Strategies for the monitoring of drugs in body fluids by micellar electrokinetic capillary chromatography

Electrokinetic capillary techniques can exploit numerous separation principles, making them flexible and easily applicable to a variety of separation problems. In recent publications, this emerging technology has been shown to be well suited for monitoring drugs and metabolites in body fluids, including serum, saliva and urine. Most attention has been focused on micellar electrokinetic capillary chromatography (MECC) because it permits the separation and determination of drugs with discrimination being largely based on differences in hydrophobicity. An overview of literature data on the MECC of drugs in body fluids and recent data obtained with antiepileptics in serum and saliva, with model mixtures of illicit drugs, and with extracts from urine specimens that tested positively for opiates and cocaine metabolites are presented. Emphasis is focused on buffer selection and simple sample preparation procedures, including direct injection of body fluids, ultrafiltration and solid-phase extraction.     

Nonlinear buckling optimization of composite structures

The paper presents an approach to nonlinear buckling fiber angle optimization of laminated composite shell structures. The approach accounts for the geometrically nonlinear behaviour of the structure by utilizing response analysis up until the critical point. Sensitivity information is obtained efficiently by an estimated critical load factor at a precritical state. In the optimization formulation, which is formulated as a mathematical programming problem and solved using gradient-based techniques, a number of the lowest buckling factors are included such that the risk of “mode switching” during optimization is avoided. The presented optimization formulation is compared to the traditional linear buckling formulation and two numerical examples, including a large laminated composite wind turbine main spar, to clearly illustrate the pitfalls of the traditional formulation and the advantage and potential of the presented approach.     

The manpower allocation problem with time windows and job-teaming constraints: A branch-and-price approach

In this paper, we consider the manpower allocation problem with time windows, job-teaming constraints and a limited number of teams (m-MAPTWTC). Given a set of teams and a set of tasks, the problem is to assign to each team a sequential order of tasks to maximize the total number of assigned tasks. Both teams and tasks may be restricted by time windows outside which operation is not possible. Some tasks require cooperation between teams, and all teams cooperating must initiate execution simultaneously. We present an integer programming model for the problem, which is decomposed using Dantzig–Wolfe decomposition. The problem is solved by column generation in a branch-and-price framework. Simultaneous execution of tasks is enforced by the branching scheme. To test the efficiency of the proposed algorithm, 12 realistic test instances are introduced. The algorithm is able to find the optimal solution in 11 of the test instances. The main contribution of this article is the addition of synchronization between teams in an exact optimization context.     

Post‐buckling optimization of composite structures using Koiter's method

Thin‐walled structures, when compressed, are prone to buckling. To fully utilize the capabilities of such structures, the post‐buckling response should be considered and optimized in the design process. This work presents a novel method for gradient‐based design optimization of the post‐buckling performance of structures. The post‐buckling analysis is based on Koiter's asymptotic method. To perform gradient‐based optimization, the design sensitivities of the Koiter factors are derived, and new design optimization formulations based on the Koiter factors are presented. The proposed optimization formulations are demonstrated on a composite square plate and a curved panel where the post‐buckling stability is optimized. Copyright © 2016 John Wiley & Sons, Ltd.     

Separation and purification of methadone enantiomers by continuous- and interval-flow electrophoresis

Continuous- or free-flow electrophoresis is based upon a thin film of fluid flowing between two parallel plates. The electrolytes and the sample are continuously admitted at one end of the electrophoresis chamber and are fractionated by an array of outlet tubes at the other. Using the Octopus apparatus in a horizontal position, continuous preparative separation of methadone enantiomers in the presence of (2-hydroxypropyl)-β-cyclodextrin as a chiral selector was investigated under conditions of continuous-flow zone electrophoresis and continuous-flow isotachophoresis. The enantiomeric composition of methadone in the collected fractions was assessed by chiral capillary electrophoresis and circular-dichroism spectroscopy. In both electrophoretic modes, partial separation of the two enantiomers with an enrichment of about 80% and a throughput of 10-20 mg of racemic methadone per hour was obtained. Operating the Octopus apparatus with interrupted buffer flow during electrophoresis, a process termed interval-flow electrophoresis, resulted in complete separation of milligram quantities of the two methadone enantiomers. Furthermore, commencing with racemic methadone, continuous multistage isotachophoretic processing is shown to be suitable to purify (R)-(-)-methadone, the enantiomer with higher pharmacological activity, on a mg/h scale and at a mM concentration in the collected product stream.     

