A tandem enzyme reaction to produce optically active halohydrins,epoxides and diols

The recombinant halohydrin dehalogenase from Agrobacterium radiobacter AD1 was used to obtain enantiomerically pure halohydrins and epoxides by kinetic resolution. By adding an excess of the recombinant epoxide hydrolase from the same organism the reversible conversion was drawn to completion. Halohydrins such as (S)-2,3-dichloro-1-propanol (E>100) and (S)-2-chloro-1-phenylethanol (E=73) were obtained with an enantiomeric excess of higher than 99%. This is a novel biocatalytic route for obtaining enantiomerically pure aromatic halohydrins and epoxides.     

Enantioconvergent hydrolysis of styrene epoxides by newly discovered epoxide hydrolases in mung bean

[reaction: see text] Two novel epoxide hydrolases were discovered in mung bean (Phaseolus radiatus L.) for the first time, either of which can catalyze enantioconvergent hydrolysis of styrene epoxides. Their regioselectivity coefficients are more than 90% for the p-nitrostyrene oxide. Furthermore, the crude mung bean powder was also shown to be a cheap and practical biocatalyst, allowing a one-step asymmetric synthesis of chiral (R)-diols from racemic epoxides, in up to >99% ee and 68.7% overall yield (after recrystallization).     

Gene Cloning, Expression, and Characterization of a Thermostable Xylanase from Nesterenkonia xinjiangensis CCTCC AA001025

An endo-β-1,4-xylanase-encoding gene, xyn11NX, was cloned from Nesterenkonia xinjiangensis CCTCC AA001025 and expressed in Escherichia coli. The gene encoded a 192-amino acid polypeptide and a putative 50-amino acid signal peptide. The deduced amino acid sequence exhibited a high degree of similarity with the xylanases from Streptomyces thermocyaneoviolaceus (68%) and Thermobifida fusca (66%) belonging to glycoside hydrolase family 11. After purification to homogeneity, the recombinant Xyn11NX exhibited optimal activity at pH 7.0 and 55 °C and remained stable at weakly acidic to alkaline pH (pH 5.0–11.0). The enzyme was thermostable, retaining more than 80% of the initial activity after incubation at 60 °C for 1 h and more than 40% of the activity at 90 °C for 15 min. The K _m and V _max values for oat spelt xylan and birchwood xylan were 16.08 mg ml^?1 and 45.66 μmol min^?1 mg^?1 and 9.22 mg ml^?1 and 16.05 μmol min^?1 mg^?1, respectively. The predominant hydrolysis products were xylobiose and xylotriose when using oat spelt xylan or birchwood xylan as substrate.     

Enantioselective Hydrolysis of Glycidyl Methylphenyl Ethers by <Emphasis Type="Italic">Botryosphaeria Dothidea</Emphasis> ZJUZQ007: Effect of Substitution Pattern on Enantioselectivity

Resolution of racemic glycidyl (o, m, p)-methylphenyl ethers by using a newly isolated Botryosphaeria dothidea ZJUZQ007 with epoxide hydrolase activity affords enantiopure epoxides with enantiomeric excesses (e.e._s) of 91–99% and enantiomeric ratios (E) of 18.1 to 48.6. The (R)-enantiomer was obtained from rac-glycidyl (o or m)-methylphenyl ethers whereas the (S)-epoxides was obtained from glycidyl p-methylphenyl ether. Substitution pattern of the methyl group exerted effects both on configurations of the remaining epoxides and enantioselectivities of epoxide hydrolase. The observations were explained by enzyme-substrate docking studies. It is the first example showing that for kinetic resolution of glycidyl methylphenyl ethers, fungal species of B. dothidea was applied.     

Deracemization of (±)-cis-dialkyl substituted oxides via enantioconvergent hydrolysis catalysed by microsomal epoxide hydrolase

Both enantiomers of cis-(±)-2,3-epoxyheptane 1a, cis-3,4-epoxyheptane 1b, cis-3,4-epoxynonane 1c, cis-3,4-epoxynonane-1-ol 1d, and cis-1-methoxy-3,4-epoxynonane 1e undergo a highly stereoselective microsomal epoxide hydrolase catalysed hydration at the (S) carbon to give the corresponding threo (R,R)-diol at complete conversion. A total kinetic resolution of racemic epoxides is also obtained with 1a and 1e.     

