CN108776183A - The method of quality control of rhei and Glycyrhizae Decoction - Google Patents
The method of quality control of rhei and Glycyrhizae Decoction Download PDFInfo
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Abstract
The invention discloses a kind of method of quality control of rhei and Glycyrhizae Decoction, the present invention filters out best flowing phase composition by many experiments, elution program, flow velocity, the analysis conditions such as chromatographic column and detector detection parameters, it is detected using UHPLC-TQ-MS/MS methods, the detection method can detect anthraquinone, saponin component in rhei and Glycyrhizae Decoction and its extract simultaneously, totally 17 kinds of reactive compounds.The experimental results showed that this method detection sensitivity is high, stability is good, can objective, comprehensive, accurately evaluate the quality of rhei and Glycyrhizae Decoction, to control quality and ensure that clinical efficacy is of great significance.
Description
Technical field
The present invention relates to a kind of method of quality control of Chinese medicine, and in particular to the method for quality control of rhei and Glycyrhizae Decoction.
Background technology
Rheum officinale and Radix Glycyrrhizae medicine are to being doctor often with purgatives medicine pair.Rheum officinale bitter cold lets out tie heat, the violent benefaction of power, Radix Glycyrrhizae in stomach clearly
Gan Ping, and rheum officinale phase 5 can be delayed under the rushing down of rheum officinale, rheum officinale stayed, with clean mansion, to exempt from rheum officinale bitter cold wound in stomach, have in so that it is adjusted benefit with
More disease.Two medicine compatibilities, mutual reinforcement between are to use, and Radix Glycyrrhizae mitigates the property that rheum officinale bitter cold is captured, and makes its stopping nausea and vomiting by lowering the adverse flow of QI without the gas that injures one's stomach, and can be clear
Stomach energy is let out, rhei and Glycyrhizae Decoction is also by this two tastes Chinese medicinal composition.
Rheum officinale contains there are many anthraquinone component, contains multiple saponin components in Radix Glycyrrhizae, current detection method, detection
Active ingredient is unable to the active ingredient of complete detection rheum officinale and Radix Glycyrrhizae medicine centering than relatively limited.Therefore, big in order to fully control
The clinical safety of yellow and Radix Glycyrrhizae medicine pair and its preparation, safeguards the interests of patient, it is necessary on the basis of the prior art
Research and design goes out the detection method of the accurate complete detection rhei and Glycyrhizae Decoction of energy and its preparation active ingredient.
Invention content
Goal of the invention：Present invention aim to address the deficiencies in the prior art, are screened by many experiments, using ultra high efficiency
Liquid chromatographic detection, the detection method can detect Anthraquinones in rhei and Glycyrhizae Decoction, steroid totally 17 kinds of active ingredients simultaneously
Object.This method detection sensitivity is high, and stability is good, can objective, comprehensive, accurately evaluate the matter of rhei and Glycyrhizae Decoction and its preparation
Amount to control quality and ensures that curative effect is of great significance.
Technical solution：In order to achieve the goal above, the technical solution that the present invention takes is：
A kind of method of quality control of rhei and Glycyrhizae Decoction, includes the following steps：
(1) preparation of sample solution
First extracting liquorice medicinal material adds the water refluxing extraction of 8 to 16 times of amounts, rhubarb medicinal material is then added, and medicinal material gross weight is added
Rhubarb glycyrrhiza Aqueous extracts are obtained by filtration in the water of 8~16 times of amount, refluxing extraction, centrifuge, Aspirate supernatant, cross 0.22 μm of filter membrane, obtain
Sample solution；
(2) preparation of reference substance solution
Accurately weighed Rhein, rheum emodin, aloe-emodin, Chrysophanol, Physcion, the Portugals Rhein -8-O- β-D-
Polyglycoside, emodin -8-O-β-D-glucosule, Sennoside A, Sennoside B, glycyrrhizic acid, enoxolone, liquiritin, different Radix Glycyrrhizae
Glycosides, glycyrrhizin, isoliquiritigenin, Licochalcone A and 17 kinds of reference substances of glabridin are each, are dissolved to obtain reference substance respectively with methanol
Then storing solution quantitatively draws each reference substance storing solution and obtains mixing reference substance storing solution, crosses 0.22 μm of filter membrane, must mix control
Product solution；
(4) quantitative analysis
By the sample solution that step (1) is prepared and the mixed reference substance solution that step (2) is prepared, carry out respectively
Ultra performance liquid chromatography detects, and obtains the UHPLC-TQ-MS/MS chromatograms of sample and reference substance respectively, it is qualitative fixed then to carry out
Amount analysis.
