CN102127574A - Method for producing fructooligosaccharide by using transfructosylase - Google Patents
Method for producing fructooligosaccharide by using transfructosylase Download PDFInfo
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- CN102127574A CN102127574A CN201010548887XA CN201010548887A CN102127574A CN 102127574 A CN102127574 A CN 102127574A CN 201010548887X A CN201010548887X A CN 201010548887XA CN 201010548887 A CN201010548887 A CN 201010548887A CN 102127574 A CN102127574 A CN 102127574A
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- oligofructose
- fructosyl transferase
- sucrose
- transfructosylase
- fructooligosaccharide
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Abstract
The invention relates to the technical field of food biotechnology, particularly a method for producing fructooligosaccharide by using transfructosylase, which comprises preparation of transfructosylase and industrial production of fructooligosaccharide by using the enzyme preparation. The method is characterized by comprising the following steps: selecting a good strain containing high-activity transfructosylase, culturing the strain in an appropriate culture medium, separating to obtain large-scale cultured mycelia containing high-activity transfructosylase, and storing the mycelia for later use; and converting sucrose into fructooligosaccharide by a batch process by using the mycelia containing high-activity transfructosylase as a biocatalyst. The method provided by the invention has the advantages of short bioconversion time, and high conversion rate of fructooligosaccharide; the mycelia can be repeatedly used; and the content of fructooligosaccharide (trisaccharide) prepared by using the transfructosylase is up to 57-62%, which is 3-5% higher than that of the fructooligosaccharide prepared by using Aspergillus niger.
Description
(1) technical field
The present invention relates to technical field of food biotechnology, particularly a kind of method of producing oligofructose with fructosyl transferase.
(2) background technology
Oligofructose is a kind of water-soluble dietary fibre, take for a long time and can reduce serum cholesterol, improve lipid metabolism, through animal and human's body experiment confirm, oligofructose has following physiological function: by probioticss such as bifidus bacillus are utilized, bifidus bacillus has the effect of two-ways regulation when promptly only breeding 10～100 times.After human body is taken in oligofructose, profitable strain bifidus bacillus quantity increases in the body, can suppress growth and breeding such as intrinsic putrefactive bacterium such as Salmonellas in external source pathogenic bacterium and the intestines, reduces the growth and the accumulation of corrupt substance in the intestines, promote intestines peristalsis, prevent constipation and diarrhoea.Oligofructose is a kind of good water-soluble dietary fibre, can effectively reduce the quantity of serum cholesterol, triglyceride, free fatty acids, for having a better role because of a series of cardiovascular disordeies such as the high hypertension that causes of blood fat, arteriosclerosis.Oligofructose is generated lactic acid by fermentation using bacteria in large intestine, can solubilize calcium, mineral substance such as magnesium, iron, promote the absorption of human body to mineral substance.
Experiment confirm, oligofructose promote the specific absorption of calcium to reach 70.8%.Therefore, oligofructose can promote to grow and prevent osteoporosis.Also can promote forming naturally of VITMAIN B1, B2, B3, B6, B12 and folic acid simultaneously, thereby improve the human body metabolism, improve immunizing power and disease resistance, the skin diseases that prevention and improvement cause owing to the body endogenous toxic material, can prevent face sore, blackspot, freckle, comedo, senile plaque, make skin beautiful, slow down aging.After bifidus bacillus absorbs oligofructose; breed rapidly; suppressing spoilage organism such as intestinal bacteria, Salmonellas and clostridium has an effect; reduce the generation of toxic metabolites (as indoles, nitroso-group ammonia), rapidly toxic metabolites is excreted simultaneously, alleviate burden of liver; play liver-protective effect; prevent effects such as various chronic diseases, cancer obviously, oligofructose seldom can be decomposed by hydrochloric acid in gastric juice in the digestive tube and enzyme, and extremely difficulty is absorbed by the body.According to surveying and determination, the calorific value of oligofructose is below the 1.5Kcal/g, and the sucrose calorific value is 4.6Kcal/g, therefore after taking in oligofructose, can not cause obesity, be ideal, low-calorie functional sweetener. oligofructose can not be generated insoluble glucan and provide oral microorganism deposition to produce acid and corrosive place (tartar) by the streptococcus mutans utilization, therefore can prevent carious tooth.
