Development of a liquid chromatography with mass spectrometry method for the determination of gelsemine in rat plasma and tissue: Application to a pharmacokinetic and tissue distribution study

Gelsemine from Gelsemium elegans Benth is a potential anesthetic and analgesic agent with no physical dependence and opiate addiction. This study was aimed at developing an ultrafast liquid chromatography coupled to tandem mass spectrometry method to quantify gelsemine in rat plasma and tissues. Plasma and tissues were processed with acetonitrile precipitation, and dendrobine was chosen as the internal standard. Sample separation was performed on an ACQUITY HSS T3 column. The mobile phase consisted of acetonitrile and 0.1% formic acid aqueous solution. Multiple reactions monitoring mode was utilized to detect the compounds of interest. The mass spectrometer was operated in the positive ion mode for detection. The MS/MS ion transitions monitored were m/z 323.2→70.5 for gelsemine and 264.2→108.05 for dendrobine, respectively. The calibration curves were linear over the range of 1–500 ng/mL in all biological matrices. The lower limit of quantification for rats plasma and tissues was 1.0 ng/mL. The values for inter‐ and intraday precision and accuracy were well within the ranges acceptable (< 15%). It was successfully applied to the pharmacokinetic and tissue distribution studies of gelsemine after intravenous doses of 5, 2, and 0.5 mg/kg in rats. These data of gelsemine would be useful for clinical application and further development.     

Liquid chromatography-mass spectrometry method for the determination of venlafaxine in human plasma and application to a pharmacokinetic study

A sensitive and selective liquid chromatographic-mass spectrometric (LC-MS) method for the determination of venlafaxine in human plasma has been developed. Samples were prepared using liquid-liquid extraction and analyzed on a C(18) column interfaced with a triple quadrupole mass spectrometer. Positive electrospray ionization was employed as the ionization source. The mobile phase was methanol-water containing 10 mmol/L ammonium acetate, pH 7.9 adjusted with aqueous ammonia (80:20, v/v) at the flow rate of 1.0 mL/min. The analyte and internal standard clozapine were both detected by use of selected ion monitoring mode. The method was linear in the concentration range of 1.0-200.0 ng/mL. The lower limit of quantification (LLOQ) was 1.0 ng/mL. The intra- and inter-day relative standard deviation across three validation runs over the entire concentration range was less than 10.1%. The accuracy determined at three concentrations (5.0, 50.0 and 150.0 ng/mL for venlafaxine) was within +/-10.0% in terms of relative error (RE). The method was successfully applied for the evaluation of pharmacokinetic profiles of venlafaxine capsule in 20 healthy volunteers. The results show AUC, T(max), C(max) and T(1/2) between the testing formulation and reference formulation have no significant difference (p > 0.05). Relative bioavailability was 103.4 +/- 14.1%.     

Development and validation of liquid chromatography-mass spectrometric method for simultaneous determination of moxifloxacin and ketorolac in rat plasma: application to pharmacokinetic study

A highly sensitive, selective and rapid liquid chromatography–electrospray ionization mass spectrometry (LC‐MS) method has been developed and validated for simultaneous determination of moxifloxacin (MFX) and ketorolac (KTC) in rat plasma. Gemifloxacin (GFX) was used as an internal standard (IS). A simple protein precipitation method was used for the extraction of analytes from rat plasma. Effective chromatographic separation of MFX, KTC and GFX was achieved on a Kromasil C₁₈ column (100 × 4.6 mm, 5 µm) using a mobile phase consisting of acetonitrile–10 mm ammonium acetate (pH 2.5)–0.1% formic acid (50:25:25) in an isocratic elution, followed by detection with positive ion electrospray ionization mass spectrometry using target ions of [M + H]⁺ at m/z 402 for MFX, m/z 256 for KTC and m/z 390 for GFX in selective ion recording mode. The method was validated over the calibration range of 5–100 ng/mL for MFX and 10–6000 ng/mL for KTC. The method demonstrated good performances in terms of intra‐ and inter‐day precision (0.97–5.33%) and accuracy (93.91–101.58%) for both MFX and KTC, including lower and upper limits of quantification. The recoveries from spiked control samples were >75% for MFX and >79% for KTC. The matrix effect was found to be negligible and the stability data were within acceptable limits. Further, the method was also successfully applied to a single‐dose pharmacokinetic study in rats. This method can be extended to measure plasma concentrations of both drugs in human to understand drug interaction and adverse effects. Copyright © 2012 John Wiley & Sons, Ltd.     

