CN104313085A - Method for extracting sesame leaf polysaccharose from sesame leaves through biological enzyme method - Google Patents
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Abstract
The invention belongs to the technical field of polysaccharose extraction and particularly relates to a method for extracting sesame leaf polysaccharose from sesame leaves through a biological enzyme method. The method for extracting the sesame leaf polysaccharose from the sesame leaves through the biological enzyme method comprises the following steps: (1) pretreatment of raw materials; (2), enzymolysis; (3), non water soluble polysaccharose conversion; (4), centrifugal separation; (5), soluble polysaccharose extraction. Due to the adoption of zymosis, conditions are mild, enzymolysis to fat and cellulose in the sesame leaves is performed by using different enzymes, so that polysaccharose is extracted, the utilization ratio of the sesame leaves is improved, and after the adoption of the method provided by the invention, the sesame leaf polysaccharose obtained is not only high in yield, but also high in purity.
Description
Technical field
The invention belongs to technical field of polysaccharide extraction, be specifically related to a kind of method that biological enzyme extracts sesame leaf polysaccharide from sesame leaf.
background technology
Sesame leaf is pedaliaceae annual herb plant sesame, the flat bitter of its property, has nourishing liver, kidney, moisturizes laxation function, can treat the illnesss such as post disease alopecia, depletion of body fluid causing lack of blood, constipation, acute and chronic pharyngitis.Sesame leaf abounds with in Huanghuai Area.In recent years, along with molecular biological development, people recognize that polysaccharide and protein, nucleic acid are the same gradually, are one of three class biomacromolecules relevant to vital movement.The functions such as polysaccharide has and improves immunological competence, hypoglycemic, antitumor, antiviral are the focuses of pharmaceutical research.
Oxyradical causes cell injury, human senility and the pathogenetic one of the main reasons of some disease.At present conventional food antioxidant is synthetics mostly, if synthetized oxidation preventive agent improper use, can cause food safety question, and what have even causes deformity, carcinogenic.Indivedual antioxidant is used by some country's forbiddings or restriction, and natural antioxidants causes the concern of people because having the advantage such as safe, nontoxic, finds efficiently, health needs that natural antioxidants meets modern, has Development volue.At present flavonoid compound aspect is mainly concentrated on to the research of sesame leaf, report is rarely had to the research of sesame leaf polysaccharide.
The method mainly water extraction extraction process of current extraction sesame leaf polysaccharide, dry sesame leaf pulverizes → sieves → ether defatting dry degreasing sample → add appropriate pure water → 75 DEG C leaching 45 minutes → centrifugal → precipitation adding distil water carry out second time extract → merge in three centrifuged supernatant → activated carbon decolorizing → concentrating under reduced pressure → Sevage method deproteinated → deproteinated concentrated solutions that to add its volume fraction of ethanol → make be 80% → precipitate and use dehydrated alcohol successively, acetone and washed with diethylether → dry Crude polysaccharides sample → dry to obtain sesame leaf polyoses extract with water-ethanol recrystallization purifying 2 times → 60 DEG C.It is not high that above-mentioned method extracts the many yield obtained, and its yield is 4.32%.
Therefore, need to improve for above-mentioned method, design a kind of reaction conditions gentle and the method extracting sesame polysaccharide from sesame leaf of its extraction yield can be improved.
Summary of the invention
In order to solve above-mentioned technical problem, the invention provides a kind of method that yield and the higher biological enzyme of purity extract sesame leaf polysaccharide from sesame leaf.
