Summary of the invention:
The present invention adopts the biological wall breaking method; Anticipating substitute is rolled in Mierocrystalline cellulose, semicellulose, pectin segment and the albuminous membranae of seed skin shell; Make the oil cells that is wrapped dissociate out; Combine the microwave treatment method again, remove the fat in the seed fast, and degreasing more thoroughly obtains deciding the oily novel method of Silymarin of content.
Silybum marianum seed after the de-oiling is made solvent through microwave reflux extracting device with ethanol, removes albumen in iso-electric point, after macroporous resin adsorption, concentrates, dryly obtains highly purified SLB.The inventive method can obtain Silymarin oil and two products of SLB simultaneously.Be different from traditional cold press oil removing, ethyl acetate extraction is had to the method for single SLB.Shortcomings such as SLB routine techniques extraction yield is relatively low, the running time is long, extracting substance is single have been overcome.
For realizing above-mentioned purpose, the technical scheme that the present invention adopts is:
A kind of to utilize biological wall breaking, Microwave Pretreatment to get oily, extracts and the macroporous resin purification technology through microwave counter current again, obtains the method for high purity SLB.Its step is following:
(1) biological wall breaking: dry silybum marianum seed soaks; Add an amount of cellulase, semicellulose enzyme solution, insulation enzymolysis 60min removes the Mierocrystalline cellulose, semicellulose, pectin segment and the albuminous membranae that are wrapped in seed skin shell; Make the oil cells that is wrapped dissociate out, improve the grease extraction yield.
(2) Microwave Pretreatment: silybum marianum seed was pulverized the 30-60 mesh sieve; Add 1-6 times of sherwood oil (boiling range 60-90 ℃) solution, carry out Microwave Pretreatment, microwave power is at 300~600W; Frequency 4000~8000MHz; Extraction time 300~400s, extraction temperature is 50~60 ℃ and reclaims sherwood oil, obtains Silymarin oil.
(3) microwave counter current extracts: the silybum marianum seed powder after the de-oiling combines microwave counter current to extract the forward charging with the ethanol of 80-85%. and flowing of backspace water inlet material and water is reverse continuous dynamic flow.Extract 50~60 ℃ of temperature, time 60-80min, 400~600W, frequency 5000~8000MHz; Slag is forced to be pushed to slag notch and is discharged through the spiral conveyer of slagging tap, and the spiral of slagging tap pushes slag simultaneously, and debris in the slag is extruded; Improve the extracting solution yield, ethanol is reclaimed in centrifuging.
(4) isoelectric precipitation: owing to be rich in acidic amino acid, isoelectric points of proteins mainly is distributed in the neutral meta-acid scope, regulates PH to 6-6.5 with hydrochloric acid, separates out protein precipitation, and separating protein is regulated pH value between 9-10 with NaOH again:
(5) macroporous resin adsorption: with the pretreated macroporous resin adsorption of process on the Silymarin alkaline solution, make adsorptive single, the resin model is S-8, AB-8, DA-201, NKA-9 resin, and the aspect ratio of resin column is 8: 1-15: 1:
(6) sour water wash-out: at normal temperatures and pressures, with the 0.5-1BV/h flow velocity, use PH is 2 phosphoric acid solution wash-out resin, and elution volume is 1 6BV;
(7) ethanol elution: at normal temperatures and pressures, with the 2-4BV/h flow velocity, with the ethanol aqueous wash deresination of 70%-95%, the wash-out body is protected and is 3-4BV;
(8) drying under reduced pressure: collect ethanol eluate, concentrating under reduced pressure reclaims ethanol, vacuum-drying, and micronization obtains SLB
Embodiment: a kind ofly utilize biological wall breaking, twice microwave extraction, and the macroporous resin purification technology, obtain the method for high purity SLB.