Development of an ultra-performance liquid chromatography-electrospray ionization-Orbitrap mass spectrometry method for determination of xanthopurpurin in rat plasma and its application to pharmacokinetic study

A rapid and sensitive method was developed and validated for the quantitative determination of xanthopurpurin (XPP) in rat plasma using ultra-performance liquid chromatography-electrospray ionization-Orbitrap mass spectrometry. XPP inhibits IgE production and prevents peanut-induced anaphylaxis. The XPP and emodin (internal standard) were determined in negative ion mode with m/z 239.0350 → 211.0400 and 269.0455 → 241.0507, respectively. The separation process was achieved using an ACQUITY UPLC HSS T₃ column with acetonitrile and 0.1% formic acid in water (85:15). The linear range was 0.5–100 ng/mL, and the correlation coefficient (r²) was > 0.993. The inter-day and intra-day precision was within an acceptable range of 15%. The extraction recovery and matrix effect were 78.9–87.2% and 94.3–98.5%, respectively. Under different conditions, the XPP was stable in the range of 5.6–10.6%. This method was successfully applied to study the pharmacokinetics of XPP with an oral dose of 10.0 mg/kg and intravenous dose of 2.0 mg/kg in rats. The absolute oral bioavailability of XPP was 4.6%.     

Development and validation of a liquid chromatography with tandem mass spectrometry method for the determination of isomangiferin in rat plasma with application to a preclinical pharmacokinetic study

A sensitive and specific liquid chromatography with tandem mass spectrometry method was firstly developed for the measurement of isomangiferin in rat plasma. Chloramphenicol was selected as the internal standard. Sample preparation was carried out through a simple one‐step protein precipitation procedure with methanol. Negative electrospray ionization was performed using multiple reaction monitoring mode with transitions of m/z 421.1/301.1 for isomangiferin, and 321.1/151.9 for chloramphenicol. The calibration curve was linear over the range of 0.1–600 ng/mL, with a lower limit of quantification at 0.1 ng/mL. The intra‐ and interday precisions (relative standard deviation) were no more than 8.2% and accuracies (relative error) were within the range of –8.4 to 2.2%. The recovery, matrix effect and stability under different conditions were all proved acceptable. The validated method has been successfully applied to a preclinical pharmacokinetic study of isomangiferin in rats for the first time.     

Simultaneous determination of norisoboldine and its major metabolite in rat plasma by ultra-performance liquid chromatography-mass spectrometry and its application in a pharmacokinetic study

Norisoboldine (NIB) is one of the main bioactive isoquinoline alkaloids in Linderae Radix. A rapid, selective and sensitive method using UPLC‐ESI/MS was first developed for simultaneous determination of NIB and norisoboldine‐9‐O‐α‐glucuronide (NIB‐Glu), its major metabolite in rat plasma. A one‐step protein precipitation with methanol was employed as sample preparation technique. Chromatographic separation was carried out on an Acquity UPLC BEH C₁₈ column (50 × 2.1 mm, i.d. 1.7 µm) with a gradient mobile phase consisting of acetonitrile and water containing 0.1% formic acid. Detection and quantification were performed using a quadrupole mass spectrometer by selective ion reaction‐monitoring mode. Good linearity was achieved using weighted (1/x²) least squares linear regression over the concentration ranges 0.01–2 µg/mL for NIB and 0.025–25 µg/mL for NIB‐Glu. The lower limit of quantification of NIB and NIB‐Glu was 0.01 and 0.025 µg/mL, respectively. The intra‐ and inter‐day precisions (relative standard deviations) of the assay at all three quality control levels were 4.6–14.1% for NIB, and 5.0–12.2% for NIB‐Glu. The accuracies (relative error) were −13.5–8.1% for NIB and −12.8–7.6% for NIB‐Glu, respectively. This developed method was successfully applied to an in vivo pharmacokinetic study in rats after a single intravenous dose of 10 mg/kg NIB. Copyright © 2010 John Wiley & Sons, Ltd.     

