CN103589759A - Method for producing sophorolipid by double carbon source fermentation of waste molasses and illegal cooking oil - Google Patents
Method for producing sophorolipid by double carbon source fermentation of waste molasses and illegal cooking oil Download PDFInfo
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- CN103589759A CN103589759A CN201310590289.2A CN201310590289A CN103589759A CN 103589759 A CN103589759 A CN 103589759A CN 201310590289 A CN201310590289 A CN 201310590289A CN 103589759 A CN103589759 A CN 103589759A
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- sophorolipid
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- waste molasses
- carbon source
- sewer oil
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- ZTOKUMPYMPKCFX-CZNUEWPDSA-N (E)-17-[(2R,3R,4S,5S,6R)-6-(acetyloxymethyl)-3-[(2S,3R,4S,5S,6R)-6-(acetyloxymethyl)-3,4,5-trihydroxyoxan-2-yl]oxy-4,5-dihydroxyoxan-2-yl]oxyoctadec-9-enoic acid Chemical compound OC(=O)CCCCCCC/C=C/CCCCCCC(C)O[C@@H]1O[C@H](COC(C)=O)[C@@H](O)[C@H](O)[C@H]1O[C@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](COC(C)=O)O1 ZTOKUMPYMPKCFX-CZNUEWPDSA-N 0.000 title claims abstract description 51
- 239000002699 waste material Substances 0.000 title claims abstract description 36
- 238000000855 fermentation Methods 0.000 title claims abstract description 35
- 230000004151 fermentation Effects 0.000 title claims abstract description 35
- 235000013379 molasses Nutrition 0.000 title claims abstract description 30
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 title claims abstract description 23
- 229910052799 carbon Inorganic materials 0.000 title claims abstract description 22
- 238000004519 manufacturing process Methods 0.000 title abstract description 13
- 239000008162 cooking oil Substances 0.000 title abstract description 6
- 239000002609 medium Substances 0.000 claims abstract description 29
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims abstract description 21
- 238000000034 method Methods 0.000 claims abstract description 21
- 240000004808 Saccharomyces cerevisiae Species 0.000 claims abstract description 19
- 239000000725 suspension Substances 0.000 claims abstract description 17
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 claims abstract description 15
- 239000001963 growth medium Substances 0.000 claims abstract description 13
- 239000000758 substrate Substances 0.000 claims abstract description 9
- 239000001888 Peptone Substances 0.000 claims abstract description 5
- 108010080698 Peptones Proteins 0.000 claims abstract description 5
- 235000019319 peptone Nutrition 0.000 claims abstract description 5
- 241000894006 Bacteria Species 0.000 claims description 18
- 239000000843 powder Substances 0.000 claims description 18
- 239000008103 glucose Substances 0.000 claims description 14
- 241000222120 Candida <Saccharomycetales> Species 0.000 claims description 12
- 238000001816 cooling Methods 0.000 claims description 12
- 239000002054 inoculum Substances 0.000 claims description 12
- 238000011218 seed culture Methods 0.000 claims description 12
- 230000001954 sterilising effect Effects 0.000 claims description 12
- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 claims description 8
- 239000004202 carbamide Substances 0.000 claims description 8
- 239000003960 organic solvent Substances 0.000 claims description 8
- FRXSZNDVFUDTIR-UHFFFAOYSA-N 6-methoxy-1,2,3,4-tetrahydroquinoline Chemical compound N1CCCC2=CC(OC)=CC=C21 FRXSZNDVFUDTIR-UHFFFAOYSA-N 0.000 claims description 6
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 claims description 6
- 238000004821 distillation Methods 0.000 claims description 6
- 239000007788 liquid Substances 0.000 claims description 6
- 229910052760 oxygen Inorganic materials 0.000 claims description 6
