CN1393563A - Process for preparing itaconic acid by fermentation and bacterial strain for it - Google Patents
Process for preparing itaconic acid by fermentation and bacterial strain for it Download PDFInfo
- Publication number
- CN1393563A CN1393563A CN 01113240 CN01113240A CN1393563A CN 1393563 A CN1393563 A CN 1393563A CN 01113240 CN01113240 CN 01113240 CN 01113240 A CN01113240 A CN 01113240A CN 1393563 A CN1393563 A CN 1393563A
- Authority
- CN
- China
- Prior art keywords
- fermentation
- acid
- substratum
- itaconic acid
- sngb9003
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
- 238000000855 fermentation Methods 0.000 title claims abstract description 41
- 230000004151 fermentation Effects 0.000 title claims abstract description 41
- 238000004519 manufacturing process Methods 0.000 title claims abstract description 17
- LVHBHZANLOWSRM-UHFFFAOYSA-N methylenebutanedioic acid Natural products OC(=O)CC(=C)C(O)=O LVHBHZANLOWSRM-UHFFFAOYSA-N 0.000 title claims description 29
- JAHNSTQSQJOJLO-UHFFFAOYSA-N 2-(3-fluorophenyl)-1h-imidazole Chemical compound FC1=CC=CC(C=2NC=CN=2)=C1 JAHNSTQSQJOJLO-UHFFFAOYSA-N 0.000 title claims description 13
- 230000001580 bacterial effect Effects 0.000 title description 18
- 241001465318 Aspergillus terreus Species 0.000 claims abstract description 3
- 238000000034 method Methods 0.000 claims description 36
- 229920002472 Starch Polymers 0.000 claims description 16
- 241000894006 Bacteria Species 0.000 claims description 11
- 235000019890 Amylum Nutrition 0.000 claims description 9
- 239000000413 hydrolysate Substances 0.000 claims description 9
- 244000005700 microbiome Species 0.000 claims description 7
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 claims description 6
- 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 description 5
- 239000008103 glucose Substances 0.000 claims description 5
- 239000007788 liquid Substances 0.000 claims description 5
- 238000002425 crystallisation Methods 0.000 claims description 4
- 230000008025 crystallization Effects 0.000 claims description 4
- 230000000813 microbial effect Effects 0.000 claims description 4
- 238000001914 filtration Methods 0.000 claims description 3
- 235000011187 glycerol Nutrition 0.000 claims description 3
- 238000005342 ion exchange Methods 0.000 claims description 3
- OWEGMIWEEQEYGQ-UHFFFAOYSA-N 100676-05-9 Natural products OC1C(O)C(O)C(CO)OC1OCC1C(O)C(O)C(O)C(OC2C(OC(O)C(O)C2O)CO)O1 OWEGMIWEEQEYGQ-UHFFFAOYSA-N 0.000 claims description 2
- 229920001353 Dextrin Polymers 0.000 claims description 2
- 239000004375 Dextrin Substances 0.000 claims description 2
- GUBGYTABKSRVRQ-PICCSMPSSA-N Maltose Natural products O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@@H](CO)OC(O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-PICCSMPSSA-N 0.000 claims description 2
- 229930006000 Sucrose Natural products 0.000 claims description 2
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 claims description 2
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 claims description 2
- 235000019425 dextrin Nutrition 0.000 claims description 2
- 235000001727 glucose Nutrition 0.000 claims description 2
- 239000005720 sucrose Substances 0.000 claims description 2
- 239000002253 acid Substances 0.000 abstract description 20
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 abstract description 4
- 229910052799 carbon Inorganic materials 0.000 abstract description 4
- 241000228212 Aspergillus Species 0.000 abstract 1
- 239000001963 growth medium Substances 0.000 abstract 1
- 230000009466 transformation Effects 0.000 description 10
- 239000008107 starch Substances 0.000 description 7
- 235000019698 starch Nutrition 0.000 description 7
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 description 6
- VWDWKYIASSYTQR-UHFFFAOYSA-N sodium nitrate Chemical compound [Na+].[O-][N+]([O-])=O VWDWKYIASSYTQR-UHFFFAOYSA-N 0.000 description 6
