CN1724995A - Determination method of balsum pear saponin - Google Patents
Determination method of balsum pear saponin Download PDFInfo
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- CN1724995A CN1724995A CN 200510014471 CN200510014471A CN1724995A CN 1724995 A CN1724995 A CN 1724995A CN 200510014471 CN200510014471 CN 200510014471 CN 200510014471 A CN200510014471 A CN 200510014471A CN 1724995 A CN1724995 A CN 1724995A
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Abstract
The invention discloses a balsam pear saponin determining method, including the following steps: 1. preparing a balsam pear saponin control sample: (1) extracting coarse product of balsam pear saponin, i.e. adding cellulose enzyme into balsam pear seed powder for microwave processing, and filtering and collecting supernatant liquor, passing the supernatant liquor through hollow fiber film, taking penetrating fluid to pass it through the other hollow fiber film for holding back molecular weight-6000 substances, taking penetrating fluid again for concentrating, and obtaining balsam pear saponin coarse extractant for purifying and drying, so as to obtain the coarse product; (2) pre-purifying for preparing front balsam pear saponin by HPCL separation; (3) preparing by HPLC separation; (4) re-crystallizing the product; 2. determining the saponin content of the sample: adopting light-splitting photometric method to measure light absorptivity in 468 nm wavelength, calculating to obtain the content. The determining method provides standard control sample, accurate and reliable.
Description
Technical field
The present invention relates to a kind of assay method of compound, particularly relate to a kind of assay method of momordica saponins.
Background technology
Momordica saponins is one of main effective constituent of balsam pear, to the existing a large amount of reports of the mensuration of momordica saponins in balsam pear and the goods thereof.At present, the mensuration to momordica saponins is reference substance with panaxoside or momordica saponins mainly both at home and abroad, adopts spectrophotometry.
People such as Chen Xun (Chen Xun, in Haining, Tang Desong etc., the research of momordica saponins rapid separation and purification method, Food Science, 2004,25 (2): with ethanol lixiviate balsam pear, be reference substance with the panaxoside A 114-117), the 548nm place surveys absorbance measurement momordica saponins total amount.Sun Lei (Sun Lei, the technical study of alcohol extract momordica saponins, the Ningxia medical journal, 2005,27 (2): also being to use the alcohol extract momordica saponins 112-114), determining optimised process, is reference substance with the panaxoside A, the 548nm place surveys absorbance.Gao Wen etc. (Gao Wen, Zhu Xiaoyun, Liu Lijun etc., the application of microwave technology in Charantin extracts, Guangxi light industry, 2004,, 5:13-15) with the panaxoside be standard items (Rb1, content 〉=98%), measure absorbance at 550nm wavelength place and calculate Charantin content.People such as Cui Henglin (Cui Henglin, Xu Bin, Dong Ying, the leaching process research of momordica saponins, Jiangsu University's journal, 2004,25 (5): be reference substance 372-375) with the homemade momordica saponins A in laboratory, the 544nm place with spectrophotometric method to balsam pear in saponin measure, but the purity of momordica saponins reference substance is not described.People such as chloroazotic acid (chloroazotic acid, Tang Lin, Guo Yiran etc., the combine content of fast measuring Charantin A of Solid-Phase Extraction and high performance liquid chromatography, chromatogram, 2001,19 (2): be reference substance 128-131), set up the method for the mensuration Charantin A content that Solid-Phase Extraction and high performance liquid chromatography combine with Charantin A.People such as Jiang Fashou (Jiang Fashou, Liu Jinrong, Wang Hangyu etc., the assay of Charantin in the balsam pear, land-reclaimable medical science, 2001,23 (5)) comparing with the cupreol standard items, measure the content of cupreol-3-glycoside in the balsam pear, is cupreol-3-glycoside and 5 by momordica saponins, 25-beans steroid dienol-3-glycoside calculates the content of momordica saponins with the potpourri of 1: 1 ratio composition.
In the said method be that the method that reference substance is measured momordica saponins content in the balsam pear is inaccurate with the panaxoside, this is that just the structure of momordica saponins and panaxoside has similarity, so can make measurement result inaccurate because do not contain panaxoside in the balsam pear.Compare with the cupreol standard items, the method of content that calculates momordica saponins is more inaccurate, because as can be known from existing research, the kind of momordica saponins is more in the balsam pear, have more than is cupreol-3-glycoside and 5, and 25-beans steroid dienol-3-glycoside is with the potpourri of 1: 1 ratio composition.With the momordica saponins is that the reference substance result is best, but the purifying of momordica saponins reference substance prepare aspect report not, the purity of the momordica saponins that can buy is too low, is not suitable for the product of comparing, the mensuration that therefore causes momordica saponins is difficulty relatively.
