Biocatalysis in Green Solvents

By Pedro Lozano

Biocatalysis in Green Solvents offers a pragmatic overview and instruction in biocatalysis and enzymology of green solvents for sustainable industries and medicine, running from concept to application. Here, international experts in the field discuss structure-function relationships of enzymes in ionic liquids (ILs) and examine how enzymes act as selective catalysts for fine biochemical synthesis in non-aqueous environments. Several integral green biochemical processes of biocatalytic transformation and pure product separation are described in detail.

Application focused chapters discuss the role of biocatalysis in creating and implementing deep eutectic solvents, biomass derived solvents, sub and supercritical fluids, carbon dioxide biphasic systems, and enzymatic membrane reactors, as well as applying these biocatalytic processes in drug discovery and production.

• Examines the structure-function relationships of enzymes in ionic liquids and biocatalytic processes for various applications across industry and medicine
• Includes clear instruction in core green biochemical processes—of (bio)catalytic transformation—and pure product separation
• Features chapter contributions from international experts across academia and industry

• Language: English
• Publisher: Academic Press
• Release date: Aug 2, 2022
• ISBN: 9780323914253

Book preview

Chapter 1

Biocatalysis, solvents, and green metrics in sustainable chemistry

Roger Arthur Sheldon¹,²,    ¹School of Chemistry, University of the Witwatersrand, Johannesburg, South Africa,    ²Department of Biotechnology, Technical University of Delft, Delft, The Netherlands

Abstract

A sustainable production process should generate minimal waste through the efficient use of (preferably renewable) resources and avoid the use of toxic and hazardous reagents and solvents. This is stimulated by the application of green metrics, such as the E-factor and atom economy, to measure the generation of waste. One way to significantly reduce waste is to replace archaic processes using stoichiometric amounts of reagents with greener catalytic alternatives. Biocatalysis, involving the use of enzymes as catalysts, has many advantages in this respect. The use of organic solvents is another major contributor to waste and substantial reductions in solvent use and avoiding the use of toxic and/or hazardous solvents is the key to sustainability. An important advantage of biocatalysis is, therefore, that it is favored in an aqueous medium. However, this can present problems with hydrophobic substrates, and various approaches to afford resource efficient and sustainable biocatalytic processes are outlined.

Keywords

Green chemistry; sustainability; metrics; waste minimization; solvents; biocatalysis; aqueous media; micellar catalysis

1.1 Introduction to green chemistry and sustainability

Thirty years ago, there was growing environmental concern regarding the generation of waste and the use of toxic and hazardous materials by the chemical industry. In particular, the production of fine chemicals, for example, flavors and fragrances, and active pharmaceutical ingredients (APIs) primarily involved the use of stoichiometric inorganic and organic reagents that generated copious amounts of waste. Catalytic processes, widely applied in bulk commodity chemicals manufacture, were almost totally absent. Synthetic organic chemists preferred the more familiar stoichiometric reagents for example, metal hydrides for reductions and stoichiometric inorganic oxidants such as CrVI reagents for oxidations, rather than for example, catalytic hydrogenation and aerobic oxidation. An important reason for clinging to these well-established reagents, such as chromium (VI) oxidants, was their broad scope in organic synthesis.

However, increasing environmental awareness created a pressing need for alternative, cleaner processes that are more resource efficient and generate less waste, with emphasis on waste prevention rather than waste remediation. A paradigm shift was clearly needed, from traditional concepts of reaction efficiency and selectivity, focusing largely on chemical yield, to one that assigns value to maximizing raw materials utilization, eliminating waste and avoiding the use of toxic and/or hazardous substances [1]. Furthermore, the US Pollution Prevention Act of 1990 [2] focused regulatory attention on the need to reduce environmental pollution and acknowledged that waste prevention at source not only eliminates the cost of waste remediation but also strengthens economic competitiveness through more efficient use of raw materials. It caused a fundamental shift in the strategy for environmental protection—from end of pipe waste treatment to waste prevention—leading to the emergence, in the early 1990s, of the term Green Chemistry at the US Environmental Protection Agency [3].

Green chemistry focuses on waste prevention rather than waste remediation. That is, Green chemistry efficiently utilizes (preferably renewable) raw materials, eliminates waste and avoids the use of toxic and/or hazardous reagents and solvents in the manufacture and application of chemical products. The overall guiding principle is benign by design. Green chemistry gained formal recognition with the publication of the 12 principles of green chemistry (Table 1.1), by Anastas and Warner in 1998 [4]:

Table 1.1

The Brundtland report, Our Common Future, published in 1987 by the World Commission on Environment and Development, launched the phrase sustainable development [5]. While acknowledging the need for industrial and societal development to provide a growing global population with a satisfactory quality of life, it emphasized that such a development must be sustainable over time. Sustainable development was defined as "development that meets the needs of the present generation without compromising the ability of future generations to meet their own needs." According to Graedel [6], to be sustainable, a technology must fulfill two conditions: (i) natural resources should be used at rates that do not unacceptably deplete supplies over the long term and (ii) residues should be generated at rates no higher than can be assimilated readily by the natural environment. It is abundantly clear that an economy based on nonrenewable fossil resources—oil, coal and natural gas—consumes natural resources at a much higher rate than they are produced in Nature and that carbon dioxide is being generated at a much higher rate than can be assimilated by the natural environment, leading to undesirable climate change.

