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Biomedical subjects

S S Ghosh

Publications and source records attributed to S S Ghosh.

At least 55 records · Page 3Linked to original sources
PubMed1990

Inhibition of human phospholipases A2 by cis-unsaturated fatty acids and oligomers of prostaglandin B1.

Inhibition of human phospholipases A2 by cis-unsaturated fatty acids and their oxidative metabolites and/or polymers was studied using partially purified human phospholipases A2 and [1-14C]oleate labelled, autoclaved E. coli as substrate. As previously reported for other phospholipases A2, oleic and arachidonic acids inhibited human synovial fluid phospholipase A2 with IC50s of 15 and 30 microM respectively. Air oxidation of arachidonic acid or hydroxylation of oleic acid (12-hydroxy-oleate) substantially relieved that inhibition. Similarly, the enzymatically oxidatized metabolite of arachidonate, prostaglandin B1 (PGB1), did not inhibit enzymatic activity. However, prostaglandin Bx (PGBx), an oligomer (n = 6) of PGB1, was a potent inhibitor of Ca(++)-dependent, neutral-active phospholipase A2 activities. Enzymatic activity in acid extracts from human neutrophils, platelets, sperm, plasma, synovial fluid, endometrium, degenerative disc, and snake venom was inhibited by PGBx with IC50s ranging from 0.5-7.0 microM. Inhibition was independent of substrate phospholipid concentration over a 24-fold range (5-120 microM) and PGBx quenched the tryptophan fluorescence of snake venom phospholipase A2 in a dose-dependent manner. Agonist-induced (A23187) release of arachidonic acid from prelabelled human neutrophils and cultured human endothelial cells was also inhibited by PGBx with IC50s of 3 and 20 microM, respectively. These results illustrate that oxidative reactions of cis-unsaturated fatty acids relieve their natural inhibitory activity, and polymerization of an inactive fatty acid metabolite yields a potent inhibitor of in vitro and in situ phospholipase A2 activity.

Arachidonic Acid↗
PubMed1990

Design of an effective mechanism-based inactivator for a zinc protease.

(R)-2-Benzyl-5-cyano-4-oxopentanoic acid (compound 4) was studied as a mechanism-based inactivator (suicide substrate) for the zinc protease carboxypeptidase A (CPA; peptidyl-L-amino-acid hydrolase, EC 3.4.17.1). This compound was designed rationally based on the knowledge of the active site topology and the reported stereospecific proton exchange on ketonic substrate analogue (R)-3-(p-methoxybenzoyl)-2-benzylpropanoic acid [Sugimoto, T. & Kaiser, E. T. (1978) J. Am. Chem. Soc. 100, 7750-7751] by CPA. It is suggested that enzymic deprotonation on the C-5 methylene moiety may result in the transient formation of a ketenimine as the key intermediate that partitions between turnover and enzyme inactivation. The enzyme inactivation exhibited pseudo-first-order kinetics, was irreversible, and could be fully prevented in the presence of the reversible inhibitor benzyl-succinate. The inactivation rate constant, kintact, was evaluated to be 0.083 +/- 0.003 min-1 and kcat was measured at 1.78 +/- 0.06 min-1. In turn, a partition ratio of 28 +/- 3 was calculated. The reversible inhibitor constant (Ki) was measured at 1.8 +/- 0.5 microM, indicative of a high affinity for compound 4 shown by CPA; however, Km for the turnover process was determined at 4.93 +/- 0.43 mM. Kinetic analysis and labeling by the radioactive form of the inactivator suggested that the stoichiometry for protein modification by compound 4 approaches a 1:1 ratio.

Carboxypeptidases↗
PubMed1990

Detection of human immunodeficiency virus type 1 in AIDS patients using amplification-mediated hybridization analyses: reproducibility and quantitative limitations.

