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@#Objective: To investigate the effect of isoimperatorin on histopathological and biochemical changes in acetic acid-induced colitis rats. Methods: Colitis was induced by intracolonic administration of acetic acid solution (4% v/v) in rats. Rats were divided into six groups including the sham group, the negative control group, the dexamethasone-treated group, and the groups treated with isoimperatorin (0.1, 1, and 10 mg/kg/d by gavage). The treatments were administered for three days and then colonic status was assessed by macroscopic, histopathological, and biochemical analyses. Results: Isoimperatorin significantly alleviated colonic damage in a dose-dependent manner and improved histological changes in rats with acetic acid-induced colitis. It also significantly reduced myeloperoxidase, TNF-α, IL-1β, and malodialdehyde levels. Conclusions: Isoimperatorin alleviates acetic acid-induced colitis in rats and may be a potential therapeutic agent for the treatment of colitis.
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Objective To optimize the supercritical CO2 extraction conditions of volatile oil from Wenjing Huoxue cataplasm. Methods On the basis of single factor investigation on the comprehensive score of extraction yield , osthole content and isoimperatorin, the effects of extraction temperature, pressure and time on the comprehensive score of extracted volatile oil were optimized by orthogonal design. Results In the single factor experiment, the factors that had a great influence on the comprehensive score of the extracted volatile oil were extraction temperature, extraction pressure and extraction time. The orthogonal experiment results showed that the extraction temperature and extraction pressure had a significant influence on the comprehensive score of volatile oil. The optimized extraction process was as follows: extraction temperature at 55 ℃, extraction pressure as 30 MPa, and extraction time as 2 h. Conclusion The extraction process optimized in this experiment is stable and feasible, which could be used for the extraction and preparation of the volatile oil.
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Objective To establish a HPLC fingerprints of Biyuanjing capsules. Methods The column was Agilent SB-C18(4.6mm×250 mm, 5 µm). The mobile phase was acetonitrile-water with gradient elution at a flow rate of 1.0 ml/min. The detection wavelength was 210 nm. The detection time was 80 min. Results The HPLC fingerprints of Biyuanjing capsules were established. Twenty common peaks were confirmed, of which, 15 peaks were belonging to each crude drug and 5 peaks were identified as chemical components. The overall similarity of the fingerprints of 10 batches of samples was above 90% comparing with the control. Conclusion This method can be used for the quality control of Biyuanjing capsules.
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Objective: To explore the difference of anti-inflammatory effects in vitro and in vivo between Canqiang, Tiaoqiang and Datouqiang, and to obtain the anti-inflammatory active substances by relation of fingerprint with pharmacodynamics. Methods: Dimethylbenzene for the induction of a mouse ear edema in vivo and LPS-stimulated RAW 264.7 cells in vitro were used to investigate the anti-inflammatory activity, inflammatory factors such as nitric oxide (NO), tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) were determined by NO kit and ELISA kit, and the gray correlation analysis method was used to analyze the correlation between the peak area and the anti-inflammatory activity of the fingerprint features of each sample. Results: Different commercial specifications presented inhibition on acute inflammation of ear edema and inflammatory factors NO, TNF-α and IL-6, and anti-inflammatory effects were Canqiang > Datouqiang > Tiaoqiang. The chemical composition represented by 40 characteristic peaks was related to anti-inflammatory activity, and 17 characteristic peaks were highly correlated with this effect, among which three peaks were known components, namely chlorogenic acid, ferulic acid and isoimperatorin. Conclusion: The anti-inflammatory effect of Canqiang was the most significant, which was consistent with Canqiang being the most expensive and effective in the market, also with traditional grading standards. This study initially obtained anti-inflammatory active substances were chlorogenic acid, ferulic acid and isoimperatorin.
