High Molar Yields Research Articles

Nitrite ion mitigates the formation of N-nitrosodimethylamine (NDMA) during chloramination of ranitidine

Ranitidine (RNT) has been an important tertiary amine precursor of N-nitrosodimethylamine (NDMA) in chlorine-based water treatment, due to reaction with monochloramine (NH2Cl) with exceptionally high molar yields up to 90%. This study examined the effects of nitrite ions (NO2−) on the kinetics of NDMA formation during the chloramination of RNT under variable concentrations of dissolved oxygen (DO, 0.7–7.5mg/L), RNT (5–30μM), NH2Cl (5–20mM), NO2− or NO3− (0–2mM) and pH (5.6–8.6). In the absence of the NO2−, the ultimate molar yield of NDMA after 6h of reaction was primarily influenced by [DO] and pH, while marginally affected by initial [RNT] and [NH2Cl]. A kinetic model, prepared in accordance with the reaction sequence of NDMA formation, suggested that the rate determining step was accelerated with increasing [NH2Cl]0, [DO], and pH. A Kinetic study together with ultra-performance liquid chromatography-quadrupole-time of flight mass spectrometer (UPLC-Q-TOF MS) and gas chromatography (GC)/TOF MS analyses in parallel demonstrated that the nitrite ion inhibited the nucleophilic substitution of the terminal amine on NH2Cl, and reduced the pseudo-steady state concentration of N-peroxyl radicals, significantly decreasing the ultimate yields of NDMA.

Paper-based diagnostics in the antigen-depletion regime: High-density immobilization of rcSso7d-cellulose-binding domain fusion proteins for efficient target capture

In this work, we report the development of a general strategy for enhancing the efficiency of target capture in immunoassays, using a bifunctional fusion protein construct which incorporates a substrate-anchoring moiety for the high-abundance immobilization of an antigen-binding domain. This approach was informed by the development of a pseudo first-order rate constant model, and tested in a paper-based assay format using a fusion construct consisting of an rcSso7d binding module and a cellulose-binding domain. These rcSso7d-CBD fusion proteins were solubly expressed and purified from bacteria in high molar yields, and enable oriented, high-density adsorption of the rcSso7d binding species to unmodified cellulose within a 30-second incubation period. These findings were validated using two distinct, antigen-specific rcSso7d variants, which were isolated from a yeast surface display library via flow cytometry. Up to 1.6 micromoles of rcSso7d-CBD was found to adsorb per gram of cellulose, yielding a volume-averaged binder concentration of up to 760μM within the resulting active material. At this molar abundance, the target antigen is captured from solution with nearly 100% efficiency, maximizing the attainable sensitivity for any given diagnostic system.

Combination of ester biosynthesis and \u03c9-oxidation for production of mono-ethyl dicarboxylic acids and di-ethyl esters in a whole-cell biocatalytic setup with Escherichia coli

BackgroundMedium chain length (C6–C12) α,ω-dicarboxylic acids (DCAs) and corresponding esters are important building blocks for the polymer industry. For DCAs of 12 carbon atoms and longer, a sustainable process based on monooxygenase catalyzed ω-oxidation of fatty-acids has been realized. For medium-chain DCAs with a shorter chain length however, such a process has not been developed yet, since monooxygenases poorly ω-oxidize medium-chain fatty acids (MCFAs). On the contrary, esterified MCFAs are ω-oxidized well by the AlkBGTHJ proteins from Pseudomonas putida GPo1.ResultsWe show that MCFAs can be efficiently esterified and subsequently ω-oxidized in vivo. We combined ethyl ester synthesis and ω-oxidation in one-pot, whole-cell biocatalysis in Escherichia coli. Ethyl ester production was achieved by applying acyl-CoA ligase AlkK and an alcohol acyltransferase, either AtfA or Eeb1. E. coli expressing these proteins in combination with the ω-oxidation pathway consisting of AlkBGTHJ, produced mono-ethyl DCAs directly from C6, C8 and C9 fatty acids. The highest molar yield was 0.75, for mono-ethyl azelate production from nonanoic acid. Furthermore, di-ethyl esters were produced. Diethyl suberate was produced most among the di-ethyl esters, with a molar yield of 0.24 from octanoic acid.ConclusionThe results indicate that esterification of MCFAs and subsequent ω-oxidation to mono-ethyl DCAs via whole-cell biocatalysis is possible. This process could be the first step towards sustainable production of medium-chain DCAs and medium-chain di-ethyl esters.

