CN105039486A - Method for extracting ferulic acid from wheat straw by biotechnology - Google Patents
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Abstract
The invention discloses a method for extracting ferulic acid from wheat straw by biotechnology. The method utilizes wheat straw as a raw material and an ultrasonic assisted digestion-microbial fermentation combined method to prepare ferulic acid so that an extraction rate is greatly improved. A detection result shows that an extraction rate of ferulic acid from wheat straw is 95% or more and the highest extraction rate is 9.17mg/g and is substantially higher than the existing level. Cellulose and lignin in wheat straw are degraded by an alkali treatment-composite microbial fermentation-ultrasound assisted enzymolysis-two-step alcohol extraction combined technology and crosslinking of ferulic acid ester bonds, and polysaccharide and lignin is broken so that a ferulic acid extraction rate is improved. Wheat straw cellulose is obtained by ultrasonic assisted digestion and microbial fermentation so that a straw feed nutrition value is substantially improved. A production cost is low, drying processes with high energy consumption are less, anhydrous ethanol can be recovered and recycled and industrialization feasibility is improved.
Description
Technical field
The present invention relates to a kind of method that biological process extracts forulic acid in wheat stalk, belong to biological chemical field.
Background technology
Wheat stalk is ripe wheat stalk, leaf, fringe part summation, is remainder after wheat maturation acquisition seed, accounts for crop quality 50%.China produces nearly 1.1 hundred million tons of wheat stalk every year, except minority is by as except animal feed, farm manure and rural area fuel, major part is stacked or also field after directly burning, and effective industrial utilization proportion is extremely low, waste precious resources, and very big pollution is also caused to environment.Modern cereal science research finds, and wheat stalk contains and multiplely has antitumor, anti-oxidant important physiologically active substance, as food fibre, araboxylan, Dui – coumaric acid and forulic acid; Wherein, in recent years general concern has been caused to the research of forulic acid both at home and abroad.Forulic acid (FerulicAcid) is organic phenolic acid, it is one of derivative of TRANSCINNAMIC ACID, forulic acid has been internationally recognized natural antioxidants now, research in recent years in pharmacological effect has found pharmacological action and the biological activity of many forulic acid and derivative, the protective effect on cancer risk of forulic acid have also been obtained high cognition degree, not only be applied to field of medicaments, be also widely used in the fields such as food, healthcare products, makeup; The main Physiological Function of forulic acid has oxidation-resistance, antisepsis and anti-inflammation, reducing blood-fat, antithrombotic, anti-mutation, anti-coronary heart disease and anti-cancer etc.; Forulic acid is except being widely used in field of medicaments, and some national approveds will it can be used as foodstuff additive to be applied to field of food, mainly as preparing antisepsis antistaling agent, food linking agent etc.The preparation method of current forulic acid mainly comprises extraction and biological synthesis process from vegetable material, from vegetable material, wherein extract methods such as mainly comprising alkaline process, enzyme process, ultrasonic wave, microwave, supercritical extraction.
Research in recent years finds, forulic acid is the phenolic acid that in wheat stalk, content is the highest, in the cell walls of vegetable cell, it is cross-linked mainly through ester bond and polysaccharide and xylogen, or self esterification or etherificate form two forulic acid, extract difficulty larger, existing extractive technique extraction yield is general lower, general less than 80%, and extraction cost is higher, govern the suitability for industrialized production utilizing wheat stalk to extract forulic acid, therefore, set up a set of efficiency higher, cost is lower, the wheat stalk total ferulic acid development and utilization measure of safe and feasible, for the further recycling of wheat stalk provides new approach, to the economic value added increasing wheat planting industry, the industrial advantage giving full play to China's wheat has great importance.
Summary of the invention
For overcoming the deficiencies in the prior art, a kind of biological process is the object of the present invention is to provide to extract the method for forulic acid in wheat stalk, this technique extraction process is simple, be easy to large-scale industrial production, contribute to solving the wheat stalk existed at present and extract the problems such as extraction rate of ferulaic acid form is lower, cost is higher, owing to improving the present situation of forulic acid still based on chemosynthesis in the market.
