Inhibition of the Na/H exchanger is a promising approach for treating ischemia-reperfusion injury, but no clinical agent is yet available. Recently, we established the structural requirements for potent inhibitors of the Na/H exchanger. In the present work, we designed N-(3-oxo-3,4-dihydro-2H-benzo[1,4]oxazine-7-carbonyl)guanidine 3a as a new lead compound for potent inhibitors with good water-solubility, based on the previous information. During the structural optimization, care was taken to keep the hydrophobicity (clogP) in the range of about 1.5-2.0, which is considered optimum for good bioavailability. Various derivatives of 3a were synthesized and the quantitative structure-activity relationship (QSAR) was studied. The QSAR result indicated that the lengths of the substituents at the 2- and the 4-positions of the 2H-benzo[1,4]oxazine ring are parabolically related to activity. The most potent compounds were (R) and/or (S)-N-(2-ethyl-4-isopropyl(or ethyl)-3-oxo-3,4-dihydro-2H-benzo[1,4]oxazine-7-carbonyl)guanidines 3q-t with IC50 values of 0.036-0.073 microM. The water-solubility of the hydrochlorides and methanesulfonates is 3-5 mg/ml, which is sufficient for therapeutic use.