1 An introduction to biotechnological innovations in the chemical industry ~ 1.1 Introduction 1.2 Production processes 1.3 Choice of production pnxess 2 Chapter 1 non-renewable fossil oil renewable biibgical material mitoperations upstream& downstream operations An introduction to biotechnological innovations in the chemical industry 1.1 Introduction Although chemistry as an emperical fundamental dicipline has a long history, its application in industry gained importance after the introduction of the use of fossil energy sources during the industrial revolution. The chemical industry withdraws materials, mainly fossil oil, from the earth's reserves to use as an energy source and as a source of raw materials for production processes. The products so produced are rather different from naturally occurring materials and mankind has become heavily dependent on them. However, other energy and raw material sources have to be sought because fossil sources are non-renewable and will eventually become depleated. Nowadays, renewable biological materials such as starch, sugar, oils and molasses are used on a relatively small scale as energy and raw materials for the chemical industry. These biological sources are mainly derived from waste products and overproduction in agriculture. With these materials less drastic conversions are applied, in comparison with fossil oil, in order to make as much as possible of the chemical structure present in the raw material. Such conversions are therefore generally performed by micruorganisms or parts of them. This means that the production process includes at least one biological conversion step. 1.2 Production processes In production processes, raw material are converted into desired products using a series of unit operations. Such unit operations may be few in number and they are linked together in a logical sequence. Typical unit operations include such activities as the transport of solids and liquids, the transfer of heat, crysallisation, collection and drymg. In a chemical production process at least one of the unit operations (the chemical reactor) is the place in which chemical conversion takes place. However, the chemical reactor is proceeded by a series of unit operations in which the new materials are prepared (the upstream operations). After conversion has taken place, the products subjected to a further series of unit operations (the downstream operations). These downstream operations include product recovery and purification steps. A typical example of a production process is illustrated in Figure 1.1. Which of the unit operations described in Figure 1.1 represents the chemical n reactor? You should have identified the "conversion step" An introduction to biotechnological innovations in the chemical industry Figure 1.1 Example of a simple production process with eight unit operations. In practice, production processes are usually rather more complex. Raw materials are usually impure and thus some pre-purification steps may be required. Obviously impurities in the raw materials will incresae the probability of impurities and byproducts occuring in the output stream from the chemical conversion step. Even using pure raw materials, most chemical conversion are incomplete and often lead to the formation of undesirable byproducts. Furthermore often additional (auxiliary) materials are used (for example catalysts, specific solvents), which have to be separated from the desired product, Thus, in typical production processes a large number of separation steps are required. To improve the efficiency of the process, raw materials and auxiliary chemicals are recycled providing it is economically viable. Similarly ways are sought to find uses for byproducts and intermediates. This usually involves using them as feeds for further reactions. Invariably, production processes produce waste streams. These must be brought to an acceptable state before being disposed of. This is especially a concern relating to chemical production processes in which the compounds produced may be incornpatable or toxic to living systems and can thus cause pollution problems. Increasing regulatory and technical burdens are being place on chemical p'ocess operators to ensure that such environmental problems do not arise from their operations. In biotechnological processes, the conversion of raw material to product is usually performed by micm-organisrns, or parts of micro-organisms (eg enzymes) known as a fermentation or bioconversion processes respectively. On a large scale, the conversion is generally carried out in a so-called bioreactor. The conditions under which the conversion is done are generally very gentle with regards to temperature, pressure and pH, when compared to those in a chemical process. Other advantages of biotechnological production processes include high reaction specificity and selectivity (therefore fewer byproducts), and the need for relatively few reaction additives. Another important difference between a chemical and a biotechnological production process is that the latter type is closely related to naturally occurring processes: byproducts may only be carbon dioxide and water. The implementation of biotechnological production methods can, therefore, be seen as an environmentally friendly production strategy. We shall compare chemical and biotechnological catalysis in more detail in the next chapter. Within the chemical industry, micruorganisms and enzymes are often used as catalysts. It is possible for a unit operation