Summary of the invention
The objective of the invention is to overcome the deficiency in the existing determination techniques, a kind of assay method of momordica saponins is provided.
Technical scheme of the present invention is summarized as follows:
A kind of assay method of momordica saponins, form by following steps:
(1) preparation of momordica saponins reference substance
1. the extraction of momordica saponins crude product
With the bitter melon seed powder that shines after 60 mesh sieves are crossed in dry grinding, ratio by weight 1: 15～40 adds distilled water, add cellulase by 2～6mg/ml again, 30 ℃～60 ℃, effect 1～3h uses 500～5000W microwave treatment, 10～60min again, filters, centrifugal, collect supernatant, cross the hollow-fibre membrane of 0.2 μ m, get through liquid, cross the hollow-fibre membrane of molecular cut off 6000, get and see through liquid in rotary evaporator, 60 ℃, 0.05～0.07Mpa, be evaporated to the 1/5-1/10 of original volume, get the momordica saponins crude extract; The sherwood oil or the ether that add 0.5～2 times of volume in described momordica saponins crude extract extract 1～3 time, remove oil-soluble impurities, take off a layer water; Use the extracting n-butyl alcohol 1～3 time of 0.5～2 times of volume again, remove hydrophilic impurities, get the normal butyl alcohol phase, behind anhydrous sodium sulfate dehydration, vacuum drying obtains pale brown sugar colour pulpous state momordica saponins crude product;
2. HPLC separates the prepurification of the preceding momordica saponins of preparation
Described momordica saponins crude product 10g is dissolved in the 400ml distilled water, last macroporous adsorbent resin column chromatography, it with percent by volume 60%～80% ethanol water wash-out, collect eluent, with eluent with being that weight percent concentration is that 40% potassium hydroxide aqueous solution or sodium hydrate aqueous solution are transferred pH to 10, placed 8～12 hours for 3 ℃～5 ℃, filter, filtrate is used anhydrous Na
2SO
4Dehydration behind the recovery solvent, drips ether and makes with extra care to such an extent that white crystal is the momordica saponins of prepurification;
3. HPLC separates preparation
The momordica saponins crystal of described prepurification is dissolved in redistilled water, is made into the aqueous solution of 5mg/ml, analyze, detect chromatographic condition: chromatographic column: C18 at 192nm with HPLC; 10 μ m; 7.8mmID * 150mm; Moving phase: acetonitrile: water=35: 65, isocratic elution; Flow velocity: 4ml/min;
4. HPLC separates the recrystallization of preparation back product
After HPLC separates, collect 60 ℃ of liquid concentrating under reduced pressure, 0.05Mpa, 4h get momordica saponins reference substance crystal;
(2) momordica saponins Determination on content in the test sample
1. the preparation of reference substance solution
It is an amount of that precision takes by weighing the momordica saponins reference substance that is dried to constant weight, increases the weight of to steam water-soluble separating, and makes the solution that every 1ml contains 0.1mg;
2. the preparation of need testing solution
It is an amount of that precision takes by weighing test sample, adds an amount of distilled water fragmentation, presses 4mg/ml again and add cellulase, 45 ℃ act on 2h down, use 750W microwave treatment 45min again, filter, centrifugal, collect supernatant, cross the hollow-fibre membrane of 0.2 μ m, get and see through liquid, cross the hollow-fibre membrane of molecular cut off 6000, get through liquid in rotary evaporator 60 ℃, 0.05～0.07Mpa is evaporated to 1/5～1/10 of original volume, water is settled to scale in the immigration volumetric flask, shakes up, promptly;
3. momordica saponins Determination on content in the test sample
Accurate respectively reference substance solution and each 1ml of need testing solution of drawing places tool plug test tube respectively, and solvent is flung in water-bath, each accurate 5% vanillic aldehyde-glacial acetic acid solution 0.2ml that adds new preparation, perchloric acid 0.8ml heats 15min in 60 ℃ of water-baths, cooling adds glacial acetic acid 5ml, shakes up, place 15min, do blank test with the aqueous solution retinue,, survey absorbance at the wavelength place of 468nm according to spectrophotometric method, calculate, promptly.
