Gian Luigi Biagi

Partition data of penicillins determined by means of reversed-phase thin-layer chromatography

Abstract The RM values of II penicillins were measured by means of a reversed-phase thin-layer chromatographic method with various concentrations of acetone in the mobile phase. RM values were calculated by interpolation or extrapolation from the ranges of linearity between RM values and composition of the mobile phase. The influence of substituent groups on the RM values of the penicillins and disagreement with Σπ values are pointed out.

Determination of lipophilicity by means of reversed-phase thin-layer chromatography: II. Influence of the organic modifier on the slope of the thin-layer chromatographic equation

The RM values measured for a series of steroids were very close to those determined more than 15 years ago using the same chemicals. This finding supports the reliability of RM values as a lipophilicity parameter. However, the point at issue in this work was the influence of the organic solvent in the mobile phase on the slopes of the TLC equations. In fact, the slopes of the TLC equations were shown to be related to the reciprocal of the solvent strength (1/E0). As a consequence, the ratio between the slopes of the TLC equations in different solvent systems are close to the ratio between the 1/E0 values for the corresponding solvent pairs. A further interesting aspect seems to arise from the analysis of the equations correlating slopes and intercepts of the TLC equations. In particular, the ratios between the b values of such equations in different solvent systems are close to the ratios between the corresponding E0 values.

β-carotene as enhancer of cell transforming activity of powerful carcinogens and cigarette-smoke condensate on BALB /c 3T3 cells in vitro

We report the ability of beta-carotene (betaC) to affect the cell transforming activity of 3-methylcholanthrene (3-MCA), benzo(a)pyrene (B(a)P) and cigarette-smoke condensate (TAR) in an in vitro medium-term (approximately 8 weeks) experimental model utilizing BALB/c 3T3 cells. Different experimental schedules were performed either in the presence or absence of betaC: (i) cultures treated for 72 h with each chemical (acute treatment), (ii) cultures grown in presence of each chemical for the whole period of the experiment (chronic treatment). These procedures suggested a possible cocarcinogenic potential of the carotenoid following interactions with other chemicals mimicking continuous human exposition to several xenobiotics. Although the pigment did not show any cell transforming potential when tested alone either in acute or chronic treatment, it did augment that of other tested agents. Induction of cell transformation by B(a)P was markedly enhanced by the presence of this carotenoid in either acute or chronic treatment. Only in presence of betaC, was TAR able to significantly act as a cell transforming agent in prolonged, chronic treatment of cultures. Enhanced cell transformation activity could be due to the boosting effect of betaC on P450 apparatus. Indeed, elsewhere we have found that the latter increased the ratio of formation of diol epoxide carcinogenic metabolites of B(a)P as well as other carcinogens present in TAR. By contrast, no differences of cell transforming activity of 3-MCA, an ultimate carcinogen, were seen either in the presence or absence of betaC under the various experimental conditions. These data, which are in keeping with the cocarcinogenic potential of betaC, may help to explain the unexpected lung cancer increases obtained in chemoprevention trials in heavy smokers supplemented with the isoprenoid. Our findings also highlight the potential risk to humans derived from interactions among xenobiotics present in the environment.

Induction and suppression of cytochrome P450 isoenzymes and generation of oxygen radicals by procymidone in liver, kidney and lung of CD1 mice

Although chronic administration of procymidone (a widely used dicarboximide fungicide) leads to an increased incidence of liver tumors in mice, short-term genotoxicity studies proved negative. As cytochrome P450 (CYP) induction has been linked to non-genotoxic carcinogenesis, we investigated whether procymidone administration causes induction of CYP-dependent monooxygenases in liver, kidney and lung microsomes of male Swiss Albino CD1 mice after single or repeated (daily for three consecutive days) i.p. treatment with either 400 or 800 (1/10 or 1/20 of the DL(50)) mgkg(-1) b.w. procymidone. CYP content and CYP3A1/2, 1A1, 1A2, 2B1/2, 2E1, 2A, 2D9 and 2C11 supported oxidations were studied using either the regio- and stereo-selective hydroxylation of testosterone as multibiomarker or highly specific substrates as probes of various CYPs. While a single dose was uneffective, multiple procymidone administration lead to marked inductions of various monooxygenases: CYP3A1/2 in liver and lung (as measured by N-demethylation of aminopyrine and testosterone 6 beta-hydroxylase); CYP2E1 in liver (p-nitrophenol hydroxylation); CYP1A1 in liver and kidney (deethylation of ethoxyresorufin). Several hydroxylations were induced in the liver, including the CYP2A-linked 7 alpha (14-fold) as well as 6 alpha (22-fold), 6 beta, 16 beta and 2 beta hydroxylases. The pattern of inductions/suppressions recorded in the three different tissues suggests that procymidone exerts complex effects on the CYP profile. Tissue-specific trends included a large number of inductions in the liver and suppressions in the lung. The main inductions were corroborated by immunoblotting analyses and Northern blotting showed that inductions of CYP3A1/2, CYP2E1 and CYP1A1/2 were paralleled by increased mRNA levels. It was also found that CYP over-expression generates large amounts of reactive oxygen species (ROS), especially in liver. These data may explain why in vitro short-term genotoxicity studies on procymidone were negative, whereas in vivo long-term carcinogenesis studies turned out positive: long-term CYP induction (e.g. oxygen centered free radicals over-production) can have a co-carcinogenic and/or promoting potential.

