CN103115976A - Method for measuring content of in-blood chemical component-allantoin in medicinal material-psammosilene tunicoides - Google Patents
Method for measuring content of in-blood chemical component-allantoin in medicinal material-psammosilene tunicoides Download PDFInfo
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Abstract
The invention discloses a method for measuring content of an in-blood chemical component-allantoin in a medicinal material-psammosilene tunicoides. The method is a high performance liquid chromatography which takes an allantoin reference substance as the reference and the ratio of acetonitrile or methanol to water as (60-100):(0-40) as the flowing phase. The method selects the in-blood chemical component-allantoin in the medicinal material-psammosilene tunicoides as the index to carry out content measurement; the measuring method is high in precision degree, good in reproducibility, good in stability, high in recovery rate and accurate in measuring result, and the quality of the medicinal material-psammosilene tunicoides can be effectively controlled, so that the safety and effectiveness in clinical medication of the psammosilene tunicoides are ensured.
Description
Technical field
The present invention relates to enter in a kind of tuniclike psammosilene root medicinal material the content assaying method of blood chemistry composition allantoin, belong to the technical field of medicinal material being carried out quality control.
Background technology
Tuniclike psammosilene root (Psammosilene tunicoides), the dry root for Caryophyllaceae tuniclike psammosilene root platymiscium tuniclike psammosilene root is used as medicine with root.Mainly be distributed in the In Southwest Chinas such as Guizhou, Yunnan, Sichuan, Tibet; it is the distinctive Mono-species genus in Southwestern China area; be also Guizhou Chinese herbal medicine commonly used among the people, now list in " Chinese Plants Red Book " as rare endangered species, belong to Chinese Second Class Key Protected Plant.The tuniclike psammosilene root effect of carbuncle apocenosis that has loose stasis of blood analgesic therapy, stops blooding and disappear is used for the treatment of traumatic injury, rheumatalgia, peratodynia, traumatic bleeding etc.
According to bibliographical information, the tuniclike psammosilene root medicinal material is in analgesia, anti-inflammatory, antibacterial and regulate the aspect such as immunologic function and all have the conspicuousness effect, and finds that in tuniclike psammosilene root, main chemical composition is triterpene saponin and cyclic peptide compound.But present, also there is no the bibliographical information about the research of tuniclike psammosilene root quality of medicinal material control method.Although tuniclike psammosilene root is recorded in " Guizhou Province's Chinese crude drug, national quality of medicinal material standard " version in 2003, both differentiate without thin-layer chromatography in this standard, also without the assay item; And " Chinese pharmacopoeia also only had the thin layer of control medicinal material to differentiate in 2010 in version.And if there is no tight quality standard and the detection means of science, just be difficult to effectively control its inherent quality, can not guarantee the safe, effective of medication, do not meet the requirement of international medical market yet, thereby may seriously restrict the development of China's Chinese herbal medicine industry.Therefore, strengthening the Chinese crude drug quality controling research is Chinese medicine standardization, standardized key issue, only has the quality control of Chinese crude drug being carried out science, guarantee traditional Chinese medicine quality, " safely, effective, stable, controlled " who realizes Chinese crude drug.
Summary of the invention
The object of the invention is to, the content assaying method that enters blood chemistry composition allantoin in a kind of tuniclike psammosilene root medicinal material is provided.The present invention is by carrying out quantitative examination to having the bioactive content that enters blood chemistry composition allantoin in tuniclike psammosilene root, and the perfect quality inspection standard of tuniclike psammosilene root medicinal material, the quality determining method that makes the tuniclike psammosilene root medicinal material be science, rationally more.
The present invention consists of like this: enter the content assaying method of blood chemistry composition allantoin in a kind of tuniclike psammosilene root medicinal material, namely take the allantoin reference substance as contrast, take acetonitrile or methyl alcohol: water=60～100:0～40 are as the high performance liquid chromatography of mobile phase.
