Enantioselective vinylation of aldehydes with the vinyl Grignard reagent catalyzed by magnesium complex of chiral BINOLs

Enantioselective vinylation of aldehydes via direct catalytic asymmetric Grignard reaction of aldehdyes and the vinyl Grinard reagent is a long‐standing challenge. This work demonstrated that the magnesium (S)‐3,3′‐dimethyl BINOLate enantioselectively catalyze the direct vinylation of aldehydes with the deactivated vinylmagnesium bromide by bis(2‐[N,N′‐dimethylamino]ethyl) ether (BDMAEE) in the addition of n‐butylmagnesium chloride. The highest ee of 63% was achieved up to date.     

The Syntheses of new Camphorsulfonylated Ligands Derived from 2-Amino-2'-Hydroxy-1,1'-Binaphthyl and Their Enantioselectivities in the Addition of Dialkylzinc Reagents to Aldehydes

A series of new camphorsulfonylated ligands derived from chiral 2‐amino‐2′‐hydroxy‐1,1′‐binaphthyl (NOBIN) and (+)‐camphorsulfonic acid were synthesized by a short and simple synthetic sequence, and their enantioselective catalytic activities were assessed in the nucleophilic addition reaction of dialkylzinc reagents to aldehydes in the presence of titanium tetraisopropoxide. The most efficient ligand, N‐hydroxycamphorsulfonylated (S)‐NOBIN, gave (S)‐addition products with good yields and up to 87% of ee value. The ¹H nuclear magnetic resonance (NMR) and ¹³C NMR results of the titanium titration experiments on this ligand indicate that the most likely catalytic reactive species involved in this catalytic asymmetric addition is a bimetallic titanium complex. A possible catalytic reaction mechanism is proposed. Chirality 24:825–832, 2012. © 2012 Wiley Periodicals, Inc.     

Synthesis and application of 3‐substituted (S)‐BINOL as chiral ligands for the asymmetric ethylation of aldehydes

A series of (S)‐BINOL ligands substituted at the 3 position with some five‐membered nitrogen‐containing aromatic heterocycles were effectively prepared and their catalytic abilities were evaluated in the asymmetric addition of diethylzinc to benzaldehyde in the presence of titanium tetraisopropoxide. Under the optimized reaction conditions, titanium complex of (S)‐3‐(1H‐benzimidazol‐1‐yl)‐1,1′‐bi‐2‐naphthol was found to be the most efficient catalyst for asymmetric ethylation of various aldehydes to generate the corresponding secondary alcohols in up to 99% yield and 91% ee. Chirality, 2010. © 2010 Wiley‐Liss, Inc.     

Synthesis of 8‐Substituted 2′‐Deoxyisoguanosines via Unprotected 8‐Brominated 2‐Amino‐2′‐deoxyadenosine

A variety of applications of 8‐alkynylated nucleosides has prompted the synthesis of new purine analogues. Bromination of unprotected 2‐amino‐2′‐deoxyadenosine with Br₂/AcOH/AcONa gives 2‐amino‐8‐bromo‐2′‐deoxyadenosine (87%). The brominated derivative is converted to 8‐alkynylated 2‐amino‐2′‐deoxyadenosines by palladium‐catalyzed Sonogashira cross‐coupling reaction via microwave assistance (81 – 95%). The resulting compounds are further transformed to 8‐alkynylated 2′‐deoxyisoguanosines (52 – 70%). The physical properties of new compounds are investigated.     

