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AbstractThe study provides new function tested on labetalol in large infusion volumes (Vinf= 80-115 µL) viaelectrospray ionization mass spectrometry, tandem MS2–MS7operation modes and collision energy: 0, 0.1, 10, 20, 25, 26, 30 and 35 eV, respectively. It is derived from the stochastic dynamic mass spectrometricequation D”SD= 2.6388.10-17.(-2), which exactly quantifies analyte concentration in solution. Also, it determines 3D conformations and electronic structures. The description of mass spectrometric intensity data as random variables and the shown relation, there are written two new linear functions among D”SDparameters, average total intensity values of fragmentation peaks, infusion volume, and collision energy. They introduce covarianceof datasets of variables pershort span of scan time of ions in any experimental conditions “l” and “m”. They are: D”SD;m,l= D”SD,l+ D”SD,m+ 5.2776.10-17.{–.}≈+ and +≈|rl,m|.sd(yEr±)1.sd(yEr±)m. Those relations yields to new formula D”SD,l+ D”SD,m= |rl,m|.sd(yEr±)l.sd(yEr±)m, providing exact function of mass spectrometric variable intensityof any peaks of analyte ion in any two sets of experimental conditions of measurements and diffusion parameters according to the first formula. Correlation between theory and experiment of fragmentation processes of labetalol shows |r|=1–0.99999. Chemometricsis used.

AbstractThe study reports quantification of metronidazole (MTZ) in clinical human urineemploying our innovative stochastic dynamic equation D”SD=2.6388.10-17.(–). There are direct analysis without presence of internal standard, using ultra-high accuracy nano-electrospray ionization (ESI) mass spectrometry (MS), quantum chemistry and chemometrics, respectively. The linear calibration D”SD=f(conc.) equations are obtained for concentration range 2.5 to 25000 ng.(mL)-1of spiked urine samples, examining analyte MS ions at m/z 171.0998, 172.0718, 172.04081, 213.1463, 181.0722and 151.1114, respectively. There is achieved exact coefficient of linear correlation (|r|=1) between theory and experiment of ion at m/z 172.04081. The major challenge to quantify MTZ in urine sample is that it stabilizes cations [M+H]+of two tautomers, which equilibrium varies drastically within the standard linear calibration dynamic range. The fragmentation patterns depend on analyte tautomers and it unambiguous assignment in multicomponent samples with complex sample matrix effect(SME) is unable to be made without mediation to an independent physico-chemical law. To solve the problem, herein, is used, again, above equation, which is designed a bridge between quantitative and multidimensional structural analytical chemistry, when is used complementary with Arrhenius’s quantum chemical diffusion parameter (DQC.) There is achieved |r|=0.98 of D”SD=f(DQC’) examining tautomeric molecular and fragment ions of MTZ.

The leaves of Dracaena steudneri yielded 6 new flavonoids–3,5,7-trihydroxy-6-methyl-3′,4′-methylenedioxyflavone (1), 5,7-dihydroxy-3-methoxy-6-methyl-3′,4′-methylenedioxyflavone (2), 3,5,7-trihydroxy-6-methoxy-3′,4′-methylenedioxyflavone (3), (2 S ,3 S)-3,7-dihydroxy-6-methoxy-3′,4′-methylenedioxyflavanone (4), 4′,5,7-trihydroxy-3,3′,8-trimethoxy-6-methylflavone (5), (2 R) 7-hydroxy-2′,8-dimethoxyflavanone (6)–together with 13 known congeners. Their structures were established using spectroscopic and spectrometric methods including NMR, CD, and HRMSn measurements. The compounds were evaluated for their anti-inflammatory potential through measurement of the levels of cytokines IL-1 β , IL-2, GM-CSF, and TNF- α in the supernatant of human peripheral blood mononuclear cells stimulated by lipopolysaccharide. Flavones derivatives 1 – 4 with a C-3′/4′ methylenedioxy substituent led to a substantial increase in the production of IL-1 β and GM-CSF out of 4 pro-inflammatory cytokines relative to LPS control. Quercetin derivatives 5, 11, and 13 with a hydroxyl group at C-4′ inhibited the production of IL-2, GM-CSF, and TNF- α. The presence of a C-2/C-3 double bond in 14 was pivotal to the significantly stronger (0.4 to 27.5% of LPS control) inhibitory effect compared to its dihydro derivative 8 (36.2 to 262.7% of LPS control) against all tested cytokines. It is important to note that the inhibitory activity of 14 was substantially higher than that of the standard drug used, ibuprofen. [ABSTRACT FROM AUTHOR]

AbstractIn the current paper, we further search for certainty of our innovative stochastic dynamic(SD) simplistic model formula D”SD= 2.6388.10-17. (-2) connecting among the so-called mass spectrometric SD diffusion parameter DSDparameter, the mass spectrometric measurable variable intensity,and the analyte concentration in solution. Since this equation has been derived more recently from our basic, but a more complex, formula for a large number of practical purposes of the analytical chemistrysuch a simplistic relationship is sufûcient significantly. Therefore, the motivation behind this work is further experimental justiûcation of the validity of this formula that is required in order to successfully apply the new model to the analytical practice. The quantification is carried out by studying reserpine (1) in solution at an analyte concentration of ng.(mL)-1. The capability of the model equation to express quantitatively the analyte concentration in solution is tested with independent chromatographic analysis. Correlation between chromatographic and mass spectrometric(MS) data is carried out by means of chemometrics. The diffusion parameters according to our stochastic dynamic concept and the simplistic model formula presented above and the corresponding diffusion data according to the current monitoring method(CMM) are correlated mutually, as well.

Melianthus majoris a medicinal plant endemic to South Africa. Its leaf extract led to the isolation of five new bufadienolides, 2β-acetoxy-3,5-di-O-acetylhellebrigenin (1), 2β-acetoxy-3-O-acetylhellebrigenin (2), 2β-acetoxy-14-deoxy-15β,16β-epoxymelianthugenin (4), 2β-acetoxy-14-deoxy-15β,16β-epoxymelianthusigenin (5), and 2β-hydroxymelianthusigenin (6), and four known analogues. The structures of the compounds were elucidated using NMR and HRESIMS data analyses. The relative configurations were defined by single-crystal X-ray crystallography and NOESY correlations. The isolated compounds exhibited strong cytotoxicity against MCF-7 breast cancer cells and sensitive CCRF-CEM and multidrug-resistant CEM/ADR5000 leukemia cells. Compound 1showed the most potent activity, with IC50values of 0.1 μM toward CCRF-CEM and CEM/ADR5000 and 0.3 μM toward MCF-7.

