Claudine Servy

Nitration of catecholamines with nitrogen oxides in mild conditions: a hypothesis for the reactivity of NO in physiological systems

Abstract Dopamine, norepinephrine and epinephrine react at room temperature, in acetate buffer (36) with sodium nitrite or in non-deaerated phosphate buffer (pH 7.4) with NO. The corresponding 6-nitro derivatives are formed.

1-Nitrosomelatonin is a spontaneous NO-releasing compound

Melatonin (N-acetyl-5-methoxytryptamin), the main hormone secreted by the pineal gland in mammals, is nitrosated by nitrite at acidic pH and by NO in the presence of oxygen under neutral conditions. Melatonin is also partly converted to 1-nitrosomelatonin by oxoperoxonitrate (ONOO-, peroxynitrite) in phosphate-buffered solutions at pH 7–10 [Blanchard, B., et al. (2000) Journal of Pineal Research 29, 184–192]. In the present report, we show that 1-nitrosomelatonin in turn behaves as an NO-donor regenerating melatonin. NO-release is evidenced by the formation of nitrite in phosphate-buffered solutions and oxidation of HbO2. No peroxynitrite was formed during that decomposition because serotonin used as a probe was converted only to 4-nitroso-serotonin as expected for a true NO-donor [Blanchard, B., et al. (2001) Free Radical Research, 34, 177–188]. The spontaneous decay of 1-nitrosomelatonin is not affected by GSH and metallic ions but its decomposition is accelerated in acidic pH or in the presence of NADH or ascorbate. Furthermore, melatonin is partially or entirely recovered in the absence or presence of ascorbate, respectively. A homolytic cleavage of 1-nitrosomelatonin is strongly suggested and discussed. Formation of 1-nitrosomelatonin from melatonin and reactive nitrogen species (RNS) followed by its decay into NO demonstrates that melatonin could reduce these RNS to NO.

Bicyclic and tetracyclic diterpenes from a Trichoderma symbiont of Taxus baccata.

Trichoderma atroviridae UB-LMA is an endophytic fungus isolated from Taxus baccata trees. Liquid-state fermentation coupled to in situ solid phase extraction (SPE) was applied, and four compounds were discovered. Compounds 2-4 belong to the harziane tetracyclic diterpene family. Bicylic compound 1 may represent the biosynthetic precursor of this scarce family of compounds.

Isolation and Characterization of α,β-Unsaturated γ-Lactono-Hydrazides from Streptomyces sp.

Two novel α,β-unsaturated γ-lactono-hydrazides, geralcin A (2) and geralcin B (3), were isolated from Streptomyces sp. LMA-545. This unusual scaffold consists of the condensation of alkyl-hydrazide with an α,β-unsaturated γ-lactone, 3-(5-oxo-2H-furan-4-yl)propanoic acid (1), which was isolated from the same broth culture. Amberlite XAD-16 solid-phase extraction was used during the cultivation step, and the trapped compounds (1-3) were eluted from the resin with methanol. The structures were elucidated using (1)H, (13)C, and (15)N NMR spectroscopic analysis and high-resolution mass spectrometry. Geralcin B (3) was cytotoxic against MDA231 breast cancer cells with an IC(50) of 5 μM.

Isolation, structure elucidation and biological activity of metabolites from Sch-642305-producing endophytic fungus Phomopsis sp. CMU-LMA.

Eight polyketide compounds were isolated from the cultivation broth of Phomopsis sp. CMU-LMA. We have recently described LMA-P1, a bicyclic 10-membered macrolide, obtained as a bioconversion derivative of Sch-642305, the major compound isolated in this study. Benquinol is the ethyl ester derivative of the 13-dihydroxytetradeca-2,4,8-trienoic acid produced by Valsa ambiens. This compound is concomitantly produced with the 6,13-dihydroxytetradeca-2,4,8-trienoic acid (DHTTA) previously isolated from Mycosphaerellarubella. The absolute configuration of the new compound, (2R,3R,4S,5R)-3-hydroxy-2,4-dimethyl-5-[(S,Z)-3-methylpentenyl]-tetrahydro-pyranone LMA-P2 was confirmed by X-ray crystallography. The δ-lactone 2,3-dihydroxytetradecan-5-olide (DHTO) was previously isolated from Seiridium unicorne. This compound may form through the cyclization of the methyl-2,3,5-trihydroxytridecanoate LMA-P3, a new linear polyketide isolated in this study. Benquoine, a new 14-membered lactone generated from the cyclization of benquinol, is proposed as the key precursor for the biosynthesis of Sch-642305. Antimicrobial activity and cancer cell viability inhibition by the new compounds were investigated. Benquoine exhibits antimicrobial activity against Gram positive bacteria, and cytotoxicity against HCT-116 cancer cell line.