Dynamic consideration of smog chamber experiments

Abstract. Recent studies of the α-pinene + ozone reaction that address particle nucleation show relatively high molar yields of highly oxidized multifunctional organic molecules with very low saturation concentrations that can form and grow new particles on their own. However, numerous smog-chamber experiments addressing secondary organic aerosol (SOA) mass yields, interpreted via equilibrium partitioning theory, suggest that the vast majority of SOA from α-pinene is semivolatile. We explore this paradox by employing a dynamic volatility basis set (VBS) model that reproduces the new-particle growth rates observed in the CLOUD experiment at CERN and then modeling SOA mass yield experiments conducted at Carnegie Mellon University (CMU). We find that the base-case simulations do overpredict observed SOA mass but by much less than an equilibrium analysis would suggest; this is because delayed condensation of vapors suppresses the apparent mass yields early in the chamber experiments. We further find that a second VBS model featuring substantial oligomerization of semivolatile monomers can match the CLOUD growth rates with substantially lower SOA mass yields; this is because the lighter monomers have a higher velocity and thus a higher condensation rate for a given mass concentration. The oligomerization simulations are a closer match to the CMU experiments than the base-case simulations, though they overpredict the observations somewhat. However, we also find that if the chemical conditions in CLOUD and the CMU chamber were identical, substantial nucleation would have occurred in the CMU experiments when in fact none occurred. This suggests that the chemical mechanisms differed in the two experiments, perhaps because the high oxidation rates in the SOA formation experiments led to rapid termination of peroxy radical chemistry.

Thermal Cycling Cascade Biocatalysis of myo-Inositol Synthesis from Sucrose

myo-Inositol belongs to the vitamin B group (vitamin B8) and is widely used in the drug, cosmetic, and food and feed industries. It is produced by acid hydrolysis of phytate, but this method suffers from costly feedstock and serious phosphorus pollution. Here a four-enzyme pathway containing thermophilic sucrose phosphorylase, phosphoglucomutase, inositol 1-phosphate synthase, and inositol monophosphatase was designed to convert sucrose to inositol and fructose. To enable the use of enzymes with different optimal temperatures and thermostabilities, we developed a thermal cycling cascade biocatalysis that can selectively add less-thermostable sucrose phosphorylase immobilized on cellulose-containing magnetic nanoparticles into the cold enzyme cocktail or remove it from the hot enzyme cocktail by using a magnetic field (ON/OFF) switch. A series of exergonic reactions push the overall reaction forward, resulting in a high product molar yield (0.98 mol of inositol/mol of sucrose). This cascade biocatalysis pl...

Isolation of six novel 7-oxo- or urso-type secondary bile acid-producing bacteria from rat cecal contents

Understanding the dynamics of secondary bile acid (SBA) formation in the gut by SBA-producing bacteria is important for host health, as SBAs have been shown to affect host pathophysiology and gut microbiota composition. However, our knowledge of SBA producers is limited in light of the diversity of gut microbes. Here, we isolated six novel SBA-producing bacteria from rat cecal contents, all of which were members of known species of gut microbes. Anaerostipes caccae D10, Bacteroides nordii C5, Clostridioides difficile D7, and Clostridium cadaveris G11 were capable of oxidizing cholic acid and chenodeoxycholic acid into 7-oxo-derivatives with varying yields. B.nordii C5 and its type strain JCM 12987T had the highest molar yield, ∼90%. Clostridium disporicum F4 and Clostridium subterminale C4 epimerized cholic acid into ursocholic acid with yields of ∼85%; the corresponding type strains lacked epimerization activity. Furthermore, although not novel as an SBA producer, Clostridium scindens G10 that produced deoxycholic acid from cholic acid was isolated for the first time from rodents. These findings will contribute to elucidation of SBA formation in the gut.