in an essentially chemical production process to be a biochemically catalysed step: giving rise to a mixed chemical/biochemical production process. The products of these reactions include organic chemicals, solvents, polymers, pharmaceuticals, and purfumes. Mixed chemical/biochemical production processes are continuously innovated and optimised, mainly for economical reasons. bypm~cts auXiliar~ mabe~ bD&&mbgM processes are mom enviro""'~~\ m,xed chemical/ biochemical pro^^^ 4 Chapter 1 economic COnsidetations specificity of readon biodegradable pro&* 1.3 Choice of production process It is possible to produce many chemical moieties (partly) by means of biotechnological production processes. For example, ethene (Cd-h) commands a large market and is produced from fossil oil. This chemical can also be produced from ethanol, which in turn can be produd by micro-organisms using agricultural wastes. Ethene is a 'building unit' for the petrochemical industry from which several other intermediates and end products such as plastics are produced. The biotechnological production mthM is, however, for economical reasons still not used in practice. The costs of producing ethene from sugar via ethanol are relatively too high. This is partly due to the cost of the raw materials and the product yields on the two different substrates. The separation of ethanol from the aqueous fermentation liquid is also relatively expensive. Unfortunately micmrganisms that make large amounts of ethene directly from glucose have not been found. Nevertheless, the biotechnological production method may become the cost effective option as fossil energy sources become depleted and relatively more expensive. Other chemicals such as gluconic acid cannot be produced by petrochemical production methods. Gluconic acid is used in the pharmaceutical industry and even as an addition to concrete. Gluconic acid can be produced from glucose, derived from potato starch, using the bacterium Gluconobucfer or the fungus Aspergillus. These micro-organisms are able to modify glucose rapidly to gluconic acid, which is slowly consumed again. In this way gluconic acid is temporarily accumulated in the fermentation fluid. Technically, it would be possible to perform this process chemically starting from glucose, but in this case the biological method is preferred as the specifity of this reaction is very high and thus no undesired byproducts are formed. For the same reason biotechnological production processes are preferred if optically pure chiral compounds, such as Lconfiguration of a certain amino acid, has to be produced. The price of such products is, however, relatively more expensive. In later chapters of this text we consider the production of organic acids and amino acids in some detail. Another example of producing a chemical in bulk from sugar with the help of a micro-organism is the polymer polyhydroxybutyric acid. Many micmorganisms accumulate this compound as a reserve material. This polymer could be substitute for polyester or polypropene plastics. The big advantage of polyhydroxybutyric acid is that it can be degraded microbially. Products such as plastics that have been used for shorter or longer times and when they are not needed any more are brought back into the environment. However, when returned to the environment they are not readily biodegraded (they are recalcitrant) and thus accumulate. The accumulation of materials that are not readily returned to natural geocycling is of mapr concern. In a world where mankind has become aware that more sustainable environmental practice have to be used to prevent pollution, biotechnology will become more and more important. A first obvious consequence of such considerations is that we should not only look at the costs of the product from an economic point of view, but that we must consider the costs of the production process in a broader sense. We must take into account the raw materials used, the amount of energy invested and the possibility to design alternatives, more environmentally friendly processes. In other words, we should not only look at the desired product, but we must consider the total life cycle of the product. The design of a production process taking into account these aspects is often referred to as integral life cycle management. An introduction to biotechnological innovations in the chemical industry 5 The application of the principles of integrated life cycle management, generally favours the replacement of products dependent upon conventional chemical and physical processes by biotechnological products and processes. As we described earlier, most biotechnological processes use biological (renewable) feedstocks and energy sources and the products are also cornpatable with biological (living) system. These products are readily biodegradable and returned to the natural geocycling and, as a consequence, do not pose the same intensity of pollution caused by the recalcitrant materials and byproducts generated by physicochemical processes. Biotechnology therefore offers a more environmentally friendly and sustainable approach to fulfilling the needs of society. It can achieve this by, for example, offering alternative routes to the manufacture of products hitherto made by potentially environmentally damaging routes. Alternative, it enables the production of novel products, which are less environmentally damaging than products made via conventional chemical routes. We will use two examples to illustrate these principles. Nitrogen