The effect of zinc and/or vitamin E supplementation on biochemical parameters of selenium-overdosed rats.

The normotensive (Wistar) and spontaneously hypertensive (SHR) rats were examined to assess the response of the organism to selenium (Se) overdose. Moreover, the effect of zinc (Zn) and vitamin E, i.e. dietary components interacting in many biochemical processes with Se, on the Se uptake was evaluated. The control group was fed an untreated diet, and the diets of two other groups were overdosed with Se in the form of sodium selenite (9 mg/kg) and supplemented with Zn (13 mg/kg). Two experimental groups were fed a diet supplemented with Zn (13 mg/kg) and Se at an adequate level (0.009 mg/kg); a half of the animals was supplemented with vitamin E. The results showed significant differences in the Se contents between the rat strains in case of Se-overdosed groups, where in the liver and kidney tissue Se contents of SHR rats exceeded 3- and 7-fold the normotensive ones. The Se uptake was altered by the vitamin E; no effect of Zn was observed. Activities of antioxidant enzymes were determined in the animal tissues indicating different patterns according to rat strain, tissue analysed, and administered Se dose. Thus, Se overdose, for instance, via an incorrectly prepared dietary supplement, can result in serious imbalances of the biochemical status of the animals.

Carbonyl and oxidative stress in patients with breast cancer--is there a relation to the stage of the disease?

UNLABELLED: Oxidative and carbonyl stress may, on one hand, contribute to the progression of cancer, on the other hand, they may have some antiproliferative effects. We examined serum levels of AGEs (advanced glycation end-products), CML (carboxymethyllysine) and AOPP (advanced oxidation protein products) in 86 patients with breast cancer subdivided based on the clinical stage (TNM classification), histologic grading, expression of hormonal and C-erb B2 receptors and in 14 healthy age-matched women as controls. Breast cancer patients had higher serum concentrations of AGEs (325,581 +/- 66,037 vs. 271,322 +/- 34,826 AU, p < 0.01) even in the early stage of the disease; patients with advanced breast cancer (stage III and IV) had significantly higher both AGEs and AOPP (113.0 +/- 44.9 vs. 78.1 +/- 28.4 micromol/l, p < 0.05) levels, not only compared to controls, but also compared to stages I and II. Serum levels of AOPP were higher in patients having only weakly positive expression of C-erb 2/Her-neu compared to controls and the patients having the highest C-erb2/Her-neu expression. Serum concentrations of AGEs in patients with breast cancer correlated with the age and also with the serum concentration of AOPP. IN CONCLUSION: breast cancer patients had an early increase of AGEs (marker of the carbonyl stress) followed by further increase of AGEs and elevation of AOPP (marker of oxidative stress) in patients with progressive disease. As the clinical significance of these observations is currently uncertain further studies are clearly warranted, especially with respect to their potential therapeutic implications.

Speciation of Cu, Se, Zn and Fe in blood serum of hemodialysed patients.

The speciation of trace elements in serum samples of hemodialysed patients was investigated using on-line connection of SEC and ICP-MS. The 0.02 mol/l TRIS-HCl buffer of pH 7.5 was used as mobile phase. The results of speciation as well as the total concentration data were compared with those of control group of healthy person. Alterations of total concentration were observed in case of selenium and zinc only. Iron was present in form of transferrin and ferritin, main amount of copper was bound to ceruloplasmin and selenium compounds were identified as selenoproteine P and glutathione peroxidase. The latter compound was detected in samples of control group only. The chromatograms of the other elements were similar and no substantial changes between both investigated groups were observed.

Oxidative stress, metabolism of ethanol and alcohol-related diseases.

