Production Of Fine Chemicals Research Articles

Polyketide synthases as a platform for chemical product design

To reduce the effects of greenhouse gas emissions on climate change, scientific efforts have sought to develop biofuels and bio‐based commodity chemicals as petrochemical replacements primarily for their environmental benefits. As the biological design space is far greater than chemical synthesis, there has been a drive to leverage this ability to create and replace fine and specialty chemicals. While polyketide synthases have traditionally been studied for their biosynthesis of pharmaceutical chemicals, the unique advantages of engineering polyketide synthases for the production of biofuels, commodity chemicals, and both pharmaceutical and nonpharmaceutical fine and specialty chemicals are discussed. With our increased understanding of polyketide biosynthetic logic, new computational tools for prospective polyketide synthesis pathways for the creation of existing and novel biochemicals are also outlined. © 2018 American Institute of Chemical Engineers AIChE J, 64: 4201–4207, 2018

In vitro culture initiation of Cleome droserifolia glandular trichomes for potential biotechnological applications

Cleome droserifolia (Forssk.) Del., belongs to family Cleomaceae. It is an aromatic shrub covered with glandular hairs that give off a distinct volatile smell of its medicinal important chemicals. The initiation of glandular hairs cultures of Cleome droserifolia was successfully achieved on Murashige and Skoog medium containing a combination of BA (1mg l-1) with 2, 4D (10 mg l-1) and IAA (6mg l-1) with Kinetin (4mg l-1) in in vitro cultures. The obtained result will open the door for new biotechnological applications for the production of fine chemicals. Stem and leaf petiole glandular hairs expanded and formed small side daughter hairs perpendicular to the mother trichome. Multiplication of single trichome is obtained from leaf lamina and added new divisions from glands. The best response for multiplications and branching of daughter glandular trichomes was observed on MS medium containing 1mg l-1BA and 10mg l-1 2, 4 D.

W(OTf)6 -Catalyzed Synthesis of γ-Lactones by Ring Contraction of Macrolides or Ring Closing of Terminal Hydroxyfatty Acids in Ionic Liquid.

γ-Lactones are an important class of fine chemical products and are widely used in perfumes, medicines, pesticides, dyes, and other fields. Herein, a new method for γ-lactones preparation based on ring contraction was developed. Starting from macrolides, W(OTf)6 was used to catalyze the ring-opening polymerization then depolymerization. The depolymerization step was not a common ring-closing process, and the carbon number of the ring was reduced one by one by rearrangement to form the most thermodynamically stable five-membered ring compounds. γ-Caprolactone (180 °C for 10 h) was obtained in a yield of 94 % when [EMIM]OTf was used as the solvent, and the yield of isolated product was up to 85 %. The interaction of various components and the reaction mechanism were studied by FTIR spectroscopy and 1 H NMR spectroscopy, respectively. Furthermore, γ-lactones could be produced when the substrate was extended to terminal hydroxyfatty acids. Unexpectedly, the catalyst was poisoned by 1 equivalent of H2 O added during the process and thus the yield decreased greatly. The reaction is green and simple, and proceeds in one pot with high atom economy (100 % for macrolides and with water as the only byproduct for terminal hydroxyfatty acid), which provides a promising approach to synthesizing γ-lactones.

Product Inhibition in Glycerol Oxidation over Au/TiO2 Catalysts Quantified by NMR Relaxation

Liquid-phase catalytic oxidation of glycerol in aqueous solutions using porous solid catalysts represents a viable strategy for the sustainable production of fine chemicals from renewable resources. Various aspects of these novel types of reactions are still under investigation. Catalyst deactivation is one of those issues that need to be understood and addressed in order to make these processes commercially viable. In a previous study, it has been reported that the catalytic activity of Au/TiO2 catalysts for the oxidation of glycerol with O2 under basic conditions can be severely inhibited by some reaction intermediates or products. It was suggested that the presence of certain species blocks the active sites of the catalyst, preventing the adsorption of glycerol, which in turn results in a decrease of reaction rate. In this work, we used NMR relaxation time measurements in order to assess surface interactions of glycerol in a Au/TiO2 catalyst pretreated with aqueous solutions of various oxygenates, incl...

