Fullerenes in asphaltenes and other carbonaceous materials: natural constituents or laser artifacts.

The presence of fullerenes as natural constituents of carbonaceous materials or their formation as laser artifacts during laser desorption ionization (LDI) mass spectrometry (MS) analysis is reinvestigated and reviewed. The results using asphaltene samples with varying composition as well as standard polycyclic aromatic hydrocarbons (PAH) and fullerene samples as models have demonstrated that indeed Cn ring fullerenes are not natural constituents but they are formed as common and often as predominant artifacts upon laser radiation, and a series of incorrect assignments based on LDI-MS data of several carbonaceous materials seems unfortunately to have been made. When the present results are evaluated also in the light of the vast literature on LDI-MS of carbonaceous materials, the formation of fullerene artifacts seems particularly common for LDI-MS analysis of asphaltenes and other carbonaceous samples with considerably high levels of PAH and varies according to the type of laser used, and the intensity of the laser beam.

The multicomponent Hantzsch reaction: comprehensive mass spectrometry monitoring using charge-tagged reagents.

A novel strategy for the ESI-MS monitoring of reaction solutions involving the alternate use of permanently charge-tagged reagents has been used for comprehensive mass spectrometry monitoring of the multicomponent Hantzsch 1,4-dihydropyridine reaction. By placing a charge tag on either, or both, of the two key reactants, ion suppression effects for ESI were eliminated or minimized, and comprehensive detection of charge-tagged intermediates was achieved. The strategy allowed the trapping and characterization of the important intermediates in the mechanism for 1,4-dihydropyridine formation.

Nanoassisted laser desorption-ionization-MS imaging of tumors.

The ability of nanoassisted laser desorption-ionization mass spectrometry (NALDI-MS) imaging to provide selective chemical monitoring with proper spatial distribution of lipid profiles from tumor tissues after plate imprinting has been tested. NALDI-MS imaging identified and mapped several potential lipid biomarkers in a murine model of melanoma tumor (inoculation of B16/F10 cells). It also confirmed that the in vivo treatment of tumor bearing mice with synthetic supplement containing phosphoethanolamine (PHO-S) promoted an accentuated decrease in relative abundance of the tumor biomarkers. NALDI-MS imaging is a matrix-free LDI protocol based on the selective imprinting of lipids in the NALDI plate followed by the removal of the tissue. It therefore provides good quality and selective chemical images with preservation of spatial distribution and less interference from tissue material. The test case described herein illustrates the potential of chemically selective NALDI-MS imaging for biomarker discovery.

Phosphatidylcholine and sphingomyelin profiles vary in Bos taurus indicus and Bos taurus taurus in vitro- and in vivo-produced blastocysts.

Lipid droplets, subspecies (Bos taurus indicus vs. Bos taurus taurus), and in vitro culture are known to influence cryopreservation of bovine embryos. Limited information is available regarding differences in membrane lipids in embryo, such as phosphatidylcholines (PC) and sphingomyelins (SM). The objective of the present study was to compare the profiles of several PC and SM species and relate this information to cytoplasmic lipid levels present in Nellore (B. taurus indicus) and Simmental (B. taurus taurus) blastocysts produced in vitro (IVP) or in vivo (ET). Simmental and IVP embryos had more cytoplasmic lipid content than Nellore and ET embryos (n = 30). Blastocysts were submitted to matrix-assisted laser desorption/ionization mass spectrometry. Differences in the PC profile were addressed by principal component analysis. The lipid species with PC (32:1) and PC (34:1) had higher ion abundances in Nellore embryos, whereas PC (34:2) was higher in Simmental embryos. IVP embryos had less abundant ions of PC (32:1), PC (34:2), and PC (36:5) compared to ET embryos. Moreover, ion abundance of PC (32:0) was higher in both Nellore and Simmental IVP embryos compared to ET embryos. Therefore, mass spectrometry profiles of PC and SM species significantly differ with regard to unsaturation level and carbon chain composition in bovine blastocysts due to subspecies and in vitro culture conditions. Because PC abundances of Nellore and Simmental embryos were distinct (34:1 vs. 34:2), as were those of IVP and ET embryos (32:0 vs. 36:5), they are potential markers of postcryopreservation embryonic survival.

On the mechanism of the Dakin-West reaction.

