CN103409492B - Method for extracting transparent low-acyl gellan gum - Google Patents
Method for extracting transparent low-acyl gellan gum Download PDFInfo
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Abstract
The invention belongs to the technical field of production of microbial polysaccharide gellan gum and discloses a method for extracting transparent low-acyl gellan gum. The method comprises the steps of fermenting, filtering, performing deacylation and flocculation precipitation, flocculating again, squeezing and drying, utilizing wastes. The invention also provides composite bacterial liquid for producing the transparent low-acyl gellan gum. By the extraction method, the yield and the transparency of the low-acyl gellan gum can be increased, waste of raw materials is reduced, pollution is avoided, and wastes are changed into treasures.
Description
Technical field
The present invention relates to microbial technology field, particularly relate to a kind of extracting method of transparent type low-acyl gellan gum.
Background technology
Gelling gum (GellanGum) is a kind of microorganism edible gum of Kelco company of the U.S. exploitation eighties in 20th century.The another novel microorganism exocellular polysaccharide of kelco company exploitation after xanthan gum, its gellifying property is more more superior than xanthan gum.It belongs to (Pseudomonaseloden) in neutral conditions by vacation list brain bacillus waterweed, take glucose as carbon source, in the substratum that ammonium nitrate is made into for nitrogenous source and some inorganic salt, the extracellular polysaccharide colloid produced through aerobic fermentation is a kind of gelifying agent of novel all-transparent.Gelling gum be followed successively by D-Glucose by four glycan molecules, polymer saccharide compound that D-Glucose aldehydic acid, D-Glucose, L-rhamnosyl are formed by connecting by glycosidic link, wherein first glucose molecule connects with β-Isosorbide-5-Nitrae glycosidic link.Mountain has superior gellifying property in gelling gum, it is widely used at field of food, as milk preparation, fruit squash, fruit spreads, pudding jelly and bread filler etc., be also applied in non food area, as the slow releasing, microbiological culture media etc. of toothpaste, medicine.Gelling gum is applied in milk preparation, gelling gum is heated to 70 DEG C ~ 75 DEG C can direct hydration in milk, in acidophilous goods, add this kind of water-sol serve as colloid protective agent, the protein flocculation in milk preparation and the effect of mouthfeel can be eliminated; For in candy, superior structure and quality can be provided to product, and shorten the time of starch jelly colloid formation; Gelling gum joins the consumption that can reduce saturated fatty acid in biscuits, and improves the level of biscuit, makes biscuit have good sedimentation; Gelling gum also alternative pectin prepares jam and jelly, also can be used in cake and fruit pies filler; In the course of processing of meat product and greengrocery, adding gelling gum, can to make up the taste of product not enough, makes it have salubrious taste feature.
Usually gelling gum product is divided into two kinds according to the content of ethanoyl in gelling gum polysaccharide molecule: one is low-acyl gellan gum, by making deacylated tRNA base to high acyl gellan gum or the process of part deacylated tRNA base obtains, after aquation, under cation sites, easily form fragility gel; Another kind is high acyl gellan gum, and namely natural gelling gum, easily forms viscoelastic gel after aquation.Low-acyl gellan gum can be divided into transparent type and nontransparent type according to transparency, and the number primarily of the residual volume of bacterial chip, impurity protein and polysaccharide causes transparency.CN201310134689 discloses the extracting method of the non-clean-type gellan gum of a low acyl group; the method is only for nontransparent type low-acyl gellan gum; and this technique exists a lot of technological deficiency, the gelling gum purity prepared of such as this technique is lower and create a large amount of refuses, causes Environment pollution.How to develop the technical problem that a kind of real prior art of the green extraction process that output is high, transparency is high and industrial energy consumption is low of gelling gum is anxious to be resolved.
