Targeted natural product isolation guided by HPLC-SPE-NMR: constituents of Hubertia species

The hyphenated technique, high-performance liquid chromatography-solid-phase extraction-nuclear magnetic resonance spectroscopy (HPLC-SPE-NMR), has been applied for rapid identification of novel natural products in crude extracts of Hubertia ambavilla and Hubertia tomentosa. The technique allowed full or partial identification of all major extract constituents and demonstrated the presence of unusual quinic acid derivatives containing the (1-hydroxy-4-oxocyclohexa-2,5-dienyl)acetyl residue that exhibit strongly coupled ABXY patterns, the parameters of which were obtained by spin simulations. Using homo- and heteronuclear 2D NMR data acquired in the HPLC-SPE-NMR mode, complete structure determination of three new natural products, i.e., 3,5-di-O-caffeoyl-4-O-[(1-hydroxy-4-oxocyclohexa-2,5-dienyl)acetyl]quinic acid (1), its 2-hydroxy derivative (2), and 3,5-di-O-caffeoyl-4-O-[(4-hydroxyphenyl)acetyl]quinic acid (3), was performed. Finally, targeted isolation of 1 was achieved by SPE fractionation and preparative HPLC, followed by evaluation of its antioxidant and antimicrobial activity. In contrast to chlorogenic acid and 3,5-di-O-caffeoylquinic acid, which act as antioxidants, compound 1 proved at the same conditions to possess prooxidant activity in an assay evaluating the oxidation of human low-density lipoprotein induced by Cu(2+).     

Spine bone mineral density and VDR polymorphism in subjects with ulcerative colitis

This study established bone mineral density in subjects with ulcerative colitis with respect to disease dissemination and severity and the association between skeletal status and vitamin D receptor (VDR) polymorphism. Forty-seven patients aged 47.6 ± 14.8 years and 47 age- and sex-matched control subjects were evaluated. Disease duration was 8.6 ± 7.2 years. Twenty-four subjects demonstrated mild, 17 moderate, and 5 severe forms of ulcerative colitis; local (proctitis and proctosigmoiditis) changes were present in 26 and disseminated changes in 21. Bone mineral density (BMD, g/cm²) was assessed at the spine, and distribution of VDR polymorphism was established. In six patients (12.8%) and in two controls (4.25%), T-score for BMD was below −2.5, but mean values of BMD did not differ between all patients and controls. Patients with moderate and severe form of disease had lower BMD measurements than patients with a mild form of colitis ulcerosa (P < 0.05), and subjects with disseminated intestinal changes had lower BMD measurements than subjects with local changes (P < 0.001). Distribution of VDR polymorphism did not differ between patients and controls. Spine Z-score was dependent on VDR polymorphism (P < 0.05) in male and female patients but not in controls. We concluded that, in patients with ulcerative colitis (UC), spine bone mineral density decreases with progression and dissemination of the disease, and that VDR polymorphism is associated with spine bone mineral density. VDR genotype bb is significantly less likely to cause low BMD in male UC patients, and VDR genotype tt is more likely to cause low BMD in female patients.     

The Metric Features of Teeth in Female-to-Male Transsexuals

The aim of the study was to evaluate the odontometric characteristics of female-to-male transsexuals (FtM) in comparison to control groups of males and females. A total of 48 FtM, aged 20–28 years, were studied. For each tooth, two standard measurements were taken: the mesiodistal (MD) and the buccolingual (BL) diameter of the crown. Significant differences between males and females were found in BL measures of all maxillary teeth as well as mandibular lateral incisor and canine. Within the MD measures, the most distinguishing were maxillary and mandibular canines and the first molars. For seven teeth, there were a total of 14 comparisons of MD (half for the maxilla and half for the mandible) between the control men and women. Of these, the men had larger diameters for 12, of which four were statistically significant. There were also a total of 14 comparisons of BL diameter; of these, the men had larger diameters for 13, of which nine were statistically significant. For seven teeth, there were a total of 14 comparisons of MD between the control men and the FtM. Of these, the men had larger diameters for 12, of which seven were statistically significant. There were also 14 comparisons of BL; of these, the men had larger diameters for 10, of which six were statistically significant. For seven teeth, there were a total of 14 comparisons of MD between the control women and the FtM. Of these, the women had larger diameters for nine, of which three were statistically significant. There were also a total of 14 comparisons of BL; of these, the FtM had larger diameters for 13, of which five were statistically significant. Our study revealed that the shift of values of metric features in teeth of FtM towards males was more frequent in the more sexually dimorphic BL diameters characterized by significantly greater width of the crown of five teeth: maxillary canine, first and second molars, as well as mandibular incisors. The results showed intermediate status of female-to-male transsexuals’ teeth between the males and females, which may suggest a genetic basis of transsexualism.     

