Biological routes to itaconic and succinic acids

1.1 Succinic acid

Succinic acid (IUPAC systematic name: butanedioic acid; traditionally known as amber acid) is a versatile platform chemical with applications in both high-valueniche personal care and food additives markets and in the large volume production of polyester, polyurethanes, plasticizers, and coatings. At present, succinic acid is predominantly produced from butane through catalytic hydrogenation of petroleum-based maleic anhydride. According to a researchandmarkets projection, the succinic acid market will reach $ 486.7 million by 2019 with a CAGR of 22.6% by volume between 2014 and 2019 [1]. The global succinic acid production capacity is between 30 000 and 50 000 tons at a compound annual growth rate of 18.7% from 2011 to 2016. Based on succinic acid applications, the global succinic acidmarket can be segmented into the following sectors and their respective market share percentages: industrial applications, 57.1%; pharmaceuticals, 15.91%; food & beverages, 13.07%; others, 13.92% (PRNewswire, June 16, 2015). Succinic acid is one of the US Department of Energy’s 12 top value-added chemicals from biomass. The estimated potential global market for succinic acid is expected to reach US $ 7–10 billion per year assuming that the price of succinic acid production will be substantially reduced in the future. Recognizing the potential of bio-based succinic acid, companies interested in manufacturing this C₄ platform chemical include BioAmber Inc., Myriant Corporation, Reverdia (a DSM-Roquette joint venture), Succinity (a BASF-Purac joint venture), Agro-Industrie Recherches et Développements (ARD), Mitsui & Co., China National BlueStar Co. and PTT Public Company Limited. Bio-succinic acid plants around the globe are summarized in Table 1 [2].

Succinic acid is a platform chemical most widely used in food ingredients, as precursors to active pharmaceutical ingredients and pharmaceutical additives and has the potential in industrial applications to replace maleic anhydride and serve as a precursor for the production of 1,4-butanediol (BDO), tertahydrofuran (THF), N-methyl pyrrolidone (NMP) and γ-bytyrolactone (GBL). 1,4-Butanediol can be further carbony-lated to synthesize adipic acid for the manufacture of nylon 6,6, lubricants, foams and food products. BDOis also raw material for the preparation of polybutylene terephtha-late (PBT), a high performance plastic. Succinates can be used as additives to animal feed and as precursors for protein synthesis. Diethyl succinate can be used for paint stripping and ethylene diaminedisuccinate is a replacement for EDTA. Other applications for succinic acid and derivatives thereof include catalysis of food seasoning preparation, cut flower preservation, soil chelation, corrosion inhibition and modification of polyester resin to improve dyeing ability. The largest existing succinic acid market involves its use as a surfactant, detergent extender and foaming agent [3]. At present, most succinic acid for industrial use is produced by petrochemical process and only natural derivatives thereof utilized in the food market are produced by fermentation. Through biological route, succinic acid is produced as an intermediate of the tricarboxylic acid cycle or as an end product of anaerobic metabolism. The leading developers of the field, BioAmber and Reverdia utilize genetically modified yeast whereas Myriant applied E. coli strains for the biosynthesis of succinic acid. Biosuccinic acid is generally considered safe. A list of maximum-level use for succinic acid in desserts, soups and broths, as well as that for starch sodium octenyl succinate in weaning food, can be found in directive 95/2/EC [4]. Bio-succinate has also been used as a flavoring enhancer for low-sodium food and emulsifiers in infant formulas and follow-on formulas. In pharmaceutical applications, succinate is used as an anti-carcinogenic and an insulinotropic agent [4, 5]. There is a wide range of applications for succinic acid and derivatives thereof [3, 6].

Table 1: Global bio-succinic acid plants (planned and in production) [2].

1.2 Itaconic acid

Itaconic acid, 2-methylidenebutanedioic acid, is an unsaturated di-carbonic acid and has been used for the production of acrylic plastics, acrylate latexes, alkyl paints, industrial adhesives, superabsorbents, detergents, and antiscaling agents. At present, synthetic latex is the largest application accounting for 55% of itaconic acid in the global market. The unsaturated vinylidene functionality of itaconic acid renders the bio-based chemical with a reactive site for generating unique molecular structures or producing a polymer with improved UV-resistant characteristics. Itaconic acid is emerging as a prospective bio-based replacement of maleic anhydride for the preparation of unsaturated polyester resins (UPRs) due to similarities between the two chemicals. Recent developments in itaconic acid applications have been in biomedicine, including those in ophthalmology, dentistry and drug delivery [7]. Chemical conversion of citric acid to itaconic acid by controlled pyrolysis/distillation processes has being described [8]. The low price difference between citric and itaconic acids limits the economic efficiency of the chemical routes. Currently, mostmanufacturers are producing itaconic acid with Aspergillus terreus using pretreated molasses under phosphate-limited conditions. Global Industry Analysts, Inc. forecasts the global itaconic acid market to reach US $ 398.3 million by 2017. In particular, the Asia-Pacific region is poised to become the fastest growing itaconic acid market at 9% CAGR, while the United States represents the largest itaconic acid market. The key players in the global market are highly concentrated in China.

