CN105938125A - Raw fleece-flower root HPLC fingerprint detection method - Google Patents

Raw fleece-flower root HPLC fingerprint detection method Download PDF

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CN105938125A
CN105938125A CN201610490084.0A CN201610490084A CN105938125A CN 105938125 A CN105938125 A CN 105938125A CN 201610490084 A CN201610490084 A CN 201610490084A CN 105938125 A CN105938125 A CN 105938125A
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radix polygoni
polygoni multiflori
detection method
glycosides
chromatographic
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CN105938125B (en
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李芸霞
龚小红
彭成
党珏
赵梦杰
赵梦君
罗林
袁岸
李燕
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Chengdu University of Traditional Chinese Medicine
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Chengdu University of Traditional Chinese Medicine
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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
    • G01N30/02—Column chromatography
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
    • G01N30/02—Column chromatography
    • G01N2030/022—Column chromatography characterised by the kind of separation mechanism
    • G01N2030/027—Liquid chromatography

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  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Immunology (AREA)
  • Pathology (AREA)
  • Investigating Or Analysing Biological Materials (AREA)
  • Medicines Containing Plant Substances (AREA)

Abstract

The invention discloses a raw fleece-flower root HPLC fingerprint detection method. The detection method comprises the following steps: (1) preparing a control substance solution; (2) preparing a reference substance solution; (3) establishing a control fingerprint; (4) preparing a test article solution; (5) detecting the test article solution. The obtained fingerprint has the advantages that there are many peaks, the resolution and repeatability are good, the precision is high, and the quality of raw fleece-flower root can be effectively monitored. By using the extraction method and chromatogram conditions of provided fingerprint detection method, contents of 16 components of raw fleece-flower root can be measured, and thus the quality of raw fleece-flower root can be monitored from more aspects.

Description

The HPLC fingerprint atlas detection method of Radix Polygoni Multiflori
Technical field
The present invention relates to Control of drug quality method and technology field, particularly relate to the HPLC finger printing detection of Radix Polygoni Multiflori Method.
Background technology
Radix Polygoni Multiflori is the dried root of polygonum multiflorum thunb (Polygonum Multiflorum Thunb.), and its property is put down, taste Sweet, bitter, GUIXIN, liver, large intestine channel, there is effect of removing toxic substances, eliminating carbuncle, preventing the attack (or recurrence) of malaria, loosening bowel to relieve constipation；For conventional Chinese medicine.Its The most widely distributed, main product in Henan, Hubei, Guangxi, Guangdong, Guizhou, the ground such as Sichuan.
Stilbene glycoside and anthraquinone component in Radix Polygoni Multiflori are its representative active component.Wherein, stilbene glucoside has Significantly neuroprotective；Anthraquinone component has antiinflammatory, resisting pathogenic microbes, antitumor, diuresis, the effect such as protects the liver, Wherein rheum emodin has antiinflammatory, resisting pathogenic microbes, antitumor, diuresis, effect, the rheum emodin-8-β-o-pyrans Portugal such as protects the liver Polyglycoside then has nootropic activity.
Use UV-spectrophotometry to Radix Polygoni Multiflori at present under " Chinese Pharmacopoeia " version Radix Polygoni Multiflori item in 2015 2,3,5,4-tetrahydroxystilbene-2-O-β-D glucosides and emodin content are determined, and establish with measurement result Quality standard.
However, it is contemplated that the composition of Chinese crude drug is various, and easily influence each other between each composition, only with one or two effective ingredient Content be difficult to the real quality of thoroughly evaluating Chinese medicine.Therefore, one is presently required and can monitor multiple compositions in Radix Polygoni Multiflori Detection method, to monitor the quality condition of Radix Polygoni Multiflori more fully hereinafter.
