CN101904882A - Preparation method of lithocarpus litseifolius total flavone - Google Patents
Preparation method of lithocarpus litseifolius total flavone Download PDFInfo
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Abstract
The invention discloses lithocarpus litseifolius total flavone extracted by using plant lithocarpus litseifolius as a raw material to develop a new hypoglycemic medicament, and provides a method for preparing the lithocarpus litseifolius total flavone. The purity of the lithocarpus litseifolius total flavone prepared by the method can reach over 80 percent, the phloridzin content reaches over 65 percent, the process is simple and has low energy consumption, and the technological condition is reasonable, stable and feasible. A hypoglycemic capsule developed by using the lithocarpus litseifolius total flavone can provide a convenient, acceptant and long-term taking oral hypoglycemic preparation for diabetics.
Description
Technical field
The present invention relates to medication preparation, the preparation method of the plant medicines and the health product of the blood sugar lowering class of more specifically saying so.
Background technology
Lithocarpus litseifolius is Fagaceae Lithocarpus plant Lithocarpus litseifolius Chun., mainly is distributed in Guangxi China Baise, reaches the clouds, in the geographic mountain region thick forest such as Pingguo, Napo County, Long Lin, Daming Shan Mountain and rivers and ponds, aboundresources.The Lithocarpus litseifolius main component is a flavones ingredient, and content is up to 12.6%, mainly comprises phlorhizin, Trilobatin, 3-hydroxyl phlorhizin.Pharmacological research is found phlorhizin and isomers Trilobatin thereof have the pharmacokinetics aspect to glycometabolic adjusting synergism, phlorhizin can reverse insulin resistance, make the carbohydrate tolerance and the insulin sensitivity of insulin resistance rat recover normal, correct the sensitivity of the glucagon of hyperglycemia Canis familiaris L. to glucose, suppress of the absorption of hyperglycemia rabbit intestinal to sugar, the expression of the glycogen metabolic gene of part reverting diabetes, increase the glucose-6-phosphate dehydrogenase (G6PD) activity, the catalysis phosphopentose pathway.U.S. Pat 4760135 in 1988 discloses the manufacturing that phlorhizin and phloretin derivant are used for hypoglycemic drug, but phlorhizin can be by glycosidase hydrolysis (Diedrich at intestinal, Arch.Biochem.Biophys., 1972,153 (1): 155), Barrow has contrasted two kinds of approach of filling harmonization of the stomach intraperitoneal injection, proof has only the interior discovery of the rat body of lumbar injection that original shape medicine (Barrow is arranged, Biochem.J.1977,125 (2): 24), this hydrolysing activity has tangible animal race difference (Ramaswamy, Comp.Biochem.Physiol.A., 1972,43 (1): 173).Because what US4760135 used is single-activity chemical compound phlorhizin, can't avoid the hydrolysis of intestinal glycosidase, therefore, must just can come into force by drug administration by injection.And diabetics is difficult to accept to the long term injections administering mode.
Summary of the invention
The objective of the invention is in order to provide a kind of to diabetics is the newtype drug of raw material with the lithocarpus litseifolius total flavone, and being beneficial to diabetics can be for a long time, take easily, and the preparation method of this medicine particularly is provided.
Discovering that Trilobatin is the inhibitor of intestinal glycosidase, is again the substrate of this enzyme.But its effect that suppresses glycosidase is much larger than the effect that is hydrolyzed (Evans, Arch.Biochem.Biophys., 1980,199 (2): 342); Suppress the hydrolysis of intestinal glycosidase to phlorhizin because Trilobatin has, the protection phlorhizin is not subjected to the hydrolysis of glycosidase, can improve the bioavailability of phlorhizin.Because phlorhizin and Trilobatin are present in the Lithocarpus litseifolius jointly, become and have the synergistic natural formula of pharmacokinetics again.Mixed extract-the lithocarpus litseifolius total flavone that extracts from Lithocarpus litseifolius comprising phlorhizin and Trilobatin, can avoid the hydrolysis of phlorhizin at intestinal, thereby oral administration is also effective.Therefore, diabetics is more prone to accept.
Lithocarpus litseifolius total flavone can suppress the absorption of intestinal to sugar, reduces the blood glucose source; Suppress kidney sugar and heavily absorb, promote the drainage of sugar, blood sugar lowering alleviates the toxic action of hyperglycemia to each organ-tissue; Improve the sensitivity of histiocyte, improve resistance of insulin and reducing insulin; Promote the synthetic of phosphopentose pathway and muscle glycogen, increase the oxidation of sugar.Because can reducing a plurality of links such as blood glucose source and increase blood glucose outlet in the carbohydrate metabolism process, it adjusts, so lithocarpus litseifolius total flavone has become a kind of main effective ingredient of novel blood sugar lowing medicine.
A kind of lithocarpus litseifolius total flavone preparation method that the present invention proposes, comprise extraction, concentrated, vacuum drying operation, it is characterized in that: be solvent with ethanol, the Lithocarpus litseifolius cured leaf is carried out heating and refluxing extraction, extracting solution is reuse polyamide adsorbing separation after treatment, is that eluant is with the lithocarpus litseifolius total flavone eluting on the polyamide again with ethanol at last.
Above-mentioned a kind of lithocarpus litseifolius total flavone preparation method is characterized in that:
1. alcohol heating reflux extracts:
A. the Lithocarpus litseifolius cured leaf adds the ethanol that 5-15 doubly measures the 20-70% volumetric concentration by weight;
B. reflux, extract, is 1--4 time, each 0.5--3 hour;
C. the reflux, extract, temperature is 50-90 ℃.
2. polyamide method separation and purification:
A. absorption: the mass volume ratio of upper prop phlorhizin amount and polyamide＜23: 1 (promptly being equivalent to 0.3g crude drug in whole/mL resin); Sample solution phlorhizin concentration is that 6-8mg/mL (is equivalent to 0.1g crude drug in whole/mL); Last column flow rate is 0.5-2BV/h;
B. eluting: with ethanol is eluant; The eluant consumption is 10-30BV; Elution flow rate is 1-3BV/h.
Above-mentioned a kind of lithocarpus litseifolius total flavone preparation method is characterized in that:
1. alcohol heating reflux extracts:
A. the Lithocarpus litseifolius cured leaf adds 10 times of ethanol of measuring 70% volumetric concentrations by weight;
B. reflux, extract, is three times, each 2 hours;
C. the reflux, extract, temperature is 80-90 ℃.
2. polyamide method separation and purification:
A. absorption: upper prop phlorhizin and polyamide mass volume ratio＜23: 1 (promptly being equivalent to 0.3g crude drug in whole/mL resin); Sample solution phlorhizin concentration is that 7mg/mL (is equivalent to 0.1g crude drug in whole/mL); Last column flow rate is 1BV/h;
B. eluting: 20% ethanol is eluant; The eluant consumption is 20BV; Elution flow rate is 2BV/h.
Above-mentioned a kind of lithocarpus litseifolius total flavone preparation method is characterized in that: the polyamide specification is the 80-100 order.
