Identification of the blue pigment formed in a D-glucose-glycine reaction system

D-Glucose (0.7 M), glycine (0.3 M), and sodium hydrogencarbonate (0.1 M) were dissolved in aqueous 30% ethanol at pH 8.0 and left at 50 °C for 4 d in a dark room under nitrogen displacement. The resulting blue pigment was isolated and purified from the blue solution by anionic exchange and gel filtration chromatography. This blue pigment, which is designated Blue-G1, was identified as 5-[1,4-bis-carboxymethyl-5-(2,3,4-trihydroxybutyl)-1,4-dihydropyrrolo[3,2-b]pyrrol-2-ylmethylene]-1,4-bis-carboxymethyl-2-(2,3,4-trihydroxybutyl)-4,5-dihydropyrrolo[3,2-b]pyrrol-1-ium. Blue-G1 had two symmetrical pyrrolopyrrole structures with a trihydroxybutyl group. Blue-G1 had a polymerizing activity, suggesting it to be an important Maillard reaction intermediate through the formation of melanoidins.     

Identification of the Blue Pigment Formed in a D-Glucose-Glycine Reaction System

D-Glucose (0.7 M), glycine (0.3 M), and sodium hydrogencarbonate (0.1 M) were dissolved in aqueous 30% ethanol at pH 8.0 and left at 50 °C for 4 d in a dark room under nitrogen displacement. The resulting blue pigment was isolated and purified from the blue solution by anionic exchange and gel filtration chromatography. This blue pigment, which is designated Blue-G1, was identified as 5-[1,4-bis-carboxymethyl-5-(2,3,4-trihydroxybutyl)-1,4-dihydropyrrolo[3,2-b]pyrrol-2-ylmethylene]-1,4-bis-carboxymethyl-2-(2,3,4-trihydroxybutyl)-4,5-dihydropyrrolo[3,2-b]pyrrol-1-ium. Blue-G1 had two symmetrical pyrrolopyrrole structures with a trihydroxybutyl group. Blue-G1 had a polymerizing activity, suggesting it to be an important Maillard reaction intermediate through the formation of melanoidins.     

A novel thiamine-derived pigment,pyrizepine, formed by the Maillard reaction

To find a Maillard pigment derived from thiamine, a solution containing glucose and thiamine was heated and analyzed with high-performance liquid chromatography equipped with diode-array detection. As a result, a unique peak showing an absorption maximum at 380 nm was detected. This peak was then isolated from a reaction solution containing glucose, lysine and thiamine, and was identified as 1-(2-methyl-6,9-dihydro-5H-pyrimido[4,5-e][1,4]diazepin-7-yl)ethan-1-one using instrumental analyses. This compound, named pyrizepine, was a novel yellow pigment having a fused ring consisting of pyrimidine and diazepine. Pyrizepine was a major low-molecular-weight pigment in the reaction solution. The structure suggests that pyrizepine is formed by condensation reaction between a degradation product of thiamine and a tetrosone derivative formed from glucose by the Maillard reaction.     

Substituted furo[3,2-b]pyridines: novel bioisosteres of 5-HT 1F receptor agonists

Synthesis and evaluation of a series of 2,3,5- and 3,5-substituted furo[3,2-b]pyridines were undertaken in order to investigate their utility as bioisosteres of 5-HT(1F) receptor agonist indole analogues, 1-3. The replacement proved to be effective, providing compounds with similar 5-HT(1F) receptor affinity and improved selectivity when compared with the indole analogues. Through these studies we identified 4-fluoro-N-[3-(1-methyl-piperidin-4-yl)-furo[3,2-b]pyridin-5-yl]-benzamide (5), a potent and selective 5-HT(1F) receptor agonist with the potential to treat acute migraine.     

