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Creatinine plays a key role in the function and maturation of fetal kidneys throughout pregnancy. It is important to identify other markers that may help in the diagnosis of renal dysfunction. Our aim was to determine the profile of and the correlation between biochemical markers to be used to assess renal function and maturation of the fetus in the amniotic fluid during pregnancy and to determine the distribution of normal values for creatinine, N-acetyl-beta-D-glucosaminidase (NAG), beta2-microglobulin, glucose, urea, sodium, potassium, phosphorus, calcium, uric acid, albumin, and osmolality in three gestational age groups. This was a cross-section study that assessed 115 samples of amniotic fluid during three different periods of pregnancy, i.e., 13 to 20, 27 to 34, and 36 to 42 weeks. Concentrations of creatinine, NAG, urea, potassium and uric acid increased during pregnancy (P0.05). Beta2-microglobulin, glucose, sodium, phosphorus, calcium, and albumin concentration and osmolality decreased (P0.05), whereas beta2-microglobulin, glucose and uric acid presented significant correlations with gestational age and creatinine, respectively (r0.6, P0.05). Urea, potassium and phosphorus showed mild correlations with both (r0.5, P0.05). NAG, sodium, albumin and osmolality did not show significant correlations (r0.5, P0.05). These tests confirmed the important role of creatinine in terms of correlation with gestational age. beta2-Microglobulin, glucose and uric acid were significant as markers of function and maturation of fetal kidneys, whereas NAG did not demonstrate a useful role for the assessment of renal maturation.