Collect the eluent of 20BV, the eluent decompression recycling ethanol concentrates, and vacuum drying obtains blood sugar lowering extract of the present invention---lithocarpus litseifolius total flavone.
Technical scheme of the present invention and operational approach are selections like this and determine.
Solvent extraction method is generally adopted in the extraction of flavone compound, as method (Chen Xiaoqing such as hot water extraction, alkaline water or alkaline rare alcohol extraction, organic solvent extraction, system's solvent extractions, Deng. medicinal herb components separate analytical technique and method, the 1st edition. Beijing: Chemical Industry Press's modern biotechnology and medical sci-tech publishing centre, 2006:62-65.).And from Lithocarpus litseifolius, extract flavone component mainly contain the water boiling and extraction method (Liao Xiaofeng, etc. from Pasania cuspidata, extract the research of dihydrochalcone (DHC) class sweeting agent. Guangzhou food industry science and technology, 1999,15 (1): 35-38.; Yu Lijing, etc. the extraction of dihydrochalcone, purification and assay in the Pasania cuspidata. Food Additives Used in China, 2007, (1): 176-179.; Dong Huaqiang, etc. microwave-assisted extracts Pasania cuspidata tender leaf flavone technical study. Transactions of the Chinese Society of Agricultural Engineering, 2007,23 (2): 213-217.) and ethanol extraction method (Xiao Kunfu, Deng. Flavonoid substances Study on extraction process in the Pasania cuspidata. Food Science, 2004,25 (5): 112-115.; Li Shenghua, etc. Pasania cuspidata total flavone extracting process and content Dynamic variation thereof. Food Science, 2008,29 (6): 139-141.; Zhao Hongfang, Yang Yaling, week is strong. the extraction of total flavones and antiallergic activity thereof research in the Folium hydrangeae strigosae of Yunnan. Hebei chemical industry, 2006,29 (5): 10-11.).The hot water extraction method is a common method of extracting flavonoid glycoside compound, is easy to industrialized great production, but impurity soluble in water is more during the hot water lixiviate, and post processing is complicated, and extraction efficiency is also not too high.
Because flavone compound has phenolic hydroxyl group mostly, therefore available alkaline water or the rare pure lixiviate of alkalescence, leachate can be separated out flavone compound after acidify.But when extracting with basic solvent, used alkali concn is too high, destroys the flavone compound parent nucleus during heating easily; And during the leachate acidify, in rare alcohol of the flavone compound of separating out or the sour water certain dissolubility is arranged, reduced product yield.So not too be suitable for suitability for industrialized production.
Ethanol is that the most frequently used flavone compound extracts solvent.Though other organic solvent selectivitys are strong, volatilization easily, how inflammable (except the chloroform), generally poisonous, price is more expensive, and equipment requirements is also than higher, action need has the ventilation installation, and when a large amount of extraction herbal raw materials or suitability for industrialized production, directly using this kind solvent has certain limitation.
Take all factors into consideration the each side factor, we select for use relatively commonly used, relatively more classical, easy operating, simple ethanol extraction of technology and water extraction to compare, thereby select reasonable extraction solvent.
1. the extraction of lithocarpus litseifolius total flavone
1.1 the extraction choice of Solvent is got Lithocarpus litseifolius tender leaf coarse powder and is used ethanol and water extraction respectively, serves as to investigate index with the main effective ingredient phlorhizin extracted amount in the lithocarpus litseifolius total flavone, optimizes and extracts solvent preferably.
1.1.1 water boiling and extraction is got Lithocarpus litseifolius tender leaf coarse powder 100g, the water boiling and extraction secondary adds 10 times of water gagings at every turn, each 2 hours, merge extractive liquid, filters, and filtrate is concentrated into thick paste, vacuum drying, weigh, measure its phlorhizin content, calculate the phlorhizin extracted amount.
1.1.2 alcohol reflux is got Lithocarpus litseifolius tender leaf coarse powder 100g, the reflux secondary, add 10 times of amount 70% ethanol at every turn, each 2 hours, merge backflow, filter, decompression filtrate recycling ethanol is concentrated into thick paste, and vacuum drying is weighed, measure its phlorhizin content, be calculated as follows the phlorhizin extracted amount.
Experimental result sees Table 1.
Table 1 water boiling and extraction and alcohol reflux are relatively
Experimental result shows that ethanol extraction is good than water extraction.
Ethanol safety, environmental protection is easy to suitability for industrialized production: ethanol is the most frequently used organic solvent of Chinese medicine extraction, and its solubility property is good, and is stronger to the penetration power of Chinese herbal medicine cell; Concentration of ethanol can also change according to the character that is extracted material, adopts Different concentrations of alcohol to extract; Lack than water consumption with ethanol extraction, extraction time is short, and it is also few to dissolve the water-solubility impurity that; Though ethanol is inflammable, toxicity is little, low price, and convenient sources has a locking equipment can reclaim repeatedly and uses; Ethanol extract is difficult for moldy metamorphism.Therefore, ethanol is the widest a kind of solvent of range of application in laboratory and the commercial production, is always to extract the most frequently used a kind of solvent.
The flavonoid chemical constituent is soluble in ethanol: the main effective ingredient of Lithocarpus litseifolius is the dihydrochalcone flavonoid, is soluble in ethanol, so few with the big impurity of ethanol extraction content.Generally speaking, ethanol is beneficial to the stripping of flavone effective ingredient, safety, and environmental protection, and technological process is simple, and the extraction ratio height is easy to suitability for industrialized production.
Take all factors into consideration, extracting solvent is good with ethanol.
1.2 the selection of extracting method is adopted the research that experimentizes of cold-maceration, percolation, circumfluence method, ultrasonic method respectively with ethanol extraction, serves as to investigate index with the extracted amount of phlorhizin, relatively these 4 kinds of methods optimize preferable extracting method.
1.2.1 cold-maceration is got Lithocarpus litseifolius tender leaf coarse powder 100g, room temperature dipping secondary adds 10 times of amount 70% ethanol, each 24 hours at every turn, and stir constantly, merge lixiviating solution, filter, decompression filtrate recycling ethanol is concentrated into thick paste, vacuum drying, weigh, measure the content of its phlorhizin, calculate the phlorhizin extracted amount.
1.2.2 percolation is got Lithocarpus litseifolius tender leaf coarse powder 100g, add an amount of 70% ethanol, flood after 24 hours, with the speed percolation (shared 20 times of amount 70% alcohol dipping, percolation) of 1～3mL/min, collect percolate, the percolate decompression recycling ethanol is concentrated into thick paste, vacuum drying, weigh, measure the content of its phlorhizin, calculate the phlorhizin extracted amount.
1.2.3 heating reflux method is got Lithocarpus litseifolius tender leaf coarse powder 100g, the reflux secondary adds 10 times of amount 70% ethanol, each 2 hours at every turn, merge backflow, filter, decompression filtrate recycling ethanol is concentrated into thick paste, vacuum drying, weigh, measure the content of its phlorhizin, calculate the phlorhizin extracted amount.
