CN102488819B - Preparing method for daylily flower extract - Google Patents
Preparing method for daylily flower extract Download PDFInfo
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- CN102488819B CN102488819B CN201110410822.3A CN201110410822A CN102488819B CN 102488819 B CN102488819 B CN 102488819B CN 201110410822 A CN201110410822 A CN 201110410822A CN 102488819 B CN102488819 B CN 102488819B
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Abstract
The invention discloses a preparing method for daylily flower extract, which comprises the steps of extracting ethanol from daylily flowers at a low temperature, enabling ethanol extraction liquid to pass through a macroporous resin column, recovering ethanol to obtain the daylily extract and the like. The preparing method for the daylily flower extract is low in cost, high in efficiency and suitable for mass production.
Description
Technical field
The present invention relates to extract of a kind of Chinese medicine and preparation method thereof, particularly Hemerocallis fulva L. flower extract and preparation method thereof.
Background technology
Hemerocallis fulva L., i.e. Radix hemerocalis plicatae (Hemerocallis citrina Baroni), have another name called " Flos Hemerocallis ", " Bulbus Lilii on the one ", be commonly called as " forgetting melancholy grass ", belong to Liliaceae hemerocallis herbaceos perennial plant, have plantation in China various places, Ming Dynasty's Li Shizhen (1518-1593 A.D.) claims effect that it has tranquilization and refreshment, increasing intelligence the chest stuffiness relieving, heat sterilization, relieving restlessness lactogenesis are nourished blood, separate in beauty treatment in Compendium of Material Medica more.Modern study Hemerocallis fulva L. has supplementing the brain and is good for the effects such as intelligence, removing stasis to stop bleeding, removing obstruction in the collateral to relieve pain, anti-inflammation, and it has enhancing immunity, blood circulation promoting and blood stasis dispelling, protects the liver antiinflammatory, the several functions such as anti-bacteria and anti-virus, Tumor suppression, scavenging activated oxygen and bacteriostasis.It has abundant nutrition, main chemical compositions has protein, fat, carbohydrate, also has multivitamin, alkaloid, glycosides, flavonoid, carotene, volatile oil, tannin and inorganic mineral calcium, phosphorus, take flavone compound as main component, can medicine-food two-purpose, protect the liver antiinflammatory, calm multiple physiologically active effect such as supplementing the brain, slow down aging antidepressant in view of flavone compound has, the research of flavone compound causes the concern of people day by day.For the plant of this medicine-food two-purpose of further development and utilization Hemerocallis fulva L., the extraction process of the total flavones of Hemerocallis fulva L. is worth research and development.
Summary of the invention
The object of the invention is to open a kind of Hemerocallis fulva L. flower extract preparation method;
Another object of the present invention is the fingerprint atlas detection method of open Hemerocallis fulva L. flower extract.
The object of the invention is to be achieved through the following technical solutions:
The preparation method of Hemerocallis fulva L. flower extract of the present invention, comprises the steps:
Step 1: Hemerocallis fulva L. ethanol less than 60 DEG C extraction;
Step 2: ethanol extract crosses macroporous resin column;
Step 3: reclaim ethanol and obtain Hemerocallis fulva L. flower extract.
Preferred Hemerocallis fulva L. ethanol less than 30 DEG C extraction in described step 1;
In described step 1, Hemerocallis fulva L. ethanol extraction is: less than 30 DEG C lixiviates or percolation;
Preferably, the amount that step 1 lixiviate adds ethanol be the 6-12 volume of medical material doubly, ethanol is 60%-95% ethanol; Most preferably be 70% ethanol;
Preferably, step 1 extraction time is 18-48 hour;
Preferably, step 1 alcohol steep, filter, filtering residue adds ethanol again, soaks, and filters; Merging filtrate;
Step 1 is more preferably: will add 8 volumes, 70% ethanol doubly in Hemerocallis fulva L., and 30 DEG C below are soaked 24 hours, and filter, filtrate is for subsequent use, and filtering residue adds 8 volumes, 70% ethanol doubly again, 30 DEG C below immersions 24 hours, filtration, merging filtrate;
Described step 1: the method for percolation is: use 70% ethanol as solvent, less than 30 DEG C flood 24 hours, with the speed percolation of 1-3ml per minute, collect percolate for less than 30 DEG C.