Enhanced biohydrogen production from corn stover by the combination of Clostridium cellulolyticum and hydrogen fermentation bacteria

Hydrogen was produced from steam-exploded corn stover by using a combination of the cellulolytic bacterium Clostridium cellulolyticum and non-cellulolytic hydrogen-producing bacteria. The highest hydrogen yield of the co-culture system with C. cellulolyticum and Citrobacter amalonaticus reached 51.9L H2/kg total solid (TS). The metabolites from the co-culture system were significantly different from those of the mono-culture systems. Formate, which inhibits the growth of C.cellulolyticum, could be consumed by the hydrogen-evolving bacteria, and transformed into hydrogen. Glucose and xylose were released from corn stover via hydrolysis by C.cellulolyticum and were quickly utilized in dark fermentation with the co-cultured hydrogen-producing bacteria. Because the hydrolysis of corn stover by C.cellulolyticum was much slower than the utilization of glucose and xylose by the hydrogen-evolving bacteria, the sugar concentrations were always maintained at low levels, which favored a high hydrogen molar yield.

Metabolic Engineering of the Actinomycete Amycolatopsis sp. Strain ATCC 39116 towards Enhanced Production of Natural Vanillin.

The Gram-positive bacterium Amycolatopsis sp. ATCC 39116 is used for the fermentative production of natural vanillin from ferulic acid on an industrial scale. The strain is known for its outstanding tolerance to this toxic product. In order to improve the productivity of the fermentation process, the strain's metabolism was engineered for higher final concentrations and molar yields. Degradation of vanillin could be decreased by more than 90% through deletion of the vdh gene, which codes for the central vanillin catabolism enzyme, vanillin dehydrogenase. This mutation resulted in improvement of the final concentration of vanillin by more than 2.2 g/liter, with a molar yield of 80.9%. Further improvement was achieved with constitutive expression of the vanillin anabolism genes ech and fcs, coding for the enzymes feruloyl-coenzyme A (CoA) synthetase (fcs) and enoyl-CoA hydratase/aldolase (ech). The transcription of both genes was shown to be induced by ferulic acid, which explains the unwanted adaptation phase in the fermentation process before vanillin was efficiently produced by the wild-type cells. Through the constitutive and enhanced expression of the two genes, the adaptation phase was eliminated and a final vanillin concentration of 19.3 g/liter, with a molar yield of 94.9%, was obtained. Moreover, an even higher final vanillin concentration of 22.3 g/liter was achieved, at the expense of a lower molar yield, by using an improved feeding strategy. This is the highest reported vanillin concentration reached in microbial fermentation processes without extraction of the product. Furthermore, the vanillin was produced almost without by-products, with a molar yield that nearly approached the theoretical maximum. Much effort has been put into optimization of the biotechnological production of natural vanillin. The demand for this compound is growing due to increased consumer concerns regarding chemically produced food additives. Since this compound is toxic to most organisms, it has proven quite difficult to reach high concentrations and molar yields. This study shows that improvements in the final vanillin concentrations and molar yields can be made through a combination of modification of the fermentation parameters and molecular strain engineering, without the need for methods such as continuous extraction from the fermentation broth. Using this approach, we were able to reach a final vanillin concentration of 22.3 g/liter, which is the highest vanillin concentration reported to date that was generated with Amycolatopsis sp. ATCC 39116 without additional extraction of the toxic product.

A continuous diethanolamine dehydrogenation fixed bed catalyst and reactor system

We have developed a fixed bed Cu/Co/ZrO2 catalyst and reactor system for the production of disodiumiminodiacetic acid by dehydrogenation of diethanolamine. This process can give high molar yields (>98%) in a series of adiabatic reactors with interstage heating. This research and development program proceeded in three stages: catalyst discovery, reactor design, and process development. The discovery of a catalyst formulation that is superior in activity and selectivity to sponge-metal copper enabled project success. Subsequent reactor scale-up proceeded based on laboratory scale measurements of the reaction kinetics and catalyst lifetime. Validation experiments demonstrated the importance of using feed compositions and operating conditions in the laboratory that are identical to those expected in the full-scale operation.

