CN103695519A - Nitrogen source, culture medium and culture method for producing riboflavin - Google Patents
Nitrogen source, culture medium and culture method for producing riboflavin Download PDFInfo
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- CN103695519A CN103695519A CN201310712200.5A CN201310712200A CN103695519A CN 103695519 A CN103695519 A CN 103695519A CN 201310712200 A CN201310712200 A CN 201310712200A CN 103695519 A CN103695519 A CN 103695519A
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Abstract
The invention relates to a method for obtaining a nitrogen source for producing riboflavin. The method comprises the following steps: performing desalination treatment on wastewater of a riboflavin fermentation broth after extraction, wherein the total salt content of the wastewater is controlled to be less than 5g/l after desalination; adding 2-5% of polymerized ferrous sulfate for flocculating for 1-2 hours, centrifuging for filtering by using a double-conical decanter centrifuge so as to obtain a solid substance, and drying the solid substance so as to obtain bacterial protein. By adopting the method, the problem that the riboflavin bacterial protein is hard to utilize because of high salt content is solved, the bacterial protein which can be alternatively used as a fermentation nitrogen source is obtained, the application of the expensive organic nitrogen source is greatly reduced, and recycling and utilization of resource are achieved. Besides, on the basis of the bacterial protein, the invention further discloses a corresponding seed culture medium formula and a fermentation culture medium formula, and the process control conditions are optimized starting from factors including temperature, pH values, residual sugar content, dissolved oxygen content, ammonia feeding amount, material supplement mode and the like which can affect the stability of fermentation.
Description
Technical field
The present invention relates to biological process and produce riboflavin.
Background technology
The main carbon source that the microorganism fermentation of riboflavin adopts is vegetables oil, glucose, molasses or rice candy etc.; Organic nitrogen source be take the good biological nitrogen of peptone, gelatine, fish meal, corn steep liquor and proportioning as main, and inorganic salt have NaCl, K2HPO4, MgSO4 etc.Cost of material used rises steadily, and causes cost high in industrial application.Meanwhile, the factory effluent of riboflavin fermentation industry is high concentrated organic wastewater, has the features such as high chroma, high salinity, high COD, high BOD, high thalline content, and control is more difficult.In waste water, contain a large amount of sugar, protein, SS and ammonia nitrogen, these materials are not hazardous and noxious substances, can in growth and breeding, be utilized by some microorganisms and animal, and resource has not only been wasted in its any discharge, and cause serious environmental pollution.Therefore, by the tropina recycling in riboflavin waste water, be, one of outlet effectively solving in the required nitrogenous source of riboflavin fermentation.
Existing riboflavin tropina is that the waste water after extracting carries out concentrate drying by multi-effect evaporation system by Lactochrome fermentation liquor, underflow material after concentrated utilizes spray-drying tower (or roller dryer) to be dried and to obtain, and adds application in a large number for feedstuff industry as solid protein feedstuff.Its shortcoming is that the too high added value that causes of tropina salt is too low, and the costs such as energy consumption are higher than value of the product; While adding application as solid protein feedstuff, need access license requirement, greatly limited riboflavin tropina and applied.
Chinese patent 201010101762.2, discloses a kind of waste cell microbial fermentation that utilizes and has realized the method that nitrogen cycle utilizes, and the method is by waste cell pre-treatment, and then hydrolysis obtains nitrogenous source.Although the method provides a kind of thinking using waste cell as nitrogenous source, it does not utilize the too high tropina of salinity, not in conjunction with the feature of riboflavin fermentation, does not solve tropina salinity too high and cause the problem that its added value is low yet.
Summary of the invention
Technical problem to be solved by this invention is how to utilize riboflavin tropina as nitrogenous source, and substratum and the cultural method of suitable riboflavin fermentation is provided.
Technical scheme of the present invention is:
A kind of nitrogenous source acquisition methods of producing riboflavin, the waste water of Lactochrome fermentation liquor after extracting is through desalting treatment, after desalination, the total salt of waste water is controlled at below 5g/l, add 2%～5% polymerization ferrous sulfate flocculation 1～2 hour, with the centrifuging of bipyramid horizontal screw centrifuge, obtain solid formation, drying obtains tropina.
Further, described desalting treatment adopts nanofiltration or electrodialysis.
