CN111679022A - Method for measuring contents of effective components atractylenolide III and atractylenolide I in atractylenolide - Google Patents
Method for measuring contents of effective components atractylenolide III and atractylenolide I in atractylenolide Download PDFInfo
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- FBMORZZOJSDNRQ-GLQYFDAESA-N Atractylenolide III Chemical compound C=C([C@@H]1C2)CCC[C@]1(C)C[C@@]1(O)C2=C(C)C(=O)O1 FBMORZZOJSDNRQ-GLQYFDAESA-N 0.000 title claims abstract description 96
- FBMORZZOJSDNRQ-UHFFFAOYSA-N Demethoxy,B,HCl-Adriamycin Natural products C1C2C(=C)CCCC2(C)CC2(O)C1=C(C)C(=O)O2 FBMORZZOJSDNRQ-UHFFFAOYSA-N 0.000 title claims abstract description 48
- OAXKIRPCKWQWOQ-UHFFFAOYSA-N atractylenolide III Natural products CC1=C2CC3C(CCCC3=C)CC2(O)OC1=O OAXKIRPCKWQWOQ-UHFFFAOYSA-N 0.000 title claims abstract description 48
- ZTVSGQPHMUYCRS-SWLSCSKDSA-N (4as,8as)-3,8a-dimethyl-5-methylidene-4a,6,7,8-tetrahydro-4h-benzo[f][1]benzofuran-2-one Chemical compound C=C([C@@H]1C2)CCC[C@]1(C)C=C1C2=C(C)C(=O)O1 ZTVSGQPHMUYCRS-SWLSCSKDSA-N 0.000 title claims abstract description 33
- OQYBLUDOOFOBPO-UHFFFAOYSA-N Asterolide Natural products C1C2C(=C)CCCC2(C)CC2C1=C(C)C(=O)O2 OQYBLUDOOFOBPO-UHFFFAOYSA-N 0.000 title claims abstract description 33
- 238000000034 method Methods 0.000 title claims abstract description 24
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims abstract description 72
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims abstract description 33
- 239000000243 solution Substances 0.000 claims abstract description 26
- 239000013558 reference substance Substances 0.000 claims abstract description 21
- 239000012088 reference solution Substances 0.000 claims abstract description 15
- 239000007788 liquid Substances 0.000 claims abstract description 14
- 239000000843 powder Substances 0.000 claims abstract description 13
- 238000005303 weighing Methods 0.000 claims abstract description 12
- 238000000967 suction filtration Methods 0.000 claims abstract description 10
- 238000010812 external standard method Methods 0.000 claims abstract description 9
- 239000004480 active ingredient Substances 0.000 claims abstract description 8
- 238000001704 evaporation Methods 0.000 claims abstract description 7
- 239000000706 filtrate Substances 0.000 claims abstract description 7
- 238000001914 filtration Methods 0.000 claims abstract description 7
- 239000012528 membrane Substances 0.000 claims abstract description 7
- 238000002791 soaking Methods 0.000 claims abstract description 7
- 238000001514 detection method Methods 0.000 claims abstract description 5
- 241000132012 Atractylodes Species 0.000 claims description 34
- 239000012085 test solution Substances 0.000 claims description 17
- 239000000047 product Substances 0.000 claims description 10
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 9
- 239000003814 drug Substances 0.000 claims description 6
- 239000012634 fragment Substances 0.000 claims description 6
- 238000000227 grinding Methods 0.000 claims description 6
- 238000005406 washing Methods 0.000 claims description 6
- 238000005520 cutting process Methods 0.000 claims description 3
- 239000000523 sample Substances 0.000 abstract description 13
- 239000012488 sample solution Substances 0.000 abstract description 7
- 238000011084 recovery Methods 0.000 abstract description 5
- 238000011156 evaluation Methods 0.000 abstract description 2
- 238000005259 measurement Methods 0.000 abstract description 2
- 238000004090 dissolution Methods 0.000 abstract 1
- 238000010298 pulverizing process Methods 0.000 abstract 1
- 238000012360 testing method Methods 0.000 description 11
- 150000002596 lactones Chemical class 0.000 description 8
- 241000092665 Atractylodes macrocephala Species 0.000 description 6
- 238000004128 high performance liquid chromatography Methods 0.000 description 6
- 238000012417 linear regression Methods 0.000 description 6
- 238000002360 preparation method Methods 0.000 description 5