1.2.4 the supersound process method is got Lithocarpus litseifolius tender leaf coarse powder 100g, supersound process (power 80W, frequency 50KHz) secondary adds 10 times of amount 70% ethanol at every turn, each 1 hour, merge extractive liquid, filters, and decompression filtrate recycling ethanol is concentrated into thick paste, vacuum drying, weigh, measure the content of its phlorhizin, calculate the phlorhizin extracted amount.Experimental result sees Table 2.
The comparison of the different ethanol extraction methods of table 2
Experimental result shows, ultrasonic method＞circumfluence method＞percolation＞cold-maceration.Ultrasonic method is the easiest, quick, and extraction effect is also better, is experimental extraction and assay method preferably.Though but the leaching process of the intervention efficient hardening total flavones effective ingredient of ultrasonic field shortens extraction time, improve extraction ratio, ultrasonic equipment is difficult to industrialization at present, only suitable laboratory research, and be not suitable for large batch of commercial production.The extraction effect of circumfluence method is only second to ultrasonic method in above-mentioned comparison, device therefor is also simpler, extracting cycle is not long, quantity of solvent is also little, quality and content generation harmful effect to effective ingredient are less, in the big production of industry, more often use, easily carry out fairly large commercial production, a kind of effective ways that the Lithocarpus litseifolius flavone effective ingredient industry of can yet be regarded as is extracted.And the effect of cold-maceration and percolation all is not so good as circumfluence method.
Take all factors into consideration, extracting method is good with heating reflux method.
1.3 the optimization of ethanol extraction process conditions serves as to investigate index with the extracted amount of phlorhizin, adopts orthogonal test, the optimum extraction process condition of preferred reflux extraction.
1.3.1 Orthogonal Experiment and Design and factor level are chosen under material degree of grinding and the solvent for use certain condition, factors such as the concentration of solvent, return time, extraction temperature, extraction time and solvent amount all can exert an influence to extraction, it is to extract solvent that ethanol is selected in this test for use, in order not destroy effective ingredient as much as possible, the reflux, extract, temperature is controlled at 80-90 ℃, therefore, choose concentration of alcohol, ethanol consumption, extraction time, extraction time as the investigation factor, 3 levels of each factor design are pressed L
9(3
4) test.The factor level table sees Table 3.
Table 3 factor level table
1.3.2 test method takes by weighing with 9 parts of a collection of Lithocarpus litseifolius tender leaf coarse powder, every part of 100g carries out reflux, extract, by orthogonal design, extracting solution reclaims ethanol and is concentrated into thick paste, and vacuum drying is weighed, calculate the rate of extract, measure the content of phlorhizin, calculate the phlorhizin extracted amount.The results are shown in Table 4.
Table 4 L
9(3
4) the orthogonal test computer chart
Analyze orthogonal experiments through direct visual comparison, actual in conjunction with producing simultaneously, the optimum process condition that lithocarpus litseifolius total flavone extracts is A2B2C2D3, it is Lithocarpus litseifolius, add 10 times of amount 70% alcohol reflux three times, each 2 hours, the reflux, extract, temperature was controlled at 80-90 ℃.
1.3.3 the optimum extraction condition property confirmed test
Get with a collection of Lithocarpus litseifolius tender leaf coarse powder, take by weighing 100g, totally 3 parts, after extracting by above-mentioned preferred preferred plan, concentrate, dry, measure the content of phlorhizin, calculate the phlorhizin extracted amount, the results are shown in Table 5.
The table 5 optimum extraction scheme property confirmed result of the test
Result of the test shows, extracts by the process conditions that preferably obtain, and the stable content of its phlorhizin illustrates that in more excellent level the process conditions reasonably stability that preferably obtains is feasible.
This extraction process is simple, and it is low to consume energy, and mainly adopts ethanol extraction in technical process, makes the less residue of hazardous solvent in the raw material, meets the requirement of country to food and medicine.
2. the separation purifying technique of lithocarpus litseifolius total flavone research
2.1 the extraction of lithocarpus litseifolius total flavone
Get Lithocarpus litseifolius tender leaf coarse powder 300g, add 10 times of amount 70% alcohol reflux three times, each 2 hours, merge extractive liquid,, filter, decompression filtrate recycling ethanol adds water to 300mL (being that original pharmaceutical content is 1g/mL) to there not being the alcohol flavor, shake up, measure its phlorhizin content and be 76.80mg/mL, standby.
2.2 the separation and purification of lithocarpus litseifolius total flavone
The separation method of flavone compound generally has: polyamide absorption method, solvent extraction, lead salt method, boric acid complexometry, PH gradient extraction, Amberlyst process, column chromatography etc.
[2]And the isolation and purification method of lithocarpus litseifolius total flavone mainly contains the macroporous adsorbent resin partition method
[9-10], ion-exchange-resin process
[3]
The required solvent of above-mentioned some method ubiquity, apparatus expensive, the shortcoming that method is loaded down with trivial details, and can't carry out large-scale separation and purification to lithocarpus litseifolius total flavone, in actual production, be difficult to be applied.
Solvent extraction is better simply separation method, and macroporous adsorbent resin method and polyamide absorption method are not only simple to operate, and price is low, the general processing of commercially available process just can be used, and generally uses the ethanol water eluting, do not use more valuable solvent, environmentally friendly, equipment requirements is not high yet, and the separation and purification effect is good, uses more in actual industrial production.So select for use above three kinds of methods to compare, optimize wherein isolation and purification method preferably.
2.2.1 the selection of isolation and purification method
With the content of main effective ingredient phlorhizin and the purity of total flavones is performance assessment criteria, and relatively solvent extraction, macroporous adsorbent resin method and polyamide absorption method are to the separation and purification effect of lithocarpus litseifolius total flavone.
2.2.1.1 macroporous adsorbent resin pretreatment
Get macroporous adsorbent resin, with 95% alcohol solution dipping resin 24 hours, fully adorn post with wet method after the swelling, (V is the resin volume with 2BV with 95% ethanol, down with)/flow velocity of h cleans, mix (1: 5) with water to effluent and be not white in color till the muddiness, the reuse distilled water cleans pillar to effluent does not have the alcohol flavor, standby.Be unit with resin volume after the swelling (mL) in water when measuring resin.
2.2.1.2 polyamide pretreatment
Get polyamide, with 95% alcohol solution dipping resin 24 hours, fully adorn post with wet method after the swelling, clean with the flow velocity of 95% ethanol with 2BV/h, it is transparent to be washed till effluent, and the reuse distilled water cleans pillar to effluent not to be had alcohol and distinguish the flavor of, standby.Be unit with resin volume after the swelling (mL) in water when measuring resin.
2.2.1.3 Solvent Extraction Separation
Get the extracting solution 20mL under 4.1, use the petroleum ether extraction secondary, each 20mL, petroleum ether extraction liquid discards, reuse water-saturated n-butanol extraction three times, each 20mL, combining extraction liquid, the reclaim under reduced pressure n-butyl alcohol, vacuum drying, the content and the total flavones purity of mensuration phlorhizin.