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Objective: To establish HPLC fingerprint of Notopterygii Rhizoma et Radix with different commercial specifications and to provide the basis for the division of commercial grades and the quality control of Notopterygii Rhizoma et Radix on the market. Method: The market investigation and literature research were used to understand the existing situation of Notopterygii Rhizoma et Radix goods.Notopterygii Rhizoma et Radix goods were divided into three commercial specifications according to the source and appearance,such as Canqiang,Tiaoqiang and Datouqiang.Fingerprint of Notopterygii Rhizoma et Radix with different commercial specifications was established by HPLC-PDA,the mobile phase was consisted of acetonitrile-0.3% acetic acid in a gradient elution mode.Similarity evaluation system for chromatographic fingerprint of traditional Chinese medicine(version of 2004A) was used to confirm the common peaks and evaluate the similarity.SPSS 19.0 statistical software was used to make principal component analysis(PCA) for HPLC fingerprint pattern. Result: The common mode of fingerprint for Canqiang,Tiaoqiang and Datouqiang were established separately.A total of 22 common peaks were marked in Canqiang,23 common peaks were marked in Tiaoqiang,29 common peaks were marked in Datouqiang.The result of similarity evaluation and PCA showed that the quality of Notopterygii Rhizoma et Radix with the same commercial specification was stable.There were great differences in chemical compositions and their contents among Notopterygii Rhizoma et Radix with different commercial specifications. Conclusion: The fingerprint method can well distinguish commercial specifications of Notopterygii Rhizoma et Radix,and it can provide the basis for the division of commercial grades and the quality control of Notopterygii Rhizoma et Radix.
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Objective: To analyze the quality of Angelicae Dahuricae Radix (ADR) from national drug evaluation sampling, continuously improve and revise the standard determination method, and ensure the safety of public drug use, six index components of ADR medicinal material and decoction pieces were determined by combining HPLC fingerprint chromatography and quantitative analysis of multi-components by single marker (QAMS). Methods: The determination and exploratory study were carried out according to the National Standard. The establishment of steamed Angelicae dahurica HPLC fingerprint, method for multiple assessment of six index components to conduct a methodological survey determination of multi-index component content of 20 batches of Angelicae dahurica tablets, the final determination of the quality of Angelicae dahurica. Result:s A total of 239 batches of ADR were received from 189 enterprises, in which 225 batches were qualified and 14 batches were unqualified. The qualified rate was 94%. A method that determined content of six kinds of coumarin in ADR decoction pieces using imperatorin as control was established, which could solve the problem of difficulty in obtaining control and consuming long determination time. It would better indicate the drug quality. Conclusio:n The method we had established could be used in qualitative and quantitative analysis of oxypeucedanin hydrate, byakangelicin, oxypeucedanin, berapten, imperatorin and isoimperatorin in ADR decoction pieces. Through this random inspection, it is found that there were still some quality problems in ADR decoction pieces. The quality supervision of traditional Chinese medicine should be strengthened further.
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Objective To investigate the chemical constituents from the ethanol extract of Yaobitong Capsule. Methods The chemical constituents were isolated by repeated silica gel column, dynamic axial compression chromatography, Sephadex LH-20 column, and prep-HPLC. Their structures were identified on the basis of NMR spectral data analysis. Results Sixteen compounds were isolated and identified to be nodakenetin (1), columbianetin (2), ferulic acid (3), umbelliferone (4), bergaptol (5), xanthotoxin (6), isoimperatorin (7), scoparone (8), (Z)-6-hydroxy-7-methoxydihydroligustilide (9), 4-hydroxy-3-butylphthalide (10), 3-(3′-hydroxy)- butylphthalide (11), senkyunolide C (12), senkyunolide H (13), senkyunolide I (14), fraxidin (15), and isofraxidin (16). Conclusion Compounds 1—16 are isolated from Yaobitong Capsule for the first time, and compound 11 is obtained by the means of separation for the first time.