Phytochemical analysis of a methanol–dichloromethane (1:1) extract of the aerial parts of Tephrosialinearisled to the isolation of 18 compounds. Seven of these, namely, lineaflavones A–D (1–4), 6-methoxygeraldone (5), 8″-acetylobovatin (6), and 5-hydroxy-7-methoxysaniculamin A (7) are new compounds. The compounds were characterized based on their NMR and HRMSndata. The anti-inflammatory effects of the crude extract and isolated compounds were evaluated by measuring the levels of interleukins (IL-1β, IL-2, and IL-6), granulocyte-macrophage colony-stimulating factor (GM-CSF), and tumor necrosis factor-α (TNF-α) in lipopolysaccharide (LPS)-stimulated peripheral blood mononuclear cells (PBMCs). The crude extract inhibited the release of all cytokines except IL-1β, which slightly increased in comparison to the LPS control. All the tested compounds suppressed the production of IL-2, GM-CSF, and TNF-α. Whereas compounds 1, 2, 4–8, 10–15, 17, and 18decreased production of IL-6, compounds 1, 2, 4, 7, 10, 13–15, and 17inhibited the release of IL-1β. It is worth noting that most of the compounds tested showed a superior reduction in cytokines release compared to the reference drug ibuprofen.

This paper details with design, synthesis, isolation, spectroscopic and structural elucidation of seven crystals of organic barbiturates (a red phase of bis(5-(hydroxyimino)pyrimidine-2,4,6(1H,3H,5H)-trione) 4H-1,2,4-triazol-4-amine co-crystal (1y), 2-methylamino-1-phenyl-propan-1-ol 2-thioxo-dihydro-pyrimidine-4,6-dione (2), 4-phenylpyridine thiobarbituric acid co-crystal (3), new monoclinic polymorph of violuric acid monohydrate (4), 2-pyrrolidin-1-yl-ethylammonium bis(violurate) dihydrate (5)and redetermination of barbituric acid dihydrate (6)and noncentrosymmetric orthorhombic violuric acid monohydrate (7)). The noncentrosymmetric crystals (1r)(2)and (7)can efficiently be employed in the field of nonlinear optical (NLO) materials research. We aim at studying relationship between molecular, respectively, crystal structure and NLO properties within a relatively large range of temperatures (T= 273–500 K). The single crystal X-ray diffraction, electronic absorption, conventional and linear polarized infrared spectroscopies have been used to provide experimental structural and spectroscopic data. The deuterated derivatives of (4)and (7)have been analysed, as well. The theoretical description of linear and NLO properties of crystals (1)–(7)is based on quantum chemical ab initio methods and density functional theory (DFT). The work contributes, importantly, to the fields of nonlinear optics, crystal engineering or materials research, reflecting in mind frequency conversion of lasers, electro-optics, modulators, waveguides and more.

Matrix-assisted laser desorption/ionization imaging high-resolution mass spectrometry (MALDI-imaging-HRMS) is an important technique for visualizing the spatial distribution of compounds directly on the surface of organisms such as microorganisms, insects, plants, animals, and human tissues. However, MALDI-imaging-HRMS and the stable isotope labeling approach have never been combined for the detection and simultaneous visualization of labeled and unlabeled compounds, their analogues and derivatives, as well as their precursors. Herein, we present a methodology that labels microbial secondary metabolites directly on agar with stable isotopes and allows concurrent spatial distribution analyses by MALDI-imaging-HRMS. Using a thin film of labeled agar supplemented with [1-13C]-l-proline, [methyl-D3]-l-methionine, 15NH4Cl, or [15N]-l-serine overlaid on unlabeled agar, we demonstrate the incorporation of labeled precursors into prodiginines and serratamolides produced by an endophytic bacterium, Serratia marcescens, by MALDI-imaging-HRMS and HPLC–HRMS. Further, we show the incorporation of CD3into prodigiosin as well as its characteristic fragments directly by MALDI-imaging-HRMS2. Our methodology has several advantages over currently existing techniques. First, both labeled and unlabeled compounds can be visualized simultaneously in high spatial resolution along with their labeled and unlabeled precursors. Second, by using a thin film of labeled agar, we utilize minimum amounts of expensive labeled compounds (1–3 mg) ensuring a cost-effective method for investigating biosynthetic pathways. Finally, our method allows in situvisualization and identification of target and nontarget compounds without the need of isolating the compounds. This is important for compounds that are produced by microorganisms in low, physiologically, or ecologically relevant concentrations.

An endophytic fungus, Eupenicilliumsp. LG41, isolated from the Chinese medicinal plant Xanthium sibiricum, was subjected to epigenetic modulation using an NAD+-dependent histone deacetylase (HDAC) inhibitor, nicotinamide. Epigenetic stimulation of the endophyte led to enhanced production of two new decalin-containing compounds, eupenicinicols C and D (3and 4), along with two biosynthetically related known compounds, eujavanicol A (1) and eupenicinicol A (2). The structures and stereochemistry of the new compounds were elucidated by extensive spectroscopic analysis using LC-HRMS, NMR, optical rotation, and ECD calculations, as well as single-crystal X-ray diffraction. Compounds 3and 4exist in chemical equilibrium with two and three cis/transisomers, respectively, as revealed by LC-MS analysis. Compound 4was active against Staphylococcus aureuswith an MIC of 0.1 μg/mL and demonstrated marked cytotoxicity against the human acute monocytic leukemia cell line (THP-1). We have shown that the HDAC inhibitor, nicotinamide, enhanced the production of compounds 3and 4by endophytic Eupenicilliumsp. LG41, facilitating their isolation, structure elucidation, and evaluation of their biological activities.

