A pulse-sequence strategy was developed for generating regional maps of alveolar oxygen partial pressure (pO2) in a single 6-sec breath hold, for use in human subjects with impaired lung function. Like previously described methods, pO2 values are obtained by measuring the oxygen-induced T1 relaxation of inhaled hyperpolarized 3He. Unlike other methods, only two 3He images are acquired: one with reverse-centric and the other with centric phase-encoding order. This phase-encoding arrangement minimizes the effects of regional flip-angle variations, so that an accurate map of instantaneous pO2 can be calculated from two images acquired a few seconds apart. By combining this phase-encoding strategy with variable flip angles, the vast majority of the hyperpolarized magnetization goes directly into the T1 measurement, minimizing noise in the resulting pO2 map. The short-breath-hold pulse sequence was tested in phantoms containing known O2 concentrations. The mean difference between measured and prepared pO2 values was 1 mm Hg. The method was also tested in four healthy volunteers and three lung-transplant patients. Maps of healthy subjects were largely uniform, whereas focal regions of abnormal pO2 were observed in diseased subjects. Mean pO2 values varied with inhaled O2 concentration. Mean pO2 was consistent with normal steady-state values in subjects who inhaled 3He diluted only with room air.