## 2 Materials and Methods  
Six sampling campaigns were conducted in the KRA River estuary (Croatia) located in front of Sibenik town (N 43 deg44''9.509'', E 15 deg52''37.145''): 28 July 2018; 11 February and 31 July 2019; 24 July and 20 September 2020; 7 April 2021. Samples of the vertical column profile were collected by a 'butterfly'-type vertical water sampler or van Dorm-type horizontal sampler (alfa or beta, Wildco) at 6 depths (M1-M6), selected according to the measured vertical salinity profiles (using the EXO2 multiparameter CTD probe, YSI, Yellow Springs, USA) and the total water column depth: M1 and M2 in FWL, M3 and M4 in FSL, M5 and M6 in SWL. Exact depths are readable in the vertical profiles of trace metals that are presented in Figure 1.  
After the sampling, samples were filtered using pre-cleaned cellulose-nitrate capsule filters of nominal pore sizes of 5, 1.2 and 0.1 \(\upmu\)m ([PERSON], [PERSON]). The truly dissolved fraction was separated by centrifugal ultrafiltration (4000 rpm, 30 min; Eppendorf centrifuge, model 5804) through a polyethersulfone (PES) ultrafiltration membrane, with a nominal molecular weight of 3 kDa ([PERSON], Vivaspin 15 Turbo ultrafiltration spin columns). An ultrafiltered sample was obtained from the previously filtered sample using a 0.1 \(\upmu\)m pore size. Repeated tests showed that the selected membrane and syringe filters were contamination free on trace metals, since the ultrapure (MilliQ) water (18.2 M\(\upmu\)  
Figure 1: Depth profiles of the concentrations of studied trace metals in the total fraction (unfiltered sample). Dashed lines separate the three distinct layers, the freshwater layer (FWL), freshwater-seawater interface (FSI) and seawater layer (SWL), as indicated in the first graph. Synthesis A10, Millipore, Burlington, VT, USA) filtered through the pre-cleaned filters showed no difference in trace metal concentrations compared to the unfiltered MilliQ water. The ultrafiltration system, however, showed the gain of dissolved organic carbon and absorbance peak during filtration, even though PES was proposed as the best membrane material for the filtration of dissolved organic matter [36]. A possible source of dissolved organic carbon contamination is leaching from the polypropylene holder under the slightly increased temperature (up to 30 \({}^{\circ}\)C) during the centrifugation. Due to this reason, planned analysis separating the colloidal and truly dissolved organic matter was not possible. Filtered and unfiltered samples were acidified by the addition of 0.2% \(v/v\) HNO\({}_{3}\) (69% \(w/w\) HNO\({}_{3}\) ROTIPURAN Supra, [PERSON]) to a pH < 2 and were UV-irradiated (250 W high pressure Hg lamp) for at least 24 h in order to decompose the natural organic matter. Samples were analyzed using stripping voltammetry (Zn, Cd, Pb, Cu, Ni and Co) and HR-ICP-MS (Zn, Cu, Mn, Fe and Al). A certified reference material \"Nearsbore seawater Reference Material for Trace Metals\" CASS-5 (National Research Council of Canada) was used for validation of the analysis. Volumetric measurements were carried out in a three-electrode cell (663 VA Stand, Metrohm, Herisau, Switzerland) controlled by Metrohm-Autolab (Eco Chemie, Utrecht, The Netherlands) potentiostat/galvanostat (\(\mu\)Autolab Type III or PGSTAT128N) in conjunction with the IME663 module (Metrohm-Autolab, Herisau, Switzerland) interfaced with a GPES (General Purpose Electrochemical System) v.4.9 software (Eco Chemie). The three-electrode system consisted of (i) a hanging mercury drop (HMDE; MME, Metrohm) (drop size 2:0.40 mm\({}^{2}\)), serving as the working electrode, (ii) a Pt wire, serving as the counter electrode and (iii) a Ag \(|\) AgCl \(|\) sat. NaCl reference electrode. A home-made quartz electroanalytical cell with a working volume of 10-50 mL was used. In order to remove reactive dissolved oxygen, the sample (10-12 mL) was purged with nitrogen, for at least 3 min prior to the first measurement, and for 15-30 s between separate measurements in the same sample.