HPLC analysis revealed that eight subsidiary colors existed in commercial Food Green No. 3 (fast green FCF, FD & C Green No. 3). Among them, four subsidiary colors C, F, G, and H were isolated by using preparative HPLC and their structures were determined by MS and NMR. They were the disodium salt of 2-[[4-[N-ethyl-N-(3-sulfophenylmethyl)amino]phenyl][4-[N-ethyl-N-(4-sulfophenylmethyl)amino]phenyl]methylio]-4-hydroxybenzenesulfonic acid (abbreviated as m,p-G-3), the sodium salt of 2-[[(4-N-ethylamino)phenyl][4-[N-ethyl-N-(3-sulfophenylmethyl)amino]-phenyl]methylio]-4-hydroxybenzenesulfonic acid [abbreviated as HSBA-(EA) (m-EBASA)], the sodium salt of 2-[[(4-N-diethylamino)phenyl][4-[N-ethyl-N-(3-sulfophenylmethyl)amino]phenyl]methylio]-4-hydroxybenzenesulfonic acid [abbreviated as HSBA-(di-EA) (m-EBASA)], and the sodium salt of 2-[[4-[N-ethyl-N-(phenylmethyl)amino]phenyl][4-[N-ethyl-N-(3-sulfophenylmethyl)amino]phenyl]methylio]-4-hydroxybenzenesulfonic acid [abbreviated as HSBA-(EBA) (m-EBASA)], respectively. HSBA-(di-EA) (m-EBASA) was a subsidiary color newly found in commercial Food Green No. 3.