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Objective To establish a method for isoimperatorin in Fengshi Bitong capsule and to study the effects of different doses of 60Co irradiation on the content of isoimperatorin. Methods The Agilent C18 column(4.6 mm×150 mm, 5μm) was used with the mobile phase consisted of acetonitrile -water (41:59) ;The flow rate was 1.0 mL?min-1 ,the detection wavelength was at 310 nm,and the column temperature was at 30 ℃ . Results Isoimperatorin showed a good linear relationship within a range of 3.66896-183.448 μg? mL-1( r = 1.0000).The average rate of recovery was 96.42％, RSD = 2.61％.The content of isoimperatorin in Fengshi Bitong capsule did not change significantly when irradiated by 2, 5 and 10 kGy of 60Co. Conclusion The established methods can be used for isoimperatorin determination in Fengshi Bitong capsule with high specificity,accuration and feasibility.The influence was not significant when the irradiation dose was less than 10 kGy.
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AIM To establish an HPLC method for the simultaneous content determination of four constituents in Liujing Toutong Tablets (Angelicae dahuricae Radix,Magnoliae Flos,Ligustici Rhizoma et Radix,etc.).METHODS The analysis of 30％ ethanol extract of this drug was performed on a 35 ℃ thermostatic Waters C18 column (4.6 mm × 250 mm,5 μm),with the mobile phase comprising of methanol-4％ acetic acid flowing at 1.0 mL/min in a gradient elution manner,and the detection wavelength was set at 320 nm.RESULTS Puerarin,ferulic acid,imperatorin and isoimperatorin showed good linear relationships within the ranges of 60.6-303 μg/mL (r=0.999 9),1.59-7.95 μg/mL (r =0.999 9),1.57-7.85 μg/mL (r =0.999 9) and 0.752 5-3.762 5 μg/mL (r =0.999 7),whose average recoveries (RSDs) were 97.75％ (1.7％),97.68％ (2.3％),97.94％ (1.0％) and 98.29％ (1.6％),respectively.CONCLUSION This stable and reliable method can be used for the quality control of Liujing Toutong Tablets.
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Objective To develop a new method for content determination of chlorogenic acid, ferulic acid, and isoimperatorin in Compound Baizhi Zhitong Capsules through quantitative analysis of multi-components by single-marker (QAMS) method by setting imperatorin as internal standard substance. Methods HPLC was adopted. The chromatographic column of Thermo Hypersil GOLD (4.6 mm×250 mm, 5 μm) was used; the mobile phase was composed of acetonitrile and 0.2％ formic acid in gradient elution; the flow speed was 1 m L/min; the detection wavelength were 320 nm (chlorogenic acid, ferulic acid) and 300 nm (imperatorin and isoimperatorin). The relative analysis correction factors of imperatorin to other three components were established by setting imperatorin as internal standard substance. Results The relative analysis correction factors of chlorogenic acid, ferulic acid, and isoimperatorin in Compound Baizhi Zhitong Capsules were 1.16, 2.00, and 1.00, which had a high reproducibility. There was no significant difference between QAMS and external standard method. Conclusion The QAMS method for Compound Baizhi Zhitong Capsules by setting imperatorin as internal standard substance is accurate and feasible, which can be used as a useful quality control method for Compound Baizhi Zhitong Capsules.
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The study was aimed to establish a liquid chromatography-tandem mass spectrometric method for the determination of the docetaxel concentration in rat plasma, and study the effect of coumarin constituents (imperatorin, isoimperatorin and oxypeucedanin) in Angelica dahurica on pharmacokinetics of docetaxel.Plasma was precipitated with acetonitrile and determined by LC-MS method with Paclitaxel as an internal standard. The specificity, linearity, range, accuracy, precision and stability of the method were suitable for the determination of docetaxel in plasma.Six sprague-dawley rats in each group received intragastric administration of docetaxel (50 mg•kg⁻¹), oxypeucedanin (8 mg•kg⁻¹) combined with docetaxel (50 mg•kg⁻¹), imperatorin (15 mg•kg⁻¹) combined with docetaxel (50 mg•kg⁻¹), and isoimperatorin(15 mg•kg⁻¹) combined with docetaxel (50 mg•kg⁻¹).Their drug plasma concentration was determined by LC-MS with Paclitaxel as an internal standard to draw plasma concentration-time curve, and the phamacokinetic parameters were calculated by DAS 2.0. The results showed that the phamacokinetic parameters of docetaxel all had notable changes when combined with imperatorin, isoimperatorin, and oxypeucedanin, respectively. The phamacokinetic parameters AUC and Cmax were significantly increased, indicating that coumarin constituents in Angelica dahurica could promote the oral bioavailability of docetaxel, and their effects were in the following order: oxypeucedanin> isoimperatorin> imperatorin.
