CN103305490B - Method for producing pectinase in fermentation manner by taking waste hemp degumming liquid as carbon source - Google Patents
Method for producing pectinase in fermentation manner by taking waste hemp degumming liquid as carbon source Download PDFInfo
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- CN103305490B CN103305490B CN201210072909.9A CN201210072909A CN103305490B CN 103305490 B CN103305490 B CN 103305490B CN 201210072909 A CN201210072909 A CN 201210072909A CN 103305490 B CN103305490 B CN 103305490B
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Abstract
The invention discloses a novel method for preparing a pectinase in a fermentation manner by taking a waste hemp degumming liquid as a carbon source. The method comprises the following steps: a, removing the majority of NaOH (sodium hydroxide) and water from the waste hemp degumming liquid by using ultrafiltration equipment; b, regulating a pH value to 4.0 by using HCl (hydrogen chloride) and precipitating, thereby obtaining a lignin; c, inoculating an aspergillus niger and producing a pectinase in the fermentation manner by taking pectin and hemicelluloses as a main carbon source and wheat bran as a main nitrogen source; d, soaking a fermentation material; separating a pectin liquid by using a liquid-slag separator; performing ultrafiltration concentration, DEAE (diethyl-aminoethanol)-cellulose adsorption, elution, concentration, acetone precipitation, ethanol dehydration and freezing drying, thereby obtaining the pectinase with the specific activity of 15000u/g, the optimum pH value of 4.3 and the optimum temperature of 43 DEG C. According to the method, the process is simple; the industrialization is realized easily; the pollution of the waste hemp degumming liquid to environment is reduced; the production cost of the pectinase is lowered. Thus, the economic and social benefits of an enterprise are increased.
Description
Technical field
The present invention relates to a kind of method utilizing hemp degumming waste liquid pectinase by fermentation, belong to field of microbial fermentation.
Background technology
Fiber is one of important weaving crudefiber crop of China with hemp.Dry into hemp phloem after being peeled off in hemp phloem portion, hemp phloem about contains the colloid of about 40%, and colloid comprises pectin, hemicellulose and xylogen, and wherein pectin accounts for more than 70% of colloid.Product after phloem comes unstuck is hemp degummed ramie, and purely can spin and also can become hemp yarn with cotton blending, the latter is made into cloth.Large ramie product have well-pressed smooth, ventilated, anti-mildew is mothproof, the advantage such as antistatic, quick-drying washable, enjoys high reputation in world market, is the superior product of China's weaving outlet, be a large amount of foreign exchange of country's creation every year.In numerous Bast Fiber Textiles, hemp textile is the product that the European Community and the U.S. do not limit.
Coming unstuck is the first working procedure that hemp is processed, general employing biological-chemical combination degumming method.First use degumming bacterium liquid (containing polygalacturonase) to soak hemp phloem at normal temperatures, the polygalacturonase of degumming bacterium secretion is by most of pectin (biological degumming); Hemicellulose and xylogen remove (chemical Degumming) by NaOH solution (6%) boiling.The waste water that biological degumming and chemical Degumming produce merges into degumming waste water, and waste water is containing NaOH, pectin, hemicellulose, xylogen and pigment etc.According to the analysis to part hemp degumming enterprise wastewater, waste water NaOH concentration is about 4.5-5.0% (w/v), COD
crabout 10.6-12.8g/L, BOD
5about 5.5-6.8g/L, SS are about 1.02-1.15g/L.Medium scale hemp degumming source mill discharges degumming waste water and be about 30m every day
3, wastewater to reach standard process needs about 600 yuan every day, and annual needs direct pollution treatment expense about 200,000 yuan, add facility investment and operational and administrative expenses, pollution treatment costly.
Polygalacturonase is one of large zymin in the world four, is the general name of the multiple enzyme of decompose pectin, is divided into Pectin depolymerase and Rohapect MPE (PE).Ester bond in PE hydrolysis of pectin macromolecule side chain; Pectin depolymerase comprises again polygalacturonase (PG) and pectin lyase (PL) two subclass, PG is by hydrolyzing alpha-1,4 glycosidic links work and macromole hydrolyzed pectin are become small molecules, PL cuts off the α-1 in pectin molecule by trans-elimination, pectin molecule is degraded to small molecules by 4 glycosidic links, industrial use mostly be PG.
The bacterium that can produce polygalacturonase has multiple, and many moulds, a small amount of bacterium and yeast can secrete polygalacturonase.Aspergillus niger is conventional polygalacturonase zymophyte, belongs to fungi, has various mutations kind, and product is PG.Because the pectinase activity of fermentation of Aspergillus niger production polygalacturonase technical maturity, unit raw material production is high, security is high, therefore both at home and abroad substantially all using aspergillus niger as pectin enzyme-producing bacteria.