2.2.1.4 macroporous adsorbent resin separates
Get macroporous adsorbent resin (DM-130) 100mL that has handled well, wet method dress post (3.4cm * 45cm), get the extracting solution 20mL under 4.1, add water to 100mL, stir evenly, placed 24 hours, filter, filtrate is passed through resin column continuously with the flow velocity of 2BV/h, with the water elution of 2BV, water elution liquid discards, with the flow velocity eluting of 50% ethanol with 2BV/h, collect the eluent of 25BV, the eluent decompression recycling ethanol is to thick paste, vacuum drying, the content and the total flavones purity of mensuration phlorhizin.
2.2.1.5 polyamide separates
Get polyamide (100-200 order) 100mL that has handled well, wet method dress post (3.4cm * 45cm), get the extracting solution 20mL under 4.1, add water to 100mL, stir evenly, placed 24 hours, filter, filtrate is passed through resin column continuously with the flow velocity of 2BV/h, with the water elution of 2BV, water elution liquid discards, with the flow velocity eluting of 50% ethanol with 2BV/h, collect the eluent of 25BV, the eluent decompression recycling ethanol is to thick paste, vacuum drying, the content and the total flavones purity of mensuration phlorhizin.
Three kinds of separation method comparative results see Table 6.
Three kinds of different separation method result of the tests of table 6
Experimental result shows, in three kinds of methods, polyamide method separating effect is preferable, lithocarpus litseifolius total flavone after polyamide separates, phlorhizin content and total flavones purity are all than the height of all the other two kinds of methods, and total flavones purity meets national effective site new drug and declares the regulation requirement greater than 50%, so the polyamide method is adopted in the separation and purification of lithocarpus litseifolius total flavone.
2.2.2 the screening of polyamide
By polyamide the suitableeest resin specification is screened in the research of the static adsorption-elution property of phlorhizin in the lithocarpus litseifolius total flavone to different size.Get the extracting solution 30mL under 4.1, add water to 150mL, stir evenly, placed 24 hours, filter, filtrate is divided equally 3 parts, as test liquid.Measure each 20mL of polyamide of the different size of having handled well (30-60 order, 80-100 order, 100-200 order), place 100mL tool plug ground conical flask respectively, add above-mentioned test liquid respectively, flood 24 hours (once), fully after the absorption every jolting in 4 hours, filter, filtrate decompression concentrates, and vacuum drying is weighed, measure index components phlorhizin content wherein, be calculated as follows static saturated extent of adsorption.Filter the gained resin and place 100mL tool plug ground conical flask in addition, add 50% ethanol 50mL, every jolting in 20 minutes 1 time, continue 4 hours, left standstill then 12 hours, filter, filtrate decompression concentrates, and vacuum drying is weighed, measure index components phlorhizin content wherein, be calculated as follows static eluting rate.The results are shown in Table 7.
The static adsorption of table 7 different size polyamide-eluting research
Experimental result shows: polyamide 80-100 order and 100-200 purpose adsorbance are apparently higher than the 30-60 order, though 100-200 purpose adsorbance is the highest, but eluting rate is lower, and because its toner is meticulous, the too slow and obstruction easily of flow velocity, therefore, take all factors into consideration the static adsorption and the elution property of each resin, select the suitable resin specification of 80-100 purpose polyamide as the lithocarpus litseifolius total flavone separation and purification.
2.2.3 the investigation of polyamide adsorption conditions
2.2.3.1 the investigation of sample solution concentration
Get the extracting solution 10mL under 4.1, totally 4 parts, dilution is made into the solution of 2.5 times, 5 times, 7.5 times, 10 times concentration respectively, shakes up, and places 24 hours, filters, and filtrate is respectively as sample solution.Measure polyamide (80-100 order) 20mL that has handled well, totally 4 parts, difference wet method dress post (1.3cm * 30cm), get above-mentioned sample solution with same flow velocity difference upper prop, collected the post effluent, concentrating under reduced pressure, vacuum drying, weigh, measure index components phlorhizin content wherein, be calculated as follows dynamic saturated extent of adsorption.The results are shown in Table 8.
The dynamic saturated extent of adsorption of the different sample solution concentration of table 8
Experimental result shows, when dilute 10 times concentration (be that every mL contains phlorhizin 7mg in the sample solution, be equivalent to the 0.1g crude drug in whole approximately), dynamic saturated extent of adsorption maximum is so selection phlorhizin concentration is that (sample solution that promptly is equivalent to 0.1g crude drug in whole/mL) is good to 7mg/mL.
2.2.3.2 the investigation of last sample flow velocity
Get the extracting solution 40mL under 4.1, add water to 400mL, stir evenly, placed 24 hours, filter, filtrate is divided into 4 parts, as sample solution.Measure polyamide (80-100 order) 20mL that has handled well, totally 4 parts, wet method is adorned post (1.3cm * 30cm), get above-mentioned sample solution and go up sample absorption respectively respectively, control certain flow rate (get flow velocity respectively be 1,2,3,4BV/h), collect effluent, concentrating under reduced pressure, vacuum drying, weigh, measure index components phlorhizin content wherein, calculate dynamic saturated extent of adsorption, the results are shown in Table 9.
The dynamic saturated extent of adsorption of not the same sample speed of table 9
Experimental result shows that along with the quickening of upper prop absorption flow velocity, dynamically saturated extent of adsorption is also along with minimizing, and when the absorption flow velocity was 1BV/h, dynamically the saturated extent of adsorption maximum was 1BV/h so select the absorption flow velocity.
2.2.3.3 the selection of applied sample amount
According to the experimental result under the 4.2.3.2 item, when select best sample solution concentration be 7.68mg phlorhizin/mL (promptly be equivalent to 0.1g crude drug in whole/mL), when upper prop absorption flow velocity is 1BV/h, dynamically saturated extent of adsorption is 22.92mg/mL, be every mL polyamide (80-100 order) to the saturated extent of adsorption of phlorhizin it is 22.92mg (being equivalent to the 0.3g crude drug in whole approximately), therefore, upper prop phlorhizin amount (mg) should＜23: 1 (promptly being equivalent to 0.3g crude drug in whole/mL resin) with the ratio of used resin (mL).
2.2.3.4 determining of optimal adsorption condition
The experimental result of comprehensive above adsorption conditions, the optimal adsorption condition of polyamide (80-100 order) purification lithocarpus litseifolius total flavone is: upper prop phlorhizin amount (mg) answered for＜23: 1 (promptly being equivalent to 0.3g crude drug in whole/mL resin) with the ratio of used resin (mL), sample solution phlorhizin concentration is that 7mg/mL (is equivalent to that the 0.1g crude drug in whole/mL), last column flow rate is 1BV/h.Can reach the optimal adsorption effect like this.
2.2.4 the investigation of polyamide elution requirement
2.2.4.1 the investigation of eluant strength
Get the extracting solution 40mL under 4.1, add water to 400mL, stir evenly, placed 24 hours, filter, filtrate is divided equally 4 parts, as sample solution.Measure polyamide (80-100 order) 20mL that has handled well, totally 4 parts, (1.3cm * 30cm), get the flow velocity respectively upper prop absorption of above-mentioned sample solution with 1BV/h carries out eluting with 20%, 40%, 60%, 80% ethanol with same elution flow rate respectively to wet method dress post then respectively, collect the eluent of 25BV respectively, the eluent concentrating under reduced pressure, vacuum drying is weighed, measure index components phlorhizin content wherein, relatively the total amount of phlorhizin in the eluent.The results are shown in Table 10.
