A total of 12 pulses were utilized to produce indigestible proteins following different processing treatments. Black-eye bean after gelation (slow-cooling) had the highest yield (67.84%) and dry heated (24 h) moong dal washed had the least (0.49%). The indigestible proteins had in vitro bile acid-binding property that was dependent on treatment but not pulse variety. Surface hydrophobicity (So) of the indigestible proteins was significantly different but there was no correlation between So and bile acid-binding ability. Native and indigestible black-eye bean proteins were selected for rats feeding experiment based on yield and bile acid-binding activity. Addition of 5% of indigestible protein reduced plasma triglyceride and total cholesterol levels when compared to casein diet in female rats but not in males. We conclude that indigestible black-eye bean protein may be effective for lowering plasma cholesterol, indicating a potential function as cholesterol-lowering reagent for certain segments of the human population.