Amino acid divergence in the ligand-binding pocket of Vibrio LuxR/HapR proteins determines the efficacy of thiophenesulfonamide inhibitors.

The quorum-sensing signaling systems in Vibrio bacteria converge to control levels of the master transcription factors LuxR/HapR, a family of highly conserved proteins that regulate gene expression for bacterial behaviors. A compound library screen identified 2-thiophenesulfonamide compounds that specifically inhibit Vibrio campbellii LuxR but do not affect cell growth. We synthesized a panel of 50 thiophenesulfonamide compounds to examine the structure-activity relationship effects on Vibrio quorum sensing. The most potent molecule identified, PTSP (3-phenyl-1-(thiophen-2-ylsulfonyl)-1H-pyrazole), inhibits quorum sensing in multiple strains of V. vulnificus, V. parahaemolyticus, and V. campbellii at nanomolar concentrations. However, thiophenesulfonamide inhibition efficacy varies significantly among Vibrio species: PTSP is most inhibitory against V. vulnificus SmcR, but V. cholerae HapR is completely resistant to all thiophenesulfonamides tested. Reverse genetics experiments show that PTSP efficacy is dictated by amino acid sequence in the putative ligand-binding pocket: F75Y and C170F SmcR substitutions are each sufficient to eliminate PTSP inhibition. Further, in silico modeling distinguished the most potent thiophenesulfonamides from less-effective derivatives. Our results revealed the previously unknown differences in LuxR/HapR proteins that control quorum sensing in Vibrio species and underscore the potential for developing thiophenesulfonamides as specific quorum sensing-directed treatments for Vibrio infections.

Hybrid solid-phase extraction to overcome interference due to phospholipids for determination of neratinib in human plasma using UPLC-MS/MS.

A reliable and robust bioanalytical method was developed to quantify neratinib, a tyrosine kinase inhibitor in human plasma, using UPLC-MS/MS. The extraction of neratinib and its deuterated internal standard, neratinib-d6, was successfully performed on hybrid solid-phase extraction ultra-cartridges to remove the interference of phospholipids and proteins. Chromatographic analysis was performed on a UPLC BEH C18 (50 × 2.1 mm, 1.7 µm) column using 0.1% formic acid and acetonitrile under gradient conditions. The total analysis time was 1.5 min. Neratinib was quantified using electrospray ionization source operated in the positive-ion multiple reaction monitoring mode. The mass transitions of neratinib and neratinib-d6 were m/z 557.3/112.1 and m/z 563.1/118.2, respectively. The linear concentration range for neratinib was 0.5-500 ng/mL, which adequately covers concentration levels expected in real subject samples. The assay was extensively validated for various validation parameters following standard guidelines for a bioanalytical assay. The intra- and inter-batch precision was ≤4.6%, and neratinib was found to be stable under various stability conditions. The mean internal standard-normalized matrix factor and recovery were 0.997 and 95.4%, respectively. The validated method was successfully applied to a pharmacokinetic study in healthy subjects with different doses.

Application of supercritical fluid chromatography for separation and quantitation of 15 co-formulated binary anti-hypertensive medications using a single elution protocol.

A facile supercritical fluid chromatography method is proposed to analyse 15 co-formulated binary anti-hypertensive drug combinations using a customized elution procedure. The effect of mobile phase composition, column back pressure and temperature was suitably optimized for adequate retention, analyte response and resolution. The chromatographic separation of the different drug combinations was performed on a DCPak poly(4-vinylpyridine) column (250 × 4.6 mm, 5 µm) at 125-bar pressure and 40°C using a photodiode array detector. A linear gradient of CO2 and 0.1% formic acid in methanol provided the best elution conditions for all drug combinations. Baseline separation of the drugs was possible with resolution factor Rs ranging from 1.42 to 12.58. The method was validated for specificity, sensitivity, accuracy and precision, recovery and robustness. The limit of detection and limit of quantitation for aliskiren, amlodipine, atenolol, candesartan, hydrochlorothiazide, lisinopril, losartan, metoprolol, olmesartan, telmisartan and valsartan were in the range of 0.26-2.56 and 0.77-7.75 µg/mL, respectively. The thermodynamic study revealed that interactions of the drugs with the stationary phase were spontaneous as evident from the negative free energy values, and the separation process was enthalpy driven. The developed method was successfully employed to analyse these drugs in their co-formulated tablet formulations.

Usability of Electronic Health Record-Generated Discharge Summaries: Heuristic Evaluation.