Preferably, the method for quality control of above-described rhei and Glycyrhizae Decoction, ultra high efficiency liquid phase used in step (3)
Chromatographic parameter setting is as follows：
Instrument：Waters ACQUITY UPLC systems；Tetons mass detectors；
Chromatographic column：Thermo Scientific Hypersil Gold C18Chromatographic column, 100mm × 2.1mm, 1.9 μm,；
Column temperature：35℃；Sampling volume：2μL；Mobile phase：A phases are 0.1% formic acid water, B phases are acetonitrile；Flow velocity：0.4ml·min-1；Ladder
Degree elution；
Mass Spectrometry Conditions are：Ion source：ESI ion sources；Ion source temperature：150℃；Scan mode：More reaction detection patterns；
Capillary voltage：3.0kV；Remove solvent temperature degree：150℃；Go solvent stream amount：1000L·h-1；Taper hole throughput：30L·h-1；Collision gas flow：0.15mL·min-1。
Preferably, the method for quality control of above-described rhei and Glycyrhizae Decoction, gradient elution program are：0~
2min, A：95%~95%, B：5%~5%；2~4min, A：95%~65%, B：5%~35%；4~8min, A：65%
~5%, B：35%~95%；8~13min, A：5%~5%, B：95%~95%；13~14min, A：5%~95%, B：
95%~5%；14~15min, A：95%~95%, B：5%~5%.
Preferably, the method for quality control of above-described rhei and Glycyrhizae Decoction, in step (3) with 90% methanol
Mixed reference substance solution is diluted step by step according to dilution method step by step, the reference substance solution point of 5 various concentration ranks is made
Not carry out ultra performance liquid chromatography detection, measure the peak area of 17 ingredients in each reference substance solution, using each constituent concentration as
Abscissa, peak area are the standard curve that ordinate draws 17 ingredients, carry out linear analysis using the Return Law, calculate each control
The linearly dependent coefficient r of product2；And LOQ and LOD are measured, such as following table
。
Preferably, the method for quality control of above-described rhei and Glycyrhizae Decoction, the system of step (2) reference substance solution
Preparation Method is：
Accurately weighed Rhein, rheum emodin, aloe-emodin, Chrysophanol, Physcion, the Portugals Rhein -8-O- β-D-
Polyglycoside, emodin -8-O-β-D-glucosule, Sennoside A, Sennoside B, glycyrrhizic acid, enoxolone, liquiritin, different Radix Glycyrrhizae
Glycosides, glycyrrhizin, isoliquiritigenin, Licochalcone A and each 5mg of glabridin reference substance or so, concentration is dissolved as with 90% methanol
Respectively 108.6 μ gmL of Rhein-1, rheum emodin, 106.0 μ gmL-1, 220.0 μ gmL of aloe-emodin-1, Chrysophanol
90.0μg·mL-1, 130.0 μ gmL of Physcion-1, Rhein -8-O- β -1347.0 μ gmL of D-Glucose glycosides-1, rheum officinale
Element -8-O- β -408.3 μ gmL of D-Glucose glycosides-1, 126.0 μ gmL of Sennoside A-1, 245.0 μ gmL of Sennoside B-1, it is sweet
1353.0 μ gmL of oxalic acid-1, 721.4 μ gmL of enoxolone-1, 505.7 μ gmL of liquiritin-1, 3320.0 μ g of isoliquiritin
mL-1, 3233.0 μ gmL of glycyrrhizin-1, 3553.0 μ gmL of isoliquiritigenin-1, 2127.0 μ gmL of licochalcone-1, light Radix Glycyrrhizae
Fixed 2647.0 μ gmL-1Reference substance storing solution；Then it quantitatively draws each reference substance storing solution and obtains mixing reference substance storing solution,
Wherein Rhein (1), rheum emodin (2), aloe-emodin (3), Chrysophanol (4), Physcion (5), Rhein -8-O- β-D-
Glucoside (6), emodin -8-O-β-D-glucosule (7), Sennoside A (8), Sennoside B (9), glycyrrhizic acid (10), Radix Glycyrrhizae
Hypo acid (11), liquiritin (12), isoliquiritin (13), glycyrrhizin (14), isoliquiritigenin (15), Licochalcone A (16), light are sweet
The content of grass fixed (17) is respectively 15.404 μ gmL-1、15.035μg·mL-1、15.603μg·mL-1、12.766μg·mL-1、
18.440μg·mL-1、19.106μg·mL-1、14.479μg·mL-1、17.872μg·mL-1、17.376μg·mL-1、
19.191μg·mL-1、15.349μg·mL-1、14.346μg·mL-1、23.546μg·mL-1、22.929μg·mL-1、
25.199μg·mL-1、15.085μg·mL-1、23.005μg·mL-1。
