In situ selective determination of methylmercury in river water by diffusive gradient in thin films technique (DGT) using baker's yeast (Saccharomyces cerevisiae) immobilized in agarose gel as binding phase

Saccharomyces cerevisiae immobilized in agarose gel as binding phase and polyacrylamide as diffusive layer in the diffusive gradient in thin films technique (DGT) was used for selective determination of methylmercury (MeHg). Deployment tests showed good linearity in mass uptake up to 48 h (3276 ng). When coupling the DGT technique with Cold Vapor Atomic Fluorescence Spectrometry, the method has a limit of detection of 0.44 ng L⁻¹ (pre concentration factor of 11 for 48 h deployment). Diffusion coefficient of 7.03 ± 0.77 × 10⁻⁶ cm² s⁻¹ at 23 °C in polyacrylamide gel (pH = 5.5 and ionic strength = 0.05 mol L⁻¹ NaCl) was obtained. Influence of ionic strength (from 0.0005 mol L⁻¹ to 0.1 mol L⁻¹ NaCl) and pH (from 3.5 to 8.5) on MeHg uptake were evaluated. For these range, recoveries of 84–105% and 84–98% were obtained for ionic strength and pH respectively. Potential interference due to presence of Cu, Fe, Mn, Zn was also assessed showing good recoveries (70–87%). The selectivity of the proposed approach was tested by deployments in solutions containing MeHg and Hg(II). Results obtained showed recoveries of 102–115 % for MeHg, while the uptake of Hg(II) was insignificant. The proposed approach was successfully employed for in situ measurements in the Negro River (Manaus-AM, Brazil).     

Preconcentration of phthalic acid esters in water samples by Saccharomyces cerevisiae immobilized on silica gel

A sensitive and selective column adsorption method is proposed for the off-line preconcentration and determination of phthalic acid esters (PAEs), namely benzyl-butyl phthalate (BBP), di-n-butyl phthalate (DBP) and di-cyclohexyl phthalate (DCHP). The PAEs was preconcentrated on Saccharomyces cerevisiae immobilized on silica gel and then determined by high performance liquid chromatography (HPLC). Several parameters on the recovery of the analytes were investigated. Recoveries of BBP, DBP and DCHP were 100±2, 98±2 and 98±3%, respectively, at 95% confidence level under optimum conditions. The detection limits (3S/N) of BBP, DBP and DCHP were 3.2, 6.3 and 3.1 μg l⁻¹, respectively. The capacity of the adsorbent was also examined and found to be 1.4 mg g⁻¹ for BBP and DBP, and 3.6 mg g⁻¹ for DCHP. S. cerevisiae immobilized on silica gel is suitable for repeated use without decreasing recovery up to about 25 adsorption-elution cycles. The proposed method was successfully applied to the determination of PAEs in river water with high precision and accuracy.     

Baker’s yeast (Saccharomyces cerevisiae) mediated transformations of C-aryl-N-phenylnitrones

C-Aryl-N-phenylnitrones are transformed to a mixture of azoxybenzene and aryl aldehydes when treated with a mixture of Baker’s yeast and sucrose in pH = 6.0 phosphate buffer medium at 32 °C.     

Effect of high voltage electrical discharges on filtration properties of Saccharomyces cerevisiae yeast suspensions

In the present work the dead-end filtration of Saccharomyces cerevisiae yeast suspensions disrupted by high voltage electrical discharges (HVED treatment) was investigated. The efficiency of disruption was evaluated using conductivity disintegration index of suspension Z (Z = 0–1) and absorbance spectra of supernatant solutions. The electronic microscopy study, particle sizing and measuring of ζ-potential and turbidity were used to characterize variation of the colloidal properties of a yeast suspension during disruption. The HVED treatment was found to cause an effective disruption of yeast cells and extraction of intracellular proteins and other bio-products. The study of filtration revealed suspension filterability deterioration after disruption. It was shown that filtration behaviour of the HVED-processed suspensions was governed by cake formation, the filtrate volume decreased and the cake resistance increased with increase of Z. For high levels of disruption (Z > 0.99), filtration was governed by membrane fouling. The optimal dosage of polycationic flocculant promoted the formation of flocks and accelerated filtration. However, selected flocculant (poly(diallyldimethylammonium chloride)) provoked binding of bio-product and was inappropriate for using as an agent enhancing extraction from disrupted yeast cells.     

New separation methodologies for the distinction of the growth phases of Saccharomyces cerevisiae cell cycle

In the present work two separation techniques, namely the gravitational field-flow fractionation (GrFFF) and the reversed-flow gas chromatography (RFGC), are proposed for the distinction of the growth phases of Saccharomyces cerevisiae (AXAZ-1) yeast cycle at different temperatures (30 °C, 25 °C, 20 °C, and 15 °C) and pH (2.0, 3.0, 4.0 and 5.0) values. During the fermentation processes, differences observed in the peak profiles, obtained by GrFFF, can be related with the unlike cell growth. The distinction of the phases of AXAZ-1 cell cycle with the GrFFF, was also confirmed with the RFGC technique, which presented similar fermentation time periods for the alcoholic fermentation phases. Simultaneously, the reaction rate constant for each phase of the fermentation process and the activation energies were determined with the aid of the RFGC technique. Finally, the application of both the GrFFF and the RFGC techniques, in combination with high-performance liquid chromatography, allowed us to find the ideal experimental conditions (temperature and pH) for the alcoholic fermentation by AXAZ-1. The results indicate that S. cerevisiae cells performed better at 30 °C, whereas at lower temperatures decreases in the fermentation rate and in the number of viable cells were observed. Moreover, the pH of the medium (pH 5.0) resulted in higher fermentation rates and ethanol productivities.     

