In this work, an integrated microfluidic device was developed for simultaneously characterization of drug metaoblites and a cytotoxicity assay. The multi-layer device was composed of a quartz substrate, a polycarbonate (PC) membran and three PDMS layers which were self-assembled layer by layer. The functional units of drug metabolism and cytotoxicity assays including metabolite generation, sample pretreatment, online detection with an electrospray ionization quadrupole time-of-flight mass spectrometer (ESI-Q-TOF MS) and incubation with cultured cells to evaluate metabolism induced cytotoxicity were all integrated on the microfluidic device. It is noted that by aligning the PC membrane with a microwell on the PDMS layer, the human liver microsomes (HLMs) could be easily immobilized inside the microwell without any damage. To verify the feasibility of our established platform, the metabolism of flavopiridol (FlAP) and the cytotoxicity of metabolites on MCF-7, MCF-10A, HepG2 and QSG7701 cells were studied. The metabolism with the increasing of concentration and time which resulted in the increased cytotoxicity was proved on this device. The total analysis time for one product was about 30 min and only less than 3 μL reaction system was required for each experiment. The results demonstrated that this platform is robust for low levels of compounds and shows potential for high-throughput drug screening in the drug development.

This work was supported by the National Natural Science Foundation of China (Nos. 21172129, 20935002)，International S&T Cooperation Program of China (No. 2011DFA30620), the National High Technology Research and Development Program of China (863 Program) (No. 2013AA092902), and Basic research projects in Shenzhen (No. JCYJ20120831165730898).

The corresponding authors: Dan Gao, Hongxia Liu