Purification and characterization of a hydroxynitrile lyase from Amygdalus pedunculata Pall

Hydroxynitrile lyases (HNLs) are widely used in the asymmetric synthesis of cyanohydrins which are organic compounds used in the production of fine chemicals and pharmaceuticals, because these enzymes exhibit high catalytic efficiency and are very economical. In the present study, seeds of A. pedunculata Pall were identified as new potential source of HNLs. The HNL from A. pedunculata Pall (APHNL) was purified 138 fold and 4.20% yield with a specific activity of 661 U/mg. SDS-PAGE result showed the enzyme to be present as a monomer and the relative molecular mass determined by MALDI-TOF MS was 61 kDa. APHNL owned highest activity at pH 6.0 and at 60 °C temperature, showing activity up to 80 °C and stable up to 60 °C. APHNL has a Km of 0.5 mM, Vmax of 665.9 μmol mg−1 min−1, Kcat of 676.5 s−1 and Kcat/Km of 1353 s−1 mM−1 using mandelonitrile as substrate. Syntheses of (R)-mandelonitrile and (R)-2-Hydroxy-2-(3-phenoxy-phenyl)-acetonitrile were carried out using APHNL and molar conversion of (R)-mandelonitrile and (R)-2-Hydroxy-2-(3-phenoxy-phenyl)-acetonitrile were 90% and 98% with 94% and 93% ee, respectively. These results indicated that APHNL was an excellent biocatalyst and has very high potential for synthesis of enantiopure cyanohydrins.

An automated Design-Build-Test-Learn pipeline for enhanced microbial production of fine chemicals

The microbial production of fine chemicals provides a promising biosustainable manufacturing solution that has led to the successful production of a growing catalog of natural products and high-value chemicals. However, development at industrial levels has been hindered by the large resource investments required. Here we present an integrated Design–Build-Test–Learn (DBTL) pipeline for the discovery and optimization of biosynthetic pathways, which is designed to be compound agnostic and automated throughout. We initially applied the pipeline for the production of the flavonoid (2S)-pinocembrin in Escherichia coli, to demonstrate rapid iterative DBTL cycling with automation at every stage. In this case, application of two DBTL cycles successfully established a production pathway improved by 500-fold, with competitive titers up to 88 mg L−1. The further application of the pipeline to optimize an alkaloids pathway demonstrates how it could facilitate the rapid optimization of microbial strains for production of any chemical compound of interest.

Limonene dehydrogenase hydroxylates the allylic methyl group of cyclic monoterpenes in the anaerobic terpene degradation by Castellaniella defragrans

The enzymatic functionalization of hydrocarbons is a central step in the global carbon cycle initiating the mineralization of methane, isoprenes, and monoterpenes, the most abundant biologically produced hydrocarbons. Also, terpene-modifying enzymes have found many applications in the energy-economic biotechnological production of fine chemicals. Here, we describe a limonene dehydrogenase that was purified from the facultatively anaerobic betaproteobacterium Castellaniella defragrans 65Phen grown on monoterpenes under denitrifying conditions in the absence of molecular oxygen. The purified limonene:ferrocenium oxidoreductase activity hydroxylated the methyl group of limonene (1-methyl-4-(1-methylethenyl)-cyclohex-1-ene) yielding perillyl alcohol ([4-(prop-1-en-2-yl)cyclohex-1-en-1-yl]methanol). The enzyme had a DTT:perillyl alcohol oxidoreductase activity yielding limonene. Mass spectrometry and molecular size determinations revealed a heterodimeric enzyme comprising CtmA and CtmB. Recently, the two proteins had been identified by transposon mutagenesis and proteomics as part of the cyclic terpene metabolism (ctm) in C. defragrans and are annotated as FAD-dependent oxidoreductases of the protein domain family phytoene dehydrogenases and related proteins (COG1233). CtmAB is the first heterodimeric enzyme in this protein superfamily. Flavins in the purified CtmAB are oxidized by ferrocenium and are reduced by limonene. Heterologous expression of CtmA, CtmB, and CtmAB in Escherichia coli demonstrated that limonene dehydrogenase activity required both subunits, each carrying a flavin cofactor. Native CtmAB oxidized a wide range of monocyclic monoterpenes containing the allylic methyl group motif (1-methyl-cyclohex-1-ene). In conclusion, we have identified CtmAB as a hydroxylating limonene dehydrogenase and the first heteromer in a family of FAD-dependent dehydrogenases acting on allylic methylene or methyl CH-bonds. We suggest placing in Enzyme Nomenclature as new entry EC 1.17.99.8.

