Quantification of homoegonol in rat plasma using liquid chromatography–tandem mass spectrometry and its pharmacokinetics application

Homoegonol is a biologically active neolignan isolated from Styrax species with cytotoxic, antimicrobial, anti‐inflammatory and anti‐asthma activities. For the quantification of homoegonol in rat plasma, a selective and sensitive liquid chromatography–tandem mass spectrometric method was developed and validated for the first time using protein precipitation with methanol as a sample clean‐up procedure. The analytes were separated in an Atlantis dC₁₈ column using a gradient elution of methanol and 0.1% formic acid, and mass‐to‐charge ratios were determined in selective reaction monitoring mode using tandem mass spectrometry with m/z 343.12 > 296.97 for homoegonol and m/z 517.30 > 282.90 for udenafil (internal standard). The standard curve was linear over the concentration ranges of 1 ? 500 ng/mL using a 30 μL rat plasma sample. The coefficient of variation and relative error for intra‐ and inter‐assay at four quality control levels were 3.9–10.0 and ‐3.3–2.7%, respectively. The overall recovery of homoegonol from rat plasma using protein precipitation was 99.7 ± 7.7%. The pharmacokinetics parameters of homoegonol were dose‐independent after both intravenous (1, 2.5 and 5 mg/kg doses) and oral (5, 10 and 20 mg/kg doses) administration in male Sprague–Dawley rats. Copyright © 2014 John Wiley & Sons, Ltd.     

Quantification of pramipexole in human plasma by liquid chromatography tandem mass spectrometry using tamsulosin as internal standard

A high-performance liquid chromatography/electrospray ionization tandem mass spectrometry method was developed and validated for the quantification of pramipexole in human plasma. Following liquid-liquid extraction, the analytes were separated using an isocratic mobile phase on a reverse-phase column and analyzed by MS/MS in the multiple reaction monitoring mode using the respective [M + H](+) ions, m/z 212/152 for pramipexole and m/z 409/228 for the IS. The method exhibited a linear dynamic range of 200-8000 pg/mL for pramipexole in human plasma. The lower limit of quantification was 200 pg/mL with a relative standard deviation of less than 8%. Acceptable precision and accuracy were obtained for concentrations over the standard curve range. A run time of 3.5 min for each sample made it possible to analyze more than 200 human plasma samples per day. The validated method has been successfully used to analyze human plasma samples for application in pharmacokinetic, bioavailability or bioequivalence studies.     

Liquid chromatography tandem mass spectrometry method for the quantification of rimonabant, a CB1 receptor antagonist, in human plasma

A sensitive high-performance liquid chromatography-tandem mass spectrometry method was developed and validated for the quantification of rimonabant in human plasma. Following liquid-liquid extraction, the analytes were separated using an isocratic mobile phase on a reverse-phase column and analyzed by MS/MS in the multiple reaction monitoring mode using the respective (M+H)+ ions, m/z 463-363 for rimonabant and m/z 408-235 for the internal standard. The assay exhibited a linear dynamic range of 0.1-100 ng/mL for rimonabant in human plasma. The lower limit of quantification was 0.1 ng/mL with a relative standard deviation of less than 6%. With dilution integrity up to 10-fold, the upper limit of quantification was extendable up to 1000 ng/mL. Acceptable precision and accuracy were obtained for concentrations over the standard curve range. A run time of 2.0 min for each sample made it possible to analyze more than 250 human plasma samples per day. The validated method was successfully used to analyze human plasma samples for application in pharmacokinetic studies.     

Quantification of granisetron in human plasma by liquid chromatography coupled to electrospray tandem mass spectrometry

A simple, sensitive and rapid high-performance liquid chromatography/electrospray ionization tandem mass spectrometry method was developed and validated for the assay of granisetron in human plasma. Following liquid-liquid extraction, the analytes were separated using an isocratic mobile phase on a reversed-phase C18 column and analyzed by MS in the multiple reaction monitoring mode using the respective [M+H]+ ions, m/z 313/138 for granisetron and m/z 409/228 for the IS. The assay exhibited a linear dynamic range of 0.1-20 ng/mL for granisetron in human plasma. The lower limit of quantification was 100 pg/mL with a relative standard deviation of less than 5%. Acceptable precision and accuracy were obtained for concentrations over the standard curve range. A run time of 2.0 min for each sample made it possible to analyze more than 400 human plasma samples per day. The validated method has been successfully used to analyze human plasma samples for application in pharmacokinetic, bioavailability or bioequivalence studies.     