Three new compounds, colletotrichones A–C (1–3), and one known compound, chermesinone B (4a), were isolated from an endophytic fungus, Colletotrichumsp. BS4, harbored in the leaves of Buxus sinica, a well-known boxwood plant used in traditional Chinese medicine (TCM). Their structures were determined by extensive spectroscopic analyses including 1D and 2D NMR, HRMS, ECD spectra, UV, and IR, as well as single-crystal X-ray diffraction, and shown to be azaphilones sharing a 3,6a-dimethyl-9-(2-methylbutanoyl)-9H-furo[2,3-h]isochromene-6,8-dione scaffold. Owing to the remarkable antibacterial potency of known azaphilones coupled to the usage of the host plant in TCM, we evaluated the antibacterial efficacy of the isolated compounds against two commonly dispersed environmental strains of Escherichia coliand Bacillus subtilis, as well as against two human pathogenic clinical strains of Staphylococcus aureusand Pseudomonas aeruginosa. Compound 1exhibited marked antibacterial potencies against the environmental strains that were comparable to the standard antibiotics. Compound 3was also active against E. coli. Finally, compound 2aexhibited the same efficacy as streptomycin against the clinically relevant bacterium S. aureus. The in vitrocytotoxicity of these compounds on a human acute monocytic leukemia cell line (THP-1) was also assessed. Our results provide a scientific rationale for further investigations into endophyte-mediated host chemical defense against specialist and generalist pathogens.

A new dimeric naphtho-γ-pyrone, 2-hydroxydihydronigerone (1), along with five compounds, nigerone (2), pyrophen (3), kojic acid (4), 4-(hydroxymethyl)-5-hydroxy-2H-pyran-2-one (5), and p-hydroxyphenylacetic acid (6), was isolated from an endophytic fungus Aspergillus nigerAKRN associated with the roots of Entandrophragma congoënse. The structure of the new compound has been elucidated using spectroscopic data including 1D and 2D NMR as well as the high-resolution mass spectrometry. Compounds 1–5showed weak antimicrobial activity on five selected Gram-negative bacteria, namely Enterobacter aerogenes(CM64), Enterobacter cloacae(BM67), Klebsiella pneumonia(K2), and Escherichia coli(ATCC8739 and ATCC10536).

A new lanostanoid, 19-nor-lanosta-5(10),6,8,24-tetraene-1α,3β,12β,22S-tetraol (1), characterized by the presence of an aromatic B ring and hydroxylated at C-1, C-3, C-12, and C-22, was isolated from an endophytic fungus, Diaporthesp. LG23, inhabiting leaves of the Chinese medicinal plant Mahonia fortunei. Six biosynthetically related known steroids were also isolated in parallel. Their structures were confirmed on the basis of detailed spectroscopic analysis in conjunction with the published data. Compound 1, an unusual fungus-derived 19-nor-lanostane tetracyclic triterpenoid with an aromatic B-ring system, exhibited pronounced antibacterial efficacy against both Gram-positive and -negative bacteria, especially the clinical isolates of Streptococcus pyogenesand Pseudomonas aeruginosaas well as a human pathogenic strain of Staphylococcus aureus. Our results reveal the potential of endophytes not only in conferring host fitness but also in contributing toward traditional host plant medicines.

Eight new triterpenoids, prototiamins A–G (1–6, 9) and seco-tiaminic acid A (10), were isolated along with four known compounds from the bark of Entandrophragma congoënse. Their structures were elucidated by means of HRMS and different NMR techniques and chemical transformations. Assignments of relative and absolute configurations for the new compounds were achieved using NOESY experiments and by chemical modification including the advanced Mosher’s method. Additionally, the structure and relative configuration of compound 3were confirmed by single-crystal X-ray diffraction analysis. Compounds 1, 3, and 5displayed significant in vitro antiplasmodial activity against the erythrocytic stages of chloroquine-sensitive Plasmodium falciparumstrain NF54. Prototiamin C (3) was the most potent of the compounds isolated, with an IC50value of 0.44 μM. All compounds tested showed low cytotoxicity for the L6 rat skeletal myoblast cell line.

Several recent studies have lent evidence to the fact that certain so-called plant metabolites are actually biosynthesized by associated microorganisms. In this work, we show that the original source organism(s) responsible for the biosynthesis of the important anticancer and cytotoxic compound maytansine is the endophytic bacterial community harbored specifically within the roots of Putterlickia verrucosaand P. retrospinosaplants. Evaluation of the root endophytic community by chemical characterization of their fermentation products using HPLC-HRMSn, along with a selective microbiological assay using the maytansine-sensitive type strain Hamigera avellanearevealed the endophytic production of maytansine. This was further confirmed by the presence of AHBA synthase genes in the root endophytic communities. Finally, MALDI-imaging-HRMS was used to demonstrate that maytansine produced by the endophytes is typically accumulated mainly in the root cortex of both plants. Our study, thus, reveals that maytansine is actually a biosynthetic product of root-associated endophytic microorganisms. The knowledge gained from this study provides fundamental insights on the biosynthesis of so-called plant metabolites by endophytes residing in distinct ecological niches.

Despite of the fact that the manufacturing of beer dated from many centuries, and nevertheless, of enormous analytical instrumental elaborations, developments and implementations for highly precise and accurate protocols for determination of foodstuffs, the quantitation of beer ingredients still represent significant challenge. It is caused by: (A) high complexity of the beer as foodstuffs matrix with significant proteins and maltooligosaccharide content as well as diversity of more than 57 low-molecular weight hop bitter acidic analytes and their oxidation products; (B) higher instability and variety of chemical oxidation processes typical for these ingredients; (C) easy isometrization and rearrangements, tautomerism and large number of different conformations of hop bitter acids; (D) structural similarity of the analytes; (E) photo-instability of the ingredients; and (F) still unavailable commercial standards for all hop acids, which difficult the analysis of major and minor components in crude hop extracts, during brewing process and storage of final food product. The diverse number of chemical and photochemical processes may be conducted at each these stages of beer manufacturing, which caused need of methodological elaborations and implementations of accurate analytical protocols for on-line analysis of beer components. For this reason the paper content encompasses quantitative solid-state analysis of hop bitter acids in different brands beer beverages employing UV–MALDI–Orbitrap MS method. Despite of its powerful instrumental capability, this mass spectrometric method has been scarce utilized in analysing of beer foodstuffs, limiting the scientific efforts and reports to mainly qualitative analysis and fingerprinting of proteins and maltooligosaccharides. In this respect the study provided novelty in the field of the quantitative analytical chemistry of low molecular weight analytes in beer demonstrating its capability for accurate determination of closely structural ingredients such as the large number of naturally occurring hop bitter acids.

