Lipase-Mediated Syntheses of Trimethylolpropane-Based Biolubricant and Cyclic Carbonate

Abstract Biocatalysis is considered as benign and efficient alternative to chemical catalysis for organic syntheses. Lipases are the most versatile biological catalysts implemented so far with great potential for production of different chemicals and materials in non-conventional reaction media. This thesis presents investigations on lipase catalyzed esterification and transesterification reactions in solvent-free media with a polyol (tri-ol), trimethylolpropane (TMP) to form TMP-trioleate and -cyclic carbonate for lubricant and polymer applications, respectively. Conventional lubricants are mineral oil based and lack biodegradability resulting in their accumulation in the environment. Synthetic esters of polyhydric alcohols and fatty acids are biodegradable and possess desirable technical properties for lubricant applications. Synthesis of TMP-trioleate from oleic acid and TMP catalyzed by commercial immobilized Candida antarctica lipase B, Novozym®435 (N435) was studied by varying reaction parameters. The product obtained possesses desirable pour point (-42 °C) for lubricant applications in sub-zero conditions. The biocatalyst was recycled in reactions at 70 °C for 7 batches, 24 h each, with a half-life of 94 h. The biocatalyst half-life was doubled by washing it with 2-propanol between the batches. A simplified kinetic model was developed for the lipase-catalyzed reaction in order to facilitate optimization and design of the process and minimize the amount of resources required for investigations of the process. The methodology used for the kinetic modeling is applicable for similar types of enzymatic reactions involving multi-substrate multi-product systems. Cyclic carbonates are potential monomers for phosgene-/isocyanate-free polycarbonates and polyurethanes that have wide range of applications. Six-membered cyclic carbonates can readily undergo ring-opening polymerization to form aliphatic polycarbonates and polyurethanes and their copolymers. Six-membered cyclic carbonate with hydroxyl functional group was obtained with 75% yield using a chemoenzymatic process involving lipase B catalyzed transesterification of dimethylcarbonate (DMC) with TMP in the presence of molecular sieve to form linear TMP carbonate followed by thermal cyclization. Performing the reaction in a recirculating flow reactor, higher conversion rates were obtained compared to the batch process, the product was recovered easily without extra separation steps, and the biocatalyst and molecular sieve remained intact for reuse. In silico evaluations of the reaction accompanied with empirical investigations confirmed that lipase B prefers DMC as acyl-donor while TMP and its derivatives, formed during the course of the reaction, serve as acyl acceptors. The formation of TMP carbonate oligomers hence found to be non-enzymatic and intensified by heat

Realizing the Continuous Chemoenzymatic Synthesis of Anilines Using an Immobilized Nitroreductase

The use of biocatalysis for classically synthetic transformations has seen an increase in recent years, driven by the sustainability credentials bio-based approaches can offer the chemical industry. Despite this, the biocatalytic reduction of aromatic nitro compounds using nitroreductase biocatalysts has not received significant attention in the context of synthetic chemistry. Herein, a nitroreductase (NR-55) is demonstrated to complete aromatic nitro reduction in a continuous packed-bed reactor for the first time. Immobilization on an amino-functionalized resin with a glucose dehydrogenase (GDH-101) permits extended reuse of the immobilized system, all operating at room temperature and pressure in aqueous buffer. By transferring into flow, a continuous extraction module is incorporated, allowing the reaction and workup to be continuously undertaken in a single operation. This is extended to showcase a closed-loop aqueous phase, permitting reuse of the contained cofactors, with a productivity of >10 g g and milligram isolated yields >50% for the product anilines. This facile method removes the need for high-pressure hydrogen gas and precious-metal catalysts and proceeds with high chemoselectivity in the presence of hydrogenation-labile halides. Application of this continuous biocatalytic methodology to panels of aryl nitro compounds could offer a sustainable approach to its energy and resource-intensive precious-metal-catalyzed counterpart. [Abstract copyright: © 2023 The Authors. Published by American Chemical Society.

Enhancing the productivity of the bi-enzymatic convergent cascade for ɛ-caprolactone synthesis through design of experiments and a biphasic system

A two-step Design of Experiments (DoE) strategy followed by a two-liquid-phase system (2LPS) was applied to enhance the ɛ-caprolactone yield in the cyclohexanone monooxygenase (CHMO)-alcohol dehydrogenase (ADH) convergent cascade system. The key reaction parameters were identified and optimized for the determination of an optimal operational window for the aqueous media. In the 2LPS system, high partitioning of the lactone product was observed in 2-MeTHF and in toluene; however, these solvents led to drastically reduced enzymatic activity. Dodecane was chosen as the non-miscible organic phase owing to the enzymes’ high residual activity, despite the low partitioning of the lactone. Cyclohexanone concentrations up to 75 mM were applied in the aqueous media. The turnover numbers for the nicotinamide cofactor and for the ADH reached up to 980 and 392,000, respectively whereas a turnover number value of 5600 was achieved for the CHMO. By employing a 2LPS, whereby 91 mM of cyclohexanone was applied in the second phase, turnover numbers were slightly increased

Multi-steps green process for synthesis of six-membered functional cyclic carbonate from trimethylolpropane by lipase catalyzed methacrylation and carbonation, and thermal cyclization.

