Yeast flocculation and its biotechnological relevance

Adhesion properties of microorganisms are crucial for many essential biological processes such as sexual reproduction, tissue or substrate invasion, biofilm formation and others. Most, if not all microbial adhesion phenotypes are controlled by factors such as nutrient availability or the presence of pheromones. One particular form of controlled cellular adhesion that occurs in liquid environments is a process of asexual aggregation of cells which is also referred to as flocculation. This process has been the subject of significant scientific and biotechnological interest because of its relevance for many industrial fermentation processes. Specifically adjusted flocculation properties of industrial microorganisms could indeed lead to significant improvements in the processing of biotechnological fermentation products such as foods, biofuels and industrially produced peptides. This review briefly summarises our current scientific knowledge on the regulation of flocculation-related phenotypes, their importance for different biotechnological industries, and possible future applications for microorganisms with improved flocculation properties.     

Anaerobic biotechnological approaches for production of liquid energy carriers from biomass

In recent years, increasing attention has been paid to the use of renewable biomass for energy production. Anaerobic biotechnological approaches for production of liquid energy carriers (ethanol and a mixture of acetone, butanol and ethanol) from biomass can be employed to decrease environmental pollution and reduce dependency on fossil fuels. There are two major biological processes that can convert biomass to liquid energy carriers via anaerobic biological breakdown of organic matter: ethanol fermentation and mixed acetone, butanol, ethanol (ABE) fermentation. The specific product formation is determined by substrates and microbial communities available as well as the operating conditions applied. In this review, we evaluate the recent biotechnological approaches employed in ethanol and ABE fermentation. Practical applicability of different technologies is discussed taking into account the microbiology and biochemistry of the processes.     

Rapid on-line protein detection in biotechnological processes by flow injection analysis (FIA)

Two rapid methods for on-line protein determination useful for control purposes in the automation of biotechnological processes such as fermentation and downstream processing are described. Both methods are derived from colorimetric laboratory biuret and Bradford protein assays adapted to a flow injection analyser.     

Metabolic effects of furaldehydes and impacts on biotechnological processes

There is a growing awareness that lignocellulose will be a major raw material for production of both fuel and chemicals in the coming decades—most likely through various fermentation routes. Considerable attention has been given to the problem of finding efficient means of separating the major constituents in lignocellulose (i.e., lignin, hemicellulose, and cellulose) and to efficiently hydrolyze the carbohydrate parts into sugars. In these processes, by-products will inevitably form to some extent, and these will have to be dealt with in the ensuing microbial processes. One group of compounds in this category is the furaldehydes. 2-Furaldehyde (furfural) and substituted 2-furaldehydes—most importantly 5-hydroxymethyl-2-furaldehyde—are the dominant inhibitory compounds found in lignocellulosic hydrolyzates. The furaldehydes are known to have biological effects and act as inhibitors in fermentation processes. The effects of these compounds will therefore have to be considered in the design of biotechnological processes using lignocellulose. In this short review, we take a look at known metabolic effects, as well as strategies to overcome problems in biotechnological applications caused by furaldehydes.     

Biotechnological solubilization of rock phosphate on media containing agro-industrial wastes

Rock phosphate (RP) is an important natural material traditionally used for the production of phosphorus (P) fertilizers. Compared with chemical treatment, microbial solubilization of RP is an alternative environmentally mild approach. An overview of biotechnological techniques, mainly based on solubilization processes involving agro-industrial residues, is presented. Potential advantages of composting, solid-state fermentation, and liquid submerged fermentation employing free and immobilized microorganisms that produce organic acids and simultaneously solubilize RP are discussed. Subsequent introduction of the final fermented products into soil-plant systems promotes plant growth and P acquisition.     

Oil cakes and their biotechnological applications--a review

Oil cakes have been in use for feed applications to poultry, fish and swine industry. Being rich in protein, some of these have also been considered ideal for food supplementation. However, with increasing emphasis on cost reduction of industrial processes and value addition to agro-industrial residues, oil cakes could be ideal source of proteinaceous nutrients and as support matrix for various biotechnological processes. Several oil cakes, in particular edible oil cakes offer potential benefits when utilized as substrate for bioprocesses. These have been utilized for fermentative production of enzymes, antibiotics, mushrooms, etc. Biotechnological applications of oil cakes also include their usages for vitamins and antioxidants production. This review discusses various applications of oil cakes in fermentation and biotechnological processes, their value addition by implementation in feed and energy source (for the production of biogas, bio-oil) as well.     

Biotechnological routes based on lactic acid production from biomass

Lactic acid, the most important hydroxycarboxylic acid, is now commercially produced by the fermentation of sugars present in biomass. In addition to its use in the synthesis of biodegradable polymers, lactic acid can be regarded as a feedstock for the green chemistry of the future. Different potentially useful chemicals such as pyruvic acid, acrylic acid, 1,2-propanediol, and lactate ester can be produced from lactic acid via chemical and biotechnological routes. Here, we reviewed the current status of the production of potentially valuable chemicals from lactic acid via biotechnological routes. Although some of the reactions described in this review article are still not applicable at current stage, due to their “greener” properties, biotechnological processes for the production of lactic acid derivatives might replace the chemical routes in the future.     