- 238000002347 injection Methods 0.000 description 3
- 239000007924 injection Substances 0.000 description 3
- 238000013112 stability test Methods 0.000 description 3
- TYPSVDGIQAOBAD-DZGCQCFKSA-N Atractylone Chemical compound C([C@]1(C)C2)CCC(=C)[C@@H]1CC1=C2OC=C1C TYPSVDGIQAOBAD-DZGCQCFKSA-N 0.000 description 2
- 239000013642 negative control Substances 0.000 description 2
- 206010000234 Abortion spontaneous Diseases 0.000 description 1
- 206010003445 Ascites Diseases 0.000 description 1
- 208000008035 Back Pain Diseases 0.000 description 1
- 206010016654 Fibrosis Diseases 0.000 description 1
- 208000027530 Meniere disease Diseases 0.000 description 1
- 208000004880 Polyuria Diseases 0.000 description 1
- 208000037063 Thinness Diseases 0.000 description 1
- 241000234314 Zingiber Species 0.000 description 1
- 235000006886 Zingiber officinale Nutrition 0.000 description 1
- 208000012876 acute enteritis Diseases 0.000 description 1
- TYPSVDGIQAOBAD-UHFFFAOYSA-N atractylone Natural products C1C2(C)CCCC(=C)C2CC2=C1OC=C2C TYPSVDGIQAOBAD-UHFFFAOYSA-N 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000007882 cirrhosis Effects 0.000 description 1
- 208000019425 cirrhosis of liver Diseases 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 239000013068 control sample Substances 0.000 description 1
- 230000035619 diuresis Effects 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 235000008397 ginger Nutrition 0.000 description 1
- 201000002364 leukopenia Diseases 0.000 description 1
- 231100001022 leukopenia Toxicity 0.000 description 1
- 201000007270 liver cancer Diseases 0.000 description 1
- 238000004949 mass spectrometry Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 208000015994 miscarriage Diseases 0.000 description 1
- 239000011259 mixed solution Substances 0.000 description 1
- 239000003921 oil Substances 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 238000003908 quality control method Methods 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 210000000952 spleen Anatomy 0.000 description 1
- 208000000995 spontaneous abortion Diseases 0.000 description 1
- 210000002784 stomach Anatomy 0.000 description 1
- 238000005728 strengthening Methods 0.000 description 1
- 230000035900 sweating Effects 0.000 description 1
- 206010048828 underweight Diseases 0.000 description 1
- 239000000341 volatile oil Substances 0.000 description 1
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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
- G01N30/62—Detectors specially adapted therefor
- G01N30/74—Optical detectors
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- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
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Abstract
The invention relates to the technical field of detection, in particular to a method for measuring contents of active ingredients, namely atractylenolide III and atractylenolide I in atractylenolide. Which comprises the following steps: pulverizing Atractylodis rhizoma into powder, and soaking in ethyl acetate; after suction filtration, evaporating to dryness, adding methanol for dissolution and fixing the volume; filtering with filter membrane to obtain filtrate; precisely weighing atractylenolide III and atractylenolide I reference substances; adding methanol to obtain solution containing 0.1mg per 1 ml; precisely sucking 10 μ L of each of the reference solution and the sample solution, and respectively injecting into a liquid chromatograph for determination; recording chromatogram, and calculating contents of atractylenolide III and atractylenolide I by external standard method. The method is designed to carry out measurement through a liquid chromatograph, record a chromatogram, calculate the contents of the atractylenolide III and the atractylenolide I by an external standard method, is simple and rapid to operate, and has good sample precision, reproducibility, stability and recovery rate by measuring peak area under set chromatographic conditions, thereby meeting evaluation standards.
