CN100510094C - Production method of konjak mannose using cellulase - Google Patents
Production method of konjak mannose using cellulase Download PDFInfo
- Publication number
- CN100510094C CN100510094C CNB2006100191729A CN200610019172A CN100510094C CN 100510094 C CN100510094 C CN 100510094C CN B2006100191729 A CNB2006100191729 A CN B2006100191729A CN 200610019172 A CN200610019172 A CN 200610019172A CN 100510094 C CN100510094 C CN 100510094C
- Authority
- CN
- China
- Prior art keywords
- cellulase
- enzymolysis
- enzyme
- mannooligo saccharide
- konjak
- 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.)
- Active
Links
- 229920002752 Konjac Polymers 0.000 title claims abstract description 31
- 108010059892 Cellulase Proteins 0.000 title claims abstract description 22
- 229940106157 cellulase Drugs 0.000 title claims abstract description 22
- 238000004519 manufacturing process Methods 0.000 title claims abstract description 13
- 235000010485 konjac Nutrition 0.000 title claims abstract description 11
- WQZGKKKJIJFFOK-QTVWNMPRSA-N D-mannopyranose Chemical compound OC[C@H]1OC(O)[C@@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-QTVWNMPRSA-N 0.000 title claims description 5
- 108090000790 Enzymes Proteins 0.000 claims abstract description 25
- 102000004190 Enzymes Human genes 0.000 claims abstract description 25
- 229940088598 enzyme Drugs 0.000 claims abstract description 25
- 238000000034 method Methods 0.000 claims abstract description 21
- 238000001914 filtration Methods 0.000 claims abstract description 9
- 230000008569 process Effects 0.000 claims abstract description 9
- 239000012528 membrane Substances 0.000 claims abstract description 6
- 150000001720 carbohydrates Chemical class 0.000 claims description 29
- 239000000203 mixture Substances 0.000 claims description 15
- 239000000843 powder Substances 0.000 claims description 13
- 238000000108 ultra-filtration Methods 0.000 claims description 10
- 229920002488 Hemicellulose Polymers 0.000 claims description 7
- 239000011259 mixed solution Substances 0.000 claims description 7
- 239000002893 slag Substances 0.000 claims description 3
- 238000001035 drying Methods 0.000 abstract description 11
- 108010002430 hemicellulase Proteins 0.000 abstract description 7
- 229940059442 hemicellulase Drugs 0.000 abstract description 6
- 239000001913 cellulose Substances 0.000 abstract description 2
- 229920002678 cellulose Polymers 0.000 abstract description 2
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 27
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 18
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 16
- 229920002581 Glucomannan Polymers 0.000 description 10
- 238000006243 chemical reaction Methods 0.000 description 10
- 229940046240 glucomannan Drugs 0.000 description 10
- 229920001542 oligosaccharide Polymers 0.000 description 10
- 150000002482 oligosaccharides Chemical class 0.000 description 10
- 239000000047 product Substances 0.000 description 9
- 239000000243 solution Substances 0.000 description 9
- 238000010828 elution Methods 0.000 description 8
- LUEWUZLMQUOBSB-FSKGGBMCSA-N (2s,3s,4s,5s,6r)-2-[(2r,3s,4r,5r,6s)-6-[(2r,3s,4r,5s,6s)-4,5-dihydroxy-2-(hydroxymethyl)-6-[(2r,4r,5s,6r)-4,5,6-trihydroxy-2-(hydroxymethyl)oxan-3-yl]oxyoxan-3-yl]oxy-4,5-dihydroxy-2-(hydroxymethyl)oxan-3-yl]oxy-6-(hydroxymethyl)oxane-3,4,5-triol Chemical compound O[C@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@@H](CO)O[C@@H](O[C@@H]2[C@H](O[C@@H](OC3[C@H](O[C@@H](O)[C@@H](O)[C@H]3O)CO)[C@@H](O)[C@H]2O)CO)[C@H](O)[C@H]1O LUEWUZLMQUOBSB-FSKGGBMCSA-N 0.000 description 7
- 238000005516 engineering process Methods 0.000 description 7
- 238000004809 thin layer chromatography Methods 0.000 description 7
- 230000008878 coupling Effects 0.000 description 6
- 238000010168 coupling process Methods 0.000 description 6
- 238000005859 coupling reaction Methods 0.000 description 6
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 6
- 241001465754 Metazoa Species 0.000 description 5
- 238000004440 column chromatography Methods 0.000 description 5
- 239000000706 filtrate Substances 0.000 description 5
- 239000006188 syrup Substances 0.000 description 5
- 235000020357 syrup Nutrition 0.000 description 5
- 241001312219 Amorphophallus konjac Species 0.000 description 4
- 235000001206 Amorphophallus rivieri Nutrition 0.000 description 4
- 238000004587 chromatography analysis Methods 0.000 description 4
- 230000000694 effects Effects 0.000 description 4
- 235000013305 food Nutrition 0.000 description 4
- 239000000252 konjac Substances 0.000 description 4
- 150000003254 radicals Chemical class 0.000 description 4
- 238000000926 separation method Methods 0.000 description 4
- 238000003756 stirring Methods 0.000 description 4
