Oxygen Equilibrium Characteristics of Abnormal Hemoglobins: Hirose (α2β237Ser), L Ferrara (α247Glyβ2), Broussais (α290Asnβ2), and Dhofar (α2β258Arg)

The oxygen equilibrium characteristics of four structural variants of hemoglobin A were correlated with their amino acid substitutions.Hemoglobin Dhofar, in which the proline at E2(58)beta is replaced by arginine, had normal oxygen equilibrium characteristics.Hemoglobin L Ferrara. in which the aspartic acid at CD5(47)alpha is replaced by glycine, and hemoglobin Broussais, in which the lysine at FG2(90)alpha is replaced by asparagine, both showed a slightly elevated oxygen affinity; nevertheless both demonstrated a normal heme-heme interaction and a normal Bohr effect.Hemoglobin Hirose, in which the tryptophan at C3 (37)beta is replaced by serine, showed abnormalities of all oxygen equilibrium characteristics; i.e., increased oxygen affinity, diminished heme-heme interaction, and reduced Bohr effect.These results suggest that aspartic acid at CD5(47)alpha and lysine at FG2(90)alpha are involved in the function of the hemoglobin molecule, despite the fact that these positions are not located directly in the heme or the alpha-beta-contact regions.Tryptophan at C3(37)beta is located at contact between alpha(1)- and beta(2)-subunits. It is suggested that the substitution by serine might disturb the quarternary structure of the mutant hemoglobin molecule during transition from oxy-form to deoxy-form resulting in an alteration of the heme function.     

Coumarin derivatives inhibit the aggregation of β-lactoglobulin

The binding of a small molecule to a protein through non-covalent interactions mainly depends on its size and electronic environment. Such binding can change the stability of the three dimensional protein structure which sometimes may destabilize it to accelerate or to inhibit protein aggregation. Coumarin is a widely used fluorescent dye with several biological applications. Different substituents (electron-donating and electron-withdrawing) at different positions of the coumarin moiety can influence its molecular volume, physical and chemical properties. Here we investigate the effect of such substituents of coumarin on the aggregation of a model protein, beta-lactoglobulin (β-lg) through a multi spectroscopic approach. It was observed that coumarin methyl ester with an 8-hydroxyl group can inhibit the β-lg aggregation. This compound can bind the hydrophobic site of beta-lactoglobulin and stabilize a particular protein conformation through the formation of hydrogen bond and hydrophobic interactions. Thus a properly designed compound can inhibit protein–protein interactions through protein–small molecule interactions. Other coumarinoid compounds also are effective in the prevention of thermal aggregation of β-lg.

Aggregation of β-lactoglobulin (β-lg) was inhibited through the stabilization of the native structure by various non-covalent interactions of coumarin derivatives. The 8-hydroxy compound was most effective against the self-assembly of β-lg.     

Catalyst-free chemoselective α-sulfenylation/β-thiolation for α,β-unsaturated carbonyl compounds

A novel, efficient, catalyst-free and product-controllable strategy has been developed for the chemoselective α-sulfenylation/β-thiolation of α,β-unsaturated carbonyl compounds. An aromatic sulfur group could be chemoselectively introduced at α- or β-position of carbonyls with different sulfur reagents under slightly changed reaction conditions. A series of desired products were obtained in moderate to excellent yields. Mechanistic studies revealed that B₂pin₂ played the key role in activating the transformation towards the β-thiolation of α,β-unsaturated carbonyl compounds. This transition-metal-catalyst-free method provides a convenient and efficient tool for the highly chemoselective preparation of α-thiolation or β-sulfenylation products of α,β-unsaturated carbonyl compounds.

This catalyst-free method provides a useful and efficient tool for the highly chemoselective preparation of α-thiolation or β-sulfenylation products of α,β-unsaturated carbonyl compounds.     

