2(S),4(R)-4-(β-d-Galactopyranosyloxy)-4-isobutyl-glutamic acid: A new amino acid in Reseda odorata

2(S),4(R)-4-(β-d-Galactopyranosyloxy)-4-isobutylglutamic acid (I) has been isolated from the flowers of Reseda odorata, wherein it occurs in substantial quantity. Hydrolysis of I gives d-galactose, 2(S),4(R)-4-hydroxy-4-isobutylglutamic acid (II) and 3(R),5(S)-3-hydroxy-3-isobutyl-2-pyrrolidone-5-carboxylic acid (III) and its treatment with nitrous acid yields a galactoside of a non-nitrogenous hydroxy acid lactone (IV). The structures of I and its degradation products are supported by PMR, ¹³C-NMR and other spectroscopic methods. ¹³C-NMR spectroscopy of the model compound 2-(β-d-galactopyranosyloxy)isobutyric acid confirmed the structure of the natural product. The S- (or l-) configuration at C(2) in the amino acid moiety of I has been established by the use of the Clough—Lutz—Jirgenson rule and the R-configuration at C(4) of the same unit has been assigned tentatively. I represents the first example of a glycoside of a higher plant amino acid in which the carbohydrate residue is linked to an aliphatic hydroxy group.     

Chemical and enzymatic synthesis of monomeric procyanidins (leucocyanidins or 3′,4′,5,7-tetrahydroxyflavan-3,4-diols) from (2R,3R)-dihydroquercetin

The major product from the reduction of (2R,3R)-dihydroquercetin with sodium borohydride is the 2,3-trans-3,4-trans isomer of leucocyanidin [(2R,3S,4R-3,3′,4,4′,5,7-hexahydroxyflavan] whereas the enzymatic reduction product is the 2,3-trans-3,4-cis isomer [(2R,3S,4S)-3,3′,4,4′,5,7-hexahydroxyflavan]. The 3,4-trans isomer may be partly converted to the 3,4-cis isomer under mild acid conditions. The 3,4-cis isomer is more acid-labile, and more reactive both chemically with thiols and enzymatically with a diol reductase, than the 3,4-trans isomer.     

Biosynthesis of α-spinasterol from (2- 14C (4R)-4-3H1) mevalonic acid by Spinacea oleracea and Medicago sativa

Leaves of Spinacea oleracea and Medicago sativa were incubated with (2-¹⁴C, (4R)-4³H₁ mevalonic acid and the sterols isolated. Cycloartenol had a ³H: ¹⁴C atomic ratio of 6:6 whilst oxidation to cycloartenone resulted in a ratio of 5:6 showing that tritium was present in the 3α-position and that the cycloartenol was symmetrically labelled. Separation of the 4-demethyl sterols gave α-spinasterol and a mixture of stigmast-7-enol and 24-methylcholest-7-enol, which had ³H: ¹⁴C atomic ratios of 3:5. Ozonolysis of α-spinastery] acetate gave the terminal side chain fragment as 2-ethyl-3-methyl butanoic acid. The acid contained ¹⁴C but no tritium thus showing that the C-24 hydrogen of cycloartenol is lost during the alkylation reactions leading to the C-24 ethyl group of α-spinasterol.     

One-pot efficient synthesis of 13(R),14(R)-epoxy-17β-methyl-20(S)-hydroxyl-18-nor-pregna-4-en-3-one via a tandem epoxidation-rearrangement-epoxidation reaction sequence

One-pot synthesis of an 18-norsteroid compound, 13(R),14(R)-epoxy-17β-methyl-20(S)-hydroxyl-18-nor-pregna-4-en-3-one has been achieved with peracetic acid/acetic acid under a mild condition, via a proved tandem epoxidation-rearrangement-epoxidation sequence. Its structure was designated on the basis of NMR and X-ray crystallography data.     