Ethanol production from biodiesel-derived crude glycerol by newly isolated Kluyvera cryocrescens

The rapidly expanding market for biodiesel has increased the supply and reduced the cost of glycerol, making it an attractive sustainable feed stock for the fuel and chemical industry. Glycerol-based biorefinery is the microbial fermentation of crude glycerol to produce fuels and chemicals. A major challenge is to obtain microbes tolerant to inhibitors such as salts and organic solvents present in crude glycerol. Microbial screening was attempted to isolate novel strain capable of growing on crude glycerol as a sole carbon source. The newly isolated bacteria, identified as nonpathogenic Kluyvera cryocrescens S26 could convert biodiesel-derived crude glycerol to ethanol with high yield and productivity. The supplementation of nutrients such as yeast extract resulted in distinguished enhancement in cell growth as well as ethanol productivity under anaerobic condition. When glycerol fermentation is performed under microaerobic condition, there is also a remarkable improvement in cell growth, ethanol productivity and yield, compared with those under strict anaerobic condition. In batch fermentation under microaerobic condition, K. cryocrescens S26 produced 27 g/l of ethanol from crude glycerol with high molar yield of 80% and productivity of 0.61 g/l/h.

Biohydrogen production from untreated and hydrolyzed potato steam peels by the extreme thermophiles Caldicellulosiruptor saccharolyticus and Thermotoga neapolitana

Production of hydrogen by the extreme thermophiles Caldicellulosiruptor saccharolyticus and Thermotoga neapolitana was studied in serum flasks and in pH-controlled bioreactors with glucose, and hydrolyzed and untreated potato steam peels (PSP) as carbon sources. Two types of PSP hydrolysates were used: one in which the starch in the PSP was liquefied with alpha-amylase, and one in which the liquefied starch was further hydrolyzed to glucose by amyloglucosidase. When the PSP hydrolysates or untreated PSP were added at circa 10–14 g/L of glucose units, both strains grew well and produced hydrogen with reasonable to high molar yields (2.4–3.8 moles H 2/mole glucose units), and no significant production of lactate. The hydrogen production rates and yields were similar with untreated PSP, hydrolyzed PSP, and pure glucose, showing that C. saccharolyticus and T. neapolitana are well equipped for the utilization of starch. When the concentrations of the substrates were increased, growth and hydrogen production of both strains were hampered. At substrate concentrations of circa 30–40 g/L of glucose units, the molar hydrogen yield of C. saccharolyticus was severely reduced due to the formation of high amounts of lactate, while T. neapolitana was unable to grow at all. The results showed that PSP and PSP hydrolysates are very suitable substrates for efficient fermentative hydrogen production at moderate substrate loadings.

Disinfection by-products from halogenation of aqueous solutions of terpenoids

We report the formation of trihalomethanes and other disinfection by-products from four polyfunctional terpenoids during simulated chlorination of natural waters. Complex suites of products were identified by closed loop stripping analysis (CLSA)/gas chromatography–mass spectrometry (GC–MS) from halogenation of β-carotene and retinol. β-Ionone appeared to be a key intermediate in the halogenation of β-carotene and retinol, reacting further under the reaction conditions to produce trans-β-ionone-5,6-epoxide and β-cyclocitral. Halogenation of the four terpenoids also produced trihalomethanes (THMs), most likely through haloform reaction on methyl ketone groups within many of the intermediates. Since halogenation of retinol produced a significant quantity of THMs at a slow reaction rate, retinol-based structures may possibly contribute to the slow reacting phase of THM formation in natural waters. Two polyhydroxyphenol model compounds were halogenated for comparison. The only products identified by CLSA/GC–MS from halogenation of 4′,5,7-trihydroxyflavanone and ellagic acid were THMs. 4′,5,7-Trihydroxyflavanone rapidly produced THMs, with an extremely high molar yield (94%) at pH 7. Terpenoids of the β-ionone and retinol type should be considered to be significant THM precursors, while 4′,5,7-trihydroxyflavanone has been shown to be an extremely significant THM precursor, potentially present within natural organic matter in water treatment processes and distribution systems.