BACKGROUND: Obtaining accurate clinical information about recent acute care visits is extremely important for outpatient providers. However, documents used to communicate this information are often difficult to use. This puts patients at risk of adverse events. Elderly patients who are seen by more providers and have more care transitions are especially vulnerable. OBJECTIVE: This study aimed to (1) identify the information about elderly patients' recent acute care visits needed to coordinate their care, (2) use this information to assess discharge summaries, and (3) provide recommendations to help improve the quality of electronic health record (EHR)-generated discharge summaries, thereby increasing patient safety. METHODS: A literature review, clinician interviews, and a survey of outpatient providers were used to identify and categorize data needed to coordinate care for recently discharged elderly patients. Based upon those data, 2 guidelines for creating useful discharge summaries were created. The new guidelines, along with 17 previously developed medical documentation usability heuristics, were applied to assess 4 simulated elderly patient discharge summaries. RESULTS: The initial research effort yielded a list of 29 items that should always be included in elderly patient discharge summaries and a list of 7 "helpful, but not always necessary" items. Evaluation of 4 deidentified elderly patient discharge summaries revealed that none of the documents contained all 36 necessary items; between 14 and 18 were missing. The documents each had several other issues, and they differed significantly in organization, layout, and formatting. CONCLUSIONS: Variations in content and structure of discharge summaries in the United States make them unnecessarily difficult to use. Standardization would benefit both patients, by lowering the risk of care transition-related adverse events, and outpatient providers, by helping reduce frustration that can contribute to burnout. In the short term, acute care providers can help improve the quality of their discharge summaries by working with EHR vendors to follow recommendations based upon this study. Meanwhile, additional human factors work should determine the most effective way to organize and present information in discharge summaries, to facilitate effective standardization.

Selective quantification of lacosamide in human plasma using UPLC-MS/MS: Application to pharmacokinetic study in healthy subjects with different doses.

A practical, sensitive, and robust UPLC-MS/MS method was developed and validated to quantify lacosamide in human plasma. A simple one-step protein precipitation was used to extract lacosamide and labeled lacosamide-13C, D3 as an internal standard (IS) from 150-µL plasma. The extracts were analyzed on an Eclipse Plus C18 column (50 × 2.1 mm, 1.8 µm) using 0.1% formic acid in water and methanol:acetonitrile (50:50, v/v) under gradient conditions. The extracts were quantified on LCMS-8040 using electrospray ionization source operated in positive ionization and multiple reaction monitoring modes. The method showed good linearity from 0.02 to 20 µg/mL, which was adequate to cover lacosamide concentration assayed in formulations with different strengths. The bioanalytical assay was fully validated as per current regulatory guidelines. The intra-batch and inter-batch precision values of lacosamide were less than 4.6%. Lacosamide was found to be stable at different storage conditions. The extraction recoveries and IS-normalized matrix factors for lacosamide ranged from 97.17 to 99.68% and from 0.973 to 1.012, respectively. The validated method was successfully applied to a pharmacokinetic study with three lacosamide formulations (50, 100, and 200 mg) in 36 healthy subjects. The assay reliability was determined by reanalysis of 81 subject samples.

Optimization of chromatography to overcome matrix effect for reliable estimation of four small molecular drugs from biological fluids using LC-MS/MS.

The article describes a systematic study to overcome the matrix effect during chromatographic analysis of gemfibrozil, rivastigmine, telmisartan and tacrolimus from biological fluids using LC-ESI-MS/MS. All four methods were thoroughly developed by the appropriate choice of analytical column, elution mode and pH of mobile phase for improved chromatography and overall method performance. Matrix effect was assessed by post-column analyte infusion, slope of calibration line approach and post-extraction spiking. The best chromatographic conditions established were: Acquity BEH C18 (50 × 2.1 mm, 1.7 µm) column with 5.0 mm ammonium acetate, pH 6.0-methanol as the mobile phase under gradient program for gemfibrozil; Luna CN (50 × 2.0 mm, 3 µm) column with a mobile phase consisting of acetonitrile-10 mm ammonium acetate, pH 7.0 (90:10, v/v) for rivastigmine; Inertsustain C18 (100 × 2.0 mm, 5 µm) column using methanol-2.0 mm ammonium formate, pH 5.5 (80: 20, v/v) as the mobile phase for isocratic elution of telmisartan; and Acquity BEH C18 (50 × 2.1 mm, 1.7 µm) with methanol-10 mm ammonium acetate, pH 6.0 (95:5, v/v) as mobile phase for tacrolimus. The methods were thoroughly validated as per European Medicines Agency and US Food and Drug Administration guidance and were successfully applied for pharmacokinetic studies in healthy subjects.