Summary of the invention
For solving the problems referred to above, the invention provides the HPLC fingerprint atlas detection method of a kind of Radix Polygoni Multiflori, it includes Following steps:
The HPLC fingerprint atlas detection method of Radix Polygoni Multiflori, it is characterised in that: it comprises the following steps:
(1) preparation of reference substance solution: take Radix Polygoni Multiflori medical material, pulverizes to obtain medicinal powder, 40～60% ethanol water carry Take, filter, obtain Radix Polygoni Multiflori reference substance solution；Preferably concentration of alcohol is 50%；
(2) preparation of object of reference solution: take gallic acid, procyanidin B1, catechin, epicatechol gallate, Aloe big Flavin glycosides, polygonin, stilbene glucoside, ponticin, resveratrol, rheum emodin glycosides, physcione glycosides, Aloe are big Flavin, chrysophanic acid, chrysophanol, rheum emodin, physcione reference substance are appropriate, mixing, add methanol, obtain object of reference Solution；
(3) foundation of reference fingerprint:
Take object of reference solution and the reference substance solution prepared by different batches Radix Polygoni Multiflori medical material respectively, inject efficient liquid phase Chromatograph detects, and obtains each batch medical material and the chromatographic data of object of reference；Chromatographic condition is as follows:
Detection wavelength: 254～275nm；
Chromatographic column: C18 chromatographic column and C18 pre-column；
Flowing phase: mobile phase A is 0.1% formic acid water, and Mobile phase B is acetonitrile；
Gradient elution program is as follows:
Gained chromatographic data is processed, sets up the reference fingerprint of Radix Polygoni Multiflori；
(4) preparation of need testing solution: take medical material to be measured, prepares test sample according to the method that step (1) is identical molten Liquid；
(5) according to the chromatographic condition that step (3) is identical, need testing solution is injected high performance liquid chromatograph detection, by institute Obtain chromatogram to compare with reference fingerprint, Radix Polygoni Multiflori and quality condition thereof can be identified.
Further, in step (1), described extraction supersound extraction.
Further, the time of described supersound extraction is 60min.
Further, in step (1), the temperature of described extraction is 40 DEG C.
Further, in step (1), the w/v of described medical material ethanol water is 1g:20mL.
Further, described C18 chromatographic column is ZORBAX C18 chromatographic column, a length of 250mm, and internal diameter is 4.6mm, The particle diameter of filler is 5 μm；Described C18 pre-column is ZORBAX C18 chromatographic column, a length of 12.5mm, and internal diameter is 4.6mm, the particle diameter of filler is 5 μm..
Further, the column temperature of described chromatographic condition is 30 DEG C.
Further, the flow velocity of described chromatographic condition is 1.0mL/min.
Further, described reference fingerprint is as shown in Figure 5.
Present invention also offers a kind of method of 16 kinds of compositions in Radix Polygoni Multiflori of mensuration simultaneously, described 16 kinds of compositions are Galla Turcica (Galla Helepensis) Acid, procyanidin B1, catechin, epicatechol gallate, Barbaloin, polygonin, stilbene glucoside, rheochrysin, Resveratrol, rheum emodin glycosides, physcione glycosides, aloe-emodin, chrysophanic acid, chrysophanol, rheum emodin and rheum emodin Methyl ether；
It comprises the following steps:
A, the foundation of 16 kinds of ingredient standard curves:
Take 16 kinds of composition mixing reference substance solution, according to the chromatographic condition detection that step (3) in aforementioned detection method is identical, Obtain 16 kinds of respective standard curves of composition；
B, take Radix Polygoni Multiflori medical material, according to the method that step (1) in aforementioned detection method is identical, prepare test sample molten Liquid, injects high performance liquid chromatography, with the chromatographic condition detection that step a is identical, how obtains life according to the standard curve of step a The content of 16 kinds of compositions in the Radix Polygoni Multiflori.
The fingerprint image of the present invention composes that peak is many, separating degree is good, reproducible, precision is high, for overall monitor Radix Polygoni Multiflori Material amount provide effective guarantee.
Meanwhile, utilize extracting method and the chromatographic condition of fingerprint atlas detection method of the present invention, it is also possible to record what head raw simultaneously 16 kinds of composition gallic acids, procyanidin B in crow1, catechin, epicatechol gallate, Barbaloin, polygonin, Stilbene glucoside, ponticin, resveratrol, rheum emodin glycosides, physcione glycosides, aloe-emodin, chrysophanic acid, big Yellow phenol, rheum emodin, the content of physcione, such that it is able to from the quality of medicinal material of more angle monitoring Radix Polygoni Multiflori.
Obviously, according to the foregoing of the present invention, according to ordinary technical knowledge and the customary means of this area, without departing from this Invent under above-mentioned basic fundamental thought premise, it is also possible to make the amendment of other various ways, replace or change.