Determination of the neuropharmacological drug nodakenin in rat plasma and brain tissues by liquid chromatography tandem mass spectrometry: Application to pharmacokinetic studies

A rapid and sensitive liquid chromatography tandem mass spectrometry detection using selected reaction monitoring in positive ionization mode was developed and validated for the quantification of nodakenin in rat plasma and brain. Pareruptorin A was used as internal standard. A single step liquid–liquid extraction was used for plasma and brain sample preparation. The method was validated with respect to selectivity, precision, accuracy, linearity, limit of quantification, recovery, matrix effect and stability. Lower limit of quantification of nodakenin was 2.0 ng/mL in plasma and brain tissue homogenates. Linear calibration curves were obtained over concentration ranges of 2.0–1000 ng/mL in plasma and brain tissue homogenates for nodakenin. Intra‐day and inter‐day precisions (relative standard deviation, RSD) were <15% in both biological media. This assay was successfully applied to plasma and brain pharmacokinetic studies of nodakenin in rats after intravenous administration.     

Rapid and simple liquid chromatography tandem mass spectrometry method for the quantification of zidovudine in rat plasma

A high-performance liquid chromatography/electrospray ionization tandem mass spectrometry method was developed and validated for the quantification of zidovudine in rat plasma. Following solid-phase extraction, the analytes were separated using an isocratic mobile phase on a reverse phase column and analyzed by MS/MS in the multiple reaction monitoring mode using the respective [M+H]+ ions, m/z 268/127 for zidovudine and m/z 230/112 for the internal standard. The method exhibited a linear dynamic range of 5-500 ng/mL for zidovudine in rat plasma. The lower limit of quantification was 5 ng/mL with a relative standard deviation of less than 8%. Acceptable precision and accuracy were obtained for concentrations over the standard curve range. A run time of 1.5 min for each sample made it possible to analyze more than 400 plasma samples per day. The validated method was applied for pharmacokinetic studies of the novel drug delivery systems of zidovudine in rats.     

Quantification of fexofenadine in human plasma by liquid chromatography coupled to electrospray tandem mass spectrometry using mosapride as internal standard

A rapid high-performance liquid chromatography/positive ion electrospray tandem mass spectrometry method was developed and validated for the quantification of fexofenadine in human plasma using mosapride as internal standard. Following solid-phase extraction, the analytes were separated using an isocratic mobile phase on a reverse-phase column and analyzed by MS/MS in the multiple reaction monitoring mode using the respective [M+H]+ ions, m/z 502/466 for fexofenadine and m/z 422/198 for the IS. The method exhibited a linear dynamic range of 1-500 ng/mL for fexofenadine in human plasma. The lower limit of quantification was 1 ng/mL with a relative standard deviation of less than 5% for fexofenadine. Acceptable precision and accuracy were obtained for concentrations over the standard curve range. The total chromatographic run time of 2 min for each sample made it possible to analyze more than 400 human plasma samples per day. The validated method has been successfully used to analyze human plasma samples for application in pharmacokinetic, bioavailability or bioequivalence studies.     

Sensitive liquid chromatography tandem mass spectrometry method for the quantification of sitagliptin, a DPP-4 inhibitor, in human plasma using liquid-liquid extraction

A sensitive high-performance liquid chromatography-positive ion electrospray tandem mass spectrometry method was developed and validated for the quantification of sitagliptin, a DPP-4 inhibitor, in human plasma. Following liquid-liquid extraction, the analytes were separated using an isocratic mobile phase on a reverse-phase column and analyzed by MS/MS in the multiple reaction monitoring mode using the respective [M + H](+) ions, m/z 408-235 for sitagliptin and m/z 310-148 for the internal standard. The assay exhibited a linear dynamic range of 0.1-250 ng/mL for sitagliptin in human plasma. The lower limit of quantification was 0.1 ng/mL with a relative standard deviation of less than 6%. Acceptable precision and accuracy were obtained for concentrations over the standard curve range. A run time of 2.0 min for each sample made it possible to analyze more than 300 human plasma samples per day. The validated method has been successfully used to analyze human plasma samples for application in pharmacokinetic studies.     

