Nitric oxide (NO) has a much stronger affinity for hemoglobin than carbon monoxide (CO); therefore, the DL(NO) (diffusing capacity for NO) is less influenced by changes in capillary blood volume than the DL(CO) (diffusing capacity for CO), and represents the true membrane diffusing capacity. We measured the combined single breath DL(NO)/DL(CO) in 124 healthy subjects, and generated reference equations for the DL(NO) and K(NO). In a subset of 21 subjects the measurements were performed on different inspiratory levels. The reference equation for DL(NO) in females is 53.47*H(height)0.077*A(age)-48.28(RSD5.22) and for males 59.84*H-0.25*A-44.20(RSD6.39). Reference equations for K(NO) in females is -2.03*H-0.025*A+11.52(RSD0.48) and for males -0.15*H-0.045*A+9.47(RSD0.65). The K(CO) (DL(CO)/V(A)) increases when V(A) (alveolar volume) decreases, probably due to an increase of blood volume per unit lung volume. The DL(NO) was much stronger related to the V(A), the K(NO) was almost independent of V(A). Because of the relative independence of the K(NO) on V(A), the K(NO) appears to be a much better index for the diffusion capacity per unit lung volume (transfer coefficient) than the K(CO).