Micellar electrokinetic chromatography of tri aza aromatic ligand compounds of iron (II): influence of bile salt type on enantiomeric separation.

Micellar electrokinetic chromatography is used with a variety of bile salt micelles to separate the enantiomers of bis(8-((pyridine-2-methylene)amino)quinoline)iron(II) hexafluorophosphate, Fe(PMAQ)2(PF6)2; bis(8-((pyridine-2-methylene)amino)lepidine iron(II) hexafluorophosphate, Fe(PMAL)2(PF6)2; and bis(1-(2-pyridinyl)ethylidine)-8-aminoquinoline iron(II) hexafluorophosphate, Fe(PEAQ)2(PF6)2. The influence of ten different bile salts on the resolution of each pair of enantiomers is investigated. Significant changes in resolution are seen depending upon the bile salt used. The dihydroxy bile salts are superior to the trihydroxy bile salts in terms of resolution, and the taurine or glycine conjugated bile salts yield better results than the unconjugated bile salts. Resolution for most enantiomers is maximized in a buffer solution containing 10-15% acetone and employing either taurochenodeoxycholic or glycochenodeoxycholic acid as the bile salt. Evidence for the separation of the corresponding Fe(III) complexes is presented.

Measurement of bilirubin partition coefficients in bile salt micelle/aqueous buffer solutions by micellar electrokinetic chromatography.

The partition coefficients for the distribution of bilirubin between aqueous phosphateborate buffer and cholic, taurocholic, taurodeoxycholic, and taurochenodeoxycholic micelles have been measured by micellar electrokinetic chromatography at pH 8.5. Determination of the partition coefficients required that the critical micelle concentration and partial specific volumes be determined for each bile salt. Critical micelle concentrations were slightly higher for the trihydroxy bile salts. Partial specific volumes of the bile salt micelles differed very little from each other, and for each bile salt they were constant over the concentration range studied, which was typically from slightly above the critical micelle concentration to 35 mM. Capacity factors were corrected for the effects of applied voltage by extrapolation of the capacity factor to zero applied volts. The free solution mobility of bilirubin, determined in the absence of bile salt, was also corrected for the effects of applied voltage. Plots of extrapolated capacity factor versus phase ratio yield the partition coefficient as the slope of a linear fit to the data. Partition coefficients for bilirubin were significantly higher for dihydroxy bile salts than for trihydroxy bile salts.

Attitudes toward colon cancer gene testing: factors predicting test uptake.

OBJECTIVES: Genetic discoveries in hereditary nonpolyposis colorectal cancer (HNPCC) have made possible genetic testing to determine susceptibility to this form of colorectal cancer (CRC). This study measured the uptake of genetic testing for HNPCC among first-degree relatives of CRC patients and conducted a preliminary analysis of the predictors of test uptake. MATERIALS AND METHODS: We compared 77 test acceptors and 181 decliners on demographic, medical history, and psychological characteristics, controlling for distance from the testing center. The psychological factors studied were risk perception for CRC, frequency of cancer thoughts, and perceived ability to cope with unfavorable genetic information. RESULTS: In the final regression model, after accounting for all variables, the significant predictors of test uptake were increased risk perception, greater perceived confidence in ability to cope with unfavorable genetic information, more frequent cancer thoughts, and having had at least one colonoscopy. The association between risk perception and uptake was dependent on frequency of cancer thoughts. Among those who thought about getting CRC more often, the probability of testing increased as perceived risk increased to approximately 50% likelihood of getting CRC and then leveled off. In contrast, among those who never or rarely thought about getting CRC, risk perception was unrelated to testing decision. CONCLUSIONS: Our findings are consistent with the associations reported between psychological factors and other cancer screening behaviors.

Increased densities of AMPA GluR1 subunit proteins and presynaptic mossy fiber sprouting in the fascia dentata of human hippocampal epilepsy.