Quantification of fenoprofen in human plasma using UHPLC-tandem mass spectrometry for pharmacokinetic study in healthy subjects.

A rapid, simple and sensitive ultra-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method has been developed to quantify fenoprofen, a nonsteroidal anti-inflammatory drug in human plasma for a pharmacokinetic study in healthy subjects. Owing to high levels of protein binding, protein precipitation followed by solid-phase extraction was employed for the extraction of fenoprofen and fenoprofen-d3 (used as internal standard) from 200 µL human plasma. Separation was performed on a BEH C18 (50 × 2.1 mm, 1.7 µm) column using methanol-0.2% acetic acid in water (75:25, v/v) under isocratic elution. Electrospray ionization was operated in the negative mode for sample ionization. Ion transitions used for quantification in the selected reaction monitoring mode were m/z 241/197 and m/z 244/200 for fenoprofen and fenoprofen-d3, respectively. Under the optimized conditions, fenoprofen showed excellent linearity in the concentration range 0.02-20 µg/mL (r2 ≥ 0.9996), adequate sensitivity, favorable accuracy (96.4-103.7%) and precision (percentage coefficient of variation ≤4.3) with negligible matrix effect. The validated method was successfully applied to a pharmacokinetic study of fenoprofen in healthy subjects. The significant features of the method include higher sensitivity, small plasma volume for processing and a short analysis time.

Simultaneous quantitation of metformin and dapagliflozin in human plasma by LC-MS/MS: Application to a pharmacokinetic study.

A single LC-MS/MS assay has been developed and validated for the simultaneous determination of metformin and dapagliflozin in human plasma using ion-pair solid-phase extraction. Chromatographic separation of the analytes and their internal standards was carried out on a reversed-phase ACE 5CN (150 × 4.6 mm, 5 µm) column using acetonitrile-15 mm ammonium acetate, pH 4.5 (70:30, v/v) as the mobile phase. To achieve higher sensitivity and selectivity for the analytes, mass spectrometric analysis was performed using a polarity switching approach. Ion transitions studied using multiple reaction monitoring mode were m/z 130.1 [M + H]+ /60.1 for metformin and m/z 467.1 [M + CH3 COO]- /329.1 for dapagliflozin in the positive and negative modes, respectively. The linear calibration range of the assay was established from 1.00 to 2000 ng/mL for metformin and from 0.10 to 200 ng/mL for dapagliflozin to achieve a better assessment of the pharmacokinetics of the drugs. The limit of detection and limit of quantitation for the analytes were 0.39 and 1.0 ng/mL for metformin and 0.03 and 0.1 ng/mL for dapagliflozin, respectively. There was no interference of plasma matrix obtained from different sources, including hemolyzed and lipemic plasma. The method was successfully applied to study the effect of food on the pharmacokinetics of metformin and dapagliflozin in healthy subjects.

Development and validation of a rapid and sensitive UPLC-MS/MS assay for the quantification of tofacitinib in human plasma.

A highly sensitive, selective and rapid ultra-performance liquid chromatography-tandem mass spectrometry method has been developed for the quantification of a Janus kinase (JAK) inhibitor, tofacitinib (TOF). The assay employed liquid-liquid extraction with methyl-tert butyl ether to extract tofacitinib and tofacitinib-13C3 15 N (as internal standard) from human plasma. The samples were analyzed on a UPLC BEH C18 (50 × 2.1 mm, 1.7 µm) column using acetonitrile and 10.0 mm ammonium acetate, pH 4.5 (75:25, v/v) as the mobile phase within 1.4 min. The precursor/product ion transitions were monitored at m/z 313.3/149.2 and 317.4/149.2 for tofacitinib and tofacitinib-13C3 15 N, respectively, in the positive electrospray ionization mode. The calibration curves were linear (r2  ≥ 0.9978) across the concentration range of 0.05-100 ng/mL. The mean extraction recovery of tofacitinib across quality controls was 98.6%. The intra- and inter-batch precision (CV) and accuracy ranged from 2.1-5.1 and 96.2-103.1%, respectively. All validation results complied well with the current guidelines. The method is amenable to high sample throughput and was applied to determine TOF plasma concentration in a pharmacokinetic study with 12 healthy Indian subjects after oral administration of 5 mg tablets.

