CN1947740A - Fingerprint spectrum quality detection method for lamiophlomis rotata medicinal material, intermediate and injection thereof - Google Patents
Fingerprint spectrum quality detection method for lamiophlomis rotata medicinal material, intermediate and injection thereof Download PDFInfo
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- 238000002347 injection Methods 0.000 title claims abstract description 49
- 238000001514 detection method Methods 0.000 title claims abstract description 17
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- 239000000543 intermediate Substances 0.000 title abstract description 33
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- 238000001228 spectrum Methods 0.000 title abstract description 3
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- 238000001914 filtration Methods 0.000 claims abstract description 19
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- Investigating Or Analysing Biological Materials (AREA)
Abstract
The invention discloses a fingerprint spectrum quality detection method of lamiophlomis rotata medicinal materials, intermediates and injection thereof. The method comprises the following steps of chromatographic condition and system applicability test: using octadecylsilane chemically bonded silica as a filler to carry out gradient elution; taking quercetin reference substance, adding methanol to make into 40 μ g/ml solution as reference solution; preparing a lamiophlomis rotata medicinal material test solution: extracting the medicinal powder under reflux, filtering, evaporating water, dissolving the residue in methanol, and centrifuging; adding the supernatant into the reference solution, adding methanol to desired volume, shaking, filtering, and collecting filtrate; preparing an intermediate and an injection test solution thereof: adding reference solution into Lamiophlomis rotata injection intermediate or injection, diluting with methanol, shaking, filtering, and collecting filtrate; the determination method comprises the following steps: sucking reference substance solution and test solution, injecting into liquid chromatograph, and measuring; the similarity between the test sample fingerprint and the comparison fingerprint is more than 0.90.
Description
Technical field
The present invention relates to the quality determining method of Chinese crude drug, intermediate and injection thereof, particularly the finger print quality detecting method of Radix Lamiophlomidis Rotatae medical material, intermediate and injection thereof.
Background technology
Main chemical constituent mainly contains flavones ingredient and becomes to grade with some fatty acids with the iridoid constituents in the Radix Lamiophlomidis Rotatae.Flavones ingredient mainly contains luteolin, luteolin-7-o-glucoside, Quercetin, Quercetin-3-o-galactoside, compositions such as apigenin.The quality standard of Chinese medicine is very lack of standardization and perfect at present, and the means of quality analysis and evaluation are also very backward, can't reflect the quality condition of existing Chinese crude drug and finished product accurately.Chinese medicine fingerprint is a notion of using for reference fingerprint identification in the prudence, through suitably handling, obtains reflecting the total chromatographic peak of Chinese medicine feature with certain analysis means, identifies the true and false of Chinese medicine by having chromatographic peak, guarantees the quality of Chinese medicine.Simultaneously do quantitative assay, not only can carry out true and false discriminating, also allow the intrinsic index components of Chinese medicine quantizes, standardization simultaneously raw medicinal material in conjunction with the content of effective in the Chinese medicine.
By relatively screening, because of Quercetin appearance time in finger printing relatively stable, and there is not chromatographic peak to occur at this time period test sample, can be used as " scale " and demarcate each characteristic peak, so the present invention use high performance liquid chromatography with Quercetin as internal standard substance matter, carry out the finger printing research of Chinese crude drug, intermediate, finished product and set up corresponding reference fingerprint, and with this important indicator as control crude drug and end product quality.
Though in No. 200410083678.7 patent applications, also related to the correlation technique content of Radix Lamiophlomidis Rotatae finger printing, but this patent application has been introduced macroporous adsorbent resin and carried out pre-treatment in the preparation process of medical material, intermediate and injection test sample, thereby may cause the influence of at least two aspects to final result: first macroporous adsorbent resin inevitably can be introduced new material in the process of absorption and eluting; It two is macroporous adsorbent resins in the process of handling, and the related substance in the test sample is adsorbed.Can not reflect real fingerprint characteristic.
Summary of the invention
The object of the invention is to provide the finger print quality detecting method of Radix Lamiophlomidis Rotatae medical material or lamivphlomis root injection liquid intermediate and injection thereof.
The present invention seeks to be achieved through the following technical solutions
The finger print quality detecting method of Radix Lamiophlomidis Rotatae medical material of the present invention, this method comprises the steps:
Chromatographic condition and system suitability test: with octadecylsilane chemically bonded silica is filler; Flow velocity: 1ml/min; The detection wavelength is 254nm; Column temperature: 30 ℃; Analysis time: 1 hour; Carry out gradient elution: preceding 10 minutes, carry out eluting by mobile phase by 10: 90 proportionate relationship of acetonitrile-0.2% phosphate aqueous solution, mobile phase becomes acetonitrile-0.2% phosphate aqueous solution 15: 85 by acetonitrile-0.2% phosphate aqueous solution at 10: 90; 10-40 minute, mobile phase became acetonitrile-0.2% phosphate aqueous solution 30: 70 by acetonitrile-0.2% phosphate aqueous solution at 15: 85; 40-50 minute, mobile phase became acetonitrile-0.2% phosphate aqueous solution 40: 60 by acetonitrile-0.2% phosphate aqueous solution at 30: 70; 50-60 minute, mobile phase remained acetonitrile-0.2% phosphate aqueous solution 40: 60; The preparation of object of reference solution: get the Quercetin reference substance, add the solution that methanol is made 40 μ g/ml, as object of reference solution; The preparation of need testing solution: get and cross the Radix Lamiophlomidis Rotatae medicinal powder 1-10 weight portion that internal diameter is the 5mm sieve after pulverizing, preferred 5 weight portions, put in the flask, add water 50-200 parts by volume, preferred 100 parts by volume, reflux, extract, 1-3 hour, preferred 2 hours, filter water liquid evaporate to dryness, residue adds methanol and dissolves in right amount, 1000r/min is centrifugal, in the supernatant dislocation 20 parts by volume volumetric flasks, adds object of reference solution 0.4-4 parts by volume, preferred 2 parts by volume, methanol constant volume shakes up to scale, with 0.45 μ m filtering with microporous membrane, get filtrate, promptly;
Algoscopy: draw each 10 μ l of object of reference solution and need testing solution, inject chromatograph of liquid, measure; Test sample finger printing and reference fingerprint similarity are greater than 0.90;
The relative retention time of the total fingerprint peaks of Radix Lamiophlomidis Rotatae medical material is respectively: No. 1 peak: 0.145 ± 0.002, No. 2 peaks: 0.154 ± 0.002, No. 3 peaks: 0.199 ± 0.001, No. 4 peaks: 0.220 ± 0.001, No. 5 peaks: 0.231 ± 0.001, No. 6 peaks: 0.242 ± 0.001, No. 7 peaks: 0.260 ± 0.002, No. 8 peaks: 0.298 ± 0.003, No. 9 peaks: 0.313 ± 0.002, No. 10 peaks: 0.336 ± 0.003, No. 11 peaks: 0.369 ± 0.003, No. 12 peaks: 0.386 ± 0.004, No. 13 peaks: 0.402 ± 0.001, No. 14 peaks: 0.414 ± 0.002, No. 15 peaks: 0.476 ± 0.006, No. 16 peaks: 0.484 ± 0.004, No. 17 peaks: 0.533 ± 0.003, No. 18 peaks: 0.550 ± 0.001, No. 19 peaks: 0.585 ± 0.002, No. 20 peaks: 0.642 ± 0.005, No. 21 peaks: 0.661 ± 0.003, No. 22 peaks: 0.760 ± 0.001, No. 23 peaks: 0.796 ± 0.007, No. 24 peaks are the S peak: 1, No. 25 peak: 1.077 ± 0.012; Relative peak area ratio is respectively: No. 1 peak: 0.136 ± 0.100, No. 2 peaks: 1.648 ± 0.564, No. 3 peaks: 1.745 ± 0.653, No. 4 peaks: 0.062 ± 0.031, No. 5 peaks: 0.728 ± 0.247, No. 6 peaks: 0.370 ± 0.141, No. 7 peaks: 0.230 ± 0.112, No. 8 peaks: 0.116 ± 0.048, No. 9 peaks: 0.815 ± 0.262, No. 10 peaks: 0.090 ± 0.025, No. 11 peaks: 0.132 ± 0.107, No. 12 peaks: 0.228 ± 0.140, No. 13 peaks: 0.154 ± 0.193, No. 14 peaks: 0.142 ± 0.183, No. 15 peaks: 1.752 ± 0.455, No. 16 peaks: 2.508 ± 1.357, No. 17 peaks: 0.541 ± 0.379, No. 18 peaks: 0.179 ± 0.152, No. 19 peaks: 2.508 ± 2.437, No. 20 peaks: 0.202 ± 0.109, No. 21 peaks: 0.330 ± 0.262, No. 22 peaks: 0.349 ± 0.137, No. 23 peaks: 0.206 ± 0.046, No. 24 peaks are the S peak: 1, No. 25 peak: 0.205 ± 0.077.