Eighty-six peripheral blood mononuclear cell (PBMC) samples from 30 patients with AIDS were analyzed using a transcription-based amplification system (TAS) and the polymerase chain reaction (PCR). Human immunodeficiency virus type 1 (HIV-1) sequences were detected by amplification-mediated hybridization in 98% of the samples, 52% of which were positive for p24 antigen by ELISA. Neither TAS (93%) nor PCR (95%) detected HIV-1 sequences in all 86 samples. The hybridization-detection methods (slot blot, bead-based sandwich, and solution) used to detect the HIV-1-specific TAS products had a clear influence on the efficiency of detecting and quantitating the levels of HIV-1 present in these samples. The reproducibility of amplification of constant amounts of HIV-1 RNA and beta-globin DNA by TAS and PCR was studied over 3 months. The results indicated that variations of 10- and 5-fold in the HIV-1 sequence levels could be detected between samples by TAS and PCR, respectively. Within the range of sensitivities for each assay used, the administration of zidovudine did not appear to reduce the amount of HIV-1 nucleic acid sequences as observed in PBMC obtained serially from six AIDS patients.

Acquired Immunodeficiency Syndrome↗
PubMed1989

Synthesis of 5'-oligonucleotide hydrazide derivatives and their use in preparation of enzyme-nucleic acid hybridization probes.

A hydrazone-based method for conjugating synthetic nucleic acids and reporter molecules for use as nonradioactive hybridization probes is presented. Oligonucleotides complementary to the hepatitis B virus were derivatized at their 5' ends with hydrazine or homobifunctional acyl hydrazides. These derivatives reacted facilely with aldehydes to give hydrazones, which were characterized by uv spectroscopy and HPLC. Coupling of aldehyde-modified alkaline phosphatase with carbohydrazide-oligonucleotide derivatives provided a mixture of two enzyme-nucleic acid conjugates in 80-85% yield. The conjugates had a 1:1 and a 2:1 oligonucleotide/enzyme ratio, respectively, and were separated by ion-exchange chromatography. Both conjugates were able to detect 7 amol of target DNA in 1 h, using a colorimetric assay. In contrast, oligonucleotide-horseradish peroxidase conjugates were 40-fold lower in sensitivity of detection.

Alkaline Phosphatase↗
PubMed1989

Comparative efficacy of four vaccines against Salmonella virchow in chicks in India.

The efficacy of four vaccines in preventing Salmonella virchow infection in poultry was assessed by determining survivors after challenge, bacteriological status of tissues and excretion through stool and eggs following artificial infection in vaccinated and control birds. The study indicated that formol killed oil-adjuvant vaccine conferred the highest degree of immunity followed by gel and Freund's complete adjuvant vaccines.

Animals↗
PubMed1987

Covalent attachment of oligonucleotides to solid supports.

Coupling efficiencies for the covalent attachment of oligonucleotides (17-29 bases in length) to solid supports derivatized with alkyl-amino and -carboxylic functionalities have been determined. Attachment efficiencies of 60-80% were obtained for coated long-chain alkylamino controlled pore glass (CPG) supports. Similar efficiencies of immobilization were observed for carboxyl-bearing supports, which additionally exhibited lower levels of non-covalent binding. The extent of terminally linked oligonucleotide was determined to be 50-55% of the overall attachment in the carbodiimide-mediated coupling reaction of a 5'-aminohexyl phosphoramidate derivative of a 29-mer to Sephacryl carboxyl support. While lower overall efficiencies of attachment were obtained in the reaction with Sephacryl N-hydroxysuccinimide-activated carboxyl support, greater than 80% of this coupling results in end-attached oligonucleotides.

Chemical Phenomena↗
PubMed1987

Effect of tolbutamide treatment on the pharmacokinetics of intravenously administered sulphamethoxazole in rabbits.

Fifteen days of tolbutamide treatment significantly decreased the elimination half life (t1/2), area under the curve (AUC0----infinity) and increased the clearance of sulphamethoxazole (SMZ) in rabbits. No significant difference was observed in the volume of distribution. The percentage of plasma protein binding to SMZ was not altered, while N-acetyltransferase activity in liver and kidney was significantly increased after tolbutamide therapy. The changes observed in the pharmacokinetic parameters of SMZ after tolbutamide treatment is due to the induction of liver N-acetyltransferase activity.

