CN109444275A - A kind of method of rapid screening Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme medicinal material - Google Patents
A kind of method of rapid screening Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme medicinal material Download PDFInfo
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Abstract
The invention discloses the methods of a kind of UPLC-MS-MS rapid screening Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme, include the following steps: that radix tetrastigme medicinal material to be distinguished is beaten to powder sieving is made test sample, it is added 10-20 times and measures 50-70% ethyl alcohol, ultrasonic extraction 20-40 minutes, test sample solution is obtained, UPLC-MS-MS network analysis is then carried out, obtains the corresponding mass spectrometric data of test sample, data analysis is carried out, chromatographic peak is carried out for test sample and extracts and Information in Mass Spectra characterization；According to the Chromatographic information and Information in Mass Spectra of test sample, judge whether contain the distinctive flavones characteristic compounds of Chinese wistaria radix tetrastigme in test sample.This method screening is quick, accuracy is high, greatly simplifies and improves the workload in radix tetrastigme difference germplasm (Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme) analytic process.
Description
Technical field
The present invention relates to Chinese medicine analysis and research technical fields, and in particular to one kind compound characterized by flavones uses
The method of UPLC-MS/MS rapid screening Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme medicinal material.
Background technique
Radix tetrastigme is the distinctive rare medicinal plant of China, is Vitaceae Tetrastigma plant tetratigma hemsleyanum
(Tetrastigma hemsleyanum Diels et Gilg), herb can be used as medicine, most with the medicinal effects of underground root tuber
It is good.Its is cool in nature, acrid flavour, hardship, nontoxic, has protection liver, antiviral, anti-inflammatory and antalgic, antitumor, clinically to measles complicated lung
A variety of diseases such as inflammation, infantile hyperpyrexia, tonsillitis also have significant curative effect.And it is widely used in a variety of Chinese patent drugs or health care product, such as China
He wind pain treasured, stone body, golden stilbene piece etc..
The place of production of radix tetrastigme is mainly distributed on the areas such as Zhejiang, Jiangxi, Fujian, Hubei, Hunan, Guangdong, Sichuan.Radix tetrastigme
Germplasm type is mainly distinguished from leaf blade size and shape, surface keratin degree, limb sawtooth, rattan color and shape, general big
Cause is divided into Chinese wistaria and two kinds of sinomenium acutum, there is biggish difference in terms of morphology therebetween, but configuration of medicinal materials is difficult to distinguish.Chinese wistaria
The new rattan of radix tetrastigme (also known as Zhejiang radix tetrastigme) is green, and the old rattan in plant lower part gradually becomes purple or brown after general 2~3 years.
Liana is relatively fine, there is corner angle, and blade is smaller, and root tuber is in calabash shaped more, is distributed mainly on the ground such as Zhejiang, Jiangxi.Sinomenium acutum radix tetrastigme
The surface Bluish white of (also known as Guangxi radix tetrastigme) old rattan, liana is more sturdy, and blade is larger, and root tuber individual is larger, is distributed mainly on
China Hunan, Fujian, Guangxi, Yunnan some areas.In contrast, sinomenium acutum radix tetrastigme distributional region is wider, grows vigorous, production
Amount is higher, and quantity is more.It is generally understood that sinomenium acutum radix tetrastigme root tuber is cold in nature, medicinal efficacy not as good as Chinese wistaria radix tetrastigme, traditionally with
Zhejiang produces Chinese wistaria radix tetrastigme as genunie medicinal materials use, therefore the price of Chinese wistaria radix tetrastigme will be much higher than sinomenium acutum radix tetrastigme in the market.