The pass of above-mentioned parts by volume and weight portion is milliliter and the relation that restrains.
The finger print quality detecting method of lamivphlomis root injection liquid intermediate of the present invention, this method comprises the steps:
Chromatographic condition and system suitability test: with octadecylsilane chemically bonded silica is filler; Flow velocity: 1ml/min; The detection wavelength is 254nm; Column temperature: 30 ℃; Analysis time: 1 hour; Carry out gradient elution: preceding 10 minutes, carry out eluting by mobile phase by 10: 90 proportionate relationship of acetonitrile-0.2% phosphate aqueous solution, mobile phase becomes acetonitrile-0.2% phosphate aqueous solution 15: 85 by acetonitrile-0.2% phosphate aqueous solution at 10: 90; 10-40 minute, mobile phase became acetonitrile-0.2% phosphate aqueous solution 30: 70 by acetonitrile-0.2% phosphate aqueous solution at 15: 85; 40-50 minute, mobile phase became acetonitrile-0.2% phosphate aqueous solution 40: 60 by acetonitrile-0.2% phosphate aqueous solution at 30: 70; 50-60 minute, mobile phase remained acetonitrile-0.2% phosphate aqueous solution 40: 60; The preparation of object of reference solution: get the Quercetin reference substance, add the solution that methanol is made 40 μ g/ml, as object of reference solution; The preparation of need testing solution: get lamivphlomis root injection liquid intermediate 0.5-2 parts by volume, preferred 1 parts by volume, put in the volumetric flask of 10 parts by volume, add object of reference solution 0.5-2 parts by volume, preferred 1 parts by volume, methanol is diluted to 10 parts by volume, shake up, with 0.45 μ m filtering with microporous membrane, get filtrate, promptly;
Algoscopy: draw each 10 μ l of object of reference solution and need testing solution, inject chromatograph of liquid, measure; Test sample finger printing and reference fingerprint similarity are greater than 0.90;
The relative retention time of the total fingerprint peaks of lamivphlomis root injection liquid intermediate is respectively: No. 1 peak: 0.143 ± 0.001, No. 2 peaks: 0.153 ± 0.001, No. 3 peaks: 0.165 ± 0.001, No. 4 peaks: 0.198 ± 0.001, No. 5 peaks: 0.217 ± 0.001, No. 6 peaks: 0.230 ± 0.001, No. 7 peaks: 0.242 ± 0.002, No. 8 peaks: 0.260 ± 0.001, No. 9 peaks: 0.312 ± 0.001, No. 10 peaks: 0.391 ± 0.008, No. 11 peaks: 0.475 ± 0.002, No. 12 peaks: 0.484 ± 0.002, No. 13 peaks: 0.532 ± 0.001, No. 14 peaks: 0.554 ± 0.004, No. 15 peaks: 0.586 ± 0.001, No. 16 peaks: 0.640 ± 0.003, No. 17 peaks: 0.660 ± 0.002, No. 18 peaks: 0.717 ± 0.001, No. 19 the peak is the S peak: 1, No. 20 peak: 1.070 ± 0.007; The relative peak area ratio of total fingerprint peaks is respectively: No. 1 peak: 0.029 ± 0.019, No. 2 peaks: 0.384 ± 0.138, No. 3 peaks: 0.028 ± 0.013, No. 4 peaks: 0.260 ± 0.013, No. 5 peaks: 0.094 ± 0.005, No. 6 peaks: 0.121 ± 0.004, No. 7 peaks: 0.058 ± 0.007, No. 8 peaks: 0.086 ± 0.010, No. 9 peaks: 0.127 ± 0.003, No. 10 peaks: 0.037 ± 0.001, No. 11 peaks: 0.352 ± 0.014, No. 12 peaks: 0.619 ± 0.026, No. 13 peaks: 0.119 ± 0.012, No. 14 peaks: 0.032 ± 0.006, No. 15 peaks: 1.402 ± 0.054, No. 16 peaks: 0.051 ± 0.006, No. 17 peaks: 0.183 ± 0.010, No. 18 peaks: 0.066 ± 0.042, No. 19 the peak is the S peak: 1, No. 20 peak: 0.036 ± 0.005.
Above-mentioned lamivphlomis root injection liquid intermediates preparation is as follows, but is not limited to this method:
Get the Radix Lamiophlomidis Rotatae medical material, decoct with water three times, the water logging that adds 15~25 times was for the first time steeped 0.5~1 hour, decocted 0.5~1.5 hour, and respectively added 10~20 times of water gagings for the second time, for the third time, decocted respectively 0.5~1.5 hour, filter, merging filtrate is concentrated into liquor strength and is 1: 1~2 concentrated solution; Add ethanol and make and contain alcohol amount and reach 60%~80%, in 5~8 ℃ of cold preservations 12~24 hours, filter, filtrate adds ethanol again to be made and contains the alcohol amount and reach 80%~90%, in 5~8 ℃ of cold preservations 12~24 hours, filters; Filtrate is transferred PH to 8~9 with 30%~50% sodium hydroxide, in 5~8 ℃ of cold preservations 12~24 hours, filters; Filtrate adds 10%~30% sulphuric acid and transfers PH to 5~6, placed 1~3 hour, and filtered, reclaim ethanol to there not being the alcohol flavor, adding water for injection is diluted in right amount, 5~8 ℃ of cold preservation 12~24 hours filters, and filtrate adds active carbon 0.05%~0.25%, boiled 15~30 minutes, filter, it is 20 volume weight parts with the medical material weight ratio extremely that filtrate adds water for injection, gets the lamiophlomis rotata injection intermediate.
