Nanoformulations based on Pistacia vera L. with immunomodulatory activity

The use of specialized plant metabolites represents a novel approach in modern medicine, aimed at harnessing naturally occurring resources, including waste products from processing industries. Given the growing interest in Pistacia vera L., in this study pistachio stems, which are typically discarded, were examined for the first time. The aim is to maximize the use of raw materials, reduce waste, and enhance their benefits, aligning with the concept of "circular bioeconomy." Furthermore, given the numerous biological properties of the plant, with particular attention to the antiinflammatory activity due to compounds such as α-pinene and myricetin (1), this study reportsthe phytotherapeutic use of two different extracts obtained from pistachio waste products with particular focus on their immunomodulatory effects. To address issues of cytotoxicity and low bioavailability, the potential for incorporating these extracts into nanoformulations, such as liposomes and transferosomes, was assessed. In this study, hydrolate and essential oil were obtained from the stems through hydro distillation. The extracts were characterized using liquid andgas chromatography coupled with mass spectrometry, identifying 21 specialized metabolites, 13 ofwhich were common among the two extracts. These extracts were then formulated into liposomesand transferosomes and characterized for polydispersity index, size, and negative charge. Notably, encapsulation of the extracts within the vesicles reduced their size compared to empty vesicles (2). The prepared transfersomes were larger and more homogeneous than liposomes. Moreover,transfersomes were found to be more stable than liposomes, as they showed no changes in size, homogeneity, and charge after 90 days from the preparation. The encapsulation efficiency of the nanoformulations was evaluated by quantifying the main compounds present in the extracts(naringenin, myristic acid, and catechin), achieving over 80% efficiency. Following this, the immunomodulatory activity of the essential oil and hydrolate on U937 cells stimulated with LPS wasinvestigated. Specifically, the incorporation of the oil into transferosomes significantly enhanced its immunomodulatory effect on macrophages by inhibiting the expression and activity of the ATP citrate lyase (ACLY) enzyme. Additionally, the hydrolate in solution and the essential oil in liposomes reduced the production of ROS and NO• induced by LPS stimulation of the cells. The extracts also demonstrated the ability to inhibit prostaglandin E2 (PGE2) production. Based on these results, it can be concluded that P. vera extracts may play a crucial role in modulating macrophages induced by various stimuli, such as LPS endotoxin, by acting on immunometabolism. Most interestingly, theformulation in transfersomes abolished the cytotoxicity of the essential oil while increasing its abilityto inhibit inflammatory mediators via the immunometabolic citrate pathway.