The detailed description of the invention of form by the following examples, is described in further detail the foregoing of the present invention again. But this should not being interpreted as, the scope of the above-mentioned theme of the present invention is only limitted to Examples below.All real based on foregoing of the present invention institute Existing technology belongs to the scope of the present invention.
Accompanying drawing explanation
Fig. 1 is the finger printing (A-F) of Radix Polygoni Multiflori difference gradient elution program.
Fig. 2 is the finger printing (A-D) of ultrasonic 60min under Radix Polygoni Multiflori condition of different temperatures.
Fig. 3 is the finger printing (A-E) under the Radix Polygoni Multiflori difference supersound extraction time.
Fig. 4 is the standard substance collection of illustrative plates of 16 compositions of Radix Polygoni Multiflori: 1: gallic acid, 2: PB1,3: catechin, 4: epicatechol gallate, 5: Aloe rheochrysin, 6: polygonin, 7: stilbene glucoside, 8: ponticin, 9: white hellebore Alcohol, 10: rheum emodin glycosides, 11: physcione glycosides, 12: aloe-emodin, 13: chrysophanic acid, 14: rheum emodin, 15: Chrysophanol, 16: physcione.
Fig. 5 is the reference fingerprint of Radix Polygoni Multiflori
Fig. 6 is the finger printing of 25 batches of commercially available Radix Polygoni Multifloris.
Detailed description of the invention
Reagent
Gallic acid (lot number: 150226)；Procyanidin B1(lot number: 140628)；Catechin (lot number: 141224)； Epicatechol gallate (lot number: 140730)；Aloe-emodin-8-O-β-D-Glucose glycosides (it is called for short: Barbaloin, Lot number: 141225)；Polygonin (lot number: 141228)；2,3,5,4-tetrahydroxystilbene-2-O-β-D-Glucose glycosides (being called for short: stilbene glucoside, lot number: 141121)；Ponticin (lot number: 141209)；Resveratrol (lot number: 150122)；Rheum emodin-8-β-O-pyranglucoside (is called for short: rheum emodin glycosides, lot number: 141209)；Rheum emodin first Ether-8-O-β-D-Glucose glycosides (is called for short: physcione glycosides, lot number: 150120)；Chrysophanol (lot number: 140325) All purchase in Chengdu clo agate biology company limited.Aloe-emodin (lot number: MUST-10112301)；Chrysophanic acid (batch Number: MUST-11032801)；Rheum emodin (lot number: MUST-12022715)；Physcione (lot number: MUST-12022005) all purchase in Man Site bio tech ltd.
Medical material
25 batches of Radix Polygoni Multiflori medical materials are accredited as polygonaceae plant Polygonum through Chengdu University of Traditional Chinese Medicine's Pharmacognosy Studies room The dried root of multiflorum Thunb..Polygonum multiflorum medicinal material sample source is shown in Table 1.
Table 1 polygonum multiflorum medicinal material sample
Experimental apparatus
Agilent 1260 high performance liquid chromatograph device (pump G1312B, DEACB07224；Injector G1329B, DEAAC25245；Column oven G1316A, DEACN27098；UV-detector G4212BDEAA307363)； KQ-500DB ultrasonic cleaner (Kunshan Ultrasonic Instruments Co., Ltd.)；11D type electronic analytical balance (Sartorius Company)；BP-2Vortex-Genie 2 turbula shaker (Scientific Industries, the U.S.)；TG16-WS type is desk-top High speed centrifuge (Xiang Yi centrifuge company limited)；Vacuum diaphragm pump (Tianjin Jin Teng experimental facilities company limited).
Experiment reagent
Dehydrated alcohol (lot number: 20120306) purchases in Chengdu chemical industry near a river.Formic acid, acetonitrile, water are chromatographic grade, purchase In Fisher company, other reagent are analytical pure.
The screening of embodiment 1 experiment condition
1. flowing phase conditional filtering
Under Radix Polygoni Multiflori same sample sampling condition, respectively using 0.1% formic acid water and acetonitrile as the A pump of elution flow phase and B pump, carries out condition of gradient elution screening, gradient elution journey with the different volumes ratio (v/v) of A and B different time sections Sequence is shown in Table 2, and the Radix Polygoni Multiflori finger printing obtaining correspondence is shown in Fig. 1 (A-F), and as shown in Figure 1, Figure 1A fails substantially Separately；In Figure 1B, C, D, part-time point collection of illustrative plates fails separately；In the middle part of Fig. 1 E, time segment baseline drift is serious, figure Presenting the separation that each composition is good in 1F, baseline is substantially steady, therefore this condition of gradient elution is set to Radix Polygoni Multiflori Fingerprint chromatogram analysis condition.
