Bioproduction of 2-phenylethanol and 2-phenethyl acetate by Kluyveromyces marxianus through the solid-state fermentation of sugarcane bagasse

Applied Microbiology and Biotechnology - 2-Phenylethanol (2-PE) and 2-phenethyl acetate (2-PEA) are important aroma compounds widely used in food and cosmetic industries due to their rose-like...     

Recent advances in biotechnological production of 2-phenylethanol

2-Phenylethanol (2-PE) is an important aromatic alcohol with a rose-like fragrance. It has been widely applied in the cosmetic, perfume, and food industries and is mainly produced by chemical synthesis. An alternative method for the production of natural flavors and fragrances is the microbial transformation process, which is attracting increasing attention because it is an environmentally friendly process and the products are considered “natural”. The production of 2-PE from L-phenylalanine by biotransformation is possible through the Ehrlich pathway and considerable progress has been made in the development of this process. The present report reviews recent advances in biotechnological production of 2-PE, with emphasis on the strategies used to increase production and the applications of in situ product removal techniques. Future research should focus on product scale-up and product recovery processes for the industrialization of microbial processes.     

Synergistic effect enhances 2-phenylethyl acetate production in the mixed fermentation of Hanseniaspora vineae and Saccharomyces cerevisiae

2-Phenylethyl acetate (2-PEA) is a desired aroma compound in wine due to its honey- and flowery-like characteristics. The effects of adding l-phenylalanine (Phe) during 2-PEA production were investigated in the co-fermentation of Hanseniaspora vineae (HV6) and Saccharomyces cerevisiae BDX. BDX and HV6 strains overproduced 2-phenylethyl alcohol (2-PE) and 2-PEA, respectively. The co-fermentation of BDX and HV6 achieved a 14.9 fold increase in 2-PEA odour activity value (OAV) but a 42.0 % reduction of 2-PE OAV compared to BDX fermentation; the 2-PEA concentration was significantly higher than the sum of BDX and HV6 pure fermentations. This suggests that BDX and HV6 have synergistic effects on 2-PEA formation in mixed culture. Adding 151.6 mg/L Phe enhanced the OAV of 2-PEA by 52.8 % compared to the control. The combination of Phe addition with the co-fermentation of S. cerevisiae and H. vineae is a potential way to increase 2-PEA production and improve wine aromatic quality.     

酵母2-苯乙醇耐受性的研究进展

2-苯乙醇(2-phenylethanol, 2-PE)是一种可食用且有玫瑰香味的高级芳香醇,常用于食品、化妆品和药品行业。由于物理和化学法制备2-PE得率低,不适用于工业生产。而作为单细胞真核微生物的酵母具有高效合成“天然” 2-PE的潜力,因此酵母作为底盘微生物合成2-PE的策略深受研究者青睐。然而,在酵母进行2-PE发酵过程中不免会受到2-PE毒害作用影响。因此,亟须研究酵母耐受2-PE的机制为生产实际提供理论基础,这也有助于选育具有较高2-PE耐受性的酵母菌株。本文综述了酵母2-PE耐受性的研究进展,从酵母2-PE合成途径、2-PE耐受性机理等方面进行阐述,主要说明提升酵母2-PE耐受性的方法。掌握酵母2-PE耐受机制,最终提升酵母2-PE产量及转化效率是今后研究的重中之重。     

Exploring the bioprospecting and biotechnological potential of white-rot and anaerobic Neocallimastigomycota fungi: peptidases,esterases, and lignocellulolytic enzymes

Fungi constitute an invaluable natural resource for scientific research, owing to their diversity; they offer a promising alternative for bioprospecting, thus contributing to biotechnological advances. For a long time, extensive information has been exploited and fungal products have been tested as a source of natural compounds. In this context, enzyme production remains a field of interest, since it offers an efficient alternative to the hazardous processes of chemical transformations. Owing to their vast biodiversity and peculiar biochemical characteristics, two fungal categories, white-rot and anaerobic Neocallimastigomycota, have gathered considerable attention for biotechnological applications. These fungi are known for their ability to depolymerize complex molecular structures and are used in degradation of lignocellulosic biomass, improvement of animal feed digestibility, biogas and bioethanol production, and various other applications. However, there are only limited reports that describe proteolytic enzymes and esterases in these fungi and their synergistic action with lignocellulolytic enzymes on degradation of complex polymers. Thus, in this minireview, we focus on the importance of these organisms in enzyme technology, their bioprospecting, possibility of integration of their enzyme repertoire, and their prospects for future biotechnological innovation.

