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Synthesis of an innovative drug 9-(2,5-dihydroxyphenyl)-2-(4-ethoxyphenyl)-2,3,7,8-tetrahydro-1H-pyrido[1,2-a]pyrazine-1,4(6H)-dione (PPI) for the treatment of peptic ulcer with a new mechanism of action has necessitated the pharmacokinetics study of the substance in the blood plasma and investigation of its absorption and elimination processes. This paper describes the development and validation of a technique for the quantitative determination of new compound PPI in rat blood plasma by HPLC/MS. The method of solid-phase extraction is applied to extract PPI from blood plasma in the process of biological sample preparation. The lower limit of quantification is 0.1 ng/mL with a linearity range from 0.1 to 1000 ng/mL. By the developed technique, the sample analysis of rat blood plasma was carried out after the introduction of PPI pharmaceutical substance in a dose of 20 mg/kg, a pharmacokinetic profile of the substance was established, and pharmacokinetic parameters were calculated in order to judge the degrees and rates of absorption and excretion of the new antiulcer drug in the blood of laboratory rats. [ABSTRACT FROM AUTHOR]

Abstract Methylisothiazolinone (MI) is a preservative used in consumer products to control bacterial and fungal growth. MI can be toxic, act as a skin sensitizer and irritant, and initiate lung diseases; therefore, it is important to understand the mechanisms underlying the toxicity of MI in the body. In this study, we developed a method to analyze plasma MI by using an LC-MS/MS-coupled multiple reaction monitoring (MRM) technique, which follows the fragments of a target metabolite in rat plasma. The MRM transition of MI was m / z 116 ➔ 101, and the lower limit of quantification (LLOQ) was set at 10 ng/mL. Including the concentration of LLOQ, a seven-point calibration curve explained much of the variation in the response, and it was strongly linear (R2 = 0.9998); its intra- and inter-day accuracy and precision values were within 15% of the standard deviation (SD%) and along with the FDA and Korea Ministry of Food and Drug Safety guidelines. For intravenous (iv) pharmacokinetic studies of MI using rats, we developed an analytical method that was useful in detecting the profile of MI in the plasma. We also determined half-life, and area under the curve (AUC) of MI using a non-compartment model, and these might be useful for the study of toxic mechanisms of MI in the body. [ABSTRACT FROM AUTHOR]

Objective: The purpose of this study was to develop a LC-MS method to determine amiodarone (AMI) and its major metabolite desethylamiodarone (DEA) from rat plasma released from the adipose tissue of AMI treated rats subjected to a weight gain/weight loss cycle. Methods: Separation of the compounds was performed on a Kinetex 2.6 µm C18 100 x 4.6 mm column under isocratic conditions using a mixture of acetonitrile: 0.1% formic acid 65:35 at a flow rate of 0.5 ml/min. Detection of the analyte was performed by electrospray positive ionization, the monitored ions being 135 m/z from 646 for AMI and 135 m/z of 618 for DEA. Analytes were extracted after plasma protein precipitation with methanol. Results: The developed method presented specificity and linearity on the concentration range of 25-2500 ng/ml plasma for AMI and 2.5-1250 ng/ml plasma for DEA and the precision and accuracy of the method at all of quality control concentration levels including LLOQ were according to official guidelines for validating analytical methods. Conclusions: A sensitive and accurate LC-MS method has been developed with a much lower LLOQ than literature data to detect the plasma concentration differences of the studied analytes that result from forced lipolysis and mobilization from the adipose tissue. [ABSTRACT FROM AUTHOR]

A sensitive, selective, and reliable LC-MS/MS method was developed and validated for simultaneous quantification of venlafaxine (VEN) and its 5 metabolites (ODV, NDV, NNDDV, OHV and NODDV) in rat plasma. The calibration ranges are 15.0 to 6000 ng/mL for VEN, 1.00 to 400 ng/mL for ODV, 5.00 to 2000 ng/mL for NDV, 1.00 to 400 ng/mL for NNDDV, 10.0 to 4000 ng/mL for OHV, and 0.200 to 20.0 ng/mL for NODDV. Briefly, 50 μL of rat plasma was extracted using liquid-liquid extraction (LLE) with methyl tert -butyl ether (MTBE). The analytes were separated on an Agilent SB-Phenyl (50 mm × 4.6 mm, 3.5 μm) column using a binary gradient of 0.1% formic acid in water versus 0.1% formic acid in acetonitrile at a flow rate of 0.8 mL/min. The method was validated following FDA guidance for bioanalytical method validation. Validated method was successfully applied to a pharmacokinetic study of VEN orally administered to rats. [ABSTRACT FROM AUTHOR]

LNA-i-miR-221, a 13-mer oligonucleotide, is a new miR-221 inhibitor that could be used as a novel drug for multiple myeloma. Herein, an ion-pair reversed phase liquid chromatography–tandem mass spectrometry (LC–MS/MS) method has been developed and validated for the quantification of LNA-i-miR-221 in rat plasma. Plasma samples were prepared with an initial phenol/chloroform/isoamyl alcohol liquid–liquid extraction followed by a solid phase extraction. Chromatographic separation was performed with a gradient system on a HALO C18 column using hexafluoro-2-propanol/triethylamine buffer and methanol as mobile phase at a flow rate of 0.4 mL/min. Under these conditions LNA-i-miR-221 and the analogue internal standard are co-eluted at 1.2 min. The detection was carried out in multiple reaction monitoring (MRM) mode using a negative electrospray ionization (ESI) interface. The assay showed a good linearity within the calibration range 10–10000 ng/mL. The precision, accuracy, and recovery values were found to be <15% (<20% at LLOQ), 100 ± 15%, and 97.6–103.7%, respectively. This method was successfully applied to measure the concentrations of LNA-i-miR-221 in plasma samples following the intravenous administration during a 4-week toxicity study in rats. [ABSTRACT FROM AUTHOR]

We present a validated liquid chromatography with tandem mass spectrometry method for simultaneous determination of 20-( S)-protopanaxatriol and its two oxidative stereoisomeric metabolites (20 S,24 S)-epoxy-dammarane-3,6,12,25-tetraol (M1) and (20 S,24 R)-epoxy-dammarane-3,6,12,25-tetraol (M2) in rat plasma. 20-( S)-Protopanaxatriol, M1, and M2 were extracted with methanol and separated on an ACQUITY HSS T3 column. The mass spectrometry detection was accomplished in selected reaction monitoring mode with precursor-to-product ion transitions of m/z 493.4→143.1 for M1 and M2, m/z 475.4→391.3 for 20-( S)-protopanaxatriol, and m/z 459.4→375.3 for 20-( S)-protopanaxadiol (internal standard). The method showed good linearity over the concentration ranges of 1-1000 ng/mL for 20-( S)-protopanaxatriol and 0.5-200 ng/mL for M1 and M2, with correlation coefficients of more than 0.995. The lower limits of quantification for 20-( S)-protopanaxatriol, M1, and M2 were 1, 0.5, 0.5 ng/mL, respectively. The intra- and interday precisions (RSD, %) were less than 10.41% while the accuracy (relative error, %) ranged from -3.14 to 8.73%. Under the current conditions, M1 and M2 were completely separated within 3 min. The validated assay was successfully applied to evaluating pharmacokinetic profiles of 20-( S)-protopanaxatriol, M1, and M2 in rat. [ABSTRACT FROM AUTHOR]

DA-9805 is a new botanical antiparkinson drug candidate formulated using an ethanolic extract of the root of Bupleurum falcatum , the root cortex of Paeonia suffruticosa , and the root of Angelica dahurica . In this study, a sensitive and rapid LC–MS/MS method was developed to simultaneously determine, saikosaponin a, paeonol, and imperatorin, three active/representative ingredients of DA-9805, in rat plasma. Plasma was extracted by mixture of ethyl acetate and methyl tertiary butyl ether. Chromatographic separation was carried out using a C 18 column and a gradient elution of mobile phases consisting of 5 mM formic acid in water and acetonitrile. Total chromatographic run time was 10.5 min. Multiple reaction monitoring mode was used for mass spectrometry; the transitions were m / z 779.5 → 617.2 for saikosaponin a in negative-ion mode, m / z 167 → 149 for paeonol and m / z 271.1 → 203 for imperatorin in positive-ion mode. Calibration curves were constructed in the range of 0.5–1000 ng/mL for saikosaponin a, 20–10000 ng/mL for paeonol, and 0.2–1000 ng/mL for imperatorin. All the validation data, including the selectivity, linearity, precision, accuracy, recovery, matrix effect, and stability satisfied the acceptance requirements. The method was successfully applied in a pharmacokinetic study of saikosaponin a, paeonol, and imperatorin following oral administration of DA-9805. [ABSTRACT FROM AUTHOR]

