CN112684053B - Method for simultaneously determining content of 7 components in anti-chronic heart failure medicine compound - Google Patents
Method for simultaneously determining content of 7 components in anti-chronic heart failure medicine compound Download PDFInfo
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Abstract
The invention relates to a method for simultaneously measuring the content of 7 components in a compound anti-chronic heart failure medicine. The 7 components are gallic acid, hydroxy safflower yellow A, loganin, linalool, psoralen glycoside, isopsoralen glycoside and specnuezhenide, the method is a high performance liquid chromatography, and the chromatographic conditions are as follows: by C 18 Chromatographic column, mobile phase acetonitrile, water, gradient elution; flow rate of 0.2-1.2 mL/min ‑1 The column temperature is 20-40 ℃, and the detection wavelength is 220-360 nm. The method can realize simultaneous determination of the content of 7 components in the anti-chronic heart failure compound, has good linear relation, high precision, strong stability, good repeatability and high sample recovery rate, is simple and feasible, and provides a basis for establishing the quality standard of the anti-chronic heart failure compound.
Description
Technical Field
The invention relates to the field of quality control of a traditional Chinese medicine compound, in particular to a method for simultaneously measuring the content of 7 components in a compound medicine for resisting chronic heart failure.
Background
The compound medicine for resisting chronic heart failure, namely the prescription for tonifying kidney and activating blood, is an empirical prescription for clinically treating chronic heart failure for many years, is applied to patients with heart failure for a long time, and is clinically reliable and effective [1,11] And has been patented. The formula aims at tonifying deficiency and seeking the basis in the aspect of treating the symptoms and root causes, and gives consideration to the symptoms and the excess such as blood stasis, water retention and the like; the specific gravity of yin and yang is treated to balance yin and yang, mainly to supplement yang, and to seek yang from yin to yang; the relationship between the heart and kidney is emphasized, so that the heart and kidney are both related to each other and the heart origin and kidney are both treated to warm the kidney and strengthen the heart. The formula consists of 7 medicinal materials of antler, safflower, epimedium, malaytea scurfpea fruit, glossy privet fruit, dogwood and Chinese eaglewood, wherein the antler in the formula warms kidney, tonifies kidney yang and benefits kidneyEssence and blood; the safflower has the pungent and warm property and can enter the heart, promote blood circulation and dredge channels, and the two medicines are monarch medicines together; herba epimedii, fructus psoraleae, glossy privet fruit and dogwood help the antler to balance yin and yang of the kidney, and are ministerial medicines; lignum Aquilariae Resinatum has effects of invigorating qi, invigorating kidney, promoting qi circulation, relieving pain, promoting blood circulation of Carthami flos, and invigorating the recipe without stagnation, and is used as adjuvant and guiding drug; the medicines are combined to play the effects of tonifying the kidney and promoting blood circulation [1] . The anti-chronic heart failure medicine compound can act on myocardial fibroblasts in a paracrine mode through activating gp130/JAK2/STAT3 passages to inhibit myocardial fibrosis [2] Also, plasma RASS products can be affected [3,4] Which is considered to be a risk factor leading to myocardial fibrosis.
Modern pharmacological research shows that antler polypeptide as the antler active component can mediate cardiac muscle stem cell differentiation and repair damaged cardiac muscle [5] (ii) a The hydroxysafflor yellow A is a compound with a single chalcone glycoside structure, is the most effective water-soluble part with pharmacological effects of safflower, has obvious protection effect on vascular tissue cells, and has the effects of reducing blood pressure, dilating blood vessels, improving blood supply of organs, resisting blood coagulation, resisting inflammation and the like [6,7] (ii) a Fructus Psoraleae has various biological activities, and coumarins as main active ingredients, wherein psoralen has protective effect on cardiovascular system [10] (ii) a Herba Epimedii main ingredient is flavone, has great effects in resisting heart failure and protecting heart function, and its representative ingredient icariin can increase heart contraction function, relieve heart load, increase heart discharge, thereby protecting cardiovascular system [8,9] (ii) a Corni fructus is the main medicinal material for treating diabetes, coronary heart disease and hypertension, iridoid is the highest content classification group in Corni fructus, Kang et al [11] Research shows that gallic acid can increase NO content by up-regulating endothelial nitric oxide synthase (eNOS) phosphorylation level, and simultaneously, gallic acid can inhibit angiotensin I converting enzyme (ACE) and remarkably reduce blood pressure of spontaneous hypertension rats, and has the same pharmacological action strength as captopril. The morroniside and loganin have certain protective effect on myocardial of diabetic mice [12] (ii) a The effective components of fructus Ligustri Lucidi can regulate endocrine system of human body, increase coronary blood flow, and inhibit myocardial contractionHas good effect in treating heart diseases such as cardiac insufficiency and arrhythmia [10,13] Specnuezhenide [14] Can obviously inhibit the apoptosis of the mesangial cells, can increase Bcl-2/Bax and reduce the high expression of clean-caspase 3 protein, thereby inhibiting too early apoptosis of the mesangial cells, playing a role in protecting the cells and achieving the purposes of preventing and treating renal failure and reducing proteinuria.
