Obtaining quantitative information for molecular species present in aerosols from real-time mass spectrometers such as an extractive electrospray time-of-flight mass spectrometer (EESI) and an aerosol mass spectrometer (AMS) can be challenging. Typically, molecular species are calibrated directly through the use of pure standards. However, in some cases (e.g., secondary organic aerosol (SOA) formed from volatile organic compounds (VOCs)), direct calibrations are impossible, as many SOA species can either not be purchased as pure standards or have ambiguous molecular identities. In some cases, bulk OA sensitivities are used to estimate molecular sensitivities. This approach is not sufficient for EESI, which measures molecular components of OA, because different species can have sensitivities that vary by a factor of more than 30. Here, we introduce a method to obtain EESI calibration factors when standards are not available, and we provide a thorough analysis of the feasibility, performance, and limitations of this new technique. In this method, complex aerosol mixtures were separated with high-performance liquid chromatography (HPLC) followed by aerosol formation via atomization. The separated aerosols were then measured by an EESI and an AMS, which allowed us to obtain sensitivities for some species present in standard and SOA mixtures. Pure compounds were used to test the method and characterize its uncertainties, and obtained sensitivities were consistent within ±20 % when comparing direct calibrations vs. HPLC calibrations for a pure standard and within a factor of 2 for a standard mixture. In some cases, species were not completely resolved by chromatography, and positive matrix factorization (PMF) of AMS data enabled further separation. This method should be applicable to other real-time MS techniques. Improvements in chromatography are possible that would allow better separation in complex mixtures.