The peripheral benzodiazepine receptor in photodynamic therapy with the phthalocyanine photosensitizer Pc 4

The peripheral benzodiazepine receptor (PBR) is an 18 kDa protein of the outer mitochondrial membrane that interacts with the voltage-dependent anion channel and may participate in formation of the permeability transition pore. The physiological role of PBR is reflected in the high-affinity binding of endogenous ligands that are metabolites of both cholesterol and heme. Certain porphyrin precursors of heme can be photosensitizers for photodynamic therapy (PDT), which depends on visible light activation of porphyrin-related macrocycles. Because the apparent binding affinity of a series of porphyrin analogs for PBR paralleled their ability to photoinactivate cells, PBR has been proposed as the molecular target for porphyrin-derived photocytotoxicity. The phthalocyanine (Pc) photosensitizer Pc 4 accumulates in mitochondria and structurally resembles porphyrins. Therefore, we tested the relevance of PBR binding on Pc 4-PDT. Binding affinity was measured by competition with 3H-PK11195, a high-affinity ligand of PBR, for binding to rat kidney mitochondria (RKM) or intact Chinese hamster ovary (CHO) cells. To assess the binding of the Pc directly, we synthesized 14C-labeled Pc 4 and found that whereas Pc 4 was a competitive inhibitor of 3H-PK11195 binding to the PBR, PK11195 did not inhibit the binding of 14C-Pc 4 to RKM. Further, 14C-Pc 4 binding to RKM showed no evidence of saturation up to 10 microM. Finally, when Pc 4-loaded CHO cells were exposed to activating red light, apoptosis was induced; Pc 4-PDT was less effective in causing apoptosis in a companion cell line overexpressing the antiapoptotic protein Bcl-2. For both cell lines, PK11195 inhibited PDT-induced apoptosis; however, the inhibition was transient and did not extend to overall cell death, as determined by clonogenic assay. The results demonstrate (1) the presence of low-affinity binding sites for Pc 4 on PBR; (2) the presence of multiple binding sites for Pc 4 in RKM and CHO cells other than those that influence PK11195 binding; and (3) the ability of high supersaturating levels of PK11195 to transiently inhibit apoptosis initiated by Pc 4-PDT, with less influence on overall cell killing. We conclude that the binding of Pc 4 to PBR is less relevant to the photocytotoxicity of Pc 4-PDT than are other mitochondrial events, such as photodamage to Bcl-2 and that the observed inhibition of Pc 4-PDT-induced apoptosis by PK11195 likely occurs through a mechanism independent of PBR.     

Selective Homogeneous Palladium(0)-Catalyzed Hydrogenation of Alkynes to (Z)-Alkenes

Zero-valent palladium precatalysts containing rigid bidentate bis(arylimino)acenaphthene ligands (shown schematically) facilitate the highly stereoselective homogeneous catalytic hydrogenation of alkynes to (Z)-alkenes. Internal, terminal, aryl-substituted, and cyclic alkynes are suitable substrates, as are some enynes, which are chemoselectively hydrogenated to dienes. E=CO(2)Me; R(1), R(2)=4-OCH(3), 4-CH(3), 2,6-(CH(3))(2).     

A critical investigation of the effects of the radio frequency potential on the trapping of externally injected ions in ion trap mass spectrometry

The sensitivity of all ion trap mass spectrometry (ITMS) methods is dependent on the trapping efficiency of the instrument. For ITMS instruments utilizing external ion sources, such as laser desorption, trapping efficiency is known to depend on the phase and amplitude of the radio frequency (RF) potential applied to the ring electrode at the time of ion introduction. It is remarkable that, in a considerable body of literature, no consensus exists regarding the effects of these parameters on the efficacy of trapping externally generated ions. In this paper, a summary of the literature is presented in order to highlight significant discrepancies. New laser desorption ion trap mass spectrometry (LD-ITMS) data are also presented, from which conclusions are drawn in our effort to clarify some of the confusion. Copyright 1999 John Wiley & Sons, Ltd.     

