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OBJECTIVE： To investigate the rationality of TLC identification method （3） of （R，S）-epigoitrin in Isatis indigotica stated in 2015 edition of Chinese Pharmacopeia （partⅠ） （later abbreviated as pharmacopeia）， and make some improvements. METHODS： Three batches I. indigotica were collected and prepared into decoction pieces according to the processing method of I. indigotica in pharmacopoeia. TLC identification of （R，S）-goitrin in I. indigotica decoction piece and medicinal material were conducted according to identification method （3） in pharmacopeia （80％ ethanol as solvent for sample treatment， ultrasound extraction）； the rationality of pharmacopoeia method was investigated. Then the method was improved by changing the extraction solvent and pretreatment method （method one： using water as solvent， ultrasound extraction； method two： soaking in water for 1 h， then adding into methanol， ultrasound extraction； method three： the sample was wetted and then dried， using 80％ methanol as solvent， ultrasound extraction） of samples， and the optimal method was verified. According to the optimal method， the TLC identification of （R，S）-goitrin was detected by using chromatographic plates from different manufacturers， under the conditions of low temperature and low humidity （7 ℃， relative humidity 48％） and high temperature and high humidity （35 ℃， relative humidity 75％） respectively，to investigate the durability of the method. RESULTS： According to the method of pharmacopeia， in the chromatograms of decoction pieces， the same color spots appeared at the corresponding chromatographic positions of reference substance， but no corresponding spots appeared in the medicinal material chromatograms. After the samples were treated by three improvement methods， in medicinal material chromatograms， the same color spots appeared in the corresponding chromatographic positions of reference substances. There were single chromatographic spot after medicinal materials were treated with method one， and there were more spots after medicinal materials were treated with method two and three， and method two consumed less time than method three. The results of validation tests and method durability tests showed that after the treatment of I. indigotica and its decoction pieces according to method two， the same color spots appeared in the corresponding positions of the decoction pieces and the medicinal materials chromatograms as those of the control. CONCLUSIONS： The improved TLC identification method is effective， the chromatographic spots are clear， and the repeatability is good.
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AIM To establish an HPLC method for the simultaneous content determination of five constituents in Maxing Kanggan Granules (Isatidis Radix,Houttuyniae Herba,Scutellariae Radix,etc.).METHODS The analysis of 70％ ethanol extract of this drug was performed on a 30 ℃ thermostatic Wondasil C18 column (250 mm × 4.6 mm,5 μm),with the mobile phase comprising of acetonitrile-0.1％ phosphoric acid flowing at 1.0 mL/min in a gradient elution manner,and the detection wavelengths were set at 245,327,280 and 250 nm.RESULTS (R,S)-goitrin,chlorogenic acid,baicalin,baicalein and glycyrrhizic acid showed good linear relationships within the ranges of 0.24-6.05 μg/mL (r =0.999 9),0.70-17.40 μg/mL (r =0.999 9),28.59-714.70 μg/mL (r =0.999 6),1.67-41.65 μg/mL (r =0.999 5) and 2.59-64.80 μg/mL (r =0.999 7),whose average recoveries were 99.53％,99.41％,98.66％,98.31％ and 99.25％ with the RSDs of 1.02％,1.05％,1.26％,0.85％ and 1.43％,respectively.CONCLUSION This simple,accurate and reproducible method can be used for the quality control of Maxing Kanggan Granules.
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OBJECTIVE:To establish a method for simultaneous determination of (R,S)-goitrin and chlorogenic acid in Banchai oral liquid.METHODS:The dual-wavelength HPLC method was adopted.The determination was performed on Wondasil C18 column with mobile phase consisted of acetonitrile-0.1％ phosphoric acid (11∶89,V/V) at the flow rate of 1.0 mL/min.The detection wavelengths were 245 nm for (R,S)-goitrin and 327 nm for chlomgenic acid.The column temperature was 30 ℃,and injection volume was 10 μL.RESULTS:The linear ranges were 4.05-40.51 μg/mL for (R,S)-goitrin (r=0.999 9),29.41-294.05 μg/mL for chlorogenic acid (r=0.999 9).The limits of quantification were 3.32,2.45 ng,limits of detection were 1.00,0.74 ng.RSDs of precision,stability and reproducibility tests were lower than 1.0％;the recoveries were 98.46％-101.06％ (RSD=0.98％,n=9) and 98.18％-100.78％ (RSD=0.86％,n=9).CONCLUSIONS:The method is simple,accurate,sensitive and reproducible,and can be used for simultaneous determination of (R,S)-goitrin and chlorogenic acid in Banchai oral liquid.
