Biomedical subjects

Z Deyl

Publications and source records attributed to Z Deyl.

At least 19 recordsLinked to original sources

PubMed1995

Capillary electrophoresis of hair proteins modified by alcohol intake in laboratory rats.

A capillary zone electrophoretic method was used to obtain profiles of solubilized rat hair keratin proteins. The same methodology was used to reveal the presence of additional protein peaks in alcohol-consuming rats. Two types of separation were investigated. Alkali-solubilized keratins from hair of rats treated for 5 weeks with 5% ethanol and 2 weeks with 10% ethanol (instead of drinking water) and from controls were analysed. Whereas under alkaline conditions (pH 9.2, 50 mM borate) an additional fraction of "low-sulphur" keratins with the highest anodic mobility of this keratin category was shown in alcohol-treated animals, acid electrophoresis carried out at pH 3.5 in phosphate buffer (50 mM) revealed the presence of two sharp peaks absent in the controls. These findings were confirmed by two-dimensional separations of carboxymethylated keratin samples. An attempt was made to identify further one of the newly occurring fractions in alcohol-consuming animals. It was revealed that the tryptic hydrolysate of "low-sulphur" proteins obtained from alcohol-consuming animals contained a peptide not found in controls.

Alcohol Drinking↗

PubMed1995

Capillary electrophoresis: a new tool in forensic toxicology. Applications and prospects in hair analysis for illicit drugs.

Capillary electrophoresis, the modern approach to instrumental electrophoresis, is probably the most rapidly expanding analytical technique that has appeared in recent years. In the hands of forensic toxicologists, capillary electrophoresis (CE) represents a powerful new analytical tool, which has proved suitable for the investigation of illicit drugs in seized preparations and also in complex biological matrices, among which is hair. CE can be applied according to different separation mechanisms, and among those that are toxicologically relevant are capillary zone electrophoresis and micellar electrokinetic capillary chromatography, which display different selectivities. For the investigation of hair for drugs of abuse, capillary electrophoresis proved effective, providing simultaneous determinations of different drugs without derivatization, with acceptable sensitivity (typically better than 1 ng of drug per mg of hair). The possibility of carrying out determinations of the same analytes, based on different separation mechanisms (capillary zone electrophoresis and micellar electrokinetic chromatography) with the same instrumentation, simply changing the buffer composition, provides an interesting possibility of 'internal' confirmation of the results.

Electrophoresis↗

PubMed1994

General strategies and selection of derivatization reactions for liquid chromatography and capillary electrophoresis.

The general strategies, reasons and the different possibilities for the derivatization of biomedically important compounds are reviewed. Different approaches apply for small versus large analyte molecules, different advantages and disadvantages are visualized with pre- and post-column arrangements. Particular interest is focused upon solid-phase derivatization reagents.

Chromatography, High Pressure Liquid↗

PubMed1994

Biomedical applications of capillary electrophoresis.

After having grown through the stages of technique development and instrumentation availability, capillary electrophoresis has reached the stage of applications. This review attempts to show the potential of this technique for biomedical analysis. Rather than going into a detailed description of the technical details of the separation conditions suitable for the separation of a particular category of compounds, the focus is on the general principles and areas in which this technique can be applied and the prospects for the future. Particular emphasis is placed on the separation of complex matrices and their simplification, a daily task in biomedical laboratories. In addition, methods for the optimization of separation conditions are considered. Considerable prospects for capillary electrophoresis lie in profiling. The applicability of the technique in peptide and protein mapping is discussed in some detail. Finally, three other topics are dealt with, namely enzymic activity microassays, drug-protein binding assays and monitoring of drugs in body fluids.

Animals↗

PubMed1994

Capillary electrophoretic profiling of rat hair: a tool for alopecia areata diagnosis.

Capillary electrophoretic profiling of hair fractions obtained by 0.25 M HCl treatment of the tissue with subsequent extraction of the solubilized fraction with chloroform-isopropanol (9:1, v/v) revealed clear differences between hair obtained from alopecia areata affected laboratory rats (both in hair obtained from non-affected areas and hair growing on once hairless patches) and controls. Differences were observed both in the organic-phase extractable material as well as in the aqueous phase after extraction. The separations were carried out in 25 mM borate buffer pH 9.2 for the chloroform-isopropanol extractable fraction while profiling of the aqueous phase was done in the same buffer at pH 10. Untreated fused-silica capillaries, 40-45 cm to the detector of I.D. 50 and 75 microns were used at a running voltage of 20 kV. Detection was done either by UV absorbance at a fixed wavelength of 200 nm or by using a diode array detector.