Investigation of the catalytic activity of an electron-deficient vanadium(IV) tetraphenylporphyrin: A new, highly efficient and reusable catalyst for ring-opening of epoxides

In this work, the catalytic activity of high-valent tetraphenylporphyrinatovanadium(IV) trifluoromethanesulfonate, [V^IV(TPP)(OTf)₂], in the nucleophilic ring-opening of epoxides is reported. This new V(IV) catalyst was used as an efficient catalyst for alcoholysis with primary (methanol, ethanol and n-propanol), secondary (iso-propanol) and tertiary alcohols (tert-butanol), hydrolysis and acetolysis of epoxides with acetic acid and also for the conversion of epoxides to 1,2-diacetates with acetic anhydride, conversion of epoxides to thiiranes with ammonium thiocyanate and thiourea, and for conversion of epoxides to acetonides with acetone. The catalyst was reused several times without loss of its activity.     

Theoretical study of the full reaction mechanism of human soluble epoxide hydrolase

The complete reaction mechanism of soluble epoxide hydrolase (sEH) has been investigated by using the B3LYP density functional theory method. Epoxide hydrolases catalyze the conversion of epoxides to their corresponding vicinal diols. In our theoretical study, the sEH active site is represented by quantum-chemical models that are based on the X-ray crystal structure of human soluble epoxide hydrolase. The trans-substituted epoxide (1S,2S)-beta-methylstyrene oxide has been used as a substrate in the theoretical investigation of the sEH reaction mechanism. Both the alkylation and the hydrolytic half-reactions have been studied in detail. We present the energetics of the reaction mechanism as well as the optimized intermediates and transition-state structures. Full potential energy curves for the reactions involving nucleophilic attack at either the benzylic or the homo-benzylic carbon atom of (1S,2S)-beta-methylstyrene oxide have been computed. The regioselectivity of epoxide opening has been addressed for the two substrates (1S,2S)-beta-methylstyrene oxide and (S)-styrene oxide.     

巨大芽胞杆菌环氧水解酶的固定化及其催化性能

考察了多种载体对巨大芽孢杆菌ECU1001环氧水解酶的固定化.以大孔DEAE-纤维素离子交换树脂为载体时,固定化酶的活力回收达70%.进一步考察了温度和pH对固定化酶活力的影响,并使用该固定化酶进行了缩水甘油苯基醚对映选择性水解批次反应.结果表明,在较低的底物浓度下该固定化酶的稳定性较好,10批反应后仍然剩余72.4%的活力.     

Impact of Pretreatment with Dilute Sulfuric Acid Under Moderate Temperature on Hydrolysis of Corn Stover with Two Enzyme Systems

Pretreatment of corn stover with dilute sulfuric acid at moderate temperature was investigated, and glucan digestibility by Cellic CTec2 and Celluclast on the pretreated biomass was compared. Pretreatments were carried out from 60 to 180 min at the temperature from 105 to 135 °C, with acid concentrations ranging from 0.5 to 2 % (w/v). Significant portion of xylan was removed during pretreatment, and the glucan digestibility by CTec2 was significantly better than that by Celluclast in all cases. Analysis showed that glucan digestibility by both two enzymes correlated directly with the extent of xylan removal in pretreatment. Confidence interval was built to give a more precise range of glucan conversion and to test the significant difference among pretreatment conditions. Response surface model was built to obtain the optimal pretreatment condition to achieve high glucan conversion after enzymatic hydrolysis. Considering the cost and energy savings, the optimal pretreatment condition of 1.75 % acid for 160 min at 135 °C was determined, and glucan conversion can achieve the range from 72.86 to 76.69 % at 95 % confidence level after enzymatic hydrolysis, making total glucan recovery up to the range from 89.42 to 93.25 %.     

Facile synthesis of Ti-TUD-1 for catalytic oxidative desulfurization of model sulfur compounds