The mechanism of the Dakin-West reaction has been thoroughly investigated by monitoring the reaction using ESI-MS/MS techniques in combination with M06-2X/6-311++G(d,p) calculations. Several of the key intermediates in the previously proposed "azlactone" mechanism have been experimentally detected and characterized. In particular, interception of the mixed anhydrides involved in the early and late stages of the mechanistic scheme, as well as of the cyclic acyl-oxazolone intermediate, supports the original pathway suggested by Dakin and West. All intermediates and transition structures involved in several competing mechanisms have been calculated. The theoretical calculations support the experimental results and corroborate the proposed "azlactone" mechanism. The pathway involving the cyclic oxazolone ("azlactone") intermediate represents an energy barrier more than 3 kcal mol(-1) lower than for the competing aldol-type mechanism, thus ruling out this alternative mechanism. The DFT calculations explain the observed ESI-MS data and assess those intermediates which the experiments cannot fully elucidate.

Venturi easy ambient sonic-spray ionization.

The development and illustrative applications of an ambient ionization technique termed Venturi easy ambient sonic-spray ionization (V-EASI) is described. Its dual mode of operation with Venturi self-pumping makes V-EASI applicable to the direct mass spectrometric analysis of both liquid (V(L)-EASI) and solid (V(S)-EASI) samples. V-EASI is simple and easy to assemble, operating solely via the assistance of a sonic stream of nitrogen or air. The sonic gas stream causes two beneficial and integrated effects: (a) the self-pumping of solutions via the Venturi effect and (b) sonic-spray ionization (SSI) of analytes either in solution or resting on solid surfaces. In its liquid mode, V(L)-EASI is applicable to analytes in solution, forming negatively and/or positively charged intact molecular species in a soft fashion with little or no fragmentation. In its solid mode, V(S)-EASI relies on Venturi self-pumping of a proper SSI solvent solution in combination with SSI to form a stream of bipolar charged droplets that bombard the sample surface, causing desorption and ionization of the analyte molecules. As for its precursor technique (EASI), V-EASI generates bipolar droplets with considerably lower average charging, which increases selectivity for ionization with high signal-to-noise ratios and clean spectra dominated by single molecular species with minimal solvent ions. V-EASI also operates in a voltage-, heat-, and radiation-free fashion and is therefore free of thermal, electrical, or discharge interferences.

Synthesis of potentially bioactive PABA-related N-(aminoalkyl)lactamic amino acids and esters via selective S(N)Ar reactions.

Potentially bioactive N-(aminoalkyl)lactamic amino acids and esters were synthesized in satisfactory to good yields by S(N)Ar reactions of aromatic acids with N-(3-aminopropyl)lactams followed by esterification with tertiary amino alcohols. The addition-elimination S(N)Ar mechanism was confirmed by NMR and MS measurements.

C-H functionalization of 1,4-naphthoquinone by oxidative coupling with anilines in the presence of a catalytic quantity of copper(II) acetate.

The oxidative addition of anilines (2) with 1,4-naphthoquinone (3) to give N-aryl-2-amino-1,4-naphthoquinones (1) was found to be catalyzed by copper(II) acetate. In the absence of the catalyst, the reactions are slower and give lower yields with the formation of many colateral products. In the presence of 10 mol % hydrated copper(II) acetate, the reactions are generally more efficient in that they are cleaner, higher yielding, and faster.

Efficient phosphodiester hydrolysis by luminescent terbium(III) and europium(III) complexes.

The synthesis and structures of two new isostructural mononuclear [Ln(L)(NO(3))(H(2)O)(3)](NO(3))(2) complexes, with Ln = Tb (complex 1) and Eu (complex 2), which display high activity in the hydrolysis of the substrate 2,4-bis(dinitrophenyl)phosphate, are reported. These complexes displayed catalytic behavior similar to the mononuclear gadolinium complex [Gd(L)(NO(3))(H(2)O)(3)](NO(3))(2) previously reported by us (Inorg. Chem. 2008, 47, 2919-2921); one hydrolysis reaction in two stages where the diesterase and monoesterase activities could be monitored separately, with the first stage dependent on and the second independent of the complex concentration. Through potentiometric studies, electrospray ionization mass spectrometry (ESI-MS) analysis, and determination of the kinetic behaviors of 1 and 2 in acetonitrile/water solution, the species present in solution could be identified and suggested a dinuclear species, with one hydroxo group, as the most prominent catalyst under mild conditions. The complexes show high activity (k(1) = 7 and 18 s(-1) for 1 and 2, respectively) and catalytic efficiency. Complexes 1 and 2 were found to be active toward the cleavage of plasmid DNA, and complete kinetic studies were carried out. Studies with a radical scavenger (dimethylsulfoxide) confirmed the hydrolytic action of 1 and 2 in the cleavage of DNA. Studies on the incubation of distamycin with plasmid DNA suggested that 1 and 2 are regio-specific, interacting with the minor groove of DNA. These complexes displayed luminescent properties. Complex 1 showed higher emission intensity than 2 due to a more efficient energy transfer between triplet and emission levels of terbium (T --> (5)D(4)), along with nonradiative deactivation mechanisms of the excited states of europium via multiphonon decays and the ligand-to-metal charge transfer state. Lifetime measurements of the (5)D(4) and (5)D(0) excited levels for 1 and 2, respectively, indicated the numbers of coordinated water molecules for the complexes.