The research on the quality control of the Chinese herbal compound is the key of the modernization and internationalization of the Chinese herbal medicine. Patent document CN111175392A, published japanese 2020.05.19, discloses a quality control method of cough-relieving loquat syrup, and establishes an HPLC method for measuring the content of amygdalin, cortex mori element B and neotuberostemonine. Patent document CN110780018A, published japanese 2020.02.11, discloses a quality control method of liver-nourishing granules, and establishes a content detection method of emodin and chrysophanol by using an HPLC method. Patent document CN112098563A, published japanese patent No. 2020.12.18, discloses a method for detecting chemical components of a formula for tonifying kidney and activating blood, wherein the traditional Chinese medicine components of the formula for tonifying kidney and activating blood comprise eucommia bark, fructus psoraleae, radix achyranthis bidentatae, salvia miltiorrhiza, radix clematidis and pawpaw, and are mainly used for treating lower back pain, and the detection method comprises the following steps: the method comprises the steps of sequentially carrying out water soaking, extraction, concentration, ethanol extraction, standing, centrifugal separation, ethanol and ethyl acetate extraction on the traditional Chinese medicine of the kidney-tonifying and blood-activating formula, carrying out n-butanol extraction on the obtained water phase, concentrating the obtained water phase to obtain a liquid to be detected, carrying out UPLC-MS detection under specific detection conditions, and comparing the obtained detection result with a standard chromatogram and a natural product database to obtain chemical components in the liquid to be detected. However, no quality control research of the anti-chronic heart failure medicine compound as described in the application is available at present, and no method for simultaneously measuring the contents of seven components of gallic acid, hydroxysafflor yellow A, loganin, linalool, psoralen, isopsoralen and specnuezhenide in the anti-chronic heart failure medicine compound is available.
Disclosure of Invention
The invention aims to provide a method for simultaneously measuring the content of 7 components in a compound anti-chronic heart failure medicine, aiming at the defects in the prior art.
The invention also aims to provide a quality control method of the anti-chronic heart failure compound medicine.
In order to achieve the first object, the invention adopts the technical scheme that:
a high performance liquid chromatography method for simultaneously determining the content of 7 components in an anti-chronic heart failure medicine compound is disclosed, wherein the 7 components comprise gallic acid, hydroxy safflower yellow A, loganin, agaroterol, psoralen glycoside, isobavarin and specnuezhenide, and the chromatographic conditions of the high performance liquid chromatography method are as follows: by C 18 A chromatographic column, wherein a mobile phase acetonitrile-0.1% phosphoric acid water is adopted, the elution gradient is 0-10 min, and the elution gradient is 5% acetonitrile; 10-15 min,5% -10% acetonitrile; 15-35 min,10% -18% acetonitrile; 35-40 min,18% -25% acetonitrile; 40-60 min, 25-35% acetonitrile; flow rate of 0.8-1.2 mL/min -1 The column temperature is 28-32 ℃, and the detection wavelength is 220-360 nm.
As a preferred embodiment of the invention, the anti-chronic heart failure compound traditional Chinese medicine comprises the following components: 10-20 parts of antler, 6-12 parts of safflower, 15-25 parts of fructus psoraleae, 25-35 parts of epimedium herb, 10-20 parts of cornus officinalis, 25-35 parts of glossy privet fruit and 4-8 parts of agilawood.