Alcohol-induced oxidative stress is linked to the metabolism of ethanol. Three metabolic pathways of ethanol have been described in the human body so far. They involve the following enzymes: alcohol dehydrogenase, microsomal ethanol oxidation system (MEOS) and catalase. Each of these pathways could produce free radicals which affect the antioxidant system. Ethanol per se, hyperlactacidemia and elevated NADH increase xanthine oxidase activity, which results in the production of superoxide. Lipid peroxidation and superoxide production correlate with the amount of cytochrome P450 2E1. MEOS aggravates the oxidative stress directly as well as indirectly by impairing the defense systems. Hydroxyethyl radicals are probably involved in the alkylation of hepatic proteins. Nitric oxide (NO) is one of the key factors contributing to the vessel wall homeostasis, an important mediator of the vascular tone and neuronal transduction, and has cytotoxic effects. Stable metabolites--nitrites and nitrates--were increased in alcoholics (34.3 +/- 2.6 vs. 22.7 +/- 1.2 micromol/l, p < 0.001). High NO concentration could be discussed for its excitotoxicity and may be linked to cytotoxicity in neurons, glia and myelin. Formation of NO has been linked to an increased preference for and tolerance to alcohol in recent studies. Increased NO biosynthesis also via inducible NO synthase (NOS, chronic stimulation) may contribute to platelet and endothelial dysfunctions. Comparison of chronically ethanol-fed rats and controls demonstrates that exposure to ethanol causes a decrease in NADPH diaphorase activity (neuronal NOS) in neurons and fibers of the cerebellar cortex and superior colliculus (stratum griseum superficiale and intermedium) in rats. These changes in the highly organized structure contribute to the motor disturbances, which are associated with alcohol abuse. Antiphospholipid antibodies (APA) in alcoholic patients seem to reflect membrane lesions, impairment of immunological reactivity, liver disease progression, and they correlate significantly with the disease severity. The low-density lipoprotein (LDL) oxidation is supposed to be one of the most important pathogenic mechanisms of atherogenesis, and antibodies against oxidized LDL (oxLDL) are some kind of epiphenomenon of this process. We studied IgG oxLDL and four APA (anticardiolipin, antiphosphatidylserine, antiphosphatidylethanolamine and antiphosphatidylcholine antibodies). The IgG oxLDL (406.4 +/- 52.5 vs. 499.9 +/- 52.5 mU/ml) was not affected in alcoholic patients, but oxLDL was higher (71.6 +/- 4.1 vs. 44.2 +/- 2.7 micromol/l, p < 0.001). The prevalence of studied APA in alcoholics with mildly affected liver function was higher than in controls, but not significantly. On the contrary, changes of autoantibodies to IgG oxLDL revealed a wide range of IgG oxLDL titers in a healthy population. These parameters do not appear to be very promising for the evaluation of the risk of atherosclerosis. Free radicals increase the oxidative modification of LDL. This is one of the most important mechanisms, which increases cardiovascular risk in chronic alcoholic patients. Important enzymatic antioxidant systems - superoxide dismutase and glutathione peroxidase - are decreased in alcoholics. We did not find any changes of serum retinol and tocopherol concentrations in alcoholics, and blood and plasma selenium and copper levels were unchanged as well. Only the zinc concentration was decreased in plasma. It could be related to the impairment of the immune system in alcoholics. Measurement of these parameters in blood compartments does not seem to indicate a possible organ, e.g. liver deficiency.

Determination of zinc in plant samples by isotope dilution inductively coupled plasma mass spectrometry.

Determination of zinc involved spiking with (68)Zn enriched solution, digestion by HNO(3)+H(2)O(2) in microwave decomposition unit, off-line separation of zinc on Chelex-100 column and measurement of ((64)Zn+(66)Zn)/(68)Zn isotope ratio on ICP-MS spectrometer with a quadrupole mass filter. After optimization of standard operation procedure (details are given) the method was validated. LOD was found to be 0.3 mug g(-1) for the procedure without zinc separation and 3.6 mug g(-1) for the procedure involving zinc separation, respectively. The accuracy of results was proved by analyses of several CRM and a primary solution of zinc, the concentration of which was verified by gravimetry and complexometric titration. Barium is the only element causing serious interferences and it must be removed from samples. The uncertainty budget is given together with the scheme of combined uncertainty calculation. The main uncertainty components are contamination during zinc separation and uncertainty of isotopic composition of natural zinc.

Custom-made interface between electrothermal vaporiser and inductively coupled plasma-mass spectrometer: determination of nickel by isotope dilution in plant samples.

ICP-mass spectrometer Perkin Elmer Elan 6000 was adapted for ETV analysis by coupling with GBC GF 3000 graphite furnace atomiser. Both instruments were connected by a 25 cm long PTFE tubing of 6 mm i.d. and a custom-made interface. The apparatus was applied to nickel determination using isotope dilution technique. The efficiency of analyte transfer through the interface was approximately 20%, however the incomplete transfer did not influence the blank values. Measured isotope ratio was very stable, whereas the magnitude of ion signal showed a little shift. CRM samples were analysed after microwave decomposition and by slurry technique as well. Obtained results were compared with those of pneumatic sample nebulisation after nickel separation on Chelex column. All tested techniques gave accurate results with comparable precision. However, the slurry technique allowed significant decreasing of the blank values and the limit of detection. The duration of analysis run can also be substantially shortened from several hours to a few minutes.