The elute effect of table 10 Different concentrations of alcohol solution
Experimental result shows, during with 20% ethanol, 40% ethanol, 60% ethanol elution, the phlorhizin amount does not have evident difference in the eluent, but all than the height with 80% ethanol elution, consider the factors such as cost of solvent, so select 20% ethanol to be the suitableeest eluant strength.
2.2.4.2 the investigation of eluant consumption
Get the extracting solution 10mL under 2.1, add water to 100mL, stir evenly, placed 24 hours, filter, filtrate is as sample solution.Measure polyamide (80-100 order) 20mL that has handled well, wet method dress post (1.3cm * 30cm), get of the flow velocity upper prop absorption of above-mentioned sample solution with 1BV/h, carry out eluting with 20% ethanol with same elution flow rate then, eluent, concentrating under reduced pressure are collected in segmentation (0-10BV, 10BV-15BV, 15BV-20BV, 20BV-25BV, 25BV-30BV), vacuum drying, weigh, measure index components phlorhizin content wherein, relatively the total amount of phlorhizin in the eluent.The results are shown in Table 11.
20% alcoholic acid elute effect of table 11 different volumes
Experimental result shows that when effluent volume increased, the amount of phlorhizin was also along with increase.After volume reaches 20BV, increase slowly.Therefore considering aspects such as the consumption of solvent, consuming time, cost, is 20BV so select the eluant consumption of the best.
2.2.4.3 the investigation of eluant flow velocity
Get the extracting solution 40mL under 4.1, add water to 400mL, stir evenly, placed 24 hours, filter, filtrate is divided equally 4 parts, as sample solution.Measure polyamide (80-100 order) 20mL that has handled well, totally 4 parts, respectively wet method dress post (1.3cm * 30cm), get of the flow velocity respectively upper prop absorption of above-mentioned sample solution with 1BV/h, use then 20% ethanol respectively with 1,2,3, the speed eluting of 4BV/h, collect the eluent of 20BV respectively, concentrating under reduced pressure, vacuum drying is weighed, measure index components phlorhizin content wherein, relatively the total amount of phlorhizin in the eluent.The results are shown in Table 12.
The elute effect of the different elution speeds of table 12
Experimental result shows that when elution flow rate was 2BV/h, the phlorhizin total amount was maximum in the eluent, and promptly elute effect is best, so determine that elution flow rate is 2BV/h.
2.2.4.4 determining of optimum washing engaging condition
The experimental result of comprehensive above elution requirement, the optimum washing engaging condition of polyamide (80-100 order) purification lithocarpus litseifolius total flavone is: with 20% ethanol is eluant, and elution flow rate is 2BV/h, and the eluant consumption is 20BV.Can reach best elute effect like this.
The preparation technology of 3 lithocarpus litseifolius total flavones determines
Process is to the systematic study and the amplification demonstration test of the separation purifying technique of lithocarpus litseifolius total flavone, the polyamide specification that filters out the separation and purification best results is the 80-100 order, its optimum process condition is: upper prop phlorhizin amount (mg) answered for＜23: 1 (promptly being equivalent to 0.3g crude drug in whole/mL resin) with the ratio of resin (mL), phlorhizin concentration is that 7.68mg/mL (promptly is equivalent to 0.1g crude drug in whole/mL) in the sample solution, the absorption flow velocity is 1BV/h, the eluant concentration of alcohol is 20%, elution flow rate is 2BV/h, the eluant consumption is 20BV, the purity of the lithocarpus litseifolius total flavone that obtains behind the purification can reach more than 80% like this, and phlorhizin content reaches more than 65%.
The present invention has following advantage:
1. the technical scheme of the preparation of lithocarpus litseifolius total flavone of the present invention, the stable content of its phlorhizin is in more excellent level, and the process conditions reasonably stability is feasible.
2. this extraction process is simple, and it is low to consume energy, and mainly adopts ethanol extraction in technical process, makes the less residue of hazardous solvent in the raw material, meets the requirement of country to food and medicine.
3. the lithocarpus litseifolius total flavone that obtains with the present invention is a primary raw material, and the preparation hypoglycemic drug can be provided convenience to the diabetes patient, the more acceptable and oral antidiabetic drug preparation taken for a long time.
Specific embodiment
Embodiment 1
Get Lithocarpus litseifolius tender leaf coarse powder 1500g, add 10 times of amounts 70% (volumetric concentration, below identical) alcohol reflux three times, each 2 hours, merge extractive liquid, filters, and decompression filtrate recycling ethanol is not to there being the alcohol flavor, and thin up is to 15000mL, stir evenly, placed 24 hours, filter, filtrate is as sample solution; Measure polyamide (80-100 order) 5100mL that has handled well respectively, wet method dress post, upper prop phlorhizin amount (mg) is 23: 1 with the ratio of resin volume (mL), sample solution adsorbs with the flow velocity of 1BV/h, water elution with 2BV, water elution liquid discards, with the flow velocity eluting of 20% ethanol, collect the eluent of 20BV, the eluent decompression recycling ethanol with 2BV/h, concentrate, vacuum drying obtains blood sugar lowering extract of the present invention, measurement result, the purity 88.77% of total flavones, phlorhizin content 75.39%.
The said extracted thing is made (capsule, tablet, granule, oral liquid etc.) hypoglycemic pharmaceutical preparation.
Embodiment 2
Get Lithocarpus litseifolius cured leaf coarse powder 100g, add 10 times of amount 70% alcohol reflux three times, each 2 hours, merge extractive liquid, filtered, and decompression filtrate recycling ethanol is not to there being the alcohol flavor, and thin up stirs evenly to 1000mL, places 24 hours, filters, and filtrate is as sample solution; Measure polyamide (80-100 order) 340mL that has handled well respectively, wet method dress post, upper prop phlorhizin amount (mg) is 23: 1 with the ratio of resin volume (mL), sample solution adsorbs with the flow velocity of 1BV/h, water elution with 2BV, water elution liquid discards, and with the flow velocity eluting of 20% ethanol with 2BV/h, collects the eluent of 20BV, the eluent decompression recycling ethanol, concentrate, vacuum drying obtains blood sugar lowering extract-lithocarpus litseifolius total flavone of the present invention.Measurement result, the purity 85.13% of total flavones, phlorhizin content 67.46%.
Embodiment 3
Get Lithocarpus litseifolius tender leaf coarse powder 300g, add 10 times of amount 70% alcohol reflux three times, each 2 hours, merge extractive liquid, filtered, and decompression filtrate recycling ethanol is not to there being the alcohol flavor, and thin up stirs evenly to 3000mL, places 24 hours, filters, and filtrate is as sample solution; Measure polyamide (80-100 order) 1050mL that has handled well respectively, wet method dress post, upper prop phlorhizin amount (mg) is 23: 1 with the ratio of resin volume (mL), sample solution adsorbs with the flow velocity of 1BV/h, water elution with 2BV, water elution liquid discards, and with the flow velocity eluting of 20% ethanol with 2BV/h, collects the eluent of 20BV, the eluent decompression recycling ethanol, concentrate, vacuum drying obtains blood sugar lowering extract-lithocarpus litseifolius total flavone of the present invention.Measurement result, the purity 86.74% of total flavones, phlorhizin content 69.53%.
