Synthesis of 3-[4'-(p-chlorophenyl)-thiazol-2'-yl]-2-[(substituted azetidinone/thiazolidinone)-aminomethyl]-6-bromoquinazolin-4-ones as anti-inflammatory agent

N-Chloroacetyl-5-bromoanthranilic acid (1), 3-[4'-(p-chlorophenyl)-thiazol-2'-yl]-2-chloromethyl-6-bromoquinazolin-4-one (2), 3-[4'-(p-chlorophenyl)-thiazol-2'-yl]-2-hydrazinomethyl-6-bromoquinazolin-4-one (3), 3-[4'-(p-chlorophenyl)-thiazol-2'-yl]-2-substitutedbenzylidene aminomethyl-6-bromoquinazolin-4-ones (4-11), 2-[(4'-oxo-3'-chloro-2'-phenylazetidin-1'-yl)aminomethyl]-3-[4'-(p-chlorophenyl)thiazol-2'-yl]-6-bromoquinazolin-4-ones (12-19) and 2-(4'-oxo-2'-phenyl-thiazolidin-3'-yl-aminomethyl)- 3-[4'-(p-chlorophenyl)-thiazol-2'-yl]-6-bromoquinazolin-4-ones (20-27) have been synthesized. All the compounds have been screened for their anti-inflammatory and analgesic activities at the dose of 50mg/kg po. Compound 21 showed maximum anti-inflammatory (38.35%) and analgesic (37.36%) activities. Compound 21 was also tested for ulcerogenic activity and the UD(50) value was found to be 195.6mg/kg po. The structure of all compounds has been evaluated by elemental analysis (C, H, N) and spectral analysis (IR, (1)H NMR and mass spectrometry).     

New phenolic compounds from Kokuto,non-centrifuged cane sugar

Five new phenolic compounds, 4-(beta-D-glucopyranosyloxy)-3,5-dimethoxyphenyl-propanone (8), 3-[5-[(threo) 2,3-dihydro-2-(4-hydroxy-3-methoxyphenyl)-3-hydroxymethyl-7-methoxybenzofuranyl]-propanoic acid (12), 2-[4-(3-hydroxy-1-propenyl)-2,6-dimethoxyphenoxy]-3-hydroxy-3-(4-hydroxy-3,5-dimethoxyphenyl)propyl-beta-D-glucopyranoside (13), 4-[(erythro) 2,3-dihydro-3(hydroxymethyl)-5-(3-hydropropyl)-7-methoxy-2-benzofuranyl]-2,6-dimethoxyphenyl-beta-D-glucopyranoside (14), 9-O-beta-D-xylopyranoside of icariol A2 (15), and known phenolic compounds were isolated from Kokuto, non-centrifuged cane sugar (Saccharum officinarum L.). Their structures were determined by a spectral investigation.     

Synthesis of some newer derivatives of substituted quinazolinonyl-2-oxo/thiobarbituric acid as potent anticonvulsant agents

5-[1'-[3"-Aminoacetyl-2"-methyl-6",8"-dihalosubstitutedquinazolin-4"(3"H)-onyl]-thiosemicarbazido]-2-oxo/thiobarbituric acids 3a-3h and 5-[2'-amino-5'-[3"-aminomethylene-2"-methyl-6",8"-dihalosubstitutedquinazolin-4"(3"H)-onyl]-1',3',4'-thiadiazol-2'-yl]-2-oxo/thiobarbituric acid 5a-5h were prepared by incorporating 1-[3'-aminoacetyl-2'-methyl-6",8"-dihalosubstituted-quinazolin-4'(3'H)-onyl]-thiosemicarbazides 2a-2d and 2-amino-5-[3'-aminomethylene-2'-methyl-6',8'-dihalosubstituted-quinazolin-4'(3'H)-onyl]-1,3,4-thiadiazoles 4a-4 h respectively at 5(th) position of 2-oxo/thiobarbituric acids (via Mannich reaction). All the newly synthesized compounds were screened for their anti-convulsant activity in MES and PTZ models and were compared with standard drugs phenytoin sodium and sodium valproate. Interestingly, these compounds were found to be devoid of sedative and hypnotic activities when tested. Out of the compounds studied, the most active compound 5h, that is 5-[2'-amino-5'-[3"-aminomethylene-2"-methyl-6",8"-dibromoquinazolin-4"(3"H)-onyl]-1',3',4'-thiadiazol-2'-yl]-2-thiobarbituric acid showed activity (90%) more potent than the standard drug.     

