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BMS-823778 is a potent 11-β-hydroxysteroid-dehydrogenase 1 (11βHSD-1) inhibitor and a potential therapeutic agent for type 2 diabetes mellitus (T2DM). A high performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay was developed and validated to enable reliable separation and quantification of both enantiomers of a chiral hydroxy metabolite (BMT-094817) in human plasma. Following liquid-liquid extraction in a 96-well plate format, chromatographic separation of the metabolite enantiomers was achieved by isocratic elution on a Chiralpak IA-3 column. Chromatographic conditions were optimized to ensure separation of both metabolite enantiomers. Metabolite enantiomers and stable isotope-labeled (SIL) internal standards were detected by positive ion electrospray tandem mass spectrometry. The LC-MS/MS assay was validated over a concentration range of 0.200-200ng/mL. Intra- and inter-assay precision values for replicate quality control samples were less than 9.9% for both enantiomers during the assay validation. Mean quality control accuracy values were within ±7.3%. Assay recoveries were high (>75%) and consistent across the assay range. The metabolite enantiomers were stable in human blood for 2h on ice. The analytes were also stable in human plasma for 25h at room temperature, 34days at -20°C and -70°C, and following five freeze-thaw cycles. No interconversion of the metabolite enantiomers was detected under any bioanalytical stress conditions, from blood collection/processing through extracted sample storage. The validated assay was successfully applied to the quantification of both metabolite enantiomers in human plasma in support of a human pharmacokinetic study.

To quantify a therapeutic PEGylated protein in monkey serum as well as to monitor its potential in vivo instability and methionine oxidation, a novel ultra high performance liquid chromatography-high resolution mass spectrometric (UHPLC-HRMS) assay was developed using a surrogate disulfide-containing peptide, DCP(SS), and a confirmatory peptide, CP, a disulfide-free peptide. DCP(SS) was obtained by eliminating the step of reduction/alkylation before trypsin digestion. It contains an intact disulfide linkage between two peptide sequences that are essential for drug function but susceptible to potential in vivo cleavages. HRMS-based single ion monitoring (SIM) on a Q Exactive™ mass spectrometer was employed to improve assay specificity and sensitivity for DCP(SS) due to its poor fragmentation and low sensitivity with SRM detection. The assay has been validated for the protein drug in monkey serum using both surrogate peptides with excellent accuracy (within ±4.4%Dev) and precision (within 7.5%CV) with a lower limit of quantitation (LLOQ) at 10 ng mL(-1). The protein concentrations in monkey serum obtained from the DCP(SS)-based assay not only provided important pharmacokinetic parameters, but also confirmed in vivo stability of the peptide regions of interest by comparing drug concentrations with those obtained from the CP-based assay or from a ligand-binding assay (LBA). Furthermore, UHPLC-HRMS allowed simultaneous monitoring of the oxidized forms of both surrogate peptides to evaluate potential ex vivo/in vivo oxidation of one methionine present in each of both surrogate peptides. To the best of our knowledge, this is the first report of using a surrogate disulfide-containing peptide for LC-MS bioanalysis of a therapeutic protein.

Herein we report a rapid, accurate and robust UHPLC–MS/MS assay for the quantitation of BMS-911453, a Janus kinase 2 inhibitor under clinical development for the treatment of myeloproliferative disorders, in human plasma. A systematic method development approach was used to optimize the mass spectrometry, chromatography, and sample extraction conditions, and to minimize potential bioanalytical risks. The validated method utilizes stable-isotope labeled 13 C 4 -BMS-911543 as the internal standard. Liquid-liquid extraction was used for sample preparation. Chromatographic separation was achieved within 2 min on a Zorbax Extend-C18 column with an isocratic elution. BMS-911543 and its internal standard were detected by positive ion electrospray tandem mass spectrometry. The assay range was from 1 to 500 ng/mL, and the standard curve was fitted with 1/ x 2 weighted linear regression. The intra-assay precision was within 5.0% CV and the inter-assay precision was within 2.6% CV. The inter-assay mean accuracy, expressed as percents of theoretical, was between 99.8% and 102.3%. The assay has high recovery (∼80%) and minimal matrix effect (0.95–1.00). BMS-911543 was stable in human plasma for at least 24 h at room temperature, 90 days at −20 °C, and following three freeze–thaw cycles. The validated method was successfully applied to sample analysis in clinical studies.

Apixaban (Eliquis™) was developed by Bristol-Myers Squibb (BMS) and Pfizer to use as an antithrombotic/anticoagulant agent and has been recently approved for the prevention of stroke and systemic embolism in patients with nonvalvular atrial fibrillation. A clinical study of apixaban, sponsored by BMS and Pfizer, included a pilot exploratory portion to evaluate the potential for future drug concentration monitoring using dried blood spot (DBS) sample collection. For DBS sample collection, a fixed blood volume was dispensed onto a DBS card by either regular volumetric pipette (venous blood collection) or capillary dispenser (finger prick blood collection). A 96-well semi-automated liquid-liquid extraction sample preparation procedure was developed to provide clean extracts for UHPLC-MS/MS quantitation. Assays using both partial-spot center punch and whole spot punch were developed and validated. The linear dynamic ranges for all the analyses were from 0.5 to 500 ng/mL. The coefficient of determination (r(2)) values was >0.9944 for all the validation runs. For the center punch approach, the intra-assay precision (%CV) was within 4.4% and inter-assay precision was within 2.6%. The assay accuracy, expressed as %Dev., was within ± 5.4% of the nominal concentrations. One accuracy and precision run was performed using the whole spot approach, the intra-assay precision (%CV) was within 7.1% and the accuracy was within ± 8.0% of the nominal concentrations. In contrast to the center punch approach, the whole spot approach eliminated the effect of hematocrit and high lipids on the analysis of apixaban in human DBS when an accurate sample blood volume was collected on DBS cards.

