CN102206689B - Method for modifying bacterial cellulose in the fermentation process - Google Patents
Method for modifying bacterial cellulose in the fermentation process Download PDFInfo
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- CN102206689B CN102206689B CN 201110063655 CN201110063655A CN102206689B CN 102206689 B CN102206689 B CN 102206689B CN 201110063655 CN201110063655 CN 201110063655 CN 201110063655 A CN201110063655 A CN 201110063655A CN 102206689 B CN102206689 B CN 102206689B
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Abstract
The invention belongs to the field of microbial fermentation, and relates to a method for modifying bacterial cellulose in the fermentation process. The method comprises the following steps of: activating strains for fermentation production of the bacterial cellulose and preparing seed liquid; adding the seed liquid into a fermentation medium added with hydroxypropylmethylcellulose and carboxymethyl cellulose sodium, and standing for fermentation; and collecting a bacterial cellulose film generated on the surface of the medium, performing alkali washing, and drying until the water content is less than 5 percent. The method is simple and practicable; aftertreatment is not needed, the net structure of the bacterial cellulose can be changed directly in the bacterial cellulose fermentation process, and the hydrogen bond action in molecules and between molecules is reduced; moreover, used reagents are low in cost, and special equipment is not needed; and the prepared bacterial cellulose product has high rehydration and the application range of the bacterial cellulose product is widened.
Description
Technical field
The invention belongs to the microbial fermentation field, relate to a kind of method of during the fermentation modified bacteria cellulose, and the bacteria cellulose that utilizes this method to prepare.
Background technology
Bacteria cellulose (Bacterial cellulose, BC) refer under different condition, by synthetic cellulosic general designation such as certain microorganism in acetic acid Pseudomonas (Acetobacter), Agrobacterium (Agrobacterium), rhizobium (Rhizobium) and the Sarcina (Sarcina) etc.Compare with natural cellulose, it has the advantageous properties such as high purity, high-crystallinity, high-polymerization degree, high retentiveness, has been widely used in every field.
Existing bacteria cellulose all is manufactures in the liquid medium within, and the bacterial cellulose gel water content of membrane that obtains is more than 98%, and water-activity is high, microorganism very easily grows breeding, and storage tolerance not is simultaneously because moisture content is high, bulky, transport and carry inconvenience, trucking costs is expensive.Now usually by compression or the dry water content that reduces gel-film, reduce volume and weight, convenient transportation.But compression or dry remove the translucent bacteria cellulose desciccator diaphragm that bacteria cellulose behind the moisture can become a very thin and hard and compact, because exist a large amount of hydroxyls can form in the very strong molecule and intermolecular hydrogen bonding in the bacteria cellulose, the effect of this hydrogen bond is so that the reconstitution rate of this desciccator diaphragm can only reach 3-5 times of bacteria cellulose deadweight, so after the drying again the gel-film that obtains of rehydration with respect to former gel-film, state and character have had very large change, some good characteristics are also destroyed, and volume reduces greatly, cost increases, and its application is restricted.
2009 disclosed Chinese patent application on December 2, (CN101591448A) a kind of preparation method of bacterial cellulose membrane with high rehydration is disclosed, the step of its preparation is: (1) will remove the bacteria cellulose aquagel film alkalinisation treatment of bacterium and substratum; (2) film after will processing carries out etherificate and processes; (3) the etherificate film is carried out neutralization reaction and process, obtain the bacteria cellulose film of hydroxypropylation; (4) bacteria cellulose film of hydroxypropylation is dry, namely get bacterial cellulose membrane with high rehydration.This invention utilizes bacteria cellulose surface hydroxypropylation modification to destroy part hydrogen bond in the Mierocrystalline cellulose, finds that by the test of reconstitution rate dried hydroxypropylation bacteria cellulose film possesses very high rehydration and swelling ability again.But this method complex disposal process, the agents useful for same cost is higher, be unfavorable for large-scale industrial production, and the etherifying reagent that uses has pungency and toxicity, in treating processes, can damage to some extent workman's health, and in the finished product, be easy to residually, made product can't be applied to health care of food product industry.