Overlapping chemical and genetic diversity in Ligularia lamarum and Ligularia subspicata. Isolation of ten new eremophilanes and a new seco-bakkane compound

Root chemicals and DNA sequences were analyzed for Ligularialamarum samples collected in Yunnan and Sichuan Provinces of China and the relationship between the plant and Ligulariasubspicata, its taxonomically closest species, was examined. Both species produce furanoeremophilanes and eremophilan-8-one derivatives as the major components. Eleven new compounds, including a new seco-bakkane compound, were isolated. The two species were found to harbor overlapping intra-specific diversities with respect to the chemicals and the DNA sequences.     

Euryjanicin A: a new cycloheptapeptide from the Caribbean marine sponge Prosuberites laughlini

A new proline-containing cycloheptapeptide, euryjanicin A (1), has been isolated from the marine sponge Prosuberites laughlini indigenous to Puerto Rico, and its structure established by an X-ray crystal structure determination. The absolute configuration of each amino acid residue was determined by Marfey’s method.     

Malbrancheamide, a new calmodulin inhibitor from the fungus Malbranchea aurantiaca

Bioassay-directed fractionation of an ethyl acetate extract (mycelia and broth) of the fungus Malbranchea aurantiaca led to the isolation of the novel phytotoxic alkaloid (5aS,12aS,13aS)-8,9-dichloro-12,12-dimethyl-2,3,11,12,12a,13-hexahydro-1H,6H-5a,13a (epiminomethano)indolizino[7,6b]carbazol-14-one (1) of the brevianamide series. The phytotoxin was given the trivial name of malbrancheamide (1). The structure of 1 was unequivocally established by UV, NMR, MS and X-ray studies. The absolute configuration was established by X-ray analysis according to the method of Flack. According to the conformational studies using molecular mechanics analyses, 1 exists in one preferred conformation, which was optimized by DFT calculations. Compound 1 caused moderate inhibition of radicle growth of Amaranthus hypochondriacus (IC₅₀=0.37 μM) and inhibited the activation of the calmodulin-dependent enzyme PDE1 (IC₅₀=3.65±0.74 μM). This effect was comparable to that of chlorpromazine (IC₅₀=2.75±0.87 μM) a well characterized CaM antagonist. The inhibition mechanism of 1 was competitive with respect to CaM according to a kinetic analysis.     

Emervaridione and varioxiranediol, two new metabolites from the endophytic fungus, Emericella variecolor

Two new compounds, emervaridione and varioxiranediol, together with the three known compounds, stellatic acid, ergosterol and dihydroterrein, were isolated from the endophytic fungus, Emericella variecolor. The structures of emervaridione and varioxiranediol were established from spectroscopic and X-ray crystallographic analysis.     

Hupermine A,a novel C16N2-type Lycopodium alkaloid from Huperzia phlegmaria

A novel C₁₆N₂-type Lycopodium alkaloid consisting of a quinolizidine with a 6-dimethylaminohexyl side chain, hupermine A (1), was isolated from the club moss of Huperzia phlegmaria, and the structure and relative stereochemistry were elucidated on the basis of spectroscopic data.     

Real-time detection of cofactor availability in genetically modified living Saccharomyces cerevisiae cells — Simultaneous probing of different geno- and phenotypes

This work describes a mediated amperometric method for simultaneous real-time probing of the NAD(P)H availability in two different phenotypes, fermentative and respiratory, of the phosphoglucose isomerase deletion mutant strain of S. cerevisiae, EBY44 [ENY.WA-1A pgi1-1D::URA3], and its parental strain, ENY.WA-1A. The developed method is based on multichannel detection using microelectrode arrays. Its versatility was demonstrated by using four microelectrode arrays for simultaneously monitoring the NAD(P)H availability of both geno- and phenotypes under the influence of two different carbon sources, glucose and fructose, as well as the cytosolic and mitochondrial inhibitor and uncoupler, dicoumarol. The obtained results indicate that the method is capable of accurately and reproducibly (overall relative standard error of mean 3.2%) mapping the real-time responses of the cells with different genotype–phenotype combinations. The ENY.WA cells showed the same response to glucose and fructose when dicoumarol was used; fermentative cells indicated the presence of cytosolic inhibition and respiratory cells a net effect of mitochondrial uncoupling. EBY44 cells showed cytosolic inhibition with the exception of respiratory cells when fructose was used as carbon source.     

