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Hederasaponin C (HSC), one of the main components of Pulsatilla chinensis, is considered as a potential drug for the treatment of inflammatory bowel disease. In the present research, we developed a pharmacokinetics-pharmacodynamics model to describe the concentration-effect course of drug action following the intraperitoneal injection of HSC in colitis rats. A sensitive UPLC-MS/MS method was established for the the determination of HSC in rat plasma to explore the pharmacokinetics properties. The separation was performed on an Accucore C 18 column (2.1 × 100 mm, 2.6 μm) with a flow phase consisting of acetonitrile and 0.1% formic acid water. The assay method was validated and demonstrated good adaptability for application in the pharmacokinetics study. Then the levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-1β (IL-1β) in colon tissues were measured using an ELISA assay. The levels of TNF-α, IL-1β and IL-6 were decreased after HSC administration, suggesting that HSC can significantly improve the level of inflammatory syndrome factor. The pharmacokinetics study showed that the time to peak concentration of HSC was 1 h. The concentration-effect curves showed a hysteresis loop. There was also a hysteresis between the peaked concentration and the maximum effect of HSC. The present study established in vivo pharmacokinetics-pharmacodynamics models and the results showed a great potential of HSC for treating ulcerative colitis., (© 2022 John Wiley & Sons Ltd.)

Luliconazole (LCZ) is a novel antifungal imidazole with broad-spectrum and high susceptibility of Aspergillus and Fusarium are the dominant species of fungal keratitis, may potentially be a new medical treatment option for ocular fungal infection. To evaluate LCZ distribution in ocular tissues after topical application for the development of ophthalmic delivery system, it is important to have a bioanalytical method for measuring the drug concentrations in different ocular tissues and aqueous humor (AH). A selective and sensitive ultrahigh performance liquid chromatography coupled with tandem mass spectrometry (UPLC-MS/MS) method was developed for the quantification of LCZ in rabbit ocular tissues, including conjunctiva, cornea, AH, iris, lens, vitreous humor (VH), retinal choroid and sclera, using lanoconazole as internal standard (IS). Chromatographic separation was achieved on a Xterra MS, C18 column (2.1 × 50 mm, 3.5 μm) using mobile phase with formic acid solution (0.2%, v/v): acetonitrile (50:50, v/v) at a flow rate of 0.2 ml/min, and the run time was 2.5 min. Detection was performed using the transitions 354.1 → 150.3 m/z for LCZ and 320.1 → 150.3 m/z for IS by positive ion electrospray ionization in multiple reaction monitoring (MRM) mode. Method validation was conducted in accordance with U.S. Food and Drug Administration's regulatory guidelines for bioanalytical method validation. The calibration curves were linear over the concentration range from 2.80 ng/ml to 2038 ng/ml for conjunctiva, cornea and sclera, 2.09 ng/ml to 1019 ng/ml for AH, 2.09 ng/ml to 509.5 ng/ml for iris, 2.09 ng/ml to 203.8 ng/ml for retinal choroid and VH, 2.04 ng/ml to 101.9 ng/ml for lens, with all the squared correlation coefficients (r 2 ) more than 0.99. The accuracy of the method was within the acceptable limit of 89.34%∼112.78% at the lower limit of quantification and other concentrations, Inter-day and intra-day precision values, expressed in terms of RSD (%), in all tissues were within 15% at all concentrations. The mean recoveries of LCZ in rabbit ocular tissues was 84.85%∼100.52%. No interference was found due to matrix components. Luliconazole was stable during the stability studies, including autosampler stability, benchtop stability, freeze/thaw stability and long-term stability. The method was successfully applied to the ocular pharmacokinetic and tissues distribution studies of LCZ in rabbit after topical administration of LCZ ophthalmic drug delivery system., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Contezolid is a novel oxazolidinone antibiotic with good antibacterial activity against gram-positive bacteria including methicillin-resistant Staphylococcus aureus. For the purpose to further characterize the pharmacokinetics of contezolid and its major metabolite M2, accurate and rapid ultra-performance liquid chromatography-tandem mass spectrometric assays (UPLC-MS/MS) were developed and validated for simultaneous quantification of contezolid and M2 in human plasma and urine. The plasma samples were pretreated by liquid-liquid extraction. The automated solid phase extraction method was used to preprocess urine samples. ACQUITY UPLC® BEH C8 (2.1 mm × 100 mm, 1.7 µm) column was used to separate the analytes with a gradient mobile phase of acetonitrile and water at a flow rate of 0.4 mL/min. The calibration curves showed good linearity over the concentration ranges of 0.0100-5.00 µg/mL for contezolid in plasma and urine, 0.00200-1.00 µg/mL in plasma and 0.0200-10.0 µg/mL in urine for M2, respectively. For both plasma and urine assays, the intra- and inter-batch accuracy and precision were within 15% for all quality control levels, including the lower limit of quantitation. The methods were fully validated and successfully applied to a pharmacokinetic study of contezolid tablets in subjects with moderate hepatic impairment., (Copyright © 2022. Published by Elsevier B.V.)

Lenvatinib is a multi-targeted tyrosine kinase inhibitor that inhibits tumor angiogenesis, but hypertension is the most common adverse reaction. Telmisartan is an angiotensin receptor blocker used to treat hypertension. In this study, a simple ultra-performance liquid chromatography-tandem mass spectrometry method was developed for the simultaneous determination of lenvatinib and telmisartan, and it was applied to the pharmacokinetic drug interaction study. Plasma samples were treated with acetonitrile to precipitate protein. Water (containing 5 mM of ammonium acetate and 0.1% formic acid) and acetonitrile (0.1% formic acid) were used as the mobile phases to separate the analytes with gradient elution using a column XSelect HSS T3 (2.1 mm × 100 mm, 2.5 μm). Multiple reaction monitoring in the positive ion mode was used for quantification. The method was validated and the precision, accuracy, matrix effect, recovery, and stability of this method were reasonable. The determination of analytes was not interfered with by other substances in the blank plasma, and the calibration curves of lenvatinib and telmisartan were linear within the range of 0.2-1000 ng/mL and 0.1-500 ng/mL, respectively. The results indicate that lenvatinib decreased the systemic exposure of telmisartan. Potential drug interactions were observed between lenvatinib and telmisartan.

A sensitive and accurate hydrophilic interaction liquid chromatography - tandem mass spectrometry method (HILIC-MS/MS) was developed and validated for the determination of phenylephrine concentration in Dried Blood Spot (DBS) samples from preterm infants, after ocular administration of an ophthalmic solution with phenylephrine. Sample preparation involved the extraction of the analyte from an 85 μL DBS sample with methanol - acetonitrile (50:50, v/v). Chromatographic separation was achieved on an ACQUITY UPLC BEH AMIDE column, under isocratic conditions within a 5 min run. Detection was achieved with a triple quadrupole MS applying electrospray ionization in positive mode. The method was fully validated and proved precise and accurate with in a linear range of 0.59-3.53 ng/ml in blood. The method was developed to provide insights on the level of exposure of infant population to phenylephrine after ocular administration., (Copyright © 2022 Elsevier B.V. All rights reserved.)

