CN104730166B - 3,3 '-dimethylellagic acid, the quantitative detecting method of 3,3 ', 4 '-trimethyl ellagic acid in a kind of Herba Polygoni Runcinati - Google Patents
3,3 '-dimethylellagic acid, the quantitative detecting method of 3,3 ', 4 '-trimethyl ellagic acid in a kind of Herba Polygoni Runcinati Download PDFInfo
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- AFSDNFLWKVMVRB-UHFFFAOYSA-N Ellagic acid Chemical compound OC1=C(O)C(OC2=O)=C3C4=C2C=C(O)C(O)=C4OC(=O)C3=C1 AFSDNFLWKVMVRB-UHFFFAOYSA-N 0.000 title claims abstract description 65
- 239000002253 acid Substances 0.000 title claims abstract description 63
- ATJXMQHAMYVHRX-CPCISQLKSA-N Ellagic acid Natural products OC1=C(O)[C@H]2OC(=O)c3cc(O)c(O)c4OC(=O)C(=C1)[C@H]2c34 ATJXMQHAMYVHRX-CPCISQLKSA-N 0.000 title claims abstract description 62
- 229920002079 Ellagic acid Polymers 0.000 title claims abstract description 62
- 229960002852 ellagic acid Drugs 0.000 title claims abstract description 62
- 235000004132 ellagic acid Nutrition 0.000 title claims abstract description 62
- FAARLWTXUUQFSN-UHFFFAOYSA-N methylellagic acid Natural products O1C(=O)C2=CC(O)=C(O)C3=C2C2=C1C(OC)=C(O)C=C2C(=O)O3 FAARLWTXUUQFSN-UHFFFAOYSA-N 0.000 title claims abstract description 62
- 238000000034 method Methods 0.000 title claims abstract description 41
- 238000001514 detection method Methods 0.000 claims abstract description 23
- 239000000463 material Substances 0.000 claims abstract description 3
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 75
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 claims description 48
- 239000000523 sample Substances 0.000 claims description 41
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims description 33
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 claims description 32
- 239000013558 reference substance Substances 0.000 claims description 18
- 229910000147 aluminium phosphate Inorganic materials 0.000 claims description 16
- 239000012567 medical material Substances 0.000 claims description 16
- 239000012488 sample solution Substances 0.000 claims description 16
- 239000000243 solution Substances 0.000 claims description 16
- 239000007788 liquid Substances 0.000 claims description 14
- 239000000047 product Substances 0.000 claims description 10
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 9
- 238000002156 mixing Methods 0.000 claims description 9
- 238000002360 preparation method Methods 0.000 claims description 9
- 239000011550 stock solution Substances 0.000 claims description 9
- 239000002904 solvent Substances 0.000 claims description 7
- UGAPHEBNTGUMBB-UHFFFAOYSA-N acetic acid;ethyl acetate Chemical compound CC(O)=O.CCOC(C)=O UGAPHEBNTGUMBB-UHFFFAOYSA-N 0.000 claims description 5
- 239000002024 ethyl acetate extract Substances 0.000 claims description 5
- 239000000284 extract Substances 0.000 claims description 5
- 239000012528 membrane Substances 0.000 claims description 5
- 230000008569 process Effects 0.000 claims description 5
- 238000010992 reflux Methods 0.000 claims description 5
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 5
- 238000010790 dilution Methods 0.000 claims description 3
- 239000012895 dilution Substances 0.000 claims description 3
- 238000012417 linear regression Methods 0.000 claims description 3
- 239000012467 final product Substances 0.000 claims description 2
- OAICVXFJPJFONN-UHFFFAOYSA-N Phosphorus Chemical compound [P] OAICVXFJPJFONN-UHFFFAOYSA-N 0.000 claims 1
- 238000012856 packing Methods 0.000 claims 1
- 229910052698 phosphorus Inorganic materials 0.000 claims 1
- 239000011574 phosphorus Substances 0.000 claims 1
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- Investigating Or Analysing Biological Materials (AREA)
- Heterocyclic Carbon Compounds Containing A Hetero Ring Having Oxygen Or Sulfur (AREA)
Abstract
The invention discloses in a kind of Herba Polygoni Runcinati 3,3' dimethylellagic acid, 3,3', the quantitative detecting method of 4' trimethyl ellagic acid, this quantitative detecting method uses high performance liquid chromatography to record with special color spectral condition and calculates in Herba Polygoni Runcinati 3 by linear equation after peak area, 3' dimethylellagic acid, 3,3', the content of 4' trimethyl ellagic acid.Having can be with 3, and 3' dimethylellagic acid, 3,3', the detection by quantitative of 4' trimethyl ellagic acid effectively controls Herba Polygoni Runcinati quality of medicinal material, the feature that method easily operates, result is stable, precision is high.
