Inhibitory Effect Of Chloramphenicol Research Articles

Evaluation of days-dependent chloramphenicol dosage on rat liver microsomal lipid peroxidation and catalase activity

Drugs and chemical agents can alter the cellular functions associated with the oxidative metabolism, thereby stimulating ROS production. The objective of the this study was the in vivo study of the inhibitory effect of chloramphenicol on hepatic microsomal enzyme system and the effect on lipid peroxidation using the thiobarbituric acid (TBA) reactive as index of peroxidation damage was investigated. The rats were randomly divided into 3 groups; group 1 serves as the control receiving saline water, group 2 receive chloramphenicol at a dose of 28.6 mg/kg body weight/day for 5 days and group 3 for 7 days. Liver protein content, lipid hydroperoxides and catalase activity were all determined. TBARS concentration increased statistically significantly with time of exposure. After 5 days of treatment, the level of TBARS increased by 103.56% with exogenous oxidant and by 141.54% for without oxidant, as compared to the respective control. On day 7, TBARS level in the liver was approximately 90.75% higher than in the control group with oxidant and 117.03% higher than the control without oxidant. The antibiotic elicited significant increase in rat liver lipid peroxidation compared to control. There were decreases in microsomal protein content of the liver in the drug-treated rats when compared with the control rats. It also showed that chloramphenicol treatment affects cytosolic catalase activities. It decreases catalase levels by 15.75% in 5 days dosage treatment and 17.81% in 7 days dosage treatment respectively. There were significance difference (P<0.05) in the treated and control irrespective of the days of dosage. The catalase level in hepatocyte in 5 days and 7 days group was significance at p<0.01. In conclusion, it was evident that treatment of rats for 5 days and 7 days with the therapeutic doses of Chloramphenicol altered antioxidant system and the intensity of effects was concentration/dosage days dependent, it resulted in membrane lipid peroxidation, protein damage, and inhibition of microsomal catalase due to increased generation of free radicals, reactive oxygen species and reactive nitrogen speciesKeywords: Chloramphenicol, rat liver, lipid peroxidation, catalase activity, antioxidant

Construction of novel pJRD215-derived plasmids using chloramphenicol acetyltransferase (cat) gene as a selection marker for Acidithiobacillus caldus.

BackgroundAcidithiobacillus caldus, a Gram-negative, chemolithotrophic sulfur-oxidizing bacterium, is widely applied in bioleaching. The absence of an ideal selection marker has become a major obstacle to achieve high efficiency of the gene transfer system for A. caldus. Plasmid pJRD215, widely used in Acidithiobacillus spp., has severe drawbacks in molecular manipulations and potential biosafety issues due to its mobility. Therefore, finding a new selection marker and constructing new plasmids have become an urgent and fundamental work for A. caldus.ResultsEffective inhibitory effect of chloramphenicol on the growth of A. caldus was elucidated for the first time. The P2-cat gene cassette, including a chloramphenicol acetyltransferase gene (cat) from plasmid pACBSR and a promoter (P2) upstream of the tetracycline resistance gene on pBR322, was designed, chloramphenicol acetyltransferase was expressed in A. caldus, and the enzyme activity was assessed. A new vector pSDU1 carrying the replication and mobilization regions derived from pJRD215, the P2-cat gene cassette and a multiple cloning site from pUC19 was successfully constructed. Compared with pJRD215, pSDU1 had a 27-fold increase in electrotransformation efficiency (30.43±0.88×104 CFU/μg DNA for pSDU1 and 1.09±0.11×104 CFU/μg DNA for pJRD215), better carrying capacity and could offer more convenience for the restriction enzyme digestion. In addition, the generated plasmid pSDU1Δmob, a novel non-mobilizable derivative of pSDU1 lacking some DNA sequences involved in the mobilization process, had increased copy number in A. caldus and lost its mobility for biosafety considerations. Both pSDU1 and pSDU1Δmob exhibited stable maintenance in A. caldus within 50 passages. However, further deletion of orfEF region involved in regulating repAC operon resulted in a negative effect on transformation efficiency, copy number and stability of plasmid pSDU1ΔmobΔorfEF in A. caldus.ConclusionChloramphenicol was proved to be an ideal selection marker for A. caldus. Novel plasmids carrying cat gene were constructed. The utilization of these vectors will undoubtedly facilitate efficient genetic manipulations and accelerate the research progress in A. caldus.