A sensitive assay was developed to evaluate inhibitory effects of aqueous solution on acetylcholinesterase (AChE) activity via measuring hydrolysis rates of acetylcholine (ACh) based on ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). Upon having identified precursor ions and product ions of the ACh and its hydrolysis products choline (Ch), the separation chromatogram for these two analytes has been established using a 50 mm reverse-phase BEH Shield RP18 column. The total chromatographic separation time is 7 min; limits of detection (LODs) for ACh and Ch are 0.14 µg L -1 and 0.12 µg L -1 , respectively. A simple method for inactivation of AChE and optimization of operational parameters were then sequentially performed. It was found that adjusting solution pH to 2.5 not only can terminate the enzymatic reaction but also solve band shifting and broadening caused by aqueous matrices in chromatographic separation during UPLC-MS/MS detection. Under conditions of 0.00075 U mL -1 AChE, initial concentration of ACh at 100 µg L -1 and 20 min observation time, IC 50 values of the proposed assay for chlorpyrifos-oxon, diazoxon, malaoxon, methidathion oxon, omethoate and paraoxon were 3.5 nM, 16.8 nM, 2.4 nM, 6.8 nM, 270 nM and 36.9 nM, respectively. They are 4.5-51.9 times smaller than those reported in a LC-MS based method, and >120 times lower than those obtained by the traditional Ellman method. The results suggested that, the proposed assay significantly increases the sensitivity of commercial AChE. In addition, inhibition efficiencies of three surface waters, a groundwater and four commercial brands of bottled drinking water samples on AChE activity were firstly measured using this UPLC-MS/MS based method. These water samples were proved to have different inhibitory effects on AChE activity, and the inhibition efficiencies dependent on concentrations of dissolved organic carbon (DOC) but are independent of UV absorbance at 254 nm (UV 254 ) values. These results indicate that the proposed method has advantages of high sensitivity over all other conventional methods. It may become a promising AChE inhibition assay for assessing toxicity of aqueous solution containing neurotoxicity contaminants such as organophosphorus pesticides (OPPs) at low levels, or used to evaluate potential inhibition effects of natural waters on AChE activity., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Pexidartinib is the first drug approved by the U.S. Food and Drug Administration specifically to treat the rare joint tumor tenosynovial giant cell tumor. In the current study, a validated, selective, and sensitive UPLC-MS/MS assay was developed for the quantitative determination of pexidartinib in plasma samples using gifitinib as an internal standard (IS). Pexidartinib and IS were extracted by liquid-liquid extraction using methyl tert-butyl ether and separated on an acquity BEH C 18 column kept at 40 °C using a mobile phase of 0.1% formic acid in acetonitrile: 0.1% formic acid in de-ionized water (70:30). The flow rate was 0.25 mL/min. Multiple reaction monitoring (MRM) was operated in electrospray (ESI)-positive mode at the ion transition of 418.06 > 165.0 for the analyte and 447.09 > 128.0 for the IS. FDA guidance for bioanalytical method validation was followed in method validation. The linearity of the established UPLC-MS/MS assay ranged from 0.5 to 1000 ng/mL with r > 0.999 with a limit of quantitation of 0.5 ng/mL. Moreover, the metabolic stability of pexidartinib in liver microsomes was estimated.

Aim: We aimed to establish and validate a simple and sensitive UPLC-MS/MS method for the determination of UNC1999, a dual inhibitor against EZH1 and EZH2 in plasma samples. Materials & methods: UNC1999 in rat plasma was processed with protein precipitation method and then separated on a C 18 column and detected under positive ionization mode. The method presented good linearity over the range of 1.0-2000 ng/ml with good accuracy and precision. UNC1999 was absorbed slowly and achieved a maximum concentration of 118.8 ± 12.0 ng/ml 1.5 h after oral administration. Conclusion: The method provides a favorable character in selectivity, linearity, accuracy, precision, recovery, matrix effects and stabilities and was suitable for describing the pharmacokinetic profile of UNC1999.

Upadacitinib, as a selective and reversible Janus kinase (JAK) inhibitor, has been widely used in the treatment of atopic dermatitis, ulcerative colitis and other inflammatory bowel diseases and other immune-mediated diseases. The combination of methotrexate and upadacitinib is a common clinical treatment strategy for rheumatoid arthritis (RA) in recent years. In this study, we established an ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) assay for quantitative measurement of upadacitinib and methotrexate, by which we successfully determined pharmacokinetic parameters of them in rat plasma. In order to pretreat the samples, we used acetonitrile as the precipitant, and for the internal standard (IS), we chose tofacitinib. The Acquity BEHC18 (2.1 mm × 50 mm, 1.7 μm) column, with acetonitrile and 0.1% formic acid aqueous solution composed mobile phases, was used to separate upadacitinib, methotrexate and tofacitinib. A Xevo TQ-S triple quadrupole tandem mass spectrometer was used as the detecting instrument in the positive ion mode. For upadacitinib, excellent linearity was shown of this assay in the calibration range with 0.1-200 ng/mL, and as for methotrexate, the range was 0.05-100 ng/mL. As the results indicated, the lower limit of quantification (LLOQ) was respectively 0.1 and 0.05 ng/mL for upadacitinib and methotrexate, the intra- and inter-day precision were ≤ 13.3%, and the accuracy of all the analytes ranged from -4.1% to 12.7%. The recovery of each analyte was > 80.2% in this experiment, and matrix effects we observed were unobvious. The establishment of this method and its successful application in rat plasma can provide a theoretical and technical support for the deeper study of pharmacodynamics and the clinical medication strategies., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Osmundacetone is a potential medicinal substance existing in ferns and has excellent antioxidant effects. This research aims to obtain the pharmacokinetic data for and metabolite products of osmundacetone. An UPLC-MS/MS quantitative method was established for the measurement of osmundacetonein in rat plasma over a linear range of 6.72-860.00 ng/ml. The signal to noise ratio of the lower limit of quantification was 60:1, the precision was <9.74% and the method had good selectivity and stability. The established method was successfully applied to the pharmacokinetic study of osmundacetone for the first time. Osmundacetone reached a peak at 0.25 h with a maximum value of 3283.33 μg/L. The apparent volume of distribution not multiplied by the bioavailability was 127.96 L/kg, and the half-life of osmundacetone was 5.20 h. At the same time, an UPLC-QE-Orbitrap-HRMS method was established to identify metabolites in plasma, urine and feces for the first time. A total of 30 metabolites were identified and the metabolic profile of osmundacetone was defined. In general, we have established a mass spectrometry quantitative method for osmundacetone for the first time and characterized its metabolic characteristics in rats., (© 2021 John Wiley & Sons, Ltd.)

Pyrotinib is an irreversible EGFR/HER2 inhibitor that has been approved for the treatment of breast cancer. The aim of this work was to establish a quantification method for the simultaneous determination of pyrotinib and its metabolite pyrotinib-lactam in rat plasma using UPLC-MS/MS. After simple protein precipitation with acetonitrile, the analytes and internal standard (neratinib) were separated on an ACQUITY BEH C 18 column (2.1 × 50 mm, 1.7 μm) using a mobile phase of water containing 0.1% formic acid and acetonitrile. The detection was performed using selected reaction monitoring mode with precursor-to-product ion transitions at m/z 583.2 > 138.1 for pyrotinib, m/z 597.2 > 152.1 for pyrotinib-lactam, and m/z 557.2 > 112.1 for internal standard. The assay exhibited excellent linearity in the concentration range of 0.5-1000 ng/mL for pyrotinib and pyrotinib-lactam. The assay met the criteria of the United States Food and Drug Administration-validated bioanalytical methods and was successfully applied to a pharmacokinetic study of pyrotinib and its metabolite for the first time. Our results demonstrated that pyrotinib rapidly converted into pyrotinib-lactam, whose in vivo exposure was 21% that of pyrotinib., (© 2021 John Wiley & Sons, Ltd.)

Background: Accurately measuring plasma aldosterone concentration is difficult but meaningful for primary aldosteronism (PA) diagnosis., Methods: In this study, we developed an ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method for plasma aldosterone detection, evaluated its performance according to guidelines issued by CLSI, including detection limit, linearity, precision, and compared it with chemiluminescence immunoassay. Then, a reference range of plasma aldosterone in young people was established by using this method., Results: The lower limit of quantitation (LOQ) was 10 pg/ml. The mean recovery rates of analyte added to serum were 100.07-102.05% in different concentrations. The linearity range was 20-2000 pg/ml. Inter-assay CVs were 2.20-3.97% at aldosterone concentrations of 65.66-854.75 pg/ml. The regression equation of UPLC-MS/MS (x) and chemiluminescence immunoassay (y) was y = 1.002x + 65.854 (r = 0.9456, n = 237). The reference range of plasma aldosterone detected by UPLC-MS/MS was 11.30-363.82 pg/ml in young people in South China, and there was no statistically significant difference in plasma aldosterone concentration between two genders., Conclusion: In conclusion, UPLC-MS/MS can rapidly and accurately detect plasma aldosterone and is appropriate for clinical application., (© 2021 The Authors. Journal of Clinical Laboratory Analysis published by Wiley Periodicals LLC.)

