CN104560783A - Bacillus subtilis enrichment culture medium and application thereof - Google Patents
Bacillus subtilis enrichment culture medium and application thereof Download PDFInfo
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- CN104560783A CN104560783A CN201410735138.6A CN201410735138A CN104560783A CN 104560783 A CN104560783 A CN 104560783A CN 201410735138 A CN201410735138 A CN 201410735138A CN 104560783 A CN104560783 A CN 104560783A
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- manioc waste
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
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Abstract
The invention relates to a Bacillus subtilis enrichment culture medium which contains manioc waste, yeast extract, potassium dihydrogen phosphate, magnesium sulfate heptahydrate, and distilled water or deionized water. The application method of the Bacillus subtilis enrichment culture medium comprises the following steps: (1) pulverizing the manioc waste until the length of the manioc waste is 0.5-2mm, and sterilizing at 121 DEG C for 20 minutes; and (2) adding the yeast extract, potassium dihydrogen phosphate, magnesium sulfate heptahydrate, and distilled water or deionized water, inoculating the strain, and culturing for 16-24 hours while keeping the temperature at 30-35 DEG C. The yield of Bacillus subtilis cultured by the culture medium provided by the invention is obviously higher than that of the conventional carbon source, such as peptone, glucose, sucrose or the like. The culture medium increases the added value of manioc processing, and reduces the environmental pollution from industry. Further more, the manioc waste is cheaper than the glucose and sucrose, thereby greatly lowering the production cost; and therefore, the culture medium lays foundation for implementing high-yield large-scale production of Bacillus subtilis.
Description
Technical field
The present invention relates to a kind of bacterial classification enrichment medium and use thereof, particularly a kind of bacillus subtilis bacterium culture medium and use thereof.
Background technology
Manioc waste is cassava at waste left after starch factory squeezes starch or grain distillery produces alcohol, but still containing abundant Mierocrystalline cellulose, hemicellulose, starch and inorganic salt and other material, is a kind of very useful biomass resource.The main growing area of China cassava in Guangxi, Guangdong, Hainan, Yunnan, the ground such as Fujian, more than 1,000 ten thousand tons, cassava is produced in the whole nation per year.The generation of manioc waste is 1:2 (with dry basis) with the ratio of starch, and manioc waste output is very huge.The development of China cassava alcohol processing industry starts from before and after the nineties in 20th century, mainly with fresh cassava and tapioca slice for raw material production edible ethanol and industrial spirit.1/3 of current China alcohol output take cassava as raw material.Along with China to produce the development of alcohol fuel, cassava process deeply industry to the increase of starch demand and cassava, the demand for cassava also increases thereupon, and current domestic cassava production cannot satisfy the demands, every year need from a large amount of cassava of external import.While cassava usage quantity increases, the waste residue of generation---manioc waste also constantly increases.
For now, the process of manioc waste has following several: (1) anaerobically fermenting produces biogas; (2) fermentative production alcohol; (3) feed is produced; (4) fertilizer is produced.Although there is the treatment process of several manioc waste at present, non-popularization and application for various reasons, a large amount of manioc wastes is also in discarded state.The conventional carbon source of subtilis is glucose, sucrose etc., and cost is higher.
The information being disclosed in this background technology part is only intended to increase the understanding to general background of the present invention, and should not be regarded as admitting or imply in any form that this information structure has been prior art that persons skilled in the art are known.
Summary of the invention
The object of this invention is to provide one utilizes manioc waste to make subtilis enrichment medium and use thereof, reduces the cost producing subtilis while being intended to improve producing bacillus subtilis amount.
For achieving the above object, technical solution of the present invention is as follows:
A kind of subtilis enrichment medium, comprises following component raw material: manioc waste 18 ~ 22g, yeast extract paste 0.8 ~ 1.2g, potassium primary phosphate 0.8 ~ 1.2g, 7 water magnesium sulfate 0.15 ~ 0.22g, distilled water or deionized water 1000ml in mass ratio.
Wherein, count in mass ratio, described manioc waste to be water content be 70% manioc waste.
Wherein, described water content to be the manioc waste of 70% be cassava is processed after fresh manioc waste under natural lighting, be dried to water content be 70%.
A use for subtilis enrichment medium, comprises following operation steps:
(1) take required each component raw material, manioc waste being crushed to manioc waste length is 0.5 ~ 2mm, then at 121 DEG C of sterilizing 20min;
(2) add yeast extract paste, potassium primary phosphate, 7 water magnesium sulfates to gained manioc waste after sterilizing in step (1), distilled water or deionized water form substratum, inoculation bacterial classification, keep temperature to be 30 ~ 35 DEG C and cultivate 16 ~ 24h.
Wherein, stir once every 5 ~ 7h during cultivating bacterial classification in step (2).