Thermal stability and oxidation characteristics of α-pinene, β-pinene and α-pinene/β-pinene mixture

Turpentine is a renewable resource, has good combustion performance, and is considered to be a fuel or promising additive to diesel fuel. This is very important for the investigation of thermal stability and energy oxidation characteristics, because evaluation of energy or fuel quality assurance and use safety are necessary. The main components of turpentine are α-pinene and β-pinene, which have unsaturated double bonds and high chemical activity. By investigating their thermal stability and oxidation reaction characteristics, we know the chemical thermal properties and thermal explosion hazard of turpentine. In this present study, the thermal stability and oxidation characteristics of α-pinene, β-pinene and α-pinene/β-pinene mixture were investigated using a high sensitivity accelerating rate calorimeter (ARC) and C80 calorimeter. The important parameters of oxidation reaction and thermal stability were obtained from the temperature, pressure and exothermic behavior in chemical reaction. The results show that α-pinene and β-pinene are thermally stable without chemical reaction under a nitrogen atmosphere even when the temperature reaches 473 K. The initial exothermic temperature of the two pinenes and their mixture is 333–338 K, and the heat release (−ΔH) of their oxidation is 2745–2973 J g⁻¹. The oxidation activation energy (E_a) of α-pinene, β-pinene and α-pinene/β-pinene mixture is 116.25 kJ mol⁻¹, 121.85 kJ mol⁻¹, and 115.95 kJ mol⁻¹, respectively. There are three steps in the oxidation of pinenes: the first is the induction period of the oxidation reaction; the second is the main oxidation stage, and the pressure is reduced; the third is thermal decomposition to produce gas.

Turpentine is a renewable resource, has good combustion performance, and is considered to be a fuel or promising additive to diesel fuel.     

Electrophilic halogenations of propargyl alcohols: paths to α-haloenones, β-haloenones and mixed β,β-dihaloenones

The Meyer–Schuster rearrangement of propargyl alcohols or alkynols leading to α,β-unsaturated carbonyl compounds is well known. Yet, electrophilic halogenations of the same alkynols and their alkoxy, ester and halo derivatives are inconspicuous. This review on the halogenation reactions of propargyl alcohols and derivatives intends to give a perspective from its humble direct halogenation beginning to the present involving metal catalysis. The halogenation products of propargyl alcohols include α-fluoroenones, α-chloroenones, α-bromoenones and α-iodoenones, as well as β-haloenones and symmetrical and mixed β,β-dihaloenones. They are, in essence, tri and tetrasubstituted alkenes carrying halo-functionalization at the α- or β-carbon. This is a potential stepping stone for further construction towards challenging substituted alkenones via Pd-catalysed coupling reactions.

This review highlights the development of α-haloenone, β-haloenone and mixed β,β-dihaloenone formations from propargyl alcohols via direct electrophilic halogenations and metal catalysed-halonium interception rearrangements.     

Equal synthesis of α- and β-globin chains in erythroid precursors in heterozygous β-thalassemia

In patients with heterozygous beta-thalassemia, the beta/alpha synthetic ratio in marrow erythroid cells incubated in vitro is 1, whereas in reticulocytes the ratio is 0.5. These ratios reflect the equal synthesis of the two chains on the polyribosomes of the bone marrow and unequal synthesis on the polyribosomes of the peripheral blood reticulocytes. alpha- and beta-chain synthesis is also equal in marrow cells in vivo. Equal synthesis is probably due both to a decrease in alpha-chain synthesis and an increase in beta-chain synthesis in bone marrow erythroid cells and may contribute to the absence of overt hemolysis due to excess alpha-globin chain accumulation in heterozygous beta-thalassemia.     

Inhibition of β-Lactamases by β-Lactam Antibiotics

The inhibitory properties of a selected number of beta-lactam antibiotics were studied, with the use of three distinct types of beta-lactamases. The three enzymes were found to be distinguishable on the basis of their susceptibility to inhibition. Not one of the potential inhibitors tested was found to be a potent inhibitor of all three enzymes, but nafcillin possessed the broadest inhibitory activity. The enzyme isolated from Enterobacter cloacae was found to be the most susceptible. In some cases, the degree of inhibition varied with the time of incubation, and, depending upon the time chosen, widely different observations could be made. It is suggested that, in studies such as these, every consideration should be given to the period of incubation and to the concentration of inhibitor employed. Mixtures of inhibitor and cephaloridine did not always act synergistically against growing bacteria, and a number of reasons for failure are suggested.     

