Natalie Foster

Training via the web: a virtual instrument

Abstract Using readily available software to capture screens from a gas chromatograph–mass spectrometer (GC–MS) while setting up and running actual experiments, we have developed a virtual GC–MS mounted on the World Wide Web to train students in instrument operation. This resource enables students to use any computer by which they have access to the Internet to learn how to run the instrument. The students learn to operate the virtual instrument using this program outside the laboratory, thus freeing the instrument for use within the laboratory periods to run meaningful experiments and collect data. The virtual instrument has proved popular with students and faculty at both, the undergraduate and graduate levels for its realism and usefulness.

Development of a molecularly imprinted polymer for the analysis of avermectin

Five molecularly imprinted polymers (MIPs) were synthesized for a large molecule, avermectin, using different preparation techniques, monomers, and polymerization solvents. Selectivities (α) of each were compared using HPLC and different mobile phases containing various levels of acetic acid. Selectivity (α) for avermectin was greatest (α estimated ≥18) when the polymer was prepared non-covalently (utilizing only non-covalent interactions between avermectin and monomer) in chloroform using methacrylic acid (MAA) monomer and evaluated in chloroform. When evaluated in acetonitrile, an MIP prepared in acetonitrile provided better selectivity (α=8.4) than the polymer prepared in chloroform. Optimizing mobile phase conditions by adding acetic acid was much more important when MIPs were evaluated in chloroform than in acetonitrile. MIPs prepared with MAA provided better selectivity than a polymer prepared with acrylamide monomer. Covalent preparation of two MIPs utilizing a covalent bond between avermectin and monomer before polymerization did not improve selectivity but did improve peak shape in chromatograms. Specificity was demonstrated by comparing the selectivity of avermectin with eprinomectin (α=3.0), a compound with a very similar structure. Results indicate that an MIP can be prepared for the large avermectin molecule, and has the potential to simplify sample preparation and to reduce the time needed for analysis.

Interactions of porphyrins and transfer RNA

The interactions of the free base porphyrin, tetra-(4N-methylpyridyl)porphyrin and its copper(II), manganese(III) and zinc(II) complexes with brewers yeast type V phenylalaninyl tRNA were evaluated by UV-visible spectroscopy, circular dichroism and melting temperature studies over a range of magnesium ion concentrations and ionic strengths. Scatchard analysis of absorption spectra of the porphyrins in the presence of tRNA showed the free base, copper and zinc porphyrins to have binding constants of 7.3 X 10(7), 1.7 X 10(6) and 2.3 X 10(8), respectively; the manganese(III) complex did not demonstrate changes in its electronic spectra that enable the calculation of a binding constant. The results of the spectroscopic studies indicate a mode of binding for the free base, copper(II) and zinc(II) complexes that is neither intercalative nor simply outside electrostatic. The magnitude of the binding constants and the UV-visible results support intercalation, but the analyses of the thermal denaturation studies and the circular dichroism evaluations suggest that the porphyrins are associating at a single site in a fold of the tertiary structure of the tRNA close to several crucial hydrogen bonds, perhaps in the vicinity of the P10 loop. That the manganese(III) complex does not bind in this site points to constraints on the axial thickness of a molecule that may be accommodated in this locus.

Separation of metalloporphyrins from metallation reactions by liquid chromatography and electrophoresis.

The analytical separation of the indium and manganese complexes of three synthetic, meso-substituted, water-soluble porphyrins from their respective free bases in metallation reaction mixtures is described. The ligands tetra-3N-methylpyridyl porphyrin, tetra-4N-methylpyridyl porphyrin and tetra-N,N,N-trimethylanilinium porphyrin are complexed with In (III) and Mn (III) and are separated from residual free base by high-performance liquid chromatography (HPLC) in acidic conditions with gradient elution on ODS bonded stationary phase. Electrophoretic separation is achieved on both cellulose polyacetate strips and polyacrylamide tube gels under basic conditions. Although analytical separations can be achieved by both HPLC and electrophoresis, only HPLC is suitable for the development of preparative scale separations. Column chromatography, ion-pairing and ion-suppression HPLC techniques fail to separate such highly charged and closely related aromatic compounds.

