Radiation chemistry of 2'-deoxycytidylyl-(3'-5')-2'-deoxyguanosine and its sequence isomer in N2O- and O2-saturated solutions

The radiation chemistry of the dinucleoside monophosphate d(CpG) and its sequence isomer, d(GpC), has been examined in aqueous solutions saturated with either N2O or O2. The products were isolated using HPLC, and the major products were identified using proton NMR spectroscopy and mass spectrometry. The major products include 5,6-dihydroxy-5,6-dihydrouracil (glycol) derivatives, 5- and 6-hydroxycytosine substitution products, 1-carbamoyl-2-oxo-4,5-dihydroxyimidazolidine products, and the 8-hydroxyguanine substitution product. Both trans stereoisomers of the imidazolidine derivatives are obtained from d(CpG) as well as from its sequence isomer. These are prominent products when the irradiation is carried out in the presence of oxygen, but they are not observed in the absence of oxygen.     

Synthesis and characterization of camphorsulfonyl acetate of cellulose

Novel cellulose derivatives were prepared from reacting (1R)-(+)-camphor-10-sulfonic chloride (CSC) with cellulose acetate (CA) in acetone and triethylamine. The reaction conditions, including reaction time and reactant molar ratios, were optimized. The structure of the products was confirmed by means of 1H NMR, 13C NMR, FT-IR and elementary analysis. The techniques were also used to determine the degree of the substitution of camphorsulfonyl groups (DSCS). The data calculated from 1H NMR, 13C NMR, percent grafting (G %) and elementary analysis coincided with those from chemical analysis. Compared to cellulose acetate, the cellulose derivatives exhibited decreased thermal stability, improved solubility in organic solvents and enhanced enantioselectivity towards tyrosine isomers. The solubility and enantioselectivity increased with increasing degrees of camphorsulfonyl substitution.     

Molecular structure of fagopyritol A1 (O-alpha-D-galactopyranosyl-(1 --> 3)-D-chiro-inositol) by NMR

The molecular structure of fagopyritol A1, a novel galactopyranosyl cyclitol from buckwheat seeds, was determined to be O-alpha-D-galactopyranosyl-(1 --> 3)-D-chiro-inositol by 1H and 13C NMR. Fagopyritol A1 is a positional isomer of fagopyritol B1 (O-alpha-D-galactopyranosyl-(1 --> 2)-D-chiro-inositol), representing a different series of fagopyritol oligomers. Trimethylsilyl derivatives of both compounds have similar mass spectra, but each may be identified by different abundance ratios of fragments with m/z 305/318 and 318/319.     

Regioselective monoacylation of cyclomaltoheptaose at the C-2 secondary hydroxyl groups by the alkaline protease from Bacillus subtilis in nonaqueous media

Transesterification of cyclomaltoheptaose (beta-CD) with divinyl butanedioate, divinyl hexanedioate, and divinyl decanedioate, catalyzed by the alkaline protease from Bacillus subtilis in anhydrous DMF for 5 days, furnished the corresponding vinyl-beta-CD derivatives. The products were characterized by ESI-MS, (1)H NMR, (13)C NMR, IR, and DSC. The results indicated the products to be monosubstituted esters, with monoacylation occurring at the C-2 secondary hydroxyl groups of beta-CD. The regioselectivity of the monoacylation as catalyzed by alkaline protease was not affected by the chain length of the acyl donor.     

Configurations of polyunsaturated sesterterpenoids from the diatom, Haslea ostrearia

The partial configurations of C25 isoprenoid alkenes isolated from the diatom Haslea ostrearia Gaillon (Simonsen) have been established. A combination of NMR spectroscopy studies of the alkenes with chiral shift reagents in conjunction with soluble silver beta-diketonate complexes and enantioselective gas chromatography of oxidation products of the alkenes was used. Unexpected differences in highly branched isoprenoid isomer configurations were observed between different laboratory cultures of the alga.     