A balance has to be found between societal equity, environmental impact, and economic development, often referred to as the three Ps—people, planet, and profit—or the triple bottom line. Sustainable development, in contrast to green chemistry, comprises an economic component. A technology will only be sustainable in the long term if it is cost-effective. Increased interest in green and sustainable growth has focused attention on resource efficiency and is driving the transition from a take-make-use-dispose economy based on a linear flow of materials, to a greener, more sustainable circular economy [7].

1.2 The role of catalysis

Most of the waste produced in the industrial production of organic compounds, for example, pharmaceuticals, consists of inorganic salts, a consequence of the use of stoichiometric inorganic reagents. As mentioned earlier, this primarily involves oxidants, such as Cr(VI) reagents, permanganate, and manganese dioxide, and reductants such as metal hydrides, in addition to stoichiometric amounts of mineral and Lewis acids, and bases such as NaOH and KOH. Hence, the generation of copious amounts of waste can be alleviated by the widespread replacement of stoichiometric reagents with catalytic alternatives [8,9]. This applies to all segments of the chemical industry.

A pertinent example from the bulk chemicals arena is provided by propylene oxide with an annual production of 9–10 mio tonnes. Traditionally this involved the reaction of propylene with hypochlorous acid (essentially chlorine in water). For every kg of propylene oxide ca. 3 kg of CaCl2 is produced (Fig. 1.1). One could say that it is a process to make calcium chloride with propylene oxide as the byproduct. More recently, this process has been largely superseded by greener alternatives involving catalytic oxidations with alkyl hydroperoxides or, in the HPPO process, hydrogen peroxide [10]. Combination with the direct production of hydrogen peroxide by catalytic reaction of hydrogen with oxygen [11] affords a process for the production of propylene oxide from propylene, hydrogen and oxygen with coproduction of one equivalent of water.

Figure 1.1 Manufacture of propylene oxide.

Another regulatory change, with far-reaching technological consequences for the pharmaceutical industry, was implemented at roughly the same time as the Pollution Prevention Act. In 1989, the FDA passed legislation that required pharmaceutical companies to market chiral drugs as the pharmacologically active enantiomer, or prove that the wrong enantiomer exhibited no detrimental side-effects [12]. This regulatory measure was long overdue [13]. It meant that the wrong enantiomer should be considered as waste which should be recycled or, preferably, not formed in the first place. It created a pressing need for clean, cost-effective, that is catalytic methods for the synthesis of pure enantiomers and led to the development of, for example, catalytic asymmetric hydrogenation. Subsequently, as a result of modern advances in molecular biology, in particular (meta)genome sequencing and directed evolution through protein engineering, biocatalysis emerged as an environmentally attractive and commercially viable technology for the highly enantioselective synthesis of pure enantiomers.

1.3 Advantages and limitations of biocatalysis

Biocatalysis conforms to 10 of the 12 principles of green chemistry [14] and has clear economic and environmental benefits:

• Enzymes are derived from readily available and inexpensive renewable resources and are biocompatible and biodegradable.

• Enzyme costs are essentially stable and predictable in contrast with the disruptive price fluctuations observed with scarce precious metals and the serious environmental costs associated with their mining.

• Costly removal of traces of noble metals from end products, to the very low levels demanded by regulatory authorities, is avoided.

• Enzymatic reactions are performed under mild conditions (near ambient temperature and atmospheric pressure) in water in standard multipurpose reactors.

• Enzymatic methods proceed without the need for functional-group activation or convoluted protection and deprotection protocols and are, hence, more step economic [15], generate less waste and are more cost-effective than conventional organic syntheses.

• Enzymes generally exhibit high chemo-, regio- and stereo- selectivities that are difficult to achieve with chemo-catalytic methods. In particular, the (near) perfect enantioselectivities observed with highly engineered enzymes are practically inimitable [16].

• Enzymatic reactions, in contrast with many chemo-catalytic reactions, usually involve roughly the same temperature and pressure, thus facilitating integration of multiple steps into cost-effective and environmentally friendly one-pot processes [17]. Telescoping multistep processes into one-pot cascades affords several economic and environmental benefits: fewer unit operations, minimized solvent and reactor volume, higher throughputs and space time yields and reduced waste [18] Moreover, it circumvents the need for wasteful and costly separation and purification of intermediates. Additionally, coupling of enzymatic steps can drive equilibria toward product formation, thus enhancing overall yields.

Biocatalysis can involve metabolically active and growing cells, that is fermentations, or isolated enzymes or metabolically inactive cells where generation of the enzyme and the biotransformation are uncoupled. In this case, deactivated enzymes have to be replaced with a fresh batch and cofactors have to be independently regenerated. Nonetheless, the significant investments and long lead times associated with the development of fermentation processes, and the preference of organic chemists for catalysts that can be stored, motivated the adoption of isolated enzymes as the preferred biocatalysts for the pharmaceutical industry [19].