The finger print quality detecting method of lamivphlomis root injection liquid of the present invention, this method comprises the steps:
Chromatographic condition and system suitability test: with octadecylsilane chemically bonded silica is filler; Flow velocity: 1ml/min; The detection wavelength is 254nm; Column temperature: 30 ℃; Analysis time: 1 hour; Carry out gradient elution: preceding 10 minutes, carry out eluting by mobile phase by 10: 90 proportionate relationship of acetonitrile-0.2% phosphate aqueous solution, mobile phase becomes acetonitrile-0.2% phosphate aqueous solution 15: 85 by acetonitrile-0.2% phosphate aqueous solution at 10: 90; 10-40 minute, mobile phase became acetonitrile-0.2% phosphate aqueous solution 30: 70 by acetonitrile-0.2% phosphate aqueous solution at 15: 85; 40-50 minute, mobile phase became acetonitrile-0.2% phosphate aqueous solution 40: 60 by acetonitrile-0.2% phosphate aqueous solution at 30: 70; 50-60 minute, mobile phase remained acetonitrile-0.2% phosphate aqueous solution 40: 60; The preparation of object of reference solution: get the Quercetin reference substance, add the solution that methanol is made 40 μ g/ml, as object of reference solution; The preparation of need testing solution: get lamivphlomis root injection liquid 0.5-2 parts by volume, preferred 1 parts by volume is put in the volumetric flask of 10 parts by volume, adds object of reference solution 0.5-2 parts by volume, preferred 1 parts by volume, methanol is diluted to 10 parts by volume, shakes up, with 0.45 μ m filtering with microporous membrane, get filtrate, promptly;
Algoscopy: draw each 10 μ l of object of reference solution and need testing solution, inject chromatograph of liquid, measure; Test sample finger printing and reference fingerprint; Similarity is greater than 0.90;
The relative retention time of the total fingerprint peaks of lamivphlomis root injection liquid is respectively: No. 1 peak: 0.141 ± 0.005, No. 2 peaks: 0.153 ± 0.001, No. 3 peaks: 0.199 ± 0.001, No. 4 peaks: 0.218 ± 0.002, No. 5 peaks: 0.233 ± 0.002, No. 6 peaks: 0.242 ± 0.001, No. 7 peaks: 0.260 ± 0.001, No. 8 peaks: 0.312 ± 0.001, No. 9 peaks: 0.475 ± 0.001, No. 10 peaks: 0.484 ± 0.001, No. 11 peaks: 0.530 ± 0.001, No. 12 peaks: 0.550 ± 0.001, No. 13 peak: 0.0.585 ± 0.001, No. 14 peaks: 0.659 ± 0.001, S peak: 1; The relative peak area ratio of total fingerprint peaks is respectively: No. 1 peak: 0.023 ± 0.028, No. 2 peaks: 0.168 ± 0.140, No. 3 peaks: 0.163 ± 0.078, No. 4 peaks: 0.054 ± 0.021, No. 5 peaks: 0.067 ± 0.029, No. 6 peaks: 0.031 ± 0.022, No. 7 peaks: 0.047 ± 0.048, No. 8 peaks: 0.057 ± 0.049, No. 9 peaks: 0.164 ± 0.110, No. 10 peaks: 0.325 ± 0.174, No. 11 peaks: 0.066 ± 0.066, No. 12 peaks: 0.048 ± 0.050, No. 13 peaks: 0.591 ± 0.423, No. 14 peaks: 0.091 ± 0.071, S peak: 1.
Above-mentioned lamivphlomis root injection liquid and preparation method thereof can be following method, but is not limited to this method:
Get the Radix Lamiophlomidis Rotatae medical material, decoct with water three times, the water logging that adds 15~25 times was for the first time steeped 0.5~1 hour, decocted 0.5~1.5 hour, and respectively added 10~20 times of water gagings for the second time, for the third time, decocted respectively 0.5~1.5 hour, filter, merging filtrate is concentrated into liquor strength and is 1: 1~2 concentrated solution; Add ethanol and make and contain alcohol amount and reach 60%~80%, in 5~8 ℃ of cold preservations 12~24 hours, filter, filtrate adds ethanol again to be made and contains the alcohol amount and reach 80%~90%, in 5~8 ℃ of cold preservations 12~24 hours, filters; Filtrate is transferred PH to 8~9 with 30%~50% sodium hydroxide, in 5~8 ℃ of cold preservations 12~24 hours, filters; Filtrate adds 10%~30% sulphuric acid and transfers PH to 5~6, placed 1~3 hour, and filtered, reclaim ethanol to there not being the alcohol flavor, adding water for injection is diluted in right amount, 5~8 ℃ of cold preservation 12~24 hours filters, and filtrate adds active carbon 0.05%~0.25%, boiled 15~30 minutes, filter, it is 10 volume weight parts with the medical material weight ratio extremely that filtrate adds water for injection, stirs evenly, add sodium chloride then, add 0.05%~0.25% active carbon, be heated to 80 ℃, boiled 15~30 minutes, filter, filtrate adds injection and is diluted with water in right amount, and with dilute sulfuric acid adjust pH 5.5~6.0, adding water for injection is 20 volume weight parts with the medical material weight ratio extremely, filter with 0.22 μ m microporous filter membrane, fill is sterilized, promptly in the 5ml ampoule.The ratio of above-mentioned volume weight part is ml/g.
Radix Lamiophlomidis Rotatae medical material in the present invention, the characteristic peak quantity of lamivphlomis root injection liquid intermediate and three kinds of test samples of lamivphlomis root injection liquid is successively decreased and has been reflected accurately that also the Radix Lamiophlomidis Rotatae medical material is in being prepared into lamivphlomis root injection liquid process, the phenomenon that unnecessary material is constantly removed with preparation process, reflect more accurately and showed Radix Lamiophlomidis Rotatae and in preparation process, constantly make with extra care, remove and the irrelevant relevant composition of treatment, be beneficial in process of production and accurately product quality controlled and monitored.
The application does not introduce macroporous adsorbent resin and carries out pre-treatment in the preparation process of medical material, intermediate and injection test sample, guaranteed that effective ingredient embodies in finger printing to greatest extent, can reflect real fingerprint characteristic.
Experimentation shows: the need testing solution of Radix Lamiophlomidis Rotatae medical material of the present invention, intermediate and injection finger print quality detecting method thereof is stable in 24 hours; The separating degree at each peak is better, and baseline is more steady, medicinal materials fingerprint and preparation finger better dependency arranged; Precision good; The repeatability of method is good.Show by the result, medicinal materials fingerprint of the present invention, has certain specificity, can obviously distinguish the medical material difference in the different places of production, for the discriminating of medical material provides foundation, difference medical material of poor quality provides precursor and guarantee for producing qualified preparation, what technology was relative stablizes, and can be used for the suitability for industrialized production of preparation; The intermediate finger printing that the present invention formulates has certain specificity, can be used for the monitoring of technology, guarantees the quality of finished product; Lamivphlomis root injection liquid finger printing of the present invention has certain specificity, can be used for the control of end product quality, guarantees the quality of finished product.
Description of drawings:
Fig. 1 is a Radix Lamiophlomidis Rotatae medical material standard finger-print;
Fig. 2 is a Radix Lamiophlomidis Rotatae intermediate standard finger-print;
Fig. 3 is a lamivphlomis root injection liquid standard finger-print.