Chemical component in radix tetrastigme has flavones, flavonoid glycoside, starch, reduced sugar, steroid, amino acid etc., wherein
Flavones and its glycoside are a kind of important active materials.And show at present about a large amount of radix tetrastigme pharmacological researches of radix tetrastigme root tuber
Its antitumor effective substance is flavones ingredient.Since market demand is increasing, so that price of medicinal material persistently soars,
However, wild Chinese wistaria radix tetrastigme resource is very short, causes its wild resource by exhaustive exploitation, do great damage.Planting
It is big to train difficulty, root tuber growth in medicinal part underground is slow, needs the requirement that can be only achieved commodity herb in 3~5 years.And it is different
There are bright for the place of production, the underground beet yield of different germplasm origins (Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme) and active constituent content
Significant difference is different.Medicinal material market Top-three Leaves blueness germplasm mixes at present, the medicine of different germplasm origins (Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme)
Material price variance great disparity.Therefore occur filling with the common adulterant such as cheap sinomenium acutum radix tetrastigme or sibling species in the market
When authentic Chinese wistaria radix tetrastigme is sold to reap staggering profits the phenomenon that, it is difficult to guarantee the safety and validity of medication.
The quickening of modernization of Chinese medicine paces puts forward new requirements Chinese medicine standardization, and standardized production is Chinese Medicine Industry hair
Exhibition and the key for entering international market, and GAP (Good Agriculturing Practice, the Chinese medicine carried out energetically at present
Material quality of production management regulation) working centre.And variety standard and the identification method of science are quality standard control and management
Premise, therefore the method for Materia Medica Identification will meet quick, accurate and standardization etc. and require.However the identification of radix tetrastigme at present
Method is mainly analyzed according to morphology and routine chemical components, or even only relies on micro-judgment, there is not yet with modern analysis
Method establishes the research report of Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme Rapid Identification.For specification radix tetrastigme market, ensures and face
Bed curative effect, protection consumer's interests have very important realistic meaning.Therefore, it is necessary to develop a kind of practicability height, energy
Quickly, the method for precise Identification difference germplasm radix tetrastigme solves to interweave truth with fiction in medicinal material market, the problems such as quality is very different.
Currently, the chemical constitution study in radix tetrastigme is it has been reported that but using clear ingredient as the Germplasm Identification of index and matter
Amount control but has not been reported.In recent years, with the development of analytical technology, the triple level four bars of ultra performance liquid chromatography (UPLC) series connection
Mass spectrum (TQD-MS/MS) is since its separating rate is fast, high sensitivity, measurement accuracy is strong and is widely used in Chinese medicine and compound
Analysis in.
Currently, the discrimination method to Chinese medicine product mainly has PCR, IR, NIR etc., exist in these methods a large amount of false
Positive and Problem of False Negative, when causing to be analyzed using these methods, may omit important material information, and then lead to institute
The result filtered out can not effectively identify, and bring difficulty to the Chinese medical herb of the same race of the different germplasm of determination.The true and false of radix tetrastigme
And there are many methods for place of production discriminating, such as: quickly identifying the PCR-RFLP method of radix tetrastigme and its a variety of pseudo- mixed product
(ZL201510016355.4), a kind of method (ZL201310373809.4) for identifying medicinal plant radix tetrastigme, a kind of identification three
The method (201710762908.X) in the place of production Ye Qing, a kind of near infrared spectrum quickly detect radix tetrastigme place of production method
(201710371389.4), however it there is no and effectively identify and distinguish different germplasm radix tetrastigme (three leaves of Chinese wistaria radix tetrastigme and sinomenium acutum
It is green) method of medicinal material.
Summary of the invention
The purpose of the present invention is in view of the above deficiencies, provide a kind of UPLC-MS-MS rapid screening quick, accuracy is high
The method of Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme medicinal material, to greatly simplify and improve radix tetrastigme difference germplasm (Chinese wistaria radix tetrastigme
With sinomenium acutum radix tetrastigme) workload in analytic process.
In order to achieve the above objectives, present invention employs following technical proposals:
A kind of method of rapid screening Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme medicinal material, comprising the following steps:
1) radix tetrastigme medicinal material to be distinguished is played into powder sieving and test sample is made；
2) test sample of step 1) is taken, 10-20 times is added and measures 50-70% ethyl alcohol, ultrasonic extraction 20-40 minutes, obtain confession
Test agent solution then carries out UPLC-MS-MS network analysis, obtains the corresponding mass spectrometric data of test sample；
3) data that step 2) acquires are introduced directly into mass spectral analysis software, then carry out data analysis, for for examination
Sample carries out chromatographic peak and extracts and Information in Mass Spectra characterization；
4) Chromatographic information and Information in Mass Spectra of the test sample obtained according to step 3), judges whether contain in test sample
Kaempferol-O- β-D- furans celery sugar -7-O- alpha-L-rhamnoside and Kaempferol -7-O- α-L- rhamnose -3-O- β-D- pyrans
Glucoside；Such as contain Kaempferol-O- β-D- furans celery sugar -7-O- alpha-L-rhamnoside and Kaempferol -7-O- α-L- sandlwood
Sugar -3-O- β-D- glucopyranoside, then be Chinese wistaria radix tetrastigme；It does not contain, is then sinomenium acutum radix tetrastigme.