Dual or triple combination regimens of novel hepatitis C direct-acting antivirals (DAA, daclatasvir, asunaprevir, or beclabuvir) provide high sustained virological response rates and reduced frequency of resistance compared to clinical monotherapy. To support pharmacokinetic (PK) assessments in clinical studies, a multiplexed liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the simultaneous quantitation of daclatasvir, asunaprevir, beclabuvir (BMS-791325) and its active metabolite (BMS-794712) in human plasma was developed and validated. Human plasma samples were extracted with methyl-t-butyl ether followed by an LC-MS/MS analysis, which was conducted in a multiple reaction monitoring (MRM) mode. The lower limits of quantitation (LLOQ) were 1 ng/mL for daclatasvir, asunaprevir, and BMS-794712, and 2 ng/mL for beclabuvir. Intra-run precision (≤4.5% CV), inter-run precision (≤2.9% CV), and accuracy (±5.3% deviation) based on different concentration levels (low, geometric mean, mid and high) of the quality control samples (QCs) provided evidence of the methods accuracy and precision. Selectivity and matrix effect on LC-MS/MS detection, stability in plasma, and potential interference of coadministered drugs (ribavirin and interferon) were all evaluated and the results were acceptable. Method reproducibility was demonstrated by the reanalysis of a portion of study samples. The cross-validation results for QCs demonstrated the equivalency between this method and two single-analyte methods which were previously validated for quantitation of daclatasvir in human plasma. This approach of using a multiplexed LC-MS/MS method for the simultaneous quantitation of three DAAs is time- and cost-effective, and can maintain good data quality in sample analysis.

Monoclonal antibody production in glycoengineered Pichia pastoris was optimized for high cell density fed-batch fermentations. Results show that antibody productivity was 1.72-fold higher when maintaining oxygen-limited conditions compared to methanol-limited conditions during the protein production phase of the bioprocess. Under oxygen-limited conditions with a 30 mmol L −1 h −1 of oxygen uptake rate, an optimum methanol dosage level of 10 gL −1 was established by comparing antibody titers under different methanol dosages during protein production. Real-time on-line NIR monitoring of glycerol concentration, methanol concentration, and relative cell density provided valuable data for process characterization and optimization. Real-time profiles of glycerol concentration and relative cell density documented run-to-run consistency during the batch and fed-batch phases by analyses of the glycerol consumption and cell growth kinetics. The volumetric methanol consumption rate was characterized for the induction process following the end of the glycerol fed-batch phase. Under oxygen-limited conditions, the volumetric methanol consumption rate increased over a 60-h period and this increase was independent of the methanol concentration over a range of 2.5–30 gL −1 when the oxygen utilization rate was maintained at 30 mmol L −1 h −1 . Utility of the on-line NIR monitor is demonstrated over the one-year period of this investigation by accurately tracking biomass, glycerol concentration, and methanol concentration without user required recalibration as well as by documenting abnormal events during high cell density bioprocesses.

Biological therapeutics can induce an undesirable immune response resulting in the formation of anti-drug antibodies (ADA), including neutralizing antibodies (NAbs). Functional (usually cell-based) NAb assays are preferred to determine NAb presence in patient serum, but are often subject to interferences from numerous serum factors, such as growth factors and disease-related cytokines. Many functional cell-based NAb assays are essentially drug concentration assays that imply the presence of NAbs by the detection of small changes in functional drug concentration. Any drug contained in the test sample will increase the total amount of drug in the assay, thus reducing the sensitivity of NAb detection. Biotin-drug Extraction with Acid Dissociation (BEAD) has been successfully applied to extract ADA, thereby removing drug and other interfering factors from human serum samples. However, to date there has been no report to estimate the residual drug level after BEAD treatment when the drug itself is a human monoclonal antibody; mainly due to the limitation of traditional ligand-binding assays. Here we describe a universal BEAD optimization procedure for human monoclonal antibody (mAb) drugs by using a LC-MS/MS method to simultaneously measure drug (a mutant human IgG4), NAb positive control (a mouse IgG), and endogenous human IgGs as an indicator of nonspecific carry-over in the BEAD eluate. This is the first report demonstrating that residual human mAb drug level in clinical sample can be measured after BEAD pre-treatment, which is critical for further BEAD procedure optimization and downstream immunogenicity testing.

A sensitive, accurate and rugged UHPLC–MS/MS method was developed and validated for the quantitation of Epothilone D (EpoD), a microtubule stabilizer in development for treatment of Alzeimer's disease, in rat plasma. The ester group in EpoD can be hydrolyzed by esterases in blood or plasma, which creates a stability concern for the bioanalysis of EpoD. Species differences in the stability of EpoD in plasma were observed. Carboxylesterases were identified as the likely esterases responsible for the hydrolysis of EpoD in plasma ex vivo, and the cause of the species different stability. Phenylmethanesulfonyl fluoride, a carboxylesterase inhibitor, was used to stabilize EpoD in rat blood during sample collection, processing, and storage. A systematic method screening and optimization strategy was used to improve the assay sensitivity and minimize potential bioanalytical risks. The stabilized plasma samples were extracted by liquid–liquid extraction. Chromatographic separation was achieved on an Acquity UPLC BEH Phenyl column with a gradient elution. EpoD and its stable isotope labeled internal standards were detected by positive ion electrospray tandem mass spectrometry. The standard curve, which ranged from 0.100 to 100 ng/mL was fitted to a 1/x2 weighted linear regression model. The intra-assay precision was within ±3.6% CV and inter-assay precision was within ±4.2% CV. The assay accuracy was within ±8.3% of the nominal values. Assay recovery of EpoD was high (∼90%) and matrix effect was minimal (1.02–1.05). EpoD was stable in stabilized rat plasma for at least 30 h at room temperature, 180 days at −20 °C, and following three freeze-thaw cycles. The validated method was successfully applied to sample analysis in toxicology studies.

