Electron Spray Ionization Mass Spectrometry Research Articles

Nanosized divalent transition metal ion‐prompted macrocyclic complexes: Synthesis, characterization, molecular modeling, and biological studies

AbstractIn the present research work, four new 14‐membered tetraazamacrocylic complexes of Cobalt(II), Nickel(II), Copper(II) and Zinc(II) with (1E,14E)‐8,8,17,17‐tetramethyl‐2,5,11,14‐tetraazatricyclo[13.3.1.16,10]icosa‐1,5,10,14‐tetraene were synthesized using the template methodology that leads to the formation of a complex of type [MLX2] in which L is a macrocyclic ligand derived from ethylenediamine (ED) and 5,5‐dimethylcyclohexanedione (DCH) and X = Cl−/CH3COO−. Spectroscopic, physical, and analytical characterization of complexes was carried out with the assistance of infra‐red, nuclear magnetic resonance, electron spin resonance, Ultraviolet‐visible, powder X‐Ray diffraction, electron spray ionization ‐ mass spectroscopy (ESI‐MS), thermogravimetric analysis, magnetic susceptibilities, and carbon hydrogen nitrogen analysis. The information regarding the monomeric and nonelectrolytic behavior was elucidated from ESI‐MS and molar conductance values. Powder X‐Ray diffractogram studies point toward the crystalline or amorphous nature of the complexes. All the compounds exhibited the nonelectrolytic nature. Semiempirical calculations were performed using Gaussian 09 software and quantum chemical parameters were determined. Newly designed macrocyclic complexes were examined for their antifungal and antibacterial potency by the Agar well diffusion method. In‐vitro DNA binding studies were carried out in order to understand the extent and nature of binding shown by the complexes with the DNA. In addition to this, in‐silico absorption distribution metabolism excretion toxicity studies were also carried out for the interpretation of drug‐like properties in the newly synthesized complexes.

A New Bioactive Cembranolide Sarcophytonolide V from Bornean Soft Coral Genus Sarcophyton

A new cembranolide diterpene, sarcophytonolide V (1), along with 6 known compounds, isosarcophytonolide D (2), (4 Z,8 S*,9 R*,12 E,14 E)-9-hydroxy-1-(prop-1-en-2-yl)-8,12-dimethyl-oxabicyclo[9.3.2]-hexadeca-4,12,14-trien-18-one (3), (7 E,11 E)-3,4-epoxy-7,11,15-cembratriene (4), (1 S*,3 S*,4 S*,7 E,11 E)-3,4-epoxy-13-oxo-7,11,15-cembratriene (5), (-)-eunicenone (6), and 2-[( E, E, E)-7′,8′-epoxy-4′,8′,12′-trimethylcyclotetradeca-1′,3′,11′-trienyl]propan-2-ol (7) were isolated from the Bornean soft coral Sarcophyton sp. Their structures were elucidated based on spectroscopic data, such as nuclear magnetic resonance (NMR) and high resolution electron spray ionization mass spectroscopy (HRESIMS). These compounds were evaluated for their biological activity against marine pathogenic fungi.

TWO NOVEL FLAVONE C-GLYCOSIDES ISOLATED FROM AFROCARPUS GRACILIOR: POM ANALYSES AND IN VITRO CYTOTOXIC ACTIVITY AGANIST HEPATOCELLULAR CARCINOMA