Plasmids for biphenyl, chlorobiphenyl and metatoluylate degradation from Pseudomonas putida

Pseudomonas putida strain SU83, harbors the pBS311 plasmid coding for the degradation of biphenyl, 2- and 4-chlorbiphenyl, meta- and paratoluylate. The insertional mutants of the plasmid obtained by the transposon Tn5 insertion were isolated. One of the mutants was used for cloning of the biphenyl degradation genes. The plasmid pBS311:: Tn5 DNA was inserted into the BamHI site of the plasmid pBR322 and cloned. 11 recombinants of 354 tested were treated with 0.1% solution of 2,3-dioxybiphenyl. One of them has acquired the yellow colour testifying to conversion of 2,3-dioxyphenyl to "2-hydroxy-6-keto-6-phenylhexa-2,4-diene acid. The recombinant plasmid pBS312 from this clone is 10.5 kb in size, the size of the insert being 6.2 kb. Escherichia coli SU185 cells harbouring pBS312 are able to support metacleavage of 2,3-dioxybiphenyl, 3-methylcatechol and catechol, but not of 4-methylcatechol. The results suggest the cloned fragment to contain a gene for 2,3-dioxybiphenyl-1,2-dioxygenase, the third enzyme for biphenyl catabolism.     

Derivatives of methanopterin, a coenzyme involved in methanogenesis

Degradational studies of methanopterin, a coenzyme involved in methanogenesis, are reported. The results of these studies are in full accordance with the proposed structure of methanopterin as N-[1'-(2'-amino-4'-hydroxy-7' -methyl-6'-pteridinyl)ethyl]-4-[2', 3', 4', 5'-tetrahydroxypent-1'-yl(5'-1' )O-alpha-ribofuranosyl-5'-phosphoric acid] aniline in which the phosphate group is esterified with alpha-hydroxyglutaric acid. Acid hydrolysis of methanopterin cleaved the 5'----1' glycosidic bond and yielded a 'hydrolytic product' which was identified as N-[1'-(2'-amino-4'-hydroxy-7' -methyl-6'-pteridinyl)ethyl]-4-[2', 3', 4', 5'-tetrahydroxypent-1'-yl]aniline. Alkaline permanganate oxidation of methanopterin yielded 7-methylpterin-6-carboxylic acid. Catalytic (or enzymatic) hydrogenation of methanopterin gave a mixture of 6-ethyl-7-methyl-7,8-dihydropterin, 6-ethyl-7-methylpterin and a third compound, named methaniline which was identified as 4-[2', 3', 4', 5'-tetrahydroxypent-1'-yl(5'----1')O-alpha -ribofuranosyl-5'-phosphoric acid]aniline, in which the phosphate group is esterified with alpha-hydroxyglutaric acid. Methanosarcina barkeri contains a closely related coenzyme called sarcinapterin, which was identified as a L-glutamyl derivative of methanopterin, where the glutamate moiety is attached to the alpha-carboxylic acid group of the alpha-hydroxyglutaric acid moiety of methanopterin via an amide linkage.     

Biosynthesis of biopterin. Studies on the mechanism of 6-pyruvoyltetrahydropteridine synthase

[1'-3H]- and [2'-3H]dihydroneopterin triphosphate (NH2TP) were prepared enzymatically from [4-3H]- and [5-3H]glucose and converted to tetrahydrobiopterin (BH4) by an extract from bovine adrenal medulla. The formation of BH4 from both [1'-3H]- and [2'-3H]-NH2TP proceeds with virtually complete loss of the respective tritium label. The breaking of the CH-bond at C-1' is characterized by a kinetic isotope effect of 2.6 +/- 0.5. A smaller kinetic isotope effect of 1.5 +/- 0.2 was found for the breaking of the CH-bond at C-2'.     

