ABSTRACT.-De Vita B., Campos L.L., Listoni A.J., Martin I., Takahira R.K., Curcio B.R., Landim-Alvarenga F. The viability and fetal maturity can be estimated by biochemical evaluation of the fetal fluids of several species; however the biochemical composition of amniotic fluid during pregnancy is not fully defined for equine. The aim of this study was to establish and compare the biochemical profile of amniotic fluid in different moments of pregnancy and at delivery, in order to better explain the peculiarities of the physiology of pregnancy in mares. The values founded for pH, osmolarity, glucose, urea, creatinine, gamma-GT, Sodium, potassium, chloride and total protein were evaluated in amniotic fluid collected from 122 mares comparing the results between the initial-third (IT), mid-third (MT) and latter--third (FT) of gestation and at delivery (D). The gestational period samples were collected at slaughterhouses. Gestational ages (IG), in days, were defined following the regression formula suggested by Naves et al. (2008) by measuring the craniocaudal (CC) distance of the fetuses. Commercial kits for biochemical evaluation were used. Due to the presence of varying degrees of asymmetry and deviations from a standard Gaussian distribution, the Kruskal-Wallis test was used to compare the median of each response variable among the study groups. When there has been significant evidence that at least one of the medians differed from the others, the Wilcoxon test was used to perform multiple comparisons between groups. The Bonferroni method was used to adjust the resulting p-value for multiple comparisons. Statistical analysis was performed using PROC Npar1way and statistical significance was defined as p<0.05. The pH and osmolarity of equine amniotic fluid did not change significantly during the stages of pregnancy and at delivery. The values found for glucose were significantly lower during late pregnancy and at delivery. Concentrations of urea tended to be statistically different in at least one of the groups. A significant increase in creatinine concentrations was observed during the initial-third, medium-third and the final-third of pregnancy and the value found at delivery remained equal to final-third. Values for Gamma GT differed only between FT and D groups and more studies should be conducted about its role in the amniotic fluid of domestic species. For the sodium and chloride 1 Recebido em 22 de setembro de 2013.Aceito para publicação em 23 de fevereiro de 2014.