Preferably, in step 4), pass through the corresponding quasi-molecular ion peak of peak area and peak retention time of chromatography
Mass number and second order ms information judge whether contain Kaempferol-O- β-D- furans celery sugar -7-O- α-L- in test sample
Rhamnoside and Kaempferol -7-O- α-L- rhamnose -3-O- β-D- glucopyranoside.
Kaempferol-O- β-D- furans celery sugar -7-O- alpha-L-rhamnoside in step 4): molecular weight 726, molecular formula
C32H38O19, second order ms fragment is 647.5,527.4,486.6,431.9,392.1,289.7.
Kaempferol -7-O- α-L- rhamnose -3-O- β-D- glucopyranoside in step 4): molecular weight 594, molecule
Formula C27H30O15, second order ms fragment is 515.5,451.7,386.5,291.0,246.5.
Preferably, test sample beats powder and crosses 60 meshes in step 1), it is kept in dark place at -20 DEG C；
Preferably, in step 2), the test sample 1.0g of step 1) is taken, it is accurately weighed, it sets in stuffed conical flask, it is accurate
50% ethyl alcohol 10ml is added, weighed weight is ultrasonically treated 30 minutes, lets cool, then weighed weight, supply weightlessness with 50% ethyl alcohol,
It shakes up, is centrifuged, precision measures supernatant 5ml, sets in 25ml brown measuring bottle, adds 50% ethyl alcohol to scale, shakes up, filter, take continuous
Filtrate filtering, obtains test sample solution.
Preferably, mass spectral analysis software is MassLynx V4.1 software in step 3).
Preferably, in step 2), the analysis condition of the UPLC-MS-MS system are as follows:
Chromatographic condition: Waters ACQUITY UPLC BEH C18 chromatographic column, 2.1 × 100mm, 1.7 μm；With 0.1% first
Acid-water is mobile phase A, and acetonitrile is Mobile phase B, and gradient elution is as shown in the table:
Flow velocity: 0.3ml/min；Column temperature is 30 DEG C；Detection wavelength is 254nm；
Mass Spectrometry Conditions:
MS system: the triple level four bars mass spectrometers of ACQUITY UPLC-TQD-MS；The use condition of tandem mass spectrum instrument
Are as follows: ionization mode: ESI (-)；Detection mode: more reaction detections；Capillary voltage: 3.0Kv；Orifice potential: 38V；Source temperature
120℃；Remove solvent temperature: 350 DEG C；Go solvent flow rate: 500L/hr；Taper hole gas velocity: 50L/hr；Collision energy: 19ev；It stays
Stay the time: 0.2s；Full scan: surface sweeping range is molecular weight 100~1000.
Compared with prior art, the present invention having the following beneficial effects:
(1) method that the present invention utilizes UPLC-MS-MS rapid screening Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme for the first time, the party
Method has many advantages, such as quick, accurate, stable；To greatly simplify and improve radix tetrastigme difference germplasm (Chinese wistaria radix tetrastigme and sinomenium acutum
Radix tetrastigme) workload in analytic process, while passing through diagnostic fragment filtering policy and other fragments and neutral loss molecule
Assistant analysis is conducive to the Fast Classification and identification of complicated ingredient.By establishing diagnostic fragment filtering policy and neutral loss mistake
Filter strategy, and be used for the rapid screening of Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme, efficiently solve identification and distinguish Chinese wistaria radix tetrastigme
With the technical problem of the method for sinomenium acutum radix tetrastigme.
