O-methyl Analogues Research Articles

Total synthesis and biological evaluation of oscillatoxins D, E, and F.

Oscillatoxins (OTXs) and aplysiatoxins are biosynthetically related polyketides produced by marine cyanobacteria. We previously developed a synthetic route to phenolic O-methyl analogs of OTX-D and 30-methyl-OTX-D during collective synthesis of these natural products. According to our synthetic strategy, we achieved total synthesis of OTX-D, 30-methyl-OTX-D, OTX-E, and OTX-F by deprotecting the O-methyl group in an earlier intermediate, and determined their biological activities. Although OTX-D and 30-methyl-OTX-D have been reported to show antileukemic activity against L1210 cell line, we found that their cytotoxicity in vitro against this cell line is relatively weak (IC50: 29-52 µm). In contrast, OTX-F demonstrated cell line-selective antiproliferative activity against DMS-114 lung cancer cells, which implies that OTXs target as yet unknown target molecules as part of this unique activity.

Large-Scale Isolation of Three O-Methyl Anthocyanins from Bilberry (Vaccinium myrtillus L.) Extract.

Three O-methyl anthocyanidin 3-O-β-D-glucopyranosides were isolated from bilberry extract on a large-scale basis together with two non O-methyl analogues. Anthocyanidin 3-O-β-D-galactopyranosides were removed from bilberry extract together with parts of anthocyanidin 3-O-α-L-arabinopyranosides after treatment with β-galactosidase. The remaining arabinopyranosides were removed by applying acid catalytic hydrolysis. The amounts of anthocyanins recovered as flavylium trifluoroacetic acid salt were as follows: 630 mg for petunidin 3-O-β-D-glucopyranoside, 423 mg for peonidin 3-O-β-D-glucopyranoside, 588 mg for malvidin 3-O-β-D-glucopyranoside, 877 mg for delphinidin 3-O-β-D-glucopyranoside, and 742 mg for cyanidin 3-O-β-D-glucopyranoside.

Substrate-Assisted Hydroxylation and O-Demethylation in the Peroxidase-like Cytochrome P450 Enzyme CYP121.

CYP121 is a P450 enzyme from Mycobacterium tuberculosis that catalyzes a C-C coupling reaction between the two aromatic rings on its native substrate cyclo(l-Tyr-l-Tyr) (cYY) to form mycocyclosin, a necessary product for cell survival. Unlike the typical P450 enzymes for hydroxylation, CYP121 is believed to behave like a peroxidase and conduct radical-mediated C-C bond formation. Here, we probe whether the phenolic hydrogen of the substrate is the site of the postulated hydrogen atom abstraction for radical formation. We synthesized a singly O-methylated substrate analogue, cYF-4-OMe, and characterized its interaction with CYP121 by ultraviolet-visible and electron paramagnetic resonance spectroscopies and X-ray crystallography. We found that cYF-4-OMe can function as a substrate of CYP121 using the established assay via the peroxide shunt. Analysis of the enzymatic reaction revealed an O-demethylation of cYF-4-OMe instead of cyclization, yielding cYY and formaldehyde. A hydroxylated substrate, cYF-4-OMeOH, is expected to be the intermediate product, which was trapped and structurally characterized by X-ray crystallography. We further determined that the deformylation reaction of cYF-4-OMeOH proceeds via an alkyl-oxygen rather than aryl-oxygen bond cleavage by the 18O-labeling studies. Finally, the pH dependence catalytic study on the native substrate and the methoxy analogue further supports the mechanistic understanding that the hydrogen atom abstraction is the critical first oxidation step exerted by a heme-based oxidant during the cyclization reaction of cYY. The switch in catalytic activity reveals the power of CYP121 as a P450 enzyme and provides insight into the peroxidase-like catalytic mechanism.