Step 2 is preferred, and ethanol extract liquid crosses macroporous resin column, and it is transparent that distilled water is washed till effluent, with 20%-95% ethanol elution 2-6 times of column volume;
Step 2 is preferred, and macroporous resin column is AB-8 macroporous resin column; The concentration of ethanol elution is: 55%-65%.
Step 2 is preferred, and ethanol cooling bath crosses AB-8 macroporous resin column, and it is transparent that distilled water is washed till effluent, with 60% ethanol elution, 3 times of column volumes, reclaims ethanol.
The present invention also comprises any one pharmaceutical preparation of making as active constituents of medicine with described Hemerocallis fulva L. flower extract.The acceptable any conventional dosage form of pharmaceutics can be made, the preparations such as such as capsule, tablet, granule, powder, oral liquid, pill by preparation process routinely.
The present invention further provides the preparation method of Hemerocallis fulva L. flower extract, it is characterized in that the method comprises the steps:
Step 1: Hemerocallis fulva L. ethanol extracts below 60 DEG C, obtains ethanol extract;
Step 2: ethanol extract crosses macroporous resin column;
Step 3: reclaim ethanol and obtain Hemerocallis fulva L. flower extract.
Step 4: Hemerocallis fulva L. flower extract HPLC finger printing detects.
Preferred Hemerocallis fulva L. ethanol less than 30 DEG C extraction in described step 1;
Described step 1: Hemerocallis fulva L. ethanol extraction is: less than 30 DEG C lixiviates or percolation;
Preferably, the amount that step 1 lixiviate adds ethanol be the 6-12 volume of medical material doubly, ethanol is 60%-95% ethanol; Most preferably be 70% ethanol;
Preferably, step 1 extraction time is 18-48 hour;
Preferably, step 1 alcohol steep, filter, filtering residue adds ethanol again, soaks, and filters; Merging filtrate;
Step 1 is more preferably: will add 8 volumes, 70% ethanol doubly in Hemerocallis fulva L., and 30 DEG C below are soaked 24 hours, and filter, filtrate is for subsequent use, and filtering residue adds 8 volumes, 70% ethanol doubly again, 30 DEG C below immersions 24 hours, filtration, merging filtrate;
Described step 1: the method for percolation is: use 70% ethanol as solvent, less than 30 DEG C flood 24 hours, with the speed percolation of 1-3ml per minute, collect percolate for less than 30 DEG C.
Step 2 is preferred, and ethanol extract liquid crosses macroporous resin column, and it is transparent that distilled water is washed till effluent, with 20%-95% ethanol elution 2-6 times of column volume;
Step 2 is preferred, and macroporous resin column is AB-8 macroporous resin column; The concentration of ethanol elution is: 55%-65%.
Step 2 is preferred, and ethanol cooling bath crosses AB-8 macroporous resin column, and it is transparent that distilled water is washed till effluent, with 60% ethanol elution, 3 times of column volumes, reclaims ethanol.
The present invention also provides the HPLC fingerprint atlas detection method of Hemerocallis fulva L. flower extract:
Chromatographic condition: C18 chromatographic column, mobile phase methanol-water (0.5% glacial acetic acid) gradient is washed, 0.5-1.5mL/min
-1, determined wavelength 200-300nm, column temperature 20-30 DEG C; The condition of described methanol-water (0.5% glacial acetic acid) gradient elution is: 0-20min: methanol be 20%, 0.5% glacial acetic acid aqueous solution be 80%; 20-30min: methanol be 30%, 0.5% glacial acetic acid aqueous solution be 70%; 30-70min: methanol be 30%, 0.5% glacial acetic acid aqueous solution be 70%; 70min is to last: methanol be 80%, 0.5% glacial acetic acid aqueous solution be 20%;
The preparation of reference substance solution: take rutin, adds Methanol and becomes reference substance solution;
The preparation of need testing solution: take Hemerocallis fulva L. flower extract powder, add methanol supersound extraction, leaves standstill, and gets supernatant and filters, get filtrate, to obtain final product.
Detection method: draw reference substance, need testing solution, sample detection.
