Planarian toxicity fluorescent assay: A rapid and cheap pre-screening tool for potential skin irritants

Abstract

Here we report a new planarian (Dugesia lugubris) fluorescent assay as a rapid and cheap pre-screening tool to predict strong skin irritants. Our aim was to provide a simple and cost-effective in vivo method that avoided use of higher vertebrates. Adapting previously reported methods for planaria mobility alongside an acute toxicity assay, different irritants at five concentrations (0.1%, 0.05%, 0.025%, 0.01% and 0.005% w/v) were tested but both methods failed to discriminate the irritation potential of the test compounds. Therefore, a new alternative fluorescence assay was developed, hypothesising that increasing damage from the irritant to the planarian outer protective membrane will increase accumulation of sodium fluorescein in the flatworm. Fourteen test chemicals were selected representing strong, moderate, mild and non-irritants. In general, results showed increasing sodium fluorescein accumulation within planaria following acute exposure to increasingly strong skin irritants; on exposure to the strong irritants, benzalkonium chloride, citronellal, methyl palmitate, 1-bromohexane and carvacrol, fluorescence within the planaria was significantly greater (P < 0.05) than the negative controls and the common non-irritants PEG-400, dipropylene glycol and isopropyl alcohol; fluorescence values of planaria tested with negative controls and non-irritants were not significantly different. For all test compounds, Fluorescence Intensity of the planaria was compared with literature Primary Irritation Index data and generated a statistically significant (P < 0.005) Pearson correlation (r) of 0.87. Thus, the planarian fluorescent assay is a promising tool for rapid early testing of potential strong skin irritants, and non-irritants, and avoids use of higher vertebrate models.