Determination of copper, selenium and zinc in human blood by inductively coupled mass spectrometry: the sources of uncertainty and variability of results.

The concentration of Cu, Se and Zn in whole human blood and plasma was analysed by inductively coupled plasma mass spectrometry. Method involved dilution of sample by diluted nitric acid and Triton X-100 and measurement of 65Cu, 77Se and 66Zn intensities. Calibration solutions were spiked by main interferents. In order to evaluate suitability of the method for recognition of alternation of trace elements concentration, the uncertainty of results was estimated. The major part of uncertainty was due to repeatability, the other source (calibration and interferences) were found to be of lesser importance. The resulting uncertainty of concentration of mentioned trace elements is low as compared to between individual variability. Short-term (within day) and long-term (between day) variability of trace elements concentration was investigated and compared with between individual variability too. The time variability was negligible except for Zn concentration. The alternations of trace elements concentration in patients with several renal diseases were investigated. Only in case of patients in end-stage renal disease: hemodialyzed patients and patient treated by continuous ambulatory peritoneal dialysis the significant alternations were proved.

Trace elements in end-stage renal disease. 1. Methodological aspects and the influence of water treatment and dialysis equipment.

For human beings trace elements are essential nutrients with a gamut of functions. They are for instance indispensable components of many enzymes, so they have some regulatory functions and they may affect immune reactions and free radical generation. Altered blood levels of different trace elements have been described in patients with advanced renal failure and especially in those treated by different kinds of renal replacement therapy. Altered renal function may result in impaired renal excretion of trace elements and their accumulation or depletion in the body. The dialysate concentrate and water used for preparing the dialysate may be an important source of the accumulation or depletion of trace elements in dialyzed patients. The gain or loss of trace elements during dialysis depends on the gradient between the ultrafiltrable fraction of a particular element in serum and its concentration in the dialysis fluid, and also on the type and permeability of the dialysis membrane. There are some methodological problems concerning the handling and storing of blood samples and measurement techniques leading to the rather inconsistent results of different studies concerning trace elements in renal disease. Geographical variations and environmental contamination of soil and water and different dietary habits may significantly influence trace elements in these patients. The abnormalities of trace elements are primarily the result of uremia, and they may be further modified and sometimes greatly exacerbated by the dialysis procedure.

Trace elements in end-stage renal disease. 2. Clinical implication of trace elements.

For human beings trace elements are essential nutrients with a gamut of functions. They are for instance indispensable components of many enzymes, so they have some regulatory functions and they may affect immune reactions and free radical generation. Abnormalities of trace elements are primarily the result of uremia, and they may be further modified and sometimes greatly exacerbated by the dialysis procedure. The role of trace elements in hemodialysis (HD) patients has not yet been fully characterized. To prevent some complications in chronic HD patients, it is very important to regulate the levels of trace elements by adequate water treatment. Reverse osmosis is able to prevent the accumulation of the majority of trace elements in the patients. Zinc supplementation may be recommended for patients with proven zinc deficiency, but for all chronic renal failure patients it is questionable. Selenium deficiency is to be suspected in dialyzed patients and selenium supplementation may be beneficial (increasing glutathione peroxidase activity, cardioprotective effect, immunostimulatory properties) for chronic renal failure patients. Supplementation with a trace element may be indicated when its depletion was unequivocally documented and when there is evidence of the positive effects of this element on the quality of life of the dialyzed patients.

Binding of lead to collagen type I and V and alpha2(I) CNBr (3,5) fragment by a modified Hummel-Dreyer method.

Binding of lead (as lead acetate) to collagen type I alpha, and alpha2 chains, collagen type V and a large cyanogen bromide fragment of type I collagen [alpha2(I)CB(3,5)] was investigated by the large-zone Hummel-Dreyer method. It was demonstrated that two categories of binding sites exist in the collagen molecule, the number of which correlates rather well with the available aspartic and glutamic acid residues. Similar results were obtained for all collagen chains (fragments) used. The number of sites thus obtained was compared with the cross-striation pattern (reflecting areas where lead is bound) of the SLS form of collagen type I (alpha1 chain); it is suggested that the number of bands seen in the SLS form reflects primarily the number of available aspartic acid residues in the molecule. The association constants obtained are comparable with the low affinity interactions seen e.g., between Cu and bovine serum albumin.