A riboflavin seed culture medium, starch glucose 20g/l; Tropina 12g/l; Yeast powder 5g/l; SODIUMNITRATE 5g/l; Ammonium nitrate 5g/l; Potassium primary phosphate 1g/l; Dipotassium hydrogen phosphate 3g/l; Magnesium sulfate heptahydrate 0.5g/l; Calcium chloride 0.1g/l; Iron protochloride 0.05g/l; Zinc Sulphate Heptahydrate 0.03g/l; Manganous chloride tetrahydrate 0.02g/l; PH6.7～6.9.
A riboflavin fermentation substratum, starch glucose 120g/l; Tropina 40g/l; Yeast powder 10g/l; SODIUMNITRATE 15g/l; Ammonium nitrate 5g/l; Potassium primary phosphate 2g/l; Dipotassium hydrogen phosphate 4.5g/l; Magnesium sulfate heptahydrate 1.0g/l; Calcium chloride 0.2g/l; Iron protochloride 0.05g/l; Zinc Sulphate Heptahydrate 0.03g/l; Manganous chloride tetrahydrate 0.02g/l; Trimethyl-glycine 0.012g/l; PH6.7～7.1.
A riboflavin fermentation cultural method, comprising:
1) seed culture medium is packed into 10 m3 seeding tanks, after having sterilized, the bacillus subtilis spore suspension that access makes, the aerobic cultivation 12～16h of 120～140rpm at 37～39 ℃.Normal in seed liquor mycelia division, microscopy, without miscellaneous bacteria, every detection data, index all reach under the prerequisite of processing requirement, and the seeding tank that disappeared, to the connecting tube of fermentor tank, is prepared culture transferring;
2) by fermention medium with pump by connecting tube in magnetizing assembly suction 200m3 fermentor tank, after sterilization, sterilizing, fill into the aseptic Glucose Liquid that sterilization has magnetized, make the initial sugar concentration of fermentation tank culture medium reach 15g/L, with liquefied ammonia, regulate PH to 6.7～7.1, controlling ventilation ratio (VVM) is 1: 0.6; Move into cultured normal seed liquor, fermentation volume is controlled at 120～150m3, starts fermentation, keeps fermentation culture temperature at 37～39 ℃, and the sugar amount in controlled fermentation process, then augments and add 30% concentration sterilizing and magnetized Glucose Liquid; Start mechanical stirring, mixing speed is cumulative to 130rpm by 110rpm, maintains the interior dissolved oxygen of tank between 20%～30%.
Further, it is 0～24 hour that the sugar amount in described fermenting process is controlled, between glucose concn 10～15g/L; 24～32 hours, between glucose concn 5～8g/L; Between 32～40 hours glucose concn 2～5g/L; Put tank and within first two hours, stop sugar, residual sugar amount requires regulation and control at 1～2g/L.W-Gum liquid glucose with 30%～60% carries out restricted flow feeding.
Further, Glucose Liquid is through the magnetic field magnetisation of 0.03～0.08T.
Further, the magneticstrength of described magnetizing assembly is 0.03～0.08T.
Compared with prior art, the invention has the beneficial effects as follows:
The invention solves the too high problem that is difficult to utilization of riboflavin tropina salinity, obtain tropina, substitute as fermentation nitrogen source, greatly reduce the use of expensive organic nitrogen source, realized resource recycling.And take this tropina as basis, and invented corresponding seed culture medium, fermentative medium formula, from affecting the factors such as temperature, pH value, residual sugar amount, dissolved oxygen amount, logical ammonia amount and feed supplement mode of fermentation stability, start with, process control condition is optimized.Utilize magnetic field effect, the physico-chemical property of fermentation materials is changed, become and have the magnetic treatment of biological effect solution, promote thalli growth, realize stable yields synergy.
Embodiment
Below in conjunction with embodiment, the present invention will be further described.
Embodiment
Pack seed culture medium into 10 m3 seeding tanks, after having sterilized, the bacillus subtilis spore suspension that access makes, the aerobic cultivation 12～16h of 120～140rpm at 37～39 ℃.Normal in seed liquor mycelia division, microscopy, without miscellaneous bacteria, every detection data, index all reach under the prerequisite of processing requirement, and the seeding tank that disappeared, to the connecting tube of fermentor tank, is prepared culture transferring.