Development and validation of a liquid chromatography–tandem mass spectrometry method for the determination of febuxostat in human plasma

A rapid and sensitive analytical method based on liquid chromatography coupled to tandem mass spectrometry detection with positive ion electrospray ionization was developed for the determination of febuxostat in human plasma using d₇‐febuxostat as the internal standard (IS). A simple protein precipitation was performed using acetonitrile. The analyte and IS were subjected to chromatographic analysis on a Capcell PAK C₁₈ column (4.6 × 100 mm, 5 µm) using acetonitrile–5 mm ammonium acetate–formic acid (85:15:0.015, v/v/v) as the mobile phase at a flow rate of 0.6 mL/min. An Agilent 6460 electrospray tandem mass spectrometer was operated in the multiple reaction monitoring mode. The precursor‐to‐product ion transitions m/z 317 → m/z 261 (febuxsotat) and m/z 324 → m/z (261 + 262) (d₇‐febuxostat, IS) were used for quantitation. The results were linear over the studied range (10.0–5000 ng/mL), and the total analysis time for each chromatograph was 3 min. The intra‐ and inter‐day precisions were less than 7.9 and 7.2%, respectively, and the accuracy was within ±4.2%. No evidence of analyte instability in human plasma was observed storage at ?20°C for 31 days. This method was successfully applied in the determination of febuxostat concentrations in plasma samples from healthy Chinese volunteers. Copyright © 2012 John Wiley & Sons, Ltd.     

Simultaneous determination of 2‐arachidonoylglycerol, 1‐arachidonoylglycerol and arachidonic acid in mouse brain tissue using liquid chromatography/tandem mass spectrometry

Endocannabinoids (ECs), such as anandamide (AEA) and 2‐arachidonoylglycerol (2‐AG), modulate a number of physiological processes, including pain, appetite and emotional state. Levels of ECs are tightly controlled by enzymatic biosynthesis and degradation in vivo. However, there is limited knowledge about the enzymes that terminate signaling of the major brain EC, 2‐AG. Identification and quantification of 2‐AG, 1‐AG and arachidonic acid (AA) is important for studying the enzymatic hydrolysis of 2‐AG. We have developed a sensitive and specific quantification method for simultaneous determination of 2‐AG, 1‐AG and AA from mouse brain and adipose tissues by liquid chromatography/tandem mass spectrometry (LC/MS/MS) using a simple brain sample preparation method. The separations were carried out based on reversed phase chromatography. Optimization of electrospray ionization conditions established the limits of detection (S/N = 3) at 50, 25 and 65 fmol for 2‐AG, 1‐AG and AA, respectively. The methods were selective, precise (%R.S.D. < 10%) and sensitive over a range of 0.02–20, 0.01–10 and 0.05–50 ng/mg tissue for 2‐AG, 1‐AG and AA, respectively. The quantification method was validated with consideration of the matrix effects and the mass spectrometry (MS) responses of the analytes and the deuterium labeled internal standard (IS). The developed methods were applied to study the hydrolysis of 2‐AG from mouse brain extracts containing membrane bound monoacylglycerol lipase (MAGL), and to measure the basal levels of 2‐AG, 1‐AG and AA in mouse brain and adipose tissues. Copyright © 2009 John Wiley & Sons, Ltd.     

