CN1132929C - Enzymatic preparation of high-purity xylo-oligosaccharide - Google Patents
Enzymatic preparation of high-purity xylo-oligosaccharide Download PDFInfo
- Publication number
- CN1132929C CN1132929C CN00109788A CN00109788A CN1132929C CN 1132929 C CN1132929 C CN 1132929C CN 00109788 A CN00109788 A CN 00109788A CN 00109788 A CN00109788 A CN 00109788A CN 1132929 C CN1132929 C CN 1132929C
- Authority
- CN
- China
- Prior art keywords
- xylo
- oligosaccharide
- enzyme
- xylan
- zytase
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
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- HEBKCHPVOIAQTA-NGQZWQHPSA-N d-xylitol Chemical compound OC[C@H](O)C(O)[C@H](O)CO HEBKCHPVOIAQTA-NGQZWQHPSA-N 0.000 title claims abstract description 46
- 238000002360 preparation method Methods 0.000 title abstract description 16
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Images
Classifications
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
- Y02P20/00—Technologies relating to chemical industry
- Y02P20/50—Improvements relating to the production of bulk chemicals
- Y02P20/52—Improvements relating to the production of bulk chemicals using catalysts, e.g. selective catalysts
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- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
The invention relates to a method for preparing xylo-oligosaccharide by enzyme conversion, which is characterized in that alkaline-resistant xylanase producing strain Pseudomonas sp.XY-11 screened and preserved in a laboratory is taken as parent strain, ultraviolet and nitrosoguanidine mutagenesis treatment is carried out according to a conventional method to obtain mutant Pseudomonas XUN024 (namely CGMCC No.0458), and the enzyme activity of fermentation enzyme production reaches 800-; the enzyme preparation is used for enzyme conversion of refined xylan prepared by an alkaline method by taking bran, bagasse or corncobs as raw materials, and the product components of xylobiose and xylotriose account for more than 90% of the total conversion product; the xylo-oligosaccharide in the enzyme conversion solution is decolorized by activated carbon, desalted by 732 and 717 ion exchange resins, ultrafiltered and concentrated in vacuum to prepare xylo-oligosaccharide syrup, or is dried in vacuum to prepare xylo-oligosaccharide powder, and the xylo-oligosaccharide product xylobiose and xylotriose content accounts for more than 90 percent of the total sugar component.
Description
The present invention relates to wheat bran, bagasse or corn cob is raw material, and the refining xylan of alkaline process preparation is made substrate, and the zytase with pseudomonas Pseudomonas sp.XUN024CGMCCNo.0458 fermentative production carries out the method that xylo-oligosaccharide is made in enzymatic conversion.
Xylo-oligosaccharide (Xylooligosaccharide) is a kind of in the functional oligose.So-called oligose claims oligosaccharides (oligosaccharide) again, and it is the polymkeric substance that is made of by glycosidic link 2-10 monose molecule.The research and development and the industrialized development thereof of functional oligose be swift and violent over nearly 20 years, and this series products is widely used in food and the fodder production as positive growth factor for bifidus.Industrialized functional oligose has kind more than ten.The Asia is except that Japan, and Korea S and China comprise the also existing functional oligose variety production in Taiwan Province.European countries are very active to this research, and US enterprise circle is also expressed great interest to oligose in recent years.The U.S. is equipped with functional oligose in the low calories drink of aspartame preparation, improving the mouthfeel of beverage, and increase the cultivation effect of bifidus bacterium.
