CRISPR-Cas9 assisted functional gene editing in the mushroom Ganoderma lucidum.

The genetic manipulation of basidiomycete mushrooms is notoriously difficult and immature, and there is a lack of research reports on clustered regularly interspaced short palindromic repeat (CRISPR) based gene editing of functional genes in mushrooms. In this work, Ganoderma lucidum, a famous traditional medicinal basidiomycete mushroom, which produces a type of unique triterpenoid-anti-tumor ganoderic acids (GAs), was used, and a CRISPR/CRISPR-associated protein-9 nuclease (Cas9) editing system for functional genes of GA biosynthesis was constructed in the mushroom. As proof of concept, the effect of different gRNA constructs with endogenous u6 promoter and self-cleaving ribozyme HDV on ura3 disruption efficiency was investigated at first. The established system was applied to edit a cytochrome P450 monooxygenase (CYP450) gene cyp5150l8, which is responsible for a three-step biotransformation of lanosterol at C-26 to ganoderic acid 3-hydroxy-lanosta-8, 24-dien-26 oic acid. As a result, precisely edited cyp5150l8 disruptants were obtained after sequencing confirmation. The fermentation products of the wild type (WT) and cyp5150l8 disruptant were analyzed, and a significant decrease in the titer of four identified GAs was found in the mutant compared to WT. Another CYP gene involved in the biosynthesis of squalene-type triterpenoid 2, 3; 22, 23-squalene dioxide, cyp505d13, was also disrupted using the established CRISPR-Cas9 based gene editing platform of G. lucidum. The work will be helpful to strain molecular breeding and biotechnological applications of G. lucidum and other basidiomycete mushrooms.

A New Class of Glucosyl Thioureas: Synthesis and Larvicidal Activities.

A novel series of glucosyl thioureas were synthesized in good overall yields (up to 37% over four steps) from d-glucose and primary amines, and their larvicidal activities toward Mythimna separata Walker were also investigated. This new class of glucosyl thioureas demonstrated low to moderate growth inhibition activity of Mythiman separata Walker, with a growth inhibitory rate of up to 47.5% at a concentration of 100.0 mg/L in acetone.

Facile access to unnatural dipeptide-alcohols based on cis-2,5-disubstituted pyrrolidines.

Well-defined unnatural dipeptide-alcohols based on a cis-2,5-disubstitued pyrrolidine backbone were synthesized from commercially available starting materials meso-diethyl-2,5-dibromoadipate, (S)-(-)-1-phenylethylamine, and phenylalaninol. The structures of these unnatural dipeptide-alcohols are supported by HRMS, 1H- and 13C-NMR spectroscopy. These unnatural dipeptide-alcohols can act as building blocks for peptidomimetics.

HDAC4/5-HMGB1 signalling mediated by NADPH oxidase activity contributes to cerebral ischaemia/reperfusion injury.

Histone deacetylases (HDACs)-mediated epigenetic mechanisms play critical roles in the homeostasis of histone acetylation and gene transcription. HDAC inhibitors have displayed neuroprotective properties in animal models for various neurological diseases including Alzheimer's disease and ischaemic stroke. However, some studies have also reported that HDAC enzymes exert protective effects in several pathological conditions including ischaemic stress. The mixed results indicate the specific roles of each HDAC protein in different diseased states. However, the subtypes of HDACs associated with ischaemic stroke keep unclear. Therefore, in this study, we used an in vivo middle cerebral artery occlusion (MCAO) model and in vitro cell cultures by the model of oxygen glucose deprivation to investigate the expression patterns of HDACs and explore the roles of individual HDACs in ischaemic stroke. Our results showed that inhibition of NADPH oxidase activity ameliorated cerebral ischaemia/reperfusion (I/R) injury and among Zn(2+) -dependent HDACs, HDAC4 and HDAC5 were significantly decreased both in vivo and in vitro, which can be reversed by NADPH oxidase inhibitor apocynin. We further found that both HDAC4 and HDAC5 increased cell viability through inhibition of HMGB1, a central mediator of tissue damage following acute injury, expression and release in PC12 cells. Our results for the first time provide evidence that NADPH oxidase-mediated HDAC4 and HDAC5 expression contributes to cerebral ischaemia injury via HMGB1 signalling pathway, suggesting that it is important to elucidate the role of individual HDACs within the brain, and the development of HDAC inhibitors with improved specificity is required to develop effective therapeutic strategies to treat stroke.

