Quantitative detection of bisphenol A and bisphenol A diglycidyl ether metabolites in human plasma by liquid chromatography–electrospray mass spectrometry

Due to the ubiquity of epoxy resin compounds and their potential role in increasing the risk for reproductive dysfunction and cancer, the need for an assessment of human exposure is urgent. Therefore, we developed a method for measuring bisphenol A (BPA) and bisphenol A diglycidyl ether (BADGE) metabolites in human blood samples using high-performance liquid chromatography–electrospray ionization mass spectrometry (LC–MS). Human blood samples were processed using enzymatic deconjugation of the glucuronides followed by a novel sample preparation procedure using a solid-phase-cartridge column. This selective analytical method permits rapid detection of the metabolites, free BPA and a hydrolysis product of BADGE (BADGE-4OH) with detection limits in the low nanogram per milliliter range (0.1 ng ml⁻¹ of BPA and 0.5 ng ml⁻¹ of BADGE-4OH). The sample extraction was achieved by Oasis HLB column on gradient elution. The recoveries of BPA and BADGE-4OH added to human plasma samples were above 70.0% with a standard deviation of less than 5.0%. This selective, sensitive and accurate method will assist in elucidating potential associations between human exposure to epoxy-based compounds and adverse health effects.     

Tail-associated structural protein gp61 of Staphylococcus aureus phage φMR11 has bifunctional lytic activity

A tailed bacteriophage, φMR11 (siphovirus), was selected as a candidate therapeutic phage against Staphylococcus aureus infections. Gene 61, one of the 67 ORFs identified, is located in the morphogenic module. The gene product (gp61) has lytic domains homologous to CHAP (corresponding to an amidase function) at its N-terminus and lysozyme subfamily 2 (LYZ2) at its C-terminus. Each domain of gp61 was purified as a recombinant protein. Both the amidase [amino acids (aa) 1–150] and the lysozyme (aa 401–624) domains but not the linker domain (aa 151–400) caused efficient lysis of S . aureus . Immunoelectron microscopy localized gp61 to the tail tip of the φMR11 phage. These data strongly suggest that gp61 is a tail-associated lytic factor involved in local cell-wall degradation, allowing the subsequent injection of φMR11 DNA into the host cytoplasm. Staphylococcus aureus lysogenized with φMR11 was also lysed by both proteins. Staphylococcus aureus strains on which φMR11 phage can only produce spots but not plaques were also lysed by each protein, indicating that gp61 may be involved in 'lysis from without'. This is the first report of the presence of a tail-associated virion protein that acts as a lysin, in an S. aureus phage.     

Effect of Clostridium perfringens Beta Toxin on Blood Pressure of Rats

Guanethidine treatment or adrenal medullectomy significantly inhibited the elevation in blood pressure induced by Clostridium perfringens beta toxin, and the combination of the two drastically reduced the pressure rise, to less than 19% of that in control rats. When rats were pretreated with tetrodotoxin or hexamethonium, the toxin-evoked rise was significantly inhibited. Elevation in blood pressure induced by the toxin in spinal rats tended to be less than that in control rats. When investigated by a microscopical technique, arteriolar constriction in the mesenteric vasculature was observed after the blood pressure elevation induced by the toxin reached a maximum. Blood flow in the skin decreased with an increase in blood pressure following intravenous injection of the toxin. It is concluded that beta toxin acts on the autonomic nervous system and produces arterial constriction.     

Isolation and Chemical Structure of New Oxidation Product of 5-Ketogluconic Acid,and a Hypothetical Pathway from Glucose to Tartaric Acid through this New Compound

In the course of study on the mechanism of the tartaric acid formation from 5-ketogluconic acid, a new intermediary substance with mauve color to Abdel-Akhel and Smith’s reagent was isolated from intact cell culture liquid. The chemical structure of this substance was determined as 1,2-dihydroxyethyl hydrogen L(+) tartrate from the results of hydrolysis experiments and from the identifications of the constituents of the molecule, and named “pretaric acid.” Tartaric acid was evidently produced from pretaric acid by intact cell culture. Clearly, then, pretaric acid appears to be an intermediate in the formation of tartaric acid from 5-ketogluconic acid. The authors assumed that in the formation of pretaric acid from 5-ketogluconic acid, a Baeyer-Villiger type oxidation occurred.     

