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Gluconobacter oxydans and method for preparing ketoxylose using the same

A technology of Gluconobacter oxidans and xylulose, applied in the fields of fermentation engineering and enzyme engineering, can solve problems such as increased production costs, separation and purification of unfavorable products, and difficulties in downstream processes

Active Publication Date: 2011-06-08
SHANDONG TIANLI PHARMA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

This is mainly because: on the one hand, cells are in a free state and cannot be recycled; on the other hand, it makes the downstream process very difficult, which is not conducive to the separation and purification of products, and the production cost is greatly increased.

Method used

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  • Gluconobacter oxydans and method for preparing ketoxylose using the same

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Incline medium: glucose 30g / L, yeast extract 10g / L, beef extract 5g / L, agar 20g / L.

[0044] Shake flask medium: glucose 30g / L, yeast extract 25g / L, beef extract 5g / L, D-arabitol 10g / L, KH 2 PO 4 5g / L.

[0045] Cultivate Staphylococcus oxidans CGMCC No.2709 on a slant medium at 30°C for 24 hours, then put a ring of this bacteria in the shake flask medium, cultivate it at 30°C for 10 hours, and the shake flask rotates at a speed of 200r / min. The content of D-ara is 20g / L, and the enzyme production is 2.5~5.0U / mL. It is transformed by free cells. Weigh 1g of wet cells after centrifugation and add them to 10mL of 30g / L D-ara solution (the amount of cells added is calculated as per 100g of cells transformed 1L 30g / L D-ara solution), under the condition of 30°C, stir and shake in a shaker flask (rotating speed 120r / min) to convert and produce D-xylulose, the reaction time is 9h, and the conversion rate of the substrate is measured after the reaction is terminated and prod...

Embodiment 2

[0047] With 30g / L maltose as the carbon source, 20g / L yeast extract, and 10g / L bean cake powder as the nitrogen source, the other components of the medium and the cell culture conditions are the same as in Example 1, and a 1.5% sodium alginate colloid solution is prepared, and the physiological After mixing the bacterial suspension prepared with saline evenly, add 8% diatomaceous earth for adsorption, and drop 2% CaCl with a syringe 2 Solidify in the solution to make spherical immobilized cells with a diameter of about 3-4mm, and put them in a refrigerator at 4°C for several hours, then pour off the supernatant, wash twice with distilled water, and replace 1g of the centrifuged wet cells with seaweed 5 g of immobilized cells were obtained by embedding in sodium phosphate, and transformed into 10 mL of 50 g / L D-arabitol solution (rotational speed 200 r / min) in a shake flask at 30 ° C. The transformation time of each batch was 13 h, and 30 batches were transformed. times, the av...

Embodiment 3

[0049] Glucose 225g, yeast extract 112.5g, beef extract 22.5g, D-arabitol 45g, KH 2 PO 4 22.5g, pH 5.0, dilute to 4.5L with water, make culture medium, put into 7.5L stirring fermenter, steam sterilize at 121℃ for 15min. Prepare seed medium: glucose 30g / L, yeast extract 10g / L, beef extract 5g / L. Put Gluconobacter oxydans CGMCCNo.2709 in the seed medium one by one, and culture it at 30°C for 8 hours to obtain the seed liquid. Put the seed liquid into the cooled fermentation medium according to the inoculation amount of 3% of the volume of the fermentation liquid, and cultivate it at 30°C. (Ventilation 0.7vvm, stirring speed 600r / min) 48h. The fermented liquid is subjected to bacterial filtration treatment in an ultrafilter, and the molecular weight cut-off of the ultrafiltration membrane is 10 6 Da, the operating pressure is 0.2MPa, the temperature is controlled at 50°C, and the membrane surface velocity is 4m / s. After immobilizing the 100g bacterium obtained by interceptio...

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Abstract

The invention discloses a Gluconobacter oxydans, which is classified and named as Gluconobacter oxydans NH-10, and preserved in China General Microbiological Culture Center of Microbial Culture Collection Management Committee with the preservation number of CGMCC No.2709. The invention further discloses a D-xylulose preparation method that uses the Gluconobacter oxydans. The strain obtained by screening can transform D-Arabitol so as to generate D-xylulose in high efficiency, the D-xylulose preparation method reaches the highest D-Arabitol transformation ratio of 99.5 percent (w / w) and the D-xylulose yield ratio of 95 percent, the D-xylulose concentration of a conversion fluid can reach 95g / L, and the conversion fluid hardly contains other ingredients.

Description

technical field [0001] The invention belongs to the technical fields of fermentation engineering and enzyme engineering, and relates to a D-arabitol dehydrogenase-producing microorganism Gluconobacter oxydans and a method for using it to produce D-xylulose. Background technique [0002] Xylulose (xylulose) is a ketopentose. L-xylulose is excreted in the urine of pentoseuria. In vivo, the L-type and D-type xylulose are first reduced to xylitol, and then reoxidized to convert each other. D-xylulose reacts with ATP to generate D-xylulose-5-phosphate and enters the pentose phosphate pathway. In addition, glucose forms D-xylulose-5-phosphate through 6-phosphogluconic acid→D-ribulose-5-phosphate, which is an important intermediate in the pentose phosphate pathway of another glycolysis pathway. In addition, D-xylulose can be used as a material for the production of xylitol, which is one of the important sweeteners in the food industry and other fields. Xylitol is a natural five...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20C12P19/02C12P7/18C12R1/01
Inventor 徐虹朱宏阳李莎赵敏蔡恒
Owner SHANDONG TIANLI PHARMA
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