Two rugged liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods for the determination of propylparaben, its major metabolite, p-hydroxybenzoic acid (pHBA), and their sulfate conjugates have been developed and validated in citric acid-treated rat plasma. To prevent propylparaben being hydrolyzed to pHBA ex vivo, rat plasma was first treated with citric acid; then collected and processed at a reduced temperature (ice bath). Stable isotope labeled internal standards, d4-propylparaben, (13)C6-pHBA, and the d4-labeled internal standards of their sulfate conjugates were used in the methods. The analytes were extracted from the matrix using protein precipitation, followed by chromatographic separation on a Waters ACQUITY UPLC HSS T3 column. Quantification using negative ion electrospray was performed on a Sciex API 4000 mass spectrometer. The analytical ranges were established from 2.00 to 200 ng/mL for propylparaben, 50.0-5000 ng/mL for pHBA, 50.0-10,000 ng/mL for the sulfate conjugate of propylparaben (SPP) and 200-40,000 ng/mL for the sulfate conjugate of pHBA (SHBA). Inter- and intra-run precision for the quality control samples were less than 5.3% and 4.4% for all analytes; and the overall accuracy was within ±5.7% of the nominal values. The validated bioanalytical methods demonstrated excellent sensitivity, specificity, accuracy and precision and were successfully applied to a rat toxicology study under the regulations of Good Laboratory Practices (GLP). Strategies have been developed and applied toward overcoming the challenges related to analyte stability, and environmental and endogenous background.

In recent years, there has been increasing interest in using LC-MS/MS to measure protein drugs (biotherapeutics) in plasma/serum to support pharmacokinetic/toxicokinetic (PK/TK) studies. A number of strategies have been proposed to quantify different types of protein drug candidates in biological matrices. However, the application of LC-MS/MS in biotherapeutics is still very limited in regulated bioanalysis due to practical challenges. In this manuscript, we propose a "data-dependent" approach for the LC-MS/MS support of protein drug candidates, focusing on operational aspects. We have developed and validated a fast, simple and reliable LC-MS/MS method to quantify a protein drug candidate in mouse serum. This method can fit into our current workflow for routine small molecule LC-MS/MS assays with an overall sample preparation time of less than two hours. The method has been demonstrated to be accurate, precise and rugged, and it has been successfully applied to analyze samples from toxicology studies. The results were verified using a ligand binding assay and tested by standard-addition sample reanalysis. In addition, general recommendations for developing and validating an LC-MS/MS assay based on surrogate peptide(s) of protein drug candidates are also proposed for regulated bioanalysis.

Asunaprevir (BMS-650032) is a potent hepatitis C virus (HCV) non-structural protein protease inhibitor currently in Phase III clinical trials for the treatment of HCV infection. A rugged and accurate LC–MS/MS method was developed and validated for the quantitation of asunaprevir in rat, dog, monkey, rabbit and mouse plasma. A systematic method screening and optimization strategy was applied to achieve optimized mass spectrometry, chromatography, and sample extraction conditions. The validated method utilized stable-isotope labeled D 9 -asunaprevir as the internal standard. The samples were extracted by liquid–liquid extraction using 10% ethyl acetate in hexane. Chromatographic separation was achieved with gradient elution on a Waters Atlantis dC18 analytical column. Analyte and its internal standard were detected by positive ion electrospray tandem mass spectrometry. The standard curve, which ranged from 5.00 to 2000 ng/mL for asunaprevir, was fitted to a 1/ x 2 weighted linear regression model. The intra-assay precision was within ±3.6% CV, inter-assay precision was within ±4.0% CV, and the assay accuracy was within ±8.1% of the nominal values in all the species. The method was successfully applied to support multiple pre-clinical toxicokinetic studies in different species.

Automated sample extraction for regulated bioanalysis by liquid chromatography/tandem mass spectrometry (LC-MS/MS) still presents significant challenges. A new sample preparation methodology with a simplified and completely automated workflow was developed to overcome these challenges using cap piercing for direct biofluid transfer and evaporation-free solid phase extraction (SPE). Using pierceable cap sample tubes, a robotic liquid handler was able to sample without uncapping or recapping during sample preparation. Evaporation for SPE was eliminated by using a mobile phase-compatible elution solvent followed by sample dilution prior to LC-MS/MS analysis. Presented here are three LC-MS/MS assays validated using this methodology to support three CNS drug development programs: (1) BMS-763534 and its metabolite, BMS-790318, in dog plasma; (2) BMS-694153 in monkey plasma; and (3) Pexacerfont (BMS-562086) and two metabolites, BMS-749241 and DPH-123554, in human plasma. These assays were linear from 1.00 to 1000 or 2.00 to 2000ng/mL for each analyte with excellent assay accuracy, precision and reproducibility. These assays met acceptance criteria for regulated bioanalysis and have been successfully applied to drug development study samples. The methodology described here successfully eliminated all manual intervention steps achieving fully automated sample preparation without compromising assay performance. Importantly, this methodology eliminates the potential exposure of the bioanalyst to any infectious biofluids during sample preparation.