Concrete content assaying method is: according to " appendix VID high effective liquid chromatography for measuring of Chinese pharmacopoeia version in 2010:
Chromatographic condition and system suitability: take amino bonded silica gel as filling agent; Take acetonitrile or methyl alcohol: water=60 ~ 100:0 ~ 40 are as mobile phase; Flow velocity is 0.5 ~ 2mLmin
-1Column temperature is 30 ~ 50 ℃; The detection wavelength is 210 ~ 240nm; Number of theoretical plate should be not less than 3000;
The preparation of reference substance solution: precision takes the allantoin reference substance, adds 10% ~ 50% ethanol and makes the solution that every 1mL contains 0.05～0.30mg allantoin, and get final product;
The preparation of need testing solution: get tuniclike psammosilene root medicinal powder 0.5 ~ 5g, accurately weighed, be placed in tool plug conical flask or triangular flask, add 10% ~ 50% ethanol 10 ~ 40mL, weighed weight, ultrasonic extraction 10 ~ 60min supplies the weight of less loss with 10% ~ 50% ethanol, shake up, filter, get subsequent filtrate, and get final product;
Determination method: precision is drawn reference substance solution and each 1 ~ 10 μ L of need testing solution respectively, injects high performance liquid chromatograph, measures, and get final product.
Preferred content assaying method is: according to " appendix VID high effective liquid chromatography for measuring of Chinese pharmacopoeia version in 2010:
Chromatographic condition and system suitability: take amino bonded silica gel as filling agent; Take acetonitrile: water=93:7 as mobile phase; Flow velocity is 1mLmin
-1Column temperature is 40 ℃; The detection wavelength is 220nm; Number of theoretical plate should be not less than 3000;
The preparation of reference substance solution: precision takes the allantoin reference substance, adds 20% ethanol and makes the solution that every 1mL contains 0.05～0.30mg allantoin, and get final product;
The preparation of need testing solution: get tuniclike psammosilene root medicinal powder 2g, accurately weighed, be placed in tool plug conical flask or triangular flask, add 20% ethanol 30mL, weighed weight, ultrasonic extraction 40min supplies the weight of less loss with 20% ethanol, shake up, and filters, and gets subsequent filtrate, and get final product;
Determination method: precision is drawn reference substance solution and each 5 μ L of need testing solution respectively, injects high performance liquid chromatograph, measures, and get final product.
Tuniclike psammosilene root (Psammosilene tunicoides) medicinal material records in " Guizhou Province's Chinese crude drug, national quality of medicinal material standard " version in 2003, but both the thin-layer chromatography without the tuniclike psammosilene root medicinal material is differentiated item in this standard, also without the assay item, simultaneously " Chinese pharmacopoeia also only had the thin layer of the control medicinal material of tuniclike psammosilene root to differentiate in 2010 in version, therefore for the quality of more effective control tuniclike psammosilene root medicinal material, the inventor specializes in its quality determining method.
Find by consulting the domestic and foreign literature data: the Research Literature report of tuniclike psammosilene root quality of medicinal material control method aspect seldom, for this reason, the inventor is studied the blood chemistry composition that enters of tuniclike psammosilene root, has determined that the bioactive ingredients allantoin of tuniclike psammosilene root belongs to into the blood chemistry composition; On the basis of determining the blood chemistry composition, then the content assaying method that enters blood chemistry composition allantoin in the tuniclike psammosilene root medicinal material has been carried out experimental study.
Below the inventor to the process of the content assaying method research experiment that enters blood chemistry composition allantoin in the tuniclike psammosilene root medicinal material:
One, the tuniclike psammosilene root medicinal material enters determining of blood chemistry composition
The inventor has carried out a large amount of research to the document of relevant tuniclike psammosilene root medicinal material, and early stage the chemical constitution study that tuniclike psammosilene root launches is found according to inventor place seminar, allantoin is the bioactive ingredients in the tuniclike psammosilene root medicinal material, and the allantoin of therefore choosing the tuniclike psammosilene root medicinal material is that index has carried out entering blood test research:
After getting the tuniclike psammosilene root medicinal material drying, pulverizing, with twice of 10% ~ 40% edible ethanol extraction of 10 ~ 50mL; Merge extract, reduced pressure concentration gets Radix Psammosilenes extract after reclaiming solvent.Get appropriate Radix Psammosilenes extract water dissolving, being made into concentration is 20mgmL
-1Aqueous solution; Getting Radix Psammosilenes extract aqueous solution 2～4mL, is the rat oral gavage administration of 180～220g to weight, at 10min, 30min, 60min and 120min, rat is taken a blood sample respectively; To get upper plasma after centrifugal blood, centrifugal after the vortex mixing with acetonitrile or methanol extraction albumen, supernatant liquor is placed on Nitrogen evaporator dries up; Get and dry up in right amount thing, add acetonitrile-water=93:7(volume ratio) solution dissolves, centrifugal, as need testing solution; Blank plasma is processed after the same method; Get the allantoin reference substance and make reference substance solution.