Following experimental example and embodiment are used to further specify but are not limited to the present invention.
The experiment of experimental example 1 Radix Lamiophlomidis Rotatae medicinal materials fingerprint
(1) source of medical material
Crude drug source is mainly Gansu Province's Wudu County, and the place of production is more fixing, and the natural environment of medical material growth is identical, therefore collects the medical material of different local medical materials in this area or different collecting seasons and commercially available medical material as test sample; The medical material of collecting the different places of production simultaneously as test sample by comparison makes medical material have enough representativenesses.See Table 1.
Table 1 Radix Lamiophlomidis Rotatae crude drug source
Sample number into spectrum | The place of production | Collecting time |
1 2 3 4 5 6 7 8 9 10 | Gansu. Gansu, Maqu. Gansu, Maqu. Gansu, Maqu. Gansu, Maqu. Gansu, Maqu. Gansu, Maqu. Gansu, Maqu. Gansu, Maqu. Gansu, Maqu. the Maqu | 2003.9 2003.10 2003.9 2003.9 2003.9 2003.10 2002.10 2002.9 2002.8 2002.9 |
Main chemical constituent mainly contains flavones ingredient and becomes to grade with some fatty acids with the iridoid constituents in the Radix Lamiophlomidis Rotatae.Flavones ingredient mainly contains luteolin, luteolin-7-o-glucoside, Quercetin, Quercetin-3-o-galactoside, compositions such as apigenin.The experimental applications high performance liquid chromatography experimentizes as detecting index with flavones ingredient in the Radix Lamiophlomidis Rotatae, sets up the finger printing of medical material.
(2) instrument and reagent
The Agilent1100 high performance liquid chromatograph, Chemistation chromatographic work station, Thermo ODS-2HYPERSIL (C
18, 250mm * 4.6mm, 5 μ m) and chromatographic column.
Sartorius type analysis balance (sensibility reciprocal: 0.1mg, 0.01mg), Sartorius AG produces.Acetonitrile, chromatographically pure, import, MERCK company.Water is ultra-pure water; All the other reagent are analytical pure.
(3) selection of chromatographic condition
1. the selection of mobile phase
Selection is that mobile phase experimentizes with methanol-0.2% phosphate aqueous solution and acetonitrile-0.2% phosphate aqueous solution, the result shows that when acetonitrile-0.2% phosphate aqueous solution was mobile phase, the peak shape of chromatographic peak was good, each peak separates more complete, so select for use acetonitrile-0.2% phosphate aqueous solution as mobile phase.
Because complicated component in the sample, under constant current conditions, each component separating degree in the sample is poor, the difficult overall permanence that embodies finger printing; Mobile phase is moved with following gradient, and each component all has separation preferably in the sample, and baseline is steady, so select for use as mobile phase.
Table 2 gradient table:
Time (min) | Acetonitrile (%) | 0.2% phosphate aqueous solution (%) |
0 10 40 50 60 | 10 15 30 40 40 | 90 85 70 60 60 |
Flow velocity is comparatively suitable with 1ml/min, and with this understanding, each peak separates better.
2. detect the selection of wavelength
Use the DAD detector test sample has been carried out the long mensuration of all-wave relatively, the result is bigger in the response value that 254nm surveys main chromatographic peak in the finger printing, and each chromatographic peak feature is outstanding, so determine that maximum absorption wavelength is 254nm.
3. the selection of chromatographic column
Because Main Ingredients and Appearance is the flavonoid composition in the sample, therefore adopt general octadecyl silane to separate for the filler chromatographic column.Thermo ODS-2 HYPERSIL (C
18, 250mm * 4.6mm, 5 μ m) and chromatographic column.
Sample introduction is investigated under above-mentioned chromatographic condition, and results sample is fine in this filler chromatographic column separating degree, determines to select for use 5u C
18(2) 100A, 4.6mm * 250mm, chromatographic column.
(4) gradient is measured lag time
Replacing chromatographic column with a zero volume adapter, is solvent orange 2 A with methanol, and the methanol that contains 0.1% acetone is solvent B, and the detection wavelength is 260nm, and the gradient operation sees the following form:
The operation of table 3 gradient
Time (min) | Flow | A% | B% |
0 20 | 1.0 1.0 | 100 0 | 0 00 |
The result: gradient lag time is 0min.
(5) preparation of need testing solution
Need testing solution mainly is to extract and the rich flavones ingredient that amasss in the medical material, for the ease of the dependency of contrast medical material and preparation, and with reference to the preparation technology of preparation, the following extracting method of experimental design:
1. get and cross the medicinal powder 5g that internal diameter is the 5mm sieve after pulverizing, put in the flask, add water 100ml, reflux, extract, 2 hours filters, and is concentrated into 20ml, adding ethanol to determining alcohol is 75%, and refrigerator was placed 12 hours, filters, reclaim ethanol, residue adds 95% dissolve with ethanol, and standardize solution shakes up in the volumetric flask of 20ml, with 0.45 μ m filtering with microporous membrane, get filtrate, promptly; 2. get and cross the medicinal powder 5g that internal diameter is the 5mm sieve after pulverizing, put in the flask, add 95% ethanol 100ml, reflux, extract, 2 hours, filter, evaporate to dryness, residue add 95% dissolve with ethanol standardize solution and shake up in the volumetric flask of 20ml, with 0.45 μ m filtering with microporous membrane, get filtrate, promptly; 3. get and cross the medicinal powder 5g that internal diameter is the 5mm sieve after pulverizing, put in the flask, add water 100ml, reflux, extract, 2 hours, filter, evaporate to dryness, residue add an amount of dissolve with methanol, centrifugal (1000r/min), the supernatant standardize solution shakes up in the volumetric flask of 20ml, with 0.45 μ m filtering with microporous membrane, get filtrate, promptly.
Drawing the 10ul sample introduction respectively measures.The result shows: each peak separating degree is better in the need testing solution chromatogram that 1. method prepares, and stronger characteristic is arranged, but the method complexity, the time is longer, therefore can not adopt; Though the method 2. separating degree at each peak is better, baseline is more steady, owing to differ bigger with preparation process, the dependency that records medicinal materials fingerprint and preparation finger is relatively poor, therefore also not employing; The method 3. separating degree at each peak is better, and baseline is more steady, records the better dependency of having of medicinal materials fingerprint and preparation finger, therefore determines to adopt this method to prepare the medical material need testing solution.
(6) analysis time
Following sample introduction at above-mentioned chromatographic condition is investigated 2 hours, and the result shows that in 60 minutes, all peaks all can be gone out by complete eluting in the sample, so be defined as 1 hour analysis time.
(7) methodological study
1. stability experiment
Get same need testing solution,, investigate finger printing similarity situation of change, the results are shown in Table 4 respectively in the different time sample detection.
Table 4. stability experiment similarity evaluation result
Experimental result shows: in 24 hours, similarity does not have obvious variation, and RSD% illustrates that less than 0.3% need testing solution was stable in 24 hours.
2. precision experiment
Get same need testing solution continuous sample introduction 5 times, investigate finger printing similarity situation of change, the results are shown in Table 5.
Table 5 precision experiment similarity evaluation result
Experimental result shows: the similarity of 5 sample introductions does not have obvious variation mutually, and RSD% shows that less than 0.3% the precision of instrument is good.