Description
Technical field
The invention belongs to medicine quantitative measurement technology field, relate in particular in a kind of Herba Polygoni Runcinati 3,3'-dimethyl tan
Flower acid, 3,3', the quantitative detecting method of 4'-trimethyl ellagic acid.
Background technology
3,3'-dimethylellagic acids and 3,3', 4'-trimethyl ellagic acid belongs to the derivant of ellagic acid, is widely present in
Plant kingdom, has stronger anti peroxidation of lipid and removes the ability of free radical, various bacteria and virus are had inhibitory action, energy
The canceration that the suppression position such as liver, kidney induces due to chemical induction, it also has coagulation function.
Herba Polygoni Runcinati derives from Polygonaceae arsesmart Herba Polygoni Runcinati (Polygonum runcinatum Buch.-Ham.var.
Sinene Hemsl.) dry rhizome, another name soil is done one's utmost, Polygonum runcinatum Buch.Ham, Polygonum runcinatum var. sinense Hemsl., snakehead knotweed etc..Early summer or autumn excavate, property
Cold, bitter in the mouth, acid puckery, micro-.There is relaxing muscles and tendons to promote blood circulation detumescence, the effect of heat-clearing and toxic substances removing, and there is good antioxidant activity and to G-
The obvious bacteriostasis of dysentery bacterium.Cure mainly dysentery, have loose bowels, spit blood spitting of blood, bleeding hemorrhoids, leucorrhea with red and white discharge, dysmenorrhea, poison
Venom, impairment caused by overstrain lumbago and traumatic injury etc., mastitis, carbuncle furuncle and phyma poison etc. are controlled in external.
Finding in the chemical constitution study of Herba Polygoni Runcinati, the content of its ellagic acid compounds is higher.Therefore, in Herba Polygoni Runcinati
The content of ellagic acid analog derivative, is the important indicator weighing its drug effect.
Herba Polygoni Runcinati now records in " Guizhou Province's Chinese crude drug, the Ethnic crude drugs quality standard " of version in 2003, the openest literary composition
Offering and only relate to Herba Polygoni Runcinati character and discriminating, for the assay of its active ingredient, at present, only spectrophotography carries out red shin
The document report of determination of total flavonoids in dissipating, the method, according to measuring absorbance after colour developing, broadly calculates substance that show color
Content is as at the bottom of the content of total flavones, specificity and accuracy, and measurement result deviation is relatively big, is unfavorable for accurately measuring.
There is because of ellagic acid compounds the multiple biological activitys such as antioxidation, antiinflammatory, blood coagulation, anti-cancer, be the medicine of Herba Polygoni Runcinati
Effective substance basis one of, with it as quantitative target, more scientific rationally, the quality of Herba Polygoni Runcinati medical material can be made more effectively to be controlled
System.
Summary of the invention
The one that it is an object of the invention to overcome aforementioned drawback and provide can effectively control Herba Polygoni Runcinati quality of medicinal material, method
3,3'-dimethylellagic acid, 3,3' in easily operation, the Herba Polygoni Runcinati that result is stable, precision is high, 4'-trimethyl ellagic acid quantitative
Detection method.
