Evaluating transit time by using 18O seasonal variation is often difficult in a Mediterranean context due to the erratic rainfall signature added to the complexity of flow mixing. The present study aims to show that using organic matter can improve interpretations of the 18O signal. The natural fluorescence of organic compounds and dissolved organic carbon (DOC) data were recorded in the underground low-noise laboratory, located within the catchment area of the Fontaine de Vaucluse karstic system, over a four-year period. By combining both total fluorescence and DOC, a systematic seasonal variation is observed only due to soil-water interaction. Therefore, when the recharge rate is enough at the time of the season concerned, a new specific seasonal tracer, independent of rainfall signature, is available. Besides, the DOC concentration is a tracer of rapid infiltration with short transit time associated with high DOC concentration, while long transit time waters are characterised by a low DOC concentration. Then this can also shed light on such a mixing, recent/old waters. A more sensitive interpretation of 18O variations is then possible: if the rainfall amount and 18O follow a seasonal trend, both tracers may be used in the same way; if the recharge is discontinuous due to discontinuous rainfall regime, 18O tracing alone is usable; in case of erratic or smooth rainfall signature during the homogeneous rainy period, fluorescent organic matter tracing alone is then usable.