2.1 Chemical synthesis of succinic acid

Succinic acid for current industrial applications is predominantly produced by the hydrogenation of petroleum-based maleic acid or maleic anhydride. The raw material, maleic anhydride, is obtained by catalytic oxidation of C₄ hydrocarbons. Petrochemical production of succinic acid is achieved by catalytic hydrogenation of maleic anhydride to prepare succinic anhydride followed by hydration. The resulting solution is then concentrated to enable crystallization of succinic acid at low temperature. The product is recovered by centrifugation and drying. For the synthesis of 1,4-BDO from maleic anhydride, succinic acid is produced as an in situ intermediate. Separation and drying are thus irrelevant. Alternative chemical routes to succinic acid include paraffin oxidation, electrolytic reduction of maleic anhydride in acidic medium, catalytic addition of acetylene and acrylic acid [5, 12]. Electrochemical synthesis has the advantage of high product purity, which is especially useful in food and pharmaceutical applications.

2.2 Biological routes to succinic acid

Current biological routes to produce succinic acidon a commercial scale employ E. coli and yeast strains. As with many biomass-derived chemicals, bio-succinic acid is still disadvantaged in terms of cost competitiveness when compared to its petroleum-based counterparts; feedstock price and the processing cost are key considerations. A broad range of carbon and nitrogen sources have been studied as potential feed-stocks. Molasses, sugar mixtures, and glycerol were evaluated as candidates for carbon sources, while yeast extracts and wheat processing byproducts were used as nitrogen sources. The downstream isolation and purification process are critical to the production cost. It was suggested that the price of succinic acid needs to fall below US $ 0.45 per kilogram to open up commodity markets thereof [3]. According to Wilke’s model, it will require 100% (w/w) yield, a productivity of 3 g/ l/h, and titers up to 250 g/l to achieve the target production cost [13, 14]. A very limited number of succinic acid biosynthesis processes achieve a productivity of greater than 3 g/l/h [13]. Such productivity strongly governs plant capacity and thus affects both variable costs and fixed costs, while yield on feedstock and feedstock price have direct impact on variable costs. Considering future projection quantities and cost, continuous integrated production is likely to outperform batch process for the biosynthesis of succinic acid.

Fermentative production of succinic acid from renewable resources is not only advantageous in terms of cost effectiveness, but also results in CO₂ fixation. However, the biological route requires considerable space and amount of water, as well as long fermentation time and a complicated product isolation process [12]. Fermentative succinate-producing microorganisms can be generally classified into natural (bacteria, fungi) and engineered (bacteria, yeast) species [5, 13]. Natural bacteria species show high tolerance to osmotic pressure caused by high succinate concentration, but their cultivation requires expensive nutrients. Fungal strains, on the other hand, show lower productivity than that of their bacterial counterparts. Notable microorganisms engineered for efficient succinate production are E. coli, Corynebacterium glutamicum, and S. cerevisiae. Strain engineering strategies to enhance succinate productivity include inactivation of branch pathways, overexpression of genes directly involved in branch pathways, redirection of metabolic flux, and the introduction of reducing power. Biosynthesis of succinate can proceed through anaerobic and aerobic conditions. Anaerobic fermentation is preferred over aerobic fermentation because of its low capital investment and operational cost.

Three formation pathways for succinate are the glyoxylate path and the oxidative and the reductive branches of the triarboxylic acid (TCA) cycle [15]. The glyoxylate and the oxidative pathways are active under aerobic conditions whereas the reductive branch of the TCA cycle proceeds under anaerobic conditions. NADH generation in the glyoxylate route alone results in an imbalance of electrons. Activation of the glyoxylate pathway in conjunction with anaerobic fermentation improves succinate yield [16]. The succinate yield and byproduct formation in anaerobic culture are strongly governed by the availability of NADH. It is critical to utilize genetic engineering tools to manipulate metabolic pathways to achieve a suitable intracellular redox balance. During the oxidative branch of the TCA cycle, the conversion of succinate to fumarate should be blocked. The theoretical yield is 1.71 moles of succinate produced per mole of glucose in the presence of CO₂. Such a theoretical yield increases to 2 moles of succinate produced per mole of glucose with electron donor supplements, such as hydrogen [17]. The nature of the fermentation pathway and CO₂/H₂ ratio are critical factors in succinate production yield.