3,3'-dimethylellagic acid, 3,3' in a kind of Herba Polygoni Runcinati of the present invention, the detection by quantitative of 4'-trimethyl ellagic acid
Method, comprises the following steps:
(1) sample solution preparation: take Herba Polygoni Runcinati medical material, pulverizes, and crosses 60 mesh sieves, weighs 1.0g, accurately weighed, is placed in container
In, add 100ml alcohol reflux 1h, reclaim the solvent in extracting solution and be concentrated into extractum shape, adding 20ml water, supersound process
20min, extracts 5 times with ethyl acetate shaking, and the consumption of ethyl acetate is each 20ml, is evaporated acetic acid ethyl acetate extract, residue
Adding methanol dissolve and be settled in 10ml measuring bottle, shake up, 0.45 μm microporous filter membrane filters, and prepares sample solution；
(2) sample size measures:
Chromatographic condition:
C18 chromatographic column；
Flowing phase: volume ratio is the methanol of 7-27: 20-50: 23-73: acetonitrile: 0.1% phosphoric acid；
Detection wavelength: 230-270nm；
Column temperature: 25-40 DEG C；
Volume flow: 0.8-1.2ml min-1；
Sample size: 10-20 μ l；
Detecting sample solution with high performance liquid chromatograph, the peak area according to recording calculates sample by linear equation
3,3'-dimethylellagic acid and 3,3' in product, the content of 4'-trimethyl ellagic acid:
The linear equation of 3,3'-dimethylellagic acid is: Y1=5598X1+ 180.8, r=0.9991；
The linear equation of 3,3', 4'-trimethyl ellagic acid is: Y2=5517.4X2+ 73.3, r=0.9990；
Wherein, Y1Representing 3,3'-dimethylellagic acid utilizes the peak area that high performance liquid chromatograph detects, Y2Represent 3,
3', 4'-trimethyl ellagic acid utilizes the peak area that high performance liquid chromatograph detects, X1Represent 3,3'-dimethylellagic acid
Sample introduction quality, X2Representing 3,3', the sample introduction quality of 4'-trimethyl ellagic acid, r is correlation coefficient.
3,3'-dimethylellagic acid, 3,3' in a kind of Herba Polygoni Runcinati of the present invention, the detection by quantitative of 4'-trimethyl ellagic acid
Method, wherein: described 3,3'-dimethylellagic acid, 3,3', the linear equation of 4'-trimethyl ellagic acid is by the following method
Arrive:
Take 3,3'-dimethylellagic acid reference substance 19.8mg and 3,3', 4'-trimethyl ellagic acid reference substance 9.82mg, add
Enter methanol to dissolve and constant volume, make every 1ml and contain 3,3'-dimethylellagic acid 0.0990mg and 3,3', 4'-trimethyl ellagic acid
The mixing reference substance stock solution of 0.0491mg；
Precision measure mixing reference substance stock solution 2,4,6,8mL, put in 10mL measuring bottle, add methanol dilution constant volume, preparation
Become gradient solution, make control series product solution together with former mixing reference substance stock solution；
Employing chromatographic condition: C18 chromatographic column, flowing phase: volume ratio is the methanol of 7-27: 20-50: 23-73: acetonitrile:
0.1% phosphoric acid, detection wavelength: 230-270nm, column temperature: 25-40 DEG C, volume flow: 0.8-1.2ml min-1, sample size: 10-
20 μ l, utilize high performance liquid chromatograph to detect, with the sample introduction quality of reference substance as abscissa, peak area as vertical coordinate, will
Record sample introduction quality and the peak area of above-mentioned variable concentrations titer, by linear regression and get final product.
3,3'-dimethylellagic acid, 3,3' in a kind of Herba Polygoni Runcinati of the present invention, the detection by quantitative of 4'-trimethyl ellagic acid
Method, wherein: described C18 chromatographic column is Dikma ODS C18 post, and the length of chromatographic column is 250mm, and diameter is 4.6mm, filler
Particle diameter is 5 μm.
3,3'-dimethylellagic acid, 3,3' in a kind of Herba Polygoni Runcinati of the present invention, the detection by quantitative of 4'-trimethyl ellagic acid
Method, wherein: described flowing is the methanol of 17:33:50 for volume ratio mutually: acetonitrile: 0.1% phosphoric acid.
3,3'-dimethylellagic acid, 3,3' in a kind of Herba Polygoni Runcinati of the present invention, the detection by quantitative of 4'-trimethyl ellagic acid
Method, wherein: methanol and acetonitrile in described flowing mutually are Chromatographic Pure Methanol and acetonitrile, and described phosphoric acid is analytical pure phosphoric acid.
3,3'-dimethylellagic acid, 3,3' in a kind of Herba Polygoni Runcinati of the present invention, the detection by quantitative of 4'-trimethyl ellagic acid
Method, wherein: described detection wavelength is 248nm.
3,3'-dimethylellagic acid, 3,3' in a kind of Herba Polygoni Runcinati of the present invention, the detection by quantitative of 4'-trimethyl ellagic acid
Method, wherein: described column temperature is 30 DEG C.