Description
Technical Field
The invention relates to the technical field of detection, in particular to a method for measuring contents of active ingredients, namely atractylenolide III and atractylenolide I in atractylenolide.
Background
The bighead atractylodes rhizome is a common and important bulk Chinese medicinal material and has the functions of tonifying spleen and strengthening stomach, eliminating dampness and promoting diuresis, stopping sweating and preventing miscarriage and the like. The rhizome of Atractylodes macrocephala contains volatile oil, the main components of the oil are atractylone, atractylol, atractylenolide, etc., and the drug has certain curative effect on ascites due to cirrhosis, primary liver cancer, Meniere's syndrome, chronic lumbago, acute enteritis, leukopenia, etc. The rhizoma atractylodis macrocephalae has wide application, and is an important raw material of more than 40 Chinese patent medicine preparations besides the medicine used in the medical formula.
The atractylenolide is a compound in the atractylenolide, and is a main lactone component in the atractylenolide together with atractylenolide III and atractylenolide I, at present, the method for determining atractylenolide in the atractylenolide mostly adopts CC, HPLC and mass spectrometry to determine one or more active ingredients, and a unified standard housin can not be formed until now in relation to the quality control of atractylenolide, so that the establishment of a stable, controllable, accurate and efficient method for determining the contents of atractylenolide III and atractylenolide I has important significance.
Disclosure of Invention
The invention aims to provide a method for measuring the content of active ingredients atractylenolide III and atractylenolide I in atractylenolide, so as to solve the problems in the background technology.
In order to achieve the purpose, the invention provides a method for measuring the content of active ingredients, namely atractylenolide III and atractylenolide I in atractylenolide, which comprises the following steps:
preparing a test solution:
s1.1, taking the bighead atractylodes rhizome, grinding into powder, and then adding ethyl acetate for soaking;
s1.2, carrying out suction filtration, evaporating to dryness, adding methanol for dissolving, and fixing the volume;
s1.3, filtering with a filter membrane, and taking a subsequent filtrate to obtain the product;
(II) preparing a reference solution:
s1.4, precisely weighing atractylenolide III and I reference substances;
s1.5, adding methanol to prepare a solution containing 0.1mg of methanol per 1ml to obtain the product;
(III) measuring:
s1.6, precisely sucking 10 mu L of reference solution and test solution respectively, and injecting the reference solution and the test solution into a liquid chromatograph for determination respectively;
s1.7, recording a chromatogram, and calculating the contents of the atractylenolide III and the atractylenolide I by an external standard method to obtain the medicine.
As a further improvement of the technical solution, the chromatographic conditions of the liquid chromatograph are as follows:
a chromatographic column: a C18 column (150 mm. times.4.6);
column temperature: 35 ℃;
mobile phase: methanol and water, the ratio of methanol to water being 72: 28;
flow rate: 1.0 ml/min;
detection wavelength: 220 nm;
sample introduction amount: 10 μ L.
As a further improvement of the technical scheme, the method for preparing the test solution comprises the following steps:
s2.1, taking the bighead atractylodes rhizome, cutting the bighead atractylodes rhizome into fragments by using a knife, grinding the fragments into powder by using a grinder, taking 18-22g of the powder, and precisely weighing the powder;
s2.2, placing the weighed white atractylodes rhizome into a 250ml conical flask with a plug, adding 100ml of ethyl acetate, soaking for 72 hours, and performing suction filtration to leave a white atractylodes rhizome solution;
s2.3, washing the filter residue after suction filtration by using ethyl acetate, and combining the washing liquid into the bighead atractylodes rhizome solution;
s2.4, evaporating the bighead atractylodes rhizome solution to dryness by using a rotary evaporator in a water bath at 40 ℃;
s2.5, adding methanol to dissolve and fixing the volume to 50 ml;
s2.6, filtering with a 0.45-micron filter membrane, and taking a subsequent filtrate to obtain the product.