Summary of the invention
The objective of the invention is to provide for the deficiencies in the prior art a kind of method of during the fermentation modified bacteria cellulose, directly in the bacteria cellulose fermenting process, change its reticulated structure, reduce in the molecule and intermolecular hydrogen bond action, and the agents useful for same cost is low, need not specific installation, do not need post-processed, the bacteria cellulose product for preparing has good rehydration.
The technical scheme that technical solution problem of the present invention adopts is: a kind of method of during the fermentation modified bacteria cellulose may further comprise the steps:
(1) activation bacteria cellulose fermentative production bacterial strain, preparation fermentative production seed liquor;
(2) 8～12% inoculum size joins seed liquor in the fermention medium of the Carbon and nitrogen sources that contains the capacity that supplies strain growth by volume; Add HPMC and Xylo-Mucine in the described fermention medium;
(3) 25～35 ℃ of lower standing for fermentation 8～20 days;
(4) collect the bacteria cellulose film that media surface produces, alkali cleaning is dried to water-content and is lower than 5%.
Method of the present invention, the bacteria cellulose fermentative production in the wherein said step (1) is preferably acetobacter xylinum or gluconate pyracetobacillus with bacterial strain.
Method of the present invention, the preparation method of seed liquor will activate inoculation to seed culture medium in the wherein said step (1), and 25～35 ℃, 150～220rpm are cultivated 20～48h; Contain the Carbon and nitrogen sources for the capacity of strain growth in the described seed culture medium.Described carbon source is preferably N.F,USP MANNITOL, sorbyl alcohol and/or sucrose.
Method of the present invention, the weight percent content of HPMC and Xylo-Mucine is respectively 0.1～10%, 0.1～8% in the fermention medium in the wherein said step (2).Preferably be added with the HPMC of weight percent 0.5～5% and 0.5～5% Xylo-Mucine.Most preferably be added with the HPMC of weight percent 1% and 1% Xylo-Mucine.The carbon source of described fermention medium is the various glucides that bacterial classification can metabolism utilizes, preferred glucose sugar, fructose, sucrose or Sucus Cocois; Based on the consideration of cost, be more preferably with 10～90% Sucus Cocois as carbon source.
Can also be added with tensio-active agent in the fermention medium in the method for the present invention, wherein said step (2), such as tween 80, glycerin fatty acid ester etc., addition is less than or equal to 0.1%.Can also add as required the inorganic nitrogen-sourced of easy utilization, such as urea etc., promote the metabolism of bacterial strain to produce film.Rehydration performance to bacteria cellulose after test is found to add also has certain improvement.
The present invention is simple, do not need post-processed, can directly in the bacteria cellulose fermenting process, change its reticulated structure, reduce in its molecule and intermolecular hydrogen bond action, and the agents useful for same cost is lower, need not specific installation, the bacteria cellulose product for preparing has good rehydration, has enlarged the scope of its application.
Embodiment
Below in conjunction with embodiment, the specific embodiment of the present invention is described in further detail.Following examples are used for explanation the present invention, but are not used for limiting the scope of the invention.