Lithocarpus litseifolius total flavone is applied to prepare the medicine or the health food of the various preparations of blood sugar lowering, as preparations such as capsule, tablet, granule, oral liquids.
Capsule with above-mentioned extract is made carries out toxicology, pharmacodynamics and clinical trial, and the result is as follows:
Long term toxicity test
Summary is given the rat oral gavage administration respectively with 120 times, 60 times, 30 times three dosage of clinical daily dose, and every day 1 time, administration is 6 days weekly, continuous 26 weeks.Each treated animal hematology, blood biochemical index, organ coefficient, body weight gain and histopathology index there is no overt toxicity and sexually revise, and each index is all in normal range.Show that it should be safe that this product is used with dosage regimen by clinical plan.
Be subjected to the reagent thing: Lithocarpus litseifolius fluid extract, the suitable crude drug 3.828g of every ml, lot number 20000801.Facing the time spent is diluted to the suspension of desired concn with water, presses the 10ml/kg body weight and irritates stomach.This test dose is represented with " crude drug g/kg body weight ".
Animal: Wistar rat body weight 100 ± 20g, 160, male and female half and half, Colleges Of Traditional Chinese Medicine Of Guangxi zoopery center provides, regular grade, the quality certification number: No. the 11005th, the moving word of osmanthus doctor.Sub-cage rearing, 5 in every cage.Feed the standard particle feedstuff, 25 ± 3 ℃ of test chamber temperature, relative humidity 70 ± 5%.
Reagent and instrument: provide reagent by this nation's biological engineering company limited of Australia, measure blood biochemical respectively with biochemical measurement instrument and blood counting instrument and learn and hematological indices.
Test method: 160 of Wistar rats, divide equally four groups at random, 40 every group, male and female half and half.If matched group, this product 28g/kg, 14g/kg and 7g/kg group (be respectively clinical daily dose 120 times, 60 times and 30 times).Rat is gastric infusion 6 days weekly, once a day, and each 10ml/kg body weight, 26 weeks of continuous use.Observe every day such as feed, activity, feces of animal etc.Preceding 8 weeks, to weigh weekly 1 time, after 8 weeks, per two weeks weigh 1 time, adjust dosage simultaneously.20/group (male and female half and half) of 24 hours execution rats after the last administration.Get hematometry hematology and blood biochemical and learn index.Cut open and get main organs (heart, liver, spleen, lung, kidney, testis or uterus, ovary) and weigh, calculate the organ coefficient of every 100g body weight and do the pathology histological examination.Compare with each index and matched group, use the significance of t ' or t value method check group difference.Remain each 10 rat of every group of male and female, drug withdrawal continues to observe for 2 weeks, in kind observes These parameters.
Result of the test:
Reach the drug withdrawal viewing duration during a administration, each outward appearance behavior, breathing, extremity activity, feces of organizing rat there is no unusually no animal dead.The body weight gain rate is in normal range.
B hematological examination index: red blood cell count(RBC) (RBC), content of hemoglobin (HGB), platelet count (PLT), numeration of leukocyte (WBC), lymphocyte count (LYM), mononuclear cell+acidophil+basophil count (MID), neutrophil cell counting (GRA) and clotting time (CT).Administration 26 week each dosage group the difference that respectively detects index and matched group there are no significant meaning, and all in normal range
(3)
The c blood biochemical is learned and is checked index: total protein content (TP), albumin content (ALB), globulin content (GLO), glutamate pyruvate transaminase (ALT), glutamic oxaloacetic transaminase, GOT (AST), alkali phosphatase (ALP), T-CHOL (T-ch), triglyceride (TG), glucose (GLU), blood urea nitrogen (Urea), creatinine (Cr).Respectively detecting in the index of administration 26 week and 2 weeks of drug withdrawal, there are no significant the meaning of the difference between each dosage group and the matched group, and all in normal range
(3)
D main organs coefficient: the difference between 26 weeks of administration and drug withdrawal 2 all each administration groups and the matched group does not have the significance meaning.
E histopathologic examination: all internal organs of each treated animal are not seen the pathological change relevant with drug toxicity.
Conclusion: 120,60 and 30 multiple doses with clinical daily dose are given the rat oral gavage administration respectively, and every day 1 time, administration is 6 days weekly, continuous 26 weeks.Each treated animal hematology, blood biochemical are learned index, organ coefficient, body weight gain and histopathology index and be there is no the pathologic change, and each index is all in normal range.Show that it should be safe that this product is used with dosage regimen by clinical plan.
Pharmacodynamics test:
60 of male mices are got in the influence of 1 pair of model induced by alloxan mice hyperglycemia, stay 10 to make the blank group, and all the other iv alloxan 80mg/kg body weight are made hyperglycemia model, survey fasting blood sugar after 7 days.Choose 50 of the mice of fasting blood sugar more than 12.0mmol/L, evenly be divided into 5 groups: model control group, glyburide matched group, extract low dosage, middle dosage and high dose group, 10 every group by blood sugar level.Except that blank group and model control group ig distilled water, all the other 4 administration treated animals are given glyburide 0.3g/kg, extract 0.2,0.4 and 0.8g/kg respectively by body weight.The administration volume is the 0.1ml/10g body weight, and be administered once every day, connect to give 7 days, and after the last administration 1.5 hours, the ophthalmic corner of the eyes was got blood, separation of serum.According to test kit description input parameter, survey serum glucose with automatic clinical chemistry analyzer.Compare with model control group, fasting blood sugar reduces the significance meaning, judges to be subjected to the reagent thing to have the blood sugar lowering effect.
As a result, the blood glucose of Lithocarpus litseifolius height, middle dosage group and glyburide group all is lower than model control group, and group difference has significance meaning (table 1).Prompting, extract has the blood sugar lowering effect.
Table 1 extract is to the influence of alloxan type hyperglycemia mice fasting glucose (x ± s)
Annotate: * P＜0.05, * * P＜0.01, compare with model control group * * * P＜0.001, down together
The influence of 2 pairs of hyperglycemia mice carbohydrate tolerance: the mice fasting 24 hours iv give alloxan, make hyperglycemia model, modeling type success back group technology is with 2.1, respectively organize mice fasting 3～5 hours after the grouping, then extract and glyburide group respectively ig give the extract and the glyburide of various dose, model control group gives distilled water, and after 20 minutes, each is organized ig and gives glucose solution 2.0g/kg.Measure 0,0.5,2 hour blood glucose value after giving glucose solution, calculate each animal blood glucose area under curve (=0.25 * (0 hour blood glucose value+4 * 0.5 hour blood glucose value+3 * 2 hours blood glucose value).Compare with model control group,, judge to be subjected to the reagent thing to have the effect of the carbohydrate tolerance of raising if the area reduction has the significance meaning under the blood glucose curve.
