Objective To explore the effect of berberine hydrochloride on human CXC chemokine ligand 16 (CXCL16) and interleukin 33 (interleukin-33, IL-33) in patients with acute cerebral infarction. Methods 118 patients with acute cerebral infarction were divided into berberine group and control group (59 cases each) according to random number table. The control group was given basic treatments such as nutritional nerves, stable plaque, and improved circulation. The berberine group was supplemented with berberine hydrochloride tablets orally 0.5g twice a day for 2 weeks. Serum CXCL16 and IL-33 levels were measured before treatment, day 7 and day 14 of treatment. Results There was no significant difference in CXCL16 and IL-33 between the two groups of patients before treatment (P>0.05). The levels of IL-33 and CXCL16 were significantly lower on day 7 and 14 of treatment than before treatment (P<0.05). The levels of CXCL16 and IL-33 in berberine group were lower than those in control group (P<0.05) on day 7 and 14 of treatment (P<0.05). Conclusion Berberine plays an important role in regulating the levels of CXCL16 and IL-33 in patients with acute cerebral infarction and can improve their prognosis. Therefore, berberine hydrochloride can be added to patients with acute cerebral infarction in clinical practice.