Development and validation of dried blood spots technique for quantitative determination of topiramate using liquid chromatography–tandem mass spectrometry

An LC‐MS/MS method for determination of the anti‐epileptic drug topiramate (TPM) in dried blood spot (DBS) samples was developed and validated. DBS samples were prepared by spotting 30 μL of spiked whole blood onto FTA^TM DMPK‐C cards and drying for at least 3 h. Six‐millimetre punched spots were then extracted by using a mixture of methanol and water (90:10, v/v) with deuterated internal standard (topiramate‐d₁₂). The extracted samples were injected into a liquid chromatograph equipped with a tandem mass spectrometric detector. Negative ions were monitored in the selected reaction monitoring mode and transitions m/z 338.2 → 78.1 and m/z 350.3 → 78.1 were used for the quantitative evaluation of TPM and internal standard, respectively. The results obtained from validation were statistically evaluated according to the requirements of the European Medicines Agency and US Food and Drug Administration regulatory guidelines. The linearity of the method was checked within a concentration range from 10 to 2000 ng/mL. The validation results indicate that the method is accurate, precise, sensitive, selective and reproducible. Copyright © 2013 John Wiley & Sons, Ltd.     

A simple and sensitive assay for the quantitative analysis of paclitaxel in human and mouse plasma and brain tumor tissue using coupled liquid chromatography and tandem mass spectrometry

The development and validation of an assay for the determination of paclitaxel in human plasma, human brain tumor tissue, mouse plasma and mouse brain tumor tissue is described. Paclitaxel was extracted from the matrices using liquid-liquid extraction with tert-butyl methyl ether, followed by chromatographic analysis using an alkaline eluent. Positive ionization electrospray tandem mass spectrometry was performed for selective and sensitive detection. The method was validated according to the FDA guidelines on bioanalytical method validation. Validation results indicate that calibration standards in human plasma can be used to quantify paclitaxel in all tested matrices. In human samples, the validated range for paclitaxel was from 0.25-1000 ng ml(-1) using 200 microl plasma aliquots and from 5 to 5000 ng g(-1) using 50 microl tumor homogenate aliquots (0.2 g tissue ml(-1) control human plasma). In mice, the ranges were 1-1000 ng ml(-1) and 5-5000 ng g(-1) using 50 microl of mouse plasma and 50 microl of tumor homogenate aliquots (0.2 g tissue ml(-1) control human plasma), respectively. The method can be applied to studies generating only small sample volumes (e.g. mouse plasma and tumor tissue), but also to studies in human plasma requiring a lower limit of quantitation. The assay was applied successfully to several studies with both human and mouse samples.     

Liquid chromatography tandem mass spectrometry method for determination of bisoprolol in human plasma using d5‐bisoprolol as the internal standard

A simple, reliable and sensitive liquid chromatography tandem mass spectrometry (LC‐MS/MS) protocol was developed and validated for quantification of bisoprolol in human plasma. The sample was pretreated with a simple procedure of protein precipitation and an isotope‐labeled d5‐bisoprolol was used as internal standard. The chromatographic separation was performed on a Capcell Pak C₁₈ MG III column (100 mm × 2.0 mm, 5 µm). The protonated ion of the analyte was detected in positive ionization by multiple reaction monitoring mode. The mass transition pairs of m/z 326.3 → 116.3 and m/z 331.3 → 121.3 were used to detect bisoprolol and the internal standard, respectively. Linearity, accuracy, precision, recovery, matrix effect, dilution test and stability were evaluated during method validation over the range of 0.5–100 ng/mL. The validated method was successfully applied to analyze human plasma samples in a bisoprolol bioavailability study. Copyright © 2009 John Wiley & Sons, Ltd.     

A liquid chromatography–tandem mass spectrometry method for preclinical pharmacokinetics and tissue distribution of hydrolyzable tannins chebulinic acid and chebulagic acid in rats

A simple and sensitive liquid chromatography–tandem mass spectrometry method was developed for the simultaneous determination of chebulinic acid and chebulagic acid in rat plasma and tissues and well used in the pharmacokinetic and tissue distribution studies after intraperitoneal injection administration. Samples were processed with methanol by protein precipitation, and chromatographic separation was performed on an Agilent Zorbax SB‐C₁₈ column (50 × 2.1 mm, 1.8 μm) with a mobile phase consisting of methanol and water containing 0.1% formic acid (60:40, v/v). Quantification was performed by selected reaction monitoring with m/z 977.1 → 806.8 for chebulagic acid, m/z 979.0 → 808.7 for chebulinic acid and m/z 851.2 → 704.9 for the internal standard. Good linearity was observed over their respective concentration range. The pharmacokinetic study showed that both compounds reached their peak concentration values (605.8 ± 35.6 ng/mL for chebulinic acid and 1327.1 ± 118.6 ng/mL for chebulagic acid) at the same time of 0.9 h following intraperitoneal injection administration. The two compounds could be detected in blood‐abundant tissues. The kidney had the highest concentrations (462.6 ± 138.5 ng/g for chebulinic acid and 1651.7 ± 167.7 ng/g for chebulagic acid) at 1 h post‐dose, followed by the heart, liver, spleen and lung.     

