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P B Taylor

Publications and source records attributed to P B Taylor.

At least 19 recordsLinked to original sources
PubMed1992

A comprehensive procedure for preparation of partially methylated alditol acetates from glycoprotein carbohydrates.

Various steps involved in the preparation of partially methylated alditol acetates (PMAAs) from glycoprotein-derived carbohydrates were improved to obtain the derivatives in a rapid manner with excellent yields. Carbohydrates were permethylated in dimethyl sulfoxide (DMSO), using a fine suspension of sodium hydroxide and methyl iodide (CH3I). The fine suspension of NaOH was prepared conveniently from commercially available 50% aqueous NaOH in DMSO by sonication and washing the precipitate with DMSO. Methylation of ovalbumin and fetuin glycopeptides using the fine suspension of NaOH and CH3I was complete within 5 min, and the methylation reaction did not generate any nonsugar artifacts. Methylated carbohydrates without any purification were hydrolyzed in a mixture of volatile organic acids, which permitted rapid removal of the acids from samples by evaporation. Acetylation of partially methylated alditols with acetic anhydride for 2-4 h at ambient temperature using 4-N,N'-dimethylaminopyridine as a catalyst and the reaction was free from generating nonsugar reaction artifacts. The reaction time course for methylation, hydrolysis, and acetylation was determined to obtain optimum reaction conditions for preparation of the PMAAs. The procedure facilitated rapid identification and quantitation of PMAAs due to diminished reaction artifacts and the quality of the chromatogram depended only on the purity of starting material and the reagents used for the methylation analysis. Utility of these simple methods for rapid methylation analysis was demonstrated in the characterization of oligosaccharides isolated in small amounts using a carbohydrate analyzer.

Acetates↗
PubMed1992

Myocardial stretch alters twitch characteristics and Ca2+ loading of sarcoplasmic reticulum in rat ventricular muscle.

OBJECTIVE: The aim was to determine the influence of diastolic muscle length on force development and timing parameters of cardiac muscle twitch contraction and to determine whether a length dependency exists for the calcium loading capacity of the sarcoplasmic reticulum. METHODS: Right ventricular papillary muscles and trabeculae were isolated from hearts of female Wistar rats weighing 220-280 g. Papillary muscles were stretched to diastolic lengths of 90, 95, and 100% Lmax and paced at 1.0 Hz. Individual twitch profiles were characterised by their peak force and the maximum rate (dF/dt) of the positive and negative force changes. Intrinsic timing was identified through waveform analysis that divided the twitch profile into time domains for the ascending limb (T0-T1; T1-T2) and the descending limb (T2-T3; T3-T4). Each domain was compared at three muscle lengths. The sarcoplasmic reticular calcium content at short (1.88 microns) and long (2.11 microns) sarcomere lengths was characterised by rapid cooling contractures after 1 s and 60 s of diastolic rest. RESULTS: Peak developed force and the maximum rate of positive and negative force development decreased as diastolic muscle length was reduced from Lmax to 90% Lmax. The intrinsic timing for the segment that reflects the relaxation phase of the twitch (T1-T4) was shortened as muscle length was reduced. The time domain that reflects the combined effects of calcium release and the early phase of contraction (T0-T1) was insensitive to diastolic muscle length. The fractional release of sarcoplasmic reticular calcium at different muscle lengths was approximately 32-35% of the total sarcoplasmic reticulum calcium pool. CONCLUSIONS: The data on the intrinsic timing of the twitch characteristics coupled with rapid cooling contracture analysis suggests a fractional calcium release that is approximately 32-35% of the total sarcoplasmic reticular capacity at either long or short muscle lengths. However, the loading capacity of the sarcoplasmic reticulum is greater when the muscle operates at a shorter diastolic length. This can be interpreted as meaning that diastolic muscle length differentially influences sarcoplasmic reticular calcium storage and release processes.

Animals↗
PubMed1991

Quantitative determination of phenyl isothiocyanate-derivatized amino sugars and amino sugar alcohols by high-performance liquid chromatography.