- KWGKDLIKAYFUFQ-UHFFFAOYSA-M lithium chloride Chemical compound [Li+].[Cl-] KWGKDLIKAYFUFQ-UHFFFAOYSA-M 0.000 description 4
- 239000000047 product Substances 0.000 description 4
- PAWQVTBBRAZDMG-UHFFFAOYSA-N 2-(3-bromo-2-fluorophenyl)acetic acid Chemical compound OC(=O)CC1=CC=CC(Br)=C1F PAWQVTBBRAZDMG-UHFFFAOYSA-N 0.000 description 3
- WTLKTXIHIHFSGU-UHFFFAOYSA-N 2-nitrosoguanidine Chemical compound NC(N)=NN=O WTLKTXIHIHFSGU-UHFFFAOYSA-N 0.000 description 3
- 229920001817 Agar Polymers 0.000 description 3
- 108090000790 Enzymes Proteins 0.000 description 3
- 102000004190 Enzymes Human genes 0.000 description 3
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- 240000008042 Zea mays Species 0.000 description 3
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 description 3
- 235000002017 Zea mays subsp mays Nutrition 0.000 description 3
- 239000008272 agar Substances 0.000 description 3
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 description 3
- 235000005822 corn Nutrition 0.000 description 3
- 238000011161 development Methods 0.000 description 3
- 230000018109 developmental process Effects 0.000 description 3
- 229940088598 enzyme Drugs 0.000 description 3
- 239000002504 physiological saline solution Substances 0.000 description 3
- 239000004317 sodium nitrate Substances 0.000 description 3
- 229940001516 sodium nitrate Drugs 0.000 description 3
- 235000010344 sodium nitrate Nutrition 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- XEEYBQQBJWHFJM-UHFFFAOYSA-N Iron Chemical compound [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 description 2
- XNPAKGMQCVYQAO-UHFFFAOYSA-N [F].CC(O)=O Chemical compound [F].CC(O)=O XNPAKGMQCVYQAO-UHFFFAOYSA-N 0.000 description 2
- 238000011534 incubation Methods 0.000 description 2
- 230000006698 induction Effects 0.000 description 2
- 238000011081 inoculation Methods 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 230000035772 mutation Effects 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 208000012788 shakes Diseases 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 description 1
- 240000005979 Hordeum vulgare Species 0.000 description 1
- 235000007340 Hordeum vulgare Nutrition 0.000 description 1
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 1
- DGAQECJNVWCQMB-PUAWFVPOSA-M Ilexoside XXIX Chemical compound C[C@@H]1CC[C@@]2(CC[C@@]3(C(=CC[C@H]4[C@]3(CC[C@@H]5[C@@]4(CC[C@@H](C5(C)C)OS(=O)(=O)[O-])C)C)[C@@H]2[C@]1(C)O)C)C(=O)O[C@H]6[C@@H]([C@H]([C@@H]([C@H](O6)CO)O)O)O.[Na+] DGAQECJNVWCQMB-PUAWFVPOSA-M 0.000 description 1
- 229910002651 NO3 Inorganic materials 0.000 description 1
- NHNBFGGVMKEFGY-UHFFFAOYSA-N Nitrate Chemical compound [O-][N+]([O-])=O NHNBFGGVMKEFGY-UHFFFAOYSA-N 0.000 description 1
- 239000001888 Peptone Substances 0.000 description 1
- 108010080698 Peptones Proteins 0.000 description 1
- ZLMJMSJWJFRBEC-UHFFFAOYSA-N Potassium Chemical compound [K] ZLMJMSJWJFRBEC-UHFFFAOYSA-N 0.000 description 1
- WCUXLLCKKVVCTQ-UHFFFAOYSA-M Potassium chloride Chemical compound [Cl-].[K+] WCUXLLCKKVVCTQ-UHFFFAOYSA-M 0.000 description 1
- 240000006394 Sorghum bicolor Species 0.000 description 1
- 235000011684 Sorghum saccharatum Nutrition 0.000 description 1
- 235000021307 Triticum Nutrition 0.000 description 1
- 244000098338 Triticum aestivum Species 0.000 description 1
- LEHOTFFKMJEONL-UHFFFAOYSA-N Uric Acid Chemical compound N1C(=O)NC(=O)C2=C1NC(=O)N2 LEHOTFFKMJEONL-UHFFFAOYSA-N 0.000 description 1
- TVWHNULVHGKJHS-UHFFFAOYSA-N Uric acid Natural products N1C(=O)NC(=O)C2NC(=O)NC21 TVWHNULVHGKJHS-UHFFFAOYSA-N 0.000 description 1
- HCHKCACWOHOZIP-UHFFFAOYSA-N Zinc Chemical compound [Zn] HCHKCACWOHOZIP-UHFFFAOYSA-N 0.000 description 1
- 150000003863 ammonium salts Chemical class 0.000 description 1
- 239000010775 animal oil Substances 0.000 description 1
- 238000005422 blasting Methods 0.000 description 1
- 239000011575 calcium Substances 0.000 description 1
- 229910052791 calcium Inorganic materials 0.000 description 1
- 239000002775 capsule Substances 0.000 description 1
