Synthesis of jasplakinolide analogues containing a novel omega-amino acid

The synthesis of the omega-amino acid 4 is described utilizing a two-dimensional synthesis strategy combined with an enzymatic differentiation of homotopic ester groups. The amino acid 4 features two non-bonded interactions that result in conformational constraints on a cyclic construct. This amino acid was incorporated into the four macrolactams 17, 22, 31, and 37. The ring in 17 and 22 is 18-membered, whereas 31 and 37 have a 19-membered ring. The pairs with the same ring size differ in a N-methyl group. For the larger macrolactams (31 and 37) conformational analysis showed that the macrocyclic rings are somewhat more rigid than in the natural lead, the depsipeptide jasplakinolide. Nevertheless, their conformations are comparable to the natural product. There are no intramolecular hydrogen bonds, neither is the cis-rotamer populated in the N-methyl compound 37. Due to the increased flexibility of the smaller macrolactams 17 and 22 and signal overlap, a distinct solution structure could not be obtained for these compounds. The amino acid 4 should be useful for restricting the conformation of other small peptides.     

Investigation of conditions allowing the synthesis of acrylamide-based monolithic microcolumns for capillary electrochromatography and of factors determining the retention of aromatic compounds on these stationary phases

The influence of the cross-linker (concentration), the porogen (lyotrophic salt) and the solvent type as well as the type and concentration of up to three "functional", i.e., interactive monomers on the morphology and the chromatographic properties of acrylamide-based hydrophilic monoliths are investigated. High total monomer concentrations favored polymers with a rigid rather than gel-like structure. High cross-linker concentrations also favor the formation of a nodular structure. The addition of a lyotrophic salt favors the formation of small nodules especially at higher monomer concentration; the pore size of the polymer can also be modulated through the salt concentration. Suitable monoliths were further investigated as potential stationary phases for capillary electrochromatography (CEC). Depending on the type and concentration of the monomers, plate numbers between 50,000 and 100,000 were routinely obtained. The standard deviation of the run-to-run reproducibility was below 2% and that of the batch-to-batch reproducibility below 5%. A set of nine hydrophobic and polar aromatic compounds (all noncharged) was used to investigate the retention mechanism. Possible candidates for chromatographic interaction and retention in these monoliths are the hydrophobic polymer backbone itself and the alkyl, carbonyl, hydroxy, amino, amide, and charged groups introduced by the various functional monomers. Judging from our results, the carbonyl and the hydroxy functions, as well as the hydrophobic polymer backbone can be supposed to be the main sites of interaction. The charged but also the alkyl functions seem to be less important in this regard. The polymerization conditions and especially the composition of the reaction mixture have a strong influence on the behavior of the final column.     

Mass spectrometric characterization of two novel inflammatory peptides from the venom of the social wasp Polybia paulista

The social wasp P. paulista is relatively common in southeast Brazil causing many medically important stinging incidents. The seriousness of these incidents is dependent on the amount of venom inoculated by the wasps into the victims, and the characteristic envenomation symptoms are strongly dependent on the types of peptides present in the venom. In order to identify some of these naturally occurring peptides available in very low amounts, an analytical protocol was developed that uses a combination of reversed-phase and normal-phase high-performance liquid chromatography (HPLC) of wasp venom for peptide purification, with matrix-assisted laser desorption/ionization time-of-flight post-source decay mass spectrometry (MALDI-Tof-PSD-MS) and low-energy collision-induced dissociation (CID) in a quadrupole time-of-flight tandem mass spectrometry (QTof-MS/MS) instrument for peptide sequencing at the sub-picomole level. The distinction between Leu and Ile was achieved both by observing d-type fragment ions obtained under CID conditions and by comparison of retention times of the natural peptides and their synthetic counterparts (with different combinations of I and/or L at N- and C-terminal positions). To distinguish the isobaric residues K and Q, acetylation of peptides was followed by Q-Tof-MS analysis. The primary sequences obtained were INWLKLGKMVIDAL-NH(2) (MW 1611.98 Da) and IDWLKLGKMVMDVL-NH(2) (MW 1658.98 Da). Micro-scale bioassay protocols characterized both peptides as presenting potent hemolytic action, mast cell degranulation, and chemotaxis of polymorphonucleated leukocyte (PMNL) cells. The primary sequences and the bioassay results suggest that these toxins constitute members of a new sub-class of mastoparan toxins, directly involved in the occurrence of inflammatory processes after wasp stinging.     

Human topoisomerase I poisoning: docking protoberberines into a structure-based binding site model

Summary Using the X-ray crystal structure of the human topoisomerase I (top1) – DNA cleavable complex and the Sybyl software package, we have developed a general model for the ternary cleavable complex formed with four protoberberine alkaloids differing in the substitution on the terminal phenyl rings and covering a broad range of the top1-poisoning activities. This model has the drug intercalated with its planar chromophore between the −1 and +1 base pairs flanking the cleavage site, with the nonplanar portion pointing into the minor groove. The ternary complexes were geometry-optimized and relative interaction energies, computed by using the Tripos force field, were found to rank in correct order the biological potency of the compounds; in addition, the model is also consistent with the top1-poisoning inactivity of berberine, a major prototype of the protoberberine alkaloids. The model might serve as a rational basis for elaboration of the most active compound as a lead structure, in order to develop more potent top1 poisons as next generation anti-cancer drugs.     