Direct β-selectivity of α,β-unsaturated γ-butyrolactam for asymmetric conjugate additions in an organocatalytic manner

The β-selective asymmetric addition of γ-butyrolactam with cyclic imino esters catalyzed by a bifunctional chiral tertiary amine has been developed, which provides an efficient access to optically active β-position functionalized pyrrolidin-2-one derivatives in both high yield and enantioselectivity (up to 78% yield and 95 : 5 er). This is the first catalytic method to access chiral β-functionalized pyrrolidin-2-one via a direct organocatalytic approach.

The asymmetric addition of γ-butyrolactam with cyclic imino esters catalyzed by (DHQD)₂AQN has been developed, which provides an access to β-position functionalized pyrrolidin-2-one derivatives in high levels yield and enantioselectivity.

Metal-free organocatalytic asymmetric transformations have successfully captured considerable enthusiasm of chemists as powerful methods for the synthesis of various kinds of useful chiral compounds ranging from the preparation of biologically important molecules through to novel materials.¹ Chiral pyrrolidin-2-ones have been recognized as important structural motifs that are frequently encountered in a variety of biologically active natural and synthetic compounds.² In particular, the β-position functionalized pyrrolidin-2-one backbones, which can serve as key synthetic precursors for inhibitory neurotransmitters γ-aminobutyric acids (GABA),³ selective GABA_B receptor agonists⁴ as well as antidepressant rolipram analogues,⁵ have attracted a great deal of attention. Therefore, the development of highly efficient, environmentally friendly and convenient asymmetric synthetic methods to access these versatile frameworks is particularly appealing.As a direct precursor to pyrrolidin-2-one derivatives, recently, α,β-unsaturated γ-butyrolactam has emerged as the most attractive reactant in asymmetric organometallic or organocatalytic reactions for the synthesis of chiral γ-position functionalized pyrrolidin-2-ones (Scheme 1). These elegant developments have been achieved in the research area of catalytic asymmetric vinylogous aldol,⁶ Mannich,⁷ Michael⁸ and annulation reactions⁹ in the presence of either metal catalysts or organocatalysts (a, Scheme 1). These well-developed catalytic asymmetric methods have been related to the γ-functionalized α,β-unsaturated γ-butyrolactam to date. However, in sharp contrast, the approaches toward introducing C-3 chirality at the β-position of butyrolactam through a direct catalytic manner are underdeveloped (b, Scheme 1)¹⁰ in spite of the fact that β-selective chiral functionalization of butyrolactam can directly build up α,β-functionalized pyrrolidin-2-one frameworks.Open in a separate windowScheme 1Different reactive position of α,β-unsaturated γ-butyrolactam in catalytic asymmetric reactions.So far, only a few metal-catalytic enantioselective β-selective functionalized reactions have been reported. For examples, a rhodium/diene complex catalyzed efficient asymmetric β-selective arylation^10a and alkenylation^10b have been reported by Lin group (a, Scheme 2). Procter and co-workers reported an efficient Cu(i)–NHC-catalyzed asymmetric silylation of unsaturated lactams (b, Scheme 2).^10c Despite these creative works, considerable challenges still exist in the catalytic asymmetric β-selective functionalization of γ-butyrolactam. First, the scope of nucleophiles is limited to arylboronic acids, potassium alkenyltrifluoroborates and PhMe₂SiBpin reagents. Second, the catalytic system and activation mode is restricted to metal/chiral ligands. To our knowledge, an efficient catalytic method to access chiral β-functionalized pyrrolidin-2-one via a direct organocatalytic approach has not yet been established. Therefore, the development of organocatalytic asymmetric β-selective functionalization of γ-butyrolactam are highly desirable. In conjunction with our continuing efforts in building upon chiral precedents by using chiral tertiary amine catalytic system,¹¹ we rationalized that the activated α,β-unsaturated γ-butyrolactam might serve as a β-position electron-deficient electrophile. This γ-butyrolactam may react with a properly designed electron-rich nucleophile to conduct an expected β-selective functionalized reaction of γ-butyrolactam under a bifunctional organocatalytic fashion, while avoiding the direct γ-selective vinylogous addition reaction or β,γ-selective annulation as outlined in Scheme 2. Herein we report the β-selective asymmetric addition of γ-butyrolactam with cyclic imino esters¹² catalyzed by a bifunctional chiral tertiary amine, which provides an efficient and facile access to optically active β-position functionalized pyrrolidin-2-one derivatives with both high diastereoselectivity and enantioselectivity.Open in a separate windowScheme 2β-Selective functionalization of γ-butyrolactam via metal- (previous work) or organo- (this work) catalytic approach.To begin our initial investigation, several bifunctional organocatalysts¹³ were firstly screened to evaluate their ability to promote the β-selective asymmetric addition of γ-butyrolactam 2a with cyclic imino ester 3a in the presence of 15 mol% of catalyst loading at room temperature in CH₂Cl₂ (entries 1–6,

