Purification of cyclosporin A (CyA), which is a specific and potent immunosuppressive produced by some fungi as a secondary metabolite, has been accomplished by chromatography techniques in this research. After submerged incubation of the mould Tolypocladium inflatum in the production medium, the fermentation mixture was extracted by ethyl acetate and underwent column liquid chromatography to purify CyA. In the first step, silica gel-40 and ethylacetate/isopropanol (95:5, v/v) were applied as stationary and mobile phases, respectively. The obtained fractions were assayed for the existence of lipids by the phosphovanilin method. The bioassay method measured the amount of cyclosporins in the fractions. For further purification, the fractions verified to have relatively pure CyA were pooled and submitted to the second step of the column liquid chromatography on Sephadex LH-20 resin. All the fractions obtained were tested by CyA bioassay and thin layer chromatography. HPLC and IR (infrared) spectrometry-confirmed the purity and identity of the product. The amount of total cyclosporin produced was 73 mg/L, including 71% cyclosporin A, which reached 98% after purification steps.