Synthesis and characterization of 4′-C-guanidinomethyl-2′-O-methyl-modified RNA oligomers

This study investigated the synthesis and properties of 4′-C-guanidinomethyl-2′-O-methyluridine and RNAs containing the analog. Thermal and thermodynamic stabilities of double-stranded RNAs (dsRNAs) containing the nucleoside analog were examined. It was found that although the analog decreased the thermal and thermodynamic stabilities of dsRNA, it had base-discrimination ability. The 4′-C-guanidinomethyl modification increased stability of RNAs in a buffer containing serum. Furthermore, small interference RNAs incorporating one analog at the passenger strand still preserved their RNA interference activities. It was suggested that the 4′-guanidinomethyl modification significantly improved cell membrane permeability of RNA. Thus, 4′-C-guanidinomethyl-2′-O-methyl analogs may be useful in improving the properties of therapeutic siRNA molecules.

A Highly Sensitive ESIPT-Based Ratiometric Fluorescence Sensor for Selective Detection of Al(3.).

An excited-state intramolecular proton transfer (ESIPT)-based highly sensitive ratiometric fluorescence sensor, 1H was developed for selective detection of aluminum (Al(3+)) in acetonitrile as well as in 90% aqueous system. Single-crystal X-ray diffraction analysis reveals almost planar and conjugated structure of 1H. Photophysical properties of the sensor as well as its selectivity toward Al(3+) are explored using UV-visible, steady-state, and time-resolved fluorescence spectroscopic studies. The bright cyan (λem = 445 nm) fluorescence of 1H in acetonitrile turns into deep blue (λem = 412 nm) with ∼2.3-fold enhancement in emission intensity, in the presence of parts per billion level Al(3+) (detection limit = 0.5 nM). Interestingly, the probe 1H exhibits increased selectivity toward Al(3+) in H2O/acetonitrile (9:1 v/v) solvent system with a change in fluorescence color from pale green to deep blue associated with ca. sixfold enhancement in emission intensity. Density functional theoretical (DFT) calculations provide the ground- and excited-state energy optimized structures and properties of the proposed aluminum complex [Al(1) (OH)]2(2+), which is in harmony with the solution-state experimental findings and also supports the occurrence of ESIPT process in 1H. The ESIPT mechanism was also ascertained by comparing the basic photophysical properties of 1H with a similar O-methylated analogue, 1'Me.

Convenient Syntheses of 3-Aryl-3,4-Dihydroisocoumarins

A convenient method for the synthesis of naturally occurring 3,4-dihydroisocoumarins by judicious use of Friedel-Craft acylation, regioselective formylation and oxidative cyclisation reactions is described. O-methylated analogues of montroumarin and thunberginols C and D and have been synthesised using 3,5-dimethoxyphenylacetic acid as a starting material. Furthermore, the starting material 3,5-dimethoxyphenylacetic acid was synthesised by a route involving an oxidative Nef reaction.

Development of a visible nanothermometer with a highly emissive 2′-O-methylated guanosine analogue

We have synthesized a fluorescent base analogue, 2-aminothieno[3,4-d]pyrimidine based G-mimic deoxyribonucleoside, 2′-OMe-thG, and investigated its photophysical properties and DNA incorporation.

Biological evaluation and SAR analysis of O-methylated analogs of quercetin as inhibitors of cancer cell proliferation.

Using a high-throughout screening approach, the anticancer activities of 16 O-methylated (OMe) analogs of quercetin were assessed. The structure-activity relationships showed that OMe moieties at the 4' and/or 7 positions were important for maintaining inhibitory activities against the 16 cancer cell lines. Furthermore, when the OH groups at the 3' and 4' positions were both replaced by OMe moieties, anticancer activity was enhanced.

99mTc-MAG3-Aptamer for Imaging Human Tumors Associated with High Level of Matrix Metalloprotease-9

The human matrix metalloprotease 9 (hMMP-9) is involved in many physiological processes such as tissue remodeling. Its overexpression in tumors promotes the release of cancer cells thus contributing to tumor metastasis. It is a relevant marker of malignant tumors. We selected an RNA aptamer containing 2'-fluoro, pyrimidine ribonucleosides, that exhibits a strong affinity for hMMP-9 (K(d) = 20 nM) and that discriminates other human MMPs: no binding was detected to either hMMP-2 or -7. Investigating the binding properties of different MMP-9 aptamer variants by surface plasmon resonance allowed the determination of recognition elements. As a result, a truncated aptamer, 36 nucleotides long, was made fully resistant to nuclease following the substitution of every purine ribonucleoside residue by 2'-O-methyl analogues and was conjugated to S-acetylmercaptoacetyltriglycine for imaging purposes. The resulting modified aptamer retained the binding properties of the originally selected sequence. Following (99m)Tc labeling, this aptamer was used for ex vivo imaging slices of human brain tumors. We were able to specifically detect the presence of hMMP-9 in such tissues.