- 239000010941 cobalt Substances 0.000 description 1
- 229910017052 cobalt Inorganic materials 0.000 description 1
- GUTLYIVDDKVIGB-UHFFFAOYSA-N cobalt atom Chemical compound [Co] GUTLYIVDDKVIGB-UHFFFAOYSA-N 0.000 description 1
- 238000012258 culturing Methods 0.000 description 1
- 239000013530 defoamer Substances 0.000 description 1
- 229940111205 diastase Drugs 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 230000008030 elimination Effects 0.000 description 1
- 238000003379 elimination reaction Methods 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 238000012262 fermentative production Methods 0.000 description 1
- 239000011790 ferrous sulphate Substances 0.000 description 1
- 235000003891 ferrous sulphate Nutrition 0.000 description 1
- 239000000706 filtrate Substances 0.000 description 1
- 230000009970 fire resistant effect Effects 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 230000035784 germination Effects 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 239000002054 inoculum Substances 0.000 description 1
- 229910017053 inorganic salt Inorganic materials 0.000 description 1
- 229910052742 iron Inorganic materials 0.000 description 1
- BAUYGSIQEAFULO-UHFFFAOYSA-L iron(2+) sulfate (anhydrous) Chemical compound [Fe+2].[O-]S([O-])(=O)=O BAUYGSIQEAFULO-UHFFFAOYSA-L 0.000 description 1
- 229910000359 iron(II) sulfate Inorganic materials 0.000 description 1
- 239000002932 luster Substances 0.000 description 1
- WPBNNNQJVZRUHP-UHFFFAOYSA-L manganese(2+);methyl n-[[2-(methoxycarbonylcarbamothioylamino)phenyl]carbamothioyl]carbamate;n-[2-(sulfidocarbothioylamino)ethyl]carbamodithioate Chemical compound [Mn+2].[S-]C(=S)NCCNC([S-])=S.COC(=O)NC(=S)NC1=CC=CC=C1NC(=S)NC(=O)OC WPBNNNQJVZRUHP-UHFFFAOYSA-L 0.000 description 1
- 230000035800 maturation Effects 0.000 description 1
- 235000013372 meat Nutrition 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 239000002480 mineral oil Substances 0.000 description 1
- 235000010446 mineral oil Nutrition 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 238000002703 mutagenesis Methods 0.000 description 1
- 231100000350 mutagenesis Toxicity 0.000 description 1
- -1 nitrogenous compound Chemical class 0.000 description 1
- 239000003921 oil Substances 0.000 description 1
- 150000007524 organic acids Chemical class 0.000 description 1
- 235000019319 peptone Nutrition 0.000 description 1
- 150000003016 phosphoric acids Chemical class 0.000 description 1
- 239000000049 pigment Substances 0.000 description 1
- 239000011591 potassium Substances 0.000 description 1
- 229910052700 potassium Inorganic materials 0.000 description 1
- 229920001592 potato starch Polymers 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 230000005855 radiation Effects 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 238000007670 refining Methods 0.000 description 1
- 239000011347 resin Substances 0.000 description 1
- 229920005989 resin Polymers 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 238000004513 sizing Methods 0.000 description 1
- 239000011734 sodium Substances 0.000 description 1
- 229910052708 sodium Inorganic materials 0.000 description 1
- 238000009923 sugaring Methods 0.000 description 1
- 229940116269 uric acid Drugs 0.000 description 1
- 235000013311 vegetables Nutrition 0.000 description 1
- 238000000207 volumetry Methods 0.000 description 1
- 230000037303 wrinkles Effects 0.000 description 1
- 239000011701 zinc Substances 0.000 description 1
- 229910052725 zinc Inorganic materials 0.000 description 1
Landscapes
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
A process for preparing itaconis acid features the microbe fermentation in the culture medium containing carbon source. The mentioned microbe is Aspergillus terrous SNGB9003 CGMCC No.0569. Its advantage is very high output rate. This invention also provides the high-output Aspergillus terreus SNGB9003 CGMCC No.0569 by itaeonic acid fermentation.