High‐performance liquid chromatography method for determination of carnosic acid in rat plasma and its application to pharmacokinetic study

A sensitive and reproducible high‐performance liquid chromatography method with ultraviolet detection (UV) was developed for the determination of carnosic acid (CA) in rat plasma. After simple acidification and liquid–liquid extraction of plasma samples using gemfibrozil as an internal standard, the supernatant was evaporated to dryness under a gentle stream of nitrogen. The residue was reconstituted in 200 µL before being injected into the chromatographic system. The analysis was performed on a C₁₈ column protected by an ODS guard column using acetonitrile–0.1% phosphoric acid (55:45, v/v) as mobile phase, and the wavelength of the UV detector was set at 210 nm. The calibration curve was linear over the range of 0.265–265.0 µg/mL with a correlation coefficient of 0.9997. The recovery for plasma samples of 0.530, 13.25, 132.5 and 265.0 µg/mL was 72.2, 87.9, 90.4 and 94.7%, respectively. The intra‐day and inter‐day relative standard deviations for the measurements of quality control samples were less than 3.1%. The stability of the plasma samples was also validated. This method was successfully used to study the pharmacokinetics and bioavailability of CA in rats. Copyright © 2009 John Wiley & Sons, Ltd.     

A sensitive method for determination of furanodiene in rat plasma using liquid chromatography/tandem mass spectrometry and its application to a pharmacokinetic study

Furanodiene, a sesquiterpene component extracted from the essential oil of the rhizome of Curcuma wenyujin Y.H. Chen et C. Ling (Wen Ezhu), is widely used in traditional Chinese medicine. A sensitive analytical method was established and validated for furanodiene in rat plasma, which was further applied to assess the pharmacokinetics of furanodiene in rats receiving a single dose of furanodiene. Liquid chromatography tandem mass spectrometry (LC/MS/MS) in multiple reaction monitoring mode was used in the method and costundide was used as internal standard. A simple protein precipitation based on methanol was employed. The simple sample cleanup increased the throughput of the method substantially. The method was validated over the range of 1–1000 ng/mL with a correlation coefficient >0.99. The lower limit of quantification was 1 ng/mL for furanodiene in plasma. Intra‐ and inter‐day accuracies for furanodiene were 88–115 and 102–107%, and the inter‐day precision less than 14.4%. After a single oral dose of 10 mg/kg of furanodiene, the mean peak plasma concentration of furanodiene was 66.9 ± 23.4 ng/mL at 1 h, the area under the plasma concentration–time curve (AUC_{0–10 h}) was 220 ± 47.8 h ng/mL, and the elimination half‐life was 1.53 ± 0.06 h. After an intravenous adminstration of furanodiene at a dosage of 5 mg/kg, the area under the plasma concentration–time curve was 225 ± 76.1 h?ng/mL, and the elimination half‐life was 2.40 ± 1.18 h. Based on this result, the oral bioavailability of furanodiene in rats at 10 mg/kg is 49.0%. Copyright © 2011 John Wiley & Sons, Ltd.     

Development and validation of a reliable high-performance liquid chromatographic method for determination of nodakenin in rat plasma and its application to pharmacokinetic study

A simple and reliable high-performance liquid chromatographic (HPLC) method has been developed for the determination of nodakenin in rat plasma. The concentration of nodakenin was determined in plasma samples after deproteinization with methanol using hesperidin as internal standard. HPLC analysis was performed on a Diamonsil C(18) analytical column using acetonitrile-water (25:75, v/v) as the mobile phase and a UV detection at 330 nm. This method was validated in terms of recovery, linearity, accuracy and precision (intra- and inter-day variation). The extraction recoveries were 91.3 ± 10, 87.8 ± 4.8 and 92.6 ± 5.1 at concentrations of 0.500, 5.00 and 40.0 μg/mL, respectively. The standard curve for nodakenin was linear (r(2) ≥ 0.99) over the concentration range 0.250-50.0 μg/mL with a lower limit of quantification of 0.250 μg/mL. The intra- and inter-day precision (relative standard deviation, RSD) values were not higher than 12% and the accuracy (relative error, RE) was within ± 5.8% at three quality control levels. The validated method was successfully applied for the evaluation of the pharmacokinetics of nodakenin in rats after oral administration of Rhizoma et Radix Notopterygii decoction and nodakenin solution.     