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OBJECTIVE:To establish a method for simultaneous determination of (R,S)-goitrin and chlorogenic acid in Banchai oral liquid.METHODS:The dual-wavelength HPLC method was adopted.The determination was performed on Wondasil C18 column with mobile phase consisted of acetonitrile-0.1％ phosphoric acid (11∶89,V/V) at the flow rate of 1.0 mL/min.The detection wavelengths were 245 nm for (R,S)-goitrin and 327 nm for chlomgenic acid.The column temperature was 30 ℃,and injection volume was 10 μL.RESULTS:The linear ranges were 4.05-40.51 μg/mL for (R,S)-goitrin (r=0.999 9),29.41-294.05 μg/mL for chlorogenic acid (r=0.999 9).The limits of quantification were 3.32,2.45 ng,limits of detection were 1.00,0.74 ng.RSDs of precision,stability and reproducibility tests were lower than 1.0％;the recoveries were 98.46％-101.06％ (RSD=0.98％,n=9) and 98.18％-100.78％ (RSD=0.86％,n=9).CONCLUSIONS:The method is simple,accurate,sensitive and reproducible,and can be used for simultaneous determination of (R,S)-goitrin and chlorogenic acid in Banchai oral liquid.
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Objective: Taking thewater extraction of Compound Banlangen Liyan Granules (IsatidisRadix, ScrophulariaeRadix, PlatycodiRadix, GlycyrrhizaeRadix, etc.)as the research object to explore the dynamic migration regular of the active ingredient population of compoundChinesemateriamedica in different apertures in ceramic membrane separation process and compare its membrane filtration flux and solid inclusion removal rate. Methods: The water extraction of Compound Banlangen Liyan Granules was separated with 200 nm and 50 nm aperture of ceramic membrane, respectively. Continuously sampling for many times was done in the ceramic membrane separation process. The optimized HPLC methods were as follows: chromatographic column was Agilent Zorbox Eclipse XDB-C18, mobile phase was acetonitrile-0.05% phosphoric acid in gradient elution at flow rate of 1 mL/min, detection wavelength was wavelength switching, injection volum was 10 μL, the column temperature was 30 ℃. The contents of sixactive ingredients[adenosine, (R,S)-goitrin, liquiritin, harpagosid, platycodinD, and monoammonium glycyrrhizinate]were determined in thecompound at the same time, and the dynamic migration rates were investigated. Results:The simultaneous determination of the sixactive ingredient in thecompound was done with HPLC method. For the water extraction in Compound Banlangen Liyan Granules, in the 200 nm ceramic membrane separation process, the dynamic migration rates of the sixactive ingredientsranged between 71%-104%, the average migration rate was 85%, the membrane filter flux attenuation was smaller, the stable flux was in 426-340 L/(m2∙h), solid inclusion removal rate was 21.0%. But in 50 nm ceramic membrane, the dynamic migration rates of the sixactiveingredients ranged between 83%-107%, the average migration rate was83%, the membrane filter flux attenuation was larger, the stable flux was in 258-228 L/(m2∙h), and solid inclusion removal rate was23.9%. Conclusion :The experiment preliminarily reflects the dynamic migration regular of the active ingredient population of compoundChinesemateriamedica in two different aperture ceramic membrane separation process. It lays a migration theoretical foundation of effective materials for popularization and application of modern ceramic membrane technology in Chinese materiamedica refining.
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Objective: To explore the relationship between chemical constituents of Isatidis Radix (IR)and the inhibition of bioactivity of neuraminidase (NA)on influenza virus in vitro. Methods: To establish the chemical fingerprints of IR extraction from different producing areas and determine the contents of common peaks by UPLC. To determine the activity of NA on influenza virus tested with IR by Kit. To analyze the correlation of the chemical information and biological effect by mathematical statistics methods such as grey correlation analysis and one-factor analysis. Results: The chemical contents and biological effect detection varied markedly in the products from different regions and by extracting methods. The result of comprehensive analysis showed that uridine and (R, S)-goitrin of common components in UPLC-fingerprints related with the inhibition of bioactivity of NA on influenza virus, and the content of (R, S)-goitrin was correlated significantly to the activity of NA. Conclusion: Based on the biological assay of NA on influenza virus, the correlation of the content of chemical components and biological effect is verified preliminarily, with a view to provide a more reliable data and reference for the selection and extraction of antiviral active ingredients fromIR.