Succinate is one key intermediate of the TCA cycle. Thus, it is possible to synthesize succinic acid by evaluating metabolic pathways leading to other TCA cycle intermediates, such as lactate, acetate, and ethanol, and genetically engineering microorganisms accordingly. Microorganisms for the production of bio-based succinic acid include Actinobacillus succinogenes, Anaerobiospirillum succiniciproducens, Mannheimia succiniciproducens, C. glutamicum, Bacteroides fragilis, E. coli, and Yarrowia lipolytica [6, 15, 18, 19]. The physiology of succinate-producing microorganisms varies significantly from one species to the next [3]. For example, both A. succiniciproducens and A. succinogenes follow the phosphoenolpyruvate (PEP) carbocykinase pathway exclusively to produce succinate. A. succiniciproducens is an obligate anaerobe, while A. succinogenes is a facultative anaerobe, able to switch between aerobic and anaerobic metabolic pathways. On the other hand, E. coli, a facultative anaerobe, proceeds through multiple pathways to produce succinate. A. succinogenes, unlike A. succiniciproducens and E. coli, has high tolerance to succinate salts. The PEP carboxykinase pathway is regulated by CO₂ concentration to a selected product distribution. Under high CO₂ levels (100mol CO₂/100 mol glucose), succinate is the major product, with traces of lactic acid or ethanol. At a low CO₂ level, A. succiniciproducens produces lactic acid as a major product, whereasA. succinogenes generates more ethanol than succinic and lactic acids.

Carboxylation of pyruvate, the most important central metabolite, is the controlling step of succinic acid biosynthesis. Typical biosynthesis of succinic acid utilizes glucose or glycerol as a carbon source while assimilating CO₂. Succinic acid can be produced in anaerobic and aerobic conditions [15]. Succinate yield through a fermentative pathway is limited by the supply of NADH. A combined anaerobic and aerobic strategy is often applied for the biosynthesis of succinic acid. Carbon sources, carbon dioxide, hydrogen and culture pH are critical factors for economic production of succinic acid. In addition to external CO₂ gas, carbonates are considered as a source of CO₂. Carbon dioxide and carbonates dissolved in water generate and The equilibrium between CO₂, and is governed by pH in fermentation broth. In a biosynthesis system, hydrogen is a potential electron donor to improve succinate yield by decreasing cellular redox potential and improving NADPH recycles. In addition to phosphoenolpyruvate, oxaloacetate, and malate, pyruvate was identified as a fourth node governing A. succinogenes fermentation pathways to succinate and alternative products [20]. At high CO₂ and H₂ concentrations, A. succinogenes produces more succinate and less formate, acetate, and ethanol. Studies suggest high NaHCO₃ concentrations minimized the tendency to push the flux from the C₄ pathway to C₃ pathway.

Most fermentation processes are carried out in batch reactors. High titers and yields of succinate, however, were often obtained in fed-batch or continuous systems. Succinate titers of 133 g/l were reported by using engineered C. glutamicum in a fermentation medium consisting of glucose, bicarbonate and formate under anaerobic condition (Fig. 1). At present, biosynthetic processes have limited potential for industrial scale application due to the use of formate which complicates downstream purification and increases production cost [6]. Okino et al. achieved a very high succinic acid titer of 146 g/l with 1.40 mol/mol yield under oxygen deprivation with intermittent addition of sodium bicarbonate and glucose [21]. Succinate titers of up to 110 g/l at 83–87 wt.% yield were achieved with A. succinogenes by maintaining the pH with magnesium [12, 22]. The recombinant E. coli AFP184 (Fig. 2) demonstrated productivity of 3 g/ l/h by applying dual-phase fermentations in high sugar concentrations whereas E. coli produced succinic acid with titers of up to 99.2 g/l at productivities of up to 1.3 g/l/h [12]. A very high productivity of 10.4 g/l/h at a titer of 83 g/l and a 1.35 mol/mol yield were obtained in a continuous system with a membrane for cell recycling and an on-line electrodialysis system to eliminate the end-product inhibition [15, 23]. Productivity reaches 14.8 g/l/h at a titer of 42 g/l when using an integrated membrane-bioreactor-electrodialysis system under a suitable culture condition [23]. The integrated fermentation system produces a highly concentrated cell-free succinate solution that can be recovered by simple water evaporation/ acidification. The production yield affects the variable costs of raw materials and utilities while productivity and titer govern fixed costs and total production investment? Low production rate and low product concentrations result in high energy input and cost. Downstream recovery and purification represent both technological and economical challenges. Product isolation processes will need to be coupled with upstream fermentation to render industrial scale-up manufacturing viable. Strategies include genetically engineering strains to reduce acid byproducts, reactive extraction to improve succinate selectivity, in situ product removal, and separation of acetic acid from succinic acid using an emulsion liquid membrane (ELM) [15, 24]. Thus, to improve the competitiveness of succinic acid biosynthesis, it is essential to address the importance of succinatetolerant microorganisms, low-cost biomass feedstocks and downstream product recovery. The succinate fermentationefficiencies of a selected number of bacteria strains are summarized in Table 2.

