Isotope hydrograph separation (IHS) is a valuable tool to study runoff generation processes. To perform an IHS, samples of baseflow (pre-event water) and streamflow are taken at the catchment outlet. For rainfall (event water) either a bulk sample is collected or it is sampled sequentially during the event. For small headwater catchment studies, event water samples are usually taken at only one sampling location in or near the catchment because the spatial variability in the isotopic composition of rainfall is assumed to be small. However, few studies have tested this assumption. In this study, we investigated the spatiotemporal variability in the isotopic composition of rainfall and its effects on IHS results using detailed measurements from a small pre-alpine headwater catchment in Switzerland. Rainfall was sampled sequentially at eight locations across the 4.3 km2 Zwäckentobel catchment and stream water was collected in three subcatchments (0.15, 0.23, and 0.70 km2) during ten events. The spatial variability in rainfall amount, average and maximum rainfall intensity and the isotopic composition of rainfall was different for each event. There was no significant relation between the isotopic composition of rainfall and total rainfall amount, rainfall intensity or elevation. For eight of the ten studied events the temporal variability in the isotopic composition of rainfall was larger than the spatial variability in the rainfall isotopic composition. The isotope hydrograph separation results, using only one rain sampler, varied considerably depending on which rain sampler was used to represent the isotopic composition of event water. The calculated minimum pre-event water contributions differed up to 60%. The differences were particularly large for events with a large spatial variability in the isotopic composition of rainfall and a small difference between the event and pre-event water isotopic composition. Our results demonstrate that even in small catchments the spatial variability in the rainfall isotopic composition can be significant and has to be considered for IHS studies. Using data from only one rain sampler can result in significant errors in the estimated pre-event water contributions to streamflow.