Summary of the invention
In order to overcome the deficiencies in the prior art, improve the output of low-acyl gellan gum and transparency, the waste reclaimation in conservation and production technique, the invention provides a kind of extracting method of transparent type low-acyl gellan gum, comprises following preparation process:
The first step, fermentation: (Sphingomonas paucimobilis liquid and bacillus pumilus liquid weight ratio are 8:1, and in two kinds of bacterium liquid, the concentration of thalline is 1 × 10 by mixed bacteria liquid
8individual/mL) cultivate according in the inoculum size access seeding tank of 8% (volume ratio), it is 30 DEG C in temperature, under the condition of pH value 7.2, cultivate and obtain liquid A in 24 hours, then by liquid A: fermentation tank culture medium is that the volume ratio of 1:10 proceeds in fermentor tank and cultivates, temperature 30 DEG C, pH value 7.2, incubation time 72 hours, obtains fermented liquid; Wherein, the component of seed tank culture base is: sucrose 25g, peptone 5g, ammonium sulfate 4.0g, yeast extract paste 3.0g, citric acid 3.5g, calcium carbonate 2.0g, distilled water 1000ml; The component of fermentation tank culture medium is: sucrose 40g, peptone 8g, potassium primary phosphate 3g, citric acid 2.5g, calcium carbonate 2.0g, magnesium sulfate 1g, distilled water 1000ml;
Second step, filter: the mixture simultaneously adding diatomite and aerosil in the fermented liquid that the first step obtains, interpolation limit, limit is stirred, and stir 10min, stirring velocity is 100r/min, then uses Plate Filtration, collects the filtrate after filtering and thalline respectively; The addition of wherein said mixture is 0.5% of fermented liquid weight, and the mass ratio of described diatomite and aerosil is 3:1; The specific surface area of described aerosil is 200m
2/ more than g;
3rd step, deacylated tRNA base and flocculation sediment: in the filtrate that second step obtains, add liquid caustic soda regulate pH to be 10, be warming up to 90 DEG C ~ 95 DEG C, maintain 30 ~ 40 minutes, carry out the process of deacylated tRNA base, be then down to room temperature; Add 70% ~ 90% ethanol of 2 ~ 3 times of volumes subsequently, mix, then through membrane filtration removing waste liquid, obtain the gelling gum that flocculates; And after the ethanol in low-temperature rotary evaporation Recycling of waste liquid, obtain liquid B; Above-mentioned flocculation gelling gum again disperseed and is dissolved in water, obtains gelling gum liquid, then adding the saturated nacl aqueous solution of gelling gum liquid volume 6% ~ 7%, Precipitation gelling gum;
4th step, again flocculate: by the 3rd step separate out gelling gum according to 1g gelling gum: 100ml water ratio again soluble in water, obtain gelling gum liquid, then 80% ethanol accounting for gelling gum liquid 2 times of volumes is added, mix, then through membrane filtration removing waste liquid, obtain the gelling gum that flocculates;
5th step, squeezing is dry: the flocculation gelling gum the 4th step obtained, through screw press, is removed unnecessary moisture, obtained wet product gelling gum; Then by dry for wet product gelling gum, pulverize and obtain transparent type low-acyl gellan gum;
6th step, utilization of waste material: the liquid B that the thalline obtain second step and the 3rd step obtain is mixed to get mixing solutions, and then add Semen Maydis powder, sorghum flour, bean dregs and wheat bran in mixing solutions, interpolation limit, limit is stirred to pasty state; Finally pass into steam and be warming up to 110 DEG C, distill 15 minutes; Then by after distillment oven dry, pulverizing, add zinc sulfate, vitamin-E and chlorogenic acid, mix, obtain pig feed; Wherein, Semen Maydis powder, sorghum flour, bean dregs and wheat bran account for 6%, 5%, 2% and 1% of mixing solutions quality respectively, zinc sulfate, vitamin-E and chlorogenic acid account for respectively mixing solutions quality ten thousand/.
The invention also discloses a kind of composite bacteria liquid producing transparent type low-acyl gellan gum, it is that 8:1 mixes by Sphingomonas paucimobilis liquid and bacillus pumilus liquid according to weight ratio, and in two kinds of bacterium liquid, the concentration of thalline is all about 1 × 10
8individual/mL.
The Sphingomonas paucimobilis that the present invention uses and the bacterial strain that bacillus pumilus is commonly used for this area, (ATCC31461, see J Ind Microbiol Biotechnol.2002Oct for preferred Sphingomonas paucimobilis Sphingomonas paucimobilis; 29 (4): 170-6.) and bacillus pumilus (Bacillus pumilus) (ATCC27142, see Journal of FoodProtection1995, Volume58, Number4).