A new rearranged and a new seco-ent-kaurane diterpenoids from Isodon parvifolius

Parvifoline X (1), a new rearranged ent-kaurane diterpenoid, and parvifoline Y (2), a new 8,15-seco-ent-kaurane diterpenoid, were isolated from the leaves of Isodon parvifolius. Their structures were elucidated by spectroscopic methods including 2D NMR analysis, and supported by a biogenetic pathway. Parvifoline X (1), possessing a new 15(8→11)-abeo-7α,20-epoxy-ent-kaurane skeleton, was found from the genus Isodon for the first time. Compounds 1 and 2 were evaluated for their inhibitory activity against A549, HT-29, and K562 cell lines. Parvifoline Y (2) was the most cytotoxic against A549 cells with an IC₅₀ value of 4.97 μM.     

Coibanoles, a new class of meroterpenoids produced by Pycnoporus sanguineus

Three new terpenoids of mixed biosynthetic origin were isolated from the culture filtrate of the endophytic fungus Pycnoporus sanguineus. Their structures were determined by extensive spectroscopic analyses. We have named these tricyclic and tetracyclic metabolites ‘coibanoles A-C’ in reference to Coiba Island and Coiba National Park, Panamá, from which the plant and endophyte were collected. The extract was inactive to the human parasites Trypanosoma cruzi, Leishmania donovani, and Plasmodium falciparum at a test concentration of 10 μg/mL.     

Daphcalycinosidines A and B, new iridoid-alkaloids from Daphniphyllum calycinum

Three new alkaloids, daphcalycinosidines A (1) and B (2) and daphcalycic acid (3) have been isolated from the seeds of Daphniphyllum calycinum. The structures and relative stereochemistries were determined on the basis of spectral studies including 2D NMR, mass spectrometry and chemical transformations. Structures 1 and 2 are characterized by an iridoid glucoside moiety linked to new Daphniphyllum alkaloid moieties.     

A pair of new atropisomeric cupressuflavone glucosides isolated from Juniperus communis var. depressa

A pair of new atropisomers, (M)- and (P)-cupressuflavone 4′-O-β-d-glucoside, were isolated from Juniperus communis var. depressa, and their absolute structures and axial configurations were determined using 2D NMR and circular dichroism. These are the first reported stable (M)- and (P)-isomers of cupressuflavone glucosides isolated from natural sources.     

Lyconadins C and F, new Lycopodium alkaloids from Lycopodium complanatum

New Lycopodium alkaloids, lyconadins C (1) and F (2), were isolated from the club moss Lycopodium complanatum. Lyconadin C (1) is a new C₁₆N₂-type Lycopodium alkaloid possessing unique fused-tetracyclic ring system consisting of a cycloheptene ring fused to a decahydroquinoline and pyridone rings. Lyconadin F (2) possesses a primary amide moiety in its molecular, which is the first example of Lycopodium alkaloids. Biogenetically, lyconadins C (1) and F (2) might be related to lyconadins A (4) and B (5). The structures and relative stereochemistry of 1 and 2 were elucidated on the basis of spectroscopic data. The absolute stereochemistry of 2 was elucidated by chemical correlations with lyconadin B (5) through hemiaminal form of lyconadin F (3).     

Aphapolynins A and B, two new limonoids from the fruits of Aphanamixis polystachya

Two new highly oxidized A,B-seco limonoids, aphapolynins A (1) and B (2), were isolated from the fruits of Aphanamixis polystachya. Their structures were elucidated by spectroscopic analysis, in particular, the absolute configuration of aphapolynin A was determined by a single-crystal X-ray study using a mirror Cu Kα radiation. Aphapolynin A exhibit moderate cytotoxicities when tested against a panel of tumor cell lines.     

A crotofolane-type diterpenoid and a rearranged nor-crotofolane-type diterpenoid with a new skeleton from the stems of Croton cascarilloides

From the stems of Croton cascarilloides collected in the Okinawa Islands, a structurally rare crotofolane-type diterpenoid (1) and a rearranged nor-crotofalane, a new skeletal diterpenoid (2) were isolated. The structures were determined by X-ray crystallographic analyses, establishing their absolute stereostructures for the first time. Compound 2 was probably biosynthesized from 1 through several steps, such as decarboxylation, oxidation, C-C bond migration.     

Fuligoic acid, a new yellow pigment with a chlorinated polyene-pyrone acid structure isolated from the myxomycete Fuligoseptica f. flava

Fuligoic acid (1), a new yellow pigment with a chlorinated polyene-pyrone acid structure, was isolated from field-collected fruit bodies of the myxomycete Fuligoseptica f. flava, and its structure was elucidated by spectral data, including its absolute configuration based on CD data.     

Exiguolide, a new macrolide from the marine sponge Geodia exigua

A new 20-membered macrolide designated exiguolide has been isolated from the marine sponge Geodia exigua, and its structure determined by interpretation of spectroscopic data. Exiguolide specifically inhibited fertilization of sea urchin (Hemicentrotus pulcherrimus) gametes but not embryogenesis of the fertilized egg.