A facile synthesis route for Ti-TUD-1 at room temperature employing silatrane and titanium glycolate as Si and Ti sources (2?C8 mol%), respectively, over a triethanolamine template is proposed. XRD, N₂ adsorption?Cdesorption isotherms, and TEM analysis confirmed disordered mesoporous structures with high surface area (715?C824 m²/g). According to the UV?Cvisible spectroscopy of the calcined materials, titanium species of ca. 2.7 mol% Ti loading were present mostly in tetrahedral coordination for a sample prepared with 4 mol% Ti in the substrate mixture. Ti-TUD-1 showed catalytic activity in cyclohexene epoxidation, which depended on the amount of tetrahedrally coordinated Ti species. The hydrophilic nature of the surface of Ti-TUD-1 was confirmed by the effect of oxidant such that tert-butyl hydroperoxide (TBHP, 5?C6 M in decane) was superior to other oxidants in water (cyclohexene conversion: TBHP in decane 36.5% vs. TBHP in water 30.6%). Ti-TUD-1 was more active in oxidative desulfurization (ODS) reaction than Ti-MCM-41 at the same Ti loading; the former produced 4,6-dimethyldibenzothiophene (4,6-DMDBT) conversion near 100% after reacting for 15 min, whereas Ti-MCM-41 produced final conversion of 4,6-DMDBT of 89% after reacting for 180 min. ODS over Ti-TUD-1 was influenced both by electron density and steric hindrance in model sulfur compounds. Partially polymerized Ti sites seemed to also contribute to the reaction.     

A direct method for the conversion of terminal epoxides into gamma-butanolides

A new and efficient process for the conversion of terminal epoxides to gamma-butanolides is described involving Lewis acid promoted epoxide ring-opening by 1-morpholino-2-trimethylsilyl acetylene. Addition of a terminal epoxide to a solution of the ynamine and boron trifluoride diethyl etherate in dichloromethane at 0 degrees C rapidly affords a cyclic keteneaminal that can be hydrolyzed and protodesilylated under mild conditions to provide the corresponding gamma-butanolide in high yield. The net transformation is equivalent to an acetate enolate opening of terminal epoxides. The formation of a cyclic keteneaminal as the direct addition product was observed by monitoring of the reaction by IR and NMR spectroscopy. Functionalized gamma-lactones were prepared by the interception of the reactive cyclic keteneaminal prior to hydrolysis. Reactions with enantiomerically enriched terminal epoxides provide the corresponding gamma-butanolides without loss of optical activity. The compatibility of the present methodology with a wide range of functional groups is noteworthy.     

Triethanolamine/KI: A Multifunctional Catalyst for CO2 Activation and Conversion with Epoxides into Cyclic Carbonates

A catalytic system of triethanolamine/potassium iodide (KI) was proved to be efficient for the chemical fixation of CO₂ with epoxide. It was found that triethanolamine with dual function could activate both CO₂ and epoxides. Effects of parameters such as catalyst molar ratio and amount, reaction time, pressure, and temperature were studied systematically. As a result, 99% propylene oxide conversion as well as 99% propylene carbonate selectivity could be obtained under the optimal reaction condition. Furthermore, the catalyst was found to be applicable to a variety of terminal epoxides, providing the corresponding cyclic carbonates in good yields and selectivity. Moreover, the catalyst could be reused five times without loss of activity. This work presents an example of a cheap and efficient catalyst for the chemical fixation of CO₂ to high-value chemicals, which could help to improve the catalytic efficiency and decrease cost of products for larger applications.

[Supplementary materials are available for this article. Go to the publisher's online edition of Synthetic Communications® for the following free supplemental resource: Full experimental and spectral details.]     

Shifting the equilibrium of a biocatalytic cascade synthesis to enantiopure epoxides using anion exchangers

Hydroxide-loaded anion exchangers have been successfully employed to shift the equilibrium of a one-pot, two-step, two-enzyme cascade reaction affording enantiopure epoxides starting from prochiral α-chloroketones. The α-chloroketones were asymmetrically reduced employing an alcohol dehydrogenase and then transformed further to the corresponding epoxides employing halohydrin dehalogenases. Each epoxide enantiomer could be obtained with up to 93% conversion in enantiomerically pure form (>99% ee). In contrast to previous studies the amount of hydride donor (2-propanol) could be reduced due to favoured halohydrin formation in the reduction of α-chloroketones.     

A Two-Step,One-Pot Enzymatic Method for Preparation of Duck Egg White Protein Hydrolysates with High Antioxidant Activity