Two new compounds containing the decalin moiety, eupenicinicols A and B (1and 2), two new sirenin derivatives, eupenicisirenins A and B (3and 4), and four known compounds, (2S)-butylitaconic acid (5), (2S)-hexylitaconic acid (6), xanthomegnin (7), and viridicatumtoxin (8), were isolated from an endophytic fungus, Eupenicilliumsp. LG41, harbored in the roots of the Chinese medicinal plant Xanthium sibiricum. Their structures were confirmed through combined spectroscopic analysis (NMR and HRMSn), and their absolute configurations were deduced by ECD calculations or optical rotation data. Since the endophytic fungus was isolated from the roots, the antibacterial efficacies of the compounds 1–6were investigated against Bacillus subtilisand Acinetobactersp. BD4, which typically inhabit soil, as well as the clinically important Staphylococcus aureusand Escherichia coli. (2S)-Butylitaconic acid (5) and (2S)-hexylitaconic acid (6) exhibited pronounced efficacy against Acinetobactersp., corroborating the notion that root-endophytes provide chemical defense to the host plants. Compound 2was highly active against the clinically relevant S. aureus. By comparing 1with 2, it was revealed that altering the substitution at C-11 could drastically increase the antibacterial efficacy of 1. Our study reveals plausible ecological roles of the endophyte and its potential pharmaceutical use as a source of antibacterial compounds.

A new spiroalkaloid, 7-epi-javaniside (1), was isolated from the Cameroonian plant Pauridiantha callicarpoides, along with seven known compounds viz., javaniside (2), sweroside (3), hymexelsin (4), scopoletin (5), 7,7`-dihydroxy-6,6`-dimethoxy-8,8`-biscoumarin (6), 7,7`-dihydroxy-6,6`- dimethoxy-3,3`-biscoumarin (7), and 4-hydroxy-3-methoxybenzoic acid (8). The structures of the new alkaloid, as well as those of the known compounds, were elucidated by detailed spectroscopic analysis including 1D and 2D NMR and by comparison with the literature data for related compounds.

Quantitative UV-MALDI-Orbitrap mass spectrometric study of naturally occurring insecticides, acaricides and, piscicides rotenone (1), rotenolone (2), sumatrol (3), deguelin (4) as well as tephrosin (5) in rapeseed oil is presented. Direct emulsion assay and solid-state sample preparation techniques in presence of matrixes of organic crystals of ions are utilized. The protocol by APCI-MS and HPLC–ESI–MS/MS methods is verified. The method performance parameters along the matrix effects is evaluated by a comparison of analytical metrology achieved for rotenoids 1–5by each of mass spectrometric methods. Advantages of UV-MALDI-Orbitrap-MS method for quantitation are emphasized. They are compared to those of core analytical hybrid methods such as GC–MS, and HPLC–ESI (or APCI)-MS/MS for lipidomics and determination of organic pollutants in oils. The statistical simple random sampling plan is utilized for analysis of non-polluted rapeseed oils sample sets. The determination of polluted sample set and validation by spiked rapeseed oils is performed.

Fruiting bodies of Mycena metatawere screened for the presence of new secondary metabolites by means of HPLC-UV, LC-HR-ESIMS, and high-resolution matrix-assisted laser desorption/ionization mass spectrometry imaging (HR-MALDI-MS imaging). Thus, a new β-carboline alkaloid, 6-hydroxymetatacarboline D (1d), was isolated from fruiting bodies of M. metata. 6-Hydroxymetatacarboline D consists of a highly substituted β-carboline skeleton, which is likely to be derived biosynthetically from l-tryptophan, 2-oxoglutaric acid, l-threonine, and l-proline. The structure of the alkaloid was established by 2D NMR spectroscopic methods and HR-ESIMS. Moreover, by extensive application of LC-HR-ESIMS, LC-HR-ESIMS/MS, and LC-HR-ESIMS3techniques we were able to elucidate the structures of a number of accompanying β-carboline alkaloids, 1a–1c, 1e–1i, and 2a–2g, structurally closely related to 6-hydroxymetatacarboline D, which are present in M. metatain minor amounts. The absolute configuration of the stereogenic centers of the β-carboline alkaloids was determined by GC-MS comparison with authentic synthetic samples after hydrolytic cleavage and derivatization of the resulting amino acids.

Four new beilschmiedic acid derivatives, cryptobeilic acids A–D (1–4), and tsangibeilin B (5) have been isolated from the bark of Beilschmiedia cryptocaryoidescollected from Madagascar. Their structures were elucidated using detailed spectroscopic and spectrometric methods. Cryptobeilic acid A (1) and tsangibeilin B (5) structures were confirmed by single-crystal X-ray diffraction analysis. Compounds 1–5displayed moderate antibacterial activity against Escherichia coli6r3, Acinetobacter calcoaceticusDSM 586, and Pseudonamas stutzeriA1501, with the minimum inhibitory concentrations ranging from 10 to 50 μM, respectively. In addition, the compounds exhibited antiplasmodial activity against erythrocytic stages of chloroquine-resistant Plasmodium falciparumstrain NF54 and weak cytotoxicity against L6 cell lines.

