Determination and pharmacokinetic study of syringin and chlorogenic acid in rat plasma after administration of Aidi lyophilizer

A high-performance liquid chromatographic (HPLC) method was developed for the first time to simultaneously quantify syringin and chlorogenic acid in rat plasma using wavelength-transfer technology. The analysis was performed on a Diamonsil C(18) column (200 x 4.6 mm i.d., 5 microm particle size) with isocratic mobile phase consisting of acetonitrile-0.05% phosphoric acid (12:88, v/v). The linear ranges were 0.20-10 and 0.25-30 microg/mL, respectively. The lower limits of quantification were 0.20 and 0.25 microg/mL, respectively. The method was shown to be reproducible and reliable with intraday precision below 8.5 and 6.1%, interday precision below 7.1 and 5.5%, accuracy within +/-7.1 and +/-8.6%, and mean extraction recovery excess of 92.1 and 80.9%, respectively, which were all calculated from the blank plasma sample spiked with syringin and chlorogenic acid at three concentrations of 0.20, 1.0 and 6.0 microg/mL for syringin and 0.25, 2.0 and 20 microg/mL for chlorogenic acid. This method was validated for specificity, accuracy and precision and was successfully applied to the pharmacokinetic study of syringin and chlorogenic acid in rat plasma after intravenous administration of Aidi lyophilizer.     

Determination of dehydrodiisoeugenol in rat tissues using HPLC method

Dehydrodiisoeugenol (DDIE) is a bioactive neolignan from the seeds of Myristica fragrans Houtt., which exhibits good anti-inflammatory activity. A rapid and simple high-performance liquid chromatographic (HPLC) method has been developed for the determination of DDIE in rat tissues after intravenous administration. The tissue samples were processed by liquid-liquid extraction. The analyses were successfully carried out on a Diamonsiltrade mark ODS C18 column (250 x 4.6 mm i.d., 5 microm) equipped with a C18 guard column (8 x 4.6 mm i.d., 5 microm). The mobile phase was the system of methanol-water (4:1, v/v). The UV detection was set at 270 nm. The calibration curves were linear from 0.4 to 200.0 microg/g with the correlation coefficients (r(2)) greater than 0.998. The intra- and inter-day precisions in quality control samples were less than 10% and the accuracies were in the range 85.4-110.3%. The average recoveries from all the tissues were between 84.4 and 106.0%. This assay method has been successfully used to study the tissues distribution of DDIE in rats after intravenous administration. The result suggests that DDIE is distributed to rat tissues rapidly with possibly greater initial concentrations in liver and brain than in other tissues.     

Determination of piceatannol in rat serum and liver microsomes: pharmacokinetics and phase I and II biotransformation

A method of analysis of piceatannol in biological fluids is necessary to study the kinetics of in vitro and in vivo metabolism and determine its concentration in foodstuffs. A novel and simple high-performance liquid chromatographic method was developed for simultaneous determination of piceatannol and products of its metabolism in rat serum and liver microsomes. Serum, or microsomes (0.1 mL), were precipitated with acetonitrile after addition of the internal standard, 4-methylumbelliferone. Separation was achieved on a phenomenex C(18) column (250 x 4.6 mm i.d., 5 microm) equipped with a phenomenex C(18) (4 x 3.0 mm i.d., 5 microm) guardcolumn with fluorescence excitation at 320 nm and emission at 420 nm. Separation was also possible with UV detection at 310 nm. The fluorescent calibration curves were linear ranging from 0.05 to 100 microg/mL. The mean extraction efficiency was >95%. Precision of the assay was <10% (coefficient of variation), and was within 10% at the limit of quantitation (0.05 ng/mL). Bias of the assay was lower than 7%. The limit of detection was 50 ng/mL for a 0.1 mL sample. The assay was applied successfully to the in vitro kinetic study of metabolism of piceatannol in rat liver microsomes and pharmacokinetics in rats. Three metabolites of piceatannol have been identified. .     