3. repeatability experiment
Get 5 parts of same lot number medical materials, the preparation method of shining need testing solution is equipped with need testing solution with legal system, detects under these conditions, to investigate the repeatability of test sample preparation method, the results are shown in Table 6.
Table 6 repeatability experiment similarity evaluation result
The result shows that the similarity of measuring behind the 5 duplicate samples sample introductions does not have obvious variation, and RSD% illustrates that less than 0.3% the repeatability of this method is good.
(8) similarity evaluation
Ten batches of Radix Lamiophlomidis Rotatae medical materials of table 7 similarity evaluation
Experimental example 2 Radix Lamiophlomidis Rotatae intermediate and injection finger printing chromatographic condition are selected
Adopt gradient elution, mobile phase situation over time sees Table 8.
Table 8 gradient table
Time (min) | Acetonitrile (%) | 0.2% phosphate aqueous solution (%) |
0 10 40 50 60 | 10 15 30 40 40 | 90 85 70 60 60 |
Flow velocity: lml/min; Detect wavelength: 254nm.
Column temperature: 30 ℃; Analysis time: 1 hour.
The result shows, measures under this chromatographic condition, and a chromatographic peak feature is outstanding in the sample, and separating degree is good, and baseline is steady.
Experimental example 3 Radix Lamiophlomidis Rotatae intermediate finger printing stability experiments
Get Radix Lamiophlomidis Rotatae intermediate 1ml, put in the dissolve measuring bottle of 10ml, it is an amount of that the Quercetin reference substance is got in adding, add methanol and make the solution 1ml of 0.4mg/ml, methanol is diluted to scale, shakes up, with 0.45 μ m filtering with microporous membrane, get filtrate, get need testing solution, detect at different time sample introduction 10 μ l respectively, the record chromatogram, investigate finger printing similarity situation of change, the results are shown in Table 9.
Table 9 stability experiment similarity evaluation result
Experimental result shows: in 24 hours, similarity is all greater than 0.98, and RSD is 0.05%, illustrates that need testing solution is basicly stable in 24 hours.
The experiment of experimental example 4 Radix Lamiophlomidis Rotatae intermediate finger printing repeatability
Table 10 repeatability experiment similarity evaluation result
The result shows that the finger printing similarity of measuring behind the 5 duplicate samples sample introductions is all greater than 0.98, and RSD is 0.09%, and similarity changes not remarkable, illustrates that the repeatability of this method is good.
Experimental example 5 Radix Lamiophlomidis Rotatae intermediate finger printing similarity evaluations
Ten batches of Radix Lamiophlomidis Rotatae intermediate of table 11 finger printing similarity evaluation
The result shows: each batch similarity is not significant to be changed, and RSD% is 0.26%, less than 3.0%, shows that the intermediate finger printing that this method is formulated has certain specificity, can be used for the monitoring of technology, guarantees the quality of finished product.
Experimental example 6 Radix Lamiophlomidis Rotatae intermediate and injection finger printing need testing solution preparation method are selected experiment
1. get Radix Lamiophlomidis Rotatae intermediate or injection stock solution as need testing solution.
2. get Radix Lamiophlomidis Rotatae intermediate or injection 1ml, put in the dissolve measuring bottle of 10ml, add methanol and be diluted to scale, as need testing solution.
Get above-mentioned need testing solution 10 μ l sample introductions respectively and measure, the record chromatogram.The result shows: sample stock solution sample introduction, and the separating degree of each chromatographic peak is relatively poor, and baseline is not steady; Behind the sample methanol dilution sample introduction, the separating degree of each chromatographic peak is better, and baseline is steady.Chromatogram and each chromatographic peak ratio of stock solution sample introduction chromatogram do not have marked difference, and 2. therefore global feature that can objective comprehensive response preparation determine to adopt method as the preparation method of need testing solution.
Experimental example 7 lamivphlomis root injection liquid finger printing stability experiments
Get same need testing solution, respectively in different time 10 μ l sample detection, the record chromatogram is investigated finger printing similarity situation of change, the results are shown in Table 12.
Table 12 stability experiment similarity evaluation result
Experimental result shows: in 24 hours, similarity changes not obvious, and RSD% illustrates that less than 3.0% test sample was stable in 24 hours.
The repeatability experiment of experimental example 8 lamivphlomis root injection liquid finger printing test sample preparation methoies
Get 5 parts in same lot number sample, the preparation method of shining need testing solution is equipped with need testing solution with legal system, and sample introduction is measured under above-mentioned chromatographic condition, calculates similarity.The results are shown in Table 13:
Table 13 repeatability experiment similarity evaluation result
The result shows that the finger printing similarity of measuring behind the 5 duplicate samples sample introductions is all greater than 0.98, and RSD is 0.79%, and RSD% illustrates that less than 3.0% the repeatability of this method is good.
Experimental example 9 lamivphlomis root injection liquid finger printing similarity evaluations
Ten batches of lamivphlomis root injection liquid of table 14 finger printing similarity evaluation
The result shows: each batch sample similarity is not significant to be changed, and RSD% is 2.65%, less than 3.0%, shows relative the stablizing of technology, can be used for the suitability for industrialized production of preparation.The lamivphlomis root injection liquid finger printing that this method is formulated has certain specificity, can be used for the control of end product quality, guarantees the quality of finished product.
Following embodiment all can realize the described effect of above-mentioned experimental example.
The specific embodiment
Embodiment 1: the finger print quality detecting method of Radix Lamiophlomidis Rotatae medical material
Chromatographic condition and system suitability test: with octadecylsilane chemically bonded silica is filler; Flow velocity: 1ml/min; The detection wavelength is 254nm; Column temperature: 30 ℃; Analysis time: 1 hour; Carry out gradient elution:
Gradient elution
Time (min) | Acetonitrile (%) | 0.2% phosphate aqueous solution (%) |
0 10 40 50 60 | 10 15 30 40 40 | 90 85 70 60 60 |
The preparation of object of reference solution: it is an amount of to get the Quercetin reference substance, adds the solution that methanol is made 40 μ g/ml, as object of reference solution;
The preparation of need testing solution: get and cross the medicinal powder 5g that internal diameter is the 5mm sieve after pulverizing, put in the flask, add water 100ml, reflux, extract, 2 hours filters water liquid evaporate to dryness, residue adds methanol and dissolves in right amount, and 1000r/min is centrifugal, in the supernatant dislocation 20ml volumetric flask, the accurate object of reference solution 2ml that adds, methanol constant volume shakes up to scale, with 0.45 μ m filtering with microporous membrane, get filtrate, promptly;
Algoscopy: draw each 10 μ l of object of reference solution and need testing solution, inject chromatograph of liquid, measure; Test sample finger printing and reference fingerprint similarity are greater than 0.90;
The preferred relative retention time of the total fingerprint peaks of Radix Lamiophlomidis Rotatae medical material is respectively: 0.145 ± 0.002 (1), 0.154 ± 0.002 (2), 0.199 ± 0.001 (3), 0.220 ± 0.001 (4), 0.231 ± 0.001 (5), 0.242 ± 0.001 (6), 0.260 ± 0.002 (7), 0.298 ± 0.003 (8), 0.313 ± 0.002 (9), 0.336 ± 0.003 (10), 0.369 ± 0.003 (11), 0.386 ± 0.004 (12), 0.402 ± 0.001 (13), 0.414 ± 0.002 (14), 0.476 ± 0.006 (15), 0.484 ± 0.004 (16), 0.533 ± 0.003 (17), 0.550 ± 0.001 (18), 0.585 ± 0.002 (19), 0.642 ± 0.005 (20), 0.661 ± 0.003 (21), 0.760 ± 0.001 (22), 0.796 ± 0.007 (23), 1 (S) (24), 1.077 ± 0.012 (25); Preferred relative peak area ratio is respectively: 0.136 ± 0.100 (1), 1.648 ± 0.564 (2), 1.745 ± 0.653 (3), 0.062 ± 0.031 (4), 0.728 ± 0.247 (5), 0.370 ± 0.141 (6), 0.230 ± 0.112 (7), 0.116 ± 0.048 (8), 0.815 ± 0.262 (9), 0.090 ± 0.025 (10), 0.132 ± 0.107 (11), 0.228 ± 0.140 (12), 0.154 ± 0.193 (13), 0.142 ± 0.183 (14), 1.752 ± 0.455 (15), 2.508 ± 1.357 (16), 0.541 ± 0.379 (17), 0.179 ± 0.152 (18), 2.508 ± 2.437 (19), 0.202 ± 0.109 (20), 0.330 ± 0.262 (21), 0.349 ± 0.137 (22), 0.206 ± 0.046 (23), 1 (S) (24), 0.205 ± 0.077 (25).