An ideal microorganism candidate for industrial production must be able to utilize a wide range of carbon sources, such as fructose, glucose, sucrose, maltose, lactose and xylose, for the synthesis of succinic acid. This capability provides the opportunity to employ agricultural and food processing wastes as feedstock to reduce production cost. However, E. coli almost exclusively uses glucose for succinic acid production. Wang et al. engineered an E. coli strain SBS550MG bearing both the pHL413 plasmid and the pUR400 plasmid for utilizing sucrose and mixed sugars to produce succinate [26]. Evaluation of different substrates indicates that the nature of sugar carbon sources affects the fermentation pathways and thus the yield and productivity of succinate. The E. coli strain SBS550MG pHL413 produced the highest succinate yield of 1.86 mol/mol with a productivity of 0.82 g/ l/h when provided with fructose as a carbon source. The strain consumed glucose preferentially and grew better in a mixture of glucose and fructose than in fructose alone. The succinate yield was 1.76 mol/mol hexose with a productivity of 1.21 g/ l/h in a glucose/ fructosemixture of 1/1 ratio. E. coli are not able to consume sucrose to produce succinate due to the lack of an invertase. E. coli strains able to metabolize sucrose were achieved via expression of the pUR400 plasmid containing scrK, Y, A, B, and R genes that can convert sucrose to β-D-fructose and α-D-glucose 6-phosphate. However, a low productivity of 0.48 g/ l/h was obtained using SBS550MG pHL413 pUR400 due to slow sucrose hydrolysis during the early stage of the fermentation process. The fermentation strategy was then changed to utilize sucrose during the aerobic phase before switching to a mixture of fructose, glucose, and sucrose during the anaerobic phase. The processes achieved a yield of 1.67 mol/mol hexose with a productivity of 0.85 g/ l/h. The strain consumed sugars in the following priority: glucose, sucrose, and then fructose. Fermentation in a sucrose hydrolysis solution achieved a similar result, with a succinate yield of 1.67 mol/mol hexose and a productivity of 0.86 g/ l/h. Studies on the effect of the dissolved oxygen (DO) level during the aerobic phase suggest a high agitation rate is conducive for increased cell growth rate and succinate productivity, while low agitation is better for increased succinate production yield. Formation of a microaerobic condition before switching to a completely anaerobic condition (due to rapid depletion of the DO level following slower agitation of the growth medium) may have contributed to differences in cell metabolism and led to improved succinate yield. Li et al. utilized corn stover hydrolysate as a carbon source and spent yeast hydrolysate as a nitrogen source to produce 56.4 g/ l succinic acid with A. succinogenes [27]. They achieved 0.73 g/ g yield and over 50% production cost reduction. Earlier studies indicated that the hydrolysis of corn stover generateda 2 : 1 (w/w) ratio of glucose to xylose. Amixedsugar feedstock resulted in delayed succinic acid production when compared with that achieved with glucose only. In general, consumption of xylose starts only when glucose is depleted. A wide range of carbon sources including corn starch, corn steep liquor, whey, cane molasses, glycerol, and lignocelluloses, have been evaluated for the fermentative production of succinic acid. Metabolic engineering approaches to facilitate simultaneous digestion of sugars are beneficial for the industrial production of bio-succinate. The delay in succinic acid production resulting from mixed sugar feedstocks may have also been caused by the presence of inhibitors, such as furfural, HMF, and acetic acid, in corn stover hydrolysate. E. coli is an ideal candidate for future commercial bio

Fig. 1:

Simplified metabolic pathway of wild-type C. glutamicum for succinate production. PEP, phosphoenolpyruvate [21].