Dissipation of carbendazim and chloramphenicol alone and in combination and their effects on soil fungal:bacterial ratios and soil enzyme activities

The dissipation of carbendazim and chloramphenicol alone and in combination and their effects on soil fungal:bacterial ratios and soil enzyme activities were investigated. The results revealed that carbendazim dissipation was little affected by chloramphenicol, whereas chloramphenicol dissipation was found to be retarded significantly by the presence of carbendazim. The inhibitory effect of carbendazim on the fungal:bacterial ratios was increased by the presence of chloramphenicol, and the inhibitory effect of chloramphenicol on neutral phosphatase was increased by the presence of carbendazim. Carbendazim increased soil catalase and urease activities, but this increase was partially diminished by the presence of chloramphenicol. Little interaction was observed between carbendazim and chloramphenicol with regard to their influence on soil invertase. The results obtained in this study suggest that combinations of fungicides and antibiotics may alter the compounds’ individual behaviors in soil and their effects on soil enzymes.

Extension of the Inhibitory Effect of Chloramphenicol on Bacteria by Incorporating It into Fe3O4 Magnetic Nanoparticles

Magnetite Fe3C4 nanoparticles were prepared by using a co-precipitation method, then functionalized by double layers of surfactants (core/oleic acid (OA) /sodium dodecyl sulfate (SDS)). For application as a drug carrier, the double-layer-coated magnetic nanoparticles were fully loaded with the antibiotic chloramphenicol (Cm) to investigate the effect of the drug release process on the bacteria Escherichia coli (E. coli). The water-soluble Cm and Cm-coated magnetic nanoparticles (Cm-NPs) (equivalent to 50 ?l of 5-200 ?g/ml Cm) were poured into small holes on agar plates spread with E. coli, after which the plates were incubated overnight. The diameters of the nonbacteria circles were shown to gain their maximal values after 14 h, then to gradually decrease in almost all samples. Nevertheless, the circles created by Cm-NPs were about 1.5 times larger than those for the control Cm. The speed of bacterial lawn re-grown in the case of the Cm-NPs was obviously slower than that of the controlled Cm. We conclude that the magnetic nanoparticles gradually release the antibiotic, thereby maintaining the stability and the effect of the antibiotic longer than that of the conventional water-soluble antibiotic. Author

Use of a “least square” optimization procedure to estimate enzyme characteristics and substrate affinities in the denitrification reactions in soil

Predicting temporal variation in the production of gaseous forms of N generated in the denitrification process over a short time scale requires a quantification of the characteristics and kinetics of the denitrification enzymes. The treatments used soil aerobically incubated for 5 days, amended with or without C 2H 2 and a solution containing 3.4 m M KNO 3 with or without 3.7 m M chloramphenicol. Samples were then anaerobically incubated for 48 h at 25°C. Gas fluxes and mineral N pools were monitored periodically from 1 to 48 h. Experiments were set up to determine the initial concentrations of the reduction enzymes involved in denitrification and their rates of de novo synthesis when anaerobiosis was induced for a soil from a permanent pasture. The dynamics of the system were explored with the DETRAN model using an iterative curve-fitting procedure based on the “least square” criterion. The NO 3 − reduction to CO 2 production ratio was ca. 0.6 but increased to 0.8 when glucose was added. The link between the C mineralization and denitrification in the model was adjusted accordingly: “readily” decomposable organic material had a ratio of NO 3 − reduced to CO 2 produced of 1.17 while soil organic matter had a ratio of 0.6. Chloramphenicol did not inhibit the reduction of nitrate so de novo synthesis of nitrate reductase did not occur. The concentration of nitrate reductase could therefore be set at 1. Best fits (lowest residual sum of squares) for the simulation of the NO 2 − concentrations were obtained when the initial concentration of nitrite reductase, relative to the nitrate reductase concentration, was ca. 0.3, its affinity for NO 2 − was between 300 and 450, and its de novo synthesis occurred between ca. 4 and 8 h after the onset of anaerobiosis. Best fits for the simulation of the N 2O and N 2 concentrations were obtained when the relative initial concentration of nitrous oxide reductase was ca. 0.1, its affinity for N 2O was between 3 and 4, and its de novo synthesis occurred between 27 and 42 h after the onset of anaerobiosis. Chloramphenicol reduced the activity of the nitrite and nitrous oxide reductases; pointing at a possible inhibitory effect of the antibiotic. It was concluded that the iterative curve-fitting procedure enabled the estimation of the characteristics of the reduction enzymes involved in the denitrification process, i.e. initial concentrations and the rates of de novo synthesis, which could be used to simulate the kinetics of the denitrification process. The curve-fitting procedure can be used to test the validity of the concepts used to simulate the denitrification process and it can reveal anomalies, e.g. the inhibitory effect of chloramphenicol on the nitrite and nitrous oxide reductase.