The method that biological enzyme of the present invention extracts sesame leaf polysaccharide from sesame leaf is realized by following technical scheme:
Biological enzyme extracts a method for sesame leaf polysaccharide from sesame leaf, comprises following step:
(1) raw materials pretreatment
Sesame leaf drying to its moisture content is 2-4%, then micronizing is to 800-1000 order;
(2) enzymolysis
By the sesame leaf after pulverizing, add water mixing, the part by weight of sesame leaf and water is 1:6-12, and adds lipase, the addition 0.3-0.5% of lipase, hydrolysis temperature 45 DEG C, adjust pH 5.5, above-mentioned raw material is placed in Microwave Extraction Equipment and extracts, the power of microwave is 400-600W, frequency is 800MHZ, enzymolysis 0.1-0.5h;
Add cellulase again, the addition of cellulase is 0.4-0.6%, hydrolysis temperature 40 DEG C, pH value 5, above-mentioned raw material is placed in Microwave Extraction Equipment and extracts, and the power of microwave is 400-600W, and frequency is 800MHZ, enzymolysis 0.1-0.5h;
Add ficin and neutral protease again, the addition of ficin is 0.3-0.8%, the addition of neutral protease is 0.2-0.5%, hydrolysis temperature is 50 DEG C, and above-mentioned raw material is placed in Microwave Extraction Equipment and extracts by pH value 6.8-7.2, the power of microwave is 400-600W, frequency is 800MHZ, enzymolysis 0.1-0.5h, and go out after enzymolysis enzyme 5-10min at 100-105 DEG C;
(3) water-insoluble polysaccharide conversion
Diastatic addition is 1.0-1.2%, hydrolysis temperature 40 DEG C, pH value 5, after enzymolysis 1.5-2.0h, and the 100-105 DEG C of enzyme 5-10min that goes out;
(4) centrifugation
By the sesame leaf after enzymolysis, centrifugal 10-20 min, centrifugal rotational speed is 4500 rpm, supernatant liquor and precipitation is collected respectively;
(5) extraction of soluble polysaccharide
The supernatant liquid filtering got in step (4) removes slag, the filtrate obtained after filtration is concentrated at 60-70 DEG C, being concentrated into its concentration is 60-70%, concentrated solution adds in 1:4 ratio that 95% alcohol settling leaves standstill 10-20 hour, centrifugal 10-20 min, centrifugal rotational speed is 4500 rpm, and collecting precipitation also cleans 2-3 time repeatedly with ethanol, and lyophilize to moisture content is after 4-6%, pulverize at-1-4 DEG C again, obtain soluble polysaccharide;
Above-mentioned enzyme concentration is the weight percent that enzyme accounts for the sesame leaf after micronizing.
Preferably, the consumption of lipase is 0.4%.
Preferably, the consumption of cellulase is 0.5%.
Preferably, diastatic addition is 1.0%.
Preferably, a kind of biological enzyme extracts the method for sesame leaf polysaccharide from sesame leaf, comprises following step:
(1) raw materials pretreatment
Sesame leaf drying to its moisture content is 2-4%, then micronizing to 800 order;
(2) enzymolysis
By the sesame leaf after pulverizing, add water mixing, the part by weight of sesame leaf and water is 1:8, and adds lipase, the addition 0.4% of lipase, hydrolysis temperature 45 DEG C, adjust pH 5.5, above-mentioned raw material is placed in Microwave Extraction Equipment and extracts, the power of microwave is 500W, frequency is 800MHZ, enzymolysis 0.2h;
Add cellulase again, the addition of cellulase be 0.5%, hydrolysis temperature 40 DEG C, pH value 5, above-mentioned raw material is placed in Microwave Extraction Equipment and extracts, the power of microwave is 500W, and frequency is 800MHZ, enzymolysis 0.2h;
Add ficin and neutral protease again, the addition of ficin is 0.5%, the addition of neutral protease is 0.4%, hydrolysis temperature is 50 DEG C, and above-mentioned raw material is placed in Microwave Extraction Equipment and extracts by pH value 7.0, the power of microwave is 500W, frequency is 800MHZ, enzymolysis 0.2h, and go out after enzymolysis enzyme 8min at 100 DEG C;
(3) water-insoluble polysaccharide conversion
Diastatic addition is 1%, hydrolysis temperature 40 DEG C, pH value 5, after enzymolysis 1.8h, and 100 DEG C of enzyme 6min that go out;
(4) centrifugation
By the sesame leaf after enzymolysis, centrifugal 15 min, centrifugal rotational speed is 4500 rpm, supernatant liquor and precipitation is collected respectively;
(5) extraction of soluble polysaccharide
The supernatant liquid filtering got in step (4) removes slag, the filtrate obtained after filtration is concentrated at 65 DEG C, being concentrated into its concentration is 65%, concentrated solution adds 95% alcohol settling in 1:4 ratio and leaves standstill 15 hours, centrifugal 15 min, centrifugal rotational speed is 4500 rpm, and collecting precipitation also cleans 2-3 time repeatedly with ethanol, and lyophilize to moisture content is after 5%, pulverize at-1-4 DEG C again, obtain soluble polysaccharide;
Above-mentioned enzyme concentration is the weight percent that enzyme accounts for the sesame leaf after micronizing.