Molecular Orientations Change Reaction Kinetics and Mechanism: A Review on Catalytic Alcohol Oxidation in Gas Phase and Liquid Phase on Size-Controlled Pt Nanoparticles

Catalytic oxidation of alcohols is an essential process for energy conversion, production of fine chemicals and pharmaceutical intermediates. Although it has been broadly utilized in industry, the basic understanding for catalytic alcohol oxidations at a molecular level, especially under both gas and liquid phases, is still lacking. In this paper, we systematically summarized our work on catalytic alcohol oxidation over size-controlled Pt nanoparticles. The studied alcohols included methanol, ethanol, 1-propanol, 2-propanol, and 2-butanol. The turnover rates of different alcohols on Pt nanoparticles and also the apparent activation energy in gas and liquid phase reactions were compared. The Pt nanoparticle size dependence of reaction rates and product selectivity was also carefully examined. Water showed very distinct effects for gas and liquid phase alcohol oxidations, either as an inhibitor or as a promoter depending on alcohol type and reaction phase. A deep understanding of different alcohol molecular orientations on Pt surface in gas and liquid phase reactions was established using sum-frequency generation spectroscopy analysis for in situ alcohol oxidations, as well as density functional theory calculation. This approach can not only explain the entirely different behaviors of alcohol oxidations in gas and liquid phases, but can also provide guidance for future catalyst/process design.

Breeding of Methanol-Tolerant Methylobacterium extorquens AM1 by Atmospheric and Room Temperature Plasma Mutagenesis Combined With Adaptive Laboratory Evolution.

Methylobacterium extorquens AM1, which can be used as a methylotrophic cell factory (MeCF) for the production of fine chemicals from methanol, is the most extensively studied model methylotrophic strain. However, its low tolerance for methanol limits the development of bioprocesses and there have been no reports of improved methanol tolerance of M. extorquens AM1. In this study, atmospheric and room temperature plasma (ARTP) mutagenesis, in combination with adaptive laboratory evolution (ALE), is used to generate a mutant with high methanol tolerance (referred to as CLY-2533). The final cell density of CLY-2533 is 7.10 times higher than that of the wild-type strain in medium containing 5% (v/v) methanol. Through comparative genomics analysis and overexpression of the exploited putative genes, seven mutated genes are identified as being closely related to the higher methanol tolerance of CLY-2533. Additionally, the mvt operon, which contains genes related to the biosynthesis of mevalonate acid (MEV), is introduced into CLY-2533. This recombinant strain shows significant improvements in both MEV production and cell growth in 5% methanol medium. These findings will be helpful in rational design of methanol-utilizing strain for an improved host platform for methanol based biomanufacturing.

Sustainability Evaluation of Alternative Routes for Fine Chemicals Production in an Early Stage of Process Design Adopting Process Simulation along with Data Envelopment Analysis

We propose a framework for the preliminary screening of chemical process designs according to sustainability criteria. Through the implementation of each flowsheet in a process simulator, several indicators based on sustainability pillars, i.e. economy, society, and environment, are calculated and then analyzed employing data envelopment analysis (DEA) in order to select the most efficient designs considering multiple criteria simultaneously. Suboptimal alternatives are further investigated by adopting a retrofit analysis, aiming to identify the parameters that most contribute to the final indicators scores, and therefore highlighting the major sources of deviation from optimal conditions. We applied this framework on a case study based on various patents regarding the production of pioglitazone hydrochloride in order to validate the capabilities of our methodology. Our approach, which embeds process simulation, sustainability indicators, and DEA within the same procedure, will support practitioners durin...