Discovery and in vitro evaluation of potent TrkA kinase inhibitors: oxindole and aza-oxindoles

The discovery, synthesis, potential binding mode, and in vitro kinase profile of 3-(3-bromo-4-hydroxy-5-(2'-methoxyphenyl)-benzylidene)-5-bromo-1,3-dihydro-pyrrolo[2,3-b]pyridin-2-one, 3-[(1-methyl-1H-indol-3-yl)methylene]-1,3-dihydro-2H-pyrrolo[3,2-b]-pyridin-2-one as potent TrkA inhibitors are discussed.     

Protective effect of Blue-M1 against oxidative stress on COS-1 cells

Blue-M1 is a blue pigment formed from xylose and glycine in the Maillard reaction. Previous work revealed that Blue-M1 scavenged hydroxyl radicals, and prevented the autoxidation of linoleic acid in vitro. We investigated the protective effect of Blue-M1 for 2,2'-azobis(2-amidino-propane)dihydrochloride (AAPH)-induced toxicity in COS-1 cells. COS-1 cells were cultured in AAPH containing DMEM medium with or without Blue-M1 at 37 degrees C for 24 h. Blue-M1 decreased the AAPH-induced toxicity in COS-1 cells, and this effect was dose-dependent. Furthermore, COS-1 cells were treated with diphenyl-1-pyrenylphosphine (DPPP), as a reagent for the detection of lipid peroxide, and then were cultured in AAPH containing DMEM medium with or without Blue-M1 at 37 degrees C for 6 h. Blue-M1 prevented the AAPH-induced peroxidation of cell membrane on COS-1 cells, and this effect was also dose-dependent. These results suggest that Blue-M1 prevents the oxidative cell injury. Therefore, Blue-M1 will be an antioxidant, which protect against the oxidative stress in living systems.     

Isolation and identification of the intermediates during pyrazole formation of some carbohydrate hydrazone derivatives

Reaction of the oxidation product of L-ascorbic acid, dehydro-L-ascorbic acid, with o-phenylenediamine, followed by 2,4,6-trichlorophenylhydrazine (3) afforded 3-[1-(2,4,6-trichlorophenylhydrazono)-L-threo-2,3,4-trihydroxybut-1-yl]quinoxalin-2(1H)one (4), whose structure was deduced from studying its periodate oxidation, which gave the glyoxal derivative 3-[1-(2,4,6-trichlorophenylhydrazono)glyoxal-1-yl]quinoxalin-2(1H)one (5) that upon reduction afforded 3-[1-(2,4,6-trichlorophenylhydrazono)-2-hydroxyethy-1-yl]quinoxalin-2(1H)one (6). The reaction of 5 with 3 afforded the bishydrazone 3-[1,2-bis(2,4,6-trichlorophenylhydrazono)glyoxal-1-yl]quinoxalin-2(1H)one. The reaction of 5 with acetic anhydride in pyridine afforded the 2,3-dihydrofuro[2,3-b]quinoxaline derivative 2-acetoxy-3-[2-acetyl-2-(2,4,6-trichlorophenyl)hydrazono)]-2,3-dihydrofuro[2,3-b]quinoxaline. Acetylation of 4 with acetic anhydride in pyridine afforded the acyclic diacetate intermediate 3-[3,4-di-O-acetyl-2-deoxy-1-(2,4,6-trichlorophenylhydra-zono)but-2-en-1-yl]quinoxalin-2(1H)one (12), which was also obtained from the reaction of 4 with boiling acetic anhydride. Compound 12 rearranged under the reaction conditions to give the pyrazole derivatives 3-[5-(ace-toxymethyl)-1-(2,4,6-trichlorophenyl)pyrazol-3-yl]quinoxalin-2(1H)one (14) and 2-acetoxy-3-[5-(acetoxymethyl)-1-(2,4,6-trichlorophenyl)pyrazol-3-yl)]quinoxaline (15), as well as the 2,3-dihydrofuro[2,3-b]quinoxaline derivative 2-(2-acetoxyethen-2-yl)-3-[2-(2,4,6-trichlorophenyl)hydrazono]-2,3-dihydrofuro[2,3-b]quinoxaline. Acetylation of 3-[5-(hydroxymethyl)-l-(2,4,6-trichlorophenyl)pyrazol-3-yl]quinoxalin-2(1H)one (16) with acetic anhydride in pyridine or 12 with boiling acetic anhydride afforded 15 and 16, respectively. Treatment of 4 with diluted sodium hydroxide afforded the pyrazolo[2,3-b]quinoxaline (flavazole) derivative 1-(2,4,6-trichlorophenyl)-3-(L-threo-glycerol-1-yl)pyrazolo[2,3-b]quinoxaline whose acetylation afforded the acetyl derivative 3-(2,3,4-tri-O-acetyl-L-threo-glycerol-1-yl)-1-(2,4,6-trichlorophenyl)pyrazolo[2,3-b]quinoxaline. The assigned structures were based on spectral analysis. The activity of compound 4 against hepatitis B virus has been studied.     

