As a continuation of our research on the development of pesticide active quinolizidine alkaloids (QAs) from the family Fabaceae, the chemical constituents of the root of Sophora tonkinensisGagnep. were systematically investigated. Seventeen new matrine-type alkaloids (1–17), including one new naturally occurring compound (17), along with 20 known ones were isolated from the EtOH extract of S. tonkinensis. Notably, compound 5possessed an unprecedented 6/6/5/4/6/6 hexacyclic system. Their structures were confirmed via comprehensive spectroscopic data analysis (IR, UV, NMR, HRESIMS), ECD calculation, and X-ray crystallography. Biological tests indicated that compounds 1, 4, 10, 12, 13, and 30displayed significant anti-tomato spotted wilt virus (TSWV) activities compared with the positive control ningnanmycin. Moreover, compound 12strongly inhibited the expression of the TSWV N, NSs, and NSmgenes and TSWV NSs protein in plant host. Furthermore, compounds 4, 10, 12, 20, and 22exhibited moderate insecticidal activities against TSWV thrip vector (Frankliniella occidentalis).