Modeling conversion and transport phenomena in solid-state fermentation: a review and perspectives.

Solid-state fermentation (SSF) is accompanied inevitably by development of concentration and temperature gradients within the substrate particles and microbial biofilms. These gradients are needed for driving the transport of substrates and products. In addition, concentration gradients have been suggested to be crucial for obtaining the characteristics that define the products of SSF; nevertheless, gradients are also known to result in reduced productivity and unwanted side reactions. Solid-state fermentations are generally batch processes and this further complicates their understanding as conditions change with time. Mathematical models are therefore needed for improving the understanding of SSF processes and allowing their manipulation to achieve the desired outcomes. Existing models of SSF processes describe coupled substrate conversion and diffusion and the consequent microbial growth. Existing models disregard many of the significant phenomena that are known to influence SSF. As a result, available models cannot explain the generation of the numerous products that form during any SSF process and the outcome of the process in terms of the characteristics of the final product. This review critically evaluates the proposed models and their experimental validation. In addition, important issues that need to be resolved for improved modeling of SSF are discussed.

Significance of bed porosity, bran and specific surface area in solid-state cultivation of Aspergillus oryzae.

In this paper, the effects of bed porosity, bran and specific surface area on the oxygen uptake rate and alpha-amylase production during growth of Aspergillus oryzae on wheat grain and wheat-flour substrate are reported. The high oxygen uptake rate found during cultivation of A. oryzae on wheat-flour substrate was not reached on wheat grain. This is mainly due to the bran of the wheat grain. Using wheat-flour substrates, it was shown that extra bed porosity increased the alpha-amylase production and oxygen uptake rates. Furthermore, the peak oxygen uptake rate decreased with increasing surface area-volume ratio of the substrate particles, while the alpha-amylase production and the cumulative oxygen uptake per gram of initial substrate dry matter increased. The present work does not support a direct correlation between aerial mycelia and enzyme production. There is, however, a correlation between the alpha-amylase yield and the cumulative oxygen uptake (not the uptake rate). This implies that aerial mycelia could accelerate alpha-amylase production even if they do not increase the yield.

Effect of low oxygen concentrations on growth and alpha-amylase production of Aspergillus oryzae in model solid-state fermentation systems.

Oxygen transfer in the fungal mat is a major concern in solid-state fermentation (SSF). Oxygen supply into the mycelial layers is hampered by diffusion limitation. For aerobic fungi, like Aspergillus oryzae, this oxygen depletion can be a severely limiting factor for growth and metabolite production. This paper describes the effects of a low oxygen concentration on growth at the levels of individual hyphae, colonies and overcultures, and on alpha-amylase production in overcultures. PDA medium was used to study the effect of a low oxygen concentration on hyphal elongation rate and branching frequency of hyphae, and radial extension rate of colonies of A. oryzae. We found similar saturation constants (K(O2)) of 0.1% (v/v in the gas phase) for oxygen concentration described with Monod kinetics, for branching frequency of hyphae and colony extension rate. When A. oryzae was grown as an over-culture on wheat-flour model substrate at 0.25% (v/v) oxygen concentration, the reduction in growth was more pronounced than as individual hyphae and a colony on PDA medium. Experimental results also showed that the specific alpha-amylase production rate under the condition of 0.25% (v/v) oxygen was reduced. Because the value of K(O2) is relatively low, it is reasonable to simplify the kinetics of growth of A. oryzae to zero-order kinetics in coupled diffusion/reaction models.

Contribution of aerial hyphae of Aspergillus oryzae to respiration in a model solid-state fermentation system.

Oxygen transfer is for two reasons a major concern in scale-up and process control in industrial application of aerobic fungal solid-state fermentation (SSF): 1) heat production is proportional to oxygen uptake and it is well known that heat removal is one of the main problems in scaled-up fermenters, and 2) oxygen supply to the mycelium on the surface of or inside the substrate particles may be hampered by diffusion limitation. This article gives the first experimental evidence that aerial hyphae are important for fungal respiration in SSF. In cultures of A. oryzae on a wheat-flour model substrate, aerial hyphae contributed up to 75% of the oxygen uptake rate by the fungus. This is due to the fact that A. oryzae forms very abundant aerial mycelium and diffusion of oxygen in the gas-filled pores of the aerial hyphae layer is rapid. It means that diffusion limitation in the densely packed mycelium layer that is formed closer to the substrate surface and that has liquid-filled pores is much less important for A. oryzae than was previously reported for R. oligosporus and C. minitans. It also means that the overall oxygen uptake rate for A. oryzae is much higher than the oxygen uptake rate that can be predicted in the densely packed mycelium layer for R. oligosporus and C. minitans. This would imply that cooling problems become more pronounced. Therefore, it is very important to clarify the physiological role of aerial hyphae in SSF.

Concomitant extracellular accumulation of alpha-keto acids and higher alcohols by Zygosaccharomyces rouxii.

Alpha-keto acids are key intermediates in the formation of higher alcohols, important flavor components in soy sauce, and produced by the salt-tolerant yeast Zygosaccharomyces rouxii. Unlike most of the higher alcohols, the alpha-keto acids are usually not extracellularly accumulated by Z. rouxii when it is cultivated with ammonium as the sole nitrogen source. To facilitate extracellular accumulation of the alpha-keto acids from aspartate-derived amino acid metabolism, the amino acids valine, leucine, threonine and methionine were exogenously supplied during batch and A-star cultivations of (routants of) Z. rouxii. It was shown that all alpha-keto acids from the aspartate-derived amino acid metabolism, except alpha-ketobutyrate, could be extracellularly accumulated. In addition, it appeared from the concomitant extracellular accumulation of alpha-keto acids and higher alcohols that in Z. rouxii, valine, leucine and methionine were converted via Ehrlich pathways similar to those in Saccharomyces cerevisiae. Unlike these amino acids, threonine was converted via both the Ehrlich and amino acid biosynthetic pathways in Z. rouxii.

Aspergillus oryzae in solid-state and submerged fermentations. Progress report on a multi-disciplinary project.

We report the progress of a multi-disciplinary research project on solid-state fermentation (SSF) of the filamentous fungus Aspergillus oryzae. The molecular and physiological aspects of the fungus in submerged fermentation (SmF) and SSF are compared and we observe a number of differences correlated with the different growth conditions. First, the aerial hyphae which occur only in SSFs are mainly responsible for oxygen uptake. Second, SSF is characterised by gradients in temperature, water activity and nutrient concentration, and inside the hyphae different polyols are accumulating. Third, pelleted growth in SmF and mycelial growth in SSF show different gene expression and protein secretion patterns. With this approach we aim to expand our knowledge of mechanisms of fungal growth on solid substrates and to exploit the biotechnological applications.