Description
Technical field
The present invention relates to the biological fermentation field, relate in particular to the field of fermentation production of organic acid.More particularly, the present invention relates to a kind of method of fermentation production of itaconic acid, and the used bacterial strain of this method.
Background of invention
China begins one's study from the eighties and uses obtained by producing itaconic acid by fermentation.In recent years, the development of this respect is very rapid, and state of the art improves constantly, and especially at the cultivation of bacterial classification, the aspects such as raising of producing sour Study on Conditions, acid production rate continuous progress has been arranged.From early shake flask test with the amylum hydrolysate of the sugar be raw material produce acid 3.5%, to sugared transformation efficiency 35% (referring to " development and application of methylene-succinic acid and derived product thereof ", Zhao Hongsheng, " Ningbo chemical industry ", in February, 1998) develop into and at present newer shake bottle acid production rate and can reach 7%, transformation efficiency is that 50-55% is (referring to " cultivation of methylene-succinic acid superior strain ", people such as Wu Jing, " medicine biotechnology " in March, 1997).The dissociant 15-UV-07 of the graduate terreus NRRL1960 of Sichuan food fermentation produces acid 7.5% in 30 liters of fermentor tanks, to sugared transformation efficiency is (referring to " development and present situation that methylene-succinic acid is produced " more than 50%, the first-class people of Zhu Guang, " chemical engineer ", in January, 1992).
U.S. Pat 5457040 has reported by increasing glycerine and has come fermentation production of itaconic acid as carbon source that it produces acid and is up to 6.97%.The method that U.S. Pat 3873425 has reported that a kind of high acid reaches 7%, sugared transformation efficiency is 55% fermentation production of itaconic acid.Another piece U.S. Pat 5231016 has reported that a kind of microbial fermentation with high yield produces the method for methylene-succinic acid, and its cycle is 80 hours, and acid production rate is 7.41%, and output capacity is 0.926 grams per liter hour.
Summary of the invention
Problem to be solved by this invention is further to improve the fermentative production level of methylene-succinic acid.An object of the present invention is to provide a kind of method of improved fermentation production of itaconic acid.This method is fit to industrial large scale fermentation jar (100 cubic metres), produces acid and can reach 8%, and transformation efficiency is 60%, and fermentation period is about 49 hours, and output capacity reaches 1.6 grams per liters hour.Another object of the present invention provides the new terreus of the method that is used for above-mentioned fermentation production of itaconic acid and produces bacterium.
For achieving the above object, one aspect of the present invention relates to a kind of method of fermentation production of itaconic acid, this method is to produce methylene-succinic acid by microbial fermentation in substratum, it is characterized in that, described microorganism is terreus (Aspergillusterreus) SNGB9003 CGMCC No.0569.
The present invention relates to a kind of itaconic acid fermentation on the other hand and produces bacterium, it is characterized in that this bacterium is terreus SNGB9003 CGMCC No.0569.
