Single-photon ionization (SPI) using vacuum ultraviolet (VUV) light produced by an electron beam pumped rare gas excimer source has been coupled to a compact and mobile time-of-flight mass spectrometer (TOFMS). The novel device enables real-time on-line monitoring of organic trace substances in complex gaseous matrixes down to the ppb range. The pulsed VUV radiation of the light source is employed for SPI in the ion source of the TOFMS. Ion extraction is also carried out in a pulsed mode with a short time delay with respect to ionization. The experimental setup of the interface VUV light source/time-of-flight mass spectrometer is described, and the novel SPI-TOFMS system is characterized by means of standard calibration gases. Limits of detection down to 50 ppb for aliphatic and aromatic hydrocarbons were achieved. First on-line applications comprised real-time measurements of aromatic and aliphatic trace compounds in mainstream cigarette smoke, which represents a highly dynamic fluctuating gaseous matrix. Time resolution was sufficient to monitor the smoking process on a puff-by-puff resolved basis. Furthermore, human breath analysis has been carried out to detect differences in the breath of a smoker and a nonsmoker, respectively. Several well-known biomarkers for smoke could be identified in the smoker's breath. The possibility for even shorter measurement times while maintaining the achieved sensitivity makes this new device a promising tool for on-line analysis of organic trace compounds in process gases or biological systems.