Alopecia Areata↗

PubMed1994

Age- and feeding-dependent production of carbonyl compounds in hypoxic heart. The role of carbonyls produced in connective tissue modification.

The concentration of reactive lipid metabolites (malondialdehyde, formaldehyde, acetaldehyde and acetone) was assayed in rat heart reperfusates after 30 min ischemia in animals of different age and kept on different feeding regimes. It was revealed that there is no difference in the concentration of reactive carbonyl compounds in reperfusates from animals of different age, but the amount of released carbonyl compounds is much lower in animals kept on 50% restricted diet. If tail tendons from young (3 months) rats are incubated in the reperfusate, their solubility after CNBr treatment is decreased so that this material resembles tendons from old animals. Also the amino acid composition of the insoluble residue cannot be distinguished from that obtained from rat tail tendons of 24- or 29-month-old rats. The results prove the ability of carbonyl containing lipid metabolites to create a CNBr-insoluble core in connective tissue.

Acetone↗

PubMed1992

Silver staining of collagen type I after sodium dodecylsulphate polyacrylamide gel electrophoresis: effect of Maillard reaction.

Differences in the acidic silver staining, after sodium dodecylsulphate polyacrylamide gel electrophoresis, between the alpha 1 and alpha 2 collagen chains, as well as between rat-tail tendon and calf-skin collagen type I, were observed. The staining conditions at which the staining differences are both most expressed and reproducible were characterized. Age differences between staining of the alpha 1 CB6 fragment from young rats (2 and 12 months) and old rats (29 months) indicated that different susceptibilities of collagen species to the silver staining can be the result of different extents of some age-dependent post-translational modification, such as glycation. In vitro incubation of acid-soluble rat-tail tendon collagen with various sugars led to an increase in silver staining compared with samples incubated in the absence of sugar. This effect was inhibited by sodium cyanoborohydride, diethylenetriamine pentaacetic acid and aminoguanidine, i.e. compounds inhibiting the Maillard reaction at various stages. It can be concluded that the enhanced silver susceptibility of glycated collagen is related to advanced-phase Maillard reaction products attached to collagen.

Animals↗

PubMed1992

Altered ratio of collagen chains in bone of a patient with non-lethal osteogenesis imperfecta.

Bone from a patient with osteogenesis imperfecta contained type III collagen which was absent in control bone. The ratio of alpha 1(I)/alpha 2(I) in type I collagen of patient's bone was increased (2.9 vs. 2.3 +/- 0.2 in controls) and the ratio of dimers beta 11/beta 12/beta 22 was altered due to the increased beta 22 content. No abnormality was observed in collagen from the patient's skin. The altered composition of collagen in bone, but the normal composition in skin suggests that the disease in the patient is due to impaired regulation of the synthesis of collagens in bone, rather than by a mutation in one of the two type I collagen genes. Unlike in skin, all the type III collagen in patient's bone was pepsin-soluble indicating an inability of the bone to incorporate type III collagen into mature highly cross-linked extracellular matrix.

Bone and Bones↗

PubMed1991

Capillary zone electrophoresis: its applicability and potential in biochemical analysis.

Recent developments in capillary zone electrophoresis (CZE) are reviewed, starting with available instrumentation, a description of different operational modes and the most commonly used detection systems. Appropriate attention is paid to CZE-mass spectrometry coupling and coupling of electrophoretic and chromatographic procedures. The possibility of separating chiral molecules is also discussed. Examples of applications concern mainly amino acids, peptides, proteins, nucleic acids and their constituents.

Amino Acid Sequence↗

PubMed1991

Separation of elastin cross-links as phenylisothiocyanate derivatives.