RM and log P values of 5-nitroimidazoles

Abstract The chromatographic RM values of a series of nitroimidazoles and their log P values were determined in view of a study of their structure-activity relationships as mutagenic agents. The equations describing the relationship between RM and log P values show a low correlation coefficient. The introduction of the molar refractivity of the R1 and R2 groups yields a significant improvement in the correlation coefficient. The molar refractivity could be an expression of the adsorption activity of the silica gel layer.

Study of the lipophilic character of xanthine and adenosine derivatives. I: RM and log P values

Abstract The RM values of a series of xanthine and adenosine derivatives were measured using silicone reversed-phase thin-layer chromatographic (TLC) and C18 reversed-phase high-performance TLC systems. The two series of data were well correlated. Both were compared with experimental log P and calculated CLOGP values. For xanthine derivatives a good linear relationship was shown between the RM values from the two chromatographic systems and the log P or CLOGP data. For adenosine derivatives the CLOGP values had to be corrected in order to fit the data to the same equation. The TLC data proved to be reliable parameters for describing the lipophilic properties of the test compounds.

Study of lipophilic character of serotonergic ligands

Abstract The R M values of a series of serotonergic derivatives were measured using a reversed-phase TLC system with acetone, acetonitrile or methanol as the organic modifier of the mobile phase. As regards the basic aspects of the chromatographic technique, the series of compounds studied here behave in quite the same way as the previoulsy investigated series of chemicals, i.e. a very good correlation is found between the experimental and extrapolated R M values as well as between the extrapolated R M values from different organic solvent systems; the relationship between intercepts and slopes of the TLC equations in series of congeneric compounds; the influence of the organic modifier on the slopes of the TLC equations. The R M values from the above system were shown to be correlated with those obtained with a C 18 high-performance TLC. Finally, both chromatographic parameters were compared with calculated octanol-water log P values.

Determination of lipophilicity by means of reversed-phase thin-layer chromatography. III: Study of the TLC equations for a series of ionizable quinolone derivatives

The RM values of a series of antibacterial quinolones were measured at pH 9.0 and 1.2 using a reversed-phase TLC system with acetone, methanol or acetonitrile as the organic modifier of the mobile phase and silicone DC 200 as the impregnating agent of the silica gel layer. The data obtained provide a further contribution to the assessment of the basic aspects of the chromatographic determination of lipophilicity for ionizable compounds. The very good correlations between experimental and extrapolated RM values support the validity of the extrapolation technique. The overlapping of the extrapolated RM values from three different systems show that they are not dependent on the nature of the organic solvent. In a series of congeneric compounds there is a relationship between intercepts (a) and slopes (b) of the TLC equations. Factors affecting chromatographic congenerity are discussed. The slopes of the TLC equations and those of the equations correlating the parameters a and b are related to the solvent strength of the organic modifiers.

Mutagenicity of a series of 25 nitroimidazoles and two nitrothiazoles in Salmonella typhimurium

Twenty-five 5-nitroimidazole and two 5-nitrothiazole derivatives were tested for mutagenicity as well as for antibacterial activity in Salmonella typhimurium TA-100 strain. Many of these compounds such as metronidazole, azanidazole, nimorazole, carnidazole, ornidazole, tinidazole, etc are extensively used in human chemotherapy, and some of them were recently synthetized for possible clinical trials as hypoxic cell specific radiosensitizers. Both mutagenic and antibacterial activity were shown for 22 of the test compounds. The high correlation between mutagenic and antibacterial activity supports the hypothesis of a same mechanism for both activities. The present results confirm that the mutagenicity of the nitroheterocyclic compounds is not separated from other biological activities, such as antimicrobial activity.

NADPH-generating system: Influence on microsomal mono-oxygenase stability during incubation for the liver-microsomal assay with rat and mouse S9 fractions ☆

Activity levels of 7-ethoxycoumarin O-deethylase (ED), aminopyrine N-demethylase (APD), p-nitroanisole O-demethylase (p-NAD) and glucose-6-phosphate dehydrogenase (G-6-PDH) were determined in incubation mixtures for the liver-microsomal assay (LMA) at time 0 and after 1 and 2 h incubation under conditions for mutagenic assay. The experiments were performed with S9 liver fractions from mice (induced with Na-phenobarbital and beta-naphthoflavone) and rats (induced with Aroclor 1254) with and without G-6-PDH in the incubation mixtures. In the absence of G-6-PDH the activities were significantly lower at time 0 in the mouse. The pattern of stability, however, was similar for the activities, with an increase of stability after 1 and 2 h of pre-incubation (an exception for p-NAD). Only ED activity showed a similar behaviour in the rat. No differences were present for APD and p-NAD activities at time 0 in the rat, but the enzyme stabilities were significantly decreased after 2 h of incubation (about 15% and 10% for APD and p-NAD respectively) in the absence of G-6-PDH. At time 0, the amounts of G-6-PDH differed between mouse and rat fractions; however, during the incubations for LMA they decreased by about 57% and 53% for the two species, respectively. In addition to the above biochemical results, the presence of exogenous G-6-PDH in the incubations for the mutagenic assay, significantly increased the mitotic gene conversion and mitotic crossing-over of dimethylnitrosamine (DMN) and AR2MNFN (a nitroimidazo[2,1-b]thiazole) in the D7 strain of Saccharomyces cerevisiae.