Xylo-oligosaccharide is with its excellent characteristic: mouthfeel be similar to well sucrose, acid acceptance and Heat stability is good, viscosity low, reduce water-activity, prevent to freeze etc., and help that non-digestibility is processed in foodstuffs industry, whole intestinal function is remarkable, dose is few (everyone 0.7g every day) etc., enable in numerous oligose products, to occupy critical role.Especially to the bifidus bacillus cultivation effect, be better than other functional oligoses.The human body oral test proves, every day intake 0.7g, the bifidus bacterium ratio is increased to 26.2% from taking in after being increased to for 17.9%, three week after preceding 8.5%, two week in the large intestine.In contrast, intake every day that reaches suitable bifidus bacterium propagation required oligomeric isomaltose of ratio or oligofructose is 10-15g, and 7-8g.Therefore its price on present Japanese oligose market is the highest, is 16 times of oligomeric isomaltoses, 6 times of oligofructose.
The xylo-oligosaccharide product of Japanese market has three kinds of specifications: wood oligose 70 (liquid contains xylo-oligosaccharide 70%, other carbohydrates 30%, 2500 yen/kilogram of prices); Wood oligose 35 (powder contains xylo-oligosaccharide 35%, other carbohydrates 65%); Wood oligose 20 (powder contains xylo-oligosaccharide 20%, other carbohydrates 80%).Product is applied to lactic drink as food ingredients, the production of finished product such as chocolate and black vinegar food flavouring.
Method for preparing lower polyxylose has following several:
1. chemical process degraded, as with acid-hydrolysis method (Mitsuish Y.:Agric, Biol.Chem, 1988,52:921-927);
2. physical method degraded comprises vapor pressure osmose process (JP6197800A2), microwave method (JP1224384A2) etc. as hot-water extraction (JP62281890A2);
3. biological degradation method or enzyme process, enzyme process preparation mainly are to utilize the zytase of microorganism system, and the hemicellulose in the enzymolysis of plants raw material is an xylan, and obtaining with xylo-bioses, xylotriose is the xylo-oligosaccharide product of principal constituent.
Physics, chemical process degradation speed are difficult to control; Or process for refining is loaded down with trivial details, and yield is low, is not suitable for suitability for industrialized production.The enzyme process preparation is that research is maximum, also is more practical method.The key of enzyme process preparation is high vigor and has the zytase that reasonable enzyme is.
Early stage zytase research mainly is to be used for feed, papermaking, textile auxiliary agent etc.Study both at home and abroad and report that the microorganism that can produce zytase has aspergillus niger, nipa palm aspergillus, Trichodermareesei, viride, Chaetomium globosum, little purple mould, paecilomyces varioti, Penicillium corylophilum, bacillus acidocldarius, Bacillus licheniformis, pseudomonas etc.
Zytase is a class prozyme system, mainly comprises inscribe-beta-xylanase, end-grain cutting zytase, xylobiase etc.The enzyme of different sources is that component is different.When being used to prepare xylo-oligosaccharide, then wish not contain in the enzymolysis product or contain less monose (wood sugar).Wish to obtain only cutting internally the restriction endonuclease of β-1.4 glycosidic link, obtain enzymolysis product xylo-bioses, xylotriose, and wish not have or reduce from the xylosidase of the end-grain cutting enzyme of end-grain cutting β-1.4 key and hydrolysis xylo-bioses, xylotriose.Early stage zytase research is paid attention to its enzyme and is lived, and wishes thorough enzymolysis, and does not pay attention to enzyme system.The zytase that bacterial strain produced of research at present or report all also produces components such as higher wood sugar, glucose when being used for the enzymolysis xylan.
The Japanese Patent spy who goes into Jiang Lifu etc. opens clear 62-155095 and JP02119790A2, has reported the manufacture method of xylan resolvent and the manufacture method of xylo-bioses.They have made extensive work to the screening that obtains producing based on the zytase of xylo-bioses bacterial strain.Finally screen Chaetomium globosum (Chaetomiumgracil) bacterial strain and produced zytase, (the enzyme work of Chaetomium gracil 1161 solid state fermentations reaches 2 to bacterial strain after the mutagenesis, the ratio that 600U/g substratum, enzyme liquid are used for hydrolyzed xylan (birch) products therefrom xylo-oligosaccharide and wood sugar 34: 66 before the mutagenesis are brought up to 72: 21.This technology is used to the suitability for industrialized production of xylo-oligosaccharide, produces zytase by new Japanese KCC, and Suntory Ltd. produces the xylo-oligosaccharide product.