As another preferred embodiment of the present invention, the method for preparing the test solution by the high performance liquid chromatography method comprises: precisely weighing the above materials, adding 8-15 times of water, decocting for 60-90min, filtering, concentrating the filtrate under reduced pressure, transferring to volumetric flask, metering volume, shaking, filtering with microporous membrane, and collecting the filtrate.
As another preferred embodiment of the invention, the method for preparing the mixed reference solution by the high performance liquid chromatography comprises the following steps: accurately weighing gallic acid, hydroxy safflower yellow A, loganin, linalool, psoralen, isopsoralen, and specnuezhenide, and adding methanol to obtain mixed reference solutions with concentrations of 2.71, 8.61, 7.61, 643, 17.42, 11.88, and 45.00 μ g/mL.
In another preferred embodiment of the present invention, the amount of sample is 1 to 12. mu.L.
More preferably, the sample size is 10. mu.L.
As another preferred embodiment of the present invention, the flow rate is 1.0 mL-min -1 。
As another preferred embodiment of the present invention, the column temperature is 30 ℃.
As another preferred embodiment of the present invention, the detection wavelength is 220 to 240 nm.
In order to achieve the second purpose, the invention adopts the technical scheme that:
a quality control method of a compound medicine for resisting chronic heart failure comprises the steps of the high performance liquid chromatography method.
The invention has the advantages that:
1. the invention establishes a method for measuring the contents of seven components of gallic acid, hydroxy safflower yellow A, loganin, agaroterol, psoralen, isobavarin and specnuezhenide in the anti-chronic heart failure medicine compound, and performs methodology verification of system adaptability, specificity, linearity, precision, repeatability and stability by a detection method of seven indexes to determine the feasibility of the seven component detection methods.
2. The parameters related to the detection method are obtained on the basis of research experience of the inventor, and the excellent detection effect is realized under the detection condition.
3. The invention provides a basis for establishing the compound quality standard of the anti-chronic heart failure medicine.
Drawings
FIG. 1: the control HPLC chart (A) and the test HPLC chart (B) are mixed. 1. Gallic acid; 2. hydroxysafflor yellow a; 3. loganin; 4. linalool; 5. psoralen glycosides; 6. isobavachin; 7. specnuezhenide.
FIG. 2: chromatograms under different mobile phases under the same chromatographic conditions. A: 0.1% formic acid- (B) methanol system; b: 0.1% formic acid-acetonitrile system.
FIG. 3: chromatograms at different elution gradients. A: the elution gradient is 0-8 min, and 5% of acetonitrile; 8-60 min,5% -50% acetonitrile; recording time is 0-60 min; b: the elution procedure is that the mobile phase is acetonitrile-0.1% phosphoric acid water, the elution gradient is 0-5 min, and the elution gradient is 5% acetonitrile; 5-15 min,5% -15% acetonitrile; 15-60 min, 15% -45% acetonitrile; recording time is 0-60 min; c: the elution procedure is that the mobile phase is acetonitrile-0.1% phosphoric acid water, the elution gradient is 0-5 min, and the elution gradient is 5% acetonitrile; 5-15 min,5% -15% acetonitrile; 15-41 min, 15% -27% acetonitrile; 41-43 min, 27% -30% acetonitrile; 43-60 min, 30-40% acetonitrile; 60-61 min, 40% -5% acetonitrile; recording time is 0-60 min; d: the elution procedure is that the mobile phase is acetonitrile-0.1% phosphoric acid water, the elution gradient is 0-10 min, and the elution gradient is 5% acetonitrile; 10-15 min,5% -10% acetonitrile; 15-35 min,10% -18% acetonitrile; 35-40 min,18% -25% acetonitrile; 40-60 min, 25-35% acetonitrile; the recording time is 0-60 min.
FIG. 4: chromatograms of the anti-chronic heart failure medicine compound under different wavelengths. A.211nm; b.230nm; c.254nm; d.265nm; e.280nm; f.300nm; g.330nm.
Detailed Description
The following detailed description of the present invention will be made with reference to the accompanying drawings.