Above-mentioned purpose of the present invention is achieved by the following technical programs:
The invention provides a kind of method that biological process extracts forulic acid in wheat stalk, it is characterized in that specifically comprising the following steps:
(1) wheat stalk pre-treatment: pulverized by wheat stalk, crosses 100 mesh sieves, adds distilled water with the amount of feed liquid mass ratio 1:5 ~ 10, heats the enzyme that goes out, obtains wheat stalk suspension;
(2) alkaline purification: the S-WAT adding total mass ratio 3 ~ 5% in wheat stalk suspension, stir process 2 ~ 3h;
(3) ultrasound-assisted enzymolysis: adjustment pH of suspension 6.0 ~ 7.0, add 3 ~ 5% cellulases of wheat stalk quality, 2 ~ 3% zytases, enzymolysis under ultrasonic assistant effect, hydrolysis temperature 50 ~ 60 DEG C, ultrasonic power 600W, period adopts stirrer at the uniform velocity to stir, and enzymolysis time is 40 ~ 60min;
(4) centrifugal: enzymolysis terminates rear centrifugal, obtain supernatant liquor a and wheat stalk residue;
(5) bio-fermentation agent preparation: get 8 ~ 10 parts of sorbent materials, 3 ~ 5 parts of monocalcium phosphates, 6 ~ 8 parts of glucose, 10 ~ 15 parts of rice brans, 10 ~ 15 portions of Semen Maydis powder, 3 ~ 5 parts of rapeseed cakes, 10 ~ 15 parts of wheat stalks, 2 ~ 3 parts of Trichodermareeseis, 1 ~ 2 part of aspergillus niger, 1-2 part subtilis, mixing and stirring; Then moisture is adjusted to 35 ~ 40%, anaerobically fermenting 7 ~ 10d at 25 DEG C ~ 30 DEG C with the aqueous solution containing 2% molasses;
(6) batch mixing: wheat stalk residue 100 parts, bio-fermentation agent 3 ~ 5 parts, cellulase 0.5 ~ 1 part, zytase 0.5 ~ 1 part of ratio mix in mass ratio, adjustment moisture to 35 ~ 40%;
(7) anaerobically fermenting: by mixed mixture temperature control anaerobically fermenting 2 ~ 3d, temperature 28 ~ 32 DEG C, periodic agitation;
(8) ultrasound-assisted enzymolysis: add 3 ~ 5 times of water after fermentation ends in fermentation residue, add the zytase of fermentation residue mass fraction 2 ~ 3%, enzymolysis under ultrasonic assistant effect, hydrolysis temperature 50 ~ 60 DEG C, ultrasonic power 300W, period adopts stirrer at the uniform velocity to stir, and enzymolysis time is 20 ~ 30min;
(9) centrifugal: enzymolysis terminates rear excessively centrifugal, centrifugal rotational speed 3000 ~ 5000rpm, separation of supernatant b and solid residue;
(10) concentrate drying: merge supernatant liquor a, b, concentrated, vacuum-drying obtains forulic acid crude product;
(11) two step alcohol extractings: to solid residue 3 ~ 5 times of dehydrated alcohols, alcohol extracting 20 ~ 30min under ultrasonic assistant effect, ultrasonic power 300W, period adopts stirrer at the uniform velocity to stir;
(12) centrifugal: alcohol extracting terminates rear centrifugal, centrifugal rotational speed 3000 ~ 5000rpm, is separated to obtain alcohol extract and solid residue;
(13) alcohol wash: rinsed 3 ~ 5 times by above-mentioned solid residue dehydrated alcohol, merges alcohol extract;
(14) purifying: the alcohol extract that the forulic acid crude product step 13 step 10 obtained obtains dissolves, centrifugal removing precipitation, obtains transparent faint yellow supernatant;
(15) concentrate drying: supernatant concentration, vacuum-drying are obtained the higher forulic acid of purity.
Sorbent material described in above-mentioned steps (5) is one or more in zeolite, gac, wilkinite, vermiculite.
The invention provides a kind of method that biological process extracts forulic acid in wheat stalk, product utilization wheat stalk is raw material, preparation forulic acid is combined by ultrasound-assisted enzymolysis and microbe fermentation method, extraction yield is greatly improved, detected result shows that the extraction rate reached of forulic acid in wheat stalk is to more than 95%, can 9.17mg/g be reached, be significantly higher than state of the art of having reported for work at present.
