Quantitative Determination of Lobeline Hydrochloride in Rabbit Plasma by LC–MS–MS and Its Application

A sensitive and selective liquid chromatography tandem mass spectrometry method for quantitative determination of lobeline hydrochloride in rabbit plasma was developed and validated. After addition of triazolam as internal standard, protein precipitation by acetonitrile was used as sample preparation. Chromatographic separation was achieved on a Zorbax SB-C18 column with acetonitrile-0.1% formic acid as mobile phase with gradient elution. Electrospray ionization source was applied and operated in positive ion mode; multiple reaction monitoring mode was used for quantification using target fragment ions m/z 338.1 → 315.8 for lobeline hydrochloride and m/z 342.9 → 308.0 for the IS. Calibration plots were linear over the range of 2–500 ng mL^?1 for lobeline hydrochloride in plasma. Lower limit of quantitation for lobeline hydrochloride was 2 ng mL^?1. Mean recovery of lobeline hydrochloride from plasma was in the range 97.5–102.3%. RSD of intra-day and inter-day precision were both <9%. This developed method is successfully used in pharmacokinetic study of lobeline hydrochloride in rabbit.     

Simultaneous Determination of Pethidine and Atropine in Rabbit Plasma by LC�CMS�CMS and Its Application to a Pharmacokinetic Study after Intramuscular Administration

A sensitive and selective liquid chromatography?Ctandem mass spectrometry method for the determination of pethidine and atropine in rabbit plasma was developed and validated. The analytes and internal standard (IS) are extracted from plasma by liquid?Cliquid extraction using ethyl acetate, and separated on a Zorbax SB-Aq column (2.1 × 150 mm, 3.5 ??m) using acetonitrile?C0.1% formic acid as mobile phase with gradient elution. Electrospray ionization source was applied and operated in positive ion mode, and multiple reaction monitoring mode was used for quantification using target fragment ions m/z 247.8 ?? 219.7 for pethidine, m/z 289.9 ?? 123.8 for atropine and m/z 295.0 ?? 266.8 for IS, respectively. The assay is linear over the range of 5?C1,000 ng mL^?1 for pethidine and atropine, with a lower limit of quantification of 3 ng mL^?1 for pethidine and 5 ng mL^?1 for atropine. Intra-day and inter-day precision are less than 11% and the accuracy are in the range of 90.4?C106.3%. Furthermore, the newly developed method is successfully used for the determination of pethidine and atropine in rabbit plasma for pharmacokinetic study.     

Determination of Asiaticoside in Rat Plasma by LC–MS–MS and Its Application to Pharmacokinetics Studies

A rapid and sensitive LC–MS–MS method was developed and validated for the determination of asiaticoside in rat plasma. Asiaticoside was extracted by protein precipitation with acetonitrile, and separated on a C₁₈ column. The total analytical time was relatively short (4 min), and the limit of quantification was 38 ng mL^?1 using 100 μL of rat plasma. Asiaticoside and the internal standard (felodipine) were monitored in the multi-reaction-monitoring mode as follows: m/z 957.4 → 469.3 and m/z 382.2 → 145.1, respectively. Calibration was linear over a concentration range from 38 to 7,600 ng mL^?1, and the correlation coefficient was greater than 0.998. The recoveries of asiaticoside from plasma were better than 85%, and RSDs of inter-day and intra-day assays were below 10.1%. The method is sensitive and specific, and suitable for pharmacokinetic studies of asiaticoside in rats.     

LC–MS–MS Determination of Nikethamide in Human Plasma

A sensitive LC–MS–MS method with electrospray ionization has been developed for determination of nikethamide in human plasma. After addition of atropine as internal standard, liquid–liquid extraction was used to produce a protein-free extract. Chromatographic separation was achieved on a 150 mm × 2.1 mm, 5 μm particle, Agilent Zorbax SB-C₁₈ column, with 45:55 (v/v) methanol–water containing 0.1% formic acid as mobile phase. LC–MS–MS was performed in multiple reaction monitoring mode using target fragment ions m/z 178.8 → 107.8 for nikethamide and m/z 289.9 → 123.8 for the internal standard. Calibration plots were linear over the range of 20.0–2,000 ng mL^?1. The lower limit of quantification was 20.0 ng mL^?1. Intra-day and inter-day precisions were better than 4.2 and 6.1%, respectively. Mean recovery of nikethamide from human plasma was in the range 65.3–71.1%.     

