Effect of a methanolic extract of Salvadora oleoides Decne. on LPS-activated J774 macrophages, its in vitro and in vivo toxicity study and dereplication of its chemical constituents

Hafiz Abdul Khaliq; Sergio Ortiz; Mireille Alhouayek; Tanguy Neyts; Giulio G Muccioli; Joëlle Quetin-Leclercq

doi:10.1016/j.toxrep.2022.09.004

Effect of a methanolic extract of Salvadora oleoides Decne. on LPS-activated J774 macrophages, its in vitro and in vivo toxicity study and dereplication of its chemical constituents

Toxicol Rep. 2022 Sep 8:9:1742-1753. doi: 10.1016/j.toxrep.2022.09.004. eCollection 2022.

Authors

Hafiz Abdul Khaliq^{1

2

3}, Sergio Ortiz¹, Mireille Alhouayek², Tanguy Neyts¹, Giulio G Muccioli², Joëlle Quetin-Leclercq¹

Affiliations

¹ Pharmacognosy Research Group, Louvain Drug Research Institute, UCLouvain, 1200 Brussels, Belgium.
² Bioanalysis and Pharmacology of Bioactive Lipids Research Group, Louvain Drug Research Institute, UCLouvain, 1200 Brussels, Belgium.
³ Department of Pharmacognosy, Faculty of Pharmacy, Bahauddin Zakariya University, Multan 60800, Pakistan.

Abstract

Salvadora oleoides is used in Pakistani traditional medicine to treat inflammatory conditions, piles, boils, and ulcers. To evaluate the anti-inflammatory potential of S. oleoides (a mixture of aerial branches, leaves, and stem bark), we prepared crude extracts in Soxhlet apparatus by successively using different solvents and found the methanolic extract (OLM) to significantly inhibit the LPS-induced expression of pro-inflammatory cytokines and enzymes in J774 macrophages, at 50 µg/mL concentration. We also analysed the chemical constituents of OLM by dereplication, performed by HPLC-MS/MS and molecular networking. The major detected constituents were flavonoids and phenolic acids glycosides, most of them identified for the first time in S. oleoides. We also evaluated the toxicity of OLM against five cell lines, namely Caco-2, HepG2, HeLa, J774, and WI-38 by MTT assay. The IC₅₀ was found to be higher than 100 µg/mL against these five cell lines after 72 h treatment. Furthermore, OLM was tested in mice for acute and sub-acute oral toxicity according to the guidelines of the Organisation for Economic Co-operation and Development (OECD). OLM was found non-toxic, except for some fibrosis observed in the spleens of treated mice in the sub-acute oral toxicity test. Our results confirm the anti-inflammatory potential of OLM and that it could be tested in in vivo inflammatory models, but its effect on the spleen should be considered before designing the experiments.

Keywords: Flavonoids; GNPS; Inflammation; OECD; OLM, Salvadora oleoides methanolic extract; Oral toxicity.