Development and validation of a liquid chromatography/tandem mass spectrometry assay for the quantification of methyl protodioscin in rat plasma: application to a pharmacokinetic study

A high performance liquid chromatography/tandem mass spectrometry assay was first developed and validated for the quantification of methyl protodioscin (MPD), a natural furostanol saponin with distinct antitumor activity, in rat plasma with 17alpha-ethinylestradiol as internal standard (IS). Methanol-mediated protein precipitation was employed for plasma sample pretreatment. The separation was achieved on a C(18) column (150 x 4.6 mm, i.d., 5 microm) by isocratic elution with methanol-water (72:28, v/v) as mobile phase at a flow rate of 1.0 mL/min. Ion acquisition was performed in selective reaction monitoring positive mode by monitoring the transition of m/z 1085.7 --> 1053.7 for MPD, and in selective ion monitoring negative mode by monitoring the deprotonated ion m/z 295.5 for IS. The assay was linear over the concentration range of 2.024-270.0 microg/mL with 2.024 microg/mL as the lower limit of quantification. It was specific, accurate, precise and reproducible with intra- and inter-run RSD <8.3% and RE between -11.5 and 12.8%. The assay was successfully applied to a preclinical pharmacokinetic study after an intravenous dose of 40 mg/kg MPD to rats.     

固相萃取-高效液相色谱法测定人血浆中的川芎嗪

建立了高效液相色谱测定人血浆中川芎嗪浓度的方法。色谱条件:分析柱为Luna C18(150 mm×4.6 mm i.d.,5 μ m),流动相为甲醇-乙腈-醋酸盐缓冲液(pH 5.0)(体积比为50∶8∶42),流速1.0 mL/min,柱温40 ℃,检测波长280 nm。 血浆样品预处理采用C8固相小柱萃取法。方法的线性范围为25～5000 μg/L,线性相关系数为0.9999。高、中、低浓度 的川芎嗪在标准血浆样品中的平均提取回收率为96.72％～100.90％,日内和日间相对标准偏差(RSD)小于8.64％,准确度 为99.59%~103.26%,检测限为10 μg/L。该方法的各项效能指标符合生物样品的分析要求,可用于川芎嗪制剂的人体药 代动力学研究。     

Determination of picroside II in dog plasma by HPLC and its application in a pharmacokinetics study

A sensitive and simple high-performance liquid chromatography method with UV detection was developed and validated for determining picroside II in dog plasma. Paeoniflorin was employed as internal standard and the sample pre-treatment procedure consists of deproteinization by addition of acetonitrile. Chromatographic separations were performed on a Shimadzu VP-ODS column (250 x 4.6 mm i.d., 5 microm). The mobile phase consisted of acetonitrile-0.1% acetic acid aqueous (v/v), 23:77, v/v, at a rate of 1 mL/min. Detection was carried out at a wavelength of 266 nm. Calibration standards ranged from 0.25 to 500 microg/mL in dog plasma and the mean correlation coefficient of 0.9981 was found for the linear calibration curves (n = 6). The limit of quantification (LOQ) was 0.25 microg/mL. Intra- and inter-assay RSD ranged from 0.70 to 7.5%. Accuracy (%bias) ranged from -6.3 to 6.0%. This method was applied to the pharmacokinetic study of picroside II in dogs. The study demonstrated the plasma picroside II concentration-time curves were fitted to the two-compartment open model and showed linear pharmacokinetics.     

Simultaneous determination of caffeine, theophylline and theobromine in human plasma by on-line solid-phase extraction coupled to reversed-phase chromatography

A reversed-phase liquid chromatographic column switching system was described for the determination of caffeine (CF), theophylline (TH) and theobromine (TB) in human plasma with a direct injection procedure. A short protein-coated mu Bondapak CN silica pre-column (20 x 3 mm, i.d.) was used for enrichment of the drugs and clean up from weakly retained plasma components using phosphate buffer saline pH 7.4. After washing step, the retained drugs were flushed into a reversed-phase column (5 microm TSK gel ODS-80 TM, 150 x 4.6 mm i.d.) with a mobile phase of methanol-0.01 M phosphate buffer, pH 3.5 (30:70, v/v) for the final separation. The eluent was monitored with a UV detector at 275 nm. The resulting chromatograms showed no interference from endogenous plasma components. A linear relationship between the concentration of drug and peak height was confirmed in the range of 0.5-20 microg/mL for all drugs. High extraction recoveries from plasma ranging from 96.12 to 100.32% were achieved. Validation of the method was examined performing intra- and inter-day accuracy and precision and was found to be satisfactory. The coefficients of variation of the three drugs were less than 3% for intra-day and less than 4% for inter-day run assays.     