In human hippocampal epilepsy, there is a consistent pathology of cell loss and reactive synaptic reorganization of 'excitatory' mossy fibers (MF) into the inner molecular layer (IML) of the fascia dentata (FD). In this study, neo-Timm's histochemistry of MFs and immunocytochemistry of GluR1 were used to determine, in patients with or without hippocampal sclerosis (HS), if there was a correlation between aberrant supragranular (IML) mossy fiber sprouting and increased densities of AMPA GluR1 subunit proteins in the IML of the FD. Computerized quantified densitometric grey values of Timm and GluR1 densities were corrected for the densities of granule cell losses using cell counts. In the IML of the HS group, despite the losses of granule cells, mossy fiber sprouting was significantly greater (P<0.000001) and GluR1 protein densities were significantly higher (P<0.0005) than those of the non-HS group. Unlike supragranular mossy fiber sprouting, which was limited to the IML, the increased GluR1 stainings were distributed throughout the whole molecular layer. For all cases, MF synaptic reorganization in the supragranular ML was correlated with GluR1 subunit protein densities in the IML (R=0.784, P<0.0093). These data demonstrate that in the human epileptic fascia dentata, there are significantly increased AMPA GluR1 subunit proteins associated with aberrant MF synaptic reorganizations. This suggests that the hyperexcitability of sclerotic hippocampus occurs, at least in part, from the associated changes of both presynaptic mossy fiber glutamatergic neoinnervation and increased GluR1 subunit proteins in the dendritic domains of the FD.

Induced expression of NMDAR2 proteins and differential expression of NMDAR1 splice variants in dysplastic neurons of human epileptic neocortex.

Immunocytochemistry was used to study the expressions of glutamate receptor subunit proteins for NMDAR2A/B, NMDAR1 splice variants, and AMPA Glu-R2/3 in human brain resected for intractable epilepsy associated with cortical dysplasia. NMDAR2A/B intensely labeled dysplastic neurons showing staining in both the cell bodies and dendritic profiles. However, nondysplastic neurons were not immunoreactive to NMDAR2A/B. The antibody selective to NMDAR1 splice variants of NR1-1a. -1b, -2a, and -2b labeled dysplastic neurons, but few nondysplastic neurons. In contrast, the antibody to splice variants of NR1-1a, -1b, 2a, -2b, -3a, -3b, -4a, and -4b labeled both dysplastic and nondysplastic neurons. The different labeling patterns by these two antibodies indicate that variants of NMDAR1-3a, -3b, -4a, and -4b are present in nondysplastic neurons. Both dysplastic neurons and nondysplastic neurons were immunoreactive to AMPA GluR2/3, but denser immunoreactivity was observed in dysplastic neurons. We also found that the locations of dysplastic neurons labeled by NMDAR2A/B were related to focal epileptic EEG seizure onsets or spiking and to focal behavioral seizure types. Our results suggest that there is hyperexcitability of dysplastic cortical regions, at least in part, from the presence of NMDAR2 subunits and selectively expressed NMDAR1 splice variants in dysplastic neurons.

Bile salt micellar electrokinetic chromatography of bilirubin and related compounds.

The interaction of bilirubin, biliverdin, bilirubin dimethyl ester, biliverdin dimethyl ester, xanthobilirubic acid, and xanthobilirubin methyl ester with trihydroxy and dihydroxy bile salt solutions is investigated by micellar electrokinetic chromatography (MEKC). The capacity factor of each compound is measured in solutions of the different bile salts over the pH range of 6.5-9.0. The capacity factor of bilirubin increases with pH below 7 in all bile salt solutions. Biliverdin and xanthobilirubin show essentially identical capacity factors for all bile salts. Biliverdin dimethyl ester and xanthobilirubin methyl ester also have very similar capacity factors, which are greater than those of the carboxy analogs, in trihydroxy bile salts. The capacity factors of these esters are higher in the dihydroxy bile salts, with the capacity factor of biliverdin dimethyl ester being twice that of xanthobilirubin methyl ester. Factors involved in the MEKC analysis of these compounds are discussed.