Syntheses of phenylpyrazolodiazepin-7-ones as conformationally rigid analogs of aminopyrazole amide scaffold and their antiproliferative effects on cancer cells

Recently, we have reported the syntheses and antiproliferative activities of N-(5-amino-1-(4-methoxybenzyl)-1H-pyrazol-4-yl amide derivatives on melanoma cells. As a continuous work for antiproliferative agents in melanoma, here we report the synthesis of conformationally rigid analogs, phenyl-6,8-dihydropyrazolo[3,4-b][1,4]diazepin-7(1H)-one derivatives 7a-g, 8a-o and their antiproliferative activities against A375P melanoma cell line and U937 hematopoietic cell line. Most compounds showed competitive antiproliferative activities to sorafenib, the reference standard. Among them, N-(3-(1-benzyl-7-oxo-1,6,7,8-tetrahydropyrazolo[3,4-b][1,4]diazepin-5-yl)phenyl)-4-chloro-3-(trifluoro methyl)benzamide-amino-1-(4-methoxybenzyl)-1H-pyrazol-4-yl)-5-(3-(4-chloro-3-(trifluoromethyl) phenyl) ureido)-2-methylbenzamide (7b) exhibited potent activities (GI(50)=0.43 μM and 0.06 μM) on both cell lines. It has been further confirmed to be a potent and selective Raf kinases inhibitor and also mild inhibitor of PI3Kα.     

A light-sensitive yellow ommochrome pigment from the house fly

A light-sensitive pigment (lambda max at 430 and 340 nm), extracted in acidic methanol from the Musca domestica heads, showed, in the absorption curve, a plateau at 480-500 nm and a new maximum at 400 nm, after visible light irradiation. The light-sensitive house fly pigment showed spectroscopic and chemical properties of the ommochrome pigments (Butenandt and Sch?fer: Recent Progress in the Chemistry of Natural and Synthetic, Colouring Matters and Related Fields, Academic Press, New York, pp 13-33, 1962; Bolognese and Scherillo: Experientia 30:225-226, 1974). The treatment of the extracted pigment with a methanol-HClsat. mixture afforded some coloured compounds; two main products were identified by comparison of their chromatographic and spectral properties with authentic samples of 1-oxo-2H-3-carbomethoxy-5-methoxy-11-(fumaroyl-methylester)-pyrido [3,2-a] phenoxazine (compound 7) and 1-oxo-2H-3-carbomethoxy-5-methoxy-9-chlorine-11-(fumaroyl-methylester )-pyrido [3,2-a] phenoxazine (compound 8), obtained from the oxidation mixture of 3-hydroxykynurenine methylester (compound 9).     