The beneficial effect that the present invention obtains is mainly as follows:
By test of many times and research, in Sphingomonas paucimobilis fermentation, with the addition of the bacillus pumilus of proper ratio pioneeringly, make bacillus pumilus can effectively degrade Sphingomonas paucimobilis produce all kinds of acidic substance, and under making fermented liquid maintain weak basic condition all the time, keep Sphingomonas paucimobilis ferment effect to maintain higher state, improve the output of gelling gum; The ratio suitably improving bacillus pumilus can improve the transparency of gelling gum, but can have a certain impact to the output of gelling gum;
The Optimal pH that Sphingomonas paucimobilis fermentation produces gelling gum is about 7.2, but multiple acidic substance can be produced during the fermentation, comprise hydroxybutyric acid etc., class acidic substance are easily entrained in gelling gum and are not easy to be precipitated, cause the transparency of gelling gum to decline, and bacillus pumilus is by the collaborative symbiosis with Sphingomonas paucimobilis, facilitates the generation of gelling gum, and reduce the content of all kinds of acidic substance in gelling gum, improve purity and the transparency of gelling gum;
Without the need to adding pH adjusting agent in fermented liquid, decreasing the waste of raw material, having saved cost; Decrease all kinds of chemical substances that in prior art, other extracting method use in leaching process, avoid and chemical substance is brought in gelling gum;
The mode that the present invention adopts the flocculation of repeated multiple times ethanol to purify in postorder treating processes avoids the use of multivalent metal cation, thus avoid gellan gum finished product and mix too much impurity, ensure that purity and the transparency of gelling gum, ethanol used in purification process can recycling, reduces production cost;
In the process extracting gelling gum, tropina, macromolecular polysaccharide etc. are reclaimed, avoids the waste liquid pollution on the environment of this type of material; Meanwhile, obtain nutritious animal feeding-stuff containing somatic protein, turn waste into wealth, increase economic benefit; Achieve the environmental protection of gelling gum leaching process simultaneously, almost arrange outward without waste water, feed liquid component makes full use of.
Embodiment
Below employing specific embodiment is further explained the present invention, but should not regards the restriction to initiative spirit of the present invention as.
Embodiment 1
An extracting method for transparent type low-acyl gellan gum, comprises following preparation process:
The first step, by mixed bacteria liquid, (Sphingomonas paucimobilis liquid (ATCC31461) and bacillus pumilus liquid (ATCC27142) weight ratio are 8:1, and in two kinds of bacterium liquid, the concentration of thalline is 1 × 10
8individual/mL) cultivate according in the inoculum size access seeding tank of 8% (volume ratio), it is 30 DEG C in temperature, under the condition of pH value 7.2, cultivate and obtain liquid A in 24 hours, then by liquid A: fermentation tank culture medium is that the volume ratio of 1:10 proceeds in fermentor tank and cultivates, temperature 30 DEG C, pH value 7.2, incubation time 72 hours, obtains fermented liquid; Wherein, the component of seed tank culture base is: sucrose 25g, peptone 5g, ammonium sulfate 4.0g, yeast extract paste 3.0g, citric acid 3.5g, calcium carbonate 2.0g, distilled water 1000ml; The component of fermentation tank culture medium is: sucrose 40g, peptone 8g, potassium primary phosphate 3g, citric acid 2.5g, calcium carbonate 2.0g, magnesium sulfate 1g, distilled water 1000ml;
Second step, in the fermented liquid that the first step obtains, add the mixture of diatomite and aerosil, interpolation limit, limit is stirred simultaneously, and stir 10min, stirring velocity is 100r/min, then uses Plate Filtration, collects the filtrate after filtering and thalline respectively; The addition of wherein said mixture is 0.5% of fermented liquid weight, and the mass ratio of described diatomite and aerosil is 3:1; The specific surface area of described aerosil is 200m
2/ more than g;
3rd step, adds liquid caustic soda and regulates pH to be 10, be warming up to 90 DEG C ~ 95 DEG C, maintain 30 ~ 40 minutes, carry out the process of deacylated tRNA base, be then down to room temperature and obtain deacylated tRNA based sols in the filtrate that second step obtains; In deacylated tRNA based sols, add 70% ~ 90% ethanol accounting for deacylated tRNA based sols 2 ~ 3 times of volumes subsequently, mix, flocculation gelling gum; Then through membrane filtration removing waste liquid, the gelling gum that flocculates is obtained; And after the ethanol in low-temperature rotary evaporation Recycling of waste liquid, obtain liquid B; Above-mentioned flocculation gelling gum again disperseed and is dissolved in water, obtains gelling gum liquid, then adding the saturated nacl aqueous solution of gelling gum liquid volume 6% ~ 7%, Precipitation gelling gum;
4th step, by the 3rd step separate out gelling gum according to 1g gelling gum: 100ml water ratio again soluble in water, obtain gelling gum liquid, then 80% ethanol accounting for gelling gum liquid 2 times of volumes is added, mix, then through membrane filtration removing waste liquid, obtain the gelling gum that flocculates;
5th step, the flocculation gelling gum the 4th step obtained, through screw press, is removed unnecessary moisture, is obtained wet product gelling gum; Then by dry for wet product gelling gum, pulverize and obtain transparent type low-acyl gellan gum;
6th step, the liquid B that the thalline obtain second step and the 3rd step obtain is mixed to get mixing solutions, and then in mixing solutions, add Semen Maydis powder, sorghum flour, bean dregs and wheat bran, interpolation limit, limit is stirred to pasty state; Finally pass into steam and be warming up to 110 DEG C, distill 15 minutes; Then by after distillment oven dry, pulverizing, add zinc sulfate, vitamin-E and chlorogenic acid, mix, obtain pig feed; Wherein, Semen Maydis powder, sorghum flour, bean dregs and wheat bran account for 6%, 5%, 2% and 1% of mixing solutions quality respectively, zinc sulfate, vitamin-E and chlorogenic acid account for respectively mixing solutions quality ten thousand/.