The preparation method of reference substance solution is preferably: take rutin, adds methanol and makes the solution of every 1mL containing 0.2176mg, shake up, to obtain final product;
The preparation method of need testing solution is preferably: precision takes sample powder 2g, puts in 25mL volumetric flask, adds methanol to scale, supersound process 30 minutes, methanol supplies the weight of loss, leaves standstill and lets cool, get supernatant, 0.22 μm of microporous filter membrane filters, and gets filtrate, to obtain final product.
Described finger printing comprises 12 common characteristic peaks, and be rutin for reference to peak with No. 3, relative retention time and relative peak area are respectively: No. 1 peak relative retention time is 0.232, and relative peak area is: 0.051 ~ 0.278; No. 2 peak relative retention time are 0.936, and relative peak area is: 0.020-0.212; No. 4 peak relative retention time are 1.039, and relative peak area is: 0.023-0.041; No. 5 peak relative retention time are 1.079, and relative peak area is: 0.008-0.178; , No. 6 peak relative retention time are 1.095, and relative peak area is: 0.005-0.029; No. 7 peak relative retention time are 1.115, and relative peak area is: 0.045-0.367; No. 8 peak relative retention time are 1.135, and relative peak area is: 0.049-0.269; No. 9 peak relative retention time are 1.159, and relative peak area is: 0.004-0.023; No. 10 peak relative retention time are 1.171, and relative peak area is: 0.003-0.014; No. 11 peak relative retention time are 1.218, and relative peak area is: 0.012-0.062; No. 12 peak relative retention time are 1.326, and relative peak area is: 0.002-0.313.
Prior art adopted ethanol refluxing process to extract concentrated medical material solution before D101 post is separated, has the precipitation of a large amount of oil-soluble impurities, and thickness is difficult to upper prop.Find in Hemerocallis fulva L. flower extract preparation method of the present invention to adopt ethanol merceration, impurity is few, particularly water-solubility impurity is less, and during large production, dipping medicinal liquid can not block macroporous resin pillar because placing generation precipitation, greatly reduces the pollution to resin, be beneficial to the regeneration of resin, economic and practical, and break through the understanding that merceration in the past can reduce than heating extraction effective ingredient, extraction efficiency is high, not only economical and have high efficiency, be suitable for large production.
The fingerprint atlas detection method of Hemerocallis fulva L. provided by the present invention is that analytical method is effectively reliable and stable by obtaining after a large amount of concrete creative experiment sieving, for the discriminating of Hemerocallis fulva L. and quality evaluation and control provide foundation.
Experimental example 1 ultraviolet spectrophotometry detects general flavone content
The selection of 1 mensuration wavelength
After rutin standard solution and Hemerocallis fulva L. extracting solution distinguish empirically method colour developing, in the interscan of wavelength 400 ~ 600nm scope, have absorption maximum at 510nm place after the colour developing of result rutin standard solution, the colour developing of Hemerocallis fulva L. extracting solution also has absorption at this wavelength, and experiment selects 510nm to measure wavelength.
The preparation of 2 need testing solutions
Precision takes preparation (being prepared by embodiment 1) 20mg, puts in 10mL measuring bottle, and add 30% ethanol to scale, supersound process 10min, to obtain final product.
The preparation of 3 reference substance solution
It is appropriate that precision takes control substance of Rutin, adds methanol and make the solution of every 1mL containing 0.2mg, shake up, to obtain final product.
The preparation of 4 standard curves
Precision measures reference substance solution 0.5ml, 1ml, 2ml, 3ml, 4ml, 5ml, 6ml, put in 25ml measuring bottle respectively, respectively add water to 6.0ml, add 5% sodium nitrite solution 1ml, mixing, place 6 minutes, add 10% aluminum nitrate solution 1ml, shake up, place 6 minutes, hydro-oxidation sodium test solution 10ml, add water to scale again, shake up, place 15 minutes, take corresponding reagent as blank, absorbance is measured at 510nm wavelength place, take absorbance as vertical coordinate, concentration is abscissa, drawing standard curve, obtaining regression equation is Y=13.65X+0.0128, r=0.9997, result shows that rutin concentration has good linear relationship at 0.004026 ~ 0.048312mg/ml and absorbance.
