension of individual/mL;
(2) preparation of seed liquor: seed culture medium composition (g/L): glucose 80, yeast powder 6, urea 0.8, pH6.0; This seed culture medium of 100mL is placed in 500mL shaking flask, and 121 DEG C of sterilizing 20min, access above-mentioned bacteria suspension according to the inoculum size of volume percent 4% after cooling, shaking flask are placed in 28 DEG C, 24h cultivated by the shaking table of rotating speed 150rpm, obtains seed liquor;
(3) above-mentioned seed liquor is for inoculating aseptic 5L fermentor tank with in waste molasses and the sewer oil fermention medium that is substrate.Fermention medium composition (g/L): sewer oil 80, waste molasses 100, yeast powder 4, pH=6.0, tap water is settled to 3L, 121 DEG C of sterilizing 20min, according to the inoculum size of volume ratio 4%, above-mentioned seed liquor is accessed in fermention medium after cooling, leavening temperature 28 DEG C, dissolved oxygen is 35%, fermentation time 84h;
(4) after fermentation ends, extract sophorolipid with the isopyknic ethyl acetate of fermented liquid, underpressure distillation removing organic solvent, obtaining sophorolipid output is 59.0g/L.
Embodiment 3
(1) by torulopsis (
torulopsisbombicola) ATCC22214 is inoculated on YPD slant medium, 30 DEG C of constant temperature culture 2 days, with for 0.85% physiological saline wash-out thalline, make cell content and be about 7.4 × 10
7the bacteria suspension of individual/mL;
(2) preparation of seed liquor: seed culture medium composition (g/L): glucose 100, yeast powder 7, urea 1, pH6.0; This seed culture medium of 100mL is placed in 500mL shaking flask, and 121 DEG C of sterilizing 20min, access above-mentioned bacteria suspension according to the inoculum size of volume percent 5% after cooling, shaking flask are placed in 30 DEG C, 24h cultivated by the shaking table of rotating speed 180rpm, obtains seed liquor;
(3) above-mentioned seed liquor is for inoculating aseptic 5L fermentor tank with in waste molasses and the sewer oil fermention medium that is substrate.Fermention medium composition (g/L): sewer oil 100, waste molasses 160, yeast powder 4, pH=6.0, tap water is settled to 3L, 121 DEG C of sterilizing 20min, according to the inoculum size of volume ratio 4%, above-mentioned seed liquor is accessed in fermention medium after cooling, leavening temperature 30 DEG C, dissolved oxygen is 45%, fermentation time 96h;
(4) after fermentation ends, extract sophorolipid with the isopyknic ethyl acetate of fermented liquid, underpressure distillation removing organic solvent, obtaining sophorolipid output is 83.4g/L.
Embodiment 4
(1) by torulopsis (
torulopsisbombicola) ATCC22214 is inoculated on YPD slant medium, 30 DEG C of constant temperature culture 2 days, with for 0.85% physiological saline wash-out thalline, make cell content and be about 8.5 × 10
7the bacteria suspension of individual/mL;
(2) preparation of seed liquor: seed culture medium composition (g/L): glucose 80, yeast powder 8, urea 0.8, pH6.0; This seed culture medium of 100mL is placed in 500mL shaking flask, and 121 DEG C of sterilizing 20min, access above-mentioned bacteria suspension according to the inoculum size of volume percent 5% after cooling, shaking flask are placed in 32 DEG C, 24h cultivated by the shaking table of rotating speed 180rpm, obtains seed liquor;
(3) above-mentioned seed liquor is for inoculating aseptic 5L fermentor tank with in waste molasses and the sewer oil fermention medium that is substrate.Fermention medium composition (g/L): sewer oil 120, waste molasses 180, yeast powder 5, pH=6.0, tap water is settled to 3L, 121 DEG C of sterilizing 20min, according to the inoculum size of volume ratio 5%, above-mentioned seed liquor is accessed in fermention medium after cooling, leavening temperature 32 DEG C, dissolved oxygen is 60%, fermentation time 120h;
(4) after fermentation ends, extract sophorolipid with the isopyknic ethyl acetate of fermented liquid, underpressure distillation removing organic solvent, obtaining sophorolipid output is 99.3g/L.
