Production Of Cutinase By Thermobifida Fusca WSH 03-11 And Its Application In The Textile Industry

Cutinase is an exteacellular enzyme which catalyses the hydrosis of cutin and belongs to aclass of serine esterases. Cutinase is specific toward the cutin polymer in the scouring andimprove the wettability of raw cotton and in coreaction with pectin lyase which facilites uniformdyeing and finishing. Bioscouring with other enzymes brings environmentally friendly effect.Cutinase from microorganisms was not reported in domestic research. In this study, thenutritional and environmental conditions for Thermobifida fusca WSH 03-11and cutinaseexcretion were were studied. The characteristics of cutinase from T. fusca WSH 03-11 were alsoinvestigated. The enzymatic scouring of cutinase and coreation with pectin lyase was evaluated.The main results were given as follows:1. Nutritional and environmental conditions for the growth of T. fuscaWSH03-11 wereinvestigated. The optimum seed culture medium contained (per liter): 90 g soluble starch, 5 gbeef extract, 5 g yeast extract, 5 g NaCl, 2 g K2HPO4 and 1% trace-elements solution at pH8.0. Temperature was 50℃. The suitable volume of medium was 100 mL in 500 mL flask andthe age of seed was 35～40 h.2. The optimal conditions for cutinase production in flask were as follows. The fermentationmedium contained (per liter): 10 mL (v/v) ethanol, 5 g peptone, 5 g yeast extract, 5 g NaCl, 2g K2HPO4 ,1 g cutin and 1% trace elements solution. The volume of medium was 100 mL in500 mL shake flasks, and initial pH 8.0.3. Glucose strongly inhibited cutinase production over the 5 d incubation period. In order toalleviate the effect of residual sugar from seed medium on cutinase synthesis, inoculationway with collected biomass was adopted. Cutinase activity increased by 2-fold when glucosewas removed from the inoculum. The pH of the culture broth also increased from 5.8 to 8.4,which would enhance pH stability of cutinase during bioscouring.4. Various cutins from natural raw materials were tested for their ability to induce cutinaseproduction by T. fusca WSH 03-11. Tomato peel induced the highest levels of cutinase andapple peel also was a good inducer. With 5~6 g/L tomato peel added at 0 h, cutinase activityreached 24U/mL. 16-hydroxypalmitic acid obviously inhibited the growth and cutinaseproduction of T. fusca WSH 03-11.5. Addition of CaCO3 influenced both cell growth and cutinase activity. With 2 g/L CaCO3,cutinase activity was increased by 2.5 U/mL than the control. The effects of amino acids anddetergent on cell growth and cutinase excretion were investigated. Cutinase activity could beimproved by adding 1 g/L Arg and 0.1g/L Tween-40, a non-ionic detergent. Cutinase activityincreased by 16% and 10% than the control, respectively.6. Cutinase showed good heat stability between 30℃ and 60 ℃. Its activity was notsignificantly reduced by heating at 50℃ and 60℃ for up to 1 h. The stability of pH rangedfrom 7.0 to 11.0, and cutinase exhibited optimal activity at pH 8.5～9.5. Among the metalions, Co2+ and Hg2+ inhibited the enzyme activity strongly and Mg2+ activated the cutinase tosome extent.7. The scouring efficiency of cutinase from T. fusca WSH 03-11 was studied. Cutinaseimproved the wettability and dying effect of cotton fabrics. The combination application ofcutinase and pectin lyase resulted in a synergistic effect. Compared with chemical scouringprocess, enzymatic scouring process showed the same scouring effect and, more important,had environmentally friendly benefits.