Bernd A Herbold

Ciprofloxacin: in vivo genotoxicity studies.

The fluoroquinolone ciprofloxacin is widely used in antimicrobial therapy. It inhibits the bacterial gyrase and in high concentrations in vitro also the functionally related eukaryotic topoisomerase-II, which resulted in genotoxic effects in several in vitro tests. In order to evaluate the relevance of these findings, ciprofloxacin was tested in vivo for genotoxic activity using the following test systems: micronucleus test in bone marrow of mice, cytogenetic chromosome analysis in Chinese hamster, dominant lethal assay in male mice and UDS tests in primary rat and mouse hepatocytes in vivo. These results are compared with already published in vitro and in vivo studies with ciprofloxacin. All in vivo genotoxicity revealed no genotoxic effect for ciprofloxacin. In addition, ciprofloxacin was found to be non-carcinogenic in two rodent long-term bioassays. Therefore, ciprofloxacin is considered to be safe for therapeutic use.

Investigations on the mutagenicity of 1,4-dichlorobenzene and its main metabolite 2,5-dichlorophenol in vivo and in vitro

The genotoxic potential of 1,4-dichlorobenzene (1,4-DCB) has been extensively evaluated in vitro and in vivo. The majority of the studies demonstrated the absence of a genotoxic potential for 1, 4-DCB. At variance are a bone marrow micronucleus test (MNT) after intraperitoneal (i.p.) treatment of NMRI mice [Mohtashamipur et al., Mutagenesis 2 (1987) 111-113] and a gene mutation assay on mouse lymphoma cells [McGregor et al., Environ. Mol. Mutagen. 12 (1988) 85-145]. Therefore, we investigated 1,4-DCB and its main metabolite 2,5-dichlorophenol (2,5-DCP) for both endpoints. In an MNT, male and female NMRI mice were treated orally with single doses of 2500mg/kg 1,4-DCB and 1500mg/kg 2,5-DCP, respectively. Smears were prepared 24, 48 and 72h thereafter. No induction of micronuclei was detected for both compounds. Also under the conditions of Mohtashamipur et al. (1987), intraperitoneal treatments of male and female mice with 2 x 177.5 and 2 x 355mg/kg 1,4-DCB failed to induce micronuclei. In addition, CHO/HPRT-gene mutation tests with 1,4-DCB and 2,5-DCP yielded negative results for both compounds with and without metabolic activation system. Therefore, 1,4-DCB and 2,5-DCP are considered to be non-mutagenic in these test systems.

Mutagenicity studies with 2,4,5‐T on bacteria and mammalian germ cells

The herbicide 2,4,5-T (2,4,5-trichlorophenoxyacetic acid) was evaluated for potential mutagenicity by a Salmonella/mammalian-microsome test, a dominant lethal test on female rats, and by a cytogenetic assay on spermatogonia of Chinese hamster. In the Salmonella/mammalian-microsome test on four Salmonella typhimurium strains (TA 1535, TA 100, TA 1537, and TA 98), doses of up to and including 2500 micrograms/plate did not cause any mutagenic effects. In a dominant lethal test on female rats, 8-week dietary administration of 2,4,5-T at doses of up to and including 10 mg/kg/day did not cause any increase in preimplantation loss or the rate of dead implants, and did not have any effect on the fertilization quota. Cytogenetic analysis of the spermatogonia of male Chinese hamsters orally dosed five times at 24-hr intervals with 2,4,5-T at levels of up to and including 100 mg/kg did not provide any indication of 2,4,5-T having chromosome-damaging effects. Therefore, none of the three test systems provided any indication of 2,4,5-T having a mutagenic effect.

A new method for the enrichment of single renal proximal tubular cells and their first use in the comet assay

A protocol was developed to isolate and enrich single renal proximal tubular cells, performing the following steps: in situ kidney perfusion; isolation of renal tissue pieces by collagenase digestion; selective enrichment of proximal tubular fragments by Percoll gradient centrifugation; and isolation of single proximal tubular cells by digestion of proximal tubular fragments with trypsin. The mean enrichment rate, determined by the glucose-6-phosphatase staining method, was 78.9% with a mean cell viability of 93.8%. After modification of the comet assay protocol, genotoxicity in proximal tubular cells could be investigated. A dose-dependent genotoxic effect of ethyl methanesulphonate in these cells was proven.

Preliminary results of an international survey on sensitivity of s. typhimurium strains in the ames test

Abstract Based on 1275 results of the Ames test, all done at least with the tester strains Salmonella typhimurium TA1535, TA1537, TA1538, TA100 and TA98, 414 tests were considered positive. 4.1% were only positive in TA1535, 8.7% in TA100, 2.7% in TA 1537, 2.9% in TA98, but only 0.7% in TA1538. Therefore, it may be justifiable for routine tests to omit TA1538.

Dimethylhydrazine: a reliable positive control for the short sampling time in the UDS assay in vivo.

In the first international guideline addressing the unscheduled DNA synthesis (UDS) assay in vivo (OECD guideline no. 486, adopted July 1997) only the genotoxic liver carcinogen N-nitrosodimethylamine (NDMA) is proposed as positive control for the short sampling time. Since NDMA is extremely volatile, alternative positive controls should be identified to facilitate handling and reduce exposure risk during routine testing. At Bayer AG and at RCC-CCR GmbH, the genotoxic but non-volatile dimethylhydrazine (DMH; as dihydrochloride) was used instead as positive control in livers of Wistar rats and to a limited extent of NRMI mice after 2-4h exposure. As shown by the data presented in this paper DMH induced a positive result in a total of 21 UDS in vivo studies over a period of 7 years. A negative result was never seen for DMH. Due to these results DMH was proven to be a suitable and reliable positive control in the UDS assay in vivo. Consequently, DMH should be considered as positive control for the short sampling time in the next issue of OECD guideline no. 486.