CN103131643B - Strain for producing mannitol and method for producing mannitol through fermentation of strain - Google Patents
Strain for producing mannitol and method for producing mannitol through fermentation of strain Download PDFInfo
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- CN103131643B CN103131643B CN201310081411.3A CN201310081411A CN103131643B CN 103131643 B CN103131643 B CN 103131643B CN 201310081411 A CN201310081411 A CN 201310081411A CN 103131643 B CN103131643 B CN 103131643B
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Abstract
The invention discloses a strain for producing mannitol and a method for producing mannitol through fermentation of the strain, and belongs to the technical field of food biologics. The invention provides Candida parapsilosis SK26.003 which is screened from sugarcane juice and is preserved in the Chinese Typical Culture Collection Center with the preservation number of CCTCC NO: M2012491. The invention further provides a method for producing mannitol through fermentation of the Candida parapsilosis strain. The method comprises the following steps of: (a) fermenting the strain in a fermentation culture medium containing glucose so as to obtain a fermentation liquid containing mannitol; and (b) separating and purifying mannitol from the fermentation liquid. Mannitol produced by using the strain and the method is safe and reliable, is a functional product with much market potential, and is widely applied to industries such as food, cosmetics, medicament and the like. The strain can be used for producing mannitol in high efficiency and is applicable to large-scale production, and a new method is provided for the industrial preparation of mannitol.
Description
Technical field
The present invention relates to produce N.F,USP MANNITOL a kind of Candida parapsilosis (
candida parapsilosis) SK26.003 screening and utilize the method for this yeast strain fermentative production N.F,USP MANNITOL, belong to technical field of food biotechnology.
Background technology
That N.F,USP MANNITOL has is non-hygroscopic, sugariness is suitable, heat is low, have no side effect, human body metabolism and Regular Insulin, have nothing to do, do not improve the features such as blood glucose value, unlikely carious tooth, can be used as sweeting agent and the functional food additives of diabetics, adiposis patient, also can be used for the antiseized of the food such as maltose, chewing gum, rice cake, and the antiseized powder of the general cake of conduct.Take N.F,USP MANNITOL as raw material can also synthesize multiple important fine-chemical intermediate.
Because the consumption of low calorie foods in recent years sharply increases, at present, in world wide, the consumption of N.F,USP MANNITOL is sharp increase trend.Up to now, the manufacture method of N.F,USP MANNITOL, mainly contains three kinds.
The first, natural extraction method.In China, from marine alga, sea-tangle, extract N.F,USP MANNITOL is one of suitability for industrialized production N.F,USP MANNITOL method always, and main recrystallization or the electrodialysis desalination of adopting extracts N.F,USP MANNITOL from marine alga, sea-tangle at present.The method principle is simple, but process is complicated, and yield less energy-consumption is large, and production cost is higher.
The second, chemical catalysis.Chemical catalysis is produced N.F,USP MANNITOL and be take sucrose, starch or glucose and by nickel shortening, prepare as raw material.This method cost is low, but its yield is lower, and has sorbyl alcohol by product, increases later separation purifying cost.
The 3rd, biotransformation method.The method of producing N.F,USP MANNITOL by biotechnology has enzyme process, fermentation method etc., and fermentation method is better than enzyme process generally.The advantage of fermentation method is that cost is low, and output is high, and reaction conditions is gentle, and by product is few, there is no the generation of toxic substance in the production process of product, convenience is provided to product post-treatment, has reduced cost, and is applicable to large-scale production.
Summary of the invention
The object of this invention is to provide a kind of new microorganism Candida parapsilosis (
candida parapsilosis) SK26.003 bacterial strain, it can be take high concentration glucose as raw material and needn't add fructose, fermentative production N.F,USP MANNITOL.
Another object of the present invention is to provide a kind of method of utilizing above-mentioned Candida parapsilosis bacterial strain to produce N.F,USP MANNITOL.
A further object of the present invention is to provide a kind of from the method containing separation and purification N.F,USP MANNITOL the fermented liquid of N.F,USP MANNITOL.
Technical scheme of the present invention: separatedly from sugar cane juice obtain the bacterial strain that N.F,USP MANNITOL is produced in a strain, according to the mushroom of certain rule learn character, the analysis of biochemical property and the mensuration of 16sRNA sequence draw its Classification And Nomenclature be Candida parapsilosis (
candida parapsilosis) SK26.003, being preserved in Chinese Typical Representative culture collection center, deposit number is: CCTCC NO:M 2012491.