As a further improvement of the present technical solution, the method for preparing the reference solution comprises the following steps:
s3.1, precisely weighing 5.4mg and 5.0mg of atractylenolide III and I reference substances, and placing the reference substances in a 5ml measuring flask;
s3.2, adding methanol to dissolve and dilute to a scale, and shaking up;
and S3.3, obtaining solutions containing 1.08mg/ml and 1.00mg/ml of atractylenolide III and I respectively.
Compared with the prior art, the invention has the beneficial effects that:
1. in the method for measuring the contents of the effective components atractylenolide III and atractylenolide I in the atractylenolide, a liquid chromatograph is used for measuring, a chromatogram is recorded, the contents of atractylenolide III and atractylenolide I are calculated by an external standard method, and the operation is simple, convenient and quick.
2. In the method for measuring the content of the active ingredients atractylenolide III and atractylenolide I in the atractylenolide, the concentration of a reference substance is a horizontal coordinate, the peak area is a vertical coordinate, a standard curve is drawn to obtain a linear regression equation, and the linear relation of the concentrations of atractylenolide III and atractylenolide I can be obtained.
3. According to the method for measuring the content of the active ingredients, namely the atractylenolide III and the atractylenolide I in the atractylenolide, the peak area is measured under the set chromatographic condition, and the sample precision, reproducibility, stability and recovery rate are all good, so that the evaluation standard is met.
Drawings
FIG. 1 is a linear regression plot of a control, atractylenolide III, provided by practice of the invention;
FIG. 2 is a second linear regression plot of a atractylenolide III control sample provided in accordance with the practice of the invention;
FIG. 3 is a linear regression plot of a comparison product of atractylenolide I provided in the practice of the invention;
FIG. 4 is a two-stage linear regression graph of atractylenolide I control provided by practice of the invention.
FIG. 5 is an HPLC chromatogram of atractylenolide in a standard Atractylodis rhizoma sample provided by the practice of the invention.
FIG. 6 is an HPLC chromatogram of atractylenolide in a sample of Atractylodis rhizoma to be tested, provided by the practice of the present invention;
FIG. 7 shows a negative control HPLC chromatogram of atractylenolide III and atractylenolide I provided by the practice of the invention.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Referring to fig. 1-4, the present invention provides a method for determining the content of atractylenolide iii and i, which are effective components in atractylodes macrocephala koidz, comprising the following steps:
preparing a test solution:
s1.1, taking the bighead atractylodes rhizome, grinding into powder, and then adding ethyl acetate for soaking;
s1.2, carrying out suction filtration, evaporating to dryness, adding methanol for dissolving, and fixing the volume;
s1.3, filtering with a filter membrane, and taking a subsequent filtrate to obtain the product;
(II) preparing a reference solution:
s1.4, precisely weighing atractylenolide III and I reference substances;
s1.5, adding methanol to prepare a solution containing 0.1mg of methanol per 1ml to obtain the product;
(III) measuring:
s1.6, precisely sucking 10 mu L of reference solution and test solution respectively, and injecting the reference solution and the test solution into a liquid chromatograph for determination respectively;
s1.7, recording a chromatogram, and calculating the contents of the atractylenolide III and the atractylenolide I by an external standard method to obtain the medicine.
In this embodiment, the liquid chromatograph uses a UV detector, and the chromatographic conditions of the liquid chromatograph are as follows:
a chromatographic column: a C18 column (150 mm. times.4.6);
column temperature: 35 ℃;
mobile phase: methanol and water, the ratio of methanol to water being 72: 28;
flow rate: 1.0 ml/min;
detection wavelength: 220 nm;
sample introduction amount: 10 mu L of the solution;
the theoretical plate number is not less than 2000 according to atractylenolide III and atractylenolide I.