(JP10215866A2) etc. the zytase produced of usefulness bacillus sp. prepares xylo-oligosaccharide for NOGUCHI NORITAKA, JP06261750A2 about the research of xylo-oligosaccharide such as Noguchi for other.(ARAKI TOSHIYOSHI JP10295372A2) waits the β-1 that produces with Alcaligenes (Alealigenes), 3-zytase to waste wood, β-1,3 key of its hydrolysis water glycan, and do not act on xylo-bioses β-1,4 keys, thereby to obtain product mainly be that wood two is to the xylo-oligosaccharides of wooden six sugar.(Ehzymic production of oligosaccharides from corncob xylan such as France Patrice Pellerin, Enzyme.Microb.Technol., 13:617-621,1991) zytase with clostridium (Clostridium) prepares xylo-oligosaccharide from corn cob.The research of other countries is less.
Domestic 1997 Cai Jing equality (microbiology circular, 24:91-94,1997) has reported that fungi decomposes the research that corn cob is produced xylo-oligosaccharide.Recently also have some investigators to report the research that they prepare xylo-oligosaccharide with mycetogenetic zytase such as aspergillus niger, Trichodermareesei, Penicillium corylophilum from the xylan of corn cob etc. successively.
The purpose of this invention is to provide the method that the simple enzymatic conversion xylan of a kind of technology prepares xylo-oligosaccharide.Xylo-oligosaccharide content reaches more than 90% in the resulting enzymatic conversion product, and monosaccharide components such as wood sugar, glucose are below 10%.
The present invention realizes by following scheme:
(1) the alkali resistance zytase with this laboratory screening and preservation produces bacterium Rhodopseudomonas Pseudomonas sp.XY-11, be the bacterium that sets out, obtain dissociant pseudomonas (Psudomonas sp.) XUNO24-CGMCCNo.0458 through ultraviolet ray and nitrosoguanidine mutagenesis, as bacterium producing multi enzyme preparation;
(2) bacterial strain is through slant activation, and enzymatic production prepares the liquid or solid zymin, as the enzyme source;
(3) be raw material with wheat bran, bagasse or corn cob, alkaline process prepares xylan, and refining back is as the enzymatic conversion substrate;
(4) enzymatic conversion generates xylo-oligosaccharide, through decolouring, desalination, concentrated, makes the xylo-oligosaccharide syrup, or is dried to the xylo-oligosaccharide powder.
Bacterial classification and mutagenesis thereof:
Take a sample from certain paper mill effluent ditch, through constantly separation and purification and screening, obtain a strain and produce zytase bacterium XY-11, this strain enzyme-producing is stable, and enzyme work shake flask fermentation 36h enzyme work on product enzyme basic medium can reach 170U/ml.With this bacterium as starting strain.
Bacterial classification preliminary evaluation result:
This bacterium of electron microscopic observation is a Gram-negative, rod-short, and cell is single, no gemma, atrichia.
On dextrose culture-medium, the slick round bacterium colony of dimpling was milk yellow in the middle of this bacterium formed, butteriness, and the 2 days colony diameters of growing are 1-1.5mm.On the meat soup flat board, this bacterium has circular folds around being, middle nick, and bacterium colony is greyish white slightly yellow, and the 2 days colony diameters of growing are 2-3mm.
The agar stab incubation growth is slow, is in line indiffusion.Draw straight line on the bouillon agar inclined-plane and cultivate, it is thread that lawn is, and do not spread to both sides.The meat soup surface forms the mycoderm of epistasis shape when cultivating, and produces alkali.
