CN104177508A - Method for comprehensively extracting tea seed saponin, tea seed polypeptide and tea seed polysaccharide from tea seed cake - Google Patents
Method for comprehensively extracting tea seed saponin, tea seed polypeptide and tea seed polysaccharide from tea seed cake Download PDFInfo
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Abstract
The invention discloses a method for comprehensively extracting tea seed saponin, tea seed polypeptide and tea seed polysaccharide by using tea seed cake as a raw material. The extracting method comprises the steps of mixing degreased tea seed cake with ethanol, treating with a colloid mill and a homogenizer, then performing water bath ethanol extraction, centrifuging, performing ultra-filtration, and concentrating to obtain the tea seed saponin; mixing the centrifuged sediment with water, extracting with ultrasonic and microwave, centrifuging, regulating the pH of the supernate, centrifuging for the second time, dissolving the sediment, and performing enzymolysis, enzyme deactivation, ultra-filtration and spraying to obtain the tea seed polypeptide; and spraying the supernate obtained by secondary centrifuging to obtain coarse tea seed polysaccharide, and then performing ultra-filtration and ethanol precipitation to obtain the high-purity tea seed polysaccharide. The method is simple, low in production and equipment cost, high in product extraction rate and high in purity, and greatly improves the additional value of the tea seed cake. A new thought and a new method are provided for efficient comprehensive utilization of the tea seed cake.
Description
Technical field
The present invention relates to a kind of method of extracting biologically active substance from leached tea oil slag.
Background technology
Thea saponin claims again theasaponin, TS, the class in saponin.Research finds that thea saponin has haemolysis and ichthyotoxin, pest-resistant germicidal action, impervious anti-inflammatory, preventing phlegm from forming and stopping coughing, analgesia, the pharmacological function such as anticancer.Thea saponin is also a kind of good natural nonionic surface active agent with stronger foaming, emulsification, dispersion, the effect such as moistening.Can be widely used in the fields such as daily use chemicals, building materials, weaving, agricultural chemicals, there is higher economic worth.
Tea seed polypeptide is easily digested, has anti-oxidant, hypotensive, to reduce cholesterol level, promotion calcium absorption physiological function.Tea seed polysaccharide is a kind of macromolecular compound (molecular weight 40,000-100,000) of a kind of similar ganoderan and panaxan.Modern medicine study is found; tea seed polysaccharide has the effect of " 2 anti-3 fall 4 anti-", i.e. kinds of protect blood phases such as radioprotective, prevention and cure of cardiovascular disease, hypoglycemic, reducing blood-fat, hypotensive, anticancer, anti-freezing, antithrombotic, atherosclerosis and strengthen effect of human body non-specific immunity.
Having thea saponin content in bibliographical information leached tea oil slag is that 11-17%, crude protein content are that 15-18%, polysaccharide content are 45-49%.According to incompletely statistics, the output of the annual camellia seed meal of China exceedes 500,000 tons, but does not obtain for a long time comprehensive reasonable utilization, has not only caused a large amount of wastes of resource also to pollute environment.In recent years, leached tea oil slag is developed and day by day causes various places great attention, but how from leached tea oil slag, comprehensively to extract thea saponin, tea seed polypeptide, tea seed polysaccharide, and be purified and obtain that a class is of many uses, the high-quality product of excellent performance, particularly important to solving the comprehensive utilization of leached tea oil slag.
At present not about the report of comprehensively preparing thea saponin, tea seed polypeptide, tea seed polysaccharide from leached tea oil slag.Therefore, be badly in need of a kind of method that can maximized raising leached tea oil slag added value, thereby realize the comprehensive Development and Production of leached tea oil slag product.
Summary of the invention
The present invention aims to provide a kind of method that can comprehensively prepare thea saponin, tea seed polypeptide, tea seed polysaccharide from leached tea oil slag, to make up the deficiencies in the prior art, improves its purity simultaneously.The method technique is simple, and production cost is low, and product extraction yield is high, purity is high, quality is good.