Further, the method for preparing the test solution comprises the following steps:
s2.1, taking the bighead atractylodes rhizome, cutting the bighead atractylodes rhizome into fragments by using a knife, grinding the fragments into powder by using a grinder, taking 18-22g of the powder, and precisely weighing the powder;
s2.2, placing the weighed white atractylodes rhizome into a 250ml conical flask with a plug, adding 100ml of ethyl acetate, soaking for 72 hours, and performing suction filtration to leave a white atractylodes rhizome solution;
s2.3, washing the filter residue after suction filtration by using ethyl acetate, and combining the washing liquid into the bighead atractylodes rhizome solution;
s2.4, evaporating the bighead atractylodes rhizome solution to dryness by using a rotary evaporator in a water bath at 40 ℃;
s2.5, adding methanol to dissolve and fixing the volume to 50 ml;
s2.6, filtering with a 0.45-micron filter membrane, and taking a subsequent filtrate to obtain the product.
Specifically, the method for preparing the reference solution comprises the following steps:
s3.1, precisely weighing 5.4mg and 5.0mg of atractylenolide III and I reference substances, and placing the reference substances in a 5ml measuring flask;
s3.2, adding methanol to dissolve and dilute to a scale, and shaking up;
and S3.3, obtaining solutions containing 1.08mg/ml and 1.00mg/ml of atractylenolide III and I respectively.
Preparation of control solutions and linear relationship experiments:
precisely weighing 5.4mg of atractylenolide III and I as reference substances and 5.0mg of atractylenolide III and I as reference substances, placing into a 5ml measuring flask, adding methanol to dissolve and dilute to scale, and shaking up to obtain solutions containing atractylenolide III and I of 1.08mg/ml and 1.00mg/ml respectively. Respectively taking appropriate amount of the above control solution to prepare atractylenolide III at 0.027mg/ml, 0.054mg/ml, 0.108mg/ml, 0.27mg/ml and 0.54 mg/ml; the mixed solution of atractylenolide I at 0.025mg/ml, 0.050mg/ml, 0.100mg/ml, 0.25mg/ml and 0.50mg/ml was injected into HPLC at 10. mu.L, and the peak area was measured, and the results are shown in the following table:
the relation between the concentration of epi-atractylenolide III reference substance and peak area
Relationship between concentration and peak area of epibiatractyloide I reference substance
Taking the concentration (mg/ml) of the atractylenolide III and I reference substances as abscissa and the peak area as ordinate, drawing a standard curve to obtain linear regression equations Y-4.02455 e +007X +0, Y-4.03779 e +007X +0 and a correlation coefficient r. The results show that the atractylenolide III has good linear relation in the concentration range of 0.027mg/ml to 0.54mg/ml, and the atractylenolide I has good linear relation in the concentration range of 0.025mg/ml to 0.50 mg/ml.
Negative test:
taking 100ml ethyl acetate, preparing according to the method for preparing the test solution, injecting sample according to the chromatographic conditions, and recording a chromatogram, wherein the result shows that no chromatographic peaks of atractylenolide III and atractylenolide I are found in the negative solution, and the negative solution is free of interference. The HPLC chromatogram of the atractylenolide III and atractylenolide I negative control is shown in FIG. 7.
And (3) precision test:
taking a reference solution with the concentration of about 0.1mg/ml and sample solutions of bighead atractylodes rhizome 3 and 51 batches of bighead atractylodes rhizome to be tested, repeating sample injection for 5 times and 6 times respectively according to the chromatographic conditions, and measuring peak areas, wherein the results are shown in the following tables three and four.
Precision test result for determining content of epiatractylenolide III
Determination precision test result of content of epi-atractylenolide I
As can be seen from tables III and IV, the RSD of the reference and test solutions was low, and the results indicated that the precision was good.
