Noninvasive imaging of cell death using an Hsp90 ligand

Cell death plays a central role in normal physiology and in disease. Common to apoptotic and necrotic cell death is the eventual loss of plasma membrane integrity. We have produced a small organoarsenical compound, 4-(N-(S-glutathionylacetyl)amino)phenylarsonous acid, that rapidly accumulates in the cytosol of dying cells coincident with loss of plasma membrane integrity. The compound is retained in the cytosol predominantly by covalent reaction with the 90 kDa heat shock protein (Hsp90), the most abundant molecular chaperone of the eukaryotic cytoplasm. The organoarsenical was tagged with either optical or radioisotope reporting groups to image cell death in cultured cells and in murine tumors ex vivo and in situ. Tumor cell death in mice was noninvasively imaged by SPECT/CT using an (111)In-tagged compound. This versatile compound should enable the imaging of cell death in most experimental settings.     

Determination of lead by the precipitation of lead molybdate from a homogeneous solution by the diffusion method

A satisfactory quantitative method for the determination of lead as lead molybdate by precipitation from a homogeneous solution has been developed. The method employs the diffusion of ammonia into a solution containing lead and sodium molybdate in nitric acid medium. The precipitation of lead in the range of 28·0–280·0 mg is complete within 8–9 hours. Anions like sulphate, arsenate, phosphate, tungstate and chromate in moderate concentrations and cations such as Zn (II), Cu (II), Ni (II), Co (II), Mn (II), Cd (II), Hg (II), Ag (I), Ti (IV), As (V), Mg (II) and Al (III) do not contaminate the precipitate. The precipitate gets constant weight at 120°. This eliminated the ignition of the precipitate,i.e., the contamination of molybdic oxide by the precipitate was reduced.     

Thermally-induced dielectric relaxation spectra in three aldohexose monosaccharides

Three aldohexose monosaccharides, d-glucose, d-mannose, and d-galactose, were examined by scanning temperature dielectric analysis (DEA) from ambient temperatures through their melts. Phase transitions, including glass transition (T _g) and melting temperature (T _m), were evaluated by differential scanning calorimetry (DSC). The monosaccharides were found to exhibit thermally-induced dielectric loss spectra in their amorphous-solid phase before melting. Activation energies for electrical charging of each of the monosaccharides were calculated from an Arrhenius plot of the tan delta (e″/e′, dielectric loss factor/relative permittivity) peak frequency versus reciprocal temperature in Kelvin. The DEA profiles were also correlated with the DSC phase diagrams, showing the changes in electrical behavior associated with solid–solid and solid–liquid transitions.     

Thermal analysis of water and magnesium hydroxide content in commercial pharmaceutical suspensions milk of magnesia

A standard protocol was developed to determine the water content by thermal analysis of milk of magnesia (MoM). Differential scanning calorimetry (DSC) and thermogravimetry (TG) were used in a novel manner for examining the physical characteristics of the commercial pharmaceutical suspensions. Moisture analyzer and oven-dry methods validate the proposed protocol. MoM consists primarily of water and magnesium hydroxide [Mg(OH)₂]. Experimental design of the thermal analysis parameters were considered including sample size, flowing atmosphere, sample pan, and heating rate for both DSC and TG. The results established the optimum conditions for minimizing heat and mass transfer effect. Sample sizes used were: (5?C15?mg) for DSC and (30?C50?mg) for TG. DSC analysis used crimped crucibles with a pinhole, which allowed maximum resolution and gave well-defined mass (water) loss. TG analysis used a heating rate of 10?°C/min^?1 in an atmosphere of nitrogen. The heat of crystallization, heat of fusion, and heat of vaporization of unbound water are 334, 334, and 2,257?Jg^?1, respectively (Mitra et al. Proc NATAS Annu Conf Therm Anal Appl 30:203?C208, 2002). The DSC average water content of (MoM) was 80?wt% for name brand and 89.5?wt% for generic brand, based on the relative crystallization, melting and vaporization heats/Jg^?1 of distilled water in the recently purchased (2011) MoM samples. The TG showed a two-step process, losing water at 80?C135?°C for unbound water and bound water (MgO·H₂O) at 376?C404?°C, yielding a total average water loss of 91.9?% for name brand and 90.7?% for generic brand by mass. The difference between the high-temperature TG and the lower-temperature DSC can be attributed for the decomposition of magnesium hydroxide or MgO·H₂O. Therefore in performing this new approach to water analysis by heating to a high temperature decomposed the magnesium hydroxide residue. It was determined that the TG method was the most accurate for determining bound and unbound water.     