Retail shopping studies often conclude that desirable shopper behaviors, such as spending more money, indicate underlying approach motivation, while undesirable behaviors, such as leaving the store, indicate underlying avoidance motivation. However, hedonic consumption would seem to provide an opportunity not only for approaching fun and excitement but also for avoiding problems and stress in everyday life. This study investigates approach and avoidance motivations in a hedonic consumption context. Results show that both approach and avoidance motivation lead to heightened hedonic motivations for shopping and to more positive shopper evaluations. Additional investigation reveals several differences by gender and across four shopping contexts. Several theoretical and managerial implications are offered.

Biological sample dilution is a rate-limiting step in bioanalytical sample preparation when the concentrations of samples are beyond standard curve ranges, especially when multiple dilution factors are needed in an analytical run. We have developed and validated a Microsoft Excel-based robotic sample preparation program (RSPP) that automatically transforms Watson worklist sample information (identification, sequence and dilution factor) to comma-separated value (CSV) files. The Freedom EVO liquid handler software imports and transforms the CSV files to executable worklists (.gwl files), allowing the robot to perform sample dilutions at variable dilution factors. The dynamic dilution range is 1- to 1000-fold and divided into three dilution steps: 1- to 10-, 11- to 100-, and 101- to 1000-fold. The whole process, including pipetting samples, diluting samples, and adding internal standard(s), is accomplished within 1 h for two racks of samples (96 samples/rack). This platform also supports online sample extraction (liquid-liquid extraction, solid-phase extraction, protein precipitation, etc.) using 96 multichannel arms. This fully automated and validated sample dilution and preparation process has been applied to several drug development programs. The results demonstrate that application of the RSPP for fully automated sample processing is efficient and rugged. The RSPP not only saved more than 50% of the time in sample pipetting and dilution but also reduced human errors. The generated bioanalytical data are accurate and precise; therefore, this application can be used in regulated bioanalysis.

A liquid chromatography and tandem mass spectrometry (LC-MS/MS) method was developed and validated to simultaneously determine the concentrations of saxagliptin (Onglyza™, BMS-477118) and its major active metabolite, 5-hydroxy saxagliptin to support pharmacokinetic analyses in clinical studies. The dynamic range of the assay was 0.1-50 ng/mL for saxagliptin and 0.2-100 ng/mL for 5-hydroxy saxagliptin. Protein precipitation (PPT) with acetonitrile was used to extract the analytes from plasma matrix before injecting on an Atlantis(®) dC18 column (50 mm × 2.1 mm, 5 μm) for LC-MS/MS analysis. The sample pre-treatment process was carefully controlled to disrupt DPP4-specific binding and non-specific binding observed at lower concentrations. The recoveries for both analytes were >90%. The assay was selective, rugged and reproducible; storage stability of at least 401 days at -20°C was demonstrated. Under these chromatographic conditions, the isomers of saxagliptin and 5-hydroxy saxagliptin were chromatographically separated from saxagliptin and 5-hydroxy saxagliptin. The assay has been used to support multiple clinical studies and regulatory approvals.

The post-entry strategic positioning in a host market is important for MNEs’ success, as firms must position properly in the marketplace to gain competitive advantage. However, little attention has been paid to firms’ strategic positioning in market center as generalist firms or market peripheries as specialist firms in a host market. This study adopts the “strategy tripod” perspective that integrates resource-, industry-, and institution-based views to investigate foreign firms’ strategic positioning (i.e. their choice of generalist or specialist strategy) in the U.S. host market. The findings of this study support the major hypotheses, suggesting that: (1) market concentration and foreign firms’ heterogeneous resources affect foreign firms’ strategic positioning; (2) institutional distance between host and home countries exerts confounding moderating effects on the relationship between firm resources and strategic positioning in the host market.

This study, using a sample of Korean multinational corporations, focuses on testing the relationships between the constructs of experiential knowledge, creativity, and performance in the context of international marketing projects. Relying on a multi-level conceptualization of experiential knowledge and creativity, our findings suggest that process-based creativity is enhanced when the team members have a higher level of experiential knowledge, but outcome-based creativity is not significantly influenced by either team- or firm-level experiential knowledge. It is concluded that, in the context of international marketing projects, the domain-relevant knowledge of the actors (i.e., the team-level experiential knowledge in the foreign markets) largely governs the level of their process-based creativity. The findings also suggest that project performance is directly influenced by firm-level experiential knowledge and process-based creativity, and is indirectly influenced by team-level experiential knowledge through the mediation of process-based creativity.

A sensitive method was developed and validated for the measurement of ixabepilone (BMS-247550, Ixempra) using a demethylated analogue of ixabepilone (BMS-212188) as an internal standard. A 0.050 mL portion of each plasma sample was extracted with 0.450 mL of acetonitrile containing the internal standard via protein precipitation. The supernatant was analyzed on a LC-MS/MS system. Chromatography was carried out on a 2.0 mm x 100 mm YMC ODS-AQ 3 microm column using an isocractic mobile phase consisting of acetonitrile:10 mM ammonium acetate, pH 5.0 (70:30, v/v) at a flow rate of 0.30 mL/min. The mass spectrometer was fitted with a TurboIonSpray source and operated in negative ionization mode. Detection of ixabepilone and BMS-212188 were accomplished using multiple reaction monitoring (MRM) of precursor>product ion pairs of m/z 505.2>405.2, and 492.1>392.1, respectively. The assay range was 2.00-500 ng/mL and was fitted to a 1/x(2) weighted quadratic regression model. Replicate sample analysis indicated that intra- and inter-day accuracy and precision are within +/-15.0%. The recovery of ixabepilone from 0.050 mL of plasma containing 5.00 and 400 ng/mL was greater than 94%. The method was demonstrated to be sensitive, selective and robust, and was successfully used to support clinical studies. This paper also discussed approaches used for resolving a curve splitting issue observed during quantitative analysis of ixabepilone in biological matrices. Finally, to adapt the methodology to pharmacokinetics of ixabepilone after oral administration, the potential interference of chemical degradants on the determination of ixabepilone was evaluated.