HPLC analysis and pharmacokinetics of icariin in rats

As a prerequisite to the determination of pharmacokinetic parameters of icariin in rats, an HPLC method using UV detection was developed and validated. Icariin and the internal standard, quercetin, were extracted from plasma samples using ethyl acetate after acidification with 0.05 mol/L NaH2PO4 solution (pH 5.0). Chromatographic separation was achieved on an Agilent XDB Cls column (250 x 4.6 mm id, 5 microm) equipped with a Shim-pack GVP-ODS C18 guard column (10 x 4.6 mm id, 5 microm) using a mobile phase of ACN/water/acetic acid (31:69:0.4 v/v/v) at a flow rate of 1.0 mL/ min. Detection was at 277 nm. The calibration curve was linear from 0.05 to 100.0 microg/mL with 0.05 microg/mL as the lower LOQ (LLOQ) in plasma. The intra- and interday precisions in terms of RSD were lower than 5.7 and 7.8% in rat plasma, respectively. The accuracy in terms of relative error (RE) ranged from -1.6 to 3.2%. The extraction recoveries of icariin and quercetin were 87.6 and 80.1%, respectively. The main pharmacokinetic parameters for rats were determined after a single intravenous administration of 10 mg/kg icariin: t1/2, 0.562 +/- 0.200 h; AUC0-infinity, 8.73 +/- 2.23 microg x h/mL; CLToT, 20.10 +/- 5.80 L/kg x h; Vz, 1.037 +/- 0.631 L/kg; MRT0-infinity, 0.134 +/- 0.040 h; and Vss, 0.170 +/- 0.097 L/kg.     

Simple determination of pirfenidone in rat plasma via high-performance liquid chromatography

A simple, rapid and reliable high-performance liquid chromatographic method was developed and validated for the determination of pirfenidone and its major metabolites in rat plasma. Plasma proteins were precipitated with perchloric acid (10%, v/v) and the supernatant after centrifugation was determined using high-performance liquid chromatography. The analysis was carried out on a Lichrospher C(18) column (250 x 4.6 mm i.d., 5 microm). The mobile phase consisted of acetonitrile-water containing 0.2% acetic acid (23:77, v/v) at a flow-rate of 1 mL/min. The eluant was detected at 310 nm. The calibration curves were linear over a concentration range from 0.15 to 76.67 microg/mL. The accuracy (relative error) of the assay ranged from -2.6 to 7.9% and the precision (coefficient of variation) was less than 4.5%. The established method has been successfully applied to a pharmacokinetic study of pirfenidone following a single oral dose to rats.     

High-performance liquid chromatographic method for the determination and pharmacokinetic study of wogonoside in rat serum after oral administration of traditional Chinese medicinal preparation Huang-Lian-Jie-Du decoction

A high-performance liquid chromatographic method for the determination of wogonoside in plasma of rats administrated orally with the traditional Chinese medicinal preparation Huang-Lian-Jie-Du decoction was developed. Sample preparation was carried out by protein precipitation with a mixture of acetonitrile and methanol (1:1, v/v). The extracted sample was separated on a Hypersil C(18) (150 x 5 mm i.d., 5 microm) analytical column by linear gradient elution using 0.05% (v/v) phosphoric acid (containing 5 mm sodium dihydrogen phosphate) and acetonitrile as mobile phase at a flow rate of 1.5 mL/min. The eluate was detected using a UV detector at 276 nm. The assay was linear over the range 0.109-7.0 microg/mL (R(2) = 0.9999, n = 5). Mean recovery was determined as 98.39%. Intra- and inter-day precisions (RSD) were < or =7.59%. The limit of quantitation was 0.109 microg/mL. After validation, the HPLC method developed was applied to investigate the preliminary pharmacokinetics of wogonoside in rat after oral administration of Huang-Lian-Jie-Du decoction.     