Puerarin, a bioactive natural C -glycoside isoflavonoid isolated from Pueraria lobata (Willd.) Ohwi, possesses many potential health benefits. However, the in vivo metabolic fates of puerarin have not been comprehensively clarified yet. In this study, an efficient strategy based on UHPLC-LTQ-Orbitrap mass spectrometer in both positive and negative ion modes was developed to profile and characterize puerarin metabolites in rat urine and plasma. Meanwhile, post-acquisition data-mining methods including high-resolution extracted ion chromatogram (HREIC) and multiple mass defect filtering (MMDF) were utilized to screen potential puerarin metabolites from HR-ESI–MS 1 stage. The mass fragmentation behaviors of five reference standards, including puerarin, daidzin, daidzein, genistin, and genistein, were comprehensively studied for construction of diagnostic product ions (DPIs), which could be employed to implement rapid identification of puerarin metabolites. Finally, a total of 66 metabolites (prototype compound included) were tentatively or positively identified based on standard substances, chromatographic retention times, accurate mass measurement, and corresponding Clog P values. Our results demonstrated that puerarin underwent multiple in vivo metabolic reactions including methylation, hydroxylation, dehydroxylation, hydrogenation, deglycosylation, glycosylation, sulfonation, glucuronidation, and their complicated reactions. In conclusion, the newly discovered puerarin metabolites significantly expanded the understanding on its pharmacological effects and built the foundation for further toxicity and safety studies. [ABSTRACT FROM AUTHOR]

A high-performance liquid chromatography tandem mass spectrometry (HPLC–MS/MS) method was developed and fully validated for simultaneous determination of puerarin, 3′-hydroxy puerarin, 6″- O -xylosyl puerarin, 3′-methoxy puerarin, mirificin, puerarin-7- O -glucoside, daidzin, daidzein, daidzein-7- O -glucoside, ginsenoside-Rd, notoginsenoside-R1, ginsenoside-Re, ginsenoside-Rg1, ginsenoside-Rb1 in rat plasma after intravenous injection of 5 mL/kg Gegen-Sanqi compatibility solution (containing Pueraria flavonid 10.8 mg/mL and Panax notoginsenosidum 5.4 mg/mL). After addition of internal standard (IS) baicalin, the analytes and IS were recovered from rat plasma by protein precipitation using acetonitrile containing 0.1% formic acid. Chromatographic separation was performed on a Capcell pak MG C18 column (3.0 mm × 75 mm, 3.0 μm) at 35 °C with a flow rate of 0.75 mL/min. Mass spectrometry was conducted using multiple reaction monitoring in positive mode. The method was linear for all of the analytes over 1000 times concentration range with correlation coefficients greater than 0.99. The precision and accuracy of the LC–MS/MS assay based on the three analytical quality control (QC) levels were well within the acceptance criteria from FDA guidance for bioanalytical method validation. Method recoveries were higher than 75% and the matrix effects were minimal. All analytes were stable under tested conditions. It was the first time to study the pharmacokinetics of daidzein-7- O -glucoside in rat plasma. To the best of our knowledge, this is the first study for simultaneous determination of so many analytes in rat plasma after intravenous injection of Gegen-Sanqi compatibility solution. It was expected that the present work would provide some helpful references for the apprehension of the mechanism of action and further clinical efficacy studies of Gegen-Sanqi herb-pair. [ABSTRACT FROM AUTHOR]

Brigatinib is the second-generation anaplastic lymphoma kinase – inhibitor in non-small cell lung cancer and it can overcome the crizotinib-resistance. Chromatographic separation was carried out on an Acquity Ultra Performance Liquid Chromatography (UPLC) unit with a BEH C18 column (2.1 mm × 50 mm, 1.7 μm). The mobile phase was composed of acetonitrile and 0.1% formic acid in water. No endogenous interfering compounds was discovered at retention time of brigatinib (0.56 min) and imatinib (IS, 1.41 min). MS/MS detection was performed in positive mode. And the MRM transitions were m / z 584.09 → 484.08 and m / z 494.3 → 394.2 for brigatinib and IS, respectively. This method was assessed to be stable, specificity, and no matrix effect in three concentrations (0.004, 0.4, 4 μg/mL). The intra-day and inter-day precisions were less than 11.09% and 6.43%. And intra-day and inter-day accuracies were ranged from −3.88% to 5.44%. The recovery of brigatinib was from 85.26% to 96.05%. Additionally, the method had a good linearity in the range of 0.002–5 μg/mL. The presented method was effectively implemented to determine the concentration of brigatinib in rat plasma. [ABSTRACT FROM AUTHOR]

The main objective of this study was to investigate the ameliorative effects of aspirin eugenol ester (AEE) in hyperlipidemic rat. After five-week oral administration of AEE in high fat diet (HFD)-induced hyperlipidemic rats, the impact of AEE on plasma and urine metabonomics was investigated to explore the underlying mechanism by UPLC-Q-TOF/MS analysis. Blood lipid levels and histopathological changes of liver, stomach and duodenum were also evaluated after AEE treatment. Without obvious gastrointestinal (GI) side effects, AEE significantly relieved fatty degeneration of liver and reduced triglyceride (TG), low density lipoprotein (LDL) and total cholesterol (TCH) ( P < 0.01). Clear separations of metabolic profiles were observed among control, model and AEE groups by using principal component analysis (PCA) and orthogonal partial least-squares-discriminate analysis (OPLS-DA). 16 endogenous metabolites in plasma and 18 endogenous metabolites in urine involved in glycerophospholipid metabolism, fatty acid metabolism, fatty acid beta-oxidation, amino acid metabolism, TCA cycle, sphingolipid metabolism, gut microflora and pyrimidine metabolism were considered as potential biomarkers of hyperlipidemia and be regulated by AEE administration. It might be concluded that AEE was a promising drug candidate for hyperlipidemia treatment. These findings could contribute to the understanding of action mechanisms of AEE and provide evidence for further studies. [ABSTRACT FROM AUTHOR]

A rapid, simple and sensitive ultra-high performance liquid chromatography-tandem mass spectrometric (UPLC-MS/MS) method was developed and validated for the determination of hederasaponin B, an active triterpenoid saponin widely existed in Hedera helix L. Plasma samples were processed by protein precipitation with acetonitrile and separated on a Thermo Hypersil GOLD C18 (2.1 mm × 50 mm,1.9 μm) at flow rate of 0.3 ml/min, with a gradient elution consisting of acetonitrile and water containing 0.1% (v/v) formic acid at 30 °C and detected by electrospray ionization mass spectrometry in the positive multiple reaction monitoring (MRM) mode. The linearity was found to be within the concentration range of 0.5-5000 ng/ml with a lower limit of quantification of 0.5 ng/ml. The absolute oral bioavailability of hederasaponin B was 0.24 ± 0.49%. This indicated that the concentration-time course of the hederasaponin B existed a double-peak phenomenon. This method was further applied to the determination of hederasaponin B in rat plasma and showed good practicability, for the first time, after intragastric (25 mg/kg) and intravenous (2 mg/kg) administration in rats. [ABSTRACT FROM AUTHOR]

In this study, various neurosteroids in brain and plasma were simultaneously determined using liquid chromatography-tandem mass spectrometry and their profile changes in a stress-induced rats were investigated. The investigated neurosteroids are as follows: progesterone (P4), 5α-dihydroprogesterone (5α-DHP), 5β-dihydroprogesterone, estrone, androstenedione (AE), cortisol, cortisone, corticosterone (CORT), dehydroepiandrosterone (DHEA), pregnanolone (3α,5β-THP), allopregnanolone (ALLO), 11-deoxycorticosterone (DOC), 11-deoxycortisol, pregnenolone (PREG), and 5α/5β-tetrahydrodeoxycorticosterone (5α/5β-THDOC). Brain and plasma samples were processed using solid-phase extraction with methanol and acetic acid (99:1), and derivatized with a hydroxylamine reagent. Separation was achieved within 13 min at a flow rate of 0.4 mL/min with a C18 column (3.0 × 50 mm, 2.7 μm). The triple quadrupole mass spectrometer was operated in the positive electrospray ionization mode. Using this method, the neurosteroid level variation was quantitated and investigated in the brain and plasma upon immobilization stress in rats. As a result, AE, CORT, DOC, P4, 5α-DHP, 5α/5β-THDOC, DHEA, 3α,5β-THP, ALLO, and PREG levels were significantly altered in both the brain and plasma samples when stress was induced. These findings demonstrated that stress leads to the alteration of the GABAergic neurosteroid profile. The present results will be helpful for furthering an understanding of the role of neurosteroids in stressed conditions. [ABSTRACT FROM AUTHOR]