3-n-Butylphthalide (NBP) is a potent drug for the treatment of ischemic stroke. The aim of this study was to develop a simple and sensitive ultra-high-performance liquid chromatography-tandem mass spectrometric (UPLC-MS/MS) method for the simultaneous determination of NBP and its circulating metabolite 10-hydroxy-NBP in rat plasma using senkyunolide I as the internal standard (IS). The analytes and IS were extracted from the plasma by ethyl acetate-ethyl ether (1:5, v/v) and then separated on an ACQUITY BEH C 18 column (2.1 × 50 mm, 1.7 μm). The mobile phase consisted of water containing 0.1% formic acid and acetonitrile containing 0.1% formic acid, which was delivered at a flow rate of 0.3 mL/min with gradient elution. MS detection was achieved under selective reaction monitoring mode with precursor-to-product transitions at m/z 191.1 > 145.1 for NBP, m/z 207.1 > 171.1 for 10-hydroxy-NBP and m/z 207.1 > 161.1 for IS, respectively. The assay showed excellent linearity over the concentration range of 0.5-1000 ng/mL for both analytes, with correlation coefficient greater than 0.998. The other validation parameters were all within the required limits. The validated UPLC-MS/MS method has been further applied to the pharmacokinetic study of NBP and 10-hydroxy-NBP in rats after they were orally administered with NBP (30 mg/kg)., (© 2021 John Wiley & Sons, Ltd.)

Isothiazolinones, a family of biocides, are used as preservatives for their fungicidal, bactericidal, and algacidal properties. These compounds can be found in a wide range of consumer and building products, such as paints, varnishes, shampoos, and liquid detergents. A robust analytical UPLC-MS/MS method to identify and quantify seven isothiazolinones (MIT, CMI, BIT, MBIT, BBIT, OIT, and DCOIT) in consumer products and their emissions was developed and validated according to ISO/IEC 17025. The limits of detection (LODs) ranged from 0.14 μg L -1 (BIT) to 0.54 μg L -1 (CMI). The method was applied for the quantification of the seven isothiazolinones in four types of consumer products (i.e., cosmetics, air fresheners, cleaning products, and building products) and the indoor emissions from a paint. Matrix effects were observed for the shampoo (63-74%), the shower gel (67-84%), and the ceramic glass cleaner (53-57%). All isothiazolinones indicated by the manufacturer (i.e., MIT, CMI, BIT, OIT, and DCOIT) were detected in the products and successfully quantified by the UPLC-MS/MS method., (© 2021. Springer-Verlag GmbH Germany, part of Springer Nature.)

Our previously reported, first validated, UPLC-MS/MS-based simultaneous analysis of five human milk B-vitamins revealed severe matrix effects. High levels of endogenous lactose fouled the electrospray ionization source affecting the analysis. We evaluated solid-phase extraction (SPE), liquid-solid extraction (LSE), protein precipitation (PPT), and liquid chromatography effluent diversion for lactose-removal. SPE failed to separate lactose from vitamins; LSE using 2-propanol reduced lactose and vitamin recoveries. PPT-solvent, milk volume, and reconstitution solvent influenced flavin adenine dinucleotide, pyridoxal and nicotinamide recoveries. Using an optimized LC-gradient enabled chromatographic separation of lactose from vitamins and its removal using a post-column switch-valve. Only 40 µL milk was subjected to methanol-PPT and non-polar matrix removal by methyl tert-butyl ether. B-vitamin recoveries were established (81.9-118.6%; CV ≤ 11.9%; precision: 4.9-13.7%) with greatly reduced matrix effects, and improved process efficiency, and recovery., (Copyright © 2021 The Authors. Published by Elsevier B.V. All rights reserved.)

A simple, selective, rapid, and reliable ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was developed and validated to determine CX3002 in human plasma using CX3002-d3 as the internal standard (IS). After a rapid protein precipitation with acetonitrile (3:1, v/v), the chromatographic separation of CX3002 and IS was performed on a Thermo Hypersil GOLD C18 column (2.1 mm × 50 mm, 1.9 μm) with gradient elution at a flow rate of 0.4 ml/min. Gradient elution was achieved with mobile phase A consisting of water containing 0.1% formic acid and 5 mmol/L ammonium formate and mobile phase B consisting of methanol containing 0.1% formic acid. The detection was performed on AB SCIEX QTRAP® 5500 tandem mass spectrometry in the positive ion mode. Multiple reactions monitoring (MRM) was used for quantitative analysis at transition of m/z 460.3 → 199.3 for CX3002 and m/z 463.3 → 202.3 m/z for IS. The method was fully validated and displayed good linearity over a concentration range of 0.2-400 ng/mL with the correlation coefficient above 0.997. The intra-run and inter-run precision (coefficient of variation, CV) ranged from 0.60%-16.46% and the accuracy bias ranged from -7.09%-9.75%. The mean IS-normalized extraction recovery ranged from 98.30% to 104.52%. The CV(%) of IS-normalized matrix factors at the low and high QC concentration were 4.09% and 1.68%, respectively. The storage stability under different conditions was in accordance with the bioanalytical guidelines. The method was successfully applied to the pharmacokinetic study of CX3002 (30 mg) in healthy Chinese subjects., (Copyright © 2021. Published by Elsevier B.V.)

Background: Upadacitinib, a novel selective Janus kinase 1 (JAK1) inhibitor, has been recently approved by the US FDA for the treatment of adult patients with moderately to severely active rheumatoid arthritis (RA). An ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method for the quantitative analysis of upadacitinib in beagle dog plasma was developed and validated., Methods: Upadacitinib and fedratinib (internal standard, IS) were extracted with ethyl acetate under alkaline condition and then separated and detected. The chromatographic column was Waters Acquity UPLC BEH C18 column (2.1 mm × 50 mm, 1.7 μm), the mobile phase was acetonitrile and 0.1% formic acid in water with gradient elution procedure, and the flow rate was 0.40 mL/min. Under the positive ion mode, upadacitinib and IS were monitored by multiple reaction monitoring (MRM) as the following mass transition pairs: m/z 447.00 → 361.94 for upadacitinib and m/z 529.82 → 141.01 for IS., Results: In the concentration range of 1-500 ng/mL, upadacitinib had good linearity, and the lower limit of quantification (LLOQ) was 1 ng/mL. The RSD of the intra- and inter-day precision was less than 10.03%, and the RE of accuracy was -3.79% to 2.58%. The extraction recovery of upadacitinib was more than 80%, the matrix effect was around 100%, and upadacitinib was found to be stable., Conclusion: The novel optimized UPLC-MS/MS assay was an effective tool for the determination of upadacitinib and had been successfully applied to the pharmacokinetic study of upadacitinib in beagle dogs, and this method would also be used to study DDIs., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2021 Wang et al.)

Qixue-Shuangbu Prescription (QSP) is an efficacious prescription for treating heart failure, myocardial ischemia and other diseases. It is composed of nine Chinese herbs. This study investigated and compared the pharmacokinetics of QSP in rats by UPLC-MS/MS between two dosage forms of traditional decoction (TD) and compound tincture (CT). Owing to the complexity of the chemicals in QSP, ginsenoside Rg1, ginsenoside Re, ferulic acid, astragaloside IV, rhein and calycosin were chosen for the pharmacokinetics study. The method established for detecting serum specimens was shown to have acceptable selectivity, linearity, lower limit of quantitation, precision, accuracy, recovery, matrix effect and stability. The peak concentration, AUC 0-t and AUC 0-∞ of ginsenoside Re, ginsenoside Rg1, ferulic acid and rhein were significantly increased after oral administration of CT (P < 0.05), the half-life of ferulic acid in the CT group was lower than that in the TD group (P < 0.05) and the half-life and AUC 0-∞ of astragaloside IV in the CT group were significantly increased (P < 0.05), which revealed that wine-processing could influence the bioavailability and the elimination of these compounds. For better clinical efficacy, we suggest that the CT dosage form of QSP should be selected., (© 2021 John Wiley & Sons, Ltd.)