This bacterium strictness is aerobic, just can grow in simple organic compound as the inorganic medium of the unique carbon source and the energy, can utilize carbon sources such as acetate, succinate, lactic acid salt and glucose, nitrate can be as nitrogenous source, the growth temperature range of this bacterium is wider, can grow between 4-48 ℃.Below PH4.5C, do not grow well-grown under the condition of PH9.4.Part physio-biochemical characteristics result of study sees Table 1.1.
Preliminary evaluation result according to the XY-11 bacterial strain, contrast " uncle Jie Shi Bacteria Identification handbook and some relevant Bacteria Identification documents, compared wherein G-, bacillar description (particularly to the experiment of atrichia and oxydase is arranged), our preliminary evaluation is Rhodopseudomonas (Pseudomonas sp.XY-11).
The part physio-biochemical characteristics of table 1.1 XY-11
| The utilization of carbon source experiment | Glucose | + |
| Sucrose | + | |
| Lactose | - | |
| Semi-lactosi | + | |
| Maltose | + | |
| Wood sugar | + | |
| Raffinose | + | |
| Pectinose | + | |
| Rhamnosyl | - | |
| Cellobiose | + | |
| Sodium acetate | + | |
| Other biochemical test | Hydrolyzed starch | - |
| The peroxidation oxygenase | - | |
| Urase | - | |
| MR-VP | - | |
| Indoles | - | |
| Litmus milk | Peptonize cow's milk | |
| Gelatin hydrolysis | + | |
| The two hydrolysis poison of arginine | - | |
| Oxydase | - |
With Pseudomonas sp.XY-11 is parental plant, carries out ultraviolet ray (UV) and nitrosoguanidine (NTG) mutagenic treatment according to a conventional method.Ultraviolet irradiation time is 60s (lethality rate is 83%), the bigger some strain bacterium of picking transparent circle from the selectivity flat board, adopt the basis to produce the enzyme substratum and shake the multiple sieve of bottle, wherein produce No. 15 higher bacterial strain called after XUV015 of enzyme, enzyme work than starting strain exceeds about 50% (enzyme 250U/ml alive), adopt NTG to carry out mutagenesis again, be that the dosage (lethality rate is 86%) of 30min carries out mutagenesis action time, the bigger bacterium of the tens of strain transparent circles of picking shakes the multiple sieve of bottle from the selectivity flat board, screen a strain enzyme very high bacterial strain alive No. 24, its enzyme work improves about 42% than XUV015, reach 354U/ml, called after XUN024.This bacterial strain is preserved in China Committee for Culture Collection of Microorganisms common micro-organisms preservation center on June 19th, 2000, and preserving number is CGMCC No.0458.
The cultivation of bacterium producing multi enzyme preparation and fermentation:
Slant culture: preparation contains glucose 1-10%, peptone 0.5-5%, and NaCl 0.5-5%, agar 2-2.5%, all the other are water, the substratum of PH6-9, each components contents is percent weight in volume, and promptly g/100ml is together following.100-120 ℃ of sterilization, 20-50 minute, the XUNO24 bacterial strain that sterilization postcooling, inoculation obtain was above cultivated 10-40 hour for 20-40 ℃.
The product enzyme is cultivated: preparation contains wheat bran 1-6%, (NH
4)
2SO
40.4-1%, K
2HPO
40.2-1%, all the other are water, the substratum of PH6-9, liquid amount 20-100ml/250ml triangular flask, 100-120 ℃ of sterilization, 20-50 minute, sterilization postcooling, inoculation slant culture, inoculum size 5-10%, 20-40 ℃, the rotary bottle cabinet rotating speed 100-220r/m that shakes.XUN024 carries out the shake flask fermentation test under these conditions, and 24-36h produces enzyme and can reach more than the 600U/ml.