Determination of sodium cromoglycate in human plasma by liquid chromatography with tandem mass

A sensitive and selective liquid chromatography-tandem mass spectrometric (LC-MS/MS) method was developed and validated for the determination of sodium cromoglycate (SCG) in human plasma after a nasal dose of 10.4 mg sodium cromoglycate nasal spray, using pravastatin sodium as the internal standard. The method was validated over a linear range of 0.300-20.0 ng/mL. SCG and I.S. were extracted from 1.0 mL of heparinized plasma by C(18) solid-phase extraction cartridges using methanol as eluting solvent. The dried residue was reconstituted with 100 microL of mobile phase, and 10 microL was injected onto the LC-MS/MS system. Chromatographic separation was achieved on a C(18) column (250 x 4.6 mm i.d., 5 microm particle size) with a mobile phase of methanol-acetonitrile-water (containing 2 mmol/L ammonium acetate; 42.5:42.5:15, v/v/v) at a flow rate of 0.4 mL/min. The analytes were detected with a triple quad LC-MS/MS using ESI with positive ionization. Ions monitored in the multiple reaction monitoring mode were m/z 469.0 (precursor ion) to m/z 245.0 (product ion) for SCG and m/z 447.2 (precursor ion) to m/z327.1 (product ion) for pravastatin sodium (internal standard) The average recovery of SCG from human plasma was 94.88% and the lower limit of quantitation was 0.3 ng/mL. Results from a 3-day validation study demonstrated excellent precision and accuracy across the calibration range of 0.3-20 ng/mL. The method was successfully applied to the pharmacokinetic study of SCG in healthy Chinese volunteers. Copyright (c) 2008 John Wiley & Sons, Ltd.     

High-performance liquid chromatographic determination of doxazosin in human plasma for bioequivalence study of controlled release doxazosin tablets

A highly sensitive high-performance liquid chromatographic quantification method with fluorescence detection was developed and validated for the determination of doxazosin in human plasma. The developed method employed one-step extraction of doxazosin from plasma matrix with ethyl acetate using propranolol as an internal standard. Chromatographic separation was obtained within 8.0 min using a reverse-phase Capcell-Pak C(18) column (150 x 4.6 mm i.d., 5 microm) and the mobile phase consisted of methanol-water containing 10 mM perchloric acid and 1.8 mM sodium heptane sulfonic acid (50:50, v/v) and was set at a flow rate of 1.5 mL/min. The calibration curve constructed was linear in the range of 0.3-50.0 ng/mL. The proposed method achieved a lower limit of quantification of 0.3 ng/mL, better than the reported HPLC methods. Average recoveries of doxazosin and the internal standard from human plasma matrix were 87.0 and 85.9%, respectively. The present method was validated by evaluating the precision and accuracy for inter- and intraday variation in the concentration range 0.3-50 ng/mL. The precision values expressed as relative standard deviations in the inter- and intraday validation were 1.17-6.29 and 0.84-5.94%, respectively. This method was successfully applied to the bioequivalence study of two doxazosin controlled release tablets in healthy, male human subjects.     

HPLC analysis and pharmacokinetic study of quercitrin and isoquercitrin in rat plasma after administration of Hypericum japonicum thunb. extract

A simple HPLC method was developed for determination of quercitrin and isoquercitrin in rat plasma. Reversed-phase HPLC was employed for the quantitative analysis using kaempferol-3-O-beta-D-glucopyranoside-7-O-alpha-L-rhamnoside as an internal standard. Following extraction from the plasma samples with ethyl acetate-isopropanol (95:5, v/v), these two compounds were successfully separated on a Luna C(18) column (250 x 4.6 mm, 5 microm) with isocratic elution of acetonitrile-0.5% aqueous acetic acid (17:83, v/v) as the mobile phase. The flow-rate was set at 1 mL/min and the eluent was detected at 350 nm for both quercitrin and isoquercitrin. The method was linear over the studied ranges of 50-6000 and 50-5000 ng/mL for quercitrin and isoquercitrin, respectively. The intra- and inter-day precisions of the analysis were better than 13.1 and 13.2%, respectively. The lower limits of quantitation for quercitrin and isoquercitrin in plasma were both of 50 ng/mL. The mean extraction recoveries were 73 and 61% for quercitrin and isoquercitrin, respectively. The validated method was successfully applied to pharmacokinetic studies of the two analytes in rat plasma after the oral administration of Hypericum japonicum thunb. ethanol extract.     