Polygonum orientale L. is a traditional Chinese medicine having extensive pharmacological activities including antimyocardial ischemia (MI) injury properties. Isoorientin, orientin, vitexin, quercitrin, astragalin and protocatechuic acid are the main compounds in P. orientale extract. The aim of this study was to establish an ultra-performance liquid chromatography-tandem mass spectrometry method for the determination of the content of these compounds in urine, feces and bile samples simultaneously and application of the method in a comparative excretion study in normal and MI model rats after oral administration of P. orientale extract. Chromatographic seperation was conducted on an Agilent Eclipse Plus C 18 column with the mobile phase consisting of 0.1% formic acid-acetonitrile and 0.1% formic acid-water. Negative ion multiple reaction monitoring mode was used for quantification. The six compounds had good linearity (r ≥ 0.9921) and acceptable accuracy ranging from 10.10 to -5.82% The relative standard deviations of within-day precision and inter-day precision were <10.45 and 13.44%, respectively. The extraction recovery of the six analytes ranged from 80.31 to 101.47% and the matrix effect was 82.56-102.88%, indicating that the preparations of sample collected form urine, feces and bile were stable throughout analysis. The excretion amount of the six analytes increased in both normal and MI model rats' urine, feces and bile in a 24 h period and became stable between 36 and 48 h after administration. The total excretion rate of six compounds was <5% in urine, feces and bile of normal and MI model rats. The excretion peak period for all compounds in MI rats was slower than that in normal rats. This excretion study provides insights for further application and research on P. orientale., (© 2021 John Wiley & Sons, Ltd.)

Given the key role of bees as indicators for environmental assessment, residues in bees and bee products have attracted great interest. In this regard, an improved, highly sensitive method for quantifying the insecticide pyriproxyfen and its four metabolites (4'-OH-Pyr, DPH-Pyr, 2-OH-PY, 4'-OH-POP) in honeybees, larvae, and bee products (honey, pollen, royal jelly and wax) should be established. For this purpose, we used ultra-performance liquid chromatography coupled with triple-quadrupole mass spectrometry for rapid quantification (≤5 min). Recoveries for various matrices ranged from 73.77% to 114.97%, with satisfactory intra-day and inter-day precision (relative standard deviations of 0.03-8.61% and 0.10-7.25%, respectively). The results demonstrated excellent linearity (R 2 > 0.9903) with a limit of quantification of 1 μg/kg for six different matrices. We collected and analyzed 597 samples (honey, bees and wax) from four major beekeeping areas in China. Only 47 of these samples contained residues of pyriproxyfen and two of its metabolites (2-OH-PY, 4'-OH-Pyr), and high levels of contamination were found in bee samples (2-739 μg/kg), with substantive accumulation in wax (levels were 9.49% higher than in other samples). The result demonstrate that the method provides a reliable and convenient means of monitoring pyriproxyfen and its metabolites in bee products for better product quality, human health, and international commercial competition and also lays a foundation for risk assessment of potential pyriproxyfen contamination in China., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

A UPLC-MS/MS method was developed to determine the levels of five traditional antipsychotics (APs) (chlorprothixene, perphenazine, fluphenazine, thioridazine, and promethazine) in human plasma with carbamazepine as the internal standard. Samples were extracted using simple liquid-liquid extraction (ethyl acetate/methyl tert-butyl ether, 2:3 v/v); then the analytes were subjected to gradient elution chromatography with a mobile phase composed of 0.1% formic acid in water and acetonitrile. The analytes were separated using a Waters XBridge BEH C 18 column (100 × 2.1 mm, 2.5 μm). The linear ranges of chlorprothixene, perphenazine, fluphenazine, thioridazine, and promethazine are 2-250 ng/mL, r > 0.995. The limit of quantitation is 2 ng/mL, and the limit of detection is in the range of 0.1-0.5 ng/mL. The inter-day and intra-day relative standard deviations are less than 10%, and the relative errors are in the range of -5.70 to 7.20%. The recoveries of the five drugs are in the range of 70-109%. The results of methodology verification indicate that this method is simple, economical, sensitive, and suitable for the simultaneous quantification of five traditional APs in human plasma., (© 2021 John Wiley & Sons, Ltd.)

The determination of psychotropic drugs and metabolites in blood is relevant in the context of both therapeutic drug monitoring and clinical and forensic toxicology. LC-MS/MS is the preferred method for these assays. However, LC-MS/MS is particularly susceptible to matrix ionization effects and appropriate sample preparation is required to minimize these effects. In this study, a simple, single-step, mini-QuEchERS extraction procedure, coupled to UPLC-MS/MS, was developed and validated for the determination of 15 toxicologically relevant compounds in whole blood, including psychoactive drugs and some metabolites. The assay was linear in the range of 25-1,000 ng ml -1 , fulfilling criteria for accuracy and precision. Extraction yields (71.9-87.7%) and matrix effects (-3.3 to +4.4%, with the exception of codeine, which had matrix effects of -35.36 to -28.14%) were acceptable for the majority of the evaluated compounds, using a single internal standard. The assay was applied to 238 clinical specimens from patients admitted to an emergency service, with 22 samples presenting quantifiable concentrations of 11 different compounds. The developed assay is a simple and efficient strategy for determination of target psychotropic drugs and metabolites in forensic and clinical toxicology., (© 2021 John Wiley & Sons, Ltd.)

In this study, we used ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) to measure the concentration of narciclasine and 7-deoxynarciclasine in mouse blood after intravenous (i.v.) and oral administration (p.o.), and we used this method to investigate their pharmacokinetics profiles in mice. Chromatographic separation of the analytes was achieved using a UPLC HSS T3 column (2.1 mm × 100 mm, 1.8 μm) with a mobile phase consisting of acetonitrile-water (0.1% formic acid) by gradient elution. Electrospray ionization (ESI positive-ion mode)-tandem mass spectrometry in multiple reaction monitoring (MRM) mode was employed for quantitative analysis of the analytes in mouse blood samples. Twelve mice were administered narciclasine and 7-deoxynarciclasine (2 mg/kg) intravenously (iv), while the other twelve mice were administered narciclasine and 7-deoxynarciclasine (10 mg/kg) orally. The mouse blood was withdrawn from the caudal vein to be processed, after which the blood was analyzed by UPLC-MS/MS, and the corresponding data were fitted using the Drug and Statistics (DAS) software. Standard curves of narciclasine and 7-deoxynarciclasine were generated over the concentration range of 5-5000 ng/mL. The intra-day accuracy of narciclasine and 7-deoxynarciclasine was 90-105%, and the corresponding inter-day accuracy was 87-108%. The intra-day precision was less than 13%, while the inter-day precision was less than 14%. Matrix effects were also observed (between 94% and 104%), and the recovery calculated was higher than 70%. The developed and validated UPLC-MS/MS method was then successfully applied in determining the mouse pharmacokinetics of narciclasine and 7-deoxynarciclasine. From this, thebioavailabilityofnarciclasine and 7-deoxynarciclasinewasdetermined to be 10.3%and35.4%, respectively., (Copyright © 2021 Elsevier B.V. All rights reserved.)

A simple, sensitive, rapid and specific method based on ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) for the simultaneous quantification of dantrolene (DAN) and paracetamol (PAR) in real human plasma was developed and validated. The preparation of sample was achieved by liquid-liquid extraction with tertiary butyl methyl ether. The analysis was performed on a reversed-phase C 18 column (1.7 µm, 2.1 × 30 mm) using acetonitrile: 0.1% formic acid (80:20, v/v) as the mobile phase and pumped in an isocratic mode at a flow rate of 0.3 mL/min using citalopram (CIT) as an internal standard. Tandem mass spectrometric detection was carried out by both positive and negative electrospray ionization (ESI) in the multiple-reaction monitoring mode (MRM). The analysis was carried out within 1 min for each sample which made it possible to analyze more than 350 human samples per day. Validation of the method was performed according to FDA guidelines for bio-analytical method. The method was found to be linear in the range of 25-2500 ng/mL and 100-10,000 ng/mL for DAN and PAR, respectively. The method was applied successfully for the determination of the two analytes in the plasma after oral administration of Dantrelax® compound capsules to healthy volunteers. The study was accomplished after approval of the ethics committee., (Copyright © 2021. Published by Elsevier B.V.)