Simple and rapid methods for the preparation of phenylthiocarbamyl (PTC) derivatives of amino sugars and amino sugar alcohols and their quantitative determination with high sensitivity (less than 10 pmol) by C18 reversed-phase high-performance liquid chromatography are described. Rapid sample preparation of the phenyl isothiocyanate (PITC)-derivatized amino sugars and amino sugar alcohols was achieved by a simple extraction of the reaction mixture with chloroform to remove the excess PITC and its adducts. Baseline separation of the PTC derivatives of amino sugars and amino sugar alcohols was obtained within 30 min, using a simple solvent system consisting of 0.2% each of n-butylamine, phosphoric acid, and tetrahydrofuran. The mobile phase containing n-butylamine, in conjunction with a C18 stationary phase, mimics the conditions for the separation of carbohydrates on an amino-bonded column. GlcNH2 and GalNH2 derived from the initial protein-sugar linkages were also separated from the amino acids for quantitative estimation of sugar chains in glycoproteins. Amino sugar alcohols gave single reaction products with PITC while the reaction with amino sugars was accompanied by the formation of secondary products. Apparently the secondary products were formed in an acid-catalyzed intramolecular cyclization of the PTC-hexosamines involving the aldehyde functional group. Conditions were developed to stop the transformations and maintain the stability of PTC derivatives for their convenient determination by HPLC.

Amino Sugars↗
PubMed1991

Rapid characterization of asparagine-linked oligosaccharides isolated from glycoproteins using a carbohydrate analyzer.

Chromatographic methods were developed for the separation and characterization of acidic (sialylated) and neutral (asialo-complex and high-mannose) oligosaccharides released from glycoproteins with peptide N-glycosidase F. endo-beta-N-acetylglucosaminidase F and endo-beta-N-acetylglucosaminidase H using a carbohydrate analyzer (Dionex BioLC). All the carbohydrate separations were carried out on a polymeric pellicular anion-exchange column HPIC-AS6/CarboPac PA-1 (Dionex) using only two eluants namely, 0.5 M NaOH and 3% acetic acid/NaOH pH 5.5, which were mixed with water to generate various gradients. Developed conditions for quantitative detection of carbohydrates with pulsed amperometry were necessary to obtain steady baselines at 0.1-0.3 microA output with suitable sensitivity (less than 5 pmol) in separations employing a variety of acidic and alkaline sodium acetate gradients. Oligosaccharides released from heat-denatured and trypsin-treated glycoproteins were purified initially from large-scale digestion (greater than 0.1 g) by extraction of peptide material into phenol/chloroform and finally by ion-exchange chromatography of the acqueous phase. Oligosaccharides isolated from the peptide N-glycosidase digests of bovine fetuin, human transferrin and alpha 1-acid glycoprotein gave multiple peaks in each charge group in separations based on the charge content at pH 5.5. Alkaline sodium acetate gradients were developed to obtain oligosaccharide maps of the glycoproteins within 60 min, in which separated oligosaccharides eluted in the order of neutral, mono-, di-, tri- and tetra-sialylated species based on both charge, size and structure. Baseline separations were obtained with neutral oligosaccharide types but mixtures of high-mannose and complex types were poorly resolved. The high-mannose peaks were eliminated specifically from complex oligosaccharides by digesting with alpha-mannosidase. Treatment with beta-galactosidase, beta-N-acetylglucosaminidase and alpha-mannosidase resulted in a decrease of the oligosaccharide elution times corresponding to the number of sugar residues lost, the profile of changes was highly reproducible. In contrast, treatment with alpha-L-fucosidase, endo-beta-N-acetylglucosaminidase F and endo-beta-N-acetylglucosaminidase H resulted in an increase in their corresponding oligosaccharide retention times similar to the presence of an additional sugar residue. Conditions developed for separation of the reduced oligosaccharides and also a mixture of monosaccharide to oligosaccharide containing about 15 sugar residues within 30 min were useful in determining the effect of endo- and exo-glycosidases on porcine thyroglobulin oligosaccharides. Changes in elution time of the oligosaccharides following specific glycosidase digestions combined with methylation analysis provided a rapid and sensitive tool for confirmation of the carbohydrate primary structures present in thyroglobulin.

Animals↗
PubMed1991

Structural classification of carbohydrates in glycoproteins by mass spectrometry and high-performance anion-exchange chromatography.