Synthesis of glycos-3-yl-α,γ-diamino acids. Synthesis of four diastereomeric spiro-3,4′-(R and S)-3-deoxy-1,2:5,6-di-O-isopropylidene-α-d-ribo-hexofuranose)-3′-(R and S)-acetamido-2′-pyrrolidinones

Treatment of (Z)-3-deoxy-1,2:5,6-di-O-isopropylidine-3-C-(methoxycarbonyl)-methylene-α-d-ribo-hexofuranose (1) with diazomethane in ether afforded the unstable Δ¹- and Δ²-pyrazolines 2 and 2a. High-pressure hydrogenation of the latter compounds over Raney nickel afforded a mixture of amines 3, 5, 7, and 9 (in 80% yield), which were separated by chromatography. Acetylation of these compounds yielded the N-acetyl derivatives 4, 6, 8, and 10. X-Ray analysis of compounds 8 and 10 showed them to be spiro-3,4′-(R)-(3-deoxy-1,2:5,6-di-O-isopropylidine-α-d-ribo-hexofuranose)-3′-(R)-[and 3′-(S)]-acetamido-2′-pyrrolidinone, respectively. The structures of compounds 4 and 6 (determined by chemical means) were the corresponding spiro-3,4′-(S)-3′-(R)-acetamido-2′-pyrrolidinone and 3′-(S)-acetamido-2′-pyrrolidinone, respectively.     

R-(-)-β-O-methylsynephrine, a natural product, inhibits VEGF-induced angiogenesis in vitro and in vivo

R-(-)-β-O-methylsynephrine (OMe-Syn) is an active compound isolated from a plant of the Rutaceae family. We conducted cell proliferation assays on various cell lines and found that OMe-Syn more strongly inhibited the growth of human umbilical vein endothelial cells (HUVECs) than that of other normal and cancer cell lines tested. In angiogenesis assays, it inhibited vascular endothelial growth factor (VEGF)-induced invasion and tube formation of HUVECs with no toxicity. The anti-angiogenic activity of OMe-Syn was also validated in vivo using the chorioallantonic membrane (CAM) assay in growing chick embryos. Expression of the growth factors VEGF, hepatocyte growth factor, and basic fibroblast growth factor was suppressed by OMe-Syn in a dose-dependent manner. Taken together, our results indicate that this compound could be a novel basis for a small molecule targeting angiogenesis.     

The molecular structure of (20R)-20-phenyl-5-pregnene-3β,20-diol,an inhibitor of cholesterol conversion to pregnenolone

The crystal and molecular structure of (20R)-20-phenyl-5-pregnene-3β, 20-diol hemihydrate has been determined by X-ray analysis in order to establish the configuration and conformation at C(20). Interest in this compound was stimulated by its high affinity inhibitory binding to cytochrome P-450_SCC, the enzyme which catalyzes the biosynthesis of pregnenolone(3β-hydroxy-5-pregnen-20-one) from cholesterol. The results of the analysis suggest a possible conformation for the cholesterol side chain in the enzyme complex.     

The structure of Λ-β1-[Co(R,R-picchxn)(S-tyr)]Br2 · 3.5H2O and solution configuration of Λ-β1-[Co(R,R-picchxn)(aromatic S-aminoacidate)] complexes: model systems for the investigation of non-covalent aromatic interactions

Intramolecular non-covalent interactions involving aromatic residues in the ternary species Λ-β₁-[Co(R,R-picchxn)(S-aa)]²⁺ (aa=Tyr, OMe-tyr or Phe) have been investigated. Such interactions are important to discriminatory processes associated with molecular recognition in chemical and biochemical systems. The single-crystal X-ray study of Λ-β₁-[Co(R,R-picchxn)(S-tyr)]Br₂ · 3.5H₂O demonstrates the influence of intramolecular π-π and bifurcated NH-π interactions in determining the molecular conformation of the complex cation in the solid state. The Co(III) complexes synthesised are diamagnetic, and have been fully assigned in solution using multidimensional NMR techniques. Remarkably, the solid state conformation observed for Λ-β₁-[Co(R,R-picchxn)(S-tyr)]²⁺ has been shown to predominate in solutions of all the complexes, as evidenced by appropriate rOe correlations. ¹H NMR measurements carried out in order to determine equilibrium rotamer distributions confirm the dominance of this conformer in solution. NMR measurements also show that rotamer populations are relatively unchanged at elevated temperatures and in a variety of solvents. The results of this detailed study, which demonstrate the significance of non-covalent interactions involving aromatic residues to the determination of the molecular conformation, serve to highlight the suitability of these simple ternary Co(III) complexes to act as models for such interactions.     