Yunaconitine and indaconitine are active ingredients from the rhizomes of Aconitum plants. In this study, an ultra-high-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method was developed to measure the concentrations of the yunaconitine and indaconitine in mouse blood, and the method was applied in measuring the pharmacokinetics of the two alkaloids after oral and intravenous administration. A UPLC HSS T3 column (2.1 mm × 100 mm, 1.8 μm particle size) was used for chromatographic separation by gradient elution using acetonitrile-water (0.1% formic acid) as the mobile phase at a flow rate of 0.4 mL/min. Multiple reaction monitoring (MRM) mode and electrospray ionization (ESI) (positive-ion mode) were used to monitor the transitions of each analyte by tandem mass spectrometry for quantitative analysis. Yunaconitine and indaconitine were administered to the mice orally at 2 mg/kg and intravenously at 0.05 mg/kg. Blood was collected at various time intervals, and the blood samples were processed after collection and analyzed by UPLC-MS/MS. The standard curve generated for each analyte was linear over the concentration range of 0.5-500 ng/mL. The intra-day and inter-day accuracy of yunaconitine and indaconitine were 90%-103% and 86%-106%, respectively, and the precision (RSD, %) was less than 15% for both intra-day and inter-day measurements. The matrix effect ranged from 96% to 109%, and the recovery was higher than 72%. The UPLC-MS/MS method developed herein was successfully applied to measuring the pharmacokinetic parameters of yunaconitine and indaconitine in mice after intravenous and oral administration. The bioavailability of yunaconitine and indaconitine were 27.4% and 25.8%, respectively., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Codonopsis pilosula (CP) is a traditional Chinese medicine used to invigorate spleen, replenish lung, nourish blood and engender fluid. A rapid, selective and sensitive ultra-performance LC-tandem mass spectrometry method was developed and validated to determine lobetyolin in rat plasma. The calibration curve showed good linearity over a concentration range of 0.46-1000 ng/mL for lobetyolin. The extraction recovery ranged from 72.5% to 89.1% with matrix effects of 81.6%-107.8%. The intra- and inter-batch precision and accuracy were 0.02-14.4% and -13.9% to -1.36%, respectively. The method was successfully applied for the bioavailability study of lobetyolin in rats after oral administration of pure lobetyolin and CP extract. Results showed that the elimination half-time (t 1/2 ) and the area under the concentration-time curve from zero to infinity of lobetyolin in CP extract were statistically different from those of the pure monomer (P < 0.05). However, the time to reach the maximum plasma concentration (T max ) and the maximum concentration (C max ) showed no significant differences between the two treatments. Furthermore, the bioavailability of lobetyolin in the experimental group was only 3.90%, significantly lower than that of the CP extract group (6.97%). The low bioavailability indicated that this component may be absorbed poorly or metabolized extensively in rats. Our results will provide useful information for further preclinical studies and formulation preparation of lobetyolin., (© 2021 John Wiley & Sons, Ltd.)

Bisphenol compounds (BPs) are a group of environmental contaminants with endocrine-disrupting effects both for humans and animals. The present work developed a sensitive analytical method for the detection of multiple BPs in the animal feed based on ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) with post-column ammonium hydroxide (NH 4 OH) infusion. A modified QuEChERS method was incorporated into the extraction and purification processes. The limit of detection (LODs) and quantification (LOQs) for the target BPs were in the ranges of 0.02-0.75 μg kg -1 and 0.04-0.95 μg kg -1 , respectively. Average recoveries were ranged between 82.6% and 112%. The proposed method was successfully applied to determine the concentrations of BPs in 20 actual feed samples, and the preliminary profiles of BPs in products from local feed factories were obtained. Each sample was simultaneously contaminated with at least 2 to 4 BPs, and bisphenol A (BPA) was the dominant analog of BPs found in animal feed., (Copyright © 2021 Elsevier B.V. All rights reserved.)

With the purpose of carrying out therapeutic drug monitoring (TDM) of nine antiepileptic drugs simultaneously in the clinic, a UPLC-MS/MS assay method was developed and validated. Separation of antiepileptic drugs and internal standard (naproxen and diazepam) was performed on an Agilent Eclipse XDB-C 18 column (50.0 × 2.1 mm, 1.7 μm) using water and acetonitrile as mobile phase for gradient elution. Polarity switch ionization mode (positive-negative-positive) was performed in a total run time of 3.0 min. The method validation was conducted based on bioanalytical method validation guidance, including specificity, calibration curve, precision, accuracy, recovery, matrix effect, stability and dilution integrity, and all of the results satisfied the requirements. Consequently, the proposed method was applied to the TDM of nine antiepileptic drugs and was proved to be sensitive, reliable and specific to determine antiepileptic drugs in human plasma simultaneously. Meanwhile, the TDM results showed that the plasma drug concentration of many patients was not within the effective therapeutic range, especially those taking topiramate, oxcarbazepine and levetiracetam, which was of great guiding significance to rational drug use and timely adjustment of the treatment plan. Therefore, individualized therapy based on advanced analytical methods and TDM theory is of great practical significance., (© 2021 John Wiley & Sons, Ltd.)

A novel, simple and rapid UPLC-MS/MS method was developed and validated for determination of favipiravir (FAV) in human plasma. Lamivudine was used as an internal standard (IS). The Xevo TQD LC-MS/MS was operated under the multiple-reaction monitoring mode using electrospray ionization. Precipitation with acetonitrile was used in sample preparation as it gives relatively cleaner plasma samples. The prepared samples were chromatographed using an Acquity UPLC ® HSS C 18 (100 × 2.1 mm, 1.8 μm) column. The mobile phase was composed of ammonium formate and methanol in a gradient mode that was pumped at a flow rate of 0.35 ml/min. The developed method was validated as per the FDA guidelines and linearity was in the range of 0.25-16 μg/ml for FAV. The intra- and inter-day precision and accuracy results were within the acceptable limits. A run time of 4.5 min and a low quantification limit of FAV allowed the application of the developed method for the determination of FAV in a bioequivalence study in healthy human volunteers., (© 2021 John Wiley & Sons, Ltd.)

A sensitive and robust method has been developed using an ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) assay to quantify Tat-K13, a novel interfering peptide for the treatment of ischemic stroke, in human plasma. Automated solid-phase extraction on a Waters Oasis WCX (30 μm, 10 mg) 96-well plate was used to extract Tat-K13 from human plasma and the extracts were separated on a Waters Acquity CSH column (2.1 × 50 mm i.d., 1.7 μm) with a gradient elution method by mobile phase A (nonafluoropentanoic acid-acetic acid-water, 1:2:1000, v/v/v) and B (nonafluoropentanoic acid-acetic acid-water-acetonitrile, 1:2:100:900, v/v/v/v). The method was fully validated following international bioanalytical guidelines and showed good linearity from 2.10 to 1,050 ng/ml. The method was successfully applied to investigate the clinical pharmacokinetics of Tat-K13 in health volunteers. Rapid elimination of Tat-K13 from the body was observed, with half-life ranging from 0.26 to 0.78 h across different dose levels. The exposure of Tat-K13 was approximately dose-dependent in terms of the area under the concentration-time curve and peak concentration., (© 2021 John Wiley & Sons, Ltd.)