     

Genetic and metabolic engineering of microorganisms for the development of new flavor compounds from terpenic substrates

Throughout human history, natural products have been the basis for the discovery and development of therapeutics, cosmetic and food compounds used in industry. Many compounds found in natural organisms are rather difficult to chemically synthesize and to extract in large amounts, and in this respect, genetic and metabolic engineering are playing an increasingly important role in the production of these compounds, such as new terpenes and terpenoids, which may potentially be used to create aromas in industry. Terpenes belong to the largest class of natural compounds, are produced by all living organisms and play a fundamental role in human nutrition, cosmetics and medicine. Recent advances in systems biology and synthetic biology are allowing us to perform metabolic engineering at the whole-cell level, thus enabling the optimal design of microorganisms for the efficient production of drugs, cosmetic and food additives. This review describes the recent advances made in the genetic and metabolic engineering of the terpenes pathway with a particular focus on systems biotechnology.     

Biosynthesis of 2-phenylethanol using tobacco waste as feedstock

Abstract

2-Phenylethanol (2-PE) is an aromatic alcohol with a rose-like odour, which is widely used in the food, drink and cosmetic industry. It also present in cigarette aromas. As a unique renewable biomass, tobacco contains abundant aromatic compounds, but is rarely used as a feedstock for synthesizing bio-based products. In this study, it was found that 2-PE can be produced from tobacco waste by Saccharomyces cerevisiae. Plackett–Burman design, steepest ascent design and Box–Behnken designs were applied to optimize the fermentation bioprocess, and the maximum titre of 2-PE reached 1.55 g/l. The study explored a new method of 2-PE production and also provided a valuable way to utilize tobacco waste.     

Biotechnological applications of penicillin acylases: state-of-the-art

This review describes the most recent developments in the biotechnological applications of penicillin acylases. This group of enzymes is involved mainly in the industrial production of 6-aminopenicillanic acid and the synthesis of semisynthetic beta-lactam antibiotics. In addition, penicillin acylases can also be employed in other useful biotransformations, such as peptide synthesis and the resolution of racemic mixtures of chiral compounds. Particular emphasis is placed on advances in detection of new enzyme specificities towards other natural penicillins, enzyme immobilization, and optimization of enzyme-catalyzed hydrolysis and synthesis in the presence of organic solvents.     

酵母菌合成2-苯乙醇的研究进展

2-苯乙醇是一种具有令人愉悦的玫瑰风味的芳香醇,在食品、化妆品和药品等领域具有广泛的应用。本文对酵母菌合成2-苯乙醇的代谢途径及其调控过程、以及提高2-苯乙醇产量的国内外研究进展进行了综述,并对通过微生物转化法合成2-苯乙醇目前存在的不足及进一步研究方向进行了讨论。     

Outdoor cultivation of microalgae for carotenoid production: current state and perspectives

Microalgae are a major natural source for a vast array of valuable compounds, including a diversity of pigments, for which these photosynthetic microorganisms represent an almost exclusive biological resource. Yellow, orange, and red carotenoids have an industrial use in food products and cosmetics as vitamin supplements and health food products and as feed additives for poultry, livestock, fish, and crustaceans. The growing worldwide market value of carotenoids is projected to reach over US$1,000 million by the end of the decade. The nutraceutical boom has also integrated carotenoids mainly on the claim of their proven antioxidant properties. Recently established benefits in human health open new uses for some carotenoids, especially lutein, an effective agent for the prevention and treatment of a variety of degenerative diseases. Consumers’ demand for natural products favors development of pigments from biological sources, thus increasing opportunities for microalgae. The biotechnology of microalgae has gained considerable progress and relevance in recent decades, with carotenoid production representing one of its most successful domains. In this paper, we review the most relevant features of microalgal biotechnology related to the production of different carotenoids outdoors, with a main focus on β-carotene from Dunaliella, astaxanthin from Haematococcus, and lutein from chlorophycean strains. We compare the current state of the corresponding production technologies, based on either open-pond systems or closed photobioreactors. The potential of scientific and technological advances for improvements in yield and reduction in production costs for carotenoids from microalgae is also discussed.     