As a result, area all is lower than model control group under the blood glucose curve of Lithocarpus litseifolius height, middle dosage group and glyburide group, and group difference has significance meaning (table 2).Prompting, extract have the effect of the mice of raising carbohydrate tolerance.
Table 2 extract is to the influence of alloxan type hyperglycemia mice carbohydrate tolerance (x ± s)
3 pairs of streptozotocins bring out the influence of rat hyperglycemia.Get 60 of male rats, stay 10 to make the blank group, all the other iv streptozotocin 80mg/kg body weight are made hyperglycemia model, survey fasting blood sugar after 7 days.Choose 50 of the rat of fasting blood sugar more than 12.0mmol/L, evenly be divided into 5 groups: model control group, glyburide matched group, extract low dosage, middle dosage and high dose group, every group 10 Mus by blood sugar level.Except that blank group and model control group ig distilled water, all the other 4 administration treated animals are given glyburide 0.15g/kg, extract 0.1g, 0.2 and 0.4g/kg respectively by body weight.The administration volume is the 1.0ml/100g body weight, and be administered once every day, connect to give 7 days, and after the last administration 1.5 hours, the ophthalmic corner of the eyes was got blood, separation of serum.According to test kit description input parameter, survey serum glucose with automatic clinical chemistry analyzer.With model control group relatively, fasting blood sugar reduces the significance meaning, judges to be subjected to the reagent thing to have the blood sugar lowering effect.
As a result, the blood glucose of Lithocarpus litseifolius height, middle dosage group and glyburide group all is lower than model control group, and group difference has significance meaning (table 3), prompting, and extract has the blood sugar lowering effect.
Table 3 extract is to the influence of streptozotocin type hyperglycemic rat fasting glucose (x ± s)
The influence of 4 pairs of hyperglycemic rat carbohydrate tolerance: the rat fasting gave streptozotocin 80mg/kg intravenous injection modeling in 24 hours, modeling type success back group technology is with 2.3, respectively organize rat fasting 3～5 hours after the grouping, the other ig of glyburide and extract components gives the extract and the glyburide of various dose then, blank and model control group give distilled water, after 20 minutes, each group is irritated stomach and is given glucose solution 2.0g/kg.Measure 0,0.5,2 hour blood glucose value after giving glucose solution, calculate each animal blood glucose area under curve (=0.25 * (0 hour blood glucose value+4 * 0.5 hour blood glucose value+3 * 2 hours blood glucose value).Compare with model control group,, judge to be subjected to the reagent thing to have the effect of the carbohydrate tolerance of raising if the area reduction has the significance meaning under the blood glucose curve.
As a result, area all is lower than model control group under the blood glucose curve of Lithocarpus litseifolius height, middle dosage group and glyburide group, and group difference has significance meaning (table 4).Prompting, extract have the effect of the rat of raising carbohydrate tolerance.
Table 4 extract is to the influence of streptozotocin type hyperglycemic rat carbohydrate tolerance (x ± s)
Clinical trial
Object choice: the age was 45～75 years old, non-insulin-dependent diabetes mellitus 20 examples.Each case was not all used insulin.Blood glucose is respectively at 11mmol/L-28mmol/L.Man's 10 examples, women 10 examples, the elder of the course of disease 12 years, the shortest person 2 years all has typical " more than three " to show blood glucose soprano 28mmol/L 5 examples.All case does not have severe cardiac, liver, kidney complication, and no ketoacidosis performance in month.
Diagnostic criteria: carry out with reference to Ministry of Public Health " the clinical research guideline of new Chinese medicine treatment diabetes ".
Observational technique: each patient measures fasting blood sugar before medication, gives the Lithocarpus litseifolius capsule oral then, each 3, every day 3 times, medication 1 week and after 1 month, check blood glucose, and write down other symptom and signs (comprise polydipsia, polyphagia, polyuria, become thin, weak etc.).
Criterion of therapeutical effect: with reference to the efficacy assessment standard of " the clinical research guideline of treatment diabetes (the diabetes) " defined in " the new Chinese medicine clinical research guideline " of Ministry of Public Health formulation in 1993.Produce effects: transference cure, fasting glucose are reduced to below the 7.2mmol/L, and 2 hours after the meal blood glucose is below 8.3mmol/L, and twenty-four-hour urine sugar is below 10.0g, and perhaps fasting glucose descends more than 55% before the treatment; Effectively: clinical symptoms is obviously improved, and fasting glucose is reduced to below the 8.3mmol/L, and 2 hours after the meal blood glucose is below 10.0mmol/L, and perhaps fasting glucose descends more than 35% before the treatment; Invalid: treatment back symptom does not have obvious improvement, and blood glucose, glucose in urine descend and do not reach above-mentioned standard.
Therapeutic outcome: 20 routine results show each 3 of Lithocarpus litseifolius capsule, every day 3 times, and continuous 1 month, significant blood sugar reducing function is arranged, do not see any toxic and side effects (table 1).
The capsular hypoglycemic effect of table 1 Lithocarpus litseifolius
| Index | The example number | Blood glucose before the medication | Blood glucose after the medication | Blood glucose decline % |
| Fasting glucose (mmol/L) | 20 | 11.31±3.31 | 5.51±3.10 | 51 |
| Post-prandial glycemia (mmol/L) | 20 | 16.5±4.23 | 6.30±3.96 | 62 |
| 24h glucose in urine (g) | 20 | 22.3±15.1 | 4.2±21.3 | 81 |
According to the efficacy assessment standard evaluation, Lithocarpus litseifolius capsule curative effect is: produce effects 10 examples (50%), effective 9 examples (45%), invalid 1 example (5%).Total effective rate 95%.
Model case:
Example 1, Cen, accounting, women, 57 years old.Because of xerostomia polydipsia 10 years, increase the weight of to go to a doctor in 1 month.Card is seen: thirst and liking drink, fatigue and weakness, spontaneous sweating, vexed night sweat, vexed insomnia, yellowish urine constipation, red tongue, corpulent tongue, tongue stripping, thready and rapid pulse.Fasting glucose 14.2mmol/L, 2 hours after the meal blood glucose 20.1mmol/L, twenty-four-hour urine sugar is 19.7g quantitatively.Tcm diagnosis: diabetes (syndrome of deficiency of both qi and yin).Western medicine diagnose: diabetes.Giving the Lithocarpus litseifolius capsule takes.Each 3, every day 3 times, after 1 week of taking medicine, xerostomia polydipsia symptom obviously alleviates, fasting glucose 11.2mmol/L, and 2 hours after the meal blood glucose 15.6mmol/L, twenty-four-hour urine sugar is 15g quantitatively.Take medicine after 4 weeks, subjective symptoms disappears substantially, fasting glucose 5.5mmol/L, and 2 hours after the meal blood glucose 5.6mmol/L, twenty-four-hour urine sugar is 1.2g quantitatively.Efficacy evaluation: produce effects.
