Distance-based detection (DbD) on paper-based microfluidic analytical devices (μPADs) has emerged as a promising, cost-effective, simple, and instrumentation-free assay method. Broadening the applicability of a new way of immobilization of reagent for DbD on μPADs (DμPADs) is presented, employing an ion exchange (IE) interaction of an anionic metallochromic reagent, 2-(5-bromo-2-pyridylazo)-5-[N-n-propyl-N-(3-sulfopropyl)amino]phenol (5-Br-PAPS), on the anion-exchange filter paper. The IE DμPADs demonstrate superiority over standard cellulose filter paper in terms of the degree of reagent immobilization, detection sensitivity, and clear detection endpoints due to the strong retention of 5-Br-PAPS. The study investigated various parameters influencing DbD, including 5-Br-PAPS concentrations (0.25-1 mM), buffer types (acetic acid-Tris, MES), buffer concentrations (20-500 mM), and auxiliary complexing agents (acetic, formic, and glycolic acids). Subsequently, the performance of 17 metals (Ag+, Cd2+, Co2+, Cr3+, Cu2+, Fe2+, Hg2+, La2+, Mn2+, Ni2+, Pb2+, Ti2+, Zn2+, Al3+, As3+, Fe3+, and V4+) was evaluated, with color formation observed for 12 metals. Additionally, the paper surface was examined using SEM and SEM-EDX to verify the suitability of certain areas in the detection channel for reagent immobilization and metal binding. This method demonstrates quantitation limits of metals in the low μg mL-1 range, showing great potential for the rapid screening of toxic metals commonly found in herbal supplements and cosmetics regulated by the Food and Drug Administration (FDA). Thus, it holds promise for enhancing safety and regulatory compliance in product quality assessment. Furthermore, this method offers a cost-effective, environmentally sustainable, and user-friendly approach for the rapid visual quantification of heavy metals for in-field analysis, eliminating the need for complex instrumentation.

Central European Institute of Technology Brno University of Technology Purkynova 123 CZ 612 00 Brno Czech Republic

Department of Chemistry and Biochemistry Mendel University in Brno Zemedelska 1 CZ 613 00 Brno Czech Republic

Department of Pharmaceutical Chemistry Faculty of Pharmacy Mahidol University 447 Sri Ayudhaya Rd Rajathevee Bangkok 10400 Thailand

School of Natural Sciences and Australian Centre for Research on Separation Science University of Tasmania Private Bag 75 Hobart 7001 Australia

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