Enzymatic syntheses of 6-(4H-selenolo[3,2-b]pyrrolyl)-L-alanine, 4-(6H-selenolo[2,3-b]pyrrolyl)-L-alanine, and 6-(4H-furo[3,2-b]pyrrolyl-L-alanine

6-(4H-Selenolo[3,2-b]pyrrolyl)-L-alanine 1, 4-(6H-selenolo[2,3-b]pyrrolyl)-L-alanine 2, and 6-(4H-furo[3,2-b]pyrrolyl)-L-alanine 3 have been synthesized via reactions of selenolo[3,2-b]pyrrole, selenolo[2,3-b]pyrrole, and furo[3,2-b]pyrrole, respectively, with L-serine. The reactions are catalyzed by Salmonella typhimurium tryptophan synthase.     

Benzoquinones from Ardisia japonica with inhibitory activity towards human protein tyrosine phosphatase 1B (PTP1B)

From the whole plant of Ardisia japonica, four [1,4]benzoquinones were isolated by means of bioassay-directed fractionation of the EtOH extract. Apart from the two known compounds maesanin (1) and its congener 2, two new benzoquinones, i.e., 5-ethoxy-2-hydroxy-3-[(10Z)-pentadec-10-en-1-yl][1,4]benzoquinone (3) and 5-ethoxy-2-hydroxy-3-[(8Z)-tridec-8-en-1-yl][1,4]benzoquinone (4), were identified. All compounds showed significant in vitro bioactivities against the PTP1B enzyme, with IC50 values in the range of ca. 3-19 microM.     

Identification of mutagenic heterocyclic amines (IQ, Trp-P-1 and AalphaC) in the water of the Danube river

Three mutagenic heterocyclic amines, 2-amino-3-methylimidazo-[4, 5-f]quinoline (IQ), 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1) and 2-amino-9H-pyrido[2,3-b]indole (AalphaC), were isolated and identified in water from the Danube River in Vienna. Heterocyclic amines were extracted from river water by the blue rayon hanging method, and analyzed by gas chromatography with a nitrogen-phosphorous detector (GC-NPD) and GC-mass spectrometry (GC-MS) after conversion into their N-dimethylaminomethylene derivatives. Identity of IQ, Trp-P-1 and AalphaC in the river water was confirmed by GC-MS. The contents of IQ, Trp-P-1 and AalphaC were estimated by GC-NPD at 1.78+/-0.17, 0.14+/-0.02 and 0.44+/-0.02 ng/g blue rayon equivalent (n=3), respectively. The total amounts of these amines accounted for 26% of the mutagenicity of blue rayon extracts evaluated by the Ames test using TA98 with metabolic activation.     

Biosynthesis of quinoxaline antibiotics: purification and characterization of the quinoxaline-2-carboxylic acid activating enzyme from Streptomyces triostinicus

A quinoxaline-2-carboxylic acid activating enzyme was purified to homogeneity from triostin-producing Streptomyces triostinicus. It could also be purified from quinomycin-producing Streptomyces echinatus. Triostins and quinomycins are peptide lactones that contain quinoxaline-2-carboxylic acid as chromophoric moiety. The enzyme catalyzes the ATP-pyrophosphate exchange reaction dependent on quinoxaline-2-carboxylic acid and the formation of the corresponding adenylate. Besides quinoxaline-2-carboxylic acid, the enzyme also catalyzes the formation of adenylates from quinoline-2-carboxylic acid and thieno[3,2-b]pyridine-5-carboxylic acid. No adenylates were seen from quinoline-3-carboxylic acid, quinoline-4-carboxylic acid, pyridine-2-carboxylic acid, and 2-pyrazinecarboxylic acid. Previous work [Gauvreau, D., & Waring, M. J. (1984) Can. J. Microbiol. 30, 439-450] revealed that quinoline-2-carboxylic acid and thieno[3,2-b]pyridine-5-carboxylic acid became efficiently incorporated into the corresponding quinoxaline antibiotic analogues in vivo. Together with the data described here, this suggests that the enzyme is part of the quinoxaline antibiotics synthesizing enzyme system. The enzyme displays a native molecular weight of 42,000, whereas in its denatured form it is a polypeptide of Mr 52,000-53,000. It resembles in its behavior actinomycin synthetase I, the chromophore activating enzyme involved in actinomycin biosynthesis [Keller, U., Kleinkauf, H., & Zocher, R. (1984) Biochemistry 23, 1479-1484].     

