A kind of method of from the brown alga oligose fermented liquid, separating brown alga oligoseTechnical field
The invention belongs to technical field of biological fermentation, particularly relate to a kind of from the synthetic method of separating brown alga oligose the fermented liquid of brown alga oligose that makes of glucose.
Background technology
Brown alga oligose is the oligomer of algal polysaccharide, is the less algal polysaccharide of the polymerization degree.Algal polysaccharide is by marine alga and a kind of polysaccharide of bacterium synthetic; Algal polysaccharide mainly plays skeleton function in the marine alga; And algal polysaccharide is a kind of composition of cell walls in the bacterium, resists bad environment, and some bacterium then is secreted into the extracellular with the synthetic algal polysaccharide.Has identical substruction from the algal polysaccharide of bacterium with algal polysaccharide from marine alga; Difference only is in the bacterium algal polysaccharide that C2 and/or C3 hydroxyl usually are acetylation to some extent, and degree of acetylation has evident difference because of the difference of bacterial strain and growth conditions.Algal polysaccharide and oligomer thereof have been widely used in a plurality of fields such as food, pharmacy, chemical industry, have the wide development prospect, as the algal polysaccharide and the oligosaccharides that are rich in mannuronic acid have effects such as antitumor and immunomodulatory.It is reported that brown alga oligose has is anticancer significantly, anti-freezing, lipopenicillinase, immunomodulatory and anti-aging effects, both can be used as protective foods, also can be used as medicine or sterilant or sanitas.Obviously receive the influence of the place of production, season and extraction process technology from the brown alga oligose The Nomenclature Composition and Structure of Complexes (G, M arrangement of fragments mode) of extracting algae, cause performance difference obvious, cause the as a result poor repeatability of microcapsule in experimentation on animals of preparation.
The Production by Microorganism Fermentation brown alga oligose can be realized stable quality control, therefore, has become the focus of brown alga oligose research.At present; Main microbe fermentation method has the microbe fermentation method that utilizes the bacterial classification that produces algin catenase to produce the enzyme liberating algin and obtain brown alga oligose; In addition pseudomonas mendocina is inoculated on the substratum that glucose is carbon source; 30 ℃, the big ventilation are with method (i.e. " pseudomonas mendocina NK-01 is used to produce the purposes and the method for brown alga oligose ", the application number 200910068579.4 of the synthetic brown alga oligose of glucose.Guo Wenbin, Wang Shufang, Cao Mingfeng; Geng Weitao, Song Cunjiang. synthetic brown alga oligose of pseudomonas mendocina Pseudomonas mendocina NK-01 and structure thereof are identified. biological journal, 25 (9); 1366-1370. No. 005 Production by Microorganism Fermentation brown alga oligose of Ningxia Hui Autonomous Region's scientific and technological achievement assay certificate peaceful section mirror word (2010)); Aforesaid method technology is simple, and fermentation period is short, but process for extracting lacks research; Therefore, it is significant to study a kind of effective process for extracting.The process for extracting of Production by Microorganism Fermentation brown alga oligose mainly contains at present: organic solvent precipitation method, the chemical precipitation method or the membrane sepn precipitator method obtain brown alga oligose.But the said extracted method is only used in the laboratory, and has cost height, complicated, the low and ropy problem of extraction product purity of method.
Goal of the invention
The objective of the invention is to overcome the defective of above-mentioned prior art, a kind of industrial production that meets is provided, technology is simple, and production cost is low, and extracts product purity height, the measured method of separating brown alga oligose from the brown alga oligose fermented liquid of matter.
For realizing that the technical scheme that the object of the invention adopted is:
1, puts brown alga oligose (pseudomonas mendocina is inoculated on the substratum that glucose is carbon source 30 degrees centigrade, ventilates, cultivated 48 hours greatly), detect brown alga oligose content in the fermented liquid, PH;
2, carry fermented liquid to ultra-filtration membrane (aperture is molecular weight 10000-100000 dalton) degerming, add deionized water gradation dialysis in the process, remove thalline and foreign protein like the 20%-70% of fermented liquid weight;
3, ultrafiltrated is gone to nf membrane (aperture is molecular weight 300-1000 dalton) system and concentrate and desalting treatment, small-molecule substances such as salt are removed in the deionized water gradation dialysis that adds the 10%-40% of ultrafiltrated weight in the process;
4, nanofiltration liquid concentrator metered volume is gone to crystallizer, the anhydrous edible ethanol with its 4 times of volumes is delivered to crystallizer simultaneously, opens the stirring and evenly mixing temperature control and leaves standstill 20-48 hour for 5 ℃-10 ℃;
5, feed liquid constant voltage in the crystallizer is delivered to Plate Filtration, filtrating is delivered to the alcohol recovery system;
6, the sheet frame filter cake is delivered to vacuum lyophilization or microwave drying to water cut reaches below 5%, make the brown alga oligose product.