Embodiment 2: the finger print quality detecting method of Radix Lamiophlomidis Rotatae intermediate
Chromatographic condition and system suitability test: with octadecylsilane chemically bonded silica is filler; Flow velocity: 1ml/min; The detection wavelength is 254nm; Column temperature: 30 ℃; Analysis time: 1 hour; Carry out gradient elution:
Gradient elution
Time (min) | Acetonitrile (%) | 0.2% phosphate aqueous solution (%) |
0 10 40 50 60 | 10 15 30 40 40 | 90 85 70 60 60 |
The preparation of object of reference solution: it is an amount of to get the Quercetin reference substance, adds the solution that methanol is made 40 μ g/ml, as object of reference solution;
The preparation of need testing solution: get lamivphlomis root injection liquid intermediate 1ml, put in the dissolve measuring bottle of 10ml, the accurate object of reference solution 1ml that adds, methanol is diluted to scale, shakes up, and with 0.45 μ m filtering with microporous membrane, gets filtrate, promptly;
Algoscopy: draw each 10 μ l of object of reference solution and need testing solution, inject chromatograph of liquid, measure; Test sample finger printing and reference fingerprint similarity are greater than 0.90;
The preferred relative retention time of the total fingerprint peaks of Radix Lamiophlomidis Rotatae intermediate is respectively: 0.143 ± 0.001 (1), 0.153 ± 0.001 (2), 0.165 ± 0.001 (3), 0.198 ± 0.001 (4), 0.217 ± 0.001 (5), 0.230 ± 0.001 (6), 0.242 ± 0.002 (7), 0.260 ± 0.001 (8), 0.312 ± 0.001 (9), 0.391 ± 0.008 (10), 0.475 ± 0.002 (11), 0.484 ± 0.002 (12), 0.532 ± 0.001 (13), 0.554 ± 0.004 (14), 0.586 ± 0.001 (15), 0.640 ± 0.003 (16), 0.660 ± 0.002 (17), 0.717 ± 0.001 (18), 1 (19) (S), and 1.070 ± 0.007 (20); The preferred relative peak area ratio of total fingerprint peaks is respectively: 0.029 ± 0.019 (1), 0.384 ± 0.138 (2), 0.028 ± 0.013 (3), 0.260 ± 0.013 (4), 0.094 ± 0.005 (5), 0.121 ± 0.004 (6), 0.058 ± 0.007 (7), 0.086 ± 0.010 (8), 0.127 ± 0.003 (9), 0.037 ± 0.001 (10), 0.352 ± 0.014 (11), 0.619 ± 0.026 (12), 0.119 ± 0.012 (13), 0.032 ± 0.006 (14), 1.402 ± 0.054 (15), 0.051 ± 0.006 (16), 0.183 ± 0.010 (17), 0.066 ± 0.042 (18), 1 (19) (S), and 0.036 ± 0.005.
Embodiment 3: the finger print quality detecting method of lamivphlomis root injection liquid
Chromatographic condition and system suitability test: with octadecylsilane chemically bonded silica is filler; Flow velocity: 1ml/min; The detection wavelength is 254nm; Column temperature: 30 ℃; Analysis time: 1 hour; Carry out gradient elution:
Gradient elution
Time (min) | Acetonitrile (%) | 0.2% phosphate aqueous solution (%) |
0 10 40 50 60 | 10 15 30 40 40 | 90 85 70 60 60 |
The preparation of object of reference solution: it is an amount of to get the Quercetin reference substance, adds the solution that methanol is made 40 μ g/ml, as object of reference solution;
The preparation of need testing solution: get lamivphlomis root injection liquid 1ml, put in the volumetric flask of 10ml, add object of reference solution 1ml, methanol is diluted to scale, shakes up, and with 0.45 μ m filtering with microporous membrane, gets filtrate, promptly;
Algoscopy: draw each 10 μ l of object of reference solution and need testing solution, inject chromatograph of liquid, measure; Test sample finger printing and reference fingerprint; Similarity is greater than 0.90;
The preferred relative retention time of the total fingerprint peaks of lamivphlomis root injection liquid is respectively: 0.141 ± 0.005 (1), 0.153 ± 0.001 (2), 0.199 ± 0.001 (3), 0.218 ± 0.002 (4), 0.233 ± 0.002 (5), 0.242 ± 0.001 (6), 0.260 ± 0.001 (7), 0.312 ± 0.001 (8), 0.475 ± 0.001 (9), 0.484 ± 0.001 (10), 0.530 ± 0.001 (11), 0.550 ± 0.001 (12), 0.0.585 ± 0.001 (13), ± 0.001 (14), 1 0.659 (S) (15); The preferred relative peak area ratio of total fingerprint peaks is respectively: 0.023 ± 0.028 (1), 0.168 ± 0.140 (2), 0.163 ± 0.078 (3), 0.054 ± 0.021 (4), 0.067 ± 0.029 (5), 0.031 ± 0.022 (6), 0.047 ± 0.048 (7), 0.057 ± 0.049 (8), 0.164 ± 0.110 (9), 0.325 ± 0.174 (10), 0.066 ± 0.066 (11), 0.048 ± 0.050 (12), 0.591 ± 0.423 (13), ± 0.071 (14), 1 0.091 (S) (15).