Evidence of regulatory myeloid dendritic cells and circulating inflammatory epidermal dendritic cells-like modulated by Toll-like receptors 2 and 7/8 in adults with atopic dermatitis.

BACKGROUD: Atopic dermatitis (AD) is a chronic, inflammatory skin disease characterized by intense pruritus and xerosis. Dendritic cells (DC) play an essential role in tissue inflammation in atopic dermatitis (AD) skin, especially the inflammatory epidermal dendritic cells (IDEC), a particular subset of myeloid dendritic cells (mDC). The aim of the present study was to assess the phenotype and function of mDC and circulating IDEC-like in peripheral blood mononuclear cells (PBMC) of adults with AD. METHODS: We selected 21 AD patients and 21 non-AD controls, age and gender matched. Expressions of FcεRI, CD36, TNF, IFN- γ, and IL-10 in mDC were analyzed by flow cytometry under various stimuli, such as staphylococcal enterotoxin B (SEB), TLR2 (Pam3CSK4), TLR4 (LPS), and TLR7/8 (CL097) agonists. RESULTS: The most prominent findings in AD patients were: (i) enhanced frequency of IL-10 under TLR4 (LPS), and decreased frequency of IFN-γ and TNF under TLR2 (Pam3CSK4) and 7/8 (CL097) stimulation in classic mDC; (ii) elevation of circulating IDEC-like frequency with TLR2 (Pam3CSK4) stimuli, augmented frequency of IFN-γ in nonstimulated condition, and of IL-10 under TLR7/8 (CL097) stimuli in IDEC-like population. CONCLUSIONS: In AD individuals, classic mDC showed an immunomodulatory profile, favoring tolerance in a combined action with IDEC-like, and inducing Th1 polarization. Our findings indicate a potential role of IDEC-like in the maintenance of inflammation in atopic dermatitis patients; moreover, IDEC-like may exert a regulatory impact on T cells of AD individuals through IL-10, often induced by agonist mimicking single stranded RNA virus.

Dataset on lipid profile of bovine oocytes exposed to Lα-phosphatidylcholine during in vitro maturation investigated by MALDI mass spectrometry and gas chromatography-flame ionization detection.

Data presented in this article are related with the research article entitled "Effect of soybean phosphatidylcholine on lipid profile of bovine oocytes matured in vitro" [1]. This article describes the differences in the relative abundance of the lipid ions detected by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) in control and Lα-phosphatidylcholine-treated oocytes. In addition, the fatty acids (FA) content in pure Lα-phosphatidylcholine supplement and oocytes was analyzed by gas chromatography-flame ionization detection (GC-FID). The dataset provides information and inputs for further studies aiming to optimize in vitro maturation conditions and cryotolerance of mammalian oocytes.

Effect of soybean phosphatidylcholine on lipid profile of bovine oocytes matured in vitro.

The phospholipid (PL) composition of embryo and oocyte membranes affects thermal phase behavior and several physicochemical properties such as fluidity and permeability. The characterization of PL profiles and the development of suitable in vitro maturation (IVM) protocols, that are able to modify membrane's composition, may result in significant improvements in oocyte developmental potential and cryotolerance. Using soybean phosphatidylcholine (PC) as a model supplement, we evaluated the effect of PL supplementation during IVM on bovine cumulus-oocyte-complex (COC). Substantial changes in the lipid profiles of oocyte membrane were observed and associated with pre-implantation data. The propensity of the PC supplement to become soluble in the maturation medium and/or diffuse into mineral oil was also assessed. Oocytes were matured in TCM without supplementation, i.e. control, (n=922) or supplemented with 50 or 100µM PC (n=994). The maturation media and mineral oil pre- and post- IVM, along with control and PC-treated oocytes were then analyzed using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS), and the lipid profiles were compared via principal component analysis (PCA). Soybean PCs are bioavailable and stable in IVM medium; further, PCs did not diffuse to the mineral oil, which also remained unaltered by the metabolism of treated oocytes. PC supplementation at 100µM resulted in substantially greater relative abundances of polyunsatured PL, namely PC (32:1), PC (34:2), PC (36:6), PC (36:4), and PC (38:6), in oocyte membrane. These differences indicated that short-term exposure to the PC supplement could indeed modify the lipid composition of IVM-oocytes in a dose-dependent manner. Membrane incorporation of polyunsaturated molecular species of PC was favored, and does so without compromising the viability of the subsequent embryo in regards to cleavage, blastocyst development and hatching rate. The reported approach will allow for the development of novel strategies to modulate oocyte membrane dynamics and structure.