Metabolism-based synthesis, biologic evaluation and SARs analysis of O-methylated analogs of quercetin as thrombin inhibitors

In blood, quercetin is mainly found in metabolized forms. In order to study the activities of these quercetin metabolites in cardiovascular disease, 17 methylquercetin derivatives were synthesized based on metabolism in vivo, their thrombin inhibition activity were evaluated through the analyzation of prothrombin time (PT), activated partial thromboplastin time (APTT), thrombin time (TT) and fibrinogen (FIB). The results showed that 6 methylquercetin derivatives had stronger inhibitory activities than that of quercetin. Preliminary SARs analysis showed that hydroxyl groups at C-3′ and C-4′ position in the B-ring and hydroxyl group at C-3 position in the C-ring played key roles in the thrombin inhibitory activity. The findings of this study would provide information for the exploitation and utilization of quercetin as thrombin inhibitor for thrombotic disease treatment.

Synthesis, free solution capillary electrophoresis separation and toxicity of seven potential impurities of dobutamine.

The seven O-methylated analogs of dobutamine [(+/-)-4-[2-[[3-(p-hydroxyphenyl)-1-methylpropyl]amino]ethyl]pyrocatechol, CAS 34368-04-2), a trihydroxy secondary amine, can be considered potential impurities of the latter, the ultimate step of the synthesis of dobutamine being the deprotection of the three phenolic groups. In order to enable the detection and the identification of such a contamination, the di- and the monomethylated derivatives of dobutamine were prepared and their mixture with dobutamine and its trimethoxy precursor completely resolved by Free Solution Capillary Electrophoresis (FSCE). Indeed, the above impurities proved to occur in dobutamine in consequence of improper demethylation conditions and to be significantly more toxic than the latter.

Synthesis, Photochemistry, and Photophysics of Butadiene Derivatives: Influence of the Methyl Group on the Molecular Structure and Photoinduced Behavior

Novel butadiene derivatives display diverse photochemistry and photophysics. Excitation of 2-methyl-1-(o-vinylphenyl)-4-phenylbutadiene leads to the dihydronaphthalene derivative, whereas photolysis of the corresponding model o-methyl analogue results in the formation of the naphthalene-like derivative, deviating from the nonmethylated analogue of the prior starting compound and producing benzobi- and -tricyclic compounds. The effect of the methyl substituents is even more dramatic in the case of the dibutadienes. The parent unsubstituted compound undergoes photoinduced intramolecular cycloaddition giving benzobicyclo[3.2.1]octadiene, whereas the photochemical reaction of the corresponding dimethylated derivative shows only geometrical isomerization due to the steric effect of the substituents. Methyl groups on the butadiene backbones reduce the extent of conjugation, causing a blue-shift of the characteristic absorption band. The fluorescence efficiency is dramatically decreased, as a consequence of nonplanarity and reduced rigidity of the molecules due to the crowding by the methyl and phenyl groups together. Four molecules of very similar structures show dramatically different photoinduced behavior, revealing how changes of the nature and position of the substituents are valuable in understanding the photophysics and photochemistry of these types of compounds.

ChemInform Abstract: Torsionally and Hydrophobically Modified 2,3-Diarylindenes as Estrogen Receptor Ligands.