5 assay methods
Get need testing solution 1ml, put in 25ml measuring bottle, the method under sighting target directrix curve preparation, from " adding water to 6.0ml ", measures absorbance in accordance with the law, reads the concentration containing rutin in need testing solution from standard curve, calculates, to obtain final product.
6 methodological study results
6.1 precision test
Obtain test liquid by the method under sample determination item to the medical material process that lot number is 20110530, UV method measures, a sample METHOD FOR CONTINUOUS DETERMINATION 6 times, and the results are shown in Table 1, RSD is 0.16%.
Table 1 Precision test result
6.2 stability test
Get same sample solution, measured an absorbance every 10 minutes, sample is good at 1 hour internal stability, the results are shown in Table 2.
Table 2 stability test result
Result shows, and sample is good at 1 hour internal stability, and RSD value is 1.20%.
6.3 repeatability tests,
Measure by the sample of the method under sample determination item to lot number 20110530, measure 6 parts altogether, the results are shown in Table 3.
Table 3 reproducible test results
Result content meansigma methods is 0.6111, RSD value is 2.90%.
6.4 average recovery tests
Precision takes sample, and (lot number is 20110530, general flavone content 0.61%) 1g, fixed 6 parts of nominal, put in 25mL measuring bottle respectively, add the control substance of Rutin solution 0.4mL of 0.9250mg/ml, prepare need testing solution as stated above, measure content, calculate average recovery, the results are shown in Table 4.
Rutin average recovery in table 4 Hemerocallis fulva L.
Result display average recovery meansigma methods is 101.28%, RSD value is 1.10%.
7 sample determinations
Precision takes three batch samples, presses the preparation method preparation of need testing solution respectively, by above-mentioned determined by ultraviolet spectrophotometry, the results are shown in following table 5:
The assay of table 5 preparation
8 conclusions
The ultraviolet spectrophotometry content assaying method that medicine of the present invention adopts its within the scope of 0.004026 ~ 0.048312mg/ml, r=0.9997, average recovery rate is 101.28%, RSD value is 1.10%, linear relationship is good, stability, precision, repeatability etc. are all good, can effectively control pharmaceutical preparation quality.
The assay of the total flavones of experimental example 2 ethanol heating extraction method sample
Ethanol heating extraction method:
Dry for 9900g Hemerocallis fulva L. is cut into the segment of 2-3 centimeter length, 95% soak with ethanol is spent the night.95% ethanol heating extraction 3 times, timing from boiling, the time is respectively 40min, 30min, 30min, filters, merging filtrate, reclaims ethanol, obtains extractum A.Residue 60% ethanol heating extraction 2 times, each 40min, filters, merging filtrate, reclaims ethanol, obtains extractum B.Filter after extractum A distilled water diluting, by processed good AB-8 macroporous resin column, it is transparent that distilled water is washed till effluent, with 60% ethanol elution, 3 times of column volumes, reclaims ethanol, obtain sample 1.Same treatment extractum B, obtains sample 2.Sample 1 is 64.3101g, and sample 2 is 60.9338g.
Assay method: precision takes the control substance of Rutin (Nat'l Pharmaceutical & Biological Products Control Institute of drying under reduced pressure to constant weight, lot number 100080-200707) 0.0101g, methanol constant volume shakes up in 50ml volumetric flask, accurately measure 0.0,1.0,2.0,3.0,4.0,5.0ml, in 25ml volumetric flask, adds 5%NaNO
26ml, mixing, places 6min, adds 10%Al (NO
3)
30.6ml, mixing, places 6min, adds 4%NaOH solution 5ml, with 60% ethanol dilution to scale, places 15min, by pharmacopeia spectrophotography, with the first pipe for blank, surveys trap at 500nm wavelength place.Take trap as abscissa, concentration is vertical coordinate, drawing standard curve.
Table 6
Regression equation is y=12.401x-0.0021 r=0.9990
Accurately take each 0.0025g of sample 1 and 2, be placed in 50ml volumetric flask, be measured in the same method trap and be respectively 0.11867 and 0.12219, be respectively 0.009739 and 0.010022mg/ml by standard curve calculating concentration.General flavone content is respectively 19.478% and 20.044%.
