Objective: To investigate the alterations of serum sexual homones level, and the expression and distribution of estrogen receptor β (ERβ) in the colon of rats with slow transit constipation (STC). Methods: Chemiluminescence, immunohistochemistry and Western blot technique were carried out to detect the serum level of sexual homones and determine the distribution and expression of ERβ in the colons of the STC rats and normal control rats, respectively. Results: Chemiluminescence showed that all the serum levels of follicle-stimulating hormone (FSH), estradiol (E2), luteinizing hormone (LH), progesterone (P) and testosterone (Testo) of rats with STC had no differences compared with those of the control rats (all P＞0.05). Immunohistochemical staining showed that the ERβ proteins were distributed mainly in myenteric nerve plexus and submucous nerve plexus of the rats′colons, and the expression level of ERβ of the STC rat was markedly lower than that of the normal rat. Western blot analysis also revealed that the expression level of ERβ protein of STC rats was deceased compared with that of the normal rats and this had a significant difference (P＜0.01). Conclusions: The expression level of ERβ in the colon of the STC rats is reduced, which may probably be associated with the pathogenesis of STC.