Embodiment 2
The performance perameter of transparent type low-acyl gellan gum prepared by embodiment 1:
Control group: only adopt Sphingomonas paucimobilis fermentative production gelling gum, other steps are with embodiment 1; Experimental group is gelling gum prepared by embodiment 1.The molecular weight determination content of acyl group (in the determination of acid-basetitration gelling gum): the molecular weight of the low acyl group of control group is 710,000 dalton, and the molecular-weight average of the gelling gum of experimental group is 720,000 dalton.
The mensuration of gel-strength: carry out the mensuration measuring transmittance with TA.TX21 property tester;
The mensuration of transmittance: take 0.5 sample, adding distil water 100ml, beaker is placed in 80 C water bath, the calcium chloride solution 2ml of 2.7% is added after sample dissolution, supplement evaporated water to original volume, while hot by sol solution impouring cuvette, the thermostat container putting into 20 degrees Celsius is immediately placed 15 minutes, measure transmittance with spectrophotometer at 497nm place, contrast with distilled water.Specifically see table 1:
Table 1
The test of embodiment 3 pig feed of the present invention:
After testing, pig feed protein content 35.2% prepared by embodiment 1, polysaccharose substance content 24.1%, inorganic mineral content 2.7%, all the other are starch, Mierocrystalline cellulose and a small amount of trace element etc.
Choose a month large weanling pig 120, be divided into two groups, often organize 60, wherein the diet prepared of experimental group the present invention, every 50kg is 210 yuan, and control group, with honest feed (SSB-25 model), is 300 yuan of calculating according to every 50kg.Raise after 6 weeks and detect indices see table 1.
Table 1
| Index (every piglet) | Control group | Of the present invention group |
| Weanling pig body weight (kg) | 5.12 | 5.08 |
| The body weight (kg) increased for 6 weeks | 13.25 | 14.86 |
| Consume feed (kg) | 17.2 | 17.3 |
| Feed for nursing cost (unit) | 103.2 | 72.7 |
Conclusion: the pig feed cost that the present invention utilizes waste material to prepare is starkly lower than market common feedstuffs, and the increase of body weight is also greater than control group.
Finally, it is also to be noted that what enumerate above is only several specific embodiments of the present invention.Obviously, the invention is not restricted to above embodiment, many distortion can also be had.All distortion that those of ordinary skill in the art can directly derive from content disclosed by the invention or associate, all should think protection scope of the present invention.