Lipidome signatures in early bovine embryo development.

Mammalian preimplantation embryonic development is a complex, conserved, and well-orchestrated process involving dynamic molecular and structural changes. Understanding membrane lipid profile fluctuation during this crucial period is fundamental to address mechanisms governing embryogenesis. Therefore, the aim of the present work was to perform a comprehensive assessment of stage-specific lipid profiles during early bovine embryonic development and associate with the mRNA abundance of lipid metabolism-related genes (ACSL3, ELOVL5, and ELOVL6) and with the amount of cytoplasmic lipid droplets. Immature oocytes were recovered from slaughterhouse-derived ovaries, two-cell embryos, and eight- to 16-cell embryos, morula, and blastocysts that were in vitro produced under different environmental conditions. Lipid droplets content and mRNA transcript levels for ACSL3, ELOVL5, and ELOVL6, monitored by lipid staining and quantitative polymerase chain reaction, respectively, increased at morula followed by a decrease at blastocyst stage. Relative mRNA abundance changes of ACSL3 were closely related to cytoplasmic lipid droplet accumulation. Characteristic dynamic changes of phospholipid profiles were observed during early embryo development and related to unsaturation level, acyl chain length, and class composition. ELOVL5 and ELOVL6 mRNA levels were suggestive of overexpression of membrane phospholipids containing elongated fatty acids with 16, 18, and 20 carbons. In addition, putative biomarkers of key events of embryogenesis, embryo lipid accumulation, and elongation were identified. This study provides a comprehensive description of stage-specific lipidome signatures and proposes a mechanism to explain its potential relationship with the fluctuation of both cytoplasmic lipid droplets content and mRNA levels of lipid metabolism-related genes during early bovine embryo development.

Staphylococcal enterotoxin B induces specific IgG4 and IgE antibody serum levels in atopic dermatitis.

BACKGROUND: Atopic dermatitis (AD) is a chronic recurrent inflammatory disease, with prevalence of about 10-20% in children and 1-3% in adults. Staphylococcus aureus is present in 80-100% of skin from atopic patients and is related to worsening of the disease by the action of enterotoxins. The aim of this study was to evaluate the profile of anti-Staphylococcus aureus enterotoxin B (SEB) antibody isotypes and IgG subclass levels in adult AD. METHODS: We selected 38 patients with AD, diagnosed by Hanifin and Rajka's criteria, aged between 18 and 65, and 26 healthy controls (HC). The severity of the disease was established according to the Eczema Area and Severity Index and patients graded as mild (28%), moderate (58%), and severe (14%). Sera were assessed for IgG subclasses, IgA, IgM, and IgE against SEB by ELISA. RESULTS: Elevated circulating IgE and IgG4 anti-SEB antibody levels associated with decreased IgA and IgM levels were detected in patients with AD, when compared to HC individuals. The severity of AD was related to low IgG1 and IgG3 levels and a high IgE antibody response to SEB. Interestingly, absence of IgG4 response to SEB was lower in patients with AD (2.63%), when compared to controls (34.6%), while a similar absence was detected for IgG1 and IgE antibodies (AD, 23.3 and 18.4% vs. HC, 38.5 and 19.2%). CONCLUSION: Our findings evidenced a contributing role for IgG4 and IgE antibodies in AD pathogenesis, which are triggered by staphylococcal superantigens.

Chemo-, regio- and stereoselective Heck arylation of allylated malonates: mechanistic insights by ESI-MS and synthetic application toward 5-arylmethyl-γ-lactones.

We describe herein a general method for the controlled Heck arylation of allylated malonates. Both electron-rich and electron-poor aryldiazonium salts were readily employed as the aryl-transfer agents in good yields and in high chemo-, regio-, and stereoselectivity without formation of decarboxylated byproducts. Reaction monitoring via ESI-MS was used to support the formation of chelated Pd species through the catalytic cycle. Additionally, some Heck adducts were successfully used in the total synthesis of pharmacologically active γ-lactones.