China is rapidly becoming an important market for consumer goods, but relatively little is known about variations in consumer shopping patterns in different regions of China. We employ a cultural materialism perspective in understanding decision-making styles of inland and coastal shoppers. Our findings reveal that consumers in the two regional markets do not differ in utilitarian shopping styles but they do in hedonic shopping styles. Marketers need to understand these differences to be able to market effectively to consumers in different regional markets within China.

Two studies investigate the relationship between promotion and prevention focus, mood regulation, and retail marketplace evaluations and behaviors. Study 1, a laboratory experiment, finds that individuals high in promotion focus regulate their moods more than individuals low in promotion focus. Study 2, a field study, investigates the relationships between retail outcomes, regulatory focus, and the three core mood regulation constructs of mood monitoring, mood clarity, and mood repair. Results suggest that mood regulation is closely related to promotion and prevention focus, having both a direct influence on retail outcomes as well as mediating the influence of regulatory focus. Implications for theory and practice, limitations, and directions for future research are discussed.

An overview of our research results on the implementation of model predictive control (MPC) on-chip is presented, with central focus the development of small-size, energy efficient controllers suitable for drug delivery systems and devices. Profiling simulations coupled with codesign techniques are used in order to reveal algorithmic bottlenecks and to effectively customize implementation designs. Hardware in-the-loop simulations using a general purpose processor and a field programmable gate array, and emulations of an application specific processor are provided. The performance measurements and estimates illustrate that MPC is suitable for on-chip real-time optimal control of complex systems with fast dynamics.

BMS-068645 is a selective adenosine 2A agonist that contains a methyl ester group which undergoes esterase hydrolysis to its acid metabolite. To permit accurate determinations of circulating BMS-068645 and its acid metabolite, blood samples must be rapidly stabilized at the time of collection. A sensitive, rapid and specific liquid chromatography-tandem mass spectrometry (LC/MS/MS) method for the simultaneous quantitation of BMS-068645 and its acid metabolite in human plasma has been developed and validated using diisopropyl fluorophosphate (DFP) as the esterase inhibitor to prevent BMS-068645 from converting to its acid metabolite. The D 5 -stable isotope labeled analogs of BMS-068645 and its metabolite were used as the internal standards (IS). Analytes and IS in plasma containing 20 mM DFP were acidified and extracted into methyl tert -butyl ether. The liquid–liquid extraction effectively eliminated the strong matrix effect caused by the esterase inhibitor. The chromatographic separation was achieved on a Waters Atlantis C18 column with a run time of 4 min. Detection was performed on a Sciex API 4000 with positive ion electrospray mode (ESI/MS/MS), monitoring the ion transitions m / z 487 > 314 and 473 > 300 for BMS-068645 and its acid metabolite, respectively. The method was validated over the range from 0.020 to 10.0 ng/mL for BMS-068645 and 0.050 to 10.0 ng/mL for its acid metabolite. Inter- and intra-run precision for the quality control samples during validation were less than 8.7% and 4.0%, respectively, for the two analytes. The assay accuracy was within ±5.4% of the nominal values. The esterase inhibitor effectively stabilized BMS-068645 during blood collection and storage. Blood collection tubes containing DFP were easily prepared and used at the clinical sites and could be stored at −30 °C for 3 months. This method demonstrated adequate sensitivity, specificity, accuracy, precision, stability and ruggedness to support the analysis of human plasma samples in pharmacokinetic studies.

Previous research on both hedonic and utilitarian shopping value has focused much effort on the antecedents of shopping value with very little emphasis on the outcomes of shopping value. This study investigates the complex interrelationships between satisfaction with the retailer, hedonic and utilitarian shopping value, and important retail outcomes. Both hedonic and utilitarian shopping values are found to influence key retail outcomes. The results also support predicted differences in the relative influence of hedonic and utilitarian shopping value. Hedonic and utilitarian shopping values are also found to moderate a number of relationships between satisfaction and retail outcomes.

We propose a framework for embedding model predictive control for Systems-on-a-Chip applications. In order to allow the implementation of such a computationally expensive controller on chip, we propose reducing the precision of the microprocessor to the minimum while maintaining near optimal control performance. Taking advantage of the low precision, a logarithmic number system based microprocessor architecture is used, that allows the design of a reduced size processor, providing further energy and computational cost savings. The design parameters for this high-performance embedded controller are chosen using a combination of finite element method simulations and bit-accurate hardware emulations in a number of parametric tests. We provide the methodology for choosing the design parameters for two particular control problems; the temperature regulation in a wafer cross-section geometry, and the control of temperature in a non-isothermal fluid flow problem in a microdevice. Finally, we provide the microprocessor architecture details and estimates for the performance of the resulting embedded model predictive controller.

The concept of delight is of great interest to practitioners who understand that to keep customers loyal, a firm must go beyond merely satisfying to truly delighting them [Bus. Mark. Dig. 17 (1992) 17; Mark. News 24 (1990) 10]. However, few studies specifically dedicated to customer delight have surfaced in the marketing literature [J. Retail. 73 (1997) 311], and no research to the authors' knowledge has explored delight in a retail setting. Therefore, the purpose of this study was to examine customer delight in a retail-shopping context. Specifically, qualitative research was conducted to determine the sources of delightful and terrible shopping experiences for retail shoppers. Critical incident analysis of 113 depth interviews with shoppers revealed several factors associated with delightful or terrible shopping experiences and the resulting consequences from these experiences. A number of strategic implications are discussed, and limitations and directions for future research are also addressed.