LC–MS Method for the Quantification of Clonazepam in Rat Plasma

A sensitive and specific high-performance liquid chromatography–tandem mass spectrometry method has been developed and validated for the determination of clonazepam in rat plasma. Clonazepam and internal standard diazepam were extracted from plasma samples by a single-step protein precipitation. The chromatographic separation was performed on a Dikma ODS-C18 reversed-phase column at 40 °C. The mobile phase composed of a premix of solvent A (0.1% formic acid–4 mM ammonium acetate–water)–solvent B (acetonitrile) (13:87, v/v) at a flow-rate of 0.7 mL min^?1. Positive electrospray ionization was utilized as the ionization source. Clonazepam and the internal standard were determined using multiple reaction monitoring of precursor → product ion transitions at m/z 316.0 → 270.0 and m/z 285.1 → 193.2, respectively. The lower limit of quantification was 0.25 ng mL^?1 using 50 μL plasma samples and the linear calibration range was from 0.25 to 128 ng mL^?1. The within- and between-batch RSDs were lower than 15% and the relative recoveries of clonazepam ranged from 97.4 to 104.7%. The mean extraction recoveries of clonazepam and IS were 79.7 and 77.6%, respectively. The method has been successfully applied to the pharmacokinetic studies in rat after oral administration of clonazepam.     

Determination of Poricoic Acid A in Rat Plasma after Oral and Intravenous Administration by LC–MS–MS and Its Application to a Pharmacokinetic Study

A sensitive and specific liquid chromatography–tandem mass spectrometry (LC–MS–MS) method was developed and validated for the quantification of poricoic acid A (PAA) in rat plasma. The plasma samples were precipitated by protein precipitation with methanol. Glycyrrhetic acid was used as the IS. Chromatography was performed on a Dionex C₁₈ 120 Å (4.6 × 250 mm, 5 μm) column with the mobile phase composed of acetonitrile–water (90:10, v/v) at a flow rate of 0.8 mL min^?1. A tandem mass spectrometer equipped with an ESI source was used as the detector and was operated in the negative ion mode. Quantification was performed using multiple reaction monitoring (MRM) of the transitions m/z 497.4 → 423.3 and m/z 469.2 → 425.1 for PAA and IS, respectively. The calibration curves were linear over the range of 5–5,000 ng mL^?1 (r ² = 0.9966) and the lower limit of quantification (LLOQ) was 5 ng mL^?1. In this range, RSDs were <10% for intra-assay and inter-assay precisions. The accuracy expressed by deviation (DEV) was <6%, and the extraction recoveries of QC samples were >78%. The validated method was successfully used to study the pharmacokinetics of PAA in rats after intravenous administration at a dose of 1.0, 2.5 and 5.0 mg kg^?1 and oral administration at a dose of 25, 50 and 100 mg kg^?1, respectively. The relative bioavailability of PAA in rats following oral administration was achieved.     

Simultaneous Determination of Tamsulosin and Dutasteride in Human Plasma by LC–MS–MS

A sensitive and selective liquid chromatographic tandem mass spectrometric (LC–MS–MS) method was developed for simultaneous identification and quantification of tamsulosin and dutasteride in human plasma, which was well applied to clinical study. The method was based on liquid–liquid extraction, followed by an LC procedure with a Gemini C-18, 50 mm × 2.0 mm (3 μm) column and using methanol:ammonium formate (97:3, v/v) as the mobile phase. Protonated ions formed by a turbo ionspray in positive mode were used to detect analytes and internal standard. MS–MS detection was by monitoring the fragmentation of 409.1 → 228.1 (m/z) for tamsulosin, 529.3 → 461.3 (m/z) for dutasteride and 373.2 → 305.3 (m/z) for finasteride (IS) on a triple quadrupole mass spectrometer. The lower limit of quantification for both tamsulosin and dutasteride was 1 ng mL^?1. The proposed method enables the unambiguous identification and quantification of tamsulosin and dutasteride for clinical drug monitoring.     