- 239000001301 oxygen Substances 0.000 claims description 6
- 239000002504 physiological saline solution Substances 0.000 claims description 6
- 238000002360 preparation method Methods 0.000 claims description 6
- 239000008399 tap water Substances 0.000 claims description 6
- 235000020679 tap water Nutrition 0.000 claims description 6
- 229920001817 Agar Polymers 0.000 claims description 4
- 239000008272 agar Substances 0.000 claims description 4
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 3
- 239000012153 distilled water Substances 0.000 claims description 2
- 238000011161 development Methods 0.000 abstract description 3
- 235000013305 food Nutrition 0.000 abstract description 3
- 241000235646 Cyberlindnera jadinii Species 0.000 abstract 1
- 230000001580 bacterial effect Effects 0.000 abstract 1
- 239000008121 dextrose Substances 0.000 abstract 1
- 239000003921 oil Substances 0.000 description 31
- 235000019198 oils Nutrition 0.000 description 31
- 239000003876 biosurfactant Substances 0.000 description 7
- 239000013543 active substance Substances 0.000 description 3
- 238000005516 engineering process Methods 0.000 description 3
- 238000012262 fermentative production Methods 0.000 description 3
- 244000005700 microbiome Species 0.000 description 3
- 239000000203 mixture Substances 0.000 description 3
- 235000000346 sugar Nutrition 0.000 description 3
- WRIDQFICGBMAFQ-UHFFFAOYSA-N (E)-8-Octadecenoic acid Natural products CCCCCCCCCC=CCCCCCCC(O)=O WRIDQFICGBMAFQ-UHFFFAOYSA-N 0.000 description 2
- LQJBNNIYVWPHFW-UHFFFAOYSA-N 20:1omega9c fatty acid Natural products CCCCCCCCCCC=CCCCCCCCC(O)=O LQJBNNIYVWPHFW-UHFFFAOYSA-N 0.000 description 2
- HVCOBJNICQPDBP-UHFFFAOYSA-N 3-[3-[3,5-dihydroxy-6-methyl-4-(3,4,5-trihydroxy-6-methyloxan-2-yl)oxyoxan-2-yl]oxydecanoyloxy]decanoic acid;hydrate Chemical compound O.OC1C(OC(CC(=O)OC(CCCCCCC)CC(O)=O)CCCCCCC)OC(C)C(O)C1OC1C(O)C(O)C(O)C(C)O1 HVCOBJNICQPDBP-UHFFFAOYSA-N 0.000 description 2
- QSBYPNXLFMSGKH-UHFFFAOYSA-N 9-Heptadecensaeure Natural products CCCCCCCC=CCCCCCCCC(O)=O QSBYPNXLFMSGKH-UHFFFAOYSA-N 0.000 description 2
- 229930186217 Glycolipid Natural products 0.000 description 2
- ZQPPMHVWECSIRJ-UHFFFAOYSA-N Oleic acid Natural products CCCCCCCCC=CCCCCCCCC(O)=O ZQPPMHVWECSIRJ-UHFFFAOYSA-N 0.000 description 2
- 239000005642 Oleic acid Substances 0.000 description 2
- 239000002253 acid Substances 0.000 description 2
- 239000012141 concentrate Substances 0.000 description 2
- 238000003912 environmental pollution Methods 0.000 description 2
- QXJSBBXBKPUZAA-UHFFFAOYSA-N isooleic acid Natural products CCCCCCCC=CCCCCCCCCC(O)=O QXJSBBXBKPUZAA-UHFFFAOYSA-N 0.000 description 2
- 150000002632 lipids Chemical class 0.000 description 2
- ZQPPMHVWECSIRJ-KTKRTIGZSA-N oleic acid Chemical compound CCCCCCCC\C=C/CCCCCCCC(O)=O ZQPPMHVWECSIRJ-KTKRTIGZSA-N 0.000 description 2
- 239000003208 petroleum Substances 0.000 description 2
- 238000007670 refining Methods 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 238000005406 washing Methods 0.000 description 2
- 235000017060 Arachis glabrata Nutrition 0.000 description 1
- 244000105624 Arachis hypogaea Species 0.000 description 1
- 235000010777 Arachis hypogaea Nutrition 0.000 description 1
- 235000018262 Arachis monticola Nutrition 0.000 description 1
- 244000068988 Glycine max Species 0.000 description 1
- 235000010469 Glycine max Nutrition 0.000 description 1
- OYHQOLUKZRVURQ-HZJYTTRNSA-N Linoleic acid Chemical compound CCCCC\C=C/C\C=C/CCCCCCCC(O)=O OYHQOLUKZRVURQ-HZJYTTRNSA-N 0.000 description 1
- 235000019483 Peanut oil Nutrition 0.000 description 1
- 235000019484 Rapeseed oil Nutrition 0.000 description 1
- 235000021355 Stearic acid Nutrition 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 239000002551 biofuel Substances 0.000 description 1