At present, the production of oligofructose is to be raw material with sucrose basically, 2～5 fructosyls of effect formation by fructosyl transferase are chain link, with the glucosyl group end group that is chain, with fructosyl → fructose connecting key is that main body framework links the carbohydrate that forms, promptly 1～4 fructosyl is connected with β-2,1 key on the D-fructosyl of sucrose and the kestose (GF2), GF3 (GF3), GF4 (GF4) and the sugarcane that the form mixture of six sugar (GF5) really.The commodity oligofructose generally also contains a little sugar, fructose, glucose.
The zymogenic bacteria that uses in the industrial production mostly is mould, as aspergillus niger ACTT20611, smelly aspergillus NRRL4337, aspergillus japonicus TITKJ1 etc., many plants, yeast and mould can both produce fructosyl transferase, but the active low and output of the enzyme that plant produces is subjected to the restriction in season; And the yeast conversion enzymic activity is strong but be unfavorable for the accumulation of oligofructose.
(3) summary of the invention
The present invention provides the method with fructosyl transferase production oligofructose that a kind of transformation time is short, transformation efficiency is high in order to remedy the deficiencies in the prior art.
The present invention is achieved through the following technical solutions:
A kind of method of producing oligofructose with fructosyl transferase comprises the preparation of fructosyl transferase and utilizes this zymin to carry out suitability for industrialized production oligofructose two portions, it is characterized in that:
(1) seed selection has the strain excellent that contains the high reactivity fructosyl transferase, cultivates in suitable medium, separates the mycelium that contains the high reactivity fructosyl transferase that obtains large scale culturing then, preserves stand-by;
(2) transforming sucrose as biological catalyst with batch process with the mycelium that contains the high reactivity fructosyl transferase is oligofructose.
It is a mould that seed selection of the present invention has the strain excellent that contains the high reactivity fructosyl transferase.
Suitable slant medium of the present invention comprises following material: the 5-30% potato, 1-5% sucrose, the 0.1-0.3% potassium primary phosphate, 0.15-0.4% sal epsom, the 0.2-0.5% SODIUMNITRATE, 0.2% agar, its cultural method are at 25-38 ℃, air flow 100-500ml/min cultivated 40-60 hour under the stirring velocity 50-200r/min condition.
The mycelium substratum that suitable macro preparation of the present invention contains the high reactivity fructosyl transferase comprises following material: 2-10% sucrose, 3-8% dregs of beans, 0.3-2% Semen Maydis powder, its cultural method is at 27-32 ℃, air flow 100-500ml/min, cultivated 10-40 hour under the stirring velocity 50-200r/min condition, centrifugation obtains a large amount of mycelium, places 0-4 ℃ to descend standby.
The present invention is in the retort that fills the 30-60% sucrose solution with glycosyltransferase production batch process that oligofructose was adopted, press the enzyme amount of fructosyl transferase 2000-5000u/kg sucrose, PH is at 4.9-7.0 in control, temperature is at 27-31 ℃, the agitation condition of 30-49r/min reacted 13-40 hour down, obtain faint yellowly through the microfiltration membrane separation of mycelial,, be condensed into oligofructose at last by macroporous resin decolouring to yellow liquid.The isolating mycelium of microfiltration membrane can repeatedly repeat above-mentioned reaction process and use.
The present invention be a kind of be raw material with sucrose, the method for suitability for industrialized production oligofructose, the strain excellent that the present invention can secrete fructosyl transferase is the some mould (Penicillium notatum) that screens from soil.
Mycelium of the present invention also can be prepared into immobilized enzyme and use.
The present invention produces the method for oligofructose with fructosyl transferase, the bio-transformation time is short, oligofructose transformation efficiency height, mycelium can be repeatedly used, and has the bacterial strain point mould of high fructosyltransferaseactivity activity, cultivates 10-20 hour through shake flask fermentation, yield of enzyme is 200-220u/g, prepare oligofructose with this enzyme, oligofructose trisaccharide content reaches 57%-62%, prepares the high 3-5 of oligofructose percentage point than aspergillus niger.
(4) description of drawings
The present invention is further illustrated below in conjunction with accompanying drawing.
Accompanying drawing is for preparing the technological process of production figure of oligofructose with fructosyl transferase.