Sensitive and Selective LC-MS-MS Assay for the Quantification of Palonosetron in Human Plasma and Its Application to a Pharmacokinetic Study

A highly sensitive and selective liquid chromatography-tandem mass spectrometry method was developed for the determination of palonosetron in human plasma samples. Chromatographic conditions and mass spectral parameters were optimized in order to achieve a limit of quantification of approximately 0.03 ng mL^?1. Palonosetron and citalopram (internal standard) were extracted by liquid–liquid extraction under alkaline conditions using saturated sodium bicarbonate. Separation was achieved with a Hanbon Lichrospher C₁₈ column and detection was carried out by tandem mass spectrometry using positive electrospray ionization in selected reaction monitoring mode. The target ions of palonosetron and citalopram were to m/z 297.00 → 297.00 and 325.00 → 325.00 respectively. Calibration curves were linear over the range of approximately 0.03–10 ng mL^?1. Precision and accuracy of this method was acceptable. The method was successfully applied to a pharmacokinetic study with healthy Chinese volunteers after intravenous administration of a single dose of 0.125, 0.25 or 0.5 mg palonosetron hydrochloride.     

Quantitative and Qualitative Analysis of CKD-501, Lobeglitazone,in Human Plasma and Urine Using LC–MS/MS and Its Application to a Pharmacokinetic Study

A simple, rapid and sensitive LC–MS/MS method in positive ion mode was developed and validated to determine CKD-501, lobeglitazone, in human plasma and urine using glipizide as an internal standard (IS). Lobeglitazone is a novel thiazolidinedione (TZDs)-based peroxisome proliferator-activated receptor (PPAR) agonist, used for the management of type-2 diabetes. After mixing the IS, dissolved in acetonitrile, with a plasma or urine sample containing lobeglitazone, 10 μL of supernatant was injected into the LC–MS/MS system. Quantification was performed in the multiple reaction monitoring (MRM) mode using transition of 481.5 → 152.2 (m/z) for lobeglitazone and 446.1 → 321.2 (m/z) for the IS. The method showed good linearity over concentration ranges of 0.5–1,000 ng mL⁻¹ for plasma and 0.2–250 ng mL⁻¹ for urine (r ² ≥ 0.9996). The mean percent extraction recovery of lobeglitazone was 90.8 % for plasma and 87.3 % for urine, while the recoveries of the IS were greater than 86.4 % for both. The intra-day and inter-day precision of plasma ranged from 1.1 to 3.7 and 2.5 to 3.3 % (RSD), respectively, and the intra- and inter-day precision of urine ranged from 1.5 to 2.7 and 3.2 to 3.5 %, respectively. This method is simple, sensitive, and applicable for the pharmacokinetic study of lobeglitazone in human plasma. Most of the urine concentrations of lobeglitazone were below the LLOQ because the lobeglitazone is extensively metabolized.

     

LC–MS–MS Analysis of Mirtazapine in Plasma, and Determination of Pharmacokinetic Data for Rats

A sensitive LC–MS–MS method with electrospray ionization has been developed for analysis of mirtazapine in rat plasma. After addition of diazepam as internal standard, liquid–liquid extraction was used to produce a protein-free extract. Chromatographic separation was achieved on a 150 × 4.6 mm, 5 μm particle, ODS column with 84:16 (v/v) methanol–water containing 0.1% ammonium acetate and 0.01% glacial acetic acid as mobile phase. LC–MS–MS was performed in selected-ion-monitoring (SIM) mode using target fragment ions m/z 195.09 for mirtazapine and m/z 192.80 for the IS. Calibration plots were linear over the range of 0.516–618.8 ng mL^?1. The lower limit of quantification was 0.516 ng mL^?1. Intra-day and inter-day precision were better than 12.6 and 8.8%, respectively. Mean recovery of mirtazapine from plasma was in the range 87.41–90.06%; average recovery was 88.40% (RSD 3.95%). Significant gender differences between mirtazapine pharmacokinetic data were observed in this study.     

Determination of Memantine in Human Plasma by LC–MS–MS: Application to a Pharmacokinetic Study

A sensitive and selective liquid chromatography–tandem mass spectrometry method for the determination of memantine was developed and validated over the linearity range 0.1–25 ng mL^?1 with 0.5 mL of plasma using procainamide as the internal standard. This analysis was carried out on a Cosmosil 5C₁₈-MS column and the mobile phase was composed of methanol: 0.5% formic acid (50:50, v/v). Detection was performed on a triple–quadrupole tandem mass spectrometer using positive ion mode electrospray ionization and quantification was performed by multiple reaction monitoring mode. The MS–MS ion transitions monitored were m/z 180 → 107 and 236 → 163 for memantine and procainamide, respectively. The between- and within-day precision was less than 10.9% and accuracy was less than 2.5%. The lower limit of quantification (LLOQ) was 0.1 ng mL^?1. The method proved to be accurate and specific, and was applied to the pharmacokinetic study of memantine in healthy Chinese volunteers.     