In method of the present invention, first sesame leaf is pulverized, adopt the method for micronizing, instead of common grinding mode, because after micronizing, the particle diameter of sesame leaf is less, be conducive in follow-up enzyme digestion reaction process, various component contacts with enzyme, makes enzyme better act on each component;
Enzymolysis fat, fiber order are also innovations of the present invention, and due to by after fatty enzymolysis, grease can not be wrapped in outside sesame leaf, thus are conducive to various composition and contact with enzyme, thus the efficiency of raising enzymolysis; Therefore, the fatty enzymolysis Mierocrystalline cellulose again of first enzymolysis is conducive to the removal of fat and fiber;
Generally adopt the mode of organic solvent of adding to the treatment process of fat in usual way, although organic solvent is also comparatively thorough to the removal of grease, but the condition of its effect is stronger, and follow-up removal process is also more complicated, also easily causes the environmentally hazardous problem of organic solvent in addition; Organic solvent is adopted to remove its cost of grease in addition also higher;
The present invention adopts enzyme to act on to remain on the grease in sesame leaf, and action condition is gentle, and also comparatively thorough to the removal of grease, and there is not the problem that follow-up organic solvent reclaims and bring environmental pollution;
Add cellulase again and the cellulose degradation in sesame leaf is generated water-soluble glucose;
After the polysaccharide enzymolysis of non-solubility is soluble polysaccharide, carbohydrate is soluble in water, then gets supernatant liquor, filters, concentrated, alcohol precipitation, by Polyose extraction wherein out.
The selection of the kind of enzyme of the present invention and the selection of consumption and enzymolysis order is not accidental, but contriver has paid performing creative labour obtains, the adjustment of enzyme and various ratio all can affect the extraction effect of final polysaccharide, only have adopt the present invention to enzyme and the corresponding proportion of enzyme sesame leaf is processed, and carry out according to order of the present invention, just can obtain result of the present invention, enzyme be carried out replace or the replacement of enzyme-added order, all can not get maximum polysaccharide extract rate.
Beneficial effect of the present invention is, adopts enzyme action condition gentle, adopts various different enzyme by the fat in sesame leaf, cellulase hydrolysis, from sesame leaf, extract polysaccharide simultaneously, utilize sesame leaf to greatest extent, and the polysaccharide adopting method of the present invention to obtain, its yield and purity high.
Embodiment
Below in conjunction with specific embodiment, the present invention is further described, so that those skilled in the art more understands the present invention, but does not therefore limit the present invention.
Following detection method, if no special instructions, adopts phend-sulphuric acid to survey the content of polysaccharide.
Embodiment 1
Biological enzyme extracts a method for sesame leaf polysaccharide from sesame leaf, comprises following step:
(1) raw materials pretreatment
Sesame leaf drying to its moisture content is 3%, then micronizing to 800 order;
(2) enzymolysis
By the sesame leaf after pulverizing, add water mixing, the part by weight of sesame leaf and water is 1:8, and adds lipase, the addition 0.4% of lipase, hydrolysis temperature 45 DEG C, adjust pH 5.5, above-mentioned raw material is placed in Microwave Extraction Equipment and extracts, the power of microwave is 500W, frequency is 800MHZ, enzymolysis 0.2h;
Add cellulase again, the addition of cellulase be 0.5%, hydrolysis temperature 40 DEG C, pH value 5, above-mentioned raw material is placed in Microwave Extraction Equipment and extracts, the power of microwave is 500W, and frequency is 800MHZ, enzymolysis 0.2h;
Add ficin and neutral protease again, the addition of ficin is 0.5%, the addition of neutral protease is 0.4%, hydrolysis temperature is 50 DEG C, and above-mentioned raw material is placed in Microwave Extraction Equipment and extracts by pH value 7.0, the power of microwave is 500W, frequency is 800MHZ, enzymolysis 0.2h, and go out after enzymolysis enzyme 8min at 100 DEG C;
(3) water-insoluble polysaccharide conversion
Diastatic addition is 1%, hydrolysis temperature 40 DEG C, pH value 5, after enzymolysis 1.8h, and 100 DEG C of enzyme 6min that go out;
(4) centrifugation
By the sesame leaf after enzymolysis, centrifugal 15 min, centrifugal rotational speed is 4500 rpm, supernatant liquor and precipitation is collected respectively;
(5) extraction of soluble polysaccharide
The supernatant liquid filtering got in step (4) removes slag, the filtrate obtained after filtration is concentrated at 65 DEG C, being concentrated into its concentration is 65%, concentrated solution adds 95% alcohol settling in 1:4 ratio and leaves standstill 15 hours, centrifugal 15 min, centrifugal rotational speed is 4500 rpm, and collecting precipitation also cleans 2-3 time repeatedly with ethanol, and lyophilize to moisture content is after 5%, pulverize at-1-4 DEG C again, obtain soluble polysaccharide;
Above-mentioned enzyme concentration is the weight percent that enzyme accounts for the sesame leaf after micronizing.