Effect of ionic liquids activation on laccase from Trametes versicolor: Enzymatic stability and activity

The stability and activity of laccase from Trametes versicolor in two water‐soluble ionic liquids (ILs), namely 1‐butyl‐3‐methylimidazolium methyl sulfate, [bmim][MeSO₄] and 1,3‐dimethylimidazolium methyl sulfate, [mmim][MeSO₄], were investigated in this study. Thermal inactivation of laccase was characterized in the presence of these both ILs and as expected first‐order kinetics was followed. Inactivation rate constant (k), half‐life time (t_1/2), and energy of activation (Ea) were determined. Kinetics of 2,2′‐azino‐bis(3‐ethylbenzthiazoline‐6‐sulfonic acid) oxidation by laccase in the presence of these ILs was studied and Michaelis–Menten parameters were calculated. There is no enzymatic inactivation since the maximum reaction rate remained constant for IL concentrations up to 25%, and surprisingly, it was found that laccase was activated for concentrations of 35% of ILs, since the reaction rate increased 1.7 times.     

Regioselective hydroxylation of daidzein using P450 (CYP105D7) from Streptomyces avermitilis MA4680

Regiospecific 3′‐hydroxylation reaction of daidzein was performed with CYP105D7 from Streptomyces avermitilis MA4680 expressed in Escherichia coli. The apparent K_m and k_cat values of CYP105D7 for daidzein were 21.83 ± 6.3 µM and 15.01 ± 0.6 min^?1 in the presence of 1 µM of CYP105D7, putidaredoxin (CamB) and putidaredoxin reductase (CamA), respectively. When CYP105D7 was expressed in S. avermitilis MA4680, its cytochrome P450 activity was confirmed by the CO‐difference spectra at 450 nm using the whole cell extract. When the whole‐cell reaction for the 3′‐hydroxylation reaction of daidzein was carried out with 100 µM of daidzein in 100 mM of phosphate buffer (pH 7.5), the recombinant S. avermitilis grown in R2YE media overexpressing CYP105D7 and ferredoxin FdxH (SAV7470) showed a 3.6‐fold higher conversion yield (24%) than the corresponding wild type cell (6.7%). In a 7 L (working volume 3 L) jar fermentor, the recombinants S. avermitilis grown in R2YE media produced 112.5 mg of 7,3′,4′‐trihydroxyisoflavone (i.e., 29.5% conversion yield) from 381 mg of daidzein in 15 h. Biotechnol. Bioeng. 2010. 105: 697–704. © 2009 Wiley Periodicals.     

Nucleosides. IX. Synthesis of Purine N 3,5′‐Cyclonucleosides and N 3,5′‐Cyclo‐2′,3′‐seconucleosides via Mitsunobu Reaction as TIBO‐like Derivatives

The Mitsunobu reaction was applied to prepare, in one step, purine N ³,5′‐cyclonucleosides 10a–d. A subsequent ring opening in the ribose moiety of the resultant N ³,5′‐nucleosides by sodium periodate led to the corresponding N ³,5′‐cyclo‐2′,3′‐seconucleosides. These products consist of 5‐, 6‐, and 7‐membered tricyclic system which is the basic skeleton of TIBO derivatives, known antiviral agents.     

Mitsunobu Alkylation of Cancerostatic 5‐Fluorouridine with (2E)‐10‐Hydroxydec‐2‐enoic Acid,a Fatty Acid from Royal Jelly with Multiple Biological Activities

5‐Fluorouridine ( 1 ) – a nucleoside antimetabolite with strong cancerostatic properties – was protected i) at the 2′‐ and 3′‐OH groups with a heptan‐4‐ylidene residue and ii) at the 5′‐OH group with a (4‐methoxyphenyl)(diphenyl)methyl residue. This fully protected compound, 3 , was submitted to a Mitsunobu reaction with the N‐hydroxysuccinimide (NHS) ester, 5 , of (2E)‐10‐hydroxydec‐2‐enoic acid ( 4 ) which gave nucleolipid 6 . The latter was detritylated with Cl₂CHCOOH to yield the co‐drug 7 as NHS ester.     