Preparation and photo-induced activities of water-soluble amyloid β-C60 complexes

We have shown that fullerene (C₆₀) becomes soluble in water by mixing fullerene and amyloid β peptide (Aβ40) whose fibril structures are considered to be associated with Alzheimer''s disease. The water-solubility of fullerene arises from the generation of a nanosized complex between fullerene and the monomer species of Aβ40 (Aβ40-C₆₀). The prepared Aβ40-C₆₀ exhibits photo-induced activity with visible light to induce the inhibition of Aβ40 fibrillation and the cytotoxicity for cultured HeLa cells. The observed photo-induced phenomena result from the generation of singlet oxygen via photoexcitation, inducing oxidative damage to Aβ40 and HeLa cells. The oxidized Aβ40 following photoexcitation of Aβ40-C₆₀ was confirmed by mass spectrometry.

We have shown that fullerene (C₆₀) becomes soluble in water by mixing fullerene and amyloid β peptide (Aβ40) whose fibril structures are considered to be associated with Alzheimer''s disease.     

Inhibition of metal-induced amyloid β-peptide aggregation by a blood–brain barrier permeable silica–cyclen nanochelator

Alzheimer''s disease (AD) is a neurodegenerative malady associated with amyloid β-peptide (Aβ) aggregation in the brain. Metal ions play important roles in Aβ aggregation and neurotoxicity. Metal chelators are potential therapeutic agents for AD because they could sequester metal ions from the Aβ aggregates and reverse the aggregation. The blood–brain barrier (BBB) is a major obstacle for drug delivery to AD patients. Herein, a nanoscale silica–cyclen composite combining cyclen as the metal chelator and silica nanoparticles as a carrier was reported. Silica–cyclen was characterized by scanning electron microscopy (SEM), transmission electron microscopy (TEM), Fourier transform infrared (FT-IR) and dynamic light scattering (DLS). The inhibitory effect of the silica–cyclen nanochelator on Zn²⁺- or Cu²⁺-induced Aβ aggregation was investigated by using a BCA protein assay and TEM. Similar to cyclen, silica–cyclen can effectively inhibit the Aβ aggregation and reduce the generation of reactive oxygen species induced by the Cu–Aβ₄₀ complex, thereby lessening the metal-induced Aβ toxicity against PC12 cells. In vivo studies indicate that the silica–cyclen nanochelator can cross the BBB, which may provide inspiration for the construction of novel Aβ inhibitors.

A BBB-passable nanoscale silica–cyclen chelator effectively reduces the metal-induced Aβ aggregates and related ROS, thereby decreasing the neurotoxicity of Aβ.     

