In the present study, an ultra performance liquid chromatography coupled with time-of-fight mass spectrometry (UPLC-TOF/MS) based chemical profiling approach was used to evaluate chemical constitution between co-decoction and mixed decoction of ginseng and Radix Aconiti Praeparata. Two different kinds of decoctions, namely co-decoction of ginseng and Radix Aconiti Praeparata: water extract of mixed two herbs, and mixed decoction of ginseng and Radix Aconiti Praeparata: mixed water extract of each individual herbs, were prepared. Batches of these two kinds of decoction samples were subjected to UPLC-TOF/MS analysis. The datasets of t(R) m/z pairs, ion intensities and sample codes were processed with supervised partial least squared discriminant analysis (OPLS-DA) to holistically compare the difference between these two decoction samples. Significant difference between the two decoction samples was showed in the results of positive ion mode. The contents of hypaconitine and deoxyaconitine decreased, while that of benzoylmesaconine, benzoylhypaconine and dehydrated benzoylmesaconine increased in the samples of co-decoction of ginseng and Radix Aconiti Praeparata. The content of diester-diterpenoid alkaloids decreased, while that of monoester-diterpenoid alkaloids increased, which is probably the basis of toxicity-attenuated action when combined ginseng with Radix Aconiti Praeparata.