The method of utilizing the bacterial strain of described product N.F,USP MANNITOL to produce N.F,USP MANNITOL: with described microorganism Candida parapsilosis (
candida parapsilosis) CCTCC NO:M 2012491 is starting strain, through seed culture, liquid submerged fermentation, produces N.F,USP MANNITOL.Concrete steps are:
(1) seed culture
Seed culture medium: glucose 5-20g/L, yeast extract paste 5-20g/L, deionized water preparation;
Seed culture condition: in 30-37 ℃, under the rotating speed of 160-200rpm, shaking table is cultivated 10-24h.
(2) fermentation culture
Fermention medium: glucose 100-500g/L, yeast extract paste 10-40g/L, urea 1-10g/L, magnesium sulfate 0-10g/L, sodium-chlor 0-10g/L, calcium chloride 0-5g/L, potassium primary phosphate 0-10g/L, adjusts pH5-7;
Fermentation condition: inoculum size 1%-10%, rotating speed 160-200rpm, 3-5d ferments under the condition of temperature 30-37 ℃ in fermention medium.
(3) extraction of N.F,USP MANNITOL
By fermented liquid centrifuging, after activated carbon decolorizing, utilize cation and anion exchange post to carry out desalination; The calcium type Zeo-karb of take is sorbent material, and 50-70 ℃ with water elution, obtains the N.F,USP MANNITOL of purifying after condensing crystal.
Beneficial effect of the present invention: the present invention relates to that a strain is screened from sugar cane juice and the Candida parapsilosis that comes (
candida parapsilosis) SK26.003, being preserved in Chinese Typical Representative culture collection center, deposit number is CCTCC NO:M 2012491.Take in the fermention medium that glucose is carbon source through the fermentation of 3-5d, residual sugar can reach below 5%, and the mannitol concentration in fermented liquid reaches 20-100g/L, for the preparation of industrialization of N.F,USP MANNITOL provides new method.The N.F,USP MANNITOL that the present invention produces is safe and reliable, is a kind of functional product that has very much market potential, is widely used in the industries such as food, makeup, medicine.The present invention can produce N.F,USP MANNITOL efficiently, is suitable for carrying out scale operation.
Biological material specimens preservation: the bacterial strain of N.F,USP MANNITOL is produced in a strain, its Classification And Nomenclature be Candida parapsilosis (
candida parapsilosis) SK26.003, be preserved in Chinese Typical Representative culture collection center, be called for short CCTCC, address: Wuhan, China Wuhan University, deposit number is CCTCC NO:M 2012491, preservation date on November 30th, 2012.
Embodiment
Be below Candida parapsilosis (
candida parapsilosis) SK26.003 ferments and produces the embodiment of N.F,USP MANNITOL, but technical scope of the present invention is not limited to listed several examples, is not changing under the prerequisite of its main points, can make various changes and implement.In addition, technical scope of the present invention prolongs and impartial scope.
Embodiment 1
The separated strain N.F,USP MANNITOL obtaining from sugar cane juice is produced to bacterium and is sent to Chinese Typical Representative culture collection center and measures the morphological specificity of this bacterium, physio-biochemical characteristics and 16sRNA gene sequencing, be accredited as Candida parapsilosis (
candida parapsilosis) SK26.003.
The fermentation culture of embodiment 2 Candida parapsilosis SK26.003 bacterial strains
In culture medium A, above-mentioned bacterial strains is cultivated 48 hours in 30 ℃.With this culture, be seeded in the seed culture medium B that contains 100mL, aerlbic culture 24h at 30 ℃, then transfer in 2L fermention medium C, 30 ℃ of bottom fermentation 3d, stirring velocity 200rpm.
Culture medium A: glucose 20g/L, yeast extract paste 10g/L, agar 2%;
Seed culture medium B: glucose 20g/L, yeast extract paste 10g/L;
Fermention medium C: glucose 150g/L, yeast extract paste 15g/L, urea 1g/L, magnesium sulfate 0.5g/L, sodium-chlor 0.1g/L, calcium chloride 0.1g/L, potassium primary phosphate 2g/L, adjusts pH6.5.
When stopping fermentation culture, frozen centrifugation fermented liquid.The concentration that detects the N.F,USP MANNITOL producing in supernatant liquor by high performance liquid chromatography (HPLC), concentration reaches 30g/L.
The separation and purification of embodiment 3 N.F,USP MANNITOL
Centrifuging: fermented liquid is removed thalline after frozen centrifugation, collects the supernatant liquor containing N.F,USP MANNITOL.