Simultaneous determination of harpagoside and cinnamic acid in rat plasma by high-performance liquid chromatography: application to a pharmacokinetic study

Radix Scrophulariae (Xuanshen) is one of the famous Chinese herbal medicines widely used to treat rheumatism, tussis, pharyngalgia, arthritis, constipation, and conjunctival congestion. Harpagoside and cinnamic acid are the main bioactive components of Xuanshen. The purpose of this study was to develop an HPLC–UV method for simultaneous determination of harpagoside and cinnamic acid in rat plasma and investigate pharmacokinetic parameters of harpagoside and cinnamic acid after oral administration of Xuanshen extract (760 mg kg⁻¹). After addition of syringin as internal standard, the analytes were isolated from plasma by liquid–liquid extraction. Separation was achieved on a Kromasil C₁₈ column, and detection was by UV absorption at 272 nm. The described assay was validated in terms of linearity, accuracy, precision, recovery, and limit of quantification according to the FDA validation guidelines. Calibration curves for both analytes were linear with the coefficient of variation (r) for both was greater than 0.999. Accuracy for harpagoside and cinnamic acid ranged from 100.7–103.5% and 96.9–102.9%, respectively, and precision for both analytes were less than 8.5%. The main pharmacokinetic parameters found for harpagoside and cinnamic acid after oral infusion of Xuanshen extract were as follows: C _max 1488.7 ± 205.9 and 556.8 ± 94.2 ng mL⁻¹, T _max 2.09 ± 0.31 and (1.48 ± 0.14 h, AUC_0–24 10336.4 ± 1426.8 and 3653.1 ± 456.4 ng h mL⁻¹, 11276.8 ± 1321.4 and 3704.5 ± 398.8 ng h mL⁻¹, and t _1/2 4.9 ± 1.3 and 2.5 ± 0.9 h, respectively. These results indicated that the proposed method is simple, selective, and feasible for pharmacokinetic study of Radix Scrophulariae extract in rats. Figure Radix Scrophulariae     

Development and validation of a quantification method for oleanolic acid and hederagenin in rat plasma: application to the pharmacokinetic study

A rapid and sensitive liquid chromatography tandem mass spectrometry (LC–MS/MS) was developed and validated for simultaneous quantification of oleanolic acid and hederagenin in rat plasma. After the two analytes were extracted with liquid–liquid extraction, chromatographic separation was performed on a C₁₈ column with acetonitrile and water (85:15, v /v) as mobile phase at a flow rate of 0.4 mL/min. Calibration curves exhibited good linearity (r > 0.995) over the ranges of 0.41–82.0 ng/mL for oleanolic acid and 0.32–64.0 ng/mL for hederagenin, respectively. The lower limit of quantifications (LLOQs) in plasma were 0.41 ng/mL for oleanolic acid and 0.32 ng/mL for hederagenin. The established LLOQs were within the concentration needed for the assay in plasma, which met the requirements to evaluate their pharmacokinetics of oleanolic acid and hederagenin. This developed assay was successfully applied in the pharmacokinetic study of oleanolic acid and hederagenin in rats after oral administration of Rhizoma Clematidis extract.     

Validated liquid chromatography mass spectrometry method for quantitative determination of strictosamide in dog plasma and its application to pharmacokinetic study

A simple, sensitive and specific high‐performance liquid chromatography mass spectrometry (LC‐MS) method was developed and validated for the quantification of strictosamide in dog plasma. Strictosamide and internal standard (IS, ranolazine) extracted by liquid–liquid extraction with ethyl acetate were separated on a C₁₈ column using a gradient elution program. The detection was performed by selected ion monitoring mode via a positive electrospray ionization interface. The LLOQ was 1.0 ng/mL and the method exhibited acceptable precision, extraction efficiency and matrix effect. Finally, this proposed method was successfully applied to dog pharmacokinetic study and yielded the most comprehensive data on systemic exposure of strictosamide to date. Copyright © 2011 John Wiley & Sons, Ltd.     

High-performance liquid chromatographic determination of 18 alpha-glycyrrhetinic acid and 18 beta-glycyrrhetinic acid in rat plasma: application to pharmacokinetic study.