The best mode of the extraction step of described momordica saponins crude product is: shine the bitter melon seed powder after 60 mesh sieves are crossed in dry grinding, add distilled water by weight 1: 30 ratio, press 4mg/ml again and add cellulase, 45 ℃, effect 2h uses 750W microwave treatment 45min again, filters, centrifugal, collect supernatant, cross the hollow-fibre membrane of 0.2 μ m, get through liquid, cross the hollow-fibre membrane of molecular cut off 6000, get and see through liquid in rotary evaporator, 60 ℃, 0.06Mpa, be evaporated to 1/7 of original volume, get the momordica saponins crude extract; In described momordica saponins crude extract, add the sherwood oil or the ether of 1 times of volume, extract 2 times, remove oil-soluble impurities, take off a layer water; Use isopyknic extracting n-butyl alcohol 2 times again, remove hydrophilic impurities, get the normal butyl alcohol phase, behind anhydrous sodium sulfate dehydration, vacuum drying obtains pale brown sugar colour pulpous state momordica saponins crude product.
The assay method of momordica saponins of the present invention, high performance liquid chromatography (HPLC) method of preparation separating and purifying high-purity momordica saponins standard reference material, for the standardization quantitative test of momordica saponins in balsam pear class health food and the Chinese herbal medicine provides standard reference material, for momordica saponins Determination on content in the test sample provides a kind of method accurately and reliably.This method with panaxoside or cupreol be reference substance measure momordica saponins content method more accurately and reliably.
Description of drawings
Fig. 1 analyzes chromatogram for the momordica saponins reference substance.
Embodiment
The present invention is further illustrated below in conjunction with specific embodiment.
Embodiment 1
A kind of assay method of momordica saponins, form by following steps:
(1) preparation of momordica saponins reference substance
1. the extraction of momordica saponins crude product
Bitter melon seed powder with shining after 60 mesh sieves are crossed in dry grinding adds distilled water by weight 1: 30 ratio, presses 4mg/ml again and adds cellulase, 45 ℃, effect 2h uses 750W microwave treatment 45min again, filter, centrifugal, collect supernatant, cross the hollow-fibre membrane of 0.2 μ m, get, cross the hollow-fibre membrane of molecular cut off 6000 through liquid, get and see through liquid in rotary evaporator, 60 ℃, 0.06Mpa, be evaporated to 1/7 of original volume, get the momordica saponins crude extract; In described momordica saponins crude extract, add the ether of 1 times of volume, extract 2 times, remove oil-soluble impurities, take off a layer water; Use the extracting n-butyl alcohol 2 times of 0.5 times of volume again, remove hydrophilic impurities, get the normal butyl alcohol phase, behind anhydrous sodium sulfate dehydration, vacuum drying obtains pale brown sugar colour pulpous state momordica saponins crude product;
2. HPLC separates the prepurification of the preceding momordica saponins of preparation
Described momordica saponins crude product 10g is dissolved in the 400ml distilled water, and last macroporous adsorbent resin column chromatography is 60%～80% ethanol water wash-out with percent by volume, collect eluent, eluent is transferred pH to 10 with sodium hydrate aqueous solution, placed 10 hours for 4 ℃, filter, filtrate is used anhydrous Na
2SO
4Dehydration behind the recovery solvent, drips ether and makes with extra care to such an extent that white crystal is the momordica saponins of prepurification;
3. HPLC separates preparation
The momordica saponins crystal of described prepurification is dissolved in redistilled water, is made into the aqueous solution of 5mg/ml, analyze, detect chromatographic condition: chromatographic column: C18 at 192nm with HPLC; 10 μ m; 7.8mmID * 150mm; Moving phase: acetonitrile: water=35: 65, isocratic elution; Flow velocity: 4ml/min;
4. HPLC separates the recrystallization of preparation back product
After HPLC separates, collect 60 ℃ of liquid, 0.05Mpa, concentrated 4h get momordica saponins reference substance crystal;
(2) proterties of momordica saponins reference substance
Momordica saponins reference substance prepared among the present invention is white powder, and is water-soluble, is not soluble in methyl alcohol and ethanol;
(3) discriminating of momordica saponins reference substance
1. a little is dissolved in the 2ml water to get this product, puts the jolting 1min that exerts oneself in the test tube with ground stopper, produces a large amount of foams, places 10min, and foam does not have obvious disappearance.
2. this product momordica saponins aqueous solution of being made into 5mg/ml is analyzed with HPLC, detects at 192nm that (acetonitrile: water=35: 65, flow velocity: 0.9ml/min), area normalization determines that product purity is 99.2% for C18 chromatographic column, 192nm.