The content adopting phend-sulphuric acid to record wherein polysaccharide is 5.96%.
Embodiment 2
Biological enzyme extracts a method for sesame leaf polysaccharide from sesame leaf, comprises following step:
(1) raw materials pretreatment
Sesame leaf drying to its moisture content is 2%, then micronizing to 800 order;
(2) enzymolysis
By the sesame leaf after pulverizing, add water mixing, the part by weight of sesame leaf and water is 1:6, and adds lipase, the addition 0.3% of lipase, hydrolysis temperature 45 DEG C, adjust pH 5.5, above-mentioned raw material is placed in Microwave Extraction Equipment and extracts, the power of microwave is 400W, frequency is 800MHZ, enzymolysis 0.1h;
Add cellulase again, the addition of cellulase be 0.4%, hydrolysis temperature 40 DEG C, pH value 5, above-mentioned raw material is placed in Microwave Extraction Equipment and extracts, the power of microwave is 400W, and frequency is 800MHZ, enzymolysis 0.1-0.5h;
Add ficin and neutral protease again, the addition of ficin is 0.3%, the addition of neutral protease is 0.2%, hydrolysis temperature is 50 DEG C, and above-mentioned raw material is placed in Microwave Extraction Equipment and extracts by pH value 6.8, the power of microwave is 400W, frequency is 800MHZ, enzymolysis 0.1h, and go out after enzymolysis enzyme 5min at 100 DEG C;
(3) water-insoluble polysaccharide conversion
Diastatic addition is 0.8%, hydrolysis temperature 40 DEG C, pH value 5, after enzymolysis 1.8h, and 100 DEG C of enzyme 6min that go out;
(4) centrifugation
By the sesame leaf after enzymolysis, centrifugal 15 min, centrifugal rotational speed is 4500 rpm, supernatant liquor and precipitation is collected respectively;
(5) extraction of soluble polysaccharide
The supernatant liquid filtering got in step (4) removes slag, the filtrate obtained after filtration is concentrated at 65 DEG C, being concentrated into its concentration is 65%, concentrated solution adds 95% alcohol settling in 1:4 ratio and leaves standstill 15 hours, centrifugal 15 min, centrifugal rotational speed is 4500 rpm, and collecting precipitation also cleans 2-3 time repeatedly with ethanol, and lyophilize to moisture content is after 5%, pulverize at-1-4 DEG C again, obtain soluble polysaccharide;
Above-mentioned enzyme concentration is the weight percent that enzyme accounts for the sesame leaf after micronizing.
The content adopting phend-sulphuric acid to record wherein polysaccharide is 5.63%.