Experimental example 3 preparation method technical study
1 instrument and material
1.1 instruments: water-bath (Guo Hua Electrical Appliances Co., Ltd, digital display thermostat water bath); TU-1810 ultraviolet-uisible spectrophotometer (Beijing Pu Xi all purpose instrument company limited); Analytical balance (Beijing Pu Xi general finite company); KQ-250B ultrasonic cleaner (Kunshan Ultrasonic Instruments Co., Ltd.).
1.2 materials and medicine: methanol (Beijing chemical analysis is pure); Water (Wahaha Pure Water, deionized water); 95% medical alcohol; Other reagent are analytical pure.Control substance of Rutin (Nat'l Pharmaceutical & Biological Products Control Institute, lot number 100080-200707).Hemerocallis fulva L., purchased from Beijing Kangrentang Medicine Co., Ltd, the market of farm produce, Baoding city, the place of production, Guangdong medical material.
2 methods
2.1.1 test method:
2.1.1.1 spectrophotometric determination method: sample thief liquid 2ml, is placed in 25ml volumetric flask, adds water to 6ml, add 5%NaNO
2solution 1ml, makes mixing, places 6min, adds 10%Al (NO
3)
31ml shakes up, and places 6min, adds 4%NaOH10ml, then the scale that adds water, and shakes up, and places 15min, surveys A at 510nm.
2.1.1.2 the drafting of standard curve:
Precision weighing rutin standard substance 11.30mg (bottle being write UV by 92.5%), close rutin 10.4525mg, be dissolved in 50ml volumetric flask completely with methanol, water standardize solution is the standard substance storing solution of 0.2091mg/ml, for subsequent use.
Precision measure reference substance solution 0,1,2,3,4,5,6ml is placed in 25ml volumetric flask respectively, respectively add water to 6ml, measure according to method under 2.1.1.1 item.Take A as vertical coordinate, concentration is abscissa, drawing standard curve.Regression equation is y=0.0067+0.0118x R=0.9990.
2.1.1.3 average recovery test: sample thief powder, precision takes 5 parts, adds mixed standard solution 1ml respectively, and extract by method for optimizing under 2.2.1 item, measure, recording average recovery rate is 98.11%.
2.1.1.4 precision measures: get a sample solution, replication 5 times, record absorbance, calculates RSD and is respectively 1.91%, show that precision is good.
3. water-boiling method: with the water of medical material 8 times amount, decoct twice, each 1h.It is 5.35mg/g that decocting in water carries total flavones.Because the sample liquid upper prop of water-boiling method process, the amount that water-solubility impurity extracts is large, seriously polluted to resin, is unfavorable for the regeneration of resin, abandons in this way.
4. cold-maceration:
By 30% alcohol dipping, 50% alcohol dipping, 70% alcohol dipping.
Method: put in triangular flask by 30g Hemerocallis fulva L. flower pesticide material (place of production, Guangdong), adds 8 times of volume respective concentration ethanol, soaks 24h, filters; Filtering residue adds 8 times amount ethanol again, soaks 24h, filters; Filtering residue adds 8 times amount ethanol again, soaks 24h, filters.Repetitive operation is to flooding 4 days; Measure the total flavones amount of each filtrate.The results are shown in Table 7.
The determination of total flavonoids of table 7 impregnation liquid
| 30% ethanol | 50% ethanol | 70% ethanol | |
| Flood 24 hours | 0.32％ | 0.37％ | 0.38％ |
| Flood 48 hours | 0.15％ | 0.14％ | 0.17％ |
| Flood 72 hours | 0.04％ | 0.04％ | 0.06％ |
| Flood 96 hours | 0.01％ | 0.02％ | 0.03％ |
| Add up to the medical material content extracted | 0.52％ | 0.57％ | 0.62％ |
Known according to upper table, the extracted amount with the increase total flavones of concentration of alcohol increases gradually, and the extracted amount of 70% ethanol to total flavones is the highest.Therefore continue investigation 80%, 95% alcohol dipping to the impact of total flavones extracted amount.The results are shown in Table 8.
The assay 2 of table 8 impregnation liquid total flavones
| 70％ | 80％ | 95％ | |
| 24 h immersion liquid | 0.38％ | 0.34％ | 0.26％ |
Result shows, and the dipping effect of 70% ethanol is best, soaks two days total flavones and reaches 5.3mg/g, can reach extraction ratio nearly 90%, consider economic factor and extraction efficiency, determines by 70% soak with ethanol 2 times, each 24 hours.
