Objective To evaluate the acute toxicity and acute eye irritation of hydroquinone. Methods i) Acute toxicity test. Specific pathogen-free (SPF) Kunming mice were randomly divided into four dose groups, 10 mice per group with equal number of males and females. Hydroquinone was administered as a single exposure via oral gavage and intraperitoneal injection at doses of 0.00, 100.00, 250.00, and 500.00 mg/kg body weight. The mice were observed for 14 days. The toxic symptoms were recorded, and median lethal dose (LD₅₀) was calculated. ii) In vitro eye irritation test. Fertilized chicken embryos were randomly divided into four dose groups, with six embryos in each group. The chorioallantoic membrane (CAM) assay was used to evaluate the acute eye irritation potential of hydroquinone in vitro. iii) SPF Kunming mice were randomly divided into three doses groups, 10 mice per group with equal numbers of males and females. Hydroquinone was administered via tail vein injection at doses of 0.00, 25.00, and 50.00 mg/kg body weight. Blood alanine aminotransferase (ALT), aspartate aminotransferase (AST), methemoglobin (MetHb), and cholinesterase levels were measured using colorimetric methods after one hour exposure. Organ coefficients of the heart, liver, spleen, lungs, and kidneys in mice were calculated. Results i) Following oral administration of 500.00 mg/kg body weight hydroquinone, the mice rapidly developed severe toxic symptoms, including agitation, cyanosis of the lips, eye closure, and limb convulsions. Trunk rigidity and curling occurred within 15-60 mins, ended up with death. After intraperitoneal injection at 500.00 mg/kg body weight hydroquinone, toxic reactions occurred more rapidly, with all mice died within five mins. The LD₅₀ values for acute oral and intraperitoneal exposure were 356.64 and 275.90 mg/kg body weight, respectively. Female mice had higher LD₅₀ values for acute intraperitioneal exposure than males (316.58 vs 276.38 mg/kg body weight). ii) The in vitro eye irritation test results showed an irritation score of 3.05 at a dose of 100.00 mg/kg body weight, indicating mild eye irritation, accompanied by vascular congestion and edema in the embryos. iii) Tail vein injection results showed that mouse serum ALT activity increased with increasing hydroquinone doses (all P<0.05), and ALT activity was higher in males than in females (P<0.01). Serum AST activity of mice in the 25.00 and 50.00 mg/kg body weight groups was higher than that in the 0.00 mg/kg body weight group (both P<0.05). With increasing hydroquinone-exposed doses, whole bood MetHb levels increased and cholinesterase activity decreased in both male and female mice (both P<0.05). Male mice had higher MetHb levels than females at corresponding doses among 25.00 and 50.00 mg/kg body weight groups (all P<0.05). Serum cholinesterase levels in male mice were higher than that in females at corresponding doses among 0.00 and 25.00 mg/kg body weight groups (both P<0.05). As the hydroquinone exposure dose increased, the liver organ coefficients decreased in both female and male mice (both P<0.05). The liver organ coefficient in male mice at the 50.00 mg/kg body weight group was higher than that in female mice (P<0.05). Compared to mice of the same sex in the 25.00 mg/kg body weight group, the kidney organ coefficients decreased in both female and male mice in the 0.00 mg/kg body weight group (all P<0.05), and decreased in the 50.00 mg/kg body weight group (all P<0.05). The male mice had lower kidney organ coefficient than female mice at 25.00 mg/kg body weight group (P<0.05). Conclusion Hydroquinone is classified as a moderately toxic substance. Increasing exposure doses result in pronounced eye irritation and affect hepatic and renal functions of mice.