Determination of atractylenolide II in rat plasma by reversed-phase high-performance liquid chromatography

A method for quantitative determination of atractylenolide II in rat plasma using reversed-phase high-performance liquid chromatography (RP-HPLC) coupled with UV spectrometry was established. From a variety of compounds and solvents tested, atractylenolide III was selected as the internal standard (IS) and ethyl acetate was found to be the best solvent for extracting atractylenolide II from plasma samples. RP-HPLC analysis of the extracts was performed on an analytical column (DIKMA ODS, 150 x 4.6 mm; i.d., 5 microm) equipped with a security guard pre-column system. There was good linearity over the range 0.05-5.0 microg/mL (r > 0.99). The recoveries were more than 90.0% in plasma, and the intra- and inter-day coefficients of variation were less than 10.0% in all cases. The limit of detection (LOD) was 0.025 microg/mL and the lower limit of quantification (LLOQ) was 0.05 microg/mL. The RP-HPLC method was applied to quantitate atractylenolide II in rat plasma within 24 h in a pharmacokinetics study where experimental rats received a single dose of atractylenolide II (60 mg/kg).     

Simultaneous determination of phenytoin and dextromethorphan in urine by solid-phase extraction and HPLC-DAD

A rapid and simple high-performance liquid chromatographic method with photodiode array detection was developed for the separation and the simultaneous determination of phenytoin and dextromethorphan in human urine. Analysis was performed in less than 4.5 min in isocratic mode on a reversed-phase C18 column (5 microm; 150 x 4.6 mm) using a mobile phase composed of acetonitrile-buffer phosphate 0.01 M (60:40, v/v) adjusted to pH 6.0, at 1 mL/min flow rate and UV absorbance at 210 nm. The elution order of analytes was dextromethorphan (DXM), Internal Standard (IS), and phenytoin (PHT). Calibration curves were linear in the 7.5-25 microg/mL range for PHT and in the 10-30 microg/mL range for DXM. Spike recoveries for urine samples prepared at three spiking levels ranged from 97.8 to 102.3% for PHT and from 94.8 to 100.4% for DXM. The detection limit (LOD) values ranged from 0.08 microg/mL for PHT to 0.5 microg/mL for DXM. The quantitation limit (LOQ) values ranged from 0.3 microg/mL for PHT to 1.6 microg/mL for DXM. The sample preparation method involves a rapid and simple procedure based on solid-phase extraction using a C18 reversed-phase column. Validation of the optimised method was carried out according to the ICH guidelines. The method developed in this study allows the reliable simultaneous analysis of PHT and DXM, drugs that were never quantified together in previously reported analytical methods. The described method has the advantage of being rapid and easy and it could be applied in therapeutic monitoring of these drugs in human urine of epileptic patients.     

高效液相色谱/四极杆-飞行时间质谱测定神经性贝毒

采用高效液相色谱/四极杆 飞行时间质谱(HPLC/Q TOFMS)联用技术对贝类样品中的短裸甲藻毒素 PbTx-2 进行了检测研究。样品经丙酮提取、C18小柱净化后,用Zorbax XDB C18色谱柱(2.1 mm i.d.×150 mm,3.5 μ m)进行分离,流动相为甲醇 水(体积比为85∶15)溶液(含0.5 mmol/L NH4Ac),流速0.20 mL/min 。电喷雾正离子模式,选择质子化PbTx 2分子离子 ［M+H］ +作为前体离子进行TOFMS扫描、测定。结果表明,样品的平均加标     

High performance liquid chromatographic method for the determination and pharmacokinetic studies of oxyresveratrol and resveratrol in rat plasma after oral administration of Smilax china extract

A sensitive and simple HPLC method has been developed and validated for the determination of oxyresveratrol (trans-2,4,3',5'-tetrahydroxystilbene, OXY) and resveratrol (trans-3,5,4'-trihydroxystilbene, RES) in rat plasma. The plasma samples were extracted with ethyl acetate and analyzed using HPLC on an Aglient Zorbax SB-C(18) column (250 x 4.6 mm, 5 microm) at a wavelength 320 nm, with a linear gradient of (A) acetonitrile and (B) 0.5% aqueous acetic acid (v/v), at a flow rate of 1.0 mL/min. The method was linear over the range of 0.1265-25.3 microg/mL for OXY and 0.117-23.4 microg/mL for RES. The extraction recovery for OXY, RES and internal standard ranged from 71.1 to 88.3%. The intra- and inter-day precisions were better than 10%, and the accuracy ranged from 89 to 108%. The validated method was used to study the pharmacokinetic profiles of OXY and RES in rat plasma after oral administration of Smilax china root extract.     

