CN110398560A - A kind of UPLC method detecting atractylenolide Ⅰ and atractylodes lactone III in Rhizoma Atractylodis Macrocephalae drug - Google Patents
A kind of UPLC method detecting atractylenolide Ⅰ and atractylodes lactone III in Rhizoma Atractylodis Macrocephalae drug Download PDFInfo
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- CN110398560A CN110398560A CN201910725324.4A CN201910725324A CN110398560A CN 110398560 A CN110398560 A CN 110398560A CN 201910725324 A CN201910725324 A CN 201910725324A CN 110398560 A CN110398560 A CN 110398560A
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- rhizoma atractylodis
- atractylodis macrocephalae
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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
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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
- G01N30/04—Preparation or injection of sample to be analysed
- G01N30/06—Preparation
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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
- G01N30/04—Preparation or injection of sample to be analysed
- G01N2030/042—Standards
- G01N2030/047—Standards external
Abstract
The present invention provides a kind of UPLC methods of atractylenolide Ⅰ and atractylodes lactone III in detection Rhizoma Atractylodis Macrocephalae drug, the following steps are included: S1, Rhizoma Atractylodis Macrocephalae sample preparation to be measured: first Rhizoma Atractylodis Macrocephalae crushing being placed in triangular flask, it is placed in progress ultrasound, centrifugation in ultrasonoscope after methanol is added, is finally filtered；S2, the measurement of standard Rhizoma Atractylodis Macrocephalae sample: standard Rhizoma Atractylodis Macrocephalae sample solution is injected in Ultra Performance Liquid Chromatography instrument and is measured, the UPLC characteristic spectrum of standard Rhizoma Atractylodis Macrocephalae sample is obtained；S3, Rhizoma Atractylodis Macrocephalae sample to be measured measurement: it will be measured in Rhizoma Atractylodis Macrocephalae sample to be measured in step S1 measurement liquid injection Ultra Performance Liquid Chromatography instrument, the UPLC characteristic spectrum of Rhizoma Atractylodis Macrocephalae sample to be measured is obtained, the UPLC characteristic spectrum of the standard Rhizoma Atractylodis Macrocephalae sample obtained by step S2 and the UPLC characteristic spectrum of Rhizoma Atractylodis Macrocephalae sample to be measured quantify the Rhizoma Atractylodis Macrocephalae sample to be measured being prepared.The analysis test of Rhizoma Atractylodis Macrocephalae sample can be rapidly completed in method provided by the invention, and sample precision and recovery of standard addition are preferable.
Description
Technical field
The present invention relates to atractylenolide Ⅰ and atractylodes lactones in detection technique field more particularly to a kind of detection Rhizoma Atractylodis Macrocephalae drug
III UPLC method.
Background technique
Rhizoma Atractylodis Macrocephalae is a kind of traditional Chinese medicine, is the dry rhizome of compositae plant Rhizoma Atractylodis Macrocephalae, is herbaceos perennial；Its property
Temperature, it is sweet in flavor, bitter, have strengthening the spleen and replenishing qi, eliminate dampness and have diuretic effect, hidroschesis, miscarriage prevention function.It is mainly used for spleen eating less, indigestion, abdominal distension
Diarrhea, lassitude, phlegm retention anti-dazzle nervous, oedema, spontaneous perspiration, threatened abortion are Chinese medicine tonifying spleen key medicine.Modern pharmacological studies have shown that white
Art has liver protection, cholagogue, diuresis, antibacterial, anti-inflammatory, antitumor, strengthen immunity, decompression, hypoglycemic, anticoagulation, strong, calm etc.
Effect.
Atractylodes lactone is one of Rhizoma Atractylodis Macrocephalae compound, and atractylenolide Ⅰ and atractylodes lactone III are the main lactone in Rhizoma Atractylodis Macrocephalae
Ingredient, Rhizoma Atractylodis Macrocephalae II are the dehydration product of atractylodes lactone III.It has been generally acknowledged that atractylenolide Ⅰ and atractylodes lactone III are the main work of Rhizoma Atractylodis Macrocephalae
Property ingredient, be that the key that its anti-inflammatory and anticancer effective component, atractylenolide Ⅰ and atractylodes lactone III are controlled as Rhizoma Atractylodis Macrocephalae quality refers to
One of mark, content can be used as the main indicator of Rhizoma Atractylodis Macrocephalae quality control.