In fermentation process of the present invention, described substratum can contain and is selected from maltose, dextrin, glycerine, glucose, sucrose and the amylum hydrolysate of the sugar one or more as carbon source.Preferably, in substratum, can adopt amylum hydrolysate of the sugar.This amylum hydrolysate of the sugar can make by sugaring program well known by persons skilled in the art from starch (such as barley starch, W-Gum, sorghum starch, oat starch, potato starch etc.).Concentration of reduced sugar in the substratum can be between 8-15% (w/v), and is preferable between 12-13%.This concentration of reduced sugar can be measured with conventional means well known by persons skilled in the art (as the fixed sugared method of the film).
In the fermention medium of the present invention except containing the required above-mentioned carbon source of fermentation production of citric acid, also can need to add other required material of microorganism culturing, they are including, but not limited to peptone, soyflour, meat extract, wheat skin, uric acid, corn steep liquor, ammonium salt, nitrate and other organic or inorganic nitrogenous compound.Also can contain inorganic salt in the fermention medium, as metal-salts such as vitriol, hydrochloride, phosphoric acid salt and sodium, potassium, calcium, zinc, manganese, iron.Also can add animal oil, vegetables oil or mineral oil in case of necessity as defoamer.
Selection for conditions such as leavening temperature, time, pH is conventional, and those skilled in the art can be optimized according to fermentation indexs such as producing acid, transformation efficiency.
In the method for the invention, leavening temperature can be between 33 to 40 ℃, and preferable is 35 to 38 ℃, and best is 37 ℃.Fermentation just pH is the natural pH of substratum.At fermenting process, need not pH is controlled, but fermentation usually can be between 2-7.In preferable embodiment of the present invention, initial pH value is about 5, and the pH value after the fermentation ends is about 2.
Microorganism of the present invention can add in the fermention medium by direct inoculation or middle jar of usual manner such as cultivation.
Used terreus SNGB9003 is that disclosed NRRL1960 from US 5457040 (available from american agriculture mechanism preservation center) is by ultraviolet rays, Co in fermentation process of the present invention
60The mutagenesis and screening obtains on lithium chloride or single fluorine acetic acid resistant panel repeatedly of conventional mutafacient system such as-γ, NTG.This bacterial strain is preserved in China Committee for Culture Collection of Microorganisms common micro-organisms center (China, Beijing) April 28 calendar year 2001, and preserving number is CGMCC No.0569.About the form of this bacterial strain and character and obtain to have in an embodiment detailed description.
After fermentation ends, can adopt this area ordinary method that fermented liquid is carried out purifying, these purification process are including, but not limited to, filtration, crystallization, centrifugal, dry, ion-exchange and decolouring etc.
By utilizing above-mentioned specific microorganism, every index of fermentation process of the present invention all has an obvious improvement (produce acid and can reach 8%, transformation efficiency is 60%, and fermentation period is 47-52 hour) than what report in the prior art.And the inventive method can be applicable to industrial large scale fermentation (100 cubic metres), has very high output capacity.
Embodiment
For the ease of understanding the present invention, the present invention is described in further detail below in conjunction with embodiment.Yet these embodiment have been the effect of explanation, and the present invention is not limited to these embodiment.Among the embodiment, unless refer in particular in addition, all percentage number averages refer to the contained gram number of per 100 milliliters of substratum hereinafter.It should be noted that the concentration of amylum hydrolysate of the sugar in substratum reducing sugar value representation.
Embodiment:
A) substratum
Substratum I: SODIUMNITRATE 0.3%; Dipotassium hydrogen phosphate 0.1%; Repone K 0.05%; Sal epsom 0.5%; Ferrous sulfate 0.01%; Glucose 2%; Agar 2%.The pH nature was sterilized 30 minutes for 121 ℃.
Medium ii: glucose 10%; Single fluorine acetic acid 8%; SODIUMNITRATE 0.3%; Dipotassium hydrogen phosphate 0.1%; Sal epsom 0.5%; Agar 2%.The pH nature was sterilized 30 minutes for 121 ℃.
Medium ii I: glucose 10%; Lithium chloride 2%; SODIUMNITRATE 0.3%; Dipotassium hydrogen phosphate 0.1%; Sal epsom 0.5%; Agar 2%.The pH nature was sterilized 30 minutes for 121 ℃.
