CN106645438B - A kind of detection method of 'Shensu ' - Google Patents
A kind of detection method of 'Shensu ' Download PDFInfo
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Abstract
The present invention relates to technical field of medicine quality control, in particular to a kind of detection method of 'Shensu '.The detection method includes: that test solution is made in 'Shensu ', and Puerarin receives the quality of 'Shensu ', chromatographic condition are as follows: chromatographic column C using high performance liquid chromatography detection as object of reference18Reverse-phase chromatographic column, mobile phase A are acetonitrile, and Mobile phase B is phosphoric acid solution, using gradient elution.The precision of the efficient liquid-phase chromatography method is higher, reproducibility is good, and analysis time is shorter, has certain specificity；Each characteristic peak separating effect is preferable in gained characteristic spectrum, and 'Shensu ' characteristic spectrum and intermediate characteristic spectrum have preferable correlation；Characteristic spectrum established by the present invention has 7 characteristic peaks, and information content is larger, more completely remains the effective component in test liquid, and method provided by the invention has advantageously ensured the whole-course quality control from raw material to terminal preparation.
Description
Technical field
The present invention relates to technical field of medicine quality control, in particular to a kind of detection method of 'Shensu '.
Background technique
'Shensu ' is by Radix Codonopsis, perilla leaf, pueraria lobata, the root of purple-flowered peucedanum, Poria cocos, the tuber of pinellia (system), dried orange peel, Fructus Aurantii (stir-fry), campanulaceae, sweet
Grass, radix aucklandiae, ginger, 13 taste medicinal material of jujube composition.The party has effects that benefiting qi for relieving superficies syndrome, expelling phlegm and arresting coughing, is mainly used for treating
Weakness type cold, anemofrigid cold, fever with aversion to cold, headache nasal obstruction, cough ant phlegm.Shensu drink comes from " the peaceful the Bureau of People's Welfare Pharmacies of Song Dynasty
Side ", clinical application is relatively broad.
In the prior art, according to ginseng Soviet Union prescription, the 'Shensu ' of a variety of dosage forms has been made, 'Shensu ' is main in the market
There are oral solution, pill, tablet, capsule etc..Common preparation process are as follows: by raw medicinal material it is cataclasm after, by existing conventional method
Oral solution, pill, tablet or capsule is made.
For 'Shensu ' as Chinese patent drug, drug effect is the coefficient result of a variety of active ingredients.But current 'Shensu '
Existing quality standard only measures the content of single index components Puerarin, is not monitored to other effective components, there are one
It settles finally sex-limited, cannot reflect the quality level of 'Shensu ' comprehensively.For the publication of the quality determining method of 'Shensu '
Only one, Chinese patent 200810069061.8 (publication number CN101474274A) controls its Puerarin only with HPLC method
With the content of aurantiamarin.
As follows for the quality determining method of 'Shensu ', the 4th phase of volume 9 " Hunan traditional Chinese medicine Leader " in April, 2003 " joins
Revive oral solution quality standard research ", " Chinese Medicine guide " the 9th phase of volume 11 in 2009, " Shensuganmao Tablets quality standard was ground
Study carefully ", in the articles such as the 3rd phase " Shensuganmao Tablets quality standard research " of volume 14 " Chinese experimental pharmacology of traditional Chinese medical formulae magazine " in March, 2008
Describe a kind of HPLC method for measuring the content of Puerarin in 'Shensu '；" northwest pharmaceutical journal " in May, 2014 of volume 29
3rd phase research of ginseng and Perillae Pill quality standard " improve " describes praeruptorin A, RADIX PEUCEDANI second in a kind of measurement ginseng and Perillae Pill
The HPLC method of the content of element；" China Dispensary " the 18th phase of volume 18 in 2007, " aurantiamarin contained in HPLC method measurement ginseng and Perillae Pill
Amount " describe a kind of HPLC method for measuring the content of aurantiamarin in ginseng and Perillae Pill.
In the prior art, it there is no the quality of more comprehensive method of quality control reflection existing Chinese medicine material, intermediate and finished product
Situation also can not effectively control production process and product quality, cannot preferably guarantee its clinical efficacy.Therefore, ginseng is established
Made in CCCP dose of quality determining method has very important significance.
Summary of the invention
In view of this, the present invention provides a kind of detection methods of 'Shensu '.The 'Shensu ' quality determining method
It establishes and is conducive to effectively control and guarantee its clinic treatment to its raw medicinal material, intermediate and its finished product effective component quality
Effect.
In order to achieve the above-mentioned object of the invention, the present invention the following technical schemes are provided:
The present invention provides a kind of detection methods of 'Shensu ', include the following steps:
Test solution is made in 'Shensu ', Puerarin is received as object of reference using high performance liquid chromatography detection
The testing result of 'Shensu ', chromatographic condition are as follows:
Chromatographic column is C18Reverse-phase chromatographic column, mobile phase are the gradient eluent of mobile phase A and Mobile phase B composition, mobile phase A
For acetonitrile, Mobile phase B is phosphoric acid solution, and using gradient elution, gradient elution program is as follows:
Preferably, the Detection wavelength of high performance liquid chromatography is 260~300nm.
In some embodiments provided by the invention, the Detection wavelength of high performance liquid chromatography is 283nm.
In other embodiments provided by the invention, the Detection wavelength of high performance liquid chromatography is 260nm.
In other embodiments provided by the invention, the Detection wavelength of high performance liquid chromatography is 300nm.
Preferably, the volumn concentration of phosphoric acid is 0.05%~0.1% in phosphoric acid solution.
In some embodiments provided by the invention, the volumn concentration of phosphoric acid is 0.05% in phosphoric acid solution.
In other embodiments provided by the invention, the volumn concentration of phosphoric acid is 0.1% in phosphoric acid solution.
Preferably, the column temperature of high performance liquid chromatography is 25 DEG C~40 DEG C.
In some embodiments provided by the invention, the column temperature of high performance liquid chromatography is 30 DEG C.
In other embodiments provided by the invention, the column temperature of high performance liquid chromatography is 25 DEG C.
In other embodiments provided by the invention, the column temperature of high performance liquid chromatography is 40 DEG C.
Preferably, 0.8~1.2mL/min of flow velocity of mobile phase.
In some embodiments provided by the invention, the flow velocity 1.0mL/min of mobile phase.
In other embodiments provided by the invention, the flow velocity 0.8mL/min of mobile phase.
In other embodiments provided by the invention, the flow velocity 1.2mL/min of mobile phase.
In some embodiments provided by the invention, the sample volume of test solution is 10 μ L.
Preferably, chromatographic column is Unitary C18(5 μm, 4.6 × 250mm), Thermo Syncronis-C18(5 μm,
4.6 × 250mm), Shimadzu Inertsil ODS-2 (5 μm, 4.6 × 250mm).
In some embodiments provided by the invention, 'Shensu ' is oral solution, and the preparation method of test solution uses
Direct diluted method；'Shensu ' is pill, tablet or capsule, and the preparation method of test solution is using refluxing extraction
Method.
In some embodiments provided by the invention, test solution the preparation method comprises the following steps: take 'Shensu ', add methanol or
Ethyl alcohol is extracted using the method that is heated to reflux, is filtered with miillpore filter, take subsequent filtrate to get test solution.