- 239000000126 substance Substances 0.000 description 4
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 3
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 3
- LPQOADBMXVRBNX-UHFFFAOYSA-N ac1ldcw0 Chemical compound Cl.C1CN(C)CCN1C1=C(F)C=C2C(=O)C(C(O)=O)=CN3CCSC1=C32 LPQOADBMXVRBNX-UHFFFAOYSA-N 0.000 description 3
- 235000008504 concentrate Nutrition 0.000 description 3
- 239000012141 concentrate Substances 0.000 description 3
- 150000004676 glycans Chemical class 0.000 description 3
- 230000003053 immunization Effects 0.000 description 3
- 150000002500 ions Chemical class 0.000 description 3
- 150000003272 mannan oligosaccharides Chemical class 0.000 description 3
- 238000005374 membrane filtration Methods 0.000 description 3
- 244000005700 microbiome Species 0.000 description 3
- 210000000496 pancreas Anatomy 0.000 description 3
- 229920001282 polysaccharide Polymers 0.000 description 3
- 239000005017 polysaccharide Substances 0.000 description 3
- 238000005507 spraying Methods 0.000 description 3
- 238000005406 washing Methods 0.000 description 3
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- 229920002907 Guar gum Polymers 0.000 description 2
- 244000275012 Sesbania cannabina Species 0.000 description 2
- 239000000654 additive Substances 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- 238000009835 boiling Methods 0.000 description 2
- HVYWMOMLDIMFJA-DPAQBDIFSA-N cholesterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)CCCC(C)C)[C@@]1(C)CC2 HVYWMOMLDIMFJA-DPAQBDIFSA-N 0.000 description 2
- 150000001875 compounds Chemical class 0.000 description 2
- 230000009849 deactivation Effects 0.000 description 2
- 238000011161 development Methods 0.000 description 2
- 238000004821 distillation Methods 0.000 description 2
- 239000012153 distilled water Substances 0.000 description 2
- 238000009826 distribution Methods 0.000 description 2
- 239000003814 drug Substances 0.000 description 2
- 239000000665 guar gum Substances 0.000 description 2
- 229960002154 guar gum Drugs 0.000 description 2
- 235000010417 guar gum Nutrition 0.000 description 2
- 230000007062 hydrolysis Effects 0.000 description 2
- 238000006460 hydrolysis reaction Methods 0.000 description 2
- 230000001900 immune effect Effects 0.000 description 2
- 239000007788 liquid Substances 0.000 description 2
- 238000012423 maintenance Methods 0.000 description 2
- 239000012452 mother liquor Substances 0.000 description 2
- 230000007935 neutral effect Effects 0.000 description 2
- OQUKIQWCVTZJAF-UHFFFAOYSA-N phenol;sulfuric acid Chemical compound OS(O)(=O)=O.OC1=CC=CC=C1 OQUKIQWCVTZJAF-UHFFFAOYSA-N 0.000 description 2
- 239000011148 porous material Substances 0.000 description 2
- 239000002244 precipitate Substances 0.000 description 2
- 238000001556 precipitation Methods 0.000 description 2
- 238000002360 preparation method Methods 0.000 description 2
- 238000011137 process chromatography Methods 0.000 description 2
- 238000000746 purification Methods 0.000 description 2
- 230000009467 reduction Effects 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 239000006228 supernatant Substances 0.000 description 2
- 240000004904 Amorphophallus paeoniifolius Species 0.000 description 1
- 241000984278 Bacillus sp. M10 Species 0.000 description 1
- 102100032487 Beta-mannosidase Human genes 0.000 description 1
- 208000024172 Cardiovascular disease Diseases 0.000 description 1
- 235000007516 Chrysanthemum Nutrition 0.000 description 1
- 244000189548 Chrysanthemum x morifolium Species 0.000 description 1
- 206010010774 Constipation Diseases 0.000 description 1
- 244000241257 Cucumis melo Species 0.000 description 1
- 235000015510 Cucumis melo subsp melo Nutrition 0.000 description 1
- 229930091371 Fructose Natural products 0.000 description 1
- RFSUNEUAIZKAJO-ARQDHWQXSA-N Fructose Chemical compound OC[C@H]1O[C@](O)(CO)[C@@H](O)[C@@H]1O RFSUNEUAIZKAJO-ARQDHWQXSA-N 0.000 description 1
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 1
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 1
- 208000001953 Hypotension Diseases 0.000 description 1
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 1
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 description 1
- FJJCIZWZNKZHII-UHFFFAOYSA-N [4,6-bis(cyanoamino)-1,3,5-triazin-2-yl]cyanamide Chemical compound N#CNC1=NC(NC#N)=NC(NC#N)=N1 FJJCIZWZNKZHII-UHFFFAOYSA-N 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 230000003213 activating effect Effects 0.000 description 1
- 230000000996 additive effect Effects 0.000 description 1
- 238000013459 approach Methods 0.000 description 1
- 238000009360 aquaculture Methods 0.000 description 1
- 244000144974 aquaculture Species 0.000 description 1
- 244000052616 bacterial pathogen Species 0.000 description 1
