Aims The aim of this study was to determine the effects of sub-chronic aluminum chloride (AlCl 3 ) on spermatogenesis and testicular enzymatic activity in male rats. Main methods Forty Wistar male rats were randomly divided into four groups: control group (CG, 0), low-dose group (LG, 64.18 mg/kg BW AlCl 3 ), mid-dose group (MG, 128.36 mg/kg BW AlCl 3 ) and high-dose group (HG, 256.72 mg/kg BW AlCl 3 ). The rats were orally administered with AlCl 3 for 120 days. At the end of the experiment, the contents of Al, Fe, Cu and Zn, the enzyme activities of testicular acid phosphatase (ACP), succinate dehydrogenase (SDH), lactate dehydrogenase (LDH), lactate dehydrogenase isoenzyme (LDH-x), the sperm count and the sperm malformation rate were examined. Key findings The results showed that the Al and Cu contents, sperm count and the enzyme activities of testicular ACP, SDH, LDH and LDH-x decreased, while the Zn and Fe contents and sperm malformation rate increased in AlCl 3 -treated rats. Significance It suggests that sub-chronic AlCl 3 disorders the balance of trace element and decreases the spermatogenesis and the activities of testicular enzymes, indicating that AlCl 3 has adverse effect on the testicular function in male rats.