Determination of Dothiepin in Human Plasma by LC–ESI–MS and its Application to Bioequivalence Studies

An LC–MS method for the determination of dothiepin in human plasma was developed and validated. Sample preparation involved extraction with n-hexane:2-propanol (95:5). Separation was on an Ultimate XB C18 column (2.1 × 150 mm, 5 μm). A single-quadrupole mass spectrometer with an electrospray interface was operated in the selected-ion monitoring mode to detect the [M+H]⁺ ions at m/z 296 for dothiepin and at m/z 278 for the internal standard (amitriptylene). The method demonstrated good linearity from 0.78 ng mL^?1 (the LOQ) to100 ng mL^?1. The mean extraction recovery was 82.4% for dothiepin and and 84.2% for the internal standard. The intra-day and inter-day precision ranged from 8.5 to 11.4% and 9.7 to 12.1% (RSD), respectively. The method was successfully applied to bioequivalence studies of dothiepin hydrochloride tablets to obtain the pharmacokinetic parameters.     

A Sensitive LC–ESI–MS–MS Method for the Determination of Huperzine A in Human Plasma: Method and Clinical Applications

A sensitive and specific liquid chromatography–electrospray ionization–tandem mass spectrometry method has been developed and validated for the quantification of huperzine A in human plasma. After the addition of trimetazidine, the internal standard (IS) and sodium hydroxide, plasma samples were extracted using 5 mL ethyl acetate. The compounds were separated on an Agilent Zorbax SB C₁₈ column (100 mm × 2.1 mm ID, dp 3.5 μm) using an elution system of 10 mM ammonium acetate solution–methanol–formic acid (18:82:0.1, v/v) as the mobile phase. The quantification of target compounds was obtained by using multiple reaction monitoring (MRM) transitions: m/z 243.1, 210.1 and 267.2, 166.0 were measured in positive mode for huperzine A and IS. Linearity was established for the range of concentrations 0.01–4.0 ng mL^?1 with a coefficient of correlation (r) of 0.9991. The lower limit of quantification (LLOQ) was identifiable and reproducible at 0.01 ng mL^?1. The method has been successfully applied to study the pharmacokinetics of huperzine A in healthy male Chinese volunteers.     

LC-DAD-ESI-MS Characterization of Carbohydrates Using a New Labeling Reagent

A sensitive and selective liquid chromatographic tandem mass spectrometric (LC–MS–MS) method was developed for simultaneous identification and quantification of tamsulosin and dutasteride in human plasma, which was well applied to clinical study. The method was based on liquid–liquid extraction, followed by an LC procedure with a Gemini C-18, 50 mm × 2.0 mm (3 μm) column and using methanol:ammonium formate (97:3, v/v) as the mobile phase. Protonated ions formed by a turbo ionspray in positive mode were used to detect analytes and internal standard. MS–MS detection was by monitoring the fragmentation of 409.1 → 228.1 (m/z) for tamsulosin, 529.3 → 461.3 (m/z) for dutasteride and 373.2 → 305.3 (m/z) for finasteride (IS) on a triple quadrupole mass spectrometer. The lower limit of quantification for both tamsulosin and dutasteride was 1 ng mL⁻¹. The proposed method enables the unambiguous identification and quantification of tamsulosin and dutasteride for clinical drug monitoring.

     