Benzoylpaeoniflorin (BP) is a potential therapeutic agent against oxidative stress related Alzheimer’s disease. In this study, a more rapid, selective, and sensitive liquid chromatography-tandem mass spectrometric (LC-MS/MS) method was developed to determine BP in rat plasma distinguishing with a monoterpene isomer, benzoylalbiflorin (BA). The method showed a linear response from 1 to 1000 ng/mL (r>0.9950). The precision of the interday and intraday ranged from 2.03 to 12.48% and the accuracy values ranged from −8.00 to 10.33%. Each running of the method could be finished in 4 minutes. The LC-MS/MS method was validated for specificity, linearity, precision, accuracy, recovery, and stability and was found to be acceptable for bioanalytical application. Finally, this fully validated method was successfully applied to a pharmacokinetic study in rats following oral administration. [ABSTRACT FROM AUTHOR]

Rheumatoid arthritis (RA), a chronic systemic inflammatory disorder affects many adults. Sinomenine, a natural product, has been clinically available for the treatment of RA in China. SND-117, a sinomenine derivative with much more potent activity, might serve as a candidate for anti-arthritis. The aim of the present study was to develop a sensitive and rapid high performance liquid chromatography tandem mass spectrometry (HPLC–MS/MS) method for quantification of SND-117 in rat plasma and to understand its absolute bioavailability. The HPLC–MS/MS method was developed and fully validated for determination of SND-117 in rat plasma, and the pharmacokinetic differences were investigated after different administration routes. The pharmacokinetics parameters were calculated by non-compartment model with DAS 3.0 software. After the oral or intravenous administration of different doses of SND-117, the time to peak is 1.5 h, half-life time is 8–10 h. The absolute oral bioavailability of SND-117 in rats was 9.60%. The results showed that SND-117 in rats was quickly absorbed, slowly eliminate, and the kinetics were linear. This method was suitable for pharmacokinetic studies of SNA-117 in rats. [ABSTRACT FROM AUTHOR]

Till date, no analytical method published to detect Cisplatin has been validated according to the U.S. Food and Drug Administration (FDA) guidance using liquid chromatography mass spectrometry (LC–MS/MS). We report, a validated LC–MS/MS method for quantitative determination of cisplatin in rat plasma and urine according to FDA guidlines. Cisplatin is a platinum containing compound used for the treatment of different types of cancers. Quantitative determination of cisplatin has been carried out using atomic absorption spectroscopy, high pressure liquid chromatography with phosphorescence, ultra-violet detection, or with inductively coupled plasma mass spectrometry. Few LC–MS/MS methods have been reported for the analysis of cisplatin either for direct quantification or indirect by derivatizing with organic compounds but none of the reported methods have validated the method. The developed and validated assay presented here is a highly sensitive LC–MS/MS method developed and validated for the quantitative determination of cisplatin following derivatization with diethyldithiocarbamate (DDTC) in order to detect platinum (Pt) of cisplatin, suitable for pharmacokinetic studies in rats and to further use it to study human toxicology. Chromatographic separation was achieved using a Poroshell 120 EC-C 18 column (3 × 50 mm, 2.7 μm) with a binary gradient mobile phase. Quantification was performed on a triple quadruple with electrospray ionization and detection was performed using multiple reaction monitoring. The method has a limit of detection of 1 ng/mL, and the quantifiable range was 3–3000 ng/mL in rat plasma and urine. The method was accurate and precise with an accuracy and precision for intra-day and inter-day of ±20% for lower limit of quantitation and of ±15% for low, mid and high quality control samples. This method was successfully applied to study the pharmacokinetic profile of cisplatin in rat plasma and urine given a range of doses from 0.5 to 3.5 mg/kg. [ABSTRACT FROM AUTHOR]

A sensitive and rapid ultra-performance liquid chromatography-electrospray ionization-mass spectrometry (UPLC–ESI–MS/MS) method was developed for the simultaneous analysis of anemoside B4, phellodendrine, berberine, palmatine, obakunone, esculin, esculetin, toosendanin (IS 1 of anemoside B4), tetrahydropalmatine (IS 2 of phellodendrine, berberine, palmatine and obakunone) and scopoletin (IS 3 of esculin and esculetin) and to compare the pharmacokinetics of these active ingredients in normal and ulcerative colitis rats. After methanol deproteinization, solvents were evaporated at 40 °C under a gentle stream of nitrogen. Chromatography was performed using a C18 column with a gradient elution of 0.1% aqueous formic acid and acetonitrile at 0.4 ml/min. Detection and measurement were performed on a 4000 QTRAP UPLC–MS/MS system from AB Sciex in the multiple reaction monitoring (MRM) mode. Phellodendrine, berberine, palmatine, obakunone, esculin, esculetin, tetrahydropalmatine (IS 2 ) and scopoletin (IS 3 ) were monitored under positive ionization conditions. Anemoside B4, and toosendanin (IS 1 ) were monitored under negative ionization conditions. The optimized mass transition ion-pairs ( m / z ) were 1221.1/750.7 for anemoside B4, 343.2/193.2 for phellodendrine, 337.1/321.0 for berberine, 353.0/336.9 for palmatine, 455.1/161.1 for obakunone, 341.2/179.2 for esculin, 179.1/123.0 for esculetin, 573.4/531.4 for toosendanin (IS 1 ), 356.2/192.2 for tetrahydropalmatine (IS 2 ) and 193.0/133.1 for scopoletin (IS 3 ). [ABSTRACT FROM AUTHOR]

A simple, sensitive and selective ultra-fast liquid chromatography-tandem mass spectrometry (UFLC–MS/MS) method was established for simultaneous determination and pharmacokinetic study of rosmarinic acid (RA), salvianolic acid D (Sal D), lithospermic acid (LA) and salvianolic acid B (Sal B) in rat plasma after intravenous administration of salvianolic acid for injection (SAFI). Three doses of administration, containing 14, 28 and 56 mg/kg, were investigated in this study. Plasma samples were pretreated using protein precipitation (PP) with pre-cooled acetonitrile. Chromatographic separation was achieved on a CORTECS™ UPLC C18 column (1.6 μm, 2.1 × 100 mm) with a mobile phase composed of 0.1% formic acid aqueous (V/V) and 0.1% formic acid acetonitrile (V/V). Analytes were detected using electrospray ionization (ESI) source in negative ionization mode and quantified in multiple reaction monitoring (MRM) mode. The validated method is stable and reliable. No significant difference of half lives ( t 1/2 ) of four analytes at three doses was observed. Area under the curve (AUC 0-∞ ) and peak concentration (C max ) of the four analytes demonstrated a linear increase in across the doses with the linear correlation r of each analyte at three doses were greater than 0.95. It indicated that the pharmacokinetic behavior of SAFI is positively related to dose at the range of 14–56 mg/kg. [ABSTRACT FROM AUTHOR]

Therapeutic monoclonal antibodies (mAbs) have heterogeneities in their structures. Multiple studies have reported that the variety of post-translational modifications could affect the pharmacokinetic profiles or pharmacological potencies of therapeutic mAbs. Taking into the account that the structural modification of mAbs would affect the efficacy, it is worth investigating the structural alteration of therapeutic mAbs in the blood and the relationship between their structures and pharmacological effects. Herein, we have developed the method to isolate rituximab from plasma in which endogenous IgGs interfere the detection of rituximab, and successfully developed the analytical method with a liquid chromatograph time-of-flight mass spectrometer to detect the structure of rituximab in plasma with errors less than 30 parts per millions. Eight types of carbohydrate chains in rituximab were detected by this method. Interestingly, time-dependent changes in carbohydrate chains such as AAF (G2F) and GnGn (G0) were observed in rats, although the amino acids were stable. Additionally, these structural changes were observed via incubation in plasma as in the rat experiment, suggesting that a certain type of enzyme in plasma caused the alterations of the carbohydrate chains. The present analytical methods could clarify the actual pharmacokinetics of therapeutic mAbs, and help to evaluate the interindividual variations in pharmacokinetics and efficacy. [ABSTRACT FROM AUTHOR]

It has been proved that the purine metabolic pathway has been implicated in various biological disorders including gout, diabetes, coronary heart diseases, and neurodegenerative diseases. The analysis of the purine metabolic pathway in organisms reveals important alterations under different physiological and pathological conditions, which contributes to the pathological study, diagnosis, and therapy of related diseases. In the present study, an ultra-high performance liquid chromatography with ultraviolet and tandem mass spectrometry (UHPLC-UV–MS/MS) method was developed for conducting the comprehensive analysis of the metabolite profiles of the purine pathway in rat plasma through a single analysis. The purine metabolites including adenosine-5′-monophosphate, guanosine-5′-monophosphate, adenosine, inosine, guanosine, inosine-5′-monophosphate, deoxyadenosine, deoxyguanosine, deoxyinosine, xanthine, hypoxanthine, and uric acid, were separated and quantified in the short running time of 10 min. After rapid chromatographic separation achieved by an Agilent Zorbax SB-Aq column, high concentration of uric acid and the remaining purine metabolites at lower levels were respectively detected by ultraviolet detector and triple quadruple mass spectrometry within a single analysis. The proposed method was validated by applying charcoal-stripped plasma as a matrix and it was proved to be linear ( R 2 > 0.982), accurate (with a relative error for accuracy < ± 15% and the relative standard deviation for intra- and inter-run precision <11%) and reproducible (with a matrix effect ranging between 86.49% and 111.44% with a maximum RSD of 8.69%). As a result, the method was successfully applied to the quantification of the endogenous purine metabolites in rat plasma. It was found that the concentration levels of the purine metabolites may keep a physiological balance as an integrated system in normal individuals and the concentration level of uric acid in rat plasma was 64 μM which was more than 200 times greater than the other purine metabolites. The established method and the measurement of the concentration of these purines in normal rat plasma may help the investigation of the action mechanisms between purine disorders and related diseases. [ABSTRACT FROM AUTHOR]