High-efficient liquid phase chromatogram condition is: take amino bonded silica gel as filling agent; Take acetonitrile: water=93:7 as mobile phase; Flow velocity is 1mLmin
-1Column temperature is 40 ℃; The detection wavelength is 220nm; Number of theoretical plate should be not less than 3000.
Measurement result such as Fig. 1～shown in Figure 6 can find out from Fig. 1～Fig. 6, and allantoin is that a kind of in the tuniclike psammosilene root medicinal material enters the blood chemistry composition.
Two, the research of the content assaying method of allantoin in the tuniclike psammosilene root medicinal material
1. instrument and reagent: the Ultimate300 high performance liquid chromatograph, it comprises quaternary pump, online degasser, automatic sampler, diode array detector and column oven; 100,000/electronic balance (being provided by plum Teller-Tuo benefit instrument Shanghai company limited); KQ5200E type Ultrasonic Cleaning (ultrasonic instrument company limited provides by the Kunming); Chromatogram acetonitrile, water (redistilled water, preparation before use), absolute ethyl alcohol, allantoin reference substance (being provided by Nat'l Pharmaceutical ﹠ Biological Products Control Institute) and tuniclike psammosilene root medicinal material (picking up from 6 different places of production).
2. the selection of chromatographic condition
The inventor especially selects chromatographic column and flow phase system when carrying out the selection of allantoin chromatographic condition.The chromatographic condition of determining allantoin according to experiment is: ZORBAX NH
2(4.6 * 150mm, 5 μ m) chromatographic column, acetonitrile-water (93: 7) is mobile phase, flow velocity is 1mLmin
-1, the detection wavelength is 220nm.Alternative condition sees Table 1.
The selection of table 1 allantoin chromatographic condition
3. the selection of column temperature
When carrying out the selection of column temperature, selected 20 ℃, 30 ℃, 40 ℃ to test.Experimental studies have found that when column temperature was 40 ℃, the allantoin peak shape was better, degree of separation is better.
4. the selection of test sample extraction conditions
(1) extract the selection of solvent
Get with a collection of tuniclike psammosilene root medicinal powder 2g, accurately weighed, be placed in tool plug conical flask or triangular flask, add 20% ethanol, 40% ethanol, 60% ethanol, each 30mL of methyl alcohol, weighed weight, ultrasonic extraction 40min, supply respectively the weight of less loss with 20% ethanol, 40% ethanol, 60% ethanol, methyl alcohol, shake up, filter, get subsequent filtrate, the content of allantoin in working sample.The results are shown in Table 2.
Table 2 extracts the selection of solvent
As can be seen from the above table, during with 20% alcohol extract tuniclike psammosilene root medicinal material, the content of allantoin is the highest.
(2) selection of extracting mode
Get with a collection of tuniclike psammosilene root medicinal powder 2g, accurately weighed three parts, be placed in tool plug conical flask or triangular flask, add respectively 20% ethanol 30mL, weighed weight.A ultrasonic processing 40min; Another part adds hot reflux 120min; The 3rd part is extracted 120min with Soxhlet.Let cool after three parts of extractions, more weighed weight.Supply less loss weight with 20% ethanol, shake up, filter, get subsequent filtrate, the content of allantoin in working sample.The results are shown in Table 3.
The selection of table 3 extracting mode
As can be seen from the above table, when adopting ultrasonic extraction, the content of allantoin is the highest, and when adopting hot reflux extraction and Soxhlet to extract two kinds of extracting method, effect is relatively poor, therefore select ultrasonic extraction.