LC–MS Method for the Quantification of Clonazepam in Rat Plasma

A sensitive and specific high-performance liquid chromatography–tandem mass spectrometry method has been developed and validated for the determination of clonazepam in rat plasma. Clonazepam and internal standard diazepam were extracted from plasma samples by a single-step protein precipitation. The chromatographic separation was performed on a Dikma ODS-C18 reversed-phase column at 40 °C. The mobile phase composed of a premix of solvent A (0.1% formic acid–4 mM ammonium acetate–water)–solvent B (acetonitrile) (13:87, v/v) at a flow-rate of 0.7 mL min⁻¹. Positive electrospray ionization was utilized as the ionization source. Clonazepam and the internal standard were determined using multiple reaction monitoring of precursor → product ion transitions at m/z 316.0 → 270.0 and m/z 285.1 → 193.2, respectively. The lower limit of quantification was 0.25 ng mL⁻¹ using 50 μL plasma samples and the linear calibration range was from 0.25 to 128 ng mL⁻¹. The within- and between-batch RSDs were lower than 15% and the relative recoveries of clonazepam ranged from 97.4 to 104.7%. The mean extraction recoveries of clonazepam and IS were 79.7 and 77.6%, respectively. The method has been successfully applied to the pharmacokinetic studies in rat after oral administration of clonazepam.

     

Validation of an LC–MS/MS Method for Simultaneous Detection of Five Selective KCNQ Channel Openers: ICA-27243, ML-213, PF-05020182, SF-0034 and Flupirtine in Mice Plasma and its Application to a Pharmacokinetic Study in Mice

A sensitive and rapid LC–MS/MS method was developed and validated for the simultaneous quantitation of five selective KCNQ channel openers, namely ICA-27243, ML-213, PF-05020182, SF-0034 and flupirtine in mice plasma as per regulatory guideline. The analytes and the internal standard (IS; flupirtine-d ₄ ) were extracted from 50 µL mice plasma by liquid–liquid extraction, followed by chromatographic separation using an Atlantis C₁₈ column with an isocratic mobile phase comprising 0.2% formic acid: acetonitrile (20:80, v/v) at a flow rate of 0.6 mL min^?1 within 2.5 min. Detection and quantitation was done by multiple reaction monitoring on a triple quadrupole mass spectrometer following the transitions: m/z 268.9 → 140.8, 258.1 → 95.1, 367.2 → 269.1, 322.2 → 248.2, 305.7 → 196.4 and 309.1 → 196.1 for ICA-27243, ML-213, PF-05020182, SF-0034, flupirtine and the IS, respectively, in the positive ionization mode. The calibration curves were linear from 1.00 to 2008 ng mL^?1 for all the analytes with r² ≥ 0.99. The intra- and inter-batch accuracy and precision (% CV) across quality controls varied from 90.0 to 113 and 2.64 to 13.0; 93.8 to 114 and 3.15 to 14.9%, respectively, for all the analytes. Analytes were found to be stable under different stability conditions. The method was applied to a pharmacokinetic study in mice.     

LC–MS Determination and Pharmacokinetic Study of Salidroside in Rat Plasma after Oral Administration of Traditional Chinese Medicinal Preparation Rhodiola crenulata Extract

A simple, rapid and sensitive liquid chromatography–mass spectrometry (LC–MS) method was developed for the quantification of salidroside in rat plasma and the study of its pharmacokinetics after oral administration of 15 g kg^?1 Rhodiola crenulata extract to Wistar rats. A 200 μL plasma sample was extracted by acetonitrile and performed on Kromasil C₁₈ column (150 mm × 4.6 mm, 5 μm) with the mobile phase of acetonitrile–water (11:89) within a run time of 8 min. The analyte was monitored with electrospray ionization (ESI) by selected ion monitoring (SIM) mode. The target ions were m/z 299.20 for salidroside and m/z 150.00 for internal standard (IS) paracetamol. A good linear relationship was obtained over the range of 100–20,000 ng mL^?1 and the lower limit of quantification was 100 ng mL^?1. The validated method was successfully applied for the pharmacokinetic study of salidroside in rat. After oral administration of Rhodiola crenulata extract, the main pharmacokinetic parameters T _max, T _1/2, C _max, AUC _0?t and AUC _0?∞ were 0.56 ± 0.21 h, 7.91 ± 4.42 h, 3,386 ± 2,138 ng mL^?1, 16,146 ± 6,558 ng h mL^?1 and 18,599 ± 6,529 ng h mL^?1, respectively.     