We propose a framework for embedding model predictive control for Systems-on-a-Chip applications. In order to allow the implementation of such a computationally expensive controller on chip, we propose reducing the precision of the operations coupled with using logarithmic number system arithmetic. Two particular control problems are examined. We provide the methodology for choosing the design parameters; we emulate the performance of the embedded controller for the examined cases and we give the microprocessor architecture details.

A simple, high throughput, direct-injection high-performance liquid chromatography tandem mass spectrometry method (LC/MS/MS) has been developed and validated for the quantitation of pioglitazone in human serum. After mixing the internal standard with a sample, a 10 microl portion of the mixture was directly injected into a high-flow LC/MS/MS system, which included an extraction column, an analytical column and a six-port switching valve. The on-line extraction was achieved on an Oasis HLB column (1 mm x 50 mm, 30 microm) with a 100% aqueous loading mobile phase containing 5 mM ammonium acetate (pH 4.0) at a flow rate of 4 ml/min. The extracted analyte was eluted by a mobile phase which contained 5 mM ammonium acetate and acetonitrile. The analytical column was a Luna C18 column (4.6 mm x 50 mm, 5 microm). Detection was achieved by positive ion electrospray tandem mass spectrometry. The lower limit of quantitation of the method was 9 ng/ml. The standard curve, which ranged from 9 to 1350 ng/ml, was fitted by a weighted (1/x2) quadratic regression model. The validation results demonstrated that this method had satisfactory precision and accuracy across the calibration range. There was no evidence of instability of the analyte in human serum following three freeze-thaw cycles, and samples could be stored for at least 2 weeks at -30 degrees C. This method was used to analyze pioglitazone concentrations in human serum samples from a bioequivalence study of a blinded Actos formulation (encapsulated 15 mg tablet) and an Actos 15 mg tablet. The blinded formulation was shown to be bioequivalent to an Actos 15 mg tablet.

The selectivity and robustness of near-infrared (near-IR) calibration models based on short-scan Fourier transform (FT) infrared interferogram data are explored. The calibration methodology used in this work employs bandpass digital filters to reduce the frequency content of the interferogram data, followed by the use of partial least-squares (PLS) regression to build calibration models with the filtered interferogram signals. Combination region near-IR interferogram data are employed corresponding to physiological levels of glucose in an aqueous matrix containing variable levels of alanine, sodium ascorbate, sodium lactate, urea, and triacetin. A randomized design procedure is used to minimize correlations between the component concentrations and between the concentration of glucose and water. Because of the severe spectral overlap of the components, this sample matrix provides an excellent test of the ability of the calibration methodology to extract the glucose signature from the interferogram data. The robustness of the analysis is also studied by applying the calibration models to data collected outside of the time span of the data used to compute the models. A calibration model based on 52 samples collected over 4 days and employing two digital filters produces a standard error of calibration (SEC) of 0.36 mM glucose. The corresponding standard errors of prediction (SEP) for data collected on the 5th (18 samples) and 7th (10 samples) day are 0.42 and 0.48 mM, respectively. The interferogram segment used for the analysis contained only 155 points. These results are compatible with those obtained in a conventional analysis of absorbance spectra and serve to validate the viability of the interferogram-based calibration.

Given the increasing importance of entertainment as a retailing strategy, this study identifies a comprehensive inventory of consumers’ hedonic shopping motivations. Based on exploratory qualitative and quantitative studies, a six-factor scale is developed that consists of adventure, gratification, role, value, social, and idea shopping motivations. Using the six-factor hedonic shopping motivation profiles, a cluster analysis of adult consumers reveals five shopper segments, called here the Minimalists, the Gatherers, the Providers, the Enthusiasts, and the Traditionalists. The utility of the proposed scale is discussed both for future research and retail strategy.

Fourier transform near-infrared interferogram data are used to determine the physiological levels of glucose in a synthetic biological matrix consisting of varying levels of bovine serum albumin and triacetin in a pH 7.4 phosphate buffer. Finite impulse response (FIR) digital filters are applied to short segments of the interferogram data, and the resulting filtered interferogram intensities are used as independent variables in constructing multivariate calibration models based on quadratic partial least-squares regression analysis. To increase the performance characteristics of the filters, the collected interferograms are decimated to reduce their frequency bandwidth. By use of a combination of optical and digital filtering, the spectral bands in the region of 4000 to 5000 cm−1 are aliased to the range of 0 to 1975 cm−1. This allows the attenuation in the stopbands of the filters to be increased from 7 to 50 dB, thereby increasing their selectivity. By use of FIR filters centered on the glucose C–H combination band at 4397 cm−1 and the triacetin C–H combination band at 4446 cm−1, a calibration model for glucose is constructed that achieves a standard error of calibration of 0.484 mM and a standard error of prediction of 0.618 mM over the 1–20 mM concentration range. This prediction performance represents a 20% improvement over the performance of a model built with the full-bandwidth data. The improved model is also observed to outperform a similar calibration model based on the conventional analysis of absorbance spectra.

A flow injection method is described for the selective measurement of chromium(VI) in aqueous solutions. This method is based on the dynamic quenching of ruthenium(II) fluorescence. The detection limit is 0.43 ppm and 40 samples can be analyzed per hour. Selectivity is demonstrated over ferrous, nickel, cupric and zinc cations and no effect is observed from sulfate, chloride, borate and phosphate. Some interference quenching was measured for cyanide and nitrate, but the method is more responsive to chromium(VI) by factors of 10.2 and 82, respectively. The effects of solution pH, carrier stream flow rate and ruthenium concentration are demonstrated. Results indicate the method is suitable for measuring chromium(VI) in effluents from electroplating baths.