An early example of replacing conventional chemistry with biocatalysis, that is green chemistry avant la lettre, dates from the late 1980s. It concerns the manufacture of 6-aminopenicillanic acid (6-APA), a key raw material in the industrial synthesis of semisynthetic penicillin and cephalosporin antibiotics, with an annual production of more than 20,000 tonnes, from penicillin G (Pen G). The latter is produced by fermentation. Until the late 1980s a conventional chemical procedure was used for the selective deacylation of Pen G, which is not a trivial reaction as it involves competition between a secondary and a more reactive tertiary amide function. It was accomplished in a one-pot process by protecting the carboxyl function with a trimethylsilyl group followed by transforming the less reactive secondary amide into an imine chloride by reaction with phosphorus pentachloride at −40°C. Subsequent reaction with n-butanol at −40°C afforded an enol ether, which was hydrolyzed to 6-APA at room temperature (Fig. 1.2). The process involved the use of environmentally unattractive reagents in dichloromethane: 0.6 kg of Me3SiCl, 1.2 kg PCl5, 1.6 kg PhNMe2, 0.2 kg NH3 and 8,41 kg of n-BuOH were used to produce 1 kg of 6-APA.

Figure 1.2 Enzymatic versus chemical deacylation of penicillin G.

The overall process is the epitome of what is wrong with many conventional organic syntheses. It is a multistep process involving protection, deprotection and functional-group activation steps. These involve stoichiometric amounts of environmentally unattractive reagents and suspect solvents, under energy intensive conditions, to produce, in addition to the desired product, copious quantities of toxic waste. This process, developed and commercialized by the Gist-Brocades company, is often referred to as the Delft Cleavage.

In stark contrast, enzymatic hydrolysis of Pen G is a one-step process, requiring no activation or (de)protection steps and conducted in water at 37°C. The only reagent involved is 0.9 kg NH3 per kg 6-APA to adjust the pH [20,21]. The enzyme, penicillin G amidohydrolase (E.C. 3.5.1.11), otherwise known as Pen G acylase, was already known and used in the early 1960s [22]. However, the production of the enzyme was very expensive, and it had poor thermal stability, low volumetric productivity, and was discarded after a single use. Hence, 6-APA was produced using the chemical procedure for the next two decades. However, Pen G acylases with improved stability were developed by screening, and cost-effective production was eventually achieved using recombinant DNA technology. Moreover, effective immobilization of the enzyme enabled multiple recycling, up to 1000 recycles [23] of the enzyme and ensuing dramatic reductions in the enzyme cost contribution [24].

1.4 The metrics of waste minimization

As Lord Kelvin (1824–1907) so wisely observed: If you can’t measure it you can’t improve it. Since a primary goal of green chemistry is to reduce or, preferably, eliminate waste generation in the production and application of chemicals, it is essential to have metrics for measuring the mass efficiency of chemical processes. The oldest two mass efficiency metrics—atom economy (AE) [25] and the E-factor [1]—dating from 30 years ago, are the most accepted and widely used.

1.5 Atom economy: every atom counts

AE is the molecular weight of the product divided by the sum of the molecular weights of the starting materials. Calculation of the AE assumes the use of stoichiometric amounts of starting materials and a 100% chemical yield. It doesn’t take auxiliary reagents and solvents, not occurring in the stoichiometric equation, into account but it is very useful for comparing different routes to a target molecule before any experiments are performed. AEs of multistep processes are derived from the overall stoichiometry and are not simply the addition product of the AEs of the individual steps.

1.6 The E-factor: the environmental footprint of chemicals

In contrast, the E-factor is the actual amount of waste produced per kg of product, succinctly defined as everything but the desired product and including solvent losses and chemicals used in work-up. E-factors of individual steps are additive and are readily calculated for single- or multi-step processes. The E-factor is essentially the environmental footprint of a process, the higher the E-factor the greater the environmental impact. Lower E-factors translate to a more efficient utilization of raw materials and show a strong positive correlation with reduced manufacturing and waste disposal costs [26,27]. The ideal E-factor is zero in line with the first principle of Green Chemistry: It is better to prevent waste than to treat or clean up waste after it is formed. Using the E-factor places emphasis firmly on designing cleaner, waste-free processes, and the ideal E-Factor of 0 clearly reflects the ultimate goal of zero waste manufacturing. Typical examples of high AE, low E-factor catalytic processes are ketone hydrogenations, alcohol carbonylation, aerobic oxidation of alcohols, and olefin hydroformylation (Fig. 1.3).

Figure 1.3 High atom economy catalytic processes.

The publication, in 1992, of the average E-factors of processes in various segments of the chemical industry (Table 1.2), from oil refining to pharmaceuticals, was a turning point [1]. It provided an important challenge to the industry, particularly the Fine Chemicals and Pharmaceuticals segments, to reduce the copious amounts of waste generated in their manufacturing processes. The Pharmaceutical industry accepted the challenge and has spent the last 2–3 decades cleaning up their manufacturing operations [28].

Table 1.2

The E-factor basically includes all reagents used, in both the process and the down-stream processing. We assumed that solvents would be recycled and if data were not known we assumed that 90% would be recovered and, hence, only 10% of the solvent used was counted as waste. In hindsight this was too optimistic. The penchant of organic chemists for optimizing the particular solvent used in individual steps in a multistep synthesis leads to cross-contamination and difficulties in solvent recycling.

We excluded water based on the argument that inclusion would create a skewing of E-factors. However, disposal or reuse of processed water will inevitably involve some sort of pretreatment, and the current method is to calculate and compare E-factors both including and excluding water [29,30]. This led to the introduction of simple E-factors (sEF), that disregard solvents and water for use in early route scouting, and complete E-factors (cEF) that include solvents and water assuming no recycling [26]. The true commercial E-factor will lie between the sEF and cEF and can be calculated when reliable data for recycling and solvent losses are known.