3,3'-dimethylellagic acid, 3,3' in a kind of Herba Polygoni Runcinati of the present invention, the detection by quantitative of 4'-trimethyl ellagic acid
Method, wherein: described volume flow is 1.0ml min-1。
3,3'-dimethylellagic acid, 3,3' in a kind of Herba Polygoni Runcinati of the present invention, the detection by quantitative of 4'-trimethyl ellagic acid
Method, wherein: described sample size is 10 μ l.
The present invention compared with prior art, has obvious beneficial effect, as can be known from the above technical solutions: provided by the present invention
Herba Polygoni Runcinati in 3,3'-dimethylellagic acid, 3,3', the quantitative detecting method of 4'-trimethyl ellagic acid uses high-efficient liquid phase color
Spectrometry, and through great many of experiments, chromatographic condition parameter, solvent etc. are made that corresponding restriction.With methanol-acetonitrile-0.1% phosphoric acid
Mutually can be well by 3 as flowing, 3'-dimethylellagic acid and 3,3', 4'-trimethyl ellagic acid separates, and selects ethanol to make
For sample extraction solvent, extraction efficiency is high and avirulence, in Sample Preparation Procedure, adopts and is extracted with ethyl acetate, it is to avoid impurity
Experimental result is interfered, improves the precision of detection.This law is used to measure the 3,3'-dimethylellagic acid in red shin medical material
With 3,3', 4'-trimethyl ellagic acid content can effectively control Herba Polygoni Runcinati medical material matter as the method for quality control of Herba Polygoni Runcinati
Amount.
Detailed description of the invention
Below in conjunction with detailed description of the invention, beneficial effects of the present invention is described in further detail.
Dimethylellagic acid, 3,3', the linear equation of 4'-trimethyl ellagic acid obtains by the following method:
Take 3,3'-dimethylellagic acid reference substance 19.8mg and 3,3', 4'-trimethyl ellagic acid reference substance 9.82mg, add
Enter methanol to dissolve and constant volume, make every 1ml and contain 3,3'-dimethylellagic acid 0.0990mg and 3,3', 4'-trimethyl ellagic acid
The mixing reference substance stock solution of 0.0491mg;
Precision measure mixing reference substance stock solution 2,4,6,8mL, put in 10mL measuring bottle, add methanol dilution constant volume, preparation
Become gradient solution, make control series product solution together with former mixing reference substance stock solution；
Employing chromatographic condition: C18 chromatographic column, flowing phase: volume ratio is the methanol of 7-27: 20-50: 23-73: acetonitrile:
0.1% phosphoric acid, detection wavelength: 230-270nm, column temperature: 25-40 DEG C, volume flow: 0.8-1.2ml min-1, sample size: 10-
20 μ l, utilize high performance liquid chromatograph to detect, 3,3'-dimethylellagic acids and 3,3', and 4'-trimethyl ellagic acid exists respectively
There is good linear relationship with peak area in 0.198-0.990 μ g and 0.098-0.491 μ g range, above-mentioned variable concentrations will be recorded
The sample introduction quality of titer and peak area, obtained by linear regression:
The linear equation of 3,3'-dimethylellagic acid is: Y1=5598X1+ 180.8, r=0.9991；
The linear equation of 3,3', 4'-trimethyl ellagic acid is: Y2=5517.4X2+ 73.3, r=0.9990；
Wherein, Y1Representing 3,3'-dimethylellagic acid utilizes the peak area that high performance liquid chromatograph detects, Y2Represent 3,
3', 4'-trimethyl ellagic acid utilizes the peak area that high performance liquid chromatograph detects, X1Represent 3,3'-dimethylellagic acid
Sample introduction quality, X2Representing 3,3', the sample introduction quality of 4'-trimethyl ellagic acid, r is correlation coefficient.