Substratum IV: methylene-succinic acid 10%; Ammonium nitrate 0.35%; Dipotassium hydrogen phosphate 0.1%; Sal epsom 0.5%.The pH nature was sterilized 15 minutes for 121 ℃.
Substratum V: amylum hydrolysate of the sugar 10%; Corn steep liquor 0.3%; Ammonium nitrate 0.35%; Dipotassium hydrogen phosphate 0.15%; Sal epsom 0.5%.The pH nature was sterilized 15 minutes for 121 ℃.
Substratum VI: amylum hydrolysate of the sugar 13%; Corn steep liquor 0.2%; Ammonium nitrate 0.3%; Dipotassium hydrogen phosphate 0.1%; Sal epsom 0.5%.The pH nature was sterilized 15 minutes for 121 ℃.
B) induction mutation of bacterium method
Method one: bacterial classification in 30 ℃ of cultivations 5 days, is collected spore in physiological saline in substratum I, (Co60-gamma-rays) radiation down in the cobalt source, and dosage is 200,000 roentgens, the time is 1 hour.
Method two: bacterial classification was cultivated 5 days in 30 ℃ in substratum I, collected spore in substratum V 37 ℃, and 250rpm sprouted 4-6 hour, and centrifugal collection spore is in physiological saline, and adding nitrosoguanidine (NTG) dosage is 1000 μ g/ml, handles 40 minutes for 30 ℃.
Method three: bacterial classification in 30 ℃ of cultivations 5 days, is collected spore in physiological saline in substratum I, and under the 40W ultraviolet lamp, 9 centimetres of distances were shaken 10 minutes.
C) induction mutation of bacterium process:
Terreus bacterial classification NRRL1960 is carried out single bacterium separate, the bacterial classification A3 of acid yield (4.3%) in the middle of obtaining handles the back with method two and cultivated in substratum IV 4～6 hours, and elimination germination mycelia is coated on medium ii I.Cultivated 4～5 days in 30 ℃, choose single bacterium, continue to cultivate 4～5 days, treat to meet substratum VI after the spore maturation, cultivated 72 hours, survey acid, obtain producing acid and be 6.9% bacterial classification A23 with the NaOH volumetry in 37 ℃, 200rpm rotary shaker in substratum I.With method same as above, the A23 bacterial classification is handled the back with method one in medium ii, screen, obtain producing acid and be 8.3% bacterial classification A23-12.Again the A23-12 bacterial classification is handled the back simultaneously with method one and method three and in medium ii, screen, obtain acid-producing bacteria kind SNGB9003.Shaking a bottle stage, the product acid of this bacterial strain can reach 10%, and transformation efficiency is 65%.This bacterial strain is preserved in China Committee for Culture Collection of Microorganisms common micro-organisms center April 28 calendar year 2001, and preserving number is CGMCC No.0569.
East China, China microbial preservation center substation is described below the form and the character of this bacterial classification:
| Big class | Project | Form is described |
| Giant colony | Diameter color and luster water colo(u)r back side section smell drop quality | 4.3cm the light yellow look in center, the left and right sides, edge white, the ginger-colored pigment of middle light brown does not have wrinkle, be yellow, dark mound, centre shape, the flourishing nothing of aerial hyphae has aerial hyphae fine hair shape |
| Microscopic morphology | Mycelia conidial head sporophore top capsule stigma spore | Have every big portion spherical, a small amount of column, 49-63 μ 260-290 * 6.5-7.9 μ semisphere 15-25 μ one-level 3.6-7.2 * 18 μ secondary 5.4-7.2 * 1.4 μ garden shapes, smooth, light brown 1.3-3 μ |
D) fermentation production of itaconic acid:
Sugaring: the starch concentration about 20% of sizing mixing, 50～60 ℃ of preheatings, every gram starch adds fire resistant alpha-diastase 12 units, stablely makes 100～105 ℃ of material outlet temperature by the vapo(u)r blasting liquefier, keeps a jar temperature and is controlled between 98～105 ℃.Be cooled to 60 ℃ after the liquefaction fully, pH transfers to 4.5, and every gram starch adds saccharifying enzyme 100 units, saccharification 18～20 hours, 80 ℃ of enzymes that go out when the reducing sugar value reaches at inferior high.It is stand-by that saccharification liquid filters the back.