Green synthesis of dihydroxybenzene from phenol with hydrogen peroxide catalyzed by iron modified FSM-16

Catechol and hydroquinone are very important fine chemical intermediate products, which have a wide range of applications. The hydroxylation of phenol with hydrogen peroxide to produce dihydroxybenzene is a mild and environmental friendly route. In this work, FSM-16 modified by iron (Fe/FSM-16) is used as a catalyst. With Fe content of 5% (wt), phenol to H2O2 ratio of 3:1, reaction temperature of 60 °C, catalyst amount of 0.1 g and reaction time of 6 h, phenol conversion of 29.1% and dihydroxybenzene selectivity (catechol/hydroquinone = 1.6) of 93.5% are achieved. The results are close to that of TS-1 which is used in industry production and shows good prospects for industrial applications. The characterizations revealed that the Fe/FSM-16 could be reusable and stable.

Gate-to-Gate Energy Consumption in Chemical Batch Plants: Statistical Models Based on Reaction Synthesis Type

Energy consumption in the chemical industry is an important operating cost and environmental impact factor and reducing it is also explicitly mentioned as one of the key principles of green chemistry. Energy consumption has thus been included in diverse process design and evaluation tools as a key metric. However, measurements of energy consumption at the process equipment level are scarce, especially in fine chemical production typically performed in multiproduct and multipurpose batch plants. In this work, we present a shortcut approach based on statistical models, such as probability density functions (PDF) and classification trees, for estimating steam consumption which typically represents the highest energy utility consumption in batch plants. The output of these models is in the form of intervals derived from PDF interquartile ranges and as classes derived from the classification trees, respectively. The validation results (i.e., goodness of fit, cross validation, and case studies) show that the mo...

Comparative Analysis of Bacterial Cytochromes P450 Involved in the Biosynthesis of 16‐ Membered Ring Macrolide Antibiotics

Members of the cytochrome P450 superfamily of monooxygenases (P450s) are some of nature's most ubiquitous and versatile enzymes for performing oxidative metabolic transformations. Their unmatched ability to selectively functionalize C–H bonds has led to their growing employment in academic and industrial settings for the production of fine and commodity chemicals. Many of the most interesting and potentially biocatalytically useful P450s come from microorganisms, where they catalyze key tailoring reactions in natural product biosynthetic pathways. While most of these enzymes act on structurally complex pathway intermediates with high selectivity, they often exhibit narrow substrate scope, thus limiting their broader application. In the present work, we biochemically and structurally characterized diverse bacterial P450s involved in the biosynthesis of 16-membered ring macrolide antibiotics with significant potential for development into robust biocatalysts for the late-stage functionalization of complex molecules. Following exploratory efforts to probe the reactivity properties of the P450 MycCI from the mycinamicin biosynthetic pathway, we discovered that the enzyme exhibits appreciable activity on several 16-membered ring macrolactones independent of their glycosylation state. These results were corroborated by performing equilibrium substrate binding and kinetics experiments along with X-ray crystallographic analysis of MycCI bound to its native substrate. We also characterized TylHI, a homologous P450 from the tylosin pathway, and showed that its substrate scope is severely restricted compared with that of MycCI. Turnover and equilibrium binding experiments with substrate analogs revealed that TylHI exhibits a strict preference for 16-membered ring macrolides bearing the deoxyamino sugar mycaminose. These results were partially explained through analysis of the X-ray crystal structure of TylHI in complex with its native substrate together with biochemical characterization of several sitedirected mutants. Comparative analysis of the MycCI/TylHI homolog ChmHI from the chalcomycin biosynthetic pathway provided a basis for constructing MycCI/TylHI chimeras in order to gain further insight into the features dictating the differences in the reactivity profiles of these two related P450s. These experiments unveiled the central role of the BC loop region in influencing the binding of 16-membered ring substrates to MycCI and TylHI. Overall, our studies have shed light on the molecular-level details underpinning the unique catalytic divergence of two homologous biosynthetic P450s and point toward specific regions in each protein that may be targeted in future efforts to rationally engineer novel P450-based biocatalysts. Support or Funding Information NSF under the CCI Center for Selective C–H Functionalization (CHE-1205646 and CHE-1700982), NIH (R01-GM078553, R35-GM118101, and T32-GM008353), University of Michigan Rackham Predoctoral Fellowship Physiological reactions catalyzed by P450s MycCI and TylHI in their respective biosynthetic pathways. This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Synthetic Escherichia coli-Corynebacterium glutamicum consortia for l-lysine production from starch and sucrose

