Accurate LC-MS analyses for microcystins using per-15N-labeled microcystins.

Per-(15)N-labeled microcystins were prepared for use as surrogates for accurate liquid chromatography-mass spectrometry analysis. Two strains of Microcystis aeruginosa were cultured in (15)NO(3)-containing TS-15 medium. To change from the incorporation of (14)N to (15)N into all cell components, cells of Microcystis aeruginosa were precultured in Na(15)NO(3)-containing medium for more than 6 months. After mass cultivation of the strains, cells of each strain were harvested and lyophilized. Microcystin variants were extracted from the lyophilized cells and per-(15)N-labeled microcystin variants were purified using high-performance liquid chromatography and high-performance thin-layer chromatography. The structures of per-(15)N-labeled microcystin variants were confirmed by their mass spectrometry spectra and NMR spectra. When per-(15)N-labeled microcystins were used as surrogates for quantitative analysis of these toxins in cyanobacterial cells, excellent accuracy (98-106%) was obtained, with the m/z of M(+), [M+1](+), and [M+2](+) of both microcystins and the per-(15)N-labeled microcystins as surrogates being completely separated. In conclusion, per-(15)N-labeled microcystins are excellent surrogates for microcystin analysis using liquid chromatography-mass spectrometry.

Analysis of triorganotin compounds in water samples by hydrophilic interaction liquid chromatography-electrospray ionization-mass spectrometry.

Hydrophilic interaction liquid chromatography (HILIC)-electrospray ionization-mass spectrometry (ESI-MS) was evaluated for the analysis of tributyltin (TBT) and triphenyltin (TPT) in water samples. Separation was performed in isocratic mode on an Atlantis HILIC silica (2.1 mm x 150 mm, 5 microm) column with a mobile phase of acetonitrile-0.1% aqueous HCOOH (86:14, v/v) at a flow rate of 0.2 mL/min. Under optimum conditions, limits of detection for TBT and TPT were 10 and 20 pg injected onto the column, respectively. The extraction of triorganotin compounds from seawater samples was carried out using a polymer-based solid phase extraction cartridge of mixed modes with reversed-phase and weak anion exchange. Tributyltin-d(27) chloride and triphenyltin-d(15) chloride were used as internal standards. The relative standard deviations for the analysis were less than 4%. Using the proposed method, it was possible to analyze concentrations of TPT and TBT in seawater at ppt levels.

NIES certified reference material for microcystins, hepatotoxic cyclic peptide toxins from cyanobacterial blooms in eutrophic water bodies.

A certified reference material (CRM) for microcystins has been prepared by the National Institute for Environmental Studies (NIES). Microcystins are hepatotoxic cyclic peptides produced by cyanobacteria in eutrophic water bodies. At least seven microcystin variants were found by HPLC analysis of the NIES CRM, of which [Dha(7)]microcystin-RR and -LR were the major microcystins present. Because of the lack of available standards we determined the total microcystin concentration in the CRM by the MMPB method, and elucidated the structures of the main individual microcystin variants following their isolation. Analyses of NMR and MS spectra indicated that the remaining minor variants in the CRM were [D-Asp(3), Dha(7)]microcystin-RR and -LR, and [Dha(7)]microcystin-YR, -ThTyrR, and -HilR. The CRM is valuable not only as a standard material for the quantitation of total microcystins but also for the identification of individual [Dha(7)]microcystin variants.

New values of molecular extinction coefficient and specific rotation for cyanobacterial toxin cylindrospermopsin.

The molecular extinction coefficient epsilon of cylindrospermopsin (CYN) purified by the anion exchange and the normal-phase HPLC procedures was determined to be 9800 at 262 nm. This epsilon is significantly higher than those (epsilon, 5800-6250) reported previously. In order to determine CYN concentrations in solutions using UV absorption, the epsilon-value of CYN should be corrected from 5800 to 9800. Further, the [alpha](D) value of CYN should be corrected from +12.5 degrees to +17.0 degrees .

Fibrous adsorbent for removal of aqueous aromatic hydrocarbons.