- 108010055059 beta-Mannosidase Proteins 0.000 description 1
- 238000010364 biochemical engineering Methods 0.000 description 1
- 108010059485 brain synaptic membrane glycoprotein gp 50 Proteins 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 208000026106 cerebrovascular disease Diseases 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 235000012000 cholesterol Nutrition 0.000 description 1
- 235000009508 confectionery Nutrition 0.000 description 1
- 230000007797 corrosion Effects 0.000 description 1
- 238000005260 corrosion Methods 0.000 description 1
- 230000006378 damage Effects 0.000 description 1
- 206010012601 diabetes mellitus Diseases 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 230000002526 effect on cardiovascular system Effects 0.000 description 1
- 230000002708 enhancing effect Effects 0.000 description 1
- 238000003912 environmental pollution Methods 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000000855 fermentation Methods 0.000 description 1
- 230000004151 fermentation Effects 0.000 description 1
- 229960002737 fructose Drugs 0.000 description 1
- 210000001035 gastrointestinal tract Anatomy 0.000 description 1
- 125000003147 glycosyl group Chemical group 0.000 description 1
- 239000001963 growth medium Substances 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 230000002218 hypoglycaemic effect Effects 0.000 description 1
- 208000021822 hypotensive Diseases 0.000 description 1
- 230000001077 hypotensive effect Effects 0.000 description 1
- 230000036039 immunity Effects 0.000 description 1
- 230000007365 immunoregulation Effects 0.000 description 1
- 238000011081 inoculation Methods 0.000 description 1
- 238000004255 ion exchange chromatography Methods 0.000 description 1
- 244000144972 livestock Species 0.000 description 1
- 230000004630 mental health Effects 0.000 description 1
- 238000010327 methods by industry Methods 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 229930014626 natural product Natural products 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 235000015097 nutrients Nutrition 0.000 description 1
- 238000005457 optimization Methods 0.000 description 1
- 244000144977 poultry Species 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- 230000001681 protective effect Effects 0.000 description 1
- 230000035484 reaction time Effects 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- 239000000741 silica gel Substances 0.000 description 1
- 229910002027 silica gel Inorganic materials 0.000 description 1
- 229940083608 sodium hydroxide Drugs 0.000 description 1
- 239000002689 soil Substances 0.000 description 1
- ZSJLQEPLLKMAKR-GKHCUFPYSA-N streptozocin Chemical compound O=NN(C)C(=O)N[C@H]1[C@@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O ZSJLQEPLLKMAKR-GKHCUFPYSA-N 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 235000011149 sulphuric acid Nutrition 0.000 description 1
- 230000004584 weight gain Effects 0.000 description 1
- 235000019786 weight gain Nutrition 0.000 description 1
Images
Landscapes
- Polysaccharides And Polysaccharide Derivatives (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
The present invention relates to a production method for preparing konjak mannoligose by using cellulase. Its technological process includes the following several procedures: enzymolysis, filtraction, decoloring, concentration and drying. It is characterized by that said invention adopts non-specific commercial enzyme (including cellulose and hemicellulase), and utilizes centrifugal filtering and membrane filtering combined process, so that said invention can obtain high-purity konjak mannoligose. Besides, said invention also provides the concrete requirements and steps of the above-mentioned every procedure.
Description
Technical field
The present invention relates to microorganism enzymolysis engineering, belong to functional oligosaccharide enzymolysis process production technology.
Technical background
The general name of oligosaccharides (oligose) the little polymkeric substance that to be 2-10 monose be formed by connecting by the glycosidic link of certain position.Mannooligo saccharide (or claiming oligo-glucomannan) is the zymolyte of mannosans (konjac glucomanna), extensively is present in konjaku, guar gum, sesbania gum and the multiple microorganism wall.Mannooligo saccharide has the absorption enteric pathogenic bacteria as a kind of novel little ecologic effect additive and brings into play the dual-use function of immunoregulation effect, is one of most promising microbiotic substitute.Simultaneously, this product all has purposes quite widely in industries such as food medicine.
1, be used for fodder additives, requirement was more than 200,000 tons in domestic year.