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OBJECTIVE:To establish a method for the simultaneous determination of(R,S)-goitrin,chlorogenic acid,luteolo-side and isochlorogenic acid A in Xiaoer ganmao granule. METHODS:HPLC was performed on the column of Hedera C18 with mo-bile phase of acetonitrile-0.1% formic acid aqueous at a flow rate of 1.0 ml/min,the detection wavelength was 254 nm,330 nm and 370 nm,column temperature was 40 ℃,and the injection volume was 5 μl,RESULTS:The linear range was 6.6-105 μg/ml for (R,S)-goitrin (r=0.999 9),9-140 μg/ml for chlorogenic acid (r=0.999 9),9-144 μg/ml for luteoloside (r=0.999 8) and 9-138 μg/ml for isochlorogenic acid A(r=0.999 6);the limits of quantitation were 330 ng,450 ng,450 ng and 450 ng,limits of detection were 66 ng,90 ng,90 ng and 90 ng,respectively;RSDs of precision,stability and reproducibility tests were lower than 3%;recoveries were 95.01%-98.77%(RSD=1.48%,n=6),95.14%-98.91%(RSD=1.52%,n=6),95.11%-97.54%(RSD=0.93%,n=6) and 95.58%-99.63%(RSD=1.73%,n=6). CONCLUSIONS:The method is simple and accurate,and suitable for the simultaneous determination of(R,S)-goitrin,chlorogenic acid,luteoloside and isochlorogenic acid A in Xiaoer ganmao granule.
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OBJECTIVE:To establish the quality standard of Banlangen dispensing granules. METHODS:Thin layer chroma-tography(TLC)was used to identify Isatis indigotica,leucine and arginine,the items according to granules preparations and the ex-traction were detected. HPLC was conducted to determine the content of (R,S)-goitrin. The Shimadzu VP-ODS C18 column was used with the mobile phase of methanol-0.05% phosphoric acid (7∶93,V/V) at the flow rate of 1.0 ml/min. The detection wave-length was set at 245 nm,and the column temperature was room temperature. The sample size was 10 μl. RESULTS:TLC showed clear spots and good resolution. Particle size,moisture,dissolubility and microbial limit test were all conformed to the specifica-tion. The extraction limit was no less than 25.0%.(R,S)-goitrin showed a good linear relationship in the range of 1.565 9-50.109 8μg/ml(r=0.999 9,n=6). The average recovery was 99.25%(RSD=2.08%,n=9);the RSDs of precision,stability and repeatabili-ty tests were no more than 1.12%. The content of(R,S)-goitrin was initially determined as 0.35 mg/g. CONCLUSIONS:The estab-lished quality standard has increased content determination,and the control programs are more comprehensive and reasonable. It can be used for the quality control of Banlangen dispensing granules.
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Objective To establish a HPLC method for simultaneous determination of uracil,hypoxanthine,guanosine, thymine,and( R,S)-goitrin in Isatidis Radix. Methods The determination was performed on a Hanbon Hedera C18 column (4. 6 mm×250 mm,5 μm). The mobile phase consisted of acetonitrile( A)and water( B)with linear gradient elution at the flow rate of 1.0 mL·min-1. The column temperature was 30 ℃. Detection wavelength was 254 nm. Results The linear range of uracil,hypoxanthine,guanosine,thymine,and(R,S)-goitrin was 1.97-39.40 mg·L-1(r=0.999 9),1.25-50.00 mg·L-1(r=0.999 9),0.25-10.40 mg·L-1(r=0. 999 6),2. 84-56. 70 mg·L-1(r=0. 999 9),and 0. 72-28.80 mg·L-1(r=0.999 8), respectively. The average recovery was 99. 36%(RSD=0. 91%),99. 79%(RSD=1. 12%),100. 90%(RSD=1. 71%),98. 67%(RSD=1. 50%),and 99. 33%(RSD=0. 94%),respectively. Conclusion The method is simple,accurate,reliable,reproducible and sensitive,which can be used as an effective method for quality control of Isatidis Radix.