Alcohol dehydrogenase isoenzymes and aldehyde dehydrogenase activity in the sera of patients with esophageal cancer

Various alcohol dehydrogenase (ADH) isoenzymes and aldehyde dehydrogenase (ALDH) exist in the human esophageal mucosa. In our last experiments we have shown that ADH and ALDH are present also in the esophageal cancer cells. Moreover, the activities of total ADH and class IV isoenzymes were significantly higher in cancer tissue than in healthy mucosa, which suggests that these changes may be reflected by enzyme activity in the serum. Therefore, we measured the activity of total alcohol dehydrogenase, and classes I–IV of this enzyme and aldehyde dehydrogenase in the sera of patients with this cancer. Serum samples were taken for routine biochemical investigation from 67 patients with esophageal cancer before treatment. Total ADH activity was measured by photometric method with p-nitrosodimethylaniline (NDMA) as a substrate and ALDH activity by the fluorometric method with 6-methoxy-2-naphtaldehyde as a substrate. For the measurement of the activity of class I and II isoenzymes, we employed the fluorometric methods, with class-specific fluorogenic substrates. The activity of class III alcohol dehydrogenase was measured by the photometric method with formaldehyde and class IV with m-nitrobenzaldehyde as a substrate. A statistically significant increase of class IV alcohol dehydrogenase isoenzymes was found in the sera of cancer patients. The median activity of this class isoenzyme in the total cancer group increased by about 26.5% (7.42 mU/l) in comparison to the control level (5.46 mU/l). The total alcohol dehydrogenase activity was significantly higher (30%) among patients with cancer. The activities of other tested ADH isoenzymes and total ALDH were unchanged. The activity of the class I ADH isoenzyme was significantly higher in the sera of drinkers with esophageal cancer than non-drinking patients. The increased total activity of alcohol dehydrogenase and class IV isoenzyme in the sera of patients with esophageal cancer probably can be caused by release of this isoenzyme from cancer cells or might be stimulated by alcohol drinking.     

Human gastrointestinal nematode infections: are new control methods required?

Gastrointestinal (GI) nematode infections affect 50% of the human population worldwide, and cause great morbidity as well as hundreds of thousands of deaths. Despite modern medical practices, the proportion of the population infected with GI nematodes is not falling. This is due to a number of factors, the most important being the lack of good healthcare, sanitation and health education in many developing countries. A relatively new problem is the development of resistance to the small number of drugs available to treat GI nematode infections. Here we review the most important parasitic GI nematodes and the methods available to control them. In addition, we discuss the current status of new anthelmintic treatments, particularly the plant cysteine proteinases from various sources of latex-bearing plants and fruits.     

Concentrations of soluble Fas and soluble Fas ligand as indicators of programmed cell death among patients coinfected with Human Immunodeficiency Virus and Hepatitis C Virus