Determination of terbinafine in human plasma using UPLC-MS/MS: Application to a bioequivalence study in healthy subjects.

A high-throughput and sensitive ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method has been developed for the determination of terbinafine in human plasma. The method employed liquid-liquid extraction of terbinafine and terbinafine-d7 (used as internal standard) from 100 µL human plasma with ethyl acetate-n-hexane (80:20, v/v) solvent mixture. Chromatography was performed on a BEH C18 (50 × 2.1 mm, 1.7 µm) column using acetonitrile-8.0 mm ammonium formate, pH 3.5 (85:15, v/v) under isocratic elution. For quantitative analysis, MS/MS ion transitions were monitored at m/z 292.2/141.1 and m/z 299.1/148.2 for terbinafine and terbinafine-d7, respectively, using electrospray ionization in the positive mode. The method was validated according to regulatory guidance for selectivity, sensitivity, linearity, recovery, matrix effect, stability, dilution reliability and ruggedness with acceptable accuracy and precision. The method shows good linearity over the tested concentration range from 1.00 to 2000 ng/mL (r2 ≥ 0.9984). The intra-batch and inter-batch precision (CV) was 1.8-3.2 and 2.1-4.5%, respectively. The method was successfully applied to a bioequivalence study with 250 mg terbinafine in 32 healthy subjects. The major advantage of this method includes higher sensitivity, small plasma volume for processing and a short analysis time.

A high-throughput LC-MS/MS method for determination of flunarizine in human plasma: Pharmacokinetic study with different doses.

A high-throughput and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method has been developed and validated for the determination of flunarizine in human plasma. Liquid-liquid extraction under acidic conditions was used to extract flunarizine and flunarizine-d8 from 100 µL human plasma. The mean extraction recovery obtained for flunarizine was 98.85% without compromising the sensitivity of the method. The chromatographic separation was performed on Hypersil Gold C18 (50 × 2.1 mm, 3 µm) column using methanol-10 mm ammonium formate, pH 3.0 (90:10, v/v) as the mobile phase. A tandem mass spectrometer (API-5500) equipped with an electrospray ionization source in the positive ion mode was used for detection of flunarizine. Multiple reaction monitoring was selected for quantitation using the transitions, m/z 405.2 → 203.2 for flunarizine and m/z 413.1 → 203.2 for flunarizine-d8. The validated concentration range was established from 0.10 to 100 ng/mL. The accuracy (96.1-103.1%), intra-batch and inter-batch precision (CV ≤ 5.2%) were satisfactory and the drug was stable in human plasma under all tested conditions. The method was used to evaluate the pharmacokinetics of 5 and 10 mg flunarizine tablet formulation in 24 healthy subjects. The pharmacokinetic parameters Cmax and AUC were dose-proportional.

Structure of the agonist-bound neurotensin receptor.

Neurotensin (NTS) is a 13-amino-acid peptide that functions as both a neurotransmitter and a hormone through the activation of the neurotensin receptor NTSR1, a G-protein-coupled receptor (GPCR). In the brain, NTS modulates the activity of dopaminergic systems, opioid-independent analgesia, and the inhibition of food intake; in the gut, NTS regulates a range of digestive processes. Here we present the structure at 2.8 Å resolution of Rattus norvegicus NTSR1 in an active-like state, bound to NTS(8-13), the carboxy-terminal portion of NTS responsible for agonist-induced activation of the receptor. The peptide agonist binds to NTSR1 in an extended conformation nearly perpendicular to the membrane plane, with the C terminus oriented towards the receptor core. Our findings provide, to our knowledge, the first insight into the binding mode of a peptide agonist to a GPCR and may support the development of non-peptide ligands that could be useful in the treatment of neurological disorders, cancer and obesity.

Determination of silodosin and its active glucuronide metabolite KMD-3213G in human plasma by LC-MS/MS for a bioequivalence study.