Novel cytotoxic bufadienolides derived from bufalin by microbial hydroxylation and their structure-activity relationships

Microbial transformation was used to prepare novel cytotoxic bufadienolides. Twelve products (3-14) were obtained from bufalin (1) by the fungus Mucor spinosus. Their structures were elucidated by high-resolution mass spectroscopy (HR-MS) and extensive NMR techniques, including 1H NMR, 13C NMR, DEPT, 1H-1H correlation spectroscopy (COSY), two dimensional nuclear Overhauser effect correlation spectroscopy (NOESY), heteronuclear multiple quantum coherence (HMQC), and heteronuclear multiple bond coherence (HMBC). Compounds 3, 4, 9 and 11-14 are new mono- or dihydroxylated derivatives of bufalin with novel oxyfunctionalities at C-1beta, C-7beta, C-11beta, C-12beta and C-16alpha positions. The in vitro cytotoxic activities against human cancer cell lines of 3-14, together with 16 biotransformed products derived from cinobufagin (15-30) were determined by the MTT method, and their structure-activity relationships (SAR) were discussed.     

Phospholipids chiral at phosphorus. Absolute configuration of chiral thiophospholipids and stereospecificity of phospholipase D

Separate diastereomers of 1,2-dipalmitoyl-sn-glycero-3- thiophosphoethanolamine ( DPPsE ) were prepared in 97% diastereomeric purity and characterized by 31P, 13C, and 1H nuclear magnetic resonance (NMR). The isomers hydrolyzed by phospholipases A2 and C specifically were designated as isomer B (31P NMR delta 59.13 in CDCl3 + Et3N ) and isomer A (59.29 ppm), respectively, analogous to the isomers B and A of 1,2-dipalmitoyl-sn-glycero-3- thiophosphocholine ( DPPsC ) [ Bruzik , K., Jiang , R.-T., & Tsai, M.-D. (1983) Biochemistry 22, 2478-2486]. Phospholipase D from cabbage was shown to be specific to isomer A of DPPsC in transphosphatidylation . The product DPPsE was shown to be isomer A. The absolute configuration of chiral DPPsE at phosphorus was elucidated by bromine-mediated desulfurization in H2 18O to give chiral 1,2-dipalmitoyl-sn-glycero-3-[18O]phosphoethanolamine ( [18O]DPPE) followed by 31 P NMR analysis [ Bruzik , K., & Tsai, M.-D. (1984) J. Am. Chem. Soc. 106, 747-754]. The absolute configuration of chiral DPPsC was elucidated by desulfurization in H2 18O mediated by bromine or cyanogen bromide to give chiral 1,2-dipalmitoyl-sn-glycero-3-[18O]phosphocholine ( [18O]DPPC), which was then converted to [18O]DPPE by phospholipase D with retention of configuration [ Bruzik , K., & Tsai, M.-D. (1984) Biochemistry (preceding paper in this issue)]. The results indicate that isomer A of both DPPsE and DPPsC is SP whereas isomer B is RP.     

Native corrinoids from Clostridium cochlearium are adeninylcobamides: spectroscopic analysis and identification of pseudovitamin B(12) and factor A

The corrinoids from the obligate anaerobe Clostridium cochlearium were extracted as a mixture of Co(beta)-cyano derivatives. From 50 g of frozen cells, approximately 2 mg (1.5 micromol) of B(12) derivatives was obtained as a crystalline sample. Analysis of the corrinoid sample of C. cochlearium by a combination of high-pressure liquid chromatography and UV-Vis absorbance spectroscopy revealed the presence of three cyano corrinoids in a ratio of about 3:1:1. The spectroscopic data acquired for the sample indicated the main components to be pseudovitamin B(12) (Co(beta)-cyano-7"-adeninylcobamide) (60%) and factor A (Co(beta)-cyano-7"-[2-methyl]adeninylcobamide) (20%). Authentic pseudovitamin B(12) was prepared by guided biosynthesis from cobinamide and adenine. Both pseudovitamin B(12) and its homologue, factor A, were subjected to complete spectroscopic analysis by UV-Vis, circular dichroism, mass spectrometry, and by one- and two-dimensional (1)H, (13)C-, and (15)N nuclear magnetic resonance (NMR) spectroscopy. The third component was indicated by the mass spectra to be an isomer of factor A and is likely (according to NMR) to be 7"-[N(6)-methyl]-adeninylcobamide, a previously unknown corrinoid. C. cochlearium thus biosynthesizes as its native "complete" B(12) cofactors the 7"-adeninylcobamides and two homologous corrinoids, in which the nucleotide base is a methylated adenine.     