Studies show that in a large number mannooligo saccharide can be widely used in various livestock and poultry, aquaculture.Can activate and improve the immunizing power of animal, the quantity discharged of nitrogen prevents to pollute in the incidence of reduction gastrointestinal tract disease and animal dead rate raising day weight gain and feed conversion rate, the reduction ight soil, improves the growing environment of animal.According to relevant departments' statistics, 2004, average addition 0.1% reckoning in the perfect compound feed was pressed in 9,500 ten thousand tons of China's perfect compound feed ultimate productions, 1,500 ten thousand tons of concentrated fodders, 5,000,000 tons of Preblendes, and the demand of mannooligo saccharide is more than 200,000 tons.
2, huge at (health care) food, pharmaceutically application potential, have vast market prospect.
Mannooligo saccharide can significantly improve the human body micro-ecological environment, strengthens the human immunological competence, promotes people's physical and mental health.According to related data, prove that through clinical trial that mannooligo saccharide has is significantly hypotensive, hypoglycemic, cholesterol regulating, prevent cardiovascular and cerebrovascular diseases, and effects such as anti-curing oncoma, relief of constipation, simultaneously, also can be used for fat-reducing.Has huge development potentiality at medicine and protective foods application facet.
At present, domestic production and development kanjak mannan oligosaccharides are mainly realized by polysaccharide hydrolysis chemical method or biologic enzymolysis method.Hydrolysis chemistry law part requires harsh; equipment corrosion is serious; environmental pollution is outstanding; and the biological enzymolysis rule can overcome these shortcomings well; be ideal approach comparatively, but be that the technical study of feedstock production mannooligo saccharide is domestic at the early-stage with the konjaku that the major technique of employing all is with degrade mannosans in the konjaku of mannase; its cost height, efficient are low, can not reach the economy and the technical requirements of scale preparation kanjak mannan oligosaccharides.
Pertinent literature:
Relate to a kind of neutral beta-mannase enzyme liberating konjaku powder production Portugal sweet dew melon and fruit sugar technology, by inoculation Bacillus sp.M10 seed liquor in culture medium selected, produce enzyme incident bottom fermentation in optimization and obtain crude enzyme liquid, again this crude enzyme liquid is used for the enzymolysis konjaku powder, obtains the oligo-glucomannan syrupy product.
Document 2, " preparation, the chemical structure of konjaku Portugal village dew oligosaccharides reach the influence to pancreas islet NO free radical burst size " quick Lv Xiu chrysanthemum of Chen Xiu etc., biochemical engineering National Key Laboratory of Chinese Academy Of Sciences Process Engineering Research Institute, Chinese biological chemistry and molecular biosciences journal .2002,18 (6)
Konjaku powder is after the 'beta '-mannase enzymolysis becomes oligosaccharides, carry out separation and purification with activated carbon column, with different concns (5,10,20) ethanol elution, study different elution fractions and streptozotocin (STZ) is induced the influence of the pancreas islet NO free radical burst size of diabetes model, find ling/ml] can make NO free radical burst size in the pancreas islet nutrient solution 25.4 (P＜=0.05) that on average descend with the oligosaccharides of 5 ethanol elutions, 0.1mg/ml makes NO free radical level 20 (P＜=0.05) that descend with the oligosaccharides of 5 ethanol elutions.
And people such as Zhang Yingqing have proposed to utilize cellulase to prepare the technical scheme of Konjac Glucomannan.
Document 3, " cellulase prepares Konjac Glucomannan "/Zhang Yingqing do letter Xie Bijun, food science and technology institute of Hua Zhong Agriculture University natural product chemistry research department, Hubei Polytichnic College's bioengineering dept, University Of Jishou's journal: natural science edition .2003,24 (3).
Utilize cellulase (viride, Ec3.2.1.4) enzymolysis Rhizoma amorphophalli glucomannan, 10g konjaku powder, the 500U cellulase, at 40 ℃, the pH value is that 5.0 o'clock reaction 2h are optimum condition, gained oligose number average relative molecular weight is 5100 under this condition, and the weight average relative molecular weight is 7160.
Its weak point is: the oligosaccharides molecule number that makes with these processing condition is greatly about 28, and molecular chain is excessive, is unfavorable for activating to greatest extent and improves the immunizing power of animal and improve the human body micro-ecological environment, strengthens the human immunological competence.
Summary of the invention
The object of the present invention is to provide a kind of production method of improved konjak mannose using cellulase, improve enzymolysis efficiency, shorten the mannooligo saccharide molecular chain, reduce its molecule number, improve the immunizing power that mannooligo saccharide activated and improved animal, with the ability of improving human body micro-ecological environment, enhancing body immunity, be that technology is simple, cost is low, the new technology of production high purity mannooligo saccharide.
Technical scheme of the present invention is: a kind of production method of konjak mannose using cellulase, its technical process is: enzymolysis, filtration, decolouring, concentrated, dry, it is characterized in that: press konjaku powder 1g: 80-120u enzyme unit alive drops into cellulase, hemicellulose enzyme mixture, and the enzyme ratio of living is 2.5-3.5: 1 in cellulase, the hemicellulose enzyme mixture; Enzymolysis adopts centrifuging, membrane filtration after stopping successively, obtains mannooligo saccharide mixed solution and filter residue; Thick slag is removed in centrifuging, and ultrafiltration membrance filter obtains the mannooligo saccharide mixed solution, removes macromole; The aperture of used filtering membrane is 2-10 microns.