(4) momordica saponins Determination on content in the test sample
1. the preparation of reference substance solution
It is an amount of that precision takes by weighing the momordica saponins reference substance that is dried to constant weight, increases the weight of to steam water-soluble separating, and makes the solution that every 1ml contains 0.1mg.
2. the preparation of need testing solution
It is an amount of that precision takes by weighing test sample, adds an amount of distilled water fragmentation, presses 4mg/ml again and add cellulase, 45 ℃ act on 2h down, use 750W microwave treatment 45min again, filter, centrifugal, collect supernatant, cross the hollow-fibre membrane of 0.2 μ m, get and see through liquid, cross the hollow-fibre membrane of molecular cut off 6000, get through liquid in rotary evaporator 60 ℃, 0.06Mpa be evaporated to 1/7 of original volume, water is settled to scale in the immigration volumetric flask, shakes up, promptly.
3. momordica saponins Determination on content in the test sample
Accurate respectively reference substance solution and each 1ml of need testing solution of drawing places tool plug test tube respectively, and solvent is flung in water-bath, each accurate 5% vanillic aldehyde-glacial acetic acid solution 0.2ml that adds new preparation, perchloric acid 0.8ml heats 15min in 60 ℃ of water-baths, cooling adds glacial acetic acid 5ml, shakes up, place 15min, do blank test with the aqueous solution retinue,, survey absorbance at the wavelength place of 468nm according to spectrophotometric method, calculate, promptly.
Embodiment 2
A kind of assay method of momordica saponins, form by following steps:
(1) preparation of momordica saponins reference substance
1. the extraction of momordica saponins crude product
Bitter melon seed powder with shining after 60 mesh sieves are crossed in dry grinding adds distilled water by weight 1: 15 ratio, presses 6mg/ml again and adds cellulase, 60 ℃, effect 1h uses 5000W microwave treatment 10min again, filter, centrifugal, collect supernatant, cross the hollow-fibre membrane of 0.2 μ m, get, cross the hollow-fibre membrane of molecular cut off 6000 through liquid, get and see through liquid in rotary evaporator, 60 ℃, 0.05Mpa, be evaporated to 1/5 of original volume, get the momordica saponins crude extract; In described momordica saponins crude extract, add the sherwood oil of 0.5 times of volume, extract 1 time, remove oil-soluble impurities, take off a layer water; Use isopyknic extracting n-butyl alcohol 1 time again, remove hydrophilic impurities, get the normal butyl alcohol phase, behind anhydrous sodium sulfate dehydration, vacuum drying obtains pale brown sugar colour pulpous state momordica saponins crude product;
2. HPLC separates the prepurification of the preceding momordica saponins of preparation
Described momordica saponins crude product 10g is dissolved in the 400ml distilled water, and last macroporous adsorbent resin column chromatography is 60%～80% ethanol water wash-out with percent by volume, collect eluent, eluent is transferred pH to 10 with potassium hydroxide aqueous solution, placed 12 hours for 3 ℃, filter, filtrate is used anhydrous Na
2SO
4Dehydration behind the recovery solvent, drips ether and makes with extra care to such an extent that white crystal is the momordica saponins of prepurification;
3. HPLC separates preparation
With embodiment 1
4. HPLC separates the recrystallization of preparation back product
With embodiment 1
(2) momordica saponins Determination on content in the test sample
1. the preparation of reference substance solution
With embodiment 1
2. the preparation of need testing solution
It is an amount of that precision takes by weighing test sample, adds an amount of distilled water fragmentation, presses 4mg/ml again and add cellulase, 45 ℃ act on 2h down, use 750W microwave treatment 45min again, filter, centrifugal, collect supernatant, cross the hollow-fibre membrane of 0.2 μ m, get and see through liquid, cross the hollow-fibre membrane of molecular cut off 6000, get through liquid in rotary evaporator 60 ℃, 0.05Mpa be evaporated to 1/5 of original volume, water is settled to scale in the immigration volumetric flask, shakes up, promptly.
