Optimization of antifungal production by Bacillus species in the presence of nanoparticle supplementation and preliminary scale-up.

Abstract

The intensive agricultural practices used to meet global crop production demands have resulted in the rigorous use of chemical pesticides. These ultimately compromise crop production as well as the environment. In order to alleviate this, cheaper and environmentally friendly, biocontrol agents have been considered as an alternative to chemical pesticides. Biosurfactants are a promising alternative to chemical pesticides due to their higher biodegradability, lower toxicity, and environmental friendliness. Amongst the many bacterial and fungal biosurfactant producers, biosurfactants from Bacillus species show promise as biocontrol agents. These biosurfactants are known for their wide biotechnological use in agricultural, industrial, and medicinal fields. However, large scale production is still faced with challenges such as low yields and high production cost thus raising the need for modelling, optimization, catalytic and scale up investigations. Hence, a study was undertaken with the aim of enhancing biosurfactant production through process modelling and optimization with subsequent assessment of the scale up potential of the optimized process. A Response Surface Methodology (RSM) using box Behnken design was used to investigate the optimal process conditions for improved biosurfactant production from B. subtilis BS20. The investigated process parameters included glucose concentration (10 – 30 g/L), incubation temperature (25 – 45℃) and incubation time (24 – 96 h). The developed model gave a high coefficient of determination (R2 ) = 0.86, p-value of 0.0279 and F-value of 4.62 for the modelled biosurfactant production. Optimized process conditions of 11.5 g/L glucose concentration, 24 h incubation time and 41o C for incubation temperature were obtained and produced a maximal antifungal activity of 68 mm. Moreover, supplementary inclusion of seven (7) different nanoparticles as a biocatalyst in the cultivation of B. subtilis BS20 was carried out using the optimal process condition to further improve antifungal (biosurfactant) production. The inclusion of nanoparticles favored increased biomass yield, but biosurfactant with high antifungal activity was not obtained. Moreover, when it comes to commercializing new bioprocess and bioproduct developments, bio – process scale-up in the biotechnology industry is an essential stage. This study therefore evaluated the scale up of biosurfactant production based on constant power consumption, Reynold number and impeller tip speed. The stirrer speed (n), impeller diameter (di), number of impellers (N), power number (Np), broth density (⍴), working volume and geometric factor (fc) were correlated with impeller tip speed (Vtip), Reynolds number (𝑅𝑅𝑅𝑅) and power consumption rate (P/V) to obtain the most suitable criterion for biosurfactant production in a 10 L bioreactor. Implementing constant Vtip value from the 1 L scale: 93 rpm, Reynold number (Re) 5.9E - 04, Power (P) 0.32 W, Power to Volume ratio (P/VL) 160 W/m3 , circulation time (tc) 5.2 s and shear stress (γ) 15.5 S-1, at 41 °C, gave the highest antifungal activity of 65 mm zone of inhibition in the 10 L scale bioreactor. The antifungal activity obtained for constant Vtip were comparable to those obtained at 1L bioreactors (57 mm), this showed that the bioprocess dynamics for achieving high antifungal activity are available, further paving the way for feasible commercialization strategies. This study has elucidated the optimum process conditions for B. subtilis BS20 metabolism for improved biosurfactant production resulting in significant antifungal activity. Furthermore, findings showed that the inclusion of nanoparticles biocatalyst to the process enhanced biomass yields. Process scale up provided preliminary data for large scale production of biosurfactant production from B. subtilis BS20.