The paper reports organosilver catalyzed C–N coupling reactions of pyridyl (py) carbaldehydes and substituted anilines, thereby producing substantially different phenazines (PZs) with basic molecular skeletal and bulk Npy-substituents. The different chemical reactivity of Npy-substituted derivatives in polar protic solvents, in the presence of the AgI-salts, with the dinuclear silver(i/ii) organometallic precursors exhibiting C–AgI/II–Cl covalently bonded species, is described. The latter are elucidated fully by the theoretical quantum chemical method and experimentally evidenced by mass spectrometry in the gas phase (GP), involving a variety of ionization techniques. To our knowledge, we first report a comprehensive study of the nature of the organometallic precursors, which represents an important step of the catalyzed C–C and C–N coupling processes. Thus impacting all fields the enormous synthetic research efforts are focused on PZs, which through the diversity of the molecular architectures are main components observed in naturally occurring products (NPs), dyes, pesticides, antibiotics, etc. The emphasis of the paper is on the mass spectrometry (MS), employing its high-resolution power and accuracy of the analytical qualitative, quantitative and structural information as well as the physico-chemical thermodynamic one, at the analyte concentrations in the range of fg g−1. The latter, which are beyond the capability of other instrumental methods for structural analysis, make the MS an irreplaceable method in the advanced catalytical research. Its combined application with the method of the quantum chemistry provides reliable thermodynamic information in the condensed phase and in the GP, which helps understand comprehensively the mechanisms of the organometallic catalyzing processes, particularly the unique coupling reactions, thus improving significantly their effectiveness.

The current discussion about the relevance of guttation as a water source for honey bees (Apis melliferaL.) draws attention to the activity of water foraging bees. However, due to the wide foraging range, field observations of water foraging bees are difficult to carry out. Therefore, samples of returning bees were collected in front of the entrance early in the morning as well as during the day in summer and autumn 2010. In the laboratory, honey sacs of bees without pollen or propolis were examined. Honey sac samples with a weight of =0.005 g were classified as successful foragers. For the determination of water or nectar foragers, the honey sac contents of successful foragers were analyzed with a refractometer. Due to the dilution and the precision of the refractometer, an accuracy of 5 % was possible. Based on the known range of sugar concentrations in nectars, the analyzed samples were divided into nectar foragers (25–65 % sugar) and water foragers (0–15 % sugar). The study reveals that nectar and water foraging bees can be discriminated by analyzing the honey sac extracts with a refractometer. In autumn, the water demand of honey bee colonies is almost exclusively covered by water collection, whereas during summer nectar is mainly used as a substitute for water. In general, water collection is not restricted to certain times of the day.

Despite enormous instrumental mass spectrometric (MS) developments, providing sufficient advantages towards highly selective and sensitive analytical protocols, including those related to different ionization techniques, the presented paper aimed to study the ionization UV-MALDI-MS process and to describe the chemical aspects and essential role of the types of the matrixes. Organic salts appeared promising as matrixes for analysis of natural products (NPs), including the imaging techniques, versusthe co-crystals of neutral polyfunctional carboxylic acids. Seven model crystals of organic salts, and their matrix–analyte polycrystals, were comprehensively studied using the MALDI-MS and ESI-MS methods. The MS data were compared and supported by theoretical and experimental data, using quantum chemical methods, optical spectroscopy, and single crystal X-ray diffraction. The role of proton transfer (PT) effects in the desorption/ionization UV-MALDI processes was clarified.

The paper reported a matrix-assisted laser desorption/ionization (MALDI) mass spectrometric (MS) protocol for determination of herbicides, improving the analytical sample preparation step, making a significant contribution to the reproducibility and precision of the method. Matrix-analyte sampling is performed, employing low molecular weight (LMW) dication-containing organic crystals. Considering environmental control applications, six of the best selling herbicides, such as aclonifen (1), bifenox (2), diclofop-methyl (3), racemic mixture of diclofop (4), alachlor (5) and metolachlor (6) as well as their mixtures are quantified successively through the replacement of the polyfunctional benzoic acids with dication-containing organic matrices. The application of classical matrices for UV-MALDI-MS for the analysis of 1–6caused their competitive chemical transformation, thus leading to spurious results and inaccurate analytical information, due to the wide variety of experimental conditions depending on the chemical, physico-chemical and physical nature of the herbicide analytes. The full method and technique validation, based on UV-MALDI-MS employing an Orbitrap analyser, are presented, achieving to our knowledge promising measurements for LMW analytes. These latter are judged by a fully validated analytical protocol based on positive electrospray ionization (ESI) mass spectrometry in a single and tandem (MS/MS) mode of operation. The chemical modifications, polymorph transformations of the UV-MALDI-MS crystal matrices and/or matrix-analyte samples, as well as ion-process formation upon solution → crystalline state and crystalline state → gas-phase transitions is controlled by joint application of unique and robust instrumental analytical methods for absolute structural determination, such as single crystal X-ray diffraction and mass spectrometry. As direct experimental qualitative, quantitative and structural evidence regarding the analytes was provided, the reported protocol is a fully validated one at each of the analytical steps. The additional quantum chemical data allowed correlation of the thermodynamic stability and MS abundances of observed ionic species in the gas-phase as well as the structure of crystals. Since environmental analysis involves a broad variety of analytes, conditions and degree of matrix complexity, only an individual certification method development may achieve meaningful analytical information. In this respect, the reported UV-MALDI-MS method for herbicides significantly impacts the field of environmental analytical chemistry, quality assurance and control; moreover the analytical protocol is constructed to deal with a real environmental problem. Therefore the reported method ensures real practical efficiency and reliability in the results for the human health safety, environmental monitoring and control programs.

Fungal endophytes inhabit healthy tissues of all terrestrial plant taxa studied and occasionally produce host-specific compounds. We recently isolated an endophytic fungus, Fusarium solani, from Camptotheca acuminata, capable of biosynthesizing camptothecin (CPT, 1), but this capability substantially decreased on repeated subculturing. The endophyte with an impaired 1biosynthetic capability was artificially inoculated into the living host plants and then recovered after colonization. Although the host−endophyte interaction could be reconstituted, biosynthesis of 1could not be restored. Using a homology-based approach and high-precision isotope-ratio mass spectrometry (HP-IRMS), a cross-species biosynthetic pathway is proposed where the endophyte utilizes indigenous G10H(geraniol 10-hydroxylase), SLS(secologanin synthase), and TDC(tryptophan decarboxylase) to biosynthesize precursors of 1. However, the endophyte requires host STR(strictosidine synthase) in order to condense the nitrogen-containing moiety (tryptamine, 2) with the carbon-containing moiety (secologanin, 3) to form strictosidine (4) and complete the biosynthesis of 1. Biosynthetic genes of 1in the seventh subculture generation of the endophyte revealed random and unpredictable nonsynonymous mutations. These random base substitutions led to dysfunction at the amino acid level. The controls, Top1gene and rDNA, remained intact over subculturing, revealing that instability of biosynthetic genes of 1was not reflected in the primary metabolic processes and functioning of the housekeeping genes. The present results reveal the causes of decreased production of 1on subculturing, which could not be reversed by host−endophyte reassociation.