Photoelastic determination of stresses in multiple-pin connectors

The design, fabrication, and testing of photoelastic models of double-lap, multiple-pin connectors are discussed. Interest is in the stresses in the inner laps. These stresses are determined by constructing models with photoelastic inner laps and transparent-acrylic outer laps. The connectors have two pins, in tandem, parallel to the load direction. A photoelastic-isotropic point is shown to permit the evaluation of load sharing between the two pins. A numerical scheme, utilizing the isochromatic- and isoclinic-photoelastic data and a finite-difference representation of the planestress equilibrium equations, is used to compute the stresses around the two pins. Representative stress distributions and stress-concentration factors are shown.     

Enantiomeric separation of R,S-naproxen by conventional and nano-liquid chromatography with methyl-beta-cyclodextrin as a mobile phase additive

Chiral separations of R,S-naproxen mixtures were obtained on an achiral column (ODS) with methyl-beta-cyclodextrin as a mobile phase additive using conventional and nano-LC. The optimised mobile phase composition was 20 mmol l(-1) methyl-beta-cyclodextrin, 20% (v/v) acetonitrile, and 50 mmol l(-1) sodium acetate buffer at pH 3 using hydrochloric acid for pH adjustment. In addition to UV detection at 232 nm, amperometric detection was also investigated. Without using any internal standard, the reproducibility of amperometric detection (+1.05 V vs. Ag/AgCl) over a long analysis cycle in LC was greatly improved by choosing the peak area ratio between R- and S-naproxen as the analytical readout (the relative standard deviation was 2.11%) and enantiomeric purity could be assessed directly. This method was successfully employed for enantiomeric purity assessment in commercial naproxen tablets. Finally, successful transfer from conventional LC to nano-LC was realised, resulting in over 1000-fold reduction in reagent consumption.     

Ion chromatography on-chip

On-chip separation of inorganic anions by ion-exchange chromatography was realized. Micro separation channels were fabricated on a silicon wafer and sealed with a Pyrex cover plate using standard photolithography, wet and dry chemical etching, and anodic bonding techniques. Quaternary ammonium latex particles were employed for the first time to coat the separation channels on-chip. Owing to the narrow depths of the channels on the chip, 0.5-10 microm, there were more interactions of the analytes with the stationary phase on the chip than in a 50-microm I.D. capillary. With off-chip injection (20 nl) and UV detection, NO2-, NO3-, I-, and thiourea were separated using 1 mM KCl as the eluent. The linear ranges for NO2- and NO3- are from 5 to 1000 microM with the detection limits of 0.5 microM.     

Evidence for TeV gamma-ray emission from a region of the galactic plane

Gamma-ray emission from a narrow band at the galactic equator has previously been detected up to 30 GeV. We report evidence for a TeV gamma-ray signal from a region of the galactic plane by Milagro, a large-field-of-view water Cherenkov detector for extensive air showers. An excess with a significance of 4.5 standard deviations has been observed from the region of galactic longitude l E (40 degrees, 100 degrees) and latitude /b/ < 5 degrees. Under the assumption of a simple power law spectrum, with no cutoff in the EGRET-Milagro energy range, the measured integral flux is phi gamma(>3.5 TeV) = (6.4 +/- 1.4 +/- 2.1) x 10(-11) cm(-2) s(-1) sr(-1). This flux is consistent with an extrapolation of the EGRET spectrum between 1 and 30 GeV in this galactic region.     

On-chip definition of picolitre sample injection plugs for miniaturised liquid chromatography

High-speed counter-current chromatography was applied to the preparative separation and purification of naphthopyranone glycosides from a crude 70% ethanolic extract of the capitula of Paepalanthus microphyllus. The solvent system used was composed of water-ethanol-ethyl acetate-hexane (10:4:10:4, v/v). This technique led to the separation of four different naphthopyranone glycosides in pure form in only 7 h.