Synthesis of D-ring-substituted (5'R)- and (5'S)-17β-pyrazolinylandrostene epimers and comparison of their potential anticancer activities

Various steroidal benzylidenes were synthetized from pregnenolone with benzaldehyde and p-substituted benzaldehydes. The resulting 17β-chalconyl derivatives of pregnenolone were reacted with hydrazine hydrate in acetic acid solution. Regardless of the starting material, the ring-closure reaction afforded (in contrast with the literature data) a mixture of two steroidal pyrazoline epimers. The epimers were critical isomer pairs, which could be separated only in their acetylated form; their structures were investigated by NMR techniques. The in vitro inhibition of rat testicular C(17,20)-lyase activity and the antiproliferative effects on four human cancer cell lines were measured, and the results obtained from the two epimer series were compared.     

Identification and characterization of a novel linkage isomerization in the reaction of trans-diamminedichloroplatinum(II) with 5'-d(TCTACGCGTTCT)

The oligonucleotide 5'-d(TCTACGCGTTCT) reacts with trans-diamminedichloroplatinum(II) to yield primarily trans-[Pt(NH3)2[d(TCTACGCGTTCT)-N7-G(6),N7-G(8)]], containing the desired trans-[Pt(NH3)2[d(GCG)]] 1,3-cross-link. A key element of the platination reaction is the use of low pH to suppress coordination at A(4). The product was fully characterized by pH-dependent NMR titrations, enzymatic degradation analysis, and 195Pt NMR spectroscopy. Interestingly, the 1,3-cross-linked adduct is unstable at neutral pH, rearranging unexpectedly to form the linkage isomer trans-[Pt(NH3)2[d-(TCTACGCGTTCT)-N3-C(5),N7-G(8)]]. This rearrangement product is more stable than the initially formed isomer and could be characterized by pH-dependent NMR titrations, enzymatic degradation analysis, liquid secondary ion mass spectrometric analysis of an enzymatically digested fragment, 195Pt NMR spectroscopy, and modified Maxam-Gilbert footprinting experiments. By contrast, the 1,3-intrastrand cross-linked isomer rearranges during the course of both pH titration and enzymatic degradation experiments to form the 1,4-adduct. The equilibrium constant for this rearrangement is approximately 3, favoring the 1,4-adduct. Kinetic studies of the linkage isomerization reaction reveal t1/2 values for the first-order disappearance of the 1,3-intrastrand cross-linked isomer ranging from 129 (at 30 degrees C) to 3.6 h (at 62 degrees C), with activation parameters delta H not equal to = 91 +/- 2 kJ/mol and delta S not equal to = -58 +/- 8 J/(mol.K). Mechanistic implications of these kinetic results as well as the general relevance of this linkage isomerization reaction to platinum-DNA chemistry are briefly discussed.     

Effects of backbone structures and stereospecificities of lipid A-subunit analogues on their biological activities

Among chemically synthesized analogues corresponding to the nonreducing sugar part of lipid A, we have found an analogue (GLA-27) which exhibits Limulus, mitogenic, polyclonal B cell activation (PBA), interferon-inducing, and tumor necrosis factor (TNF)-inducing activities but not pyrogenic activity. The structure of GLA-27 comprises 4-O-phosphono-D-glucosamine with tetradecanoyl and 3-tetradecanoyloxytetradecanoyl (C14-O-(C14] groups as the 3-O- and 2-N-acyl substituents, respectively. Derivatives of GLA-27 with different backbone structures, such as the 1-deoxy, 3-epimeric, 3-amino, and 1-deoxy-3-epimeric derivatives of glucosamine, were chemically synthesized, and their mediator-inducing activities such as interferon- and TNF-inducing activities were investigated in comparison with their B cell activation activities including mitogenic and PBA activities. Among these derivatives, a derivative with a 1-deoxyglucosamine backbone (GLA-40) exhibited stronger B cell activation activities than those of GLA-27 while the mediator-inducing activities of GLA-40 were weaker than those of GLA-27. In addition to these derivatives, stereoisomers of GLA-27 which possess the (R) and (S) forms of C14-O-(C14) as the 2-N-acyl substituent were also synthesized and their biological activities compared. The (S) isomer exhibited much stronger mediator-inducing activities than the (R) isomer. On the other hand, B cell activation activities of the (R) isomer were strong and those of the (S) isomer weak. These results clearly demonstrate that mediator-inducing activities and B cell activation activities can be selectively expressed by modifying the structures of lipid A analogues.     