Preferably, the method for quality control of above-described rhei and Glycyrhizae Decoction, the sample solution described in step (1)
Preparation method be：
Then after adding 8 times of water refluxing extraction 1h measured rhubarb medicinal material is added, and medicinal material gross weight is added in first extracting liquorice medicinal material
The water of 8 times of amount, refluxing extraction 2 times merge extracting solution, rhubarb glycyrrhiza Aqueous extracts are obtained by filtration, and centrifuge, Aspirate supernatant, mistake
0.22 μm of filter membrane, obtains sample solution.The weight ratio of rheum officinale and Radix Glycyrrhizae is 4:1,4:2,4:3,4:4,3:4,2:4 or 1:4.
Condition optimizing is tested
The present invention, can be to all the components by being investigated to conditions such as mobile phase, gradient elution program, flow velocitys
It can be detached in short period and preferable peak shape can be obtained.It was found that it is big to can be good at separation with 0.1% formic acid water-acetonitrile
17 kinds of active constituents in Huang-Radix Glycyrrhizae, formic acid, which is added, can reduce the trailing phenomenon of fragrant acrylic component；It is washed by optimizing gradient
It is preferable that de- program finds that high proportion organic phase detaches active constituent in rheum officinale-Radix Glycyrrhizae, determines gradient；Flow rate of mobile phase is
0.4ml·min-1It is 0.2mlmin that each ingredient separating effect and peak shape, which are better than flow velocity,-1.Pattern (cation and negative is swept using complete
Ion mode) 17 kinds of compounds of optimization mass spectrometry parameters, instrument analyzing the letter of each ingredient under cation and negative ion mode
Number intensity and sensitivity select detection pattern, and orifice potential and capillary voltage are by software Automatic Optimal.In chromatography and mass spectrum item
The UHPLC-TQ-MS/MS chromatograms that 17 kinds of active constituents in rheum officinale-Radix Glycyrrhizae are obtained after piece optimization are as shown in Figure 1.
Advantageous effect：Compared to the prior art the method for quality control of rhei and Glycyrhizae Decoction provided by the invention has following excellent
Point：
The present invention is according to 2 class heterogeneity of anthraquinone in rhei and Glycyrhizae Decoction and saponin(e, the work of totally 17 kinds of different structures and property
The structure and its property feature of property compound, filter out best flowing phase composition by many experiments, elution program, flow velocity,
The analysis conditions such as chromatographic column, detector detection parameters.Show that the present invention can detect anthraquinone and soap simultaneously through many experiments verification
2 class heterogeneity of glycosides totally 17 kinds of compounds, this method detection sensitivity is high, and stability is good, can it is objective, comprehensively and accurately comment
The quality of valence rhei and Glycyrhizae Decoction and its preparation to control quality and ensures that clinical efficacy is of great significance.
Description of the drawings
Fig. 1 is more reaction detection pattern (MRM) chromatograms of the UHPLC-TQ-MS/MS of 17 ingredients in rhei and Glycyrhizae Decoction.
Specific implementation mode
With reference to specific embodiment, the present invention is furture elucidated, it should be understood that these embodiments be merely to illustrate the present invention and
It is not used in and limits the scope of the invention, after having read the present invention, various of equal value shapes of the those skilled in the art to the present invention
The modification of formula falls within the application range as defined in the appended claims.
