For a meaningful interpretation of toxic effects in fish, knowledge of sub-lethal events is essential and for this reason, we have conducted a series of studies with a relevant environmental concentration of a mixture of herbicides; Atrazine, simazine, diuron and isoproturon (ASDI) pollutants and using immune (Fatima et al. 2007) and endocrinological changes as principal endpoints under controlled laboratory conditions. Goldfish (Carassius auratus) were exposed to a cumulative concentration of 50µg/l of ASDI mixture for 12 weeks which was reported above the permissible level in the drinking water of Belgium (EAEW report, 1998) and nearby aquatic environments, confirming some of these herbicides still persistence despite restriction policies (Pesce et al. 2010;Agbohessi et al. 2014).Control and exposed fish were sampled at week-4, 8, and 12. We investigated the endocrine modulatory effect of ASDI which shows a low degradation rate in water. Thus, at the realistic level of ASDI concentration, significantly increased levels of cortisol and aromatase after 4-weeks were recorded. Reduction in aromatase activity at 8 and 12 weeks is followed by estradiol (E2) showing a positive correlation between them. Hence, 11-Keto reduced significantly at 4-8 weeks and enhanced at 12 weeks. Remarkable reduction in E2 and 11-Keto may be correlated with the reduced egg production in stressed fish by these herbicides. However, no remarkable change in testosterone is observed at 4 and 12 weeks as compared to the control. ASDI mixture treatment induced remarkable induction in vitellogenin (Vtg) biomarker protein in male stressed fish. Furthermore, it may confer that modulation in the pathway of the reproductive cycle may alter the gamete quality, quantity and consequently may disrupt spawning and mating behaviour in fishes even at a realistic level of pollutant in the environment.