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Protoplast release in pectinolytic strain mutants of Aspergillus sp. CH-Y-1043 (A13) and Aspergillus flavipes ATCC-16795 (F7) is described. Optimum yield of protoplasts A13 was obtained in a lapse of 1 h when commercially lytic enzymes of Trichoderma harzanium (2 mg ml-1) were added in 0.05 mol l-1 citrate-phosphate buffer pH 5.0 containing 0.7 mol l-1 KCl and 10 mg ml-1 BSA. Best results in F7 were obtained when the protoplasting system of A13 was supplemented with 10 mg ml-1Aureobasidium sp. lytic enzymes. Isolated protoplasts in A13 and F7 were capable of a high regeneration frequency of 87% and 53% when 0.7 mol l-1 KCl and sorbitol were used as osmotic stabilizers. Endo-P, Exo-P and pectin lyase production were not modified during the process of regeneration.