Enarodustat, a potent, orally bioavailable, selective inhibitor of hypoxia inducible factor-Prolyl hydroxylase (HIF-PH), has been approved recently in Japan for the treatment of anemia in patients with chronic kidney disease (CKD). To evaluate the pharmacokinetics of enarodustat, a bioanalytical assay in human plasma was needed for the quantitation of enarodustat for both healthy subjects and patients with CKD. The UPLC-MS/MS method for the quantitation of enarodustat was initially validated in a bioanalytical laboratory in Japan to support clinical studies conducted in Japan, and then was transferred and validated in a bioanalytical laboratory in United States to support clinical studies conducted here. A cross-validation was successfully performed between the two bioanalytical laboratories using both quality control (QC) samples and incurred study samples. Enarodustat was fortified with its isotopically labeled internal standard in a 25 µL plasma aliquot and extracted with protein precipitation. The chromatographic separation was achieved on an Acquity UPLC BEH C18 (1.7 µm, 2.1 × 50 mm) column with gradient elution. The calibration curve range for the assay was 1.00-500 ng/mL. Assay precision, accuracy, linearity, selectivity, sensitivity and analyte stability covering sample storage and analysis were established. No interferences were observed from medications that may be co-administered along with enarodustat. The validated UPLC-MS-MS method at the US bioanalytical laboratory has been successfully applied to eight clinical studies for the determination of enarodustat concentrations in human plasma for both healthy subjects and patients with CKD., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Background: An efficient, fast and sensitive ultra high-performance liquid chromatography-mass spectrometry (UPLC-MS/MS) method for simultaneous determination of celecoxib (CEL), dezocine (DEZ) and dexmedetomidine (DEX) in beagle plasma were established., Methods: The beagle dogs plasmawas precipitated by acetonitrile. The column was Acquity UPLC BEH C18 column and the mobile phase was acetonitrile-formic acid with gradient mode, and the flow rate was set at 0.4 mL/min. Under the positive ion mode, CEL, DEZ, DEX and Midazolam (internal standard, IS) were monitored by multiple reaction monitoring (MRM) as the following mass transition pairs: m/z 381.10→282.10 for CEL, m/z 246.20→147.00 for DEZ, m/z 201.10→94.90 for DEX, and m/z 326.10→291.10 for IS., Results: This UPLC-MS/MS method had good linearity for CEL, DEZ and DEX. The RSDs of inter-day and intra-day precision were the values of 0.31-7.66% and 0.11-9.63%, respectively; the RE values were from -6.05% to 10.98%. The extraction recovery was more than 79%, and the matrix effect was around 100%. The RSDs of stability were less than 8.96%. All of them met the acceptance standard of biological analysis method recommended by FDA., Conclusion: This UPLC-MS/MS method is an effective tool for the simultaneous determination of CEL, DEX and DEX, and has been successfully applied to the study of pharmacokinetics in beagle dogs., Competing Interests: The authors have declared that no competing interests exist., (© 2021 Hu et al.)

Tedizolid phosphate (TZP) a prodrug of Tedizolid (TDZ) is a novel oxazolidinone antibiotic, used for the treatment of acute bacterial skin, skin structure infections and other serious gram positive and MRSA infections. In the present study, a sensitive UPLC-MS/MS analytical method was developed and validated for the quantification of TDZ in rabbit's aqueous humor (AqH) by using linezolid as internal standard (IS). Both TDZ and IS were separated on an Acquity™ HILIC column using an isocratic mobile phase comprising of acetonitrile: 20 mM ammonium acetate (85:15, v/v), eluted at 0.3mLmin -1 flow rate with total run time of 3 min. The AqH samples were processed by protein precipitation method by using acetonitrile as precipitating agent. TDZ and IS were detected in positive mode using electrospray ionization source. The precursor to product ion transitions at m/z 371.15 to 343.17 for TDZ and m/z 338.18 to 296.22 for IS were used for the quantification in multiple reaction monitoring mode. The calibration curve was linear in the concentration range of 4.98-1000ngmL -1 and the lower limit of detection was 1.97ngmL -1 only. The method was validated following US-FDA-guidelines and the results of validation parameter were found within the set limits. The developed UPLC-MS/MS method was fast, sensitive and reliable for the quantification of TDZ in the rabbit AqH and was successfully employed for ocular pharmacokinetic study of TDZ in AqH after topical ocular application of TZP-containing formulations in rabbit eyes., (Copyright © 2021 Elsevier B.V. All rights reserved.)

A method was established for the simultaneous determination of 20 kinds of bile acids in human serum employing ultraperformance liquid chromatography-tandem mass spectrometry. Chromatographic conditions and sample preparation were optimized to achieve good separation and maximum sensitivity for these analytes. The linearity, accuracy and repeatability of the development method were validated with a series of experiments. Under the optimum conditions, good linearities were achieved in the quantitative range for each bile acid with the correlation coefficients (r 2 ) >0.9901. The limit of detections (signal-noise ratio 3) of the method were in a range from 0.02 to 0.57 nmol/L. The recoveries were in the range of 88.1-109.9%, RSD < 6.12%. This method was successfully applied for the determination of bile acids in a human serum sample with simple operation, high sensitivity and good accuracy, and provides a reference for the clinical determination of bile acid content., (© 2021 John Wiley & Sons, Ltd.)

Lots of studies showed the combination therapy of perindopril, indapamide and amlodipine could increase BP lowering efficacy and the benefits of high-risk patients. To evaluate potential pharmacokinetic interaction, a simultaneous UPLC-MS/MS quantification method of perindopril, perindoprilat and indapamide in human plasma was developed and validated. The plasma samples were prepared by solid phase extraction, and then separated on an X-terra MS C 18 (2.1 mm × 150 mm, 3.5 μm) with isocratic elution. The ion transitions at m/z 369.165 → 172.000 (perindopril), m/z 341.146 → 170.112 (perindoprilat), m/z 366.010 → 132.100 (indapamide), m/z 389.120 → 206.200 (S10211-1, IS1) and m/z 394.080 → 160.200 (S1641, IS2) were monitored under the positive ion mode of electrospray ionization with multiple reaction monitoring. This method exhibited great sensitivity, linearity, accuracy, and precision for the determination of perindopril, perindoprilat and indapamide over the range of 0.250-50.0 ng/mL. The average extraction recovery of perindopril, perindoprilat and indapamide samples at low, medium, and high concentration levels were between 85.9% and 93.6%, respectively. The stability of analytes over different storage and processing conditions in the whole study was also validated. The method is fast, accurate, sensitive and reproducible, which is suitable for the detection of the concentration of perindopril, perindoprilat and indapamide in human plasma., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Perillyl alcohol (POH) is a monocyclic terpene that has strong antitumor activity. Brain tumors are particularly difficult to treat with therapeutic agents, and clinical trials have shown their low tolerance through oral administration. We proposed the entrapment of POH into an oil-in-water chitosan nanoemulsion aiming its intranasal administration for brain targeting. An ultra-performance liquid chromatography/tandem mass spectrometry (UPLC-MS/MS) method was developed and validated for the quantitation of total metabolite perillic acid (PA) in plasma and brain of rats. The rat samples containing the metabolite were treated by liquid-liquid extraction with acetonitrile. The mobile phase was 0.1% formic acid in water (solvent A) and 0.1% formic acid in methanol (solvent B), at a flow rate of 0.3 mL min -1 in gradient elution. The chromatography was run for 10 min, and analytical curves were built in acetonitrile, plasma, and brain. The PA was detected in positive ion mode with multiple reaction monitoring. The method has shown high selectivity, sensitivity, and throughput. The low quantification limits of 162, 178, and 121 ng mL -1 for acetonitrile, brain, and plasma, respectively, indicate a good detectability of the method. The repeatability and precision observed were within the limits recommended in the literature. The accuracy of the method was verified through high recovery rates (106-118%). The validated method was successfully applied to the pharmacokinetic study of the metabolite PA after the intranasal administration of free or POH-loaded nanoemulsion in rats. The results showed that chitosan nanoemulsion improved the plasma and brain bioavailability of POH, representing a promising alternative to free POH treatment., (© 2020 John Wiley & Sons, Ltd.)

Asarinin, β-eudesmol, and wogonin have common antiangiogenic activities and have the potential for use in chemotherapy. Besides, they are multivalent substances that are combined in various herbal medicines. The purpose of this study was to develop a method for simultaneous analysis of asarinin, β-eudesmol, and wogonin, which are representative pharmacological components of Asarum heterotropoides, Atractylodes lancea, and Scutellaria baicalensis, respectively, in rat biosamples using ultraperformance liquid chromatography-tandem mass spectrometry. The three components were separated using 5 mm aqueous ammonium acetate containing 0.1% formic acid and acetonitrile as a mobile phase, equipped with a KINETEX core-shell C 18 column. The analysis was quantitated on a triple-quadrupole mass-spectrometer employing electrospray ionization, and operated in the multiple reaction monitoring mode. The chromatograms showed high resolution, sensitivity, and selectivity with no interference with plasma, urine, and feces constituents. The developed analytical method satisfied international guidance criteria and could be successfully applied to the pharmacokinetic (PK) studies evaluating oral bioavailability of asarinin, β-eudesmol, and wogonin after oral and intravenous administration and their urinary and fecal excretion ratios after oral administration to rats. Furthermore, the analysis was extended to PK studies following oral administration of Gumiganghwal-tang. This study was the first simultaneous analysis of the aforesaid three constituents in rat plasma, urine, and feces that also determined their PK parameters., (© 2020 John Wiley & Sons, Ltd.)