Concrete production method of the present invention is:
Enzymolysis: in the enzymolysis device, in 100-140g: the 1000ml ratio adds konjaku powder and water, press konjaku powder 1g:80-120u enzyme unit alive and drop into cellulase, hemicellulose enzyme mixture, the enzyme ratio of living is 2.5-3.5: 1 in cellulase, the hemicellulose enzyme mixture, the pH value of control enzymolysis is 4.5-5.5,45 ℃ ± 2 ℃ of temperature, enzymolysis reaction after 5-7 hours reaction times;
Filter: enzymolysis adopts centrifuging, membrane filtration after stopping successively, obtains mannooligo saccharide mixed solution and filter residue; Thick slag is removed in centrifuging, and ultrafiltration membrance filter obtains the mannooligo saccharide mixed solution, removes macromole; The aperture of used filtering membrane is 2-10 microns;
Decolouring: the mannooligo saccharide mixed solution, adopt the column chromatography for separation coupling, the elutriant activated carbon decolorizing;
Concentrate: vacuum concentration, 60 ℃ of controlled temperature, vacuum tightness-0.8mPa obtains the mannooligo saccharide syrup;
Dry: as to adopt spraying drying, 75-80 ℃ of drying temperatures.
The present invention has the following advantages:
1, adopted non-specific commercial enzyme (cellulase and hemicellulase), cost is low, and the efficient height is fit to industrialization;
2, take the filter method of centrifuging, membrane filtration, optimize ultra-filtration membrane separation coupling purification technique and obtain mannooligo saccharide, purity height, the kanjak mannan oligosaccharides that obtains mainly are straight chain or the branched chain that 2-5 monose are formed by connecting by glycosidic link, and its molecular weight is 300-2000;
3, enzymolysis process is simple, and the yield height reaches 77.7%.
Description of drawings
Fig. 1 is the molecular structure of 6 kinds of products in the embodiments of the invention
Fig. 2 is thin layer chromatography analytical results figure.
Fig. 3, Fig. 4 are ion chromatography analysis figure as a result.
Concrete embodiment
One, Technology
(1) main raw:
Konjaku powder, cellulase, hemicellulase, sodium hydroxide (NaOH), industrial alcohol.
(2) main laboratory apparatus:
Water bath with thermostatic control, electric mixer, whizzer, ultrafiltration membrance filter device, activated carbon chromatography post, rotation concentration evaporator, spray-drier, silica gel vacuum drier.
(3) technical process:
Enzymolysis, filtration, decolouring, concentrated, dry.
(4) know-why of project:
The general name of the oligosaccharides little polymkeric substance that to be 2-10 monose be formed by connecting by the glycosidic link of certain position.Mannooligo saccharide is the zymolyte of mannosans (konjac glucomanna), extensively is present in konjaku, guar gum, sesbania gum and the multiple microorganism wall.The main component of konjaku is a Rhizoma amorphophalli glucomannan, and molecular formula is (C
6H
10O
5) n, the averagemolecular wt amount is between 20-200 ten thousand, be a kind of by D-glucose and D-seminose by the mol ratio of 2:3 or 1:1.6 by β-1,4 and the natural polysaccharide that is formed by connecting of β-1, glycosidic links such as 3, content is generally 44-65%, and the content of wherein white konjaku, elephant-foot yam kind can reach 50-65%.Rhizoma amorphophalli glucomannan and cellulosic structure are similar, all be to connect by β-1,4 glycosidic links, utilize cellulase and hemicellulase to carry out enzymolysis, be intended to utilize it to β-1,4 and the destruction of β-1, glycosidic links such as 3, Rhizoma amorphophalli glucomannan is cut into many short chains, be degraded to oligose.Why selected cellulase and hemicellulase blended biological enzyme can work to Rhizoma amorphophalli glucomannan, be because the substrate of Glycosylase (polysaccharide that β-1,4 glycosidic links connect) specificity, comprise the specificity of glycosyl one side, the α of glycosidic link-, β-different specificity.
Two, experimental technique:
1) enzymolysis:
In 1000 ml distilled waters, add 125 gram konjaku powders, stir, regulate its temperature to 45 ℃, and with NaOH (sodium hydroxide) the solution adjusting potential of hydrogen of 0.5M (volumetric molar concentration) to pH5.Add total enzyme live cellulase and hemicellulase (the enzyme ratio of living the is 3:1) mixture of 12500u (international unit), stirring reaction is 7 hours under 45 ℃, the condition of pH5.In the reaction process, regulate maintenance pH5 with the NaOH solution of 0.5M.After reaction finishes, boiling water bath deactivation in 15 minutes, standing over night under the room temperature.