Its concrete technology scheme is as follows:
From leached tea oil slag, a comprehensive method of extracting thea saponin, tea seed polypeptide, tea seed polysaccharide, is characterized in that comprising the steps:
(1) leached tea oil slag is crushed to 300-400 order, it is carried out to skimming treatment with sherwood oil;
(2) thea saponin extracts: it is miscible that the leached tea oil slag in will (1) after degreasing and dehydrated alcohol are pressed mass volume ratio g/mL 1:2-1:6, and 50 DEG C of water-baths extraction 2h, extract 6 times; Centrifugal, get the filter of supernatant liquor supermembrane, the thea saponin solution that trapped fluid is purifying, is concentrated into the fast completely dissolve of ethanol, and heating water dissolves, and freeze-drying, obtains thea saponin;
(3) tea seed polypeptide extracts: the centrifugal precipitation obtaining in (2) is dried naturally, add water by solid-liquid ratio g/mL 1:2; Adjust pH to 10.0 with the NaOH solution of 1moL/L, adopt ultrasonic-microwave double wave assisted extraction 2 times, extract 2h 1 time, ultrasonic wave and microwave power are 200-400w; Centrifugal, get supernatant liquor, adjust pH to 4.5, for the second time centrifugal must precipitation and supernatant liquor; To precipitate soluble in waterly, adding trypsinase amount is 0.6% of substrate again, enzymolysis 2.5h, the boiling water bath enzyme 6-10min that goes out; Adopt supermembrane filter, trapped fluid is the polypeptide solution after purifying, and spraying is dried to obtain tea seed polypeptide;
(4) tea seed polysaccharide extracts: will be in (3) for the second time the centrifugal supernatant liquor obtaining spray to be dried and to obtain slightly tea polysaccharide; Be dissolved in dehydrated alcohol by solid-liquid ratio 1:4 (g/mL), supermembrane filter, trapped fluid is the tea seed polysaccharide solution after purifying, precipitation 12h, precipitation freeze-drying is tea seed polysaccharide.
In order to reach better effect, before 2h is extracted in (2) 50 DEG C of water-baths of described step, first cross colloidal mill and 40Mpa clarifixator;
Step (3) ultrasonic wave and microwave best power are 600w;
Step (2) supermembrane filter is specially and selects molecular weight cut-off is that the ultra-filtration membrane of 2000Da carries out separation and purification, and tangential flow velocity is 4L/ m
2min, pressure is 60psi;
Step (3) supermembrane filter is specially and selects molecular weight cut-off is that the ultra-filtration membrane of 3000Da carries out separation and purification, and tangential flow velocity is 6L/ m
2min, pressure is 60psi;
Step (4) supermembrane filter is specially and selects molecular weight cut-off is that the ultra-filtration membrane of 50000Da carries out separation and purification, and tangential flow velocity is, 8L/ m
2min, pressure is 60psi;
182 DEG C of the dry inlet temperature of step (3) spraying, 80～100 DEG C of air outlet temperatures;
182 DEG C of the dry inlet temperature of step (4) spraying, 80～100 DEG C of air outlet temperatures.
The present invention has the following advantages:
(1) realize the comprehensive preparation of thea saponin, tea seed polypeptide, tea seed polysaccharide in leached tea oil slag, raw material has been carried out to maximized comprehensive utilization, improved the added value of leached tea oil slag;
(2) adopt ultrasonic-microwave double wave ancillary technique to prepare tea seed polypeptide and tea seed polysaccharide, and adopted ultra-filtration membrane filter technology purifying, greatly improved yield and the purity of tea seed polypeptide and tea seed polysaccharide.Wherein, the extraction yield of thea saponin is 96.3%, and purity is 95.1%, and the extraction yield of tea seed polypeptide is 93.6%, and purity is 98.6%, and tea seed polysaccharide extraction yield is 95.7%, and purity is 95.3%.
Embodiment
Embodiment mono-
Tea seed cake was pulverized 300-400 mesh sieve, and petroleum ether degreasing adds dehydrated alcohol after processing, solid-liquid ratio is 1:2(g/mL), cross each 2 times of colloidal mill (JM-LB) and 40Mpa clarifixator (SHP4000-25), extract 2h in 50 DEG C of water-baths, extract 6 times.Centrifugal, getting supernatant liquor, to select molecular weight cut-off be that the ultra-filtration membrane of 2000Da carries out separation and purification, and tangential flow velocity is 4L/ m
2min, pressure is 60psi, the thea saponin solution that trapped fluid is purifying is concentrated into the fast completely dissolve of ethanol, adds hot water dissolving, freeze-drying.Gained thea saponin yield is 92.1%, and purity is 90.3%;
Naturally dry extracting thea saponin leached tea oil slag afterwards, by solid-liquid ratio 1:2(g/mL) add water, with NaOH solution tune pH to 10.0 left and right of 1MoL/L, adopt ultrasonic-microwave double wave (XH-300A, under the auspicious swan in Beijing with) auxiliary extraction technology extract 2 times, extract 2h 1 time, ultrasonic wave and microwave power are 200w.Centrifugal, get supernatant liquor, adjust pH to 4.5 left and right, centrifugal must precipitation and supernatant liquor.Precipitate soluble in waterly again, adjust pH to 4.5 left and right, adding trypsinase amount is 0.6% of substrate, enzymolysis 2h, the boiling water bath enzyme that goes out.Adopt the ultra-filtration membrane that molecular weight cut-off is 3000Da to carry out separation and purification, tangential flow velocity is 6L/ m
2min, pressure is 60psi, and trapped fluid is the polypeptide solution after purifying, and spraying is dried to obtain tea polypeptide, 182 DEG C of inlet temperature, 80～100 DEG C of air outlet temperatures, flow is depending on air outlet temperature.Gained tea seed polypeptide yield is 90.1%, and purity is 91.1%;
Centrifugal for the second time supernatant liquor is sprayed and is dried to obtain thick tea seed polysaccharide, 182 DEG C of inlet temperature, 80～100 DEG C of air outlet temperatures, flow is depending on air outlet temperature.Add dehydrated alcohol by solid-liquid ratio 1:1.5 (g/mL), adopt the ultra-filtration membrane that molecular weight cut-off is 50000Da to carry out separation and purification, tangential flow velocity is 8L/ m
2min, pressure is 60psi, and trapped fluid is the tea polysaccharide solution after purifying, leaves standstill 12h, and precipitation freeze-drying had both obtained tea seed polysaccharide.Gained tea seed polysaccharide yield is 90.0%, and purity is 90.1%.