The present invention compares with prior art has following technical superiority:
1, through adopting the ceramic membrane system below the 100000D aperture can effectively remove thalline (the fermentation strain size is at the 0.1-0.2 micron), the filtered liq microscopy visual field there is not thalline.And traditional technology organic solvent precipitation method or chemical precipitation method are difficult to thalline is thoroughly eliminated.
2,The nf membrane system of employing 300-1000D has concentrated with the volume-diminished of feed liquid about 5 times; Practiced thrift the alcohol of 16 times of liquid concentrator volumes accordingly; And the situation (molecular weight product is at 1000D-2000D) that does not exist product to cross film to lose; The above nf membrane in 300D aperture can be so that most salt sees through, and detected result shows: do not use nf membrane product ash content more than 70%, use the back ash content to stablize and be controlled at below 20%.
3, conventional high temperature drying way can make that product blackout, Jiao stick with paste, taste bad will.The present invention adopts low temperature or microwave drying to make that product property is good, steady quality.
In sum, brown alga oligose novel technology for extracting of the present invention can effectively be removed impurity in the fermented liquid, and the extraction product purity is high, quality is good, and technology is simple, and production cost is low.
Embodiment
Embodiment 1
With tank body volume 500L fermentor tank is example, takes the present invention to extract oligosaccharides technology, and step is following:
1, detects by the false pseudomonas bacillus of Mendoza through the fermented liquid of cultivating, fermentation makes, record that brown alga oligose content is 5.3g/100mL, PH 6.7, volume 354L in the fermented liquid.
2, carry the degerming of fermented liquid to 50000D ultra-filtration membrane, add deionized water 180L in the process and divide 3 dialysis, remove thalline and foreign protein 35L.
3, ultrafiltrated 499L is gone to 300D nf membrane system and concentrates and desalting treatment, add deionized water 40L divide dialyse for 3 times liquid concentrator 66L.
4, nanofiltration liquid concentrator 66L is gone to crystallizer, simultaneously the anhydrous edible ethanol of 264 L is delivered to crystallizer, open stirring and evenly mixing and left standstill 20-48 hour at 5 ℃-10 ℃.
5, feed liquid constant voltage in the crystallizer is delivered to Plate Filtration.Filtrating is delivered to the alcohol recovery system, the evaporation recycling.
6, the sheet frame filter cake is delivered to vacuum lyophilization or microwave drying to water cut, make brown alga oligose product 16.5Kg.
Embodiment 2
With tank body volume 500L fermentor tank is example, takes the present invention to extract oligosaccharides technology, and step is following:
1, detects by the false pseudomonas bacillus of Mendoza through the fermented liquid of cultivating, fermentation makes, record that brown alga oligose content is 5.5g/100mL, PH 6.6, volume 350L in the fermented liquid.
2, carry the degerming of fermented liquid to 10000D ultra-filtration membrane, add deionized water 180L in the process and divide 3 dialysis, remove thalline and foreign protein 38L.
3, ultrafiltrated 492L is gone to 1000D nf membrane system and concentrates and desalting treatment, add deionized water 40L divide dialyse for 3 times liquid concentrator 50L.
4, nanofiltration liquid concentrator 50L is gone to crystallizer, simultaneously the anhydrous edible ethanol of 200 L is delivered to crystallizer, open stirring and evenly mixing and left standstill 20-48 hour at 5 ℃-10 ℃.
5, feed liquid constant voltage in the crystallizer is delivered to Plate Filtration.Filtrating is delivered to the alcohol recovery system, the evaporation recycling.
6, the sheet frame filter cake is delivered to vacuum lyophilization or microwave drying to water cut, make brown alga oligose product 16.7Kg.
Embodiment 3
With tank body volume 500L fermentor tank is example, takes the present invention to extract oligosaccharides technology, and step is following:
1, detects by the false pseudomonas bacillus of Mendoza through the fermented liquid of cultivating, fermentation makes, record that brown alga oligose content is 5.2g/100mL, PH 6.7, volume 361L in the fermented liquid.
2, carry the degerming of fermented liquid to 100000D ultra-filtration membrane, add deionized water 180L in the process and divide 3 dialysis, remove thalline and foreign protein 37L.
3, ultrafiltrated 504L is gone to 1000D nf membrane system and concentrates and desalting treatment, add deionized water 40L divide dialyse for 3 times liquid concentrator 60L.
4, nanofiltration liquid concentrator 60L is gone to crystallizer, simultaneously the anhydrous edible ethanol of 240 L is delivered to crystallizer, open stirring and evenly mixing and left standstill 20-48 hour at 5 ℃-10 ℃.
5, feed liquid constant voltage in the crystallizer is delivered to Plate Filtration.Filtrating is delivered to the alcohol recovery system, the evaporation recycling.
6, the sheet frame filter cake is delivered to vacuum lyophilization or microwave drying to water cut, make brown alga oligose product 16.5Kg.
The invention is not restricted to the scheme that above-mentioned embodiment provides.