Hair fiber as a nanoreactor in controlled synthesis of fluorescent gold nanoparticles.

The synthesis and detailed characterization of gold nanoparticles (AuNPs) inside human hair has been achieved by treatment of hair with HAuCl(4) in alkaline medium. The AuNPs, which show a strong red fluorescence under blue light, are generated inside the fiber and are arranged in the cortex in a remarkably regular pattern of whorls based on concentric circles, like a fingerprint. It opens an area of genuine nanocomposites with novel properties due to AuNPs inside the hair shaft.

Organocatalyzed enantioselective desymmetrization of aziridines and epoxides.

Enantioselective desymmetrization of meso-aziridines and meso-epoxides with various nucleophiles by organocatalysis has emerged as a cutting-edge approach in recent years. This review summarizes the origin and recent developments of enantioselective desymmetrization of meso-aziridines and meso-epoxides in the presence of organocatalysts.

One-pot tandem cyclization of enantiopure asymmetric cis-2,5-disubstituted pyrrolidines: Facile access to chiral 10-heteroazatriquinanes.

A series of chiral 10-heteroazatriquinanes were synthesized from enantiopure asymmetric cis-2,5-disubstituted pyrrolidines through a one-pot tandem cyclization procedure. The structures and configurations of these new chiral 10-heteroazatriquinanes are confirmed by X-ray single-crystal diffraction analysis.

Chiral bifunctional organocatalysts for enantioselective synthesis of 3-substituted isoindolinones.

A series of chiral bifunctional organocatalysts were prepared and used for enantioselective synthesis of 3-substituted isoindolinones from 2-formylarylnitriles and malonates through aldol-cyclization rearrangement tandem reaction in excellent yields and enantioselectivites (up to 87% yield and 95% ee) without recrystallization. In this investigation, we found that chiral tertiary-amine catalysts with a urea group can afford 3-substituted isoindolinones both in higher yields (87% vs. 77%) and enantioselectivities (95% ee vs. 46% ee) than chiral bifunctional phase-transfer catalysts.

Efficient synthesis of unnatural dipeptides based on cis-2,5-disubstituted pyrrolidine.

The well-defined unnatural dipeptides based on cis-2,5-disubstituted pyrrolidine backbone were synthesized from commercially available starting materials meso-diethyl-2,5-dibromoadipate, (S)-(-)-1-phenylethylamine, and phenylalanine. The configurations of all the chiral centers in these unnatural dipeptides are confirmed by X-ray crystal diffraction analysis.

Chiral Cyclopropenimine-catalyzed Asymmetric Michael Addition of Bulky Glycine Imine to α,ß-Unsaturated Isoxazoles.

A highly efficient asymmetric Michael addition of bulky glycine imine to α,ß-unsaturated isoxazoles has been achieved by using 5 mol% of chiral cyclopropenimine as a chiral organo-superbase catalyst under mild conditions. Michael adducts were obtained in excellent yields (up to 97%) and stereoselectivities (up to >99 : 1 dr and 98% ee). A significant solvent effect was found in these chiral organosuperbase catalyzed asymmetric Michael reactions. Gram-scale preparation of Michael adducts and their transformations are realized to provide corresponding products without loss of stereoselectivities. The configurations of Michael adduct was determined by single-crystal X-ray diffraction analysis.

(S)-1-Ferrocenyl-3-hy-droxy-3-phenyl-propan-1-one.

In the title compound, [Fe(C(5)H(5))(C(14)H(13)O(2))], the dihedral angle between the phenyl ring and the unsubstituted cyclo-petadienyl ring is 85.0 (2)°while that between the phenyl ring and the substituted cyclo-petadienyl ring is 83.6 (2)°. The dihedral angle between the two cyclo-penta-1,3-diene rings of the ferrocene unit is 2.2 (2)°. The mol-ecules are stabilized by inter-molecular O-H⋯O hydrogen-bonding inter-action within the crystal lattice.

2,2-Dichloro-1-(4-methyl-phen-yl)ethanone.

The mol-ecule of the title compound, C(9)H(8)Cl(2)O, is almost planar: the dihedral angle between the benzene ring and the plane defined by the carbonyl O and ethane C atoms is 15.5 (2)°. The crystal packing is stabilized by inter-molecular C-H⋯O hydrogen bonds.

1-(4-Amino-3,5-dichloro-phen-yl)ethanol.