Two new isopropenyl-dihydrofuranoisoflavones exhibiting antimicrobial properties have been isolated along with eight known compounds from the Cameroonian medicinal plant Crotalaria lachnophora. The structures of the new compounds were elucidated by 1D and 2D NMR spectroscopy and high-resolution mass spectrometry as 7,2′,4′-trihydroxy-5′′-isopropenyl-4′′,5′′-dihydrofurano[2′′,3′′:5,6]isoflavone (1) and 4,8-dihydroxy-2-isopropenyl-2,3-dihydro-5H-[1]benzofuro[2,3-b]furo[3,2-g]chromen-5-one (2). The CH2Cl2/MeOH (1:1) extract and the compounds isolated were subjected to in vitro antimicrobial assays against a panel of pathogenic microorganisms, including Gram-positive and Gram-negative bacteria and fungi. The new compounds, named lachnoisoflavones A (1) and B (2), showed moderate inhibitory activities against Escherichia coliand Klebsiella pneumoniae.

Exophialin, a previously unknown indole alkaloid, was isolated from the humanpathogenic fungus Exophiala dermatitidisupon cultivation on a medium containing tryptophan as sole nitrogen source. Its structure has been elucidated by 1D and 2D NMR experiments and mass spectrometry. When the mutant strain Mel1 was used in place of the wildtype strain, the new indole alkaloid 8hydroxyexophialin was generated instead of exophialin. This result indicates that the aromatic moiety of exophialin originates from 2hydroxyjuglone, a shunt product of the 1,8dihydroxynaphthalene melanin biosynthesis, which is only produced by the wildtype strain,but not by the mutant. In contrast, the mutant accumulatesflavioline, which acts as precursor leading to the generation of 8hydroxyexophialin instead of exophialin.

Soil, the “Earths thin skin” serves as the delicate interface between the biosphere, hydrosphere, atmosphere, and lithosphere. It is a dynamic and hierarchically organized system of various organic and inorganic constituents and organisms, the spatial structure of which defines a large, complex, and heterogeneous interface. Biogeochemical processes at soil interfaces are fundamental for the overall soil development, and they are the primary driving force for key ecosystem functions such as plant productivity and water quality. Ultimately, these processes control the fate and transport of contaminants and nutrients into the vadose zone and as such their biogeochemical cycling. The definite objective in biogeochemicalinterface research is to gain a mechanistic understanding of the architecture of these biogeochemical interfaces in soils and of the complex interplay and interdependencies of the physical, chemical, and biological processes acting at and within these dynamic interfaces in soil. The major challenges are 1 to identify the factors controlling the architecture of biogeochemical interfaces, 2 to link the processes operative at the individual molecular andor organism scale to the phenomena active at the aggregate scale in a mechanistic way, and 3 to explain the behavior of organic chemicals in soil within a general mechanistic framework. To put this in action, integration of soil physical, chemical, and biological disciplines is mandatory. Indispensably, it requires the adaption and development of characterization and probing techniques adapted from the neighboring fields of molecular biology, analytical and computational chemistry as well as materials and nanosciences. To shape this field of fundamental soil research, the German Research Foundation DFG has granted the Priority Program “Biogeochemical Interfaces in Soil”, in which 22 individual research projects are involved.

The possible microbial mechanism of hypericin (1) and emodin (2) biosynthesis was studied in axenic submerged culture conditions in the endophytic fungus Thielavia subthermophila, isolated from Hypericum perforatum. The growth and secondary metabolite production of the endophyte remained independent of the illumination conditions. This production remained unaltered on spiking the medium with 3 or 5 mM 2, although the biomass accumulation was reduced. Neither emodin anthrone (3) nor protohypericin (4) could be detected at any stage of fermentation, irrespective of either spiking or illumination conditions. The endophytic metabolites exhibited photodynamic cytotoxicity against the human acute monocytic leukemia cell line (THP-1), at 92.7 vs 4.9%, and 91.1 vs 1.0% viability by resazurin and ATPlite assays, in light and in the dark, respectively. In trying to ascertain the presence/expression of the candidate hyp-1gene in the endophyte, it was revealed that the hyp-1gene was absent in T. subthermophila, indicating that the biosynthetic pathway in the endophytic fungus might be different and/or governed by a different molecular mechanism than the host plant or host cell suspension cultures. We have discussed the biosynthetic principles and evolutionary implications relating to endophytic T. subthermophilabased on the results obtained.

In the present study, we report the application of LC‐MS based on two different LC‐MS systems to mycotoxin analysis. The mycotoxins were extracted with an ACN/water/acetic acid mixture and directly injected into a LC‐MS/MS system without any dilution procedure. First, a sensitive and reliable HPLC‐ESI‐MS/MS method using selected reaction monitoring on a triple quadrupole mass spectrometer (TSQ Quantum Ultra AM) has been developed for determining 32 mycotoxins in crude extracts of wheat and maize. This method was operated both in positive and in negative ionization modes in two separate chromatographic runs. The method was validated by studies of spiked recoveries, linearity, matrix effect, intra‐assay precision and sensitivity. Further, we have developed and evaluated a method based on accurate mass measurements of extracted target ions in full scan mode using micro‐LC‐LTQ‐Orbitrap as a tool for fast quantitative analysis. Both instruments exhibited very high sensitivity and repeatability in positive ionization mode. Coupling of micro‐LC to Orbitrap technology was not applicable to the negatively ionizable compounds. The LC triple quadrupole MS method has proved to be stable in quantitation, as it is with respect to the matrix effects of grain samples.

