Abstract

Zearalenone (ZEA) is a well-known non-steroidal oestrogenic mycotoxin produced by common soil fungi of Fusarium genera. India being a tropical country provides the suitable conditions for fungal invasion to the cereals. Several toxicological manifestations of ZEA are well known; however dermal toxicity is not yet explored. Therefore, the aim of this study is to analyze the contamination levels of ZEA in different cereal samples consumed by Indian population and to investigate the dermal toxic potential of ZEA in experimental animals.

Experimental methods: 117 samples of wheat (30), rice (30), maize (27) and oat (30) were collected from the northern states of India. A sample of 10 gm each was grinded and ZEA was extracted with acetonitrile – water (90:10 v/v) mixture. The extracted filtrate was analyzed through HPLC with fluorescence detector and positive samples were validated through LC-MS along with standard ZEA. To evaluate dermal toxic potential of ZEA, dermal hyperplasia and skin edema induction were performed using a single topical application of ZEA (40 and 80 nmol). Further, skin tumor initiation potential of ZEA at a higher dose (120 and 240 nmol) was studied using a two-stage mouse skin tumor protocol for 24 weeks.  

Results and discussion: The results of this study showed that out of 117 cereal samples, 70 (84%) were positive for ZEA, among which 24 (33% approximate) exceeded the permissible limits proposed by European Union. The positive samples were further validated by LC- MS analysis. Based on the quantitative estimation of ZEA contamination in cereals and their daily consumption values, the probable daily intake of ZEA was found to be 6.2 to 6.4 fold higher in rice and wheat samples than the tolerable daily intake prescribed by European Food Safety Authority (EFSA). Rice and wheat are the leading cereals being produced and consumed in India This study demonstrates high contamination level of ZEA in Indian cereal grains which may be the resultant of poor agricultural proceedings and storage conditions. Further, single topical application of ZEA (40 & 80 nmol) significantly enhanced dermal hyperplasia and skin edema. It was also observed that a single topical application of ZEA (120 and 240 nmol) followed by twice weekly application of 12-O-tetradecanoylphorbol-13-acetate for 24 week leads to tumor formation suggesting its tumor initiating potential.

Conclusion: Presence of ZEA in higher level indicates the higher exposure risk for the Indian population as the cereals analyzed in the study are the major food being consumed. The in vivo studies conducted in mice suggests that ZEA have the cell proliferation activity causing edema and hyperplasia as an early event and skin tumor initiating potential as a late event. The present findings are relevant towards the health risks of population where dermal exposure may occur especially in agricultural fields. Unfortunately, there are no regulatory limits for ZEA in cereal grains in India. Thus, it is an important issue for the Indian regulatory body to take proper action to safeguard the health of common people. The farmers should be made aware about good agricultural practices on one hand and regulatory limits for mycotoxins in cereal grains may also be framed so as to safeguard the health of consumers.

Keywords: Zearalenone, HPLC, LCMS, cereals, hyperplasia, Skin edema, 12-Otetradecanoylphorbol-13-acetate