The main raw material of fermentation of Aspergillus niger is wheat bran under normal circumstances, and crack rice on a small quantity and other auxiliary materials, wheat bran mainly provides nitrogenous source and carbon source, crack rice supplementary carbon source and partial nitrogen source, other auxiliary materials supplementing as trace element.Fermentation of Aspergillus niger is solid fermentation, about the 30 DEG C about 3-4 days that ferment.With physiological saline soaking fermentation material, centrifugation goes out liquid pectin, obtains polygalacturonase after ultrafiltration and concentration through DEAE-cellulose adsorption, wash-out, concentrated, acetone precipitation, ethanol dehydration, lyophilize.
Summary of the invention
Hemp degumming waste liquid contains NaOH up to 4.5-5.0%, the yeasting of aspergillus niger is acid, therefore NaOH will be removed, if not only increase cost with in HCl with NaOH, and generate a large amount of NaCl, impact is come unstuck the addition of waste liquid, addition exceed a certain amount of will osmotic pressure too high, black-koji mould can not grow.Useless come unglued liquid contains xylogen, and xylogen can not be decomposed by aspergillus niger.Xylogen has been widely used, and as the raw material preparing resol, can add HCl and adjust pH to 4.0 precipitation, centrifugal acquisition xylogen.The present invention is intended to utilize the hemp degumming waste liquid (containing the composition such as pectin and hemicellulose) removing most of NaOH and xylogen to replace starch, and other techniques and ordinary method are substantially identical.The present invention not only solves the pollution problem of hemp degumming waste liquid to environment, again reduces the production cost of polygalacturonase.Technique effect of the present invention is embodied in several aspect:
1, reduce the pollution of hemp degumming waste liquid, environmental benefit is obvious
First hemp degumming waste liquid is removed most of NaOH and water by ultrafiltration apparatus, add new solid NaOH and recycle (for chemical Degumming); Xylogen wherein obtains by adjusting pH to 4.0 to precipitate, and xylogen has been widely used, can as the raw material preparing resol; Last remaining pectin and hemicellulose are as the carbon source of fermentation of Aspergillus niger.Therefore, useless come unglued liquid all utilizes by patent of the present invention substantially, and environmental benefit is obvious.
2, decrease the pollution treatment expense of enterprise, improve the economic benefit of enterprise
Degumming waste water 30m is discharged every day according to general hemp degumming enterprise
3calculate, wastewater to reach standard process about needs 600 yuan every day, annual direct requirement pollution treatment expense about 200,000 yuan, facility investment probably needs 2,000,000 yuan, annual operational management expense about 100,000 yuan, comprehensive depreciation of fixed assets, direct expenses and operation management expense, the expense every year in wastewater treatment about 500,000 yuan.Although also there is effluent part to need process in polygalacturonase production process, only need about 150,000 yuan every year, the present invention can reduce the pollution treatment expense about 350,000 yuan of enterprise every year.
3, reduce the production cost of polygalacturonase, enhance the competitive power of product.
The present invention utilizes the hemp degumming waste liquid (containing the composition such as pectin and hemicellulose) removing most of NaOH and xylogen to replace starch, wheat bran provides nitrogenous source, add Secondary ammonium phosphate simultaneously and supplement nitrogenous source and phosphoric, add ferrous sulfate and supplement ferro element.Reduce the production cost of polygalacturonase than ordinary method with useless degumming water replacement part material, produce the fermentation raw material pricing of 6.8kg polygalacturonase with 1 ton of fermentation material, the cost compare of two kinds of methods sees the following form, and proferment cost reduces very remarkable.
The polygalacturonase raw materials cost analysis (producing 6.8kg polygalacturonase for 1 ton of fermentation material, unit: unit) that two kinds of methods are produced
Fermentation material conservation cost 253 yuan per ton as can be seen from the above table, hemp factory discharges degumming waste water about 9000 tons every year, can prepare fermentation material 6000 tons, can manufacture polygalacturonase 40.8 tons, and the whole year can conservation cost 151.1 ten thousand yuan.
Embodiment
Remove the useless come unglued liquid of most of NaOH and xylogen containing pectin 12.5-13.0%; Hemicellulose 6.2-6.4%; Xylogen 1.1-1.2%; Other solids components 1.9-2.1%; Water 74.6-75.8%.