The raw material that adopts in following examples is as follows:
HPMC (HPMC): substituting group is that methoxy substitution 23% and propoxyl replace 10%;
Na-CMC (Xylo-Mucine): substituting group is that carboxymethoxyl replaces 65%;
Tween 80: HLB 15;
Glycerin fatty acid ester: HLB 3.6～4.0;
Urea (purity 〉=99.5%)
Embodiment one
A kind of method of during the fermentation modified bacteria cellulose may further comprise the steps:
(1) activation bacteria cellulose fermentative production bacterial strain gluconate pyracetobacillus, preparation fermentative production seed liquor: the seed liquid nutrient medium contains N.F,USP MANNITOL 25g/l, yeast extract 5g/l, peptone 3g/l, dress 500ml liquid nutrient medium in per 1 liter of shaking flask 180 turns, the lower 24h of cultivation of 30 degree gets seed liquor;
(2) 10% inoculum size adds seed liquor in the fermention medium by volume, and described fermention medium contains: acetic acid: 0.2%, and sucrose: 10%, HPMC:1%, Na-CMC:1%, all the other are Sucus Cocois;
(3) 30 ℃, leave standstill in the 250ml culture plate and cultivated 14 days;
(4) collect the mycoderm that media surface forms, in 0.5N NaOH, boil 10min, at room temperature soak 24h among the 0.5N NaOH, extremely neutral with rinsed with deionized water, measuring its weight in wet base is Wa, and then high speed (3000rpm) is centrifugal 30 minutes, and the supernatant liquor quality measurement is Wb, be weighed as Wc after the throw out lyophilize, the water holding capacity of the bacteria cellulose product that then can obtain producing is:
Water holding capacity (%)=[(Wa-Wc)-Wb]/Wc * 100%
Again the bacteria cellulose after the lyophilize (Wc) is immersed in the deionized water (W/V=1: 2), rehydration 10min～7h, until the quality of rehydration sample (Wd) place also no longer increased in 7 hours till.Reconstitution rate is calculated as follows:
Reconstitution rate (%)=(Wd-Wc)/(Wa-Wb) * 100%
The bacteria cellulose product water holding capacity that this example obtains is 10000%, and reconstitution rate is 35%.
Embodiment two
A kind of method of during the fermentation modified bacteria cellulose may further comprise the steps:
(1) activation bacteria cellulose fermentative production bacterial strain gluconate pyracetobacillus, preparation fermentative production seed liquor: the seed liquid nutrient medium contains N.F,USP MANNITOL 25g/l, yeast extract 5g/l, peptone 3g/l, dress 500ml liquid nutrient medium in per 1 liter of shaking flask 180 turns, the lower 24h of cultivation of 30 degree gets seed liquor;
(2) 8% inoculum size adds seed liquor in the fermention medium by volume, and described fermention medium contains acetic acid: 0.2%, and glucose: 10%, HPMC:5%, Na-CMC:1%, all the other are Sucus Cocois;
(3) 30 ℃, leave standstill in the 250ml culture plate and cultivated 16 days;
(4) collect the mycoderm that media surface forms, in 0.5N NaOH, boil 10min, at room temperature soak 24h among the 0.5N NaOH, extremely neutral with rinsed with deionized water.Measuring its water holding capacity by method among the embodiment one is 9000%, and reconstitution rate is 25%.
Embodiment three
A kind of method of during the fermentation modified bacteria cellulose may further comprise the steps:
(1) activation bacteria cellulose fermentative production bacterial strain gluconate pyracetobacillus, preparation fermentative production seed liquor: the seed liquid nutrient medium contains N.F,USP MANNITOL 25g/l, yeast extract 5g/l, peptone 3g/l, dress 500ml liquid nutrient medium in per 1 liter of shaking flask 180 turns, the lower 24h of cultivation of 30 degree gets seed liquor;
(2) 12% inoculum size adds seed liquor in the fermention medium by volume, and described fermention medium contains acetic acid: 0.2%, and sucrose: the heavy % of 5%, fructose: 5%, HPMC:1, Na-CMC:5%, all the other are Sucus Cocois;
(3) 30 ℃, leave standstill in the 250ml culture plate and cultivated 12 days;
(4) collect the mycoderm that media surface forms, boil 10min in 0.5N NaOH, at room temperature soak 24h among the 0.5N NaOH, to neutral, measuring its water holding capacity by method among the embodiment one is 8500% with rinsed with deionized water, and reconstitution rate is 22%.
Embodiment four
A kind of method of during the fermentation modified bacteria cellulose may further comprise the steps:
(1) activation bacteria cellulose fermentative production bacterial strain gluconate pyracetobacillus, preparation fermentative production seed liquor: the seed liquid nutrient medium contains N.F,USP MANNITOL 25g/l, yeast extract 5g/l, peptone 3g/l, dress 500ml liquid nutrient medium in per 1 liter of shaking flask 180 turns, the lower 24h of cultivation of 30 degree gets seed liquor;
(2) 10% inoculum size adds seed liquor in the fermention medium by volume, and described fermention medium contains acetic acid: 0.2%, and sucrose: 10%, HPMC:0.1%, Na-CMC:8%, all the other are Sucus Cocois;
(3) 30 ℃, leave standstill in the 250ml culture plate and cultivated 18 days;
(4) collect the mycoderm that media surface forms, in 0.5N NaOH, boil 10min, at room temperature soak 24h among the 0.5N NaOH, extremely neutral with rinsed with deionized water.Measuring its water holding capacity according to method among the embodiment one is 8000%, and reconstitution rate is 20%.
