Acid whey (AW), a by-product from the production of acidified dairy products, contains high amounts of lactic acid and minerals that can be recovered by electrodialysis (ED). To better understand the process and improve its efficiency, the objective of this study was to investigate fouling of ion-exchange membranes (IEMs) during ED of AW and concentrated AW by reverse osmosis (ROAW), underlimiting (ULCD), limiting (LCD) and overlimiting current density operating conditions (OLCD). The structure, hydrophobicity, and chemical composition of membranes showed differences regarding fouling on anion- (AEM) and cation- (CEM) exchange membranes facing the diluate and the concentrate, both for AW and ROAW. Furthermore, operating at OLCD tends to reduce fouling compared to ULCD, due to the expected generation of electroconvective vortices. 2D fluorescence spectroscopy and Fourier-transform infrared spectroscopy (FTIR) provided complementary and more detailed information regarding the fouling and efficiency of the cleaning procedure. The 2D fluorescence spectra showed that the AEM surfaces in contact with the diluate change more than those in contact with the concentrate. The FTIR analyses showed the presence of lactose and lactic acid on the AEM surfaces in contact with the concentrate, which could not be detected by fluorescence.