Aqueous-phase ozonolysis of methacrolein and methyl vinyl ketone: a potentially important source of atmospheric aqueous oxidants

Abstract. Recent studies indicate that isoprene and its gas-phase oxidation products could contribute a considerable amount of aerosol through aqueous-phase acid-catalyzed oxidation with hydrogen peroxide (H2O2), although the source of H2O2 is unclear. The present study revealed a potentially important route to the formation of aqueous oxidants, including H2O2, from the aqueous-phase ozonolysis of methacrolein (MAC) and methyl vinyl ketone (MVK). Laboratory simulation was used to perform the atmospheric aqueous-phase ozonolysis at different pHs and temperatures. Unexpectedly high molar yields of the products, including hydroxylmethyl hydroperoxide (HMHP), formaldehyde (HCHO) and methylglyoxal (MG), of both of these reaction systems have been seen. Moreover, these yields are almost independent of pH and temperature and are as follows: (i) for MAC–O3, 70.3±6.3% HMHP, 32.3±5.8% HCHO and 98.6±5.4% MG; and (ii) for MVK–O3, 68.9±9.7% HMHP, 13.3±5.8% HCHO and 75.4±7.9% MG. A yield of 24.2±3.6% pyruvic acid has been detected for MVK–O3. HMHP is unstable in the aqueous phase and can transform into H2O2 and HCHO with a yield of 100%. We suggest that the aqueous-phase ozonolysis of MAC and MVK can contribute a considerable amount of oxidants in a direct and indirect mode to the aqueous phase and that these compounds might be the main source of aqueous-phase oxidants. The formation of oxidants in the aqueous-phase ozonolysis of MAC and MVK can lead to substantial aerosol formation from the aqueous-phase acid-catalyzed reaction of H2O2 with MAC, even if there are no other sources of oxidants.

Continuous production of biosurfactant with foam fractionation

AbstractA glucose‐limited chemostat was used to determine the growth parameters of BBK006 for continuous production of the biosurfactant surfactin. The continuous cultivation exhibited low maintenance metabolism (m = 0.39 mmolglucose gbacteria−1 h−1) and high molar growth yield ($Y_{x/S}^{\max} = 30.8$ gbacteria molglucose−1). It was found that the surfactin production rate in continuous culture was not only a function of dilution rate but also varied with the initial concentration of glucose in the feed. A high steady state concentration of surfactin (18 mg L−1) was maintained in the culture at a dilution rate of 0.2 h−1 when glucose concentration in the feed was 0.25 g L−1. This is the first demonstration of continuous surfactin production and recovery using glucose as a carbon source. The production of surfactin is known to be related to the age of the microorganisms and a simple mathematical model has been constructed to show how the age‐related production can be quantified. Copyright © 2006 Society of Chemical Industry

Enhanced anaerobic biotransformation of carbon tetrachloride with precursors of vitamin B12 biosynthesis

Relatively low concentrations of Vitamin B(12) are known to accelerate the anaerobic biotransformation of carbon tetrachloride (CT) and chloroform (CF). However, the addition of vitamin B(12) for field-scale bioremediation is expected to be costly. The present study considered a strategy to generate vitamin B(12) by addition of biosynthetic precursors. One of the precursors, porphobilinogen (PB) involved in the formation of the corrin ring, significantly increased the CT biotransformation rates by 2.7-, 8.8- and 10.9-fold when supplemented at 160, 500 and 900 microM, respectively. A positive control with 10 microM of vitamin B(12) resulted in a 5.9-fold increase in the CT-bioconversion rate. PB additions provided high molar yields of inorganic chloride (57% of CT organochlorine), comparable to that obtained with vitamin B(12) supplemented cultures. The primary substrate, methanol, known to induce vitamin B(12) production in methanogens and acetogens, was required for PB to have a significant impact on CT conversion. The observation suggests that PB's role was due to stimulating vitamin B(12) biosynthesis. The present study therefore provides insights on how to achieve vitamin B(12) enhanced rates of CT bioremediation through the use of less complex compounds that are precursors of vitamin B(12). Although PB is a costly chemical, its large impact points to corrin ring formation as the rate-limiting step.

Mechanistic evidences of the synthesis of methyl formate from methane and air on oxide catalysts

The direct conversion of methane into methyl formate with high molar yields (>12%) by a two reactor system has been attained. The reaction sequence consists of the partial oxidation of methane to HCHO at 600–700°C (MPO) on a “precipitated” SiO 2 catalyst and the subsequent dimerization of HCHO to methyl formate (FD) at 130–170°C on various oxide systems (TiO 2, SiO 2, ZrO 2, SO 4 2−/ZrO 2). FTIR characterization of the oxide catalysts allowed to highlight the superior performance of amphoteric oxides like TiO 2 and ZrO 2 in the FD reaction arising from the presence of surface acid and base sites playing a synergetic role in the reaction pathway.