Preferably, the solvent of refluxing extraction is 0%~50% methanol or 0%~50% ethyl alcohol.
In some embodiments provided by the invention, object of reference the preparation method comprises the following steps: Puerarin is dissolved in 50%~
100% methanol, is made object of reference.
Preferably, the analysis time of high performance liquid chromatography is not less than 60min.
The present invention also provides a kind of characteristic spectrum detection method of 'Shensu ', detection method includes the following steps:
Test solution is made in 'Shensu ', Puerarin is received as object of reference using high performance liquid chromatography detection
The characteristic spectrum of 'Shensu ', chromatographic condition are as follows:
Chromatographic column is C18Reverse-phase chromatographic column, mobile phase are the gradient eluent of mobile phase A and Mobile phase B composition, mobile phase A
For acetonitrile, Mobile phase B is phosphoric acid solution, and using gradient elution, gradient elution program is as follows:
Preferably, characteristic spectrum has 7 characteristic peaks, wherein No. 2 peaks are that Puerarin is used as referring to peak, No. 6 peaks are shaddock
Skin glycosides, No. 7 peaks are aurantiamarin, the relative retention time and relative standard deviation of each characteristic peak are as follows:
No. 1 peak: relative retention time 0.529 ± 0.026；
No. 2 peaks: relative retention time 1.000 ± 0.000；
No. 3 peaks: relative retention time 1.322 ± 0.066；
No. 4 peaks: relative retention time 1.784 ± 0.089；
No. 5 peaks: relative retention time 1.830 ± 0.091；
No. 6 peaks: relative retention time 1.910 ± 0.095；
No. 7 peaks: relative retention time 1.962 ± 0.098.
In some embodiments provided by the invention, characteristic spectrum has 7 characteristic peaks, wherein No. 2 peaks are Puerarin work
For referring to peak, No. 6 peaks are aurantiin, No. 7 peaks are aurantiamarin, the relative retention time and relative standard deviation of each characteristic peak are as follows:
No. 1 peak: relative retention time 0.529 ± 0.026, relative standard deviation 0.20%；
No. 2 peaks: relative retention time 1.000 ± 0.000, relative standard deviation 0.00%；
No. 3 peaks: relative retention time 1.322 ± 0.066, relative standard deviation 0.13%；
No. 4 peaks: relative retention time 1.784 ± 0.089, relative standard deviation 0.18%；
No. 5 peaks: relative retention time 1.830 ± 0.091, relative standard deviation 0.18%；
No. 6 peaks: relative retention time 1.910 ± 0.095, relative standard deviation 0.19%；
No. 7 peaks: relative retention time 1.962 ± 0.098, relative standard deviation 0.19%.
In other embodiments provided by the invention, characteristic spectrum has 7 characteristic peaks, wherein No. 2 peaks are Puerarin
As referring to peak, No. 6 peaks are aurantiin, and No. 7 peaks are aurantiamarin, the relative retention time and relative standard deviation of each characteristic peak
Are as follows:
No. 1 peak: relative retention time 0.529 ± 0.026, relative standard deviation 0.12%；
No. 2 peaks: relative retention time 1.000 ± 0.000, relative standard deviation 0.00%；
No. 3 peaks: relative retention time 1.322 ± 0.066, relative standard deviation 0.15%；
No. 4 peaks: relative retention time 1.784 ± 0.089, relative standard deviation 0.10%；
No. 5 peaks: relative retention time 1.830 ± 0.091, relative standard deviation 0.16%；
No. 6 peaks: relative retention time 1.910 ± 0.095, relative standard deviation 0.14%；
No. 7 peaks: relative retention time 1.962 ± 0.098, relative standard deviation 0.14%.
In other embodiments provided by the invention, characteristic spectrum has 7 characteristic peaks, wherein No. 2 peaks are Puerarin
As referring to peak, No. 6 peaks are aurantiin, and No. 7 peaks are aurantiamarin, the relative retention time and relative standard deviation of each characteristic peak
Are as follows:
No. 1 peak: relative retention time 0.529 ± 0.026, relative standard deviation 0.15%；
No. 2 peaks: relative retention time 1.000 ± 0.000, relative standard deviation 0.00%；
No. 3 peaks: relative retention time 1.322 ± 0.066, relative standard deviation 0.18%；
No. 4 peaks: relative retention time 1.784 ± 0.089, relative standard deviation 0.08%；
No. 5 peaks: relative retention time 1.830 ± 0.091, relative standard deviation 0.21%；
No. 6 peaks: relative retention time 1.910 ± 0.095, relative standard deviation 0.17%；
No. 7 peaks: relative retention time 1.962 ± 0.098, relative standard deviation 0.19%.
In other embodiments provided by the invention, characteristic spectrum has 7 characteristic peaks, wherein No. 2 peaks are Puerarin
As referring to peak, No. 6 peaks are aurantiin, and No. 7 peaks are aurantiamarin, the relative retention time and relative standard deviation of each characteristic peak
Are as follows:
No. 1 peak: relative retention time 0.529 ± 0.026, relative standard deviation 0.16%；
No. 2 peaks: relative retention time 1.000 ± 0.000, relative standard deviation 0.00%；
No. 3 peaks: relative retention time 1.322 ± 0.066, relative standard deviation 0.10%；
No. 4 peaks: relative retention time 1.784 ± 0.089, relative standard deviation 0.02%；
No. 5 peaks: relative retention time 1.830 ± 0.091, relative standard deviation 0.14%；
No. 6 peaks: relative retention time 1.910 ± 0.095, relative standard deviation 0.10%；
No. 7 peaks: relative retention time 1.962 ± 0.098, relative standard deviation 0.09%.
In other embodiments provided by the invention, characteristic spectrum has 7 characteristic peaks, wherein No. 2 peaks are Puerarin
As referring to peak, No. 6 peaks are aurantiin, and No. 7 peaks are aurantiamarin, the relative retention time and relative standard deviation of each characteristic peak
Are as follows:
No. 1 peak: relative retention time 0.529 ± 0.026, relative standard deviation 0.21%；
No. 2 peaks: relative retention time 1.000 ± 0.000, relative standard deviation 0.00%；
No. 3 peaks: relative retention time 1.322 ± 0.066, relative standard deviation 0.08%；
No. 4 peaks: relative retention time 1.784 ± 0.089, relative standard deviation 0.06%；
No. 5 peaks: relative retention time 1.830 ± 0.091, relative standard deviation 0.13%；
No. 6 peaks: relative retention time 1.910 ± 0.095, relative standard deviation 0.14%；
No. 7 peaks: relative retention time 1.962 ± 0.098, relative standard deviation 0.14%.
Preferably, the Detection wavelength of high performance liquid chromatography is 260~300nm.
Preferably, the volumn concentration of phosphoric acid is 0.05%~0.1% in phosphoric acid solution.
Preferably, the volumn concentration of phosphoric acid is 0.05% in phosphoric acid solution.
Preferably, the column temperature of high performance liquid chromatography is 25 DEG C~40 DEG C.
Preferably, 0.8~1.2mL/min of flow velocity of mobile phase.