LC–MS Simultaneous Determination of Itopride Hydrochloride and Domperidone in Human Plasma

A rapid, simple, sensitive and specific liquid chromatography–tandem mass spectrometry method was developed and validated for simultaneous quantification of itopride hydrochloride and domperidone in human plasma. Both drugs were extracted by liquid–liquid extraction with ethyl acetate and saturated borax solution. The chromatographic separation was performed on a reversed-phase C18 column with a mobile phase of water–methanol (2:98, v/v) containing 0.5% formic acid. The protonated analyte was quantitated in positive ionization by multiple reaction monitoring with a mass spectrometer. The assay exhibited linearity over the concentration range of 3.33–500 ng mL^?1 for itopride hydrochloride and 3.33–100 ng mL^?1 for domperidone in human plasma. The precursor to product ion transitions of m/z 359.1–72.3 and 426.0–147.2 were used to measure itopride hydrochloride and domperidone respectively. The method was found suitable for the analysis of plasma samples collected during phase 1 pharmacokinetics study of itopride HCl 50 mg and domperidone 20 mg in 12 healthy volunteers after single oral doses of the combination drug.     

LC–MS/MS Method for Detection of a Highly Selective KCa3.1 Blocker,TRAM-34, in Rat Plasma

1-[(2-Chlorophenyl)diphenylmethyl]pyrazole (TRAM-34) is a highly selective K_Ca3.1 channel blocker. TRAM-34 was commonly used to study the role of K_Ca3.1 in the pathogenesis of disease in vivo, but there was no validated analytical method. Here, we describe the first validated LC–MS/MS analytical method for TRAM-34. Solid-phase extraction (SPE) was performed to extract TRAM-34 from the rat plasma. Chromatographic separation was achieved on the phenyl column. A triple quadrupole mass spectrometer was operated in positive-mode electrospray ionization. There were two multiple-reaction monitoring (MRM) transitions for TRAM-34: m/z 277.2 → 165.1 (for quantification) and m/z 277.2 → 241.2 (for qualification). Bifonazole was used as an internal standard. The lower limit of quantification (LLOQ) achieved was 1 ng mL^?1 and the run time was 7.5 min. The linear range was from 1 to 1,000 ng mL^?1. The pharmacokinetics profile was acquired for rats following an intraperitoneal injection of TRAM-34, with the following pharmacokinetics parameters found: C _max 17.03 ± 1.34 ng mL^?1; T _max 8.67 ± 3.06 h; and T _1/2 13.45 ± 2.72 h. In addition, a suspected metabolite of TRAM-34 was found using this LC–MS/MS method. Given the results of the detailed validation process and its application to TRAM-34 pharmacokinetics, it is clear that a fast, selective, precise, and reproducible TRAM-34 LC–MS/MS analytical method was successfully established.     

Determination of Dydrogesterone in Human Plasma by Tandem Mass Spectrometry: Application to Therapeutic Drug Monitoring of Dydrogesterone in Gynecological Disorders

A sensitive and specific tandem mass spectrometric (MS–MS) method was developed and validated for the determination of dydrogesterone (Duphaston^®), an orally active synthetic progestogen, in human plasma. Multiple reaction monitoring (MRM) scans at m/z 313.1 > 105.5 (dydrogesterone) and m/z 393 > 147 (dexamethasone, internal standard) were selected to determine dydrogesterone by the internal standard method. Linear correlations (r: ～0.99 ± 0.05) of the calibration curves were established over the concentration range 10–60 ng mL^?1 with a lower limit of quantification (LLQ) of 10 ng mL^?1 (RSD% 14.9 and %DEVs ?10.5 to +15.6). Solid-phase extraction (SPE) technique was used for extraction of dydrogesterone and internal standard from patient plasma samples using Oasis^® Max C18 cartridges. Ion suppression studies indicated negligible effects of plasma matrix on the mass ions detection of dydrogesterone and IS, when measured in MRM mode. Validation data showed that RSD% values were <22.0%, whereas %DEV values were in the range of ?20.2 to +13.3 for intra- and inter-day precision and accuracy, respectively. Analytical recoveries of dydrogesterone from supplemented plasma samples with the drug were in the range of 100.7–112%, indicating the efficiency of the SPE for separation of dydrogesterone from human plasma. Stability studies conducted at ?20 °C, showed that dydrogesterone was stable in plasma as indicated from the measured degradation kinetic parameters. The developed method was applied for monitoring plasma levels of dydrogesterone in 25 patients treated with Duphaston^® tablets at a dose of 10 mg three times daily. Mean plasma concentration of 16.1 ± 3.5 ng mL^?1 of dydrogesterone were measured at the steady state. The data suggest the utility of tandem mass method in therapeutic drug monitoring of plasma levels of dydrogesterone in gynecological disorders treated with Duphaston^® tablets.