And (3) repeatability test:
taking the same two batches of samples (2 and 8 batches of largehead atractylodes rhizome), preparing 6 parts in parallel according to the preparation method of the test solution, precisely measuring 10ul of the sample solution, respectively injecting the sample solution into a high performance liquid chromatograph, measuring the peak area, and calculating the contents of largehead atractylodes rhizome lactone III and largehead atractylodes rhizome lactone I by using an external standard method, wherein the result is shown in the fifth table:
reproducibility test result for determining content of epibola atractylodis lactone III
| Numbering | 1 | 2 | 3 | 4 | 5 | 6 | RSD(％) |
| Atractylodes macrocephala Koidz 2 | 0.28 | 0.28 | 0.28 | 0.28 | 0.28 | 0.29 | 1.4 |
| Atractylodes macrocephala 8 | 0.22 | 0.22 | 0.22 | 0.22 | 0.22 | 0.22 | 0 |
Reproducibility test result for determining content of epihexa-atractylenolide I
| Numbering | 1 | 2 | 3 | 4 | 5 | 6 | RSD(％) |
| Atractylodes macrocephala Koidz 2 | 0.69 | 0.67 | 0.70 | 0.69 | 0.70 | 0.70 | 1.7 |
| Atractylodes macrocephala 8 | 0.77 | 0.77 | 0.77 | 0.77 | 0.77 | 0.77 | 0 |
And (3) stability test:
taking the reference solution with the concentration of 0.108mg/ml and 0.100mg/ml and the sample solution of 51 batches of the largehead atractylodes rhizome, and respectively measuring the peak areas of the largehead atractylodes rhizome lactone III and the largehead atractylodes rhizome lactone I in 0,12, 24, 48 and 72 hours according to the chromatographic conditions, wherein the results are shown in tables seven and eight:
stability test result of content determination of epi-hepta-atractylenolide III
Stability test result of content determination of epiocta-atractylenolide I
The results show that the RSD of the reference substance and the test solution are respectively shown in the seven and eight tables, so that the reference substance and the test solution are stable within 72 hours.
Recovery rate test:
precisely weighing 6 parts of about 5g of ginger sample with known content (largehead atractylodes rhizome 51, the average content of atractylenolide III is 0.43g/kg, and the content of atractylenolide I is 0.76g/kg), respectively adding 1.95mg of atractylenolide III reference substance and 1.75mg of atractylenolide I reference substance, treating the sample with the preparation method of a sample solution, carrying out sample injection according to the chromatographic conditions to determine peak areas, and calculating the contents of atractylenolide III and I by an external standard method, wherein the results are shown in tables nine and ten:
content determination of epi-nona-atractyloide III recovery test results (n ═ 6)
TABLE TEN-LARGE CACTOBACTONE I CONTENT DETERMINATION RECOVERY RATE TEST RESULT (n ═ 6)
And (3) sample determination:
20 batches of largehead atractylodes rhizome samples are purchased from the market, the samples are processed by the preparation method of the test sample solution, the peak area is determined by the sample injection according to the chromatographic conditions, the contents of largehead atractylodes rhizome lactones III and largehead atractylodes rhizome lactones I are calculated by an external standard method, and the result is shown in the eleventh table:
TABLE UNDERWEIGHT MEASUREMENT OF LACTOLIDE III AND I IN SAMPLES OF THE UNDERWEAR-DETABLES
The foregoing shows and describes the general principles, essential features, and advantages of the invention. It will be understood by those skilled in the art that the present invention is not limited to the embodiments described above, and the preferred embodiments of the present invention are described in the above embodiments and the description, and are not intended to limit the present invention. The scope of the invention is defined by the appended claims and equivalents thereof.