Objective: Cancer is considered as one of the top reasons of death and the number of cases increasing gradually. Cancer is severe clinical difficulty to the health caution system. This study explored two novel polyphenols of Afrocarpus gracilior Pilger growing in Egypt and evaluated their cytotoxic activity.
 Methods: Methanolic (80%) extract of the leaves of A. gracilior was subjected to column chromatography; the chemical structures of the isolated compounds were established by advanced spectral techniques: UV, 1H, 13C NMR, two dimensional NMR (2D NMR) and electron spray ionization mass spectroscopy (ESI-MS). Compounds 1 and 2 were studied for their cytotoxic activity against hepatocellular carcinoma (Hep-G2) using sulforhodamine B (SRB) assay. Furthermore the pharmacokinetics profiles of these molecules were accessed by employing Petra/Osiris/Molinspiration (POM) analyses.
 Results: Two novel C-flavonoid glycosides were isolated [1: Apigenin 8-C-β-D-glucopyranosyl-(1```→4``)-O-β-D-glucopyranoside] and [2: 7-O methyl-luteolin 8-C-β-glucopyranosyl-(1```→4``)-O-β-D-glucopyranoside]. They exhibited significant cytotoxic activity (IC50 = 9.02 and 15.61 µg/ml, respectively) against Hep-G2 cells. The POM analyses revealed that the activity of these two compounds depends on the presence of glucosyl and alkyl groups at the internal and terminal atmosphere of the compounds.
 Conclusion: These findings demonstrated that the leaves of A. gracilior contain a series of bioactive polyphenolic compounds with significant cytotoxic properties against hepatocellular carcinoma and may be used as alternative anticancer agents for doxorubicin. On the basis of POM calculations, it will be interesting to develop some alternative flavones because the deglucosylated derivatives have a better drug score than parent molecules. This preliminary study will be extended to other strains of cancer.

Mass Fingerprints of Tomatoes Fertilized with Different Nitrogen Sources Reveal Potential Biomarkers of Organic Farming.

Direct-injection electron spray ionization mass spectrometry (DIESI-MS) can be used to quantify the whole set of positive and negative ions in complex biological samples. A cherry tomato cultivar was grown inside a greenhouse in soil pots supplemented with different nitrogen sources. Organic cultivation increased fruit dry matter while conventional chemical fertilizers increased yield due to higher water content. While soluble sugars were unaltered, secondary metabolism of tomato fruit was highly sensitive to compost soil supplied to the roots. From a total of ~1647 DIESI-MS signals, 725 revealed significant differences between treatments. Heatmap biclustering showed that ionomic differences were robustly maintained in independent experiments carried out during three consecutive years. The ionomic fingerprints allowed reproducible sample classification, reflecting the effect of organic farming on tomato fruit quality. Specific biomarker ions could be identified for various nitrogen sources. We propose DIESI-MS as an up-front strategy for plant food characterization aiming to identify the ions with the most significant differences across genotypes or agronomic conditions.

Characterization of Intracellular Peptides from Zebrafish (Danio rerio) Brain.

Peptides represent a large class of cell signaling molecules, and they are mainly produced by the classical secretory pathway or during protein degradation. The peptide profile of Danio rerio (zebrafish) shows a lack of information when compared with other consolidated animal models. The aim of this work was to characterize the peptide profile of zebrafish brain by using triplex reductive methylation of amines labeling and liquid chromatography coupled to electron spray mass spectrometry. A total of 411 peptide fragments were detected and 125 peptide sequences could be solved. Further analysis suggested that most of the peptides were fragments of intracellular cytosolic and mitochondrial proteins, and that 60% of the precursor proteins were cleaved at either their N- or C-terminal. The most common residue in the P1 position was leucine whereas other common residues were lysine, alanine, arginine, and phenylalanine. Rare cleavage sites at P1 position were histidine, glutamic acid, and isoleucine. The peptide profile of zebrafish brain has similarities with results previously described in mice brain peptidome studies. Thus, this study represents an important basis for the molecular understanding of zebrafish and its use as a model for human diseases.

Document ink dye age estimation by direct injection-mass spectrometry and correlation analysis

Unsupervised Linkage Threshold (ULT) method was applied for the analysis of ballpoint pen inks with the purpose of estimating the date on which manuscripts were launched on documents. In order to verify ink degradation patterns over time, manuscript samples were removed from forensic expert reports produced along the years ranging from 1962 to 2014, and which had been stored in the archives of the National Institute of Criminalistics (NIC) of the Brazilian Federal Police. A total of 505 ink samples were analyzed by Electron Spray Ionization (ESI) mass spectrometry, and these showed a significant variation in the ink formulation over time. From the normalized mass spectrometry peak areas, the ULT method was employed in the correlation analysis of ink manuscripts using the cosine squared function as a similarity measure. The method allowed a simple, fast and reliable determination of threshold correlation values as well as ink dating confidence intervals for each kind of formulation studied. It has been observed that dye stability may change drastically depending upon the chemical group (methyl or ethyl) lost in the degradation process. Due to its statistical nature, the developed method does not rely upon prior knowledge of factors such as storage conditions or initial dye formulation. This feature can be an advantage in real forensic applications since this type of information is not usually available to forensic experts.

