Chromene Flavonones from Dalea boliviana as Xanthine Oxidase Inhibitors: in vitro biological evaluation and molecular docking studies

Einy Nallybe Bedoya Aguirre ^1,2 Maria Daniela Santi ³ Melisa Fabiana Negro ^1,2 Javier Echeverria ^4* Margot Paulino Zunini ⁵ María Andrea Peralta ^1,2 María Gabriela Ortega ^1,2*

• ¹ Unidad de Investigación y Desarrollo en Tecnología Farmacéutica (UNITEFA-CONICET), Ciudad Universitaria, Cordoba, Argentina
• ² Farmacognosia, Departamento de Ciencias Farmacéuticas, Facultad de Ciencias Químicas, Universidad Nacional de Córdoba, Ciudad Universitaria, Haya de la torre y Medina Allende, Edificio Ciencias II, Cordoba, Argentina
• ³ Max Planck Institute for Multidisciplinary Sciences, Göttingen, Lower Saxony, Germany
• ⁴ Departamento de Ciencias del Ambiente, Facultad de Química y Biología, Universidad de Santiago de Chile, Santiago, Santiago Metropolitan Region (RM), Chile
• ⁵ Área Bioinformática, DETEMA, Facultad de Química, Universidad de la República, Montevideo, Uruguay

Background: Prenylated flavanones represent a structurally diverse class of natural compounds with significant biological potential. Among them, chromene flavanones constitute a rare and specialized subgroup with promising therapeutic applications.These molecules have gained attention for their potential to inhibit xanthine oxidase (XO), a key enzyme involved in oxidative stress-related disorders such as gout and hyperuricemia. Given their potential relevance, this study focuses on the in vitro and in silico evaluation of three chromene flavanones (CFs) isolated from Dalea boliviana Britton [Fabacea], assessing their capacity to inhibit XO and elucidating key structure-activity relationships that contribute to their biological effectiveness.Purpose: This work attempts to study in vitro and in silico interactions of the chromene flavanones (2S) 5,2′-dihydroxy-6″,6″-dimethylchromeno-(7,8:2″,3″)-flavanone (1), (2S) 5,2'-dihydroxy-6'',6''-dimethylchromeno-(7,8:2'',3'')-3'-prenylflavanone(2), and obovatin (3), obtained from Dalea boliviana, with the XO to propose them as candidates for inhibiting its activity, and potential therapeutic approaches for hyperuricemic-diseases.Material and Methods: XO inhibition of three chromene flavanones was measured spectroscopically. The relationships between the structure and the inhibitory activity were evaluated. Moreover, molecular docking studies were performed to propose the binding mode of the most active natural compound.Results and Discussion: Compounds 1 and 2 exhibited potent inhibition, with IC50 values in the nanomolar range (0.5 ± 0.01 nM and 1.7 ± 0.46 nM, respectively), demonstrating significantly higher activity compared to allopurinol, the reference inhibitor (IC50 = 247 ± 4 nM). In contrast, compound 3 displayed only weak inhibition. Structure-activity relationship (SAR) analysis revealed that the presence of a chromene moiety in the A-ring, in combination with hydroxyl and prenyl groups in the B-ring, played a crucial role in enhancing inhibitory activity. Molecular docking studies confirmed the strong binding affinities of compounds 1 and 2 within the active site of XO (PDB ID: 3NVY), with binding energies of -6.1687 and -6.7820 kcal/mol, respectively. Key stabilizing interactions involved π-π interactions with Phe914 and hydrogen bonding with residues such as Leu873 and Leu1014. Conclusion: These findings highlight the structural features essential for potent XO inhibition and suggest that chromene flavanones represent a valuable scaffold for the development of novel inhibitors.