Coordinated β-globin expression and α2-globin reduction in a multiplex lentiviral gene therapy vector for β-thalassemia

A primary challenge in lentiviral gene therapy of β-hemoglobinopathies is to maintain low vector copy numbers to avoid genotoxicity while being reliably therapeutic for all genotypes. We designed a high-titer lentiviral vector, LVβ-shα2, that allows coordinated expression of the therapeutic β^A-T87Q-globin gene and of an intron-embedded miR-30-based short hairpin RNA (shRNA) selectively targeting the α2-globin mRNA. Our approach was guided by the knowledge that moderate reduction of α-globin chain synthesis ameliorates disease severity in β-thalassemia. We demonstrate that LVβ-shα2 reduces α2-globin mRNA expression in erythroid cells while keeping α1-globin mRNA levels unchanged and β^A-T87Q-globin gene expression identical to the parent vector. Compared with the first β^A-T87Q-globin lentiviral vector that has received conditional marketing authorization, BB305, LVβ-shα2 shows 1.7-fold greater potency to improve α/β ratios. It may thus result in greater therapeutic efficacy and reliability for the most severe types of β-thalassemia and provide an improved benefit/risk ratio regardless of the β-thalassemia genotype.     

Hydrophilic and organophilic pervaporation of industrially important α,β and α,ω-diols

The distillation-based purification of α,β and α,ω-diols is energy and resource intensive, as well as time consuming. Pervaporation separation is considered to be a remarkable energy efficient membrane technology for purification of diols. Thus, as a core pervaporation process, hydrophilic polyvinyl alcohol (PVA) membranes for the removal of water from 1,2-hexanediol (1,2-HDO) and organophilic polydimethylsiloxane–polysulfone (PDMS–PSF) membranes for the removal of isopropanol from 1,5 pentanediol (1,5-PDO) were employed. For 1,2-HDO/water separation using a feed having a 1 : 4 weight ratio of 1,2-HDO/water, the membrane prepared using 4 vol% glutaraldehyde (GA4) showed the best performance, yielding a flux of 0.59 kg m⁻² h⁻¹ and a separation factor of 175 at 40 °C. In the organophilic pervaporation separation of the 1,5-PDO/IPA feed having a 9 : 1 weight ratio of components, the PDMS membrane prepared with a molar ratio of TEOS alkoxy groups to PDMS hydroxyl groups of 70 yielded a flux of 0.12 kg m⁻² h⁻¹ and separation factor of 17 638 at 40 °C. Long term stability analysis found that both hydrophilic (PVA) and organophilic (PDMS) membranes retained excellent pervaporation output over 18 days'' continuous exposure to the feed. Both the hydrophilic and organophilic membranes exhibited promising separation performance at elevated operating conditions, showing their great potential for purification of α,β and α,ω-diols.

The distillation-based purification of α,β and α,ω-diols is energy and resource intensive, as well as time consuming.     

Optical and dielectric properties of NaCoPO4 in the three phases α, β and γ

In this work, we are interested in the synthesis of monophosphate α-NaCoPO₄, β-NaCoPO₄ and γ-NaCoPO₄ compounds by mechanochemical method and their characterization by X-ray powder diffraction patterns. These compounds are crystallized in the orthorhombic, hexagonal and monoclinic system, in Pnma, P6₅ and P2₁/n space groups, respectively. The optical properties were measured by means of the UV-vis absorption spectrometry in order to deduce the absorption coefficient α and optical band gap E_g. The calculated values of the indirect band gaps (E_gi) for three samples were estimated at 4.71 eV, 4.63 eV and 3.8 for compounds α, β and γ, respectively. The Tauc model was used to determine the optical gap energy of the synthesized compounds. Then, the results of the dielectric proprieties measured by varying the frequency are described.

In this work, we are interested in the synthesis of monophosphate α-NaCoPO₄, β-NaCoPO₄ and γ-NaCoPO₄ compounds by mechanochemical method and their characterization by X-ray powder diffraction patterns.     

Nanoscopic insights into the surface conformation of neurotoxic amyloid β oligomers

Raman spectroscopy assisted by localized plasmon resonances generating effective hot spots at the gaps between intertwined silver nanowires is herein adopted to unravel characteristic molecular motifs on the surface of Aβ₄₂ misfolded oligomers that are critical in driving intermolecular interactions in neurodegeneration.

Unraveling characteristic structural determinants at the basis of Aβ₄₂ oligomers'' neurotoxicity by a sub-molecular SERS investigation of their surface.     