Embodiment 1-10
In a kind of Herba Polygoni Runcinati medical material 3,3'-dimethylellagic acid, 3,3', the quantitative detecting method of 4'-trimethyl ellagic acid,
Wherein, 10 batches that Herba Polygoni Runcinati medical material is gathered respectively autumn, comprise the following steps:
(1) sample solution preparation: take Herba Polygoni Runcinati medical material, pulverizes, and crosses 60 mesh sieves, weighs 1.0g, accurately weighed, is placed in container
In, add 100ml alcohol reflux 1h, reclaim the solvent in extracting solution and be concentrated into extractum shape, adding 20ml water, supersound process
20min, extracts 5 times with ethyl acetate shaking, and the consumption of ethyl acetate is each 20ml, is evaporated acetic acid ethyl acetate extract, residue
Adding methanol dissolve and be settled in 10ml measuring bottle, shake up, 0.45 μm microporous filter membrane filters, and prepares sample solution；
(2) sample size measures:
Chromatographic condition:
C18 chromatographic column, enlightening horse Dikma ODS C18 post, the length of chromatographic column is 250mm, and diameter is 4.6mm, filler grain
Footpath is 5 μm；
Flowing phase: volume ratio is the methanol of 17:33:50: acetonitrile: 0.1% phosphoric acid, and wherein, methanol and acetonitrile are chromatographically pure first
Alcohol and acetonitrile, phosphoric acid is analytical pure phosphoric acid；
Detection wavelength: 248nm；
Column temperature: 30 DEG C；
Volume flow: 1.0ml min-1；
Sample size: 10 μ l；
Detecting sample solution with high performance liquid chromatograph, the peak area according to recording calculates sample by linear equation
3,3'-dimethylellagic acid and 3,3' in product, the content of 4'-trimethyl ellagic acid.
The assay of 10 batch sample the results are shown in Table 1.
Embodiment 11-12
In Herba Polygoni Runcinati medical material 3,3'-dimethylellagic acid, 3,3', the quantitative detecting method of 4'-trimethyl ellagic acid, its
In, 2 batches that Herba Polygoni Runcinati medical material is gathered respectively summer, comprise the following steps:
(1) sample solution preparation: take Herba Polygoni Runcinati medical material, pulverizes, and crosses 60 mesh sieves, weighs 1.0g, accurately weighed, is placed in container
In, add 100ml alcohol reflux 1h, reclaim the solvent in extracting solution and be concentrated into extractum shape, adding 20ml water, supersound process
20min, extracts 5 times with ethyl acetate shaking, and the consumption of ethyl acetate is each 20ml, is evaporated acetic acid ethyl acetate extract, residue
Adding methanol dissolve and be settled in 10ml measuring bottle, shake up, 0.45 μm microporous filter membrane filters, and prepares sample solution；
(2) sample size measures:
Chromatographic condition:
C18 chromatographic column, enlightening horse Dikma ODS C18 post, the length of chromatographic column is 250mm, and diameter is 4.6mm, filler grain
Footpath is 5 μm；
Flowing phase: volume ratio is the methanol of 15:40:45: acetonitrile: 0.1% phosphoric acid；
Detection wavelength: 230nm；
Column temperature: 25 DEG C；
Volume flow: 0.8ml min-1；
Sample size: 15 μ l；
Detecting sample solution with high performance liquid chromatograph, the peak area according to recording calculates sample by linear equation
3,3'-dimethylellagic acid and 3,3' in product, the content of 4'-trimethyl ellagic acid.
The assay of 2 batch sample the results are shown in Table 2.
Embodiment 13-14
In Herba Polygoni Runcinati medical material 3,3'-dimethylellagic acid, 3,3', the quantitative detecting method of 4'-trimethyl ellagic acid, its
In, 2 batches that Herba Polygoni Runcinati medical material is gathered respectively spring, comprise the following steps:
(1) sample solution preparation: take Herba Polygoni Runcinati medical material, pulverizes, and crosses 60 mesh sieves, weighs 1.0g, accurately weighed, is placed in container
In, add 100ml alcohol reflux 1h, reclaim the solvent in extracting solution and be concentrated into extractum shape, adding 20ml water, supersound process
20min, extracts 5 times with ethyl acetate shaking, and the consumption of ethyl acetate is each 20ml, is evaporated acetic acid ethyl acetate extract, residue
Adding methanol dissolve and be settled in 10ml measuring bottle, shake up, 0.45 μm microporous filter membrane filters, and prepares sample solution；
(2) sample size measures:
Chromatographic condition:
C18 chromatographic column, enlightening horse Dikma ODS C18 post, the length of chromatographic column is 250mm, and diameter is 4.6mm, filler grain
Footpath is 5 μm；
Flowing phase: volume ratio is the methanol of 20: 30: 50: acetonitrile: 0.1% phosphoric acid；
Detection wavelength: 270nm；
Column temperature: 40 DEG C；
Volume flow: 1.2ml min-1；
Sample size: 20 μ l；
Detecting sample solution with high performance liquid chromatograph, the peak area according to recording calculates sample by linear equation
3,3'-dimethylellagic acid and 3,3' in product, the content of 4'-trimethyl ellagic acid.