The document report of the measuring method of atractylodes lactone is more in Rhizoma Atractylodis Macrocephalae, but mostly uses GC, HPLC, mass spectroscopy wherein one
Kind or several effective components, the larger and extraction step of GC extraction sample loss is cumbersome, and HPLC analytical cycle is longer, solvent consumption
Larger, mass spectrometer is expensive.Quality control about Rhizoma Atractylodis Macrocephalae fails to form unified standards system, version " China in 2015 so far
Pharmacopeia " content determination item of Rhizoma Atractylodis Macrocephalae is not yet recorded in (one).Therefore, establish stably and controllable, precise and high efficiency atractylenolide Ⅰ and
The measuring method of III content of atractylodes lactone is of great significance.
Summary of the invention
In order to solve the above technical problems, the present invention provides atractylenolide Ⅰ and atractylodes lactones in a kind of detection Rhizoma Atractylodis Macrocephalae drug
III UPLC method, comprising the following steps:
S1, Rhizoma Atractylodis Macrocephalae sample preparation to be measured: first Rhizoma Atractylodis Macrocephalae crushing is placed in triangular flask, is placed in ultrasonic wave after methanol is added
Ultrasound, centrifugation are carried out in instrument, are finally filtered；
S2, the measurement of standard Rhizoma Atractylodis Macrocephalae sample: standard Rhizoma Atractylodis Macrocephalae sample solution is injected in Ultra Performance Liquid Chromatography instrument and is measured, is obtained
The UPLC characteristic spectrum of standard Rhizoma Atractylodis Macrocephalae sample；
S3, Rhizoma Atractylodis Macrocephalae sample to be measured measurement: Rhizoma Atractylodis Macrocephalae sample to be measured in step S1 measurement liquid is injected into Ultra Performance Liquid Chromatography instrument
Middle measurement obtains the UPLC characteristic spectrum of Rhizoma Atractylodis Macrocephalae sample to be measured, passes through the UPLC feature of the obtained standard Rhizoma Atractylodis Macrocephalae sample of step S2
The UPLC characteristic spectrum of map and Rhizoma Atractylodis Macrocephalae sample to be measured quantifies the Rhizoma Atractylodis Macrocephalae sample to be measured being prepared.
Wherein, the chromatographic condition of ultra performance liquid chromatography are as follows:
Chromatographic column: BEH C18 column (2.1 μ m 100mm, 1.7mm)；
Flow velocity: 0.3mL/min；
Column temperature: 30 DEG C；
Sample volume: 5 μ L；
Ultraviolet detection wavelength: 221nm；
Mobile phase: acetonitrile and water, wherein the volume ratio of acetonitrile and water is 54:46.
Wherein, the content of atractylodes lactone is calculated as follows in the Rhizoma Atractylodis Macrocephalae sample to be measured:
In formula:
The content of atractylodes lactone in X-Rhizoma Atractylodis Macrocephalae sample to be measured；
Cs- from the content for obtaining atractylodes lactone in Rhizoma Atractylodis Macrocephalae sample to be measured measurement liquid in standard curve；
miThe quality of-sample to be tested；
V-Rhizoma Atractylodis Macrocephalae sample to be measured measures liquid constant volume.
Wherein, in the step S1 Rhizoma Atractylodis Macrocephalae sample preparation to be measured method specifically: weigh 5g after first crushing Rhizoma Atractylodis Macrocephalae and set
In 50mL triangular flask, it is placed in ultrasound 20-45min in ultrasonoscope after 20-30mL methanol is added, control revolving speed is 2500-
3500r/min is centrifuged 10-30min, finally with 0.45 μm of membrane filtration.
Wherein, the step S2 Plays Rhizoma Atractylodis Macrocephalae sample solution is prepared by following steps:
(1) preparation of atractylodes lactone I standard working solution:
The preparation of 1.1 100g/L atractylodes lactone I standard reserving solutions: 1.02g atractylodes lactone I standard items (content is weighed
98%) it, with methanol constant volume to 10mL, shakes up；
The preparation of 1.2 100mg/L atractylodes lactone I standard working solutions: the Rhizoma Atractylodis Macrocephalae for the 100g/L for taking step 1.1 to be prepared
10 μ L of lactone I standard reserving solution is shaken up with methanol constant volume to 10mL；
1.3 1,5,10,20, the preparation of 50mg/L atractylodes lactone I standard working solution: step 1.2 is taken to be prepared respectively
10,50,100,200, the 500 μ L of atractylodes lactone I standard working solution of 100mg/L is shaken up with methanol constant volume to 1mL；
(2) preparation of atractylodes lactone III standard working solution:
The preparation of 1.1 100g/L atractylodes lactone III standard reserving solutions: 1.02g atractylodes lactone III standard items (content is weighed
98%) it, with methanol constant volume to 10mL, shakes up；
The preparation of 1.2 100mg/L atractylodes lactone III standard working solutions: the 100g/L's for taking step 1.1 to be prepared is white
10 μ L of art lactone II I standard reserving solution is shaken up with methanol constant volume to 10mL；
1.3 1,5,10,20, the preparation of 50mg/L atractylodes lactone III standard working solution: step 1.2 is taken to be prepared respectively
10,50,100,200, the 500 μ L of atractylodes lactone III standard working solution of 100mg/L shaken up with methanol constant volume to 1mL.
