Objective To investigate the acute toxicity, irritation and effects of Shexiang Anhe Hemorrhoids ointment on hemorrhoid-phased model rats via TLR4/p38 MAPK/NF-κB signaling pathway. Methods Sixteen New Zealand rabbits were randomly divided into the intact skin Shexiang Anhe Hemorrhoids ointment, the intact skin control group, the broken skin Shexiang Anhe Hemorrhoids ointment group and the broken skin control group, with four rabbits in each group. In the experimental group, 20 g of Shexiang Anhe Hemorrhoids ointment (1 g/mL) was evenly applied to the hair removal area of rabbits, and an equal volume of solvent (a mixture of glycerol, lanolin and water) was evenly applied to the hair removal area of the back of rabbits in the control group once a day for 14 d. Another 40 rats were taken and randomly divided into a normal group, a model group, a maillard group, and a hemorrhoid cream group of 10 animals in each group. The cream was applied once a day for 14 d. The acute toxicity of the cream in intact and broken skin of rabbits was observed by HE staining and other methods after treatment. In situ photographs were taken of the perianal tissues of rats with similar hemorrhoids to observe the efficacy of Shexiang Anhe Hemorrhoids ointment, and the length and width of the ulcers were measured with vernier calipers to calculate the area of the ulcers, and RT-qPCR was used to detect the expression of TLR4, p38 MAPK, and NF-κB mRNA in the perianal tissues of the rats. Results Compared with the control group with intact skin and broken skin, there was no significant change in the body mass of rabbits in the administered group; compared with the control group, the mean values of the irritation evaluation points of Shexiang Anhe Hemorrhoids ointment on rabbits with broken skin with the administration of the drug for 1 h, 24 h, 48 h, and 72 h were 1.5, 1, 0.5 and 0.25, respectively, and there was no obvious skin irritation.On the effect of hemorrhoidal similar rats, compared with the model group, Shexiang Anhe Hemorrhoids ointment group rats after 14 d administration of symptoms significantly reduced; rats ulcer area are significantly reduced, the difference is statistically significant (P<0.05); TLR4, P38MAPK, NF-κB mRNA levels were significantly reduced, the difference is statistically significant (P<0.05).Conclusions Shexiang Anhe Hemorrhoids ointment is a safe topical ointment with less acute toxicity and irritation to the skin, and has achieved good efficacy in the treatment of hemorrhoidal-adjacent rats, and its mechanism of action may be related to the inhibition of the TLR4/p38 MAPK/NF-κB signaling pathway.