The production of l-lysine by fermentation

Over 40 000 tons of L-lysine are used each year, chiefly as a nutritional supplement in animal feeds. Two major biotechnological processes are used to manufacture lysine: the enzymatic conversion of dl-α-amino-ε-caprolactam, and the direct fermentation of molasses or starch by microbes, especially species of Brevibacteria and Corynebacterium glutamicum. This review focuses on the fermentative processes and describes the development of various mutants which over-produce l-lysine.     

Biotechnological production of enantiomerically pure <Emphasis Type="Italic">d</Emphasis>-lactic acid

The fermentation process of l-lactic acid is well known. Little importance was attached to d-lactic acid, but in the past 10 years, d-lactic acid gained significantly in importance. d-Lactic acid is an interesting precursor for manufacturing heat-resistant polylactic acid (PLA) bioplastics which can be widely used, for example as packaging material, coatings, for textiles or in the automotive industry.This review provides a comprehensive overview of the most recent developments, including a spectrum of studied microorganisms and their capabilities for the production of d-lactic acid. Additionally, the technological achievements in biotechnological d-lactic acid production including fermentation techniques like fed batch, simultaneous saccharification, and fermentation and continuous techniques are presented. Attention is also turned to suitable alternative substrates and their applicability in fermentation processes. Furthermore, advantages and disadvantages of product recovery and purification are discussed. Economic aspects of PLA are pointed out, and the present industrial producers of lactic acid are briefly introduced.     

Desaturase-defective fungal mutants: useful tools for the regulation and overproduction of polyunsaturated fatty acids

Increasing demand for biologically important polyunsaturated fatty acids has led to the search for alternative sources, especially fungi. Although successes in this area have induced interest in developing fungal fermentation processes, manipulation of their lipid composition requires new biotechnological strategies to achieve high yields of the desired polyunsaturated fatty acids. Fungal desaturase mutants with unique enzyme systems are useful not only for the regulation and overproduction of valuable polyunsaturated fatty acids but also because they are excellent models for the elucidation of fungal lipogenesis.     

Progress in the development of methods used for the abatement of microbial contaminants in ethanol fermentations: a review

Biofuel research and development roadmap is currently underway in several countries and is expected to pave a way for the establishment of a viable renewable energy sector that can compete with petroleum-based fuels. Ethanol fermentation has garnered increasing attention amongst various stakeholders (industries, governments, and academia) due to its economic and environmental merits. However, microbial contamination continues to be one of the major barriers in ethanologenic processes, resulting in low ethanol yields and thereby translating into economic losses. To this end, technological innovations geared towards effective elimination of microbial contamination are constantly being developed. This review explores and discusses the fermentation conditions that facilitate the growth of undesired microorganisms during ethanol fermentation processes. It highlights the methods that are currently used in biorefineries as well as innovative and advanced biotechnological methods currently being evaluated as viable alternative strategies to control or eliminate microbial contaminants in ethanol fermentations. These methods have the potential to minimize or control the contamination problem and could pave a way for the development of an efficient biofuel sector.

     

New insights and novel developments in clostridial acetone/butanol/isopropanol fermentation

Clostridial acetone/butanol fermentation used to rank second only to ethanol fermentation by yeast in its scale of production and thus is one of the largest biotechnological processes known. Its decline since about 1950 has been caused by increasing substrate costs and the availability of much cheaper feedstocks for chemical solvent synthesis by the petrochemical industry. The so-called oil crisis in 1973 led to renewed interest in novel fermentation and product recovery technologies as well as in the metabolism and genetics of the bacterial species involved. As a consequence, almost all of the enzymes leading to solvent formation are known, their genes have been sequenced (in fact, Clostridium acetobutylicum has been recently included in the microbial genome sequencing project), the regulatory mechanisms controlling solventogenesis have begun to emerge and recombinant DNA techniques have been developed for these clostridia to construct specific production strains. In parallel, cheap agricultural-waste-based feedstocks have been exploited for their potential as novel substrates, continuous culture methods have been successfully established and new on-line product recovery technologies are now available, such as gas stripping, liquid/liquid extraction, and membrane-based methods. In combination with these achievements, a reintroduction of acetone/butanol fermentation on an industrial scale seems to be economically feasible, a view that is supported by a new pilot plant in Austria recently coming into operation. Received: 18 December 1997 / Received revision: 27 January 1998 / Accepted: 27 January 1998     

Development of Suitable Solvent System for Downstream Processing of Biopolymer Pullulan Using Response Surface Methodology

Downstream processing is an important aspect of all biotechnological processes and has significant implications on quality and yield of the final product. Several solvents were examined for their efficacy on pullulan precipitation from fermentation broth. Interactions among four selected solvents and their effect on pullulan yield were studied using response surface methodology. A polynomial model was developed using D-optimal design and three contour plots were generated by performing 20 different experiments and the model was validated by performing optimization experiments. The results indicated that lower concentration of ethanol in combination with the other three solvents has resulted in higher yield of polymer from fermentation broth and the optimized solvent system was able to recover 1.44 times more pullulan as compared to the conventional ethanolic precipitation method. These observations may help in enhancing efficiency of pullulan recovery from fermentation broth and also result in reduced cost of production for the final product.     