The production of hybrid seeds: press substratum V batching in 10 cubic metres of standard fermentor tanks, 8.5 cubic metres of liquid amounts were sterilized 15 minutes for 121 ℃, spore inoculating, and concentration is 10
8～10
9Individual/milliliter, culture condition is 37 ℃ of temperature, rotating speed 120～150rpm, 80～120 cubic metres/hour of air quantity, tank pressure 0.05Mpa.Incubation time is 24～36 hours.
Fermentation: press substratum VI batching in 100 cubic metres of standard fermentor tanks, sterilized 15 minutes for 121 ℃, 85 cubic metres of inoculation back liquid amounts, inoculum size 10%.Culture condition is 37 ℃ of temperature, rotating speed 75rpm, 600～800 cubic metres/hour of air quantity, tank pressure 0.05Mpa.Incubation time is 48～55 hours.
E) extraction is refining:
Fermentation stops 80 ℃ of enzymes that go out of back intensification, and through Plate Filtration, filtrate is in 680mmHg, and 60 ℃ of vacuum concentration to 30% dopes of temperature are cooled to 5-10 ℃ with refrigerated water, centrifugal coarse crystallization in crystallizer; Coarse-grain is dissolved to 18-20%, adds 2% gac, 90 ℃ of decolourings; Through the ion-exchange of 732# sun resin; Vacuum concentration to 30% gets finished product with quadrat method crystallization, centrifugal, 65 ℃ of dry sub-sieves.
F) result:
Table 1 has been reported the result of terreus superior strain SNGB9003 of the present invention in 100 cubic metres of fermentor tank industrial experimentations.
100 cubic metres of industrial experimentation results of table 1 terreus SNGB9003
| The test lot number | Produce bacterium | Produce acid (%) | Transformation efficiency (%) | Fermentation period |
| ????1 | ??SNGB9003 | ????7.81 | ????60.8 | ????47 |
| ????2 | ????8.15 | ????61.3 | ????48 | |
| ????3 | ????8.09 | ????62.2 | ????52 | |
| Mean value | ????8.02 | ????61.4 | ????49 |
Test (average extract yield is 85.1%) through continuous three batches are done to extract, the result shows that quality product is qualified.As can be seen, fermentation process of the present invention produces acid and reaches 8.02% from last table 1, and transformation efficiency reaches 61.4%, and the average fermentation cycle is 49 hours, and output capacity is about 1.63 grams per liters hour, and every index all is better than prior art.
Claims (8)
1. the method for a fermentation production of itaconic acid, this method is to produce methylene-succinic acid by microbial fermentation in substratum, it is characterized in that, described microorganism is terreus (Aspergillus terreus) SNGB9003 CGMCC No.0569.
2. method according to claim 1 is characterized in that, described substratum contains and is selected from maltose, dextrin, glycerine, glucose, sucrose and the amylum hydrolysate of the sugar one or more.
3. method according to claim 1 is characterized in that, contains amylum hydrolysate of the sugar in the described substratum.
4. method according to claim 1 is characterized in that, the concentration of reduced sugar in the described substratum is 8-15%.
5. method according to claim 1 is characterized in that described fermentation is carried out between 33-40 ℃.
6. method according to claim 1 is characterized in that, this method also randomly is included in the fermentation back fermented liquid is carried out purifying.
7. method according to claim 6 is characterized in that, described purifying comprises filtration, crystallization, centrifugal, dry, ion-exchange and decolouring.