Example 1
1 materials and methods
1.1 materials and instruments
An Agilent 1100series model high performance liquid chromatograph (Agilent DAD detector, Agilent technologies, inc.); BSA124S-CW electronic analytical balance (parts per million), BT25S electronic analytical balance (parts per hundred thousand, sedolis scientific instruments (beijing) ltd). Hydroxysafflor yellow A (batch No. AF9081601, content 98%), loganin (batch No. AF7110703, content 98%), psoralen (batch No. AF9102107, content 98%), isopsoralen (batch No. AF9102108, content 98%) were purchased from Shanghai Hongheng Yongsheng Biotech Co., Ltd; linalool (lot number DST190801-042, content 99%), gallic acid (lot number DST190715-008, content 99%) were purchased from chengdale pharmaceutical science and technology limited; a control of specnuezhenide (batch No. 2018011201, 95% content) was purchased from Doriplas Biotech, Inc. Cornu cervi, safflower, fructus psoraleae, herba epimedii, dogwood, glossy privet fruit and agilawood decoction pieces are purchased from Shanghai Kangqiao traditional Chinese medicine decoction piece Limited company and are identified as certified products by Wanghong researcher of Shanghai traditional Chinese medicine standardization research center. Acetonitrile (Fisher corporation, USA); formic acid (batch No. G0960200, content 98%) was purchased from Shanghai' an Spectroscopy scientific Instrument Co., Ltd; other reagents are analytically pure; the water is ultrapure water.
1.2 test methods and results
1.2.1 chromatographic conditions
The chromatographic column is ZORBAX SB-Aq (4.6mm multiplied by 250mm,5 mu m), the mobile phase is acetonitrile-0.1% phosphoric acid water, and the elution gradient is 0-10 min and 5% acetonitrile; 10-15 min,5% -10% acetonitrile; 15-35 min,10% -18% acetonitrile; 35-40 min,18% -25% acetonitrile; 40-60 min, 25-35% acetonitrile; flow rate 1.0 mL/min -1 The column temperature is 30 ℃, the detection wavelength is 230nm, 10 mu L of sample is injected, and the HPLC chromatogram of the mixed reference solution and the test solution is shown in figure 1.
1.2.2 preparation of Mixed control solutions
Accurately weighing appropriate amount of gallic acid, hydroxy safflower yellow A, loganin, linalool, psoralen glycoside, isobavarin, and specnuezhenide, and adding methanol to obtain mixed reference solutions with concentrations of 2.71, 8.61, 7.61, 643, 17.42, 11.88, and 45.00 μ g/mL.
1.2.3 preparation of test solutions
Precisely weighing appropriate amount of each medicinal material according to prescription amount (15 g of antler, 9g of safflower, 20g of fructus psoraleae, 30g of epimedium herb, 15g of cornus officinalis, 30g of glossy privet fruit and 6g of agilawood), adding 12 times of water, decocting for 75min, filtering, concentrating the filtrate under reduced pressure, transferring to a 500mL measuring flask, fixing the volume, shaking uniformly, filtering with a 0.45-micron microporous membrane, and taking the subsequent filtrate to obtain the traditional Chinese medicine.
1.2.4 Linear relationship investigation
Mixing the reference solution under the term of 1.2.2, diluting, shaking, and injecting 10 μ L of the sample under the chromatographic condition of the term of 1.2.1. The solution mass concentration was used as the abscissa (X) and the peak area integral value was used as the ordinate (Y) to conduct regression, and a standard curve was drawn, and the results are shown in Table 1.
TABLE 1 Linear relationship examination result of 7 components in anti-chronic heart failure drug compound
1.2.5 precision test
Sampling control solutions with high (H), medium (M) and low (L) mass concentrations for 6 times under the condition of 1.2.1 chromatography in 1d, and continuously measuring for 3 days, wherein the measured peak areas RSD of the components indicate that the precision of the instrument is good in each day, and the results are shown in Table 2.
TABLE 2 precision investigation results of 7 components in anti-chronic heart failure drug compound
1.2.6 repeatability tests
The method for preparing the test solution according to the item '1.2.3' is characterized in that 6 parts of the test solution are paralleled, and the content of the gallic acid, the hydroxysafflor yellow A, the loganin, the agaroterol, the psoralen glycoside, the isopsoralen glycoside and the specnuezhenide RSD are respectively measured to be 2.04%, 2.78%, 2.51%, 2.78%, 0.75%, 2.49% and 0.96% under the condition of 1.2.1 chromatographic samples, which indicates that the method has good repeatability and meets the test requirements.