Preferably, the flow velocity 1.0mL/min of mobile phase.
Preferably, the sample volume of test solution is 10 μ L.
In some embodiments provided by the invention, 'Shensu ' is oral solution, and the preparation method of test solution uses
Direct diluted method；'Shensu ' is pill, tablet or capsule, and the preparation method of test solution is using refluxing extraction
Method.
In some embodiments provided by the invention, the solvent of refluxing extraction is 0%~50% methanol or 0%~50% second
Alcohol.
In some embodiments provided by the invention, object of reference the preparation method comprises the following steps: Puerarin is dissolved in 50%~
100% methanol, is made object of reference.
Preferably, the analysis time of high performance liquid chromatography is not less than 60min.
The present invention provides a kind of detection methods of 'Shensu '.The detection method of the 'Shensu ' include: will join it is made in CCCP
Test solution is made in agent, and Puerarin receives the detection knot of 'Shensu ' using high performance liquid chromatography detection as object of reference
Fruit, chromatographic condition are as follows: chromatographic column C18Reverse-phase chromatographic column, mobile phase are the gradient eluent of mobile phase A and Mobile phase B composition,
Mobile phase A is acetonitrile, and Mobile phase B is phosphoric acid solution, using gradient elution.Compared with prior art, beneficial effects of the present invention
It is:
(1) the characteristic spectrum detection method of 'Shensu ' of the present invention is simple to the pre-treating method of each test sample, characteristic
Ingredient retains completely, and test solution has good stability；
(2) precision of the efficient liquid-phase chromatography method is higher, reproducibility is good, and analysis time is shorter, has centainly
Specificity；Each characteristic peak separating effect is preferable in gained characteristic spectrum, and 'Shensu ' characteristic spectrum and intermediate characteristic spectrum
With preferable correlation；
(3) characteristic spectrum established by the present invention has 7 characteristic peaks, and information content is larger, more completely remains test liquid
In effective component, specify 3 characteristic peaks, compared using retention time it is moderate, separating degree is preferable, the biggish Pueraria lobota of peak area
Root element object of reference is used as referring to peak；
(4) application of 'Shensu ' intermediate characteristic spectrum can increase the controllability of 'Shensu ' production process, join made in CCCP
The application of agent characteristic spectrum then advantageously ensures that the effective component quality stabilization and clinical efficacy of 'Shensu ', provided by the invention
Method has advantageously ensured the whole-course quality control from raw material to terminal preparation.
Detailed description of the invention
Fig. 1 is Detection wavelength 3D figure；
Fig. 2 is that object of reference is superimposed chromatogram；S1 shows 5- ethyoxyl person's base furfural reference substance；S2 shows aurantiamarin reference substance；S3
Show Puerarin reference substance；S4 shows aurantiin reference substance；
Fig. 3 is 'Shensu ' characteristic spectrum；1~No. 7 peak is characterized peak, wherein No. 2 peaks are referring to peak；
Fig. 4 is 10 batches of 'Shensu ' characteristic spectrums；
Fig. 5 is 'Shensu ' intermediate characteristic spectrum；
Fig. 6 is 10 batches of 'Shensu ' intermediate characteristic spectrums；
Fig. 7 is ginseng Soviet Union capsule characteristics map；
Fig. 8 is 10 batches of ginsengs Soviet Union capsule characteristics maps；
Fig. 9 is ginseng Soviet Union piece characteristic spectrum；
Figure 10 is 10 batches of ginsengs Soviet Union piece characteristic spectrums；
Figure 11 is ginseng and Perillae Pill characteristic spectrum；
Figure 12 is 10 batches of ginseng and Perillae Pill characteristic spectrums；
Figure 13 is that each characteristic peak of 'Shensu ' characteristic spectrum belongs to map；
Figure 14 is that 'Shensu ' shares peak figure spectrum.
Specific embodiment
The invention discloses a kind of 'Shensu ' characteristic spectrum and its detection method, those skilled in the art can use for reference this
Literary content, is suitably modified realization of process parameters.In particular, it should be pointed out that all similar substitutions and modifications are to art technology
It is it will be apparent that they are considered as being included in the present invention for personnel.Method and application of the invention has passed through preferably
Embodiment is described, related personnel obviously can not depart from the content of present invention, in spirit and scope to side as described herein
Method and application are modified or appropriate changes and combinations, carry out implementation and application the technology of the present invention.
Drug used, reagent, the city instrument Jun Keyou in 'Shensu ' characteristic spectrum provided by the invention and its detection method
Field is bought.
Medicinal material of the present invention, the source of drug and reference substance source are as follows:
Codonopsis pilosula (producer: Hunan Tian Hong pharmaceutcal corporation, Ltd, lot number: YP1505016)；Perilla leaf medicinal material (producer: lake
Macro pharmaceutcal corporation, Ltd, Nan Tian, lot number: YP1505006)；Pueraria lobata medicinal material (producer: Hunan Tian Hong pharmaceutcal corporation, Ltd, lot number:
YP1505007)；Root of purple-flowered peucedanum medicinal material (producer: Hunan Tian Hong pharmaceutcal corporation, Ltd, lot number: YP1505008)；Poria cocos medicinal material (producer:
Hunan Tian Hong pharmaceutcal corporation, Ltd, lot number: YP1505014)；The tuber of pinellia (system) medicinal material (producer: Hunan Tian Hong pharmaceutcal corporation, Ltd,
Lot number: YP1505009)；Dried orange peel medicinal material (producer: Hunan Tian Hong pharmaceutcal corporation, Ltd, lot number: ZY1406025)；Fructus Aurantii (stir-fry) medicine
Material (producer: Hunan Tian Hong pharmaceutcal corporation, Ltd, lot number: YP1505010)；Campanulaceae medicinal material (producer: the limited public affairs of Hunan Liu Hong medicine company
Department, lot number: YP1505006)；Licorice medicinal materials (producer: Hunan Liu Hong pharmaceutcal corporation, Ltd, lot number: ZY1401003)；Radix aucklandiae medicine
Material (producer: Hunan Tian Hong pharmaceutcal corporation, Ltd, lot number: YP1505012)；Ginger medicinal material (producer: the macro limited public affairs of medicine company in Hunan day
Department, lot number: ZY1505013)；Jujube medicinal material (producer: Hunan Tian Hong pharmaceutcal corporation, Ltd, lot number: YP1505011)；It is natural clear
(producer: Tianjin day is at Science and Technology Ltd., lot number: 20150428) for clear agent component A；Natural clarifying agent B component (producer: Tianjin
It is at Science and Technology Ltd., lot number: 20150428)；Potassium sorbate (producer: Hunan that health pharmacy, lot number: 1054-
20150201)；Potassium sorbate reference substance (Chinese pharmaceutical biological product examines and determine research institute, lot number: 101075-200901)；Pueraria lobata
Plain reference substance (National Institute for Food and Drugs Control)；(the Chinese food drug assay research of 5- ethyoxyl person's base furfural reference substance
Institute)；Aurantiamarin reference substance (National Institute for Food and Drugs Control)；(the Chinese food drug assay research of aurantiin reference substance
Institute)；Neohesperidin reference substance (National Institute for Food and Drugs Control)；Shensu Oral Liquid" (Hu'nan Kangshou Pharmaceutical Co., Ltd.,
Lot number be 20130406,20131101,20141205,20141011,20140101,20140709,20140606,150601,
150301,150108)；Ginseng and Perillae Pill agent (Hu'nan Time Sun Pharmaceutical Co., Ltd., lot number 20120111,20141214,
Remaining 8 batches for laboratory make by oneself), ginseng Soviet Union tablet, capsule (laboratory self-control)；(Hunan Kang Shou pharmacy is limited for ginseng Soviet Union intermediate
Company, 150601).