2) separate the coupling purifying:
With the product of last step, under 3600 rev/mins of conditions, removed precipitation in centrifugal 10 minutes.Can be used as feed after this drying precipitate, also can drop into enzymolysis once more.Centrifuge mother liquor is the ultrafiltration membrance filter by 2-10 micron pore size further, returns enzymolysis step next time after the filter residue and drying, and filtrate is separated the mixture solution that coupling obtains the mannooligo saccharide behind the purifying by the activated carbon column chromatography method.Before carrying out column chromatography, with 8 liters of distillation washing posts, use the linear tonsure wash-out of ethanol of 40L0-30% (W/W) then, elution speed is 500 milliliters/hour, collects and numbers and preserve with automatic distribution collector.Oligosaccharide content in the separated product and composition use phenol-sulfuric acid process and TLC method to detect respectively.According to the detected result merging filtrate.
3) concentrate, decolouring, drying
The mannooligo saccharide mixture solution that previous step is rapid is used activated carbon decolorizing, and the supernatant liquor after the filtration in temperature is below 60 ℃, carry out vacuum concentration under the condition of vacuum tightness-0.8mPa, obtains the mannooligo saccharide syrup.Syrup obtains 97.2g mannooligo saccharide powder-like product, yield 77.7% through spraying drying.In this way, the mannooligo saccharide that obtains has six kinds of compositions, is followed successively by according to the elution order of activated carbon column: I (A), II (B), III (C), IV (D), V (E), VI (F).(see figure 1)
Embodiment 2
1) enzymolysis:
In 1000 ml distilled waters, add 110g konjaku powder, 15 gram centrifugation filter residues, 3 gram ultra-filtration membranes separation filter residues, stir, add cellulase and hemicellulose enzyme mixture (cellulose enzyme activity: hemicellulase enzyme work=3:1, total enzyme 12500u alive), fully mixing, regulate its temperature to 50 ℃, and regulate pH to 5 with sodium hydroxide (NaOH) solution of 0.5M (volumetric molar concentration), stirring reaction is 7 hours under 50 ℃, the condition of pH5, enzymolysis reaction.In reaction process, regulate maintenance pH5 with the NaOH solution of 0.5M.Behind the reaction terminating, boiling water bath 15min deactivation, standing over night under the room temperature.
2) separate the coupling purifying:
With the product of last step, under 3600 rev/mins of conditions, removed precipitation in centrifugal 10 minutes.Can be used as feed after this drying precipitate, drop into enzymolysis after also can handling in proportion once more.Centrifuge mother liquor is the ultrafiltration membrance filter by 2-10 micron pore size further, returns enzymolysis step next time after the filter residue and drying, and filtrate is separated the mixture solution that coupling obtains the mannooligo saccharide behind the purifying by the activated carbon column chromatography method.Before carrying out column chromatography, with 8 liters of distillation washing posts, use the linear tonsure wash-out of ethanol of 40L0-30% (W/W) then, elution speed is 500 milliliters/hour, collects and numbers and preserve with automatic distribution collector.Oligosaccharide content in the separated product and composition use phenol-sulfuric acid process and TLC method to detect respectively.According to the detected result merging filtrate.
3) concentrate, decolouring, drying
The filtrate that previous step is rapid is used activated carbon decolorizing, and the supernatant liquor after the filtration in temperature is below 60 ℃, carry out vacuum concentration under the condition of vacuum tightness-0.8mPa, obtains the mannooligo saccharide syrup.Syrup obtains 88.6g mannooligo saccharide powder-like product through spraying drying, relatively (konjaku powder charging capacity) yield 80.5%, definitely yield (total charging capacity) 69.2%.In this way, the mannooligo saccharide that obtains has six kinds of compositions, is followed successively by according to the elution order of activated carbon column: I (A), II (B), III (C), IV (D), V (E), VI (F).The relative yield 80.5% of (see figure 1), absolute yield 69.2%
Three, interpretation of result
1, analytical procedure:
(1) chromatography of ions (HPAEC) method:
Instrument: chromatography of ions IC2500, adopt ED50 detector, GP50 pump, CarboPac chromatographic column.
Condition: mobile phase A: NaOH (A.R.) 100mM, B: water;
Gradient 40mM-70mM drip washing 30 minutes; 30 ℃;
(2) thin-layer chromatography (TLC) method:
Developping agent: acetonitrile: water=8:2 (V/V); Launch number of times: three times
Developer: the methanol solution of 5% H2SO4 (V/V)
Color condition: 110 ℃, 10 minutes
2, detected result:
(1) thin-layer chromatography (TLC) method: the results are shown in Figure 2.
(2) chromatography of ions (HPAEC) method: the results are shown in Figure 3, Fig. 4.
