Short-chain fatty acids (SCFAs) are important compounds produced during gut microbiota fermentation of indigestible carbohydrates. SCFA are recognized for anti-inflammatory action, cancer prevention, and cholesterol pathways inhibition. Acetic, propionic and n-butyric acid are the most important SCFA produced from carbohydrates. This work aimed to develop and validate a rapid and low-cost capillary electrophoretic (CE) method for determination of acetic, propionic and n-butyric acids in rats and mice faeces. Electrophoretic separation was performed using a MDQ Beckman Coulter CE system, fused silica capillary, 60 cm × 75 µm i.d., applied voltage 25 kV, hydrodynamic injection for 3 s, electrolyte system 160 mM tris(hydroxymethyl)aminomethane 10 mM benzoic acid at pH 8.5, indirect UV detection at 228 nm. Samples were prepared by extraction of SCFA from faeces in water and direct injection in CE. An internal standard (2-ethylbutyric acid) was added to all samples and, prior to analysis, they were diluted 1:10 in separation buffer. An adequate separation of the SCFA peaks was achieved in 10 min. The method showed good linearity with range of 5.00 to 30.0 mM (r² > 0.98) for acetic acid, 1.00 to 10.0 mM (r² > 0.98) for propionic acid, and 1.00 to 6.00 mM (r² > 0.99) for n-butyric acid. Average recovery varied from 74.1 to 109.8% for mice and 81.3 to 120.5% for rats and intraday and interday precision essays presented RSD lower than 10%. The method provides LODs and LOQs of 0.13 – 0.43 mM for acetic acid, 0.09 – 0.29 mM for propionic acid and 0.03 – 0.09 mM for n-butyric acid. After validation, this methodology was successfully applied to perform SCFA analysis in samples of rats and mice fed with different diets composition.