Regiospecific glycosidase-assisted synthesis of lacto-N-biose I (Galbeta1-3GlcNAc) and 3'-sialyl-lacto-N-biose I (NeuAcalpha2-3Galbeta1-3GlcNAc).

The all-transglycolytic synthesis of lacto-N-biose I (Galbeta1-3GlcNAc) and 3'-sialyl-lacto-N-biose I (NeuAcalpha2-3Galbeta1-3GlcNAc) was performed. The disaccharide lacto-N-biose I was obtained by use of p-nitrophenyl beta-D-galactopyranoside as the donor, 2-acetamido-2-deoxy-D-glucopyranose as the acceptor and Xanthomonas manihotis beta-D-galactosidase as the catalyst. The reaction was shown to be regiospecific, with a high molar yield (about 55%) with respect to the donor. Lacto-N-biose I obtained by this method was used as the acceptor for a subsequent enzymatic reaction catalyzed by Trypanosoma cruzi trans-sialidase in which 2'-(4-methylumbellyferyl)-alpha-D-N-acetylneuraminic was used as the donor of the N-acetylneuraminil moiety. The reaction generated the product, 3'-sialyl-lacto-N-biose I, regiospecifically and with a molar yield of about 35%.

Reduction of aryl acids by white-rot fungi for the biocatalytic production of aryl aldehydes and alcohols.

Ligninolytic basidiomycetes were screened for their ability to reduce aryl acids to the corresponding aldehydes and alcohols. Seven fungal strains converted p-anisic acid in high molar yields to the reduced products. The white-rot fungus Bjerkandera sp. strain BOS55 was one of the best reducing strains and was highly tolerant towards high concentrations of different aromatic acids. It was tested for the reduction of p-anisic, veratric, 3-chloro-4-methoxybenzoic, 3,5-dichloro-4-methoxybenzoic, 3,4-dichlorobenzoic, 4-fluorobenzoic, and 3-nitrobenzoic acids. All of these compounds were reduced to their corresponding aldehydes and alcohols.

Stably Sustained Hydrogen Production with High Molar Yield through a Combination of a Marine Green Alga and a Photosynthetic Bacterium.

Stably sustained continuous production of hydrogen with high molar yield was achieved through a combination of dark fermentative hydrogen evolution by Chlamydomonas sp. strain MGA161 and hydrogen photoevolution by a marine photosynthetic bacterium W-1S in an alternating light-dark cycle as a model of the day-night cycle. The newly isolated strain W-1S could use acetic acid and ethanol excreted by strain MGA161 as electron donors for hydrogen photoevolution. The fermentation broth of strain MGA161 stimulated the hydrogen photoproduction of strain W-1S. This alga-bacterial combination had a high conversion yield of 8 mol H2/mol of glucose of starch, with the possibility of improvement up to 10.5.

Optimization of L‐malic acid production by Aspergillus flavus in a stirred fermentor

Effects of various nutritional and environmental factors on the accumulation of organic acids (mainly L-malic acid) by the filamentous fungus Aspergillus flavus were studied in a 16-L stirred fermentor. Improvement of the molar yield (moles acid produced per moles glucose consumed) of L-malic acid was obtained mainly by increasing the agitation rate (to 350 rpm) and the Fe(z+) ion concentration (to 12 mg/L) and by lowering the nitrogen (to 271 mg/L) and phosphate concentrations (to 1.5 mM) in the medium. These changes resulted in molar yields for L-malic acid and total C(4) acids (L-malic, succinic, and fumaric acids) of 128 and 155%, respectively. The high molar yields obtained (above 100%) are additional evidence for the operation of part of the reductive branch of the tricarboxylic acid cycle in L-malic acid accumulation by A. flavus. The fermentation conditions developed using the above mentioned factors and 9% CaCO(3) in the medium resulted in a high concentration (113 g/L L-malic acid from 120 g/L glucose utilized) and a high overall productivity (0.59 g/L h) of L-malic acid. These changes in acid accumulation coincide with increases in the activities of NAD(+)-malate dehydrogenase, fumarase, and citrate synthase.