Near-infrared calibration models are described for the measurement of total protein, albumin protein, globulin protein, triglycerides, cholesterol, urea, glucose, and lactate. Spectra are collected in triplicate over the 5000–4000 cm −1 spectral range with a 2.5 mm optical path length for 242 undiluted human serum samples. Calibration models are generated for each analyte by performing partial least-squares (PLS) regression on raw and digitally filtered spectra. Models are optimized individually for each analyte by considering spectral range, number of model factors and width/position of a Gaussian shaped filter response function for a digital Fourier filter. Accurate measurements are demonstrated for each analyte except lactate which is below the detection limit under our experimental conditions. Standard error of prediction (SEP) for glucose is 23.3 mg/dl (1.29 mM). Relevance and stability of the glucose calibration model are examined by evaluating the accuracy of glucose predictions from 50 human serum samples collected on a modified spectrometer nineteen months after the calibration data were collected. In addition, these 50 subsequent samples were treated in a blind manner by withholding their glucose values until all predictions were complete. Results indicate a slight positive bias in the predictions corresponding to a minor instability in the model. This instability is likely due to changes in the spectrometer hardware. Nevertheless, the strong correlation between predicted and actual glucose levels in these blind samples strongly suggests that this calibration model is based on information particular to glucose.

A mathematical model is derived from the Stern–Volmer equation as a practical calibration function for various optical oxygen sensors based on quenching of ruthenium fluorescence. The model is simple and flexible, and is ideal for computing oxygen partial pressure from 0 to 760 Torr in gaseous samples, and 0 to 200 Torr in aqueous samples. Feasibility of this model for dissolved oxygen calibration is demonstrated by applying the model to data acquired from a set of three radioluminescence-based optical oxygen sensors. The model is applicable to various sensor designs and adequately accounts for stray radiation and other sources of non-quenchable light.

Poly(hydroquinone) (PHQ), synthesized from glucose-β- d -hydroquinone by peroxidase-catalyzed polymerization in aqueous solution and placed on glassy carbon electrodes, behaves as a redox mediator for glucose sensing. The highly selective nature of enzymatic catalysis leads to PHQ with a unique structure which is more soluble in organic solvents and more electrochemically active, as compared to that prepared via electrochemical methods. A glucose sensor is constructed in a pellet form with PHQ, glucose oxidase (GOD) and graphite powder. PHQ retains its redox activity and reversibility in the solid state and effectively mediates the electron transfer between the electrode and GOD. Resulting glucose biosensors possess sub-minute response times over a dynamic range from 1 to 30 mM. The PHQ mediator permits sensor operation at 100 mV (versus SCE), thereby reducing susceptibility toward common endogenous, easily oxidizable interferences.

An absorbance based flow-through fiber-optic probe is developed for measurements of ammonia in neurochemical samples. The probe is constructed by trapping a small volume of an internal indicator solution between two sets of optical fibers. This indicator solution is separated from a flowing sample stream by a microporous gas permeable membrane. The flow-through design permits analysis of small aliquots of sample which are injected and pass across the gas permeable membrane. Ammonia in the sample diffuses across this membrane, enters the internal solution, and alters the distribution of a pH sensitive chromophore. The resulting change in absorbance is measured and related to the sample ammonia concentration. The final ammonia sensor possesses a 0.2–20 μM dynamic range with a detection limit of 0.2 ± 0.1 μM and response time of 8 min. The utility of this ammonia sensor is illustrated by measuring extracellular ammonia levels from perfused rat hippocampal tissue. These measurements indicate extracellular ammonia levels fluctuate during tissue depolarization, which is consistent with our findings with other neurological tissues.

An in vitro, flow-through optical system was designed to measure hemoglobin (Hb) concentrations in the pulp space. The system included lightemitting diodes and a silicon photodetector positioned on opposing surfaces of human teeth. A syringe pump allowed a controlled flow of blood through the pulp chamber. The Hb concentration was computed as a nonlinear function of transmitted light intensity. Transmitted light intensities were also used as indicators of oxygenation level. Optical measurements correlated with Hb values measured by the conventional cyanmethemoglobin method (r = 0.993). The mean percentage error was 5.8%, and the standard error of prediction was 0.77 g/dl for Hb concentrations ranging from 4 to 20 g/dl. Deoxygenated blood exhibited up to 31% lower transmitted intensity. Light transmission through teeth may be useful in the assessment of total Hb and blood oxygenation within the pulp chamber.

A mathematical model is derived to describe the steady-state response properties of the fiber-optic ammonia sensor. Unlike our previous model, the present model permits any concentration of total ammonia nitrogen initially present in the internal solution of the sensor. A cubic equation is solved to give the magnitude of the nonprotonated form of the indicator dye under equilibrium conditions. For the first time, the effect of the initial ammonia nitrogen concentration in the internal solution is evaluated. In addition, the response equation is differentiated with respect to the sample ammonia concentration to give an expression that can be used to evaluate the effects of critical experimental parameters on the measurement sensitivity. This model is verified by comparing predicted and actual responses for several different fiber-optic sensor configurations. The model is used to generate surface maps that allow evaluation of the interrelationships between key experimental parameters such as the indicator concentration, sample ammonia concentration, internal ammonium chloride concentration and indicator acid dissociation constant. Results of this analysis indicate that optimal responses and maximal sensitivity require careful selection of values for these parameters.