In the current work, a rapid, specific, sensitive and validated liquid chromatography tandem mass-spectrometric method was developed for the quantification of ponatinib (PNT) in human plasma and rat liver microsomes (RLMs) with its application to metabolic stability. Chromatographic separation of PNT and vandetanib (IS) were accomplished on Agilent eclipse plus C18 analytical column (50 mm × 2.1 mm, 1.8 μm particle size) maintained at 21±2°C. Flow rate was 0.25 mLmin-1 with run time of 4 min. Mobile phase consisted of solvent A (10 mM ammonium formate, pH adjusted to 4.1 with formic acid) and solvent B (acetonitrile). Ions were generated by electrospray (ESI) and multiple reaction monitoring (MRM) was used as basis for quantification. The results revealed a linear calibration curve in the range of 5–400 ngmL-1 (r2 ≥ 0.9998) with lower limit of quantification (LOQ) and lower limit of detection (LOD) of 4.66 and 1.53 ngmL-1 in plasma, 4.19 and 1.38 ngmL-1 in RLMs. The intra- and inter-day precision and accuracy in plasma ranged from1.06 to 2.54% and -1.48 to -0.17, respectively. Whereas in RLMs ranged from 0.97 to 2.31% and -1.65 to -0.3%. The developed procedure was applied for quantification of PNT in human plasma and RLMs for study metabolic stability of PNT. PNT disappeared rapidly in the 1st 10 minutes of RLM incubation and the disappearance plateaued out for the rest of the incubation. In vitro half-life (t1/2) was 6.26 min and intrinsic clearance (CLin) was 15.182± 0.477. [ABSTRACT FROM AUTHOR]

A rapid and sensitive ultra-fast liquid chromatography tandem mass spectrometry method, followed by simple protein precipitation and solid-phase extraction, has been developed and validated for the quantitative determination of four azo dyes (Para red, Solvent yellow 2, Solvent red 1 and Sudan red 7B) in rat plasma using D5-Sudan I as the internal standard. The optimal separation was accomplished on an Agilent Eclipse Plus C18 column (100 ? 2.1 mm, 1.8 µm) with gradient elution using the mobile phase including acetonitrile and water (containing 0.1% formic acid). The flow rate was 0.45 mL/min. The detection was conducted by means of electrospray ionization mass spectrometry in positive ion mode with multiple reaction monitoring. The calibration curves showed good linearity, with correlation coefficients >0.998 for all of the analytes within the concentration range. The lower limits of quantification (LLOQs) of Para red, Solvent yellow 2, Solvent red 1 and Sudan red 7B in rat plasma were 1.0, 0.1, 0.1 and 0.1 µg/L, respectively. The intra- and interday relative standard deviations were ≤9.6 and ≥12.4%, respectively, and the accuracy was in the range of -5.8 to -9.5%. The average recoveries were between 81.49 and 118.65%, and the matrix effects were satisfactory in the biological matrices. The fully validated method has been successfully applied in measuring levels of the four azo dyes in rat plasma following oral administration of 20.0 mg/kg of analytes in rats, which was suitable for the pharmacokinetic studies of the azo dyes. [ABSTRACT FROM AUTHOR]

A rapid and sensitive bioassay based on liquid chromatography-tandem mass spectrometry (LC-MS-MS) has been developed and validated to measure arbutin in rat plasma. Sample preparation of plasma after the addition of indapamide as internal standard (IS) involved solid-phase extraction (SPE) on C18 cartridges. Reversed-phase chromatography using acetonitrile and 0.5% formic acid solution (pH 2.56) was used for separation in a run time of 4.0 min. The analytes were detected in the negative ion mode using selective reaction monitoring (SRM) of the transitions at m/z 271.2→107.8 for arbutin and 364.3→189.0 for indapamide. The method has the following performance characteristics: a reliable response range of 7.5-5250.0 ng/mL with correlation coefficients (r) of >0.995. The lower limit of quantitation (LLOQ) was 7.5 ng/mL. The intra- and inter-day precision and accuracy of the quality control (QC) samples at low, medium and high concentration levels showed ≤8.79% relative standard deviation (RSD) and −1.15 to 1.49% relative error (RE). The method was successfully applied to a pharmacokinetic study to measure arbutin in rats after extracts of Vaccinium vitis-idaea was orally administered. [ABSTRACT FROM AUTHOR]

Phellodendrine, a quaternary ammonium alkaloid extracted from the dried bark of Phellodendrom chinensis Schneid and Phellodendrom amurense Rupr, has the effect of suppressing cellular immune response, reducing blood pressure and antinephritis. However, few investigations have been conducted for the pharmacokinetic study of phellodendrine. Thus, a rapid, simple and reliable ultra-high performance liquid chromatography–tandem quadrupole mass spectrometry (UHPLC–QQQ MS/MS) method has been established for quantification of phellodendrine in rat plasma and tissues by using magnoflorine as internal standard. The chromatographic separation was achieved on an Agilent ZORBAX SB-C18 column (4.6 mm × 50 mm, 1.8 μm) by gradient elution using 0.1% aqueous formic acid (A) and methanol (B). Triple quadrupole mass detection with multiple reaction monitoring mode was used to monitor the ion transitions, at m / z 342.20 → 192.20 for phellodendrine and m / z 342.20 → 58.20 for internal standard, respectively. The developed method was fully validated and successfully applied to the pharmacokinetics and tissue distribution study of phellodendrine after intravenous administration. The lower limits of quantification were 0.5 ng/mL for plasma samples, 2.5 ng/g for brain and 1 ng/g for other tested tissues. Precisions and accuracy values were within the Food and Drug Administration acceptance criteria, the recovery and matrix effects were between 87.8–113.5%. The area under the curve (AUC 0−t ) ranged from 15.58 to 57.41 mg/L min and Cmax were between 1.63–4.93 mg/L. The results showed that phellodendrine was eliminated in 120 min in plasma and most of tissues and the highest concentrations of phellodendrine were found in the kidney. This study may provide a basis for the further study of phellodendrine. [ABSTRACT FROM AUTHOR]

A rapid, sensitive and selective ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS-MS) was developed and validated for the determination and pharmacokinetic investigation of apremilast in rat plasma. Sample preparation was accomplished through a simple one-step deproteinization procedure with 0.2 mL of acetonitrile to a 0.1 mL plasma sample. Plasma samples were separated by UPLC on an Acquity UPLC BEH C18 column using a mobile phase consisting of acetonitrile-0.1% formic acid in water with gradient elution. The total run timewas 3.0 min, and the elution of apremilast was at 1.27 min. The detection was performed on a triple quadrupole tandem mass spectrometer in the multiple reaction-monitoring mode using the respective transitions m/z 461.3→257.1 for apremilast and m/z 237.2→194.2 for carbamazepine (internal standard). The calibration curve was linear over the range of 0.1-100 ng/mL with a lower limit of quantitation of 0.1 ng/mL. The mean recovery of apremilast in plasmawas in the range of 83.2-87.5%. Both intraday and interday precision were <9.6%. This method was successfully applied in the pharmacokinetic study after oral administration of 6.0 mg/kg apremilast in rats. [ABSTRACT FROM AUTHOR]

Tamoxifen (TAM) is a non-steroidal estrogen receptor antagonist that enhances erlotinib (ERL)-induced cytotoxicity in the treatment of NSCLC. ERL and TAM are metabolized by CYP3A4 enzymes. In addition, both drugs have the potential of altering the enzymatic activity through either inhibition (ERL) or induction (TAM). Thus it was expected that pharmacokinetics (PK) drug–drug interactions (DDIs) could be encountered following their co-administration. In this respect, a bioanalytical UPLC–MS/MS method has been developed and validated for the simultaneous determination of ERL and TAM in rat plasma samples, using ondansetron (OND) as an internal standard (IS). Plasma samples were prepared using mixed mode cationic solid phase extraction (SPE) STRATA™ −X-C 33 μm cartridges with good extraction recovery of both drugs from rat plasma (E r % from −13.92 to −3.32). The drugs were separated on a Waters BEH™ C18 column with an isocratic elution using a mobile phase composed of a mixture of acetonitrile and water, each with 0.15% formic acid, in the ratio of 80: 20, v/v. Quantitation was carried out using the positive ionization mode with multiple reaction monitoring (MRM) at m / z 394.20 > 278.04 (ERL), m / z 372.25 > 72.01 (TAM), and m / z 294.18 > 170.16 (OND). The method was fully validated as per the FDA guidelines over the concentration range of 0.2–50 ng/mL with very low lower limit of quantification (LLOQ) of 0.2 ng/mL for both ERL and TAM. The intra- and inter-day assay precision (in terms of relative standard deviation, RSD) and accuracy (in terms of percentage relative error, % E r ) were evaluated for both drugs and the calculated values evaluated at four different concentration levels were within the acceptable limits (<15%) for concentrations other than LLOQ and 20% for LLOQ. The method was successfully applied to the study of possible PK-DDI following the oral administration of ERL and TAM in a combination, compared to their single administration. [ABSTRACT FROM AUTHOR]