Embodiment 1
1, a kind of method of quality control of rhei and Glycyrhizae Decoction comprising following steps：
Instrument and reagent
1.1 instrument
WatersACQUITY UHPLC systems are purchased from Waters, US；Tetons mass detectors are purchased from the U.S.
Waters companies；MassLynxTMMass spectrum workstation software is purchased from Waters, US；Ultra-pure water instrument is purchased from Nanjing Yi Puyi
Up to development in science and technology Co., Ltd；Micofuge 22R Centrifuge type desk centrifuges, purchased from the learned Creative Science and Technology Co. Ltd's hair in Beijing
Open up Co., Ltd；Ultrasonic washing instrument is purchased from Kunshan He Chuan ultrasonic instruments Co., Ltd；
1.2 drugs and reagent
Rheum officinale (the place of production：Gansu, lot number：20160319), the Radix Glycyrrhizae (place of production：Ningxia, lot number：151108), rheum officinale is that Tang is ancient
Especially big Huang, Radix Glycyrrhizae is Glycyrrhiza Uralensis, and meets standards of pharmacopoeia.
Chemical standard product Rhein (1), rheum emodin (2), aloe-emodin (3), Chrysophanol (4), Physcion (5),
Rhein -8-O- β-D-Glucose glycosides (6), emodin -8-O-β-D-glucosule (7), Sennoside A (8), Sennoside B (9),
Glycyrrhizic acid (10), enoxolone (11), liquiritin (12), isoliquiritin (13), glycyrrhizin (14), isoliquiritigenin (15), Radix Glycyrrhizae
Chalcone A (16), glabridin (17) are purchased from the Chengdu bio tech ltd Rui Fensi, and purity is more than 98%.Methanol, chromatography
It is pure, it is purchased from Merck Reagent Companies of Germany；Acetonitrile, chromatographically pure are purchased from Merck Reagent Companies of Germany.
2 experimental methods
The preparation of 2.1 reference substance solutions
Each 5mg of accurately weighed above-mentioned 17 kinds of reference substances or so, with 90% methanol be dissolved as concentration be respectively Rhein (1,
108.6μg·mL-1), rheum emodin (2,106.0 μ gmL-1), aloe-emodin (3,220.0 μ gmL-1), Chrysophanol (4,
90.0μg·mL-1), Physcion (5,130.0 μ gmL-1), Rhein -8-O- β-D-Glucose glycosides (6,1347.0 μ g
mL-1), emodin -8-O-β-D-glucosule (7,408.3 μ gmL-1), Sennoside A (8,126.0 μ gmL-1), Sennoside B
(9,245.0 μ gmL-1), glycyrrhizic acid (10,1353.0 μ gmL-1), enoxolone (11,721.4 μ gmL-1), liquiritin
(12,505.7 μ gmL-1), isoliquiritin (13,3320.0 μ gmL-1), glycyrrhizin (14,3233.0 μ gmL-1), different Radix Glycyrrhizae
Element (15,3553.0 μ gmL-1), licochalcone (16,2127.0 μ gmL-1), glabridin (17,2647.0 μ gmL-1)
Reference substance storing solution.It quantitatively draws each reference substance storing solution and obtains mixing reference substance storing solution, wherein Rhein (1), rheum officinale
Plain (2), aloe-emodin (3), Chrysophanol (4), Physcion (5), Rhein -8-O- β-D-Glucose glycosides (6), rheum officinale
Element -8-O- β-D-Glucose glycosides (7), Sennoside A (8), Sennoside B (9), glycyrrhizic acid (10), enoxolone (11), liquiritin
(12), the content of isoliquiritin (13), glycyrrhizin (14), isoliquiritigenin (15), Licochalcone A (16), glabridin (17)
Respectively 15.404 μ gmL-1、15.035μg·mL-1、15.603μg·mL-1、12.766μg·mL-1、18.440μg·mL-1、19.106μg·mL-1、14.479μg·mL-1、17.872μg·mL-1、17.376μg·mL-1、19.191μg·mL-1、
15.349μg·mL-1、14.346μg·mL-1、23.546μg·mL-1、22.929μg·mL-1、25.199μg·mL-1、
15.085μg·mL-1、23.005μg·mL-1。
The preparation of 2.2 test solutions
Then after adding 8 times of water refluxing extraction 1h measured rhubarb medicinal material is added, and medicinal material gross weight is added in first extracting liquorice medicinal material
The water of 8 times of amount, refluxing extraction 2 times merge extracting solution, rhubarb glycyrrhiza Aqueous extracts are obtained by filtration, and centrifuge, Aspirate supernatant, mistake
0.22 μm of filter membrane, obtains sample solution.The weight ratio of rheum officinale and Radix Glycyrrhizae is 4:1.