Imbalance in α and β Globin Synthesis Associated with a Hemoglobinopathy

In contrast to findings in the thalasemia syndromes, studies of globin synthesis in subjects with structurally abnormal hemoglobins have generally revealed equal production of alpha and beta polypeptide chains. However, in the present investigation of globin biosynthesis in vitro in blood and marrow from two subjects heterozygous for unstable hemoglobin Leiden, beta6 or 7 Glu --> O, a significant excess of alpha-chain production was revealed. A mother and daughter of northern European ancestry with mild compensated hemolytic anemia were found to have 25% hemoglobin Leiden. Increased hemolysis occurred after the ingestion of a sulfonamide and during infections. Normal levels of hemoglobin A2, 3.0 and 2.7%, and hemoglobin F, 0.8 and 0.6%, were found in the two subjects. Similar percentages of the minor hemoglobins were demonstrated in other family members without hemoglobin Leiden. After incubation of peripheral blood with [(3)H]-leucine, the beta(A)/beta(Leiden) synthesis ratio was 1.3, and the specific activity of beta(Leiden) was 1.3-2 times beta(A). These results indicate preferential destruction of the unstable hemoglobin Leiden. However, in contrast to previous studies of other unstable hemoglobins, there was excess synthesis of alpha-chains. The total beta/alpha synthesis ratio was 0.47-0.63 in peripheral blood and 0.82 in marrow. A pool of free alpha-chains was demonstrated by starch gel electrophoresis and DEAE column chromatography. The synthesis of globin chains was balanced in family members without hemoglobin Leiden. This degree of predominance of alpha-chain synthesis in subjects with hemoglobin Leiden resembles the findings in heterozygous beta-thalassemia. However, the relatively normal hemoglobin content of the cells with this abnormal hemoglobin suggests the possibility of an absolute excess alpha-chain production in the hemoglobin Leiden syndrome.     

Direct access to multi-functionalized benzenes via [4 + 2] annulation of α-cyano-β-methylenones and α,β-unsaturated aldehydes

An efficient [4 + 2] benzannulation of α-cyano-β-methylenones and α,β-unsaturated aldehydes was achieved under metal-free reaction conditions selectively delivering a wide range of polyfunctional benzenes in high yields respectively (up to 94% yield).

An efficient [4 + 2] benzannulation of α-cyano-β-methylenones and α,β-unsaturated aldehydes was achieved under metal-free reaction conditions selectively delivering a wide range of polyfunctional benzenes in high yields respectively (up to 94% yield).