The invention provides a kind of method that biological process extracts forulic acid in wheat stalk, it is advantageous that: one is the Mierocrystalline cellulose, xylogen etc. in the process degradation wheat stalk adopting alkaline purification+compound microorganism ferments+ultrasound-assisted enzymolysis+two step alcohol extracting to combine, break the crosslinked of forulic acid ester bond and polysaccharide and xylogen, improve the extraction yield of forulic acid; Two be by ultrasound-assisted enzymolysis and fermentable after wheat stalk Mierocrystalline cellulose, Lignin degradation rate reach 67.4%, significantly improve the Forage Nutritive Value of stalk; Three is that explained hereafter cost is lower, and the high dry link that consumes energy is few, and dehydrated alcohol recoverable, is beneficial to suitability for industrialized production.
Embodiment
Below in conjunction with embodiment, the present invention will be further described.
embodiment 1:
Biological process extracts a method for forulic acid in wheat stalk, is prepared by following steps:
(1) wheat stalk pre-treatment: pulverized by wheat stalk, crosses 100 mesh sieves, adds distilled water with the amount of feed liquid mass ratio 1:8, heats the enzyme that goes out, obtains wheat stalk suspension;
(2) alkaline purification: the S-WAT adding total mass ratio 3 ~ 5% in wheat stalk suspension, stir process 2.5h;
(3) ultrasound-assisted enzymolysis: adjustment pH of suspension 6.0 ~ 7.0, adds 4% cellulase of wheat stalk quality, 2.5% zytase, enzymolysis under ultrasonic assistant effect, hydrolysis temperature 50 ~ 60 DEG C, ultrasonic power 600W, period adopts stirrer at the uniform velocity to stir, and enzymolysis time is 50min;
(4) centrifugal: enzymolysis terminates rear centrifugal, obtain supernatant liquor a and wheat stalk residue;
(5) bio-fermentation agent preparation: get 9 parts of zeolites, 4 parts of monocalcium phosphates, 7 parts of glucose, 12 parts of rice brans, 12 portions of Semen Maydis powder, 4 parts of rapeseed cakes, 12 parts of wheat stalks, 2.5 parts of Trichodermareeseis, 1.5 parts of aspergillus nigers, 1.5 parts of subtilises, mixing and stirring; Then moisture is adjusted to 35 ~ 40%, anaerobically fermenting 8d at 25 DEG C ~ 30 DEG C with the aqueous solution containing 2% molasses;
(6) batch mixing: wheat stalk residue 100 parts, bio-fermentation agent 4 parts, cellulase 0.8 part, zytase 0.8 part of ratio mix in mass ratio, adjustment moisture to 35 ~ 40%;
(7) anaerobically fermenting: by mixed mixture temperature control anaerobically fermenting 2.5d, temperature 28 ~ 32 DEG C, periodic agitation;
(8) ultrasound-assisted enzymolysis: add 4 times of water after fermentation ends in fermentation residue, add the zytase of fermentation residue mass fraction 2.5%, enzymolysis under ultrasonic assistant effect, hydrolysis temperature 50 ~ 60 DEG C, ultrasonic power 300W, period adopts stirrer at the uniform velocity to stir, and enzymolysis time is 25min;
(9) centrifugal: enzymolysis terminates rear excessively centrifugal, centrifugal rotational speed 3000 ~ 5000rpm, separation of supernatant b and solid residue;
(10) concentrate drying: merge supernatant liquor a, b, concentrated, vacuum-drying obtains forulic acid crude product;
(11) two step alcohol extractings: to solid residue 4 times of dehydrated alcohols, alcohol extracting 25min under ultrasonic assistant effect, ultrasonic power 300W, period adopts stirrer at the uniform velocity to stir;
(12) centrifugal: alcohol extracting terminates rear centrifugal, centrifugal rotational speed 3000 ~ 5000rpm, is separated to obtain alcohol extract and solid residue;
(13) alcohol wash: above-mentioned solid residue dehydrated alcohol is rinsed 4 times, merges alcohol extract;
(14) purifying: the alcohol extract that the forulic acid crude product step 13 step 10 obtained obtains dissolves, centrifugal removing precipitation, obtains transparent faint yellow supernatant;
(15) concentrate drying: supernatant concentration, vacuum-drying are obtained the higher forulic acid of purity.