Marsdenia tenacissima (Roxb.) Wight et Arn. (M. tenacissima) is considered an anticancer medicine in traditional Chinese medicine, which is extensively used in clinical application since it has great therapeutic effects. Currently, although a number of articles have examined M. tenacissima in terms of its pharmacology and quality control, few have investigated the in vivo mechanism of M. tenacissima active ingredients. Previously, we have studied the pharmacokinetics of eight active ingredients after oral administration of M. tenacissima extracts in rat plasma. This study constructed a new scientific ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) approach to simultaneously quantify the contents of tenacissosides B, G, H and I, cryptochlorogenic acid, chlorogenic acid, neochlorogenic acid and caffeic acid in rats orally administered M. tenacissima extract. The proposed approach was successfully used for investigating the distributions of those eight analytes in rat tissues, with digoxin being used as an internal control. The Eclipse Plus C 18 RRHD column was used for determination at a column temperature of 30°C. The mobile phase system consisted of acetonitrile and water (supplemented with 0.1% formic acid) under optimal gradient elution conditions. Afterwards, this approach was validated according to the requirements for the analysis of biological samples developed by the US Food and Drug Administration, including precision, accuracy, stability and matrix effects. Based on tissue distribution analysis, those eight analytes showed rapid distribution within all the tested tissues. With regard to organic acid distribution, it followed the order stomach > liver > kidney > small intestine > lung > spleen > heart, whereas the four steroids followed the order stomach > lung > spleen > small intestine > liver > kidney > heart. The present study lays the theoretical foundation for the use and development of M. tenacissima in clinical practice., (© 2020 John Wiley & Sons, Ltd.)

A simple, rapid, and selective ultra-performance liquid chromatography-tandem mass spectrometry method for determination of l-carnitine (LC) and acetyl-l-carnitine (ALC) in human serum was developed. Acetyl-l-carnitine-d 3 (ALC-d 3 ) was selected as internal standard (IS). After protein precipitation with acetonitrile-water (1 mL, 2:1, v/v), the analytes and IS were separated on a 2.5-μm XSelect HSS T3 C 18 column by gradient elution with methanol-water (containing 0.01% ammonia water) as the mobile phase at a flow rate of 0.2 mL/min. Analytes were detected with multiple reaction monitoring using a positive scan mode with electrospray ionization. Good linearity (R 2  > 0.999) was observed in the concentration range for LC and ALC. The inter- and intra-day values of relative error were -10.4% to 10.0% with CVs less than 9.84%. The average recoveries of the two analytes were 91.29%-98.23%. Blood samples containing LC and ALC were stable under various storage conditions. Normal, haemolytic, and hyperlipidaemic serum had no significant effect on the quantification of LC and ALC. This method was successfully applied to study the concentrations of endogenous LC and ALC in the serum of patients with first-episode depression., (© 2020 John Wiley & Sons, Ltd.)

Gamma-aminobutyric acid (GABA) and its precursor glutamic acid are important neurotransmitters. Both are also present in peripheral tissues and the circulation, where abnormal plasma concentrations have been linked to specific mental disorders. In addition to endogenous synthesis, GABA and glutamic acid can be obtained from dietary sources. An increasing number of studies suggest beneficial cardio-metabolic effects of GABA intake, and therefore GABA is being marketed as a food supplement. The need for further research into their health effects merits accurate and sensitive methods to analyze GABA and glutamic acid in plasma. To this end, an ultra-pressure liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS) method was developed and validated for the quantification of GABA and glutamic acid in human plasma. Samples were prepared by a protein precipitation step and subsequent solid phase extraction using acetonitrile. Chromatographic separation was achieved on an Acquity UPLC HSS reversed phase C18 column using gradient elution. Analytes were detected using electrospray ionization and selective reaction monitoring. Standard curve concentrations for GABA ranged from 3.4 to 2500 ng/mL and for glutamic acid from 30.9 ng/mL to 22,500 ng/mL. Within- and between-day accuracy and precision were <10% in quality control samples at low, medium and high concentrations for both GABA and glutamic acid. GABA and glutamic acid were found to be stable in plasma after freeze-thaw cycles and up to 12 months of storage. The validated method was applied to human plasma from 17 volunteers. The observed concentrations ranged between 11.5 and 20.0 ng/ml and 2269 and 7625 ng/ml for respectively GABA and glutamic acid. The reported method is well suited for the measurement of plasma GABA and glutamic acid in pre-clinical or clinical studies., (Copyright © 2020 The Author(s). Published by Elsevier B.V. All rights reserved.)

As one of the featured products in northeast China, Oviductus Ranae has been widely used as a nutritious food, which contains a variety of bioactive unsaturated fatty acids (UFAs). It is necessary to establish a scientific and reliable determination method of UFA contents in Oviductus Ranae . In this work, six principal UFAs in Oviductus Ranae , namely eicosapentaenoic acid (EPA), linolenic acid (ALA), docosahexaenoic acid (DHA), arachidonic acid (ARA), linoleic acid (LA) and oleic acid (OA), were identified using UPLC-MS/MS. The UFAs identified in Oviductus Ranae were further separated based on the optimized RP-HPLC conditions. Quantitative analysis of multi-components by single-marker (QAMS) method was implemented in content determination of EPA, ALA, DHA, ARA and OA, where LA was used as the internal standard. The experiments based on Taguchi design verified the robustness of the QAMS method on different HPLC instruments and chromatographic columns. The QAMS and external standard method (ESM) were used to calculate the UFA content of 15 batches of Oviductus Ranae samples from different regions. The relative error (r < 0.73%) and cosine coefficient showed that the two methods obtained similar contents, and the method validations met the requirements. The results showed that QAMS can comprehensively and effectively control the quality of UFAs in Oviductus Ranae which provides new ideas and solutions for studying the active components in Oviductus Ranae .

C-005 is a novel third-generation EGFR tyrosine kinase inhibitor for the treatment of non-small cell lung cancer (NSCLC). To support its clinical trial, we developed a rapid and sensitive bioanalytical method based on ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) technique for the quantification of C-005 and its major metabolite in NSCLC patients following international bioanalytical guidelines. After a simple and quick protein precipitation step, the supernatant was injected to a Waters Acquity BEH C 18 column (2.1 × 50 mm i.d., 1.7 mm), and the column was eluted with a gradient of buffer A (5 mM ammonium acetate and 0.1% formic acid in water) and buffer B (formic acid-acetonitrile (1:1000, v/v)). The eluates were subsequently detected by an AB QTRAP 5500 mass spectrometer with electrospray ionization using multiple-reaction monitoring mode. The method showed good linearity from 2.00 to 1000 ng/mL for C-005 and 1.00 to 500 ng/mL for M1. In conclusion, the validation results demonstrated the robustness of the method and its well-poised to support the first-in-patient study of C-005 in NSCLC patients., (Copyright © 2020 Elsevier B.V. All rights reserved.)

A method involving cloud point-salting out extraction (CPSOE) coupled with UHPLC-MS/MS was developed for the determination of eleven sulfonamides in milk. In this study, the type and concentration of the surfactant, de-emulsification condition, pH value, volume of n-butanol, equilibration temperature and time were optimized. For this developed method, the linear range of SAs was from 0.05 to 50 μg L -1 , and the correlation coefficients were higher than 0.997. The average recoveries for SAs were from 61.32 to 91.67%, and the LOQs were less than 0.06 μg kg -1 .

In this study, a specific and quick ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method was fully developed and validated for simultaneous measurement of the rat plasma levels of vortioxetine (VOR), Lu AA34443 (the major metabolite of VOR), fluoxetine and its metabolite norfluoxetine with diazepam as the internal standard (IS). After a simple protein precipitation with acetonitrile for sample preparation, the separation of the analytes were performed on an Acquity UPLC BEH C18 (2.1 × 50 mm, 1.7 μm) column, with acetonitrile and 0.1% formic acid in water as mobile phase by gradient elution. The detection was achieved on a triple quadrupole tandem mass spectrometer by multiple reaction monitoring mode via an electrospray ionization source. Good linearity was observed in the calibration curve for each analyte. The data of precision, accuracy, matrix effect, recovery and stability all conformed to the bioanalytical method validation of acceptance criteria of US Food and Drug Administration recommendations. The newly developed UPLC-MS/MS method allowed simultaneous quantification of VOR, fluoxetine and their metabolites for the first time and was successfully applied to a pharmacokinetic study in rats., (© 2020 John Wiley & Sons, Ltd.)