Optimization of methods and treatment conditions for studying effects of fatty acids on cell growth

Antiproliferative properties of molecular regulators of lipid metabolism have been increasingly studied during recent years. Discussion is ongoing concerning optimal treatment conditions and assays used for monitoring proliferation and cytotoxicity. The objective of the present work was to optimize methods and treatment conditions used for studying antiproliferative effects of fatty acids and analogs, represented by palmitic acid (PA) and the β-oxidation-restricted fatty acid analog tetradecylthioacetic acid (TTA), in rat (BT4Cn) and human (D54Mg and GaMg) glioma cell lines. Changes in [³H]thymidine incorporation preceded changes in cell number in TTA-treated glioma cell cultures, and the growth inhibition was more significantly expressed by [³H]thymidine incorporation than cell number. Addition of bovine serum albumin decreased cellular fatty acid uptake and reduced the effects of TTA and PA on [³H]thymidine incorporation. Determination of the antiproliferative effect of TTA in BT4Cn cells by MTT conversion and [³H]thymidine incorporation yielded concordant results. TTA-mediated reduction in cell number corresponded to reduction in cellular protein and total DNA content in BT4Cn cells. Reduced growth potential in TTA-treated multicellular D54Mg and GaMg spheroids supported the findings from monolayer cultures. In conclusion, cell density, treatment period, fatty acid administration, and methods for growth determination may profoundly influence the outcome of cell growth experiments. Thus, experimental conditions should be carefully controlled when performing cell growth experiments, and effects on cell growth should preferably be confirmed by different methods. Karl Johan Tronstad and Kjetil Berge contributed equally to this article.     

Analysis of illicit drugs in human urine by micellar electrokinetic capillary chromatography with on-column fast scanning polychrome absorption detection.

Using micellar electrokinetic capillary chromatography (MECC) with a borate/phosphate buffer containing 75 mM SDS (pH 9.1), common drugs of abuse and/or their metabolites, including opioids, benzoylecgonine, amphetamines, and methaqualone, can easily be analyzed. After solid-phase extraction of 5 mL of urine, drug concentrations down to about 100 ng/mL can be unambiguously monitored with on-column multiwavelength detection. Peak assignment is achieved through comparison of retention times and absorption spectra of eluting peaks with those of computer-stored model runs. The effectiveness of the approach is demonstrated with data obtained from different patient urines which tested positively for one or several drugs using nonisotopic immunoassays. Results suggest that MECC of illicit drugs is a highly specific and sensitive instrumental approach suitable for confirmation testing following a positive response of a toxicological screening procedure.     

Engineering the S1' subsite of trypsin: design of a protease which cleaves between dibasic residues

The serine protease trypsin was converted into a site-specific protease which hydrolyzes peptides between dibasic residues. Trypsin exhibits a high S1 specificity for Arg and Lys residues. However, the S1' specificity of trypsin is very broad, with only a slight preference for hydrophobic residues in P1'. We replaced Lys60 with Glu and Asp to introduce a high specificity for basic residues into the S1' site of trypsin. Both mutations cause a dramatic increase in the S1' specificity for Arg and Lys as measured by acyl transfer reactions. In K60E, the preference for Arg increases 70-fold while the preference for P1'-Lys increases 12-fold. In contrast, the preferences for other P1' residues either decrease slightly or remain the same. Thus, K60E prefers P1'-Arg over most other P1' residues by 2 orders of magnitude. Similar results are obtained when P1' specificity is measured in peptide cleavage assays. K60D exhibits an S1' specificity profile very similar to that of K60E, although the P1'-Arg preference is reduced by a factor of 2.5. Molecular modeling studies suggest that the high S1' specificity for Arg in K60E may be due to the formation of a salt bridge between Glu60 and the P1'-Arg of the substrate.     

Capillary electrophoresis in clinical and forensic analysis

During the past decade, capillary electrophoresis (CE) emerged as a promising, effective and economic approach for separation of a large variety of substances, including those encountered in clinical and forensic analysis. Reliable and automated CE instruments became commercially available and promoted the exploration of an increasing number of CE methods and fields of application. The widespread applicability of CE, its enormous separation power and high-sensitivity detection schemes make this technology an attractive and promising tool. This review discusses the principles and important aspects of CE-based assays and provides an overview of the key achievements encountered with CE in clinical and forensic analysis, including those associated with the analysis of serum proteins, hemoglobin variants, drugs and nucleic acids. Validated assays, interesting applications and future trends in clinical and forensic analysis are also discussed.