Beneficial effects of the present invention:
The UPLC method of atractylenolide Ⅰ and atractylodes lactone III, passes through ultra high efficiency in detection Rhizoma Atractylodis Macrocephalae drug provided by the invention
Liquid chromatography analysis measures the composition and content of atractylodes lactone I and atractylodes lactone III in Rhizoma Atractylodis Macrocephalae, simple and efficient to handle.It is setting
It under fixed chromatographic condition, is detected through UV detector, qualitative with retention time, peak area quantification is completed within 6min
The analysis of atractylodes lactone I and atractylodes lactone III in sample are tested, and sample precision and recovery of standard addition are preferable, are able to satisfy white
The requirement of art quality evaluation has stronger practical value.
Detailed description of the invention
It, below will be simple to needing attached drawing to be used to make in embodiment in order to illustrate more clearly of technical solution of the present invention
Singly introduce, it should be apparent that, the accompanying drawings in the following description is only some embodiments of the present invention, corresponds to the general of this field
For logical technical staff, without creative efforts, it is also possible to obtain other drawings based on these drawings.
Fig. 1 is the UPLC map of standard Rhizoma Atractylodis Macrocephalae sample provided in an embodiment of the present invention；
Fig. 2 is the UPLC map of Rhizoma Atractylodis Macrocephalae sample to be measured provided in an embodiment of the present invention.
Specific embodiment
It is the preferred embodiment of the present invention below, it is noted that for those skilled in the art,
Various improvements and modifications may be made without departing from the principle of the present invention, these improvements and modifications are also considered as this hair
Bright protection scope.
The present invention provides a kind of UPLC methods of atractylenolide Ⅰ and atractylodes lactone III in detection Rhizoma Atractylodis Macrocephalae drug, including with
Lower step:
S1, the chromatographic condition for setting ultra performance liquid chromatography:
Chromatographic column: BEH C18 column (2.1 μ m 100mm, 1.7mm)；
Flow velocity: 0.3mL/min；
Column temperature: 30 DEG C；
Sample volume: 5 μ L；
Ultraviolet detection wavelength: 221nm；
Mobile phase: acetonitrile and water, wherein the volume ratio of acetonitrile and water is 54:46.
The preparation and measurement of S2, standard Rhizoma Atractylodis Macrocephalae sample solution:
(1) preparation of atractylodes lactone I standard working solution:
The preparation of 1.1 100g/L atractylodes lactone I standard reserving solutions: 1.02g atractylodes lactone I standard items (content is weighed
98%) it, with methanol constant volume to 10mL, shakes up；
The preparation of 1.2 100mg/L atractylodes lactone I standard working solutions: the Rhizoma Atractylodis Macrocephalae for the 100g/L for taking step 1.1 to be prepared
10 μ L of lactone I standard reserving solution is shaken up with methanol constant volume to 10mL；
1.3 1,5,10,20, the preparation of 50mg/L atractylodes lactone I standard working solution: step 1.2 is taken to be prepared respectively
10,50,100,200, the 500 μ L of atractylodes lactone I standard working solution of 100mg/L is shaken up with methanol constant volume to 1mL；
(2) preparation of atractylodes lactone III standard working solution:
The preparation of 1.1 100g/L atractylodes lactone III standard reserving solutions: 1.02g atractylodes lactone III standard items (content is weighed
98%) it, with methanol constant volume to 10mL, shakes up；
The preparation of 1.2 100mg/L atractylodes lactone III standard working solutions: the 100g/L's for taking step 1.1 to be prepared is white
10 μ L of art lactone II I standard reserving solution is shaken up with methanol constant volume to 10mL；
1.3 1,5,10,20, the preparation of 50mg/L atractylodes lactone III standard working solution: step 1.2 is taken to be prepared respectively
10,50,100,200, the 500 μ L of atractylodes lactone III standard working solution of 100mg/L shaken up with methanol constant volume to 1mL.