Altered synthetic response of Campylobacter jejuni to cocultivation with human epithelial cells is associated with enhanced internalization

Campylobacter jejuni has been shown to bind to and enter epithelial cells in culture. The interaction of C. jejuni with INT 407 epithelial cells was examined to determine whether bacterial protein synthesis is required for either binding or internalization. Chloramphenicol, a selective inhibitor of bacterial protein synthesis, significantly reduced the internalization, but not binding, of C. jejuni compared with untreated controls as determined by protection from gentamicin. Electrophoretic analysis of metabolically labeled proteins revealed that C. jejuni cultured with INT 407 cells synthesized 14 proteins that were not detected in organisms cultured in medium alone. The inhibitory effect of chloramphenicol on internalization was reduced by preincubation of C. jejuni with INT 407 cells. The results indicate that C. jejuni, like some other enteric pathogens, engages in a directed response to cocultivation with epithelial cells by synthesizing one or more proteins that facilitate internalization and suggest that this phenomenon is relevant to the pathogenesis of enteritis caused by C. jejuni.

Replication of plasmid RK2 in vitro by a DNA-membrane complex: evidence for initiation of replication and its coupling to transcription and translation

The following results with an in vitro replication system utilizing a plasmid RK2 DNA-membrane complex indicate that the essential trfA-encoded replication protein of RK2 is present and active in the complex. (i) A complex extracted from a conditional replication mutant of RK2, which contains a temperature-sensitive mutation in trfA, displayed extensive DNA synthesis at the permissive temperature but little activity at the restrictive temperature. A control wild-type RK2 complex showed no inhibition of DNA synthesis at the restrictive temperature. (ii) Analysis of plasmid-encoded proteins revealed that the trfA-specified replication protein and other proteins which may be involved in the replication and maintenance of RK2 are located physically in the complex. Semiconservative plasmid DNA replication by the DNA-membrane complex was indicated by density shift experiments; DNA synthesized in the presence of a heavy-density precursor banded primarily in a heavier-density area of a neutral CsCl density gradient and consisted mostly of heavy- and light-density single-stranded DNA as determined by alkaline CsCl density gradient centrifugation. Plasmid RK2 DNA replication by the DNA-membrane complex appears to be coupled to transcription and translation as indicated by the following results: the inhibitory effects of chloramphenicol on both DNA and protein synthesis by the complex; the stimulation of replication by components normally required for protein synthesis (tRNA and all the common amino acids); the synthesis of RNA and protein by the complex; and the synthesis of specific RK2-encoded proteins.

Protein Synthesis and Photosynthetic Recovery in the Resurrection Plant, Selaginella lepidophylla

The effects of inhibition of protein synthesis on whole-plant CO(2) exchange and on protein synthesis during hydration of the resurrection plant Selaginella lepidophylla (Hook and Grev.) were examined. Both chloramphenicol and cycloheximide inhibited the redevelopment of photosynthetic capacity which normally occurs within 24 hours of hydration in the light. The inhibitory effect of chloramphenicol was greater than that of cycloheximide. The onset of chloramphenicol inhibition of net photosynthesis occurred only after 12 hours of hydration. Cycloheximide stimulated net CO(2) influx early after rehydration and inhibited net CO(2) influx after 14 hours of hydration. Total protein synthesis, as measured by l-[(35)S]methionine incorporation, increased through 24 hours of hydration. Based upon the results obtained with the protein synthesis inhibitors, most protein synthesis within the first 12 hours of hydration was cytoplasm-directed, whereas the rate of organelle-directed protein synthesis remained low until 12 hours of hydration and increased rapidly thereafter. These data suggest that both organelle- and cytoplasm-directed protein synthesis are necessary for full photosynthetic recovery during rehydration of S. lepidophylla.