A general strategy has been developed for determining the structural class (oligomannose, hybrid, complex), branching types (biantennary, triantennary, etc.), and molecular microheterogeneity of N-linked oligosaccharides at specific attachment sites in glycoproteins. This methodology combines mass spectrometry and high-performance anion-exchange chromatography with pulsed amperometric detection to take advantage of their high sensitivity and the capability for analysis of complex mixtures of oligosaccharides. Glycopeptides are identified and isolated by comparative HPLC mapping of proteolytic digests of the protein prior to, and after, enzymatic release of carbohydrates. Oligosaccharides are enzymatically released from each isolated glycopeptide, and the attachment site peptide is identified by fast atom bombardment mass spectrometry (FAB-MS) of the mixture. Part of each reaction mixture is then permethylated and analyzed by FAB-MS to identify the composition and molecular heterogeneity of the carbohydrate moiety. Fragment ions in the FAB mass spectra are useful for detecting specific structural features such as polylactosamine units and bisecting N-acetylhexosamine residues, and for locating inner-core deoxyhexose residues. Methylation analysis of these fractions provides the linkages of monomers. Based on the FAB-MS and methylation analysis data, the structural classes of carbohydrates at each attachment site can be proposed. The remaining portions of released carbohydrates from specific attachment sites are preoperatively fractionated by high-performance anion-exchange chromatography, permethylated, and analyzed by FAB-MS. These analyses yield the charge state and composition of each peak in the chromatographic map, and provide semiquantitative information regarding the relative amounts of each molecular species. Analytically useful data may be obtained with as little as 10 pmol of derivatized carbohydrate, and fmol sensitivity has been achieved. The combined carbohydrate mapping and structural fingerprinting procedures are illustrated for a recombinant form of the CD4 receptor glycoprotein.

Amino Acid Sequence↗
PubMed1991

Inotropic interventions and myocardial force-interval relation: a quantitative approach.

OBJECTIVES: To analyze inotropic influence on the early and late phases of cardiac sarcoplasmic reticulum calcium loading. DESIGN: Papillary muscles with parallel edges and no evidence of tissue branching were selected from the heart. Only muscle preparations that maintained stable passive diastolic and developed forces were used for analysis. Muscles were stretched to their maximum length and stimulated at 0.2 Hz. The early and late phases of sarcoplasmic reticulum calcium loading were evaluated quantitatively by mathematical fitting of the force-interval relation. Increasing the extracellular calcium or decreasing the extracellular sodium was used to increase the inotropic state. ANIMALS: Right ventricular papillary muscles were isolated from female Wistar rats weighing 200 to 220 g. Electrical stimulation and data acquisition were controlled through a microcomputer. MAIN RESULTS: Increasing the extracellular calcium concentration from 0.5 to 1.0 mM produced a 90% increase in developed tension. This was accounted for by a 41% increase in the early phase of sarcoplasmic reticulum loading and a 29% increase in the late phase. A 20% reduction in the extracellular sodium concentration increased contractile force 100% and shifted the force-interval curve to the left. This was accounted for through an increase in both early and late phases of sarcoplasmic reticulum loading. CONCLUSION: These results are consistent with the current model of excitation-contraction coupling and clearly indicate that various positive inotropic interventions have selective effects on each process of the force-interval relation that cooperatively interact with each other. Mathematical fitting of data clearly improves the quantitative aspect of the force-interval response.

Animals↗
PubMed1990

Spiritual healing.

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Attitude to Death↗
PubMed1990

Pathologic observations made by retinal biopsy.

The authors report four cases in which retinal biopsy findings yielded unexpected or previously unreported diagnoses in patients with inflammatory retinitis. The tissue diagnoses included Wegener's retinal vasculitis in an immunosuppressed patient with a clinical diagnosis of cytomegalovirus retinitis, a novel viral form in the retina of a patient with cytomegalovirus retinitis, a case of acute retinal necrosis due to cytomegalovirus infection in an immunologically normal adult, and a case of ganciclovir-resistant herpes family viral retinitis. These cases illustrate the use of retinal biopsy in obtaining tissue for diagnosis and guiding treatment in selected cases of retinitis.

Acquired Immunodeficiency Syndrome↗
PubMed1989

Effect of catecholamine-induced cardiac hypertrophy on the force-interval relationship.

Cardiac hypertrophy was induced in adult female Wistar rats following 12 days of daily subcutaneous injections of isoproterenol (ISO). The left atria responded with a 13-14% increase in tissue growth, while the ventricles achieved a 34-39% increased tissue mass. Maximum force generation and twitch characteristics in 1.0 mM external Ca2+ for the left atria or the right papillary muscle were unchanged in the ISO-treated animals. The force-interval relation was determined at 26 degrees C between 0.5 and 120 s. The development of maximum force clearly passed through two phases identified as alpha and beta. To characterize these two processes the data were fitted to a two-term linear combination of exponentials (two-compartment model). The time constant and capacity of each process to contribute to the whole force-interval curve was determined by a four-parameter least square fit method. In control atrial muscle the time constants for the alpha and beta processes were 0.47 and 11.23 s, respectively. The contribution of each process to the total force curve in control atrial muscle was approximately 50% alpha and 50% beta. Following ISO-induced growth the time constants were 0.38 and 13.33 s with a shift of contributions towards 60% alpha and 40% beta. Control papillary muscle from the right ventricle had a similar alpha time constant of 0.49 s compared with atrial muscle but possessed a considerably slower beta time constant of 26.17 s. The contribution of each process to interval-dependent force development was 44.5 and 55.5%, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1988

Conjunctival lymphocyte subsets in trachoma.

