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In vivo and in vitro destruction of rat liver cytochrome P-450 by a monoterpene ketone, pulegone.

In vivo administration of pulegone once daily decreased the levels of liver microsomal cyt. P-450 to the extent of 32 and 76% at the end of 24 and 96 hrs respectively. However, cyt. b5 and NAD(P)H-cyt. c reductase activities remained unchanged. In vitro incubation (15 min) of liver microsomes from phenobarbitol (PB)-treated rats with pulegone (10 mM), aerobically or anaerobically resulted in the loss (approximately 60%) of cyt. P-450 in the presence or absence of NADPH. Destruction of cyt. P-450 was more in PB-treated microsomes as compared to 3-methylcholanthrene (MC)-treated and control microsomes. The loss of cyt. P-450 was accompanied by a concomitant loss of microsomal heme. In contrast, menthone or carvone upon incubation with PB-induced microsomes resulted in the conversion (25-40%) of cyt. P-450 to cyt. P-420 without any loss of microsomal heme. The destructive process is irreversible, time dependent, linear upto a substrate concentration of 10 mM and follows first order kinetics.

Administration, Oral↗

Intracellular potentials of salamander mitral/tufted neurons in response to odor stimulation.

Intracellular responses of salamander mitral/tufted neurons to defined odor pulses are described. Responses to odor stimulation, which generally are more complex than responses to olfactory nerve or tract electrical stimulation, show periods of depolarization and hyperpolarization that are influenced by odor concentration and quality. The way these periods coincide with different types of spike patterns in individual cells supports the hypothesis that the temporal patterning of odor responses is generated by differential activation of bulbar circuits.

Animals↗

Pulegone mediated hepatotoxicity: evidence for covalent binding of R(+)-[14C]pulegone to microsomal proteins in vitro.

Incubation of R(+)-[14C]pulegone with rat liver microsomes in the presence of NADPH resulted in covalent binding of radioactive material to macromolecules. Covalent binding was much higher in phenobarbital-treated microsomes as compared to 3-methylcholanthrene treated or control microsomes. The Km and Vmax of covalent binding was 0.4 mM and 1.7 nmol min-1 mg-1, respectively. Covalent binding was drastically inhibited (93%) in the presence of piperonyl butoxide. Antibodies to phenobarbital-induced cytochrome P-450 and NADPH-cytochrome P-450 reductase inhibited covalent binding to an extent of 72% and 47%, respectively. Cysteine and semicarbazide also inhibited NADPH dependent binding of radiolabel from R(+)-[14C]pulegone to microsomal proteins. The results suggest the involvement of liver microsomal cytochrome P-450 in the bioactivation of R(+)-pulegone to reactive metabolite(s) which might be responsible for covalent binding to macromolecules resulting in toxicity.

Animals↗

A radiographic evaluation of the periapical status of teeth treated by the gutta-percha--eucapercha endodontic method: a one-year follow-up study of 458 root canals. Part II.

In Part I, which appeared in the last issue, the authors discussed the variations that exist in nonsurgical endodontic therapy with respect to methods of treatment and analyses of success and failure. They then reported on the materials and methods that were used in their clinical study of 458 root canals treated by the gutta-percha--eucapercha method and gave the criteria that were used for the radiographic analysis of success and failure in those cases. In Part II, the results of the one-year follow-up of those 458 root canals is presented. Tables and illustrations are used to substantiate the results.

Dental Pulp Cavity↗

A radiographic evaluation of the periapical status of teeth treated by the gutta-percha-eucapercha endodontic method: a one-year follow-up study of 458 root canals. Part III.

In Part I which appeared two issues previously, the authors discussed the variations in treatment methods of nonsurgical endodontic therapy and in the determination of success and failure. They then reported on the materials and methods used and the radiographic criteria for success and failure in their clinical study of 458 root canals treated by the gutta-percha-eucapercha method. In Part II which appeared in the last issue, the one-year follow-up results were presented. Some of these results differ from previous investigations. For example, better success was found in (1) necrotic cases with areas of pathosis than in necrotic cases without "areas and in (2) overfilled cases than in underfilled cases. In Part III the authors discuss the possible reasons for these and other findings and give some implications for clinical practice.

Biocompatible Materials↗

A comparative tissue toxicity evaluation of established and newer root canal sealers.

Grossman's sealer, eucapercha, Endo-Fill, CRCS, Sealapex, Hypocal, and sterile saline solution (0.3 ml of each) were injected into specific dorsal subdermal tissue sites of 12 guinea pigs. The animals were killed after 6 days, 15 days, and 80 days (four per time period). Analysis of tissue response showed that, overall, Sealapex and Endo-Fill had less severe inflammatory reactions than any of the other test materials. Grossman's sealer, CRCS, and Hypocal showed principally severe inflammatory responses at both 6 and 15 days, but mild reactions at 80 days. Overall, eucapercha showed less severe inflammatory responses than Grossman's sealer, CRCS, and Hypocal. Diffuse calcification was induced by the three calcium hydroxide preparations (CRCS, Sealapex, and Hypocal). Eucapercha and Endo-Fill had minute local areas of calcification. Both Grossman's sealer and CRCS did not have overall favorable histologic reactions; however, Grossman's sealer and CRCS have been used successfully clinically. Further clinical studies are needed.

