Toxin produced by Colletotrichum falcatum Went, the incitant of red rot of sugarcane was isolated, purified and assayed to determine host specificity and identify its chemical nature. The toxin was found to be not host specific as it inhibited germination of various seeds(gram, greengram, blackgram, pea, cowpea, rice and sugarcane) as well as different seedlings viz. tomato, coriander, pea and rice. The toxin consists of two distinct fraction-one fraction having $R_f$, value at 0.36 producing identical red rot lesion when inoculated at leaf midrib of sugarcane, and the other having $R_f$, value at 0.72 not showing any red rot lesion. Chromatogram of high performance liquid chromatography(HPLC) of the red rot lesion causing fraction showed a sharp peak at 1.62 min of retention time(RT), and spectral analysis indicated the presence of following chemical $CH_3$ - groups-C-H, C=O, C-N, $-CH_3,;-CH_2$ -CH and molecular mass of the compound was 203. - ($M^+,;C_{11}H_{11}N_2O_2$).