fixation via the Haber-Bosch process is a well established chemical process in which dinitrogen gas (N2) and hydrogen are combined to produce ammonia (NEG). The major use of this produce is as a nitrogen fertiliser. Several million tonnes are produced annually. On the positive side, use of ammonia (usually as an ammonium salt) has undoubtedly increased the yields of crops. In strictly limited economic terms, the increase in crop yields achieved by the use of ammonia from the Haber-Bosch process more than outweighs the cost of producing the ammonia. With this limited perspective, the Haber-Bosch process is undoubtedly successful. If, however, we take an integrated life management approach, the issue is not so clear cut. The reduction of dinitrogen is an energy expensive process. Energy is needed to split the stable N=N bond. In the Haber-Bosch process, high temperatures (400°C) and pressures are used to achieve significant conversions. This energy input is invariably derived from non-renewable energy sources. However, the environmentally damaging effects of this activity is not limited to the production of ammonia. Much of the ammonia-based fertilisers applied to land is washed out (leached) from soils. This ends up in rivers and in impounded water, causing eutrophication (increase in organic content). The consequence of this, is these waters support greater 'blooms' of algae, which in time die and decompose. This decomposition is accompanied by the consumption of oxygen, which tends to lead to anoxia. Thus the waters lose amenity value because they no longer support fish life, are more difficult to treat to become potable; they become odourous and are no longer suitable for bathing. Thus, if one adds to the cost of the Haber-Bosch process the true environmental costs, then the virtue of this process is less than clear cut. Biotechnology, however, offers an alternative approach to achieving the same objective as the Haber-Bosch process. It has long been known that bacteria capable of utilising atmospheric nitrogen can supply plants with nitrogen in a form that the plants can use and very little of this "fixed" nitrogen is leached from the soil. In essence, what biotechnology offers is the potential to widen the range of crops that can be supported using bioiogically generated nitrogen fertilisers. These biological nitrogen-fixers use biological energy sources (carbohydrates) to drive fixation and do not lead to the same levels of entrophication as does the application of chemically produced ammonia. Even on the rather simplified arguments described here, it should be clear that biotechnological approaches are generally more environmentally friendly and that we can apply biotechnological strategies to inorganic, as well as organic chemicals. We nitrogen fixation eutrophication 6 Chapter 1 pesticides biomagnification biodegradable pesticides campetition in the market Place legislation could, for example, cite the use of micrmrganisms in the mining of a range of metals from low grade ores, by processes generally referred to as "acid mining". In the example above we have illustrated how biotechnology may, in integrated life management terms, offer environmentally better routes of manufacture. It may also lead to environmentally more acceptable products. As an example consider the production and use of pesticides. The majority of pesticides are made by synthetic organic chemisry leading to molecules that are distinctly non-biological. Often they contain functional groups (for example halogens) and are made by reaction mechanisms (for example using free radicles) that are, in general, incompatable with biodegradation. These products, although used in low concentration, tend to accumulate in the environment especially within biological systems. Of particular importance is the accumulation of these materials in relatively high concentrations in organisms at the end of food chains. Thus an insecticide may be present only in low levels in particular insects but when these are eaten by birds, the biological part of the insect is metabolised, while the recalcitrant insecticide remains. Thus the concentration of the insecticide becomes greater in the birds than in its food. This process, called biomagnification, may result in the concentrations of the insectide in the birds reaching toxic levels. Furthermore, chemically produced pesticides generally have wide ranging activities, killing both beneficial as well as pest species. The advent of contemporary biotechnology has enabled development of new strategies to achieve the same objectives: protecting crops using biologically-produced, biodegradable pesticides. A typical example is the production and use of proteinaceous insect toxins encoded by Bad0 virus. These types of pesticides are readily biogradable and are target specific. We will not enlarge on the environment potential of biotechnology any further at this stage. We will, however, raise some environmental issues in later section of this text If you would like to learn more, we recommend the BIOTOL text "Biotechnological Innovations in Environmental and Energy Management". The development of new products based on cleaner production processes and alternative raw resources is not only a question of technological development. The products have to compete in the market place and have to be acceptable to potential customers. Also the introduction of new processes and products depends upon gaining both the confidence and the financial resources of