1. Rat kidneys which were perfused with saline contained both kininogenase (KGA) and kininase activity. These activities were separated by gel filtration on a Sephadex G-100 column. The kininase activity was excluded from the column whereas the KGA activity was retained. Kidney KGA activity was primarily found in the sedimentable fraction of the homogenate.2. The kidney KGA activity was compared with the urinary KGA activity, and the following properties were found to be the same: molecular dimension, pH optimum, effect of inhibitors, and ability to liberate kinins from kininogens.3. A urinary sample collected over 24 h contained about 8 times the KGA activity found in the corresponding kidneys at the end of the collection period. The urine: kidney ratio for alkaline phosphatase was about 0.01.4. The ability of kidney and urinary samples to hydrolyse N-alpha-benzoyl-L-arginine ethyl ester (BAEE) at pH 8.5 paralleled the KGA activity.