A highly functionalized six-membered cyclic carbonate, methacrylated trimethylolpropane (TMP) cyclic carbonate, which can be used as a potential monomer for bisphenol-free polycarbonates and isocyanate-free polyurethanes, was synthesized by two steps transesterifications catalyzed by immobilized Candida antarctica lipase B, Novozym®435 (N435) followed by thermal cyclization. TMP was functionalized as 70-80% selectivity of mono-methacrylate with 70% conversion was achieved, and the reaction rate was evaluated using various acyl donors such as methacrylic acid, methacrylate-methyl ester, -ethyl ester and -vinyl ester. As a new observation, the fastest rate obtained was for the transesterfication reaction using methacrylate methyl ester. By-products resulted from leaving groups were adsorbed on the molecular sieves (4Å) to minimize the effect of leaving group on the equilibrium. The difference of reaction rate was explained by molecular dynamic simulations on interactions between carbonyl oxygen and amino acid residues (Thr(40) and Gln(157) ) in the active site of lipase. Our docking studies revealed that as acyl donor, methyl ester was preferred for the initial conformation of the 1(st) tetrahederal intermediate with hydrogen bonding interactions. TMP-monomethacrylate (TMP-mMA) cyclic carbonate was obtained in 63% yield (74.1% calculated in 85% conversion) from the lipase-catalyzed carbonation reaction of TMP-mMA with dimethylcarbonate, and followed by thermal cyclization of the monocarbonate at 90°C. From the multiple reactions demonstrated in gram scale, TMP-mMA cyclic carbonate was obtained as a green process without using chlorinated solvent and reagent. This article is protected by copyright. All rights reserved

A Bi-enzymatic Convergent Cascade for epsilon-Caprolactone Synthesis Employing 1,6-Hexanediol as a "Double-Smart Cosubstrate'

A bi-enzymatic cascade consisting of a Baeyer-Villiger monooxygenase and an alcohol dehydrogenase (ADH) was designed in a convergent fashion to utilise two molar equivalents of cyclohexanone (CHO) and one equivalent of 1,6-hexanediol as a 'double-smart cosubstrate' to produce epsilon-caprolactone (ECL) with water as sole by-product. The convergent enzymatic cascade reaction reported herein, is performed at ambient conditions in water, is self-sufficient with respect to cofactor, and incorporates all starting materials into the desired product, ECL. Among different enzymes explored, the reaction catalysed by cyclohexanone monooxygenase from Acinetobacter sp. NCIMB 9871 coupled with ADH from Thermoanaerobacter ethanolicus showed the best results, reaching 91% conversion of CHO after 24h with a product titre of 2gL(-1). Scale-up of the coupled system (50mL) performed better than the small-scale reactions and >99% conversion of CHO and ECL concentration of 20mM were achieved within 18h

Six‐membered cyclic carbonates from trimethylolpropane: Lipase‐mediated synthesis in a flow reactor and in silico evaluation of the reaction

Six-membered cyclic carbonates with hydroxyl and methoxycarbonyloxy functional groups were prepared by transesterification of trimethylolpropane (TMP) with dimethylcarbonate (DMC) by solvent-free lipase-mediated flow reaction followed by thermal cyclization. The flow reaction efficiency was evaluated using different configurations of reactor consisting of packed beds of Novozym 435 (immobilized Candida antarctica lipase B—CalB—a.k.a. N435) and molecular sieves, flow rate, and biocatalyst loads. The mixed column of the bio-catalyst and molecular sieves, allowing rapid and efficient removal of the by-product—methanol—was the most efficient setup. Higher conversion (81.6%) in the flow reaction com-pared to batch process (72%) was obtained using same amount of N435 (20% (w/w)N435:TMP) at 12 h, and the undesirable dimer and oligomer formation were suppressed.Moreover, the product was recovered easily without extra separation steps, and the biocatalyst and the molecular sieves remained intact for subsequent regeneration and recycling.The reaction of CalB with DMC and the primary transesterification product, monocarbonated TMP, respectively, as acyl donors was evaluated by in silico modeling and empirically to determine the role of the enzyme in the formation of cyclic carbonates and other side products. DMC was shown to be the preferred acyl donor, suggesting that TMP and its carbonated derivatives serve only as acyl acceptors in the lipase-catalyzed reaction. Subsequent cyclization to cyclic carbonate is catalyzed at increased temperature and not by the enzyme

Optimization of a two-step process comprising lipase catalysis and thermal cyclizationimproves the efficiency of synthesis of six-membered cyclic carbonate from trimethylolpropane and dimethylcarbonate.

Six-membered cyclic carbonates are potential monomers for phosgene and/or isocyanate free polycarbonates and polyurethanes via ring-opening polymerization. A two-step process for their synthesis comprising lipase-catalyzed transesterificationofa polyol, trimethylolpropane(TMP) with dimethylcarbonate(DMC)in a solvent-free system followed by thermal cyclization was optimized to improve process efficiency and selectivity. Using full factorial designed experiments and partial least squares (PLS) modeling for thereaction catalyzed by Novozym®435 (N435; immobilized Candida antarctica lipase B), the optimum conditions for obtaining either high proportion of mono-carbonated TMP and TMP-cyclic-carbonate (3 and 4), or di-carbonated TMP and monocarbonated TMP-cyclic-carbonate (5 and 6) were found. The PLS model predicted that the reactions using 15-20% (w/w) N435 at DMC:TMP molar ratio of 10-30 can reach about 65% total yield of 3 and 4 within 10 h, and 65-70% total yield of 5 and 6 within 32-37 h, respectively. High consistency between the predictedresults and empirical data was shown with 66.1% yield of 3 and 4 at 7 h and 67.4% yield of 5 and 6 at 35 h, using 18% (w/w) biocatalyst and DMC:TMPmolar ratio of20. Thermal cyclization of the product from 7 h reaction, at 110 °C in the presence of acetonitrile increased the overall yield of cyclic carbonate 4 from about 2% to more than 75%within 24 h.N435 was reused for 5 consecutive batches, 10 h each, to give 3+4 with a yield of about 65% in each run. © 2012 American Institute of Chemical Engineers Biotechnol. Prog., 2012