Inclusion of the energy requirements of a process in the E-factor was always implicit as energy consumption generates waste, which can be accounted for as carbon dioxide equivalents. In practice fine chemicals and pharmaceuticals are generally produced in multipurpose facilities and energy consumption is not allocated to particular processes. This makes assigning waste derived from energy to individual products difficult. In contrast, bulk chemicals are produced in dedicated units and energy forms an important part of total resources used. To improve the energy accounting of processes, the E+ factor, which considers the greenhouse gas emissions generated from electricity used for unit operations such as cooling, heating, stirring and pumping, was recently proposed [31].

The strength of the E-factor is its simplicity—in terms of both concept and use—and, because it was introduced 30 years ago, it is familiar to many and is widely used. Other mass-based green metrics have been proposed and they can be divided into two types: those constituting a percentage of the ideal analogous to AE and those expressed as kg/kg analogous to the E-factor. An example of the former is reaction mass efficiency [32], a refinement of AE taking reaction yield and excess of reagents into account. An example of the latter type is process mass intensity (PMI) which is the mass ratio of the total input of materials (including solvents and water) to product [33]. However, such metrics have not reached the same broad acceptance as the E-factor [34,35], and their use is largely confined to small-molecule pharmaceuticals [26]. The E-factor has been widely adopted in chemistry courses at undergraduate and postgraduate level [36–41] and high-school chemistry classes [42] and consistently used as a yardstick for assessing the greenness of competing synthetic routes to various chemicals [43–48].

1.7 Intrinsic E-factors and system boundaries

Traditionally, the E-factor is based on gate-to-gate system boundaries, that is the process conducted at the manufacturing site, rather than cradle-to-grave. This means that the E-factor is dependent on the starting point of the synthesis and the E-factor of a multistep synthesis can be significantly reduced overnight by farming out an early intermediate instead of making it in-house. This means that it is necessary to agree on a definition of starting material. For example, in pharmaceuticals manufacture this has been defined as being readily available at a price of < $100 per kg from a reputable commercial supplier [24]. Since E-factors are additive (in contrast to PMIs), the intrinsic E-factor for the ASM synthesis may simply be added to the main synthesis E-factor to obtain an unbiased E-factor value for a complete synthetic pathway.

For example, Dunn and coworkers at Pfizer [49] calculated the E-factor of the commercial process for the production of sildenafil citrate (Viagra). The traditional E-factor, excluding water and including 10% of the mass of solvents used, was found to be 6.4 and the sEF (excluding solvents and water) and cEF were 3.9 and 50.3, respectively. However, one of the starting materials did not meet the requirement of being commercially available at a price of < $100 per mole, and its inclusion led to significant increases: the sEF increased from 3.9 to 9.9, the cEF from 50.3 to 85.5, and the E-factor from 6.4 to 13.8. Hence, an agreement is required on an industry-wide starting point concept for assessing process greenness.

1.8 The climate factor

Thirty years ago, the number one environmental problem was ozone-depleting chemicals in the atmosphere. Thirty years later greenhouse gases, in particular carbon dioxide, have become the number one priority for climate change mitigation. This is motivating the transition from an economy based on fossil resources to a bio-based economy based on renewable energy and raw materials, which is manifest in the defossilization of chemicals manufacture.

Christensen and coworkers [50] proposed the use of the climate factor, defined as the total mass of CO2 emitted divided by the mass of product formed (kg CO2/kg product), to compare the CO2 burdens of different processes. It is the sum of kg CO2 emitted in the production of the raw material(s) and conversion of the latter to the product(s). It is useful for comparing biomass- versus fossil resource-based processes [51] but system boundaries will need to be defined and accepted. It should not be confused with carbon economy which is the % of carbon in a molecule that is derived from renewable resources. C factor is to carbon economy as the E-factor is to AE.

1.9 The nature and environmental impact of wastes

Not only the amount of waste generated but also the environmental impact of that waste is of paramount importance. The E-factor assigns the same weighting to all types of waste and the E-factor must, therefore, be considered in conjunction with other metrics [52]. We recognized this when we introduced the E-factor thirty years ago and proposed [53] the environmental quotient where Q is an unfriendliness multiplier representing the nature of the waste. The problem then becomes how to quantify Q. A decade later, Eissen and Metzger [54] developed the simple and easy-to-use Environmental Assessment Tool for Organic Synthesis software to assess the potential environmental impact, that is the Q value, of waste by assigning penalty points based on human and eco-toxicity.

Graedel [55] noted, in 1999, that adding a life-cycle perspective to green chemistry enlarges its scope and enhances its environmental benefits. However, conducting a full scale cradle-to-grave Life Cycle Assessment (LCA) in the design or development phase of a process is generally too difficult and time consuming. Nonetheless, many groups [33,56–59] have addressed the problem of integrating mass efficiency with LCA-based assessment of the environmental impact of waste. However, the application of the LCA methodology to fine chemicals and pharmaceuticals is a challenge owing to the paucity of life cycle inventory data. Consequently, many chemical companies have adopted their own customized metrics, for example, BASF’s eco-efficiency [60].

Pharmaceutical and Flavor and Fragrance companies have been particularly active in this respect. For example, GSK developed FLASC (Fast Life Cycle Assessment of Synthetic Chemistry) [61] and the cosmetic ingredients producer Chimex, a subsidiary of L’Oreal, introduced Eco-footprint which covers the supply chain from the supplier’s gate to the product leaving the production facilities [62]. It comprises a manufacturing footprint and an eco-design footprint. The former is based on five indicators: water footprint, carbon footprint of the transportation of raw materials to the production site, aqueous waste and used solvents valorization and energy consumption. The eco-design footprint comprises the E-factor, synthetic pathway efficiency, raw materials of renewable origin, and environmental impacts of raw materials and waste.