Ethanol cold-maceration proposes impurity and is obviously less than heat reflow method, flood medicinal liquid during large production and can not block macroporous resin pillar because placing generation precipitation, and total flavones extraction ratio nearly 90%, therefore final extraction scheme is decided to be: by 70% ethanol 8 times amount room temperature immersion twice, each 24 hours, merge twice impregnation liquid, less than 80 DEG C decompression recycling ethanols.
Experimental example 4HPLC finger printing is tested
1 instrument and material
SD-20 high performance liquid chromatograph (Japanese Shimadzu Corporation); LC-20ATVP quaternary pump (Japanese Shimadzu); Control substance of Rutin (lot number 100080-200707, Nat'l Pharmaceutical & Biological Products Control Institute); Methanol is chromatographically pure (Fisher company); Water is Wahaha Pure Water, and all the other reagent are analytical pure, tests 10 batch sample sources in table 9.
Table 9 Hemerocallis fulva L. crude drug source
Test 10 batches of medical materials and prepare sample by embodiment 1 method.
2 methods and result
2.1 chromatographic condition
Chromatographic column is Agilent (specification C18,4.6*250mm, 5 μm) mobile phase methanol one water (0.5% glacial acetic acid) gradient elution, condition in table 10, flow velocity 1mL/min-1, determined wavelength 254nm, column temperature 25 DEG C.
Table 10 condition of gradient elution
The preparation of 2.2 reference substance solution: it is appropriate that precision takes control substance of Rutin, adds methanol and makes the solution of every 1mL containing 0.2176mg, shake up, to obtain final product.
The preparation of 2.3 need testing solutions: precision takes sample powder 2g, puts in 25mL volumetric flask, adds methanol to scale, supersound process 30 minutes, and methanol supplies the weight of loss, leaves standstill and lets cool, get supernatant, and 0.22 μm of microporous filter membrane filters, and gets subsequent filtrate, to obtain final product.
2.4 methodological study
2.4.1 precision test: the same need testing solution 10 μ l of accurate absorption, continuous sample introduction 6 pin, measures its RP-HPLC collection of illustrative plates, is with reference to peak (No. 3 peaks) with rutin, calculate the RSD value of each total peak relative retention time for 0.02-2.75, show that method precision is good.
2.4.2 repeatability test: get with a collection of Hemerocallis fulva L. flower pesticide material 6 parts, according to 2.3 below legal system available test sample solutions, sample introduction respectively, records the RSD value of each total peak relative retention time for 0.02-5.94, shows that method repeatability is good.
2.4.3. stability test: accurate draw need testing solution, respectively at 0,2,4,6,8 hour sample introduction after preparation, records the RSD value of each total peak relative retention time for 0.03-2.61, shows that the method is stablized in 8 hours.
The mensuration of 2.5 sample finger printing:
The finger printing of the Hemerocallis fulva L. flower pesticide material of 10 provinces is detected by above-mentioned liquid phase chromatogram condition, respectively the finger printing of 10 batches of medical materials is inputted in " Chinese medicine fingerprint similarity evaluation systematic study version (2004A) " of Chinese Pharmacopoeia Commission's formulation, carry out total peak match, finally obtain 12 common characteristic peaks, see Fig. 1.
2.6 similarity evaluation
2.6.1 according to similarity evaluation software, select rutin to do with reference to peak, each peak relative retention time, relative peak area result, see the following form 11:
The HPLC fingerprint map analyzing result of table 11 10 batches of Hemerocallis fulva L. flower pesticide materials
2.6.2 by the AIA data importing " Chinese medicine fingerprint similarity evaluation systematic study version (2004A) " of 10 batches of medicinal materials fingerprints, calculate each finger printing and the similarity contrasting collection of illustrative plates, the results are shown in following table 12:
The similarity result of the finger printing of table 12 10 batch sample
3 conclusions
10 batches of medical materials in different province are under this experiment liquid-phase condition, and totally 12 common characteristic peaks, this analytical method is reliable and stable, for the discriminating of Hemerocallis fulva L. flower pesticide material and quality evaluation and control provide foundation.