Synthesis of New Potential Lipophilic Co‐Drugs of 2‐Chloro‐2′‐deoxyadenosine (Cladribine, 2‐CdA,Mavenclad®, Leustatin®) and 6‐Azauridine (z6U) with Valproic Acid

2‐Chloro‐2′‐deoxyadenosine (cladribine, 1 ) was acylated with valproic acid ( 2 ) under various reaction conditions yielding 2‐chloro‐2′‐deoxy‐3′,5′‐O‐divalproyladenosine ( 3 ) as well as the 3′‐O‐ and 5′‐O‐monovalproylated derivatives, 2‐chloro‐2′‐deoxy‐3′‐O‐valproyladenosine ( 4 ) and 2‐chloro‐2′‐deoxy‐5′‐O‐valproyladenosine ( 5 ), as new co‐drugs. In addition, 6‐azauridine‐2′,3′‐O‐(ethyl levulinate) ( 8 ) was valproylated at the 5′‐OH group (→ 9 ). All products were characterized by ¹H‐ and ¹³C‐NMR spectroscopy and ESI mass spectrometry. The structure of the by‐product 6 (N‐cyclohexyl‐N‐(cyclohexylcarbamoyl)‐2‐propylpentanamide), formed upon valproylation of cladribine in the presence of N,N‐dimethylaminopyridine and dicyclohexylcarbodiimide, was analyzed by X‐ray crystallography. Cladribine as well as its valproylated co‐drugs were tested upon their cancerostatic/cancerotoxic activity in human astrocytoma/oligodendroglioma GOS‐3 cells, in rat malignant neuro ectodermal BT4Ca cells, as well as in phorbol‐12‐myristate 13‐acetate (PMA)‐differentiated human THP‐1 macrophages. The most important result of these experiments is the finding that only the 3′‐O‐valproylated derivative 4 exhibits a significant antitumor activity while the 5′‐O‐ as well as the 3′,5′‐O‐divalproylated cladribine derivatives 3 and 5 proved to be inactive.     

Effective Anomerisation of 2′‐Deoxyadenosine Derivatives During Disaccharide Nucleoside Synthesis

The formation of a disaccharide nucleoside (11) by O3′‐glycosylation of 5′‐O‐protected 2′‐deoxyadenosine or its N ⁶‐benzoylated derivative has been observed to be accompanied by anomerisation to the corresponding α‐anomeric product (12). The latter reaction can be explained by instability of the N‐glycosidic bond of purine 2′‐deoxynucleosides in the presence of Lewis acids. An independent study on the anomerisation of partly blocked 2′‐deoxyadenosine has been carried out. Additionally, transglycosylation has been utilized in the synthesis of 3′‐O‐β‐d‐ribofuranosyl‐2′‐deoxyadenosines and its α‐anomer.     

A kinetic study on the Novozyme 435‐catalyzed esterification of free fatty acids with octanol to produce octyl esters

Octyl esters can serve as an important class of biolubricant components replacing their mineral oil counterparts. The purpose of the current work was to investigate the enzymatic esterification reaction of free fatty acids (FFA, from waste cooking oil) with octanol in a solvent‐free system using a commercial lipase Novozyme 435. It was found that the esterificaton reaction followed the Ping‐pong bi‐bi kinetics with no inhibition by substrates or products within the studied concentration range. The maximum reaction rate was estimated to be 0.041 mol L^?1 g^?1 h^?1. Additionally, the stability of Novozyme 435 in the current reaction system was studied by determining its activity and final conversion of FFA to esters after 12 successive utilizations. Novozyme 435 exhibited almost 100% enzyme activity up to 7 cycles of reaction and gradually decreased (by 5%) thereafter. The kinetic parameters evaluated from the study shall assist in the design of reactors for large‐scale production of octyl esters from a cheap biomass source. The enzyme reusability data can further facilitate mass production by curtailing the cost of expensive enzyme consumption. © 2015 American Institute of Chemical Engineers Biotechnol. Prog., 31:1494–1499, 2015     

Synthesis of D‐Altritol Nucleosides with a 3′‐O‐Tert‐Butyldimethylsilyl Protecting Group

Four D‐altritol nucleosides with a 3′‐O‐tert‐butyldimethylsilyl protecting group are synthesized (base moieties are adenine, guanine, thymine and 5‐methylcytosine). The nucleosides are obtained by ring opening reaction of 1,5:2,3‐dianhydro‐4,6‐O‐benzylidene‐D‐allitol. Optimal reaction circumstances (NaH, LiH, DBU, phase transfer, microwave irridation) for the introduction of the heterocycles are base‐specific. For the introduction of the 3′‐O‐silyl protecting group, long reaction times and several equivalents of tert‐butyldimethylsilyl chloride are needed.     