A method has been developed for the separation and quantitation of desmosines in tissue samples. The tissue is treated with cold 10% trichloroacetic acid to remove collagen and hydrolysed in HCl vapours in sealed vials. Preseparation of desmosines from tissue acid hydrolysates is performed on a cellulose column, first eluted with n-butanol-acetic acid-water to wash out other amino acids and then with water to recover desmosines. Separated desmosines are then derivatized with phenylisothiocyanate and determined by reversed-phase high-performance liquid chromatography using a gradient system with sodium acetate pH 6.4 and acetonitrile. Desmosines were detected spectrophotometrically at 254 nm. The method was applied to the determination of desmosine in elastin, rat aorta and bovine ligamentum nuchae.

Animals↗

PubMed1991

Change with age of UV absorbance and fluorescence of collagen and accumulation of epsilon-hexosyllysine in collagen from Wistar rats living on different food restriction regimes.

Accumulation of glycation products (as revealed by the thiobarbituric test and hexosyllysine assay) and the pigmented products (350 nm UV absorbance and 370ex/440em nm fluorescence) in aortal and skin collagen was investigated under the conditions of different nutritional regimes. Four groups of animals were tested: (1) ad libitum fed controls, (2) animals which were food restricted throughout their whole life (50% food intake), (3) animals fed ad libitum during their first year of life and then food restricted and (4) animals food restricted when young and fed ad libitum from the age of 1 year onwards. It was shown that all food-restricted animals showed lower levels of glycation and pigmentation products in collagen preparations from skin and aorta. The lowest accumulation was observed in group 4 which exhibited the longest 50% survival (29.4 months, as compared with 18.3 months in normally-fed controls). Of particular interest is the fact that in this group the decreased rate of accumulation of the glycated and pigmented products was preserved even after 1 year of life, i.e., when the animals had a free access to food. Though not directly supporting the glycation theory of aging (Cerami, 1985), our data are indicative of the involvement of glucose metabolism in the ageing process. Correlation between the levels of glycated and pigmented products in aortal and skin collagen as well as the correlation between the rate of accumulation of these products and 50% survival was impossible to establish. Nevertheless, each time that food restriction was imposed on the animals it always resulted in decreased accumulation of glycated and pigmented products and increased 50% survival. Possible mechanisms for this process are discussed.

Aging↗

PubMed1991

Change in the amount of epsilon-hexosyllysine, UV absorbance, and fluorescence of collagen with age in different animal species.

Skin and aorta collagen specimens of Wistar rats, white mice, beagle dogs, cats, horses, and human necropsies of different ages were examined with respect to the content of glycated products. The data presented show that (a) glycation and accumulation of the chromophore(s) are comparable in collagen samples from different species of comparable age; (b) glycation and pigmented accumulation increase markedly during the first 5-10 years of age; (c) the extent of glycation is different in different tissues (in particular, glycation of aortal collagen is about twice that of skin collagen); and (d) collagen pigmentation as followed by fluorescence is comparable in aortal and skin collagen (except below 10 years); pigmentation measured by absorbance at 350 nm is, on the contrary, lower in aortal than in skin collagen. Based on the assumption of constant blood glucose level during the life span, it appears feasible to conclude that the degree of nonenzymatic collagen glycation reflects the time period for which the protein was exposed to the action of sugars. This period, because of increased cross-linking, is likely to be extended in older animals. Other factors, such as differences in collagen turnover between different tissues and the intensity of the removal process of the glycated products, should be taken into consideration as well.

Aging↗

PubMed1990

Separation and partial characterization of Maillard reaction products by capillary zone electrophoresis.

Capillary zone electrophoresis proved useful for separating small amounts of both charged and uncharged solutes that are otherwise difficult to analyse. A typical complex mixture that had previously resisted all analytical approaches, including reversed-phase separations, is the products arising from the reaction of free amino acids with aldehydic sugars (Maillard reaction products). By using capillary zone electrophoresis [untreated capillary 50 cm x 75 microns I.D., 18 kV, 0.02 mol/l phosphate buffer (pH 7.5)], a number of products resulting from the reaction of glucose or ribose with glycine, alanine and isoleucine were separated and partially characterized. They were separated (1) without derivatization (and profiles of compounds absorbing at 220 nm were obtained), (2) as phenylthiocarbamyl derivatives in a search for reactive amino groups and (3) after derivatzation with 2,4-dinitrophenylhydrazine in a search for a method for compounds with a free aldehydic group. Phenylthiocarbamyl derivatives were separated in 0.005 mol/l borate buffer (pH 9.6) at 20 kV and 25 microA. Separation of 2,4-dinitrophenylhydrazones was effected by electrokinetic micellar chromatography in the same apparatus using a 50 cm x 75 microns I.D. capillary at 10 kV in 0.01 mol/l Na2HPO4-0.006 mol/l tetraborate, 0.050 mol/l with respect to sodium dodecyl sulphate. The results are compared with those given by high-performance liquid and thin-layer chromatography.