Table 2 gradient elution time (T) change and the change of aqueous phase (A pump) ratio
2. temperature screening when Radix Polygoni Multiflori extracts
Take 4 parts of the Radix Polygoni Multiflori sample of equal in quality, add 50% ethanol water of 20 times of quality, after soaking 30min, point Not at 20 DEG C, 40 DEG C, 60 DEG C, ultrasonic 60min under the conditions of 80 DEG C, sample introduction after filtration, in the gradient elution journey screened Set up the finger printing of the Radix Polygoni Multiflori of different Extracting temperature under sequence, see Fig. 2 (A-D).Record stilbene at each temperature Glycosides, rheum emodin glycosides, physcione glycosides, with rheum emodin peak area, are changed to screening conditions with what these 4 kinds of composition front covers amassed, Obtain table 3.Under the conditions of understanding 40 DEG C in conjunction with Fig. 2 and Biao 3, the peak area of 4 kinds of compositions is the most maximum, and prompting obtains under the conditions of being somebody's turn to do The component content arrived is the highest, and therefore arranging Extracting temperature is 40 DEG C.
Under table 3 condition of different temperatures, ultrasonic 60min obtains the peak area of 4 kinds of compositions
3. Radix Polygoni Multiflori extraction time screening
Take 5 parts of the Radix Polygoni Multiflori sample of equal in quality, add 50% ethanol water of 20 times of quality, after soaking 30min, Ultrasonic 10min, 20min, 40min, 60min and 80min under the conditions of screening obtain 40 DEG C, sample introduction after filtration, Set up the finger printing of the Radix Polygoni Multiflori of different Extracting temperature under the gradient elution program screened, see Fig. 4 (A-E).Record Stilbene glucoside at each temperature, rheum emodin glycosides, physcione glycosides, with rheum emodin peak area, amass with these 4 kinds of composition front covers Be changed to screening conditions, obtain table 4.Understanding as time went in conjunction with Fig. 4 and Biao 4, the peak area of each composition increases, Peak area during 60min reaches maximum, increase the most in time and increase, point out the most available composition of ultrasonic 60min Content is the highest, and therefore arranging Extracting temperature is 40 DEG C, and ultrasonic time is 60min.
Under the conditions of 4 40 DEG C of table, ultrasonic different time obtains the peak area of 4 kinds of compositions
The method of embodiment 2 present invention and Method validation thereof
The preparation of 1 sample solution
25 batches of commercially available Radix Polygoni Multifloris are broken into coarse powder respectively standby.Precision weighs each coarse powder 0.25g, puts in tool plug triangular flask, adds 20 times amount 50% ethanol waters, after soaking 30min, supersound extraction 60min at 40 DEG C, constant volume to 5mL.0.45μm Microporous filter membrane filters, and takes subsequent filtrate sample introduction.
The preparation of 2.2 reference substance solution
Precision weighs reference substance gallic acid, procyanidin B1, catechin, epicatechol gallate, Barbaloin, tiger Cane glycosides, stilbene glucoside, ponticin, resveratrol, rheum emodin glycosides, physcione glycosides, aloe-emodin, Radix Et Rhizoma Rhei Acid, chrysophanol, rheum emodin, physcione reference substance are in 10mL volumetric flask, and methanol constant volume, to scale, is configured to dense Degree be respectively 131,98,185,142,78,125,5400,142,94,750,86,71,81,500,25, The storing solution of 88 μ g/mL, 4 DEG C of storages are standby.
2.3 chromatographic condition
Chromatographic column ZORBAX C18 (4.6mm*250mm, 5 μm), pre-column ZORBAX C18 (4.6mm*12.5mm, 5μm)；Flow velocity 1.0mL/min, column temperature 30 DEG C, sample size 5 μ L, gradient elution A are 0.1% formic acid water, and B is second Nitrile, flow phase system proportioning is shown in Table 5.
Table 5 condition of gradient elution
The foundation of 2.4 finger printing
The Chinese medicine fingerprint area of computer aided similarity evaluation software A version using Chinese Pharmacopoeia Commission to recommend carries out each The similarity analysis of concocted time sample.
