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Application and method of using pseudomonas mendocina NK-01 for production of brown alginate oligosaccharides

A technology for the production of fucoidan oligosaccharides and Pseudomonas spp. is applied in the direction of microorganism-based methods, biochemical equipment and methods, microorganisms, etc., which can solve the problems of high price, easy inactivation of enzymes, and low yield, and achieve a fermentation cycle Short, simple process, overcome the cumbersome effect of separation and purification

Inactive Publication Date: 2011-09-14
NINGXIA WANSHENG BIO ENG
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The main disadvantage of the chemical synthesis method is that multiple group protection and deprotection steps are required, the yield is low, and a large number of synthetic complex compounds cannot be obtained by conventional chemical methods, so there is no practical industrial value
To prepare fucoidan oligosaccharides by enzymatic method, most of them are separated and purified to obtain fucoidan oligosaccharide lyase first, and then a series of fucoidan oligosaccharide products are prepared. Synthesis is also more expensive

Method used

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  • Application and method of using pseudomonas mendocina NK-01 for production of brown alginate oligosaccharides

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] Example 1, one-step fermentation to prepare fucoidan oligosaccharides (fermentation under low carbon and low nitrogen conditions)

[0022] The pre-cultivation is in the L-test tube, add 5ml of LB medium by aseptic operation, insert a single colony of NK-01 strain with a sterile toothpick, and cultivate at 30°C and 120rpm for 12h. Inoculate 0.5 ml of the pre-culture into a 500 ml Erlenmeyer flask containing 100 ml of LB medium, and culture with shaking at 150 rpm at 30°C for 24 hours. Sterile centrifuge at 6000g for 10min at 4°C, discard the supernatant, shake and mix with sterile phosphate buffer in a centrifuge tube, centrifuge again at 4°C, 6000g for 12min, discard the supernatant. The bacteria without LB medium were aseptically inserted into a 1-liter fermentation device with controllable oxygen stirring, which contained 1 liter of medium 1 (pH 7), and cultured at 30° C. for 48 hours with aeration. After the fermentation, the cells were removed by centrifugation, an...

Embodiment 2

[0023] Example 2, two-step fermentation to prepare fucoidan oligosaccharides (fermentation under low carbon and nitrogen-free conditions)

[0024] The pre-cultivation is in the L-test tube, add 5ml of LB medium by aseptic operation, insert a single colony of NK-01 strain with a sterile toothpick, and cultivate at 30°C and 120rpm for 12h. Inoculate 0.5 ml of the pre-culture into a 500 ml Erlenmeyer flask containing 100 ml of LB medium, and culture with shaking at 150 rpm at 28°C for 24 hours. Sterile centrifuge at 6000g for 10min at 4°C, discard the supernatant, shake and mix with sterile phosphate buffer in a centrifuge tube, centrifuge again at 4°C, 6000g for 12min, discard the supernatant. The bacterial cells without LB medium were aseptically inserted into 10 bottles of 500ml Erlenmeyer flasks containing 100ml of medium 2 (pH 7), cultured at 30°C with shaking at 150rpm. Finish fermentation after 48 hours, following steps are with embodiment 1.

Embodiment 3

[0025] Example 3, Preparation of Fucoidan Oligosaccharides by One-step Fermentation (Fermentation under Low Carbon and Low Nitrogen Conditions)

[0026] The pre-cultivation is in the L-test tube, add 5ml of LB medium by aseptic operation, insert a single colony of NK-01 strain with a sterile toothpick, and cultivate at 30°C and 120rpm for 12h. Inoculate 0.5 ml of the pre-culture into a 500 ml Erlenmeyer flask containing 100 ml of LB medium, and culture with shaking at 150 rpm at 30°C for 24 hours. Sterile centrifuge at 6000g for 10min at 4°C, discard the supernatant, shake and mix with sterile phosphate buffer in a centrifuge tube, centrifuge again at 4°C, 6000g for 12min, discard the supernatant. The bacteria without LB medium were aseptically inserted into a 1-liter fermentation device with controllable oxygen stirring, which contained 1 liter of medium 3 (pH 6), and cultured at 28°C for 60 hours with aeration. Following steps are with embodiment 1.

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Abstract

The invention discloses applications and a method of using pseudomonas mendocina NK-01 for the production of brown alginate oligosaccharides. The method comprises the following steps of: firstly carrying out activation culture on the bacteria in an L tube containing 5ml LB culture medium, then inoculating the activated microbial strain onto a fermentation culture medium for culture, centrifugallyremoving the bacteria after the completion of the fermentation, using 3 times of volume of anhydrous ethanol for carrying out sedimentation on supernatant liquid, using water for dissolving sediment after removing the supernatant liquid, then using qualitative filter paper for filtration, freezing and drying the filtrate, and obtaining the product of brown alginate oligosaccharides. The brown alginate oligosaccharides can be widely applied a plurality of fields such as foods, pharmaceutical, chemical and the like. The method overcomes the shortcomings that chemical method which is usually usedfor preparing the brown alginate oligosaccharides needs multi-group protection and the de-protection steps, the operation is complicated, the enzymatic method is easy to be inactivated and the priceis expensive. The production method of the invention has simple process and short fermentation cycle; furthermore, the fermentation conditions and the process lead the large-scale industrial production to be possible.

Description

technical field [0001] The present invention relates to the use of Pseudomonas mendoza to produce polysaccharide biodegradable macromolecules, especially the use of Pseudomonas mendocina (Pseudomonas mendocina) NK-01CCTCC M 208005 (referred to as Pseudomonas mendoza NK -01) Use and method for producing fucoidan oligosaccharides. Background technique [0002] Fucoidan is an oligomer of fucoidan, which is a fucoidan with a degree of polymerization of less than 20. Fucoidan is a polysaccharide synthesized by seaweed and bacteria. In seaweed, fucoidan mainly acts as a skeleton, while in bacteria, fucoidan is a component of the cell wall, which prevents antibiotics from entering the cell, and some bacteria synthesize it. Fucoidan is secreted extracellularly. Sodium alginate or sodium alginate is a product in which the carboxyl hydrogen of uronic acid on the alginate polysaccharide chain is replaced by sodium, and they are also known as alginate. [0003] Fucoidan from seaweed ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12R1/38C12P19/04
Inventor 宋存江郭文斌王淑芳曹名锋耿伟涛
Owner NINGXIA WANSHENG BIO ENG
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