Abstract: This paper studied proanthocyanidins in callus of Acacia mearnsii De Wilde. Callus were obtained by inoculating leaf explants on MS medium, and supplemented with the combination of 6-BA and 2,4-D. The content of total phenolic compounds and proanthocyanidins in the 12 different treatments were tested by Folin-Ciocalteu assay and vanillin assay, respectively. Proanthocyanidins in callus were characterized by ESI-MS. The results showed that the leaf explants of A.mearnsii treated with 0.25 mg/L 6-BA in combination with 2.0 mg/L 2,4-D resulted in a higher rate of callus formation, better growth and maximum content of total phenolic compounds 20.19 mg/g and proanthocyanidins 11.10 mg/g, which is suitable for proliferation culture. We detected monomer, dimer and trace trimer in callus of A.mearnsii, too.