Similarly, Green Motion, a gate-to-gate green metric tool for evaluating the efficiency and health, safety, and environmental impacts of manufacturing processes, was developed by flavor and fragrance company, Mane [63]. Their starting point was to group the 12 Principles of Green Chemistry into seven fundamental concepts: raw material, solvent selection, hazard and toxicity of reagents, reaction efficiency, process efficiency, hazard and toxicity of final product and waste generation. Penalty points are then allocated within each category based on well-defined criteria, such as origin of raw materials (renewable or synthetic), yield, number of steps and solvents involved, and amount of waste as expressed by the E-factor. The latter was favored over PMI because it fitted better with the objective of zero points for an E-factor of zero. For a given criterion, the higher the impact on health, safety and environment, the higher the number of penalty points. Hazard and toxicity ratings are based on the hierarchy of GHS pictograms used on labels and safety data sheets that denote various hazards and toxicity.

Each process is assessed by means of a questionnaire requiring simple yes/no answers, pictograms, numerical values or a selection of multiple choice options that then assigns penalty points based on the answers. Deduction of the penalty points from 100 affords an overall score, meaning the higher the score the more sustainable and the lower the environmental impact of the process. The method was used to rate more than a thousand products in 12 months, thus building a useful database for benchmarking, and each new product which was scaled to production level was rated. The authors concluded that Green Motion is a simple and quantitative method that is well suited to the evaluation of flavor and fragrance ingredients, noting that a full assessment can be made in only half an hour.

We recently used the E-factor and other metrics in selecting a route for the synthesis of a key intermediate for HIV protease inhibitors [64]. The inclusion of so-called intrinsic E-factors to account for the synthesis of advanced starting materials was essential for comparison on a level playing field. Green motion was the most useful and conceptually simple method for comparing the three routes. Radial polygon is a useful visual tool that provides an overview of multivariable performance indicators [65]. An ideal green synthesis corresponds to a regular polygon, while distortions toward the center identify weak points in a synthesis and provide guidance for optimization.

1.10 The role of solvents: the medium is the message

In 1992 we remarked that "So many of the solvents favored by organic chemists are now on the black list that the whole question of solvents in organic synthesis requires rethinking. Not only do organic chemists generally use too much solvent, they very often choose the wrong ones. In the first place, is a solvent really necessary? If a solvent (diluent) is needed it should preferably be water" [1]. The last two decades have been devoted to rethinking the question of solvents in chemical processes [66]. The best solvent is no solvent. Reactions can be performed with neat liquid substrates, for example, in biodiesel production from triglycerides [67].

Solvents constitute 80%–90% of the total mass of nonaqueous material used in pharmaceutical manufacture and account for the majority of waste formed and 75%–80% of the environmental life cycle impacts [68]. A survey of solvent usage, described in 388 publications in the period 1997–2012, revealed much room for improvement across the global pharmaceutical industry [69].

The eight most commonly used solvents of concern were three chlorinated hydrocarbons (dichloromethane, 1,2-dichloroethane, chloroform), four ethers (diethyl ether, diisopropyl ether, 1,2-dimethoxyethane and 1,4-dioxane) and one hydrocarbon (n-hexane). Many of these solvents were still being selected despite the availability of greener alternatives. The clear need for the greening of organic solvents encouraged pharmaceutical companies to develop Solvent Selection Guides (SSGs) to stimulate replacement of environmentally undesirable solvents, in particular chlorinated hydrocarbons [70]. Traffic-light inspired color coding was used, whereby green, amber and red signify preferred, usable and undesirable solvents, respectively, [71–75]. GSK was the first to incorporate LCA into solvent selection [76]. Solvents were assigned a score of 1–10 in four categories, waste disposal, environmental impact, health and safety (EHS) issues, each of which consisted of multiple subcategories. Each solvent is given a composite score corresponding to the geometric mean of the four scores: <3.5 is red, 3.5–<7.5 is amber and 7.5–10 is green.

The concept of scores based on EHS statements was adopted by the ACS Green Chemistry Institute’s Pharmaceutical Round Table consortium [77] and the European collaborative research project known as CHEM21 [78]. 51 solvents were ranked and placed in four categories: recommended, problematic, hazardous and highly hazardous. In contrast with the GSK method, greener solvents have a lower score in the CHEM21 SSG. As Welton [79] remarked, appropriate solvent selection can improve both the environmental and commercial performance of a process and, therefore, its overall sustainability. Hence, the quest for improved solvents continues, for example, aided by in silico machine learning methods [80]. In addition to the need for replacing chlorinated solvents there is a pressing need to replace many hydrocarbons and ethers that are classed as hazardous or highly hazardous on the basis of toxicity and/or flammability issues. In addition, the four most commonly used dipolar aprotic solvents—dimethyl formamide, dimethylacetamide, N-methylpyrrolidone and sulfolane—are reproductive toxicity hazards the first three of which are classified as substances of very high concern.