Enantiomers of 41,1,2trimethylhexylphenol, a chiral isomer of the endocrine disrupting chemical nonylphenol, have been resolved and isolated by preparative chiral HPLC. The absolute configurations of the enantiomers were then determined by an Xray crystallographic study of the −camphanoyl derivative of the first eluted enantiomer NP35E1. The first enantiomer NP35E1 and the second enantiomer NP35E2 eluted were found to have the Sand Rabsolute configurations, respectively. The estrogenic potencies of the Sand Renantiomers were tested by the Escreen assay. A slight difference was observed in the relative proliferative effect between the Senantiomer and Renantiomer in the Escreen assay. Chirality, 2009. © 2008 WileyLiss, Inc.

The pentacyclic quinoline alkaloid camptothecin (1) is a potent antineoplastic agent. Two of its analogues, 9-methoxycamptothecin (2) and 10-hydroxycamptothecin (3), exhibit similar potency but do not have the potential therapeutic drawbacks produced by unmodified 1. We have established methodology for the isolation and unequivocal identification and characterization of a novel endophytic fungus isolated from the inner bark of the medicinal plant Camptotheca acuminata, which produced 1−3in rich mycological medium (Sabouraud dextrose broth), under shake-flask fermentation conditions. The fungus was identified by its morphology and authenticated by ITS analysis (ITS1 and ITS2 regions and the intervening 5.8S rDNA region). Camptothecin (1) and its analogues were identified by 1H NMR spectroscopy and LC-HRMS and confirmed by comparison with authentic standards. The production pattern of the metabolites over seven successive subculture generations of this endophyte was studied. A sharp attenuation in the production of 1and 2was observed from the first- through to the seventh-generation subculture. Therefore, these results offer a caution as to the possibility of using endophytic fungi as alternate sources of plant secondary metabolite production. Further studies have been initiated on the analysis of the upstream metabolic intermediates to understand the steps at which the production of the metabolites in question is constrained.

Crystal structure and spectroscopic properties of 4-acetaminopyridine and its protonated form4-Acetaminopyridine dihydrate and its protonated form, stabilized as the hydrochloride salt have been synthesized and spectroscopic elucidated in solution and in the solid-state by means of the inear-polarized solid state IR-spectroscopy (IR-LD), UV-spectroscopy, TGA, DSC, and the positive and negative ESI MS. Quantum chemical calculations were used to obtain the electronic structure, vibrational data and the electronic spectra. The spectroscopic and theoretical data are compared with the structure of the first compound obtained by single crystal X-ray diffraction. The effect of Npyprotonation on the optical and magnetic properties of a 4-acetaminopyridine is discussed.

For the first time, an endophytic fungus has been isolated from the stems of the medicinal herb Hypericum perforatum(St. John’s Wort). The fungus produced the napthodianthrone derivative hypericin ( 1) in rich mycological medium (potato dextrose broth) under shake flask and bench scale fermentation conditions. Emodin ( 2) was also produced simultaneously by the fungus under the same culture conditions. We propose 2as the main precursor in the microbial metabolic pathway to 1. The fungus was identified by morphology and authenticated by 28S (LSU) rDNA sequencing. Compounds 1and 2were identified by LC-HRMS, LC-MS/MS, and LC-HRMS/MS and confirmed by comparison with authentic standards. In bioassays with a panel of laboratory standard pathogenic control strains, including fungi and bacteria, both fungal 1and 2possessed antimicrobial activity comparable to authentic standards. This endophytic fungus has significant scientific and industrial potential to meet the pharmaceutical demands for 1in a cost-effective, easily accessible, and reproducible way.

A multimethod based on liquid–liquid extraction and solid–liquid extraction for the analysis of persistent organic pollutants in water and sludge from sewage treatment plants has been established. Traces of 22 organic compounds used in industry and personal care products (PCPs) were analyzed by GC/MS. The LODs for the analytes were less than 2.3 ng/L for wastewater and 31 μg/kg (dry weight matter) for sewage sludge. Satisfactory recoveries (70–130%) were achieved. The validated method permits the analysis of water and sludge samples at various stages of the treatment from different sewage treatment plants. Thus, the distribution between water and sludge as well as the dissipation of the compounds analyzed were balanced. By this means, the efficiency of different wastewater treatment plants (WWTPs) can be evaluated and measures can be taken to optimize the treatment process at different stages.

The objective of this study was to identify the effect of different land uses in peri‐urban agriculture on the soil properties. Soil health indicators were evaluated in the top 10 cm at five tilled agricultural sites involving different cropping systems and use of agrochemicals within the peri‐urban agricultural areas of Yaounde, Cameroon, and compared with a native forest land. The experimental data showed that the selected indicators were sensitive to cropping practice. Most cropped land had significantly higher total C, available N and P concentrations, soil pH, electrical conductivity, salinity, biomass C and P, dehydrogenase, β‐glucosidase, and acid phosphatase activities. Land producing corn (Zea maysL.) and sugarcane (Saccharum officinarumL.) differed from that producing tomatoes (Lycopersicon esculentumMill.), but cultivation of these crops has significantly impacted native soil quality. However, phenol oxidase, microbal biomass C/organic C (Cmic/Corg), and microbial biomass C/microbial biomass P (Cmic/Pmic) were negatively affected. These appeared to be more consistent indicators of negative management causing changes to soil health and may be suitable for an early appraisal of soil health.

Abstract: Valinamide ester amide of squaric acid has been synthesized and structurally characterized by single crystal X-ray diffraction. Individual molecules are linked into zig-zag helix chains running parallel to [0 1 0] by N–H ⋯ O hydrogen bonds of length 2.7908(19) Å and 2.980(2) Å involving the valinamide nitrogen atoms and the amide carbonyl oxygen atom. An experimental IR spectroscopic characterization and theoretical vibrational analysis have also been performed, using the possibilities of polarized linear-dichroic IR-spectroscopy (IR-LD), based on an orientation technique of the molecules as a suspension in a nematic liquid crystal.

