Optimisation Of Size-Exclusion Chromatography With Diode Array Detector For Aquatic Organic Matter Profiling Using Design Of Experiments: Application To Tropical Reservoirs

In this article, a size-exclusion chromatography with diode array detector (HPSEC-DAD) method was developed to determine the molecular weight distribution of natural aquatic organic matter. First, a variable screening was performed using sodium polystyrene sulphonate (PSS) standards, verifying the significant effects of ionic strength and flow rate of the mobile phase and the injection volume on resolution and final retention time. The method was then optimised using a Box - Behnken design (BBD), obtaining optimal values of 60 mM (ionic strength), 70 mu L (injection volume) and 1.2 mL/min (flow rate). The process was repeated with the same design, using natural water as analyte and introducing peak area as a response, achieving optimum values of 30 mM (ionic strength), 60 mu L (injection volume) and 1.17 mL/min (flow rate). In view of this discrepancy, the results of both optimal methods, using standard humic acid, were compared with the results reported in the literature, resulting in lower relative errors with the PSS method than with the natural water method. Under PSS method conditions, different samples of natural water were analysed, and it was possible to distinguish between the molecular weight profiles of organic matter coming from different environments.