(5) embodiment
Embodiment 1:
The penicillium notatum kind is linked into contains 5% sucrose, 3.5% dregs of beans, the fermentor tank of 0.5% Semen Maydis powder nutrient solution 50L, culture condition is 30 ℃, PH5.20, air flow 200ml/min, stirring velocity 150r/min, through fermentation in 20 hours, centrifugation obtained a large amount of mycelium, detecting enzyme by HPLC lives, the enzyme amount of pressing 3500u/kg sucrose drops into the enzyme reaction jar of the sucrose solution of mycelium to 45%, 30 ℃ of control reaction temperature, PH5.2 ± 0.2, reacted 24 hours, detect oligofructose content by HPLC and reach more than 50%, reaction solution is crossed the ceramic microfiltration membrane separation of mycelial, obtains the thick liquid of transparent faint yellow oligofructose, by macroporous resin decolouring, concentrate and obtain colourity less than 0.05 oligofructose.
Embodiment 2:
The penicillium notatum kind is linked into contains 8% sucrose, 4.0% dregs of beans, the fermentor tank of 1.5% Semen Maydis powder nutrient solution 5000L, culture condition is 32 ℃, air flow 500ml/min, stirring velocity 120r/min, through fermentation in 24 hours, centrifugation obtains a large amount of mycelium, detects enzyme by HPLC and lives, and the enzyme amount of pressing 4500u/kg sucrose drops into the enzyme reaction jar of the sucrose solution of mycelium to 50%, 31 ℃ of control reaction temperature, reacted 20 hours PH6.0 ± 0.2, detects oligofructose content by HPLC and reach more than 50%, reaction solution is crossed the ceramic microfiltration membrane separation of mycelial, obtain the thick liquid of transparent faint yellow oligofructose,, concentrate and obtain colourity less than 0.05 oligofructose by macroporous resin decolouring.
Embodiment 3:
The penicillium notatum kind is linked into contains 20% potato, 3% sucrose, 0.2% potassium primary phosphate, 0.3% sal epsom, 0.3% SODIUMNITRATE, the fermentor tank of 0.2% agar nutrient solution 50L, culture condition are 27 ℃, air flow 300ml/min, stirring velocity 100r/min, through fermentation in 50 hours, centrifugation obtained a large amount of mycelium, detecting enzyme by HPLC lives, the enzyme amount of pressing 3000u/kg sucrose drops into the enzyme reaction jar of the sucrose solution of mycelium to 40%, 27 ℃ of control reaction temperature, PH6.3 ± 0.2, reacted 30 hours, detect oligofructose content by HPLC and reach more than 50%, reaction solution is crossed the ceramic microfiltration membrane separation of mycelial, obtains the thick liquid of transparent faint yellow oligofructose, by macroporous resin decolouring, concentrate and obtain colourity less than 0.05 oligofructose.
Claims (5)
1. a method of producing oligofructose with fructosyl transferase comprises the preparation of fructosyl transferase and utilizes this zymin to carry out suitability for industrialized production oligofructose two portions, it is characterized in that:
(1) seed selection has the strain excellent that contains the high reactivity fructosyl transferase, cultivates in suitable medium, separates the mycelium that contains the high reactivity fructosyl transferase that obtains large scale culturing then, preserves stand-by;
(2) transforming sucrose as biological catalyst with batch process with the mycelium that contains the high reactivity fructosyl transferase is oligofructose.
2. method of producing oligofructose with fructosyl transferase according to claim 1, it is characterized in that: it is a mould that seed selection has the strain excellent that contains the high reactivity fructosyl transferase.
3. method of producing oligofructose with fructosyl transferase according to claim 1, it is characterized in that: described suitable medium comprises following material: the 5-30% potato, 1-5% sucrose, the 0.1-0.3% potassium primary phosphate, 0.15-0.4% sal epsom, 0.2-0.5% SODIUMNITRATE, 0.2% agar, its cultural method is at 25-38 ℃, and air flow 100-500ml/min cultivated 40-60 hour under the stirring velocity 50-200r/min condition.
4. method of producing oligofructose with fructosyl transferase according to claim 1, it is characterized in that: described suitable medium comprises following material: 2-10% sucrose, 3-8% dregs of beans, 0.3-2% Semen Maydis powder, its cultural method is at 27-32 ℃, air flow 100-500ml/min cultivated 10-40 hour under the stirring velocity 50-200r/min condition.
