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.     

A liquid chromatography/tandem mass spectrometry method for determination of aristolochic acid‐I in rat plasma

A sensitive, rapid and specific liquid chromatography–electrospray ionization–tandem mass spectrometry method was developed and validated for the determination of aristolochic acid‐I (AA‐I) in rat plasma. Finasteride was used as the internal standard (IS). The analyte was separated on a Zorbax Eclipse XDB‐C₁₈ column by isocratic elution with methanol‐10 mM ammonium acetate (75:25, v/v, pH = 7.3) at a flow rate of 0.25 mL/min, and analyzed by mass spectrometry in positive multiple reaction monitoring mode. The precursor‐to‐product ion transitions of m/z 359.0 → 298.2 and m/z 373.1 → 305.2 were used to detect AA‐I and IS, respectively. Good linearity was achieved over a range of 0.4–600 ng/mL. Intra‐ and inter‐day precisions measured as relative standard deviation were less than 13.5%, and accuracy ranged from 94.2 to 97.5%. The developed method was successfully applied in the pharmacokinetic study of AA‐I in rats. Copyright © 2009 John Wiley & Sons, Ltd.     

Development of a sensitive liquid chromatography/tandem mass spectrometry method for the determination of fenofibric acid in rat plasma

A rapid and sensitive LC‐MS/MS method for the quantification of fenofibric acid in rat plasma was developed and validated. Plasma samples were prepared by liquid–liquid extraction with a mixture of N‐hexane–dichloromethane–isopropanol (100:50:5, v/v/v). Isocratic chromatographic separation was performed on a reversed‐phase Discovery C₁₈ column (2.1 × 50 mm, 5 µm). The mobile phase was methanol–water–formic (75:25:0.25, v/v/v). Detection of fenofibric acid and the internal standard (IS) diclofenac acid was achieved by ESI MS/MS in the negative ion mode using m/z 317 → m/z 213 and m/z 294 → m/z 250 transitions, respectively. The method was linear from 0.005 to 1.250 µg/mL when 100 μL plasma was analyzed. The lower limit of quantification was 0.005 µg/mL. The intra‐ and inter‐day precision values were below 8.2%, and accuracy ranged from ?0.9 to 2.1% in all quality control samples. The recovery was 90.3–94.7% and 83.3% for fenofibric acid and IS, respectively. Total run time for each sample analysis was 2.5 min. The validated method was successfully applied to a pharmacokinetic study in six rats after oral administration of fenofibrate, the ester prodrug of fenofibric acid (equivalent to fenofibric acid 5 mg/kg). The method permits laboratory scientists with access to the appropriate instrumentation to perform rapid fenofibric acid determination. Copyright © 2011 John Wiley & Sons, Ltd.     

A sensitive and specific liquid chromatography/tandem mass spectrometry method for determination of echinacoside and its pharmacokinetic application in rats

A rapid and sensitive method based on liquid chromatography/tandem mass spectrometry (LC/MS/MS) for the determination of echinacoside in rat plasma was established and fully validated. A single step of liquid–liquid extraction with n‐butanol was utilized. Chromatographic separation of the analyte and the internal standard (IS), chlorogenic acid, from the sample matrix was performed using a Capcell‐MG C₁₈ analytical column (100 2.0 mm × 5 µm), with a gradient of acetonitrile and water containing 0.1% acetic acid as the mobile phase. Detection was performed on a triple quadrupole tandem mass spectrometer equipped with electrospray ionization source operated in negative ion selected reaction monitoring mode. The method was linear in the concentration range 10–2500 ng/mL. The deviations of both intra‐ and inter‐day precisions (RSD) were 7.1% and the assay accuracies were within 99.2–106.5%. Echinacoside proved to be stable during sample storage, preparation and analysis when an antioxidant solution was used. The method was successfully applied to a pharmacokinetic study in rats after an intragastric administration of echinacoside (100 mg/kg). With the lower limit of quantification at 10 ng/mL, this method proved to have sufficient selectivity, sensitivity and reproducibility for the pharmacokinetic study of echinacoside. Copyright © 2009 John Wiley & Sons, Ltd.     