Human placental estradiol 17 beta-dehydrogenase. Identification of a single histidine residue affinity-labeled by both 3-bromoacetoxyestrone and 12 beta-bromoacetoxy-4-estrene-3,17-dione

Human placental estradiol 17 beta-dehydrogenase (EC 1.1.1.62) was affinity-labeled at pH 6.3 by 3-bromo[2'-14C]acetoxyestrone and 12 beta-bromo-[2'-14C] acetoxy-4-estrene-3,17-dione (both are substrates) in separate incubations. The affinity-alkylated enzyme samples were then treated separately as described below. Amino acid compositions of both samples revealed radioactive 3-carboxymethylhistidine. Tryptic digests of each sample were prepared, applied to Sephadex G-50, and 3-carboxymethylhistidine-bearing fractions identified. These peptides were further purified by cation exchange chromatography, gel filtration, and paper electrophoresis. The purified, 3-carboxymethylhistidine-bearing peptides labeled by the two steroids had identical electrophoretic mobilities at pH 6.5, 3.5, and 1.9. The amino acid sequence of the radioactive peptide alkylated by 3-bromo[2'-14C]acetoxyesterone was determined as: Leu-Ala-3-[14C]CmHis-Ser-Lys. The smaller quantity of peptide obtained from the inactivation with 12 beta-bromo[2'-14C]acetoxy-4-estrene-3,17-dione precluded the determination of its complete sequence. However, the first 3 residues were found to be Leu-Ala-3-[14C]CmHis and the amino acid composition showed that serine and lysine were also present. It is concluded that the steroid-binding site of human placental estradiol 17 beta-dehydrogenase contains a histidine residue which proximates the upper A-ring region of the steroid as it undergoes the reversible binding step.     

Novel TIPP (H-Tyr-Tic-Phe-Phe-OH) analogues displaying a wide range of efficacies at the δ opioid receptor. Discovery of two highly potent and selective δ opioid agonists

Analogues of the δ opioid antagonist peptide TIPP (H-Tyr-Tic-Phe-Phe-OH; Tic=1,2,3,4-tetrahydroisoquinoline3-carboxylic acid) containing various 4'-[N-(alkyl or aralkyl)carboxamido]phenylalanine analogues in place of Tyr(1) were synthesized. The compounds showed subnanomolar or low nanomolar δ opioid receptor binding affinity and various efficacy at the δ receptor (antagonism, partial agonism, full agonism) in the [(35)S]GTPγS binding assay. Two analogues, [1-Ncp(1)]TIPP (1-Ncp=4'-[N-(2-(naphthalene-1-yl)ethyl)carboxamido]phenylalanine) and [2-Ncp(1)]TIPP (2-Ncp=4'-[N-(2-(naphthalene-2-yl)ethyl)carboxamido]phenylalanine), were identified as potent and selective δ opioid agonists.     

Antimycobacterial activity: a facile three-component [3+2]-cycloaddition for the regioselective synthesis of highly functionalised dispiropyrrolidines

A series of twelve dispiropyrrolidines were synthesized using [3+2]-cycloaddition reactions. The synthesized compounds were screened for their antimycobacterial activity against M. tuberculosis H(37)Rv and INH resistant M. tuberculosis strains using agar dilution method, four of them showed good activity with MIC of less than 1 μM. Compound 4'-[5-(4-fluorophenyl)pyridin-3-yl]-1'-methyldispiro[indan-2,2' pyrrolidine-3',2″-indan]-1,3,1″-trione (4b) was found to be the most active with MIC of 0.1215 and 5.121 μM, respectively.     

Synthesis and monoamine transporter binding properties of 2beta-[3'-(substituted benzyl)isoxazol-5-yl]- and 2beta-[3'-methyl-4'-(substituted phenyl)isoxazol-5-yl]-3beta-(substituted phenyl)tropanes

