Youxin Li

Characterization of tyrosine kinase and screening enzyme inhibitor by capillary electrophoresis with laser-induced fluoresce detector

An effective, rapid and reliable capillary electrophoresis-laser induced fluorescence (CE-LIF) procedure was built to study the characterization of tyrosine kinase (TK), which was a target for drug screening. In this procedure, CE separated the sample of the TK reaction and LIF selectively detected the fluorescence-labeled polypeptide substrate and product. The precise TK activity was quantitated by introducing the transformation ratio of the substrate (T%) to avoid the deviation resulted from the detection sensitivity and the injection amounts in different runs and different capillaries. By measuring the T%, the effects of various reaction conditions were optimized. Meanwhile, the progression of the enzyme reaction was monitored. The K(m) and V(max) were calculated for TK under the optimized experimental conditions. In addition, the inhibition effectiveness of two model inhibitors, Staurosporine and SU6656 were evaluated. The results indicated that the screening platform based on electrophoresis was suitable for TK analysis and laid a foundation for the HTS of TK inhibitors.

An approach to the determination of the enantiomeric excess at the extreme case by capillary electrophoresis

Capillary electrophoresis (CE) has been applied to determine the percentage of enantiomeric excess (ee%) of chiral compounds. In such assays, the quality of chiral selectors (CSs) plays vital roles in resolving the enantiomers for accurate determination of the ee%. Selecting an efficient CS is usually by trial and error, and is, if ever possible, time-consuming and costly. Here we propose a new approach by using the velocity gap mode of CE (VGCE) method, to simplify the method development process for ee% determination. With VGCE, it is still possible to measure ee% even when the CS has a weak resolving power. This is especially important at the extreme cases where one of the enantiomers is significantly higher than the other one. The key point of VGCE in this case is to fractionate the small part of the mixture containing both enantiomers from the major component of the enantiomer, which is already enantiopure. Baseline separations can be achieved between the two enantiomers for the small mixture due to less longitudinal dispersion, making it possible to determine the ee%. The feasibility of this VGCE approach was confirmed by the ee% measurements of amlodipine and ofloxacin, respectively. And the practical application of VGCE was tested by analyzing levamlodipine besylate tablet.

Development of a novel 96-well format for liquid-liquid microextraction and its application in the HPLC analysis of biological samples.

A novel 96-well liquid-liquid microextraction system combined with modern HPLC was developed and used for the simultaneous analysis of 96 biological samples. The system made use of hollow fibers, a 96-well plate, and a plastic base with a center hole and a side hole. One end of the hollow fiber was sealed, while the other end was attached to one of the holes positioned at the center for the plastic base. The needle was inserted into the liquid from inside or outside of the hollow fiber through the center or the side holes, respectively. The system was tested with plasma samples containing three compounds, acidic indomethacin, neutral dexamethasone, and basic propafenone. Some parameters, such as the kind and dimension of hollow fiber, pH and salt concentration of the donor phase, the selection of organic solvent for the acceptor phase, and the extraction time were investigated. Under the optimization conditions, the Log D and drug concentration of indomethacin, dexamethasone, and propafenone in plasma and urine samples were analyzed. Then, the methodology was validated. The results demonstrated that ng/mL levels could be exactly and rapidly analyzed by our system, which was equipped with an auto-injection sampler, making sample analysis more convenient.

Velocity gap mode of capillary electrophoresis developed for high-resolution chiral separations

A new CE method based on velocity gap (VG) theory has been developed for high‐resolution chiral separations. In VG, two consecutive electric fields are adopted to drive analytes passing through two capillaries, which are linked together through a joint. The joint is immersed inside another buffer vial which has conductivity communication with the buffer inside the capillary. By adjusting the field strengths onto the two capillaries, it is possible to observe different velocities of an analyte when it passes through those two capillaries and there would be a net velocity change (NVC) for the same analyte. Different analytes may have different NVC which may be specifically meaningful for enantioseparations because enantiomers are usually hard to resolve. By taking advantage of this NVC, it is possible to enhance the resolution of a chiral separation if a proper voltage program is applied. The feasibility of using NVC to enhance chiral separation was demonstrated in the separations of three pairs of enantiomers: terbutaline, chlorpheniramine, and promethazine. All separations started with partial separation in a conventional CE and were significantly improved under the same experimental conditions. The results indicated that VG has the potential to be used to improve the resolving power of CE in chiral separations.