Determination of bakkenolide A in rat plasma using liquid chromatography/tandem mass spectrometry and its application to a pharmacokinetic study

A sensitive rapid analytical method was established and validated to determine the bakkenolide A (BA) in rat plasma. This method was further applied to assess the pharmacokinetics of BA in rats receiving a single dose of BA. Liquid chromatography tandem mass spectrometry 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 combination of a simple sample cleanup and short chromatographic running time (2.4 min) 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 BA in plasma. Intra‐ and inter‐day accuracies for BA were 93–112% and 103–104%, respectively, and the inter‐day precision was less than 15%. After a single oral dose of 20 mg/kg of BA, the mean peak plasma concentration (C_max) of BA was 234.7 ± 161 ng/mL at 0.25 h. The area under the plasma concentration–time curve (AUC_{0–24 h}) was 535.8 ± 223.7 h·ng/mL, and the elimination half‐life (T_1/2) was 5.0 ± 0.36 h. In case of intravenous administration of BA at a dosage of 2 mg/kg, the area under the plasma concentration–time curve (AUC_{0–24 h}) was 342 ± 98 h?ng/mL, and the elimination half‐life (T_1/2) was 5.8 ± 0.7 h. Based on the results, the oral bioavailability of BA in rats at 20 mg/kg is 15.7%. Copyright © 2012 John Wiley & Sons, Ltd.     

Rapid quantification of lisinopril in human plasma by liquid chromatography/tandem mass spectrometry

An assay based on protein precipitation and liquid chromatography/tandem mass spectrometry (LC-MS/MS) has been developed and validated for the quantitative analysis of lisinopril in human plasma. After the addition of enalaprilat as internal standard (IS), plasma samples were prepared by one-step protein precipitation using perchloric acid followed by an isocratic elution with 10 mm ammonium acetate buffer (pH adjusted to 5.0 with acetic acid)-methanol (70:30, v/v) on a Phenomenex Luna 5 mu C(18) (2) column. Detection was performed on a triple-quadrupole mass spectrometer utilizing an electrospray ionization (ESI) interface operating in positive ion and selected reaction monitoring (SRM) mode with the precursor to product ion transitions m/z 406 --> 246 for lisinopril and m/z 349 --> 206 for enalaprilat. Calibration curves of lisinopril in human plasma were linear (r = 0.9973-0.9998) over the concentration range 2-200 ng/mL with acceptable accuracy and precision. The limit of detection and lower limit of quantification in human plasma were 1 and 2 ng/mL, respectively. The validated LC-MS/MS method has been successfully applied to a preliminary pharmacokinetic study of lisinopril in Chinese healthy male volunteers.     

A sensitive and specific liquid chromatography/tandem mass spectrometry method for determination of pinaverium bromide in human plasma: application to a pharmacokinetic study in healthy volunteers

A sensitive and specific method using liquid chromatography–electrospray tandem mass spectrometry (LC‐MS/MS) for the determination of pinaverium bromide in human plasma was developed and validated. Pinaverium bromide and an internal standard (paclitaxel) were isolated from plasma samples by precipitating plasma, and determined by LC‐MS/MS in multiple‐reaction monitoring mode. The main metabolite of pinaverium bromide and endogenous substances in plasma did not show any interference. The calibration curve was linear over the plasma concentration range of 10.0–10000.0 pg/mL with a correlation coefficient of 0.9979. The relative standard derivations intra‐ and inter‐day at 30.0, 300.0 and 8000.0 pg/mL in plasma were less than 15%. The absolute recoveries of pinaverium bromide and the internal standard were 99.7–111.7 and 106.2%, respectively. The lower limit of quantitation was 10 pg/mL. The analytical method was successfully applied to study the pharmacokinetics of pinaverium bromide tablets in healthy Chinese volunteers. Copyright © 2011 John Wiley & Sons, Ltd.     

Rapid and sensitive liquid chromatography tandem mass spectrometry method for the quantification of ambroxol in human plasma

A sensitive, specific and rapid high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was described and validated for the quantification of ambroxol in human plasma using enalaprilat as the internal standard (IS). Chromatographic separation was performed on a Lichrospher CN column with a mobile phase of methanol and water (containing 0.1% formic acid) (70:30, v/v). The total run time was 5.0 min for each sample. The analytes was detected by mass spectrometry with electrospray ionization source in positive selected reaction monitoring mode. The precursor-fragment ion reaction for ambroxol was m/z 378.9 --> 263.8, and for IS was m/z 349.0 --> 205.9. The linearity was established over the concentration range of 1.56-400.00 ng/mL. The inter-day and the intra-day precisions were all within 10%. A simple protein precipitation with methanol was adopted for sample preparation. The extraction recoveries of ambroxol and IS were higher than 90.80%. The validated method was successfully applied in pharmacokinetic study after oral administration of 90 mg ambroxol to 24 healthy volunteers.     