Embodiment 3
Biological enzyme extracts a method for sesame leaf polysaccharide from sesame leaf, comprises following step:
(1) raw materials pretreatment
Sesame leaf drying to its moisture content is 4%, then micronizing to 800 order;
(2) enzymolysis
By the sesame leaf after pulverizing, add water mixing, the part by weight of sesame leaf and water is 1:12, and adds lipase, the addition 0.5% of lipase, hydrolysis temperature 45 DEG C, adjust pH 5.5, above-mentioned raw material is placed in Microwave Extraction Equipment and extracts, the power of microwave is 600W, frequency is 800MHZ, enzymolysis 0.5h;
Add cellulase again, the addition of cellulase be 0.6%, hydrolysis temperature 40 DEG C, pH value 5, above-mentioned raw material is placed in Microwave Extraction Equipment and extracts, the power of microwave is 600W, and frequency is 800MHZ, enzymolysis 0.5h;
Add ficin and neutral protease again, the addition of ficin is 0.8%, the addition of neutral protease is 0.5%, hydrolysis temperature is 50 DEG C, and above-mentioned raw material is placed in Microwave Extraction Equipment and extracts by pH value 7.2, the power of microwave is 600W, frequency is 800MHZ, enzymolysis 0.5h, and go out after enzymolysis enzyme 10min at 100 DEG C;
(3) water-insoluble polysaccharide conversion
Diastatic addition is 1.2%, hydrolysis temperature 40 DEG C, pH value 5, after enzymolysis 1.8h, and 100 DEG C of enzyme 6min that go out;
(4) centrifugation
By the sesame leaf after enzymolysis, centrifugal 15 min, centrifugal rotational speed is 4500 rpm, supernatant liquor and precipitation is collected respectively;
(5) extraction of soluble polysaccharide
The supernatant liquid filtering got in step (4) removes slag, the filtrate obtained after filtration is concentrated at 65 DEG C, being concentrated into its concentration is 65%, concentrated solution adds 95% alcohol settling in 1:4 ratio and leaves standstill 15 hours, centrifugal 15 min, centrifugal rotational speed is 4500 rpm, and collecting precipitation also cleans 2-3 time repeatedly with ethanol, and lyophilize to moisture content is after 5%, pulverize at-1-4 DEG C again, obtain soluble polysaccharide;
Above-mentioned enzyme concentration is the weight percent that enzyme accounts for the sesame leaf after micronizing.
The content adopting phend-sulphuric acid to record wherein polysaccharide is 5.57%.
Claims (5)
1. biological enzyme extracts a method for sesame leaf polysaccharide from sesame leaf, comprises following step:
(1) raw materials pretreatment
Sesame leaf drying to its moisture content is 2-4%, then micronizing is to 800-1000 order;
(2) enzymolysis
By the sesame leaf after pulverizing, add water mixing, the part by weight of sesame leaf and water is 1:6-12, and adds lipase, the addition 0.3-0.5% of lipase, hydrolysis temperature 45 DEG C, adjust pH 5.5, above-mentioned raw material is placed in Microwave Extraction Equipment and extracts, the power of microwave is 400-600W, frequency is 800MHZ, enzymolysis 0.1-0.5h;
Add cellulase again, the addition of cellulase is 0.4-0.6%, hydrolysis temperature 40 DEG C, pH value 5, above-mentioned raw material is placed in Microwave Extraction Equipment and extracts, and the power of microwave is 400-600W, and frequency is 800MHZ, enzymolysis 0.1-0.5h;
Add ficin and neutral protease again, the addition of ficin is 0.3-0.8%, the addition of neutral protease is 0.2-0.5%, hydrolysis temperature is 50 DEG C, and above-mentioned raw material is placed in Microwave Extraction Equipment and extracts by pH value 6.8-7.2, the power of microwave is 400-600W, frequency is 800MHZ, enzymolysis 0.1-0.5h, and go out after enzymolysis enzyme 5-10min at 100-105 DEG C;
(3) water-insoluble polysaccharide conversion
Diastatic addition is 0.8-1.2%, hydrolysis temperature 40 DEG C, pH value 5, after enzymolysis 1.5-2.0h, and the 100-105 DEG C of enzyme 5-10min that goes out; Above-mentioned raw material is placed in Microwave Extraction Equipment extract, the power of microwave is 400-600W, and frequency is 800MHZ, enzymolysis 0.1-0.5h, and go out after enzymolysis enzyme 5-10min at 100-105 DEG C;
(4) centrifugation
By the sesame leaf after enzymolysis, centrifugal 10-20 min, centrifugal rotational speed is 4500 rpm, supernatant liquor and precipitation is collected respectively;
(5) extraction of soluble polysaccharide
The supernatant liquid filtering got in step (4) removes slag, the filtrate obtained after filtration is concentrated at 60-70 DEG C, being concentrated into its concentration is 60-70%, concentrated solution adds in 1:4 ratio that 95% alcohol settling leaves standstill 10-20 hour, centrifugal 10-20 min, centrifugal rotational speed is 4500 rpm, and collecting precipitation also cleans 2-3 time repeatedly with ethanol, and lyophilize to moisture content is after 4-6%, pulverize at-1-4 DEG C again, obtain soluble polysaccharide;
Above-mentioned enzyme concentration is the weight percent that enzyme accounts for the sesame leaf after micronizing.