Gout is a common metabolic disorder caused by the deposition of monosodium urate crystals within joints. A new kind of xanthine oxidase inhibitor, WSJ-537, was developed as a potential drug. In order to investigate the pharmacokinetic behavior in vivo , a liquid chromatography-tandem mass spectrometry (LC–MS/MS) method for determination the concentration of WSJ-537 in rat plasma was developed. After extraction by protein precipitation method with acetonitrile, the chromatographic separation was accomplished on a Venusil ASB C 18 column(2.1 mm × 50 mm, 3 mm)at a flow rate of 0.3 mL min −1 with the mobile phase consisting of acetonitrile-ammonium acetate (33:67, v/v ). An electrospray ionization (ESI) source was applied and operated in the positive ion mode. The plasma concentration was detected by multiple reactions monitoring (MRM) mode with the target fragment ions m / z 410.2 → m / z 368.1 for WSJ-537 and m / z 244.1 → m / z 185.0 for the IS. Good linearity was observed in the range of 20–800 ng mL −1 (r = 0.9947). The recovery of WSJ-537 in rats plasma was more than 85%. This method was suitable for pharmacokinetic studies after oral administration of 10 mg/kg WSJ-537 in rats. [ABSTRACT FROM AUTHOR]

Acotiamide is a new prokinetic drug that is used to treat functional dyspepsia (FD). A sensitive and specific LC-MS-MS method has been developed and validated for the analysis of acotiamide in rat plasma. The assay involved a simple protein precipitation (PPT) step with methanol-acetonitrile (50:50, v/v) and a gradient elution using a mobile phase consisting of water containing 0.1% formic acid and methanol containing 0.1% formic acid. The analytes were chromatographed on a reversephase Agilent Zorbax XDB C18 column (2.1 mm× 50 mm, 3.5 μm) with a flowrate of 0.50 mL/min. The analytes were monitored by tandem-mass spectrometry with positive electrospray ionization. The precursor-product transitions (m/z) in the positive ion mode were 451.4→271.3 and 386.2→122.2 for acotiamide and buspirone (internal standard, IS), respectively. The assay was shown to be linear over the range of 0.10-200 ng/mL, with a lower limit of quantification of 0.10 ng/mL. The method was shown to be reproducible and reliable with the inter- and intra-batch accuracy and precision were within ±15%. The assay has been successfully used for pharmacokinetic evaluation of acotiamide after intravenous and oral administration of 10 mg/kg acotiamide in rats. The oral absolute bioavailability (F) of acotiamide in rats was estimated to be 38.4 ± 13.5% with an elimination half-life (t1/2) value of 9.11 ± 0.40 h. [ABSTRACT FROM AUTHOR]

AM-2201 is a popular synthetic cannabinoid first synthesized in 2000. AM-2201 pharmacokinetic and pharmacodynamic data are scarce, requiring further investigation. We developed a sensitive method for quantifying AM-2201 and 13 metabolites in plasma to provide a tool to further metabolic, pharmacokinetic and pharmacodynamic studies. Analysis was performed by liquid chromatography-tandem mass spectrometry. Chromatographic separation was performed by gradient elution on a biphenyl column with 0.1% formic acid in water/0.1% formic acid in acetonitrile:methanol 50:50 ( v/v ) mobile phase. Sample preparation (75 μL) consisted of an enzymatic hydrolysis and a supported liquid extraction. The method was validated with human plasma with a 0.025 or 0.050–50 μg/L working range, and cross-validated for rat plasma. Analytical recovery was 88.8–110.1% of target concentration, and intra- (n = 30) and inter-day (n = 30) imprecision < 11.9% coefficient of variation. Method recoveries and matrix effects ranged from 58.4–84.4% and −62.1 to −15.6%, respectively. AM-2201 and metabolites were stable (±20%) at room temperature for 24 h, at 4 °C for 72 h, and after three freeze-thaw cycles, and for 72 h in the autosampler after extraction. The method was developed for pharmacodynamic and pharmacokinetic studies with controlled administration in rats but is applicable for pre-clinical and clinical research and forensic investigations. Rat plasma specimen analysis following subcutaneous AM-2201 administration demonstrated the suitability of the method. AM-2201, JWH-018 N -(5-hydroxypentyl), and JWH-018 N -pentanoic acid concentrations were 4.8 ± 1.0, 0.15 ± 0.03, and 0.34 ± 0.07 μg/L, respectively, 8 h after AM-2201 administration at 0.3 mg/kg (n = 5). [ABSTRACT FROM AUTHOR]

Rhodamine B is an illegal and potentially carcinogenic food dye. The aim of this study was to develop a convenient, rapid, and sensitive UHPLC-MS/MS method for pharmacokinetic studies in rats. Rat plasma samples were deproteinized with acetonitrile and separated by UHPLC on a reverse-phase C18e column (100 mm × 2.1 mm, 2 μm) using a mobile phase consisting of methanol—5 mM ammonium acetate (90:10, v/v). Detection was performed using a triple quadrupole tandem mass spectrometer in the selected reaction monitoring mode at [M] + ion m / z 443.39 → 399.28 for rhodamine B and [M+H] + ion m / z 253.17 → 238.02 for 5-methoxyflavone as the internal standard. This method was specific and produced linear results over a concentration range of 0.5–100 ng/mL, with a lower limit of quantitation of 0.5 ng/mL. All validation parameters, including the inter-day, intra-day, matrix effect, recovery, and stability in rat plasma, were acceptable according to the biological method validation guidelines developed by the FDA (2001). This method was successfully applied to a pharmacokinetic study in rats; oral administration of 1 mg/kg of rhodamine B yielded a time to maximum concentration ( T max ) of 1.3 ± 0.4 h and an elimination half-life of 8.8 ± 1.4 h, with a clearance of 229.7 ± 19.4 mL/h/kg. These pharmacokinetic results provide a constructive contribution to our understanding of the absorption mechanism of rhodamine B and support additional food safety evaluations. [ABSTRACT FROM AUTHOR]

A simple, sensitive and accurate ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method for the determination of AKI603 in rat plasma has been firstly developed and validated. After a simple liquid–liquid extraction (LLE) with ethyl acetate, the analytes were separated on C18 column (2.1 × 100 mm, 1.9 μm, Thermo) by gradient elution with mobile phase of water (A) (containing 5 mM ammonium acetate and 0.1% formic acid) and methanol (B) with a flow rate of 0.3 mL min −1 and then analyzed by mass spectrometry in the positive multiple reactions monitoring (MRM) mode. The mass transitions monitored were m / z 410.0 → 352.9, m / z 457.1 → 367.9 for AKI603 and internal standard (Ly-7z), respectively. The developed method was validated for specificity, linearity and lower limit of quantification, intra- and inter-day precision and accuracy, extraction recovery, matrix effect and stability whose values satisfied the acceptable limits. The calibration curves for AKI603 was linear in concentration ranges of 0.025–5000 ng mL −1 . The method has been successfully used to the bioavailability study of AKI603 administered to rats intravenously (2.5 mg/kg) or orally (25 mg/kg). The oral bioavailability of AKI603 in rats was calculated as 28.7 ± 9.7%. [ABSTRACT FROM AUTHOR]

Sarsasapogenin, a natural compound from Chinese medical herb Anemarrhena asphodeloides Bge., has recently received a great deal of attention due to its various bioactivities. In this study, an easy and applicable liquid chromatography tandem mass spectrometry method for the quantification of sarsasapogenin in rat plasma was developed. Sample preparation was accomplished through a simple one-step protein precipitation procedure with methanol. Negative electrospray ionization was performed using multiple reactions monitoring (MRM) mode with transitions of m / z 417.4/273.2 for sarsasapogenin, and 415.2/271.4 for diosgenin (internal standard). The calibration curve was linear over the range of 0.5–500 ng/mL (r = 0.9994), with a lower limit of quantification at 0.5 ng/mL. The RSD of intra- and inter-day precision was below 6.41%, and accuracy ranged from 87.60% to 99.20%. The RSD of matrix effect and recovery yield was within ±15% of nominal concentrations and sarsasapogenin was stable during stability tests. This validated method had been successfully applied to the preclinical pharmacokinetic studies of sarsasapogenin in rats. The half-life (t 1/2 ) was (15.1 ± 2.3), (16.1 ± 3.0) and (15.4 ± 3.9) h after single intragastric administration of 25, 50 and 100 mg/kg sarsasapogenin, respectively. And it was found that, the area under the plasma concentration versus time curve (AUC 0-72h ) and the maximum plasma concentration (C max ) were linearly related to dose. [ABSTRACT FROM AUTHOR]