Claims (1)
1. the production method of a konjak mannose using cellulase, its technical process is: enzymolysis, filtration, decolouring, concentrated, dry, it is characterized in that: press konjaku powder 1g:80-120u enzyme unit alive and drop into cellulase, hemicellulose enzyme mixture, the enzyme ratio of living is 2.5-3.5:1 in cellulase, the hemicellulose enzyme mixture; Enzymolysis adopts centrifuging, ultrafiltration membrance filter after stopping successively, obtains mannooligo saccharide mixed solution and filter residue; Thick slag is removed in centrifuging, and ultrafiltration membrance filter obtains the mannooligo saccharide mixed solution, removes macromole; The aperture of used ultra-filtration membrane is 2-10 microns.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB2006100191729A CN100510094C (en) | 2006-05-29 | 2006-05-29 | Production method of konjak mannose using cellulase |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB2006100191729A CN100510094C (en) | 2006-05-29 | 2006-05-29 | Production method of konjak mannose using cellulase |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1904059A CN1904059A (en) | 2007-01-31 |
| CN100510094C true CN100510094C (en) | 2009-07-08 |
Family
ID=37673423
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CNB2006100191729A Active CN100510094C (en) | 2006-05-29 | 2006-05-29 | Production method of konjak mannose using cellulase |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN100510094C (en) |
Families Citing this family (11)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101816386B (en) * | 2010-04-07 | 2012-10-10 | 云南富源金田原农产品开发有限责任公司 | Konjak oligosaccharide buccal tablets |
| CN101870993A (en) * | 2010-05-16 | 2010-10-27 | 湖北楚天魔芋科技开发有限公司 | Process method for extracting konjac glucomannan by combining enzyme acid with hydrolysis |
| CN103330083A (en) * | 2013-06-28 | 2013-10-02 | 句容市郭庄镇南河农庄 | Composite feed for improving immunity of herring and preparation method thereof |
| CN105453918A (en) * | 2015-12-03 | 2016-04-06 | 丁玉琴 | Method for preparing glyceollin by means of induction effect of konjak composite oligosaccharide |
| CN106136232A (en) * | 2016-06-30 | 2016-11-23 | 成都永安缘和生物科技有限公司 | A kind of preparation method of beneficial bacteria factor composite chewable tablet |
| CN109750070B (en) * | 2019-01-31 | 2022-04-26 | 广东省农业科学院蚕业与农产品加工研究所 | Functional mulberry leaf oligosaccharide and preparation method and application thereof |
| CN111264813A (en) * | 2020-03-24 | 2020-06-12 | 陕西理工大学 | Multifunctional konjac flour and preparation method thereof |
| CN111513273A (en) * | 2020-04-01 | 2020-08-11 | 陕西理工大学 | Konjak functional food and preparation method thereof |
| CN111840248B (en) * | 2020-08-07 | 2021-11-30 | 武汉理工大学 | Konjac pullulan hollow capsule and preparation method thereof |
| CN112280814A (en) * | 2020-10-29 | 2021-01-29 | 武汉芘芘薇莎生物科技有限公司 | High-content medium-low molecular weight konjac glucomannan, preparation method thereof and functional composition thereof |
| CN112587535B (en) * | 2021-01-06 | 2024-06-21 | 澳门大学 | Glucomannan oligosaccharide series composition, preparation method and application thereof |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1280989A (en) * | 2000-08-18 | 2001-01-24 | 干信 | Process for preparing dodecanoyl Konjak-glucomannoligose-sodium sulfonate |
-
2006
- 2006-05-29 CN CNB2006100191729A patent/CN100510094C/en active Active
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1280989A (en) * | 2000-08-18 | 2001-01-24 | 干信 | Process for preparing dodecanoyl Konjak-glucomannoligose-sodium sulfonate |
Non-Patent Citations (4)
| Title |
|---|
| 湖北广华生物有限公司的科研成果通过鉴定. 李章涛，易晓辉.广水市科学技术局（科协）工作动态. 2006 |
| 湖北广华生物有限公司的科研成果通过鉴定. 李章涛，易晓辉.广水市科学技术局（科协）工作动态. 2006 * |
| 纤维素酶制备魔芋葡甘低聚糖. 张迎庆等.吉首大学学报(自然科学版)，第24卷第3期. 2003 |
| 纤维素酶制备魔芋葡甘低聚糖. 张迎庆等.吉首大学学报(自然科学版)，第24卷第3期. 2003 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN1904059A (en) | 2007-01-31 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN100510094C (en) | Production method of konjak mannose using cellulase | |
| CN100549019C (en) | With the stalk is the method that raw material application enzyme and membrane technique prepare high-purity oligoxylose | |
| CN102559810B (en) | Method for preparing high-purity isomaltooligosacharide from wheat starch | |
| CN103497983B (en) | A kind of method using alpha-glucosidase to prepare oligomeric isomaltose | |
| CN103484512B (en) | Method for producing high-functional-trisaccharide-content isomaltooligosaccharide by using immobilized cells | |