The homemade 'Shensu ' of the present invention is mainly made by the crude drug of following portions by weight: 90 parts of Radix Codonopsis, 90 parts of purple perillas
Leaf, 90 parts of pueraria lobatas, 90 parts of roots of purple-flowered peucedanum, 90 parts of Poria cocos, 90 parts of tuber of pinellia (system), 60 parts of dried orange peels, 60 parts of Fructus Aurantiis (stir-fry), 60 parts of campanulaceaes, 60
Part Radix Glycyrrhizae, 60 parts of radix aucklandiaes, 36 portions of gingers, 36 portions of jujubes.
'Shensu ' is the 'Shensu ' as made from following methods:
1, when 'Shensu ' is oral solution, half summer grade, 70% ethanol percolation is extracted.Perilla leaf, dried orange peel, Fructus Aurantii, radix aucklandiae, life
Ginger is extracted volatile oil 3 hours, spare；The dregs of a decoction add water decoction 0.5 hour, merge gained filtrate, spare.It is Radix Codonopsis, pueraria lobata, preceding
Recklessly, Poria cocos, campanulaceae, Radix Glycyrrhizae, jujube add 80~90 DEG C of water Dynamic Extraction 2 hours, and filtrate is spare.The above decocting liquid is merged, it is dense
ZTC 1+1 natural clarifying agent is added in contracting, and it is 30% that ethyl alcohol to alcohol content, which is added, refrigerated overnight, alcohol precipitation filtrate and tuber of pinellia percolate
Merge, recycle ethyl alcohol, gained medical fluid and volatile oil, mix, add water to 1000ml to obtain the final product.
2, 'Shensu ' be pill, tablet, capsule when, by crude drug it is cataclasm after, by existing conventional method be made pill,
Tablet, capsule.It when 'Shensu ' is oral solution, mixes, measures 1.0~5.0ml；The preparation is pill, tablet, capsule
When agent, 7~9g of powder is weighed.
Below with reference to embodiment, the present invention is further explained:
The detection method of 1 'Shensu ' of embodiment
The detection method of 'Shensu ', includes the following steps:
(a) preparation of reference solution: weighing Puerarin object of reference, be placed in volumetric flask, adds 50%~100% methanol molten
Scale is solved and be settled to, is shaken up, reference solution is made；
(b) preparation of test solution: taking 'Shensu ', add 0%~50% methanol or 0%~50% ethyl alcohol 25~
125mL is extracted using conventional method, is filtered with miillpore filter, takes subsequent filtrate to get test solution；
(c) chromatographic condition: chromatographic column C18Reverse-phase chromatographic column；Using gradient elution, mobile phase is mobile phase A and mobile phase
The gradient eluent of B composition, mobile phase A are acetonitrile, and Mobile phase B is 0.05~0.1% phosphoric acid solution；Detection wavelength be 260~
300nm is shown in Fig. 1；25 DEG C~40 DEG C of column temperature；Flow velocity 0.8mL/min~1.2mL/min；Analysis time 60min.Gradient is such as
Shown in table 1.
1 gradient table of table
| Time (min) | Acetonitrile (%) | 0.05~0.1% phosphoric acid solution (%) |
| 0~5 | 1 | 99 |
| 5~13 | 1~8 | 99~92 |
| 13~15 | 8~12 | 92~88 |
| 15~36 | 12~14 | 88~86 |
| 36~38 | 14~17 | 86~83 |
| 38~60 | 17~26 | 83~74 |
(d) measure: precision draws reference solution and test solution injects high performance liquid chromatograph, according to high-efficient liquid phase color
Spectrometry measurement, obtains characteristic spectrum.
The foundation of 2 'Shensu ' oral solution characteristic spectrum of embodiment
(1) instrument and reagent
1260 high performance liquid chromatograph of Agilent, be contained in line vacuum degassing machine (G-1311C), binary pump (G-1311C),
Standard autosampler (G-1329B), intelligent column oven (G-1316A), DAD detector (G-1314B),
Agilent1260Infinity chromatographic work station (Anjelen Sci. & Tech. Inc, the U.S.)；SB-5200D ultrasonic washing instrument is (peaceful
The biotech inc Bo Xinzhi)；FAZ004B assay balance (section is helped in Shanghai)；BT125D electronic balance (Sai Duolisi
Scientific instrument (Beijing) Co., Ltd).
(2) chromatographic condition
Chromatographic column is Thermo Syncronis-C18(5 μm, 4.6 × 250mm) reverse-phase chromatographic column；Column temperature is 25 DEG C；Detection
Wavelength is 285nm；Mobile phase A is acetonitrile, and Mobile phase B is 0.08% phosphoric acid solution, overall flow rate 1.0mL/min；Analysis time
60 minutes；Gradient is as shown in table 2.
2 gradient table of table
| Time (min) | Acetonitrile (%) | 0.05% phosphoric acid solution (%) |
| 0~5 | 1 | 99 |
| 5~13 | 1~8 | 99~92 |
| 13~15 | 8~12 | 92~88 |
| 15~36 | 12~14 | 88~86 |
| 36~38 | 14~17 | 86~83 |
| 38~60 | 17~26 | 83~74 |
(3) preparation of reference solution
Puerarin object of reference is weighed, is placed in volumetric flask, adds methanol to dissolve and dilutes every 1mL is made containing the molten of 0.08mg
Liquid shakes up to get reference solution.See Fig. 2.
(4) preparation of test solution
Shensu Oral Liquid" 5 are taken, is mixed, precision measures 5mL in 50mL measuring bottle, is diluted with water to scale, shakes up, and filters
It crosses, takes subsequent filtrate to get Shensu Oral Liquid" test solution.
(5) analysis time
Sample introduction is investigated 120 minutes under above-mentioned chromatographic condition, the results showed that, in 60 minutes, all peaks in sample
It is eluted completely, therefore analysis time is determined as 60 minutes.
(6) methodological study
1. stability test
Same test solution is taken, under above-mentioned liquid phase chromatogram condition, was examined respectively at 0,2,4,6,12,24 hour
It surveys, each 10 μ L of sample introduction investigates the relative retention time of main chromatographic peak and the consistency of relative peak area.As a result main chromatography
The relative retention time at peak and the RSD < 3% of relative peak area show more stable in test solution 24 hours.
2. precision test
It takes with a test solution, under above-mentioned liquid phase chromatogram condition, repeats sample introduction 6 times, each 10 μ L of sample introduction, investigate
The main relative retention time of chromatographic peak and the consistency of relative peak area.As a result the relative retention time and phase of main chromatographic peak
To the RSD < 3% of peak area, show that instrument precision is good.