By fermention medium with pump by connecting tube in magnetizing assembly suction 200m3 fermentor tank, after sterilization, sterilizing, fill into the aseptic Glucose Liquid that sterilization has magnetized, make the initial sugar concentration of fermentation tank culture medium reach 15g/L, with liquefied ammonia, regulate PH to 6.7～7.1, controlling ventilation ratio (VVM) is 1: 0.6.Move into cultured normal seed liquor, fermentation volume is controlled at 120～150m3, start fermentation, keep fermentation culture temperature at 37～39 ℃, after fermentation is carried out 4 hours, measure residual sugar content, make the glucose concn in tank remain on 15g/L left and right, then with the flow velocity of 500kg/hr, at the uniform velocity increase progressively and add 30% concentration sterilizing and magnetized Glucose Liquid.Start mechanical stirring, mixing speed is cumulative to 130rpm by 110rpm, maintains the interior dissolved oxygen of tank between 20%～30%.Sugar amount in fermenting process is controlled: 0～24 hour, between glucose concn 10～15g/L; 24～32 hours, between glucose concn 5～8g/L; Between 32～40 hours glucose concn 2～5g/L.Until mycelia is old and feeble, the glucose consumption filling into is complete, and dissolved oxygen rises to 70%, PH rises at 7.5 o'clock and stops fermentation, and fermented liquid enters abstraction process and extracts riboflavin.In fermenting process, the aseptic Glucose Liquid adding with aseptic magnetized water and stream supplements the water yield of evaporation, makes volume be controlled at 150 m3 left and right, and keeps tank pressure 0.02～0.05MPa.
Claims (8)
1. a nitrogenous source acquisition methods of producing riboflavin, it is characterized in that, the waste water of Lactochrome fermentation liquor after extracting is through desalting treatment, after desalination, the total salt of waste water is controlled at below 5g/l, add 2%～5% polymerization ferrous sulfate flocculation 1～2 hour, with the centrifuging of bipyramid horizontal screw centrifuge, obtain solid formation, drying obtains tropina.
2. a kind of nitrogenous source acquisition methods of producing riboflavin according to claim 1, is characterized in that, described desalting treatment adopts nanofiltration or electrodialysis.
3. a riboflavin seed culture medium, is characterized in that, starch glucose 20g/l; Tropina 12g/l; Yeast powder 5g/l; SODIUMNITRATE 5g/l; Ammonium nitrate 5g/l; Potassium primary phosphate 1g/l; Dipotassium hydrogen phosphate 3g/l; Magnesium sulfate heptahydrate 0.5g/l; Calcium chloride 0.1g/l; Iron protochloride 0.05g/l; Zinc Sulphate Heptahydrate 0.03g/l; Manganous chloride tetrahydrate 0.02g/l; PH6.7～6.9.
4. a riboflavin fermentation substratum, is characterized in that, starch glucose 120g/l; Tropina 40g/l; Yeast powder 10g/l; SODIUMNITRATE 15g/l; Ammonium nitrate 5g/l; Potassium primary phosphate 2g/l; Dipotassium hydrogen phosphate 4.5g/l; Magnesium sulfate heptahydrate 1.0g/l; Calcium chloride 0.2g/l; Iron protochloride 0.05g/l; Zinc Sulphate Heptahydrate 0.03g/l; Manganous chloride tetrahydrate 0.02g/l; Trimethyl-glycine 0.012g/l; PH6.7～7.1.
5. a riboflavin fermentation cultural method, is characterized in that, comprising:
1) seed culture medium is packed into 10 m3 seeding tanks, after having sterilized, the bacillus subtilis spore suspension that access makes, the aerobic cultivation 12～16h of 120～140rpm at 37～39 ℃.Normal in seed liquor mycelia division, microscopy, without miscellaneous bacteria, every detection data, index all reach under the prerequisite of processing requirement, and the seeding tank that disappeared, to the connecting tube of fermentor tank, is prepared culture transferring;
2) by fermention medium with pump by connecting tube in magnetizing assembly suction 200m3 fermentor tank, after sterilization, sterilizing, fill into the aseptic Glucose Liquid that sterilization has magnetized, make the initial sugar concentration of fermentation tank culture medium reach 15g/L, with liquefied ammonia, regulate PH to 6.7～7.1, controlling ventilation ratio (VVM) is 1: 0.6; Move into cultured normal seed liquor, fermentation volume is controlled at 120～150m3, starts fermentation, keeps fermentation culture temperature at 37～39 ℃, and the sugar amount in controlled fermentation process, then augments and add 30% concentration sterilizing and magnetized Glucose Liquid; Start mechanical stirring, mixing speed is cumulative to 130rpm by 110rpm, maintains the interior dissolved oxygen of tank between 20%～30%.