A series of 2beta-[3'-(substituted benzyl)isoxazol-5-yl]- and 2beta-[3'-methyl-4'-(substituted phenyl)isoxazol-5-yl]-3beta-(substituted phenyl)tropanes were prepared and evaluated for affinities at dopamine, serotonin, and norepinephrine transporters using competitive radioligand binding assays. The 2beta-[3'-(substituted benzyl)isoxazol-5-yl]-3beta-(substituted phenyl)tropanes (3a-h) showed high binding affinities for the dopamine transporter (DAT). The IC(50) values ranged from 5.9 to 22nM. On the other hand, the 2beta-[3'-methyl-4'-(substituted phenyl)isoxazol-5-yl]-3beta-(substituted phenyl)tropanes (4a-h), with IC(50) values ranging from 65 to 173nM, were approximately 3- to 25-fold less potent than the corresponding 2beta-[3'-(substituted benzyl)isoxazol]tropanes. All tested compounds were selective for the DAT relative to the norepinephrine transporter (NET) and serotonin transporter (5-HTT). 3Beta-(4-Methylphenyl)-2beta-[3'-(4-fluorobenzyl)isoxazol-5-yl]tropane (3b) with IC(50) of 5.9nM at the DAT and K(i)s of 454 and 113nM at the NET and 5-HTT, respectively, was the most potent and DAT-selective analog. Molecular modeling studies suggested that the rigid conformation of the isoxazole side chain in 4a-h might play an important role on their low DAT binding affinities.     

Mechanistic and stereochemical studies of a unique dehydration catalyzed by CDP-4-keto-6-deoxy-D-glucose-3-dehydrase: a pyridoxamine 5'-phosphate dependent enzyme isolated from Yersinia pseudotuberculosis.

CDP-4-keto-6-deoxy-D-glucose-3-dehydrase (E1) purified from Yersinia pseudotuberculosis is a pyridoxamine 5'-phosphate (PMP) dependent enzyme which catalyzes the C-O bond cleavage at C-3 of a CDP-4-keto-6-deoxy-D-glucose substrate, a key step in the formation of 3,6-dideoxyhexoses. Since enzyme E1 utilizes the PMP cofactor in a unique manner, it is essential to establish its role in E1 catalysis. When an incubation was conducted in [18O]H2O, incorporation of 18O into positions C-3 and C-4 of the recovered substrate was observed. This result not only provided the evidence necessary to reveal the reversibility of E1 catalysis but also lent credence to the formation of a delta 3,4-glucoseen intermediate. In view of E1 catalysis being initiated by a C-4' deprotonation of the PMP-substrate complex, the stereochemical course of this step was examined using chemically synthesized (4'S)- and (4'R)-[4'-3H]PMP as probes. Our results clearly demonstrated that the stereochemistry of this deprotonation is pro-S specific, which is in agreement with the stereochemical consistency found with other vitamin B6 phosphate dependent enzymes. The fact that reprotonation at C-4' of the PMP-delta 3,4-glucoseen complex in the reverse direction of E1 catalysis was also found to be pro-S stereospecific strongly suggested that enzyme E1, like most of its counterparts, has the si face of its cofactor-substrate complex exposed to solvent and accessible to active-site catalytic groups as well.(ABSTRACT TRUNCATED AT 250 WORDS)     

Elucidation of the structure of methanopterin, a coenzyme from Methanobacterium thermoautotrophicum, using two-dimensional nuclear-magnetic-resonance techniques

Methanopterin is a coenzyme involved in methanogenesis. From 2 kg wet cells of Methanobacterium thermoautotrophicum about 35 mumol methanopterin were isolated. The structure of this compound was elucidated by various two-dimensional nuclear-magnetic-resonance techniques. Methanopterin was identified as N-[1'-(2"-amino-4"-hydroxy-7" - methyl-6"- pteridinyl) ethyl]-4-[2',3',4',5'- tetrahydroxypent-1'- yl (5' leads to 1") O-alpha-ribofuranosyl-5"-phosphoric acid] aniline, in which the phosphate group is esterified with alpha-hydroxyglutaric acid. The molecular formula of the sodium salt of methanopterin at pH 7.0 is C30H38O16N6PNa3 X chiH2O (chi is about 4). The anhydrous sodium salt of methanopterin has a molecular mass of 838.60 Da and the molar absorption coefficient at 342 nm is 7.4 mM-1 cm-1 at pH 7.0.     

Stereochemistry of the dTDP-glucose oxidoreductase reaction.