Accompanying drawing explanation
Hemerocallis fulva L. contrast collection of illustrative plates (No. 3 is rutin) that Fig. 1 system generates;
The finger printing of Fig. 2 different provinces Hemerocallis fulva L. medicinal substances extract sample.
Detailed description of the invention
Embodiment 1:
To add 8 volumes, 70% ethanol doubly in Hemerocallis fulva L., 20 DEG C of lixiviates 24 hours, filter, filtrate is for subsequent use, and filtering residue adds 8 volumes, 70% ethanol doubly again, and merceration 24 hours filters, merging filtrate; Filtrate crosses AB-8 macroporous resin column, and it is transparent that distilled water is washed till effluent, with 60% ethanol elution, 3 times of column volumes, reclaims ethanol, obtains Hemerocallis fulva L. flower extract.
Embodiment 2:
To add 8 volumes, 70% ethanol doubly in Hemerocallis fulva L., 10 DEG C of lixiviates 22 hours, filter, filtrate is for subsequent use, and filtering residue adds 8 volumes, 70% ethanol doubly again, and merceration 24 hours filters, merging filtrate; Filtrate crosses AB-8 macroporous resin column, and it is transparent that distilled water is washed till effluent, with 60% ethanol elution, 4 times of column volumes, reclaims ethanol, obtains Hemerocallis fulva L. flower extract.
Embodiment 3: prepared by tablet
To add 8 volumes, 70% ethanol doubly in Hemerocallis fulva L., 15 DEG C of lixiviates 24 hours, filter, filtrate is for subsequent use, and filtering residue adds 8 volumes, 70% ethanol doubly again, and merceration 24 hours filters, merging filtrate; Filtrate crosses AB-8 macroporous resin column, and it is transparent that distilled water is washed till effluent, with 60% ethanol elution, 3 times of column volumes, reclaims ethanol, obtains Hemerocallis fulva L. flower extract; Hemerocallis fulva L. flower extract adds starch, lactose, dextrin are made after granule through common process, adds lubricant routinely, is pressed into tablet.
Embodiment 4: the preparation of Hemerocallis fulva L. flower extract capsule
To add 8 volumes, 70% ethanol doubly in Hemerocallis fulva L., 53 DEG C of lixiviates 24 hours, filter, filtrate is for subsequent use, and filtering residue adds 8 volumes, 70% ethanol doubly again, and merceration 24 hours filters, merging filtrate; Filtrate crosses AB-8 macroporous resin column, and it is transparent that distilled water is washed till effluent, with 60% ethanol elution, 3 times of column volumes, reclaims ethanol, obtains Hemerocallis fulva L. flower extract; Hemerocallis fulva L. flower extract adds lactose (or starch or dextrin) by 1: 1-5, mix homogeneously, dry, makes granule or directly cannedly in capsule, namely obtains capsule.
Embodiment 5: embodiment 1 Hemerocallis fulva L. flower extract HPLC finger printing
Chromatographic condition: chromatographic column is Agilent (specification C18,4.6*250mm, 5 μm) mobile phase methanol one water (0.5% glacial acetic acid) gradient elution, condition is shown in surface low speed 1mL/min-1, determined wavelength 254nm, column temperature 25 DEG C.
Condition of gradient elution
The preparation of reference substance solution: it is appropriate that precision takes control substance of Rutin, adds methanol and makes the solution of every 1mL containing 0.2176mg, shake up, to obtain final product.
The preparation of need testing solution: precision takes sample powder 2g, puts in 25mL volumetric flask, adds methanol to scale, supersound process 30 minutes, and methanol supplies the weight of loss, leaves standstill and lets cool, get supernatant, and 0.22 μm of microporous filter membrane filters, and gets filtrate, to obtain final product.
Embodiment 6:
To add 8 volumes, 70% ethanol doubly after Hemerocallis fulva L. coarse crushing, 20 DEG C are soaked 24 hours, with the speed percolation of 1-3ml per minute at 20 DEG C, collect just filtrate about 20 times amount medical material volume; Percolate crosses AB-8 macroporous resin column, and it is transparent that distilled water is washed till effluent, with 60% ethanol elution, 3 times of column volumes, reclaims ethanol, obtains Hemerocallis fulva L. flower extract.Adopt the ultraviolet detection method of experimental example 1, the content 30% of total flavones, rutin content 20%.