3. repetitive test
Same batch of sample is taken, prepares 6 parts of test solutions by preparation method of test article, under above-mentioned liquid phase chromatogram condition,
Sample introduction is analyzed, investigates the relative retention time of main chromatographic peak and the consistency of relative peak area.As a result the phase of main chromatographic peak
To the RSD < 3% of retention time and relative peak area, show that this method is reproducible.
(7) formulation of Shensu Oral Liquid" standard feature map
Precision draws above-mentioned reference substance solution and each 10 μ L of test solution, is injected separately into high performance liquid chromatograph, shines
High effective liquid chromatography for measuring records chromatogram；According to the characteristic spectrum of resulting 10 batches of Shensu Oral Liquid"s, characteristic pattern is formulated
Spectrum.See Fig. 3,4.
Compare 'Shensu ' chromatogram, characteristic spectrum has 7 characteristic peaks, wherein it is reference that No. 2 peaks, which are puerarin peak,
Peak, the relative retention time and relative standard deviation of each characteristic peak are as follows:
No. 1 peak: relative retention time 0.529 ± 0.026, relative standard deviation 0.20%；
No. 2 peaks: relative retention time 1.000 ± 0.000, relative standard deviation 0.00%；
No. 3 peaks: relative retention time 1.322 ± 0.066, relative standard deviation 0.13%；
No. 4 peaks: relative retention time 1.784 ± 0.089, relative standard deviation 0.18%；
No. 5 peaks: relative retention time 1.830 ± 0.091, relative standard deviation 0.18%；
No. 6 peaks: relative retention time 1.910 ± 0.095, relative standard deviation 0.19%；
No. 7 peaks: relative retention time 1.962 ± 0.098, relative standard deviation 0.19%；
(8) evaluation of each characteristic peak of characteristic spectrum.
'Shensu ' characteristic spectrum, which has, all has 7 characteristic peaks, by the opposite reservation of each feature of 'Shensu ' characteristic spectrum
Time is compared with each characteristic peak above-mentioned standard relative retention time of formulation, and relative retention time relative deviation should advise
Within ± the 5% of definite value.Concrete outcome see the table below.
3 10 batches of 'Shensu ' HPLC characteristic spectrum relative deviation result tables of table
The foundation of 3 'Shensu ' intermediate characteristic spectrum of embodiment
(1) instrument and reagent
1260 high performance liquid chromatograph of Agilent, be contained in line vacuum degassing machine (G-1311C), binary pump (G-1311C),
Standard autosampler (G-1329B), intelligent column oven (G-1316A), DAD detector (G-1314B),
Agilent1260Infinity chromatographic work station (Anjelen Sci. & Tech. Inc, the U.S.)；SB-5200D ultrasonic washing instrument is (peaceful
The biotech inc Bo Xinzhi)；FAZ004B assay balance (section is helped in Shanghai)；BT125D electronic balance (Sai Duolisi
Scientific instrument (Beijing) Co., Ltd).
(2) chromatographic condition
Chromatographic column is Thermo Syncronis-C18(5 μm, 4.6 × 250mm) reverse-phase chromatographic column；Column temperature is 30 DEG C；Detection
Wavelength is 283nm；Mobile phase A is acetonitrile, and Mobile phase B is 0.05% phosphoric acid solution, overall flow rate 1.0mL/min；Analysis time
60 minutes；Gradient is as shown in table 4.
4 gradient table of table
| Time (min) | Acetonitrile (%) | 0.05% phosphoric acid solution (%) |
| 0~5 | 1 | 99 |
| 5~13 | 1~8 | 99~92 |
| 13~15 | 8~12 | 92~88 |
| 15~36 | 12~14 | 88~86 |
| 36~38 | 14~17 | 86~83 |
| 38~60 | 17~26 | 83~74 |
(3) preparation of reference solution
Puerarin object of reference is weighed, is placed in volumetric flask, adds methanol to dissolve and dilutes every 1mL is made containing the molten of 0.08mg
Liquid shakes up to get reference solution.
(4) preparation of test solution
Precision measures 'Shensu ' intermediate 1mL in 10mL measuring bottle, is diluted with water to scale, shakes up, and filters, takes continuous filter
Liquid is to get 'Shensu ' intermediate test solution.
(5) methodological study
1. stability test
Same test solution is taken, under above-mentioned liquid phase chromatogram condition, was examined respectively at 0,2,4,6,12,24 hour
It surveys, each 10 μ L of sample introduction investigates the relative retention time of main chromatographic peak and the consistency of relative peak area.As a result main chromatography
The relative retention time at peak and the RSD < 3% of relative peak area show more stable in test solution 24 hours.
2. precision test
It takes with a test solution, under above-mentioned liquid phase chromatogram condition, repeats sample introduction 6 times, each 10 μ L of sample introduction, investigate
The main relative retention time of chromatographic peak and the consistency of relative peak area.As a result the relative retention time and phase of main chromatographic peak
To the RSD < 3% of peak area, show that instrument precision is good.
3. repetitive test
Same batch of sample is taken, prepares 6 parts of test solutions by preparation method of test article, under above-mentioned liquid phase chromatogram condition,
Sample introduction is analyzed, investigates the relative retention time of main chromatographic peak and the consistency of relative peak area.As a result the phase of main chromatographic peak
To the RSD < 3% of retention time and relative peak area, show that this method is reproducible.
(6) 'Shensu ' intermediate is formulated with the characteristic spectrum that Puerarin is reference peak.
Precision draws above-mentioned reference solution and each 10 μ L of test solution, is injected separately into high performance liquid chromatograph, shines
High effective liquid chromatography for measuring records chromatogram；According to the characteristic spectrum of resulting 10 batches of 'Shensu ' intermediates, standard is formulated
Characteristic spectrum.See Fig. 5,6.
Compare 'Shensu ' intermediate chromatogram, characteristic spectrum has 7 characteristic peaks, wherein No. 2 peaks are puerarin peak
For referring to peak, the relative retention time and relative standard deviation of each characteristic peak are as follows:
No. 1 peak: relative retention time 0.529 ± 0.026, relative standard deviation 0.12%；
No. 2 peaks: relative retention time 1.000 ± 0.000, relative standard deviation 0.00%；
No. 3 peaks: relative retention time 1.322 ± 0.066, relative standard deviation 0.15%；
No. 4 peaks: relative retention time 1.784 ± 0.089, relative standard deviation 0.10%；
No. 5 peaks: relative retention time 1.830 ± 0.091, relative standard deviation 0.16%；
No. 6 peaks: relative retention time 1.910 ± 0.095, relative standard deviation 0.14%；
No. 7 peaks: relative retention time 1.962 ± 0.098, relative standard deviation 0.14%；
(7) evaluation of each characteristic peak of characteristic spectrum.
'Shensu ' characteristic spectrum, which has, all has 7 characteristic peaks, by the opposite reservation of each feature of 'Shensu ' characteristic spectrum
Time is compared with each characteristic peak standard relative retention time of formulation, and relative retention time relative deviation should be in specified value
± 5% within.Concrete outcome see the table below.