3 experimental results
Fig. 4 is the liquid chromatogram of the standard substance of 16 kinds of compositions in Radix Polygoni Multiflori.
3.1 methodological study
3.1.1 precision
Preparation concentration is followed successively by the gallic acid of basic, normal, high 3 concentration, procyanidin B1, catechin, gallic acid Ester, Barbaloin, polygonin, stilbene glucoside, ponticin, resveratrol, rheum emodin glycosides, physcione Glycosides, aloe-emodin, chrysophanic acid, chrysophanol, rheum emodin, physcione quality-control sample, parallel 3 parts of each concentration, It is analyzed according to chromatographic condition under " 2.3 " item.Investigate withinday precision, day to day precision for three days on end, measure face, peak Amass and calculate the RSD% value of each composition, be shown in Table 6.
The each composition of table 6 Radix Polygoni Multiflori in a few days and day to day precision
As shown in Table 6, gallic acid, procyanidin B1, catechin, epicatechol gallate, Barbaloin, Rhizoma Polygoni Cuspidati Glycosides, stilbene glucoside, ponticin, resveratrol, rheum emodin glycosides, physcione glycosides, aloe-emodin, chrysophanic acid, The each composition of chrysophanol, rheum emodin, physcione in a few days and day to day precision RSD% is respectively less than 6.00%, method essence Density is good.
3.1.2 repeatability
Take with a collection of Radix Polygoni Multiflori medical material 6 parts, prepare according to need testing solution preparation method under " 2.1 " item, with color under " 2.3 " item Spectral condition is measured, and measures each composition gallic acid, procyanidin B in 6 parts of samples1, catechin, epicatechol gallate, Barbaloin, polygonin, stilbene glucoside, ponticin, resveratrol, rheum emodin glycosides, physcione glycosides, Aloe-emodin, chrysophanic acid, chrysophanol, rheum emodin, the peak area of physcione, standard curve calculates the dense of each composition Degree, calculates its RSD%, is shown in Table 7.
3.1.3 stability
Take with a collection of Radix Polygoni Multiflori medical material, prepare according to need testing solution preparation method under " 2.1 " item, with chromatostrip under " 2.3 " item Part is measured, and respectively 0,3,6,9,12 and 24h sample introduction, measures gallic acid, procyanidin B in sample1、 Catechin, epicatechol gallate, Barbaloin, polygonin, stilbene glucoside, ponticin, resveratrol, Radix Et Rhizoma Rhei Element glucosides, physcione glycosides, aloe-emodin, chrysophanic acid, chrysophanol, rheum emodin, the peak area of physcione, Standard curve calculates the concentration of each composition, calculates RSD%, is shown in Table 7.
The repeatability of each composition of table 7 Radix Polygoni Multiflori, study on the stability
As shown in Table 7, each composition of Radix Polygoni Multiflori, RSD% value is respectively less than 2.00%, meets 2015 editions States Pharmacopoeia specifications, show The favorable reproducibility of each composition under the conditions of this extracting method, sample is good at 12h internal stability analysis time.
3.1.4 the response rate
Taking the same a collection of Radix Polygoni Multiflori medical material of known content, be divided into 2 parts, isopyknic 50% ethanol dilution of a addition becomes former Having the 50% of concentration, another part undiluted adds the gallic acid of known and close concentration, procyanidin B1, catechin, Epicatechol gallate, Barbaloin, polygonin, stilbene glucoside, ponticin, resveratrol, rheum emodin glycosides, big Flavin methyl ether glycosides, aloe-emodin, chrysophanic acid, chrysophanol, rheum emodin, the equal-volume of physcione, HPLC measures After in sample after dilution, the concentration of each composition and equal-volume add reference substance each composition peak area, substitute into standard curve Calculate, use C%=CSample+mark-CSample/CMark* 100%, computing formula calculates response rate % of each composition, and Radix Polygoni Multiflori is each The response rate of composition is all between 90%～110%, and the response rate is higher, the results are shown in Table 8.
The response rate of each composition of table 8 Radix Polygoni Multiflori is investigated
As shown in Table 8, Radix Polygoni Multiflori each components recoveries value, all between 90.00%～120.00%, shows in this analysis method Condition has the preferable response rate.