Phosphorylation of Isocarbostyril‐ and Difluorophenyl‐Nucleoside Thymidine Mimics by the Human Deoxynucleoside Kinases

The thymidine mimics isocarbostyril nucleosides and difluorophenyl nucleosides were tested as deoxynucleoside kinase substrates using recombinant human cytosolic thymidine kinase (TK1) and deoxycytidine kinase (dCK), and mitochondrial thymidine kinase (TK2) and deoxyguanosine kinase (dGK). The isocarbostyril nucleoside compound 1‐(2‐deoxy‐β‐D‐ribofuranosyl)‐isocarbostyril (EN1) was a poor substrate with all the enzymes. The phosphorylation rates of EN1 with TK1 and TK2 were < 1% relative to Thd, where as the phosphorylation rates for EN1 were 1.4% and 1.1% with dCK and dGK relative to dCyd and dGuo, respectively. The analogue 1‐(2‐deoxy‐β‐D‐ribofuranosyl)‐7‐iodoisocarbostyril (EN2) showed poor relative‐phosphorylation efficiencies (k _cat /K _m ) with both TK1 and dGK, but not with TK2. The k _cat /K _m value for EN2 with TK2 was 12.6% relative to that for Thd. Of the difluorophenyl nucleosides, 5‐(1′‐(2′‐deoxy‐β‐D‐ribofuranosyl))‐2,4‐difluorotoluene (JW1) and 1‐(1′‐(2′‐deoxy‐β‐D‐ribofuranosyl))‐2,4‐difluoro‐5‐iodobenzene (JW2) were substrates for TK1 with phosphorylation efficiencies of about 5% relative to that for Thd. Both analogues were considerably more efficient substrates for TK2, with k _cat /K _m values of 45% relative to that for Thd. 2,5‐Difluoro‐4‐[1‐(2‐deoxy‐β‐L‐ribofuranosyl)]‐aniline (JW5), a L‐nucleoside mimic, was phosphorylated up to 15% as efficiently as deoxycytidine by dCK. These data provide a possible explanation for the previously reported lack of cytotoxicity of the isocarbostyril‐ and difluorophenyl nucleosides, but potential mitochondrial effects of EN2, JW1 and JW2 should be further investigated.     

Characterisation of two wheat enolase cDNA showing distinct patterns of expression in leaf and crown tissues of plants exposed to low temperature

Seasonal low temperature (LT) adversely affects growth of plants. The onset of LT in temperate zones also entails the process of cold acclimation, preparing the plants to withstand freezing temperatures. During this process of cold acclimation a number of physiological, biochemical and molecular changes occur. A differentially expressed enolase gene in wheat plants exposed to LT was previously identified by cDNA‐amplified fragment length polymorphism. In this study, two wheat enolase cDNA, TaENO‐a and TaENO‐b amplified by 5′,3′ rapid amplification of cDNA end (RACE)‐PCR (polymerase chain reaction), were isolated and characterised. Quantitative real‐time PCR (QPCR) was done to assess their expression patterns in leaf and crown tissues of wheat plants exposed to LT. BLAST searches and bioinformatic analyses were done to determine the structure, domains and phylogeny of the cloned sequences. The two cDNA sequences differed mostly in the 5′ and 3′ untranslated regions. Deduced amino acid sequence showed high identity to bacteria, yeast, fungi, human and plant enolases with conserved putative DNA‐binding and repressor domains. A genomic clone containing 17 exons distributed over 4.5 kb structurally shared a high degree of similarity to rice enolase. QPCR revealed combined effects of LT and ageing on expression of TaENO‐a and TaENO‐b. Down‐regulation of TaENO‐a was observed with age in the crown tissues upon exposure to LT, but in leaf initial up‐regulation was followed by down‐regulation. Expression of TaENO‐b was similar to expression patterns previously reported for cold‐regulated (COR) genes in wheat, wherein the recessive vrnA‐1 allele influenced its expression in the leaf and genetic background determines its expression in the crown.     