Embodiment five
A kind of method of during the fermentation modified bacteria cellulose may further comprise the steps:
(1) activation bacteria cellulose fermentative production bacterial strain gluconate pyracetobacillus, preparation fermentative production seed liquor: the seed liquid nutrient medium contains N.F,USP MANNITOL 25g/l, yeast extract 5g/l, peptone 3g/l, dress 500ml liquid nutrient medium in per 1 liter of shaking flask 180 turns, the lower 24h of cultivation of 30 degree gets seed liquor;
(2) 10% inoculum size adds seed liquor in the fermention medium by volume, and described fermention medium contains acetic acid: 0.2%, and sucrose: 10%, HPMC:10%, Na-CMC:0.1%, all the other are Sucus Cocois;
(3) 30 ℃, leave standstill in the 250ml culture plate and cultivated 18 days;
(4) collect the mycoderm that media surface forms, in 0.5N NaOH, boil 10min, at room temperature soak 24h among the 0.5N NaOH, extremely neutral with rinsed with deionized water.Measuring its water holding capacity according to method among the embodiment one is 7500%, and reconstitution rate is 21%.
Embodiment six
A kind of method of during the fermentation modified bacteria cellulose may further comprise the steps:
(1) activation bacteria cellulose fermentative production bacterial strain gluconate pyracetobacillus, preparation fermentative production seed liquor: the seed liquid nutrient medium contains N.F,USP MANNITOL 25g/l, yeast extract 5g/l, peptone 3g/l, dress 500ml liquid nutrient medium in per 1 liter of shaking flask 180 turns, the lower 24h of cultivation of 30 degree gets seed liquor;
(2) 10% inoculum size adds seed liquor in the fermention medium by volume, and described fermention medium contains acetic acid: 0.2%, and sucrose: 10%, HPMC:1%, Na-CMC:1%, tween 80: 0.1 % by weight, urea: 0.05%, all the other are Sucus Cocois;
(3) 30 ℃, leave standstill in the 250ml culture plate and cultivated 14 days;
(4) collect the mycoderm that media surface forms, in 0.5N NaOH, boil 10min, at room temperature soak 24h among the 0.5N NaOH, extremely neutral with rinsed with deionized water.Measuring its water holding capacity with method among the embodiment 1 is 11000%, and reconstitution rate is 38%.
Embodiment seven
A kind of method of during the fermentation modified bacteria cellulose may further comprise the steps:
(1) activation bacteria cellulose fermentative production bacterial strain gluconate pyracetobacillus, preparation fermentative production seed liquor: the seed liquid nutrient medium contains N.F,USP MANNITOL 25g/l, yeast extract 5g/l, peptone 3g/l, dress 500ml liquid nutrient medium in per 1 liter of shaking flask 180 turns, the lower 24h of cultivation of 30 degree gets seed liquor;
(2) 10% inoculum size adds seed liquor in the fermention medium by volume, and described fermention medium contains acetic acid: 0.2%, and sucrose: 10%, HPMC:1%, Na-CMC:1%, glycerin fatty acid ester: 0.1%, urea: 0.1%, all the other are Sucus Cocois;
(3) 30 ℃, leave standstill in the 250ml culture plate and cultivated 14 days;
(4) collect the mycoderm that media surface forms, in 0.5N NaOH, boil 10min, at room temperature soak 24h among the 0.5N NaOH, extremely neutral with rinsed with deionized water.Measuring its water holding capacity with method among the embodiment one is 11500%, and reconstitution rate is 40%.
