A kind of online evaluation method of cellulosic production Acetobacter xylinum cell concentration

Acetobacter xylinum, bacteria concentration technology, applied in the direction of biochemical equipment and methods, microbial measurement/inspection, material capacitance, etc., can solve the problems of interfering light transmission, too large deviation of detection results, and bacterial contamination, etc., to improve Bacterial concentration, shortening the culture cycle, and reducing the effect of entanglement

Inactive Publication Date: 2019-10-18
东莞市东阳光生物合成药有限公司
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AI Technical Summary

Problems solved by technology

[0005] 1) Traditional offline detection methods, such as spectrophotometry and dry-wet weight method, the detection principle of the former is to quantitatively analyze the bacteria by measuring the absorbance or luminous intensity of light at a specific wavelength or within a certain wavelength range, but due to The culture medium of Acetobacter xylinum has a darker color and the cellulose filaments produced will interfere with the absorption of light, resulting in large deviations in the test results; Weight or dry weight, but because the cellulose filaments produced by the culture of Acetobacter xylinum will also be centrifuged down with the bacteria, and occupy a relatively large proportion, the test results are not accurate
Another commonly used off-line detection method is the plate colony counting method. The detection results are relatively accurate, but this method is time-consuming and laborious to operate. It usually takes 3 to 4 days to get the results, and there is a large time lag.
In addition, the above methods all need to take samples once in a while, and the bioreactor needs to be opened during the sampling process, which is very easy to bring in miscellaneous bacteria and cause bacterial contamination.
[0006] 2) On-line detection methods, such as on-line optical turbidimetry, but the color of the culture medium of Acetobacter xylinum, insoluble particles and a large number of air bubbles will interfere with the transmission of light. Although there is no trouble of sampling, the test results can also be updated online from time to time, but the detection The deviation of the result is too large, so it cannot be applied to the evaluation of the concentration of Acetobacter xylinum

Method used

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  • A kind of online evaluation method of cellulosic production Acetobacter xylinum cell concentration
  • A kind of online evaluation method of cellulosic production Acetobacter xylinum cell concentration
  • A kind of online evaluation method of cellulosic production Acetobacter xylinum cell concentration

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] (1) Strain activation

[0039] Preparation of solid medium: glucose 2%-3%, polypeptone 0.5%-1%, citric acid 0.1%-0.2%, magnesium sulfate heptahydrate 0.03%-0.05%, ammonium sulfate 0.5%-1%, diphosphate Potassium hydrogen 0.3%-0.5%, sodium acetate 0.1%-0.2%, agar powder 2%-2.5%.

[0040] The freeze-dried preserved Acetobacter xylinum was spread on the plate solid medium, and cultured at 28°C for 4 days to form a single colony. Select a single colony cultivated above, spread it evenly on the solid medium on the slope with an inoculation loop, and cultivate it in a constant temperature incubator at 28°C for 4 days.

[0041] (2) shake flask culture

[0042] Preparation of liquid medium: 2%-3% glucose, 0.5%-1% yeast powder, 0.115%-0.2% citric acid, 0.34%-0.5% sodium hydrogen phosphate dodecahydrate, 0.5%-1% soybean peptone, Diammonium hydrogen phosphate 0.1%-0.2%.

[0043]Use a sterile spatula to take out the cultured slant film, put it into the shake flask liquid culture...

Embodiment 2

[0053] (1) Strain activation

[0054] Preparation of solid medium: glucose 2%-3%, polypeptone 0.5%-1%, citric acid 0.1%-0.2%, magnesium sulfate heptahydrate 0.03%-0.05%, ammonium sulfate 0.5%-1%, diphosphate Potassium hydrogen 0.3%-0.5%, sodium acetate 0.1%-0.2%, agar powder 2%-2.5%.

[0055] The freeze-dried preserved Acetobacter xylinum was spread on the plate solid medium, and cultured at 30°C for 6 days to form a single colony. Select the single colony cultured above, spread it evenly on the solid medium on the slope with an inoculation loop, and cultivate it in a constant temperature incubator at 30°C for 3 days.

[0056] (2) shake flask culture

[0057] Preparation of liquid medium: 1%-4% fructose, 0.5%-2% corn steep liquor, 0.2%-0.5% citric acid, 0.2%-0.5% sodium hydrogen phosphate dodecahydrate, 0.03%-0.05 magnesium sulfate heptahydrate %, diammonium hydrogen phosphate 0.4%-1%.

[0058] Use a sterile spatula to take out the cultured slant film, insert it into the s...

Embodiment 3

[0068] (1) Strain activation

[0069] Preparation of solid medium: glucose 2%-3%, polypeptone 0.5%-1%, citric acid 0.1%-0.2%, magnesium sulfate heptahydrate 0.03%-0.05%, ammonium sulfate 0.5%-1%, diphosphate Potassium hydrogen 0.3%-0.5%, sodium acetate 0.1%-0.2%, agar powder 2%-2.5%.

[0070] The freeze-dried and preserved Acetobacter xylinum was spread on the plate solid medium, and cultured at 32°C for 8 days to form a single colony. Select a single colony cultivated above, spread it evenly on the solid medium on the slope with an inoculation loop, and cultivate it in a constant temperature incubator at 32°C for 2 days.

[0071] (2) shake flask culture

[0072] Preparation of liquid medium: glucose 2-3%, corn steep liquor dry powder 0.5%-1.5%, citric acid 0.1%-0.3%, sodium hydrogen phosphate dodecahydrate 0.1%-0.3%, magnesium sulfate heptahydrate 0.02%-0.04% , Diammonium hydrogen phosphate 0.2%-0.6%.

[0073] Use a sterile spatula to take out the cultured slant film, ins...

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Abstract

The invention relates to an online evaluation method of the concentration of acetobacter xylinus capable of generating cellulose. According to the online evaluation method, a model used for representing the relationship between bacteria concentration and capacitivity is established; an online capacitivity detection device is adopted for real time monitoring of the capacitivity value of a culturing solution; the model used for representing the relationship between bacteria concentration and capacitivity is established, so that real time observing on the change trend of bacteria concentration can be realized, and culturing endpoint is determined based on a bacteria concentration change trend chart. The online evaluation method is capable of avoiding influences of culture medium, cellulose filament, insoluble particles, and foam on bacteria concentration measuring results, and possesses excellent application effect in experimental production, pilot scale production, and industrial scale production.

Description

technical field [0001] The invention belongs to the field of biological fermentation, in particular to an online evaluation method for cellulosic Acetobacter xylinum concentration. Background technique [0002] Cellulose (Bacterial Cellulose, BC) is a kind of inert polymer material synthesized by bacteria. Similar to the structure of natural cellulose, it is composed of glucose connected by β-1,4-glucosidic bonds. Bacterial cellulose has a unique physical structure and properties, such as excellent bioaffinity, biocompatibility, biocompatibility and good biodegradability, and is recognized as a new type of biological material with excellent performance in the world. . At present, BC has been successfully applied in food, medicine, chemical industry, papermaking, beauty treatment and membrane filtration. At present, the bacteria capable of producing BC mainly include Acetobacter, Rhizobium, Agrobacterium, and Sarcina, among which Acetobacter xylinus has the highest yield a...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/04G01N27/22
Inventor 贾永峰封海生毛兴艳宁荣良黄成潭王欢
Owner 东莞市东阳光生物合成药有限公司
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