In this study, we developed and validated a new, rapid, specific and sensitive ultra-performance liquid chromatography–tandem mass spectrometric (UPLC–MS/MS) method to simultaneously determine parecoxib sodium (PX) and its active metabolite, valdecoxib (VX), in rat plasma. Plasma samples were prepared by plasma protein precipitation combined with a liquid-liquid extraction method. The separation was carried out on a Kinetex C 18 column (2.1 mm × 50 mm, 2.6 μm) with a gradient elution using methanol (A) and a 2 mM ammonium acetate aqueous solution (B). The analysis was performed in less than 3 min with a flow rate of 0.2 mL/min. Ketoprofen was used as an internal standard (IS). Mass spectrometric detection was conducted with a triple quadrupole detector equipped with electrospray ionization in the negative ion mode (ESI − ) using multiple reaction monitoring (MRM). The calibration curves were linear over the concentration ranges of 5–4000 ng/mL for PX and 5–2000 ng/mL for VX with all correlation coefficients greater than 0.998. The intra- and inter-day relative standard deviations (RSD) for both analytes were within 15% and the accuracy was within 85–115% at all quality control levels. The mean extraction recoveries for all analytes obtained from three concentrations of QC plasma samples were more than 89.0% efficient. Selectivity, matrix effect, dilution integrity and stability were also validated. The method was successfully used to investigate the pharmacokinetics of PX and VX in rat plasma after intravenous and intramuscular administration of PX. [ABSTRACT FROM AUTHOR]

7′( Z )-(8″ S , 8‴ S )-epi-Salvianolic acid E (compound 1 ) and (7 ′R , 8′ R , 8″ S , 8‴ S )-epi-salvianolic acid B (compound 2 ), two novel analogs of salvianolic acid B (Sal B), have been recently isolated from Salvianolic acid for injection. They both show powerful antioxidant effects, including inducing NQO1 activity and scavenging DPPH free radical, and potential protecting effects for cerebral ischemia. However, no reports have been described the pharmacokinetic study of them. In this study, an ultra-fast liquid chromatography-tandem mass spectrometry (UFLC-MS/MS) method was developed and validated for the determination of compound 1 , compound 2 and Sal B in rat plasma, respectively. Plasma samples were pretreated by liquid–liquid extraction with ethyl acetate. Chromatographic separation was achieved on a Waters Acquity UPLC ® HSS T3 column (1.7 μm particles, 2.1 mm i.d. × 100 mm) with the mobile phase of 0.1% aqueous formic acid (A)-acetonitrile (B) (65:35, v/v). Quantification was performed on a triple quadruple tandem mass spectrometry with electrospray ionization (ESI) by multiple reaction monitoring (MRM) in the negative ion mode. Monitored transitions were set at m / z 717.0 → 519.0, 717.1 → 519.1, 717.2 → 518.9 and 320.9 → 152.1 for compound 1 , compound 2 , Sal B and chloramphenicol (internal standard, IS), respectively. Linear calibration curves were acquired over the concentration range of 2.0–1000 ng/mL for the three analytes in rat plasma. The extraction recoveries, matrix effects, intra- and inter-day precisions and accuracies of the three analytes were all within acceptable limits. The validated method was successfully applied to the pharmacokinetic study of compound 1 , compound 2 and Sal B after intravenous administration of 6.0 mg/kg in rats, respectively. The results indicated that compound 1 and compound 2 were both eliminated more slowly than Sal B. Exposure levels of both compound 1 and Sal B were higher than compound 2 in the same dosage range. This study provided critical reference for the pharmacokinetic study of compound 1 and compound 2 . [ABSTRACT FROM AUTHOR]

A sensitive, simple and rapid analytical method based on a liquid chromatography–tandem mass spetrometry (LC–MS/MS) has been established and validated for the determination of stachyose in rat plasma. Plasma samples were prepared by protein precipitation with acetonitrile. Separation of stachyose and nystose (internal standard, IS) was achieved using acetonitrile-water (55:45, v/v) as the mobile phase at a flow rate of 1 ml/min for 6 min on an Asahipak NH 2 P-50 4E column with an Asahipak NH2P-50G 4A guard column. Detection and quantification were conducted by LC–MS/MS method in the negative ion mode using multiple reaction monitoring (MRM) transitions at m / z [M–H] − 665.4 → 383.1 for stachyose and 665.5 → 485.0 for IS, respectively. The method was linear over the concentration ranges of 100–30000 ng/ml with a lower limit of quantification (LLOQ) of 100 ng/ml. The intra- and inter- day precision were all within 8.7% and the accuracy ranged from 97.2–108.4% and 98.3–102.4%, respectively. Stability studies indicated that stachyose was stable under short-term, long-term and three freeze-thaw storage conditions. The method was successfully applied to a pharmacokinetic study involving pulmonary administration of micronized Rehmannia glutinosa oligosaccharides (RGOS) to rats. [ABSTRACT FROM AUTHOR]

Lurbinectedin (PM01183) is a new synthetic tetrahydroisoquinoline alkaloid that binds to selected sequences in the minor groove of DNA, inducing PM01183-DNA adducts that stall replication, DNA repair and transcription and gives rise to double-strand breaks and finally, caspase-dependent apoptotic cell death. PM01183 has demonstrated clinical antitumor activity in platinum resistant/refractory ovarian cancer patients. A rapid and sensitive liquid chromatography/tandem mass spectrometry assay was developed and validated to quantify PM01183 in plasma from nonclinical species. The bioanalysis consisted of a supported liquid extraction, followed by a gradient phase chromatography and, detection by positive ion electrospray tandem mass spectrometry. The calibration range for PM01183 was established using PM01183 standards from 0.1 to 100 ng/mL in blank plasma. The multiple reaction monitoring, based on the transition m / z 767.3 → 273.0, was specific for PM01183, and that based on the transition m / z 771.4 → 277.0 was specific for the internal standard (deuterated PM01183). No endogenous material interfered with the analysis of PM01183 and the internal standard from blank plasma. The limit of detection (LOD) of the assay was calculated as 0.025 ng/mL. The correlation coefficients for the calibration curves ranged from 0.9937 to 0.9987. The mean inter-day accuracies for all calibration standards ranged from 92 to 108% (≤8% bias), and the mean inter-day precision for calibration standards was always less than 12%. The mean intra and inter-day assay accuracy for all quality control replicates remained between 91 and 109%. The mean intra and inter-day assay precision was less than 10% for all QC levels. The method was validated to demonstrate the specificity, recovery, limit of quantification, accuracy and precision of measurements. The assay has been used to support preclinical pharmacokinetic and toxicokinetic studies of PM01183 in nonclinical species. The main PK parameters in dogs (3 male and 3 female, respectively) were calculated as follows: maximum concentration ( C max , 12.9 ± 0.6 and 10.2 ± 3.0 ng/mL) and the area under the plasma concentration-time curve (AUC, 24.9 ± 0.7 and 22.6 ± 6.1 ng h/mL). The results showed that plasma samples could be monitored for PM01183 for long enough to accurately estimate pharmacokinetics information. [ABSTRACT FROM AUTHOR]

Semen Oroxyli, a Traditional Chinese Medicine, has many significant pharmacological activities such as analgesic, apoptotic, anti-inflammatory, anticancer, and immunostimulant activities. A sensitive and selective liquid chromatography–tandem mass spectrometry (LC–MS/MS) method was developed and validated for the simultaneous quantitation of four flavonoids (oroxin A, oroxin B, baicalin, and chrysin) of Semen Oroxyli in rat plasma. After the addition of internal standard, plasma samples were pretreated with acetonitrile via a single-step protein precipitation. Chromatographic separation was performed on a Capcell Pak C 18 column (100 mm × 2.0 mm, 5 μm particles) with isocratic elution using a mobile phase of methanol and 2 mM ammonium acetate buffer solution (75:25, v/v) at a flow rate of 0.45 mL/min. The analytes were detected without interference in the selection reaction monitoring mode with negative electrospray ionization. The validated method exhibited good linearity over a wide concentration range (r ≥ 0.9958), and the lower limits of quantification were 1.0–5.5 ng/mL for all the analytes. The mean extraction recoveries of the analytes from rat plasma exceeded 80.6%. The intra- and inter-day precisions at three QC levels were both less than 11.5%, and the accuracies ranged from −6.2% to 10.3%. The LC–MS/MS method was successfully applied to a pharmacokinetic study of the four flavonoids in rat plasma after oral administration of Semen Oroxyli extract. [ABSTRACT FROM AUTHOR]