Claims (4)
1. A method for measuring the content of active ingredients atractylenolide III and atractylenolide I in atractylenolide is characterized by comprising the following steps:
preparing a test solution:
s1.1, taking the bighead atractylodes rhizome, grinding into powder, and then adding ethyl acetate for soaking;
s1.2, carrying out suction filtration, evaporating to dryness, adding methanol for dissolving, and fixing the volume;
s1.3, filtering with a filter membrane, and taking a subsequent filtrate to obtain the product;
(II) preparing a reference solution:
s1.4, precisely weighing atractylenolide III and I reference substances;
s1.5, adding methanol to prepare a solution containing 0.1mg of methanol per 1ml to obtain the product;
(III) measuring:
s1.6, precisely sucking 10 mu L of reference solution and test solution respectively, and injecting the reference solution and the test solution into a liquid chromatograph for determination respectively;
s1.7, recording a chromatogram, and calculating the contents of the atractylenolide III and the atractylenolide I by an external standard method to obtain the medicine.
2. The method for measuring the contents of the effective components atractylenolide III and atractylenolide I in the rhizoma Atractylodis Macrocephalae according to claim 1, which is characterized in that: the chromatographic conditions of the liquid chromatograph are as follows:
a chromatographic column: a C18 column (150 mm. times.4.6);
column temperature: 35 ℃;
mobile phase: methanol and water, the ratio of methanol to water being 72: 28;
flow rate: 1.0 ml/min;
detection wavelength: 220 nm;
sample introduction amount: 10 μ L.
3. The method for measuring the contents of the effective components atractylenolide III and atractylenolide I in the rhizoma Atractylodis Macrocephalae according to claim 1, which is characterized in that: the method for preparing the test solution comprises the following steps:
s2.1, taking the bighead atractylodes rhizome, cutting the bighead atractylodes rhizome into fragments by using a knife, grinding the fragments into powder by using a grinder, taking 18-22g of the powder, and precisely weighing the powder;
s2.2, placing the weighed white atractylodes rhizome into a 250ml conical flask with a plug, adding 100ml of ethyl acetate, soaking for 72 hours, and performing suction filtration to leave a white atractylodes rhizome solution;
s2.3, washing the filter residue after suction filtration by using ethyl acetate, and combining the washing liquid into the bighead atractylodes rhizome solution;
s2.4, evaporating the bighead atractylodes rhizome solution to dryness by using a rotary evaporator in a water bath at 40 ℃;
s2.5, adding methanol to dissolve and fixing the volume to 50 ml;
s2.6, filtering with a 0.45-micron filter membrane, and taking a subsequent filtrate to obtain the product.
4. The method for measuring the contents of the effective components atractylenolide III and atractylenolide I in the rhizoma Atractylodis Macrocephalae according to claim 1, which is characterized in that: the method for preparing the reference substance solution comprises the following steps:
s3.1, precisely weighing 5.4mg and 5.0mg of atractylenolide III and I reference substances, and placing the reference substances in a 5ml measuring flask;
s3.2, adding methanol to dissolve and dilute to a scale, and shaking up;
and S3.3, obtaining solutions containing 1.08mg/ml and 1.00mg/ml of atractylenolide III and I respectively.
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106668111A (en) * | 2016-12-31 | 2017-05-17 | 江西百神药业股份有限公司 | Rhizoma atractylodis macrocephalae formula granule preparation and quality standard identification detection method thereof |
| CN110398560A (en) * | 2019-08-07 | 2019-11-01 | 江西省农业科学院农产品质量安全与标准研究所 | A kind of UPLC method detecting atractylenolide Ⅰ and atractylodes lactone III in Rhizoma Atractylodis Macrocephalae drug |
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| CN106668111A (en) * | 2016-12-31 | 2017-05-17 | 江西百神药业股份有限公司 | Rhizoma atractylodis macrocephalae formula granule preparation and quality standard identification detection method thereof |
| CN110398560A (en) * | 2019-08-07 | 2019-11-01 | 江西省农业科学院农产品质量安全与标准研究所 | A kind of UPLC method detecting atractylenolide Ⅰ and atractylodes lactone III in Rhizoma Atractylodis Macrocephalae drug |
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