Finite difference analysis of torsional vibrations of pretwisted,rotating, cantilever beams with effects of warping

Theoretical natural frequencies of the first three modes of torsional vibration of pre-twisted, rotating cantilever beams are determined for various thickness and aspect ratios. Conclusions concerning individual and collective effects of warping, pretwist, tension-torsion coupling and tennis racket effect (twist-rotational coupling) terms on the natural frequencies are drawn from numerical results obtained by using a finite difference procedure with first order central differences. The relative importance of structural warping, inertial warping, pretwist, tension-torsion and twist-rotational coupling terms is discussed for various rotational speeds. The accuracy of results obtained by using the finite difference approach is verified by a comparison with the exact solution for specialized simple cases of the equation of motion used in this paper.     

Virtual and experimental high-throughput screening (HTS) in search of novel inosine 5′-monophosphate dehydrogenase II (IMPDH II) inhibitors

IMPDH (Inosine 5??-monophosphate dehydrogenase) catalyzes a rate-limiting step in the de novo biosynthesis of guanine nucleotides. IMPDH inhibition in sensitive cell types (e.g., lymphocytes) blocks proliferation (by blocking RNA and DNA synthesis as a result of decreased cellular levels of guanine nucleotides). This makes it an interesting target for cancer and autoimmune disorders. Currently available IMPDH inhibitors such as mycophenolic acid (MPA, uncompetitive inhibitor) and nucleoside analogs (e.g., ribavirin, competitive inhibitor after intracellular activation by phosphorylation) have unfavorable tolerability profiles which limit their use. Hence, the quest for novel IMPDH inhibitors continues. In the present study, a ligand-based virtual screening using IMPDH inhibitor pharmacophore models was performed on in-house compound collection. A total of 50,000 virtual hits were selected for primary screen using in vitro IMPDH II inhibition up to 10???M. The list of 2,500 hits (with >70?% inhibition) was further subjected to hit confirmation for the determination of IC₅₀ values. The hits obtained were further clustered using maximum common substructure based formalism resulting in 90 classes and 7 singletons. A thorough inspection of these yielded 7 interesting classes in terms of mini-SAR with IC₅₀ values ranging from 0.163???M to little over 25???M. The average ligand efficiency was found to be 0.3 for the best class. The classes thus discovered represent structurally novel chemotypes which can be taken up for further development.     

Solid-state mechanical properties of crystalline drugs and excipients

Thermal mechanical analysis (TMA) of crystalline drugs and excipients in their pre-melt temperature range performed in this study corroborate their newly found linear dielectric conductivity properties with temperature. TMA of crystalline active pharmacy ingredients (APIs) or excipients shows softening at 30–100 °C below the calorimetric melting phase transition, which is also observed by dielectric analysis (DEA). Acetophenetidin melts at 135 °C as measured calorimetrically by DSC, but softens under a low mechanical stress at 95 °C. At this pre-melting temperature, the crystals collapse under the applied load, and the TMA probe shows rapid displacement. The mechanical properties yield a softening structure and cause a dimensionally slow disintegration resulting in a sharp dimensional change at the melting point. In order to incorporate these findings into a structure–property relationship, several United States Pharmacopeia (USP) melting-point standard drugs were evaluated by TMA, DSC, and DEA, and compared to the USP standard melt temperatures. The USP standard melt temperature for vanillin (80 °C) [1], acetophenetidin (135 °C) [2], and caffeine (235 °C) [3] are easily verified calorimetrically via DSC. The combined thermal analysis techniques allow for a wide variety of the newly discovered physical properties of drugs and excipients.