Embodiment 1: a kind of to utilize biological wall breaking, Microwave Pretreatment to get oily, again through microwave counter current and macroporous resin purification technology, obtains the method for high purity SLB, and its step is following:
(1) biological wall breaking: 1000g is dry, and silybum marianum seed soaks; Add an amount of cellulase, semicellulose enzyme solution; Insulation separate 60min; Remove the Mierocrystalline cellulose, semicellulose, pectin segment and the albuminous membranae that are wrapped in seed skin shell, the oil cells that bale broken is wrapped up in dissociates out, improves the grease extraction yield;
(2) Microwave Pretreatment: silybum marianum seed was pulverized 30 mesh sieves, add 4-6 times of sherwood oil (boiling range 60-90 ℃) solution, and carried out Microwave Pretreatment; Microwave power is at 300 W, and frequency 1000MHz extracts time 300s; Extraction temperature is 60 ℃; Reclaim sherwood oil, obtain Silymarin oil 195.3g, de-oiling Herba Silybi mariani powder 793g;
(3) microwave counter current extracts: the de-oiling Herba Silybi mariani powder combines microwave counter current to extract with 80% ethanol, the forward charging, and reverse water inlet, flowing of material and water is reverse continuous dynamic flow.Extract 50 ℃ of temperature, time 60min, 400W, frequency 5000MHz extrudes debris in the slag, and the protein moiety of precipitation separation reclaims ethanol;
(4) isoelectric precipitation: owing to be rich in acidic amino acid, isoelectric points of proteins mainly is distributed in the neutral meta-acid scope, regulates PH to 6 with hydrochloric acid, separates out protein precipitation, separating protein, and using NaOH to regulate pH value again is 9:
(5) macroporous resin adsorption: with the pretreated macroporous resin adsorption of process on the nonprotein Silymarin solution, make the adsorptive list, the resin model is the S-8 resin, and the aspect ratio of resin column is 8: 1;
(6) sour water wash-out: at normal temperatures and pressures, with the 0.5BV/h flow velocity, use PH is 2 phosphoric acid solution wash-out resin, and elution volume is 1BV;
(7) ethanol elution: at normal temperatures and pressures, with the 2BV/h flow velocity, the ethanol aqueous wash deresination with 70%, elution volume is 3BV;
(8) drying under reduced pressure: collect ethanol eluate, concentrating under reduced pressure reclaims ethanol, vacuum-drying, and micronization obtains SLB 51g, purity 80.8%.
Case study on implementation 2: a kind of to utilize biological wall breaking, Microwave Pretreatment to get oily, again through microwave counter current and macroporous resin purification technology, obtains the method for high purity SLB, and its step is following:
(1) biological wall breaking: 1000g is dry, and silybum marianum seed soaks; Add an amount of cellulase, semicellulose enzyme solution; Insulation enzymolysis 60min; Remove the Mierocrystalline cellulose, semicellulose, pectin segment and the albuminous membranae that are wrapped in seed skin shell, make the oil cells that is wrapped dissociate out, improve the grease extraction yield;
(2) Microwave Pretreatment: silybum marianum seed was pulverized 40 mesh sieves, add 1-6 times of sherwood oil (boiling range 60-90 ℃) solution, and carried out Microwave Pretreatment; Microwave power is at 400W, and frequency 6000MHz extracts time 400s; Extraction temperature is 60 ℃; Reclaim sherwood oil, obtain silybum marianum seed oil 195g, de-oiling Herba Silybi mariani powder 797.3g;
(3) microwave counter current extracts: the de-oiling Herba Silybi mariani powder combines microwave counter current to extract with 85% ethanol, the forward charging, and reverse water inlet, flowing of material and water is reverse continuous dynamic flow.Extract 60 ℃ of temperature, time 70min, 500W, frequency 6000MHz extrudes debris in the slag, and the protein moiety of precipitation separation reclaims ethanol:
(4) isoelectric precipitation: owing to be rich in acidic amino acid, isoelectric points of proteins mainly is distributed in the neutral meta-acid scope, regulates PH to 6.5 with hydrochloric acid, separates out protein precipitation, and it is 10 that separating protein uses NaOH to regulate pH value again;
(5) macroporous resin adsorption: with the pretreated macroporous resin adsorption of process on the nonprotein Silymarin solution, make adsorptive single, the resin model is the AB-8 resin, and the aspect ratio of resin column is 10: 1;
(6) sour water wash-out: at normal temperatures and pressures, with the 1BV/h flow velocity, use PH is 2 phosphoric acid solution wash-out resin, and elution volume is 5BV;
(7) ethanol elution: at normal temperatures and pressures, with the 3BV/h flow velocity, the ethanol aqueous wash deresination with 80%, elution volume is 4BV:
(8) drying under reduced pressure: collect ethanol eluate, concentrating under reduced pressure reclaims ethanol, and the vacuum-drying micronization obtains SLB 51.7g, purity 81.2%.
