Nicholas V. Callow

Cellulase production by continuous culture of Trichoderma reesei Rut C30 using acid hydrolysate prepared to retain more oligosaccharides for induction

An acid hydrolysate was prepared by a procedure chosen for retaining more oligosaccharides to improve the cellulase-inducing capability when used as substrate in the fungal fermentation for cellulase production. The effect was evaluated with continuous culture of Trichoderma reesei Rut C30 at the dilution rates of 0.03-0.08 h(-1). The specific cellulase production rates were found to be relatively constant at 8.9+/-0.3 (FPU/g dry cells-h), except for the lower rate, i.e., 7.2 (FPU/g-h), at the lowest dilution rate investigated (0.03 h(-1)). The former value was slightly higher than the rate obtained with a lactose-based medium, i.e., 8.2 (FPU/g-h). The maximum specific cell growth rate supported by the hydrolysate-based medium was 0.096 (h(-1)) and the apparent cell yield increased from 0.44 to 0.57 (g dry cells)/(g consumed reducing sugars) with increasing dilution rates. The best-fit maximum/ideal cell yield (without endogenous metabolism) was 0.68 (g/g), the endogenous substrate consumption rate was 0.023 (g reducing sugars)/(g dry cells-h), and the specific cell death rate was 0.016 h(-1).

Promoting pellet growth of Trichoderma reesei Rut C30 by surfactants for easy separation and enhanced cellulase production

It is desirable to modify the normally filamentous Trichoderma reesei Rut C-30 to a pellet form, for easy biomass separation from the fermentation medium containing soluble products (e.g., cellulase). It was found in this study that this morphological modification could be successfully achieved by addition of the biosurfactant rhamnolipid (at ≥ 0.3g/L) and the synthetic Triton X-100 (at ≥ 0.1g/L) to the fermentation broth before the cells started to grow actively. Thirteen other surfactants tested were not as effective. Furthermore, the added rhamnolipid and Triton X-100 increased the maximum cellulase activity (Filter Paper Units) produced in the fungal fermentation; the increase was 68 ± 7.8% for rhamnolipid and 73 ± 12% for Triton X-100. At the concentrations required for pellet formation, rhamnolipid had negative effect on the cell growth: with increasing rhamnolipid concentrations, the growth rate decreased and the lag-phase duration increased linearly. Triton X-100 caused no significant differences in growth rate or lag phase.

Rhamnolipids as platform molecules for production of potential anti-zoospore agrochemicals.

Rhamnolipid biosurfactants have potential applications in the control of zoosporic plant pathogens. However, rhamnolipids have not been closely investigated for the anti-zoospore mechanism or for developing new anti-zoospore chemicals. In this study, RhL-1 and RhL-3 groups of rhamnolipids were used to generate the corresponding RhL-2 and RhL-4 groups and the free diacids. Conversion of RhL-3 to RhL-1 was also accomplished in vitro with cellobiase as the catalyst. The anti-zoospore effects of RhL-1-RhL-4 and the diacids were investigated with zoospores of Phytophthora sojae. For RhL-1-RhL-4, approximately 20, 30, 40, and 40 mg/L, respectively, were found to be the lowest concentrations required to stop movement of all zoospores, which indicates that the anti-zoospore effect remains strong even after RhL-1 and RhL-3 are hydrolyzed into RhL-2 and RhL-4. The free diacids required a significantly higher critical concentration of about 125 mg/L. Rhamnose can be obtained as a co-product.

Nutrient control for stationary phase cellulase production in Trichoderma reesei Rut C-30.

This work describes the use of nutrient limitations with Trichoderma reesei Rut C-30 to obtain a prolonged stationary phase cellulase production. This period of non-growth may allow for dependable cellulase production, extended fermentation periods, and the possibility to use pellet morphology for easy product separation. Phosphorus limitation was successful in halting growth and had a corresponding specific cellulase production of 5±2 FPU/g-h. Combined with the addition of Triton X-100 for fungal pellet formation and low shear conditions, a stationary phase cellulase production period in excess of 300 h was achieved, with a constant enzyme production rate of 7±1 FPU/g-h. While nitrogen limitation was also effective as a growth limiter, it, however, also prevented cellulase production.