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Objective To establish an HPLC fingerprint and to determine six compounds in Chitong Xiaoyanling Granules (CXG) for reference of the effective quality control. Methods The analysis was carried out on an analytical column Dikma Luster ODS (250 mm × 4.6 mm, 5 μm) with gradient elution by methanol (A)-0.1% phosphoric acid solution (B) (0-15 min, 20%-30% A; 15-30 min, 30% A; 30-40 min, 30%-60% A; 40-55 min, 60% A), at the detection wavelengths of 254, 283, 274, and 300 nm and a flow rate of 1.0 mL/min. The column temperature was 30 ℃. Similarity evaluation software was used to evaluate the fingerprint of 10 batches of CXG, and the six marker components were quantified. Results The common mode of the fingerprint was set up with 18 common peaks, and six of them were identified by comparison with the reference. The similar degrees of 10 batches of samples were over 0.9, they were prim-O-glucosylcimifugin, 4'-O-β-D-glucosyl-5-O-methylvisamminol, pulegone, hesperidin, paeonol, and isoimperatorin. The linear ranges were 0.013-0.505 mg/mL (r = 0.999 8), 0.052-2.097 mg/mL (r = 0.999 2), 0.019-0.772 mg/mL (r = 0.998 9), 0.025-1.003 mg/mL (r = 0.999 1), 0.006-0.251 mg/mL (r = 0.999 5), and 0.014-0.576 mg/mL (r = 0.999 4) for prim- O-glucosylcimifugin, 4'-O-β-D-glucosyl-5-O-methylvisamminol, pulegone, hesperidin, paeonol, and isoimperatorin, respectively. The contents of prim-O-glucosylcimifugin, 4'-O-β-D-glucosyl-5-O-methylvisamminol, pulegone, hesperidin, paeonol, and isoimperatorin were 7.267-7.333, 4.260-4.522, 2.033-2.093, 12.234-12.771, 19.023-19.334, and 11.152-11.291 mg/g in 10 batches of samples, respectively. Conclusion The established method has high sensitivity and specificity, and can be used for the quality control of CXG.
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Objective To study the absorption features of isoimperatorin in intestine of rat.Methods Establish a single-pass intestinal perfusion model of rat,take phenolsulfonphthalein as a marker for the detection of isoimperatorin concentration from crude extracts of notopterygium,and observe the absorption features of isoimperatorin and its reference substance in intestine of rat.Results The content of isoimperatorin in crude extracts was (0.43±0.02)% (n=5).The absorption parameters of duodenum,jejunum,ileum,colon were (5.28±0.82),(4.47±0.56),(4.17±0.94),(4.32±0.68)×10-5 cm·s-1,respectively.There were no significant differences among them (P>0.05).Compared with the reference substance of isoimperatorin,crude extracts showed better absorption features.Conclusion Isoimperatorin from crude extracts have better characteristics of absorption.This study can provide theoretical basis for design of notopterygium oral formulation.