Simultaneous quantification of oleuropein and its metabolites in rat plasma by liquid chromatography electrospray ionization tandem mass spectrometry

Oleuropein (OE) is the cardinal bioactive compound derived from Olea europaea and possesses numerous beneficial properties for human health. However, despite the plethora of analytical methods that have studied the biological fate of olive oil‐derived bioactive compounds, no validated methodology has been published to date for the simultaneous determination of OE, along with all its major metabolites. In this study, a liquid chromatography‐electrospray ionization‐tandem mass spectrometry (LC‐ESI MS/MS) method has been developed and validated for the quantification of OE, simultaneously with its main metabolites hydroxytyrosol, 2‐(3,4‐dihydroxyphenyl)acetic acid, 4‐(2‐hydroxyethyl)‐2‐methoxy‐phenol or homovanillyl alcohol, 2‐(4‐hydroxy‐3‐methoxyphenyl)acetic acid or homovanillic acid, and elenolic acid in rat plasma matrix. Samples were analyzed by LC‐ESI MS/MS prior to and after enzymatic treatment. A solid‐phase extraction step with high mean recovery for all compounds was performed as sample pretreatment. Calibration curves were linear for all bioactive compounds over the range studied, while the method exhibited good accuracy, intra‐ and inter‐day precision. The limit of detection was in the picogram range (per milliliterof plasma) for HT and OE and in the nanogram range (per milliliter of plasma) for the other analytes, and the method was simple and rapid. The developed methodology was successfully applied for the simultaneous quantification of OE and its aforementioned metabolites in rat plasma samples, thus demonstrating its suitability for pharmacokinetics, as well as bioavailability and metabolism studies. Copyright © 2009 John Wiley & Sons, Ltd.     

Sensitive liquid chromatography tandem mass spectrometry method for the quantification of Quetiapine in plasma

A sensitive high-performance liquid chromatography-tandem mass spectrometry method was developed and validated for the quantification of quetiapine in rat plasma. Following liquid-liquid extraction, the analyte was separated using a gradient mobile phase on a reverse-phase column and analyzed by MS/MS in the multiple reaction monitoring mode using the respective [M + H]+ ions, m/z 384 to m/z 221 for quetiapine and m/z 327 to m/z 270 for the internal standard. The assay exhibited a linear dynamic range of 0.25-500 ng/mL for quetiapine in rat plasma. The lower limit of quantification was 0.25 ng/mL with a relative standard deviation of less than 7%. Acceptable precision and accuracy were obtained for concentrations over the standard curve range. The validated method was successfully used to analyze rat plasma samples for application in pre-clinical pharmacokinetic studies. This method in rodent plasma could be adapted for quetiapine assay in human plasma.     

Quantitative determination of sparfloxacin in rat plasma by liquid chromatography/tandem mass spectrometry

Sparfloxacin, a fluoroquinolone antibiotic, is used for the treatment of bacterial infection. A quantification method using mass spectrometry was developed for the determination of sparfloxacin in rat plasma. After simple protein precipitation with acetonitrile, the analytes were chromatographed on a reversed‐phase C₁₈ column and detected by liquid chromatography/tandem mass spectrometry with electrospray ionization. The accuracy and precision of the assay were in accordance with FDA regulations for validation of bioanalytical methods. This method was applied to measure the plasma sparfloxacin concentrations after a single oral administration of sparfloxacin in rats. Copyright © 2010 John Wiley & Sons, Ltd.     