Molecular changes of phenolic-protein interactions in isolated proteins from flaxseed and soybean using Native-PAGE, SDS-PAGE, RP-HPLC, and ESI-MS analysis.

The effects of protein-phenolic interactions on the molecular characteristics of soybean and flaxseed proteins were investigated. Proteins were isolated from soybean and flaxseed using isoelectric precipitation, followed by extraction of free and bound phenolics. The effects of elimination of the phenolic compounds on molecular characteristics of the protein isolates were studied using reversed phase-high performance liquid chromatography (RP-HPLC), Native and sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis (PAGE) and electron spray ionization-mass spectrometry (ESI-MS). The Native-PAGE fractions from isolated proteins from full-fat flaxseed and soybean revealed that protein migration was affected by removal of bound phenolics. SDS-PAGE from full-fat and defatted protein isolates of flaxseed and soybean revealed that the removal of bound phenolics affected the molecular characteristics of protein subunits. Soybean protein isolates had protein-phenolic interactions through acidic and basic subunits. RP-HPLC and ESI-MS showed that the removal of bound and free phenolics had only minor effects on the molecular characteristics of isolated proteins from defatted and full-fat soybean. With respect to isolated proteins from flaxseed, the removal of bound phenolics showed little effect on the electrophoretic behavior of the proteins or the protein subunits. PRACTICAL APPLICATIONS: Phenolic-protein and phenolic-lipid-protein interactions may affect the nutritional, physicochemical, and functional properties of isolated proteins from food in flaxseed and soybean.

Qualitative and quantitative analysis of precipitate formation following interaction of chlorhexidine with sodium hypochlorite, neem, and tulsi.

Introduction:The interaction between chlorhexidine (CHX) and sodium hypochlorite (NaOCl) yields a thick precipitate capable of occluding dentinal tubules. Previous studies are unclear as to the above-mentioned precipitate contains para-chloroaniline (PCA) or not. PCA is a known toxic and carcinogenic compound which may lead to methemoglobinemia in humans.Aim:This study aims to evaluate the precipitate formed on combination of different irrigants, weigh the amount of precipitate formed and to analyze the precipitate for PCA by using thin layer chromatography (TLC), high performance liquid chromatography (HPLC), column chromatography (CC), electron spray ionization mass spectrometry (ESI-MS), Ultraviolet (UV), and nuclear magnetic resonance (1H-NMR and C-13 NMR).Materials and Methods:Four different irrigants namely 2% CHX gluconate, 3% NaOCl, 5% neem and 5% tulsi were taken in different test tubes. Group 1, 2 and 3 included 1 ml 2% CHX combined with 1 ml each of 3% NaOCl, 5% neem and 5% tulsi. Group 4 and 5 comprised of 1 ml 3% NaOCl in combination with 1 ml 5% each of neem and tulsi. Finally, group 6 constituted 1 ml 5% neem mixed with 1 ml 5% tulsi. Each group was observed for 2 min for the formation of any precipitate, and the formed precipitate was weighed and analyzed using 1H-NMR and C-13 NMR, TLC, CC, HPLC, ESI-MS, and UV.Statistical Analysis:One-way ANOVA and Post hoc–Tukey test were used.Results:Presence of PCA was detected in group 1 (CHX + NaOCl), group 2 (CHX + neem) and group 3 (CHX + tusli) in all the sensitive methods employed.Conclusion:The presence of PCA in precipitate was confirmed by TLC, CC, HPLC, ESI-MS, and UV. Based on the results of the present study, we assume that components in CHX are responsible for precipitate formation which contains PCA as well. Extrusion of precipitate beyond the apex may cause periapical tissue damage and delay wound healing at the same time.

Visible-light-induced degradation of rhodamine B by nanosized Ag2S-ZnS loaded on cellulose.