embodiment 2:
Biological process extracts a method for forulic acid in wheat stalk, is prepared by following steps:
(1) wheat stalk pre-treatment: pulverized by wheat stalk, crosses 100 mesh sieves, adds distilled water with the amount of feed liquid mass ratio 1:10, heats the enzyme that goes out, obtains wheat stalk suspension;
(2) alkaline purification: the S-WAT adding total mass ratio 5% in wheat stalk suspension, stir process 2h;
(3) ultrasound-assisted enzymolysis: adjustment pH of suspension 6.0 ~ 7.0, adds 5% cellulase of wheat stalk quality, 3% zytase, enzymolysis under ultrasonic assistant effect, hydrolysis temperature 50 ~ 60 DEG C, ultrasonic power 600W, period adopts stirrer at the uniform velocity to stir, and enzymolysis time is 40min;
(4) centrifugal: enzymolysis terminates rear centrifugal, obtain supernatant liquor a and wheat stalk residue;
(5) bio-fermentation agent preparation: get 10 parts of gacs, 3 parts of monocalcium phosphates, 8 parts of glucose, 10 parts of rice brans, 15 portions of Semen Maydis powder, 3 parts of rapeseed cakes, 15 parts of wheat stalks, 2 parts of Trichodermareeseis, 2 parts of aspergillus nigers, 1 part of subtilis, mixing and stirring; Then moisture is adjusted to 35 ~ 40%, anaerobically fermenting 10d at 25 DEG C ~ 30 DEG C with the aqueous solution containing 2% molasses;
(6) batch mixing: wheat stalk residue 100 parts, bio-fermentation agent 5 parts, cellulase 1 part, zytase 1 part of ratio mix in mass ratio, adjustment moisture to 35 ~ 40%;
(7) anaerobically fermenting: by mixed mixture temperature control anaerobically fermenting 2d, temperature 28 ~ 32 DEG C, periodic agitation;
(8) ultrasound-assisted enzymolysis: add 5 times of water after fermentation ends in fermentation residue, add the zytase of fermentation residue mass fraction 3%, enzymolysis under ultrasonic assistant effect, hydrolysis temperature 50 ~ 60 DEG C, ultrasonic power 300W, period adopts stirrer at the uniform velocity to stir, and enzymolysis time is 20min;
(9) centrifugal: enzymolysis terminates rear excessively centrifugal, centrifugal rotational speed 3000 ~ 5000rpm, separation of supernatant b and solid residue;
(10) concentrate drying: merge supernatant liquor a, b, concentrated, vacuum-drying obtains forulic acid crude product;
(11) two step alcohol extractings: to solid residue 5 times of dehydrated alcohols, alcohol extracting 20min under ultrasonic assistant effect, ultrasonic power 300W, period adopts stirrer at the uniform velocity to stir;
(12) centrifugal: alcohol extracting terminates rear centrifugal, centrifugal rotational speed 3000 ~ 5000rpm, is separated to obtain alcohol extract and solid residue;
(13) alcohol wash: above-mentioned solid residue dehydrated alcohol is rinsed 5 times, merges alcohol extract;
(14) purifying: the alcohol extract that the forulic acid crude product step 13 step 10 obtained obtains dissolves, centrifugal removing precipitation, obtains transparent faint yellow supernatant;
(15) concentrate drying: supernatant concentration, vacuum-drying are obtained the higher forulic acid of purity.