Synthesis of 1,2,4-tri-O-acetyl-5-deoxy-3-O-methyl-5-C-[(R)- and -(S)-phenylphosphinyl]-β-d-ribopyranose

5-Deoxy-1,2-O-isopropylidene-5-C-(methoxyphenylphosphinyl)-3-O-methyl-α-d-ribofuranose (4) was prepared from 1,2-O-isopropylidene-3-O-methyl-α-d-ribo-pentodialdo-1,4-furanose by an addition reaction with methyl phenylphosphinate, followed by deoxygenation of the terminal HOCHP group of the adduct by successive reaction with 1,1′-thiocarbonyldiimidazole and tributyltin hydride. Treatment of 4 with sodium dihydrobis(2-methoxyethoxy)aluminate, followed by deacetonation with mineral acid, and acetylation with acetic anhydride—pyridine, gave mainly the two title compounds, which were isolated by column chromatography on silica gel, and characterized by 90-MHz, ¹H-n.m.r.-spectral analysis.     

Study of the inclusion of the (R)- and (S)-camphor enantiomers in α-cyclodextrin by X-ray crystallography and molecular dynamics

The inclusion of (R)- and (S)-camphor compounds in α-cyclodextrin has been studied by X-ray crystallography. The crystal structures of the complexes reveal that one guest molecule is accommodated inside the cavity formed by a head-to-head cyclodextrin dimer. In the crystal lattice, the dimers form layers which are successively shifted by half a dimer. In both (R)- and (S)-cases, the camphor molecule exhibits disorder and occupies three major sites with orientations that can be described as either ‘polar’ or ‘equatorial’. Molecular dynamics simulations performed for the observed complexes indicate that although the carbonyl oxygen of both (R)- and (S)-camphor switches between different hydrogen bonding partners, it maintains the observed mode of ‘polar’ or ‘equatorial’ alignment.     

八肋游仆虫大核基因组中组蛋白基因 H4A和H4B的克隆及分析

组蛋白作为核小体的基本组分,是染色质的结构和功能必需的。组蛋白的变体和修饰共同参与染色质修饰及基因的表达调控。真核生物细胞中的5种组蛋白在进化中高度保守,然而纤毛虫的组蛋白H4与其他真核生物相比有较大的差异。本实验应用PCR技术从八肋游仆虫(Euplotes octocarinatus)中获得了2种组蛋白H4基因,分别为H4A和H4B,GenBank登录号为:JN715068和JN715069。序列分析表明,H4A基因开放阅读框324 bp,预测编码107个氨基酸,分子量为11.6 ku,等电点为10.99。而H4B基因编码框384 bp,编码127个氨基酸,分子量为14.4 ku,等电点为9.93。Blast结果显示,H4A序列与其他生物中H4的一致性相对较高,达81%～94%,而H4B的一致性为36%～70%。H4A和H4B的一致性仅为44.7%。实时荧光定量PCR表明,H4A的转录本高于H4B。结果提示:在进化过程中八肋游仆虫可能进化出特殊的组蛋白H4基因,不同的组蛋白H4可能发挥不同的功能。     

Photo-conversion of two epimers (20R and 20S) of pregna-5,7-diene-3β, 17α, 20-triol and their bioactivity in melanoma cells

Pregna-5,7-dienes and their hydroxylated derivatives can be formed in vivo when there is a deficiency in 7-dehydrocholesterol (7-DHC) Δ-reductase function, e.g., Smith-Lemli-Opitz syndrome (SLOS). Ultraviolet B (UVB) radiation induces photoconversion of 7-DHC to vitamin D₃, lumisterol₃ and tachysterol₃. Two epimers (20R and 20S) of pregna-5,7-diene-3β,17α,20-triol (4R and 4S, respectively) were synthesized and their UVB photo-conversion products identified as corresponding 9,10-secosteroids with vitamin D-like and tachysterol-like structures, and 5,7-dienes with inverted configuration at C-9 and C-10 (lumisterol-like). The number and character of the products and the dynamics of the process were dependent on the UVB dose. At high UVB doses, the formation of multiple oxidized derivatives of the primary products, and the formation of 5,7,9(11)-triene, were observed. The production of vitamin D-like, tachysterol-like and lumisterol-like derivatives was also observed in human skin treated with 4R and 4S, and subjected to UV irradiation, as shown by RP-HPLC. Newly synthesized compounds inhibited melanoma growth in dose dependent manner, and some of them showed equal or higher potency than 1,25(OH)₂D₃. In summary, we have characterized for the first time the products of UV induced conversion of pregna-5,7-diene-3β,17α,20-triols and documented that the newly synthesized compounds have antiproliferative properties against melanoma cells.     