Quantitative analysis of acetaminophen and its six metabolites in rat plasma using liquid chromatography/tandem mass spectrometry

Acetaminophen (APAP) is a widely used analgesic and antipyretic drug. It is mainly metabolized by phase 1 and 2 reactions in the liver, and thus it could be involved in many drug–drug interactions. Therefore, the study of APAP metabolism is important in toxicological and pharmacokinetic studies. The objective of this study was to develop a rapid and sensitive method for the determination of APAP and its six metabolites in rat plasma for the pharmacokinetic studies. APAP and its metabolites were separated through a Capcell Pak MGII C₁₈ column and quantitated with a 16 min run in a triple‐quadruple mass spectrometer. The mobile phases were composed of 0.1% formic acid in either 95% water or 95% acetonitrile and analysis was performed twice in positive and negative modes. Validations such as accuracy, precision, recovery, matrix effect and stability were found to be within acceptance criteria of validation guidelines, indicating that the assay was applicable to the determination of the plasma concentrations of drug and its six metabolites. In conclusion, we developed an LC‐MS/MS method for the quantitative analysis of APAP and its six metabolites in rat plasma, and this method appears to be useful for pharmacokinetic/toxicokinetic studies of APAP and its metabolites in rats. Copyright © 2012 John Wiley & Sons, Ltd.     

Simultaneous determination of four shanzhiside methylester derivatives in rabbit plasma by liquid chromatography/tandem mass spectrometry

A simple and sensitive high‐performance liquid chromatography/tandem mass spectrometry (LC/MS/MS) method was developed and validated for simultaneous determination of shanzhiside methylester and its three derivatives in rabbit plasma. The method showed good linearity and no endogenous material interfered with the marked compounds and internal standard (IS) capatol peaks. Samples were processed by acetonitrile precipitation. Chromatography was performed using a C₁₈ column (150 × 3.9 mm i.d., 4 µm). The mobile phase consisted of methanol and water (60:40, v/v) during a total run time of 7 min. The main mass parent ions and daughter ions pairs (m/z) for monitoring were: shanzhiside methylester, 429.0/267.4; 8‐O‐acetyl shanzhiside methylester, 470.9/411.3; loganin, 413.2/251.4; phloyoside II, 479.2/281.3; and IS 385.2/203.3. Finally, the method was applied to a pharmacokinetic study of rabbits following intravenous administration of iridoid glycosides extracted from traditional herb Lamiophlomis rotata. Copyright © 2012 John Wiley & Sons, Ltd.     

Rapid and sensitive determination of acetylsalicylic acid and salicylic acid in plasma using liquid chromatography–tandem mass spectrometry: application to pharmacokinetic study

A simple and sensitive analytical method using liquid chromatography–tandem mass spectrometry (LC/MS/MS) for determination of acetylsalicylic acid (aspirin, ASA) and its major metabolite, salicylic acid (SA), in animal plasma has been developed and validated. Both ASA and SA in plasma samples containing potassium fluoride were extracted using acetonitrile (protein precipitation) with 0.1% formic acid in it. 6‐Methoxysalicylic acid was used as the internal standard (IS). The compounds were separated on a reversed‐phase column. The multiple reaction monitoring mode was used with ion transitions of m/z 178.9 → 136.8, 137.0 → 93.0 and 167.0 → 123.0 for ASA, SA and IS, respectively. The lower limits of quantification for ASA and SA were 3 and 30 ng/mL, respectively. The developed method was successfully applied for the evaluation of pharmacokinetics of ASA and SA after p.o. and i.v. administration of 1 mg/kg to rats. Copyright © 2009 John Wiley & Sons, Ltd.     