Hepatitis C Virus (HCV) and Human Immunodeficiency Virus (HIV) coinfections can affect mechanisms of programmed cell death and therefore influence acquired immunodeficiency syndrome (AIDS) development as well as the course of chronic hepatitis C. The aim of the study was to assess soluble Fas (sFas) and soluble Fas ligand (sFasL) concentrations in HIV- and HCV-coinfected patients and, moreover, to establish their relationships with HIV viral load, CD4+ T lymphocyte count, as well as liver function tests. Seventy-eight patients were included in the study, among them 30 coinfected with HIV and HCV, 10 infected only with HIV, and 38 infected only with HCV. HIV infection was confirmed by means of Western blot analysis; HIV viral load was measured by RTPCR; and CD3+, CD4+, and CD8+ T lymphocyte counts were established by means of flow cytometry. HCV infection was confirmed through HCV RNA isolation, using RT-PCR. sFas and sFasL concentrations were measured in duplicate by ELISA. The mean CD4+ T lymphocyte count decreased in HIV- and HCV-coinfected patients versus HIV-infected individuals (429 versus 279/ml). sFasL protein was detectable principally in HIV-infected individuals without HCV infection (90%), whereas in those with HCV infection it occurred only in 11% of cases. The highest sFas concentration was observed in HCV-infected patients (25.9 ng/ml) as well as in HIV- and HCV-coinfected individuals (20.3 ng/ml). This concentration was negatively proportional to sFasL prevalence. The results of our study suggest that HCV infection in HIV-positive individuals may suppress processes of programmed cell death. There was no correlation between sFas, sFasL, and HIV-1 viral load. On the other hand, sFas concentration and the presence of sFasL were related to CD4+ T lymphocyte count.     

Biochemical background of the VO2 on-kinetics in skeletal muscles

This review discusses the present knowledge on the oxygen uptake kinetics at the onset of exercise in skeletal muscle and the contribution of a previously developed computer model of oxidative phosphorylation in intact skeletal muscle to the understanding of the factors determining this kinetics on the biochemical level. It has been demonstrated recently that an increase in the total creatine pool [PCr + Cr] and in glycolytic ATP supply lengthen the half-transition time of the VO2 on-kinetics, while an increase in mitochondria content, in parallel activation of ATP supply and ATP usage, in muscle oxygen concentration, in proton leak, in resting energy demand, in resting cytosolic pH, and in initial alkalization diminish this parameter. It has also been shown that the half-transition time is near-linearly proportional to the absolute difference between the phosphocreatine concentration during work and at rest (deltaPCr). The present review discusses whether the V/O2 on-kinetics on the muscle level is strictly or only approximately exponential. Finally, it is postulated that a short transition time of the VO2 on-kinetics in itself does not need be profitable for the skeletal muscle functioning during exercise, but usually a short transition time is correlated with factors that improve exercise capacity. The transition time is a phenomenological parameter resulting from the biochemical properties of the system and not a physical factor that can cause anything in the system.     

Fine-needle aspiration cytology combined with flow cytometry immunophenotyping is a rapid and accurate approach for the evaluation of suspicious superficial lymphoid lesions

The authors report a prospective study on 88 samples of superficial lesions (lymph nodes, skin nodules, and breast tumors), performed by fine-needle aspiration cytology (FNAC) in 74 patients, allowing (i) morphologic analysis combined with immunophenotyping by flow cytometry (FCM) and (ii) a cytogenetic study in 33 cases. Thirty-nine FNAC (44.3%) were performed at the time of diagnosis. The cytology results were correlated with histopathologic examination in 32 cases. Forty-nine FNAC (55.7%) were performed in the context of follow-up of a lymphoma and the results were correlated with those of histopathologic examination in 14 cases.In this study, the concordance between FNAC plus FCM and histopathologic examination was 90% for low-grade non-Hodgkin's malignant lymphomas (NHLs) and 83% for high-grade NHL. The limits of this morphologic and phenotypic approach are (i) partial tumor infiltrations, (ii) Hodgkin lymphoma, and (iii) T-cell NHL.In conclusion, it may be said that this combined approach is very useful for diagnosis and follow-up of patients but requires teams experienced in the sampling technique and the morphologic diagnosis of the various types of low-grade NHL in which supplementary ancillary studies may be performed when morphology and flow cytometry immunophenoyping are not conclusive.     

Synthesis and microbiological activity of thiourea derivatives of 4-azatricyclo[5.2.2.0<Superscript>2,6</Superscript>]undec-8-ene-3,5-dione

A series of thiourea derivatives of 4-azatricyclo[5.2.2.0^2,6]undec-8-ene-3,5-dione were synthesized. The compounds were investigated for antibacterial activity, including Gram-positive cocci, Gram-negative rods, and antifungal activity. Compounds 1b, 2b, 4b showed significant inhibition against Gram-positive cocci. Research was carried out over 10 standard strains and 20 hospital strains. Synthesized compounds were evaluated for their cytotoxicity and anti-HIV-1 activity in MT-4 cells.