A sensitive and selective liquid chromatography-tandem mass spectrometry (LC-MS/MS) method is described for the simultaneous determination of silodosin (SLD) and its active metabolite silodosin ß-d-glucuronide (KMD-3213G) in human plasma. Liquid-liquid extraction of plasma samples was carried out with ethyl acetate and methyl tert-butyl ether solvent mixture using deuterated analogs as internal standards. The extraction recoveries of SLD and KMD-3213G were in the ranges 90.8-93.4 and 87.6-89.9%, respectively. The extracts were analyzed on a Symmetry C18 (50 × 4.6 mm, 5 µm) column under gradient conditions using 10 mm ammonium formate in water and methanol-acetonitrile (40:60, v/v), within 6.0 min. For MS/MS measurements, ionization of the analytes was carried out in the positive ionization mode and the transitions monitored were m/z 496.1 → 261.2 for SLD and m/z 670.2 → 494.1 for KMD-3213G. The method showed good linearity, accuracy, precision and stability in the range 0.10-80.0 ng/mL for SLD and KMD-3213G. The IS-normalized matrix factors obtained were highly consistent, ranging from 0.962 to 1.023 for both analytes. The method was used to support a bioequivalence study of SLD and its metabolite in healthy volunteers after oral administration of 8 mg silodosin capsules.

Quantification of metronidazole in human plasma using a highly sensitive and rugged LC-MS/MS method for a bioequivalence study.

A highly sensitive, selective and rugged method has been described for the quantification of metronidazole (MTZ) in human plasma by liquid chromatography-tandem mass spectrometry using metronidazole-d4 as the internal standard (IS). The analyte and the IS were extracted from 100 µL plasma by liquid-liquid extraction. The clear samples obtained were chromatographed on an ACE C18 (100 × 4.6 mm, 5 µm) column using acetonitrile and 10.0 mm ammonium formate in water, pH 4.00 (80:20, v/v) as the mobile phase. A triple quadrupole mass spectrometer system equipped with turbo ion spray source and operated in multiple reaction monitoring mode was used for the detection and quantification of MTZ. The calibration range was established from 0.01 to 10.0 µg/mL. The results of validation testing for precision and accuracy, selectivity, matrix effects, recovery and stability complied with current bioanalytical guidelines. A run time of 3.0 min permitted analysis of more than 300 samples in a day. The method was applied to a bioequivalence study with 250 mg MTZ tablet formulation in 24 healthy Indian males.

Simultaneous quantitation of rosuvastatin and ezetimibe in human plasma by LC-MS/MS: Pharmacokinetic study of fixed-dose formulation and separate tablets.

A simple, high-throughput and highly sensitive liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) method has been developed for the simultaneous estimation of rosuvastatin and free ezetimibe. Liquid-liquid extraction was carried out using methyl-tert butyl ether after prior acidification from 300 µL human plasma. The recovery for both the analytes and their deuterated internal standards (ISs) ranged from 95.7 to 99.8%. Rosuvastatin and ezetimibe were separated on Symmetry C18 column using acetonitrile and ammonium formate buffer, pH 3.5 (30:70, v/v) as the mobile phase. The analytes were well resolved with a resolution factor of 3.8. Detection and quantitation were performed under multiple reaction monitoring using ESI(+) for rosuvastatin (m/z 482.0 → 258.1) and ESI(-) for ezetimibe (m/z 407.9 → 271.1). A linear response function was established in the concentration ranges of 0.05-50.0 ng/mL and 0.01-10.0 ng/mL for rosuvastatin and ezetimibe, respectively, with correlation coefficient, r2 ≥ 0.9991. The IS-normalized matrix factors for the analytes ranged from 0.963 to 1.023. The developed method was successfully used to compare the pharmacokinetics of a fixed-dose combination tablet of rosuvastatin-ezetimibe and co-administered rosuvastatin and ezetimibe as separate tablets to 24 healthy subjects. The reliability of the assay was also assessed by reanalysis of 115 subject samples.

Quantification of metronidazole in healthy subjects' feces using an LC-MS/MS method.

A highly selective and sensitive liquid chromatography-tandem mass spectrometry (LC MS/MS) method was developed for the quantification of metronidazole (MTZ) in human feces. The analyte was recovered from feces after liquid-liquid extraction with ethyl acetate and separated on Waters Symmetry® C18 (100 × 4.6 mm, 5µm) column using 0.1% formic acid in water and acetonitrile (40:60, v/v) as the mobile phase. A stable-deuterated internal standard metronidazole-d4 (MTZ-d4) was used in the study. Mass analysis was performed on a triple quadrupole mass spectrometer in the positive electrospray ionization mode. A linear response function of MTZ was established in the concentration range of 0.50-250 ng/g, based on dry mass. The mean extraction recovery of MTZ (97.28%) and MTZ-d4 (96.76%) from spiked feces samples was consistent at higher as well as lower concentrations. Post-column infusion analysis showed no ion-suppression/enhancement effects and the mean IS-normalized matrix factor ranged from 0.986 to 1.013. Spiked feces samples stored at -20 and - 70°C for long-term stability were stable for at least 3 months, while extracted samples (dry and wet extracts) were stable up to 24 h. The method was applied to determine MTZ in feces of 12 healthy Indian subjects.