Simultaneous quantification of atenolol and chlorthalidone in human plasma by ultra‐performance liquid chromatography–tandem mass spectrometry

A simple, sensitive and reproducible ultra‐performance liquid chromatography–tandem mass spectrometry method has been developed for the simultaneous determination of atenolol, a β‐adrenergic receptor‐blocker and chlorthalidone, a monosulfonamyl diuretic in human plasma, using atenolol‐d7 and chlorthalidone‐d4 as the internal standards (ISs). Following solid‐phase extraction on Phenomenex Strata‐X cartridges using 100 μL human plasma sample, the analytes and ISs were separated on an Acquity UPLC BEH C₁₈ (50 mm × 2.1 mm, 1.7 µm) column using a mobile phase consisting of 0.1% formic acid–acetonitrile (25:75, v/v). A tandem mass spectrometer equipped with electrospray ionization was used as a detector in the positive ionization mode for both analytes. The linear concentration range was established as 0.50–500 ng/mL for atenolol and 0.25–150 ng/mL for chlorthalidone. Extraction recoveries were within 95–103% and ion suppression/enhancement, expressed as IS‐normalized matrix factors, ranged from 0.95 to 1.06 for both the analytes. Intra‐batch and inter‐batch precision (CV) and accuracy values were 2.37–5.91 and 96.1–103.2%, respectively. Stability of analytes in plasma was evaluated under different conditions, such as bench‐top, freeze–thaw, dry and wet extract and long‐term. The developed method was superior to the existing methods for the simultaneous determination of atenolol and chlorthalidone in human plasma with respect to the sensitivity, chromatographic analysis time and plasma volume for processing. Further, it was successfully applied to support a bioequivalence study of 50 mg atenolol + 12.5 mg chlorthalidone in 28 healthy Indian subjects. Copyright © 2015 John Wiley & Sons, Ltd.     

Liquid chromatography tandem mass spectrometry method for the quantification of clonidine with LLOQ of 10 pg/mL in human plasma

A sensitive high-performance liquid chromatography-positive ion electrospray tandem mass spectrometry method was developed and validated for the quantification of clonidine in human plasma. Following liquid-liquid extraction, the analytes were separated using an isocratic mobile phase on a reverse-phase column and analyzed by MS/MS in the multiple reaction monitoring mode using the respective [M + H](+) ions, m/z 230 to 44 for clonidine and m/z 254 to 44 for the internal standard. The assay exhibited a linear dynamic range of 10-2000 pg/mL for clonidine in human plasma. The lower limit of quantification was 10 pg/mL with a relative standard deviation of less than 6.8%. Acceptable precision and accuracy were obtained for concentrations over the standard curve range. A run time of 2.5 min for each sample made it possible to analyze more than 250 human plasma samples per day. The validated method was successfully used to analyze human plasma samples for application in pharmacokinetic studies. Copyright (c) 2008 John Wiley & Sons, Ltd.     

Quantitative determination of domperidone in human plasma by ultraperformance liquid chromatography with electrospray ionization tandem mass spectrometry

A simple and sensitive liquid chromatography-tandem mass spectrometry method was developed and validated for determining domperidone in human plasma. The analyte and internal standard (IS; mosapride) were isolated from plasma samples by protein precipitation with methanol (containing 0.1% formic acid). The chromatographic separation was performed on an Xterra MS C(18) Column (2.1 x 150 mm, 5.0 microm) with a gradient programme mobile phase consisting of 0.1% formic acid and acetonitrile at a flow rate of 0.30 mL/min. The total run time was 4.0 min. The analyses were carried out by multiple reaction monitoring using the parent-to-daughter combinations m/z 426 --> 175 and m/z 422 --> 198 (IS). The areas of peaks from the analyte and IS were used for quantification of domperidone. The method was validated according to the FDA guidelines on bioanalytical method validation. Validation results indicated that the lower limit of quantification was 0.2 ng/mL, and the assay exhibited a linear range of 0.2-60.0 ng/mL and gave a correlation coefficient (r(2)) of 0.999 or better. Quality control samples (0.4, 0.8, 15 and 50 ng/mL) in six replicates from three different analytical runs demonstrated an intra-assay precision (RSD) 4.43-6.26%, an inter-assay precision 5.25-7.45% and an overall accuracy (relative error) of <6.92%. The method can be applied to pharmacokinetic and bioequivalence studies of domperidone.