In the course of our research on the relationship between medicinal application and alkaloid constituents of Chinese drugs, we previously reported the presence of twelve known lupin alkaloids in the roots of Sophora tonkinensis, which have been used as the Chinese drug Shan-Dou-Gen to treat fever, throat inflammation, pain, hemorrhoids, tumors, etc., and reported that a major lupin alkaloid of this plant, (+)-matrine, and its stereoisomer (+)-allomatrine possesses significant antinociceptive activities which are mediated mainly by κ-opioid receptors. As a continuation of these studies, we have examined the constituents of the fresh leaves of this plant and characterized two new alkaloids, (-)-14β-acetoxymatrine and (+)-14α-acetoxymatrine, which are the first ester derivatives of matrine-type lupin alkaloids, together with fourteen known lupin alkaloids: 17-oxo-α-isosparteine, (-)-sophocarpine, (+)-matrine, (-)-14β-hydroxymatrine, (+)-14α-hydroxymatrine, 13,14-dehydrosophoranol (5α-hydroxysophocarpine), (+)-sophoranol, (+)-9α-hydroxymatrine, lamprolobine, (+)-5α,9α-dihydroxymatrine, (-)-baptifoline, (+)-matrine N-oxide, (+)-sophocarpine N-oxide, and (+)-sophoranol N-oxide. This paper describes the structural elucidation of the two new lupin alkaloids and the variations in alkaloid content in the roots and the leaves of S. tonkinensis.