1.11 Bio-based solvents

A variety of mono- and bi-phasic solvent systems have been proposed as alternatives for volatile organic solvents with undesirable ecological properties [66,81,82]. Various less common organic solvents, many of which are derived from renewable feedstocks [83], have also been ranked in the CHEM21 project as recommended or problematic [78]. These include a variety of bio-based alcohols, such as bio-ethanol [84], isobutanol and glycerol [85], bio-based ethers such as 2-methyltetrahydrofuran (Me-THF) [86], dimethyl tetrahydrofuran (diMeTHF), cyclopentyl methyl ether (CPME), dimethylsorbide, cyrene (dihydrolevoglucosenon) [87] and the cygnet [88] family of ethers, and bio-based esters such as isobutyl acetate, ethyl lactate, ethyl levulinate, diethyl carbonate, glycerol carbonate and γ-valerolactone [89] (see Fig. 1.4 for structures). Interestingly, lipases dissolve in glycerol and glycerol carbonate [90] with retention of activity. Bio-based ethers, such as Me-THF, CPME and cyrene [91], are also useful solvents for biocatalysis [92].

Figure 1.4 Bio-based solvents.

1.12 Water as a reaction medium

Significant reductions in waste produced in chemicals manufacture have been achieved by replacing archaic stoichiometric methodologies with atom efficient, low E-factor, catalytic alternatives, notably hydrogenation, hydroformylation, carbonylation and oxidation [34]. However, inefficiencies associated with the containment, recovery, and reuse of solvents still remain. Alternatives for classical organic solvents should, therefore, not only have acceptable EHS properties. Moreover, both the catalyst and the solvent should be readily recovered and recycled to afford an environmentally acceptable and cost-effective process.

Water has much to offer: it is nontoxic, noninflammable, abundantly available and inexpensive and can be used in catalytic processes by employing water soluble catalysts. Indeed, in Nature many reactions are conducted with both water miscible and immiscible substrates in aqueous media. However, when water is used as the reaction medium, after the removal of the product(s) and catalysts, the remaining water will be contaminated with small amounts of organics and hence need to undergo some sort of treatment before it can be discharged to effluent treatment facilities. This problem is often aggravated by using organic solvents to extract the product from the water [93]. Another problem is that water has the highest heat capacity of all liquids. While this property is very useful for maintaining life on earth it is a serious disadvantage for chemical processes.

Nonetheless, there is a noticeable trend toward the use of water as a reaction medium, and with remarkable results. For example, Bailey and coworkers [94] at Takeda Pharmaceutical developed a five step synthesis of an investigational 5-HT4 receptor conducted almost exclusively in an aqueous medium with dramatic improvements in process efficiency compared with the first generation process in organic solvents. The overall yield was improved from 35% to 56%, the overall PMI from 350 to 79, and the solvent intensity from 223 to 14, representing a 94% reduction in organic solvent use. Even more remarkable was the reduction in water intensity from 106 to 55, that is the process in water used 48% less water than the organic solvent-based process. Both processes delivered the API with an overall purity of ca. 99.8%.

1.13 Aqueous biphasic catalysis

Catalytic reactions can be performed in water by employing water soluble catalysts. When the substrate is immiscible with water the reactions are performed as aqueous biphasic catalysis whereby the catalyst is dissolved in the water phase and the substrate in a separate organic phase. The reaction takes place in the aqueous phase which means that the substrate needs to be slightly soluble in water. The product is recovered by phase separation and the water phase, containing the catalyst, can remain in the reactor and be recycled. All of the atom economical catalytic processes discussed earlier can be performed as aqueous biphasic catalytic processes. The most well-known process is probably the Ruhr-Chemie/Rhone Poulenc process for propylene hydroformylation employing a water-soluble Rh (I) tppts complex (Fig. 1.5) as the catalyst [95]. This process works well because propylene is slightly soluble in water. The principle is also readily translated to other noble metal catalyzed processes, such as hydrogenation, carbonylation, olefin metathesis and aerobic oxidations of alcohols [96,97].

Figure 1.5 The RuhrChemie/Rhone Poulenc hydroformylation process.

1.14 Surfactants in water: aqueous micelles as nanoreactors

The rates of the biphasic Rh/tppts catalyzed hydroformylations of olefins decrease dramatically with increasing molecular weight of the olefin, for example, the practically water insoluble 1-tetradecene afforded only traces of aldehydes [97]. The envisaged approach to circumvent this problem was to use tenside phosphines that were specifically designed to combine the functions of a ligand and an amphiphile (surfactant) to create micelles in which the water insoluble substrate is solubilized. Although the concept was sound it did not lead to broad application.

Lipshutz and coworkers [98] used a different strategy that took the concept to a new level of sophistication and has proven more successful. They used small amounts (e.g., 2 wt.%) of designer amphiphiles above their critical micelle concentration (ca. 10−4 M) that spontaneously self-assemble in water to form nanomicelles with lipophilic interiors in which reactions can occur between water-insoluble substrates and catalysts. The choice of amphiphile is critical as it determines the size, shape and internal lipophilicity of the nano-micellar reactors.

Amphiphiles, PTS and TPGS-750-M (see Fig. 1.6 for structures), derived from vitamin E, for example, were used to perform a variety of noble metal catalyzed reactions [99], [100–102], such as hydrogenation [103] and palladium catalyzed C-C coupling reactions [104] at room temperature in organic solvent free water. The product was recovered by extraction with a minimum amount of an environmentally acceptable solvent, such as ethyl acetate, or precipitated as an insoluble solid with the amphiphile remaining in the water phase. However, it will be necessary to remove traces of amphiphiles, such as TPGS-750-M, remaining in the aqueous effluent [105].