Case study on implementation 3: a kind of to utilize biological wall breaking, Microwave Pretreatment to get oily, again through microwave counter current and macroporous resin purification technology, obtains the method for high purity SLB.Its step is following:
(1) biological wall breaking: 1000g is dry, and silybum marianum seed soaks; Add an amount of cellulase, semicellulose enzyme solution; Insulation enzymolysis 60min; Remove the Mierocrystalline cellulose, semicellulose, pectin segment and the albuminous membranae that are wrapped in seed skin shell, make the oil cells that is wrapped dissociate out, improve the grease extraction yield;
(2) Microwave Pretreatment: silybum marianum seed was pulverized 50 mesh sieves, add 4-6 times of sherwood oil (boiling range 60-90 ℃) solution, and carried out Microwave Pretreatment; Microwave power is at 500W, and frequency 7000MHz extracts time 400s; Extraction temperature is 60 ℃; Reclaim sherwood oil, obtain silybum marianum seed oil 197g, de-oiling Herba Silybi mariani powder 795g;
(3) microwave counter current extracts: the de-oiling Herba Silybi mariani powder combines microwave counter current to extract with 80% ethanol, the forward charging, and reverse water inlet, flowing of material and water is reverse continuous dynamic flow.Extract 50 ℃ of temperature, time 80min, 600W, frequency 7000MHz extrudes debris in the slag, and the protein moiety of precipitation separation reclaims ethanol;
(4) isoelectric precipitation: owing to be rich in acidic amino acid, isoelectric points of proteins mainly is distributed in the neutral meta-acid scope, regulates PH to 6.5 with hydrochloric acid, separates out protein precipitation, protein isolates, and using NaOH to regulate pH value again is 9;
(5) macroporous resin adsorption: with the pretreated macroporous resin adsorption of process on the nonprotein Silymarin solution, make the adsorptive list, the resin model is the DA-201 resin, and the aspect ratio of resin column is 12: 1;
(6) sour water wash-out: at normal temperatures and pressures with the 1BV/h flow velocity, use PH is 2 phosphoric acid solution wash-out resin, and elution volume is 5BV;
(7) ethanol elution: at normal temperatures and pressures, with the 3BV/h flow velocity, the ethanol aqueous wash deresination with 80%, elution volume is 4BV;
(8) drying under reduced pressure: collect ethanol eluate, alkali is pressed and is concentrated, and reclaims ethanol, and the vacuum-drying micronization obtains SLB 50.5g, purity 82%.
Case study on implementation 4: a kind of to utilize biological wall breaking, Microwave Pretreatment to get oily, through microwave counter current and macroporous resin purification technology, obtains the method that high-purity severe edema due to hypofunction of the spleen flies silibin again.Its step is following:
(1) biological wall breaking: 1000g is dry, and silybum marianum seed soaks; Add an amount of cellulase, semicellulose enzyme solution; Insulation enzymolysis 60min; Remove the Mierocrystalline cellulose, semicellulose, pectin segment and the albuminous membranae that are wrapped in seed skin shell, make the oil cells that is wrapped dissociate out, improve the grease extraction yield;
(2) Microwave Pretreatment: silybum marianum seed was pulverized 60 mesh sieves, add 1-6 times of sherwood oil (boiling range 60-90 ℃) solution, and carried out Microwave Pretreatment; Microwave power is at 600W, and frequency 8000MHz extracts time 360s; Extraction temperature is 50 ℃; Reclaim sherwood oil, obtain silybum marianum seed oil 189g, de-oiling Herba Silybi mariani powder 790.6g;
(3) microwave counter current extracts: the de-oiling Herba Silybi mariani powder combines microwave counter current to extract with 80% ethanol, the forward charging, and reverse water inlet, flowing of material and water is reverse continuous dynamic flow.Extract 60 ℃ of temperature, time 80min, 600W, frequency 8000MHz extrudes debris in the slag, and the protein moiety of precipitation separation reclaims ethanol;
(4) isoelectric precipitation: owing to be rich in acidic amino acid, isoelectric points of proteins mainly is distributed in the neutral meta-acid scope, regulates PH to 6.5 with hydrochloric acid, separates out protein precipitation, separating protein, and using NaOH to regulate pH value again is 9;
(5) macroporous resin adsorption: with the pretreated macroporous resin adsorption of process on the nonprotein Silymarin solution, make adsorptive single, the resin model is the NKA-9 resin, and the aspect ratio of resin column is 15: 1;
(6) sour water wash-out: at normal temperatures and pressures, with the 1BV/h flow velocity, use PH is 2 phosphoric acid solution wash-out resin, and elution volume is 5BV;
(7) ethanol elution: at normal temperatures and pressures, with the 4BV/h flow velocity, the ethanol aqueous wash deresination with 95%, elution volume is 4BV;
(8) drying under reduced pressure: collect ethanol eluate, concentrate, reclaim ethanol, drying under reduced pressure, micronization obtains SLB 49.7g, purity 83.1%.