5 10 batches of 'Shensu ' intermediate HPLC characteristic spectrum relative deviation result tables of table
The foundation of 4 'Shensu ' characteristic spectrum of embodiment (by taking 'Shensu ' capsule as an example)
(1) instrument and reagent
1260 high performance liquid chromatograph of Agilent, be contained in line vacuum degassing machine (G-1311C), binary pump (G-1311C),
Standard autosampler (G-1329B), intelligent column oven (G-1316A), DAD detector (G-1314B),
Agilent1260Infinity chromatographic work station (Anjelen Sci. & Tech. Inc, the U.S.)；SB-5200D ultrasonic washing instrument is (peaceful
The biotech inc Bo Xinzhi)；FAZ004B assay balance (section is helped in Shanghai)；BT125D electronic balance (Sai Duolisi
Scientific instrument (Beijing) Co., Ltd).
(2) chromatographic condition
Chromatographic column is Thermo Syncronis-C18(5 μm, 4.6 × 250mm) reverse-phase chromatographic column；Column temperature is 25 DEG C；Detection
Wavelength is 300nm；Mobile phase A is acetonitrile, and Mobile phase B is 0.1% phosphoric acid solution, overall flow rate 0.8mL/min；Analysis time 60
Minute；Gradient is as shown in table 6.
6 gradient table of table
| Time (min) | Acetonitrile (%) | 0.10 phosphoric acid solution (%) |
| 0~5 | 1 | 99 |
| 5~13 | 1~8 | 99~92 |
| 13~15 | 8~12 | 92~88 |
| 15~36 | 12~14 | 88~86 |
| 36~38 | 14~17 | 86~83 |
| 38~60 | 17~26 | 83~74 |
(3) preparation of reference solution
Puerarin object of reference is weighed, is placed in volumetric flask, adds methanol to dissolve and dilutes every 1mL is made containing the molten of 0.08mg
Liquid shakes up to get reference solution.
(4) preparation of test solution
Ginseng Soviet Union capsule powders 7g is taken, it is accurately weighed, it sets in stuffed conical flask, water 100mL is added in precision, and close plug is weighed heavy
Amount, is heated to reflux 1 hour, lets cool, then weighed weight, the weight of less loss is supplied with water, is shaken up, and filters, takes subsequent filtrate to get ginseng
Soviet Union's capsule test solution.
(5) methodological study
1. stability test
Same test solution is taken, under above-mentioned liquid phase chromatogram condition, was examined respectively at 0,2,4,6,12,24 hour
It surveys, each 10 μ L of sample introduction investigates the relative retention time of main chromatographic peak and the consistency of relative peak area.As a result main chromatography
The relative retention time at peak and the RSD < 3% of relative peak area show more stable in test solution 24 hours.
2. precision test
It takes with a test solution, under above-mentioned liquid phase chromatogram condition, repeats sample introduction 6 times, each 10 μ L of sample introduction, investigate
The main relative retention time of chromatographic peak and the consistency of relative peak area.As a result the relative retention time and phase of main chromatographic peak
To the RSD < 3% of peak area, show that instrument precision is good.
3. repetitive test
Same batch of sample is taken, prepares 6 parts of test solutions by preparation method of test article, under above-mentioned liquid phase chromatogram condition,
Sample introduction is analyzed, investigates the relative retention time of main chromatographic peak and the consistency of relative peak area.As a result the phase of main chromatographic peak
To the RSD < 3% of retention time and relative peak area, show that this method is reproducible.
(6) formulation of 'Shensu ' characteristic spectrum.
Precision draws above-mentioned reference solution and each 10 μ L of test solution, is injected separately into high performance liquid chromatograph, shines
High effective liquid chromatography for measuring records chromatogram；According to the characteristic spectrum of resulting 10 batches of 'Shensu 's, characteristic spectrum is formulated.
See Fig. 7,8.
Compare 'Shensu ' chromatogram, characteristic spectrum has 7 characteristic peaks, wherein it is reference that No. 2 peaks, which are puerarin peak,
Peak, the relative retention time and relative standard deviation of each characteristic peak are as follows:
No. 1 peak: relative retention time 0.529 ± 0.026, relative standard deviation 0.15%；
No. 2 peaks: relative retention time 1.000 ± 0.000, relative standard deviation 0.00%；
No. 3 peaks: relative retention time 1.322 ± 0.066, relative standard deviation 0.18%；
No. 4 peaks: relative retention time 1.784 ± 0.089, relative standard deviation 0.08%；
No. 5 peaks: relative retention time 1.830 ± 0.091, relative standard deviation 0.21%；
No. 6 peaks: relative retention time 1.910 ± 0.095, relative standard deviation 0.17%；
No. 7 peaks: relative retention time 1.962 ± 0.098, relative standard deviation 0.19%；
(7) evaluation of each characteristic peak of characteristic spectrum.
'Shensu ' characteristic spectrum, which has, all has 7 characteristic peaks, by the opposite reservation of each feature of 'Shensu ' characteristic spectrum
Time is compared with each characteristic peak standard relative retention time of formulation, and relative retention time relative deviation should be in specified value
± 5% within.Concrete outcome see the table below.
7 10 batches of ginsengs Soviet Union capsule HPLC characteristic spectrum relative deviation result tables of table
The foundation of 5 'Shensu ' characteristic spectrum of embodiment (by taking ginseng revives tablet as an example)
(1) instrument and reagent
1260 high performance liquid chromatograph of Agilent, be contained in line vacuum degassing machine (G-1311C), binary pump (G-1311C),
Standard autosampler (G-1329B), intelligent column oven (G-1316A), DAD detector (G-1314B),
Agilent1260Infinity chromatographic work station (Anjelen Sci. & Tech. Inc, the U.S.)；SB-5200D ultrasonic washing instrument is (peaceful
The biotech inc Bo Xinzhi)；FAZ004B assay balance (section is helped in Shanghai)；BT125D electronic balance (Sai Duolisi
Scientific instrument (Beijing) Co., Ltd).
(2) chromatographic condition
Chromatographic column is Shimadzu Inertsil ODS-2 (5 μm, 4.6 × 250mm) reverse-phase chromatographic column；Column temperature is 40 DEG C；Detection
Wavelength is 260nm；Mobile phase A is acetonitrile, and Mobile phase B is 0.05% phosphoric acid solution, overall flow rate 1.2mL/min；Analysis time
60 minutes；Gradient is as shown in table 8.
8 gradient table of table
| Time (min) | Acetonitrile (%) | 0.05% phosphoric acid solution (%) |
| 0~5 | 1 | 99 |
| 5~13 | 1~8 | 99~92 |
| 13~15 | 8~12 | 92~88 |
| 15~36 | 12~14 | 88~86 |
| 36~38 | 14~17 | 86~83 |
| 38~60 | 17~26 | 83~74 |
(3) preparation of reference substance solution
Puerarin reference substance is weighed, is placed in volumetric flask, adds methanol to dissolve and dilutes every 1mL is made containing the molten of 0.08mg
Liquid shakes up to get reference substance solution.