Multimeric bivalent immunogens from recombinant tetanus toxin H<Subscript>C</Subscript> fragment,synthetic hexasaccharides,and a glycopeptide adjuvant

Using recombinant tetanus toxin H_C fragment (rTT-H_C) as carrier, we prepared multimeric bivalent immunogens featuring the synthetic hexasaccharide fragment of O-PS of Vibrio cholerae O:1, serotype Ogawa, in combination with either the synthetic hexasaccharide fragment of O-PS of Vibrio cholerae O:1, serotype Inaba, or a synthetic disaccharide tetrapeptide peptidoglycan fragment as adjuvant. The conjugation reaction was effected by squaric acid chemistry and monitored in virtually real time by SELDI-TOF MS. In this way, we could prepare well-defined immunogens with predictable carbohydrate–carrier ratio, whose molecular mass and the amount of each saccharide attached could be independently determined. The ability to prepare such neoglycoconjugates opens unprecedented possibilities for preparation of conjugate vaccines for bacterial diseases from synthetic carbohydrates.     

Density dependent growth of corneal endothelial cells cultured in vitro

Using the cornea of macaque monkey, we demonstrated the relationship between cell density and growth of endothelial cells in vitro. Corneal endothelial cells in a cell sheet grow most actively in regions with cell density of 1000 to 1800 cells/mm2, in explant cultures and cell sheets and in concentrated inocula dissociated cells. Cell morphology was well sustained in these cultures. Cells cultured at a higher cell density retained their potential to proliferate actively, showing clear contrast to cells cultured at a density lower than 200 cells/mm2. When dissociated cells were cultured at a low density and maintained for more than 4 weeks, they gradually lost their growth potential, altered into polymorphonuclear giant cells and eventually dedifferentiated. In addition, cells with no contact with each other did not express growth potential. Density dependent growth was confirmed by measuring the mitotic index against the cell density per square mm from the center to the peripheral regions in cultured explants. It is concluded that the growth pattern of corneal endothelial cells is closely related to cell density, and that growth of these cells might be regulated through intercellular communications.     

Existence of a polar digitalis-like factor in mammalian hypothalamus

We were able to partially purify a polar digitalis-like factor from rat and bovine hypothalami based on the capacity to inhibit [3H]ouabain binding to intact human erythrocytes. This factor was characterized in reference to the digitalis-like factor that we have isolated and reported on. Hypothalamic factor shared digitalis-like activities and physicochemical properties with the one derived from human urine and mammalian plasma. These findings strongly suggest that a polar digitalis-like factor identical to the circulatory factor does exist in mammalian hypothalamus.     

A disease with features of cutis laxa and Ehlers-Danlos syndrome

Summary A mother and daughter are described with light and electron microscopic, and biochemical abnormalities of their connective tissue characteristic of both cutis laxa and the Ehlers-Danlos syndrome. The mother was clinically normal, while her 8-year-old daughter exhibited loose, wrinkled skin and other clinical features of cutis laxa, and also fragility, bruisability and hyper-extensibility of the skin and poor healing of wounds, leaving cigarette paper scars, features characteristic of the Ehlers-Danlos syndrome. Light and electron microscopic studies of skin biopsy specimens and cultured skin fibroblasts from both individuals revealed reduced and distorted elastic fibres, a finding usually seen in cutis laxa. Electrophoretic studies of collagen excreted from cultured skin fibrobasts revealed in both individuals and alpha 2(I) chain with a molecular size smaller than usual. The father and elder daughter were normal by clinical, light and electron microscopic and electrophoretic studies. It was concluded from these findings that the mother and daughter represented a hitherto undescrbed disease of the connective tissue with dominant inheritance and variable expressivity.     

CAM Photosynthesis under Drought Conditions in Kalanchoe blossfeldiana Grown with Nitrate or Ammonium as the Sole Nitrogen Source

K. blossfeldiana Poelln. cv. Hikan was grown in vermiculite,supplied daily with nutrient solution containing 1 mM (or 10mM) nitrate or ammonium as the sole nitrogen source. The nitrate-grownplants had more activity of CAM (Crassulacean acid metabolism)photosynthesis (nocturnal CO₂ uptake in the shoot and nocturnalincreases of titratable acidity and malate content in the leaves)than the ammonium-grown plants. Interruption of the solutionsupply for 5 or more days (drought conditions) increased theactivity of CAM photosynthesis in nitrate- or ammonium-grownplants, and the diurnal CO₂ uptake pattern in the nitrate-grownplants shifted from ‘weak-CAM’ to ‘full-CAM’.The difference in the activity of CAM photosynthesis betweennitrate- and ammonium-grown plants increased under the droughtconditions. When the solution was resupplied, the activity ofCAM photosynthesis rapidly decreased to the levels before theinterruption. The physiological mechanism and ecological significanceof the effect of the nitrogen source on CAM photosynthesis arediscussed (Received January 5, 1988; Accepted April 13, 1988)