The response characteristics and ideal operating conditions are described for a novel fiber-optic internal enzyme biosensor for hydrogen peroxide. The sensing mechanism involves hydrogen peroxide entering an enzyme containing internal solution by crossing a gas-permeable membrane. Once in the internal solution, the enzyme horseradish peroxidase (HRP) catalyzes the chemiluminescent reaction between hydrogen peroxide and luminol. The resulting light intensity is monitored and related to the analyte concentration. Sensor response properties strongly depend on the composition of the internal enzyme solution. Specifically, the magnitude, rate, and stability of the sensor response depend on the pH, buffer salts, and the concentrations of luminol and horseradish peroxidase. Under optimal conditions, the steady-state signal is linearly related to the square of the hydrogen peroxide concentration according to the kinetics of the HRP catalyzed reaction. The resulting hydrogen peroxide biosensor is used as the internal sensing element of a glucose biosensor by immobilizing glucose oxidase on the outer surface of the gas-permeable membrane. This glucose biosensor possesses a linear dynamic range from 2 to 18 mM with response times shorter than 60 s. Accurate measurements of glucose in blood are demonstrated.

A novel cylindrical shape geometry is proposed for fiber-optic ammonia sensors based on chromophoric indicator dyes. Sensors are constructed by trapping the internal indicator solution inside a short segment of a gas-permeable tube. Fiber-optic probes are used to supply incident radiation and to collect light that transverses through the internal solution. This cylindrical sensor geometry provides large optical path lengths which permits the use of chromophoric indicator dyes. Unlike the conventional distal tip geometry, the diffusion path is independent of the optical path which results in short response times coupled with high sensitivity and low limits of detection. Our experiments indicate that stray radiation is negligible for this sensor design, and that the optical path length essentially equals the distance between the fiber-optic probes. Sensors constructed with Bromothymol Blue as the indicator dye are evaluated. As part of this evaluation, three different modes of operation are tested. The best analytical performance obtained when a single discrete aliquot of the internal solution is used. Steady-state responses are achieved within 13-16 min for 200 nM levels of ammonia from sensors with limits of detection ranging from 150 to 20 nM.

Feasibility is demonstrated for a novel gas-sensing, internal enzyme biosensing scheme for the selective measurement of hydrogen peroxide. Two horseradish peroxidase catalysed reactions are evaluated for the detection of hydrogen peroxide as it crosses a microporous Teflon membrane at 37 degrees C. The rate at which hydrogen peroxide crosses the membrane is determined by either a fluorescence or chemiluminescence measurement and this rate is related to the concentration of hydrogen peroxide in the sample solution. Detection limits of 0.7 mM and 10 muM are estimated for the fluorescence and chemiluminescence methods, respectively. Selectivity is demonstrated for hydrogen peroxide over ascorbic acid, uric acid and tyrosine.

A novel sensor for the selective detection of hydrogen peroxide is described. This sensor is based on the internal enzyme concept configured in a gas-sensing arrangement. Hydrogen peroxide diffuses across a microporous PTFE membrane and enters the internal solution. The entering hydrogen peroxide is then electrochemically reduced by a horseradish peroxidase (HRP) catalyzed reaction with ferrocyanide as a mediator and the corresponding cathodic current is monitored. This sensing arrangement provides excellent selectivity over ionic species in solution that cannot penetrate the gas-permeable barrier. Critical design parameters include the pH and concentrations of electrolyte, HRP and ferrocyanide in the internal solution as well as the porosity and thickness of the gas-permeable membrane. The response properties are strongly influenced by temperature; where as, they are relatively insensitive to sample pH. The dynamic range for this sensor can be controlled by adjusting the composition of the internal solution, the physical properties of the membrane, and the temperature. The detection limit is μM for sensors with an internal solution composed of 0.5 M potassium chloride, 0.05 M pH 7.4 phosphate buffer, 3 U ml −1 HRP and 0.5 mM ferrocyanide and a 0.45 μm microporous PTFE membrane. Sensors possess response times of 30 s and recovery times less than a minute.

A novel fiber-optic gas sensing arrangement based on an air-gap design is evaluated. In this arrangement, a small gap of air separates the internal solution from the sample. In addition, a second air-gap separates the internal solution from a fiber-optic probe which measures the fluorescence of the internal solution. A series of gas sensors for ammonia is used to investigate several critical design parameters. The length of the air-gap between the internal solution and the fiber-optic probe affects the magnitude of response. The length of the air-gap separating the internal and sample solutions has minimal effect on either magnitude or rate of response. As with membrane-type gas sensors, thickness of the internal solution and concentration of the indicator dye are the most important sensor parameters to consider when designing a fiber-optic gas sensor.

Reduced nicotinamide adenine dinucleotide (NADH) is shown to quench the fluorescence of thionine. Quenching of thionine is extremely efficient with a half quenching concentration of only 16.1 × 10−6 M NADH. A Stern—Volmer plot is linear over the NADH concentration range from 1 to 20μM. The corresponding Stern—Volmer quenching constant is 6.2 × 104 M−1 and the limit of detection for NADH measurements is 1.6 × 10−6 M. Process of quenching is attributed to the formation of an exciplex between thionine and NADH. Potential analytical features of this system are discussed.

A fluorescence-type fiber-optic ammonia gas sensor has been developed for the determination of ammonia in untreated serum. An internal solution has been designed to provide both the selectivity and limit of detection required for this measurement. Selectivity over carbon dioxide is accomplished by adjusting the level of ammonium chloride in the internal solution. A sub-micromolar detection limit is obtained by using a combination of 2′,7′-dichlorofluorescein and 5-carboxy-2′,7′-dichlorofluorescein as the indicator dyes. The limit of detection for the resulting sensor is 0.09 μM and response times range from 2 to 6 min. When applied to the determination of ammonia in eighteen untreated serum samples, the results from the sensor compare favorably with those from the conventional glutamate dehydrogenase assay.