Chromium levels in patients with internal diseases.

Chromium (Cr), an essential element, mainly affects saccharide (potentiated insulin action via interaction with insulin receptor on the cell surface) and lipid metabolism (inhibition of hydroxymethylglutaryl-CoA reductase with a hypolipidemic effect). The aim of the study was to describe Cr serum levels in different diseases (malignant, metabolic, renal) using an advanced analytical technique with correlation to other biochemical parameters. The concentration was measured using atomic absorption spectrometry with electrothermal atomization. The Cr levels were increased in hemodialysis patients-HD (3.67 +/- 0.35 micrograms/L) compared to controls-C (0.40 +/- 0.12 microgram/L), in significantly changed in diabetic patients-DM (0.29 +/- 0.08 microgram/L) and patients with lymphoproliferative disease-LP (0.24 +/- 0.07 microgram/L), and decreased in hyperlipidemic patients-HL (0.15 +/- 0.03 microgram/L). There were no differences in Cr concentration between DM treated by diet or peroral antidiabetic drugs; likewise hypolipidemic drugs in HL did not change the Cr concentration. The biochemical parameters-total protein, transferrin in LP group, glucose in DM group, total cholesterol, triacylglycerols, LDL-cholesterol, apolipoprotein B and A-I did not correlate with serum Cr concentration. However, the HDL-cholesterol concentration marginally significantly (p < 0.07) correlated with it. The role of Cr in humans has not yet been fully characterized. To prevent some complications in patients, it may be important to monitor the Cr levels. Chromium supplementation may be indicated in some diseases with no controversy concerning the importance of decreased serum and/or tissue levels and documented positive effects of Cr supplementation on the quality of life (e.g. hyperlipidemia).

Accumulation of lead in tissues after its administration in drinking water to laboratory rats.

Lead administered to laboratory rats in drinking water (0.1-0.8%) as lead acetate solution tends to accumulate in collagen-rich tissues such as tendons and the skin. The amount of lead deposited (and also zinc present in the tissue without its supplementation) correlates with the blood supply to the tissue investigated. The highest deposits of lead were observed in placenta and chorionic membranes, though here only about 60% are collagen-bound. No differences in the drinking habits of the animals were observed and also at lower concentrations of lead in the drinking water no dose dependence was revealed. However, at 0.8% of lead in drinking water considerable accumulation of lead was observed in all tissues investigated.

[Linear discrimination analysis in blood pressure monitoring]. / Vyuzití lineární diskriminacní analýzy pri monitorování krevního tlaku.

The 24-hours blood pressure monitoring (BPM) is a reliable technique largely used for hypertension diagnosis in clinical practice. Presence of the lack of a nocturnal decrease in blood pressure values is one differentiating criterion between essential and secondary hypertension. New useful method of mathematical and statistical processing of BPM data using the linear discriminant analysis (LDA) was described. This statistical analysis was applied for 123 BPM measurements from patients with essential hypertension without and with different kidney diseases including hemodialyzed patients. The control group was normotensive healthy volunteers. The LDA method successfully separate group of patients with essential hypertension from patients with secondary renal hypertension but the error of return ranking to these groups was 28.5%. The recalculation for groups-controls, hypertensive patients and hypertensive hemodialyzed patients leads to decreasing of the error of return ranking to 5.7%. This retrospective study of nonhomogeneous groups could not clearly differentiate groups of hypertensive patients. The new prospective study on accurately definite groups of patients can eliminate these problems.

Trace elements in hemodialysis and continuous ambulatory peritoneal dialysis patients.