2. a kind of biological enzyme extracts the method for sesame leaf polysaccharide from sesame leaf as claimed in claim 1, it is characterized in that, the consumption of described lipase is 0.4%.
3. a kind of biological enzyme as claimed in claim 1 extracts the method for sesame leaf polysaccharide from sesame leaf, it is characterized in that, the consumption of described cellulase is 0.5%.
4. a kind of biological enzyme as claimed in claim 1 extracts the method for sesame leaf polysaccharide from sesame leaf, it is characterized in that, described diastatic addition is 1.0%.
5. a kind of biological enzyme according to any one of claim 1-4 extracts the method for sesame leaf polysaccharide from sesame leaf, it is characterized in that, described method comprises following step:
(1) raw materials pretreatment
Sesame leaf drying to its moisture content is 2-4%, then micronizing to 800 order;
(2) enzymolysis
By the sesame leaf after pulverizing, add water mixing, the part by weight of sesame leaf and water is 1:8, and adds lipase, the addition 0.4% of lipase, hydrolysis temperature 45 DEG C, adjust pH 5.5, above-mentioned raw material is placed in Microwave Extraction Equipment and extracts, the power of microwave is 500W, frequency is 800MHZ, enzymolysis 0.2h;
Add cellulase again, the addition of cellulase be 0.5%, hydrolysis temperature 40 DEG C, pH value 5, above-mentioned raw material is placed in Microwave Extraction Equipment and extracts, the power of microwave is 500W, and frequency is 800MHZ, enzymolysis 0.2h;
Add ficin and neutral protease again, the addition of ficin is 0.5%, the addition of neutral protease is 0.4%, hydrolysis temperature is 50 DEG C, and above-mentioned raw material is placed in Microwave Extraction Equipment and extracts by pH value 7.0, the power of microwave is 500W, frequency is 800MHZ, enzymolysis 0.2h, and go out after enzymolysis enzyme 8min at 100 DEG C;
(3) water-insoluble polysaccharide conversion
Diastatic addition is 1.0%, hydrolysis temperature 40 DEG C, pH value 5, after enzymolysis 1.8h, and 100 DEG C of enzyme 6min that go out;
(4) centrifugation
By the sesame leaf after enzymolysis, centrifugal 15 min, centrifugal rotational speed is 4500 rpm, supernatant liquor and precipitation is collected respectively;
(5) extraction of soluble polysaccharide
The supernatant liquid filtering got in step (4) removes slag, the filtrate obtained after filtration is concentrated at 65 DEG C, being concentrated into its concentration is 65%, concentrated solution adds 95% alcohol settling in 1:4 ratio and leaves standstill 15 hours, centrifugal 15 min, centrifugal rotational speed is 4500 rpm, and collecting precipitation also cleans 2-3 time repeatedly with ethanol, and lyophilize to moisture content is after 5%, pulverize at-1-4 DEG C again, obtain soluble polysaccharide;
Above-mentioned enzyme concentration is the weight percent that enzyme accounts for the sesame leaf after micronizing.
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Citations (1)
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| CN103467612A (en) * | 2013-09-09 | 2013-12-25 | 山东省农业科学院农产品研究所 | Method for synchronously extracting polysaccharides and proteins from high-temperature peanut meal |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN103467612A (en) * | 2013-09-09 | 2013-12-25 | 山东省农业科学院农产品研究所 | Method for synchronously extracting polysaccharides and proteins from high-temperature peanut meal |
Non-Patent Citations (2)
| Title |
|---|
| 岳燕霞等: "芝麻叶多糖提取工艺研究", 《农产品加工》 * |
| 许春平等: "植物叶多糖的提取和生物活性综述", 《食品研究与开发》 * |
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