Simultaneous quantitation of acetylsalicylic acid and clopidogrel along with their metabolites in human plasma using liquid chromatography tandem mass spectrometry

The interest in therapeutic drug monitoring has increased over the last few years. Inter‐ and intra‐patient variability in pharmacokinetics, plasma concentration related toxicity and success of therapy have stressed the need of frequent therapeutic drug monitoring of the drugs. A sensitive, selective and rapid liquid chromatography coupled with tandem mass spectrometry (LC‐MS/MS) method was developed for the simultaneous quantification of acetylsalicylic acid (aspirin), salicylic acid, clopidogrel and carboxylic acid metabolite of clopidogrel in human plasma. The chromatographic separations were achieved on Waters Symmetry Shield^TM C₁₈ column (150 × 4.6 mm, 5 µm) using 3.5 mm ammonium acetate (pH 3.5)–acetonitrile (10:90, v/v) as mobile phase at a flow rate of 0.75 mL/min. The present method was successfully applied for therapeutic drug monitoring of aspirin and clopidogrel in 67 patients with coronary artery disease. Copyright © 2015 John Wiley & Sons, Ltd.     

Quantification of lurasidone, an atypical antipsychotic drug, in rat plasma with high-performance liquid chromatography with tandem mass spectrometry

A liquid chromatography–tandem mass spectrometric (LC/MS/MS) method was developed for the determination of an atypical antipsychotic drug, lurasidone, in rat plasma. The method involves the addition of acetonitrile and ziprasidone (internal standard) solution to plasma samples, followed by centrifugation. An aliquot of the supernatant was diluted with water and directly injected into the LC/MS/MS system. The separations were performed on a column packed with octadecylsilica (5 μm, 2.0 × 50 mm) with 0.1% formic acid and 0.1% formic acid in acetonitrile as mobile phase and the detection was performed using tandem mass spectrometry by multiple‐reaction monitoring via an electrospray ionization source. The standard curve was linear (r = 0.9982) over the concentration range 0.002–1 μg/mL. The intra‐ and inter‐assay precisions were 1.7 and 8.6%, respectively. The accuracy range was from 90.3 to 101.8%. The lower limit of quantification was 2.0 ng/mL using 50 μL of rat plasma sample. The developed analytical method was successfully applied to the pharmacokinetic study of lurasidone in rats. Copyright © 2011 John Wiley & Sons, Ltd.     

Determination of clopidogrel in human plasma by liquid chromatography/tandem mass spectrometry: application to a clinical pharmacokinetic study

A rapid and sensitive LC/MS/MS assay was developed and validated for the determination of clopidogrel in human plasma. Clopidogrel was extracted by single liquid-liquid extraction with pentane, and chromatographic separations were achieved on a C(18) column. The method was validated to demonstrate the specificity, linearity, recovery, lower limit of quantification (LLOQ), stability, accuracy and precision. The multiple reaction monitoring was based on m/z transition of 322.2 --> 211.9 for clopidogrel and 264.1 --> 125.1 for ticlopidine (internal standard). The total analytical run time was relatively short (3 min), and the LLOQ was 10 pg/mL using 0.5 mL of human plasma. The assay was linear over a concentration range from 10 to 10,000 pg/mL (r > 0.999). The intra- and inter-day accuracies were 101.3-108.8 and 98.4-103.5%, respectively, and the intra- and inter-day assay precisions were 1.9-5.5 and 4.4-8.1%, respectively. The developed assay method was applied to a pharmacokinetic study in human volunteers after oral administration of clopidogrel at a dose of 150 mg.     

Determination of ecliptasaponin A in rat plasma and tissues by liquid chromatography‐tandem mass spectrometry

A sensitive, rapid and specific high‐performance liquid chromatography tandem mass spectrometry method (HPLC‐MS/MS) was developed to determine ecliptasaponin A in rat plasma and tissues after oral administration. Ginsenoside Rg1 was used as the internal standard (IS). The plasma and tissues samples were prepared by liquid‐liquid extraction with ethyl acetate and separated on an Eclipse Plus C₁₈ column (2.1 mm × 150 mm, 5 µm) at a flow rate of 0.4 mL/min using acetonitrile and water (containing 0.05% acetic acid) as the mobile phase. The tandem mass detection was carried out with eletrospray ionization in negative mode. Quantification was performed by using multiple reaction monitoring (MRM), which monitored the fragmentation of m/z 633.4→587.2 for ecliptasaponin A and m/z 859.4→637.4 for the IS. The calibration curves obtained were linear in different matrices, and the lower limit of quantification (LLOQ) achieved was 0.5 ng/mL both for rat plasma and tissues. The intra‐ and inter‐day precisions were below 15%. This method was successfully applied to pharmacokinetic study of ecliptasaponin A in rat plasma and tissues after oral administration. Copyright © 2015 John Wiley & Sons, Ltd.     

