Changes in phosphatic metabolites and intralenticular pH of the intact crystalline rabbit lens induced by ouabain (10<sup>––3</sup>, 10<sup>––4</sup>, 10<sup>––5</sup> M) were measured during time-course incubations using phosphorus-31 nuclear magnetic resonance spectroscopy (<sup>31</sup>P NMR) in order to study the metabolic manifestations of this steroid on lens glycolytic activity. Significant alterations in the glycolytic intermediates of the crystalline lens only occurred with ouabain at the concentration of 10<sup>––3</sup> M. A significant time-dependent increase in α-glycerophosphate, an unidentified pentose phosphate, the nucleoside diphosphosugars (uridine diphosphoglucose and uridine diphosphomannose), inorganic orthophosphate, and inosine monophosphate, and decline in glucose 6-phosphate and ATP were detected in 10<sup>––</sup><sup>3</sup> M ouabain-treated lenses. The uridine diphosphosugars exhibited a maximum at approximately the time when the lens ATP content was 50% of control levels (12.5 h). A progressive decrease in intralenticular pH indicative of hydrogenion-pump damage was detected concomitant with the changes in lens metabolite levels. Lenses maintained transparency throughout the time-course. Since metabolite manifestations effecting the intermediates of glycolysis occur in lens tissue with its high glycolytic and low oxidative activity, we hypothesize the effects of 10<sup>––3</sup> M ouabain may represent metabolic actions of ouabain which are independent of its effects on tissue oxygen consumption as is presumed from studies involving tissues more dependent on oxidative metabolism.