8. an itaconic acid fermentation is produced bacterium, it is characterized in that this bacterium is terreus SNGB9003 CGMCC No.0569.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 01113240 CN1245518C (en) | 2001-07-04 | 2001-07-04 | Process for preparing itaconic acid by fermentation and bacterial strain for it |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 01113240 CN1245518C (en) | 2001-07-04 | 2001-07-04 | Process for preparing itaconic acid by fermentation and bacterial strain for it |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1393563A true CN1393563A (en) | 2003-01-29 |
| CN1245518C CN1245518C (en) | 2006-03-15 |
Family
ID=4659975
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN 01113240 Expired - Fee Related CN1245518C (en) | 2001-07-04 | 2001-07-04 | Process for preparing itaconic acid by fermentation and bacterial strain for it |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN1245518C (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102517344A (en) * | 2012-01-09 | 2012-06-27 | 山东金洋药业有限公司 | Fermentation production method of itaconic acid by using waste liquid from production of dextran |
| CN104419650A (en) * | 2013-09-02 | 2015-03-18 | 中国科学院青岛生物能源与过程研究所 | Transformed aspergillus terreus engineering bacteria and application thereof |
| CN111733085A (en) * | 2020-07-30 | 2020-10-02 | 江西省科学院微生物研究所 | Aspergillus terreus variant KY-AT-002 capable of producing itaconic acid AT high yield and application thereof |
-
2001
- 2001-07-04 CN CN 01113240 patent/CN1245518C/en not_active Expired - Fee Related
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102517344A (en) * | 2012-01-09 | 2012-06-27 | 山东金洋药业有限公司 | Fermentation production method of itaconic acid by using waste liquid from production of dextran |
| CN104419650A (en) * | 2013-09-02 | 2015-03-18 | 中国科学院青岛生物能源与过程研究所 | Transformed aspergillus terreus engineering bacteria and application thereof |
| CN111733085A (en) * | 2020-07-30 | 2020-10-02 | 江西省科学院微生物研究所 | Aspergillus terreus variant KY-AT-002 capable of producing itaconic acid AT high yield and application thereof |
Also Published As
| Publication number | Publication date |
|---|---|
| CN1245518C (en) | 2006-03-15 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN101899410B (en) | Streptomyces parvus and application thereof in preparation of daptomycin | |
| CN115637278B (en) | Application of Aureobasidium pullulan in fermentation production of pullulan | |
| CN101914478B (en) | Bacillus subtilis and application thereof | |
| CN102965416A (en) | Method for producing cordycepin through semi-continuous liquid fermentation of cordyceps militaris | |
| CN102533889B (en) | Method for continuously fermenting lysine | |
| CN110564580B (en) | Method for producing vinegar containing pyrroloquinoline quinone through microbial co-culture fermentation | |
| CN1136393A (en) | Agricultural chemical for preventing and controlling plant virus prepared by liquid fermentation and its producing technology | |
| CN1291237A (en) | Strain of Penicillium oxalicum var. armenia and its application | |
| CN1245518C (en) | Process for preparing itaconic acid by fermentation and bacterial strain for it | |
| CN114276941A (en) | Method for producing erythritol by fermentation | |
| CN120158404A (en) | A Priesterol bacterium and a method for producing PHA by semi-solid fermentation using the bacterium | |
| CN117210368B (en) | Slender lysine bacillus and application thereof in preparing seaweed liquid fertilizer by kelp degradation | |
| CN116218690B (en) | Curvularia sturtii producing brefeldin A and fermentation method thereof | |
| CN1053697C (en) | Fermentation method for producing D-ribose novel strain, and method for prepn. of D-ribose using said strain | |
| CN118374377A (en) | Marine bacterium for degrading south Africa bark algae and application thereof | |
| CN117683672A (en) | Streptomyces albus and application thereof | |
| CN108676743A (en) | A kind of preparation method of efficient Tiny ecosystem mixed fertilizer | |
| CN114395520A (en) | Method for efficiently producing conidia in large scale by solid state fermentation of trichoderma harzianum | |
| US20060048444A1 (en) | Process for producing matsutake mushroom mycelium | |
| CN107641602A (en) | One plant of candida utili and its fermentation lay eggs it is white in application | |
| CN1683520A (en) | A compound enzyme producing strain and method for fermenting and producing compound enzyme with the strain | |
| CN107118972B (en) | Can liquid fermentation produce pectin Solidago Canadensis endogenetic fungus and its application | |
| CN105154360A (en) | Method for cultivating comonas testosteroni HY-08D | |
| CN112746026A (en) | Candida weissensis and application thereof | |
| CN104805028A (en) | Micro-ecological fertilizer |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| C17 | Cessation of patent right | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20060315 |