Claims (1)
1. an extracting method for transparent type low-acyl gellan gum, is characterized in that, said method comprising the steps of:
The first step, fermentation: mixed bacteria liquid is cultivated according in the inoculum size access seeding tank of 8%, it is 30 DEG C in temperature, under the condition of pH value 7.2, cultivate and obtain liquid A, then by liquid A in 24 hours: fermentation tank culture medium be 1: 10 volume ratio proceed in fermentor tank and cultivate, temperature 30 DEG C, pH value 7.2, incubation time 72 hours, obtains fermented liquid; Wherein, the component of seed tank culture base is: sucrose 25g, peptone 5g, ammonium sulfate 4.0g, yeast extract paste 3.0g, citric acid 3.5g, calcium carbonate 2.0g, distilled water 1000mL; The component of fermentation tank culture medium is: sucrose 40g, peptone 8g, potassium primary phosphate 3g, citric acid 2.5g, calcium carbonate 2.0g, magnesium sulfate 1g, distilled water 1000mL;
Second step, filter: the mixture simultaneously adding diatomite and aerosil in the fermented liquid that the first step obtains, interpolation limit, limit is stirred, and stir 10min, stirring velocity is 100r/min, then uses Plate Filtration, collects the filtrate after filtering and thalline respectively; The addition of wherein said mixture is 0.5% of fermented liquid weight, and the mass ratio of described diatomite and aerosil is 3: 1;
3rd step, deacylated tRNA base and flocculation sediment: in the filtrate that second step obtains, add liquid caustic soda regulate pH to be 10, be warming up to 90 DEG C ~ 95 DEG C, maintain 30 ~ 40 minutes, carry out the process of deacylated tRNA base, be then down to room temperature; Add 70% ~ 90% ethanol of 2 ~ 3 times of volumes subsequently, mix; Then through membrane filtration removing waste liquid, the gelling gum that flocculates is obtained; And after the ethanol in low-temperature rotary evaporation Recycling of waste liquid, obtain liquid B; Above-mentioned flocculation gelling gum again disperseed and is dissolved in water, obtains gelling gum liquid, then adding the saturated nacl aqueous solution accounting for gelling gum liquid volume 6% ~ 7%, Precipitation gelling gum;
4th step, again flocculate: by the 3rd step separate out gelling gum according to 1g gelling gum: 100mL water ratio again soluble in water, obtain gelling gum liquid, then 80% ethanol accounting for gelling gum liquid 2 times of volumes is added, mix, then through membrane filtration removing waste liquid, obtain the gelling gum that flocculates;
5th step, squeezing is dry: the flocculation gelling gum the 4th step obtained, through screw press, is removed unnecessary moisture, obtained wet product gelling gum; Then by dry for wet product gelling gum, pulverize and obtain transparent type low-acyl gellan gum;
6th step, utilization of waste material: the liquid B that the thalline obtain second step and the 3rd step obtain is mixed to get mixing solutions, and then add Semen Maydis powder, sorghum flour, bean dregs and wheat bran in mixing solutions, interpolation limit, limit is stirred to pasty state; Finally pass into steam and be warming up to 110 DEG C, distill 15 minutes; Then by after distillment oven dry, pulverizing, add zinc sulfate, vitamin-E and chlorogenic acid, mix, obtain pig feed; Wherein, Semen Maydis powder, sorghum flour, bean dregs and wheat bran account for 6%, 5%, 2% and 1% of mixing solutions quality respectively, zinc sulfate, vitamin-E and chlorogenic acid account for respectively mixing solutions quality ten thousand/;
Mixed bacteria liquid in the described the first step is to mix at 8: 1 by Sphingomonas paucimobilis liquid and bacillus pumilus liquid according to weight ratio, and in described Sphingomonas paucimobilis liquid or described bacillus pumilus liquid, the concentration of thalline is 1 × 10
8individual/mL;
Described Sphingomonas paucimobilis is Sphingomonas paucimobilis (Sphingomonas paucimobilis) ATCC 31461, and described bacillus pumilus is bacillus pumilus (Bacillus pumilus) ATCC 27142.
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| CN103740624B (en) * | 2014-01-17 | 2015-06-17 | 威海光洋生物科技有限公司 | Bacillus pumilus and application thereof to preparation of exopolysaccharides |
| CN106350554A (en) * | 2016-08-30 | 2017-01-25 | 新疆阜丰生物科技有限公司 | Extraction method of non-transparent type low-acyl gellan gum |
| CN116023520B (en) * | 2023-01-17 | 2023-08-22 | 河北沣川生物科技有限公司 | Preparation method of low-acyl sanzan gum |
| CN116135886A (en) * | 2023-04-18 | 2023-05-19 | 广州市乾相生物科技有限公司 | Extraction method of transparent low-acyl gellan gum |
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| CN101182089A (en) * | 2007-11-15 | 2008-05-21 | 山东阜丰生物科技开发有限公司 | Xanthan gum waste water treatment process |
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| CN101182089A (en) * | 2007-11-15 | 2008-05-21 | 山东阜丰生物科技开发有限公司 | Xanthan gum waste water treatment process |
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