Multi-substituted benzenes are privileged structural units ubiquitous in pharmaceuticals,¹ natural products² and advanced functional materials.³ Various excellent methodologies have been investigated for the construction of functionalized aromatics including nucleophilic or electrophilic substitution,⁴ transition metal-catalyzed coupling reactions⁵ and directed metalation.⁶ However, the widespread application of these strategies established thus far suffer from the limitations of functional groups introduced on the pre-existing benzene and regioselectivity issues. Among various synthetic methods, tandem benzannulation reactions arguably represent an attractive alternative to classical methods for rapid construction of polysubstituted benzenes in an atom-economical fashion.⁷ This protocol featuring an efficient transformation of acyclic building blocks into structurally valuable benzene skeletons. In this context, α-cyano-β-methylenones has been employed as substrates to format six-membered ring in tandem cyclization reactions due to the activation of the pronucleophile methyl group. In 2015, Tong and co-workers developed a phosphine-catalyzed addition/cycloaddition domino reactions of β′-acetoxy allenoate with 2-acyl-3-methyl-acrylonitriles to give 2-oxabicyclo[3.3.1]nonanes (Scheme 1a).⁸ Soon after that, the construction of benzonitrile derivatives and 1,3,5-trisubstituted benzenes via N-heterocyclic carbene catalysis has been reported by the groups of Wang and Ye independently (Scheme 1b).⁹ Then the synthesis of 1,3,5-trisubstituted benzenes by 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU)-mediated annulation of α-cyano-β-methylenones and α,β-unsaturated carboxylic acids was also developed by Ye and co-workers (Scheme 1c).¹⁰ Shi et al. reported a base-promoted tandem cyclization reaction of α-cyano-β-methylenones and α,β-unsaturated enones, which have electron-withdrawing group (EWG), accessing to a wide range of benzonitriles in a different C–C bond formation process (Scheme 1d).¹¹ As part of our ongoing interest in harnessing enones for developing new methodologies for the construction of functionalized benzenes, we have recently demonstrated NHC-catalyzed convenient benzonitrile assembly in the presence of oxidant.^9a While the same reaction of enals and α-cyano-β-methylenones was conducted in the basic condition without NHC, a novel polyfunctionalized benzene product was obtained (Scheme 1e). The result inspired us to extend the synthetic potential of benzannulation strategy to access diverse benzonitriles, particularly from simpler, abundantly available starting materials.Open in a separate windowScheme 1α-Cyano-β-methylenones in cycloaddition domino reactions.At the outset, model reaction of 2-benzoyl-3-phenylbut-2-enenitrile 1a and cinnamaldehyde 2a was used to evaluate reaction parameters. Key results of condition optimization are summarized in 12 The configuration of products were assigned unambiguously by X-ray analysis of the product 3a. A quick solvent screening demonstrated that chloroform is the best choice to produce the benzannulation product 3a in a desirable yield (entries 10–13, †). Reducing the loading of the cinnamaldehyde or NaOH to 1.2 equivalence led to dramatical loss of the yield (entries 14 &15, EntryBaseSolventTime (h)Yield^b (%)1Cs₂CO₃Toluene24702Na₂CO₃Toluene24423K₂CO₃Toluene24384NaOHToluene12785NaOAcToluene24526KOHToluene12747K₃PO₄Toluene24588DBUToluene24339Et₃NToluene484610NaOHDCM1288 11 NaOH CHCI ₃ 12 94 12NaOHDCE128413NaOHH₂O48014^cNaOHCHCI₃128515^dNaOHCHCI₃128416^eNaOHCHCI₃1280Open in a separate window^aReaction conditions: 1a (0.1 mmol, 1.0 equiv.), 2a (0.15 mmol, 1.5 equiv.), base (0.2 mmol, 2.0 equiv.), and solvent (1 mL) for 12 h.^bIsolated yields.^c1a : 2a = 1 : 1.2.^dNaOH used 1.2 equiv.^e50 °C.Finally, the standard reaction conditions for the base-promoted synthesis of the multi-functionalized benzene derivatives identified as follows: 1.5 equivalence of NaOH and CHCl₃ as the solvent under an atmosphere of air for 12 hours at room temperature.With the optimized reaction conditions in hand, we explored the scope of the reaction. A series of enones were examined, variation of the electronic nature of the aromatic ring (R¹, including the substituted phenyl or thienyl) has little influence on the reaction efficiency (3b–f, 86–93% yields, Open in a separate window^aReaction conditions: 1a (0.1 mmol, 1.0 equiv.), 2a (0.15 mmol, 1.5 equiv.), NaOH (0.2 mmol, 2.0 equiv.), and CHCl₃ (1 mL) for 12 h.We next turned our attention to examine the scope of enals. Different substituents on the phenyl ring of cinnamaldehydes were tolerated even disregarding the position and properties, giving 4a–g in satisfying yields (82–92% yields, Open in a separate window^aReaction conditions: 1a (0.1 mmol, 1.0 equiv.), 2a (0.15 mmol, 1.5 equiv.), NaOH (0.2 mmol, 2.0 equiv.), and CHCl₃ (1 mL) for 12 h.To highlight the practicality of this mild and efficient method, the reaction of 2-benzoyl-3-phenylbut-2-enenitrile 1a at 4.0 mmol scale proceed well under the standard conditions to generate the desired product in 88% yield (Scheme 2).Open in a separate windowScheme 2Gram-Scale Synthesis of 3a.The formyl group could be easily reduced by using LiAlH₄ in THF at reflux, leading to the formation of the benzyl alcohol product 5 in 95% yield while keeping the CN group intact. Suzuki coupling of 3o with phenylboronic acid furnished derivative 6 in 90% yield¹³ (Scheme 3).Open in a separate windowScheme 3Synthetic transformation.To gain insight into the role of air in this reaction, a control experiment was designed and investigated (Scheme 4). When the reaction of 1a and 2a was carried out under an argon atmosphere, the desired product 3a was obtained in 10% yield and product 7 could be isolated in 82% yield. The results indicate that oxygen is necessary for the oxidation process and played a key role in this reaction.Open in a separate windowScheme 4Control experiment.A postulated reaction course is illustrated in Scheme 5. Briefly, α-deprotonation of enone 1a in the presence of bases, subsequent 1,4-addition of deprotonated enone I to enal 2a generates intermediate II, which undergoes an intramolecular aldol reaction to yield the adduct 7.¹⁴ Lastly, dehydration of 7 followed by spontaneous oxidative aromatization affords the polysubstituted benzonitrile 3a.Open in a separate windowScheme 5The proposed mechanism.     