Fig. 2:

Simplified anaerobic production of succinate in E. coli AFP184. The blocked reactions are shown by crosses [25].

Table 2: Summary of succinate fermentation efficiency using different bacteria strains.

succinate production due to in-depth knowledge on the genetic manipulation thereof, a high possible yield of 1.72 mol/mol glucose, and the general acceptance of its usage in industrial process [16].

Applying solid-state fermentation (SSF) with two fungi strains and using wheat milling byproducts, Dorado et al. were able to produce enough carbon and nitrogen nutrients for the production of succinic acid [28]. Agricultural and food wastes are more easily used in solid-state fermentation (SSF) than in submerged fermentation. Other low cost raw materials used for succinic acid fermentation include corncobs, corn straw and stalk, rapeseed meal, cassava starch, spirit-based distiller grains, and cane molasses [18, 29]. Typical strains employed for the production of succinic acid include isolated microorganisms, recombinant microorganisms and or improved strain (screened mutations). Biomass derived media and fermentation strategies affect the yield and productivity of bio-based succinic acid production. Genetic engineering and fermentation strategies provide a range of economic approaches for flexible and integrated production. Succinic acid was co-produced with other value-added products, such as malic acid[30], isoamyl acetate [31] and polyhydroxybutyrate [32, 33]. For fresh water conservation, seawater was successfully used for the fermentation of wheat-derived substrates to produce succinic acid [34]. Separation of succinic acid from fermentation broth accounts for a major fraction of succinic acid production cost. Typical isolation methods include precipitation, reactive extraction, and direct crystallization while integrated membrane filtration-electrodialysis is the most promising approach with improved titer and productivity. Utilization of agro-waste and integration of processes results in significant cost reduction. It is possible to produce succinic acid by fermentation for about US $ 0.55–1.10 per kg according to McKinlay et al. [17]. Producing succinic acid with biological routes is expected to save up to 40% energy consumption and result in lower carbon emissions when compared to production by typical chemical processes [29]. Life cycle assessment of bio-succinic acid prepared by low pHyeast-based fermentation technology and direct crystallization downstream processing indicate a reduction of 90% non-renewable energy use and reduced greenhouse gas (GHG) emissions by a factor of two when compared to succinic acid prepared with petro-based maleic anhydride [35].

Downstream recovery and purification account for considerable costs when producing succinic acid. The separation process includes removal of cells and proteins, concentration of succinate, conversion of succinic salt to acid, and purification of succinic acid [3, 5]. Downstream processing cost can as account for up to 60–70% of the total succinic acid production cost [12]. At first, microbial cells are separated from the fermentation broth by centrifugation or filtration followed by ultrafiltration to remove proteins, polysaccharides, and other oligomers. Several approaches to isolate succinate from fermentation broth were evaluated [3, 17]. Succinic acid can be obtained by isolating succinate from fermentation broth by electrodialysis followed by a conversion process using a bipolar, water-splitting membrane stack. The resulting succinic acid can then be purified by ion exchange chromatographies to achieve 80% recovery in dry weight. The remaining 20% of the recovered material consists of acetic acid. Datta et al. described in their patent for a precipitate succinate hydroxide process to maintain the pH of the fermentation broth in the range of 5.8 to 6.4 by addition of calcium oxide or calcium hydroxide [36]. The resulting aqueous slurry of calcium succinate was then transformed to succinic acid by introducing sulfuric acid. The resulting product was then purified with ion exchange resin to obtain succinic acid containing less than 1% nitrogenous impurities. The disadvantage of this process is that it requires handling large amounts of slurry and calcium sulfate waste. Alternatively, the pH of the fermentation broth can also be maintained by adding NH₄OH and later reacting the resulting diammonium succinate with ammonium bisulfate to form ammonium sulfate and succinic acid. Addition of sulfic acid to this mixture of ammonium sulfate and succinic acid resulted in the crystallization of succinic acid. The succinic acid was then recovered in 90% dry weight by dissolution in methanol to precipitate contaminating sulfates. Succinic acid was also purified by reactive extraction with tri-n-octylamine. There is still a need for considerable efforts to lower the recovery cost of succinic acid to achieve economically viable industrial scale production thereof.

