Comparative Assessment of Physicochemical and Cytotoxic Characterisation, in Khaini and Filtered Khaini Smokeless Tobacco Products

Abstract Objectives Smokeless tobacco (SLT) products such as Cool Lip, marketed in India in filter-pouch form and resembling Swedish snus, are widely promoted as reduced-risk alternatives to traditional SLT such as Khaini(HANS). This study aims to systematically characterize the physicochemical, chemical, and biological toxicity profile of Cool Lip in comparison to a conventional SLT brand, Hans, to provide evidence on their health risks. Study Design Comprehensive analyses were performed to assess physical properties (pH, moisture, particle size using SEM imaging), chemical constituents (nicotine, TSNA, heavy metals, PAHs, alkalis, pesticides, and microplastic contamination), and microbiological load. Cytotoxicity was evaluated via MTT assay and apoptosis by AO staining in primary human gingival fibroblasts exposed to product extracts. Results Cool Lip exhibited a finer, more porous, and fragmented microstructure than Hans, which can accelerate chemical dissolution and absorption. Both products contained high levels of carcinogenic TSNAs and PAHs, and demonstrated higher concentrations of NNN, NNK, and nicotine, while Cool Lip had a substantially greater benzo[a]pyrene burden. Cytotoxicity and apoptosis were dose-dependent for both, with Hans consistently causing greater cellular toxicity. Conclusions Despite being marketed as a harm-reduced alternative, Cool Lip exhibits comparable or even greater toxicological risks relative to established Indian SLT products. The findings emphasize the urgent need for regulatory scrutiny and consumer education to counter misleading safety perceptions.