Racemization and hydrolysis of tropic acid alkaloids in the presence of cyclodextrins

Because of the constantly increasing demand for optically pure drugs it is of great importance to elucidate factors affecting stereochemistry, in order to provide a stable formulation with a high chiral quality of the desired isomer. Therefore, the effects of cyclodextrins (CyDs) and their alkylated and hydroxyalkylated derivatives on racemization and hydrolysis of (?)-(S)-hyoscyamine and (?)-(S)-scopolamine were examined kinetically and spectroscopically (NMR). Direct methods, based on a chiral and achiral chromatographic phase system, were used to determine their degradation products and enantiomer composition during stability tests. All different CyDs, except α-CyD, retarded racemization and hydrolysis. The inclusion of the drug substances in CyDs inhibits the attack of hydroxyl ions and/or water molecules and thus retards the racemization and hydrolysis. The racemization of the tropic acid alkaloids is dependent on the pH and temperature. NMR studies were used to evidence the formation of a soluble 1:1 complex in aqueous solution. © 1993 Wiley-Liss, Inc.     

Unexpected Shift to a 4-Imino Tautomer Upon N4-Acylation of 5-Methylcytosin-1-yl Nucleoside Analogues

Abstract

In contrast with the behaviour of 5-unsubstituted cytosine nucleoside analogues, 5-methylcytosine derivatives show upon N⁴-benzoylation, commonly used as base protection in oligonucleotide synthesis, a tautomeric change of the base moiety from a 4-amino- into a 4-imino isomer. In the latter form, which is easily diagnosticized by ¹³C NMR and confirmed by X-ray data, the compounds seem to be hydrolytically less stable.     

Preparation of soluble p-aminobenzoyl chitosan ester by Schiff's base and antibacterial activity of the derivatives

Schiff's base of chitosan (BCTS) was obtained by the reaction of chitosan (CTS) and benzaldehyde. Then BCTS reacted with acyl chloride which was synthesized by p-aminobenzoic acid and thionyl chloride to get N-benzoyl-O-aminobenzoyl chitosan ester (BABCTSE), removing the groups of amino protection of BABCTSE to get the final product (ABCTSE). The structures of the derivatives were characterized by FT-IR, (1)H NMR, (13)C NMR and elemental analysis. The elemental analysis results indicated that the degrees of substitution (DS) of the products were 16.8% and 40.4%. The synthesized compounds exhibited an excellent solubility in organic solvents. TG and DTG results showed that thermal stability of the derivatives was lower than that of chitosan. In addition, the existence of two different amido in the molecular structures contributed to forming more -NH(3)(+) in the acid solution which could make the derivatives have a greater advantage in the field of bacteriostasis.     

Proton and carbon-13 nuclear magnetic resonance spectroscopy of diastereoisomeric 3- and 17 beta-tetrahydropyranyl ether derivatives of estrone and estradiol

Protection of 3- and 17 beta-hydroxyl groups of estrone and estradiol as tetrahydropyranyl ether derivatives led to mixtures of 2'(R)- and 2'(S)-diastereoisomers which were separated by crystallization (3-tetrahydropyranyl ethers), or by thin-layer chromatography (17-tetrahydropyranyl ethers), and characterized by 1H and 13C nuclear magnetic resonance (NMR). Assignments for NMR signals of estradiol 3,17 beta-ditetrahydropyranyl ether were facilitated by comparison with those of its 15 zeta, 16 zeta-dideuterio analog and by 2D 1H-13C heteroshift correlation experiments. Diastereoisomers of 3-tetrahydropyranyl ether derivatives could be identified through the 13C NMR doublet signals of the anomeric C-2' and the aromatic C-4 carbon atoms in CDCl3. Diastereoisomers of 17-tetrahydropyranyl ether derivatives were recognized from characteristic modifications of 1H NMR signals of H-2', H-6', H-1, H-17, and 18-CH3 protons as well as from the 13C NMR doublet signals corresponding to C-2', C-4', C-6', C-12, C-13, C-16, and C-17 carbon atoms. Low-temperature experiments showed a splitting of the C-2', C-6', and C-17 13C NMR signals of each of the two 17-tetrahydropyranyl ether isomers. The downfield signal (equatorial conformer) of the three resulting doublets was more intense for the 17-tetrahydropyranyl ether 2'(S)-isomer, whereas the upfield signal (axial conformer) was more intense for the 2'(R)-isomer.     