A simple, rapid and sensitive method based on liquid chromatography–tandem mass spectrometry (LC–MS/MS) has been developed and validated for the quantitative determination in rat plasma of a new candidate for AD treatment, namely PC 48 (a 7-MEOTA-donepezil like compound) in rat plasma. Sample preparation involved pH adjustment with sodium hydroxide followed by solvent extraction with ethyl acetate:dichloromethane (80:20, v/v). The chromatographic separation was achieved on an Ascentis Express RP-Amide column (75 mm × 2.1 mm, 2.7 μm) with a gradient mobile phase consisting of 0.05 M aqueous formic acid and acetonitrile. Detection was carried out using positive-ion electrospray tandem mass spectrometry on an LTQ XL system using the MS/MS CID (collision-induced dissociation) mode. The method was linear in the range 0.1–1000 ng/ml (r 2 = 0.999) with a lower limit of quantitation of 0.1 ng/mL. Extraction recovery was in the range 63.5–72.1% for PC 48 and 70.5% for reserpine (internal standard, IS). Intra- and inter-day precisions measured as relative standard deviation were below 10.8% and accuracy was from −7.2% to 7.4%. The method was successfully applied to a pharmacokinetic study involving intramuscular application of 3.86 mg/kg PC 48 to rats for the first time. Pharmacokinetic parameters for PC 48 include C max 39.09 ± 4.45 ng/mL, T max 5.00 ± 3.08 min, AUC 0 − t 23374 ± 4045 min ng/mL and t 1/2 1065 ± 246 min. [ABSTRACT FROM AUTHOR]

A rapid, sensitive and specific ultra performance liquid chromatography-tandem mass spectrometry (UPLC–MS/MS) method for the quantification of shanzhiside methyl ester, 8- O -acetylshanzhiside methyl ester and luteolin-7- O -β- d -glucopyranoside of Lamiophlomis rotata Pill in rat plasma was developed and validated. After liquid–liquid extraction with n -butyl alcohol/ethyl acetate (70:30, v/v ), analytes and paeoniflorin (internal standard, IS) were separated on an Acquity BEH UPLC C 18 column (100 × 2.1 mm, 1.7 μm) with gradient elution at a flow rate of 0.2 mL/min. All calibration curves had good linearity (r > 0.9929) over the concentration ranges of 1–1000 ng/mL for shanzhiside methyl ester and 8- O -acetylshanzhiside methyl ester, 0.3–150 ng/mL for luteolin-7- O -β- d -glucopyranoside. The intra- and inter-day precisions were all within 11.1% and the accuracy (relative error, RE%) all ranged from −13.6% to 5.3%. The method also guaranteed an acceptable selectivity, recovery and stability, which was successfully applied to a pharmacokinetic study of the three analytes in rats after oral administration of Lamiophlomis rotata Pill. [ABSTRACT FROM AUTHOR]

For the first time, a rapid and sensitive high-performance liquid chromatography-tandem mass spectrometry (LC–MS/MS) method was developed for the simultaneous determination of fraxin and its metabolite, fraxetin, in rat plasma, using esculin as the internal standard (IS). The plasma samples were precipitated with methanol before separation on an Nova-Pak C 18 column (150 mm × 3.9 mm, 3 μm) using a mobile phase consisting of 0.1% formic acid and methanol (55:45) at a flow rate of 0.8 mL/min. The analytes were detected by multiple reaction monitoring in the negative ion mode with the mass transitions at m / z 368.9→ m / z 191.9 (fraxin), m / z 206.9→ m / z 191.8 (fraxetin) and m / z 339.0→ m / z 176.9 (esculin, IS). The results demonstrated that the calibration curves for both analytes have good linearity (r ≥ 0.995) over a concentration range of 5.00–3000 ng/mL. The assay was validated according to the regulatory bioanalytical guidelines and proved acceptable. The intra- and inter-day precisions (R.S.D.%) were within 10.9% for both analytes, whereas the deviation of assay accuracies (R.E.%) ranged from −5.3 to 1.0%. The method was successfully applied to a pharmacokinetic study after a single oral dose of fraxin at 50 mg/kg to rats. [ABSTRACT FROM AUTHOR]

Rapid, sensitive, selective and accurate ultra performance liquid chromatography with tandem mass spectrometry (UPLC–MS/MS) method was developed and validated for the quantification of elbasvir (ELB) in rat plasma with deuterated elbasvir (ELB-D6) as internal standard (IS).Sample preparation was done by protein precipitation using acetonitrile containing 50 ng/mL IS. Chromatographic separation was achieved by an UPLC BEH C18 column (2.1 mm × 50 mm, 1.7 μm) column with a gradient mobile phase consisting of acetonitrile–water (containing 5.0 mM ammonium acetate with 0.01% acetic acid, pH 4.5) as mobile phase at a flow rate of 0.3 mL/min for 3 min. ELB was monitored using positive electrospray triple quadrupole mass spectrometer (Waters Xevo TQ-S) via multiple reaction monitoring (MRM) mode. The monitored transitions were set at m/z 882.51 → 656.42 and m / z 888.49 → 662.43 for ELB and ELB-D6, respectively. The achieved lower limit of quantification was 1.0 ng/mL. The validated method had an excellent linearity in the range of 1.0–2000 ng/mL (r 2 > 0.996). Recovery efficiency at three levels QC concentrations of 2.0 (low), 160 (medium) and 1600 (high) ng/mLwas in the range of 98.29–106.40% for ELB. Matrix effect was found to be minimal. The intra- and inter-day precisions were less than 7.01%. The intra- and inter-day accuracies were determined to be within ±6.23% for all accuracy measurements. The validated simple and rapid UPLC–MS/MS method was successfully used to the pharmacokinetics study of ELB in rats, providing its applicability in relevant preclinical studies. [ABSTRACT FROM AUTHOR]

Pinostilbene (3-methoxyresveratrol or trans -3,4′-dihydroxy-5-methoxystilbene) is a naturally occurring monomethylether analogue of resveratrol ( trans -3,5,4′-trihydroxystilbene) that exhibits various pharmacological activities. To further examine its medicinal potential, a sensitive LC–MS/MS method was developed and validated for the determination of pinostilbene in rat plasma. Heavy Isotope labelled resveratrol was used as an internal standard. The ESI was operated in its negative ion mode while pinostilbene and resveratrol were measured by multiple reaction monitoring (MRM) using precursor-to-product ion transitions of m / z 241 → 181 and m / z 233 → 191, respectively. This LC–MS/MS method had excellent selectivity, sensitivity (LLOQ = 1 ng/ml), accuracy (both intra- and interday analytical recovery within 100 ± 15%) and precision (both intra- and interday RSD < 15%). The matrix effect was insignificant. The pharmacokinetics of pinostilbene was subsequently profiled in Sprague–Dawley rats. Upon intravenous administration (5 or 10 mg/kg), pinostilbene displayed rapid clearance (Cl = 129 ± 42 or 107 ± 31 ml/min/kg) and extremely short mean transit time (MTT = 6.24 ± 0.41 or 8.52 ± 1.38 min). After oral dosing (50 mg/kg), the bioavailability of pinostilbene was limited but highly erratic ( F = 1.87 ± 2.67%). Pharmacokinetic comparison among pinostilbene, resveratrol and some resveratrol analogues suggested that stilbenes with meta-hydroxyl group(s) may be associated with metabolic instability and subsequently suffer from rapid clearance and low oral bioavailability. The information obtained from this study will facilitate further exploration on pinostilbene as well as other resveratrol analogues. [ABSTRACT FROM AUTHOR]

A simple and rapid quantitative UPLC–MS/MS method for simultaneous determination of picfeltarraenins IA and IB in rat plasma was developed and validated in accordance with the US FDA Bioanalytical Guidance (2001). Analytes were extracted from rat plasma by using methanol and separated on Agilent ZORBAX SB-C 18 (50 mm × 2.1 mm, 1.8 μm) column by using a mobile phase composed of methanol and water (70:30, v/v). Eluents were monitored by ESI tandem mass spectrometry detection with SRM mode using ion transitions m / z 785.4 → 639.5, m / z 815.5 → 669.5, and m / z 763.5 → 455.3 for picfeltarraenin IA, picfeltarraenin IB, and internal standard, respectively. The method was validated over the linear range of 11.5–1150 ng/mL and 13.0–1300 ng/mL. The developed analytical method was applied to support a pharmacokinetic study on simultaneous estimation of picfeltarraenins IA and IB in rats. [ABSTRACT FROM AUTHOR]

