Improving protein stability is an important goal for basic research as well as for clinical and industrial applications but no commonly accepted and widely used strategy for efficient engineering is known. Beside random approaches like error prone PCR or physical techniques to stabilize proteins, e.g. by immobilization, in silico approaches are gaining more attention to apply target-oriented mutagenesis. In this review different algorithms for the prediction of beneficial mutation sites to enhance protein stability are summarized and the advantages and disadvantages of FoldX are highlighted. The question whether the prediction of mutation sites by the algorithm FoldX is more accurate than random based approaches is addressed.

Polyunsaturated fatty acids (PUFAs) of the -3 and -6 class (e.g. -linolenic acid, linoleic acid) are essential for maintaining biofunctions in mammalians like humans. Due to the fact that humans cannot synthesize these essential fatty acids, they must be taken up from different food sources. Classical sources for these fatty acids are porcine liver and fish oil. However, microbial lipids or single cell oils, produced by oleaginous microorganisms such as algae, fungi and bacteria, are a promising source as well. These single cell oils can be used for many valuable chemicals with applications not only for nutrition but also for fuels and are therefore an ideal basis for a bio-based economy. A crucial point for the establishment of microbial lipids utilization is the cost-effective production and purification of fuels or products of higher value. The fermentative production can be realized by submerged (SmF) or solid state fermentation (SSF). The yield and the composition of the obtained microbial lipids depend on the type of fermentation and the particular conditions (e.g. medium, pH-value, temperature, aeration, nitrogen source). From an economical point of view, waste or by-product streams can be used as cheap and renewable carbon and nitrogen sources. In general, downstream processing costs are one of the major obstacles to be solved for full economic efficiency of microbial lipids. For the extraction of lipids from microbial biomass cell disruption is most important, because efficiency of cell disruption directly influences subsequent downstream operations and overall extraction efficiencies. A multitude of cell disruption and lipid extraction methods are available, conventional as well as newly emerging methods, which will be described and discussed in terms of large scale applicability, their potential in a modern biorefinery and their influence on product quality. Furthermore, an overview is given about applications of microbial lipids or derived fatty acids with emphasis on food applications.

Malic acid derived from fossil resources is currently applied in the food and beverage industries with a medium global production capacity. However, in the transition from a fossil‐based to a bio‐based economy, biotechnologically produced l‐malic acid may become an important platform chemical with many new applications, especially in the field of biopolymers. In this review, currently used petrochemical production routes to dl‐malic acid are outlined and insights into possible bio‐based alternatives for microbial l‐malic acid production are provided. Besides ecological reasons, the possibility to produce enantiopure l‐malic acid by microbial fermentation is the biggest advantage over chemical synthesis. State‐of‐the‐art and open challenges concerning production host engineering, substrate choice and downstream processing are addressed. With regard to production hosts, a literature overview is given covering the leading natural production strains of Aspergillus, Ustilago and Aureobasidium, as well as Escherichia coli as the most important engineered recombinant host. The utilization of renewable substrates as an alternative to glucose is emphasized in particular as a key aspect for a competitive bio‐based production. Out of the alternative substrates discussed in this review, the industrial side‐streams crude glycerol and molasses seem to be most promising for large‐scale l‐malic acid production. © 2019 The Authors. Journal of Chemical Technology & Biotechnology published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

The organic acid producer Aspergillus oryzae and Rhizopus delemar are able to convert several alternative carbon sources to malic and fumaric acid. Thus, carbohydrate hydrolysates from lignocellulose separation are likely suitable as substrate for organic acid production with these fungi. Before lignocellulose hydrolysate fractions were tested as substrates, experiments with several mono- and disaccharides, possibly present in pretreated biomass, were conducted for their suitability for malic acid production with A. oryzae. This includes levoglucosan, glucose, galactose, mannose, arabinose, xylose, ribose, and cellobiose as well as cheap and easy available sugars, e.g., fructose and maltose. A. oryzae is able to convert every sugar investigated to malate, albeit with different yields. Based on the promising results from the pure sugar conversion experiments, fractions of the organosolv process from beechwood (Fagus sylvatica) and Miscanthus giganteus were further analyzed as carbon source for cultivation and fermentation with A. oryzae for malic acid and R. delemar for fumaric acid production. The highest malic acid concentration of 37.9 ± 2.6 g/L could be reached using beechwood cellulose fraction as carbon source in bioreactor fermentation with A. oryzae and 16.2 ± 0.2 g/L fumaric acid with R. delemar. We showed in this study that the range of convertible sugars for A. oryzae is even higher than known before. We approved the suitability of fiber/cellulose hydrolysate obtained from the organosolv process as carbon source for A. oryzae in shake flasks as well as in a small-scale bioreactor. The more challenging hemicellulose fraction of F. sylvatica was also positively evaluated for malic acid production with A. oryzae.

2-Phenylethanol (2-PE), an important flavor and fragrance compound with a rose-like smell has been widely used in the cosmetics, perfume, and food industries. Traditional production of 2-PE was mainly through the extraction from plant materials or by chemical synthesis. However, the increasing demand of consumers for natural flavors cannot be met by these methods. Biological production of 2-PE has emerged to be an appealing solution due to an environmental friendly process and the definition of a “natural” product. In this review, we have comprehensively summarized the current status and perspectives for biological 2-PE production in terms of its advantages over classical chemical synthesis and extraction from natural plants. A comprehensive description of 2-PE synthetic pathways and global regulation mechanisms, strategies to increase 2-PE production, and the utilization of agro-industrial wastes as feedstocks has been systematically discussed. Furthermore, the application of in situ product removal techniques have also been highlighted.