A diacylglycerol lipase that produces the endocannabinoid 2-arachidonoylglycerol was purified from rat brain. Among the proteins identified from the partially purified preparation, only DDHD2, conventionally regarded as phospholipase A1, showed DG lipase activity. Enzymatic properties and immunohistochemical analysis concluded that DG lipase activity was derived from DDHD2.