Optimal single-embryo mass spectrometry fingerprinting.

In pre-implantation embryos, lipids play key roles in determining viability, cryopreservation and implantation properties, but often their analysis is analytically challenging because of the few picograms of analytes present in each of them. Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) allows obtaining individual phospholipid profiles of these microscopic organisms. This technique is sensitive enough to enable analysis of individual intact embryos and monitoring the changes in membrane lipid composition in the early stages of development serving as screening method for studies of biology and biotechnologies of reproduction. This article introduces an improved, more comprehensive MALDI-MS lipid fingerprinting approach that considerably increases the lipid information obtained from a single embryo. Using bovine embryos as a biological model, we have also tested optimal sample storage and handling conditions before the MALDI-MS analysis. Improved information at the molecular level is provided by the use of a binary matrix that enables phosphatidylcholines, sphingomyelins, phosphatidylserines, phosphatidylinositols and phosphoethanolamines to be detected via MALDI(±)-MS in both the positive and negative ion modes. An optimal MALDI-MS protocol for lipidomic monitoring of a single intact embryo is therefore reported with potential applications in human and animal reproduction, cell development and stem cell research.

Microorganisms in cryopreserved semen and culture media used in the in vitro production (IVP) of bovine embryos identified by matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS).

Commercial cattle breeders produce their own herd offspring for the dairy and beef market using artificial insemination. The procedure involves sanitary risks associated with the collection and commercialization of the germplasm, and the in vitro production and transfer of the bovine embryos must be monitored by strict health surveillance. To avoid the spreading of infectious diseases, one must rely on using controlled and monitored germplasm, media, and reagents that are guaranteed free of pathogens. In this article, we investigated the use of a new mass spectrometric approach for fast and accurate identification of bacteria and fungi in bovine semen and in culture media employed in the embryo in vitro production process. The microorganisms isolated from samples obtained in a commercial bovine embryo IVP setting were identified in a few minutes by their conserved peptide/protein profile, obtained applying matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS), matched against a commercial database. The successful microorganisms MS identification has been confirmed by DNA amplification and sequencing. Therefore, the MS technique seems to offer a powerful tool for rapid and accurate microorganism identification in semen and culture media samples.

Separation of isomeric disaccharides by traveling wave ion mobility mass spectrometry using CO2 as drift gas.

The use of CO(2) as a massive and polarizable drift gas is shown to greatly improve peak-to-peak resolution (R(p-p) ), as compared with N(2) , for the separation of disaccharides in a Synapt G2 traveling wave ion mobility cell. Near or baseline R(p-p) was achieved for three pairs of sodiated molecules of disaccharide isomers, that is, cellobiose and sucrose (R(p-p) = 0.76), maltose and sucrose (R(p-p) = 1.04), and maltose and lactose (R(p-p) = 0.74). Ion mobility mass spectrometry using CO(2) as the drift gas offers therefore an attractive alternative for fast and efficient separation of isomeric disaccharides.

Wood typification by Venturi easy ambient sonic spray ionization mass spectrometry: the case of the endangered Mahogany tree.

Venturi easy ambient sonic spray ionization mass spectrometry in both its liquid (V(L) -EASI-MS) and solid sample modes (V(S) -EASI-MS) is shown to provide nearly immediate and secure typification of woods, as demonstrated for Mahogany, an endangered and most valuable type of tropical wood. This reddish wood displays unique phytochemical markers (phragmalin-type limonoids) which are rapidly detected from the wood surface by V(S) -EASI-MS or from a simple methanol extract of a tiny wood chip by V(L) -EASI-MS. Unique profiles were obtained for Mahogany (Swietenia macrophylla) whereas genuine samples of six other similar types of woods, which are commonly falsified by artificial coloring and commercialized as Mahogany, display also typical but dissimilar pythochemical profiles as compared to that of the authentic wood. Variable and atypical chemical profiles were observed for artificially colored woods. Secure chemical characterization via V(S) -EASI-MS or V(s) -EASI-MS fingerprints of Mahogany and other types of woods with similar appearance should help to control the illegal logging and trade of this and other endangered woods and their falsification, and to create certified standards.

On the mechanism of the aza-Morita-Baylis-Hillman reaction: ESI-MS interception of a unique new intermediate.

Solutions of aza-Morita-Baylis-Hillman (aza-MBH) reactions were directly monitored by ESI(+)-MS(/MS) spectrometry to obtain information on their mechanism. A unique bis-sulfonamide intermediate was intercepted and characterized and, based on this novel species, a mechanism that rationalizes the uniqueness of aza-MBH reactions is proposed.