Adenosine deaminase (ADA) is suggested to be an important enzyme for modulating the bioactivity of insulin, but its clinical significance in diabetes mellitus (DM) is not yet characterized. We measured the serum levels of ADA isoenzymes (ADA1 and ADA2) in healthy donors (HD, n = 52), insulin-dependent diabetes mellitus (IDDM, n = 53) patients and non-insulin-dependent diabetes mellitus (NIDDM, n = 65) patients. The mean serum level of ADA1 in HD, IDDM or NIDDM patients was, respectively 6.5, 8.1 or 9.5 units/l (P < 0.001 vs. HD) and that of ADA2 in HD, IDDM or NIDDM patients was 7.0, 14.9 (P < 0.001 vs. HD) or 11.2 units/l (P < 0.001 vs. HD), respectively. Normalization of the blood glucose level by the hospitalization was associated with the decrease in ADA2 (but not ADA1) activity in 6 of 8 IDDM or 11 of 12 NIDDM poorly controlled patients. ADA2 (but not ADA1) activity in the poorly controlled NIDDM patients directly correlated with the hemoglobin A1c level (P < 0.002). Measurement of serum ADA2 activity may be important to better understand the clinical aspects of both IDDM and NIDDM. The pathogenic role of elevated ADA activity in the sera of DM patients was addressed.