Embodiment bis-
Tea seed cake was pulverized 300-400 mesh sieve, and petroleum ether degreasing adds dehydrated alcohol after processing, solid-liquid ratio is 1:4(g/mL), cross each 2 times of colloidal mill (JM-LB) and 40Mpa clarifixator (SHP4000-25), extract 2h in 50 DEG C of water-baths, extract 6 times.Centrifugal, getting supernatant liquor, to select molecular weight cut-off be that the ultra-filtration membrane of 2000Da carries out separation and purification, and tangential flow velocity is 4L/ m
2min, pressure is 60psi, the thea saponin solution that trapped fluid is purifying is concentrated into the fast completely dissolve of ethanol, adds hot water dissolving, freeze-drying.Gained thea saponin yield is 96.3%, and purity is 95.1%;
Naturally dry extracting thea saponin leached tea oil slag afterwards, by solid-liquid ratio 1:2(g/mL) add water, with NaOH solution tune pH to 10.0 left and right of 1MoL/L, adopt ultrasonic-microwave double wave (XH-300A) auxiliary extraction technology to extract 2 times, extract 2h 1 time, ultrasonic and microwave power is 300w.Centrifugal, get supernatant liquor, adjust pH to 4.5 left and right, centrifugal must precipitation and supernatant liquor.Precipitate soluble in waterly again, adjust pH to 4.5 left and right, adding trypsinase amount is 0.6% of substrate, enzymolysis 2.5h, the boiling water bath enzyme that goes out.Adopt the ultra-filtration membrane that molecular weight cut-off is 3000Da to carry out separation and purification, tangential flow velocity is 6L/ m
2min, pressure is 60psi, and trapped fluid is the polypeptide solution after purifying, and spraying is dried to obtain tea seed polypeptide, 182 DEG C of inlet temperature, 80～100 DEG C of air outlet temperatures, flow is depending on air outlet temperature.Gained tea polypeptide yield is 93.6%, and purity is 98.6%;
Centrifugal for the second time supernatant liquor is sprayed and is dried to obtain thick tea polysaccharide, 182 DEG C of inlet temperature, 80～100 DEG C of air outlet temperatures, flow is depending on air outlet temperature.Add dehydrated alcohol by solid-liquid ratio 1:2 (g/mL), adopt the ultra-filtration membrane that molecular weight cut-off is 50000Da to carry out separation and purification, tangential flow velocity is 8L/ m
2min, pressure is 60psi, and trapped fluid is the tea seed polysaccharide solution after purifying, leaves standstill 12h, and precipitation freeze-drying had both obtained tea seed polysaccharide.Gained tea polysaccharide yield is 95.7%, and purity is 95.3%.