Figure 1.6 Structures of designer amphiphiles.

Water is an excellent solvent for conducting enzymatic reactions and the product can be extracted into an environmentally acceptable solvent, such as ethyl acetate. Alternatively, the extraction can be back-integrated into the reaction step by conducting the process as aqueous biphasic catalysis. The company Codexis, for example, developed cost-effective biocatalytic processes to a variety of pharmaceutical intermediates using highly engineered enzymes in this way [106].

Alternatively, enzymatic reactions of hydrophobic substrates in water could be facilitated by designer amphiphiles. For example, ketoreductases (KREDs) were fully compatible with aqueous solutions containing micelles derived from TPGS-750-M [107]. This enabled the one-pot production of chiral secondary alcohols in high enantioselectivities through a combination of noble-metal catalyzed synthesis of ketones and subsequent enantioselective KRED catalyzed reductions (Fig. 1.7A). Moreover, the activity of the KRED was increased in the presence of the amphiphile, compared to in buffer alone. This was attributed to a reservoir effect in which the micelles control the supply of both substrate and product to the active site of the enzyme, thereby limiting substrate and product inhibition.

Figure 1.7 One-pot chemo-enzymatic conversions in water containing designer surfactants.

Hastings and coworkers [108] similarly reported one-pot combinations of chemo- and biocatalysis, involving Heck coupling and olefin metathesis reactions with enzymatic hydrolysis catalyzed by CaLB and pig liver esterase in water containing 2 wt.% of TPGS-750-M as the surfactant (Fig. 1.7B and C).

1.15 Neoteric solvents: ionic liquids and deep eutectic solvents

The last decade has witnessed the emergence of the use of neoteric solvents—ionic liquids (ILs) and Deep Eutectic Solvents (DESs), alone or together with water—as reaction media for conducting biocatalytic reactions. Following initial reports in 2000, biocatalysis in ILs, including protic ILs, has been widely studied and is the subject of numerous reviews [109–117]. The introduction of DESs, as novel reaction media for biocatalysis, was a decade later than that of ILs, but they have also been widely used as reaction media for biocatalysis with both whole cells and isolated enzymes [118–121]. In particular, natural deep eutectic solvents, prepared from combinations of relatively simple, primary metabolites, including sugars, amino acids and organic acids, such as citric, itaconic, malic, lactic and succinic acids, are potentially interesting green solvents for conducting biocatalytic processes. They play the role of reaction media in the intracellular synthesis of secondary metabolites, such as flavonoids and steroids, that are sparingly soluble in water [122] and have been called Solvents for the 21st century [123].

1.16 Concluding remarks

To be green and sustainable, a process must generate minimum waste, avoid the use of toxic and hazardous reagents and solvents, and efficiently utilize (preferably) renewable raw materials. Biocatalysis is green and sustainable, and biocatalysts perform admirably in water as solvent. However, this can lead to problems with hydrophobic substrates owing to their low water solubility. As discussed in this chapter, various approaches are possible for solving this problem. Moreover, the ongoing transition from a linear, fossil resource-based economy to a circular bio-based economy based on renewable raw materials will facilitate further widespread application of biocatalytic processes.

References

1. Sheldon RA. Organic synthesis: past, present and future. Chem Ind. 1992;903–906 (London).

2. http://www.epa.gov/laws-regulations/summary-pollution-prevention-act (accessed 23.05.21).

3. Amato I. The slow birth of green chemistry. Science. 1993;259:1538–1541.

4. Anastas P, Warner JC. Green Chemistry: Theory and Practice Oxford: Oxford University Press; 1998.

5. Brundtland CG. Our Common Future, The World Commission on Environmental Development Oxford: Oxford University Press; 1987.

6. Graedel TE. Green chemistry and sustainable development. In: Clark J, Macquarrie DJ, eds. Handbook of Green Chemistry and Technology. New York: Wiley; 2002;56–61.

7. Clark JH, Farmer TJ, Herrero-Davila L, Sherwood J. Circular economy design considerations for research and development in the chemical sciences. Green Chem. 2016;18:3914–3934.

8. Sheldon RA. Catalysis and pollution prevention. Chem Ind. 1997;12–15.

9. Sheldon RA. Catalysis: the key to waste minimization. J Chem Tech Biotechnol. 1997;68:381–388.

10. Russo V, Tesser R, Santacesaria E, Di Serio M. Kinetics of propene oxide production via hydrogen peroxide with TS-1. Ind Eng Chem Res. 2014;53(15):6274–6287.

11. Ranganathan S, Sieber V. Recent advances in the direct synthesis of hydrogen peroxide using chemical catalysis-a review. Catalysts 2018; article 379.

12. De Camp W. The FDA perspective on the development of stereoisomers. Chirality. 1989;1:2–6.

13. Ariëns EJ. Stereochemistry, a basis for sophisticated nonsense in pharmacokinetics and clinical pharmacology. Eur J Clin Pharmacol. 1984;26:663–668.

14. Sheldon RA, Woodley JM. Role of biocatalysis in sustainable chemistry. Chem Rev. 2018;118:801–838.

15. Wender PA. Toward the ideal synthesis and molecular function through synthesis-informed design. Nat Prod Rep. 2014;31:433–440.

