Diuron-induced photosynthesis inhibition of freshwater biofilms and subsequent EC50s (half maximal effective concentrations) are determined using a 3-hour exposure in average. However, given the known rapid action of diuron, earlier responses can be expected. In this study, we performed different time-response experiments to confirm the short time lag for diuron to reach its cellular target, and to assesss whether time-response curves (i.e. photosynthesis inhibition as a function of time, when diuron exposure is kept constant) can improve the assessment of short-time (pulse) pre-exposure to diuron. Our results demonstrated that: i) diuron reaches its cellular target in a very short time-span (100µg.L-1) to 7:20 minutes (exposure to EC25). We were also able to demonstrate previous pulsed (3 hours) exposure to increasing concentrations of diuron (0, 5 and 50µg.L-1) using time-responses curves, providing a new, easy-to-implement, method to assess short-term exposure to PSII inhibitors based on community photosynthesis.