In the present work, new visible-light-active nanosized Ag2S-ZnS loaded on cellulose (AZCE) was synthesized by a precipitation method. The AZCE composite was systematically characterized using powder X-ray diffraction (XRD), scanning electron microscopy (SEM), transmission electron microscopy (TEM), Brunauer-Emmett-Teller (BET) surface area, and UV-visible diffuse reflectance spectroscopy (UV-DRS). The activities of the photocatalysts were evaluated for rhodamine B dye (RhB) degradation under simulated sunlight and the amounts of the dye samples were analysed using a UV-vis spectrophotometer at λmax 554 nm. The effects of the reaction conditions such as pH, catalyst, hydrogen peroxide and dye concentration on the photodegradation rate have been investigated. The degradation profile reveals that 30 ppm of the dye could be effectively oxidized using 30 mg of the AZCE dose in the pH range 4-12 within 90 min. The oxidation of the RhB dye follows first-order kinetics and the rate constant was calculated to be 6.4 × 10-3 min-1. Various organic intermediates were identified during degradation using high performance liquid chromatography (HPLC), total organic content (TOC) and electron-spray ionization-mass spectrometry (ESI-MS). In order to determine the effectiveness of AZCE photocatalytic activity, other catalysts such as Ag2S loaded on cellulose (AZE) and ZnS loaded on cellulose (ZCE) were used as photocatalysts. The results show that photocatalytic activity follows the order AZCE > ACE > ZCE and this is due to the fact that a cellulose network is used as a catalyst carrier. The alkali pre-treated cellulose provides an activated surface hydroxyl groups to enhance the deposition efficiencies of Ag2S and ZnS and thereby a large amount of visible light can be absorbed and the photocatalytic activity is increased.

Qualitative and quantitative determination of Atractylodes rhizome using ultra-performance liquid chromatography coupled with linear ion trap-Orbitrap mass spectrometry with data-dependent processing.

A quick and effective workflow based on ultra-performance liquid chromatography coupled with electron spray ionization and LTQ-Orbitrap mass spectrometry (UPLC-LTQ-Orbitrap MS) was established for compositional analysis and screening of the characteristic compounds of three species of Atractylodes rhizome for quality evaluation. This technique was employed to determine the seven main components in Atractylodes rhizome samples. Ultimately, 78 constituents were identified; of these, seven characteristic compounds were selected for species discrimination, comprising atractylodin (63), atractylenolide I (43), atractylenolide II (49), atractylenolide III (53), atractylon (69), methyl-atractylenolide II (54) and (4E,6E,12E)-tetradecadecatriene-8,10-diyne-1,3-diacetate (59). The seven main compounds, including six characteristic compounds, were simultaneously determined in 29 batches of Atractylodes rhizome samples. Thus, the method validation showed acceptable results. Quantitative analysis showed significantly different contents of the seven main components among the three species of Atractylodes rhizome, which indicates possible distinctions in the pharmacological effects. This established method can simultaneously provide qualitative and quantitative results for compositional characterization of Atractylodes rhizomes and for quality control.

Use of ESI-MS for semi-quantitative estimation of inactive precursor in no-carrier-added 131I- meta-Iodobenzylguanidine radiopharmaceutical preparation

No-carrier-added (nca)-131I-meta-Iodobenzylguadine (mIBG) is a clinical agent used for the therapy of Neuroendocrine tumors. It is prepared by reaction of radioiodine with precursors that are chemically different from mIBG. The precursor in few cases is structurally similar and may co-elute along during purification step. Presence of these precursors in final radiolabeled formulation may affect the clinical behaviour of the radiopharmaceutical. The present paper describes the use of Electron-spray ionization-Mass Spectrometry (ESI-MS) where up to nano-molar concentrations of these precursors could be estimated with high precision in the final radiolabeled formulation.

Antiviral 4-Hydroxypleurogrisein and Antimicrobial Pleurotin Derivatives from Cultures of the Nematophagous Basidiomycete Hohenbuehelia grisea.

4-Hydroxypleurogrisein, a congener of the anticancer-lead compound pleurotin, as well as six further derivatives were isolated from the basidiomycete Hohenbuehelia grisea, strain MFLUCC 12-0451. The structures were elucidated utilizing high resolution electron spray ionization mass spectrometry (HRESIMS) and 1D and 2D nuclear magnetic resonance (NMR) spectral data and evaluated for their biological activities; for leucopleurotin, we provide Xray data. While most congeners showed moderate antimicrobial and cytotoxic activity, 4-hydroxypleurogrisein emerged as an inhibitor of hepatitis C virus infectivity in mammalian liver cells.