(4) preparation of test solution
Ginseng Soviet Union tablet powder 9g is taken, it is accurately weighed, it sets in stuffed conical flask, 50% methanol 25mL is added in precision, and close plug claims
Determine weight, be heated to reflux 2 hours, let cool, then weighed weight, the weight of less loss is supplied with water, is shaken up, filters, take subsequent filtrate, i.e.,
Soviet Union's tablet test solution must be joined.
(5) methodological study
1. stability test
Same test solution is taken, under above-mentioned liquid phase chromatogram condition, was examined respectively at 0,2,4,6,12,24 hour
It surveys, each 10 μ L of sample introduction investigates the relative retention time of main chromatographic peak and the consistency of relative peak area.As a result main chromatography
The relative retention time at peak and the RSD < 3% of relative peak area show more stable in test solution 24 hours.
2. precision test
It takes with a test solution, under above-mentioned liquid phase chromatogram condition, repeats sample introduction 6 times, each 10 μ L of sample introduction, investigate
The main relative retention time of chromatographic peak and the consistency of relative peak area.As a result the relative retention time and phase of main chromatographic peak
To the RSD < 3% of peak area, show that instrument precision is good.
3. repetitive test
Same batch of sample is taken, prepares 6 parts of test solutions by preparation method of test article, under above-mentioned liquid phase chromatogram condition,
Sample introduction is analyzed, investigates the relative retention time of main chromatographic peak and the consistency of relative peak area.As a result the phase of main chromatographic peak
To the RSD < 3% of retention time and relative peak area, show that this method is reproducible.
(6) formulation of 'Shensu ' particle characteristic map.
Precision draws above-mentioned reference solution and each 10 μ L of test solution, is injected separately into high performance liquid chromatograph, shines
High effective liquid chromatography for measuring records chromatogram；According to the characteristic spectrum of resulting 10 batches of 'Shensu 's, characteristic spectrum is formulated.
See Fig. 9,10.
Compare 'Shensu ' particulate chromatography figure, characteristic spectrum has 7 characteristic peaks, wherein No. 2 peaks are that puerarin peak is
Referring to peak, the relative retention time and relative standard deviation of each characteristic peak are as follows:
No. 1 peak: relative retention time 0.529 ± 0.026, relative standard deviation 0.16%；
No. 2 peaks: relative retention time 1.000 ± 0.000, relative standard deviation 0.00%；
No. 3 peaks: relative retention time 1.322 ± 0.066, relative standard deviation 0.10%；
No. 4 peaks: relative retention time 1.784 ± 0.089, relative standard deviation 0.02%；
No. 5 peaks: relative retention time 1.830 ± 0.091, relative standard deviation 0.14%；
No. 6 peaks: relative retention time 1.910 ± 0.095, relative standard deviation 0.10%；
No. 7 peaks: relative retention time 1.962 ± 0.098, relative standard deviation 0.09%；
(7) evaluation of each characteristic peak of characteristic spectrum.
'Shensu ' characteristic spectrum, which has, all has 7 characteristic peaks, by the opposite reservation of each feature of 'Shensu ' characteristic spectrum
Time is compared with each characteristic peak standard relative retention time of formulation, and relative retention time relative deviation should be in specified value
± 5% within.Concrete outcome see the table below.
9 10 batches of ginsengs Soviet Union tablet HPLC characteristic spectrum relative deviation result tables of table
The foundation of 6 'Shensu ' characteristic spectrum of embodiment (by taking ginseng and Perillae Pill agent as an example)
(1) instrument and reagent
1260 high performance liquid chromatograph of Agilent, be contained in line vacuum degassing machine (G-1311C), binary pump (G-1311C),
Standard autosampler (G-1329B), intelligent column oven (G-1316A), DAD detector (G-1314B),
Agilent1260Infinity chromatographic work station (Anjelen Sci. & Tech. Inc, the U.S.)；SB-5200D ultrasonic washing instrument is (peaceful
The biotech inc Bo Xinzhi)；FAZ004B assay balance (section is helped in Shanghai)；BT125D electronic balance (Sai Duolisi
Scientific instrument (Beijing) Co., Ltd).
(2) chromatographic condition
Chromatographic column is Unitary C18(5 μm, 4.6 × 250mm) reverse-phase chromatographic column；Column temperature is 35 DEG C；Detection wavelength is
290nm；Mobile phase A is acetonitrile, and Mobile phase B is 0.05% phosphoric acid solution, overall flow rate 1.0mL/min；Analysis time 60 minutes；
Gradient is as shown in table 10.
10 gradient table of table
| Time (min) | Acetonitrile (%) | 0.05% phosphoric acid solution (%) |
| 0~5 | 1 | 99 |
| 5~13 | 1~8 | 99~92 |
| 13~15 | 8~12 | 92~88 |
| 15~36 | 12~14 | 88~86 |
| 36~38 | 14~17 | 86~83 |
| 38~60 | 17~26 | 83~74 |
(3) preparation of reference substance solution
Puerarin reference substance is weighed, is placed in volumetric flask, adds methanol to dissolve and dilutes every 1mL is made containing the molten of 0.08mg
Liquid shakes up to get reference substance solution.
(4) preparation of test solution
Ginseng and Perillae Pill is taken, crushes, takes 8g, it is accurately weighed, it sets in stuffed conical flask, precision 50% ethyl alcohol 50mL of addition, close plug,
Weighed weight reflow treatment 2 hours, is let cool, then weighed weight, and the weight of less loss is supplied with water, is shaken up, and filtration takes subsequent filtrate,
Up to Shensu Oral Liquid" test solution.
(5) methodological study
1. stability test
Same test solution is taken, under above-mentioned liquid phase chromatogram condition, was examined respectively at 0,2,4,6,12,24 hour
It surveys, each 10 μ L of sample introduction investigates the relative retention time of main chromatographic peak and the consistency of relative peak area.As a result main chromatography
The relative retention time at peak and the RSD < 3% of relative peak area show more stable in test solution 24 hours.
2. precision test
It takes with a test solution, under above-mentioned liquid phase chromatogram condition, repeats sample introduction 6 times, each 10 μ L of sample introduction, investigate
The main relative retention time of chromatographic peak and the consistency of relative peak area.As a result the relative retention time and phase of main chromatographic peak
To the RSD < 3% of peak area, show that instrument precision is good.
3. repetitive test
Same batch of sample is taken, prepares 6 parts of test solutions by preparation method of test article, under above-mentioned liquid phase chromatogram condition,
Sample introduction is analyzed, investigates the relative retention time of main chromatographic peak and the consistency of relative peak area.As a result the phase of main chromatographic peak
To the RSD < 3% of retention time and relative peak area, show that this method is reproducible.
(6) formulation of 'Shensu ' oral solution characteristic spectrum.
Precision draws above-mentioned reference solution and each 10 μ L of test solution, is injected separately into high performance liquid chromatograph, shines
High effective liquid chromatography for measuring records chromatogram；According to the characteristic spectrum of resulting 10 batches of 'Shensu 's, characteristic spectrum is formulated.
See Figure 11,12.