2,3-Diarylindenes are ligands for the estrogen receptor which display intrinsic fluorescence. In order to optimize the receptor binding affinity of these compounds while preserving their desirable fluorescence properties, a series of torsionally modified analogues were prepared. A fluorine or methyl group was introduced on either of the two phenyl substituents ortho to their attachment site to the indene nucleus, in order to increase the out-of-plane twist of the appended rings. The analogues were prepared by the benzylation of appropriate deoxybenzoins, followed by Friedel-Crafts cyclic alkylation-dehydration. Comparison of the X-ray crystal structure of one analogue with unsubstituted analogues confirms the torsional perturbation effected by the ortho substituent. The torsional disposition of the C-2 aryl group in the substituted diphenylindenes is further investigated by UV (absorbance maxima and molar absorptivities), fluorescence (Stokes' shift), and NMR (chemical shifts). These spectroscopic measurements indicate increasing twisting between the C-2 aryl substituent and the indene system according to the following order: 3-ring o-Me-indene 9f less than diphenylindene 15 = 20 degrees less than 3-ring o-F-indene 9c less than 1-Me-indene 16 less than 2-ring o-F-indene 9b less than 2-ring o-Me-indene 9e = 63 degrees. The binding affinity of these analogues to the estrogen receptor was evaluated by a competitive radiometric receptor binding assay. While o-fluoro or o-methyl substitution on the 3-ring increases binding only slightly, binding of the o-fluoro 2-ring analogue is increased ca. 6-fold and the o-methyl analogue 11-fold, giving, in the latter case, a compound with an affinity equivalent to that of estradiol. The increase in binding affinity afforded by ortho substitution correlates with the increase in the torsion angle of the C-2 aryl ring. A thermodynamic evaluation of the receptor fit (Andrews, P. R.; Craik, D. J.; Martin, J. L. J. Med. Chem. 1984, 27, 1648) indicates that, for the o-methyl 2-ring analogue, the effect of the ortho substitution on increasing receptor binding appears to be a combination of increased surface area due to the substituent itself, together with a change in surface area of the ligand that results from the increased torsion of the two aryl rings. An o-fluoro substituent on the 2-ring provides a compromise between the relative planarity required for high fluorescence intensity and the molecular shape needed for increased estrogen receptor binding affinity.(ABSTRACT TRUNCATED AT 400 WORDS)

Semisynthetic latrunculin B analogs: Studies of actin docking support a proposed mechanism for latrunculin bioactivity

Latrunculins are unique macrolides containing a thiazolidinone moiety. Latrunculins A, B and T and 16-epi-latrunculin B were isolated from the Red Sea sponge Negombata magnifica. N-Alkylated, O-methylated analogs of latrunculin B were synthesized and biological evaluation was performed for antifungal and antiprotozoal activity. The natural latrunculins showed significant bioactivity, while the semisynthetic analogs did not. Docking studies of these analogs into the X-ray crystal structure of G-actin showed that, in comparison with latrunculins A and B, N-alkylated latrunculins did not dock satisfactorily. This suggests that the analogs do not fit well into the active site of G-actin due to steric clashes and provides an explanation for the absence of bioactivity.

Selective Protection of Nuclease-Sensitive Sites in siRNA Prolongs Silencing Effect

Small interfering RNAs (siRNAs) are considered as potent agents for specific gene silencing; however, nuclease sensitivity of siRNA limits their biomedical applications. Till date, no universal methodology has been developed to improve the nuclease resistance of siRNA, preserving low toxicity and high activity. In this study, we proposed an algorithm for the site-specific modification of siRNAs based on the mapping of their nuclease-sensitive sites in the presence of serum followed by the incorporation of 2'-O-methyl analogs of ribonucleotides at the identified positions of cleavage. We found that the protection of nuclease-sensitive sites considerably enhanced nuclease resistance of siRNA and only slightly reduced the efficiency of silencing. Modification of all nuclease-sensitive sites prolonged the duration of the silencing effect of the siRNA compared to nonmodified, partially modified, or randomly modified siRNA of the same sequence. This study showed that the targeted chemical modification of nuclease-sensitive sites could provide highly efficient siRNA-based therapeutics for the control of disease-related genes.