Tetrathienoanthracene and Tetrathienylbenzene Derivatives as Hole‐Transporting Materials for Perovskite Solar Cell

The synthesis and characterization of two related families of star‐shaped thiophene‐containing hole‐transporting materials (HTMs) based on fused tetrathienoanthracene and nonfused tetrathienylbenzene cores are reported. All of them are endowed with four terminal (4,4′‐dimethoxy)diphenylamino groups that are either linked directly to the core or showed a different type of bridges (i.e., thiophene‐phenyl or phenyl rings). The novel HTMs are tested in mixed‐ion perovskite (Cs_0.1FA_0.74MA_0.13PbI_2.48Br_0.39) solar cells, and power conversion efficiencies of up to 18.8% are measured under 1 sun irradiation, comparable with the efficiency obtained for the reference cell using 2,2′,7,7′‐tetrakis(N,N′‐di‐p‐methoxyphenylamine)‐9,9′‐spirobifluorene as an HTM.     

(S)‐6‐Bromo‐BINOL‐based phosphoramidite ligand with C1 symmetry for enantioselective hydrogenation and allylic substitution

(S)‐6‐Br‐BINOL‐derived phosphoramidite, a s imple monodentate ligand with a stereogenic center at the phosphorus atom, was synthesized for the first time. This stereoselector generated a high level of enantioselectivity (80–95% ee) in the rhodium‐catalyzed hydrogenation of α‐dehydrocarboxylic acid esters and was also successfully employed in the asymmetric palladium‐catalyzed allylic substitution of (E)‐1,3‐diphenylallyl acetate. The optical yield also showed significant dependence with reaction type: up to 70% ee for allylic amination, up to 75% ee for allylic sulfonylation, and up to 90% ee for allylic alkylation. Chirality, 2010. © 2010 Wiley‐Liss, Inc.     

Kinetic Modeling of Amino Acid Oxidation Catalyzed by a New D‐Amino Acid Oxidase from Arthrobacter protophormiae

Amino acid oxidases, which enantiospecifically catalyze the oxidative deamination of either D‐ or L‐amino acids, belong to the class of oxidoreductases functioning with a tightly bound cofactor. This cofactor favors industrial applications of D‐amino acid oxidases (D‐AAO). Hence, the enzyme is very important for the industrial application in the purification and determination of certain amino acids. In developing the enzyme‐catalyzed reaction for large‐scale production, modeling of the reaction kinetics plays an important role. Therefore, the subject of this study was the kinetics of the oxidative deamination, a very complex reaction system, which is catalyzed by D‐AAO from Arthrobacter protophormiae using its natural substrate D‐methionine and the aromatic amino acid 3,4‐dihydroxyphenyl‐D‐alanine (D‐DOPA). The kinetic parameters determined by the measurement of the initial rate and nonlinear regression were verified in batch reactor experiments by comparing calculated and experimental concentration‐time curves. It was found that the enzyme is highly specific towards D‐methionine (K_m = 0.24 mM) and not as specific to D‐DOPA as a substrate (K_m = 9.33 mM). The enzyme activity towards D‐methionine ( = 3.01 U/mL) was approx. seven times higher than towards D‐DOPA ( = 20.01 U/mL). The enzyme exhibited no activity towards L‐methionine and L‐DOPA. Batch and repetitive batch experiments were performed with both substrates in the presence and in the absence of catalase for hydrogen peroxide decomposition. Their comparison made it possible to conclude that hydrogen peroxide has no negative influence on the enzyme activity.     