Monoclonal antibodies were used to assess the lymphocyte populations in conjunctival biopsy specimens obtained from patients with trachoma (active or inactive) undergoing tarsotomy for the correction of trachoma-induced entropion and in three control patients. Peroxidase-labelled monoclonal antibodies OKT4 (identifies T-helper/inducer lymphocytes), OKT8 (identifies T-suppressor/cytotoxic lymphocytes), OKIal (identifies B-lymphocytes) and antisera specific for IgG, IgA and IgM were used to identify lymphocyte subpopulations and immunoglobulins in the conjunctival biopsy specimens. When grouped by disease activity, conjunctival tissue specimens revealed predominant T-helper/inducer lymphocytes in the substantia propria of patients with active trachoma while inactive trachoma patients had predominant T-suppressor/cytotoxic lymphocytes in the conjunctival biopsy specimens. B-lymphocytes were seen in moderate numbers in all conjunctival biopsy specimens from patients with active trachoma and all specimens from active cases stained for IgG, IgM, and IgA. Immunoglobulin staining was strongest with IgG and IgM.

Adult↗
PubMed1988

Minocycline effects in patients with active trachoma.

Minocycline has a unique solubility in lipids and may reach therapeutic concentrations in tears and saliva. In two consecutive prospective double-masked clinical trials that were carried out in two villages in Saudi Arabia, we assessed the effects of oral minocycline in the treatment of trachoma and compared its effects with those of topical tetracycline ointment in the first study and to tetracycline ointment and placebo in the second study. A total of 178 eyes in 96 patients were included. The age range was 7 to 14 years, with a mean age of 9 years. All patients underwent complete ophthalmologic evaluation. School children were divided into two groups in a double-masked fashion. The first group received either oral minocycline or topical tetracycline 1% ointment and the second group was divided into three subgroups, each receiving one of the following therapeutic modalities: oral minocycline, topical tetracycline ointment, or placebo ointment. All patients were evaluated before initiation of therapy, at three weeks and at 12 months following treatment. Therapy was continued for a period of five weeks. These two double-masked field-based clinical trials have shown both minocycline given orally and tetracycline ointment given topically were effective in decreasing the intensity of inflammation due to trachoma. Oral minocycline was found to be equally effective as topical tetracycline ointment in the treatment of trachoma at three weeks. Minocycline, however, was found to be superior to topical tetracycline when patients were evaluated one year after therapy (p less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Administration, Oral↗
PubMed1988

Monoclonal antibodies in the laboratory diagnosis of trachoma.

Various techniques which use monoclonal antibodies to detect Chlamydia trachomatis in clinical specimens are reviewed. An investigation comparing the efficacy of immunofluorescent staining with Giemsa staining in detecting Chlamydia in conjunctival scrapings from cases of active trachoma is presented. Sixty-two eyes of schoolboys with moderate to severe trachoma were studied. Giemsa staining detected chlamydial inclusion bodies in 34 percent of the specimens. Free elementary bodies were detected by fluorescent monoclonal antibody in 21 percent. Eleven percent were positive by both Giemsa and immunofluorescence and 55 percent were positive by either Giemsa and/or immunofluorescence. The addition of fluorescent monoclonal antibody assay to routine Giemsa staining resulted in an increase in the yield of positive specimens by 29 percent.

Antibodies, Monoclonal↗
PubMed1988

Effect of preoperative fusidic acid on the normal eyelid and conjunctival bacterial flora.