Alterations in blood and tissue concentrations of trace elements in patients with chronic renal failure have been extensively investigated. Selenium, zinc and copper are elements which play an important role in biological systems as components of proteins, enzymes and antioxidants. The concentrations of selenium, zinc and copper were determined in the plasma, erythrocytes and whole blood of patients on regular hemodialysis (HD) and continuous ambulatory peritoneal dialysis (CAPD) treatment using the method of inductively coupled plasma mass spectrometry (ICP-MS). Analysis of isotopes 77Se, 66Zn and 65Cu was performed. Methodology presents the major limitation to valid studies on trace element levels in biological materials. One of the widely used contemporary techniques is ICP-MS. It is the most sensitive one and has a high dynamic range. The selenium concentration in the studied compartments (plasma 46.1 +/- 3.0 vs. 78.0 +/- 3.4 microgram/l, p < 0.001; erythrocytes 90.4 +/- 6.5 vs. 134.2 +/- 7.6 microgram/l, p < 0.01; whole blood 67.3 +/- 3.1 vs. 106.4 +/- 3.4 microgram/l, p < 0.001) was significantly lower in HD patients compared to healthy controls. The same result was observed in plasma (63.2 +/- 5.8 vs. 78.0 +/- 3.4 microgram/l, p < 0.05) and whole blood (82.7 +/- 7.4 vs. 106.4 +/- 3.4 microgram/l, p < 0.01) from CAPD patients, but the selenium level of erythrocytes in CAPD patients was the same as in the control group (126.0 +/- 8.8 vs. 134. 2 +/- 7.6 microgram/l). The cooper content of erythrocytes was lower in HD patients than in controls (0.55 +/- 0.02 vs. 0.66 +/- 0.01 mg/l, p < 0.01) and CAPD groups (0.55 +/- 0.02 vs. 0.68 +/- 0.02 mg/l, p < 0.001). There were no differences in copper content in plasma (HD 1. 02 +/- 0.06; CAPD 1.11 +/- 0.09; controls 1.02 +/- 0.05 mg/l) and whole blood (HD 0.87 +/- 0.04; CAPD 0.90 +/- 0.05; controls 0.85 +/- 0.02 mg/l) in HD and CAPD patients and healthy controls. The zinc concentration was increased in the whole blood of CAPD patients (6. 68 +/- 0.36 vs. 5.52 +/- 0.11 mg/l, p < 0.001) and erythrocytes of HD (12.30 +/- 0.23 vs. 10.11 +/- 0.42 mg/l, p < 0.001), and CAPD groups (13.71 +/- 0.56 vs. 10.11 +/- 0.42 mg/l, p < 0.001) compared to controls. However, the plasma zinc concentration was lower in HD patients compared to blood donors (0.69 +/- 0.03 vs. 0.92 +/- 0.03 mg/l, p < 0.001) and CAPD patients (0.69 +/- 0.03 vs. 0.95 +/- 0.04 mg/l, p < 0.001). We did not find a significant increase in trace elements in whole blood after HD. These results suggest differences between plasma, erythrocytes and whole blood concentrations of the studied trace elements. The levels of trace elements are altered by HD and CAPD. A modern precise method with high accuracy, ICP-MS, which was used in our study, eliminated analytical errors and possible interferences.

Application of infrared spectroscopy to the assessment of authenticity of tea.

Diffusion reflectance fourier transform infrared spectroscopy in the mid-IR region was used to assess the authenticity of tea varieties. The differences between the spectra of 12 different tea varieties (seven black, two green, three semifermented grades) were sufficient to allow their discrimination by the soft independent modelling of class analogy classification method or linear discriminant analysis, despite a significant heterogeneity of the samples as revealed by variance analysis.

[Use of multivariate analysis in medical chemistry]. / Vyuzití vícerozmerné analýzy rozptylu v lékarské chemii.

Alternations in trace elements concentrations are intensively studied because of their possible role in pathogenesis and progress of diseases. Three groups of patients were investigated: haemodialysis patients (HD) (n = 17), continual ambulatory peritoneal dialysis patients (CAPD) (n = 11), and control group of blood donors (n = 12). They were analyzed for Se, Zn and Cu concentration in plasma and erythrocytes by inductively--coupled plasma mass spectroscopy. Distribution of analyzed elements between these blood compartments was found different in all groups of studied patients. Erythrocytes were enriched by Se (twofold higher concentration compared to plasma) and by Zn (10-20 fold higher concentrations compared to plasma). On the other hand, human plasma was enriched by Cu (approximately twofold higher concentration then in erythrocytes). Results of analyses were processed by multivariate analysis of variance (MANOVA). When only results of plasma analysis were involved into MANOVA, differences between HD patients and other two groups were found whereas CAPD and control group were not distinguished each other. However, these two groups were mutually differenced when MANOVA comprised all trace elements concentrations: both plasma and erythrocytes. Methods of multivariate statistic are able to study not only the individual variables but even their mutual relations and their medical applications are very useful.