potential investors. Regulations may also greatly influence (both positively and negatively) the adoption of these new processes. Restrictive regulations may deter investors and may, by raising the spectra of potential hazards, alienate the general public and reduce the accetability of the products. We site for example the EC Directives and National Legislation concerning the safe handling of netically mani ulated organisms. To many workers this is seen Such legislation is seen as a constraint on the development of new processes and products. To others, this leglation is a positive bonus to biotechnology because it reduces the prospect of there being a major biocatastrophy from these activities and it reassures the public that the work is undertaken in a safe manner and leads to "safe" products, thereby making them more acceptable to the public. This in turn encourages investment and development. However the reverse effect may also be true. To some, if biotechnology is "safe" it would not need to be regulated in this way. In effect, for some individuals the intmduction of legislation indicates that biotechnology is inherently "unsafe", as a result this legislation may in some circumstances, make biotechnology and its products less publically acceptable. as inhibitoxy to the r evelopment an B exploitation of genetically modified organisms. An introduction to biotechnological innovations in the chemical industry 7 role of legi shtions , politicians, investors, pressure groups on biotechnobgical development Thus we can see that legislators, politicians, investors and society as a whole are important influences on the development and adoption of biotechnology. Biotechnological development is not only dependant upon technological/scientific advance but also economic, political and sociological developments. We could, for example, envisage that specialist lobby groups (eg "Green" groups, animal rights activists) may, through influencing public opinion, greatly influence biotechnology. It is also incuberent on education to ensure that public opinion (and thus investment and legislation) is developed upon knowledge and not upon emotive and ill-founded claims. In the following chapters, we predominantly use a case study approach to illustrate a range of issues that arise from using biologically-based approaches to the production of chemicals. There are such an enormous range to choose from that we have had to be selective. Our selection has been made predominantly to ensure that the reader develops an understanding of the range and potential of biotechnology in this area, and developes an appreciation of the major advantages and limitations of this approach. In Chapter 2, we provide an overview of the types of chemical transformations that can be mediated by organisms or their constituent parts, particularly enzymes. This chapter provides a context for later chapters. In Chapter 3, we examine the cellular energetic consequences of metabolite overproduction by organisms. We use this chapter to consider the limitations on yield of products. Chapter 4 considers the production of single cell protein, particularly using fossil fuel (methane/methanol) as substrate. This chapter enables us to explain how changing market values and social acceptability atly influences the success or otherwise of biotechnological processes. In the case of !& from methane, we use economic data from the 1960s (when the process appeared to be profitable) and the 1970s (when it became uneconomic) to illustrate this point. We feel further justification for including this case study because it was this project that led to the commercial development of large scale air-lift bioreactors which find ever increasing use in a wide range of biotechnological processes. We have included chapters on large volume organic aad production to illustrate how intermediary metabolism may be manipulated to achieve overproduction of metabolic intermediates. Chapters onantibiotics, amino acids, polysaccharides and lipids are used to illustrate the application of biological systems to achieve specific transformations. Each has been chosen to enlarge on particular aspects. Thus within the amino acids chapter you will for example compare the technology and the economics of fermentative and enzymological strategies to produce stereospecific forms of particular amino acids. In the chapter on antibiotics, we illustrate how biological systems can be manipulated or used to diversify the range and characteristics of particular groups of molecules. Similarily, in the chapter on lipids, we use sterol/steroid interconversions in the health care sector to enlarge on the concepts of biological speafiaty. Later in the same chapter we turn attention to the bulk lipid market associated with food manufacture and show how biological systems can be used to convert a low prices lipid into a higher value lipid with desirable organoleptic ("mouth feel") properties. The chapter is also used to introduce the reader to the problems posed by attempting to use biological system (which are largely aqueously based) to carry out conversion with substances which have only limited compatability with water. By the end of the text you should appreciate the enormous potential that biological system have for malung a wide range of products and to achieve a variety of objectives. You should also have knowledge and be able to ate specific examples, of how economic, social and political attitudes may impinge upon the adoption of the technology.