Diarrhoea is an alteration in bowel movement characterized by increased water content and frequency of stooling. The aqueous extract of Mangifera indica leaves have been validated for their efficacy as an antidiarrhoeal agent. However, the probable mechanism(s) of action remain unknown. The objectives of the study were to: (i) partition the aqueous extract using solvent-solvent extraction; (ii) determine the antidiarrhoeal activity of the solvent-solvent and column chromatographic fractions in female Wistar rats; (iii) establish the probable mechanism of action of the sub-fractions; (iv) evaluate the toxicity of sub-fraction B (SFB) on selected organs; and (v) identify the probable bioactive constituents of SFB. rnThe aqueous extract of M. indica leaves was successively fractionated into ethyl-acetate fraction (EAF), n-butanol fraction (NBF) and residual fraction (RF). The EAF, NBF and RF were evaporated to dryness at 40â„ƒ. They were reconstituted and administered to screen for antidiarrhoeal activity using castor oil-induced diarrhoea; enteropooling; and gastrointestinal motility models. Each of the sub-fractions of the RF were also screened for their antidiarrhoeal activity and evaluated for probable mechanism(s) of action. The SFB was also evaluated for its effects on the liver, kidney and small intestine. Data were analysed using One-way Analysis of Variance and Tukeyâ€™s Post-hoc test at p < 0.05.rnThe findings revealed that the:rni. RF at 14.09 and 56.35 mg/kg body weight (bw) significantly prolonged the onset time of diarrhoea, decreased the total number of faeces, wet faeces, water content and increased the inhibition of defecation by 94.29 and 96.34% respectively; rnii. mass and volume of intestinal content significantly decreased at 14.09 mg/kg bw of the RF with an inhibition of 45.20% on intraluminal fluid accumulation;rniii. distance travelled by charcoal meal was shortest in diarrhoeal rats treated with 14.09 mg/kg bw of the RF with 39.79% inhibition of peristalsis; rniv. the antidiarrhoeal index (ADI) was highest at 14.09 mg/kg bw of the RF (55.19%). At 56.35 mg/kg bw, the ADI (49.87%) compared favourably with that of the reference drug (48.50%);rnv. highest inhibition of defecation was exhibited by sub-fraction B at 6.47 mg/kg bw while the onset of diarrhoea was most prolonged by sub-fraction A;rnvi. antienteropooling and antimotility activities were highest in the sub-fraction A and B treated rats, respectively;rnvii. activities of Na-K ATPase, alkaline phosphatase and the concentrations of glucose, Na+ and K+ were increased by SFB while the levels of Cl , protein carbonyl and nitrite were decreased; rnviii. levels of acetylcholinesterase, glutathione, superoxide dismutase and intestinal protein were all increased by SFB;rnix. sub-fraction B at 12.94 mg/kg bw exhibited structural and functional toxicity on the liver kidney and small intestine; andrnx. mangiferin and catechin were identified as the probable bioactive constituents of SBF.rnrnThe study concluded that the antidiarrhoeal principles were present in the RF while the established mechanisms included antisecretory, anti-inflammatory, antioxidant and modulation of intestinal permeability. The antidiarrhoeal principle rich-SBF may therefore be explored as a lead for drug discovery in the treatment of diarrhoea.