2.3 chromatographiccondition
Chromatographic column：Thermo Scientific Hypersil Gold C18Chromatographic column (100mm × 2.1mm, 1.9 μm)；
Column temperature：35℃；Sampling volume：2μL；Mobile phase：A (0.1% formic acid water), B (acetonitrile)；Flow velocity：0.4ml·min-1；Gradient is washed
It is de-：0~1min, A：95%~95%, B：5%~5%；2~4min, A：95%~65%, B：5%~35%；4~8min,
A：65%~5%, B：35%~95%；8~13min, A：5%~5%, B：95%~95%；13~14min, A：5%~
95%, B：95%~5%；14~15min, A：95%~95%, B：5%~5%.
2.4 Mass Spectrometry Conditions
Ion source：ESI ion sources；Ion source temperature：150℃；Scan mode：More reaction detection patterns (MRM)；Capillary
Voltage：3.0kV；Remove solvent temperature degree：150℃；Go solvent stream amount：1000L·h-1；Taper hole throughput：30L·h-1；Collision
Throughput：0.15mL·min-1.Each ingredient retention time and mass spectrometry parameters are shown in Table 1.
1 each ingredient retention time of table and mass spectrometry parameters
2.5 methodological study
2.5.1 linear relationship, quantitative limit (LOD), detection limit (LOQ)
Reference substance storing solution is diluted step by step according to dilution method step by step with 90% methanol, various concentration rank is made
Reference substance solution according under 2.3 and 2.4 chromatography and Mass Spectrometry detection method measure in each reference substance solution 17 ingredients
Peak area, using each constituent concentration as abscissa, peak area is the standard curve that ordinate draws this 17 ingredients, using recurrence
Method carries out linear analysis, calculates the linearly dependent coefficient (r of each reference substance2)；Reference substance concentration difference when signal-to-noise ratio is 3,10
For LOQ, LOD.Such as the following table 2：
2 each ingredient standard curve of table and the range of linearity and LOQ, LOD
2.5.2 precision, repeatability, stability
Precision is investigated：A reference substance solution is measured into each reference substance solution according to the above chromatography and Mass Spectrometry detection method
In 17 ingredients peak area, repeat sample introduction 3 times daily, for three days on end and on the same day in continuous sample introduction 6 times, with each at swarming face
Long-pending relative standard deviation (RSD) evaluation sample is in the daytime and withinday precision.
Repeatability is investigated：6 parts of rheum officinale-Radix Glycyrrhizaes 1 of parallel preparation:The test solution of 1 proportioning, is examined by above-mentioned chromatography and mass spectrum
Survey method measures the peak area of each ingredient in test sample, is evaluated with the relative standard deviation (RSD) of each Component peak area to comment
The repeatability of valence instrument.
Study on the stability：Take rheum officinale-Radix Glycyrrhizae 1:The test solution of 1 proportioning 1,2,4,8,12, for 24 hours when distinguish sample introduction,
The changes of contents for measuring 17 kinds of compounds in different time test sample evaluates sample with the RSD values of this 17 kinds of compound peaks areas
The stability of product.
2.5.3 the rate of recovery is tested
The reference substance solution of its content 80%, 100%, 120% is added in the sample of known 17 component contents, for examination
Product solution presses above-mentioned chromatography and Mass Spectrometry detection method determination sample peak area, obtains sample concentration according to standard curve, calculates and add
The sample rate of recovery, sample recovery rate=(mark-on sample solution concentration-sample solution concentration)/mark-on amount.The result shows that 17 kinds in rheum officinale and Radix Glycyrrhizae
For the active constituent rate of recovery in 98.16%~100.64% range, accuracy is good.