The assay of 2 batch sample the results are shown in Table 3.
According to the assay result of above 14 batch samples, the autumn in this product suitable summer plucks, and considers the fluctuation of medical material content, really
Determining this product 3, the content of 3'-dimethylellagic acid must not be less than 0.5mg/g, and the content of 3,3', 4'-trimethyl ellagic acid must not lack
In 0.17mg/g(in terms of dry product).
It is measured below by indexs such as testing precision, stability and ruggedness to the present invention.
1. precision test: precision measures the 10 same standard solution of μ L, continuous 6 sample introductions, measures by above-mentioned chromatographic condition,
Record chromatographic peak peak area.3,3'-dimethylellagic acid and 3,3', the RSD of 4'-6 sample introduction peak areas of trimethyl ellagic acid divides
It is not 0.61% and 1.64%, shows that instrument precision is good, be shown in Table 4.
4. average recovery test: taking 6 parts of the Herba Polygoni Runcinati sample of known content, every part of 0.5g, accurately weighed, precision adds
A certain amount of 3,3'-dimethylellagic acid and 3,3', 4'-trimethyl ellagic acid reference substance, by need testing solution preparation method system
Available test sample solution, is measured by above-mentioned chromatographic condition, calculates the response rate, and measurement result is shown in Table 7 and table 8.
5. the impact of wavelength change: the accurate absorption 10 same need testing solutions of μ L, under the conditions of 246nm, 248nm, 250nm
Sample introduction twice, measures according to above-mentioned chromatographic condition respectively, records chromatographic peak peak area.3,3'-dimethylellagic acid and 3,3',
The RSD of 4'-6 sample introduction peak areas of trimethyl ellagic acid is respectively 0.44% and 0.51%.Show that method has less change at wavelength
During change, measurement result has good stability, and is shown in Table 10.
6. the impact of different brands chromatographic column: select Shimadzu, Di Ma and Yi Li top grade different brands C18 chromatographic column to red
Shin dissipates the separation situation of medical material and compares, and finds that three kinds of chromatographic columns all can preferably separate 3,3'-dimethylellagic acid and 3,
3', 4'-trimethyl ellagic acid.
Knowable to the result of above test, utilize the detection method of the present invention program, precision, stability and ruggedness etc.
Index is preferable.
The above, be only presently preferred embodiments of the present invention, and the present invention not makees any pro forma restriction, appoints
What is without departing from technical solution of the present invention content, any is simply repaiied according to what above example made by the technical spirit of the present invention
Change, equivalent variations and modification, all still fall within the range of technical solution of the present invention.
Claims (9)
1. in a Herba Polygoni Runcinati 3,3'-dimethylellagic acid, 3,3', the quantitative detecting method of 4'-trimethyl ellagic acid, including with
Lower step:
(1) sample solution preparation: take Herba Polygoni Runcinati medical material, pulverizes, and crosses 60 mesh sieves, weighs 1.0g, accurately weighed, is placed in container,
Add 100ml alcohol reflux 1h, reclaim the solvent in extracting solution and be concentrated into extractum shape, adding 20ml water, supersound process
20min, extracts 5 times with ethyl acetate shaking, and the consumption of ethyl acetate is each 20ml, is evaporated acetic acid ethyl acetate extract, residue
Adding methanol dissolve and be settled in 10ml measuring bottle, shake up, 0.45 μm microporous filter membrane filters, and prepares sample solution；
(2) sample size measures:
Chromatographic condition:
C18 chromatographic column；
Flowing phase: volume ratio is the methanol of 7-27: 20-50: 23-73: acetonitrile: 0.1% phosphoric acid；
Detection wavelength: 230-270nm；
Column temperature: 25-40 DEG C；
Volume flow: 0.8-1.2ml min-1；
Sample size: 10-20 μ l；
With high performance liquid chromatograph, sample solution is detected, calculated in sample by linear equation according to the peak area recorded
3,3'-dimethylellagic acid and 3,3', the content of 4'-trimethyl ellagic acid:
The linear equation of 3,3'-dimethylellagic acid is: Y1=5598X1+ 180.8, r=0.9991；
The linear equation of 3,3', 4'-trimethyl ellagic acid is: Y2=5517.4X2+ 73.3, r=0.9990；
Wherein, Y1Representing 3,3'-dimethylellagic acid utilizes the peak area that high performance liquid chromatograph detects, Y2Represent 3,3',
4'-trimethyl ellagic acid utilizes the peak area that high performance liquid chromatograph detects, X1Represent the sample introduction of 3,3'-dimethylellagic acid
Quality, X2Representing 3,3', the sample introduction quality of 4'-trimethyl ellagic acid, r is correlation coefficient.
