Baits were used to investigate the toxicity of methanol extracts from the fern Pronephrium megacuspe (Thelypteridaceae) on workers of the red imported fire ant, Solenopsis invicta Buren (Hymenoptera: Formicidae). The methanol extract of P. megacuspe had toxic effects on workers. With macrergates, the LC50 values of the methanol extract and ethyl acetate fraction were 524.0 and 145.9 μg/g, respectively, after 48 h of treatment and 362.5 and 99.0 μg/g, respectively, after 72 h of treatment. With micrergates, the LC50 values of the methanol extract and ethyl acetate fraction were 321.9 and 90.0 μg/g, respectively, after 48 h of treatment and 235.4 and 79.1 μg/g, respectively, after 72 h of treatment. NMR spectral data revealed that the isolated compound from the ethyl acetate soluble fraction of methanol extract was phenol-3-O-beta-D-glucoside. The LC50 values of this compound with macrergates after 48 and 72 h were 21.6 and 9.7 μg/g, respectively. Similarly, the LC50 values of this compound with micrergates after 48 and 72 h were 3.4 and 2.4 μg/g, respectively. A low dose of methanol extract and ethyl acetate fractions of P. megacuspe, and of phenol-3-Obeta-D-glucoside diminished the walking and grasping abilities of S. invicta micrergates and macrergates.