3.1.5 prepared by Radix Polygoni Multiflori each ingredient standard curve
Gallic acid, procyanidin B1, catechin, epicatechol gallate, aloe-emodin-glycosides, polygonin, stilbene Glycosides, ponticin, resveratrol, rheum emodin glycosides, physcione glycosides, aloe-emodin, chrysophanic acid, chrysophanol, big Flavin, physcione storing solution carry out gradient dilution, obtain the standard solution of a series of concentration, by the liquid under " 2.3 " item The peak area of each composition is measured by phase condition, and the standard curve obtaining each composition is shown in Table 9.
The standard curve of each composition of table 9 Radix Polygoni Multiflori and the range of linearity
The assay of 3.2 25 batches of commercially available Radix Polygoni Multiflori compositions
25 batches of commercially available Radix Polygoni Multifloris press the preparation carrying out sample under " 2.1 " item, measure with chromatographic condition sample introduction under " 2.3 " item.Each Each sample replication 3 times, to be all worth to gallic acid in sample, procyanidin B1, catechin, epicatechol gallate, Barbaloin, polygonin, stilbene glucoside, ponticin, resveratrol, rheum emodin glycosides, physcione glycosides, Aloe-emodin, chrysophanic acid, chrysophanol, rheum emodin, the content of physcione, the results are shown in Table 10.
3.3 fingerprint similarities calculate
The finger printing of 25 batches of what commercially available Radix Polygoni Multifloris is obtained, by the AIA of 25 batches of polygonum multiflorum medicinal material sample finger printing through experiment Formatted data file imports Chinese medicine fingerprint similarity evaluation systematic study version (2004A version), carries out Similarity Measure, With the highest Daba Mountain of each component content (numbering: S5) place of production be finger printing be with reference to collection of illustrative plates, use median method, Time window width is 0.1min, uses Supplements coupling, it is thus achieved that Radix Polygoni Multiflori reference fingerprint, sees Fig. 5, calculates Machine auxiliary similarity appraisal report the results are shown in Table 1.The finger printing of 25 batches of polygonum multiflorum medicinal material samples is shown in Fig. 6.
11 25 batches, table commercially available Radix Polygoni Multiflori Similarity Measure result
The reference fingerprint set up by the evaluation system of Chinese medicine fingerprint (is retained with sample finger printing globality Time and peak area) carry out similarity system design, thus Chinese medicine quality is evaluated and controls, thus draw concordance and difference The opposite sex.Similarity is typically thought between 0.85～1.0 and is met the requirements.Can substantially find out that each composition is certain from Fig. 6 In retention time range of error (0.1min) can have preferable overlapping with the reference fingerprint generated, similarity between Between 0.95～0.99, and 1 and No. 13 medical material similarity is between 0.45～0.70, using similarity more than 0.85 as Judge the limit that whether qualified polygonum multiflorum medicinal material is, then 1 and No. 13 medical material is judged to defective Radix Polygoni Multiflori sample.
To sum up peak is many, separating degree is good, reproducible, precision it can be seen that the fingerprint image of the present invention is composed for result of the test Height, the material amount for overall monitor Radix Polygoni Multiflori provides effective guarantee.
Meanwhile, utilize extracting method and the chromatographic condition of fingerprint atlas detection method of the present invention, it is also possible to record what head raw simultaneously 16 kinds of composition gallic acids, procyanidin B in crow1, catechin, epicatechol gallate, Barbaloin, Rhizoma Polygoni Cuspidati Glycosides, stilbene glucoside, ponticin, resveratrol, rheum emodin glycosides, physcione glycosides, aloe-emodin, chrysophanic acid, Chrysophanol, rheum emodin, the content of physcione, such that it is able to from the quality of medicinal material of more angle monitoring Radix Polygoni Multiflori.