Animals↗

Temporal integration and reaction times in human smell.

A model description of intensity perception in human taste and smell developed earlier has now been verified experimentally to determine parameter values for odorants. The final objective is to quantify and understand odour-odour interaction phenomena in e.g., masking, deo-perfumes and flavour enhancement. Five types of olfactometer experiments were carried out, viz. determination of thresholds, determination of reaction times, scaling of perceived intensity after 5 sec stimulation, scaling of perceived intensity of a fixed concentration at variable duration, and measurement of intensity/time relationships. Four subjects were used and the odorants cineole, geraniol and hexane.

Acyclic Monoterpenes↗

Linalool and cineole type glucosides from Cunila spicata.

The leaves of Cunila spicata yielded a monoterpenetriol and six glycosidic terpenoids derived from linalool, hydroxylated linalool and 1,8-cineole: 3,7-dimethyl-oct-1-ene-3,6,7-triol, linalool-O-beta-D-glucopyranoside, 3,7 dimethyl-octa-1,6-diene-3,8-diol-3-O-beta-D-glucopyranoside as well as 3,7-dimethyl-octa-1,5-diene-3,7-diol-3-O-beta-D-glucopyranoside, 3,7-dimethyl-octa-1,7-diene-3,6-diol-7-O-beta-D-glucopyranoside, 3,7- dimethyl-oct-1-ene-3,6,7-triol-6-O-beta-D-glucopyranoside and (1S,4R,6R)-1,3,3-trimethyl-2-oxabicyclo[2.2.2]octan-6-O-beta- glucopyranoside. The structures of the glucosides were established by chemical and spectroscopic methods especially high field NMR techniques.

Acyclic Monoterpenes↗

Effect of phenolic compounds on the polymerization of methyl methacrylate.

The inhibitory effects of six phenolic compounds and two analogues on the polymerization of methyl methacrylate (MMA) by azobisisobutyronitrile (AIBM) were examined. Differential scanning calorimetry was used to determine the induction period (IP) and initial rate of polymerization (IRP). The IP values (minutes) decreased in the order: eugenol > thymol > hydroquinone > cresol > guaiacol > phenol >>> safrole > methol. The IRP values of all compounds tested ranged from 0.8 to 1.2 percent per minute, except for hydroquinone (0.2 percent per minute). Phenolic compounds inhibited the polymerization of MMA by scavenging radicals, and eugenol was the most potent inhibitor.

Cresols↗

Revalidation of the in vitro alkaline elution/rat hepatocyte assay for DNA damage: improved criteria for assessment of cytotoxicity and genotoxicity and results for 81 compounds.

The in vitro alkaline elution/rat hepatocyte assay is a sensitive assay for genotoxicity, measured as DNA strand breaks induced in primary cultures of rat hepatocytes after 3-h treatments with test compounds. Since DNA degradation can be rapid and extensive in dead and/or dying cells, the original criteria for a positive result in the assay were that a compound induce a 3.0-fold or greater increase in the elution slope (for the terminal phase of alkaline elution from 3 to 9 h) in the absence of significant cytotoxicity (defined as relative cell viability of less than 70% by trypan blue dye exclusion; TBDE). Recently we have shown that false-positive results can still be obtained due to cytotoxicity when loss of membrane integrity is a late event in toxic cell death relative to the induction of endonucleolytic DNA degradation. To improve the ability of the assay to discriminate between genotoxic vs. cytotoxic effects of chemicals, we have evaluated additional assays of cytotoxicity including cell adenosine triphosphate (ATP) and potassium (K+) content, tetrazolium dye reduction (MTT), TBDE after a further 3-h recovery incubation without test chemicals (delayed toxicity), cell blebbing and endonucleolytic DNA degradation (double-strand breaks; DSBs) assessed by pulsed-field gel electrophoresis (PFGE). We have also evaluated 2 parameters derived from the elution data which can indicate extensive, cytotoxicity-induced DNA degradation: the fraction of the DNA recovered in the neutral lysis/rinse fraction and the gamma-intercept of the extrapolation of the 3-9-h segment of the elution curve. Twenty-eight rodent non-carcinogens that are negative (or inconclusive) in the Ames assay with no, or limited, other evidence of genotoxicity, and 33 genotoxins, most of which are also carcinogens, were evaluated. The results showed that DNA degradation as measured by a 1-h PACE (Programmed Autonomously Controlled Electrodes)/PFGE assay was a sensitive indicator of cytotoxicity which correlated well with results of the other cytotoxicity indicators. The delayed TBDE (after a 3-h recovery), intracellular potassium and ATP assays as well as the gamma-intercept parameter were also shown to be sensitive and in some cases complementary measures of cytotoxicity. Using new criteria based on these data of an induced slope (treatment slope-negative control slope) of 0.020 for the 3- to 9-h elution period and cytotoxicity limits of 70% relative viability for the delayed TBDE assay and 50% for intracellular ATP content, the assay scores the genotoxicity of these 61 reference compounds with an overall accuracy of 92%. Test results using these new criteria are provided for an additional 20 compounds (5 non-genotoxic carcinogens and 15 compounds whose genotoxic and carcinogenic potential are unknown or equivocal).

Adenosine Triphosphate↗