Be 1 by the above-mentioned concentration being prepared, 5,10,20, the atractylodes lactone I standard work of 50mg/L and concentration be 1,
5,10,20, the atractylodes lactone III standard working solution of 50mg/L, which is injected separately into Ultra Performance Liquid Chromatography instrument, measures, and obtains standard
The UPLC characteristic spectrum of Rhizoma Atractylodis Macrocephalae sample, as shown in Figure 1.
The preparation and measurement of S3, Rhizoma Atractylodis Macrocephalae sample solution to be measured:
(1) preparation of Rhizoma Atractylodis Macrocephalae sample solution: weighing 5g and be placed in 50mL triangular flask after first crushing Rhizoma Atractylodis Macrocephalae, 25mL first is added
Ultrasound 30min in ultrasonoscope is placed in after alcohol, control revolving speed is that 3000r/min is centrifuged 10min, finally with 0.45 μm of filter membrane
Filtering；
(2) it will be measured in the above-mentioned Rhizoma Atractylodis Macrocephalae sample solution injection Ultra Performance Liquid Chromatography instrument being prepared, obtain Rhizoma Atractylodis Macrocephalae sample
The UPLC characteristic spectrum of product solution, as shown in Figure 2.
The content of atractylodes lactone is calculated as follows in the Rhizoma Atractylodis Macrocephalae sample to be measured:
In formula:
The content of atractylodes lactone, unit mg/g in X-Rhizoma Atractylodis Macrocephalae sample to be measured；
Cs- from the content for obtaining atractylodes lactone in Rhizoma Atractylodis Macrocephalae sample to be measured measurement liquid in standard curve, unit mg/g；
miThe quality of-sample to be tested, unit g；
V-Rhizoma Atractylodis Macrocephalae sample to be measured measures liquid constant volume, unit L.
By calculating, show that the content of atractylodes lactone I in Rhizoma Atractylodis Macrocephalae sample to be measured is 0.0101%, atractylodes lactone III's contains
Amount is 0.0087%.
Precision Experiment:
In order to verify detection method provided in an embodiment of the present invention to atractylodes lactone I in Rhizoma Atractylodis Macrocephalae sample and atractylodes lactone III
The precision of content detection, a Rhizoma Atractylodis Macrocephalae sample take 5 parts, by progress ultra high efficiency liquid phase color after above-mentioned setting condition and method processing
The reproducibility of spectrum analysis, retention time and content to atractylodes lactone I and atractylodes lactone III is analyzed, as a result such as 1 institute of table
Show.
1 atractylodes lactone I of table and atractylodes lactone III precision test (n=5)
Recovery test
It carries out preparing sample extracting solution by above-mentioned pre-treating method, takes two parts respectively from same sample extracting solution, respectively
1mL, a copy of it are settled to 10mL with methanol solution, and the atractylodes lactone I and Rhizoma Atractylodis Macrocephalae that 2mL concentration is 0.1mg/mL is added in another
Lactone II I standard solution, is equally settled to 10mL.Liquid-phase chromatographic analysis is carried out to solution before and after mark-on, according to molten before and after mark-on
The percentage of the variation of liquid concentration and scalar quantity calculates the rate of recovery of each ingredient, is measured in parallel 6 times, the results are shown in Table 2.
The rate of recovery of atractylodes lactone I and atractylodes lactone III in 2 sample of table
Above embodiments only express specific embodiment of the present invention, and the description thereof is more specific and detailed, but can not
Therefore limitations on the scope of the patent of the present invention are interpreted as.It should be pointed out that for those of ordinary skill in the art,
Without departing from the inventive concept of the premise, various modifications and improvements can be made, these are all to 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 (5)
1. a kind of UPLC method of atractylenolide Ⅰ and atractylodes lactone III in detection Rhizoma Atractylodis Macrocephalae drug, which is characterized in that including following step
It is rapid:
S1, Rhizoma Atractylodis Macrocephalae sample preparation to be measured: first Rhizoma Atractylodis Macrocephalae crushing is placed in triangular flask, is placed in ultrasonoscope after methanol is added
Ultrasound, centrifugation are carried out, is finally filtered；
S2, the measurement of standard Rhizoma Atractylodis Macrocephalae sample: standard Rhizoma Atractylodis Macrocephalae sample solution is injected in Ultra Performance Liquid Chromatography instrument and is measured, standard is obtained
The UPLC characteristic spectrum of Rhizoma Atractylodis Macrocephalae sample；
S3, Rhizoma Atractylodis Macrocephalae sample to be measured measurement: it will be surveyed in Rhizoma Atractylodis Macrocephalae sample to be measured in step S1 measurement liquid injection Ultra Performance Liquid Chromatography instrument
It is fixed, the UPLC characteristic spectrum of Rhizoma Atractylodis Macrocephalae sample to be measured is obtained, the UPLC characteristic spectrum of the obtained standard Rhizoma Atractylodis Macrocephalae sample of step S2 is passed through
The Rhizoma Atractylodis Macrocephalae sample to be measured being prepared is quantified with the UPLC characteristic spectrum of Rhizoma Atractylodis Macrocephalae sample to be measured.