Xylanase activity measuring method: reference literature Bailey et al.J.Biotechnol, 1992,23:257-270.After the calmine damping fluid dilution suitable multiple, add 0.5ml dilution enzyme liquid successively, 1ml, 1% normal wood glycan suspension, 50 ℃ of water bath heat preservation 30min, add DNS reagent 1ml immediately, the cooling of boiling water bath 5min ice-water bath is settled to 25ml, surveys OD value (is standard with the wood sugar) in 540nm place colorimetric.Enzyme work is defined as under the above-mentioned condition, and the enzyme amount that per minute discharges the reducing sugar amount (in wood sugar) of 1 μ mol is 1 enzyme unit (U) alive.
When above enzyme activity unit calculates, be that the wood sugar amount that is produced with per minute is calculated, but the product of xylanase hydrolysis xylan is generally xylo-bioses, xylotriose and the xylo-oligosaccharide of high-polymerization degree more, and does not have or wood sugar seldom, therefore calculates with the wood sugar amount and may have certain error.Measure wood sugar, xylo-bioses reducing sugar typical curve respectively, contrasted and the relation between relatively O.D value and micromole measure, find that both differ very little.Explanation is when calculating enzyme activity, and replacing xylo-bioses or xylotriose amount to calculate with the wood sugar amount is reasonably, and owing to all be to calculate enzyme activity unit with the wood sugar amount on the document, also is convenient to contrast.
The xylan preparation:
(hemicellulose that extracts in wheat bran, bagasse or the corn cob raw material with alkaline process is an xylan to reference for J.D.Breccia et al.J.Appl.Bacteriol.1995, method 78:469-472), and refining back is as the enzymatic conversion substrate.
Bagasse powder (cross 40 mesh sieves) is with the NaOH solution dissolving of 0.1-2mol/l, under the pressure of 0-0.2Npa boiling 0.5-3 hour, filters.Filtrate is used ethanol sedimentation, the centrifugal supernatant liquor that inclines of 20000 * g, the precipitation NaClO of 0.1-2mol/l
3The boiling water bath dissolving.In the centrifugal supernatant liquor that inclines of 20000 * g, the precipitation dissolve with ethanol of collection.This operation repeats twice again, and the precipitation that obtains in oven dry 4-36 hour, obtains xylan.
Obtain xylan and should carry out refinement treatment before enzymolysis, purpose is the impurity that may influence the zytase effect in the xylan extracting solution in order to remove, and helps extraction, the purifying of xylo-oligosaccharide product in the enzymolysis solution, can guarantee the quality of product.
Enzymatic conversion reaction:
With the xylan is substrate, and concentration of substrate is 1-2.0%, and enzyme concentration is the 100-500U/g substrate, and temperature of reaction is 30-40 ℃, and enzymolysis time is 10-30 hour.The xylan enzymolysis liquid of gained carries out the thin-layer chromatography stratographic analysis and shows that main component is xylo-bioses and xylotriose in the enzymolysis solution with this understanding.
Xylo-oligosaccharide extracts refining:
The xylo-oligosaccharide enzymolysis solution main component of gained is xylo-bioses and xylotriose behind the enzymolysis, also has unconverted xylan, also contain higher inorganic salt impurity and darker color, must separate, extract it and carry out desalination and decolouring and handle and to be made with extra care, thus the high purity of obtaining, high-quality xylo-oligosaccharide product.
Adopt hyperfiltration process to separate unconverted macromole xylan, adopt gac, 717
#Reinforcing yin essence ion exchange resin, 732
#Strong cation-exchanging resin carries out desalination bleaching to be handled, and like this, can reach refining effect and xylo-oligosaccharide yield preferably.
Xylan enzymolysis fluid component and Determination on content
Adopt HPLC (high pressure liquid chromatography) method, chromatographic instrument: Waters 510 types, chromatographic column: Spherisorb nh 2 column, detector: Waters 2410, temperature: 30 ℃, moving phase: 66.6% (V/V) acetonitrile/water solution, flow rate of mobile phase: 1.0ml/min.