Bundles of a strongly hydrophobic fibrous material (p-phenylene-2,6-benzobisoxazole; PBO; Zylon) were employed as an adsorbent for the removal of aqueous aromatic compounds, because the PBO fibers are too rigid to be woven and did not entrap suspended solids. The removal performance for nine kinds of polyaromatic hydrocarbons (PAHs) and di-(2-ethylhexyl) phthalate (DEHP) was evaluated. PAHs and DEHP at initial concentrations of 50 microg L(-1) were removed at 72.5-99.9% and ca. 95%, respectively, although the removal efficiencies were affected by the phase ratio (fiber weight/solution volume). The logarithm of the partition coefficient (log K) for planar PAHs was linearly correlated with the logarithm of the n-octanol/water partition coefficient (log P), but nonplanar PAHs, such as cis-stilbene, p-terphenyl, and o-terphenyl, showed significantly lower adsorption performance. The adsorbed PAHs were not desorbed effectively with CH3CN, CH2Cl2, and toluene. On the other hand, DEHP was effectively desorbed with methanol.

A novel biosurfactant, 2-acyloxyethylphosphonate, isolated from waterblooms of Aphanizomenon flos-aquae.

A novel biosurfactant, 2-acyloxyethylphosphonate, was isolated from waterblooms of Aphanizomenon flos-aquae. Its structure was elucidated by chemical degradation and HRFABMS, GC/EI-MS and 1D- and 2D-NMR spectral analyses. The surfactant contained one mole of 2-hydroxyethylphosphonate and one mole of fatty acid, with hexadecanoic acid accounting for 84.1% of the total fatty acid content. The structure was confirmed by synthesis of 2-oleoyloxyethylphosphonate from ethylene oxide, phosphorus acid and oleic acid chloride. Considering the isolated surfactant molecule as hexadecanoyloxyethylphosphonic acid (mw. 364), the critical micelle concentration (CMC) was about 22 mM.

Leucine aminopeptidase M inhibitors, cyanostatin A and B, isolated from cyanobacterial water blooms in Scotland.

Two leucine aminopeptidase M inhibitors, cyanostatin A and B, were isolated from cyanobacterial water blooms at Loch Rescobie in Scotland, and specifically from a Microcystis species. Both inhibitors were lipopeptides containing 3-amino-2-hydroxydecanoic acid and weak inhibitors of protein phosphatase (PP2A). Both strongly inhibited the activity of leucine aminopeptidase M with IC50 values of 40 and 12 ng/ml, respectively.

Microbial production and vaporization of mono-(2-ethylhexyl) phthalate from di-(2-ethylhexyl) phthalate by microorganisms inside houses.

Laboratory workers were bothered by an irritation that caused coughing during the cultivation of microorganisms that degraded di-(2-ethylhexyl) phthalate (DEHP). The authors found that mono-(2-ethylhexyl) phthalate (MEHP), a known cause of asthma, was released during the degradation of DEHP. At its highest production and vaporization rate, the amount was almost equal to that of the DEHP starting material. It appeared that transport into the atmosphere depended on its adsorption on dust particles. The authors attempted to cultivate several microorganisms from house materials, especially those composed of rotting polyvinyl chloride. And microorganisms produced MEHP in the culture medium. In addition, MEHP was produced from DEHP by several stock microorganisms. Thus, MEHP could easily be produced from DEHP by microorganisms in the environment. In Japan, there are many cases of asthma with unknown causes. If MEHP is one of causes, then preventive measures against some cases of asthma could be taken.

A Dhb-microcystin from the filamentous cyanobacterium Planktothrix rubescens.

A Dhb-microcystin variant was isolated from the filamentous cyanobacterium Planktothrix rubescens. Its structure was elucidated as (E)-Dhb-microcystin-HilR ([D-Asp3, (E)-Dhb7]microcystin-HilR) on the basis of spectral data and amino acid analysis after acid hydrolysis.

Interval immobilization technique for recognition toward a highly hydrophilic cyanobacterium toxin.

A novel adsorption medium containing selective molecular recognition site for one of the powerful cyanobacterium toxins, Cylindrospermopsin (CYN) was developed using a special technique, namely interval immobilization technique. The adsorption medium was prepared using molecular assembly derived from an alternative-template molecule coupled with functional monomers for fixing the interval between the ionic functional groups in CYN. As results of liquid chromatographic evaluations, selective molecular recognition ability for CYN was observed as expected. Further studies proved that the association constant for CYN on this medium was slightly higher than that on blank polymer.

Spiroidesin, a novel lipopeptide from the cyanobacterium Anabaena spiroides that inhibits cell growth of the cyanobacterium Microcystis aeruginosa.

Spiroidesin (1), a novel D-amino acid-containing linear lipopeptide, was isolated from waterblooms of the cyanobacterium Anabaena spiroides. The structure was identified by 2D NMR and chemical degradation analyses. Spiroidesin inhibited cell growth of the toxic cyanobacterium Microcystis aeruginosa (IC(50), 1.6 x 10(-6) M).