2. the UPLC method of atractylenolide Ⅰ and atractylodes lactone III in a kind of detection Rhizoma Atractylodis Macrocephalae drug according to claim 1,
It is characterized in that, the chromatographic condition of ultra performance liquid chromatography are as follows:
Chromatographic column: BEH C18 column (2.1 μ m 100mm, 1.7mm)；
Flow velocity: 0.3mL/min；
Column temperature: 30 DEG C；
Sample volume: 5 μ L；
Ultraviolet detection wavelength: 221nm；
Mobile phase: acetonitrile and water, wherein the volume ratio of acetonitrile and water is 54:46.
3. the UPLC method of atractylenolide Ⅰ and atractylodes lactone III in a kind of detection Rhizoma Atractylodis Macrocephalae drug according to claim 1,
It is characterized in that, the content of atractylodes lactone is calculated as follows in the Rhizoma Atractylodis Macrocephalae sample to be measured:
In formula:
The content of atractylodes lactone in X-Rhizoma Atractylodis Macrocephalae sample to be measured；
Cs- from the content for obtaining atractylodes lactone in Rhizoma Atractylodis Macrocephalae sample to be measured measurement liquid in standard curve；
miThe quality of-sample to be tested；
V-Rhizoma Atractylodis Macrocephalae sample to be measured measures liquid constant volume.
4. the UPLC method of atractylenolide Ⅰ and atractylodes lactone III in a kind of detection Rhizoma Atractylodis Macrocephalae drug according to claim 1,
It is characterized in that, the method for Rhizoma Atractylodis Macrocephalae sample preparation to be measured in the step S1 specifically: weigh 5g after first crushing Rhizoma Atractylodis Macrocephalae and be placed in
In 50mL triangular flask, it is placed in ultrasound 20-45min in ultrasonoscope after 20-30mL methanol is added, control revolving speed is 2500-
3500r/min is centrifuged 10-30min, finally with 0.45 μm of membrane filtration.
5. the UPLC method of atractylenolide Ⅰ and atractylodes lactone III in a kind of detection Rhizoma Atractylodis Macrocephalae drug according to claim 1,
It is characterized in that, the step S2 Plays Rhizoma Atractylodis Macrocephalae sample solution is prepared by following steps:
(1) preparation of atractylodes lactone I standard working solution:
The preparation of 1.1 100g/L atractylodes lactone I standard reserving solutions: weighing 1.02g atractylodes lactone I standard items (content 98%), uses
Methanol constant volume shakes up to 10mL；
The preparation of 1.2 100mg/L atractylodes lactone I standard working solutions: the atractylodes lactone I for the 100g/L for taking step 1.1 to be prepared
10 μ L of standard reserving solution is shaken up with methanol constant volume to 10mL；
1.3 1,5,10,20, the preparation of 50mg/L atractylodes lactone I standard working solution: step 1.2 is taken to be prepared respectively
10,50,100,200, the 500 μ L of atractylodes lactone I standard working solution of 100mg/L is shaken up with methanol constant volume to 1mL；
(2) preparation of atractylodes lactone III standard working solution:
The preparation of 1.1 100g/L atractylodes lactone III standard reserving solutions: 1.02g atractylodes lactone III standard items (content is weighed
98%) it, with methanol constant volume to 10mL, shakes up；
The preparation of 1.2 100mg/L atractylodes lactone III standard working solutions: in the Rhizoma Atractylodis Macrocephalae for the 100g/L for taking step 1.1 to be prepared
10 μ L of ester III standard reserving solution is shaken up with methanol constant volume to 10mL；
1.3 1,5,10,20, the preparation of 50mg/L atractylodes lactone III standard working solution: step 1.2 is taken to be prepared respectively
10,50,100,200, the 500 μ L of atractylodes lactone III standard working solution of 100mg/L is shaken up with methanol constant volume to 1mL.
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| CN111679022A (en) * | 2020-07-15 | 2020-09-18 | 磐安县食品药品检验检测中心 | Method for measuring contents of effective components atractylenolide III and atractylenolide I in atractylenolide |
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Application publication date: 20191101 |