Isolation and Characterization of Unusual Hydrazides from Streptomyces sp. Impact of the Cultivation Support and Extraction Procedure

Three novel hydrazides, geralcins C-E (1-3), were isolated from Streptomyces sp. LMA-545, together with MH-031 and geralcins A and B. This unusual family of compounds was isolated from liquid-state and agar-supported fermentation using Amberlite XAD-16 solid-phase extraction during the cultivation step. The use of such neutral resin during the cultivation step allowed the specific adsorption of microbial secondary metabolites, avoiding any contamination of the crude extracts by the constituents of the culture medium. The trapped compounds were eluted from the resin with methanol, and their structures elucidated using (1)H, (13)C, and (15)N NMR spectroscopic analysis and high-resolution mass spectrometry. Molecular modeling calculations were applied in order to support structural attributions. No antimicrobial, cytotoxic, or DnaG-inhibition activities were detected for geralcins D and E. Geralcin C has no antimicrobial activity but exhibited an IC(50) of 0.8 μM against KB and HCT116 cancer cell lines. Furthermore, geralcin C inhibited the E. coli DnaG primase, a Gram-negative antimicrobial target, with an IC(50) of 0.7 mM.

Rearranged Diterpenoids from the Biotransformation of ent-Trachyloban-18-oic Acid by Rhizopus arrhizus

In our search for inhibitors of the antiapoptotic protein Bcl-xL, investigation of Xylopia caudata afforded a new diterpenoid, ent-trachyloban-4beta-ol (2), and five known ent-trachylobane or ent-atisane compounds. Only ent-trachyloban-18-oic acid (1) exhibited weak binding activity to Bcl-xL. These compounds exhibited cytotoxicity against KB and HCT-116 cell lines with IC(50) values between 10 and 30 microM. Bioconversion of compound 1 by Rhizopus arrhizus afforded new hydroxylated metabolites (3-7) of the ent-trachylobane and ent-kaurene type and compound 8, with a rearranged pentacyclic carbon framework that was named rhizopene. Compounds 3-8 were noncytotoxic to the two cancer cell lines, and compounds 3 and 5 exhibited only weak binding affinity for Bcl-xL.

Chemical evaluation of compounds as nitric oxide or peroxynitrite donors using the reactions with serotonin

The aim of this work was to assess the capacities of some ·NO-donors to release ·NO, and consequently NOx in aerobic medium, or to give peroxynitrite. The method was based on the differential reactivity of serotonin (5-HT) with either NOx or peroxynitrite, leading in phosphate-buffered solutions to 4-nitroso- and 4-nitro-5-HT formation, respectively. Yields and formation rates of 5-HT derivatives with ·NO-donor were compared to those obtained with authentic ·NO or peroxynitrite in similar conditions. Aside from the capacity of diazenium diolates (SPER/NO and DEA/NO) to release ·NO spontaneously, converting 5-HT exclusively to 4-nitroso-5-HT, all other ·NO donors must undergo redox reactions to produce ·NO. S-nitrosoglutathione (GSNO) and sodium nitroprus-side (SNP) modified 5-HT only in the presence of Cu2+, GSNO yielding 6 times more 4-nitroso-5-HT than SNP. Furthermore, in the presence of Cu+, the yield of ·NO-release from GSNO was 45%. The molsidomine metabolite (SIN-1), which was presumed to release both ·NO and O2/·- at pH 7.4, reacted with 5-HT differently, depending on the presence of reductant or oxidant. Under aerobic conditions, SIN-1 acted predominantly as a 5-HT oxidant and also as a poor ·NO and peroxynitrite donor (15% yield of ·NO-release and 14 % yield of peroxynitrite formation). The strong oxidant Cu2+, even in the presence of air oxygen, accelerated oxidation and increased ·NO release from SIN-1 up to 86%. Only a small part of SIN-1 gave simultaneously ·NO and O2/·- able to link together to give peroxynitrite, but other oxidants could enhance ·NO release from SIN-1.

Agar-supported cultivation of Halorubrum sp. SSR, and production of halocin C8 on the scale-up prototype Platotex

Halorubrum sp. SSR was isolated from a solar saltern in Algeria. The strain exhibited a high antibiotic activity against the indicator strain Natronorubrum aibiense G23, and the bioactive compound showed thermal, acid and alkali stability. SSR was grown on agar-supported cultivation (AgSF) to compare yields and applicability with traditional submerged cultivation. AgSF scale-up was implemented taking benefit from the solid-state cultivation prototype Platotex. This technology leads to high amounts of the target Halocin and facilitate the downstream steps. The antibiotic compound was purified according to a fast efficient procedure including ion exchange chromatography followed by a fractionation on C18 Sep-Pack cartridge. The compound was identified as Halocin C8 according to N-terminal amino acid sequencing and high-resolution mass spectrometry.

Biotransformation of natural compounds. Oxido-reduction of Sch-642305 by Aspergillus ochraceus ATCC 1009

Sch-642305 is the major compound produced by the endophytic fungi Phomopsis sp. CMU-LMA. Incubation of Sch-642305 with Aspergillus ochraceus ATCC 1009 resting cells leads to three new derivatives through an oxido-reduction of the six-membered ring of the molecule. Reduction of the double bound leads to compound (1), which subsequently undergoes carbonyl reduction to (2) and ring hydroxylation to (3). According to the previously solved crystal structure of Sch-642305 coupled with (1)H NMR NOE correlation and the crystal structure of compound 1, the absolute configurations of the new derivatives were established. In contrast to the parent compound Sch-642305, compound (1) exhibits antimicrobial activity against gram-negative bacteria. Furthermore, while all derivatives exhibit cytotoxic activity against various cancer cell lines, compound (2) achieved an IC(50) of 4 nM against human myelogenous leukemia K 562, compared to 20 nM for the parent Sch-642305.