Two new flavonoids, 2',4',6',2,3,4-hexamethoxychalcone (1) and 5-hydroxy-7,2',5'-trimethoxyflavone (2) together with a known flavone glycoside, echioidinin 5-O-beta-D-glucopyranoside (3) were isolated from the whole plant of Andrographis neesiana, and the structures were elucidated by electrospray ionization tandem mass spectrometry (ESI-MS/MS) and one- and two-dimensional (1D- and 2D)-NMR spectral studies including 1H-1H correlation spectroscopy (COSY), heteronuclear single quantum coherence (HSQC), heteronuclear multiple bond connectivity (HMBC) and nuclear Overhauser enhancement spectroscopy (NOESY) experiments.