- 239000006227 byproduct Substances 0.000 description 1
- 239000000306 component Substances 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 239000006185 dispersion Substances 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 239000008157 edible vegetable oil Substances 0.000 description 1
- 230000001804 emulsifying effect Effects 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 230000032050 esterification Effects 0.000 description 1
- 238000005886 esterification reaction Methods 0.000 description 1
- 239000012847 fine chemical Substances 0.000 description 1
- IPCSVZSSVZVIGE-UHFFFAOYSA-N hexadecanoic acid Chemical group CCCCCCCCCCCCCCCC(O)=O IPCSVZSSVZVIGE-UHFFFAOYSA-N 0.000 description 1
- 230000002209 hydrophobic effect Effects 0.000 description 1
- 238000011835 investigation Methods 0.000 description 1
- 229960004232 linoleic acid Drugs 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 239000012533 medium component Substances 0.000 description 1
- 238000005272 metallurgy Methods 0.000 description 1
- 230000000050 nutritive effect Effects 0.000 description 1
- QIQXTHQIDYTFRH-UHFFFAOYSA-N octadecanoic acid Chemical compound CCCCCCCCCCCCCCCCCC(O)=O QIQXTHQIDYTFRH-UHFFFAOYSA-N 0.000 description 1
- OQCDKBAXFALNLD-UHFFFAOYSA-N octadecanoic acid Natural products CCCCCCCC(C)CCCCCCCCC(O)=O OQCDKBAXFALNLD-UHFFFAOYSA-N 0.000 description 1
- 235000021313 oleic acid Nutrition 0.000 description 1
- 235000020232 peanut Nutrition 0.000 description 1
- 239000000312 peanut oil Substances 0.000 description 1
- 239000010773 plant oil Substances 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 238000011084 recovery Methods 0.000 description 1
- 238000004064 recycling Methods 0.000 description 1
- 238000010992 reflux Methods 0.000 description 1
- 238000005067 remediation Methods 0.000 description 1
- 238000012827 research and development Methods 0.000 description 1
- 238000007127 saponification reaction Methods 0.000 description 1
- 239000008117 stearic acid Substances 0.000 description 1
- 150000008163 sugars Chemical class 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 239000011782 vitamin Substances 0.000 description 1
- 229940088594 vitamin Drugs 0.000 description 1
- 229930003231 vitamin Natural products 0.000 description 1
- 235000013343 vitamin Nutrition 0.000 description 1
- 150000003722 vitamin derivatives Chemical class 0.000 description 1
Landscapes
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention discloses a method for producing sophorolipid by double carbon source fermentation of waste molasses and illegal cooking oil. The method is characterized by comprising the following steps: (1) inoculating torulopsis utilis to a YPD (yeast peptone dextrose) slant culture medium to prepare a bacterial suspension; (2) preparing a seed solution; (3) inoculating the seed solution to a fermentation medium taking the waste molasses and the illegal cooking oil as substrates for fermenting; (4) extracting the sophorolipid by ethyl acetate. According to the method, wastes are recycled, the production cost of the sophorolipid is greatly reduced, the problems of high emission pollution and large harm in the field of foods for the illegal cooking oil are solved, illegal cooking oil resources are reasonably utilized, complex equipment is not required, a fermentation process is simple and easy to implement, and the development, application and popularization of the sophorolipid are facilitated.
Description
Technical field
The invention belongs to the preparing technical field of bio-surfactant, be specifically related to a kind of method of producing sophorolipid with waste molasses and the two carbon source through fermentation of sewer oil.