2.3 Catalytic conversion of succinic acid

Typical chemical transformations of succinic acid include esterifications, amidations and hydrogenations. Esterifications of succinic acid were used to prepare diester intermediates for polymerization. High esterification yields of greater than 75% were achieved for low purity succinic acid recovered from fermentation broths by means of an approach combining both conductive heating and microwave irradiation [37]. This in situ esterification in fermentation broth was also utilized as a method for downstream isolation of succinic acid. Catalysts for the in situ transformation process must be tolerant to water and salts, as well as resistant to organic contaminates. Hydrogenation of succinic acid produces a range of value-added chemicals, including tetrahydrofuran (THF) (used as a solvent), γ-bytyrolactone (GBL) (used as a solvent), as an intermediate in the manufacture of pyrrolidones, as an ingredient in the production of pesticides and herbicides, and 1,4-butanediol (BDO) for polymer synthesis.

Direct downstream catalytic transformation of succinic acid into valuable derivatives in filtered aqueous fermentation broth eliminates the need for isolation and thus reduces production cost [38]. Petrochemicals are mostly in a low oxidation state and therefore require oxidative derivatization, whereas renewable chemicals are often in a high oxidation state and need to be reduced. Hydrogenation of maleic acid results in the formation of succinic acid. Thus, hydrogenation of maleic acid in the aqueous phase can be easily adapted to produce succinic acid in a reduction process. Group VIII metals are the most active catalysts for the hydrogenation of succinic acidto produce γ-butyrolactone (GBL), tetrahydrofuran (THF), and 1,4-butanediol (BDO). Combinations of Group VIII metals, or their combinations with other metals, such as rhenium and tin, are often deposited on carbon, TiO₂ and ZrO₂ substrates to serve as catalysts. In addition to the nature of metals and properties of supports, dispersion and repartition of the metals on and in their catalyst supports are also critical for the catalytic efficiency of the hydrogenation process. A high degree of dispersion in bimetallic catalysts prevents the formation of unwanted microstructures that lead to catalyst aging and loss of activity. On the other hand, reaction parameters have a strong impact on product distribution and yield. Reductive amination of succinic acid using amine, ammonium, or ammonia produces pyrrolidones and derivatives. 2-Pyrrolidone is an intermediate in the preparation of nylon-4, pharmaceuticals, medicines and agrochemicals. N-methyl-2-pyrrolidone (NMP) is an important solvent in the polymer and electronic industries. The reaction selectivity appears to be sensitive to the reactant ratios and an excess of alcohol is required for the production of NMP. Direct aqueous synthesis of pyrrolidones from succinic acid without producing GBL was achieved [38]. Catalytic conversion of succinic acid into valuable derivatives is illustrated in Fig. 3.

Fig. 3:

Catalytic transformation of succinic acid into valuable derivatives.

The hydrogenation of succinates is similar to the Davy–Mackee process for the commercial production of 1,4-butanediol from maleate. Copper-based catalysts were studied for the hydrogenation of succinates to produce BDO [39, 40]. Alternatively, the hydrogenation of succinic anhydride over Cu–Zn, Cu–Zn–Zr, Cu–Zn–Cr–Zr, Cu–Mn, Re–Cu–Zn was performed using vaper-phase fixed-bed rectors [41–46]. Ruthenium complexes were utilized as homogeneous catalysts for the hydrogenation of succinic anhydride to GBL under mild conditions [47–50]. Other homogeneous catalysts include Ru acetylacetonate, trioctylphosphine and HBF₄ [51–53]. Although Ru complexes provide high product selectivity, drawbacks of using homogeneous catalysts include difficulty of separation from the reaction medium and the presence of unfavorable halogen ligands. Aqueous phase hydrogenation of succinic acid is environmentally benign and enjoys reduced production costs. However, acidic media poses a challenge for metal catalysts. Ly et al. evaluated Pd–Re bimetal catalysts with various Re content for aqueous phase hydrogenation [54]. It was observed that the Re position and oxidation states (Re³⁺, Re⁰) on the reduced bimetallic samples were related to their preparation procedures [55]. The presence of a small amount of Re in a Pd/ C catalyst enhances production of γ-butyrolactone, whereas further increase of Re loading resulted in a high THF yield [56]. Additional studies also indicate hydrogenation efficiencies can be improved by using palladium-rhenium catalysts supported by carbon materials [57, 58]. The modified Ru catalysts also provide high conversions of succinic acid in the aqueous solution to produce THF and 1,4-butanediol over Ru–Mo, Ru–Sn, Ru–Re, and Ru–Re–Sn trimetall catalyst [59–62]. Succinic acid can be transformed into 1,4-butanediol with a yield of over 70% in the presence of a Pd-5FeO_x/C catalyst under the relatively mild conditions of 200 °C and 5 MPa H₂ [63].