Repair and plasmid R46 mediated mutation requires inducible functions in Proteus mirabilis.

In Proteus mirabilis nalidixic acid or a predose of UV induce Rec protein formation, a portion of post-UV replication repair and "post-UV replication enhancement." These inducible functions are not significantly affected by the plasmid R46, which renders P. mirabilis efficiently UV-mutable. The R46-mediated UV induction of rif mutations requires additional inducible functions, as existing after nalidixic acid treatment in rec+ strains. After a nalidixic acid pretreatment UV efficient induction of rif mutations occurs without an otherwise obligatory period of post-UV incubation prior to plating on rifampicin agar. THe inducible character of this "qualification" of plasmid R46-mediated UV mutagenesis in P. mirabilis is evident from the inhibitory effects of chloramphenicol and starvation. Constitutive high-level synthesis of Rec protein in cells harboring the recombinant (multi-copy) rec+ plasmid pPM1 reduced plasmid R46-mediated UV mutagenesis, probably by preventing (inducible?) functions required by the plasmid R46 repair-mutator.

Isolation of ribosome containing synaptosome subpopulation with active in vitro protein synthesis

Subpopulations of synaptosomes harvested from neonatal rat brain cortices revealed a differential ability to synthesize protein in vitro. Incubation of synaptosomes with radiolabeled leucine, followed by continuous sucrose gradient centrifugation produced an asymmetric shift in the radioactivity toward the higher density sucrose fractions. The bulk of the mitochondrial cytochrome c-oxidase activity was also found in these fractions, however, subfractionation studies of osmotically-lysed synaptosomes suggested that the newly-synthesized proteins reside in an osmotically sensitive, non-mitochondrial compartment. The ability of each subpopulation of synaptosomes to synthesize protein in vitro was assessed after their isolation from linear continuous sucrose gradients. There was an enrichment of highly active protein synthesizing particles in the "heavy" subpopulations of neonatal synaptosomes. The inhibitory effects of chloramphenicol and cycloheximide on the protein synthesis in these particles were similar to those of the original synaptosome fraction. Electron microscopic analysis revealed an increase in the numbers of ribosome-containing structures resembling dendritic and axonal growth cones.

Phosphate-transport appearance in the sea-urchin egg. I. Effects of protein-synthesis inhibitors on fertilized eggs and embryos.

When fertilized eggs of Strongylocentrotus purpuratus, the purple sea urchin, are incubated in sea water containing 32PO43− and L-14C-valine, valine incorporation is inhibited in the presence of puromycin, cycloheximide, or chloramphenicol, but only the last inhibits the active transport of phosphate. Since chloramphenicol does not depress the egg respiration, it is likely that this drug acts specifically. This is in line with a lack of inhibitory effect of chloramphenicol on phosphate uptake at later developmental stages, except at the time of gastrulation, when the appearance of new phosphate carriers might well occur. It is suggested that the active transport carrier for phosphate, or some element required for its operation, is synthesized after fertilization, with the likely participation of the mitochondrial protein-synthesis machinery. A similar proposal holds for valine uptake.

Suppression of phage nonsense and temperature-sensitive mutants by an suA mutant E. coli.

BECKWITH and Scaife1,2 showed that the suppressor allele suA of Escherichia coli does not suppress nonsense codons in the lactose operon, but does relieve the polarity effect of UAG, UAA and possibly frameshift mutations. It is shown here that suA E. coli strongly suppresses phage T4 amber mutations and weakly suppresses phage T4 temperature-sensitive and amber mutations. Suppression of the inhibitory effect of chloramphenicol on phage T4 growth by suA was also shown. In addition, amber mutants of phage T7 were observed to be suppressed by suA. These results suggest that suA in addition to suppressing polarity in E. coli also increases translational ambiguity in T4 and T7-infected cells, especially for the amber codon.