Synthesis and applications of sulfopropyl ether γ-cyclodextrin polymer as chiral selector in capillary electrophoresis

AbstractA novel sulfopropyl ether γ-cyclodextrin polymer (SPE-γ-CDP) through polycondensating sulfated cyclodextrins (SCDs) was synthesized. This synthesis approach also has the potential of preparing other derived cyclodextrins (CDs) polymers. The polymerized SCDs took on both the properties of SCDs and certain characteristics of polymers, such as chiral selectivity and high viscosity. Synthesis parameters, including reactions sequence, sulfation, and polycondensation conditions were investigated systematically. The product was characterized by elemental analysis, infrared spectroscopy (IR), and indirect UV detections prior to use as background electrolytes additive. The separation conditions, including the concentration of SPE-γ-CDP, the concentration and pH of running buffer, separation voltage, as well as the additional organic solution were optimized during chiral separation of neutral, acidic, and basic enantiomers in capillary electrophoresis (CE). SPE-γ-CDP was proven to be an effective chiral resolving agent in CE with the advantages of simple synthesis process, low cost, similar ratio of charge-to-mass, low current, great reproducibility, and reusability. Graphical AbstractSynthesis and applications of sulfopropyl ether γ-cyclodextrin polymer

The development of a high-throughput measurement method of octanol/water distribution coefficient based on hollow fiber membrane solvent microextraction technique.

This paper describes the development of a novel high-throughput hollow fiber membrane solvent microextraction technique for the simultaneous measurement of the octanol/water distribution coefficient (logD) for organic compounds such as drugs. The method is based on a designed system, which consists of a 96-well plate modified with 96 hollow fiber membrane tubes and a matching lid with 96 center holes and 96 side holes distributing in 96 grids. Each center hole was glued with a sealed on one end hollow fiber membrane tube, which is used to separate the aqueous phase from the octanol phase. A needle, such as microsyringe or automatic sampler, can be directly inserted into the membrane tube to deposit octanol as the accepted phase or take out the mixture of the octanol and the drug. Each side hole is filled with aqueous phase and could freely take in/out solvent as the donor phase from the outside of the hollow fiber membranes. The logD can be calculated by measuring the drug concentration in each phase after extraction equilibrium. After a comprehensive comparison, the polytetrafluoroethylene hollow fiber with the thickness of 210 μm, an extraction time of 300 min, a temperature of 25 °C and atmospheric pressure without stirring are selected for the high throughput measurement. The correlation coefficient of the linear fit of the logD values of five drugs determined by our system to reference values is 0.9954, showed a nice accurate. The -8.9% intra-day and -4.4% inter-day precision of logD for metronidazole indicates a good precision. In addition, the logD values of eight drugs were simultaneously and successfully measured, which indicated that the 96 throughput measure method of logD value was accurate, precise, reliable and useful for high throughput screening.

A carbosilane dendrimer bonded silica as the stationary phase for capillary electrochromatography

A novel carbosilane dendrimer bonded silica (G2-silica) was synthesized and evaluated as a potential chromatographic resin. The G2-silica was characterized by using various analytical techniques, such as infrared spectroscopy, elemental analysis and loss on ignition analysis. The G2-silica was packed into a capillary and used as the stationary phase for capillary electrochromatography (CEC). The packing characteristics such as the electric current vs. the applied voltage, the effects of voltage, pH, and acetonitrile concentration on EOF, the hydrophobic selectivity, and the repeatability were all investigated. Seven model compounds including thiourea, benzene, ethyl benzene, butyl benzene, benzyl alcohol, toluene and naphthalene were well separated on the CEC column with the separation efficiencies ranging from 40 000 to 90 000 plates per meter. The repeatability was satisfactory with the RSD of the migration time (2.4%). These results indicate that carbosilane dendrimer bonded silica can be used as a novel packing material which has the characteristics of a typical reversed phase.

Preliminary studies of a novel multifunctional wide-bore electrophoresis system

This paper presents the results obtained from a novel multifunctional analysis platform established on the basis of wide‐bore electrophoresis (WBE) and CE. The WBE–CE system integrated various analytical steps including separation, transfer, reaction, detection, and storage into a single system. During the WBE–CE process, a distinct three‐electrode format was adopted to continuously separate and transfer samples between WBE and CE without the interruption of switching on‐and‐off the power suppliers. This continuous mode of operation also helped to eliminate the need for exactly timing the transfer of specific samples zone from WBE to CE and avoided the danger of missing specific samples. Samples representing mixtures of acids, bases, or proteins were analyzed on this system for evaluating its feasibility and applicability. The results indicated that the resolution achieved on this WBE–CE system was better than either the WBE or the CE alone. Further, samples transferred out of the WBE system could participate in online reaction, such as enzymatic reaction in the CE. Alternatively, samples from the WBE system could be transferred out and stored offline in a vial for post‐transfer reaction. The results demonstrated that this WBE–CE system has the potential to be a multifunctional platform for a range of applications.