Liquid chromatography tandem mass spectrometry method for determination of fulvestrant in rat plasma and its application to pharmacokinetic studies of a novel fulvestrant microcrystal

Fulvestrant (‘Faslodex’), an estrogen receptor antagonist, is available for the treatment of advanced breast cancer. The oil-based vehicle of Faslodex can lead to various adverse effects. A novel fulvestrant microcrystal (aqueous suspension) was developed in this study to eliminate these adverse effects. A sensitive and robust liquid chromatography tandem mass spectrometry method was developed and validated for the determination of fulvestrant in rat plasma using supported-liquid extraction. The separation of fulvestrant was achieved on an Agilent SB-C₁₈ column (2.1 × 50 mm, 3.5 μm) with isocratic elution using fulvestrant-d3 as internal standard. Mass spectrometric detection was conducted in negative multiple reaction monitoring mode. Ion transitions were at m/z 605.5 → 427.5 for fulvestrant and m/z 608.5 → 430.5 for fulvestrant-d3. The excellent linearity was demonstrated over the range 0.05–100.0 ng/ml (r² = 0.99). The lower limit of quantitation was 0.05 ng/ml, which was superior to that reported in literature The method validation was evaluated by selectivity, accuracy, precision, recovery and matrix effect in agreement with the US Food and Drug Administration guidance. The method was successfully applied to a pharmacokinetic study of a novel fulvestrant microcrystal in rats after intramuscular administration. It revealed that the rate of absorption increases and the extent of absorption is constant with a decrease in microcrystal diameter.     

Pharmacokinetic study of puerarin in rat serum by liquid chromatography tandem mass spectrometry

A highly sensitive and specific atmospheric pressure chemical ionization liquid chromatography-tandem mass spectrometry method was developed for serum pharmacokinetic studies of puerarin in rats. Chromatography was carried out on a reversed-phase Phenomenex Synergi 4 microm Fusion-RP80 column (150 x 2.0 mm i.d.) using a mobile phase consisting of acetonitrile-water (10:90, v/v) in 10 mm NH(4)OAc with a flow rate of 0.2 mL/min. Puerarin was analyzed in the multiple reaction monitoring mode with a precursor/product ion transition of m/z 415/267. The method was demonstrated to be specific and sensitive, and a linear response was observed over a range of 2-5000 ng/mL in rat serum. The validated method was successfully applied to the characterization of the pharmacokinetics of puerarin in rat serum after oral administration to spontaneously hypertensive rats. The blood concentration-time profile of puerarin showed a rapid initial increase, reaching a maximum and then declining within 1 h. Puerarin could not be detected after 24 h. The main pharmacokinetic parameters for puerarin after oral administration were as follows: C(max) (3.54 +/- 2.03 mg/L), T(max) (0.68 +/- 0.37 h), AUC(0-t) (7.29 +/- 3.79 mg h/L), AUC(0-infinity) (9.17 +/- 4.87 mg h/L), T(1/2) (1.7 +/- 0.6 h), CL/F (7.24 +/- 4.27 L/h/kg) and V/F (17.88 +/- 13.55 L/h/kg).     

A rapid and sensitive liquid chromatography/positive ion tandem mass spectrometry method for the determination of cimetropium in human plasma by liquid-liquid extraction

