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.     

Effect of Gibberellic Acid on the Ultrastructure and {alpha}-Amylase Activity of Aleurone Cells of Avena sativa L.

The -amylase activity and ultrastructure of aleurone cells inseeds of Avena sativa L. were studied using seed halves withembryo (embryo seeds) which had imbibed water and seed halveswithout embryo (embryo-less seeds) which had imbibed water withor without GA₃. -Amylase activity was detected in the aleurone layers of embryoseeds that had imbibed water and embryo-less seeds that hadimbibed GA₃-water. The ultrastructure of aleurone cells withdetectable -amylase activity showed marked changes in the roughsurfaced endoplasmic reticulum (rER), in the flattened sacculesforming stacks and in the aleurone grains. The progressive changesin the rER were as follows: first, the number of slender rERincreased; then, the inner space became wider and showed roundor oval profile; and finally, the rER became slender again witha reduced number of adhering ribosomes. The flattened sacculesforming stacks were appressed to the surface of aleurone grains.With time, they decreased in number and finally disappeared.In parallel with the decrease of flattened saccules, digestionof proteinaceous material inside the aleurone grains proceeded. (Received February 24, 1987; Accepted September 3, 1987)     

Effect of UV-B (290–320 nm) irradiation on growth and metabolism of cucumber cotyledons

Cucumber ( Cucumis sativus L. cv. Natsusairaku 3) seedlings were grown in a growth cabinet under UV-B (290–320 nm) irradiation (equivalent to the UV-B radiation normally incident at Tokyo, 36°N latitude, during clear sky conditions in mid-april on a weighted daily fluence basis) and a UV-B-free control condition. UV-B irradiation inhibited the growth of the cotyledons, i.e. the increase in area, and increase in fresh and dry weights of the cotyledons. The greatest inhibition rate was observed in the increase in area, causing a significant increase in specific leaf weight (the ratio of weight to area). UV-B irradiation had no significant effect on DNA and RNA contents in the cotyledons, but decreased protein content slightly. In contrast, the irradiation reduced the amounts of organic acids and soluble sugars, indicating that primary carbon metabolism was very sensitive to UV-B radiation. UV-B irradiation lowered the photosynthetic activity in the cotyledons without any effect on chlorophyll content and respiratory activity. These results indicate that UV-B radiation at the ambient level may act as a physiological stress in some UV-sensitive plants.     

Proportionality of ELF electric field-induced growth inhibition to induced membrane potential inZea mays andVicia faba roots

Summary The postulate that 60-Hz electric field-induced bioeffect severity is proportional to induced transmembrane potential [ ] magnitude was tested and supported using a plant root model cell system. Statistically significant correlations were obtained upon regression of the relative rates of exposedVicia faba andZea mays root segment growth on the average (calculated) arising in those segments under specified 60 Hz field exposure conditions. The associated with the apparent threshold for growth inhibition was similar inZea andVicia roots (2.5 vs 2.4 mV, respectively). At greater than this threshold,Zea root growth declined by about 9% per mV, andVicia root growth by about 19% per mV induced potential.Abbreviations EF electric field - RGR relative growth rate - RSGR relative segmental growth rate - induced membrane potential - segmental-average induced membrane potential - VC _d region of root tip in which a complete, nascent vascular cylinder is first distinguishable in histological sections     

Correction of inverted nipple with periductal fibrous flaps.

I devised a method to correct the inverted nipple considering the preservation of the lactiferous ducts, sensory fibers to the nipple, and the contracting function of the areolar muscle. Excision of the excess skin at the base of the nipple was done in three diamonds fashion, and they were located at 2, 6, and 10 o'clock positions not to jeopardize the sensory fibers to the nipple. To release the fastened nipple, the periductal fibrous tissue was thoroughly dissected and made into three flaps pedicled inferiorly. These three flaps were sutured to the dermis of the periareolar skin to pull up the nipple base by means of traction in three directions. The purse-string suture, the dermal stitch on the shorter diagonals of the diamond-shaped defects, anchors the skin-muscle bridges caught at the base of the ductal column, makes the nipple base narrower, obtains stable anchoring, helps the areolar muscle contraction to resume, and prevents the recurrence of the inversion. The use of the periductal tissue as flaps to bring in areolar skin for easier anchoring and for more prominent eversion of the nipple has not been described in the literature.     