Mechanistic studies on the synthesis of bicalutamide

Bicalutamide, a therapeutically important anti-androgen used in the treatment of hormone-sensitive cancers, may be synthesised from the appropriate halohydrin or epoxide. We report here studies aimed at demonstrating unambiguously that preparation of bicalutamide and its thioether analogue from the chlorohydrin under basic conditions proceeds via opening of an intermediate epoxide by the appropriate sulfinate or thiolate nucleophile, that the analogous anionic sulfur nucleophiles react under the same conditions and that the S(N)2 pathway involving direct displacement of chloride by the nucleophile does not operate. The proposed mechanism is confirmed by the quantitative fitting of sequential reaction kinetics, taking into account the competing dimerisation of the thiolate nucleophile that occurs under basic conditions. The O-methyl analogue of the chlorohydrin is unreactive towards thiolate under the same conditions, although a slower cyclisation to the beta-lactam was observed. The implications of these observations for the analogous preparation of thioethers and sulfones are discussed.

Effect on both aglycone and sugar moiety towards Phase II metabolism of anthocyanins

The effect of sugar moiety on anthocyanin metabolism was studied using anthocyanidin 3-rutinosides (cyanidin 3-O-rutinoside (Cy3R) and delphinidin 3-O-rutinoside (Dp3R)) and 3-O-glucosides (delphinidin 3-O-glucoside (Dp3G)). O-methylated Cy3R and Dp3R were detected in rat blood plasma after oral administration of Cy3R and Dp3R (100mg/kg body weight). On the basis of HPLC retention time and UV–visible spectra together with the data of our previous studies on the hydrophobic metabolites of anthocyanidin 3-O-glucosides, it was concluded that both 3′- and 4′-O-methyl Cy3R were metabolites of Cy3R. On the other hand, only 4′-O-methyl Dp3R was detected as hydrophobic metabolite of Dp3R. A group of hydrophilic metabolites was also detected in rat blood plasma after oral administration of anthocyanins (Dp3G, Cy3R and Dp3R) and their structures were determined to be extended glucuronides and their O-methyl analogues by tandem MS analysis. The amounts of extended glucuronides of Dp3G, Cy3R and Dp3R were less than those of cyanidin 3-O-glucoside (Cy3G) reported in our previous study. On the other hand, anthocyanidin–glucuronides (both cyanidin–glucuronide and delphinidin–glucuronide) were not detected after oral administration of Cy3R, Dp3R and Dp3G. These results indicated that both the type of sugar moiety and stability of aglycone largely affected phase II metabolism of anthocyanins, and also indicated that the type of sugar moiety did not affect the O-methylation metabolism but affected glucuronyl conjugation in both liver and small intestine.