Embodiment eight
A kind of method of during the fermentation modified bacteria cellulose may further comprise the steps:
(1) activation bacteria cellulose fermentative production bacterial strain gluconate pyracetobacillus, preparation fermentative production seed liquor: the seed liquid nutrient medium contains N.F,USP MANNITOL 25g/l, yeast extract 5g/l, peptone 3g/l, dress 500ml liquid nutrient medium in per 1 liter of shaking flask 180 turns, the lower 24h of cultivation of 30 degree gets seed liquor;
(2) 10% inoculum size adds seed liquor in the fermention medium by volume, and described fermention medium contains acetic acid: 0.2%, and glucose: 10%, HPMC:2%, Na-CMC:2%, tween 80: 0.1%, urea: 0.1%, all the other are Sucus Cocois;
(3) 30 ℃, leave standstill in the 250ml culture plate and cultivated 14 days;
(4) collect the mycoderm that media surface forms, in 0.5N NaOH, boil 10min, at room temperature soak 24h among the 0.5N NaOH, extremely neutral with rinsed with deionized water.Measuring its water holding capacity with method among the embodiment one is 10500%, and reconstitution rate is 33%.
The per-cent that relates among the application all refers to weight percent as not specifying.
The above only is preferred implementation of the present invention; should be pointed out that for those skilled in the art, under the prerequisite that does not break away from the technology of the present invention principle; can also make some improvements and modifications, these improvements and modifications also should be considered as protection scope of the present invention.
Claims (8)
1. the method for modified bacteria cellulose during the fermentation is characterized in that, may further comprise the steps:
(1) activation bacteria cellulose fermentative production bacterial strain, preparation fermentative production seed liquor;
(2) 8～12% inoculum size joins seed liquor in the fermention medium of the Carbon and nitrogen sources that contains the capacity that supplies strain growth by volume; Add HPMC and Xylo-Mucine in the described fermention medium, the weight percent content of HPMC and Xylo-Mucine is respectively 0.5～5%, 0.5～5% in the described fermention medium;
(3) 25～35 ℃ of lower standing for fermentation 8～20 days;
(4) collect the bacteria cellulose film that media surface produces, alkali cleaning is dried to water-content and is lower than 5%.
2. method according to claim 1, it is characterized in that: described bacteria cellulose fermentative production bacterial strain is acetobacter xylinum or gluconate pyracetobacillus.
3. method according to claim 1 is characterized in that: the preparation method of described seed liquor be for will activate inoculation to seed culture medium, and 25～35 ℃, 150～220rpm are cultivated 20～48h; Contain the Carbon and nitrogen sources for the capacity of strain growth in the described seed culture medium.
4. method according to claim 3 is characterized in that: the carbon source in the described seed culture medium is a kind of in N.F,USP MANNITOL, sorbyl alcohol or the sucrose or mixes.
5. method according to claim 1, it is characterized in that: the weight percent content of HPMC and Xylo-Mucine is respectively 1%, 1% in the described fermention medium.
6. method according to claim 1 is characterized in that: the carbon source of described fermention medium is a kind of in glucose sugar, fructose, sucrose or the Sucus Cocois or mixes.
7. each described method according to claim 1～6, it is characterized in that: be added with tensio-active agent in the described fermention medium, addition is less than or equal to 0.1%.
8. the modified bacteria cellulose for preparing of each described method of claim 1～7.
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| DK2836581T3 (en) | 2012-04-13 | 2023-05-15 | Cp Kelco Us Inc | HIGHLY EFFICIENT AND PRACTICAL FORM OF MICROFIBER CELLULOSE |
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| CN103566785B (en) * | 2012-08-01 | 2017-07-04 | 天津科技大学 | A kind of method that utilization oxidizing bacteria cellulose prepares Pickering emulsions |
| CN105017566B (en) * | 2014-04-28 | 2018-06-12 | 海南光宇生物科技有限公司 | A kind of HPMC composite membranes containing biology cellulose and Zinc oxide nanoparticle |
| CN105316373A (en) * | 2014-08-03 | 2016-02-10 | 浙江理工大学 | Method of utilizing bacterial fermentation to prepare bacterial cellulose with three-dimensional grid structure |
| CN105586376B (en) * | 2014-10-21 | 2020-07-21 | 财团法人食品工业发展研究所 | Method for preparing bacterial cellulose film by using surfactant |
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