AIM: To product protoplasts of pathogenic Aspergillus fumigatus,and to observe biological properties of protoplasts from stripped the cell wall to return to full form·METHODS: 1mol/L sorbitol was used as the osmotic stabilizer to product complex enzyme including 1g/dL snailase,1g/dL cellulase and 0.1g/dL lysing enzyme.Complex enzyme was confected with 5×109/L Aspergillus fumigatus conidia liquid by volume ratio of 1∶1 to digest 2 hours in gas bath thermostatic oscillator at 30℃ and 80r/min.The activity of protoplasts was detected by the neutral red solution·RESULTS: The average generation capacity of Aspergillus fumigatus conidia protoplasts was 2.84×109/L.After digestion the activity of protoplasts at 0,6,12,18,24 hours were 86.6%,77.4%,73.9%,70.6%,66.2%.The regeneration phenomenon of protoplasts began at 18 hours after the digestion,and basically completed within 36-48 hours·CONCLUSION: The generation rate of Aspergillus fumigatus protoplasts was 56.8% with complex enzyme.The time window that we can interfere was at least 18 hours.Protoplasts could maintain a high activity in 24 hours.