Compare 'Shensu ' oral solution chromatogram, characteristic spectrum has 7 characteristic peaks, wherein No. 2 peaks are puerarin peak
For referring to peak, the relative retention time and relative standard deviation of each characteristic peak are as follows:
No. 1 peak: relative retention time 0.529 ± 0.026, relative standard deviation 0.21%；
No. 2 peaks: relative retention time 1.000 ± 0.000, relative standard deviation 0.00%；
No. 3 peaks: relative retention time 1.322 ± 0.066, relative standard deviation 0.08%；
No. 4 peaks: relative retention time 1.784 ± 0.089, relative standard deviation 0.06%；
No. 5 peaks: relative retention time 1.830 ± 0.091, relative standard deviation 0.13%；
No. 6 peaks: relative retention time 1.910 ± 0.095, relative standard deviation 0.14%；
No. 7 peaks: relative retention time 1.962 ± 0.098, relative standard deviation 0.14%；
(7) evaluation of each characteristic peak of characteristic spectrum.
'Shensu ' characteristic spectrum, which has, all has 7 characteristic peaks, by the opposite reservation of each feature of 'Shensu ' characteristic spectrum
Time is compared with each characteristic peak standard relative retention time of formulation, and relative retention time relative deviation should be in specified value
± 5% within.Concrete outcome see the table below.
11 10 batches of ginsengs Soviet Union tablet HPLC characteristic spectrum relative deviation result tables of table
7 finished product of embodiment and the correlation and characteristic peak of each raw medicinal material belong to
(1) instrument and reagent
1260 high performance liquid chromatograph of Agilent, be contained in line vacuum degassing machine (G-1311C), binary pump (G-1311C),
Standard autosampler (G-1329B), intelligent column oven (G-1316A), DAD detector (G-1314B),
Agilent1260Infinity chromatographic work station (Anjelen Sci. & Tech. Inc, the U.S.)；SB-5200D ultrasonic washing instrument is (peaceful
The biotech inc Bo Xinzhi)；FAZ004B assay balance (section is helped in Shanghai)；BT125D electronic balance (Sai Duolisi
Scientific instrument (Beijing) Co., Ltd).
(2) chromatographic condition
Chromatographic column is Thermo Syncronis-C18(5 μm, 4.6 × 250mm) reverse-phase chromatographic column；Column temperature is 30 DEG C；Detection
Wavelength is 280nm；Mobile phase A is acetonitrile, and Mobile phase B is 0.05% phosphoric acid solution, overall flow rate 1.0mL/min；Analysis time
60 minutes；Gradient is as shown in table 12.
12 gradient table of table
| Time (min) | Acetonitrile (%) | 0.05% phosphoric acid solution (%) |
| 0~5 | 1 | 99 |
| 5~13 | 1~8 | 99~92 |
| 13~15 | 8~12 | 92~88 |
| 15~36 | 12~14 | 88~86 |
| 36~38 | 14~17 | 86~83 |
| 38~60 | 17~26 | 83~74 |
(3) preparation of reference solution
Puerarin reference substance is weighed, is placed in volumetric flask, adds methanol to dissolve and dilutes every 1mL is made containing the molten of 0.08mg
Liquid shakes up to get reference substance solution.
(4) preparation of test solution
Shensu Oral Liquid" 5 are taken, is mixed, precision measures 5mL in 50mL measuring bottle, is diluted with water to scale, shakes up, and filters
It crosses, takes subsequent filtrate to get Shensu Oral Liquid" test solution.
The preparation method of medicinal material solution: each single medicinal material is taken respectively, is prepared into solution by prescription, by 'Shensu ' intermediate
Sample solution preparation method prepares corresponding single medicinal material solution, totally 13 taste medicinal material solution.
The preparation method of negative medicinal material solution: other medicinal materials of wherein certain taste medicinal material are removed respectively, are prepared by prescription molten
Liquid prepares corresponding negative medicinal material solution by 'Shensu ' intermediate sample solution preparation method, and totally 13 taste feminine gender medicinal materials are molten
Liquid.
(5) formulation of each characteristic peak map ownership of single medicinal material, feminine gender, finished product.
Precision draws above-mentioned reference solution and each 10 μ L of test solution, is injected separately into high performance liquid chromatograph, shines
High effective liquid chromatography for measuring records chromatogram；According to resulting map, the ownership of each characteristic peak is formulated.13 are shown in Table, attached drawing
13。
Each characteristic peak map of table 13 ownership
The HPLC of the 'Shensu ' has 13 shared peaks (see Figure 14) in the characteristic spectrum of 283nm, wherein belonging to trifoliate orange
The shared peak of shell are as follows: peak 1, peak 2, peak 10, peak 11, peak 12, peak 13；Belong to the shared peak of dried orange peel are as follows: peak 2, peak 10, peak 12；
Belong to the shared peak of pueraria lobata are as follows: peak 3, peak 4, peak 5, peak 6, peak 7；Belong to the shared peak of perilla leaf are as follows: peak 9.Wherein choose
The shared peak that retention time and peak area are suitable for is characterized peak and establishes characteristic spectrum, wherein having: peak 2, peak 4 (S), peak 7, peak 9, peak
10, peak 11, peak 12.
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 (8)
1. a kind of detection method of 'Shensu ', which comprises the steps of:
Test solution is made in 'Shensu ', Puerarin receives ginseng Soviet Union as object of reference, using high performance liquid chromatography detection
The testing result of preparation, chromatographic condition are as follows:
Chromatographic column is C18Reverse-phase chromatographic column, mobile phase are the gradient eluent of mobile phase A and Mobile phase B composition, and mobile phase A is second
Nitrile, Mobile phase B are phosphoric acid solution, and the volumn concentration of phosphoric acid is 0.05%~0.1% in the phosphoric acid solution, using gradient
Elution, gradient elution program are as follows:
2. detection method according to claim 1, which is characterized in that the Detection wavelength of the high performance liquid chromatography is 260
~300nm.
3. detection method according to claim 1, which is characterized in that the chromatographic column is Unitary C18Or Thermo
Syncronis-C18Or Shimadzu InertsilODS-2, the column temperature of the high performance liquid chromatography are 25 DEG C~40 DEG C.
4. detection method according to claim 1, which is characterized in that 0.8~1.2mL/min of flow velocity of the mobile phase.
5. detection method according to claim 1, which is characterized in that the 'Shensu ' is oral solution, test solution
Preparation method using direct diluted method.
6. detection method according to claim 1, which is characterized in that the 'Shensu ' be pill, tablet or capsule,
The method that the preparation method of test solution uses refluxing extraction.
7. detection method according to claim 6, which is characterized in that the solvent of the refluxing extraction is water, volume basis
Content is not more than 50% ethanol water no more than 50% methanol aqueous solution or volumn concentration.
8. detection method according to claim 1, which is characterized in that the analysis time of the high performance liquid chromatography is not less than
60min。
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101474274A (en) * | 2008-12-24 | 2009-07-08 | 贵州信邦制药股份有限公司 | Quality control method of Shensu Chinese medicine preparation |
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2015
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| CN101474274A (en) * | 2008-12-24 | 2009-07-08 | 贵州信邦制药股份有限公司 | Quality control method of Shensu Chinese medicine preparation |
| CN102183590A (en) * | 2011-01-31 | 2011-09-14 | 山东沃华医药科技股份有限公司 | Measuring method of Xinkeshu preparation finger-print |
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