Nitrogen-Doped Graphene Quantum Dot-Combined Sodium 10-Amino-2-methoxyundecanoate: Studies of Proinflammatory Gene Expression and Live Cell Imaging.

Marine cyanobacteria are renowned for producing bioactive secondary metabolites with great structural diversity via mixed biosynthetic pathways. Lyngbya sp., a marine cyanobacterium, produces many metabolites with anti-inflammatory potentials; nevertheless, its bioactive metabolites exercising providing protection against inflammation has been deciphered inadequate. In this study, the ethanolic fraction of the Lyngbya sp. extract was purified and identified as sodium 10-amino-2-methoxyundecanoate (SAM) using Fourier-transform infrared spectroscopy, nuclear magnetic resonance, and electron spray ionization-mass spectroscopy. SAM showed prominent inhibition of inflammation, which was analyzed by reactive oxygen species generation and nitric oxide (NO) inhibition assay. Furthermore, the anti-inflammatory potentials of SAM were evaluated in lipopolysaccharide (LPS)-induced RAW 264.7 macrophage cell lines by fluorescence-activated cell sorting analysis, which evidenced prominent decrease in COX-2 expression (∼90%) with SAM-treated cells than the control. Subsequently, a semiquantitative real-time polymerase chain reaction analysis also revealed the downregulation of COX-2, iNOS, TNF-α, NF-κß, IL-1α, IL-1ß, IL-4, and IL-6 gene expression in SAM-treated LPS-induced RAW 264.7 cells. To further enhance the delivery of SAM into the cells, it was combined with N-doped graphene quantum dots (N-GQDs) for the anti-inflammatory potentials. It resulted in improved downregulation of COX-2, iNOS, TNF-α, NF-κß, IL-1α, IL-1ß, IL-4, and IL-6 than cells treated with SAM alone. Conclusively, N-GQDs combined with SAM have the effective therapeutic potential as an inhibitor of inflammation by modulating the expression of different cytokine genes.

Anti-proliferative potential of cyclotetrapeptides from Bacillus velezensis RA5401 and their molecular docking on G-Protein-Coupled Receptors

Elucidation of bioactive chemical compounds from rhizobacteria is highly utilized in pharmaceuticals and naturopathy, due to their health benefits to human and plants. In current study, four cyclopeptides along with one phenyl amide were isolated from the ethyl acetate extract of Bacillus velezensis sp. RA5401. Their structures were determined and characterized as cycle (L-prolyl-L-leucyl)2 (1), cyclo (L-prolyl-l-valine)2 (2), cycle (L-phenylanalyl-L-propyl)2 (3), cyclo (D-pro-L-tyr-L-pro-L-tyr)2 (4) and N-(2-phenylethyl)acetamide (5) on the basis of electron spray ionization mass spectrometry (ESI-MS), nuclear magnetic resonance (NMR) techniques and comparison with the literature data. The five compounds have been isolated for the first time from this species. The effect of various concentrations of these compounds on the proliferation of MDA-MB-231 breast cancer cells was examined. It was found that 1 and 2 induced concentration-independent anti-proliferative effects, while 3, 4 and 5 inhibited cancer cell proliferation in a concentration-dependent manner. Furthermore, to determine the suitable binding targets of these compounds within cancer cell line, detailed target prediction and comparative molecular-docking studies were performed. The compounds 1 and 2 hit intracellular anti-cancer targets of proteases family, while compounds 3, 4 and 5 interacted with different membrane receptors of G-Protein-Coupled Receptors (GPCRs). In conclusion, the Bacillus velezensis RA5401 can be an ideal strain to produce anti-proliferative constituents at industrial scale.

Characterization of a red pigment from Fusarium chlamydosporum exhibiting selective cytotoxicity against human breast cancer MCF-7 cell lines.