In the biorefinery concept renewable feedstocks are converted to a multitude of value-added compounds irrespective of seasonal or other variations of the complex biomass substrates. Conceptionally, this can be realized by specialized single microbial strains or by co-culturing various strain combinations. In the latter approach strains for substrate conversion and for product formation can be combined. This study addressed the construction of binary microbial consortia based on starch- and sucrose-based production of l-lysine and derived value-added compounds. A commensalism-based synthetic consortium for l-lysine production from sucrose was developed combining an l-lysine auxotrophic, naturally sucrose-negative E. coli strain with a C. glutamicum strain able to produce l-lysine that secretes fructose when grown with sucrose due to deletion of the fructose importer gene ptsF. Mutualistic synthetic consortia with an l-lysine auxotrophic, α-amylase secreting E. coli strain and naturally amylase-negative C. glutamicum strains was implemented for production of valuable fine chemicals from starch.

An Iron Complex with a Bent, O‐Coordinated CO2 Ligand Discovered by Femtosecond Mid‐Infrared Spectroscopy

AbstractThe activation of carbon dioxide by transition metals is widely recognized as a key step for utilizing this greenhouse gas as a renewable feedstock for the sustainable production of fine chemicals. However, the dynamics of CO2 binding and unbinding to and from the ligand sphere of a metal have never been observed in the time domain. The ferrioxalate anion is used in aqueous solution as a unique model system for these dynamics and femtosecond UV‐pump mid‐infrared‐probe spectroscopy is applied to explore its photoinduced primary processes in a time‐resolved fashion. Following optical excitation, a neutral CO2 molecule is expelled from the complex within about 500 fs to generate a highly intriguing pentacoordinate ferrous dioxalate that carries a bent carbon dioxide radical anion ligand, that is, a reductively activated form of CO2, which is end‐on‐coordinated to the metal center by one of its two oxygen atoms.

An Iron Complex with a Bent, O-Coordinated CO2 Ligand Discovered by Femtosecond Mid-Infrared Spectroscopy.

The activation of carbon dioxide by transition metals is widely recognized as a key step for utilizing this greenhouse gas as a renewable feedstock for the sustainable production of fine chemicals. However, the dynamics of CO2 binding and unbinding to and from the ligand sphere of a metal have never been observed in the time domain. The ferrioxalate anion is used in aqueous solution as a unique model system for these dynamics and femtosecond UV-pump mid-infrared-probe spectroscopy is applied to explore its photoinduced primary processes in a time-resolved fashion. Following optical excitation, a neutral CO2 molecule is expelled from the complex within about 500 fs to generate a highly intriguing pentacoordinate ferrous dioxalate that carries a bent carbon dioxide radical anion ligand, that is, a reductively activated form of CO2 , which is end-on-coordinated to the metal center by one of its two oxygen atoms.

Discovery, Characterisation, Engineering and Applications of Ene Reductases for Industrial Biocatalysis.

Recent studies of multiple enzyme families collectively referred to as ene-reductases (ERs) have highlighted potential industrial application of these biocatalysts in the production of fine and speciality chemicals. Processes have been developed whereby ERs contribute to synthetic routes as isolated enzymes, components of multi-enzyme cascades, and more recently in metabolic engineering and synthetic biology programmes using microbial cell factories to support chemicals production. The discovery of ERs from previously untapped sources and the expansion of directed evolution screening programmes, coupled to deeper mechanistic understanding of ER reactions, have driven their use in natural product and chemicals synthesis. Here we review developments, challenges and opportunities for the use of ERs in fine and speciality chemicals manufacture. The ER research field is rapidly expanding and the focus of this review is on developments that have emerged predominantly over the last 4 years.