Claims (4)
1. produce a method for sophorolipid with waste molasses and the two carbon source through fermentation of sewer oil, it is characterized in that, described method comprises the steps:
(1) preparation of bacteria suspension: by torulopsis bacterium
aTCC22214 is inoculated on the YPD slant medium that is made up of glucose, peptone, yeast powder and agar, is settled to 1L, pH=6.0 with distilled water; In 30 DEG C of constant temperature culture 2-3 days, be the physiological saline wash-out thalline of 0.85% by appropriate mass concentration, obtained bacteria suspension; Slant medium composition (g/L): glucose 20, peptone 20, yeast powder 10, agar 15-20; In bacteria suspension, mycetocyte content is 10
7-10
8individual/mL;
(2) seed liquor processed: the shaking flask seed culture medium 100ml be made up of glucose, yeast powder and urea being placed in 500ml, 121 DEG C of sterilizing 20min, above-mentioned bacteria suspension is accessed according to the inoculum size of volume percent 3-5% after cooling, then shaking flask is placed in 25-32 DEG C, 24h cultivated by the shaking table of 120-180rpm, obtain seed liquor; Seed culture medium composition (g/L): glucose 60-100, yeast powder 5-8, urea 0.5-1;
(3) ferment: be used for above-mentioned seed liquor inoculating aseptic 5L fermentor tank with in waste molasses and the sewer oil fermention medium that is substrate, 3L is settled to tap water, pH=6.0, by fermentation culture based on 121 DEG C of sterilizing 20min, by in above-mentioned seed liquor access fermention medium after cooling, keep leavening temperature 25-32 DEG C, dissolved oxygen 20-60%, ferments; Fermention medium composition (g/L): sewer oil 60-120, waste molasses 60-180, yeast powder 3-5;
(4) sophorolipid processed: after fermentation ends, extracts sophorolipid with the isopyknic organic solvent ethyl acetate of fermented liquid, and underpressure distillation removing organic solvent, obtains sophorolipid.
2. a kind of waste molasses according to claim 1 and the two carbon source through fermentation of sewer oil produce the method for sophorolipid, it is characterized in that, the inoculum size of seed liquor according to volume ratio 3-5% being accessed in fermention medium in described step (3).
3. a kind of waste molasses according to claim 1 and the two carbon source through fermentation of sewer oil produce the method for sophorolipid, and it is characterized in that, the fermentation time in described step (3) is 72-120h.
4. a kind of waste molasses according to claim 1 and the two carbon source through fermentation of sewer oil produce the method for sophorolipid, and it is characterized in that, in described step (4), the output of sophorolipid is 30-100g/L.
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| CN105154050B (en) * | 2015-07-29 | 2018-06-29 | 中国石油化工股份有限公司 | A kind of heat-resistant salt-resistant viscous crude biology thinner and preparation method thereof |
| CN105671106B (en) * | 2016-02-16 | 2021-06-11 | 珀莱雅化妆品股份有限公司 | Method for preparing glycosphingolipid by adopting olive oil fermentation method |
| CN113151020B (en) * | 2021-05-12 | 2023-10-20 | 南京工业大学 | High-acid-yield sophorolipid feed-supplementing fermentation method |
| CN114807271B (en) * | 2022-05-06 | 2023-06-23 | 河南省科学院高新技术研究中心 | Method for preparing sophorolipid by fermentation and application of sophorolipid in reverse osmosis membrane medicament |
| CN115992184B (en) * | 2023-03-23 | 2023-06-16 | 科乐美(广州)生物科技有限公司 | Biological fermentation surfactant, cosmetic and preparation method thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102329833A (en) * | 2011-06-23 | 2012-01-25 | 天津实发中科百奥工业生物技术有限公司 | Method for producing sophorose ester through fermenting waste molasses and waste glycerin |
| CN102492753A (en) * | 2011-12-07 | 2012-06-13 | 山东大学 | Method for producing sophorolipid through fermentation of lignocellulose material |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102329833A (en) * | 2011-06-23 | 2012-01-25 | 天津实发中科百奥工业生物技术有限公司 | Method for producing sophorose ester through fermenting waste molasses and waste glycerin |
| CN102492753A (en) * | 2011-12-07 | 2012-06-13 | 山东大学 | Method for producing sophorolipid through fermentation of lignocellulose material |
Non-Patent Citations (1)
| Title |
|---|
| Utilization of Restaurant Waste Oil as a Precursor for Sophorolipid Production;Vishal Shah, et al.;《Biotechnol. Prog.》;20070430;第23卷(第2期);摘要，第513页左栏第2-3段，右栏第2段，第515页左栏第1段 * |
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