This research aims to characterize the pigment produced by isolated fungi and to evaluate its anticancer activities. Pigment-producing fungi was isolated and identified as Fusarium chlamydosporum. The pigment was extracted with chloroform, purified by preparative TLC and characterized by Fourier transmittance infrared, electron spray ionization mass spectroscopy, liquid chromatography mass spectroscopy and nuclear magnetic resonance (NMR) (1 HNMR, 13 C NMR) spectral analysis, which revealed the pigment to be 'long chain hydrocarbons with poly unsaturated groups' (m/z 702). Pigment stability varied with different pH, temperature and sunlight conditions. The pigment-induced cell death in human breast adenocarcinoma cells MCF-7 and showed no significant toxicity in CHOK 1 cells. Lipid peroxidation assay revealed that treatment with pigment was able to reduce the lipid peroxidation caused by H2 O2 in MCF-7 cells. The F. chlamydosporum pigment is a compound of long-chain hydrocarbons with poly unsaturated groups, possessing selective cytotoxicity in MCF-7 cancer cell lines. The pigment can be used as a colouring agent in cosmetics. Its anticancer potential can be used in production of therapeutics in increasing demand cancer research.

Chemical constituents from Canarium album Raeusch and their anti-influenza A virus activities.

Two new dyhydrophaseic acid glucoside isomers, (1'S, 3'R, 5'S, 8'R, 2Z, 4E)-dihydrophaseic acid-3'-O-β-D-glucopyranoside (2) and (1'R, 3'S, 5'R, 8'R, 2Z, 4E)-dihydrophaseic acid-3'-O-β-D-glucopyranoside (4), together with 10 known compounds [myo-inositol (1), 3,4-dihydroxybenzoic acid (3), 3-O-galloyl quinic acid (5), ellagic acid (6), gallic acid (7), ethyl gallate (8), scopoletin (9), ellagic acid-4-O-β-D-glucopyranoside (10), ellagic acid-4-O-α-L-rhamnopyranoside (11), and isocorilagin (12)] were isolated from the chloroform extract of Canarium album Raeusch fruits by repeated chromatography on macroporous adsorption resin, silica gel, Sephadex LH-20, Toyopearl HW-40F, and reverse-phase C18 columns, etc. Their structures and absolute configurations were determined by comprehensive analysis of 1D- and 2D-nuclear magnetic resonance (NMR), high-resolution electron spray ionization mass spectrometry (HR-ESI-MS), ESI-MS, optical rotation, circular dichroism spectra, and comparison of NMR data with data of known compounds. Bioassay of their anti-influenza virus A activities showed that compounds 9 and 12 displayed a significant inhibitory effect with IC50 values of 22.9 ± 3.7 and 5.42 ± 0.97μg/ml, respectively.

RP-HPLC-UV Method for Estimation of Fluorouracil-Epirubicin-Cyclophosphamide and Their Metabolite Mixtures in Human Plasma (Matrix).

A combination of 5-fluorouracil (FU), epirubicin (EP) and cyclophosphamide (CP) is routinely employed in the treatment of breast cancer. The objective of this study was to develop a reverse phase high-performance liquid chromatography (HPLC)-UV method for simultaneous quantitative analysis of the triple-drug and their metabolites in plasma. RP-HPLC system with a C18 column and a diode array detector was employed. The plasma samples were precipitated with acetonitrile and the supernatant was dried under a flow of nitrogen gas. The mobile phase comprised of two combinations, water (pH 4.0) and methanol (98:2 v/v), and water (pH 4.0):methanol:acetonitrile (70:13:17 v/v/v). The retention times for the compounds were determined and the parameters of validation established in plasma indicated the robustness and reliability. The corresponding HPLC peaks were confirmed using electron spray ionization mass spectrometry. FU and metabolites had a recovery of >93%; EP, epirubicinol and CP were >78% from plasma. Stability at 28-30°C in water (pH 4.0) of FU, 5,6-dihydro-5-fluorouracil and EP were higher followed by CP, EPol, fluorodeoxyuridine and fluorouridine (FUR). Storage of the drug-spiked plasma at -80°C assessed for 72 h showed a small but significant (P < 0.05) change in the recovery of FUR and EP. The method was validated in patient's plasma samples (n = 6).