The supporting substrate plays an important role in governing catalyst particle size and thus hydrogenation performance. A Pd/SiO₂–NH₂ catalyst prepared by co-condensation method exhibits high catalytic activity in converting succinic acid to γ-butyrolactone with 100% conversion and 94% selectivity using 1,4-dioxane as a solvent at 240 °C and 60 bar for 4 h [64]. Kim et al. evaluated the catalytical activity of Pd-WO₃/Al₂O₃ for hydrogenation reactions in a mixture selected from dioxane, ethanol, and H₂O at 100–300 °C [69]. Studies of supported ruthenium and rehenium catalysts treated with acid for the hydrogenation of succinic acid indicate that the yields of THF increase as catalyst particle size decreases [65–68]. It was identified that mesoporous carbon and Alumina xerogel provide improved hydrogenation efficiency due to the fine dispersion of catalyst on these supports [69, 70]. Avery small amount of Pt (100 ppm) in gold catalysts facilitates H₂ dissociation and achieves 97% selectivity to GBL at 97% conversion [71]. The composition of Ru-Co bimetallic catalyst governs the reaction pathway and therefore the distribution of products [72].

3.1 Biological routes to itaconic acid

Although some Aspergillus species and other microorganisms were identified as candidates to produce itaconic acid, A. terreus remains the dominant host for industrial itaconic acid production [88]. Biosynthesis of itaconic acid is an aerobic process requiring 1.5 mol of O₂ for every mole of bio-based diacid generated (Fig. 4) [8]. It is, therefore, a challenge to balance between the competing concerns of sufficient oxygen to the growth medium and limiting cell damage of filamentous microorganisms, such as A. terreus, resulting from the hydromechanical stress of aeration. The influence was reduced to some degree by using an air lift bioreactor and suitable control of medium pH.

Fig. 4:

Biosynthesis of itaconic acid in A. terreus [89]. MTT, mitochondrial tricarboxylate transporter.

High itaconic acid yields were obtained by fermentation of glucose. For commercial production, product yield per substrate is critical for profitability. Theoretically, 1 g of glucose can be converted to 0.72 g itaconic acid. Yahiro et al. achieved yields of up to 0.57 g itaconic acid per gram of glucose, but observed a reduction in productivity and yield at titers above 60 g/ l [90]. Subsequent optimization strategies to gain cost competitiveness are enhancing productivity and titer as well as using low cost substrates. Studies indicate some limitation on the chemical mutation of A. terreus to achieve high titer. It has been proposed that the transfer into A. terreus of certain genes in more robust microbial producers (such as A. niger which is known to produce high yields and titers of citric acid) may lead to higher titers. Productivity of industrial batch fermentation using molasses as a substrate is in the range of 1 g/ l/h [8]. To be competitive with petroleum-based chemicals, productivity of a typical biosynthesis process needs to reach at least 2.5 g/ l/h. Continuous fermentations may help achieve high fermentation productivity. Various sugars, such as glucose, sucrose, xylose, saccharose, lactose, starch, molasses, and hydrolysate derived from lignocellulosic materials, can be carbon sources for the biosynthesis of itaconic acid using A. terreus. Utilization of wood waste as a feedstock for production of bio-based acids proves less efficient, due to the presence of inhibitory compounds; making matters worse, xylose leads to lower yields than those of other sugars. A process to separate inhibitors in the biomass hydrolysate might be inevitable. A. terreus can also utilize pure glycerol for the production of itaconic acid at yields of up to 55.9 wt.% after 233 hours [8]. In addition to A. terreus and A. niger, non-filamentousmicroorganisms, such as Pseudozyma anatarctica and U. maydis, various yeasts, such as Y. lipolytica, and bacteria, such as E. coli, were evaluated for the production of itaconic acid with some degree of success. Methods for downstream recovery and purification of itaconic acid include crystallization, reactive extraction using organophosphorous compounds or quaternary amine, and electrodialysis. There is a critical step to separate itaconic acid and residual glucose which is known to interfere with the crystallization.