2. 3,3'-dimethylellagic acid, 3,3', determining of 4'-trimethyl ellagic acid in a kind of Herba Polygoni Runcinati as claimed in claim 1
Quantity measuring method, wherein: described 3,3'-dimethylellagic acid, 3,3', the linear equation of 4'-trimethyl ellagic acid is by following
Method obtains:
Take 3,3'-dimethylellagic acid reference substance 19.8mg and 3,3', 4'-trimethyl ellagic acid reference substance 9.82mg, add first
Alcohol dissolves and constant volume, makes every 1ml and contains 3,3'-dimethylellagic acid 0.0990mg and 3,3', 4'-trimethyl ellagic acid
The mixing reference substance stock solution of 0.0491mg；
Precision measure mixing reference substance stock solution 2,4,6,8mL, put in 10mL measuring bottle, add methanol dilution constant volume, be configured to ladder
Degree solution, makes control series product solution together with former mixing reference substance stock solution；
Employing chromatographic condition: C18 chromatographic column, flowing phase: volume ratio is the methanol of 7-27: 20-50: 23-73: acetonitrile: 0.1% phosphorus
Acid, detection wavelength: 230-270nm, column temperature: 25-40 DEG C, volume flow: 0.8-1.2ml min-1, sample size: 10-20 μ l, profit
Detect with high performance liquid chromatograph, with the sample introduction quality of reference substance as abscissa, peak area as vertical coordinate, will record above-mentioned
The sample introduction quality of variable concentrations titer and peak area, by linear regression and get final product.
3. 3,3'-dimethylellagic acid, 3,3', 4'-trimethyl ellagic acid in a kind of Herba Polygoni Runcinati as claimed in claim 1 or 2
Quantitative detecting method, wherein: described C18 chromatographic column is Dikma ODS C18 post, and the length of chromatographic column is 250mm, and diameter is
4.6mm, packing material size is 5 μm.
4. 3,3'-dimethylellagic acid, 3,3', 4'-trimethyl ellagic acid in a kind of Herba Polygoni Runcinati as claimed in claim 1 or 2
Quantitative detecting method, wherein: described flowing is the methanol of 17:33:50 for volume ratio mutually: acetonitrile: 0.1% phosphoric acid.
5. 3,3'-dimethylellagic acid, 3,3', 4'-trimethyl ellagic acid in a kind of Herba Polygoni Runcinati as claimed in claim 1 or 2
Quantitative detecting method, wherein: methanol and acetonitrile in described flowing mutually are Chromatographic Pure Methanol and acetonitrile, described phosphoric acid for point
Analysis pure phosphoric acid.
6. 3,3'-dimethylellagic acid, 3,3', 4'-trimethyl ellagic acid in a kind of Herba Polygoni Runcinati as claimed in claim 1 or 2
Quantitative detecting method, wherein: described detection wavelength is 248nm.
7. 3,3'-dimethylellagic acid, 3,3', 4'-trimethyl ellagic acid in a kind of Herba Polygoni Runcinati as claimed in claim 1 or 2
Quantitative detecting method, wherein: described column temperature is 30 DEG C.
8. 3,3'-dimethylellagic acid, 3,3', 4'-trimethyl ellagic acid in a kind of Herba Polygoni Runcinati as claimed in claim 1 or 2
Quantitative detecting method, wherein: described volume flow is 1.0ml min-1。
9. 3,3'-dimethylellagic acid, 3,3', 4'-trimethyl ellagic acid in a kind of Herba Polygoni Runcinati as claimed in claim 1 or 2
Quantitative detecting method, wherein: described sample size is 10 μ l.
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| CN101788537A (en) * | 2009-09-28 | 2010-07-28 | 贵州益康制药有限公司 | Method for measuring content of ellagic acid ingredients in euscaphis japonica medicinal materials |
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