Synthesis of a Complete Series of O-Methyl Analogues of Naringenin and Apigenin

Flavonoids have a variety of biological effects in numerous mammalian systems in vitro as well as in vivo. Most of the in vitro studies are performed on flavonoid itself and not on its metabolites as these latter are not readily available from commercial sources. However, no systematic preparation of flavonoid metabolites has been reported presumably due to the difficulty in controlling the regioselectivity. Particularly, regioselective synthesis of the 5-Omethylflavones or 5-O-methylflavanones has rarely been accomplished. It is well-known that the hydroxyl function on the 5 position of flavonoids resists to alkylation under basic conditions due to the acid-weakening effect of an intramolecular H-bond between the 5-hydroxyl group and the 4-keto group. The aim of this study was thus to prepare a complete series of O-methyl naringenins (flavanones) and apigenins (flavones) with emphasis on the syntheses of 5-Omethyl analogues. For the purpose of preparing every possible O-methyl analogues of naringenin and apigenin, various regioselective methylation and protection was attempted by using narinenin as a starting material (Scheme 1). The hydroxyl function on the 7 position of naringenin is the most acidic site among its three hydroxyl groups (7-OH, 5-OH and 4'OH). Thus, treatment of naringenin with equimolar amount of the alkylating reagents such as Me2SO4 and BzCl under basic conditions gave the corresponding 7-O-alkylated products 1 and 11, from which a variety of O-methyl naringenins and apigenins were prepared. Oxidation of 1 with DDQ provided 7-O-methylapigenin 2. On the other hand, treatment of 1 with Me2SO4 and TBDMSCl provided 7,4'di-O-methyl naringenin 3 and 7-O-methyl-4'-O-tert-butyldimethylsilyl naringenin 4, respectively. It is worth to note that, compared with dimethylation of naringenin by use of excess amount of Me2SO4, stepwise methylation of naringenin via 7-O-methyl analogue 1 always provided the 7,4'O-dimethylated naringenin 3 in higher yield. With these monoand dimethylated naringenin analogues 3 and 4 in our hands, we set out divergent syntheses of 5-O-methyl apigenins 9 and 10: methylation followed by oxidation (upper routes: 3 → 5 → 9 and 4 → 7 → 10; Scheme 2) and the other way around (lower routes: 3 → 6 → 9 and 4 → 8 → 6,10; Scheme 2). First, treatment of 3 and 4 with excess amount of Me2SO4 provided the corresponding 5-O-methyl naringenin analogues 5 and 7, respectively, which were then treated with DDQ in refluxing 1,4-dioxane to give clean conversions to the corresponding 5-O-methyl apigenin analogues 9 and 10. 5,7,4'-Trimethyl apigenin 9 was also prepared by methylation of 6 which was obtained by DDQoxidation of 3. The H NMR spectrum of 9 was found identical to the literature data. The structure of 9 was further characterized by 2DNOESY experiment in which irradiations of 5-O-Me, 7-OMe and 4'-O-Me resulted in increases of H-6, H-6/H-8, and H-3' resonance frequencies, respectively. Throughout this study, these characteristic nuclear Overhauser effects between adjacent protons were used to unequivocally identify the positions of the O-methyl groups. On the other hand, methylation of 8 gave an inseparable mixture of apigenin analogues which were purified after removal of protecting groups by TBAF in THF to give two dimethylated apigenin analogues 6 and 10. Presumably, the compound 6 was formed by partial loss of the 4'-O-TBS group followed by methylation. Thus, we tried to install other protecting groups such as benzyl or benzoyl groups at the 4' position but, in these cases, DDQ-oxidation did not proceed at all (data not shown), which suggests the 4'-OTBS group as the orthogonal protecting group of choice to the 7-O-Me of naringenin.

Syntheses of Morphine and Codeine (1992‐2002): Templates for Exploration of Synthetic Tools

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Chemical transformations of oxyresveratrol ( trans-2,4,3′,5′-tetrahydroxystilbene) into a potent tyrosinase inhibitor and a strong cytotoxic agent

From oxyresveratrol ( trans-2,4,3′,5′-tetrahydroxystilbene 1), seven derivatives were prepared, including trans-2-methoxy-4,3′,5′-trihydroxystilbene ( 2), trans-2,3′-dimethoxy-4,5′-dihydroxystilbene ( 3), trans-4,3′-dimethoxy-2,5′-dihydroxystilbene ( 4), trans-2,4,3′,5′-tetramethoxystilbene ( 5) and cis-2,4,3′,5′-tetramethoxystilbene ( 6), 2,4,3′,5′-tetrahydroxybibenzyl ( 7), and 2,4,3′,5′-tetramethoxybibenzyl ( 8). The tetrahydroxybibenzyl 7, a hydrogenation product of 1, exhibited more potent tyrosinase inhibitory activity than the parent compound, without cytotoxicity. A kinetic study revealed that 7 was a reversible and non-competitive inhibitor of mushroom tyrosinase with l-dopa as the substrate. Analysis of the K i values indicated that 7 has a slightly higher affinity to the enzyme than 1. Compound 6, a tetra- O-methylated analogue of 1 with cis-configuration, was deprived of inhibitory effect on the enzyme tyrosinase, but showed very strong cytotoxicity against the human cancer cells KB, BC, and NCI-H187, with potency comparable to those of the anticancer agents ellipticine and doxorubicin. Data on the tyrosinase inhibitory activity and cytotoxicity of 1– 8 indicated that O methylation on stilbene 1 destroyed anti-tyrosinase activity but generated cytotoxicity. Thus, facile preparations of a potent tyrosinase inhibitor ( 7) and a strong cytotoxic agent ( 6) from the natural product 1 were achieved through simple chemical reactions.