Three further naphthoquinones produced by Fusarium solani

Three naphthoquinone pigments are described which were produced by Fusarium solani. They are 2,3-dihydro-5,8-dihydroxy-6-methoxy-2-hydroxymethyl-3-(2-hydroxypropyl)-1,4-naphthalenedione, 2,3-dihydro-5-hydroxy-4-hydroxymethyl-8-methoxy-naphtho[1,2-b]furan-6,9-dione and 5,8-dihydroxy-2-methoxy-6-hydroxymethyl-7-(2-hydroxypropyl)-1,4-naphthalenedione. One of these pigments was shown to be the precursor of the other two.     

Synthesis of new 2-substituted 6-bromo-3-methylthiazolo[3,2-alpha]-benzimidazole derivatives and their biological activities

1-(6-Bromo-3-methyl-1,3-thiazolo[3,2-alpha]benzimidazol-2-yl)ethanone (2) was prepared by bromination at ambient temperature of 1-(3-methylthiazolo[3,2-alpha]benzimidazol-2-yl)ethanone (1). The structure of 2 was determined by single-crystal X-ray diffraction. The precursor 5 was synthesized by heating a mixture of acetyl 2 and bromine. Various 2-substituted 6-bromo-3-methylthiazolo[3,2-alpha]benzimidazoles containing 1,3-thiazole, 1,4-benzothiazine, quinoxaline or imidazo[1,2-alpha]pyridine moieties were prepared starting from bromoacetyl 5. Taken together from the biological investigations, 2, 5, and 7a were potent immunosuppressors against both macrophages and T-lymphocytes, and 7b, 11b, and 14 were potent immunostimulators towards both types of immune cells. The results also revealed that, among others, 2 and 14 were the most significant inhibitors of LPS-stimulated NO generation, and that 5, 7a, and 7b had a weak radical scavenging activity against DPPH radicals. Moreover, 2, 5, and 7a had a concomitant strong cytotoxicity against colon carcinoma, hepatocellular carcinoma, and lymphoblastic leukemia cells. Collectively, compounds 2, 5, and 7a are multipotent compounds with promising biological activities.     

The synthesis and vasopressin (AVP) antagonist activity of a novel series of N-aroyl-2,4,5,6-tetrahydropyrazolo[3,4-d]thieno[3,2-b]azepines

Synthesis and SAR of N-[4-[(4,5-dihydropyrazolo[3,4-d]thieno[3,2-b]azepin-6(2H)-y l)carbonyl]phenyl]benzamides as arginine vasopressin (AVP) receptor antagonists are discussed. Potent orally active AVP receptor antagonists are produced when the benzamide moiety contains a phenyl group at the 2-position. Similar analogues of 4,6,7,8-tetrahydro-5H-thieno[3,2-b]azepine and VPA-985 are reported.     

Synthesis and biological evaluation of 2-thiopyrimidine derivatives

Various 2-thiopyrimidine derivatives have been synthesized by an efficient, one-pot reaction of functionalized amines with either 4-isothiocyanato-4-methyl-2-pentanone or 3-isothiocyanatobutanal. All the synthesized compounds were fully characterized by elemental analysis (CHN), FT-IR, (1)H NMR, and mass spectral data. One of the compounds, 7,7,8a-trimethyl-hexahydro-thiazolo[3,2-c]pyrimidine-5-thione (17) showed good anti-inflammatory (37.4% at 100 mg/kg p.o.) and analgesic activity (75% at 100 mg/kg p.o.). 7-(1-Mercapto-3,3,4a-trimethyl-4,4a,5,9b-tetrahydro-3H-pyrido[4,3-b]indol-7-yl)-3,3,4a-trimethyl-3,4,4a,5-tetrahydro-benzo[4,5]imidazo[1,2-c]pyrimidine-1-thiol (3) showed moderate activity against CDK-1 (IC(50)=5 microM). The other compounds showed moderate anti-inflammatory (5-20%), analgesic (25-75%) and protein kinase (CDK-5, GSK-3) inhibitory activities (IC(50)> 10 microM).     

