The lowland tapir is the biggest Brazilian terrestrial mammal, which belongs to the order Perissodactyla, suborder Hippomorpha, superfamily Tapiroidea and a member of the family Tapiridae. At tropical forest the tapir is involved with seed dispersal. The knowledge of this wild animal reproductive cycle is one way to help its preservation. The stress due to restrain of captive or free-ranging wild animal in order to sample collection limits endocrine study once it can be hazard for the estrous cycle. One possibility is to quantify gonadal hormones at the excreta. Progesterone milk levels were studied in a tapir housed at the Araçatuba Zoo, in São Paulo, Brazil. Milk samples, vaginal cytology and rectal temperature were collected during lactation. The progesterone was quantified by radioimmunoassay solid phase (Coat-a-Count, DPC®). The standard was supplied by CENA-FAO and the assay showed sensitivity of 1.25 nmol/L with intra-assay variation of 15.36%. During most of the lactation (November to June) the female showed no detectable levels of progesterone. After 158 days (from November to April) it was detected the first progesterone peak with 2.3 nmol/L that lasted for 5 days. The second progesterone peak of 3.54 nmol/L lasted for 23 days. The lactation ceased 74 days after the first milk progesterone surge. This animal showed a prolonged lactational anestrous period (nearly 5 months) and the return of gonadal cycle by fall suggested no positive photo-period influence. The milk progesterone quantification showed to be useful for reproductive cycle evaluation of this animal, although vaginal cytology and temperature fluctuation had no relationship with hormonal levels.