The asymmetric unit of the title compound, C(8)H(9)Cl(2)NO, contains two crystallographically independent mol-ecules which are connected via an N-H⋯O hydrogen bond . There is aromatic π-π stacking in the crystal, with a centroid-centroid distance between benzene rings of 3.48 (2)Å. The crystal packing is stabilized by intermolecular hydrogen bonds.

Ethyl 4-benzamido-5-phenyl-4H-1,2,4-triazole-3-carboxyl-ate monohydrate.

In the title compound, C(18)H(16)N(4)O(3)·H(2)O, the dihedral angles between the triazole ring and the phenyl rings are 84.8 (4) and and 39.8 (4)°. The phenyl rings make a dihedral angle of 84.5 (9)°. In the crystal, the molecules are linked by N-H⋯O and O-H⋯N hydrogen bonds. An intra-molecular O⋯N inter-action also occurs [2.827 (3) Å].

Synthesis and anti-HCV activity of a new 2'-deoxy-2'-fluoro-2'-C-methyl nucleoside analogue.

2'-Deoxy-2'-fluoro-2'-C-methyl nucleoside analogue 4 was designed and synthesized. Initial biological studies indicated that this compound showed promising activity against HCV replication.

[Novel synthetic method and analgesic activity of tepoxalin].

Tepoxalin is a potent inhibitor of both the cyclooxygenase and lipoxygenase pathways of the arachidonic acid cascade, as well as a potent anti-inflammatory and control-pain (postoperation, arthritis et. al.) agent. The new method about the use of novel synthesis reagents and the first using ionic liquid as reactive solvent to synthesize tepoxalin were presented in this paper. The ionic liquid can be easily recycled and reused for several runs efficiently. The analgesic activity of tepoxalin was detected by acetic acid test on mice. The analysis of variance showed that oral administration of tepoxalin could significantly inhibit the number of writhing response within 1 hour and prolong the latent time in a dose dependent manner as compared with CMC control group (P < 0.05). At the same time, tepoxalin had the same analgesic activity as diclofenac sodium.

(3S,4S)-1-Benzyl-pyrrolidine-3,4-diol.

In the title compound, C(11)H(15)NO(2), the pyrrolidine ring adapts a twisted envelope conformation and the two hydroxyl groups are arranged in a trans conformation. The crystal packing is stabilized by inter-molecular O-H⋯N and O-H⋯O hydrogen bonds. A weak C-H⋯π inter-action also occurs.

Bifunctional phase-transfer catalysts for synthesis of 2-oxazolidinones from isocyanates and epoxides.

A series of bifunctional phase-transfer catalysts (PTCs) were synthesized to catalyze the [3 + 2] coupling reaction of isocyanates and epoxides to afford 2-oxazolidinones in good to high yields (up to 92% yield) using PhCl as a solvent at 100 °C within 12 h. These bifunctional PTCs were easily prepared from commercially available tertiary-primary diamines and isocyanates (or isothiocyanates, mono-squaramides, respectively) in two simple steps with good modularity and demonstrated high efficiency (2.5 mol% catalyst-loading). The synergistic interaction of the quaternary ammonium salt center and hydrogen-bond donor group in the catalyst with the substrate is crucial to this atom-economic reaction.

Selectivity differences between alkyl and polar-modified alkyl phases in reversed phase high performance liquid chromatography.

Compared to conventional C18 phases, polar-modified phases have distinct differences with regards to chromatographic behavior. In the present study, ODS phases and polar-modified phases were synthesized. The columns containing these new packings demonstrated satisfactory stability under both acidic (pH 1.5) and basic (pH 10) conditions. We evaluated the selectivity differences between alkyl and polar-modified alkyl RP columns by using a range of neutral analytes. The polar-modified alkyl phases showed excellent peak shapes for almost all compounds. We also compared the selectivity differences between them for separating nucleotides by using 100% aqueous mobile phase and tricyclic antidepressants in the intermediate pH mobile phases. The results demonstrated that polar-modified phases display a significantly reduced hydrophobic nature and a significantly reduced silanol activity compared to the conventional C18 phases.

2-Benz-yloxy-1-naphthaldehyde.

In the title compound, C(18)H(14)O(2), the dihedral angle between the phenyl and naphthyl ring systems is 21.8 (3)°. The packing of mol-ecules in the crystal structure is stabilized by weak inter-molecular C-H⋯O hydrogen bonds.