Embodiment tri-
Tea seed cake was pulverized 300-400 mesh sieve, and petroleum ether degreasing adds dehydrated alcohol after processing, and solid-liquid ratio is 1:6(g/mL), process with colloidal mill (JM-LB) and clarifixator (SHP4000-25), extract 2h in 50 DEG C of water-baths, extract 6 times.Centrifugal, getting supernatant liquor, to select molecular weight cut-off be that the ultra-filtration membrane of 2000Da carries out separation and purification, and tangential flow velocity is 4L/ m
2min, pressure is 60psi, the thea saponin solution that trapped fluid is purifying is concentrated into the fast completely dissolve of ethanol, adds hot water dissolving, freeze-drying.Gained thea saponin yield is 89.1%, and purity is 90.1%;
Naturally dry extracting tea saponin leached tea oil slag afterwards, by solid-liquid ratio 1:2(g/mL) add water, with NaOH solution tune pH to 10.0 left and right of 1MoL/L, adopt ultrasonic-microwave double wave (XH-300A) auxiliary extraction technology to extract 2 times, extract 2h 1 time, ultrasonic wave and microwave power are 400w.Centrifugal, get supernatant liquor, adjust pH to 4.5 left and right, centrifugal must precipitation and supernatant liquor.Precipitate soluble in waterly again, adjust pH to 4.5 left and right, adding trypsinase amount is 0.6% of substrate, enzymolysis 3h, the boiling water bath enzyme that goes out.Adopt the ultra-filtration membrane that molecular weight cut-off is 3000Da to carry out separation and purification, tangentially flow velocity is, 6L/ m
2min, pressure is 60psi, and trapped fluid is the polypeptide solution after purifying, and spraying is dried to obtain tea seed polypeptide, 182 DEG C of inlet temperature, 80～100 DEG C of air outlet temperatures, flow is depending on air outlet temperature.Gained tea seed polypeptide yield is 89.8%, and purity is 90%;
Centrifugal for the second time supernatant liquor is sprayed and is dried to obtain thick tea polysaccharide, 182 DEG C of inlet temperature, 80～100 DEG C of air outlet temperatures, flow is depending on air outlet temperature.By solid-liquid ratio 1:2.5(g/mL) add dehydrated alcohol, adopt the ultra-filtration membrane that molecular weight cut-off is 50000Da to carry out separation and purification, tangentially flow velocity is, 8L/ m
2min, pressure is 60psi, and trapped fluid is the tea polysaccharide solution after purifying, leaves standstill 12h, and precipitation freeze-drying had both obtained tea seed polysaccharide.Gained tea seed polysaccharide yield is 90.4%, and purity is 91.3%.
Claims (8)
1. a comprehensive method of extracting thea saponin, tea seed polypeptide, tea seed polysaccharide from leached tea oil slag, is characterized in that comprising the steps:
(1) leached tea oil slag is crushed to 300-400 order, it is carried out to skimming treatment with sherwood oil;
(2) thea saponin extracts: it is miscible that the leached tea oil slag in will (1) after degreasing and dehydrated alcohol are pressed mass volume ratio g/mL 1:2-1:6, and 50 DEG C of water-baths extraction 2h, extract 6 times; Centrifugal, get the filter of supernatant liquor supermembrane, the thea saponin solution that trapped fluid is purifying, is concentrated into the fast completely dissolve of ethanol, and heating water dissolves, and freeze-drying, obtains thea saponin;
(3) tea seed polypeptide extracts: the centrifugal precipitation obtaining in (2) is dried naturally, add water by solid-liquid ratio g/mL 1:2; Adjust pH to 10.0 with the NaOH solution of 1moL/L, adopt ultrasonic-microwave double wave assisted extraction 2 times, extract 2h 1 time, ultrasonic wave and microwave power are 200-400w; Centrifugal, get supernatant liquor, adjust pH to 4.5, for the second time centrifugal must precipitation and supernatant liquor; To precipitate soluble in waterly, adding trypsinase amount is 0.6% of substrate again, enzymolysis 2.5h, the boiling water bath enzyme 6-10min that goes out; Adopt supermembrane filter, trapped fluid is the polypeptide solution after purifying, and spraying is dried to obtain tea seed polypeptide;
(4) tea seed polysaccharide extracts: will be in (3) for the second time the centrifugal supernatant liquor obtaining spray to be dried and to obtain slightly tea polysaccharide; Be dissolved in dehydrated alcohol by solid-liquid ratio 1:4 (g/mL), supermembrane filter, trapped fluid is the tea seed polysaccharide solution after purifying, precipitation 12h, precipitation freeze-drying is tea seed polysaccharide.
2. method according to claim 1, before it is characterized in that 2h is extracted in (2) 50 DEG C of water-baths of described step, first crosses colloidal mill and 40Mpa clarifixator.
3. method according to claim 1, is characterized in that described step (3) ultrasonic wave and microwave best power are 600w.
4. method according to claim 1, is characterized in that described step (2) supermembrane filter is specially that to select molecular weight cut-off be that the ultra-filtration membrane of 2000Da carries out separation and purification, and tangential flow velocity is 4L/ m
2min, pressure is 60psi.
5. method according to claim 1, is characterized in that described step (3) supermembrane filter is specially that to select molecular weight cut-off be that the ultra-filtration membrane of 3000Da carries out separation and purification, and tangential flow velocity is 6L/ m
2min, pressure is 60psi.
6. method according to claim 1, is characterized in that described step (4) supermembrane filter is specially that to select molecular weight cut-off be that the ultra-filtration membrane of 50000Da carries out separation and purification, and tangential flow velocity is, 8L/ m
2min, pressure is 60psi.
7. method according to claim 1, is characterized in that 182 DEG C of the dry inlet temperature of described step (3) spraying, 80～100 DEG C of air outlet temperatures.
8. method according to claim 1, is characterized in that 182 DEG C of the dry inlet temperature of described step (4) spraying, 80～100 DEG C of air outlet temperatures.
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