Or employing flash chromatography column method, i.e. pressurized column chromatography technology.Eluent is a propyl carbinol: Glacial acetic acid: water=can satisfy R at 4: 1: 1
fThe requirement of value.Dress column method: get an amount of tlc silica gel (G60) in mortar, add eluent, after suitably grinding, slowly add in the chromatography column.The skim quartz sand is evenly spread on bottom and silica gel surface at post.Sample introduction and collection: with the dry sample of crossing of a spot of eluent dissolving, carefully add in the chromatography column with dropper, add eluent, lower piston is opened in pressurization, collects elutriant with every pipe 5ml.
Analyze and measure: with the elutriant of collecting, every a pipe sampling, point sample with behind the ascending method chromatography, develops the color in chromatography cylinder on the tlc silica gel plate.Separate liquid thin-layer chromatography color atlas relatively with former zytase, each pipe of same component is merged, vacuum concentration promptly obtains the sample of required component.
Fig. 1 is the seed selection pedigree of strain X Y-11; Fig. 2 is a process flow diagram of the present invention;
The present invention has overcome physics, the chemical process degradation speed is difficult to control, or process for refining is loaded down with trivial details, and yield is low, is not suitable for the shortcoming of suitability for industrialized production.Advantage of the present invention is seed selection, and the pseudomonas Pseudomonas sp.XUN024-CGMCCNo.0458 of inscribe-beta-xylanase is lived, mainly comprised to one plant height enzyme, provides the simple enzymatic conversion xylan of a kind of technology to prepare the method for xylo-oligosaccharide.Xylo-oligosaccharide content reaches more than 90% in the resulting enzymatic conversion product, and monosaccharide components such as wood sugar, glucose are below 10%, and obtains highly purified xylo-oligosaccharide product thus.
The following examples elaborate to the present invention.
The preparation of embodiment 1 zytase
Slant culture: substratum is a glucose 1%, peptone 0.5%, NaCl0.5%, agar 2%, PH8.5 sterilized 20 minutes for 110 ℃, the inoculation of sterilization postcooling, bacterial classification is pseudomonas Pseudomonassp.XUN024-CGMCCNo.0458, cultivates 17 hours for 37 ℃, as the slant activation seed.
Seed culture: substratum is a glucose 1%, peptone 0.5%, and PH8.5, liquid amount 20ml/250ml sterilized 20 minutes for 110 ℃, and sterilization postcooling inoculation inclined-plane seed 1 ring was cultivated 17 hours for 37 ℃, as liquid seeds.
The product enzyme is cultivated: substratum is a wheat bran 6%, (NH
4)
2SO
40.4%, K
2HPO
40.2%, pH8.5, liquid amount 20ml/250ml, 110 ℃ of sterilizations, 20 minutes, sterilization postcooling, inoculation seed liquor, 5%, 37 ℃ of inoculum size, the rotary bottle cabinet rotating speed 220r/m that shakes.XUN024 carries out the shake flask fermentation test under these conditions, and it is 825U/ml that 36h produces xylanase activity.
Embodiment 2 wheat bran xylan enzymolysis
The wheat bran pre-treatment: after wheat bran is crossed 60 mesh sieves, be to add water at 4: 1 by the liquid material than V/W, every gram wheat bran adds the high temperature resistant Alpha-starch of 10 units, and 95 ℃ of hydrolysis again with hot wash wheat bran twice, are filtered, and remove starch and a part of soluble protein.
Alkali is carried: get wheat bran after the 100 gram pre-treatment, add NaOH solution, alkali concn is 5%, the liquid material is 5: 1 than (V/W), and temperature is 40 ℃, static immersion, time is 4 hours, and is after alkali is carried, centrifugal, centrifugal back residue is centrifugal twice with distilled water wash, merges supernatant liquor, in 6.0mol/L hydrochloric acid and after, precipitated with the twice industrial spirit, placement is spent the night, and is centrifugal, under this condition, the xylan extraction yield is 33.75%.(dry to constant weight, calculate xylan and extract yield).