(2) present invention passes through extraction using the test sample of Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme medicinal material as test sample
The optimization of technique is investigated, and research obtains the preparation process of Chinese wistaria radix tetrastigme Yu sinomenium acutum radix tetrastigme test sample；It is built in conjunction with the present invention
The Chinese wistaria radix tetrastigme of different germplasm and the screening method of sinomenium acutum radix tetrastigme are stood, can be promoted and applied in Chinese wistaria radix tetrastigme and three leaf of sinomenium acutum
In the selection of the raw material of green medicine materical crude slice and its related preparations and preparation process.
Detailed description of the invention
Fig. 1 is the UPLC chromatography comparison chart of Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme test sample；
Fig. 2 is the TIC comparison diagram of Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme test sample；
Fig. 3 is Main Flavonoids constituent structure figure in Chinese wistaria radix tetrastigme；
Fig. 4 is Main Flavonoids constituent structure figure in sinomenium acutum radix tetrastigme；
Fig. 5 is the principal component analysis PCA figure of Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme test sample.
Specific embodiment
The present invention will be further described in detail with reference to the accompanying drawings and detailed description.
A kind of method of rapid screening Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme medicinal material, comprising the following steps:
1) radix tetrastigme medicinal material to be distinguished is played into powder sieving and test sample is made；
2) test sample of step 1) is taken, 10-20 times is added and measures 50-70% ethyl alcohol, ultrasonic extraction 20-40 minutes, obtain confession
Test agent solution then carries out UPLC-MS-MS network analysis, obtains the corresponding mass spectrometric data of test sample；
3) data that step 2) acquires are introduced directly into mass spectral analysis software, then carry out data analysis, for for examination
Sample carries out chromatographic peak and extracts and Information in Mass Spectra characterization；
4) Chromatographic information and Information in Mass Spectra of the test sample obtained according to step 3), judges whether contain in test sample
Kaempferol-O- β-D- furans celery sugar -7-O- alpha-L-rhamnoside and Kaempferol -7-O- α-L- rhamnose -3-O- β-D- pyrans
Glucoside；Such as contain Kaempferol-O- β-D- furans celery sugar -7-O- alpha-L-rhamnoside and Kaempferol -7-O- α-L- sandlwood
Sugar -3-O- β-D- glucopyranoside, then be Chinese wistaria radix tetrastigme；It does not contain, is then sinomenium acutum radix tetrastigme.
Above-mentioned Kaempferol-O- β-D- furans celery sugar -7-O- alpha-L-rhamnoside and Kaempferol -7-O- α-L- rhamnose -
3-O- β-D- glucopyranoside is the otherness compound of Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme, by the two flavones features
Compound can quickly and effectively distinguish Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme.
Specifically, the present embodiment obtains Chinese wistaria radix tetrastigme and the process of the otherness compound of sinomenium acutum radix tetrastigme is as follows:
A kind of method of UPLC-MS-MS rapid screening Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme, comprising the following steps:
1) Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme medicinal material to be distinguished are collected, by the Chinese wistaria three of each 20 batch of different sources
Corresponding Chinese wistaria radix tetrastigme test sample and sinomenium acutum radix tetrastigme test sample is respectively prepared in leaf blueness medicinal material and sinomenium acutum radix tetrastigme medicinal material,
It crushes, crosses 60 meshes, be kept in dark place at -20 DEG C；
Wherein, Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme test sample information are as shown in the table:
2) the Chinese wistaria radix tetrastigme of step 1) and test sample each 1.0g of sinomenium acutum radix tetrastigme are taken, it is accurately weighed, tool is set respectively
It fills in conical flask, 50% ethyl alcohol 10ml is added in precision, and weighed weight is ultrasonically treated 30 minutes, lets cool, then weighed weight, uses
50% ethyl alcohol supplies weightlessness, shakes up, and centrifugation, precision measures supernatant 5ml, sets in 25ml brown measuring bottle, adds 50% ethyl alcohol to quarter
Degree, shakes up, and filters, takes subsequent filtrate to filter, obtain test solution, then carries out UPLC-MS-MS network analysis, respectively obtains
Each Chinese wistaria radix tetrastigme mass spectrometric data corresponding with sinomenium acutum radix tetrastigme test sample；
Wherein, the analysis condition of UPLC-MS-MS system are as follows:
Chromatographic condition: Waters ACQUITY UPLC BEH C18 chromatographic column, 2.1 × 100mm, 1.7 μm；With 0.1% first
Acid-water is mobile phase A, and acetonitrile is Mobile phase B, and gradient elution is as shown in the table:
Flow velocity: 0.3ml/min；Column temperature is 30 DEG C；Detection wavelength is 254nm；
Mass Spectrometry Conditions: MS system: the triple level four bars mass spectrometers of ACQUITY UPLC-TQD-MS.