Cloning and expression of a cDNA encoding mouse indoleamine 2,3-dioxygenase

The depletion of an essential amino acid (aa), tryptophan, caused by interferon-gamma (IFN-gamma)-mediated induction of indoleamine 2,3-dioxygenase (IDO) in mouse allografted tumor cells, has been suggested as a reason for the allograft rejection. To elucidate the mechanism of this IDO induction, attempts were made to isolate cDNA clones encoding mouse IDO. In seven of 25 mouse cell lines, IDO was induced by IFN-gamma, and the highest IDO induction was observed in the case of rectal cancer (CMT-93) cells, which were further stimulated two- to threefold by the simultaneous addition of dibutyryl cyclic AMP (Bt2cAMP). A cDNA library was prepared from poly(A)+ RNA isolated from CMT-93 cells treated with IFN-gamma/Bt2cAMP. The cDNA clones were isolated using the cDNA encoding human IDO as a probe. The mouse IDO cDNA encodes a 407-aa protein with an Mr of 45,639. The deduced aa sequence agreed with partial aa sequences derived from endopeptidase digestion of purified mouse IDO and revealed 61% homology with that of human IDO. Transient expression of the mouse IDO cDNA in COS-7 cells yielded a high level of IDO activity in the cells. Northern hybridization analysis of RNA in CMT-93 cells indicated that IFN-gamma induced the IDO mRNA, and that the level of RNA was increased by simultaneous addition of Bt2cAMP, while Bt2cAMP itself had no effect on mRNA induction.     

Comparison of the affinities of newly identified human bile acid binder and cationic glutathione S-transferase for bile acids

The bile acid binding properties of the newly identified bile acid binder (Mr = 36,000) (FEBS Lett. 1984. 177: 31-35) and the major cationic glutathione (GSH) S-transferase (Mr = 50,000) in human liver cytosol were compared. Binding affinities were measured by the competitive displacement by bile acids of 1-anilino-8-naphthalene sulfonate (ANS) bound to the proteins and, in some cases, by direct methods of flow dialysis and equilibrium dialysis. The binding affinities for various bile acids by the human bile acid binder were 2-5 orders of magnitude greater than those by human cationic GSH S-transferase. This suggests an important physiologic role for the former protein in intracellular transfer of bile acids in human liver.     

Human indolylamine 2,3-dioxygenase. Its tissue distribution, and characterization of the placental enzyme.

The presence of indolylamine 2,3-dioxygenase was examined in human subjects by determining its activity with L-tryptophan as substrate. Enzyme activity was detected in various tissues, and was relatively high in the lung, small intestine and placenta. Human indolylamine 2,3-dioxygenase, partially purified from the placenta, had an Mr of about 40 000 by gel filtration and exhibited a single pI of 6.9. The human enzyme required a reducing system, ascorbic acid and Methylene Blue, for maximal activity and was able to oxidize D-tryptophan, 5-hydroxy-L-tryptophan as well as L-tryptophan, but kinetic studies indicated that the best substrate of the enzyme was L-tryptophan.     

Inactivation and sedimentation of mercury from organomercurials by Klebsiella pneumoniae

Abstract A susceptibility of 63 clinical isolates of Klebsiella pneumoniae to inorganic and organic mercuric compounds was determined. 18 of them were found to be resistant to fluorescein mercuric acetate (FMA) and merbromin (MB). Moreover, all the resistant strains inactivate the antibacterial effect of FMA. The changes in the amount of organic mercury at the time of inactivation of the drug and the structures of the end products were examined in detail with the plasmid-bearing strain JK9 and its transconjugants of Escherichia coli .
The results showed that FMA was inactivated by an intracellular enzyme produced inducively and was degraded to fluorescein (sodium salt, uranine), which led to the sedimentation of metallic mercury. The discovery of the genes conferring inducible organic mercury-inactivating enzymes determined by plasmids was the next step and their application in the recovery of metallic mercury from organomercurials is now imminent.     

IFN-gamma is the inducer of indoleamine 2,3-dioxygenase in allografted tumor cells undergoing rejection

The depletion of an essential amino acid, tryptophan, caused by induction of indoleamine 2,3-dioxygenase (IDO), has been shown to be a mechanism involving self-defense against inhaled microorganisms and tumor growth. We recently reported that the IDO is dramatically (approximately 50-fold) induced in allografted tumor (3-methylcholanthrene-induced ascites type tumor cells) cells undergoing rejection, and that the enzyme is induced by factor(s) released through the interaction of allografted tumor cells with infiltrating leukocytes. The culture supernatant of infiltrating leukocytes, which were harvested on day 7 after tumor transplantation, induced the highest IDO activity in the tumor cells. The inducer activity was completely neutralized by the addition of antibody to IFN-gamma but not by antibody to IFN-alpha/beta. Approximately 6 U/ml of IFN-gamma was detected by an ELISA assay in the 12-h culture supernatant with 2 x 10(6) leukocytes/ml, and rIFN-gamma at 6 U/ml induced IDO in 3-methylcholanthrene-induced ascites type tumor cells to the same extent as IFN-gamma in the culture supernatant. Moreover, i.p. administration of antibody to IFN-gamma almost completely inhibited the induction of IDO in the allografted tumor cells. These observations indicate that the factor responsible for IDO induction in the allografted tumor cells is IFN-gamma.