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 by microorganisms, compares with the tensio-active agent that chemosynthesis or refining of petroleum method are produced, and except total characteristic, also has himself advantage.Sophorolipid is that glycolipid is bio-surfactant, has toxicity low, good with human body and Environmental compatibility, has the characteristics such as good emulsifying, dispersion, solubilising; In fields such as food, medicine, makeup, metallurgy, environment protection, waste oil recovery, having research and development is worth.The structural stability of sophorolipid is high, by it, can make various derivatives, is that glycolipid is in bio-surfactant, to have application future most.At present in foodstuffs industry, petroleum industry, environment protection, makeup, washing composition and pharmacy, obtained application in various degree, in the world the theoretical investigation of sophorolipid and application and development have been carried out for many years, current this field of China is also in the starting stage.
Fermention medium component is the important factor that determines 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 contains the fermentable sugars up to 40-55%, and contains Na
+, K
+ca
2+the nutritive substance that is conducive to microorganism growth in 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 for sophorolipid fermentative production.Along with the development of China's sugar industry, the output of waste molasses increases day by day, and source is abundant, is the very good material of large-scale industry fermentative production.
Sewer oil also claims waste cooking oil or discarded food oils, 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 for sophorolipid fermentative production.At present, along with the flat raising of people's unboiled water, 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 may reflux and enter edible oil market by the processing of some illegal enterprises, to people are healthy, cause serious harm.
Bio-surfactant is as a kind of novel green tensio-active agent, in fields such as daily-use chemical industry, oil production, environment remediation, have a good application prospect, at present, high production cost is that restriction sophorolipid bio-surfactant is in the key of each industrial trade widespread use.
the report that adopts sewer oil to make hydrophobic carbon source at present concentrates in detection method on the one hand, mainly concentrates on the other hand in the esterification and saponification of biofuel and lipid acid, and what as substrate, ferment have not been reported.therefore, how developing and take waste and prepare the method for sophorolipid as fermenting substrate, when turning waste into wealth, improve 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 overcome the weak point of above-mentioned prior art, a kind of method of producing sophorolipid with the two carbon source through fermentation of waste molasses and sewer oil is provided, 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 producing 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 (
torulopsis bombicola) ATCC 22214 is inoculated on the YPD slant medium being comprised of glucose, peptone, yeast powder and agar, with distilled water, is settled to 1L, pH=6.0;
In 30 ℃ of constant temperature culture 2-3 days, the physiological saline wash-out thalline that is 0.85% by appropriate mass concentration, makes bacteria suspension;
(2) seed liquor processed: the shaking flask that the seed culture medium 100ml being comprised of glucose, yeast powder and urea is placed in to 500ml, 121 ℃ of sterilizing 20min, after cooling, according to the inoculum size of volume percent 3-5%, access above-mentioned bacteria suspension, then shaking flask is placed on 25-32 ℃, the shaking table of 120-180rpm and cultivates 24h, obtain seed liquor;
(3) fermentation: above-mentioned seed liquor be take to the fermention medium that waste molasses and sewer oil be substrate for inoculating aseptic 5L fermentor tank, with tap water, be settled to 3L, pH=6.0, by fermentation culture based on 121 ℃ of sterilizing 20min, after cooling, above-mentioned seed liquor is accessed in fermention medium, keep leavening temperature 25-32 ℃, dissolved oxygen 20-60%, ferments;
(4) sophorolipid processed: after fermentation ends, use with the isopyknic organic solvent ethyl acetate of fermented liquid and extract sophorolipid, organic solvent is removed in underpressure distillation, obtains sophorolipid.
Slant medium in above-mentioned steps (1) forms (g/L): 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 in above-mentioned steps (2) forms (g/L): glucose 60-100, yeast powder 5-8, urea 0.5-1.
Fermention medium in above-mentioned steps (3) forms (g/L): sewer oil 60-120, waste molasses 60-180, yeast powder 3-5.
Seed liquor is accessed in fermention medium according to the inoculum size of volume ratio 3-5% 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)
sewer oil has adopted two kinds: the one, and refining sewer oil (buying on the market), the 2nd, in the trench from dining room, reclaim, only pass through simple centrifugal treating, get upper strata oil reservoir and make raw material.