We have developed and validated a simple detection system with high-performance liquid chromatography (HPLC) with positive ion electrospray ionization tandem mass spectrometry (ESI-MS/MS) for determining cimetropium levels in human plasma using scopolamine butyl bromide as an internal standard (I.S.). The acquisition was performed in the multiple reaction monitoring (MRM) mode, by monitoring the transitions: m/z 357.9 > 103.1 for cimetropium and m/z 359.9 > 103.1 for butyl-scopolamine. The method involves a simple single-step liquid-liquid extraction with dichloromethane. The analyte was chromatographed on an YMC C18 reversed-phase chromatographic column by isocratic elution with 10 mM ammonium formate buffer-methanol (19:81, v/v; adjusted to pH 4.0 with formic acid). The results were linear over the studied range (0.2-100 ng ml(-1)), with r2 = 1.0000, and the total analysis time for each run was 2 min. Intra- and interassay precisions were 0.70-8.54% and 1.08-4.85%, respectively, and intra- and interassay accuracies were 97.56-108.23% and 97.48-103.91%, respectively. The lower limit of quantification (LLOQ) was 0.2 ng ml(-1). At this concentration, mean intra- and interassay precisions were 8.54% and 4.85%, respectively, and mean intra- and interassay accuracies were 97.56% and 98.91%, respectively. The mean recovery ranged from 62.71 +/- 4.06 to 64.23 +/- 2.32%. Cimetropium was found to be stable in plasma samples under typical storage and processing conditions. The devised assay was successfully applied to a pharmacokinetic study of cimetropium bromide administered as a single oral dose (150 mg) to healthy volunteers.     

Liquid chromatography/tandem mass spectrometry method for quantification of trans-stilbene glycoside in rat plasma and its pharmacokinetic application

A rapid and sensitive liquid chromatography/tandem mass spectrometry (LC‐MS/MS) method was developed and validated for the quantification of trans‐stilbene glycoside (SG) in rat plasma. As trans‐SG can be rapidly isomerized under light exposure, trans‐SG plasma samples were prepared in the dark and assayed immediately. Trans‐SG and internal standard were extracted by protein precipitation. Chromatographic separation was achieved on a C₁₈ column with a gradient elution program. The detection of analytes was performed by negative ion via multiple reaction monitoring mode. The precursor‐to‐product ions of m/z 405.1 → 242.9 for trans‐SG and m/z 389.1 → 226.9 for polydatin (internal standard) were monitored. No interference of endogenous components was observed for any plasma samples that we studied.The method was linear over the concentration range of 1.0–1000.0 ng/mL with a good correlation coefficient. The lower limit of quantification was 1.0 ng/mL for trans‐SG. The intra and inter‐batch accuracy for trans‐SG in stable rat plasma samples ranged from 93.3 to 102.7% and the variation was less than 8.1%. The extraction recoveries of trans‐SG in rat plasma were from 102.8 to 112.4% and the matrix effects were also acceptable. This method was successfully applied to pharmacokinetic study of trans‐SG in rats after intravenous administration. Copyright © 2012 John Wiley & Sons, Ltd.     

Simultaneous determination of harpagoside and cinnamic acid in rat plasma by liquid chromatography electrospray ionization mass spectrometry and its application to pharmacokinetic studies

A simple and sensitive method was developed for the simultaneous quantification of harpagoside and cinnamic acid in rat plasma using high-performance liquid chromatography system coupled to a negative ion electrospray mass spectrometric analysis. The plasma sample preparation was a simple deproteinization by the addition of two volumes of acetonitrile. The analytes were separated on an Intersil C8-3 column (2.1 mm i.d.x250 mm, 5 microm) with acetonitrile-5 mm ammonium formate aqueous solution (60:40, v/v) as mobile phase at a flow-rate of 0.2 mL/min. Detection was performed on a quadrupole mass spectrometer equipped with electrospray ionization (ESI) source operated under selected ion monitoring (SIM) mode. [M+HCOO]- at m/z 539 for harpagoside, [M-H]- at m/z 147 for cinnamic acid and [M-H]- at m/z 137 for salylic acid (internal standard) were selected as detecting ions, respectively. The method was validated over the concentration range 7-250 ng/mL for harpagoside and 5-500 ng/mL for cinnamic acid. The lower limits of quantitation for harpagoside and cinnamic acid were 7 and 5 ng/mL, respectively. The intra- and inter-day precisions (RSD%) were within 9.5% and the assay accuracies (RE%) ranged from -5.3 to 3.0% for both analytes. Their average recoveries were greater than 86%. Both analytes were proved to be stable during all sample storage, preparation and analysis procedures. The method was successfully applied to the pharmacokinetic study of harpagoside and cinnamic acid following oral administration of Radix Scrophulariae extract to rats.