Biotechnological production of 2-phenylethanol

2-Phenylethanol (2-PE) is an important flavour and fragrance compound with a rose-like odour. Most of the world's annual production of several thousand tons is synthesised by chemical means but, due to increasing demand for natural flavours, alternative production methods are being sought. Harnessing the Ehrlich pathway of yeasts by bioconversion of L-phenylalanine to 2-PE could be an option, but in situ product removal is necessary due to product inhibition. This review describes the microbial production of 2-PE, and also summarizes the chemical syntheses and the market situation.     

Modeling of an integrated fermentation/membrane extraction process for the production of 2-phenylethanol and 2-phenylethylacetate

An unstructured model for an integrated fermentation/membrane extraction process for the production of the aroma compounds 2-phenylethanol and 2-phenylethylacetate by Kluyveromyces marxianus CBS 600 was developed. The extent to which this model, based only on data from the conventional fermentation and separation processes, provided an estimation of the integrated process was evaluated. The effect of product inhibition on specific growth rate and on biomass yield by both aroma compounds was approximated by multivariate regression. Simulations of the respective submodels for fermentation and the separation process matched well with experimental results. With respect to the in situ product removal (ISPR) process, the effect of reduced product inhibition due to product removal on specific growth rate and biomass yield was predicted adequately by the model simulations. Overall product yields were increased considerably in this process (4.0 g/L 2-PE+2-PEA vs. 1.4 g/L in conventional fermentation) and were even higher than predicted by the model. To describe the effect of product concentration on product formation itself, the model was extended using results from the conventional and the ISPR process, thus agreement between model and experimental data improved notably. Therefore, this model can be a useful tool for the development and optimization of an efficient integrated bioprocess.     

New developments in solid state fermentation: I-bioprocesses and products

The last decade has witnessed an unprecedented increase in interest in solid state fermentation (SSF) for the development of bioprocesses, such as bioremediation and biodegradation of hazardous compounds, biological detoxification of agro-industrial residues, biotransformation of crops and crop-residues for nutritional enrichment, biopulping, and production of value-added products, such as biologically active secondary metabolites, including antibiotics, alkaloids, plant growth factors, etc. enzymes, organic acids, biopesticides, including mycopesticides and bioherbicides, biosurfactants, biofuel, aroma compounds, etc. SSF systems, which during the previous two decades were termed as a ‘low-technology’ systems, appear to be a promising one for the production of value-added ‘low volume-high cost’ products such as biopharmaceuticals. SSF processes offer potential advantages in bioremediation and biological detoxification of hazardous and toxic compounds. With the advent of biotechnological innovations, mainly in the area of enzyme and fermentation technology, many new avenues have opened for the application of SSF. This review discusses more recent developments in the area of SSF leading to the developments of bioprocesses and products.     

Photobioreactors: production systems for phototrophic microorganisms

Microalgae have a large biotechnological potential for producing valuable substances for the feed, food, cosmetics and pharmacy industries as well as for biotechnological processes. The design of the technical and technological basis for photobioreactors is the most important issue for economic success in the field of phototrophic biotechnology. For future applications, open pond systems for large-scale production seem to have a lower innovative potential than closed systems. For high-value products in particular, closed systems of photobioreactors seem to be the more promising field for technical developments despite very different approaches in design.     