The stereochemical course of the dTDP-glucose oxidoreductase (EC 4.2.1.46) reaction was studied using enzyme partially purified from Escherichia coli and dTDP-(6R)- and (6S)-[4-2H, 6-3H]glucose as substrate. The latter was prepared enzymatically by reduction of (3R)- and (3S)-3-P-[3-3H]glycerate to the 1-deuterated 3-P-glyceraldehyde with (4S)-[4-2H]NADH, followed first by conversion to glucose-1-P with the glycolytic enzymes, and then by transformation into the dTDP derivative. The stereospecifically labeled dTDP-glucose samples were mixed with nonlabeled carrier material and converted to dTDP-4-keto-6-deoxyglucose, which contained a chiral methyl group as shown by chirality analysis of the acetic acid resulting from Kuhn-Roth oxidation of the sugar nucleotide. These results confirm that the hydrogen transfer from C4 to C6 is intramolecular and show that the migrating hydrogen replaces the 6-hydroxyl group with inversion of configuration. Assuming that the hydrogen transfer, since it is intramolecular, must be suprafacial, it follows that the elimination of water from C5 and C6 is formally syn, whereas the reduction of the resulting delta5,6-double bond formally involves an anti addition of H+ and H-.     

3-p-Toluoyl-2-[4'-(3-diethylaminopropoxy)-phenyl]-benzofuran and 2-[4'-(3-diethylaminopropoxy)-phenyl]-benzofuran do not act as surfactants or micelles when inhibiting the aggregation of beta-amyloid peptide

The cmc and IC50 values of the beta-amyloid (Abeta) aggregation inhibitors, 3-p-toluoyl-2-[4'-(3-diethylaminopropoxy)-phenyl]-benzofuran 1, and 2-[4'-(3-diethylaminopropoxy)-phenyl]-benzofuran 2 have been determined. After comparison of the cmc data and biological data (IC50 values), we conclude that these active benzofurans do not act as surfactants or micelles at the concentration required to inhibit beta-amyloid-peptide aggregation.     

Synthesis of tritiated abscisic acid of high specific activity

Stable abscisic acid (RS)-[³H] was synthesized at a specific activity of 21 Ci/mmol using a basic alumina catalyzed proton exchange of 1-hydroxy-4-keto-α-ionone with T₂O followed by a Wittig reaction. Abscisic acid -[³H] of specific activity 102 mCi/mmol was synthesized after carrying out a base catalyzed tritium exchange in solution.     

Antibacterial bromophenols from the marine red alga Rhodomela confervoides

Two bromophenols, together with three known compounds, were isolated from the methanolic extract of the marine alga, Rhodomela confervoides. By means of MS and NMR spectroscopic analyses, they were identified as 3-bromo-4-[2,3-dibromo-4,5-dihydroxyphenyl] methyl-5-(hydroxymethyl) 1,2-benzenediol (1) and 3-bromo-4-[2,3-dibromo-4,5-dihydroxyphenyl] methyl-5- (ethoxymethyl) 1,2-benzenediol (2). Three known compounds were also isolated, namely 3-bromo-4-[2,3-dibromo-4,5-dihydroxyphenyl] methyl-5-(methoxymethyl) 1,2-benzenediol (3), 4,4'- methylenebis [5,6-dibromo-1,2-benzenediol] (4) and bis (2,3-dibromo-4,5-dihydroxybenzyl) ether (5). Compound 5 was the most active against five strains of bacteria with the MIC less than 70 microg/ml, while compounds 2, 3 and 4 exhibited moderate activity.     

Synthesis and biological activity of derivatives of ubiquinone: photoaffinity analogues containing the 4-azido-2-nitroanilino group

The photoaffinity analogues of ubiquinone 2,3-dimethoxy-5-methyl-6-[2-[1-oxo-3-(4-azido-2-nitroanilino) propoxy]-3-methylbutyl]-1,4-benzoquinone (2'-ANAP-Q-1) and 2,3-dimethoxy-5-methyl-6-[3-[1-oxo-3-(4-azido-2-nitroanilino) propoxy]-3-methylbutyl]-1,4-benzoquinone (3'-ANAP-Q-1) have been synthesized. The required intermediate alcohols 2,3-dimethoxy-5-methyl-6-(2-hydroxy-3-methylbutyl)-1,4-benzoquinone and 2,3-dimethoxy-5-methyl-6-(3-hydroxy-3-methylbutyl)-1,4-benzoquinone were prepared in good yield from ubiquinone 1 by hydration of the side-chain double bond via hydroboration or acid catalysis, respectively. These alcohols were then coupled with 3-(4-azido-2-nitroanilino)propanoic acid, with p-toluenesulfonyl chloride in dry pyridine, to give 2'- and 3'-ANAP-Q-1. The synthetic methods presented should be of general utility in the preparation of derivatives of ubiquinone in which a reactive or reporter group is relatively close to the ubiquinone ring. By use of membrane vesicles prepared from a ubi-men-strain of Escherichia coli described previously [Wallace, B., & Young, I. G. (1977) Biochim. Biophys. Acta 461, 84-100], it has been shown that 2'- and 3'-ANAP-Q-1 substitute for ubiquinone 8 in the NADH, succinate, and D-lactate oxidase systems. Thus, these compounds may be of value in labeling respiratory chain proteins that interact with ubiquinone.     