Wherein, in step (2) amount of inoculation bacterial classification with bacterial classification source different and difference to some extent.
Compared with prior art, the present invention has following beneficial effect:
The substratum that the present invention utilizes manioc waste to make can provide the carbon source needed for subtilis in a large number, with the output of culture medium culturing subtilis of the present invention obviously the conventional carbon source such as such as peptone, glucose, sucrose want high; Further, the present invention increases the mode that manioc waste diversification is recycled, and adds the added value of wooden heat processing, decreases the pollution of this industry to environment; Further, manioc waste is more cheap than glucose, sucrose, greatly reduces production cost, therefore the present invention be realize the high yield of subtilis, large-scale production lays a good foundation.
Embodiment
Below the specific embodiment of the present invention is described in detail, but is to be understood that protection scope of the present invention not by the restriction of embodiment.
Embodiment 1
Being dried to manioc waste water content under fresh manioc waste after being processed by cassava is placed on sunlight is 70%, and then pulverizing manioc waste to length is 0.5 ~ 2mm, for subsequent use; Take each component raw material: above-mentioned the manioc waste 18g handled well, yeast extract paste 0.8g, potassium primary phosphate 0.8g, 7 water magnesium sulfate 0.15g, distilled water 1000ml, then adds yeast extract paste, potassium primary phosphate in manioc waste, 7 water magnesium sulfates, distilled water forms substratum, for subsequent use.
The subtilis followed these steps to preserving 1 month under 4 DEG C of conditions carries out pilot scale culture test; get this bacillus subtilis suspended liquid of 1ml and be inoculated in 250ml by the above-mentioned substratum prepared; constant temperature culture at 30 DEG C, every 7h shake once, cultivates 16h.Draw 1ml liquid production substratum bacterium liquid, gradient dilution is 10
7, 10
8, 10
9, draw appropriate above-mentioned each gradient dilution liquid and count under biomicroscope on blood counting chamber, if 3 parallel group.Concrete outcome is in table 1:
Table 1
Note: ※ represents that bacterium colony is too much, cannot count.
Result shows, the bacillus subtilis bacterium culture medium of what the present invention set up with manioc waste is primary carbon source by this formula to after the preservation subtilis of 1 month carries out pilot scale culture under 4 DEG C of conditions, viable bacteria concentration is 5.7 × 10
9cFU/ml.
Embodiment 2
Being dried to manioc waste water content under fresh manioc waste after being processed by cassava is placed on sunlight is 70%, and then pulverizing manioc waste to length is 0.5 ~ 2mm, for subsequent use; Take each component raw material: above-mentioned the manioc waste 22g handled well, yeast extract paste 1.2g, potassium primary phosphate 1.2g, 7 water magnesium sulfate 0.22g, distilled water 1000ml, then adds yeast extract paste, potassium primary phosphate in manioc waste, 7 water magnesium sulfates, distilled water forms substratum, for subsequent use.
Follow these steps to carry out pilot scale culture test to certain subtilis liquid fermentation agent on the market, get this bacillus subtilis suspended liquid of 1ml and be inoculated in the above-mentioned substratum prepared of 250ml, constant temperature culture at 35 DEG C, every 6h shake once, cultivate 20h.Draw 1ml liquid production substratum bacterium liquid, gradient dilution is 10
7, 10
8, 10
9, draw appropriate above-mentioned each gradient dilution liquid and count under biomicroscope on blood counting chamber, if 3 parallel group.Concrete outcome is in table 2:
Table 2
Result shows, what the present invention set up take manioc waste as the bacillus subtilis bacterium culture medium of primary carbon source and by this formula to after certain subtilis liquid fermentation agent carries out pilot scale culture on the market, viable bacteria concentration is 1.7 × 10
9cFU/ml.
Embodiment 3
Being dried to manioc waste water content under fresh manioc waste after being processed by cassava is placed on sunlight is 70%, and then pulverizing manioc waste to length is 0.5 ~ 2mm, for subsequent use; Take each component raw material: above-mentioned the manioc waste 20g handled well, yeast extract paste 1g, potassium primary phosphate 1g, 7 water magnesium sulfate 0.2g, distilled water 1000ml, then add yeast extract paste, potassium primary phosphate in manioc waste, 7 water magnesium sulfates, and distilled water forms substratum, for subsequent use.
Follow these steps to carry out pilot scale culture test to certain bacillus subtilis bacteria solid fermentation microbial inoculum on the market; getting this bacillus subtilis suspended liquid of 6g is inoculated in the substratum of 1000ml by formulated of the present invention; constant temperature culture at 32 DEG C, every 6h shake once, cultivates 24h.Draw 1ml liquid production substratum bacterium liquid, gradient dilution is 10
7, 10
8, 10
9, draw appropriate above-mentioned each gradient dilution liquid and count under biomicroscope on blood counting chamber, if 3 parallel group.Concrete outcome is in table 3:
Table 3
Result shows, the present invention set up take manioc waste as the bacillus subtilis bacterium culture medium of primary carbon source and after carrying out pilot scale culture by this formula to certain bacillus subtilis bacteria solid fermentation microbial inoculum, viable bacteria concentration is 4.3 × 10
9cFU/ml.