Claims (6)
1, a kind of method of quality control of Radix Lamiophlomidis Rotatae medical material is characterized in that the detection step of finger printing in this method is:
Chromatographic condition and system suitability test: with octadecylsilane chemically bonded silica is filler; Flow velocity: 1ml/min; The detection wavelength is 254nm; Column temperature: 30 ℃; Analysis time: 1 hour; Carry out gradient elution: preceding 10 minutes, carry out eluting by mobile phase by 10: 90 proportionate relationship of acetonitrile-0.2% phosphate aqueous solution, mobile phase becomes acetonitrile-0.2% phosphate aqueous solution 15: 85 by acetonitrile-0.2% phosphate aqueous solution at 10: 90; 10-40 minute, mobile phase became acetonitrile-0.2% phosphate aqueous solution 30: 70 by acetonitrile-0.2% phosphate aqueous solution at 15: 85; 40-50 minute, mobile phase became acetonitrile-0.2% phosphate aqueous solution 40: 60 by acetonitrile-0.2% phosphate aqueous solution at 30: 70; 50-60 minute, mobile phase remained acetonitrile-0.2% phosphate aqueous solution 40: 60; The preparation of object of reference solution: get the Quercetin reference substance, add the solution that methanol is made 40 μ g/ml, as object of reference solution; The preparation of need testing solution: get and cross the Radix Lamiophlomidis Rotatae medicinal powder 1-10 weight portion that internal diameter is the 5mm sieve after pulverizing, put in the flask, add water 50-200 parts by volume, reflux, extract, 1-3 hour, filter water liquid evaporate to dryness, residue adds methanol and dissolves in right amount, and 1000r/min is centrifugal, in the supernatant dislocation 20 parts by volume volumetric flasks, add object of reference solution 0.4-4 parts by volume, methanol constant volume shakes up to scale, with 0.45 μ m filtering with microporous membrane, get filtrate, promptly; Algoscopy: draw each 10 μ l of object of reference solution and need testing solution, inject chromatograph of liquid, measure; The relative retention time of total fingerprint peaks is respectively: No. 1 peak: 0.145 ± 0.002, No. 2 peaks: 0.154 ± 0.002, No. 3 peaks: 0.199 ± 0.001, No. 4 peaks: 0.220 ± 0.001, No. 5 peaks: 0.231 ± 0.001, No. 6 peaks: 0.242 ± 0.001, No. 7 peaks: 0.260 ± 0.002, No. 8 peaks: 0.298 ± 0.003, No. 9 peaks: 0.313 ± 0.002, No. 10 peaks: 0.336 ± 0.003, No. 11 peaks: 0.369 ± 0.003, No. 12 peaks: 0.386 ± 0.004, No. 13 peaks: 0.402 ± 0.001, No. 14 peaks: 0.414 ± 0.002, No. 15 peaks: 0.476 ± 0.006, No. 16 peaks: 0.484 ± 0.004, No. 17 peaks: 0.533 ± 0.003, No. 18 peaks: 0.550 ± 0.001, No. 19 peaks: 0.585 ± 0.002, No. 20 peaks: 0.642 ± 0.005, No. 21 peaks: 0.661 ± 0.003, No. 22 peaks: 0.760 ± 0.001, No. 23 peaks: 0.796 ± 0.007, No. 24 peaks are the S peak: 1, No. 25 peak: 1.077 ± 0.012; The relative peak area ratio of total fingerprint peaks is respectively: No. 1 peak: 0.136 ± 0.100, No. 2 peaks: 1.648 ± 0.564, No. 3 peaks: 1.745 ± 0.653, No. 4 peaks: 0.062 ± 0.031, No. 5 peaks: 0.728 ± 0.247, No. 6 peaks: 0.370 ± 0.141, No. 7 peaks: 0.230 ± 0.112, No. 8 peaks: 0.116 ± 0.048, No. 9 peaks: 0.815 ± 0.262, No. 10 peaks: 0.090 ± 0.025, No. 11 peaks: 0.132 ± 0.107, No. 12 peaks: 0.228 ± 0.140, No. 13 peaks: 0.154 ± 0.193, No. 14 peaks: 0.142 ± 0.183, No. 15 peaks: 1.752 ± 0.455, No. 16 peaks: 2.508 ± 1.357, No. 17 peaks: 0.541 ± 0.379, No. 18 peaks: 0.179 ± 0.152, No. 19 peaks: 2.508 ± 2.437, No. 20 peaks: 0.202 ± 0.109, No. 21 peaks: 0.330 ± 0.262, No. 22 peaks: 0.349 ± 0.137, No. 23 peaks: 0.206 ± 0.046, No. 24 peaks are the S peak: 1, No. 25 peak: 0.205 ± 0.077.
2, the method for quality control of Radix Lamiophlomidis Rotatae medical material according to claim 1, the preparation process that it is characterized in that need testing solution in this method is: get and cross Radix Lamiophlomidis Rotatae medicinal powder 5 weight portions that internal diameter is the 5mm sieve after pulverizing, put in the flask, add water 100 parts by volume, reflux, extract, 2 hours, filter, water liquid evaporate to dryness, residue adds methanol and dissolves in right amount, 1000r/min is centrifugal, in the supernatant dislocation 20 parts by volume volumetric flasks, add object of reference solution 2 parts by volume, methanol constant volume is to scale, shake up, with 0.45 μ m filtering with microporous membrane, get filtrate, promptly.
3, a kind of method of quality control of lamiophlomis rotata injection intermediate is characterized in that the detection step of finger printing in this method is:
Chromatographic condition and system suitability test: with octadecylsilane chemically bonded silica is filler; Flow velocity: 1ml/min; The detection wavelength is 254nm; Column temperature: 30 ℃; Analysis time: 1 hour; Carry out gradient elution: preceding 10 minutes, carry out eluting by mobile phase by 10: 90 proportionate relationship of acetonitrile-0.2% phosphate aqueous solution, mobile phase becomes acetonitrile-0.2% phosphate aqueous solution 15: 85 by acetonitrile-0.2% phosphate aqueous solution at 10: 90; 10-40 minute, mobile phase became acetonitrile-0.2% phosphate aqueous solution 30: 70 by acetonitrile-0.2% phosphate aqueous solution at 15: 85; 40-50 minute, mobile phase became acetonitrile-0.2% phosphate aqueous solution 40: 60 by acetonitrile-0.2% phosphate aqueous solution at 30: 70; 50-60 minute, mobile phase remained acetonitrile-0.2% phosphate aqueous solution 40: 60; The preparation of object of reference solution: get the Quercetin reference substance, add the solution that methanol is made 40 μ g/ml, as object of reference solution; The preparation of need testing solution: get lamivphlomis root injection liquid intermediate 0.5-2 parts by volume, put in the volumetric flask of 10 parts by volume, add object of reference solution 0.5-2 parts by volume, methanol is diluted to 10 parts by volume, shakes up, and with 0.45 μ m filtering with microporous membrane, gets filtrate, promptly; Algoscopy: draw each 10 μ l of object of reference solution and need testing solution, inject chromatograph of liquid, measure; The relative retention time of total fingerprint peaks is respectively: No. 1 peak: 0.143 ± 0.001, No. 2 peaks: 0.153 ± 0.001, No. 3 peaks: 0.165 ± 0.001, No. 4 peaks: 0.198 ± 0.001, No. 5 peaks: 0.217 ± 0.001, No. 6 peaks: 0.230 ± 0.001, No. 7 peaks: 0.242 ± 0.002, No. 8 peaks: 0.260 ± 0.001, No. 9 peaks: 0.312 ± 0.001, No. 10 peaks: 0.391 ± 0.008, No. 11 peaks: 0.475 ± 0.002, No. 12 peaks: 0.484 ± 0.002, No. 13 peaks: 0.532 ± 0.001, No. 14 peaks: 0.554 ± 0.004, No. 15 peaks: 0.586 ± 0.001, No. 16 peaks: 0.640 ± 0.003, No. 17 peaks: 0.660 ± 0.002, No. 18 peaks: 0.717 ± 0.001, No. 19 the peak is the S peak: 1, No. 20 peak: 1.070 ± 0.007; The relative peak area ratio of total fingerprint peaks is respectively: No. 1 peak: 0.029 ± 0.019, No. 2 peaks: 0.384 ± 0.138, No. 3 peaks: 0.028 ± 0.013, No. 4 peaks: 0.260 ± 0.013, No. 5 peaks: 0.094 ± 0.005, No. 6 peaks: 0.121 ± 0.004, No. 7 peaks: 0.058 ± 0.007, No. 8 peaks: 0.086 ± 0.010, No. 9 peaks: 0.127 ± 0.003, No. 10 peaks: 0.037 ± 0.001, No. 11 peaks: 0.352 ± 0.014, No. 12 peaks: 0.619 ± 0.026, No. 13 peaks: 0.119 ± 0.012, No. 14 peaks: 0.032 ± 0.006, No. 15 peaks: 1.402 ± 0.054, No. 16 peaks: 0.051 ± 0.006, No. 17 peaks: 0.183 ± 0.010, No. 18 peaks: 0.066 ± 0.042, No. 19 the peak is the S peak: 1, No. 20 peak: 0.036 ± 0.005.