1.2.7 stability test
The sample solution is prepared according to the method under the item '1.2.3', the peak areas RSD of the gallic acid, the hydroxysafflor yellow A, the loganin, the agaroterol, the psoralen glycoside, the isobavarin and the specnuezhenide are respectively 1.6%, 2.7%, 2.5%, 2.9%, 0.9%, 1.8% and 1.3% by sample injection measurement under the chromatographic condition of 1.2.1 in 0, 1, 4, 6, 12, 16 and 24 hours, which indicates that the solution has good stability in 24 hours.
1.2.8 sample recovery test
Precisely measuring 5mL of a test solution with a known content under the item '1.2.3', placing the test solution in a 10mL volumetric flask, adding reference solution with the levels of 50%, 100% and 150%, carrying out sample injection measurement under the chromatographic condition of the item 1.2.1, and calculating the recovery rate. As a result, the average sample recovery rates of gallic acid, hydroxysafflor yellow A, loganin, agaroterol, psoralen, isopsoralen and specnuezhenide are 102.58%, 98.85%, 100.17%, 98.87%, 98.61%, 97.32% and 98.53%, and the RSD are 3.72%, 3.08%, 3.92%, 2.02%, 1.21%, 2.02% and 1.14%, respectively. The results are shown in Table 3.
TABLE 3 sample application and recovery rate test for content determination of index components in anti-chronic heart failure medicine compound recipe
1.2.9 sample determination
The test solution was prepared according to the method of item 1.2.3, and the sample injection was measured under the chromatographic conditions of item 1.2.1, and the results are shown in Table 4.
TABLE 4 determination of contents of 7 index components in anti-chronic heart failure medicine compound (n ═ 3)
Discussion of 2
(1) Chronic Heart failure can be divided into 4 stages A, B, C, D by severity [15] Wherein, the treatment of the traditional Chinese medicine in the B stage still belongs to the market blank stage, and the clinical application and animal experiments prove that the anti-chronic heart failure medicine compound can not only improve symptoms, improve the life quality,it also delays the development of myocardial remodeling. The formula consists of 7 medicinal materials of antler, safflower, epimedium, malaytea scurfpea fruit, glossy privet fruit, dogwood and Chinese eaglewood, wherein the antler in the formula warms the kidney to tonify the kidney yang and benefit the essence and blood; the safflower is pungent and warm in property and enters the heart, and promotes blood circulation and clears the channels, and the two medicines are monarch medicines together; herba epimedii, fructus psoraleae, glossy privet fruit and dogwood help the antler to balance yin and yang of the kidney, and are ministerial medicines; chenxiang is used as adjuvant and guiding drug for receiving qi and entering kidney to help tonify kidney, promoting qi circulation and relieving pain to help safflower to activate blood, and to make the recipe invigorate without stagnation.
(2) According to the 2015 edition of Chinese pharmacopoeia, hydroxysafflorin A, psoralen, isopsoralen, icariin, loganin, morroniside and specnuezhenide are all quality control indexes of corresponding medicinal materials; the linalol is used as main effective component of linalol, and the content of linalol is first determined as quality control index in 2015 edition of Chinese pharmacopoeia [16] The agilawood has a treatment effect on arrhythmia and coronary heart disease, and quantitative analysis taking the content measurement of the agarotetrol as an index component is an important component for establishing an effective quality evaluation system of the agilawood. According to the previous experimental results, the psoralen, isopsoralen, icariin and morroniside have low content in the compound decoction of the anti-chronic heart failure medicine, and are not suitable to be used as the quality control indexes of the compound, and the psoralen and isopsoralen can be converted into psoralen and isopsoralen in vivo [17] And the content of the extract in the water decoction is higher, so that gallic acid, hydroxy safflor yellow A, loganin, agaroterol, psoralen glycoside, isopsoralen glycoside and specnuezhenide are selected as the quality control indexes of the compound anti-chronic heart failure medicine.
(3) Selection of mobile phase: through comparison and screening, the 0.1% formic acid-acetonitrile system is eluted by linear gradient, the baseline of the obtained liquid chromatogram is relatively stable, the peak shape of the chromatogram is symmetrical, and the separation degree is relatively good, so the 0.1% formic acid-acetonitrile system is selected as a mobile phase, and the comparison result is shown in figure 2.