3. momordica saponins Determination on content in the test sample
With embodiment 1
Embodiment 3
A kind of assay method of momordica saponins, form by following steps:
(1) preparation of momordica saponins reference substance
1. the extraction of momordica saponins crude product
Bitter melon seed powder with shining after 60 mesh sieves are crossed in dry grinding adds distilled water by weight 1: 40 ratio, presses 2mg/ml again and adds cellulase, 30 ℃, effect 3h uses 500W microwave treatment 60min again, filter, centrifugal, collect supernatant, cross the hollow-fibre membrane of 0.2 μ m, get, cross the hollow-fibre membrane of molecular cut off 6000 through liquid, get and see through liquid in rotary evaporator, 60 ℃, 0.07Mpa, be evaporated to 1/10 of original volume, get the momordica saponins crude extract; In described momordica saponins crude extract, add the sherwood oil of 2 times of volumes, extract 3 times, remove oil-soluble impurities, take off a layer water; Use the extracting n-butyl alcohol 3 times of 2 times of volumes again, remove hydrophilic impurities, get the normal butyl alcohol phase, behind anhydrous sodium sulfate dehydration, vacuum drying obtains pale brown sugar colour pulpous state momordica saponins crude product;
2. HPLC separates the prepurification of the preceding momordica saponins of preparation
Described momordica saponins crude product 10g is dissolved in the 400ml distilled water, and last macroporous adsorbent resin column chromatography is 60%～80% ethanol water wash-out with percent by volume, collect eluent, eluent is transferred pH to 10 with potassium hydroxide aqueous solution, placed 8 hours for 5 ℃, filter, filtrate is used anhydrous Na
2SO
4Dehydration behind the recovery solvent, drips ether and makes with extra care to such an extent that white crystal is the momordica saponins of prepurification;
3. HPLC separates preparation
With embodiment 1
4. HPLC separates the recrystallization of preparation back product
With embodiment 1
(2) momordica saponins Determination on content in the test sample
1. the preparation of reference substance solution
With embodiment 1
2. the preparation of need testing solution
It is an amount of that precision takes by weighing test sample, adds an amount of distilled water fragmentation, presses 4mg/ml again and add cellulase, 45 ℃ act on 2h down, use 750W microwave treatment 45min again, filter, centrifugal, collect supernatant, cross the hollow-fibre membrane of 0.2 μ m, get and see through liquid, cross the hollow-fibre membrane of molecular cut off 6000, get through liquid in rotary evaporator 60 ℃, 0.07Mpa be evaporated to 1/10 of original volume, water is settled to scale in the immigration volumetric flask, shakes up, promptly.
Claims (2)
1. the assay method of a momordica saponins is characterized in that being made up of following steps:
(1) preparation of momordica saponins reference substance
1. the extraction of momordica saponins crude product
With the bitter melon seed powder that shines after 60 mesh sieves are crossed in dry grinding, ratio by weight 1: 15～40 adds distilled water, add cellulase by 2～6mg/ml again, 30 ℃～60 ℃, effect 1～3h uses 500～5000W microwave treatment, 10～60min again, filters, centrifugal, collect supernatant, cross the hollow-fibre membrane of 0.2 μ m, get through liquid, cross the hollow-fibre membrane of molecular cut off 6000, get and see through liquid in rotary evaporator, 60 ℃, 0.05～0.07Mpa, be evaporated to 1/5～1/10 of original volume, get the momordica saponins crude extract; The sherwood oil or the ether that add 0.5～2 times of volume in described momordica saponins crude extract extract 1～3 time, remove oil-soluble impurities, take off a layer water; Use the extracting n-butyl alcohol 1～3 time of 0.5～2 times of volume again, remove hydrophilic impurities, get the normal butyl alcohol phase, behind anhydrous sodium sulfate dehydration, vacuum drying obtains pale brown sugar colour pulpous state momordica saponins crude product;
2. HPLC separates the prepurification of the preceding momordica saponins of preparation
Described momordica saponins crude product 10g is dissolved in the 400ml distilled water, last macroporous adsorbent resin column chromatography, it with percent by volume 60%～80% ethanol water wash-out, collect eluent, eluent is transferred pH to 10 with potassium hydroxide aqueous solution or sodium hydrate aqueous solution, placed 8-12 hour for 3 ℃～5 ℃, filter, filtrate is used anhydrous Na
2SO
4Dehydration behind the recovery solvent, drips ether and makes with extra care to such an extent that white crystal is the momordica saponins of prepurification;
3. HPLC separates preparation
