Simultaneous liquid chromatography-tandem mass spectrometric determination of albendazole sulfoxide and albendazole sulfone in plasma

This paper describes a simple, fast, sensitive and reliable method for the simultaneous determination of albendazole sulfoxide (ASOX) and albendazole sulfone (ASON), the two most important metabolites of the drug albendazole (ABZ), in plasma samples using liquid chromatography and tandem mass spectrometry. After liquid-liquid extraction with dichloromethane, the two albendazole metabolites and the internal standard phenacetin were resolved in a CN column using the mobile phase methanol-water (4:6, v/v) acidified with 1% acetic acid. Detection by electrospray mass spectrometry was carried out in the positive ion mode. The method was linear up to 2500 and 250 ng/ml for ASOX and ASON, respectively, with mean recoveries of more than 85%. The precision and accuracy data, based on within- and between-day variations over 5 days, were lower than 15%. The quantitation limits of 0.5 and 5.0 ng/ml for ASON and ASOX are low enough for the method to be suitable for pharmacokinetic studies. Pharmacokinetic data obtained with the proposed method following oral administration of ABZ to a patient with neurocysticercosis are also reported.     

Nitrite-mediated decarboxylative conjugation of caffeic acid with glutathione under mildly acidic conditions

In acetate buffer, pH 4, at room temperature, nitrite ions can mediate an unusual decarboxylative conjugation of caffeic acid with glutathione leading to novel isomeric 2-(3,4-dihydroxyphenyl)-2-S-glutathionylacetaldehyde oximes. These results hint at a possible role of endogenous and/or dietary glutathione in the mechanisms by which caffeic acid can affect the burden of carcinogenic N-nitroso compounds in the digestive tract.     

Aqueous and solid complexes of iron(III) with hyaluronic acid. Potentiometric titrations and infrared spectroscopy studies

The coordination of iron(III) ion to hyaluronic acid (Hyal) in aqueous solutions and solid state was accomplished by potentiometric titrations and infrared spectroscopy. The potentiometric titration studies provided the binding constants for the complexes found in the systems and the speciation of these species according to the variation of pH values. The complexes found presented a complexing ability through both the chelating moieties of Hyal (via the N-glucosamine and D-glucoronic acid), showing no special preference for either one while in solid state, but when in aqueous solution the complexation via the N-glucosamine moiety was the preferred, forming two complexed species, ML and ML(2) (log K(ML)=8.2 and log K(ML2)=7.9). The presence of a mu-oxo complex via the D-glucoronic acid was also detected in both aqueous (log K=6.7) and solid states via the N-glucosamine and D-glucoronic acid simultaneously linked to two Hyal chains. A structure for this latter complex was suggested. The results indicated that these complexes could be used in eliminating the excess iron(III) in living organisms.     

Synergistic effect of the anti-HER-2/neu antibody and cisplatin in immortalized and primary mesothelioma cell lines

Malignant mesothelioma (MM) still remains a therapeutic and diagnostic problem to which new therapeutic perspectives are being continuously tried and tested. Three different primary cultures (MMGe-1, MES MM 98, and MES 1) and one immortalized cell line (MSTO 211 H) of human MM were studied in order to evaluate the HER-2/neu expression. Three out of four cell lines showed a different level of c-erbB-2 expression, the highest being detected on the MSTO 211 H cell line (fibroblastic phenotype), whereas MMGe-1 resulted negative. The effect of the anti-HER-2/neu antibody (Trastuzumab) alone, and in combination with cisplatin (CDDP) at different doses (ranging from 0.1 to 100 microg/ml), was studied on all the c-erB-2 positive cell lines. Trastuzumab was able to inhibit cell proliferation in a time-dependent manner, with growth inhibition also obtained at low concentrations (0.1-1 microg/ml). Combined treatment with Trastuzumab (10 microg/ml) and CDDP (1 microg/ml) showed synergism. Our results were encouraging, and suggest a rationale for further investigations in a clinical setting.     