16. Tucker JL, Faul MM. Industrial research: drug companies must adopt green chemistry. Nature. 2016;534:27–29.

17. Sheldon RA. Enzyme-catalyzed cascade reactions. In: Eduardo Garcia-Junceda, ed. Chapter 6 in Multi-Step Enzyme Catalysis: Biotransformations and Chemoenzymatic Synthesis. Weinheim: Wiley-VCH; 2008;109–135.

18. Bruggink A, Schoevaart R, Kieboom T. Concepts of nature in organic synthsis: cascade catalysis and multistep conversions in comcert. Org Proc Res Dev. 2003;7:622–640.

19. Truppo MD. Biocatalysis in the pharmaceutical industry: the need for speed. ACS Med Chem Lett. 2017;8:476–480.

20. Bruggink A, Roos EC, de Vroom E. Penicillin acylase in the industrial production of β-lactam antibiotics. Org Proc Res Dev. 1998;2:128–133.

21. Wegman MA, Janssen MHA, van Rantwijk F, Sheldon RA. Toward biocatalytic synthesis of β-lactam antibiotics. Adv Synth Catal. 2001;343:559–576.

22. Rolinson GN, Batchelor FR, Butterworth D, et al. Formation of 6-aminopenicillanic acid from penicillin by enzymatic hydrolysis. Nature. 1960;187:236–237.

23. Basso A, Serban S. Industrial applications immobilized enzymes—a review. Mol Catal. 2019;479:1–20 article 110607.

24. Parmar A, Kumar H, Marwaha SS, Kennedy JF. Advances in enzymatic transformation of penicillins to 6-aminopenicillanic acid (6-APA). Biotechnol Adv. 2000;18:289–301.

25. Trost BM. The atom economy: a search for synthetic efficiency. Science. 1991;254:1471–1477.

26. Roschangar F, Sheldon RA, Senanayake CH. Overcoming barriers to green chemistry in the pharmaceutical industry—the green aspiration level concept. Green Chem. 2015;17:752–768.

27. Hayashi Y. Pot economy and one-pot synthesis. Chem Sci. 2016;7:866–880.

28. Cue BW, Chemistry Green. Strategies for medicinal chemists. In: Zhang W, Cue BW, eds. Green Techniques for Organic Synthesis and Medicinal Chemistry. Chichester (UK): Wiley; 2012;553–569.

29. Ma SK, Gruber J, Davis C, et al. A green-by-design biocatalytic process for atorvastatin intermediate. Green Chem. 2010;12:81–86.

30. Constable DJC, Curzons AD, Cunningham VL. Metrics to green chemistry-which are the best?. Green Chem. 2002;4:521–527.

31. Tieves F, Tonin F, Fernandez-Fueyo E, et al. Energising the E-factor: The E+-factor. Tetrahedron. 2019;75:1311–1314.

32. Curzons AD, Constable DJC, Mortimer DN, Cunningham VL. So you think your process is green, how do you know?- using principles of sustainability to determine what is green—a corporate perspective. Green Chem. 2001;3:1–6.

33. Jiménez-González C, Ponder CS, Broxterman QB, Manley J. Using the right green yardstick: why process mass intensity is used in the pharmaceutical industry to drive more sustainable processes. Org Process Res Dev. 2011;15:912–917.

34. Sheldon RA. The E factor 25 years on: the rise of green chemistry and sustainability. Green Chem. 2017;19:18–43.

35. Sheldon RA. Metrics of green chemistry and sustainability: past, present, future. ACS Sust Chem Eng. 2018;6:32–48.

36. Etzkorn FA. Green Chemistry: Principles and Case Studies Cambridge: Royal Society of Chemistry; 2020.

37. Andraos J, Sayed M. On the use of green metrics in the undergraduate organic chemistry lecture and lab to assess the mass efficiency of organic reactions. J Chem Educ. 2007;84:1004–1010.

38. Mooney M, Vreugdenhil AJ, Shetranjiwalla S. A toolkit of green chemistry and life-cycle analysis for comparative assessment in undergraduate organic chemistry experiments: synthesis of (E)-stilbene. J Chem Educ. 2020;97:1336–1344.

39. Hudson R, Leaman D, Kawamura KE, et al. Exploring green chemistry metrics with interlocking building block molecular models. J Chem Educ. 2016;93:691–694.

40. Lam CH, Escande V, Mellor KE, Zimmerman JB, Anastas PT. Teaching atom economy and E-factor concepts through a green laboratory experiment: aerobic oxidative cleavage of meso-hydrobenzoin to benzaldehyde using a heterogeneous catalyst. J Chem Educ. 2019;96:761–765.

41. Fennie MW, Roth JM. Comparing amide-forming reactions using green chemistry metrics in an undergraduate organic laboratory. J Chem Educ. 2016;93:1788–1793.

42. See: https://resources4rethinking.ca/en/resource/what-is-green-chemistry https://www.beyondbenign.org/lessons/e-factor/ (accessed 25.09.21).

43. van Beurden K, de Koning S, Molendijk D, van Schijndel J. The Knoevenagel reaction: a review of the unfinished treasure map to forming carbon–carbon bonds. Green Chem Lett Rev. 2020;13:349–364.

44. Bédard A-C, Longstreet AR, Britton J, et al. Minimizing E-factor in the continuous-flow synthesis of diazepam and atropine. Bioorg Med Chem. 2017;25:6233–6241.

45. Margalef J, Samec JSM. Assessing methodologies to synthesize α-sulfenylated carbonyl compounds by green chemistry metrics. ChemSusChem.