Biocatalytic hydrolysis reactions were designed for preparation of bioactive hydrolysate of duck egg white protein (DEWP) employing two enzymes in one pot. Firstly, the fresh DEWP was thermal treated at 95 °C, for 40 min at pH 10, to effectively deactivate enzyme inhibitors thus facilitating the following two-step enzymatic hydrolysis. Compared with single-enzyme processes, the two-step enzymatic procedures showed much higher reaction efficiency. The first enzymatic step (in the presence of Alcalase or hydrolase SEEP) allowed to hydrolyze DEWP with degree of hydrolysis (DH) of 8.8–10 % and soluble peptide yield (SEP) of 60.5–70.2 % in a short period (4 h). The second enzymatic step (in the presence of Trypsin or Alcalase) gave a further degradation of DEWP with DH and SEP being more than 26.2 % and 90.4 %, respectively. The final hydrolysates exhibited high antioxidant activity in an evident DH dependent manner. The hydrolysates achieved by sequential addition of the proteinase SEEP and Alcalase at DH value 21 % gave the highest antioxidant activity, which was mainly ascribed to the changes in the amino acid compositions that the contents of some key amino acids and total hydrophobic amino acids were significantly improved by the enzymatic hydrolysis.     

A Biomass-Ligand-Based Ru(III) Complex as a Catalyst for Cycloaddition of CO2 and Epoxides to Cyclic Carbonates and a Study of the Mechanism

Aiming highly efficient conversion of greenhouse gas CO₂ to cyclic carbonates, a biomass Ru(III) Schiff base complex catalyst ( SalRu ) was constructed by employing a derivative of Lignin degradation (5-aldehyde vanillin). The SalRu catalyst had a remarkable conversion for epoxides into corresponding cyclic carbonates even at atmospheric pressure of CO₂ without the presence of co-catalyst. As the condition at 120 °C and 2 MPa CO₂ the conversion reached to 94 % with selectivity at 99 % after 8 h. 32 % cyclic carbonate production was obtained even under 0.2 MPa CO₂ pressure. The epoxide activation and ring opening, CO₂ insertion and cyclic carbonate formation were illuminated explicitly through the of characteristic absorption peaks changing, which further providing direct and visual evidence for the mechanism proposing. This study has important theoretical significance for the comprehensive utilization of environmental pollutants and energy.     

Enantioselective trans dihydroxylation of nonactivated C-C double bonds of aliphatic heterocycles with Sphingomonas sp. HXN-200

The bacterial strain Sphingomonas sp. HXN-200 was used to catalyze the trans dihydroxylation ofN-substituted 1,2,5,6-tetrahydropyridines 1 and 3-pyrrolines 4 giving the corresponding 3,4-dihydroxypiperidines 3 and 3,4-dihydroxypyrrolidines 6, respectively, with high enantioselectivity and high activity. The trans dihydroxylation was sequentially catalyzed by a monooxygenase and an epoxide hydrolase in the strain with epoxide as intermediate. While both epoxidation and hydrolysis steps contributed to the overall enantioselectivity in trans dihydroxylation of 1, the enantioselectivity in trans dihydroxylation of the symmetric substrate 4 was generated only in the hydrolysis of meso-epoxide 5. The absolute configuration for the bioproducts (+)-3 and (+)-6 was established as (3R,4R) by chemical correlations. Preparative trans dihydroxylation of 1a and 4b with frozen/thawed cells of Sphingomonas sp. HXN-200 afforded the corresponding (+)-(3R,4R)-3,4-dihydroxypiperidine 3a and (+)-(3R,4R)-3,4-dihydroxy pyrrolidine 6b in 96% ee both and in 60% and 80% yield, respectively. These results represent first examples of enantioselective trans dihydroxylation with nonterpene substrates and with bacterial catalyst, thus significantly extending this methodology in practical synthesis of valuable and useful trans diols. Enantioselective hydrolysis of racemic epoxide 2a with Sphingomonas sp. HXN-200 gave 34% of (-)-2a in >99% ee, which is a versatile chiral building block. Further hydrolysis of (-)-2a with the same strain afforded (-)-(3S,4S)-3a in 96% ee and 92% yield. Thus, both enantiomers of 3a can be prepared by biotransformation with Sphingomonas sp. HXN-200.     

99mTc-labeling of a dithiocarbamate-DWAY fragment using [99mTcN]2+ core for the preparation of potential 5HT1A receptor imaging agents

1-(2-methoxy phenyl) piperazine fragment of WAY100635 or its phenolic analogue, derived from DWAY is used to design the desired structure of 5HT_1A receptor imaging agents. In this study a DWAY analogue was labeled with ^99mTc-nitrido ([^99mTcN]²⁺) core via dithiocarbamate. 2-(piperazin-1-yl) phenol dithiocarbamate was synthesized by the reaction of 2-(piperazin-1-yl) phenol with an equivalent amount of carbon disulfide in KOH solution then radiolabeled with ^99mTc-nitrido core. The final complex was characterized by HPLC and its radiochemical purity was more than 90 %. In vitro stability studies have shown the complex was stable at least 4 h after labeling at room temperature. The n-octanol/water partition coefficient experiment demonstrated log p = 1.34 for ^99mTcN–OHPP–DTC. Biodistribution results showed that radio tracer had moderate brain uptake (0.39 ± 0.03 %ID/g at 15 min and 0.29 ± 0.02 %ID/g at 120 min) and good retention, suggesting that this complex may lead to a further development of a radiotracer with specific binding to 5-HT_1A receptor.     