Progress in antischistosomal N,N′-diaryl urea SAR

N,N′-Diaryl ureas have recently emerged as a new antischistosomal chemotype. We now describe physicochemical profiling, in vitro ADME, plasma exposure, and ex vivo and in vivo activities against Schistosoma mansoni for twenty new N,N′-diaryl ureas designed primarily to increase aqueous solubility, but also to maximize structural diversity. Replacement of one of the 4-fluoro-3-trifluoromethylphenyl substructures of lead N,N′-diaryl urea 1 with azaheterocycles and benzoic acids, benzamides, or benzonitriles decreased lipophilicity, and in most cases, increased aqueous solubility. There was no clear relationship between lipophilicity and metabolic stability, although all compounds with 3-trifluoromethyl-4-pyridyl substructures were metabolically stable. N,N′-diaryl ureas containing 4-fluoro-3-trifluoromethylphenyl, 3-trifluoromethyl-4-pyridyl, 2,2-difluorobenzodioxole, or 4-benzonitrile substructures had high activity against ex vivo S. mansoni and relatively low cytotoxicity. N,N-diaryl ureas with 3-trifluoromethyl-4-pyridyl and 2,2-difluorobenzodioxole substructures had the highest exposures whereas those with 4-fluoro-3-trifluoromethylphenyl substructures had the best in vivo antischistosomal activities. There was no direct correlation between compound exposure and in vivo activity.     

Visual learning and memory of Drosophila melanogaster wild type CS and the mutants dunce1, amnesiac, turnip and rutabaga

Dunce¹, amnesiac, turnip and rutabaga, mutants of Drosophila melanogaster deficient in olfactory learning and/or memory, were tested for visual learning ability and memory. All these mutants are able to learn conditioned visual information, but not as well as the corresponding wildtype CS. Memory of all four mutants is reduced during the first 30 min after training, but indistinguishable from that of the wildtype two hours after conditioning.     

Identification of fluorinated (R)-(−)-aporphine derivatives as potent and selective ligands at serotonin 5-HT2C receptor

A series of novel aporphine derivatives were synthesized for initial screening at the 5-HT₂ receptor subtypes. Among them, Compounds 11a and 11b were identified as potent 5-HT_2C hit ligands with high selectivity over other 5-HT₂ receptor subtypes. Molecular docking study revealed that compounds 11a and 11b formed two key interactions with the binding site of 5-HT_2C receptor, including a salt-bridge to D3.32 and a H-bond interaction with N6.55.     

Asymmetric incorporation of 4-(2′-carboxyphenyl)-4-oxobutyrate into phylloquinone by Zea mays

Radioactivity from 4-(2′-carboxyphenyl)-4-oxobutyrate-[2-¹⁴C] and 4-(2′-carboxyphenyl)- 4-oxobutyrate-[3-¹⁴C] was incorporated into C-3 and C-2 respectively of phylloquinone in maize shoots. These results show that this substrate is incorporated in the same asymmetric manner into phylloquinone as it is into the bacterial menaquinones.     

In vitro, ex vivo and in vivo models: A comparative analysis of Paracoccidioides spp. proteomic studies

Members of the Paracoccidioides complex are human pathogens that infect different anatomic sites in the host. The ability of Paracoccidioides spp. to infect host niches is putatively supported by a wide range of virulence factors, as well as fitness attributes that may comprise the transition from mycelia/conidia to yeast cells, response to deprivation of micronutrients in the host, expression of adhesins on the cell surface, response to oxidative and nitrosative stresses, as well as the secretion of hydrolytic enzymes in the host tissue. Our understanding of how those molecules can contribute to the infection establishment has been increasing significantly, through the utilization of several models, including in vitro, ex vivo and in vivo infection in animal models. In this review we present an update of our understanding on the strategies used by the pathogen to establish infection. Our results were obtained through a comparative proteomic analysis of Paracoccidioides spp. in models of infection.     