Embodiment one:
The first step: the NaOH in useless come unglued liquid is removed in ultrafiltration
Get 1500kg to give up come unglued liquid, the ultra-filtration membrane selecting Hangzhou Ou Kai membrane technique company limited to produce concentrates demineralization plant (DK8048 type membrane module, molecular weight cut-off 150-300 dalton) ultrafiltration.Ultrafiltration goes out solution 900kg (accounting for 60% of cumulative volume, containing NaOH, water and other small molecules), recycles, concentrated solution 600kg (accounting for 40% of cumulative volume) after adding solid NaOH as chemical Degumming liquid.
Second step: the extraction of xylogen
Add industrial HCl (concentration about 21%) under agitation to pH 4.0, static 24h precipitate lignin, xylogen (rotary drum rotating speed 6000rpm is isolated by liquid-solid continuous centrifuge, output pressure 200Kpa), the useless stoste of coming unstuck of liquid (containing pectin and hemicellulose) (not doing ultrafiltration and delignification process, containing about 4.5%NaOH) after removing xylogen is adjusted pH to 5.6.Record scrap rubber liquid containing pectin 12.5%, hemicellulose 6.2%, xylogen 1.1%, other solids components 1.9%, water 78.3%.
3rd step: inoculation and fermentation
To be given up come unglued liquid 600kg, wheat bran 395kg, (NH
4)
2hPO
44kg, FeSO
4.7H
2o 1kg, stir after sterilizing.Compound is cooled to room temperature, sprays aspergillus niger spore and mix (about 10
6individual spore/gram), 30 ± 2 DEG C of fermentations in the fermenting house of sterilizing, the activity of certain intervals sampling and measuring polygalacturonase, reaches polygalacturonase maximum activity (84 hours) and terminates fermentation, pectinase activity 1900U/g butt matter.
4th step: the extraction of polygalacturonase, purifying and determination of activity
With deionized water soaking fermentation material, solid-liquid ratio is 1: 4 for the first time, and lixiviate was separated (rotary drum rotating speed 6000rpm, output pressure 200Kpa) by liquid-slag separating machine after 30 minutes; Second time solid-liquid ratio is 1: 3, and lixiviate was separated (rotary drum rotating speed 6000rpm, output pressure 200Kpa) by liquid-slag separating machine after 30 minutes; Slag is mainly wood chip, can reuse.Remove most of water and salt by ultrafiltration and concentration, concentrate again after adding 0.05M NaAC-HAC damping fluid.Process domestic DEAE-Mierocrystalline cellulose (Shanghai Heng Xin chemical reagent company limited) to specifications, add DEAE-Mierocrystalline cellulose in concentrated solution to adsorb, with 0.05M NaAC-HAC (containing 0.2M NaCl) wash-out foreign protein, with 0.05M NaAC-HAC (containing 0.3M NaCl) wash-out polygalacturonase.Most of water and salt (DK8048 type membrane module is removed by ultrafiltration and concentration, molecular weight cut-off 150-300 dalton), be concentrated into 1/4 of original volume, add the acetone concentration to 66% of precooling, precipitate (rotary drum rotating speed 7000rpm centrifugal after 4 hours, output pressure 250Kpa) obtain polygalacturonase precipitation, dehydrated alcohol dewaters, lyophilize (-20 DEG C).Fermentation material per ton produces polygalacturonase 6.83kg, and yield is 38%.
The polygalacturonase feature of preparation is as follows: outward appearance is buff powder; The enzyme activity measured according to national standard (QB1502-92) is 15000u/g; Optimal pH is 4.3; Optimum temperuture is 43 DEG C.
Embodiment two:
The first step: the NaOH in useless come unglued liquid is removed in ultrafiltration
Get 1500kg to give up come unglued liquid, other operations are with embodiment one.
Second step: the extraction of xylogen
Operation is with embodiment one.Scrap rubber liquid is recorded containing pectin 13.0%, hemicellulose 6.4%, xylogen 1.2%, other solids components 2.1%, water 77.3% after removing most of NaOH and xylogen.
3rd step: inoculation and fermentation
To be given up come unglued liquid 600kg, wheat bran 395kg, (NH
4)
2hPO
44kg, FeSO
4.7H
2o 1kg, stir after sterilizing.Compound is cooled to room temperature, sprays aspergillus niger spore and mix (about 10
6individual spore/gram), 30 ± 2 DEG C of fermentations in the fermenting house of sterilizing, the activity of certain intervals sampling and measuring polygalacturonase, reaches polygalacturonase maximum activity (84 hours) and terminates fermentation, pectinase activity 1960U/g butt matter.
4th step: the extraction of polygalacturonase, purifying and determination of activity
Operation is with embodiment one.Fermentation material per ton produces polygalacturonase 6.54kg, and yield is 37.5%.