Synthesis of HMF from fructose using Purolite® strong acid catalyst: Comparison between BTR and PBR reactor type for kinetics data acquisition

5-hydroxymethyl-2-furfural (HMF) is an important furanic compound that can be obtained from renewable raw materials. HMF has wide application in the industry, acting as an intermediate in the production of fine chemicals, polymer, biofuels, and pharmaceuticals. In this research, HMF was obtained from the catalytic fructose dehydration. Purolite® SGC650H, a gel-type strongly acidic resin, was applied as a catalyst using dimethylsulfoxide (DMSO) as a solvent. All experiments were carried out in a batch reactor (BTR) and a continuous packed bed reactor (PBR). The reactor performance data showed that increase in temperature resulted in higher HMF yield. In contrast, the higher the fructose concentration, the lower the HMF conversion. The HMF production rates in the PBR were higher because of the concentration of the catalyst. The rate constants were calculated by means of a pseudo-first-order reaction model, and the Arrhenius kinetic parameters obtained changed with regards to the reactants concentrations. The kinetic parameters displayed linear correlation in the Cremer-Constable diagram, which means that the system was under a kinetic compensation effect (KCE); therefore, the model was not suitable to represent the reaction. Langmuir-Hinshelwood (LHHW) models for rate equation were also tested. However all of them failed to adjust to the data, and thus a rate law was not determined. Due to the observed KCE, it was believed that there is at least a set of parallel dehydration reactions initiated from the different fructose isomers which has an influence on kinetic parameters.

Selective depolymerization of industrial lignin-containing stillage obtained from cellulosic bioethanol processing

Reductive depolymerization of lignin-containing stillage (LA lignin) from 2nd generation bioethanol industrial plants have been studied over two different families of catalysts: conventional porous Ru catalysts supported on ZrO2 or activated carbon, and 1-dimensional Ru catalysts supported on multi-walled carbon nanotubes (MWCNT). The lignin depolymerization degree using conventional porous catalysts resulted in values of 42 and 53%, for Ru/ZrO2 and Ru/C, respectively. Strikingly, when using 1D-supported catalysts (Ru/MWCNT and Ru/ZrO2/MWCNT), the extent of lignin depolymerization measured by 1H13C HSQC NMR, reached values of c.a. 80% of β-O-4 bond disappearance and a reduction of the average molecular weight from 3600 Da to ~ 1900 Da. The higher accessibility of the lignin agglomerates to the 1-D catalyst nanoparticles substantially improves the lignin depolymerization degree, which make this approach of great interest in the production of fine chemicals and fuel additives.

Catalytic Conversion of CO2 to Cyclic Carbonates through Multifunctional Zinc‐Modified ZSM‐5 Zeolite

The production of fine chemicals using CO2 as C1 building block through inexpensive heterogeneous catalysts with high efficiency under low pressure is challenging. Herein we propose for the first time the utilization of a multifunctional heterogeneous zinc‐modified HZSM‐5 (ZnHZSM‐5) catalyst for upgrading CO2 by incorporation into cyclic carbonates from CO2 and epoxides. Owing to the nice surface properties such as abundant Lewis acid, Brønsted acid and Lewis base sites, large surface area, and plenty of micropores, CO2 could be concentrated and well activated in ZnHZSM‐5 verified by CO2‐TPD, TG‐MS, etc. Meanwhile, the epoxides were also activated through metal center and hydrogen bond. Therefore, the reaction can easily assemble at the catalyst interface and show exceptional performance, affording the aimed products with high yield of up to 99% in the presence of commercial tetra‐n‐propylammonium bromide (90% in kilogram scale with 0.004 mol% ZnHZSM‐5 and 0.015 mol% nPr4NBr).