Secondary polyphenol metabolites, urolithins (UROs), have anti-oxidative, anti-inflammatory, and antidiabetic properties. Therefore, their biological activity relies on blood transport via human serum albumin (HSA) and tissue distribution. The main goal we set was to investigate the interaction between HSA and different URO (URO A, URO B, URO C, URO D, and glucuronidated URO A and B) using a combination of multi-spectroscopic instrumental and in silico approaches. The fluorescence spectroscopy revealed that URO can quench the naturally occurring fluorescence of HSA in a concentration-dependent manner. The HSA fluorescence was quenched by both a static and dynamic mechanism. The results showed that free UROs bind to HSA with higher affinity than their conjugated forms. CD spectroscopy and FTIR revealed that the alpha-helical structure of HSA is preserved. The calculated Gibbs free energy change indicates that the URO–HSA complex forms spontaneously. There is a single binding site on the HSA surface. The molecular docking results indicated that unconjugated Uro binds to Sudlow I, while their conjugation affects this binding site, so in the conjugated form, they bind to the cleft. Docking experiments indicate that all UROs are capable of binding to both thyroxine recognition sites of ligand-bound HSA proteins. Examining interactions under the following conditions (298 K, 303 K, and 310 K, pH 7.4) is of great importance for determining the pharmacokinetics of these bioactive compounds, as the obtained results can be used as a basis for modulating the potential dosing regimen.

Over the past decade, research on urolithins has expanded significantly due to their role as mediators between polyphenol-rich diets and human health. Understanding the relationships between ellagitannin intake, gut microbiota composition, and urolithin production is essential for evaluating their biological effects and nutraceutical potential. The primary objective of this review is to critically summarise current knowledge on urolithins, bioactive metabolites derived from ellagitannins in plant-based foods, with a focus on their biosynthesis, bioavailability, protein interactions, and potential therapeutic applications. A comprehensive literature search was conducted using PubMed, Scopus, and Google Scholar to identify studies on urolithin biosynthesis, absorption, transport mechanisms, protein binding, and incorporation into extracellular vesicles. Relevant articles were critically analysed to synthesise current evidence and highlight emerging concepts. Key findings indicate that after absorption, urolithins bind to serum albumin, which facilitates their transport to target tissues, exerting anti-inflammatory and antioxidant actions. Recent evidence also shows that urolithins can be packaged into extracellular vesicles, suggesting novel mechanisms for intracellular transport and potential therapeutic applications. This review highlights gaps in current knowledge and proposes directions for future research to optimise their therapeutic potential.