There has been a growing interest in the beneficial effects of simple phenolic acids and their ability to exhibit various biological activities. The aim of this study was to assess in vitro biological activities of 2-, 3-, and 4-hydroxybenzoate lithium (HBLi) complexes on HT-1080 human fibrosarcoma cells by methods of using a metabolic activity assay, immunochemical and morphological techniques. Results showed that HBLi complexes exert their cytotoxic activities in a concentration- and chemical structure-dependent manner in the following order: 4-HBLi > 3-HBLi > 2-HBLi. Flow cytometry displayed evidence of apoptosis induced by 3-HBLi (21.8%) and 4-HBLi (33.2%). These results were verified by SEM, which revealed the formation of apoptotic bodies. In addition, these 3-HBLi and 4-HBLi caused an increase in HT-1080 cell cycle arrest in G0/G1 phase when compared to the controls (25% and 30.6%, resp.) when cells were treated with 6？mM for 24 hours. Immunochemical studies related to the molecular mechanism of apoptosis indicated that HBLi complexes downregulated the expression of Bcl-2 and upregulated Bax, p53, and caspases-3 in a concentration-dependent manner. HBLi complexes lowered Bcl-2/Bax ratios and induced the expression of p53 and caspase-3. These results suggest that HBLi complexes may exert their apoptotic effects through mitochondrial-mediated, caspase-dependent, apoptotic mechanisms. 1. Introduction Hydroxybenzoic acids (HBA) are a group of molecules that belong to simple phenolic acids, a major class of plant secondary metabolites widely distributed in plants. The parent molecule in this group is benzoic acid, encompassing three substituted hydroxyl groups analogous at ortho-, meta-, and para- positions, or C1, C2, and C3, respectively. These compounds exert various biological activities, including antioxidant, anticancer, and antimicrobial properties [1–3]. In addition, they exhibit different physicochemical properties and interactions with the primary metabolites and display various biological properties [4–6]. HBAs readily interact with metal ions to form hydroxybenzoate (HB) complexes, which are more biologically effective than HBAs or the free metal ions. Zinc salicylate, for example, was found to be more potent than salicylic acid and acetylsalicylic acid in terms of its ability to inhibit cell proliferation and induce apoptosis in cancer cells [7, 8]. Metal complexes, particularly transition element metals, interact with DNA noncovalently producing changes in the structure of the DNA, thereby interfering with replication and transcription
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