5. method of producing oligofructose with fructosyl transferase according to claim 1, it is characterized in that: with glycosyltransferase production batch process that oligofructose was adopted is in the retort that fills the 30-60% sucrose solution, press the enzyme amount of fructosyl transferase 2000-5000u/kg sucrose, PH is at 4.9-7.0 in control, temperature is at 27-31 ℃, the agitation condition of 30-49r/min reacted 13-40 hour down, obtain faint yellow to yellow liquid through the microfiltration membrane separation of mycelial, by the macroporous resin decolouring, be condensed into oligofructose at last.
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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102504994A (en) * | 2011-12-19 | 2012-06-20 | 南宁纵联科技有限公司 | Method for preparing fructooligosaccharide by using baffled bioreactor |
| CN103980382A (en) * | 2014-05-28 | 2014-08-13 | 南京工业大学 | Method for purifying fructose oligosaccharide |
| CN104059857A (en) * | 2014-06-30 | 2014-09-24 | 广西大学 | Aspergillus and application of aspergillus in fructosyltransferase preparing |
| CN104116000A (en) * | 2014-08-08 | 2014-10-29 | 山东百龙创园生物科技有限公司 | Preparation method for fructo-oligo saccharide feed additive |
| CN106467899A (en) * | 2015-08-17 | 2017-03-01 | 中国科学院天津工业生物技术研究所 | A kind of Aspergillus niger strain of high yield fructose-transferring enzyme and its application |
| CN104419688B (en) * | 2013-09-02 | 2017-12-22 | 中国科学院微生物研究所 | A kind of transfructosylase and its gene, its secreted expression carrier and application |
| CN107683934A (en) * | 2017-08-25 | 2018-02-13 | 广西南宁华鑫糖业技术有限责任公司 | A kind of preparation method of prebiotics jujube sugar |
| CN108330150A (en) * | 2018-05-16 | 2018-07-27 | 量子高科(中国)生物股份有限公司 | A kind of oligofructose prepared as raw material using brown sugar |
Citations (1)
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| CN1335402A (en) * | 2001-08-12 | 2002-02-13 | 广西大学 | Production process of cane-fruit oligosaccharide with immobilized fructose-base transferase |
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2010
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Patent Citations (1)
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| CN1335402A (en) * | 2001-08-12 | 2002-02-13 | 广西大学 | Production process of cane-fruit oligosaccharide with immobilized fructose-base transferase |
Non-Patent Citations (1)
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Cited By (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102504994A (en) * | 2011-12-19 | 2012-06-20 | 南宁纵联科技有限公司 | Method for preparing fructooligosaccharide by using baffled bioreactor |
| CN102504994B (en) * | 2011-12-19 | 2014-04-02 | 南宁纵联科技有限公司 | Method for preparing fructooligosaccharide by using baffled bioreactor |
| CN104419688B (en) * | 2013-09-02 | 2017-12-22 | 中国科学院微生物研究所 | A kind of transfructosylase and its gene, its secreted expression carrier and application |
| CN103980382A (en) * | 2014-05-28 | 2014-08-13 | 南京工业大学 | Method for purifying fructose oligosaccharide |
| CN104059857A (en) * | 2014-06-30 | 2014-09-24 | 广西大学 | Aspergillus and application of aspergillus in fructosyltransferase preparing |
| CN104116000A (en) * | 2014-08-08 | 2014-10-29 | 山东百龙创园生物科技有限公司 | Preparation method for fructo-oligo saccharide feed additive |
| CN106467899A (en) * | 2015-08-17 | 2017-03-01 | 中国科学院天津工业生物技术研究所 | A kind of Aspergillus niger strain of high yield fructose-transferring enzyme and its application |
| CN106467899B (en) * | 2015-08-17 | 2020-05-05 | 中国科学院天津工业生物技术研究所 | Aspergillus niger strain capable of producing fructose transferase in high yield and application thereof |
| CN107683934A (en) * | 2017-08-25 | 2018-02-13 | 广西南宁华鑫糖业技术有限责任公司 | A kind of preparation method of prebiotics jujube sugar |
| CN108330150A (en) * | 2018-05-16 | 2018-07-27 | 量子高科(中国)生物股份有限公司 | A kind of oligofructose prepared as raw material using brown sugar |
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Application publication date: 20110720 |