A randomised trial comparing the topical application of 1% fusidic acid with 0.3% gentamicin solution in the reduction of the normal preoperative lid and conjunctival microbial flora was performed. Forty patients awaiting cataract surgery were randomly divided into two groups consisting of 20 patients each. The first group received a 1% microcrystalline suspension of fusidic acid, the second 0.3% gentamicin to the preoperative eye every two hours between 0600 and 2400 daily for 48 hours preoperatively. Cultures were obtained from both the lid margins and the conjunctival sac of both groups prior to antibiotic therapy and again in the operating theatre before surgery. Microbiological identification and colony counts were performed by standard laboratory methods. Staphylococcus epidermidis was the commonest micro-organism isolated. Statistical analysis revealed no significant differences in the ability of a 1% microcrystalline suspension of fusidic acid and 0.3% gentamicin in eliminating or reducing the normal preoperative conjunctival or lid flora.

Administration, Topical↗
PubMed1987

Corneal and intraocular penetration of topical and subconjunctival fusidic acid.

Corneal tissue absorption and intraocular penetration of fusidic acid were assessed in the rabbit after topical or subconjunctival application. Corneal tissue levels of fusidic acid one hour after the last topical application of the drug were well above the minimum inhibitory concentrations (MICs) for most Gram-positive and many Gram-negative organisms. Adequate levels were achieved in the aqueous at one hour following the last topical application, but no significant levels were detected in the vitreous. The corneal tissue and aqueous levels declined at 12 and 24 hours following the last drug application, however, corneal tissue levels at 24 hours were considered to be above the MICs for most Gram-positive organisms. A single subconjunctival injection of 100 mg of fusidic acid produced levels above the MICs of most organisms in the cornea, aqueous, and vitreous which persisted over 24 hours, but subconjunctival injection of fusidic acid at this concentration resulted in conjunctival necrosis and corneal decompensation. Fusidic acid penetrates well into avascular tissue and fully penetrates corneas with both intact and debrided epithelium, as evidenced by the intracameral drug levels. Good corneal penetration and absence of known topical toxicity make fusidic acid suitable for the treatment of microbial keratitis caused by susceptible organisms.

Absorption↗
PubMed1987

Catecholamine induced cardiac hypertrophy.

Cardiac hypertrophy was induced in adult female Wistar rats by daily subcutaneous injections of isoproterenol (0.3 mg/kg body weight). Heart weight increased 39% after eight days of treatment. Left ventricular pressure development (positive dP/dt) in hearts four days after hypertrophy induction was significantly increased, while negative dP/dt remained unchanged. RNA polymerase activity in isolated myocyte and nonmyocyte nuclei was stimulated 29 and 23%, respectively 24 h after a single isoproterenol injection. In the myocyte fraction, RNA polymerase activation progressively increased up to four days of treatment and then returned to control values after eight days. In the nonmyocyte nuclear subset, RNA polymerase activity showed no further stimulation and gradually returned to control values after eight days of treatment. Chromatin template function was substantially stimulated in the early stage (one to four days) of hypertrophy in both myocyte and nonmyocyte fractions. Titration of chromatin against a fixed amount of RNA polymerase (5 micrograms) in the presence of rifampicin and heparin showed that less chromatin from hypertrophied hearts was required to saturate the enzyme. These results indicate that both myocyte and nonmyocte chromatin from hypertrophied hearts can support greater enzyme binding than normal chromatin. The alkaline sucrose density centrifugation profile of DNA in myocyte and nonmyocyte chromatin from day 4 hypertrophied hearts was less fragmented. These observations suggest that during the early phase of isoproterenol-induced cardiac hypertrophy, enhanced RNA polymerase activity and chromatin template function play a coordinated role in RNA synthesis. The increased template activity could be due to alterations in chromatin composition which was indicated by the change in their enzyme binding capacity and DNA fragmentation profile.

Animals↗
PubMed1987

Precision of the doubly labelled water method using the two-point calculation.

The intrasubject precision of the doubly labelled water method was investigated as a function of increasing length of the metabolic period and decreasing initial isotope enrichment. This analysis was performed in seven subjects following a single dose of isotope and three subjects who received multiple doses of isotope. The intrasubject precision was 5-6 per cent for periods of up to 14 d. This is only slightly larger than that predicted from our previously published model in which we assumed that analytical error was limiting precision. Intrasubject precision worsened with increasing length of the metabolic period and with decreasing initial isotope enrichment, but the loss of precision was two to ten times greater than that predicted by the model. The use of the intercept method to determine the isotope dilution spaces did not improve precision. We speculate that one cause of the failure of the model is that it does not consider unmeasured changes in natural isotopic abundances associated with dietary intake. Results demonstrate the importance of choosing a dose that provides an initial isotope enrichment that is 500-600 times the analytical error, and a metabolic period that is between one and three biological half-lives of the isotopic tracers.

Adult↗