Introduction

Dicyclanil (Figure 1), 4,6-diamino-2-cyclopropylaminopyrimidine-5-carbonitrile, is a pyrimidine-derived insect growth regulator used in a veterinary medicine for the prevention of myiasis (fly-strike) on sheep. As a result, small amounts of the parent drug and its metabolites are sometimes detected as residues in edible tissues from these animals. In order to evaluate the safety of this chemical, many toxicity studies using experimental animals have been performed. In an 18-monthcarcinogenicity study of dicyclanil, the incidence of hepatocellular carcinomas increased in female mice of the high-dose group (1500 mg/kg feed).Hepatocellular necrosis was also observed in mice given low doses (100 mg/kg feed) or more. However,negative results were obtained in an in vivo micronucleus test, as well as in 4 in vitro tests, reverse mutation test, DNA repair test, gene mutation test, and cytogenesis test. On the basis of these data, the Food and Agriculture Organization (FAO)/World Health Organization (WHO) Joint Expert Committee on Food Additives (JECFA) concluded that the hepatocarcinogenicity of dicyclanil in mice was probably due to a non-genotoxic mechanism resulting from a repeated necrosis-regeneration response of hepatocytes. However, the actual mechanism of this hepatocarcinogenicity is still not understood.[1]

In vivo genotoxicity of dicyclanil

In order to clarify the in vivo genotoxicity of dicyclanil with the potential of hepatocarcinogenicity, the stomach, colon, liver, kidney, urinary bladder, lung, brain and bone marrow of male ddY mice given a single oral administration of 100 and 200 mg/kg body weight of dicyclanil were evaluated in an alkaline single-cell gel electrophoresis (comet) assay. In addition, to investigate its possible initiation activity, partially hepatectomized male F344 rats given a single oral administration of 75 mg/kg body weight of dicyclanil were examined by a short-term liver initiation assay. Three and 24 hr after administration, cell migration, as a marker of DNA damage in comet assay, was not observed in any of the tissues of dicyclanil-treated mice. There were no significant differences in the number and area of glutathione Stransferase placental form (GST-P) positive foci, as a marker of hepatocellular preneoplastic lesions in rats, between treated and control groups. These results indicate that dicyclanil has neither in vivo genotoxicity nor initiation activity, and suggest that the hepatocarcinogenicity in mice induced by dicyclanil is attributable to a non-genotoxic mechanism.[1]

Risk assessment of dicycranil by different agences

Food Safety Commission of Japan (FSCJ) conducted a risk assessment of dicycranil, a pyrimidine derived insect growth regulator, using the evaluation reports from the Joint FAO/WHO Expert Committee on Food Additives (JECFA), the European Medicines Agency (EMEA), and also the Australian government. In an 18-month chronic toxicity/ carcinogenicity study in mice, increased incidences of hepatocellular adenomas and carcinomas were observed in females in the 500 ppm group. In spite of a recent experiment implying the possible indirect genotoxicity of dicyclanil on the carcinogenicity, dicyclanil is unlikely to exert the carcinogenicity in vivo through the genotoxic mechanism judging from other studies. FSCJ recognized it as feasible to set the threshold value. Adverse effects detected at the lowest dose in various toxicological studies were the increased plasma levels of cholesterol and phospholipid at 100 ppm (equivalent to 2.7 mg/kg bw/day in males and 3.5 mg/kg bw/day in females) in a 90-day subacute toxicity study in dogs. No-observed-adverse-effect level (NOAEL) of this study was 20 ppm (equivalent to 0.61 mg/kg bw/day in males and 0.71 mg/kg bw/day in females). On the other hand, the NOAEL in a long term study, a 12-month chronic toxicity study in dogs was 25 ppm (equivalent to 0.71 mg/kg bw/day in males) based on increased level of plasma cholesterol observed only in males at 150 ppm (equivalent to 4.4 mg/kg bw/day in males and 5.1 mg/kg bw/day in females). The increased cholesterol levels in plasma were common in both studies in dogs. It was appropriate to choose the NOAEL for the effect on cholesterol in the longer term treatment, and thus FSCJ adopted the NOAEL of 0.71 mg/kg bw/day. Consequently, FSCJ specified the ADI of 0.0071 mg/kg bw/day for dicyclanil based on the NOAEL of 0.71 mg/kg bw/day in the 12-month chronic toxicity study in dogs, by applying a safety factor of 100.[2]

Degradation Pathway of Dicyclanil in Water

Abiotic degradationby photolysis in aqueous solution is of interest to determining the impact of dicyclanil on the environment. Dicyclanil has no aquatic uses; however, it could potentially enter surface water by spray drift during application or runoff after application. As study relates that approximately 37-59% of the total dose remained on the animals, the remainder being collected as runoff. Therefore, determination of the rate and route of photolytic degradation in water is crucial in defining the environmental impact of dicyclanil application.

The dicyclanil is degraded both by photolysis and photocatalysis. The use of titanium dioxide allows to accelerate more than 40 times the degradation and leads to the total destruction of the initial molecule of dicyclanil in less than1 h. The use of HPLC-MS and LC-UV has permitted establishing a degradation scheme of dicyclanil under irradiation with and without the photocatalyst, including four different pathways. The first one (pathway a) results from th eoxidative opening of the cyclopropyl ring producing thealdehyde 2a (tR=6.601 min; [M+H]; [M+Na]). Pathway b leads directly to descyclopropyl dicyclanil 10a (tR=9.600 min; [M+H]) and consequently to an oxidative cleavage of the bond between the cyclopropyl ring and the exocyclic nitrogen.Pathway c rises from the oxidation of the cyano group of the dicyclanil into amide 2c (tR=9.710min; [M+H]). Then OH?radicals attack the cyclopropyl group to form the β-hydroxy-aldehyde 3c.In pathway d the first photoproduct results from the cleavage between the cyano group and the pyrimidic ring. This cleavage produces CN^- ions that are oxidized in solution into CNO^- ions. Some intermediates are observed both by photolysis and photocatalysis.[3]

References

1. Moto M, Sasaki YF, Okamura M, et al. Absence of in vivo genotoxicity and liver initiation activity of dicyclanil. J Toxicol Sci. 2003;28(3):173-179. doi:10.2131/jts.28.173

2. Food Safety Commission of Japan. Dicyclanil (Veterinary Medicinal Products). Food Saf (Tokyo). 2018;6(3):136-138. Published 2018 Sep 28. doi:10.14252/foodsafetyfscj.2018007s

3. Goutailler G, Guillard C, Faure R, Pa?ssé O. Degradation pathway of dicyclanil in water in the presence of titanium dioxide. Comparison with photolysis. J Agric Food Chem. 2002;50(18):5115-5120. doi:10.1021/jf0202943