The aqueous-ethanolic extract (AEE) of high altitude Podophyllum hexandrum has earlier been reported to render a radioprotective effect against lethal gamma radiation in in vitro model. AEE has also been reported to possess metal chelating and DNA protecting properties. The present study was undertaken to isolate and characterize the bioactive principle present in AEE and investigate its role in radiation protection. A novel molecule was found to be present in AEE and was assigned as 3-O-ß-?-galactoside of quercetin by acid hydrolysis, LC-MS, LC-APCI-MS/MS and 13C NMR spectra. Various biological activities were investigated at in vitro level. The antioxidant potential of AEE in lipid and aqueous phase was determined against numerous stresses. AEE was found to be significantly (p < 0.05) protective, i.e., against Fe2+and Cu2+-induced linoleic acid degradation, respectively. Radiationinduced lipid oxidation studies revealed that AEE maximally works at a [lignan]/0.25 kGy ratio 400 (ratio of concentration of AEE divided by the radiation dose, i.e., 0.25 kGy) and no drug-induced lipid oxidation at all concentrations tested was found. In a time-dependent study, total antioxidant activity was maximally exhibited at 1 mg/ml. The site-specific and non-site-specific deoxyribose degradation assay exhibited a dose-dependant hydroxyl scavenging potential of AEE (0.05-500 µg/ml). The anti-lipid peroxidation ability of AEE against radiation (0.25 kGy)-induced lipid peroxidation was higher in case of neural tissue homogenate as compared to kidney homogenate [activity ratio: 0.039 (brain) < 0.24 (kidney)]. The protein protection study using bovine serum albumin was also done for two time intervals (2 h and 4 h) and significant (p < 0.05) protection was observed at 500 µg/ml (> 97%). This study implies that 3-O-ß-?-galactoside present in AEE renders radioprotection by protecting lipids, proteins in renal and neural model system against supra-lethal (0.25 kGy) gamma radiation.

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Abstract. The objective of the study was to compare ecotoxicological data obtained from laboratory experiments on the side-effects of three phenylamide fungicides, pure metalaxyl (racemic mixture of R- and S-enantiomers), formulated metalaxyl and mefenoxam (only active R-enantiomer) on the chemical and biochemical parameters of two soils of different type and origin. The purpose of the comparison was to determine to what extent mefenoxam, developed as alternative to metalaxyl, can affect the activity of soil micro-organisms and their processes, and to elucidate the differences between the effects of pure and formulated metalaxyl. The dynamics of the quantitative changes in biochemical parameters induced by the addition of these fungicides at their recommended field rate were determined in a sandy clay soil from Cameroon and a sandy loam soil from Germany, during a 120-day incubation experiment. The type of soil significantly influenced the effect of these fungicides on the soil parameters studied. Incorporation of these fungicides generally stimulated the activity of phosphatases and ß-glucosidase, mineralization and the availability of N and most plant nutrients in soils. The activity of dehydrogenase and the availability of NO₃^- were generally adversely affected. Among the fungicides tested, the stimulation was more pronounced with mefenoxam followed by formulated metalaxyl.

¹H, ¹³C and two-dimensional NMR analyses were applied to determine the NMR parameters of 6-(2',3'-dihydro-1'H-inden-1'-yl)-1H-indene. The measurements were accomplished with 0.5 mg of the substance, this quantity being sufficient to determine the chemical shifts of all the H and C atoms, and also the appropriate coupling constants and to give the complete NMR resonance assignments of the molecule. The predicted patterns of the four different H atoms of the methylene groups of the indane structural element coincided completely with the complex patterns in the NMR spectra. Copyright © 2002 John Wiley & Sons, Ltd.

A method for the determination of sulfonamides and trimethoprim in the complex matrix liquid manure has been developed using reversed‐phase liquid chromatography and atmospheric pressure chemical ionization (APCI) tandem mass spectrometry. A comparison was made between electrospray and atmospheric pressure chemical ionization. APCI proved to be more robust and less sensitive to matrix effects. High‐performance liquid chromatographic (HPLC) separation of the analytes was achieved in less than 7 min. The compounds were extracted with ethyl acetate and the extracts were cleaned up by solid‐phase extraction on an aminopropyl column. Recoveries were not dependent on the concentration level. The mean recoveries were as follows: trimethoprim 79.0%, sulfadiazine 80.5%, N4‐acetylsulfadiazine 91.0%, sulfamerazine 78.6%, sulfadimidine 77.2% and sulfamethoxazole 82.8%. Linearity was established over a concentration range of 5 to 5000 µg/kg with correlation coefficients greater than 0.99. The method had a limit of quantitation (LOQ) of 5 µg/kg manure. Copyright © 2002 John Wiley & Sons, Ltd.

Reducing pesticide loads in surface waters implies identifying the pathways responsible for the pollution. The current study documents the pesticide contamination of the river Zwester Ohm, a 4917‐ha catchment in Germany with 41% of the land used for crop production. Discharges and concentrations of 19 pesticides were measured continuously at three locations for 15 mo. The load detected at the outlet of the catchment amounted to 9048 g a.i. The losses represent 0.22% of the pesticides applied by the farmers. The contamination showed a seasonal pattern following the pesticide application times. The wastewater treatment plant system (WWTPS) in the catchment (two wastewater treatment plants [WWTP], 14 combined sewer overflows (CSO), four CSO tanks) emits during dry weather periods purified sewage and during storm events sewage mixed with stormwater runoff into the river. The contribution by the WWTPS to the pesticide load was defined as point‐source pollution (PSP). The load was dominated by PSP with at least 77% of the total pollution. No significant interdependencies between intrinsic properties of the pesticides, hydrometeorological factors, and the loads occurring in the stream could be found. Therefore, it is not possible to predict PSP for other catchments based on the results from this study. Whereas 65% of the total load entered the river via the WWTP, a portion of 12% was attributed to the CSO. The study points out that the influence of CSO on PSP should be taken into account in future catchment studies in areas with comparable agricultural structure.

A new polyene pigment, melanocrocin, has been isolated either from fruit bodies or myce­lial cultures of the subterranean fungus Melanogaster broomeianus. The structure of the pig­ment was determined by spectroscopic methods and chemical transformations. Melanocrocin is the N-acyl derivative of ?-phenylalanine methyl ester with a polyolefinic carboxylic acid

The statements and discussions in this commentary are based on the disastrous situation near Baia Mare in Romania, where a dam of a cyanide basin burst on January 30, 2000, with serious consequences.

The conjugates of glycine with furan 2,5-dicarboxylic acid and of picolinic acid as well as a conjugate of pyroglutamic acid with phenylacetic acid, previously unknown as natural metabolites, were detected to be normal metabolites in human urine.