Translation of β-Globin m-RNA in β-Thalassemia and the S and C Hemoglobinopathies

Genetic and biochemical evidence indicates that in beta-thalassemia there is impaired synthesis of the beta-globin chains of hemoglobin A. In patients heterozygous for the hemoglobinopathies, hemoglobin S and hemoglobin C, the mutant beta-chain is produced in smaller amounts than normal beta(A). Defective m-RNA translation has been suggested as a possible cause of decreased beta-globin polypeptide synthesis in thalassemia and the hemoglobinopathies. In the present study, the ribosomal assembly of beta-globin chains was examined in the peripheral, nucleated red blood cells and reticulocytes of patients with Cooley's anemia, thalassemia intermedia, sickle thalassemia, sickle cell anemia, hemoglobin C disease, and in hemolytic anemias not associated with a hemoglobinopathy. The translation times of beta(A), beta(S), and beta(C) did not differ significantly (average times; beta(A) = 75 sec, range 43-114, beta(S) = 69 sec, beta(C) = 92 sec). In thalassemia, no evidence was found for a delay in translation as the cause of the marked impairment of beta-globin synthesis. In several specimens of peripheral blood from thalassemic patients, the translation time of the beta-chain was even shorter than in nonthalassemic specimens (average time = 45 sec, range 35-59). The results suggest that the defect in beta-globin synthesis in beta-thalassemia is due to impaired initiation of beta-globin chain assembly or a quantitative deficiency in m-RNA.     

Selection and Characterization of β-Lactam–β-Lactamase Inactivator-Resistant Mutants following PCR Mutagenesis of the TEM-1 β-Lactamase Gene

Mechanism-based inactivators of β-lactamases are used to overcome the resistance of clinical pathogens to β-lactam antibiotics. This strategy can itself be overcome by mutations of the β-lactamase that compromise the effectiveness of their inactivation. We used PCR mutagenesis of the TEM-1 β-lactamase gene and sequenced the genes of 20 mutants that grew in the presence of ampicillin-clavulanate. Eleven different mutant genes from these strains contained from 1 to 10 mutations. Each had a replacement of one of the four residues, Met69, Ser130, Arg244, and Asn276, whose substitutions by themselves had been shown to result in inhibitor resistance. None of the mutant enzymes with multiple amino acid substitutions generated in this study conferred higher levels of resistance to ampicillin alone or ampicillin with β-lactamase inactivators (clavulanate, sulbactam, or tazobactam) than the levels of resistance conferred by the corresponding single-mutant enzymes. Of the four enzymes with just a single mutation (Ser130Gly, Arg244Cys, Arg244Ser, or Asn276Asp), the Asn276Asp β-lactamase conferred a wild-type level of ampicillin resistance and the highest levels of resistance to ampicillin in the presence of inhibitors. Site-directed random mutagenesis of the Ser130 codon yielded no other mutant with replacement of Ser130 besides Ser130Gly that produced ampicillin-clavulanate resistance. Thus, despite PCR mutagenesis we found no new mutant TEM β-lactamase that conferred a level of resistance to ampicillin plus inactivators greater than that produced by the single-mutation enzymes that have already been reported in clinical isolates. Although this is reassuring, one must caution that other combinations of multiple mutations might still produce unexpected resistance.