Minimal toxicity data are available for 1-alkylquinolinium bromide ionic liquids. Here, their toxicity to NIH 3T3 murine fibroblast cells, of relevance to their potential antimicrobial application, is presented. Toxicity data, presented by time-point analysis with a particular focus on the immediate toxicity upon short term cellular exposure, indicate a link between the length of the alkyl chain cationic substituent and resultant biological toxicity. 1-Tetradecylquinolinium bromide was found to exhibit cellular toxicity comparable to benzalkonium chloride over all time points tested. By comparison, 1-octylquinolinium bromide initially exerted significantly lower cytotoxicity at one hour; however, toxicity was found to have a cumulative effect over time-course analysis up to three days. This illustrates that cationic substituent alkyl chain length may govern not only overall toxicity, but also the rate of toxicity. Fluorescence microscopy was utilised to examine destabilisation of the plasma membrane by 1-tetradecylquinolinium bromide and benzalkonium chloride after one hour, with membrane destabilisation not observed for 1-octylquinolinium bromide, or the base constituent quinoline.