Amino Acids↗

PubMed1990

Increased glycation and pigmentation of collagen in aged and young parabiotic rats and mice.

Changes of non-enzymatic collagen glycosylation were followed in 2- and 24-month-old rats and mice and in parabiotic animals of the same age. With advancing age increased glycation of collagen was observed in both old male Wistar rats and white mice. Further it was demonstrated that both aortal and skin collagen of young animals is rapidly non-enzymatically glycosylated in the common milieu created by parabiotic animals and the proportion of non-enzymatically incorporated glucose approaches in the young counterparts the level found in old individuals. Similar trends as with non-enzymatic glycosylation were found with a fluorescent (370/440 nm) product present in both categories of collagen preparations. This fluorescence was higher in old animals and was considerably increased in the young counterpart of the parabiotic couple 6 weeks after operation. The nature of the fluorescent product appears different from pyridinoline and remains to be elucidated.

Aging↗

PubMed1990

[The effect of chronic exposure to lead on lysosomal levels in amniotic fluid in an experimental model].

Groups of female rats were exposed for prolonged periods to different lead concentrations in drinking water; in one group was also added to the drinking water, in the amniotic fluid of exposed females greatly reduced lysozyme values were found, as compared with controls. Concurrent Zn administration considerably reduced the effect of lead. Long-term exposure to low concentrations of lead caused a greater drop of lysozyme levels in amniotic fluid than that found in previous experiments after short-term exposure to higher lead concentration.

Amniotic Fluid↗

PubMed1989

Separation of collagens by capillary zone electrophoresis.

Collagen (types I, II, V, IX and XI) constituting polypeptide chains and their polymers and cyanogen bromide-cleaved peptides of collagen type I and type III were investigated by means of capillary zone electrophoresis. Separations were effected in 2.5 mM sodium tetraborate buffer in less than 15 min. A 50 cm x 0.1 mm I.D. fused-silica capillary was used. The separations were run at 18 kV per capillary. The results of the separation were monitored at 220 nm with an on-tube detection system. Using the Offord equation, relative retention times of cyanogen bromide cleavage fragments were plotted against M(2-3)/Z, where M is the molecular mass of a polypeptide and Z its valency. A linear relationship was observed. Collagen alpha-chains and their polymers were also satisfactorily resolved.

Collagen↗

PubMed1989

Simple apparatus for capillary zone electrophoresis and its application to protein analysis.

The construction of a simple apparatus for capillary zone electrophoresis is described, consisting of an optical system allowing direct absorbance measurement in the capillary in UV light, an evaluating electronic module and a high-potential source. An attempt was made to achieve maximum sensitivity with a simple construction. In the electronics, care was taken to obtain a quiet baseline and to optimize the signal-to-noise ratio. That part of the noise which is of the frequency band of the signal is filtered off. Both suction and electrophoretic sample introduction are possible. According to operator's choice, the apparatus can be run under constant voltage or current and is protected against overloading. The high-potential electrode chamber contains separate buffer and sample compartments and its construction offers an easy interchange between the running and sampling positions. The applicability of the system to the separation of amino acids as phenylthiocarbamyl derivatives, peptides and both artificial and naturally occurring protein mixtures is demonstrated.

Adsorption↗

PubMed1989

Separation methods for the study of collagen and treatment of collagen disorders.

Liquid chromatographic and electrophoretic methods applicable to the separation of collagen and its fragments are reviewed. Special attention is paid to the separation of both stabile and labile crosslinking elements. Identification procedures exploiting the mapping of either collagen alpha-chains or of cyanogen bromide fragments are discussed. These methods can be used for diagnosing inborn errors of collagen metabolism using bioptic or necroptic samples. Analysis of urinary hydroxyproline-containing peptides or the determination of peptidically bound pyridinoline is suitable for measuring the intensity of collagen metabolism.

Amino Acids↗