A high-performance liquid chromatographic method for the separation and determination of 18 alpha-glycyrrhetinic acid and 18 beta-glycyrrhetinic acid has been developed. Indomethacin was used as an internal standard. The drugs were separated on a reversed-phase column and detected by UV detection at a wavelength of 254 nm. Methanol-water-perchloric acid-ammonia (80:20:0.4:0.4, v/v) was used as the mobile phase at pH of 7.0-7.5. The detection limit of both compounds was 0.1 microgram/ml in rat plasma. The method was applied to pharmacokinetic studies of glycyrrhetinic acids in rats. The results suggest that the pharmacokinetics appeared to be non-linear in nature.     

A high-performance liquid chromatographic method for determination of scopolin in rat plasma: application to pharmacokinetic studies

An analytical method based on high-performance liquid chromatographic (HPLC) with ultraviolet (UV) detection was developed for determination of scopolin in rat plasma using aesculin as internal standard (IS). After protein precipitation of plasma sample with methanol, the supernatant was directly injected and analyzed. Chromatographic separation was achieved on a C18 column using methanol and distilled water (22:78, v/v) containing 0.2% (v/v) glacial acetic acid as mobile phase with a column temperature of 30 degrees C. The UV detector was set at 338 nm. The calibration curve was linear over the range of 0.105-13.125 microg/mL with a correlation coefficient of 0.9998. The retention times of aesculin and scopolin were 10.4 and 12.8 min, respectively. The recoveries for plasma samples of 0.105, 4.725 and 13.125 microg/mL were 91.08, 95.30 and 96.10%, respectively. The RSD of intra- and inter-day assay variations was less than 7.35%. The lower limit of detection was 0.03 microg/mL .This HPLC assay is a simple, sensitive and accurate and was successfully applied to the pharmacokinetic study of scopolin in rats.     

Quantitative determination of sauchinone in rat plasma by liquid chromatography-mass spectrometry

A rapid, sensitive and specific method using liquid chromatography with tandem mass spectrometric detection (LC‐MS) was developed for the analysis of sauchinone in rat plasma. Di‐O‐methyltetrahydrofuriguaiacin B was used as internal standard (IS). Analytes were extracted from rat plasma by liquid–liquid extraction using ethyl acetate. A 2.1 mm i.d. × 150 mm, 5 µm, Agilent Zorbax SB‐C₁₈ column was used to perform the chromatographic analysis. The mobile phase was methanol–deionized water (80:20, v/v). The chromatographic run time was 7 min per injection and the flow‐rate was 0.2 mL/min. The tandem mass spectrometric detection mode was achieved with electrospray ionization interface in positive‐ion mode (ESI⁺). The m/z ratios [M + Na]⁺, m/z 379.4 for sauchinone and m/z 395.4 for IS were recorded simultaneously. Calibration curve were linear over the range of 0.01–5 µg/mL. The lowest limit of quantification was 0.01 µg/mL. The intra‐day and inter‐day precision and accuracy of the quality control samples were 2.94–9.42% and 95.79–108.05%, respectively. The matrix effect was 64.20–67.34% and the extraction recovery was 93.28–95.98%. This method was simple and sensitive enough to be used in pharmacokinetic research for determination of sauchinone in rat plasma. Copyright © 2011 John Wiley & Sons, Ltd.     

Highly sensitive method for the determination of omeprazole in human plasma by liquid chromatography-electrospray ionization tandem mass spectrometry: application to a clinical pharmacokinetic study

A highly sensitive, rapid assay method has been developed and validated for the estimation of omeprazole (OPZ) in human plasma with liquid chromatography coupled to tandem mass spectrometry with electrospray ionization in the positive-ion mode. The assay procedure involves alkalinization of plasma followed by simple liquid-liquid extraction of OPZ and lansoprazole (internal standard, IS) from human plasma with acetonitrile. Chromatographic separation was achieved with 0.01 M ammonium acetate:acetonitrile (40:60, v/v) at a flow rate of 0.25 mL/min on an Inertsil ODS 3 column with a total run time 2.5 min. The MS/MS ion transitions monitored were 346.1 --> 198.1 for OPZ and 370.1 --> 252.1 for IS. Method validation and clinical sample analysis were performed as per FDA guidelines and the results met the acceptance criteria. The lower limit of quantitation achieved was 0.05 ng/mL and the linearity was observed from 0.05 to 10.0 ng/mL. The intra-day and inter-day precisions were in the ranges 2.09-8.56 and 5.29-8.19%, respectively. This novel method has been applied to a pharmacokinetic study of OPZ in humans.     

