以紫外線為誘變劑，對於Trichoderma koningi iW-10菌梂進行變異試驗。由分離菌株中選出二株變異株， A- 15及A-16，其產生之纖維素分解酵素活性均較其母株為高。使用麩皮固體培養基於30 ℃下培養4天，可分別獲得纖維素分解酵素活性，A- 15變異株為每克17.5Fpu及32 Sau ; A- 16變異株為每克19 Fpu及35Sau。本試驗中分別測定FPA及β-Glucosidase之最適反應溫度及pH，結果顯示：W- 10菌株為55℃及4.8和4.0 ; A-15變異株為60℃及4.8和4.0 ; A-16變異株為55 ℃及4.8。母株及二變異株之較佳溫度安定性範圍皆低於50℃，較佳PH安定性範圍則為4.0至5.4。
糖化試驗中，使用0.5M,PH4.0檸檬酸鹽緩衝液可降低酵素之鈍化，使用18%，經4 %NaOH，室溫下前處理24小時之稻殼，於50℃及pH 4.0之下進行酵素糖化，可於48小時得到29. 3mg /ml還原糖之水解液。 The strain of Trichoderma koningii W-10 was treated with ultraviolet irradiation for the purpose of production of higher cellulase activity. Two mutants, A-15 and A-16 were selected for further studies. After cultivation of mutants A-15 and A-16 on wheat-bran solid media at 30℃ for 4 days, the crude extracts containing 17.5 FPu (32 Sau) and19 FPu (35 Sau) of cellulase activity per gram of solid wheat bran were obtained respectively.
The kinetic parameters of cellulase for strains of W-10, A-15 and A-16 were as followed (in order):
(1) Optimal reaction temperature-55℃, 60℃ and 55℃ for EPA and β-glucosidase activity.
(2) Optimal reaction pH-4.8, 4.8 and 4.8 for FPA; 4.0, 4.0 and 4.8 for β-glucosidase activity.
(3) Thermostability-under 50℃ for FPA and β-glucosidase.
(4) pH stability-from 4.0 to 5.4 for FPA and β-glucosidase.
Use of citrate buffer (0.5 M, pH 4.0) could attenuate the enzyme inactivation during long-time saccharification of cellulosic materials. As high as 29.3 mg/ml reducing sugar could be obtained from a reaction mixture which containing 18% pretreated rice hull with 1.4 FPu (3.0 Sau) per ml of aude enzyme extract of A-15 mutant when incubated at 50℃ for 48 hrs.