Simultaneous Determination of Trichothecenes A, B, and D in Maize Food Products by LC–MS–MS

Two new methods of analysis, based on liquid chromatography–tandem mass spectrometry, for simultaneous determination of trichothecenes A, B, and D in maize flour and oil have been developed and validated in accordance with European Commission decision 2002/657/EC (recovery, CCα, CCβ, and precision). The trichothecenes were extracted from maize flour by matrix solid-phase dispersion, with recoveries ≥79%, and from maize oil by liquid–liquid extraction, with recoveries ≥78%. Limits of quantitation ranged between 0.03 and 50 μg kg^?1, depending on the electrospray response to each analyte and on the matrix. Monitoring of flour and oil samples with this HPLC–MS–MS method revealed the presence of deoxynivalenol, T-2 toxin, and diacetoxyscirpenol at very low concentrations.     

Capillary isotachophoresis of some isoquinoline alkaloids

The isotachophoretic behavior of quaternary benzo/c/phenanthridine, protoberberine and aporphine alkaloids in different electrolyte systems is described. The concentration of the leading ion and the pH value of the leading electrolyte affect the relative effective mobilities of the alkaloids. The system of pH 4.7, containing the leading ion K⁺ (0.005 M) counter ion acetate, and the terminating ion β-alanine (0.02 M), has been selected for the quantitative determination of the studied alkaloids in model mixtures and plant extracts.     

Simultaneous determination of the urinary metabolites of benzene, toluene, xylene and styrene using high-performance liquid chromatography/hybrid quadrupole time-of-flight mass spectrometry

A simple and rapid method using reversed-phase liquid chromatography/tandem mass spectrometry (LC/MS/MS) for the simultaneous determination of the urinary metabolites of benzene, toluene, xylene and styrene in human urine specimens and standard solutions is described. A hybrid quadrupole/time-of-flight (QqTOF) mass spectrometer was compared for the determination of metabolite of aromatic solvents in urine samples. The metabolites selected were: trans,trans-muconic acid, hippuric acid, o-, m- and p-methylhippuric acid and phenylglyoxylic acid. The compounds were well separated from each other on narrow-bore 1-mm i.d. reversed-phase LC C-18 columns. Average recoveries for loading 100 microL of urine samples varied from 88-110% and the quantification limits were less than 30 ng/mL for each analyte (3 ng/mL for trans,trans-muconic acid). The qualitative information obtained (mass accuracy, resolution and full-scan spectra) with the QqTOF mass spectrometer allows a secure identification of analytes in biological matrices.     

Determination of metal concentration in fat supplements for swine nutrition by atomic absorption spectroscopy

The presence of some essential and toxic metals in fat supplements for swine diet was investigated. Collected samples represented a relevant production of the Italian industry. In particular, some samples were enriched with antioxidants or waste cooking oils. The method for the determination of Ca, Cu, Cd, Fe, Mg, Mn, Ni, Pb, and Zn in fat samples was developed by means of a certified reference material (CRM 186) and a representative fatty sample (RFS). All samples were digested in closed vessels in a microwave oven and then analyzed by flame atomic absorption or graphite furnace atomic absorption spectrometry. The entire analytical method provided a satisfactory repeatability and reproducibility confirmed by agreement between the experimental recovery data obtained for the CRM 186 sample and, with the method of standard additions, for the RFS material. The samples generally showed a small amount of metals compared with the recommended daily intake for the essential elements. On the other hand, some samples contained a significant concentration, from an analytical point of view, of Cd, Ni, and Pb. Principal component analysis (PCA) was applied to inspect the experimental data obtained from samples analysis. Basically no differences were detected in terms of metal concentration among the fat supplements analyzed.     

Synthesis and preliminary biological evaluation of a new pyridocarbazole derivative covalently linked to a thymidine nucleoside as a potential targeted antitumoral agent. I

The therapy of human cancer is one of the more pursued goals by medicinal chemistry research. Most of the compounds clinically used as a treatment owe their efficacy to their cytotoxic interaction (direct or indirect) with nuclear DNA. This interaction results in the inhibition of DNA synthesis and the degradation of nucleic strands. Ellipticine is a naturally occurring 6H-pyrido[4,3-b]carbazole alkaloid endowed with antitumor activity, and several ellipticine derivatives have been used in clinical trials. We previously reported some 1,4-dimethyl-9H-carbazole derivatives structurally related to ellipticine. The purpose of our research was to transform the pyridocarbazole in a prodrug so that it would have more penetration in the tumor cells and block their replication. Our prodrug is slowly hydrolyzed in human plasma in the corresponding acid. From these preliminary results, we deduce that our compound can block cellular replication. Our hypothesis is that the antitumoral activity is probably related to the induction of damage to DNA, without cellular lysis in the short term.     

Reactivation pathway of the hydrogenase H‐cluster: Density functional theory study

This work puts forth a reaction pathway for the reactivation of exogenous ligand inhibited H‐cluster, the active site of Fe‐only hydrogenases. The H‐cluster is a dimetal complex, Fe–Fe, with the metal centers bridged by di(thiomethyl)amine. Exogenous ligands, H₂O, and OH^?, are bound to the distal iron (Fe_d). Density functional theory (DFT) calculations on the native and ruthenium‐modified H‐cluster have been performed using the B3LYP functional with 6‐31+G** and 6‐311+G** basis sets. We have ascertained that there is a thermodynamically favorable pathway for the reactivation of the OH^? inhibited H‐cluster, which proceeds by an initial protonation of the Fe_d–OH^? complex. The proposed reaction pathway has all its intermediate reactions ensue exothermically. © 2006 Wiley Periodicals, Inc. Int J Quantum Chem, 2007