A possible ribosomal-directed regulatory system in Euglena gracilis. Chlorophyll synthesis.

Cycloheximide at concentrations of 0.1-100mum stimulated chlorophyll synthesis when dark-grown cells of Euglena were illuminated. Chloramphenicol (1-4mm) inhibited chlorophyll synthesis. The effect of cycloheximide on the incorporation of [(14)C]leucine into material insoluble in trichloroacetic acid, and its failure to affect the incorporation of [(32)P]orthophosphate into such material in short incubations, are interpreted as evidence that cycloheximide specifically inhibits protein synthesis by 80S ribosomes. Since the inhibitory effect of chloramphenicol on chlorophyll synthesis is counteracted by the presence of cycloheximide, it is suggested that chlorophyll synthesis is subject to control by a cytoplasmic repressor synthesized on 80S ribosomes, and to a de-repressor synthesized on 70S ribosomes.

ACTH and the differentiation of rat adrenal cortical cells grown in primary tissue culture.

Fetal rat adrenals have been grown in tissue culture. Some specialized properties of the adrenal cortical cells have been studied as an index of their degree of differentiation. Cells transformed morphologically by ACTH differ from controls in their ability to respond rapidly to a single dose of ACTH, as indicated by the yield of isocaproic acid from [26-14C]cholesterol. A correlation was also found between the structure and the steroidogenic activity of the cortical cell mitochondria. Chloramphenicol (0.3 μmole) was found to inhibit the ACTH-induced morphological differentiation of the mitochondria specifically. This inhibition was correlated with a block in the rise of steroid 11β-hydroxylation (mitochondrial) normally induced by ACTH. The rise in steroid 21-hydroxylation (cytoplasmic) was unaffected. The inhibitory effects of chloramphenicol were reversible if the cells were subsequently treated with ACTH alone after removal of chloramphenicol. The present results are consistent with the hypothesis tha...

Biosynthesis of vitamin B 12 by Propionibacterium shermanii. V. Interrelationship between vitamin B 12 and porphyrin synthesis.

Iron and cobalt inhibited the porphyrin synthesis in presence and absence of δ-amino-laevulinic acid (ALA). ALA greatly stimulated the porphyrin synthesis and inhibited the vitamin B12 synthesis. Addition of ALA removed the inhibitory effect of chloramphenicol and azaguanine on porphyrin synthesis. Azaxanthine decreased the porphyrin synthesis in the bacterial cells in presence of ALA. Sodium azide and 2,4-dinitrophenol inhibited growth and vitamin B12 but had no effect on porphyrin synthesis. Malonate had no effect. It is deduced that ALA is the structural material suitable for porphyrin synthesis.

Inhibition of auxin-induced cell expansion in Jerusalem artichoke tuber slices by low concentrations of chloramphenicol

D-threo chloramphenicol (CAP) at 5×10(-5) M, given continuously during a 24-hr aging period and subsequent post-age treatment with 2,4-dichlorophenoxyacetic acid (2,4-D)±kinetin markedly depressed cell expansion in Jerusalem artichoke (Helianthus tuberosus) tuber slices. Both the rate and total amount of expansion were reduced. An inhibitory effect of CAP could be detected at a concentration as low as 6.2×10(-6)M with 2,4-D alone and 1.6×10(-6) M with 2,4-D+kinetin. CAP also inhibited if given with 2,4-D to unaged tissue, and partially inhibited growth of aged tissue when supplied only during or only after aging. Expansion was inhibited when IAA was used in place of 2,4-D. Growth of tissue slices free of detectable bacteria was depressed by CAP, eliminating a possible indirect action of the antibiotic through inhibition of beneficial bacteria. CAP also prevented appearance of pink and brown pigments which normally occur in association with auxin-treated tissues. L-threo CAP did not inhibit growth or pigment formation. Cell division in the tuber slices was not inhibited, and was possibly even stimulated, by D-threo CAP, even at a concentration of 2×10(-4) M. It is concluded that the use of CAP for bacterial control in plant cultures can be hazardous and needs careful checking. Presumably the inhibitory action of CAP results from inhibition of growth-dependant protein metabolism in mitochondria and/or plastids which occurs both during aging and post-aging growth. Partial suppression of metabolic changes during aging would maintain the tissue in a state favouring relatively high mitotic activity and slow growth in response to auxin.