A Comparison of Human Semen from Healthy, Subfertile, and Post‐Vasectomy Donors by 31P NMR Spectroscopy

Preliminary characterizations of the phosphorylated compounds present in human semen were obtained using a JEOL FX90-Q Multinuclear NMR with a variable temperature accessory. Semen samples of 2.0 ml containing 12.5% D20 in a 10.0 mm sample tube were maintained at approximately 28.5 degrees Celsius. The spectra were recorded with a 250 msec pulse delay 1 sec acquisition time 0.95 sec acquisition delay and over the range of -16 ppm to 11 ppm. 3 distinguishable peaks were identified as glycerylphosphorycholine (GPC) at 0.00 ppm (this point was used as an internal standard) inorganic phosphate (Pi) at -2.45 ppm to -2.00 ppm and phosphorycholine (PC) at -3.4 ppm in accord with Arrata et al. A temporal study showed that in the hour immediately following ejaculation the original ratio of PC to total phosphate (P) decreased by 60% while the original ratio of GPC to total P did not change by a significant amount. These data contrast with those of arrata et al. who reported that PC hydrolyzed to Pi within 30 minutes and that GPC was stable. Data reported below were collected after a minimum of 60 minutes postejaculation following relative stabilization of GPC content. Semen specimens were evaluated either within 5 hours of collection (fresh) or quickly frozen without cryoprotectant and stored at -76 degrees Celsius (frozen). Ratios of GPC to total P differed with sample type. The authors observed the following ratios for GPC to total P: (0.17 0.15 0.16 0.19 0.099 0.16 0.15) N=7 with a mean of 0.15 +or- 0.028 for fresh healthy semen (0.085 0.070 0.086) N=3 with a mean of 0.080 +or- 0.008 for frozen health semen (0.074 0.074) N=2 with a mean of 0.074 for fresh subfertile semen (fig. 2) (0.11 0.040 0.060 0.00 0.060 0.00) N=6 with a mean of 0.045 +or- 0.042 for frozen subfertile semen and a ratio of (0.00 0.00 0.00 0.00) N=4 with a mean of 0.00 for postvasectomy specimens. While the data are generally consistent with those of Arrata et al. the authors detect a difference between fresh and frozen samples. This discrepancy may reflect partial biological hydrolysis of GPC and PC to Pi. The simple reliable quantification of GPC is important for further investigation of its biological significance. The authors suggest that 31P NMR analyses of human semen will play a valuable role in understanding detection and treatment of male infertility. (full text)

Chapter 30. Progress in the Development of Radioimaging Agents

Publisher Summary One recent classification system views radioimaging agents as either substrate nonspecific in which the compound does not participate in a specific chemical reaction or substrate specific in which the substrate must participate in a definite chemical reaction or take part in a specific ligand-substrate interaction. Radioactive gases for lung ventilation, labeled cells as blood pool or spleen imaging agents, tagged particles for delineating capillary beds, and radiocolloids for uptake in the reticuloendothelial system are examples of substrate non-specific radiodiagnostics, Enzyme substrates or inhibitors, receptor binding ligands, metabolically trapped biomolecules and their models, and antibodies to site-associated antigens are prime examples of substrate specific radiopharmaceuticals. At present, there are 48 radiopharmaceuticals available. Technetium labeled (Tc-99m) ligands, colloids, and particles are predominate on the NDA list, but many of these substances would be classified as substrate nonspecific agents. A few organic radiopharmaceuticals, both substrate nonspecific and substrate specific, have achieved commercialization; these include radioiodinated rose bengal, fibrinogen, oleic acid, human serum albumin, hippuric acid, as well as radiomercury chlormerodrin and Se-75 selenomethionine. Contemporary research concerns of radiopharmaceutical chemists are the development of convenient and rapid labeling methods for introducing radiohalogens into biologically important carrier molecules, preparation and evaluation of radio halogenated imaging agents, synthesis and structural studies on metal ion radionuclide-containing pharmaceuticals, rapid incorporation of positron-emitting isotopes into imaging agents, antibody transport of imaging nuclides to target tissues, and quantitative structure-distribution studies of radioactive diagnostics.

THE DESIGN OF POTENTIAL RADIOPHARMACEUTICALS BASED ON STRUCTURE-ACTIVITY RELATIONSHIPS

Abstract The past 20 years have seen the codification of several sets of comprehensive structure-partitioning mechanisms. Radiopharmaceutical scientists nave sought and continued to seek rational guidelines to predict the in vivo localization of newly developed radioactive diagnostic agents. Several of these codification systems are discussed with illustrations of their application to new radiopharmaceutical design. The authors show how design principles which have long been applied in the development of therapeutic agents (e.g., Hansch correlations, lipohilicity, chirality, enzyme substrate mimicry, etc.) are proving useful in preparation of a new generation of radiodiagnostics. The special advantages and difficulties in the application of short-lived positron emitters as well as problems in animal evaluation of new radiopharmaceutical candidates are also discussed.