Overcoming interference of plasma phospholipids using HybridSPE for the determination of trimetazidine by UPLC-MS/MS.

An improved, precise and reliable ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method has been developed for the quantification of trimetazidine, using trimetazidine-d8 as the internal standard (IS). Interference owing to plasma phospholipids during sample preparation was overcome using a hybrid solid-phase extraction-phospholipid ultra cartridge. The mean extraction recovery of trimetazidine (98.66%) and trimetazidine-d8 (97.63%) from spiked plasma was consistent and reproducible. Chromatographic analysis was performed on a UPLC Ethylene Bridged Hybrid (BEH) C18 (50 × 2.1 mm, 1.7 µm) column with isocratic elution using acetonitrile-5 mm ammonium formate, pH 3.5 (40:60, v/v) as the mobile phase. The parent → product ion transitions for trimetazidine (m/z 267.1 → 181.1) and trimetazidine-d8 (m/z 275.2 → 181.1) were monitored on a triple quadrupole mass spectrometer with electrospray ionization functioning in the positive multiple reaction monitoring mode. The linearity of the method was established in the concentration range of 0.05-100 ng/mL for trimetazidine. The intra-batch and inter-batch accuracy and precision (CV) were 97.3-103.1 and 1.7-5.3%, respectively. Qualitative and quantitative assessment of matrix effect showed no interference of endogenous/exogenous components. The developed method was used to measure plasma trimetazidine concentration for a bioequivalence study with 12 healthy subjects.

LC-MS/MS analysis of metformin, saxagliptin and 5-hydroxy saxagliptin in human plasma and its pharmacokinetic study with a fixed-dose formulation in healthy Indian subjects.

A specific and rapid liquid chromatography-tandem mass spectrometry method is proposed for the simultaneous determination of metformin (MET), saxagliptin (SAXA) and its active metabolite, 5-hydroxy saxagliptin (5-OH SAXA) in human plasma. Sample preparation was accomplished from 50 µL plasma sample by solid-phase extraction using sodium dodecyl sulfate as an ion-pair reagent. Reversed-phase chromatographic resolution of analytes was possible within 3.5 min on ACE 5CN (150 × 4.6 mm, 5 µm) column using acetonitrile and10.0 mm ammonium formate buffer, pH 5.0 (80:20, v/v) as the mobile phase. Triple quadrupole mass spectrometric detection was performed using electrospray ionization in the positive ionization mode. The calibration curves showed good linearity (r2 ≥ 0.9992) over the established concentration range with limit of quantification of 1.50, 0.10 and 0.20 ng/mL for MET, SAXA and 5-OH SAXA respectively. The extraction recoveries obtained from spiked plasma samples were highly consistent for MET (75.12-77.84%), SAXA (85.90-87.84%) and 5-OH SAXA (80.32-82.69%) across quality controls. The validated method was successfully applied to a bioequivalence study with a fixed-dose formulation consisting of 5 mg SAXA and 500 mg MET in 18 healthy subjects. The reproducibility of the assay was demonstrated by reanalysis of 87 incurred samples.

Task distribution, work environment, and perceived health discomforts among Indian ceramic workers.

BACKGROUND: The study examined the work environment of Indian ceramics workers and assessed associations between work hazards, work category, and self-reported symptoms. METHODS: The multi-method ergonomic review technique (MMERT) checkpoints was used for work analysis and prevalence of self-reported symptoms among 329 male workers. RESULTS: Ambient temperature and relative humidity in ceramic industries were 39.9°C and 17.4% respectively. Musculoskeletal discomfort was observed as a primary complaint; especially lower extremity pain (45%). Load handlers and machine operators had the highest levels of work hazards, including high skill requirement, strenuous work posture, poor commitment by the organization. Poor job autonomy, task clarity, hot workplace, inappropriate workplace design, inadequate auxiliary support, and mental overload were significantly associated with self-reported symptoms (P < 0.001). CONCLUSION: Work categories are associated with work hazards and may lead to various health symptoms among ceramic workers. Control of workplace hazards may lower rates of symptoms and thus may lead to improved health, productivity, and well-being. Am. J. Ind. Med. 59:1145-1155, 2016. © 2016 Wiley Periodicals, Inc.

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

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