(3) selection of extraction time
Get with a collection of tuniclike psammosilene root medicinal powder 2g, accurately weighed, be placed in tool plug conical flask or triangular flask, add 20% ethanol 30mL, weighed weight.Ultrasonic processing 10min, 20min, 30min, 40min, 50min, 60min let cool, more weighed weight.Supply less loss weight with 20% ethanol, shake up, filter, get subsequent filtrate, the content of allantoin in working sample.The results are shown in Table 4.
The selection of table 4 extraction time
| Extraction time | 10min | 20min | 30min | 40min | 50min | 60min |
| The content of allantoin (%) | 0.065 | 0.069 | 0.074 | 0.082 | 0.080 | 0.084 |
As can be seen from the above table, during ultrasonic processing 40min, allantoin extracts fully.
5. the investigation of linear relationship
Precision takes the allantoin 1.66mg that is dried to constant weight through phosphorus pentoxide and is placed in the 10mL volumetric flask, adds 20% ethanol and is made into 0.166mgmL
-1The reference substance storing solution.Get the reference substance storing solution, make respectively 0.0104mgmL
-1, 0.0208mgmL
-1, 0.0415mgmL
-1, 0.083mgmL
-1, 0.166mgmL
-1The reference substance solution of 5 variable concentrations, respectively accurate each the 5 μ L of aforementioned reference substance solution that draw, inject high performance liquid chromatograph, measure by above-mentioned chromatographic condition, and take the peak area of reference substance as horizontal ordinate, concentration (mgmL
-1) Y is ordinate, drawing standard curve, regression equation: Y=0.0137X+0.0012, r
2=0.9994(n=5).Result shows, allantoin is linear relationship good (seeing Table 5) between 0.104 ~ 1.66mg.
Table 5 allantoin linear relationship examination table
| Concentration (mgmL -1） | 0.0104 | 0.0208 | 0.0415 | 0.0830 | 0.1660 |
| Peak area | 0.7562 | 1.4675 | 2.9062 | 5.8533 | 12.3972 |
6. precision test
Withinday precision is drawn allantoin reference substance solution 5 μ L, according to above-mentioned chromatographic condition continuous sample introduction 6 times, measures peak area, and calculate the mean relative deviation result: RSD% is 0.46%, illustrates that the method has good precision (seeing Table 6).
Table 6 withinday precision examination result
Day to day precision is drawn allantoin reference substance solution 5 μ L, and according to above-mentioned chromatographic condition, every day, sample introduction twice, METHOD FOR CONTINUOUS DETERMINATION three days, and calculate the mean relative deviation result: RSD% is 0.95%, illustrates that the method has good precision (seeing Table 7).
Table 7 day to day precision examination result
7. replica test
Get with a collection of each 2g of tuniclike psammosilene root medicinal material, totally six parts, prepare test liquid by the preparation method of the test sample in content assaying method of the present invention.The accurate need testing solution 5 μ L that draw inject high performance liquid chromatograph, measure the peak area integrated value of allantoin by chromatographic condition of the present invention, calculate content, ask relative standard deviation.Result shows, it is good that the method is measured the reappearance of allantoin, RSD=2.10% (seeing Table 8).
Table 8 replica test result
8. stability test
Get with a collection of tuniclike psammosilene root medicinal material 2.0g, prepare test liquid by the test sample preparation method in content assaying method of the present invention, at room temperature respectively at 0h, 2h, 4h, 8h, the accurate 5 μ L sample introductions of drawing of 12h, 24h, measure the peak area integrated value of allantoin, calculate content, ask relative standard deviation.Result shows that allantoin is basicly stable in 24h, RSD=1.60% (seeing Table 9).
Table 9 stability test result
9. average recovery test
Adopt the application of sample absorption method, get the sample of known content, accurately weighed, precision adds a certain amount of allantoin respectively, prepares test liquid according to the preparation method of the test sample described in technical solution of the present invention, measures according to the said determination method, calculate recovery rate the results are shown in Table 10.
