ISSN print edition: 0366-6352 ISSN electronic edition: 1336-9075 Registr. No.: MK SR 9/7 Published monthly

In vitro and in silico evaluation of human serum albumin binding, blood-brain barrier permeability, and acetylcholinesterase inhibition by multi-target compounds with potential anti-Alzheimer’s disease properties

Mai Zahran, Adrian Guin-Rizzo, Irina Solomon, Johnny Guevara, Rosemary Pichardo-Bueno, Gabriel Ortiz, Ebbe Nordlander, and Alberto Martínez

Department of Biological Sciences, New York City College of Technology, The City University of New York (CUNY), Brooklyn, USA

E-mail: Mzahran@citytech.cuny.edu

Received: 8 December 2025  Accepted: 2 February 2026

Abstract:

Alzheimer’s disease (AD) continues to pose a major global health challenge, with incidence rising sharply and current therapies offering only symptomatic relief. This study builds on prior work into six structurally diverse multi-target compounds (1–6) designed to modulate key pathological processes in AD. We have now combined experimental and computational approaches to determine a set of pharmacological properties, focusing on interactions with human serum albumin (HSA) as a potential plasma carrier, blood-brain barrier (BBB) permeability, and acetylcholinesterase (AChE) inhibition. Fluorescence and circular dichroism (CD) spectroscopic assays revealed stable single-site binding to HSA without perturbation of protein secondary structure, while calculated unbound fractions remained within therapeutic ranges. BBB permeability and potential efflux were assessed using both in vitro (PAMPA-BBB) and in silico approaches, and showed that transport was strongly modulated by structural features within the compound series. Molecular docking and molecular dynamics simulations predicted preferential binding within the hydrophobic cavity of HSA subdomain IIIA, consistent with typical binding sites of numerous pharmaceuticals. Among the series, compounds 5 and 6 displayed the most favorable profiles, combining effective BBB permeability, suitable unbound fractions, and moderate low-micromolar AChE inhibition supported by predicted interactions with active-site residues of the enzyme. Together, these findings expand upon earlier work by allowing to further establish structure-activity relationships and highlighting compounds 5 and 6 as promising scaffolds for the rational design of next-generation AD therapeutics.

Graphical abstract

Keywords: Polyphenol; Multi-target heterocyclic compound; Molecular design; Docking; MM/GBSA binding free energy ΔG; Alzheimer’s disease

Full paper is available at www.springerlink.com.

DOI: 10.1007/s11696-026-04713-9

Chemical Papers 80 (5) 5331–5350 (2026)