A quick and efficient method was optimized and validated to determine chlorpyrifos in biobeds using ultra-performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS). Chlorpyrifos was extracted from the matrix with 30 mL of a mixture of acetone, phosphoric acid and water 98:1:1 (v/v/v). After homogenization, centrifugation and filtration, 125 µL of the extract was evaporated and reconstituted in 5 mL of methanol acidified with 0.1% acetic acid. Validation was performed by studying analytical curve linearity (r 2 ), estimated instrument and method limits of detection and limits of quantification (LOD i , LOD m , LOQ i and LOQ m , respectively), accuracy, precision (expressed as relative standard deviation, RSD), and matrix effect. Accuracy and precision were determined from the amount of pesticide recovered from biobed blank samples (i.e. without pesticide residue) spiked with chlorpyrifos at three different concentrations (2, 10 and 50 mg kg -1 ), with seven replicates at each concentration. For all three concentrations studied, the average recovery values obtained were between 96 and 115% with RSD values lower than 20%. The validated LOQ obtained was 2 mg kg -1 (from recovery studies) and the matrix effect observed was lower than ±20%, which demonstrated that there was neither considerable suppression nor enhancement of the analyte signal. The biobed system efficiently degraded chlorpyrifos in both 1) simulation of accidental spillage and 2) application of diluted pesticide solution. In the latter case, all the values obtained at the final sampling time (14 months) were below the validated LOQ m ., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Acamprosate is a medication used to treat alcohol dependence. Therapeutic drug monitoring is important in drugs for the treatment of substance-related disorders. Therefore, in this study, a new selective, very simple and rapid ultra-performance liquid chromatography-tandem mass spectrometer method was developed for the therapeutic drug monitoring of acamprosate. The developed method allows the determination of acamprosate in human plasma. The method was validated in terms of selectivity and linearity, which was in the range of 100-1,200 ng/ml for acamprosate. Intra-assay and inter-assay accuracy and precision were within the acceptable limits of the Eueopean Medicines Agency guideline. The lower limit of quantitation was 100 ng/ml for acamprosate. The developed method was successfully applied for therapeutic drug monitoring in patient plasma samples., (© 2020 John Wiley & Sons, Ltd.)

Abiraterone acetate efficacy against prostate cancer is dependent on the circulating levels of abiraterone and its active metabolites, which present significant pharmacokinetic variability among patients. Thus, therapeutic drug monitoring can be performed to improve treatment outcomes. To support such studies, there are only a limited number of bioanalytical methods in current literature. This work presents a fast method to quantify abiraterone and D4A in plasma in 4 min by UPLC-MS/MS. Bioanalytical method validation was performed according to the recommendations of the US Food and Drug Administration. The method was linear within the range of 1-400 ng/ml for abiraterone and 0.2-20 ng/ml for D4A (r 2  > 0.99). Based on the analysis of quality control samples at the lower limit of quantification, low, medium and high concentrations, the method was precise (CV abiraterone  ≤ 9.72%; CV D4A  ≤ 14.64%) and accurate (CV abiraterone 95.51-107.59%; CV D4A 98.04-99.89%). Application of the method to the quantification of abiraterone and D4A in 10 clinical samples revealed important variability in the conversion ratio of abiraterone to D4A (CV 90.85%). Considering the current literature, this is the fastest method to quantify abiraterone and D4A in plasma, allowing for optimization of the analytical routine., (© 2020 John Wiley & Sons, Ltd.)

Maximum residue limits (MRLs) for pesticides have been established in teas for quality control and protection of consumer health in many countries and regions, including Europe. It is, however, challenging to quantify multi-pesticide residues at low concentrations in complex matrices, such as tea. In this study, highly sensitive, efficient, and rugged analytical methods were adapted for simultaneous determination of 400 pesticide residues in green tea products using ultra performance liquid and gas chromatography coupled to tandem mass spectroscopy (UPLC-MS/MS and GC-MS/MS). Matrix effects were minimized by combining QuEChERS (quick, easy, cheap, effective, rugged, and safe) extraction and mixed-mode SPE clean-up with different sorbents in sample preparation. A C18 SPE cartridge paired with SPE GCB/PSA proved to be the most effective clean-up method and enabled 225 pesticide residues to be quantified, based on solvent calibration curves (154 residues using UPLC-MS/MS and 71 residues using GC-MS/MS). The analytical methods were validated fully in accordance with the SANTE/11945/2015. LOQs for most pesticides (386/400 or 96.5%) were below 10 µg/kg, i.e., less than the EU MRL (5-70 mg/kg). Thus, these approaches can be applied for routine analysis of multi-pesticide residues in green tea., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Hydroxyurea (HU) is the first-ever approved drug by the United States Food and Drug Administration (USFDA) for the management of sickle cell anemia (SCA). However, its treatment is associated with severe liabilities like myelosuppression. Therefore, the aim of the present investigation was to identify phytotherapeutics through assessment of the pharmacokinetic interaction of HU with dietary bioflavonoids followed by elucidation of the same phytoconstituents for their ability to protect HU-induced toxicity in hematological profile. In this direction, we developed a sensitive ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method to estimate HU in rat plasma at first and then validated as per USFDA guidelines as there is no such precedent in the literature. A simple plasma protein precipitation method was employed for plasma sample processing. The separation was achieved in gradient mode using Syncronis HILIC column (100 × 4.6 mm, 3 μm) with a mobile phase composition of water containing 0.1% (v/v) formic acid and acetonitrile. Ionization was carried out in positive heated-electrospray ionization (H-ESI) mode. Detection was done in selected reaction monitoring (SRM) mode with m/z 77.1 > 44.4 and m/z 75.1 > 58.2 for HU and methylurea (internal standard), respectively. All the validation parameters were within the acceptable criteria. This bioanalytical method was found to be useful in assessing the preclinical pharmacokinetic interaction of HU. Concomitant administration of chrysin or quercetin with HU in rats significantly enhanced the oral exposure of HU. Lowering of total red blood cells (RBC) and hemoglobin (Hb) level by HU in rats was significantly improved in the presence of chrysin, quercetin, and naringenin. Overall, both chrysin and quercetin showed potential to be a promising phytotherapeutics for concomitant therapy with HU to combat its dose-dependent side effects., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Ethoxyresorufin (ER)-O-deethylation (EROD) activity has been widely used to assess cytochrome P450 1A (CYP1A) activity. The kinetics of CYP1A activity have been well characterized in the liver microsomes. However, studies in kidney microsomes are limited due to the much lower EROD activity in this organ. Here, we developed and validated a sensitive UPLC-MS/MS assay for the characterization of the EROD activity in the rat kidney microsomes. In a 50 µL reaction mixture, rat kidney microsomes (0.25 mg/mL) were incubated with ER (0.1-5 µM) and NADPH (1 mM) for 10 min. Acidic solvents, such as trichloroacetic acid or formic acid, used for quenching of the metabolic reactions and precipitation of the proteins, unexpectedly caused a spontaneous formation of resorufin (RES) from ER. Therefore, the metabolic reactions were terminated by adding acetonitrile, containing a deuterated internal standard (IS). Chromatographic separation was achieved on a C 18 UPLC column, and the MS/MS ion transitions were 213.9/185.9 for RES and 220.0/192.0 for IS. The assay was validated in the linear range of 0.5 nM to 75 nM of RES and had a lower limit of quantitation of 0.5 nM. The overall recoveries of RES (90%-99%) and IS (85%-103%) were relatively high, with minimal matrix effect. The assay was successfully applied to the estimation of the Michaelis-Menten (MM) kinetics of EROD activity in the rat kidney microsomes (n = 3), which showed a maximum velocity of 2.68 ± 0.17 pmol/min/mg and a MM constant of 1.72 ± 0.24 µM (mean ± SD). It is concluded that our sensitive and specific analytical method, coupled with the optimized microsomal incubation conditions, provides a robust platform for further investigations of the effects of xenobiotics, environmental factors, or pathophysiologic conditions on the kinetics of EROD activity in the kidney microsomes., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)