Determination of paeoniflorin in rat plasma by a liquid chromatography-tandem mass spectrometry method coupled with solid-phase extraction

A rapid, sensitive and selective liquid chromatography-tandem spectrometry method was developed and validated for determination of paeoniflorin in rat plasma using geniposide as the internal standard. The samples were pretreated with solid-phase extraction using Extract-Clean cartridges. Separation of paeoniflorin and IS was achieved on a reversed-phase C18 column (50x4.6 mm i.d.) with a mobile phase made up of acetonitrile and 0.05% formic acid (25:75, v/v) at a flow rate of 0.5 mL/min. Detection was carried out on a triple quadrupole tandem mass spectrometer by multiple-reaction monitoring and an electrospray ionization source was employed as the ionization source. The lower limit of quantification obtained was 4 ng/mL (n=6) using 200 microL plasma with an accuracy of -3.67% (relative error) and a precision of 4.13% (relative standard deviation). A good linearity was found in the range of 4-1000 ng/mL. The intra- and inter-day relative standard deviations in the measurement of quality control samples 10, 150 and 800 ng/mL ranged from 3.73 to 4.94% and from 4.31 to 6.56%, respectively. The accuracy was from -3.93 to -1.11% in terms of relative error. The analyte and IS were stable in the battery of stability studies. This method was successfully applied to a pharmacokinetic study of paeoniflorin after a single oral administration of 53.36 mg/kg paeoniflorin to rats.     

Direct-injection high performance liquid chromatography ion trap mass spectrometry for the quantitative determination of olanzapine,clozapine and N-desmethylclozapine in human plasma

A specific and sensitive direct-injection high performance liquid chromatography electrospray ionization tandem mass spectrometry (HPLC/ESI-MS/MS) method has been developed for the rapid identification and quantitative determination of olanzapine, clozapine, and N-desmethylclozapine in human plasma. After the addition of the internal standard dibenzepin and dilution with 0.1% formic acid, plasma samples were injected into the LC/MS/MS system. Proteins and other large biomolecules were removed during an online sample cleanup using an extraction column (1 x 50 mm i.d., 30 microm) with a 100% aqueous mobile phase at a flow rate of 4 mL/min. The extraction column was subsequently brought inline with the analytical column by automatic valve switching. Analytes were separated on a 5 microm Symmetry C18 (Waters) analytical column (3.0 x 150 mm) with a mobile phase of acetonitrile/0.1% formic acid (20:80, v/v) at a flow rate of 0.5 mL/min. The total analysis time was 6 min per sample. The inter- and intra-assay coefficients of variation for all compounds were <11%. By eliminating the need for extensive sample preparation, the proposed method offers very large savings in total analysis time.     

HPLC determination and pharmacokinetic study of tenatoprazole in dog plasma after oral administration of enteric-coated capsule

A simple, sensitive and selective high-performance liquid chromatographic (HPLC) method with UV detection (306 nm) was developed and validated for determination of tenatoprazole, a novel proton-pump inhibitor, in dog plasma. Tenatoprazole and internal standard (pantoprazole) were extracted into diethyl ether and separated using an isocratic mobile phase of 10 mm phosphate buffer (pH4.7)-acetonitrile (70:30, v/v) on a Diamonsil C(18) column (150 x 4.6 mm, 5 microm). The retention times for tenatoprazole and internal standard were 7.1 and 12.3 min, respectively. No endogenous interferences were observed. This HPLC method was fully validated. The lower limit of quantitation was 20 ng/mL, with a relative standard deviation of less than 20%. A linear range of 0.02-5.0 microg/mL was established. The interday and intraday precisions were within RSD 13.4-10.1 and 4.6-1.4%, respectively. This method developed can be easily applied to the pharmacokinetic study of tenatoprazole in dog plasma after oral administration of an enteric-coated capsule. The plasma concentration of tenatoprazole from six dogs showed a mean C(max) of 2.63 microg/mL at T(max) of 1.89 h. The bioavailability of tenatoprazole was improved by administration of enteric-coated capsule.