Beneficial effect of the present invention:
1, the present invention adopts the Technology that waste molasses and the two carbon source through fermentation of sewer oil are produced sophorolipid, realizes the recycling of waste, reduces environmental pollution improvement's expense, greatly reduces the production cost of sophorolipid.
2, solved sewer oil exhaust emission and greatly, again entered edible field and endanger large problem, opened " bottleneck " that sewer oil is recycled, effective guarantee people healthy, realized the rational utilization of sewer oil resource.
3, this technique does not need complicated equipment, and fermenting process is simple, is conducive to exploitation, application and the popularization of sophorolipid.
4, the method can also be fermented and be carried out the production of sophorolipid by continuous feeding, and actual application value is higher, and market outlook are wide.
Embodiment
Embodiment 1
(1) by torulopsis (
torulopsis bombicola) ATCC 22214 is inoculated on YPD slant medium, 30 ℃ of constant temperature culture 2 days, with the physiological saline wash-out thalline that is 0.85%, make cell content and are about 5.3 * 10
7the bacteria suspension of individual/mL;
(2) preparation of seed liquor: seed culture medium forms (g/L): glucose 60, yeast powder 5, urea 0.5, pH 6.0; This seed culture medium of 100mL is placed in to 500mL shaking flask, and 121 ℃ of sterilizing 20min, access above-mentioned bacteria suspension according to the inoculum size of volume percent 3% after cooling, and shaking flask is placed in to 25 ℃, on the shaking table of rotating speed 120rpm, cultivate 24h, obtain seed liquor;
(3) above-mentioned seed liquor take for inoculating aseptic 5L fermentor tank the fermention medium that waste molasses and sewer oil be substrate.Fermention medium forms (g/L): sewer oil 60, waste molasses 60, yeast powder 3, pH=6.0, tap water is settled to 3L, 121 ℃ of sterilizing 20min, after cooling, according to the inoculum size of volume ratio 3%, above-mentioned seed liquor is accessed in fermention medium, 30 ℃ of leavening temperatures, dissolved oxygen is 20%, fermentation time 72h;
(4) after fermentation ends, use with the isopyknic ethyl acetate of fermented liquid and extract sophorolipid, organic solvent is removed in underpressure distillation, and obtaining sophorolipid output is 33.5g/L.
Embodiment 2
(1) by torulopsis (
torulopsis bombicola) ATCC 22214 is inoculated on YPD slant medium, 30 ℃ of constant temperature culture 2 days, with the physiological saline wash-out thalline that is 0.85%, make cell content and are about 6.2 * 10
7the bacteria suspension of individual/mL;
(2) preparation of seed liquor: seed culture medium forms (g/L): glucose 80, yeast powder 6, urea 0.8, pH 6.0; This seed culture medium of 100mL is placed in to 500mL shaking flask, and 121 ℃ of sterilizing 20min, access above-mentioned bacteria suspension according to the inoculum size of volume percent 4% after cooling, and shaking flask is placed in to 28 ℃, on the shaking table of rotating speed 150rpm, cultivate 24h, obtain seed liquor;
(3) above-mentioned seed liquor take for inoculating aseptic 5L fermentor tank the fermention medium that waste molasses and sewer oil be substrate.Fermention medium forms (g/L): sewer oil 80, waste molasses 100, yeast powder 4, pH=6.0, tap water is settled to 3L, 121 ℃ of sterilizing 20min, after cooling, according to the inoculum size of volume ratio 4%, above-mentioned seed liquor is accessed in fermention medium, 28 ℃ of leavening temperatures, dissolved oxygen is 35%, fermentation time 84h;
(4) after fermentation ends, use with the isopyknic ethyl acetate of fermented liquid and extract sophorolipid, organic solvent is removed in underpressure distillation, and obtaining sophorolipid output is 59.0g/L.