A sensitive and rapid ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method was developed to determine tedizolid and linezolid in rat plasma simultaneously. Chromatographic separation was carried out on an Acquity UPLC BEH C18 column and mass spectrometric analysis was performed using a XEVO TQD triple quadruple mass spectrometer coupled with an electrospray ionization (ESI) source in the positive ion mode. Multiple reaction monitoring (MRM) mode was used for quantification using target fragment ions m/z 371.4 → 343.2 for tedizolid, and m/z 338.3 → 56.1 for linezolid. This assay method has been fully validated in terms of selectivity, linearity, recovery and matrix effect, accuracy, precision and stability. The linearity of this method was found to be within the concentration range of 5–5000 ng/mL for tedizolid, and 10–10,000 ng/mL for linezolid in rat plasma, respectively. Only 3.0 min was needed for an analytical run. This assay was used to support a preclinical study where multiple oral doses were administered to rats to investigate the pharmacokinetics of tedizolid and linezolid. [ABSTRACT FROM AUTHOR]

Psoralea corylifolia L. has long been used in traditional Chinese medicine for treating and preventing many diseases. A group of flavonoid components are regarded as the active principals within the seeds. In this research, a rapid, accurate and sensitive ultra high pressure liquid chromatography tandem mass spectrometry (UHPLC/MS/MS) method has been established for simultaneous quantification of its 5 main components, namely, neobavaisoflavone, bavachin, isobavachalcone, bavachinin and corylifol A in rats’ plasma after the rats were orally administrated with Buguzhi extract. Negative ion electrospray mode was applied in the detection process. Multiple reactions monitoring (MRM) mode was utilized for simultaneous quantitative analyzing of neobavaisoflavone ( m / z 321.1 → m / z 265.1), bavachin ( m / z 323.1 → m / z 119.0), isobavachalcone ( m / z 323.2 → m / z 119.0), bavachinin ( m / z 337.2 → m / z 119.0), corylifol A ( m / z 389.2 → m / z 277.0) and liquiritigenin (Internal Standard, m / z 255.1 → m / z 119.0). Chromatographic separation of the above mentioned components was conducted on a Waters BEH-C18 column (100 mm × 2.1 mm, 1.7 μm) with gradient elution system at flow rate of 0.3 mL/min. The mobile phase was composed of acetonitrile and 0.1% formic acid solution. The lower limit of quantification (LLOQ) for each of the above analytes was 0.5 ng/mL. Each of the analytes exhibited good linearity within the concentration range of 0.5–100 ng/mL. The method was fully validated for its selectivity, accuracy, precision, stability, matrix effect and extraction recovery. The validated method has been successfully applied for simultaneous determination of the 5 flavonoids in rat plasma for the first time. [ABSTRACT FROM AUTHOR]

A chiral UFLC-MS/MS method was established and validated for quantifying d -threo-methylphenidate ( d -threo-MPH), l -threo-methylphenidate ( l -threo-MPH), d -threo-ethylphenidate ( d -threo-EPH), l -threo-ethylphenidate ( l -threo-EPH) and d , l -threo-ritalinic acid ( d , l -threo-RA) in rat plasma over the linearity range of 1–500 ng/mL. Chiral separation was performed on an Astec Chirobiotic V2 column (5 μm, 250 × 2.1 mm) with isocratic elution using methanol containing 0.003% ammonium acetate (w/v) and 0.003% trifluoroacetic acid (v/v) at a flow of 0.3 mL/min. All analytes and IS were extracted from rat plasma by a one-step liquid–liquid extraction (LLE) method. The intra- and inter-run accuracies were within 85–115%, and the intra- and inter-run precision were <10% for all analytes. Extraction recoveries were 55–62% for d -threo-MPH, 54–60% for l -threo-MPH, 55–60% for d -threo-EPH, 53–57% for l -threo-EPH and 25–30% for d , l -threo-RA. The validated UFLC-MS/MS method successfully applied to the pharmacokinetic interaction study of oral d -threo-MPH and l -threo-MPH (alone or in combination) in female Sprague Dawley rats. The EPH was not detected in rat plasma following oral administrated MPH without EtOH. As far as it is known to the authors, this study is the first one step liquid–liquid extraction method to extract and UFLC-MS/MS method to quantify d -threo-MPH, l -threo-MPH, d -threo-EPH, l -threo-EPH and d , l -threo-RA simultaneously. [ABSTRACT FROM AUTHOR]

The adrenal gland is the most common toxicological target in the endocrine system, and inhibition of adrenal steroidogenesis by drugs can be fatal in humans. However, methods to evaluate the drug effect are limited. Recently, simultaneous measurement of multiple steroids, including precursors, has become possible. Here, we evaluated the usefulness of this simultaneous measurement for the evaluation of drug effects on adrenal steroidogenesis in vivo. For this purpose, we measured plasma concentrations of adrenal steroids in rats dosed with ketoconazole, a known inhibitor of adrenal steroidogenesis, and examined its relationship with the changes in histopathology and mRNA expression of steroidogenic enzymes in the adrenal gland. Ketoconazole (150 mg/kg/day) was orally administered to male rats for 7 days. The adrenal weight was high, and the zona fasciculata/reticularis were hypertrophic with an accumulation of lipid droplets. mRNA expression of CYP11A1, a rate-limiting enzyme in adrenal steroidogenesis, was slightly high in the adrenal gland. Plasma concentration of deoxycorticosterone was markedly high, while there were no significant changes in that of corticosterone, progesterone, or pregnenolone. The changes in the adrenal gland and plasma concentration of steroids were thought to reflect inhibited metabolism of deoxycorticosterone to corticosterone through inhibition of CYP11B1, and compensatory reaction for the inhibition. The compensatory reaction was thought to have masked decrease of corticosterone. These results suggest that simultaneous measurement of multiple steroids can enable sensitive evaluation of drug effects on adrenal steroidogenesis in vivo, while providing insight into the underlying mechanism of the effect. [ABSTRACT FROM AUTHOR]

TM-2 known as a potential antitumor drug is a novel semi-synthetic taxane derivative. As drug–protein interactions contribute to insights into pharmacokinetic and pharmacodynamic properties, we elucidated the binding of TM-2 to plasma protein. In this study, a simple, rapid and reliable method was developed and validated employing equilibrium dialysis for the separation of bound and unbound drugs and ultra-performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) for the quantitation. Protein binding reached equilibrium within 24 h of incubation at 37 °C. After liquid–liquid extraction with methyl tert-butyl ether, the samples were separated on Thermo Syncronis UPLC ® C 18 (2.1 mm×50 mm, 1.7 µm), and acquisition of mass spectrometric data was performed in multiple reaction monitoring (MRM) mode via positive electrospray ionization. The assay was linear over the concentration rang of 5–2000 ng/mL. The intra- and inter-day precisions were 0.1%–14.8%, and the accuracy was from −6.4% to 7.0%. This assay has been successfully applied to a protein binding study of TM-2 in rat, human and beagle dog plasma. TM-2 showed high protein binding of 81.4%±6.5% (rat), 87.9%±3.6% (human) and 79.4%±4.0% (beagle dog). The results revealed that there was an insignificant difference among the three species. [ABSTRACT FROM AUTHOR]

3- O -Acetyloleanolic acid (OAA) is a triterpenoid compound, and exerts an apoptosis in cancer cell lines, an inhibition of both atopic and allergic contact dermatitis in murine model, and a suppression of inflammatory bone loss in mice. OAA can be converted into oleanolic acid (OA) by hydrolysis in vivo , and OA exhibits several pharmacological effects as well. A liquid chromatographic method using tandem mass spectrometry (MS/MS) was developed for the simultaneous determination of OAA and OA in rat plasma. After liquid–liquid extraction with ethylacetate, both substances were chromatographed on a reversed phase column with a mobile phase of 0.1% formic acid aqueous solution and acetonitrile (1:9, v/v). The accuracy and precision of the assay were in accordance with FDA regulations for the validation of bioanalytical methods. This analytical method was successfully applied to monitor plasma concentrations of both substances over time following an intravenous administration of OAA in rats. [ABSTRACT FROM AUTHOR]

A sensitive and accurate liquid chromatography–tandem mass spectrometry (LC–MS/MS) method has been developed and validated for determination of tulobuterol in rat plasma for the first time. Plasma samples were extracted by liquid–liquid extraction method with methyl tert -butyl ether and the analyte and clenbuterol (IS) were separated on a Venusil MP C 18 column (100 mm × 2.1 mm, 3 μm) using 0.1% formic acid–water–methanol as mobile phase, with a runtime of 5 min. The analyte was detected in multiple reaction monitoring (MRM) mode with positive electrospray ionization. Transitions of m / z 228.2 → 154.0 for tulobuterol and m / z 277.1 → 203.0 for the clenbuterol were monitored. The linear range was 0.5–100 ng/ml ( r = 0.9967) for tulobuterol with the lower limit of quantitation of 0.5 ng/ml. The intra-day and inter-day precisions were less than 10.3% for the analyte and the accuracy was less than −8.6%. The RSD of matrix effect and recovery yield were within ±15% of nominal concentrations and tulobuterol was stable during stability studies. The validated method has been successfully applied to a pharmacokinetic study of three doses of tulobuterol patch in rats for the first time. [ABSTRACT FROM AUTHOR]