Table 10 recovery test result (n=9)
10. sample determination
Prepare test sample and reference substance solution by the method described in technical solution of the present invention, sample introduction, record chromatogram respectively, calculates the content of allantoin, the results are shown in Table 11.
Content of Allantoin measurement result (n=2) in table 116 batch medicinal material
Compared with prior art, the present invention selects to enter that the blood chemistry composition---allantoin is that index is carried out assay in the tuniclike psammosilene root medicinal material, the method precision is high, favorable reproducibility, good stability, the recovery are high, measurement result is accurate, can effectively control the quality of tuniclike psammosilene root medicinal material, thereby guarantee the safe, effective of its clinical application.
Description of drawings
Fig. 1 is the high-efficient liquid phase chromatogram of allantoin reference substance;
Fig. 2 is the high-efficient liquid phase chromatogram of the blood plasma that gathers after gavage rat 10min;
Fig. 3 is the high-efficient liquid phase chromatogram of the blood plasma that gathers after gavage rat 30min;
Fig. 4 is the high-efficient liquid phase chromatogram of the blood plasma that gathers after gavage rat 60min;
Fig. 5 is the high-efficient liquid phase chromatogram of the blood plasma that gathers after gavage rat 120min;
Fig. 6 is the high-efficient liquid phase chromatogram of blank plasma;
Fig. 7 is that in the tuniclike psammosilene root medicinal material, allantoin is measured the system flexibility collection of illustrative plates.
Embodiment
Embodiment 1: to the concrete grammar that blood chemistry composition allantoin carries out assay that enters in the tuniclike psammosilene root medicinal material be: according to " appendix VID high effective liquid chromatography for measuring of Chinese pharmacopoeia version in 2010:
Chromatographic condition and system suitability: take amino bonded silica gel as filling agent; Take acetonitrile: water=93:7 as mobile phase; Flow velocity is 1mLmin-1; Column temperature is 40 ℃; The detection wavelength is 220nm; Number of theoretical plate should be not less than 3000;
The preparation of reference substance solution: precision takes the allantoin reference substance, adds 20% ethanol and makes the solution that every 1mL contains 0.05～0.30mg allantoin, and get final product;
The preparation of need testing solution: get tuniclike psammosilene root medicinal powder 2g, accurately weighed, put in tool plug conical flask or triangular flask, add 20% ethanol 30mL, weighed weight, ultrasonic extraction 40min supplies the weight of less loss with 20% ethanol, shake up, and filters, and gets subsequent filtrate, and get final product;
Determination method: precision is drawn reference substance solution and each 5 μ L of need testing solution respectively, injects high performance liquid chromatograph, measures, and get final product.Measurement result is seen Fig. 7.
Embodiment 2: enter the content assaying method of blood chemistry composition allantoin in a kind of tuniclike psammosilene root medicinal material, be specially: according to " appendix VID high effective liquid chromatography for measuring of Chinese pharmacopoeia version in 2010:
Chromatographic condition and system suitability: take amino bonded silica gel as filling agent; Take acetonitrile: water=80:20 as mobile phase; Flow velocity is 2mLmin
-1Column temperature is 50 ℃; The detection wavelength is 240nm; Number of theoretical plate should be not less than 3000;
The preparation of reference substance solution: precision takes the allantoin reference substance, adds 50% ethanol and makes the solution that every 1mL contains the 0.30mg allantoin, and get final product;
The preparation of need testing solution: get tuniclike psammosilene root medicinal powder 5g, accurately weighed, put in tool plug conical flask or triangular flask, add 50% ethanol 40mL, weighed weight, ultrasonic extraction 60min supplies the weight of less loss with 50% ethanol, shake up, and filters, and gets subsequent filtrate, and get final product;
Determination method: precision is drawn reference substance solution and each 10 μ L of need testing solution respectively, injects high performance liquid chromatograph, measures, and get final product.
