Two different endopeptidases which digest the chromogenic substrate Azocoll were found in soybean leaves. Azocoilase A has a molecular weight of 17,500 and a pI of 6.0. Azocollase B has a molecular weight of 52,000 and a pl of 9.0. Both digest Azocoll optinally at pH 9.0. Azocollase A is inhibited by 3 milimlar ethylenediamine tetraacetate (EDTA) the characteristics of the proteinases, leaf age was not rigorously controlled. Young leaves are defined as those which have just expanded, mature (eaves are those which have been on the plant at least 2 weeks after reaching full size, and senescent leaves are those which show the first visible signs of yellowing. The leaves of greenhouse-grown plants were cut into small pieces and homogenized in a Polytron homogenizer (Kinematica, Luzerne, Switzerland) for 1 min at 3 C in 4 volumes of extraction medium. The extraction medium consisted of 50 mm K-phosphate (pH 7.5) with 1% insoluble PVP (Sigma) and 20 mM sodium metabisulfite.The leaves of field-grown plants were cooled to 3 C, transported to the laboratory in ice chests and processed immediately. Each sample was composed of the leaves from eight plants and four replicated samples were obtained and analyzed. Nodes 5 through 8 (see under "Results") represent the oldest leaves, whereas nodes 13 through 16 represent the youngest leaves on the same plant. The leaves were punched with a cork borer (9-mm diameter) and a known number of discs (200-300) were weighed. The weight of 100 leaf discs was relatively constant throughout the season, increasing about 10% from the 9th week to the 14th week, then dropping 20%1o in the 15th week. The weight of 100 leaf discs taken from leaves at the top of the canopy (nodes 13-16) was 5 to 10% greater than the weight of leaf discs taken from nodes 5 through 8. The leaf discs were frozen in liquid N2, and ground in a cold mortar and pestle. The resulting powder was mixed with 4 volumes of extraction medium containing 50 mm K-phosphate buffered at pH 7.5, 1% insoluble PVP, and 0.1% fB-mercaptoethanol. The homogenates were strained through cheesecloth, centrifuged for 10 min at 1,200g and the supernatant used as a source of enzyme.For studies which did not necessitate determinations of Chl and protein content of the leaves, homogenates were centrifuged at 857 www.plantphysiol.org on May 9, 2018 -Published by Downloaded from