Micellar electrokinetic capillary chromatography analysis of the behavior of bilirubin in micellar solutions.

The capacity factor of bilirubin is determined by micellar electrokinetic capillary chromatography (MECC) techniques in three different surfactant systems. The capacity factor of bilirubin in cholic acid, taurocholic acid, and taurochenodeoxycholic acid solutions are compared to each other as a function of pH. The pH range studied is 6.5 to 9.5 which includes the pH range of bile, and includes the most likely pKa values of bilirubin carboxyl groups. MECC techniques are used to estimate these apparent pKa values for bilirubin as well as to determine the capacity factors for the separate ionization states of bilirubin in the three different surfactants. Due to the complexity of the bilirubin-bile salt system, it appears as though it is not possible to use MECC to accurately determine the bilirubin apparent pKa values. Separations are performed in 75 microns capillaries, typically 36 to 52 cm in length. UV detection, electrokinetic injection, and run voltages of 7 kV are typical. Solutions of 25 mM of each bile salt are prepared in a 20 mM phosphate-borate buffer system.

Automated instrumentation for comprehensive two-dimensional high-performance liquid chromatography of proteins.

A comprehensive two-dimensional (2-D) liquid chromatographic separation system is presented. The system uses a microbore cation exchange column, operated under gradient conditions, as the first dimension separation. Effluent from this first column alternately fills one of two loops on a computer-controlled eight-port valve. A second pump then forces loop material onto a second column, a size exclusion column. UV detection is used, and the system is applied to the separation of protein standards and serum proteins. The 2-D system has a higher resolving power and peak capacity than either of the two columns used alone. The entire first column effluent is analyzed on the second column in virtually the same time it takes to complete the first column separation, without the use of stopped flow methods. The entire system is automated and operated under computer control. Three-dimensional (3-D) data representation provides a means of viewing peak profiles in either separation dimension and contour mapping of the 3-D data provides a more reliable means of peak identification from run to run than that provided by single-column elution times.

Capillary electrophoresis of proteins in buffers containing high concentrations of zwitterionic salts.

A method for improving protein separations in capillary zone electrophoresis utilizing high concentrations of zwitterionic buffer additives was examined. Lysozyme and alpha-chymotrypsinogen A were used as test proteins in untreated fused-silica capillaries in buffers of pH ca. 7.0 and 9.0 The zwitterion-containing buffers were compared with buffers containing high ionic salt concentrations and a buffer containing a combination of high ionic salt and high zwitterion concentrations. Over 100,000 theoretical plates were obtained in less than 30 min. for both test proteins in a pH 7 buffer containing both trimethylglycine and potassium sulfate. The advantages and disadvantages of this technique compared with those of other methods used to prevent protein adsorption are discussed.

Fracture of the clavicle following radical neck dissection and postoperative radiotherapy: a case report and review of the literature.

The treatment of head and neck cancer with radiotherapy and radical neck dissection has many recognized complications. Radiotherapy in therapeutic doses can produce devascularization and weakening of bone. Radical neck dissection results in altered mechanics of the shoulder girdle and a disruption of normally balanced forces acting on the clavicle. An unusual case of clavicle fracture which is considered to have resulted from an interaction of the effects of these therapies is discussed. An approach for recognizing and distinguishing this entity by its time course, and radiographic and nuclide bone scan appearance is presented.

Prolonged orotracheal intubation in the newborn.

One hundred seventy-four consecutive cases of newborn infants who required orotracheal intubation for longer than three days were studied retrospectively. The duration of intubation varied from 3 to 211 days with an overall mortality rate of 22.9%. There was one complication directly related to orotracheal intubation. One tracheotomy was performed. Orotracheal intubation is felt to be superior to nasal tracheal intubation and tracheotomy in managing infants in this age group who require prolonged assisted ventilation.