Embodiment 3: enter the content assaying method of blood chemistry composition allantoin in a kind of tuniclike psammosilene root medicinal material, be specially: according to " appendix VID high effective liquid chromatography for measuring of Chinese pharmacopoeia version in 2010:
Chromatographic condition and system suitability: take amino bonded silica gel as filling agent; Take acetonitrile: water=99:1 as mobile phase; Flow velocity is 0.5mLmin
-1Column temperature is 30 ℃; The detection wavelength is 210nm; Number of theoretical plate should be not less than 3000;
The preparation of reference substance solution: precision takes appropriate allantoin reference substance, adds 10% ethanol and makes the solution that every 1mL contains the 0.05mg allantoin, and get final product;
The preparation of need testing solution: get tuniclike psammosilene root medicinal powder 0.5g, accurately weighed, put in tool plug conical flask or triangular flask, add 10% ethanol 10mL, weighed weight, ultrasonic extraction 10min supplies the weight of less loss with 10% ethanol, shake up, and filters, and gets subsequent filtrate, and get final product;
Determination method: precision is drawn reference substance solution and each 1 μ L of need testing solution respectively, injects high performance liquid chromatograph, measures, and get final product.
Embodiment 4: enter the content assaying method of blood chemistry composition allantoin in a kind of tuniclike psammosilene root medicinal material, be specially: according to " appendix VID high effective liquid chromatography for measuring of Chinese pharmacopoeia version in 2010:
Chromatographic condition and system suitability: take amino bonded silica gel as filling agent; Take methyl alcohol: water=93:7 as mobile phase; Flow velocity is 1mLmin
-1Column temperature is 40 ℃; The detection wavelength is 220nm; Number of theoretical plate should be not less than 3000;
The preparation of reference substance solution: precision takes appropriate allantoin reference substance, adds 20% ethanol and makes the solution that every 1mL contains the 0.15mg allantoin, and get final product;
The preparation of need testing solution: get tuniclike psammosilene root medicinal powder 2g, accurately weighed, put in tool plug conical flask or triangular flask, add 20% ethanol 30mL, weighed weight, ultrasonic extraction 20min supplies the weight of less loss with 20% ethanol, shake up, and filters, and gets subsequent filtrate, and get final product;
Determination method: precision is drawn reference substance solution and each 5 μ L of need testing solution respectively, injects high performance liquid chromatograph, measures, and get final product.
Embodiment 5: enter the content assaying method of blood chemistry composition allantoin in a kind of tuniclike psammosilene root medicinal material, be specially: according to " appendix VID high effective liquid chromatography for measuring of Chinese pharmacopoeia version in 2010:
Chromatographic condition and system suitability: take amino bonded silica gel as filling agent; Take methyl alcohol: water=60:40 as mobile phase; Flow velocity is 0.5mLmin
-1Column temperature is 30 ℃; The detection wavelength is 210nm; Number of theoretical plate should be not less than 3000;
The preparation of reference substance solution: precision takes appropriate allantoin reference substance, adds 10% ethanol and makes the solution that every 1mL contains the 0.05mg allantoin, and get final product;
The preparation of need testing solution: get tuniclike psammosilene root medicinal powder 0.5g, accurately weighed, put in tool plug conical flask or triangular flask, add 10% ethanol 10mL, weighed weight, ultrasonic extraction 15min supplies the weight of less loss with 10% ethanol, shake up, and filters, and gets subsequent filtrate, and get final product;
Determination method: precision is drawn reference substance solution and each 5 μ L of need testing solution respectively, injects high performance liquid chromatograph, measures, and get final product.
Embodiment 6: enter the content assaying method of blood chemistry composition allantoin in a kind of tuniclike psammosilene root medicinal material, be specially: according to " appendix VID high effective liquid chromatography for measuring of Chinese pharmacopoeia version in 2010:
Chromatographic condition and system suitability: take amino bonded silica gel as filling agent; Take methyl alcohol as mobile phase; Flow velocity is 2mLmin
-1Column temperature is 50 ℃; The detection wavelength is 240nm; Number of theoretical plate should be not less than 3000;
The preparation of reference substance solution: precision takes appropriate allantoin reference substance, adds 50% ethanol and makes the solution that every 1mL contains the 0.30mg allantoin, and get final product;
The preparation of need testing solution: get tuniclike psammosilene root medicinal powder 5g, accurately weighed, put in tool plug conical flask or triangular flask, add 50% ethanol 40mL, weighed weight, ultrasonic extraction 40min supplies the weight of less loss with 50% ethanol, shake up, and filters, and gets subsequent filtrate, and get final product;
Determination method: precision is drawn reference substance solution and each 10 μ L of need testing solution respectively, injects high performance liquid chromatograph, measures, and get final product.