(4) Exploration of mobile phase elution gradient: through systematic exploration and optimization, the following linear gradient elution procedures are found to be capable of comprehensively detecting chemical components in the preparation and achieving ideal separation effects of the components: the elution procedure is determined to be that the mobile phase is acetonitrile-0.1% phosphoric acid water, the elution gradient is 0-10 min, and the elution gradient is 5% acetonitrile; 10-15 min,5% -10% acetonitrile; 15-35 min,10% -18% acetonitrile; 35-40 min,18% -25% acetonitrile; 40-60 min,25% -35% acetonitrile, recording time of 0-60 min, and comparison results are shown in figure 3.
(5) Selection of chromatographic wavelength: through comparative analysis of a liquid chromatography-ultraviolet spectrum three-dimensional map, uneven base line and poor separation degree under 211nm and 330nm are found; the baseline is relatively flat in preparation spectra of 254nm, 265nm, 280nm and 300nm, the peak shape is clear, the number of the peaks is small, the components of the preparation cannot be comprehensively reflected, the baseline is relatively flat under 230nm, besides the known components of gallic acid, hydroxysafflor yellow A, loganin, linalool, psoralen, isopsoralen, specnuezhenide, quercetin and the like can be well detected, the fingerprint peak separation degree of other components is also good, the corresponding strength of the number, the baseline and signals of chromatographic peaks is comprehensively considered, 230nm is determined as the optimal detection wavelength of the preparation fingerprint spectrum, the wavelength is more than the number of peaks under other wavelengths, the response and the separation degree of each component are good, and the comparison result is shown in figure 4.
Reference to the literature
[1] Xujie, dianhuiyan, Jingfeng, deer red side clinical study on chronic heart failure with myocardial fibrosis [ J ]. Shanghai J.J. Med.2017, 51(S1):48-51.
[2]QU H，WANG Y，WANG Y，et al.Luhong formula inhibits myocardial fibrosis in a paracrine manner by activating the gp130/JAK2/STAT3 pathway in cardiomyocytes[J].J Ethnopharmacol，2017，202(56):28-37.
[3] Xujie, Dian Hui Yan, Jing Feng, deer Red granule has effects on the hemodynamics of rats with heart failure and the effects of plasma angiotensin II and aldosterone [ J ]. reported by Shanghai university of medicine 2015, 29(05):53-56.
[4] Xujie, Dian Hui Yan, Jing Feng, etc. the effect of deer Red grain on the ACE2-Ang (1-7) axis of rats with heart failure [ J ]. proceedings of Shanghai university of medicine 2015, 29(6):41-44, 49.
[5] Yellow dawn Wei, Xuyan, Handong, influence of pilose antler polypeptide-mediated cardiac stem cell differentiation on the expression of terminal cardiac differentiation genes ANP and MLC-2v [ J ]. the university of Jilin school report (medical edition), 2018, 44(02):249-253+464.
[6]Liu Y N.Zhou Z M.Chen P.Evidencethat hydroxysafflor yellow A protects the heart against ischemia-reperfusion injury by inhibiting mitoehondrial permeability transition pore opening[J].Clinical and Experimental Pharmacology and Physiology,2008,35(2):211-216.
[7]Zhu H B.Zhang L.Wang Z H，et al.Therapeutic effects of hydroxysafflor yellow A on focal cerebral ischemic injury in rats and its primary mechanisms[J].Journal of Asian Natural Products Research.2005，7(4):607-613.
[8]ZHOU H，YUAN Y，LIU Y，et al.Icariin attenuates angiotensin IIinduced hypertrophy and apoptosis in H9c2 cardiomyocytes by inhibiting reactive oxygen species-dependent JNK and p38 pathways[J].Exp Ther Med，2014，7(5):1116-1122.
[9]ZHANG Q，LI H，WANG S，et al.Icariin protects rat cardiac H9c2 cells from apoptosis by inhibiting endoplasmic reticulum stress[J].Int J Mol Sci，2013，14(9):17845-17860.
[10] The effect of the extract of the skin and the seed of the Malong grape on the blood lipid level of the rat model with hyperlipidemia [ J ] academic newspaper of Xinjiang medical university, 2005(6): 521-.