| CN104540957A (en) | Compositions and methods for biomass liquefaction | |
| CN101220381A (en) | Process for preparing xylitol by using corn core or agriculture and forestry castoff | |
| CN103468766A (en) | Preparation method of high-purity mannan oligosaccharide | |
| CN112515032A (en) | Extraction method of selenoprotein in cardamine violifolia, selenoprotein obtained by extraction method and application of selenoprotein | |
| Montgomery et al. | Isolation of 6-[α-D-glucopyranosyl]-D-glucose (isomaltose) from enzymic hydrolyzates of starch2 | |
| CN103849665A (en) | Method for pretreating lignocellulose by using carboxyl functionalized ionic liquid solution | |
| CN101597628A (en) | The method of preparing xylo-oligosaccharide by corn husk enzymatic hydrolysis | |
| CN109295131B (en) | Receptor positioning solid-phase enzymolysis preparation method of dendrobe active oligosaccharide | |
| CN101701235A (en) | Method for producing low DE value malt oligosaccharide syrup by potato starch | |
| CN107217080B (en) | Method for preparing jerusalem artichoke fructo-oligosaccharide by using immobilized enzyme | |
| US10351888B2 (en) | Highly efficient method for synthesizing difructose anhydride III | |
| CN101792785A (en) | Method for preparing phosphoryl oligosaccharide by holoenzyme method | |
| CN110903333A (en) | Preparation method of glucoside and derivatives thereof | |
| CN1680571A (en) | Preparation of maltotriose | |
| CN109369731A (en) | A kind of method of glucose during removing xylose production | |
| CN102876756B (en) | Process for co-producing lactic acid with lower polyxylose | |
| CN106046066B (en) | A method of purifying prepares high-purity xylobiose | |
| CN102199641A (en) | Method for extracting sugar from enzymic hydrolysate of jerusalem artichoke residues | |
| CN103937855A (en) | Method for synthesizing isomalto oligosaccharides through fully enzymatic transformation path | |
| JPS63226294A (en) | Production of cellobiose |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| ASS | Succession or assignment of patent right |
Owner name: ACER COMPUTER (CHINA) CO., LTD. Free format text: FORMER OWNER: BEIDA FANGZHENG SCIENCE + TECHNOLOGY COMPUTER SYSTEM CO., LTD., SHANGHAI Effective date: 20101029 |
|
| C41 | Transfer of patent application or patent right or utility model | ||
| COR | Change of bibliographic data |
Free format text: CORRECT: ADDRESS; FROM: 100085 ROOM 204, SECONDARY BUILDING, FANGZHENG BUILDING, NO.9, SHANGDI STREET 5, HAIDIAN DISTRICT, BEIJING TO: 200001 3/F, NO.168, XIZANG MIDDLE ROAD, HUANGPU DISTRICT, SHANGHAI |
|
| TR01 | Transfer of patent right |
Effective date of registration: 20101103 Address after: 430074 Hubei city of Wuhan province Kuanshan East Lake development zone two Road No. 1 International Business Center Patentee after: Wuhan Sunhy Biology Co., Ltd. Address before: 432700 No. ten Li Industrial Park, No. 9, Hubei, Guangshui Patentee before: Hubei Guanghua Biology Co., Ltd. |
|
| DD01 | Delivery of document by public notice |
Addressee: Gu Lingfang Document name: Notification of Passing Examination on Formalities |
|
| C56 | Change in the name or address of the patentee | ||
| CP02 | Change in the address of a patent holder |
Address after: 430074 Hubei city of Wuhan province East Lake New Technology Development Zone, Ling Shan Road No. 5 Patentee after: Wuhan Sunhy Biology Co., Ltd. Address before: 430074 Hubei city of Wuhan province Kuanshan East Lake development zone two Road No. 1 International Business Center Patentee before: Wuhan Sunhy Biology Co., Ltd. |
|
| DD01 | Delivery of document by public notice |
Addressee: Gu Lingfang Document name: Notice of conformity |
|
| EE01 | Entry into force of recordation of patent licensing contract |
Assignee: Zhengzhou Hongzhan Feed Additive Co., Ltd. Assignor: Wuhan Sunhy Biology Co., Ltd. Contract record no.: 2012410000011 Denomination of invention: Production method of konjak mannose using cellulase Granted publication date: 20090708 License type: Exclusive License Open date: 20070131 Record date: 20120228 |
|
| CP02 | Change in the address of a patent holder | ||
| CP02 | Change in the address of a patent holder |
Address after: No.98, guangguba Road, Donghu New Technology Development Zone, Wuhan City, Hubei Province Patentee after: Wuhan Sunhy Biology Co., Ltd. Address before: 430074, No. 5, Ling Nan Road, East Lake New Technology Development Zone, Wuhan, Hubei, China Patentee before: Wuhan Sunhy Biology Co., Ltd. |