Improvement of the Purification Process for Earthworm DNase and Cytotoxicity Analysis.

OBJECTIVES: Deoxyribonuclease (DNase) is widely used in treating obstructed purulent sputum drainage caused by bronchiectasis, lung abscess, cystic fibrosis, and atelectasis and inhibits biofilm formation. METHODS: This study optimized the earthworm DNase extraction process. Tissue homogenates were prepared using mechanical homogenization, followed by isoelectric precipitation and acetone precipitation. Crude earthworm extracts (EWE) were purified by ion exchange chromatography. DNase activity was evaluated by Bradford assay and UV absorption method. RESULTS: Method 1 took 42-45 h, while method 2 took 15-18 h. The specific activity of EWE1 was 677.48 ± 52.84 U/mg. The specific activity of EWE2 was 622.03 ± 49.76 U/mg. Following weak cation exchange chromatography and heparin affinity chromatography, the enzyme activity of the extract was 4443.33 ± 266.59 U/mg. The minimum working concentration was 1.88 µg/mL. CONCLUSION: The improved extraction process markedly reduced the preparation time and enhanced DNase specific activity of earthworm extract.