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AIM To investigate the effects of imperatorin and isoimperatorin on the expression of mouse liver cytochrome P450s and hepatic toxicity in mice.METHODS C57BL/6 mice were randomly divided into control and administration groups,which were treated with imperatorin or isoimperatorin by intragastric administration for two weeks.The effects of two compounds on mRNA expressions of major P450s isoforms were analyzed by RT-PCR.The P450 expression was determined by Western blot.The serum levels of glutamicpyruvic transaminase (GPT),glutamic-oxalacetic transaminase (GOT) and blood total bilirubin (TBIL) were detected by kits.The change of liver tissue was observed with HE staining.RESULTS The Cyp1a2 mRNA expression was significantly induced by 40 mg/kg imperatorin as compared with the control group.For isoimperatorin,the Cyp2c37 mRNA expression was significantly induced.Western blot results showed that CYP1 A2 expression was significantly induced by imperatorin.For isoimperatorin,the CYP2C and CYP2E1 expressions were significantly induced.Blood biochemical indices showed that 40 mg/kg isoimperatorin led to increased serum GOT and TBIL levels.Pathological analysis showed that both compounds (at the doses of 20 mg/kg and 40 mg/kg) could cause liver edema to a certain degree.CONCLUSION Imperatorin is the inducer of CYP1A2,while isoimperatorin is the inducer of CYP2C and CYP2E1.These two compounds (at the doses of 20 mg/kg and 40 mg/kg) can lead to damage for mouse liver.The toxicity of isoimperatorin is stronger than that of imperatorin.
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Objective: To establish an HPLC method for the determination of gastrodin, prim-O-glucosylcimifugin, 5-O-methylvisammioside, imperatorin and isoimperatorin in Yuzhen powder.Methods: A Phenomenex Gemini C18 column(250 mm×4.6 mm, 5μm) was used and the column temperature was 25℃.The flow rate was 1.0 ml·min-1 and the mobile phase consisted of methanol-water with gradient elution.The detection wavelength was 230 nm and the sample size was 5 μl.Results: The linear range of gastrodin was 2.68-214.0 μg·ml-1(r=0.999 9), and the average recovery was 100.2%(RSD=0.9%, n=6);the linear range of prim-O-glucosylcimifugin was 5.22-418.0 μg·ml-1(r=0.999 7), and the average recovery was 99.9%(RSD=0.9%, n=6);the linear range of 5-O-methylvisammioside was 4.57-365.8 μg·ml-1(r=0.999 5), and the average recovery was 99.7%(RSD=1.0%, n=6);the linear range of imperatorin was 5.22-417.2 μg·ml-1(r=0.999 6), and the average recovery was 99.0%(RSD=0.9%, n=6);the linear range of isoimperatorin was 5.29-423.2 μg·ml-1(r=0.999 8), and the average recovery was 100.2%(RSD=0.8%, n=6).Conclusion: The method is simple and accurate, and the contents of the five ingredients can be determined simultaneously.The method can be used for the quality control of Yuzhen powder.
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Objective: To study the effects of coumarins in Angelicae Dahuricae Radix (imperatorin, isoimperatorin, bergapten, and oxypeucedanin) on the activities of adenosine triphosphate (ATPase) in P-glycoprotein (P-gp) in vitro and to investigate the P-gp mediated meschanism in transport of drug. Methods: The ATPase activity assay was used to identify whether the coumarins in Angelicae Dahuricae Radix were P-gp substrates or not. Results: The average fluorescent values (ΔRLUTC) of imperation and oxypeucedanin at each concentration were greater than average basic fluorescence value (ΔRLUbasal). The average fluorescent values (ΔRLUTC) in medium and low concentration of isoimperatorin and bergapten groups were lower than average basic fluorescence value (ΔRLUbasal). Conclusion: Imperation and oxypeucedanun are non-concentration dependent P-gp ATP inhibitors. Isoimperatorin and bergamot lactones could inhibit the ATPase activity at low concentration and have the inhibitory effect on P-gp. At high concentration, the activity of ATPase can be induced so as to induce P-gp.