The momordica saponins crystal of described prepurification is dissolved in redistilled water, is made into the aqueous solution of 5mg/ml, analyze, detect chromatographic condition: chromatographic column: C18 at 192nm with HPLC; 10 μ m; 7.8mm ID * 150mm; Moving phase: acetonitrile: water=35: 65, isocratic elution; Flow velocity: 4ml/min;
4. HPLC separates the recrystallization of preparation back product
After HPLC separates, collect 60 ℃ of liquid, 0.05Mpa, concentrated 4h get momordica saponins reference substance crystal;
(2) momordica saponins Determination on content in the test sample
1. the preparation of reference substance solution
It is an amount of that precision takes by weighing the momordica saponins reference substance that is dried to constant weight, increases the weight of to steam water-soluble separating, and makes the solution that every 1ml contains 0.1mg;
2. the preparation of need testing solution
It is an amount of that precision takes by weighing test sample, adds an amount of distilled water fragmentation, presses 4mg/ml again and add cellulase, 45 ℃ act on 2h down, use 750W microwave treatment 45min again, filter, centrifugal, collect supernatant, cross the hollow-fibre membrane of 0.2 μ m, get and see through liquid, cross the hollow-fibre membrane of molecular cut off 6000, get through liquid in rotary evaporator 60 ℃, 0.05～0.07Mpa is evaporated to 1/5～1/10 of original volume, water is settled to scale in the immigration volumetric flask, shakes up, promptly;
3. momordica saponins Determination on content in the test sample
Accurate respectively reference substance solution and each 1ml of need testing solution of drawing places tool plug test tube respectively, and solvent is flung in water-bath, each accurate 5% vanillic aldehyde-glacial acetic acid solution 0.2ml that adds new preparation, perchloric acid 0.8ml heats 15min in 60 ℃ of water-baths, cooling adds glacial acetic acid 5ml, shakes up, place 15min, do blank test with the aqueous solution retinue,, survey absorbance at the wavelength place of 468nm according to spectrophotometric method, calculate, promptly.
2. the assay method of a kind of momordica saponins according to claim 1, the extraction step that it is characterized in that described momordica saponins crude product is: shine the bitter melon seed powder after 60 mesh sieves are crossed in dry grinding, add distilled water by weight 1: 30 ratio, press 4mg/ml again and add cellulase, 45 ℃, effect 2h, use 750W microwave treatment 45min again, filter, centrifugal, collect supernatant, cross the hollow-fibre membrane of 0.2 μ m, get and see through liquid, cross the hollow-fibre membrane of molecular cut off 6000, get through liquid in rotary evaporator, 60 ℃, 0.06Mpa, be evaporated to 1/7 of original volume, get the momordica saponins crude extract; In described momordica saponins crude extract, add the sherwood oil or the ether of 1 times of volume, extract 2 times, remove oil-soluble impurities, take off a layer water; Use isopyknic extracting n-butyl alcohol 2 times again, remove hydrophilic impurities, get the normal butyl alcohol phase, behind anhydrous sodium sulfate dehydration, vacuum drying obtains pale brown sugar colour pulpous state momordica saponins crude product.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102648925A (en) * | 2011-02-25 | 2012-08-29 | 苏州宝泽堂医药科技有限公司 | Preparation method of achyranthes bidentata saponin |
| CN103304490A (en) * | 2013-06-18 | 2013-09-18 | 聊城大学 | Method for separating and purifying five purine and pyrimidine bases from trichosanthes bark |
| CN103389321A (en) * | 2013-08-13 | 2013-11-13 | 华南农业大学 | Physical model testing method for pre-cooling and freshness preservation of fruits |
| CN104931293A (en) * | 2015-07-10 | 2015-09-23 | 辽宁大学 | Method for recycling overflowed solution sample |
-
2005
- 2005-07-12 CN CNB2005100144719A patent/CN100408999C/en not_active Expired - Fee Related
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102648925A (en) * | 2011-02-25 | 2012-08-29 | 苏州宝泽堂医药科技有限公司 | Preparation method of achyranthes bidentata saponin |
| CN103304490A (en) * | 2013-06-18 | 2013-09-18 | 聊城大学 | Method for separating and purifying five purine and pyrimidine bases from trichosanthes bark |
| CN103389321A (en) * | 2013-08-13 | 2013-11-13 | 华南农业大学 | Physical model testing method for pre-cooling and freshness preservation of fruits |
| CN103389321B (en) * | 2013-08-13 | 2015-06-17 | 华南农业大学 | Physical model testing method for pre-cooling and freshness preservation of fruits |
| CN104931293A (en) * | 2015-07-10 | 2015-09-23 | 辽宁大学 | Method for recycling overflowed solution sample |
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| Publication number | Publication date |
|---|---|
| CN100408999C (en) | 2008-08-06 |
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