In-vitro cytotoxicity and cell cycle analysis of two novel bis-1,2, 4-triazole derivatives: 1,4-bis[5-(5-mercapto-1,3,4-oxadiazol-2-yl-methyl)-thio-4-(p-tolyl)-1,2,4-triazol-3-yl]-butane (MNP-14) and 1,4-bis[5-(carbethoxy-methyl)-thio-4-(p-ethoxy phenyl)-1,2,4-triazol-3-yl]-butane (MNP-16)

In the present study, we have tested the cytotoxic and DNA damage activity of two novel bis-1,2,4 triazole derivatives, namely 1,4-bis[5-(5-mercapto-1,3,4-oxadiazol-2-yl-methyl)-thio-4-(p-tolyl)-1,2,4-triazol-3-yl]-butane (MNP-14) and 1,4-bis[5-(carbethoxy-methyl)-thio-4-(p-ethoxy phenyl) -1,2,4-triazol-3-yl]-butane (MNP-16). The effect of these molecules on cellular apoptosis was also determined. The in-vitro cytotoxicity was evaluated by a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay as well as Trypan blue dye exclusion methods against human acute lymphoblastic leukemia (MOLT4) and lung cancer cells (A549). Our results showed that MNP-16 induced significant cytotoxicity (IC(50) of 3-5 μM) compared with MNP-14. The cytotoxicity induced by MNP-16 was time and concentration dependent. The cell cycle analysis by flow cytometry (fluorescence-activated cell sorting [FACS]) revealed that though there was a significant increase in the apoptotic population (sub-G(1) phase) with an increased concentration of MNP-14 and 16, there was no cell cycle arrest. Further, the comet assay results indicated considerable DNA strand breaks upon exposure to these compounds, thereby suggesting the possible mechanism of cytotoxicity induced by MNP-16. Hence, we have identified a novel molecule (MNP-16) which could be of great clinical relevance in cancer therapeutics.     

Cross-linking of proteins by Maillard processes--characterization and detection of a lysine-arginine cross-link derived from D-glucose.

Covalently cross-linked proteins are among the major modifications caused by the advanced Maillard reaction. So far, the chemical nature of these aggregates is largely unknown. Investigations are reported on how the cross-linking unit 2-ammonio-6-[2-[(4-ammonio-5-oxido-5-oxopentyl)amino]-6,7-dihydrox y-4,5,6,7,8,8a-hexahydroimidazo[4,5-b]azepin-4-yl] hexanoate (7), designated as glucosepan, can be identified and quantified from D-glucose-bovine serum albumin (BSA) incubations. Independent synthesis and unequivocal structural characterization are given for glucosepan 7. A protocol was established for its determination by LC-MS with electrospray ionization (ESI). BSA and D-glucose were incubated at 37 degrees C, pH 7.4 for eight weeks and the time-dependent formation of 7 was observed. Since glucosepan 7 is unstable under acid proteolytic conditions, BSA was cleaved enzymatically. The maximum value obtained from a solution containing 50 g/L BSA and 100 mM D-glucose after eight weeks incubation time corresponds to an arginine derivatization rate of 1.38 +/- 0.07 mmol 7/mol Arg (equivalent to 31.7 +/- 1.6 mmol 7/mol BSA). From these results, it seems justified to expect 7 to play an important role in the cross-linking of proteins in vivo as well as in foodstuffs. The structural similarity of glucosepan 7 and pentosidine 1 made it obvious to also look for an eventual parallelism in the respective formation pathways.