A liquid chromatography-mass spectrometry method for the quantitation of aristololactam-I in rat plasma

A sensitive method with liquid chromatography-electrospray ionization mass spectrometry has been developed and validated for the determination of aristololactam-I in rat plasma after oral administration of aristolochic acid-I using finesteride as the internal standard. Chromatographic separation was achieved on a Lichrospher C(18) column using methanol:0.05% acetic acid in water (71:29, v/v) as a mobile phase delivered at a flow rate of 1 mL/min. The assay was linear for aristololactam-I over the range 0.3-300 ng/mL. The analysis of quality control samples demonstrated precision with coefficient of variation less than 20% (n = 5). Absolute recovery of aristololactam-I was 90.4-97.3%. The LC-MS method for the determination of aristololactam-I is sensitive, specific and can be used to investigate the toxicokinetics of aristololactam-I.     

A solid-phase extraction method for high-performance liquid chromatographic determination of salvianolic acid B in rabbit plasma: application to pharmacokinetic study

A sensitive solid-phase extraction/high-performance liquid chromatographic method with ultraviolet detection was established for the analysis of salvianolic acid B in rabbit plasma. The analyte was separated on a reversed-phase column with trifluoroacetic acid-methanol-acetonitrile (70:10:20, v/v/v) as mobile phase at a flow rate of 1 mL/min, and ultraviolet detection at 315 nm. The calibration curve for salvianolic acid B was linear over the range 35-1400 microg/L with coefficients of correlation >0.999. The inter-day and intra-day precisions of analysis were <15%, and assay accuracy ranged from 95.3 to 109.1%. This method is suitable for determining salvianolic acid B in plasma and thus investigating the pharmacokinetics of salvianolic acid B.     

A high performance liquid chromatography-tandem mass spectrometric method for the determination of mefenamic acid in human plasma: application to pharmacokinetic study

In the present study, the development and validation of an LC‐MS/MS method for quantifying mefenamic acid in human plasma is described. The method involves liquid–liquid extraction using diclofenac as an internal standard (IS). Chromatographic separation was achieved on a Thermo Hypurity C₁₈, 50 × 4.6 mm, 5 µm column with a mobile phase consisting of 2 m m ammonium acetate buffer and methanol (pH 4.5 adjusted with glacial acetic acid; 15:85, v/v) at a flow‐rate of 0.75 mL/min and the total run time was 1.75 min. Analyte was introduced to the LC‐MS/MS using an atmospheric pressure ionization source. Both the drug and IS were detected in negative‐ion mode using multiple reaction monitoring m/z 240.0 → 196.3 and m/z 294.0 → 250.2, respectively, with a dwell time of 200 ms for each of the transitions. The standard curve was linear from 20 to 6000 ng/mL. This assay allows quantification of mefenamic acid at a concentration as low as 20 ng/mL in human plasma. The observed mean recovery was 73% for the drug. The applicability of this method for pharmacokinetic studies has been established after successful application during a 12‐subject bioavailibity study. Copyright © 2012 John Wiley & Sons, Ltd.     

Development and validation of a reversed-phase HPLC method for determination of nitrendipine in rat plasma: application to pharmacokinetic studies

A simple and sensitive method for the determination of nitrendipine in rat plasma was developed using high-performance liquid chromatography (HPLC). The procedure involves extraction of nitrendipine in dichloromethane/sodium hydroxide, followed by reversed phase HPLC using a Waters, Spherisorb ODS2 (250 x 4.6 mm, 5 microm) column and UV detection at 238 nm. The retention times of nitrendipine and internal standard (felodipine) were 5.0 min and 7.5 min, respectively. The calibration curves were linear over the range of 5 ng/mL (lower limit of quantification, LOQ) to 200 ng/mL for nitrendipine. The intra- and inter-day coefficients of variation for all criteria of validation were less than 15% over the linearity range. The sensitivity and precision of the method were within the accepted limits (< 15%) throughout the validation period. The present method was also successfully applied for the study of plasma pharmacokinetics of nitrendipine loaded solid lipid nanoparticles (SLN) in rats.