Can find out that the present invention is applied to different sources, different strain forms bacterial classification all can turn out the subtilis of high yield by above three case study on implementation, substratum wide adaptability of the present invention is described, effective, further the present invention has broad applicability in high yield subtilis.
The aforementioned description to concrete exemplary of the present invention is to illustrate and the object of illustration.These descriptions not want the present invention to be defined as disclosed precise forms, and obviously, according to above-mentioned instruction, can much change and change.The object selected exemplary embodiment and describe is to explain certain principles of the present invention and practical application thereof, thus those skilled in the art can be realized and utilize various different exemplary of the present invention and various different selection and change.Scope of the present invention is intended to limited by claims and equivalents thereof.
Claims (6)
1. a subtilis enrichment medium, is characterized in that, comprises following component raw material in mass ratio: manioc waste 18 ~ 22g, yeast extract paste 0.8 ~ 1.2g, potassium primary phosphate 0.8 ~ 1.2g, 7 water magnesium sulfate 0.15 ~ 0.22g, distilled water or deionized water 1000ml.
2. subtilis enrichment medium according to claim 1, is characterized in that: count in mass ratio, described manioc waste to be water content be 70% manioc waste.
3. subtilis enrichment medium according to claim 1, is characterized in that: described water content be 70% manioc waste be that under natural lighting, be dried to water content be 70% for fresh manioc waste after being processed by cassava.
4. a use for subtilis enrichment medium as claimed in claim 1, is characterized in that, comprises following operation steps:
(1) take required each component raw material, manioc waste being crushed to manioc waste length is 0.5 ~ 2mm, then at 121 DEG C of sterilizing 20min;
(2) add yeast extract paste, potassium primary phosphate, 7 water magnesium sulfates, distilled water or deionized water, inoculation bacterial classification to gained manioc waste after sterilizing in step (1), keep temperature to be 30 ~ 35 DEG C and cultivate 16 ~ 24h.
5. the use of subtilis enrichment medium according to claim 4, is characterized in that: stir once every 5 ~ 7h during cultivating bacterial classification in step (2).
6. the use of subtilis enrichment medium according to claim 4, is characterized in that: in step (2) amount of inoculation bacterial classification with bacterial classification source different and difference to some extent.
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| CN201410735138.6A CN104560783A (en) | 2014-12-05 | 2014-12-05 | Bacillus subtilis enrichment culture medium and application thereof |
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| CN201410735138.6A CN104560783A (en) | 2014-12-05 | 2014-12-05 | Bacillus subtilis enrichment culture medium and application thereof |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106222233A (en) * | 2016-08-03 | 2016-12-14 | 四川剑南春（集团）有限责任公司 | A kind of daqu fermentation power detection culture medium, the method for its detection daqu fermentation power of preparation method and application |
| CN114854634A (en) * | 2022-05-18 | 2022-08-05 | 四川洪光农业开发有限责任公司 | Culture medium formula of bacillus |
| CN115678809A (en) * | 2022-11-17 | 2023-02-03 | 塔里木大学 | Microbial agent for soil improvement |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103013961A (en) * | 2012-12-17 | 2013-04-03 | 华南理工大学 | Method for producing neutral protease and feed additive by using fermentation of manioc wastes |
| CN103652328A (en) * | 2013-12-13 | 2014-03-26 | 广西科技大学 | Method for producing high-quality high-protein feed through mixed strain solid-state fermentation of rice dregs and manioc waste |
-
2014
- 2014-12-05 CN CN201410735138.6A patent/CN104560783A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103013961A (en) * | 2012-12-17 | 2013-04-03 | 华南理工大学 | Method for producing neutral protease and feed additive by using fermentation of manioc wastes |
| CN103652328A (en) * | 2013-12-13 | 2014-03-26 | 广西科技大学 | Method for producing high-quality high-protein feed through mixed strain solid-state fermentation of rice dregs and manioc waste |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106222233A (en) * | 2016-08-03 | 2016-12-14 | 四川剑南春（集团）有限责任公司 | A kind of daqu fermentation power detection culture medium, the method for its detection daqu fermentation power of preparation method and application |
| CN114854634A (en) * | 2022-05-18 | 2022-08-05 | 四川洪光农业开发有限责任公司 | Culture medium formula of bacillus |
| CN115678809A (en) * | 2022-11-17 | 2023-02-03 | 塔里木大学 | Microbial agent for soil improvement |
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