The use condition of tandem mass spectrum instrument are as follows: ionization mode: ESI (-)；Detection mode: more reaction detections；Capillary
Voltage: 3.0Kv；Orifice potential: 38V；120 DEG C of source temperature；Remove solvent temperature: 350 DEG C；Go solvent flow rate: 500L/hr；Taper hole gas
Flow velocity: 50L/hr；Collision energy: 19ev；Residence time: 0.2s；Full scan: surface sweeping range is molecular weight 100~1000.
3) mass spectrometric data that step 2) acquires is introduced directly into MassLynx V4.1 software, then carries out data analysis,
Chromatographic peak is carried out for each sample to extract and Information in Mass Spectra characterization；
4) the compound chromatographic peak for selecting two kinds of radix tetrastigmes shared is as reference, using Dynamic Programming calculating method correcting sample
Between chromatographic peak drift, alignment belongs to the chromatographic peak of same substance, then using closest clustering method by retention time more
Close substance is registered as a compound, finally establishes the accreditation charts of a sample and chromatographic peak；It is corrected by external standard
Afterwards, it is used for subsequent analysis；The high performance liquid chromatography ratio of the test sample of Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme as shown in figure 1
To figure, the results show that Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme have notable difference on UPLC chromatogram.
5) Chinese wistaria three is determined by the mass number of the peak area of chromatography and the corresponding quasi-molecular ion peak of peak retention time
The original document of the Information in Mass Spectra of gained otherness substance is importing directly by the otherness compound of Ye Qingyu sinomenium acutum radix tetrastigme
MassLynx V4.1 further studies its second level Information in Mass Spectra, determines the structure of matter, to obtain otherness substance.
Shown in the following Tables 1 and 2 of Main Flavonoids class compound contained in Chinese wistaria radix tetrastigme:
The relative peak area (mAU/g) of Main Flavonoids class compound in 1 Chinese wistaria radix tetrastigme of table
The Main Flavonoids constituents that 2 Chinese wistaria radix tetrastigme UPLC-MS/MS of table is analyzed
Main Flavonoids class compound is as shown in the following table 3 and table 4 in sinomenium acutum radix tetrastigme:
The relative peak area (mAU/g) of Main Flavonoids class compound in 3 sinomenium acutum radix tetrastigme of table
The Main Flavonoids constituents that 4 sinomenium acutum radix tetrastigme UPLC-MS/MS of table is analyzed
Fig. 2 shows the TIC comparison diagrams of Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme test sample, utilize principal component analysis
(PCA) Chinese wistaria radix tetrastigme and the total ion current diagram data of sinomenium acutum radix tetrastigme test sample are analyzed, as the result is shown (as schemed
5), every group of data are gathered in respective 95% confidence limit ellipse figure, and two groups of data can be kept completely separate, and illustrate that two is various
The Chinese wistaria radix tetrastigme of matter with sinomenium acutum radix tetrastigme test sample is available is totally separated from.Then by the peak area of chromatography and out
The mass number of the corresponding quasi-molecular ion peak of peak retention time determines the otherness compound of Chinese wistaria radix tetrastigme Yu sinomenium acutum radix tetrastigme,
The Information in Mass Spectra of main difference substance is used and carries out second mass analysis in MassLynxV4.1 software, and combines correlationization
Close object report determine the structure of matter (with reference to J.Nat.Med., 2011,65 (2), 360-363, Chinese herbal medicine, 2017,48 (5),