2.6 statistical analysis
All results are handled with 22.0 softwares of SPSS, and it is poor to carry out statistics using Dunnett methods in ANOVA
Different analysis, P<0.05 indicates that there is difference statistical significance, all results to be indicated using mean+SD (mean ± SD).
3. experimental result
Such as table 3, precision of the present invention, repeatability, stability, the results showed that its RSD is respectively less than 3%, and precision is good, rheum officinale
And in Radix Glycyrrhizae 17 kinds of active constituent rate of recovery in 98.16%~100.64% range.
3 precision of table, repeatability, stability and sample-adding recovery experiment
Detection method provided by the invention can accurately detect 17 compounds, and rhubarb glycyrrhiza provided by the invention simultaneously
The method of quality control of soup, precision are high, high sensitivity, and stability and accuracy are high, can it is objective, comprehensive, accurately evaluate
The quality of rhei and Glycyrhizae Decoction, the quality to accurately controlling rhei and Glycyrhizae Decoction are of great significance.
The above is only a preferred embodiment of the present invention, it is noted that for the ordinary skill people of the art
For member, various improvements and modifications may be made without departing from the principle of the present invention, these improvements and modifications are also answered
It is considered as protection scope of the present invention.
Claims (6)
1. a kind of method of quality control of rhei and Glycyrhizae Decoction, which is characterized in that include the following steps：
(1) preparation of sample solution
First extracting liquorice medicinal material, add 8 to 16 times amount water refluxing extractions, rhubarb medicinal material is then added, and be added medicinal material total weight 8~
Rhubarb glycyrrhiza Aqueous extracts are obtained by filtration in 16 times of water, refluxing extraction, centrifugation, and Aspirate supernatant crosses 0.22 μm of filter membrane, obtains sample
Solution；
(2) preparation of reference substance solution
Accurately weighed Rhein, rheum emodin, aloe-emodin, Chrysophanol, Physcion, Rhein -8-O- β-D-Glucose
It is glycosides, emodin -8-O-β-D-glucosule, Sennoside A, Sennoside B, glycyrrhizic acid, enoxolone, liquiritin, isoliquiritin, sweet
Careless element, isoliquiritigenin, Licochalcone A and 17 kinds of reference substances of glabridin are each, are dissolved to obtain reference substance deposit respectively with methanol
Then liquid quantitatively draws each reference substance storing solution and obtains mixing reference substance storing solution, crosses 0.22 μm of filter membrane, it is molten must to mix reference substance
Liquid；
(3) quantitative analysis
By the sample solution that step (1) is prepared and the mixed reference substance solution that step (2) is prepared, superelevation is carried out respectively
Effect liquid phase chromatogram detects, and obtains the UHPLC-TQ-MS/MS chromatograms of sample and reference substance respectively, then carries out qualitative, quantitative point
Analysis.
2. the method for quality control of rhei and Glycyrhizae Decoction according to claim 1, which is characterized in that
Ultra performance liquid chromatography parameter setting is as follows used in step (3)：
Instrument：WatersACQUITYUPLC systems；Tetons mass detectors；
Chromatographic column：ThermoScientificHypersilGoldC18Chromatographic column, 100mm × 2.1mm, 1.9 μm,；Column temperature：35
℃；Sampling volume：2μL；Mobile phase：A phases are 0.1% formic acid water, B phases are acetonitrile；Flow velocity：0.4ml·min-1；Gradient elution；
Mass Spectrometry Conditions are：Ion source：ESI ion sources；Ion source temperature：150℃；Scan mode：More reaction detection patterns；Capillary
Tube voltage：3.0kV；Remove solvent temperature degree：150℃；Go solvent stream amount：1000L·h-1；Taper hole throughput：30L·h-1；It touches
Hit throughput：0.15mL·min-1。
3. the method for quality control of rhei and Glycyrhizae Decoction according to claim 2, which is characterized in that
Gradient elution program is：0~2min, A：95%~95%, B：5%~5%；2~4min, A：95%~65%, B：5%
~35%；4~8min, A：65%~5%, B：35%~95%；8~13min, A：5%~5%, B：95%~95%；13~
14min, A：5%~95%, B：95%~5%；14~15min, A：95%~95%, B：5%~5%.