CD80+Gr-1+ myeloid cells inhibit development of antifungal Th1 immunity in mice with candidiasis

To find out whether polymorphonuclear neutrophils (PMN), abundantly recruited in disseminated Candida albicans infection, could directly affect the activation of Th cells we addressed the issues as to whether murine PMN, like their human counterparts, express costimulatory molecules and the functional consequence of this expression in terms of antifungal immune resistance. To this purpose, we assessed 1) the expression of CD80 (B7-1) and CD86 (B7-2) molecules on peripheral, splenic, and inflammatory murine Gr-1+ PMN; 2) its modulation upon interaction with C. albicans in vitro, in vivo, and in human PMN; 3) the effect of Candida exposure on the ability of murine PMN to affect CD4+ Th1 cell proliferation and cytokine production; and 4) the mechanism responsible for this effect. Murine PMN constitutively expressed CD80 molecules on both the surface and intracellularly; however, in both murine and human PMN, CD80 expression was differentially modulated upon interaction with Candida yeasts or hyphae in vitro as well as in infected mice. The expression of the CD86 molecule was neither constitutive nor inducible upon exposure to the fungus. In vitro, Gr-1+ PMN were found to inhibit the activation of IFN-gamma-producing CD4+ T cells and to induce apoptosis through a CD80/CD28-dependent mechanism. A population of CD80+Gr-1+ myeloid cells was found to be expanded in conventional as well as in bone marrow-transplanted mice with disseminated candidiasis, but its depletion increased the IFN-gamma-mediated antifungal resistance. These data indicate that alternatively activated PMN expressing CD80 may adversely affect Th1-dependent resistance in fungal infections.     

Reproductive performance of early-weaned female swine according to their estrus profile and frequency of artificial insemination

We determined the estrus profile (weaning-to-estrus interval (WEI), estrus duration (ED), and frequency of estrus per detection period) in 184 female swine and estimated the effect of the WEI, ED and frequency of artificial insemination (AI) on farrowing rate (FR) and litter size. Estrus detection was done at 8:30 a.m. and 5:00 p.m. The WEI was categorized as short (<100 h), medium (100-120 h) and long (>120 h). The ED was categorized as short (<60 h), medium (60-72 h) and long (>72 h). Mean lactation length was 14.6 days, mean WEI was 124.5 h and mean ED was 69 h. In each weaning group, females received either one or two AI, following a breeding schedule based on the estrus profile. In single-mated females, Al was performed 36 h after the beginning of estrus. In double-mated females, the first AI was done 24 h after the beginning of estrus and the second AI occurred 12 h later. The period of estrus detection had no effect (P > 0.05) on WEI, ED, FR, total born (TB) and live born litter size (LB). Mean FR was 82.6%, mean TB was 10.0% and mean LB was 9.2%. Mean ED was shorter (P < 0.03) for females having medium and long WEI (67.0 and 65.4 h, respectively) than for those having short WEI (72.2 h). A linear regression analysis identified a weak (R2 = 0.02) but significant negative association between ED and WEI (P = 0.05). The WEI did not influence FR (P > 0.05). Total litter size for females having short WEI (9.4) was lower (P < 0.03) than for those having long WEI (10.4). Also, LB for females having medium and long WEI (9.7-9.8) was higher (P < 0.05) than for those having short WEI (8.7). AI frequency had no effect on FR (P > 0.05). TB and LB litter size were lower (P < 0.05) for single-mated females (9.6 and 9.0, respectively) than for double-mated females (10.7 and 9.6, respectively). Double Al was associated with higher subsequent litter size. However, breeding schedules based only on estrus profile may not be precise in determining ideal breeding time, since females having short WEI had the longest ED and produced the lowest litter size.     

Structural organisation of the gene encoding the α-subunit of the human amiloride-sensitive epithelial sodium channel

The human amiloride-sensitive epithelial sodium channel (ENaC) is a member of the degenerin/ENaC family of ion channels and regulates fluid and electrolyte absorption across a number of epithelia, including kidney, colon and lung. Native ENaC has been shown to be a multimer made up of at least three homologous subunits (α, β, γ) and mutations affecting the channel complex have been identified in various human diseases. “Gain of function” mutations in one of the three ENaC subunits have been found to cause pseudoaldosteronism (Liddle’s syndrome) and ENaC “reduction of function” mutations are found in patients affected with the recessive form of pseudohypoaldosteronism (PHA) type 1. In this report, we describe the genomic organisation of the humanαENaC gene. Human αENaC consists of 13 exons spanning 17 kb on chromosome 12p13 and contains at least eight Alu sequences. In addition to the intron/exon boundaries, we have deciphered almost all the intron sequences and 475 bp of the CCAAT-less and TATA-less 5′ flanking region. Received: 23 December 1997 / Accepted: 5 March 1998     

Solution structure and stability of the full-length excisionase from bacteriophage HK022.

