We are in a phase of the evolution of biotechnology in which the true and potential commercial importance of carbohydrates is becoming appre- ated more fully. Progress in providing hard facts to establish the commercial value ofpolysaccharides and oligosaccharides is limited, as always, by lack of funding and by a relative shortage of skilled practitioners in the production and analysis of those materials. Carbohydrate science has a reputation, not unmerited, for technical difficulty owing to the structural similarity of the many monosaccharide monomers and the potential, and real, complexity of oligosaccharides and polysaccharides, particularly heterosaccharides conta- ing many different monomers. Modem analytical and synthetic methods, in many cases using enzyme technology, are beginning to allow this complexity to be unraveled. Carbohydrate Biotechnology Protocols is aimed at those newcomers who have an interest in the production and use of carbohydrate materials, but have shied away from involvement for lack of detailed descriptions of appropriate methods, including the type of practical hints that may be provided by those skilled in those methods, but that are rarely described in research papers. The majority of the contributions to this book conform to the established format of the Methods in Biotechnology series. They begin with the theoretical and c- mercial background to the method or group of methods, provide a list of the reagents and equipment required for the procedure, then give a detailed st- by-step description of how to carry out the protocol.

Enzymes and whole cells are able to catalyze the most complex chemical processes under the most benign experimental and environmental conditions. In this way, enzymes and cells could be excellent catalysts for a much more sustainable chemical industry. However, enzymes and cells also have some limitations for nonbiological applications: fine chemistry, food chemistry, analysis, therapeutics, and so on. Enzymes and cells may be unstable, difficult to handle under nonconventional conditions, poorly selective toward synthetic substrates, and so forth. From this point of view, the transformation—from the laboratory to industry—of chemical processes catalyzed by enzymes and cells may be one of the most complex and exciting goals in biotechnology. For many industrial applications, enzymes and cells have to be immobilized, via very simple and cost-effective protocols, in order to be re-used over very long periods of time. From this point of view, immobilization, simplicity, and stabilization have to be strongly related concepts. Over the last 30 years, a number of protocols for the immobilization of cells and enzymes have been reported in scientific literature. However, only very few protocols are simple and useful enough to greatly improve the functional properties of enzymes and cells, activity, stability, selectivity, and related properties.

A collection of readily reproducible classic and emerging molecular methods for the laboratory isolation and identification of the pathogens, viruses, and parasites that cause food-borne disease. Among the pathogens covered are specific bacteria, including Salmonella spp, Campylobacter spp., Listeria spp., and Bacillus spp.; viruses, including noroviruses and enteroviruses; and parasites, including Cryptosporidium and seafood nematode worms. The protocols follow the successful Methods in BiotechnologyTM series format, each offering step-by-step laboratory instructions, an introduction outlining the principles behind the technique, lists of the necessary equipment and reagents, and tips on troubleshooting and avoiding known pitfalls.

A mixture of two polymers, or one polymer and a salt, in an aqueous medium separates into two phases: this phenomenon is useful in biotechn- ogy for product separations. Separation of biological molecules and particles in these aqueous two-phase systems (ATPS) was initiated over 40 years ago by P.-Å. Albertsson, and later proved to be of immense utility in biochemical and cell biological research. A boost in the application of ATPS was seen when problems of separations in biotechnology processes were encountered. Its simplicity, biocompatibility, and amenability to easy scaleup operations make the use of ATPS very attractive for large-scale bioseparations. Despite the advantages ATPS enjoys over other separation techniques, the application of two-phase systems has for a long time been confined to selected labora- ries. Recent years have, however, shown a trend in which increasing numbers of researchers employ two-phase partitioning techniques in both basic and applied research.

Enzymatic catalysis has gained considerable attention in recent years as an efficient tool in the preparation of natural products, pharmaceuticals, fine chemicals, and food ingredients. The high selectivity and mild reaction con- tions associated with enzymatic transformations have made this approach an attractive alternative in the synthesis of complex bioactive compounds, which are often difficult to obtain by standard chemical routes. However, the maj- ity of organic compounds are not very soluble in water, which was traditi- ally perceived as the only suitable reaction medium for the application of biocatalysts. The realization that most enzymes can function perfectly well under nearly anhydrous conditions and, in addition, display a number of useful properties, e. g. , highly enhanced stability and different selectivity, has d- matically widened the scope of their application to the organic synthesis. Another great attraction of using organic solvents rather than water as a reaction solvent is the ability to perform synthetic transformations with re- tively inexpensive hydrolytic enzymes. It is worth reminding the reader that in vivo, the synthetic and hydrolytic pathways are catalyzed by different enzymes. However, elimination of water from the reaction mixture enables the “reversal” of hydrolytic enzymes and thus avoids the use of the expensive cofactors or activated substrates that are required for their synthetic count- parts.

The methods included in Environmental Microbiology: Methods and Pro- cols can be placed in the categories “Communities and Biofilms,” “Fermented Milks,” “Recovery and Determination of Nucleic Acids,” and the review s- tion, containing chapters on the endophytic bacterium, Bacillus mojavensis, the engineering of bacteria to enhance their ability to carry out bioremediation of aromatic compounds, using the hemoglobin gene from a strain of Vitreoscilla 23 spp., and the use of chemical shift reagents and Na NMR to study sodium gradients in microorganisms, all of which should be of interest to investigators in these fields. The subjects treated within the different categories also cover a wide range, with methods ranging from those for the study of marine organisms, through those for the investigation of microorganisms occurring in ground waters, including subsurface ground waters, to other types of environmental waters, to as varied subjects as the biodiversity of yeasts found in northwest Argentina. The range of topics described in the Fermented Milks section is smaller, but significant for investigators in areas concerned with milk as an item of foods for infants, small children, and even adults.

Considerable effort and time is allocated to introducing cell culture and fermentation technology to undergraduate students in academia, generally through a range of courses in industrial biotechnology and related disciplines. Similarly, a large number of textbooks are available to describe the appli- tions of these technologies in industry. However, there has been a general lack of appreciation of the significant developments in downstream processing and isolation technology, the need for which is largely driven by the stringent re- latory requirements for purity and quality of injectable biopharmaceuticals. This is particularly reflected by the general absence of coverage of this s- ject in many biotechnology and related courses in educational institutions. For a considerable while I have felt that there is increasing need for an introductory text to various aspects of downstream processing, particularly with respect to the needs of the biopharmaceutical and biotechnology ind- try. Although there are numerous texts that cover various aspects of protein purification techniques in isolation, there is a need for a work that covers the broad range of isolation technology in an industrial setting. It is anticipated that Downstream Processing of Proteins: Methods and Protocols will play a small part in filling this gap and thus prove a useful contribution to the field. It is also designed to encourage educational strategists to broaden the coverage of these topics in industrial biotechnology courses by including accounts of this important and rapidly developing element of the industrial process.