Decolouring: add 1% gac in filtrate, keep 40min at 40 ℃, filtering separation is until filtrate clarification;
Desalination: destainer is successively carried out to desalination by cationic exchange coloum 001 * 7 and anionite-exchange resin 313;
Separation and purification: be 60 ℃ in separation temperature, take calcium type Zeo-karb under the condition that sorbent material, the water of take are eluent, N.F,USP MANNITOL is carried out to separation and purification, obtain being continuously rich in the component of N.F,USP MANNITOL;
Concentrated: by rotary evaporation, to be concentrated into solid content more than 70%;
Crystallization: in concentrated solution, add the dehydrated alcohol of 4 times of volumes, and add N.F,USP MANNITOL crystal seed, temperature to remain on 4 ℃, a standing night, crystallizable.Crystallization of mannitol is filtered, use a small amount of alcohol flushing, then, in 40 ℃ of dry 10h, obtain N.F,USP MANNITOL crystal.
Claims (2)
1. the bacterial strain of N.F,USP MANNITOL is produced in a strain, its Classification And Nomenclature be Candida parapsilosis (
candida parapsilosis) SK26.003, being preserved in Chinese Typical Representative culture collection center, deposit number is CCTCC NO:M 2012491.
2. a method of utilizing the bacterial strain of the product N.F,USP MANNITOL described in claim 1 to produce N.F,USP MANNITOL, it is characterized in that with described Candida parapsilosis (
candida parapsilosis) SK26.003 is starting strain, through seed culture, liquid submerged fermentation, produces N.F,USP MANNITOL, step is:
(1) seed culture
Seed culture medium: glucose 5-20g/L, yeast extract paste 5-20g/L, deionized water preparation;
Seed culture condition: in 30-37 ℃, under the rotating speed of 160-200rpm, shaking table is cultivated 10-24h;
(2) fermentation culture
Fermention medium: glucose 100-500g/L, yeast extract paste 10-40g/L, urea 1-10g/L, magnesium sulfate 0-10g/L, sodium-chlor 0-10g/L, calcium chloride 0-5g/L, potassium primary phosphate 0-10g/L, adjusts pH5-7;
Fermentation condition: inoculum size 1%-10%, rotating speed 160-200rpm, 3-5d ferments under the condition of temperature 30-37 ℃ in fermention medium;
(3) extraction of N.F,USP MANNITOL
By fermented liquid centrifuging, after activated carbon decolorizing, destainer is successively carried out to desalination by cationic exchange coloum 001 * 7 and anionite-exchange resin 313; The calcium type Zeo-karb of take is sorbent material, and 50-70 ℃ with water elution, obtains the N.F,USP MANNITOL of purifying after condensing crystal.
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| CN201310081411.3A CN103131643B (en) | 2013-03-14 | 2013-03-14 | Strain for producing mannitol and method for producing mannitol through fermentation of strain |
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| CN201310081411.3A CN103131643B (en) | 2013-03-14 | 2013-03-14 | Strain for producing mannitol and method for producing mannitol through fermentation of strain |
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| CN103131643B true CN103131643B (en) | 2014-03-26 |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN104651419A (en) * | 2015-03-16 | 2015-05-27 | 南京工业大学 | Method for combined production of mannitol and D-lactic acid by virtue of microorganism anaerobic fermentation |
| CN109321613B (en) * | 2018-09-30 | 2020-11-06 | 江南大学 | Method for producing D-mannose |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1788076A (en) * | 2003-06-19 | 2006-06-14 | 科学与工业研究委员会 | Fungus strain and a method of obtaining mannitol from the same |
| CN102154126A (en) * | 2010-11-19 | 2011-08-17 | 广西大学 | Strain and method for producing mannitol by strain |
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- 2013-03-14 CN CN201310081411.3A patent/CN103131643B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1788076A (en) * | 2003-06-19 | 2006-06-14 | 科学与工业研究委员会 | Fungus strain and a method of obtaining mannitol from the same |
| CN102154126A (en) * | 2010-11-19 | 2011-08-17 | 广西大学 | Strain and method for producing mannitol by strain |
Non-Patent Citations (2)
| Title |
|---|
| 吴国荃 等.我国甘露醇的生产状况与发展趋势.《化工技术经济》.2004,第22卷(第4期), |
| 我国甘露醇的生产状况与发展趋势;吴国荃 等;《化工技术经济》;20040430;第22卷(第4期);4-5 * |
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