Process design and economic studies of alternative fermentation methods for the production of ethanol

Cell recycle and vacuum fermentation processes are described for the continuous production of ethanol. Preliminary process design studies are employed to make an economic comparison of these alternative fermentation schemes with continuous and batch fermentation technologies. Designs are based on a production capacity of 78,000 gal 95% ethanol (EtOH)/day employing molasses as the fermentation substrate. The studies indicate that a 57% reduction in fixed capital investment is realized by continuous rather than batch operation. Further decreases in required capital investment of 68 and 71% over batch fermentation were obtained for cell recycle and vacuum operation, respectively. However, ethanol production costs were dominated by the cost of molasses, representing over 75% of the total manufacturing cost. But, when a reasonable yeast by-product credit was assumed, the net production cost for 95% ethanol was estimated at 82.3 and 80.6 cent/gal, for the cell recycle and vacuum processes, respectively.     

Biofuels – challenges & chances: How biofuel development can benefit from advanced process technology

The presented article highlights the process of biofuel production with a special focus on bioethanol. After a short introduction to the “problems” of biofuels – the “first generation” biofuels (in regards to their competition to feed and food production) and the “second generation” biofuels (in regards to the required more complex process technology) − the different steps in the process from natural resources towards the final product are presented and the underlying biotechnological challenges discussed: the pre-treatment of the natural resources followed by the biotechnological processes of hydrolysis and fermentation. Topics such as enzyme screening for efficient or even multi-step hydrolysis as well as microbial strain selection under process conditions and the optimization of the anaerobic fermentative conversion of the saccharides to bioethanol are discussed. Optimizing the production of bioethanol to be competitive with petrochemical fuels is the main challenge for the underlying process development.     

By-product inhibition effects on ethanolic fermentation by Saccharomyces cerevisiae

Inhibition by secondary fermentation products may limit the ultimate productivity of new glucose to ethanol fermentation processes. New processes are under development whereby ethanol is selectively removed from the fermenting broth to eliminate ethanol inhibition effects. These processes can concentrate minor secondary products to the point where they become toxic to the yeast. Vacuum fermentation selectively concentrates nonvolatile products in the fermentation broth. Membrane fermentation systems may concentrate large molecules which are sterically blocked from membrane transport. Extractive fermentation systems, employing nonpolar solvents, may concentrate small organic acids. By-product production rates and inhibition levels in continuous fermentation with Saccharomyces cerevisiae have been determined for acetaldehyde, glycerol, formic, lactic, and acetic acids, 1-propanol, 2-methyl-1-butanol, and 2,3-butanediol to assess the potential effects of these by-products on new fermentation processes. Mechanisms are proposed for the various inhibition effects observed.     

An impaired ubiquitin ligase complex favors initial growth of auxotrophic yeast strains in synthetic grape must

We used experimental evolution in order to identify genes involved in the adaptation of Saccharomyces cerevisiae to the early stages of alcoholic fermentation. Evolution experiments were run for about 200 generations, in continuous culture conditions emulating the initial stages of wine fermentation. We performed whole-genome sequencing of four adapted strains from three independent evolution experiments. Mutations identified in these strains pointed to the Rsp5p-Bul1/2p ubiquitin ligase complex as the preferred evolutionary target under these experimental conditions. Rsp5p is a multifunctional enzyme able to ubiquitinate target proteins participating in different cellular processes, while Bul1p is an Rsp5p substrate adaptor specifically involved in the ubiquitin-dependent internalization of Gap1p and other plasma membrane permeases. While a loss-of-function mutation in BUL1 seems to be enough to confer a selective advantage under these assay conditions, this did not seem to be the case for RSP5 mutated strains, which required additional mutations, probably compensating for the detrimental effect of altered Rsp5p activity on essential cellular functions. The power of this experimental approach is illustrated by the identification of four independent mutants, each with a limited number of SNPs, affected within the same pathway. However, in order to obtain information relevant for a specific biotechnological process, caution must be taken in the choice of the background yeast genotype (as shown in this case for auxotrophies). In addition, the use of very stable continuous fermentation conditions might lead to the selection of a rather limited number of adaptive responses that would mask other possible targets for genetic improvement.     

Pervaporative butanol fermentation using a new bacterial strain

Fermentation processes for the production of butanol had an economic importance in the first part of this century. Today butanol is commercially produced from the Oxo reaction of propylene because relatively low priced propylene during the cracking of petroleum. Efforts have been made during the past decade or two to improve the productivity of butanol fermentation processes. It includes strain improvements, continuous fermentation processes, cell immobilization and simultaneous product separation. This review introduces a new butanol fermentation process using pervaporative product separation and a new bacterial strain producing less amount of organic acids. This review also compares the new process with chemical processes. This kind of new fermentation process may be able to compete with the chemical synthesis of butanol and revitalize the butanol fermentation process.