Embodiment 3
(1) by torulopsis (
torulopsis bombicola) ATCC 22214 is inoculated on YPD slant medium, 30 ℃ of constant temperature culture 2 days, with the physiological saline wash-out thalline that is 0.85%, make cell content and are about 7.4 * 10
7the bacteria suspension of individual/mL;
(2) preparation of seed liquor: seed culture medium forms (g/L): glucose 100, yeast powder 7, urea 1, pH 6.0; This seed culture medium of 100mL is placed in to 500mL shaking flask, and 121 ℃ of sterilizing 20min, access above-mentioned bacteria suspension according to the inoculum size of volume percent 5% after cooling, and shaking flask is placed in to 30 ℃, on the shaking table of rotating speed 180rpm, cultivate 24h, obtain seed liquor;
(3) above-mentioned seed liquor take for inoculating aseptic 5L fermentor tank the fermention medium that waste molasses and sewer oil be substrate.Fermention medium forms (g/L): sewer oil 100, waste molasses 160, yeast powder 4, pH=6.0, tap water is settled to 3L, 121 ℃ of sterilizing 20min, after cooling, according to the inoculum size of volume ratio 4%, above-mentioned seed liquor is accessed in fermention medium, 30 ℃ of leavening temperatures, dissolved oxygen is 45%, fermentation time 96h;
(4) after fermentation ends, use with the isopyknic ethyl acetate of fermented liquid and extract sophorolipid, organic solvent is removed in underpressure distillation, and obtaining sophorolipid output is 83.4g/L.
Embodiment 4
(1) by torulopsis (
torulopsis bombicola) ATCC 22214 is inoculated on YPD slant medium, 30 ℃ of constant temperature culture 2 days, with the physiological saline wash-out thalline that is 0.85%, make cell content and are about 8.5 * 10
7the bacteria suspension of individual/mL;
(2) preparation of seed liquor: seed culture medium forms (g/L): glucose 80, yeast powder 8, urea 0.8, pH 6.0; This seed culture medium of 100mL is placed in to 500mL shaking flask, and 121 ℃ of sterilizing 20min, access above-mentioned bacteria suspension according to the inoculum size of volume percent 5% after cooling, and shaking flask is placed in to 32 ℃, on the shaking table of rotating speed 180rpm, cultivate 24h, obtain seed liquor;
(3) above-mentioned seed liquor take for inoculating aseptic 5L fermentor tank the fermention medium that waste molasses and sewer oil be substrate.Fermention medium forms (g/L): sewer oil 120, waste molasses 180, yeast powder 5, pH=6.0, tap water is settled to 3L, 121 ℃ of sterilizing 20min, after cooling, according to the inoculum size of volume ratio 5%, above-mentioned seed liquor is accessed in fermention medium, 32 ℃ of leavening temperatures, dissolved oxygen is 60%, fermentation time 120h;
(4) after fermentation ends, use with the isopyknic ethyl acetate of fermented liquid and extract sophorolipid, organic solvent is removed in underpressure distillation, and obtaining sophorolipid output is 99.3g/L.
Claims (8)
1. a method of producing sophorolipid with waste molasses and the two carbon source through fermentation of sewer oil, is characterized in that, described method comprises the steps:
(1) preparation of bacteria suspension: by torulopsis bacterium (
torulopsis bombicola) ATCC 22214 is inoculated on the YPD slant medium being comprised of glucose, peptone, yeast powder and agar, with distilled water, is settled to 1L, pH=6.0;
In 30 ℃ of constant temperature culture 2-3 days, the physiological saline wash-out thalline that is 0.85% by appropriate mass concentration, makes bacteria suspension;
(2) seed liquor processed: the shaking flask that the seed culture medium 100ml being comprised of glucose, yeast powder and urea is placed in to 500ml, 121 ℃ of sterilizing 20min, after cooling, according to the inoculum size of volume percent 3-5%, access above-mentioned bacteria suspension, then shaking flask is placed on 25-32 ℃, the shaking table of 120-180rpm and cultivates 24h, obtain seed liquor;
(3) fermentation: above-mentioned seed liquor be take to the fermention medium that waste molasses and sewer oil be substrate for inoculating aseptic 5L fermentor tank, with tap water, be settled to 3L, pH=6.0, by fermentation culture based on 121 ℃ of sterilizing 20min, after cooling, above-mentioned seed liquor is accessed in fermention medium, keep leavening temperature 25-32 ℃, dissolved oxygen 20-60%, ferments;
(4) sophorolipid processed: after fermentation ends, use with the isopyknic organic solvent ethyl acetate of fermented liquid and extract sophorolipid, organic solvent is removed in underpressure distillation, obtains sophorolipid.
