Copper sulfate and jasper herbicide are heavily used in the aquatic environment to kill algae, bacteria, Mollusca, and unwanted plants. Therefore, this study was carried out to evaluate the effect of the chronic toxicity of those compounds, separately and mixed, on some biomarkers in the Nile tilapia, Oreochromis niloticus. Results showed that a 33.50 mg/1 found of the 96h-LC₅₀ of copper sulfate decreased in the mixture to record 11.70 mg/l. While in the case of a 3.80 mg/1 found of jasper, it marked a decreasing record of 2.20 mg/l in the mixture. Afterward, fish were exposed to the sublethal concentrations (1/10 of the 96h-LC50) separately and in a mixture for 28 days to detect the changes in biomarkers. Whereas the rest of the fish were transferred to water free of those chemicals for 30 days to study the recovery. Superoxide dismutase (SOD), glutathione peroxidase (GPx),  catalase (CAT), and malondialdehyde (MDA) decreased in all treatments. The SOD and MDA didn't recover in fish exposed to the mixture. The GPX and CAT exceeded the control during the recovery. Alanine transaminase (ALT) and alkaline phosphatase (ALP) elevated in all treatments on the 4^th day, and declined on the 28^th of exposure, and didn't recover. Acetyl-cholinesterase (AChE) decreased in all treated groups and still impacted in fish exposed previously to the mixture during recovery. Uric acid increased in all treated groups and recovered in fish exposed alone to jasper. Creatinine increased in all groups and recovered in those treated with copper sulfate and jasper separately. Albumin decreased in all treated groups during all periods and didn't recover. In general, the mixture of copper sulfate and jasper herbicide is more toxic for O. niloticus and the recovery state is difficult to occur for fish previously treated with the mixture.