The Huangqi-Sanqi herb pair (HS), a compatibility formula of Astragalus membranaceus (HQ) and Panax notoginseng (SQ) with a weight ratio of 3 : 1, has been used in the treatment of cardiovascular and kidney diseases. In this study, an ultra-high-performance liquid chromatography coupled with triple quadrupole electrospray tandem mass spectrometry (UHPLC-TQ-MS/MS) method has been developed and validated for the simultaneous determination of fourteen constituents, including six isoflavonoids, three astragalosides and five panax notoginsenosides in HS, HQ and SQ for the first time. Quantification was carried out using multiple reaction monitoring (MRM) via an electrospray ionization source operating in both positive and negative ionization mode. All of the constituents were separated and determined within 9 minutes with linear gradient elution of acetonitrile-0.1% (v/v) formic acid in water. The approach was validated in terms of linearity, sensitivity, precision, and repeatability as well as recovery. The developed method was successfully applied to inspect the content changes of individual components in HS with different ratios. The results showed that water extracts of HS could better increase the dissolution of isoflavonoids and astragalosides in comparison with a single herb. Additionally, water extracts of HS exhibited rising and then falling content of total panax notoginsenosides with increasing proportionality coefficient. The study might provide a basis for drug interaction during the decoction process of herb pairs based on the content changes of the marker components.