Isolation and Characterization of Two Geometric Allene Oxide Isomers Synthesized from 9S-Hydroperoxylinoleic Acid by Cytochrome P450 CYP74C3: STEREOCHEMICAL ASSIGNMENT OF NATURAL FATTY ACID ALLENE OXIDES*

Specialized cytochromes P450 or catalase-related hemoproteins transform fatty acid hydroperoxides to allene oxides, highly reactive epoxides leading to cyclopentenones and other products. The stereochemistry of the natural allene oxides is incompletely defined, as are the structural features required for their cyclization. We investigated the transformation of 9S-hydroperoxylinoleic acid with the allene oxide synthase CYP74C3, a reported reaction that unexpectedly produces an allene oxide-derived cyclopentenone. Using biphasic reaction conditions at 0 °C, we isolated the initial products and separated two allene oxide isomers by HPLC at −15 °C. One matched previously described allene oxides in its UV spectrum (λ_max 236 nm) and NMR spectrum (defining a 9,10-epoxy-octadec-10,12Z-dienoate). The second was a novel stereoisomer (UV λ_max 239 nm) with distinctive NMR chemical shifts. Comparison of NOE interactions of the epoxy proton at C9 in the two allene oxides (and the equivalent NOE experiment in 12,13-epoxy allene oxides) allowed assignment at the isomeric C10 epoxy-ene carbon as Z in the new isomer and the E configuration in all previously characterized allene oxides. The novel 10Z isomer spontaneously formed a cis-cyclopentenone at room temperature in hexane. These results explain the origin of the cyclopentenone, provide insights into the mechanisms of allene oxide cyclization, and define the double bond geometry in naturally occurring allene oxides.     

Synthesis of sugar-lactams from azides of glucuronic acid

Sugar-lactams have found application as glycosidase inhibitors, synthetic precursors of iminosugars and they are structural components of natural products. The synthesis of beta-D-glucopyranosidurono-6,1-lactams from glucuronic acid derivatives are described. NMR data and X-ray crystal structures indicate that the sugar-lactams adopt distorted (1)C4 conformations in solution and in the solid state.     

Detection of D-erythro and L-threo sphingosine bases in preparative sphingosylphosphorylcholine and its N-acylated derivatives and some evidence of their different chemical configurations

Sphingosylphosphorylcholine prepared from native sphingomyelin by the Kaller procedure was found to comprise about 70% of the L-threo (2S, 3S) isomer and 30% of the D-erythro (2S, 3R) isomer. This analytical result was obtained by gas-liquid chromatography (GLC) of trimethylsilyl derivatives of N-acetylsphingosines which were prepared by enzymatic hydrolysis of synthetic N-acetylsphingosylphosphorylcholines with Clostridium perfringens phospholipase C. Some other evidence of the different chemical configuration between the erythro and threo isomers of synthetic N-acylated sphingosylphosphorylcholines was also provided by thin layer chromatography (TLC), optical rotatory dispersion (ORD), and fast atom bombardment (FAB) mass spectrometry.     

Homogeneous modification of cellulose with succinic anhydride in ionic liquid using 4-dimethylaminopyridine as a catalyst

Cellulose, extracted from sugarcane bagasse, was successfully succinylated in ionic liquid 1-buty-3-methylimidazolium (BMIMCl) using 4-dimethylaminopyridine (DMAP) as a catalyst. Parameters investigated included the mass ratio of DMAP/succinic anhydride in a range from 0% to 15%, reaction time (from 30 to 120 min), reaction temperature (from 60 to 110 °C). The succinylated cellulosic derivatives had a degree of substitution (DS) ranging from 0.24 to 2.34. It was found that the DS of succinylated cellulosic derivatives using DMAP as a catalyst was higher than that without any catalyst under the same reaction conditions. The products were characterized by FT-IR, solid-state CP/MAS ¹³C NMR, and thermal analysis. FT-IR and solid-state CP/MAS ¹³C NMR spectra showed that succinoylation occurred at C-6, C-2 and C-3 positions. The thermal stability of the succinylated cellulose decreased upon chemical modification.     

A direct synthetic approach to uracil anhydrothionucleoside derivatives

Direct conversion of peracylated N¹-(β-d-glucopyranosyl)-2-thiouracil derivatives into the corresponding anhydrothionucleosides has been studied under various conditions including: gas-phase pyrolysis, heating without a solvent, and by heating in a solvent of high boiling point (DPE) in the presence of a base (DABCO) and reaction in a microwave reactor. Heating at 210-220 °C was found to give the best yield of a single isomer. The structures of the new anhydrothionucleosides were confirmed by NMR techniques.