embodiment 3:
Biological process extracts a method for forulic acid in wheat stalk, is prepared by following steps:
(1) wheat stalk pre-treatment: pulverized by wheat stalk, crosses 100 mesh sieves, adds distilled water with the amount of feed liquid mass ratio 1:5, heats the enzyme that goes out, obtains wheat stalk suspension;
(2) alkaline purification: the S-WAT adding total mass ratio 3% in wheat stalk suspension, stir process 3h;
(3) ultrasound-assisted enzymolysis: adjustment pH of suspension 6.0 ~ 7.0, adds 3% cellulase of wheat stalk quality, 2% zytase, enzymolysis under ultrasonic assistant effect, hydrolysis temperature 50 ~ 60 DEG C, ultrasonic power 600W, period adopts stirrer at the uniform velocity to stir, and enzymolysis time is 60min;
(4) centrifugal: enzymolysis terminates rear centrifugal, obtain supernatant liquor a and wheat stalk residue;
(5) bio-fermentation agent preparation: get 8 parts of wilkinites, 5 parts of monocalcium phosphates, 6 parts of glucose, 15 parts of rice brans, 10 portions of Semen Maydis powder, 5 parts of rapeseed cakes, 10 parts of wheat stalks, 3 parts of Trichodermareeseis, 1 part of aspergillus niger, 2 parts of subtilises, mixing and stirring; Then moisture is adjusted to 35 ~ 40%, anaerobically fermenting 10d at 25 DEG C ~ 30 DEG C with the aqueous solution containing 2% molasses;
(6) batch mixing: wheat stalk residue 100 parts, bio-fermentation agent 3 parts, cellulase 0.5 part, zytase 0.5 part of ratio mix in mass ratio, adjustment moisture to 35 ~ 40%;
(7) anaerobically fermenting: by mixed mixture temperature control anaerobically fermenting 3d, temperature 28 ~ 32 DEG C, periodic agitation;
(8) ultrasound-assisted enzymolysis: add 3 times of water after fermentation ends in fermentation residue, add the zytase of fermentation residue mass fraction 2%, enzymolysis under ultrasonic assistant effect, hydrolysis temperature 50 ~ 60 DEG C, ultrasonic power 300W, period adopts stirrer at the uniform velocity to stir, and enzymolysis time is 30min;
(9) centrifugal: enzymolysis terminates rear excessively centrifugal, centrifugal rotational speed 3000 ~ 5000rpm, separation of supernatant b and solid residue;
(10) concentrate drying: merge supernatant liquor a, b, concentrated, vacuum-drying obtains forulic acid crude product;
(11) two step alcohol extractings: to solid residue 3 times of dehydrated alcohols, alcohol extracting 30min under ultrasonic assistant effect, ultrasonic power 300W, period adopts stirrer at the uniform velocity to stir;
(12) centrifugal: alcohol extracting terminates rear centrifugal, centrifugal rotational speed 3000 ~ 5000rpm, is separated to obtain alcohol extract and solid residue;
(13) alcohol wash: above-mentioned solid residue dehydrated alcohol is rinsed 3 times, merges alcohol extract;
(14) purifying: the alcohol extract that the forulic acid crude product step 13 step 10 obtained obtains dissolves, centrifugal removing precipitation, obtains transparent faint yellow supernatant;
(15) concentrate drying: supernatant concentration, vacuum-drying are obtained the higher forulic acid of purity.
Above embodiment only for illustration of technical scheme of the present invention, but not is limited; Although be described in detail by invention with reference to previous embodiment, for the person of ordinary skill of the art, still the technical scheme described in previous embodiment can be modified, or equivalent replacement has been carried out to wherein portion of techniques feature; And to these amendments or replacement, do not make the essence of appropriate technical solution depart from the spirit and scope of the present invention's technical scheme required for protection.