Epoxy-based monoliths for capillary liquid chromatography of small and large molecules

A versatile epoxy-based monolith was synthesised by polycondensation polymerisation of glycidyl ether 100 with ethylenediamine using a porogenic system consisting of polyethylene glycol, M _w?=?1000, and 1-decanol. Polymerisation was performed at 80 °C for 22 h. A simple acid hydrolysis of residual epoxides resulted in a mixed diol-amino chemistry. The modified column was used successfully for hydrophilic interaction liquid chromatography (HILIC) of small molecule probes such as nucleic acid bases and nucleosides, benzoic acid derivatives, as well as for peptides released from a tryptic digest of cytochrome c. The mixed-mode chemistry allowed both hydrophilic partitioning and ion-exchange (IEX) interactions to contribute to the separation, providing flexibility in selectivity control. Residual epoxide groups were also exploited for incorporating a mixed IEX chemistry. Alternatively, the surface chemistry of the monolith pore surface rendered hydrophobic via grafting of a co-polymerised hydrophobic hydrogel. The inherent hydrophilicity of the monolith scaffold also enabled high performance separation of proteins under IEX and hydrophobic interaction modes and in the absence of non-specific interactions.     

Stereoregular polycarbonate synthesis: Alternating copolymerization of CO2 with aliphatic terminal epoxides catalyzed by multichiral cobalt(III) complexes

Completely stereoregular polycarbonate synthesis was achieved with the use of unsymmetric multichiral cobalt‐based complexes bearing a derived chiral BINOL and an appended 1,5,7‐triabicyclo[4.4.0] dec‐5‐ene as catalyst for the copolymerization of CO₂ and aliphatic terminal epoxides at mild conditions. The (S,S,S)‐Co(III) complex 1c with sterically hindered substituent group is more stereoregular catalyst for the copolymerization of CO₂ and racemic propylene oxide to afford a perfectly regioregular poly(propylene carbonate) (PPC), with >99% head‐to‐tail linkages, >99% carbonate linkages, and a K_rel of 24.4 for the enchainment of (R)‐epoxide over (S)‐epoxide. The isotactic PPC exhibits an enhanced glass transition temperature of 47 °C, which is 10–12 °C higher than that of the corresponding irregular polycarbonate. © 2011 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem, 2011.     

Enantioselective synthesis of fatty acid amide hydrolase inhibitors with 1,3-disubstituted butan-2-one scaffold

Fatty acid amide hydrolase is a key enzyme in the inactivation of the analgesic and anti-inflammatory endocannabinoid anandamide. Previously, the chiral compound 1-(1H-benzotriazol-1-yl)-3-(4-phenylphenoxy)butan-2-one was identified as a potent inhibitor of fatty acid amide hydrolase and is therefore of interest as a potential agent against pain and inflammation. Two different approaches for the enantioselective synthesis of fatty acid amide hydrolase inhibitors with a 1,3-disubstituted butan-2-one scaffold were carried out. The first one uses the chiral epoxide 2-[1-(4-phenylphenoxy)ethyl]oxirane with an (R)- or (S)-configuration at the exocyclic stereocenter as central intermediates. These substances were obtained by separation of the non-stereoselectively synthesized epoxide into its racemic diastereomers by reversed phase chromatography followed by Jacobsen’s hydrolytic kinetic resolution of each enantiomer with the (S)-configured oxirane ring. Furthermore, a chiral pool based enantioselective synthesis was developed. In that case, the starting compound for both target enantiomers was methyl 3,4-O-isopropylidene-l-threonate. In comparison to the first approach, the chiral pool synthesis consisted of more steps, but generated the enantiomers with much better enantiomeric excess. Biological evaluation showed that the (R)-enantiomer inhibits isolated fatty acid amide hydrolase with a 200-fold higher activity than the (S)-enantiomer.