Bred strains of A. terreus can secrete a significant concentration of itaconate (> 80 g/l) in a media. However, this is still far from the titers in excess of 200 g/l achieved in the citric acid industrial process and the estimated maximum theoretical titer of 240 g/ l [91]. Although the itaconic acid production pathway is not clear, CAD was found to be the key enzyme of the pathway. A high transcriptional level of the cad gene is crucial for the efficiency of itaconic acid biosynthesis. Due to poor stability of the CAD protein, the amino acid sequencing thereof was not completed until a substantial amount of the enzyme was purified in 2008. In addition to screening existing strains, high titers can be achieved by employing targeted genetic engineering. On the other hand, media compositions and engineering approaches related to oxygen distribution and bioreactor design also contribute to itaconic acid production efficiency. Introduction of copper ions and steady aeration were found to positively influence itaconic acid production. Various reactors including bubble column reactors, tubular reactors, packed bubble column reactors, and Air-Lift reactors have been evaluated for biosynthesis. Starch was utilized as a carbon source for fermentation to reduce production cost. Hydrolysis of starch with glucoamylase (5000AUN/ml) or nitric acid at pH 2 led to itaconic acid yields of 0.36 and 0.35 g/ g starch, respectively [7]. Relatively high titers were also achieved with agricultural residuals, such as rejected bananas (28.5 g/l), apples (31.0 g/l), Jatropha seed cakes (24.45 g/l), and sago starch (48.2 g/l).

Chemical routes for itaconic acid synthesis are impractical, while fermentative itaconic acid production using A. terreus is economically challenging [80]. Utilization of itaconic acid, therefore, is limited by its high production cost and limited number of suppliers [78]. To lower itaconate production cost, fermentation was enhanced by genetic manipulation and physiological regulation of E. coli. Okamoto et al. achieved 4.34 g/ l itaconic acid production by icd inactivation and acnB overexpression in cad-expressing E. coli [86]. They also developed a recombinant E. coli expressing α-amylase on its cell surface to saccharify starch near the cell surface and utilize the resulting sugar for the biosynthesis of itaconic acid [92].

A novel synthetic pathway in E. coli to produce itaconate is shown in Fig. 5. Itaconate is produced by a submerged-cultured fermentation with a strain of E. coli in a medium containing a carbohydrate source such as glycerol, glucose, or molasses. Nutrients contain amino acids, phosphates and metal salts, as well as sulfuric acid, hydrochloric acid or sodium hydroxide can be added as pH adjustment agent. A medium containing 6wt.% of glycerol is incubated at 30–35° and pH of 6.0–7.0 for 48 hours. Itaconate was produced at 1.5 g/ l/h and a 99.5 wt.% pure product was produced in a continuous purification section, which includes double-effect evaporators, two-stage crystallization, activated carbon treatment, and rotary dryer operation units. Successful production of itaconate from the engineered E. coli opens up the possibility of using non-native, easily manipulated organisms for itaconate production [93]. Economic analyses of a fermentation process using E. coli yielded a production cost of $ 2.0/kg for an itaconate plant with annual capacity of 20 000 MT. The calculation of such a production cost was based on experimental results of 80 g/ l titer and 1.5 g/ l/h productivity, as well as on assumptions of a 95% recovery rate, crude glycerol cost of $ 0.2/kg, a nutrient cost of $ 0.5/kg and a 10-year plant life.

Fig. 5:

Schematic representation of itaconate production in engineered E. coli (ppc, phosphenolpyruvate carboxylase; gltA, citrate synthase; acn, aconitase; cad, cis-aconitic acid decarboxylase; PEP, phosphoenolpyruvate; OAA, oxaloacetate) [93].

3.2 Catalytic conversion of itaconic acid

The catalytic conversions of itaconic acid to 2-methyl-1,4-butanediol, 3-methyltetra-hydrofuran, and 2-methyl γ-butyrolactone as well as 3-methyl γ-butyrolactone should be similar to those associated with the transformation of succinic acid to related derivatives [94–100]. The hydrogenation of itaconic acid to 2-methylsuccinic acid, a chemical with pharmaceutical applications, was performed with Ru-Starbon, Ru/TiO₂ or Ru-complex as a catalyst [100, 101]. The enantioselective hydrogenation of itaconic acid to (R)- or (S)-methylsuccinic acid using palladium and rhodium catalysts is also described. Decarboxylation of itaconic acid to bio-based methacrylic acid (MAA) was achieved with transition-metal catalysts in aqueous phase [102, 103]. Catalytic conversion of succinic acid into valuable derivatives is illustrated in Fig. 6.

Fig. 6:

Catalytic transformation of succinic acid into valuable derivatives.