Claims (10)

1. the HPLC fingerprint atlas detection method of Radix Polygoni Multiflori, it is characterised in that: it comprises the following steps:
(1) preparation of reference substance solution: take Radix Polygoni Multiflori medical material, pulverizes to obtain medicinal powder, 40～60% ethanol water carry Take, filter, obtain Radix Polygoni Multiflori reference substance solution；Preferably concentration of alcohol is 50%；
(2) preparation of object of reference solution: take gallic acid, procyanidin B1, catechin, epicatechol gallate, Aloe big Flavin glycosides, polygonin, stilbene glucoside, ponticin, resveratrol, rheum emodin glycosides, physcione glycosides, Aloe are big Flavin, chrysophanic acid, chrysophanol, rheum emodin, physcione reference substance are appropriate, mixing, add methanol, obtain object of reference Solution；
(3) foundation of reference fingerprint:
Take object of reference solution and the reference substance solution prepared by different batches Radix Polygoni Multiflori medical material respectively, inject efficient liquid phase Chromatograph detects, and obtains each batch medical material and the chromatographic data of object of reference；Chromatographic condition is as follows:
Detection wavelength: 254～275nm；
Chromatographic column: C18 chromatographic column and C18 pre-column；
Flowing phase: mobile phase A is 0.1% formic acid water, and Mobile phase B is acetonitrile；
Gradient elution program is as follows:
Gained chromatographic data is processed, sets up the reference fingerprint of Radix Polygoni Multiflori；
(4) preparation of need testing solution: take medical material to be measured, prepares test sample according to the method that step (1) is identical molten Liquid；
(5) according to the chromatographic condition that step (3) is identical, need testing solution is injected high performance liquid chromatograph detection, by institute Obtain chromatogram to compare with reference fingerprint, Radix Polygoni Multiflori and quality condition thereof can be identified.
Detection method the most according to claim 1, it is characterised in that: in step (1), described extraction supersound extraction.
Detection method the most according to claim 2, it is characterised in that: the time of described supersound extraction is 60min.
4. according to the detection method described in any one of claim 1-3, it is characterised in that: in step (1), described extraction Temperature be 40 DEG C.
Detection method the most according to claim 1, it is characterised in that: in step (1), described medical material ethanol is water-soluble The w/v of liquid is 1g:20mL.
Detection method the most according to claim 1, it is characterised in that: described C18 chromatographic column is ZORBAX C18 Chromatographic column, a length of 250mm, internal diameter is 4.6mm, and the particle diameter of filler is 5 μm；Described C18 pre-column is ZORBAX C18 chromatographic column, a length of 12.5mm, internal diameter is 4.6mm, and the particle diameter of filler is 5 μm..
7. according to the detection method described in any one of claim 1-5, it is characterised in that: the column temperature of described chromatographic condition is 30℃。
Detection method the most according to claim 7, it is characterised in that: the flow velocity of described chromatographic condition is 1.0mL/min.
9. according to the detection method described in any one of claim 1-8, it is characterised in that: described reference fingerprint such as figure Shown in 5.
10. one kind measures the method for 16 kinds of compositions in Radix Polygoni Multiflori simultaneously, it is characterised in that: described 16 kinds of compositions are no food Son acid, procyanidin B1, catechin, epicatechol gallate, Barbaloin, polygonin, stilbene glucoside, Radix Et Rhizoma Rhei Glycosides, resveratrol, rheum emodin glycosides, physcione glycosides, aloe-emodin, chrysophanic acid, chrysophanol, rheum emodin and big Flavin methyl ether；
It comprises the following steps:
A, the foundation of 16 kinds of ingredient standard curves:
Take 16 kinds of composition mixing reference substance solution, identical according to step (3) in claim 1-9 any one detection method Chromatographic condition detects, and obtains 16 kinds of respective standard curves of composition；
B, take Radix Polygoni Multiflori medical material, according to the method that step (1) in claim 1-9 any one detection method is identical, system The standby need testing solution that obtains, injection high performance liquid chromatography, with the chromatographic condition detection that step a is identical, according to the mark of step a Directrix curve obtains the content of 16 kinds of compositions in Radix Polygoni Multiflori.
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CN107490645A (en) * 2017-07-03 2017-12-19 中国人民解放军第三〇二医院 A kind of safe mass control method of prepared fleece flower root and application
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CN107179374A (en) * 2017-06-29 2017-09-19 江苏海昇药业有限公司 The detection method of tonic tablet for essence and blood finger-print
CN107179374B (en) * 2017-06-29 2019-10-22 江苏海昇药业有限公司 The detection method of tonic tablet for essence and blood finger-print
CN107490645A (en) * 2017-07-03 2017-12-19 中国人民解放军第三〇二医院 A kind of safe mass control method of prepared fleece flower root and application
CN113092623A (en) * 2021-04-07 2021-07-09 江西省科学院应用化学研究所 HPLC detection method for main flavonoid compound content in polygonum multiflorum leaves

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