874 etc.), to obtain main difference substance；As the result is shown:
1, in addition to shared compound catechin, Kaempferol-O- β-D- furans celery sugar -7-O- α-in Chinese wistaria radix tetrastigme sample
L- rhamnoside, Kaempferol -7-O- α-L- rhamnose -3-O- β-D- glucopyranoside, Kaempferol-O- β-D- mulberry cloth disaccharide
Glycosides, Quercetin -7-O- alpha-L-rhamnoside, Kaempferol -7-O- alpha-L-rhamnoside are main flavones ingredient (such as Fig. 3 and table
2).Nicotifiorin in sinomenium acutum radix tetrastigme sample, Quercetin -3-O- β-D- xylosyl glucose -7-O- alpha-L-rhamnoside, locust tree
Glycosides, Isorhamnetin -3-O- β-D-Glucose -7-O- alpha-L-rhamnoside, rutin, isoquercitrin, astragalin are Main Flavonoids
Constituents (Fig. 4 and table 4)；
B. wherein the higher flavones of content is Kaempferol-O- β-D- furans celery sugar -7-O- α-L- mouse in Chinese wistaria radix tetrastigme
Lee's glucosides and Kaempferol -7-O- α-L- rhamnose -3-O- β-D- glucopyranoside, and do not contained in sinomenium acutum radix tetrastigme, therefore
It can be by Kaempferol-O- β-D- furans celery sugar -7-O- alpha-L-rhamnoside combination Kaempferol -7-O- α-L- rhamnose -3-O- β -
Flavones characteristic compounds of the D- glucopyranoside as Chinese wistaria radix tetrastigme.
In addition, the influence of comparative measurements different solvents of the present invention and extracting mode to medicinal material general flavone content, specific method
As follows, experimental result see the table below.
1, ultrasonic extracting method
Take Chinese wistaria radix tetrastigme medicinal material, sort and remove impurity, grinding medicinal materials simultaneously cross No. 4 sieves, take 4 parts of 1.0g medicinal powder, add pure water,
50% ethyl alcohol, 70% ethyl alcohol and straight alcohol distinguish 10ml, ultrasonic extraction 30min, extracting solution filtration to get.
2, reflux extraction method
Take Chinese wistaria radix tetrastigme medicinal material, sort and remove impurity, grinding medicinal materials simultaneously cross No. 4 sieves, take 4 parts of 1.0g medicinal powder, add pure water,
50% ethyl alcohol, 70% ethyl alcohol and straight alcohol distinguish 10ml, start timing, and carry out heating water bath reflux, take out medical fluid after 1h, mention
Take liquid to filter to get.
The different extracting modes of table 5 and solvent influence result to test sample parameter
The experimental results showed that flavones content is related to the mode of extraction and solvent, but extract under identical extraction conditions molten
Agent is 50% and 70% ethyl alcohol without too big difference；Refluxing extraction flavones content is compared with ultrasonic extraction height, but UPLC-MS-MS is analyzed
The ratio of flavones is substantially higher compared to refluxing extraction in ultrasonic extraction liquid as the result is shown, which may be flavonoid glycoside in high temperature item
Unstable or other impurities are easier to be extracted at high temperature under part.It therefore, is raising Chinese wistaria radix tetrastigme test sample
The ratio of middle flavones, the extraction process of Chinese wistaria radix tetrastigme test sample are set to 50% EtOH Sonicate and extract 30min.
The embodiments described above only express several embodiments of the present invention, and the description thereof is more specific and detailed, but simultaneously
It cannot therefore be construed as limiting the scope of the patent.It should be pointed out that coming for those of ordinary skill in the art
It says, without departing from the inventive concept of the premise, various modifications and improvements can be made, these belong to protection of the invention
Range.Therefore, the scope of protection of the patent of the invention shall be subject to the appended claims.