[11] Clinical and experimental studies on the treatment of heart failure by the prescription of Fuwanying, Zhouyanhua, Dong Yan, antler [ J ]. J.Shanghai J.J.Med.1992 (11): 8-11).
[12] The effective components of dogwood have protective effect on the myocardium of mice with diabetes [ J ]. J. Chen-L.J., 2016, 36(19):4681-4682. fig..
[13] Influence of Wang Hushu, Liu Yulan and Oleanolic acid on platelet function [ J ]. Shenyang pharmaceutical university report, 1993, 10(4):275.
[14] Zhang Anna, Lipeng, hong Xiao Hua, Chen Feiyu, Wang Yang Hui, Liu Xin Yu, ren Jun Guo, Liu Jian, Special Ligustrum lucidum has protective effect on glomerular mesangial cell apoptosis stimulated by high sugar [ J ]. J.China J.Experimental & Chemicals, 2015, 21(08): 116-.
[15] Daboudoir pillars in 2005 shallow analysis of guidelines for treatment of chronic systolic heart failure [ J ]. J.China cardiovascular disease, 2005, 33(12):1065.
[16] National pharmacopoeia committee, pharmacopoeia of the people's republic of China, part [ M ]. Beijing, pharmaceutical science and technology publishing Co., China 2015, 185.
[17] Research on extraction process of effective components of Bakuchiba in Diqingfeng, Guangdong chemical industry, 2019,46(23):45-46.
The above description is only a preferred embodiment of the present invention, and it should be noted that, for those skilled in the art, several modifications and additions can be made without departing from the method of the present invention, and these modifications and additions should also be regarded as the protection scope of the present invention.
Claims (8)
1. A high performance liquid chromatography method for simultaneously determining the content of 7 components in an anti-chronic heart failure medicine compound is characterized in that the 7 components are gallic acid, hydroxysafflor yellow A, loganin, agaroterol, psoralen glycoside, isopsoralen glycoside and specnuezhenide, and the chromatographic conditions of the high performance liquid chromatography method are as follows: adopting a C18 chromatographic column, wherein the mobile phase acetonitrile-0.1% phosphoric acid water is adopted, the elution gradient is 0-10 min, and the elution gradient is 5% acetonitrile; 10-15 min,5% -10% acetonitrile; 15-35 min,10% -18% acetonitrile; 35-40 min,18% -25% acetonitrile; 40-60 min, 25-35% acetonitrile; the flow rate is 0.8-1.2 mL/min < -1 >, the column temperature is 28-32 ℃, the detection wavelength is 220-360 nm, and the anti-chronic heart failure compound traditional Chinese medicine comprises the following components in percentage by weight: 10-20 parts of antler, 6-12 parts of safflower, 15-25 parts of fructus psoraleae, 25-35 parts of epimedium herb, 10-20 parts of cornus officinalis, 25-35 parts of glossy privet fruit and 4-8 parts of agilawood, and the preparation method of the test solution by the high performance liquid chromatography method comprises the following steps: precisely weighing the above materials, adding 8-15 times of water, decocting for 60-90min, filtering, concentrating the filtrate under reduced pressure, transferring to volumetric flask, metering volume, shaking, filtering with microporous membrane, and collecting the filtrate.
2. The high performance liquid chromatography method according to claim 1, wherein the mixed reference solution is prepared by: accurately weighing gallic acid, hydroxy safflower yellow A, loganin, linalool, psoralen, isopsoralen, and specnuezhenide, and adding methanol to obtain mixed reference solutions with concentrations of 2.71, 8.61, 7.61, 643, 17.42, 11.88, and 45.00 μ g/mL.
3. The high performance liquid chromatography method according to claim 1, wherein the amount of sample is 1 to 12. mu.L.
4. The high performance liquid chromatography method according to claim 3, wherein the amount of the sample is 10. mu.L.
5. The high performance liquid chromatography method according to claim 1, wherein the flow rate is 1.0mL _ min "1.
6. The high performance liquid chromatography method of claim 1, wherein the column temperature is 30 ℃.
7. The high performance liquid chromatography method according to claim 1, wherein the detection wavelength is 220 to 240 nm.
8. A quality control method of a compound anti-chronic heart failure medicine, which is characterized by comprising the steps of the high performance liquid chromatography method of any one of claims 1 to 7.
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