Heteronuclear high-resolution NMR spectroscopy was employed to determine the solution structure of the excisionase protein (Xis) from the lambda-like bacteriophage HK022 and to study its sequence-specific DNA interaction. As wild-type Xis was previously characterized as a generally unstable protein, a biologically active HK022 Xis mutant with a single amino acid substitution Cys28-->Ser was used in this work. This substitution has been shown to diminish the irreversibility of Xis denaturation and subsequent degradation, but does not affect the structural or thermodynamic properties of the protein, as evidenced by NMR and differential scanning calorimetry. The solution structure of HK022 Xis forms a compact, highly ordered protein core with two well-defined alpha-helices (residues 5-11 and 18-27) and five beta-strands (residues 2-4, 30-31, 35-36, 41-44 and 48-49). These data correlate well with 1H2O-2H2O exchange experiments and imply a different organization of the HK022 Xis secondary structure elements in comparison with the previously determined structure of the bacteriophage lambda excisionase. Superposition of both Xis structures indicates a better correspondence of the full-length HK022 Xis to the typical 'winged-helix' DNA-binding motif, as found, for example, in the DNA-binding domain of the Mu-phage repressor. Residues 51-72, which were not resolved in the lambda Xis, do not show any regular structure in HK022 Xis and thus appear to be completely disordered in solution. The resonance assignments have shown, however, that an unusual connectivity exists between residues Asn66 and Gly67 owing to asparagine-isoaspartyl isomerization. Such an isomerization has been previously observed and characterized only in eukaryotic proteins.     

EFFECTS OF VINE COMPETITION ON AVAILABILITY OF LIGHT,WATER, AND NITROGEN TO A TREE HOST (LIQUIDAMBAR STYRACIFLUA)

Competitive effects of vines on their tree hosts are well documented, but the mechanisms involved in these interactions are poorly understood. The objectives of this study were to measure the effects of below- and/or aboveground competition from the vines Lonicera japonica and Parthenocissus quinquefolia on availability of light, water, and nitrogen to the host tree Liquidambar styraciflua, and to examine the relationship between resource availability and tree growth. Light penetration through tree canopies, pre-dawn leaf water potential, and leaf nitrogen concentration were used as predictors of light, water, and nitrogen availability to the tree, respectively. Vine presence significantly reduced light penetration through the tree canopies, but this reduction was not clearly related to the growth responses of trees. Vines did not reduce the pre-dawn leaf water potential of competing trees, which was consistently above -0.5 MPa for the duration of the study. Leaf nitrogen concentration of trees, on the other hand, was significantly reduced by belowground competition with L. japonica. The positive correlation between the annual average leaf nitrogen concentration and tree diameter growth suggested that competition for nitrogen mediated the effects of belowground competition of vines on tree growth.     

Current and future distribution of a parasite with complex life cycle under global change scenarios: Echinococcus multilocularis in Europe

Global change is expected to have complex effects on the distribution and transmission patterns of zoonotic parasites. Modelling habitat suitability for parasites with complex life cycles is essential to further our understanding of how disease systems respond to environmental changes, and to make spatial predictions of their future distributions. However, the limited availability of high quality occurrence data with high spatial resolution often constrains these investigations. Using 449 reliable occurrence records for Echinococcus multilocularis from across Europe published over the last 35 years, we modelled habitat suitability for this parasite, the aetiological agent of alveolar echinococcosis, in order to describe its environmental niche, predict its current and future distribution under three global change scenarios, and quantify the probability of occurrence for each European country. Using a machine learning approach, we developed large-scale (25 × 25 km) species distribution models based on seven sets of predictors, each set representing a distinct biological hypothesis supported by current knowledge of the autecology of the parasite. The best-supported hypothesis included climatic, orographic and land-use/land-cover variables such as the temperature of the coldest quarter, forest cover, urban cover and the precipitation seasonality. Future projections suggested the appearance of highly suitable areas for E. multilocularis towards northern latitudes and in the whole Alpine region under all scenarios, while decreases in habitat suitability were predicted for central Europe. Our spatially explicit predictions of habitat suitability shed light on the complex responses of parasites to ongoing global changes.