The polygalacturonase feature of preparation is basic identical with embodiment 1, and outward appearance is buff powder; Enzyme activity is 15200u/g; Optimal pH is 4.3; Optimum temperuture is 43 DEG C.This process stabilizing is described, can not changes because component of effluent has certain change.
Claims (1)
1. utilize hemp degumming waste liquid to prepare a method for polygalacturonase as carbon source through fermentation, it is characterized in that comprising the following steps:
A, hemp degumming waste liquid is removed most of NaOH and water by molecular weight cut-off 150-300 dalton ultrafiltration apparatus;
B, adjust pH to 4.0 precipitate lignin, centrifugal acquisition xylogen with HCl, liquid phase, in liquid phase, is adjusted pH to 5.6 by pectin and hemicellulose;
C, using above-mentioned solution as primary carbon source, using wheat bran as major nitrogen source, add Secondary ammonium phosphate and ferrous sulfate, ferment after mixing, sterilizing, inoculated aspergillus niger spore;
D, with deionized water soaking fermentation material, liquid pectin isolated by liquid-slag separating machine, and ultrafiltration and concentration removes most of water and salt, DEAE-cellulose adsorption, wash-out, concentrated, acetone precipitation, ethanol dehydration, lyophilize.
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| CN201210072909.9A CN103305490B (en) | 2012-03-12 | 2012-03-12 | Method for producing pectinase in fermentation manner by taking waste hemp degumming liquid as carbon source |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN105176838B (en) * | 2015-09-09 | 2018-02-09 | 青岛农业大学 | One plant of Aspergillus niger strain and fermenting agent and its application |
| CN105369633A (en) * | 2015-11-18 | 2016-03-02 | 德清县伊得利丝绸有限公司 | Cotton and linen bleaching and dyeing deoxygenization finishing agent |
| CN105369634A (en) * | 2015-11-18 | 2016-03-02 | 德清县伊得利丝绸有限公司 | Cotton and linen interwoven fabric bleaching dyeing deoxygenization process |
Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1271774A (en) * | 2000-06-01 | 2000-11-01 | 中国科学院生态环境研究中心 | Process for preparing pectinase by solid-state fermentation of fruit or vegetable dregs |
| CN1343771A (en) * | 2001-09-19 | 2002-04-10 | 中国科学院生态环境研究中心 | Process for preparing pectinase from waste dregs containing pection after liquid fermentation of large mucor |
| CN1398967A (en) * | 2002-09-06 | 2003-02-26 | 中国科学院生态环境研究中心 | Solid fermentation of waste residue and waste water to produce pectase |
| CN101338333A (en) * | 2008-08-12 | 2009-01-07 | 南京林业大学 | Method for conversing agricultural and forest residues to be fermentable sugars |
| CN101624727A (en) * | 2009-08-06 | 2010-01-13 | 华中科技大学 | Manufacturing method of ramie based on sectioned circulation |
| CN101787571A (en) * | 2009-12-31 | 2010-07-28 | 李耀亭 | Resource utilization process for clean hemp biological degumming and waste thereof |
| WO2010107944A1 (en) * | 2009-03-17 | 2010-09-23 | Alltech, Inc. | Compositions and methods for conversion of lignocellulosic material to fermentable sugars and products produced therefrom |
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2012
- 2012-03-12 CN CN201210072909.9A patent/CN103305490B/en not_active Expired - Fee Related
Patent Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1271774A (en) * | 2000-06-01 | 2000-11-01 | 中国科学院生态环境研究中心 | Process for preparing pectinase by solid-state fermentation of fruit or vegetable dregs |
| CN1343771A (en) * | 2001-09-19 | 2002-04-10 | 中国科学院生态环境研究中心 | Process for preparing pectinase from waste dregs containing pection after liquid fermentation of large mucor |
| CN1398967A (en) * | 2002-09-06 | 2003-02-26 | 中国科学院生态环境研究中心 | Solid fermentation of waste residue and waste water to produce pectase |
| CN101338333A (en) * | 2008-08-12 | 2009-01-07 | 南京林业大学 | Method for conversing agricultural and forest residues to be fermentable sugars |
| WO2010107944A1 (en) * | 2009-03-17 | 2010-09-23 | Alltech, Inc. | Compositions and methods for conversion of lignocellulosic material to fermentable sugars and products produced therefrom |
| CN101624727A (en) * | 2009-08-06 | 2010-01-13 | 华中科技大学 | Manufacturing method of ramie based on sectioned circulation |
| CN101787571A (en) * | 2009-12-31 | 2010-07-28 | 李耀亭 | Resource utilization process for clean hemp biological degumming and waste thereof |
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