G‐quadruplex DNA‐based asymmetric catalysis of michael addition: Effects of sonication,ligands, and co‐solvents

There is an escalating interest of using double stranded DNA molecules as a chiral scaffold to construct metal‐biomacromolecule hybrid catalysts for asymmetric synthesis. Several recent studies also evaluated the use of G‐quadruplex DNA‐based catalysts for asymmetric Diels‐Alder and Friedel‐Crafts reactions. However, there is still a lack of understanding of how different oligonucleotides, salts (such as NaCl and KCl), metal ligands and co‐solvents affect the catalytic performance of quadruplex DNA‐based hybrid catalysts. In this study, we aim to systematically evaluate these key factors in asymmetric Michael addition reactions, and to examine the conformational and molecular changes of DNA by circular dichroism (CD) spectroscopy and gel electrophoresis. We achieved up to 95% yield and 50% enantiomeric excess (ee) when the reaction of 2‐acylimidazole 1a and dimethylmalonate was catalyzed by 5′‐G₃(TTAG₃)₃?3′ (G4DNA1) in 20 mM MOPS (pH 6.5) containing 50 mM KCl and 40 µM [Cu(dmbipy)(NO₃)₂], and G4DNA1 was pre‐sonicated in ice bath for 10 min prior to the reaction. G‐quadruplex‐based hybrid catalysts provide a new tool for asymmetric catalysis, but future mechanistic studies should be sought to further improve the catalytic efficiency. The current work presents a systematic study of asymmetric Michael addition catalyzed by G‐quadruplex catalysts constructed via non‐covalent complexing, and an intriguing finding of the effect of pre‐sonication on catalytic efficiency. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:891–898, 2016     

Synthesis and application of the first planar chiral strong Brønsted acid organocatalysts

The synthesis of planar chiral strong Brønsted acid organocatalysts derived from [2.2]paracyclophane is described. Resolution was accomplished according to a modified protocol involving pseudo‐ortho‐substituted [2.2]paracyclophane‐based sulfoxides for the synthesis of three new sulfonic acids. The first planar chiral phosphoric acid diester was obtained from the corresponding phenyl‐substituted diol derived from enantiopure 4‐bromo‐12‐hydroxy [2.2]paracyclophane. These new classes of catalysts were tested in an enantioselective Friedel–Crafts reaction as well as in a direct asymmetric Mannich reaction and gave yields of up to 93% and ee‐values of up to 38%. Chirality, 2012. © 2012 Wiley Periodicals, Inc.     

Evidence for the generation of myristylated FMN by bacterial luciferase

The genes responsible for the light production in bioluminescent bacteria are present as an operon, luxCDABEG. Many strains of Photobacteria carry an additional gene, termed luxF. X‐ray crystallographic analysis of LuxF revealed the presence of four molecules of a flavin derivative, i.e. 6‐(3′‐(R)‐myristyl) flavin adenine mononucleotide (myrFMN) non‐covalently bound to the homodimer. In the present study, we exploited the binding of myrFMN to recombinant apo‐LuxF to explore the occurrence of myrFMN in various bioluminescent bacteria. MyrFMN was detected in all bacterial strains tested including Vibrio and Aliivibrio indicating that it is more widely occurring in bioluminescent bacteria than previously assumed. We also show that apo‐LuxF captures myrFMN and thereby relieves the inhibitory effect on luciferase activity. Thus our results provide support for the hypothesis that LuxF acts as a scavenger of myrFMN in bioluminescent bacteria. However, the source of myrFMN remained obscure. To address this issue, we established a cofactor regeneration enzyme‐catalyzed cascade reaction that supports luciferase activity in vitro for up to 3 days. This approach enabled us to unambiguously demonstrate that myrFMN is generated in the bacterial bioluminescent reaction. Based on this finding we postulate a reaction mechanism for myrFMN generation that is based on the luciferase reaction.