4, the method for quality control of lamiophlomis rotata injection intermediate as claimed in claim 3, the preparation process that it is characterized in that need testing solution in this method is: get lamivphlomis root injection liquid intermediate 1 parts by volume, put in the volumetric flask of 10 parts by volume, add object of reference solution 1 parts by volume, methanol is diluted to 10 parts by volume, shakes up, with 0.45 μ m filtering with microporous membrane, get filtrate, promptly.
5, a kind of method of quality control of lamivphlomis root injection liquid is characterized in that the detection step of finger printing in this method is:
Chromatographic condition and system suitability test: with octadecylsilane chemically bonded silica is filler; Flow velocity: 1ml/min; The detection wavelength is 254nm; Column temperature: 30 ℃; Analysis time: 1 hour; Carry out gradient elution: preceding 10 minutes, carry out eluting by mobile phase by 10: 90 proportionate relationship of acetonitrile-0.2% phosphate aqueous solution, mobile phase becomes acetonitrile-0.2% phosphate aqueous solution 15: 85 by acetonitrile-0.2% phosphate aqueous solution at 10: 90; 10-40 minute, mobile phase became acetonitrile-0.2% phosphate aqueous solution 30: 70 by acetonitrile-0.2% phosphate aqueous solution at 15: 85; 40-50 minute, mobile phase became acetonitrile-0.2% phosphate aqueous solution 40: 60 by acetonitrile-0.2% phosphate aqueous solution at 30: 70; 50-60 minute, mobile phase remained acetonitrile-0.2% phosphate aqueous solution 40: 60; The preparation of object of reference solution: get the Quercetin reference substance, add the solution that methanol is made 40 μ g/ml, as object of reference solution; The preparation of need testing solution: get lamivphlomis root injection liquid 0.5-2 parts by volume, put in the volumetric flask of 10 parts by volume, add object of reference solution 0.5-2 parts by volume, methanol is diluted to 10 parts by volume, shakes up, and with 0.45 μ m filtering with microporous membrane, gets filtrate, promptly; Algoscopy: draw each 10 μ l of object of reference solution and need testing solution, inject chromatograph of liquid, measure; The relative retention time of total fingerprint peaks is respectively: No. 1 peak: 0.141 ± 0.005, No. 2 peaks: 0.153 ± 0.001, No. 3 peaks: 0.199 ± 0.001, No. 4 peaks: 0.218 ± 0.002, No. 5 peaks: 0.233 ± 0.002, No. 6 peaks: 0.242 ± 0.001, No. 7 peaks: 0.260 ± 0.001, No. 8 peaks: 0.312 ± 0.001, No. 9 peaks: 0.475 ± 0.001, No. 10 peaks: 0.484 ± 0.001, No. 11 peaks: 0.530 ± 0.001, No. 12 peaks: 0.550 ± 0.001, No. 13 peak: 0.0.585 ± 0.001, No. 14 peaks: 0.659 ± 0.001, S peak: 1; Total fingerprint peaks relative peak area ratio is respectively: No. 1 peak: 0.023 ± 0.028, No. 2 peaks: 0.168 ± 0.140, No. 3 peaks: 0.163 ± 0.078, No. 4 peaks: 0.054 ± 0.021, No. 5 peaks: 0.067 ± 0.029, No. 6 peaks: 0.031 ± 0.022, No. 7 peaks: 0.047 ± 0.048, No. 8 peaks: 0.057 ± 0.049, No. 9 peaks: 0.164 ± 0.110, No. 10 peaks: 0.325 ± 0.174, No. 11 peaks: 0.066 ± 0.066, No. 12 peaks: 0.048 ± 0.050, No. 13 peaks: 0.591 ± 0.423, No. 14 peaks: 0.091 ± 0.071, S peak: 1.
6, the method for quality control of lamivphlomis root injection liquid as claimed in claim 5, the preparation process that it is characterized in that need testing solution in this method is: get lamivphlomis root injection liquid 1 parts by volume, put in the volumetric flask of 10 parts by volume, add object of reference solution 1 parts by volume, methanol is diluted to 10 parts by volume, shakes up, with 0.45 μ m filtering with microporous membrane, get filtrate, promptly.
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Cited By (3)
Publication number | Priority date | Publication date | Assignee | Title |
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CN104237441A (en) * | 2014-10-09 | 2014-12-24 | 成都中医药大学 | Method for simultaneous detection of iridoid glycoside, phenylethanoid glycoside, flavone and dicaffeoyl ingredients in lamiophlomis rotata |
CN107045035A (en) * | 2017-04-18 | 2017-08-15 | 普正药业股份有限公司 | A kind of method for building up of Duyiwei effervescent tablet HPLC finger-prints |
CN109975439A (en) * | 2017-12-28 | 2019-07-05 | 九芝堂股份有限公司 | A kind of UPLC fingerprint atlas detection method of lamiophlomis rotata medicinal material and its preparation |
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CN100419423C (en) * | 2004-10-15 | 2008-09-17 | 成都优他制药有限责任公司 | Method for controlling quality of simply single drug |
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Cited By (4)
Publication number | Priority date | Publication date | Assignee | Title |
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CN104237441A (en) * | 2014-10-09 | 2014-12-24 | 成都中医药大学 | Method for simultaneous detection of iridoid glycoside, phenylethanoid glycoside, flavone and dicaffeoyl ingredients in lamiophlomis rotata |
CN104237441B (en) * | 2014-10-09 | 2015-07-22 | 成都中医药大学 | Method for simultaneous detection of iridoid glycoside, phenylethanoid glycoside, flavone and dicaffeoyl ingredients in lamiophlomis rotata |
CN107045035A (en) * | 2017-04-18 | 2017-08-15 | 普正药业股份有限公司 | A kind of method for building up of Duyiwei effervescent tablet HPLC finger-prints |
CN109975439A (en) * | 2017-12-28 | 2019-07-05 | 九芝堂股份有限公司 | A kind of UPLC fingerprint atlas detection method of lamiophlomis rotata medicinal material and its preparation |
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