4. the method for quality control of rhei and Glycyrhizae Decoction according to claim 2, which is characterized in that
Mixed reference substance solution is diluted step by step according to dilution method step by step with 90% methanol in step (3), is made 5 not
Ultra performance liquid chromatography detection is carried out respectively with the other reference substance solution of concentration level, measures in each reference substance solution 17 ingredients
Peak area, using each constituent concentration as abscissa, peak area is the standard curve that ordinate draws 17 ingredients, using the Return Law
Linear analysis is carried out, the linearly dependent coefficient r of each reference substance is calculated2；And LOQ and LOD are measured, such as following table
5. the method for quality control of rhei and Glycyrhizae Decoction according to claim 1, which is characterized in that
The preparation method of step (2) reference substance solution is：
Accurately weighed Rhein, rheum emodin, aloe-emodin, Chrysophanol, Physcion, Rhein -8-O- β-D-Glucose
It is glycosides, emodin -8-O-β-D-glucosule, Sennoside A, Sennoside B, glycyrrhizic acid, enoxolone, liquiritin, isoliquiritin, sweet
Careless element, isoliquiritigenin, Licochalcone A and each 5mg of glabridin reference substance or so, being dissolved as concentration with 90% methanol is respectively
108.6 μ gmL of Rhein-1, rheum emodin, 106.0 μ gmL-1, 220.0 μ gmL of aloe-emodin-1, 90.0 μ g of Chrysophanol
mL-1, 130.0 μ gmL of Physcion-1, Rhein -8-O- β -1347.0 μ gmL of D-Glucose glycosides-1, rheum emodin -8-O-
408.3 μ gmL of β-D-Glucose glycosides-1, 126.0 μ gmL of Sennoside A-1, 245.0 μ gmL of Sennoside B-1, glycyrrhizic acid
1353.0μg·mL-1, 721.4 μ gmL of enoxolone-1, 505.7 μ gmL of liquiritin-1, 3320.0 μ gmL of isoliquiritin-1、
3233.0 μ gmL of glycyrrhizin-1, 3553.0 μ gmL of isoliquiritigenin-1, 2127.0 μ gmL of licochalcone-1, glabridin
2647.0μg·mL-1Reference substance storing solution；Then it quantitatively draws each reference substance storing solution and obtains mixing reference substance storing solution,
Middle Rhein (1), rheum emodin (2), aloe-emodin (3), Chrysophanol (4), Physcion (5), the Portugals Rhein -8-O- β-D-
Polyglycoside (6), emodin -8-O-β-D-glucosule (7), Sennoside A (8), Sennoside B (9), glycyrrhizic acid (10), Radix Glycyrrhizae time
Sour (11), liquiritin (12), isoliquiritin (13), glycyrrhizin (14), isoliquiritigenin (15), Licochalcone A (16), light Radix Glycyrrhizae
The content of fixed (17) is respectively 15.404 μ gmL-1、15.035μg·mL-1、15.603μg·mL-1、12.766μg·mL-1、
18.440μg·mL-1、19.106μg·mL-1、14.479μg·mL-1、17.872μg·mL-1、17.376μg·mL-1、
19.191μg·mL-1、15.349μg·mL-1、14.346μg·mL-1、23.546μg·mL-1、22.929μg·mL-1、
25.199μg·mL-1、15.085μg·mL-1、23.005μg·mL-1。
6. the method for quality control of rhei and Glycyrhizae Decoction according to claim 1, which is characterized in that the sample described in step (1)
The preparation method of product solution is：
Then after adding 8 times of water refluxing extraction 1h measured rhubarb medicinal material is added, and 8 times of medicinal material total weight is added in first extracting liquorice medicinal material
Water, refluxing extraction 2 times, merge extracting solution, rhubarb glycyrrhiza Aqueous extracts are obtained by filtration, centrifuge, Aspirate supernatant, cross 0.22 μm filter
Film obtains sample solution.The weight ratio of rheum officinale and Radix Glycyrrhizae is 4:1,4:2,4:3,4:4,3:4,2:4 or 1:4.
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