2. the method that a kind of use waste molasses according to claim 1 and the two carbon source through fermentation of sewer oil are produced sophorolipid, it is characterized in that, the slant medium in described step (1) forms (g/L): glucose 20, peptone 20, yeast powder 10, agar 15-20.
3. a kind of method of producing sophorolipid with waste molasses and the two carbon source through fermentation of sewer oil according to claim 1, is characterized in that, in the bacteria suspension in described step (1), mycetocyte content is about 10
7-10
8individual/mL.
4. a kind of method of producing sophorolipid with waste molasses and the two carbon source through fermentation of sewer oil according to claim 1, is characterized in that, the seed culture medium in described step (2) forms (g/L): glucose 60-100, yeast powder 5-8, urea 0.5-1.
5. a kind of method of producing sophorolipid with waste molasses and the two carbon source through fermentation of sewer oil according to claim 1, is characterized in that, the fermention medium in described step (3) forms (g/L): sewer oil 60-120, waste molasses 60-180, yeast powder 3-5.
6. a kind of method of producing sophorolipid with waste molasses and the two carbon source through fermentation of sewer oil according to claim 1, is characterized in that seed liquor is accessed in fermention medium according to the inoculum size of volume ratio 3-5% in described step (3).
7. a kind of method of producing sophorolipid with waste molasses and the two carbon source through fermentation of sewer oil according to claim 1, is characterized in that, the fermentation time in described step (3) is 72-120h.
8. a kind of method of producing sophorolipid with waste molasses and the two carbon source through fermentation of sewer oil according to claim 1, is characterized in that, in described step (4), the output of sophorolipid is 30-100g/L.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310590289.2A CN103589759B (en) | 2013-11-22 | 2013-11-22 | A kind of waste molasses and the two carbon source through fermentation of sewer oil produce the method for sophorolipid |
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN105154050A (en) * | 2015-07-29 | 2015-12-16 | 中国石油化工股份有限公司 | Temperature-resistant salt-tolerant viscous-oil biological viscosity reducer and preparation method thereof |
| CN105671106A (en) * | 2016-02-16 | 2016-06-15 | 珀莱雅化妆品股份有限公司 | Method for preparing glycosphingolipid through olive oil fermentation method |
| CN113151020A (en) * | 2021-05-12 | 2021-07-23 | 南京工业大学 | Feeding fermentation method for high-yield acid sophorolipid |
| CN114807271A (en) * | 2022-05-06 | 2022-07-29 | 河南省科学院高新技术研究中心 | Method for preparing sophorolipid by fermentation method and application of sophorolipid in medicament for reverse osmosis membrane |
| CN115992184A (en) * | 2023-03-23 | 2023-04-21 | 科乐美(广州)生物科技有限公司 | Biological fermentation surfactant, cosmetic and preparation method thereof |
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| 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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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105154050A (en) * | 2015-07-29 | 2015-12-16 | 中国石油化工股份有限公司 | Temperature-resistant salt-tolerant viscous-oil biological viscosity reducer and preparation method thereof |
| 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 |
| CN105671106A (en) * | 2016-02-16 | 2016-06-15 | 珀莱雅化妆品股份有限公司 | Method for preparing glycosphingolipid through olive oil fermentation method |
| 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 |
| CN113151020A (en) * | 2021-05-12 | 2021-07-23 | 南京工业大学 | Feeding fermentation method for high-yield acid sophorolipid |
| CN113151020B (en) * | 2021-05-12 | 2023-10-20 | 南京工业大学 | High-acid-yield sophorolipid feed-supplementing fermentation method |
| CN114807271A (en) * | 2022-05-06 | 2022-07-29 | 河南省科学院高新技术研究中心 | Method for preparing sophorolipid by fermentation method and application of sophorolipid in medicament for reverse osmosis membrane |
| CN115992184A (en) * | 2023-03-23 | 2023-04-21 | 科乐美(广州)生物科技有限公司 | Biological fermentation surfactant, cosmetic and preparation method thereof |
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