Claims (3)
1. enter the content assaying method of blood chemistry composition allantoin in a tuniclike psammosilene root medicinal material, it is characterized in that, described content assaying method is take the allantoin reference substance as contrast, and take acetonitrile or methyl alcohol: water=60～100:0～40 are as the high performance liquid chromatography of mobile phase.
2. enter the content assaying method of blood chemistry composition allantoin in tuniclike psammosilene root medicinal material according to claim 1, it is characterized in that, concrete grammar is: according to " appendix VID high effective liquid chromatography for measuring of Chinese pharmacopoeia version in 2010:
Chromatographic condition and system suitability: take amino bonded silica gel as filling agent; Take acetonitrile or methyl alcohol: water=60 ~ 100:0 ~ 40 are as mobile phase; Flow velocity is 0.5 ~ 2mLmin
-1Column temperature is 30 ~ 50 ℃; The detection wavelength is 210 ~ 240nm; Number of theoretical plate should be not less than 3000;
The preparation of reference substance solution: precision takes the allantoin reference substance, adds 10% ~ 50% ethanol and makes the solution that every 1mL contains 0.05～0.30mg allantoin, and get final product;
The preparation of need testing solution: get tuniclike psammosilene root medicinal powder 0.5 ~ 5g, accurately weighed, be placed in tool plug conical flask or triangular flask, add 10% ~ 50% ethanol 10 ~ 40mL, weighed weight, ultrasonic extraction 10 ~ 60min supplies the weight of less loss with 10% ~ 50% ethanol, shake up, filter, get subsequent filtrate, and get final product;
Determination method: precision is drawn reference substance solution and each 1 ~ 10 μ L of need testing solution respectively, injects high performance liquid chromatograph, measures, and get final product.
3. enter the content assaying method of blood chemistry composition allantoin in tuniclike psammosilene root medicinal material according to claim 2, it is characterized in that, concrete grammar is: according to " appendix VID high effective liquid chromatography for measuring of Chinese pharmacopoeia version in 2010:
Chromatographic condition and system suitability: take amino bonded silica gel as filling agent; Take acetonitrile: water=93:7 as mobile phase; Flow velocity is 1mLmin
-1Column temperature is 40 ℃; The detection wavelength is 220nm; Number of theoretical plate should be not less than 3000;
The preparation of reference substance solution: precision takes the allantoin reference substance, adds 20% ethanol and makes the solution that every 1mL contains 0.05～0.30mg allantoin, and get final product;
The preparation of need testing solution: get tuniclike psammosilene root medicinal powder 2g, accurately weighed, be placed in tool plug conical flask or triangular flask, add 20% ethanol 30mL, weighed weight, ultrasonic extraction 40min supplies the weight of less loss with 20% ethanol, shake up, and filters, and gets subsequent filtrate, and get final product;
Determination method: precision is drawn reference substance solution and each 5 μ L of need testing solution respectively, injects high performance liquid chromatograph, measures, and get final product.
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| CN111551665A (en) * | 2020-06-23 | 2020-08-18 | 贵州医科大学 | GC-MS technology-based method for analyzing volatile oil and fatty acid in psammosilene tunicoides |
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Cited By (3)
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|---|---|---|---|---|
| CN109839470A (en) * | 2019-02-25 | 2019-06-04 | 亚宝药业贵阳制药有限公司 | A kind of thin-layer identification method of invigorating the spleen gel products |
| CN110579559A (en) * | 2019-10-26 | 2019-12-17 | 无锡济民可信山禾药业股份有限公司 | derivatization combined LC-MS method for determining allantoin concentration in cell matrix |
| CN111551665A (en) * | 2020-06-23 | 2020-08-18 | 贵州医科大学 | GC-MS technology-based method for analyzing volatile oil and fatty acid in psammosilene tunicoides |
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