Animals↗
PubMed1986

Modification of the active site of alkaline phosphatase by site-directed mutagenesis.

The catalytically essential amino acid in the active site of bacterial alkaline phosphatase (Ser-102) has been replaced with a cysteine by site-directed mutagenesis. The resulting thiol enzyme catalyzes the hydrolysis of a variety of phosphate monoesters. The rate-determining step of hydrolysis, however, is no longer the same for catalysis when the active protein nucleophile is changed from the hydroxyl of serine to the thiol of cysteine. Unlike the steady-state kinetics of native alkaline phosphatase, those of the mutant show sensitivity to the leaving group of the phosphate ester.

2,4-Dinitrophenol↗
PubMed1986

Comparative pharmacokinetic study of four different sulfonamides in combination with trimethoprim in human volunteers.

The pharmacokinetics of four different sulfonamides i. e., sulfamethoxazole (SMZ). Sulfamoxole (SMO), Sulfadiazine (SDZ) and Sulfadimidine (SDD) in combination with trimethoprim (TMP) were studied in 12 healthy volunteers. Plasma and urine concentrations of sulfonamides were measured at different time intervals. No significant difference was observed in the area under the plasma curve (AUC) of SMZ, SMO and SDZ, while AUC of SMO was significantly higher than SDD only. Free (unmetabolized) SDZ urinary excretion during a 10-25 h period was significantly higher than SMZ, SMO and SDD. The results suggest that SDZ alone or in combination with TMP would be more effective in urinary tract infections as compared to other sulfonamides studied.

Adult↗
PubMed1986

Assessment of bio(in)equivalence of deriphyllin-digoxin in human volunteers. II. Evaluation of rabbits as qualitative animal model.

In rabbits, a three way cross-over test was carried out to assess bioavailability of digoxin from commercially available 'Deriphyllin-Digoxin' tablets. The in vitro dissolution test showed that these tablets had low dissolution even at the end of 4 hr. The in vivo tests in rabbits compared bioavailability of digoxin from Deriphyllin-Digoxin tablets with that from Lanoxin tablets and intravenous digoxin injection. The treatments were given in randomized order with a minimum of 14 days wash-out period between the treatments. After the drug administrations, periodic blood samples were collected and plasma digoxin concentrations were analysed using radioimmunoassay. As indicated by the results of in vitro dissolution tests, Deriphyllin-Digoxin tablets showed poor and delayed absorption of digoxin in vivo. A parallel study on comparative bioavailability for the same batches of digoxin tablets was also carried out in human volunteers. The study in human volunteers involved 14 subjects and had a cross-over dosing. The bioavailability results in rabbits were qualitatively similar to human bioquivalence studies. This is the first report showing digoxin bioavailability in rabbits corresponding to that in humans. The importance of the rabbit as a secondary model for bioequivalence testing of digoxin formulations has been emphasized.

Administration, Oral↗
PubMed1986

Assessment of bio(in)equivalence of deriphyllin-digoxin in human volunteers.

Bioavailability of digoxin from the formulations of a fixed dose combination of the glycoside with xanthines was compared with that of "Lanoxin". The in vitro analysis of the fixed dose "Deriphyllin-Digoxin" tablets showed that these tablets had low dissolution even at the end of four hours. The in vivo study had a randomized cross-over design with a 14 days wash-out period. The formulations were administered to 14 healthy adult volunteers and periodic blood samples were collected up to 24 hours. The samples were analyzed for digoxin concentration using radioimmunoassay. Results indicate poor and delayed absorption of digoxin from the fixed dose combination. It is concluded that a multiple dose study with pharmacodynamic assessment, in patients or in volunteers, would be adequate to critically reassess the need for the marketed fixed dose combination.

Adult↗