6. a kind of riboflavin fermentation cultural method according to claim 5, is characterized in that, it is 0～24 hour that the sugar amount in described fermenting process is controlled, between glucose concn 10～15g/L; 24～32 hours, between glucose concn 5～8g/L; Between 32～40 hours glucose concn 2～5g/L; Put tank and within first two hours, stop sugar, residual sugar amount requires regulation and control at 1～2g/L.W-Gum liquid glucose with 30%～60% carries out restricted flow feeding.
7. a kind of riboflavin fermentation cultural method according to claim 5, is characterized in that, Glucose Liquid is through the magnetic field magnetisation of 0.03～0.08T.
8. according to a kind of riboflavin fermentation cultural method claimed in claim 5, it is characterized in that, the magneticstrength of described magnetizing assembly is 0.03～0.08T.
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Cited By (9)
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| CN106244650A (en) * | 2016-07-29 | 2016-12-21 | 山东鲁抗生物制造有限公司 | Tylosin new process for fermenting |
| CN106434818A (en) * | 2016-12-22 | 2017-02-22 | 广济药业（孟州）有限公司 | Fermentation medium for synthesizing riboflavin by bacillus subtilis |
| CN108795811A (en) * | 2018-06-21 | 2018-11-13 | 赤峰制药股份有限公司 | A kind of riboflavin production bacterium culture medium |
| CN108913747A (en) * | 2018-07-28 | 2018-11-30 | 广济药业（孟州）有限公司 | A kind of high density fermentation vitamin B2Method |
| CN109081478A (en) * | 2018-09-15 | 2018-12-25 | 南京霄祥工程技术有限公司 | A kind for the treatment of process of fermentation waste water |
| CN109609580A (en) * | 2018-12-26 | 2019-04-12 | 河南巨龙生物工程股份有限公司 | A kind of fermentation medium and its fermentation process of riboflavin |
| CN110564804A (en) * | 2019-09-12 | 2019-12-13 | 河南巨龙生物工程股份有限公司 | Clear liquid fermentation medium for producing riboflavin and fermentation method |
| CN112210577A (en) * | 2020-11-04 | 2021-01-12 | 赤峰蒙广生物科技有限公司 | Method for producing beta-thymidine by fermentation method |
| CN112280679A (en) * | 2020-11-04 | 2021-01-29 | 赤峰制药股份有限公司 | Method for producing vitamin B2 by fermentation method |
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Cited By (11)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106244650A (en) * | 2016-07-29 | 2016-12-21 | 山东鲁抗生物制造有限公司 | Tylosin new process for fermenting |
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| CN108795811A (en) * | 2018-06-21 | 2018-11-13 | 赤峰制药股份有限公司 | A kind of riboflavin production bacterium culture medium |
| CN108913747A (en) * | 2018-07-28 | 2018-11-30 | 广济药业（孟州）有限公司 | A kind of high density fermentation vitamin B2Method |
| CN109081478A (en) * | 2018-09-15 | 2018-12-25 | 南京霄祥工程技术有限公司 | A kind for the treatment of process of fermentation waste water |
| CN109081478B (en) * | 2018-09-15 | 2021-12-07 | 德州迈科生物技术有限公司 | Treatment process of fermentation wastewater |
| CN109609580A (en) * | 2018-12-26 | 2019-04-12 | 河南巨龙生物工程股份有限公司 | A kind of fermentation medium and its fermentation process of riboflavin |
| CN109609580B (en) * | 2018-12-26 | 2022-04-01 | 河南巨龙生物工程股份有限公司 | Fermentation medium and fermentation method of riboflavin |
| CN110564804A (en) * | 2019-09-12 | 2019-12-13 | 河南巨龙生物工程股份有限公司 | Clear liquid fermentation medium for producing riboflavin and fermentation method |
| CN112210577A (en) * | 2020-11-04 | 2021-01-12 | 赤峰蒙广生物科技有限公司 | Method for producing beta-thymidine by fermentation method |
| CN112280679A (en) * | 2020-11-04 | 2021-01-29 | 赤峰制药股份有限公司 | Method for producing vitamin B2 by fermentation method |
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