High-throughput cytochrome p450 inhibition assays by ultrafast gradient liquid chromatography with tandem mass spectrometry using monolithic columns

A generic method employing ultrafast liquid chromatography with tandem mass spectrometry (LC/MS/MS) was developed and employed for routine screening of drug candidates for inhibition of five major human cytochrome p450 (CYP) isozymes, CYP3A4, CYP2D6, CYP2C9, CYP2C19, and CYP1A2. The method utilized a monolithic silica rod column to allow fast flow rates to significantly reduce chromatographic run time. The major metabolites of six CYP-specific probe substrates for the five p450 isoforms were monitored and quantified to determine IC(50) values of five drug compounds against each p450 isozyme. Human liver microsomal incubation samples at each test compound concentration were combined and analyzed simultaneously by the LC/MS/MS method. Each pooled sample containing six substrates and an internal standard was separated and detected in only 24 seconds. The combination of ultrafast chromatography and sample pooling techniques has significantly increased sample throughput and shortened assay turnaround time, allowing a large number of compounds to be screened rapidly for potential p450 inhibitory activity, to aid in compound selection and optimization in drug discovery.     

A kinematical study of Kirchhoff-Rayleigh flow

Summary A method of calculating the separated flow of a viscous fluid is proposed, which allows to split up properly the boundary condition problem from the viscous phenomena. The theory is developed for the flow past a plate and yields wakes of finite extension having an underpressure which depends directly on the amount of vorticity diffusion and dissipation occurring in the fluid. Application of the method to real flows shows good agreement between the calculated and the measured velocity distributions in front of the plate and in the wake.

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Résumé Une méthode de calcul de l'écoulement décollé d'un fluide visqueux est proposée qui permet de séparer clairement le problème aux limites des phénomènes visqueux. La théorie est développée pour l'écoulement autour d'une plaque et donne des sillages de longueur finie ayant une dépression de culot directement dépendante de l'intensité de la diffusion et dissipation de la vorticité se produisant dans le fluide. L'application de la méthode à des écoulements réels montre une bonne concordance entre les répartitions de vitesse calculées et mesurées sur le devant de la plaque et dans le sillage.

     

Gas phase activation energy for unimolecular dissociation of biomolecular ions determined by focused RAdiation for gaseous multiphoton ENergy transfer (FRAGMENT)

We present a novel approach for the determination of activation energy for the unimolecular dissociation of a large (>50 atoms) ion, based on measurement of the unimolecular dissociation rate constant as a function of continuous-wave CO(2) laser intensity. Following a short ( approximately 1 s) induction period, CO(2) laser irradiation produces an essentially blackbody internal energy distribution, whose 'temperature' varies inversely with laser intensity. The only currently available method for measuring such activation energies is blackbody infrared radiative dissociation (BIRD). Compared with BIRD, FRAGMENT: (a) eliminates the need to heat the surrounding ion trap and vacuum chamber to each of several temperatures (each requiring hours for temperature equilibration); (b) offers a three-fold wider range of effective blackbody temperature; and (c) extends the range of applications to include initially cold ions (e.g., gas-phase H/D exchange). Our FRAGMENT-determined activation energy for dissociation of protonated bradykinin, 1.2 +/- 0.1 eV, agrees within experimental error to the value, 1.3 +/- 0.1 eV, previously reported by Williams et al. from BIRD experiments. Copyright 1999 John Wiley & Sons, Ltd.     

Self-chemical ionization of diethylzinc

The self-chemical ionization of diethylzinc is examined by Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometry and semiempirical molecular orbital calculations. Electron impact of diethylzinc neutral produces the radical cation, C(4)H(15)Zn(+) (m/z x 122), which reacts further with the neutral (C(2)H(5))(2)Zn to give the following product ions: Zn(+) (m/z x 64), C(2)H(5)Zn(+) (m/z x 93), C(4)H(9)Zn(+) (m/z x 121), C(4)H(11)Zn(2)(+) (m/z x 187), and C(6)H(15)Zn(2)(+) (m/z x 215). To determine the structure and pathways for production of these ions, monoisotopic (12)C(4)H(15)(64)Zn(+), (64)Zn(+) and (12)C(2)H(5)(64)Zn(+) were individually isolated and reacted with the neutral background. We also performed semiempirical molecular orbital calculations (ZINDO/1). The molecular orbital calculations and experimental data are consistent in predicting that the ethyl group on the diethylzinc cation carries the positive charge. Copyright 1999 John Wiley & Sons, Ltd.     

Amplitude degradation of time-reversed pulses in nonlinear absorbing thermoviscous fluids

The linear wave equation in a lossless medium is time reversible, i.e., every solution p(x, t) has a temporal mirror solution p(x, -t). Analysis shows that time reversal also holds for the lossless nonlinear wave equation. In both cases, time-reversal invariance is violated when losses are present. For nonlinear propagation loses cannot normally be ignored; they are necessary to prevent the occurrence of multivalued waveforms. Further analysis of the nonlinear wave equation shows that amplification of a time-reversed pulse at the array elements also leads to a violation of time reversal even for lossless nonlinear acoustics. Numerical simulations are used to illustrate the effect of nonlinearity on the ability of a time-reversal system to effectively focus on a target in an absorbing fluid medium. We consider both the amplitude and arrival time of retrodirected pulses. The numerical results confirm that both shock generation (with the accompanying absorption) and amplification at the array, adversely affect the ability of a time-reversal system to form strong retrodirective sound fields.     

Active suppression of radiating wavenumbers in the dynamic response of submerged plates

The sound radiated from an electrically excited piezoelectric plate is computed. Voltage distributions obtainable from an array of finite sized electrodes are used to examine the possibility of actively suppressing sound radiated at particular spatial wavenumbers. An application to canceling the reflection of a plane wave at oblique incidence is presented. The results presented here can also be found in Barbone & Braga (1992).Published in Izvestiya Vysshikh Uchebnykh Zavedenii, Radiofizika, Vol. 36, No. 8, pp. 848–852, August, 1993.