The metabolic sulfonation and side-chain oxidation of 3'-hydroxyisosafrole in the mouse and its inactivity as a hepatocarcinogen relative to 1'-hydroxysafrole

The chemically synthesized sulfuric acid esters of 1'-hydroxysafrole and 3'-hydroxyisosafrole, 1'-sulfooxysafrole and 3'-sulfooxyisosafrole, respectively, are both strong electrophiles. Each ester reacted with deoxyguanosine (dGuo) in aqueous solution to form both safrol-1'-yl- and isosafrol-3'-yl-deoxyguanosine adducts. Both 1'-hydroxysafrole and 3'-hydroxyisosafrole were also formed from each ester in the presence of water. When either 1'-[3H]hydroxysafrole or 3'-[3H]hydroxyisosafrole was incubated with mouse liver cytosols fortified with 3'-phosphoadenosine-5'-phosphosulfate (PAPS) and RNA, similar levels of RNA- and protein-bound adducts were formed; thus, the hepatic sulfotransferase activities for these two substrates appear to be similar. In contrast, the levels of hepatic nucleic acid and protein adducts formed after administration of 3'-[3H]hydroxyisosafrole to mice were only 2-4% and 8-14%, respectively, of those obtained after an equimolar dose of 1'-[3H]hydroxysafrole. Likewise, when 3'-hydroxyisosafrole was injected into 12-day-old male B6C3F1 mice at a level of 0.1 or 2.5 mumol/g body wt., the average numbers of hepatomas per mouse (0.2 and 0.4, respectively) were not significantly increased over the average number for mice treated only with the solvent (0.2). By contrast, mice that received 0.1 mumol of 1'-hydroxysafrole/g body wt. developed about 2 hepatomas per mouse. The metabolism of 3'-hydroxyisosafrole in the rat and mouse differed markedly from that of 1'-hydroxysafrole. 3'-Hydroxyisosafrole rapidly underwent side-chain oxidation to yield 3,4-methylenedioxycinnamic acid and 3,4-methylenedioxybenzoic acid. In the first 4 h, 3,4-methylenedioxybenzoyl glycine and 3,4-methylenedioxycinnamoyl glycine, the major urinary metabolites, together accounted for 39% and 63% of the dose administered to rats and mice, respectively. The glucuronide of 3'-hydroxyisosafrole was not detected in the urine, whereas urinary excretion of the glucuronide of 1'-hydroxysafrole at 2 h accounted for approx. 40% of a dose of 1'-hydroxysafrole.     

Biosynthetic studies on the tropane ring system of the tropane alkaloids from Datura stramonium

Isotopic labelling experiments have been carried out in Datura stramonium root cultures with the following isotopically labelled precursors; [2H3]- [2-13C, 2H3]-, [1-13C, 18O2]-acetates, 2H2O, [2H3-methyl]-methionine, [2-13C]-phenyllactate, [3-2H]-tropine and [2'-13C, 3-2H]-littorine. The study explored the incorporation of isotope into the tropane ring system of littorine 1 and hyoscyamine 2 and revealed that deuterium from acetate is incorporated only into C-6 and C-7, and not into C-2 and C-4 as previously reported. Oxygen-18 was not retained at a detectable level into the C(3)-O bond from [1-13C, 18O2]-acetate. The intramolecular nature of the rearrangement of littorine 1 to hyoscyamine 2 is revealed again by a labelling study using [2'-13C, 3-2H]-littorine, [2-13C]-phenyllactate and [3-2H]-tropine.