Enzymes of the glycollate pathway in relation to greening in Euglena gracilis

The kinetics of chlorophyll formation during the greening of dark-grown Euglena gracilis was accompanied by marked increases in activity of the enzymes of glycollate metabolism; glycollate: DCPIP(1) oxidoreductase and phosphoglycollate phosphatase (E.C. 3.1.3.18.). Inhibitors of protein synthesis indicated a de novo synthesis of these enzymes during the development of the photosynthetic system. The inhibitory effects of chloramphenicol and cycloheximide, together with a non-aqueous localisation of glycollate: DCPIP oxidoreductase in photoautotrophically-grown cells, indicated that this enzyme is synthesized and located in the cytoplasm, while phosphoglycollate phosphatase was synthesized in the chloroplast. Glycollate: DCPIP oxidoreductase did not increase above the low level in heterotrophic cells when exogenous glycollate, in the presence or absence of glucose, was supplied in the dark.

Action of hydroxyurea on Anabaena variabilis

Low concentrations of hydroxyurea stimulated the growth of the blue-green alga Anabaena variabilis that had been pretreated with sublethal concentrations of chloramphenicol or of certain nucleic acid base analogues. When supplemented to the culture medium, hydroxyurea also counteracted the growth inhibitory effect of chloramphenicol on this organism. In contrast, when A. variabilis cells grown in the presence of hydroxyurea were subsequently treated with chloramphenicol, they were found to have become highly susceptible to the growth inhibitory effects of chloramphenicol. The growth of hydroxyurea pretreated cells in basal medium was attended by a lag that was shorter than that of untreated controls; on the other hand, when hydroxyurea pretreated cells were inoculated into chloramphenicol-supplemented medium, they exhibited a longer lag than that shown by untreated cells in chloramphenicol. The results obtained are discussed in terms of the probable effects of hydroxyurea and chloramphenicol on certain enzyme systems.

SUPPRESSION OF ANTIBODY SYNTHESIS AND PROLONGATION OF HOMOGRAFT SURVIVAL BY CHLORAMPHENICOL

Chloramphenicol suppresses primary antibody synthesis in vivo without affecting the ability to develop a normal anamnestic response. Chloramphenicol also prolongs homograft survival in rabbits. The survival of homografts is related to the duration as well as to the amount of chloramphenicol administered. The mechanism of action of chloramphenicol in suppressing immune responses is correlated with its ability to inhibit protein synthesis in proliferating mammalian cells. These observations suggest that the inhibitory effect of chloramphenicol on protein synthesis may be applicable to mammalian cells in vivo as well as to cell-free systems and to intact mammalian cells in vivo.

INHIBITION OF PROTEIN SYNTHESIS IN MAMMALIAN CELL-FREE SYSTEMS BY CHLORAMPHENICOL.

Chloramphenicol can inhibit protein synthesis in mammalian cell-free systems as effectively as it inhibits protein synthesis in analogous microbial systems. Significant inhibition in mammalian systems was obtained only when protein synthesis was stimulated by the addition of template RNA to the system, there being comparatively little inhibition of protein synthesis by ribosomes in the absence of stimulatory RNA. It is postulated that chloramphenicol may inhibit the function of messenger or template RNA by successfully competing for ribosomal binding sites, thereby preventing the attachment of RNA to ribosomes. The apparent discrepancy in the sensitivity of mammalian and microbial cells to chloramphenicol may be related to differences in turnover or stability of ribosomal template RNA rather than to intrinsic differences in mechanism of protein synthesis. These observations are in accord with the suggestion that protein synthesis in intact mammalian cells may be susceptible to chloramphenicol inhibition only at the time that new messenger or template RNA is being deposited on ribosomes and that more mature cells may be resistant because informative RNA already deposited on ribosomes is not accessible to chloramphenicol inhibition. The inhibitory effect of chloramphenicol on protein synthesis in proliferating cells may be an important factor in hematologic toxicity attributable to chloramphenicol.