Example 2, Liao, cadre, male, 62 years old.Because of xerostomia polydipsia 2 years, increase the weight of to go to a doctor in 2 months.Card is seen: thirst and liking drink, fatigue and weakness, yellowish urine constipation, spontaneous sweating, red tongue, tongue stripping, thready and rapid pulse.Laboratory examination: fasting glucose 12.2mmol/L, 2 hours after the meal blood glucose 19.1mmol/L, twenty-four-hour urine sugar is 29.7g quantitatively.Tcm diagnosis: diabetes (syndrome of deficiency of both qi and yin).Western medicine diagnose: diabetes (II type).Give the Lithocarpus litseifolius capsule, continuous 1 month.Take medicine after 1 week, xerostomia polydipsia symptom obviously alleviates, fasting glucose 9.2mmol/L, and 2 hours after the meal blood glucose 10.6mmol/L, twenty-four-hour urine sugar is 6g quantitatively.Take medicine after 4 weeks, subjective symptoms disappears substantially, fasting glucose 5.2mmol/L, and 2 hours after the meal blood glucose 6.6mmol/L, twenty-four-hour urine sugar is 0.8g quantitatively.Efficacy evaluation: produce effects.
Example 3, Zhang, technician, male, 56 years old.Because of polydipsia, polyphagia, polyuria surplus prescription on individual diagnosis in 2 years.The main suit is polydipsia, polyphagia, polyuria, dry mouth and throat over 2 years, and gomphiasis is easily starved fatigue and weakness.Check, red tongue with a little fluid, corpulent tongue is big, thin fur and white, thready and rapid pulse; Laboratory examination: fasting glucose 13.2mmol/L, 2 hours after the meal blood glucose 18.1mmol/L, twenty-four-hour urine sugar is 17.9g quantitatively.Tcm diagnosis: diabetes (syndrome of deficiency of both qi and yin).Western medicine diagnose: diabetes (II type).Give the Lithocarpus litseifolius capsule, continuous 1 month.Take medicine after 1 week, xerostomia, polydipsia, polyphagia, polyuria symptom obviously alleviate, fasting glucose 8.2mmol/L, and 2 hours after the meal blood glucose 12.6mmol/L, twenty-four-hour urine sugar is 10g quantitatively.Take medicine after 4 weeks, subjective symptoms disappears substantially, fasting glucose 5.9mmol/L, and 2 hours after the meal blood glucose 6.6mmol/L, twenty-four-hour urine sugar is 1.0g quantitatively.Efficacy evaluation: produce effects.
Example 4, deep certain, skilled industrial worker, male, 66 years old.Because of polydipsia, polyphagia, polyuria surplus prescription on individual diagnosis in 6 years.The main suit is polyuria, polydipsia, polyphagia over 6 years, and gomphiasis is easily starved fatigue and weakness.Check that corpulent tongue is big, red tongue with a little fluid, thready and rapid pulse; Laboratory examination: fasting glucose 18.2mmol/L, 2 hours after the meal blood glucose 22.1mmol/L, twenty-four-hour urine sugar is 27.9g quantitatively.Tcm diagnosis: diabetes (syndrome of deficiency of both qi and yin).Western medicine diagnose: diabetes (II type).Give the Lithocarpus litseifolius capsule, continuous 1 month.Take medicine after 1 week, xerostomia, polydipsia, polyphagia, polyuria symptom obviously alleviate, fasting glucose 9.2mmol/L, and 2 hours after the meal blood glucose 14.2mmol/L, twenty-four-hour urine sugar is 12.3g quantitatively.Take medicine after 4 weeks, subjective symptoms disappears substantially, fasting glucose 6.3mmol/L, and 2 hours after the meal blood glucose 7.2mmol/L, twenty-four-hour urine sugar is 1.5g quantitatively.Efficacy evaluation: produce effects.Example 5, beam, cadre, male, 62 years old.Because of xerostomia polydipsia more than 10 year, increase the weight of to go to a doctor in 1 month.Card is seen: thirst and liking drink, and polyphagia and unable, hyperhidrosis, vexed insomnia, the yellowish urine constipation, swelling of the tongue, matter are red, tongue, thready pulse.Fasting glucose 16.5mmol/L, 2 hours after the meal blood glucose 23.1mmol/L, twenty-four-hour urine sugar is 29.2g quantitatively.Tcm diagnosis: diabetes.Western medicine diagnose: diabetes.Giving the Lithocarpus litseifolius capsule takes.Each 3, every day 3 times, after 1 week of taking medicine, xerostomia polydipsia symptom obviously alleviates, fasting glucose 12.2mmol/L, and 2 hours after the meal blood glucose 15.1mmol/L, twenty-four-hour urine sugar is 12g quantitatively.Take medicine after 4 weeks, subjective symptoms disappears substantially, fasting glucose 6.5mmol/L, and 2 hours after the meal blood glucose 6.6mmol/L, twenty-four-hour urine sugar is 1.6g quantitatively.Efficacy evaluation: produce effects.
Claims (5)
1. lithocarpus litseifolius total flavone preparation method, comprise extraction, concentrated, vacuum drying operation, it is characterized in that: be solvent with ethanol, the Lithocarpus litseifolius cured leaf is carried out heating and refluxing extraction, extracting solution is reuse polyamide adsorbing separation after treatment, is that eluant is with the lithocarpus litseifolius total flavone eluting on the polyamide again with ethanol at last.
2. a kind of lithocarpus litseifolius total flavone preparation method according to claim 1 is characterized in that:
(1) alcohol heating reflux extracts:
A. the Lithocarpus litseifolius cured leaf adds the ethanol that 5-15 doubly measures the 20-70% volumetric concentration by weight;
B. reflux, extract, is 1--4 time, each 0.5--3 hour;
C. the reflux, extract, temperature is 50-90 ℃;
(2) polyamide method separation and purification:
A. absorption: the mass volume ratio of upper prop phlorhizin and polyamide＜23: 1; Sample solution phlorhizin concentration is 6-8mg/mL; Last column flow rate is 0.5-2BV/h;
B. eluting: with ethanol is eluant; The eluant consumption is 10-30BV; Elution flow rate is 1-3BV/h.
3. lithocarpus litseifolius total flavone preparation method according to claim 1 is characterized in that:
(1) alcohol heating reflux extracts:
A. the Lithocarpus litseifolius cured leaf adds 10 times of ethanol of measuring 70% volumetric concentrations by weight;
B. reflux, extract, is three times, each 2 hours;
C. the reflux, extract, temperature is 80-90 ℃;
(2) polyamide method separation and purification:
A. absorption: upper prop phlorhizin and polyamide mass volume ratio＜23: 1; Sample solution phlorhizin concentration is 7mg/mL; Last column flow rate is 1BV/h;
B. eluting: 20% ethanol is eluant; The eluant consumption is 20BV; Elution flow rate is 2BV/h.
4. a kind of lithocarpus litseifolius total flavone preparation method according to claim 1 is characterized in that: the polyamide specification is the 80-100 order.
5. the application of lithocarpus litseifolius total flavone aspect preparation blood sugar lowering medicine or health food.
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