The xylan enzymolysis: carry the wheat bran xylan with alkali, the water compound concentration is 2.0% substrate; Add the zytase by the preparation of example 1 method, enzyme concentration is the 350U/g xylan; Temperature of reaction is 37 ℃, and enzymolysis time is 24 hours.The xylan enzymolysis liquid of gained carries out the thin-layer chromatography stratographic analysis and shows that main component is xylo-bioses and xylotriose in the enzymolysis solution with this understanding, does not detect wood sugar, glucose.Analyze the dry thing of gained enzymolysis solution with the flash chromatography post, in its xylo-oligosaccharide component, xylobiose content is 30.45%, and xylotriose content is 36.32%, and the content that xylo-oligosaccharide accounts for total dry thing is 60.94%.
Embodiment 3 bagasse xylan enzymolysis
Press the method for embodiment 2, carry bagasse xylan 100g (dry-matter meter) with alkali, compound concentration is 2.0% xylan aqueous suspensions, adds the zytase by the preparation of example 1 method, presses the method enzymolysis of embodiment 2.The enzymolysis solution of gained carries out ultrafiltration (membrane molecule amount 5000), and filtrate is used activated carbon decolorizing, and 717
#, 732
#The resin desalting treatment, again in 50 ℃ ,-0.09MPa, vacuum concentration obtains xylo-oligosaccharide syrup 34.2g (dry syrup is heavy), is 34.2% to the xylan total recovery.The HPLC analysis revealed, gained xylo-oligosaccharide component: xylo-bioses 44.7%, xylotriose 48.2%, xylo-oligosaccharide content are 92.9%.
Claims (6)
1. the bacterial strain of a high yield zytase, it is pseudomonas (Pseudomonas sp.) XUNO24CGMCC No.0458.
2. method for preparing zytase comprises the described bacterial strain of claim 1 through slant activation, seed culture, ferment zytase liquid.
3. method according to claim 2 is characterized in that described fermentation condition is temperature 20-40 ℃, rotating speed 100-220r/m.
4. method according to claim 2 is characterized in that described fermention medium is: wheat bran 1-6%, (NH
4)
2SO
40.4-1%, K
2HPO
40.2-1%, all the other are water, PH6-9, each components contents is percent weight in volume.
5. make the method for xylo-oligosaccharide, comprising:
(1) be raw material with wheat bran, bagasse or corn cob, use xylan extracted with alkali, refining;
(2) product with (1) is a substrate, adds right and requires the 2 enzyme liquid that obtain, temperature of reaction 30-40 ℃, enzymolysis time 10-30 hour;
(3) adopt hyperfiltration process to separate unconverted macromole xylan, adopt gac, the desalination of 717# reinforcing yin essence ion exchange resin, the ultrafiltration final vacuum concentrates, and make the xylo-oligosaccharide syrup, or drying is made the xylo-oligosaccharide powder.
6. method according to claim 5, wherein (2) described enzyme concentration is a 100-500U/ gram substrate.
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| CN1884569B (en) * | 2006-05-19 | 2010-06-30 | 南京师范大学 | Xylose enzyme method preparing method |
| CN101381753B (en) * | 2008-10-28 | 2012-05-30 | 上海师范大学 | Method for preparing rice husk xylo-oligosaccharides |
| CN103589761B (en) * | 2013-11-08 | 2016-01-20 | 无锡群硕谷唐生物科技有限公司 | A kind of method utilizing wheat bran to prepare xylo-oligosaccharide and the product obtained by the method |
| CN105255965B (en) * | 2015-10-14 | 2023-04-25 | 山东龙力生物科技股份有限公司 | Method for preparing high-purity xylo-oligosaccharide by using cotton seed hulls as raw materials |
| CN106912964A (en) * | 2015-12-25 | 2017-07-04 | 山东龙力生物科技股份有限公司 | Soluble dietary fiber and preparation method thereof |
| CN107044065B (en) * | 2017-04-12 | 2018-06-01 | 潍坊科技学院 | A kind of paper-making pulping process |
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