Enatiomerically pure hydroxycarboxylic acids: current approaches and future perspectives

The growing awareness of the importance of chirality in conjunction with biological activity has led to an increasing demand for efficient methods for the industrial synthesis of enantiomerically pure compounds. Polyhydroxyalkanotes (PHAs) are a family of polyesters consisting of over 140 chiral R-hydroxycarboxylic acids (R-HAs), representing a promising source for obtaining chiral chemicals from renewable carbon sources. Although some R-HAs have been produced for some time and certain knowledge of the production processes has been gained, large-scale production has not yet been possible. In this article, through analysis of the current advances in production of these acids, we present guidelines for future developments in biotechnological processes for R-HA production.     

Biosurfactant production: emerging trends and promising strategies

Biosurfactants are economically most sought after biotechnological compounds of the 21st century. However, inefficient bioprocessing has mitigated the economical commercial production of these compounds. Although much work is being done on the use of low-cost substrates for their production, a paucity of literature exists on the upcoming bioprocess optimization strategies and their successes and potential for economical biosurfactant production. This review discusses some of the latest developments and most promising strategies to enhance and economize the biosurfactant production process. Recent market analysis, developments in the field of optimally formulated cost credit substrates for enhanced product formation and subsequent process economization are few of the critical aspects highlighted here. Use of nanoparticles and coproduction of biosurfactant along with other commercially important compounds like enzymes, are other upcoming bioprocess intensification strategies. The recent developments discussed here would not only give an overview of pertinent parameters for economic biosurfactant production but would also bring to fore multiple strategies that would open up new avenues of research on biosurfactant production. This would go a long way in making biosurfactants a commercially successful compound of the current century.     

Expression and export: recombinant protein production systems for Aspergillus

Several Aspergillus species, in particular Aspergillus niger and Aspergillus oryzae, are widely used as protein production hosts in various biotechnological applications. In order to improve the expression and secretion of recombinant proteins in these filamentous fungi, several novel genetic engineering strategies have been developed in recent years. This review describes state-of-the-art genetic manipulation technologies used for strain improvement, as well as recent advances in designing the most appropriate engineering strategy for a particular protein production process. Furthermore, current developments in identifying bottlenecks in the protein production and secretion pathways are described and novel approaches to overcome these limitations are introduced. An appropriate combination of expression vectors and optimized host strains will provide cell factories customized for each production process and expand the great potential of Aspergilli as biotechnology workhorses to more complex multi-step industrial applications.     

Production and purification of 2-phenylethanol by Saccharomyces cerevisiae using tobacco waste extract as a substrate

2-phenylethanol (2-PE), which is extracted naturally from plant or biotechnology processing, is widely used in the food and cosmetics industries. Due to the high cost of 2-PE production, the valorization of waste carbon to produce 2-PE has gained increasing attention. Here, 2-PE was produced by Saccharomyces cerevisiae using tobacco waste extract (TWE) as the substrate. Considering the toxicity of nicotine and its inhibition of 2-PE, the tolerance of S. cerevisiae was first evaluated. The results suggested that the production of 2-PE by S. cerevisiae in TWEs could be carried out at 2·0 mg ml⁻¹ nicotine concentrations and may be inhibited by 1·0 mg ml⁻¹ 2-PE. Thus, the compounds in the TWEs prepared at different temperatures were detected, and the results revealed that the TWEs prepared at 140°C contained 2·18 mg ml⁻¹ of nicotine, had total sugar concentrations of 26·8 mg ml⁻¹ and were suitable for 2-PE production. Due to feedback regulation, the 2-PE production was only 1·11 mg ml⁻¹, and the remaining glucose concentration remained at 13·78 mg ml⁻¹, which indicated insufficient glucose utilization. Then, in situ product recovery was further implemented to remove this inhibition; the glucose utilization (the remaining concentration decreased to 3·64 mg ml⁻¹) increased, and the 2-PE production increased to 1·65 mg ml⁻¹. The 2-PE produced in the fermentation broth was first isolated by elution from the resin with 75% ethanol and then by removing the impurities with 2·5% activated charcoal, and pure 2-PE was identified by gas chromatography mass spectrometry. The results of this study suggest that TWE could be an alternative carbon source for 2-PE production. This could provide an outlet tobacco waste as well as reducing the price of natural 2-PE, although more strategies need to be explored to improve the production yield of 2-PE by using TWE.