Nitric oxide (NO) is readily oxidized to nitrate and nitrite and NO activates guanylyl cyclase, increasing cyclic GMP levels. To determine if nitric oxide synthase (NOS) is present in urine collected daily from patients following renal transplantation, we evaluated NOS activity in the leukocyte-rich particulate fraction and measured nitrate, nitrite, and cyclic GMP levels in the supernatant fraction of the urine. Reverse transcriptase-PCR and cDNA sequencing confirmed the presence of inducible NOS (iNOS) in cells obtained from the urine of renal transplant patients with rejection. NOS activity was elevated significantly in renal transplant patients with rejection (6.40 +/- 1.47 pmol citrulline/min/mg protein) or with urinary tract infection (29.56 +/- 11.00 pmol citrulline/min/mg protein), when compared to post-renal transplantation patients without rejection or urinary tract infection (0.51 +/- 0.21 pmol citrulline/min/mg protein). Nitrate levels increased in renal transplant patients with rejection and nitrite levels increased in renal transplant patients with urinary tract infection (UTI). Cyclic GMP levels increased with both rejection and UTI. This study demonstrates the presence of NOS activity and inducible NOS-mRNA in cells isolated from the urine of patients undergoing renal allograft rejection.