Acute toxicity,critical body residues,Michaelis–Menten analysis of bioaccumulation,and ionoregulatory disturbance in response to waterborne nickel in four invertebrates: Chironomus riparius, Lymnaea stagnalis, Lumbriculus variegatus and Daphnia pulex

We investigated the bioaccumulation and acute toxicity (48 h or 96 h) of Ni in four freshwater invertebrate species in two waters with hardness of 40 (soft water) and 140 mg L^− 1 as CaCO₃ (hard water). Sensitivity order (most to least) was Lymnaea stagnalis > Daphnia pulex > Lumbriculus variegatus > Chironomus riparius. In all cases water hardness was protective against acute Ni toxicity with LC50 values 3–3.5 × higher in the hard water vs. soft water. In addition, higher water hardness significantly reduced Ni bioaccumulation in these organisms suggesting that competition by Ca and Mg for uptake at the biotic ligand may contribute to higher metal resistance. CBR50 values (Critical Body Residues) were less dependent on water chemistry (i.e. more consistent) than LC50 values within and across species by ~ 2 fold. These data support one of the main advantages of the Tissue Residue Approach (TRA) where tissue concentrations are generally less variable than exposure concentrations with respect to toxicity. Whole body Ni bioaccumulation followed Michaelis–Menten kinetics in all organisms, with greater hardness tending to decrease B_max with no consistent effect on K_d. Across species, acute Ni LC50 values tended to increase with both K_d and B_max values — i.e. more sensitive species exhibited higher binding affinity and lower binding capacity for Ni, but there was no correlation with body size. With respect to biotic ligand modeling, log K_NiBL values derived from Ni bioaccumulation correlated well with log K_NiBL values derived from toxicity testing. Both whole body Na and Mg levels were disturbed, suggesting that disruption of ionoregulatory homeostasis is a mechanism of acute Ni toxicity. In L. stagnalis, Na depletion was a more sensitive endpoint than mortality, however, the opposite was true for the other organisms. This is the first study to show the relationship between Na and Ni.     

1,3-Bis(α-aminoisopropyl)benzene, meta-C6H4(CMe2NH2)2: An N,N-bridging and N,C,N-cyclometalating ligand

Saponification of the bis(carbamic acid ester) 1,3-C₆H₄(CMe₂NHCO₂Me)₂ (1), made by the addition of methanol to commercial 1,3-C₆H₄(CMe₂NCO)₂, yielded the meta-phenylene-based bis(tertiary carbinamine) 1,3-C₆H₄(CMe₂NH₂)₂ (2). Dinuclear [{(η⁴-1,5-C₈H₁₂)RhCl}₂{μ-1,3-C₆H₄(CMe₂NH₂)₂}] (3) resulted from the action of 2 on [{(η⁴-1,5-C₈H₁₂)Rh(μ-Cl)}₂] in toluene. Combination of 2 with PdCl₂ or K₂[PdCl₄] gave the dipalladium macrocycle trans,trans-[{μ-1,3-C₆H₄(CMe₂NH₂)₂}₂(PdCl₂)₂] (4) along with cyclometalated [{2,6-C₆H₃(CMe₂NH₂)₂-κN,κC¹,κN′}PdCl] (5). Substitution of PEt₃ for the labile chlorido ligand of 5 afforded [{2,6-C₆H₃(CMe₂NH₂)₂-κN,κC¹, κN′}Pd(PEt₃)]Cl (6). The crystal structures of the following compounds were determined: bis(carbamic acid ester) 1, ligand 2 as its bis(trifluoroacetate) salt [1,3-C₆H₄(CMe₂NH₃)₂](O₂CCF₃)₂, 2 · (HAc_f)₂, complexes 3 and 6, as well as 1,3-C₆H₄(CMe₂OH)₂ (the diol analogue of 2).     

X-ray crystal,and molecular,structure of 1,2,3-tri-O-acetyl-4,5-dideoxy-4-C-[(R)-phenylphosphinyl]-α-l-lyxofuranose: The first x-ray analysis of a pentofuranose having phosphorus in the hemiacetal ring

X-Ray crystallographic analysis was performed on the compound to which had been assigned the structure of 1,2,3-tri-O-acetyl-4,5-dideoxy-4-C-[(R)-phenylphosphinyl]-α-l-lyxofuranose. The results showed that the compound has the proposed configuration, the five-membered ring is in the E₃ conformation, with a tendency towards the ³T₂ form, the substituents at P-5 and C-5 are linked bisectionally, the acetoxyl group at C-2 and the methyl group at C-4 are linked quasiequatorially, and the acetoxyl group at C-3 is linked axially.