Chemical composition, total phenolic content, antioxidant and antinutritional characterisation of exudate gums

Abstract Present study was conducted to determine the chemical composition, antioxidant potential and antinutritional factors in some plant gums that have been commercially used as additives in different food formulations. For this purpose, exudate gums were chosen which included commercially used acacia and karaya gum. The properties were compared with an underutilised gum exuded from apricot tree, called apricot gum. Monosaccharide composition and linkages in gum polysaccharides were studied using Electron spray ionization-mass spectrometry coupled with time-of-flight analyser (ESI-MS-TOF). The assay involves ionization of gum polysaccharides by passing their organic solution into electric field and converting them into a mist of charged droplets that are further analysed and detected using Time-of-flight mass spectrometry. Relative array of peaks elucidated the presence of glucosyl, galactosyl and a small proportion of uronic acid units in all gums. Spectral fragmentation distribution was bimodal with polydispersity values greater than or equal to two (Mw/Mn ≥ 2). In all the antioxidant assays, methanol and ethanol were separately used for extraction. Total phenolic content, ferrous ion chelating ability and reducing power of the gums varied from 0.44 to 1.25 mg GAE/g, 11.80–60.42% and 0.12–0.23 AAE/g, respectively. The DPPH radical scavenging ability (12.03–27.23%) and bile acid binding activity (40.98–46.60%) of the gums was also high. Presence of tannins (0.20–0.73 TAE/g), phytic acid (0.03–0.60 mg/g) and haemagglutinins (0.006–0.11 HU/mg protein) was observed at a very small concentration. On the other hand, the antinutritional factors like terpenoids, saponins and alkaloids were found completely absent.

Thermodynamic, Structural, and Computational Investigation on the Complexation between UO22+ and Amine-Functionalized Diacetamide Ligands in Aqueous Solution.

The stability constants (log β), enthalpies of complexation (ΔH), and entropies of complexation (ΔS) for the complexes of uranium(VI) with a series of amine-functionalized diaetamide ligands, 2,2'-benzylazanediylbis(N,N'-dimethylacetamide) (BnABDMA), 2,2'-azanediylbis(N,N'-dimethylacetamide) (ABDMA), and 2,2'-methylazanediylbis(N,N'-dimethylacetamide) (MABDMA), in aqueous solution were determined by potentiometry and calorimetry. Electronspray ionization mass spectrometry was used to verify the presence of uranium(VI) complexes in solution. The thermodynamic data indicate that the binding strengths of the three ligands with UO22+ follow the order BnABDMA < ABDMA < MABDMA, parallel to the order of the protonation constants as well as the order of the stability of the Nd3+ complexes, suggesting that the complexation of UO22+ with the ligands consist predominantly of electrostatic interactions. Denisty functional theory calculations were conducted to reveal the structures, electronic charge distribution, and energetics of the uranium(VI) complexes, providing insight into the thermodynamic trends of the complexation. Extended X-ray absorption fine structure spectroscopy was used to identify the structures of the uranium(VI) complexes in aqueous solution.

Structural Studies of Alpha-Synuclein Amyloid Filaments

Amyloid filaments of the alpha-synuclein (alpha-Syn) protein are considered the hallmark of Parkinson's disease (PD) and no biomarkers for early diagnosis have yet been developed. Thioflavin T (ThT) is the most frequently used dye to detect amyloids. We are using cryo-EM of ThT-bound alpha-Syn fibrils of PD familial variants (e.g., A30P, E46K and, A53T) to understand its mechanism of binding. The heterogeneity of amyloid filaments has been one of the greatest obstacles to structural studies. We used the fluorescence of ThT to follow the kinetics of alpha-Syn fibril formation at different ionic strengths to control fibril length and sample homogeneity for cryo-EM studies. The results show we can modulate secondary nucleation events involved in filament growth (i.e., the fragmentation rate) using sodium chloride concentrations below 1mM. Controlled proteinase-K digestion coupled with electron-spray ionization mass spectrometry of the fibril variants revealed distinctly different filament structures. Additionally, the power spectra of A30P and A53T filaments showed the 1/4.7 Å-1 layer line typical of cross-beta amyloids but the two have distinctly different helical organization. The A53T power spectrum revealed that the 1/4.7 Å-1 layer line is not a meridional but arises from a 1-start helix. This shows that the two strands in the filament are staggered, with a rise per subunit of 4.7 Å within each protofilament. We have confirmed this symmetry using the Iterative Helical Real Space Reconstruction (IHRSR) method to generate near-atomic resolution density maps of these amyloid filaments. Understanding the molecular mechanism in which ThT binds to amyloids represents a first step for structure-guided design of small molecules that may act as biomarkers for early PD detection.