Claims (8)
1. a kind of method of rapid screening Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme medicinal material, which comprises the following steps:
1) radix tetrastigme medicinal material to be distinguished is played into powder sieving and test sample is made；
2) test sample of step 1) is taken, 10-20 times is added and measures 50-70% ethyl alcohol, ultrasonic extraction 20-40 minutes, obtain for sample
Product solution then carries out UPLC-MS-MS network analysis, obtains the corresponding mass spectrometric data of test sample；
3) data that step 2) acquires are introduced directly into mass spectral analysis software, data analysis are then carried out, for test sample
Chromatographic peak is carried out to extract and Information in Mass Spectra characterization；
4) Chromatographic information and Information in Mass Spectra of the test sample obtained according to step 3), judges whether contain kaempferia galanga in test sample
Phenol -3-O- β-D- furans celery sugar -7-O- alpha-L-rhamnoside and Kaempferol -7-O- α-L- rhamnose -3-O- β-D- glucopyra
Glucosides；Such as contain Kaempferol-O- β-D- furans celery sugar -7-O- alpha-L-rhamnoside and Kaempferol -7-O- α-L- rhamnose -3-
O- β-D- glucopyranoside is then Chinese wistaria radix tetrastigme；It does not contain, is then sinomenium acutum radix tetrastigme.
2. a kind of method of rapid screening Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme medicinal material according to claim 1, feature
It is, in step 4), passes through the mass number and two of the corresponding quasi-molecular ion peak of peak area and peak retention time of chromatography
Grade Information in Mass Spectra judges whether contain Kaempferol-O- β-D- furans celery sugar -7-O- alpha-L-rhamnoside and mountain in test sample
Naphthol -7-O- α-L- rhamnose -3-O- β-D- glucopyranoside.
3. a kind of method of rapid screening Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme medicinal material according to claim 1, feature
It is, the Kaempferol-O- β-D- furans celery sugar -7-O- alpha-L-rhamnoside in step 4): molecular weight 726, molecular formula
C32H38O19, second order ms fragment is 647.5,527.4,486.6,431.9,392.1,289.7.
4. a kind of method of rapid screening Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme medicinal material according to claim 1, feature
It is, the Kaempferol -7-O- α-L- rhamnose -3-O- β-D- glucopyranoside in step 4): molecular weight 594, molecular formula
C27H30O15, second order ms fragment is 515.5,451.7,386.5,291.0,246.5.
5. a kind of method of rapid screening Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme medicinal material according to claim 1, feature
It is, in step 1), test sample beats powder and crosses 60 meshes, is kept in dark place at -20 DEG C.
6. a kind of method of rapid screening Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme medicinal material according to claim 1, feature
It is, in step 2), takes the test sample 1.0g of step 1), it is accurately weighed, it sets in stuffed conical flask, 50% ethyl alcohol is added in precision
10ml, weighed weight are ultrasonically treated 30 minutes, let cool, then weighed weight, supply weightlessness with 50% ethyl alcohol, shake up, and are centrifuged, essence
Close measurement supernatant 5ml, sets in 25ml brown measuring bottle, adds 50% ethyl alcohol to scale, shakes up, filter, take subsequent filtrate to filter, obtain
Test sample solution.
7. a kind of method of rapid screening Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme medicinal material according to claim 1, feature
It is, in step 3), mass spectral analysis software is MassLynx V4.1 software.
8. a kind of method of rapid screening Chinese wistaria radix tetrastigme and sinomenium acutum radix tetrastigme medicinal material according to claim 1, feature
It is, in step 2), the analysis condition of the UPLC-MS-MS system are as follows:
Chromatographic condition: Waters ACQUITY UPLC BEH C18 chromatographic column, 2.1 × 100mm, 1.7 μm；With 0.1% formic acid-
Water is mobile phase A, and acetonitrile is Mobile phase B, and gradient elution is as shown in the table:
Flow velocity: 0.3ml/min；Column temperature is 30 DEG C；Detection wavelength is 254nm；
Mass Spectrometry Conditions:
MS system: the triple level four bars mass spectrometers of ACQUITY UPLC-TQD-MS；The use condition of tandem mass spectrum instrument are as follows: from
Sub-ization mode: ESI (-)；Detection mode: more reaction detections；Capillary voltage: 3.0Kv；Orifice potential: 38V；120 DEG C of source temperature；
Remove solvent temperature: 350 DEG C；Go solvent flow rate: 500L/hr；Taper hole gas velocity: 50L/hr；Collision energy: 19ev；Residence time:
0.2s；Full scan: surface sweeping range is molecular weight 100~1000.
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