Claims (2)
1. biological process extracts a method for forulic acid in wheat stalk, it is characterized in that specifically comprising the following steps:
(1) wheat stalk pre-treatment: pulverized by wheat stalk, crosses 100 mesh sieves, adds distilled water with the amount of feed liquid mass ratio 1:5 ~ 10, heats the enzyme that goes out, obtains wheat stalk suspension;
(2) alkaline purification: the S-WAT adding total mass ratio 3 ~ 5% in wheat stalk suspension, stir process 2 ~ 3h;
(3) ultrasound-assisted enzymolysis: adjustment pH of suspension 6.0 ~ 7.0, add 3 ~ 5% cellulases of wheat stalk quality, 2 ~ 3% zytases, enzymolysis under ultrasonic assistant effect, hydrolysis temperature 50 ~ 60 DEG C, ultrasonic power 600W, period adopts stirrer at the uniform velocity to stir, and enzymolysis time is 40 ~ 60min;
(4) centrifugal: enzymolysis terminates rear centrifugal, obtain supernatant liquor a and wheat stalk residue;
(5) bio-fermentation agent preparation: get 8 ~ 10 parts of sorbent materials, 3 ~ 5 parts of monocalcium phosphates, 6 ~ 8 parts of glucose, 10 ~ 15 parts of rice brans, 10 ~ 15 portions of Semen Maydis powder, 3 ~ 5 parts of rapeseed cakes, 10 ~ 15 parts of wheat stalks, 2 ~ 3 parts of Trichodermareeseis, 1 ~ 2 part of aspergillus niger, 1-2 part subtilis, mixing and stirring; Then moisture is adjusted to 35 ~ 40%, anaerobically fermenting 7 ~ 10d at 25 DEG C ~ 30 DEG C with the aqueous solution containing 2% molasses;
(6) batch mixing: wheat stalk residue 100 parts, bio-fermentation agent 3 ~ 5 parts, cellulase 0.5 ~ 1 part, zytase 0.5 ~ 1 part of ratio mix in mass ratio, adjustment moisture to 35 ~ 40%;
(7) anaerobically fermenting: by mixed mixture temperature control anaerobically fermenting 2 ~ 3d, temperature 28 ~ 32 DEG C, periodic agitation;
(8) ultrasound-assisted enzymolysis: add 3 ~ 5 times of water after fermentation ends in fermentation residue, add the zytase of fermentation residue mass fraction 2 ~ 3%, enzymolysis under ultrasonic assistant effect, hydrolysis temperature 50 ~ 60 DEG C, ultrasonic power 300W, period adopts stirrer at the uniform velocity to stir, and enzymolysis time is 20 ~ 30min;
(9) centrifugal: enzymolysis terminates rear excessively centrifugal, centrifugal rotational speed 3000 ~ 5000rpm, separation of supernatant b and solid residue;
(10) concentrate drying: merge supernatant liquor a, b, concentrated, vacuum-drying obtains forulic acid crude product;
(11) two step alcohol extractings: to solid residue 3 ~ 5 times of dehydrated alcohols, alcohol extracting 20 ~ 30min under ultrasonic assistant effect, ultrasonic power 300W, period adopts stirrer at the uniform velocity to stir;
(12) centrifugal: alcohol extracting terminates rear centrifugal, centrifugal rotational speed 3000 ~ 5000rpm, is separated to obtain alcohol extract and solid residue;
(13) alcohol wash: rinsed 3 ~ 5 times by above-mentioned solid residue dehydrated alcohol, merges alcohol extract;
(14) purifying: the alcohol extract that the forulic acid crude product step 13 step 10 obtained obtains dissolves, centrifugal removing precipitation, obtains transparent faint yellow supernatant;
(15) concentrate drying: supernatant concentration, vacuum-drying are obtained the higher forulic acid of purity.
2. a kind of biological process according to claim 1 extracts the method for forulic acid in wheat stalk, and the sorbent material that it is characterized in that in step (5) is one or more in zeolite, gac, wilkinite, vermiculite.
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| CN108084017A (en) * | 2017-12-31 | 2018-05-29 | 青岛嘉瑞生物技术有限公司 | A kind of method for improving compound Chinese herb forulic acid extracting solution oxidation resistance |
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| WO2023191756A1 (en) * | 2022-03-29 | 2023-10-05 | Akdeniz Universitesi Doner Sermaye Isletme Mudurlugu | Production of skin friendly hand disinfectant with ferulic acid obtained from wheat bran by supercritical fluid extraction method using extracts from valuable plants such as syzygium aromaticum (clove), lavendula (lavender), thymus (thyme), citrus (orange, lemon) |
| CN118325578A (en) * | 2024-04-12 | 2024-07-12 | 慢半步(重庆)新材料科技有限公司 | Floor tile anti-slip agent and preparation method thereof |
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| CN118325578A (en) * | 2024-04-12 | 2024-07-12 | 慢半步(重庆)新材料科技有限公司 | Floor tile anti-slip agent and preparation method thereof |
| CN118325578B (en) * | 2024-04-12 | 2024-09-27 | 慢半步(重庆)新材料科技有限公司 | Floor tile anti-slip agent and preparation method thereof |
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