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Objective: To study the chemical constituents of ethyl acetate soluble parts in 70% ethanol aqueous extract from the roots of Angelica dahurica var. formosana. Methods: The compounds were separated and purified by repeated column chromatography on silica gel and HPLC, and their structures were determined by spectroscopic data analyses of MS and NMR. Results: Forty-two compounds were obtained and identified as imperatorin (1), phellopterin (2), bergaptol (3), heraclenol (4), falcarindiol (5), isoimperatorin (6), byakangelicin (7), psoralen (8), bergapten (9), isogosferol (10), japoangelol A (11), atractylenolactam (12), columbianadin (13), 5-hydroxy-8-methoxypsoralen (14), (8E)-1,8-heptadecadiene-4,6-diyne-3,10-diol (15), pabulenol (16), byakangelicin ethoxide (17), isobyakangelicol (18), umbelliferone (19), xanthotoxol (20), osthole (21), alloimperatorin (22), 5-(2- acetoxy-3-hydroxy-3-methylbutoxy) psoralen (23), oxypeucedanin hydrate (24), oxypeucedanin ethanolate (25), alloisoimperatorin (26), pabularinone (27), oxyalloimperatorin (28), isooxypeucedanin (29), adenine (30), trans-ferulic acid (31), vanillic acid (32), 2-ethoxy-2-(4-hydroxyphenyl)-ethanol (33), suberosin (34), isopimpinellin (35), xanthotoxin (36), ulopterol (37), 2,4- dihydroxybenzoic acid methylate (38), scoparone (39), swietenol (40), marmesin (41), and 5-hydroxymethylfurfural (42), respectively. Conclusion: Except for the compounds 1, 3, 6, 7, 10, 13, 14, 16, 21, 24, and 26, other compounds were isolated from the roots of A. dahurica var. formosana for the first time.
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OBJECTIVE:To establish a method for the simultaneous determination of coumarin compounds in Yifu pill. METH-ODS:HPLC was performed on the column of Inertsil ODS-3 with mobile phase of acetonitrile-water(49∶51,V/V) at a flow rate was 1.0 ml/min,the detection wavelength was 320 nm,column temperature was 30 ℃,injection volume was 20 μl. RESULTS:The linear range was 5.16-206.4μg/ml for osthole (r=0.999 9),37.40-1 495.89μg/ml for isoimperatorin(r=0.999 9) and 9.95-318.34 μg/ml for columbianadin(r=0.999 9);RSDs of precision,stability and reproducibility tests were lower than 3.0%;recov-eries were 96.05%-103.19%(RSD=2.18%,n=9) for osthole,96.90%-103.09%(RSD=2.07%,n=9) for isoimperatorin, 95.50%-103.57%(RSD=2.15%,n=9)for columbianadin. CONCLUSIONS:The method is simple with good precision,stability and reproducibility,and can be used for the simultaneous determination of coumarin compounds in Yifu pill.
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OBJECTIVE:To establish a method for the simultaneous determination of imperatorin and isoimperatorin in Qingxu-an tablet. METHODS:UPLC was performed on the column of ACQUITY UPLC BEH C18 with mobile phase of methanol-water (gradient elution)at a flow rate of 0.3 ml/min,the detection wavelength was 248 nm,column temperature was 30 ℃,and the in-jection volume was 3 μl. RESULTS:The linear range was 1.305-13.05 μg/ml for imperatorin(r=0.999 9)and 1.096-10.96 μg/ml for isoimperatorin(r=0.999 9);RSDs of precision,stability and reproducibility tests were lower than 1%;recoveries were 97.45%-100.10%(RSD=1.00%,n=9)and 98.33%-101.20%(RSD=1.01%,n=9),respectively. CONCLUSIONS:The method is accurate,fast and reliable,and can be used for the simultaneous determination of imperatorin and isoimperatorin in Qingxuan tablet.
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To study the heating treatment of Angelica dahurica under different temperature and time conditions on the stability of coumarins and tyrosinase activity. HPLC method was used to determine the contents of imperatorin and isoimperatorin, and tyrosinase activity was assayed by measuring the oxidation rate of L-DOPA in vitro. After heated, the contents of imperatorin increased. Expect for being heated at 90 ℃ for 2 h, the content of isoimperatorin was higher than crude one. Before and after being heated, A. dahurica showed an activating effect on tyrosinase. In the same temperature and time conditions, the activation rate increased with the rise of concentration of tyrosinase extracts. Heating process for A. dahurica could change the contents of imperatorin and isoimperatorin, mainly increasing their concentrations.