Embodiment 7:
To add 9 volumes, 70% ethanol doubly after Hemerocallis fulva L. coarse crushing, 10 DEG C are soaked 28 hours, with the speed percolation of 1-3ml per minute at 20 DEG C, collect just filtrate about 20 times amount medical material volume; Percolate crosses AB-8 macroporous resin column, and it is transparent that distilled water is washed till effluent, with 60% ethanol elution, 3 times of column volumes, reclaims ethanol, obtains Hemerocallis fulva L. flower extract.Adopt the ultraviolet detection method of experimental example 1, the content 29% of total flavones, rutin content 20%.
Embodiment 8:
To add 9 volumes, 70% ethanol doubly after Hemerocallis fulva L. coarse crushing, 5 DEG C are soaked 28 hours, with the speed percolation of 1-3ml per minute at 10 DEG C, collect just filtrate about 20 times amount medical material volume; Percolate crosses AB-8 macroporous resin column, and it is transparent that distilled water is washed till effluent, with 60% ethanol elution, 3 times of column volumes, reclaims ethanol, obtains Hemerocallis fulva L. flower extract.Adopt the ultraviolet detection method of experimental example 1, the content 30% of total flavones, rutin content 21%.
Claims (2)
1. the preparation method of Hemerocallis fulva L. flower extract, is characterized in that the method comprises the steps:
Step 1: Hemerocallis fulva L. 70% ethanol lixiviate or the speed seepage pressure effects with 1-3ml per minute below 30 DEG C, obtain ethanol extract;
Step 2: ethanol extract crosses AB-8 macroporous resin column, and it is transparent that distilled water is washed till effluent, with 55%-65% ethanol elution 2-6 times of column volume, obtains ethanol elution;
Step 3: ethanol elution reclaims ethanol and obtains Hemerocallis fulva L. flower extract.
2. preparation method as claimed in claim 1,4: Hemerocallis fulva L. flower extract HPLC finger printing detects to it is characterized in that the method comprising the steps of;
Wherein, Hemerocallis fulva L. flower extract HPLC fingerprint atlas detection method comprises the steps:
Chromatographic condition: C18 chromatographic column, the aqueous solution gradient of mobile phase methanol-0.5% glacial acetic acid is washed, 0.5-1.5mL/min
-1, determined wavelength 200-300nm, column temperature 20-30 DEG C; The condition of the aqueous solution gradient elution of described methanol-0.5% glacial acetic acid is: 0-20min: methanol be 20%, 0.5% glacial acetic acid aqueous solution be 80%; 20-30min: methanol be 30%, 0.5% glacial acetic acid aqueous solution be 70%; 30-70min: methanol be 30%, 0.5% glacial acetic acid aqueous solution be 70%; 70min is to last: methanol be 80%, 0.5% glacial acetic acid aqueous solution be 20%;
The preparation of reference substance solution: take rutin, adds Methanol and becomes reference substance solution;
The preparation of need testing solution: take Hemerocallis fulva L. flower extract powder, add methanol supersound extraction, leaves standstill, and gets supernatant and filters, get filtrate, to obtain final product;
Detection method: draw reference substance, need testing solution, sample detection.
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| CN104435648B (en) * | 2014-07-14 | 2018-07-31 | 张士舜 | A kind of composition of food and medicament dual-purpose and application thereof and preparation method |
| CN105853299A (en) * | 2016-05-24 | 2016-08-17 | 广州丹奇日用化工厂有限公司 | Method of extracting daylily extract from dry daylily and application thereof |
| CN110585324A (en) * | 2019-09-24 | 2019-12-20 | 上海应用技术大学 | Extraction and purification method of hemerocallis fulva flower polyphenol |
| CN112705728B (en) * | 2021-01-18 | 2023-05-23 | 山西大同大学 | Green preparation method of silver nanorods |
| CN114081922A (en) * | 2021-11-22 | 2022-02-25 | 兰州大学 | Hemerocallis plant leaf total phenol extract and preparation method and application thereof |
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