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Biomedical subjects

F J van Kuijk

Publications and source records attributed to F J van Kuijk.

At least 37 records · Page 2Linked to original sources
PubMed1995

Quantitative measurement of 4-hydroxyalkenals in oxidized low-density lipoprotein by gas chromatography-mass spectrometry.

Previously, we reported on the determination of 4-hydroxyalkenals as pentafluorobenzyl oxime derivatives by gas chromatography-mass spectrometry. In this study, an improved method is presented which allows quantitative detection of 4-hydroxyalkenals in tissues by using two stable isotope-labeled internal standards, 9D3-4-hydroxynonenal and 9D3-4-hydroxyhexenal. This assay was used to quantitate 4-hydroxyalkenals in copper-oxidized human low-density lipoprotein, and we found more 4-hydroxynonenal than 4-hydroxyhexenal. This is consistent with the fact that there are more omega-6 than omega-3 fatty acids in human low-density lipoprotein samples, which are the sources of 4-hydroxynonenal and 4-hydroxyhexenal, respectively.

Aldehydes↗
PubMed1995

Vitamin E in macular and peripheral tissues of the human eye.

This study was undertaken to investigate the distribution of vitamin E in the macular and peripheral regions of the human retina, retinal pigment epithelium (RPE) and choroid as a function of age. High-performance liquid chromatography (HPLC) was used to measure alpha- and gamma-tocopherol levels quantitatively using tocol as an internal standard. In 57 out of 70 donor eyes (ages 9-104) the macular region was isolated and the tocopherols analyzed. The conventional brush method and a new vortex method were used to isolate the retinal pigment epithelium cells. Similar trends for the vitamin E levels (increase to the 5th decade, decrease after 7th decade) were found for the macular and peripheral retina and for the macular RPE. In the peripheral RPE a slight continuous increase with age was found. The vitamin E levels are higher in the RPE than in the retina, for both macular and peripheral regions. The amounts of vitamin E/mg protein are lower in the macular retina than in the peripheral retina, whereas in the RPE there is no difference in vitamin E content between macular and peripheral regions. A simple method based on a gentle vortex step was found to offer several advantages over the more generally used isolation of RPE cells based on brushing, and there was no difference in recovery of vitamin E in RPE cells when they were isolated by either isolation technique. It was also found that denominators, used to express the values of vitamin E in tissues should, be used with care since age dependent trends in parameters/denominators could be caused by trends in the denominators only.

Adolescent↗
PubMed1995

EPR studies of spin-trapped free radicals in paraquat-treated lung microsomes.

Electron Spin Resonance and Spin Trapping techniques were used to demonstrate the generation of free radicals during incubation of paraquat with lung microsomes. Aerobic incubation of paraquat resulted in the production of superoxide radical (.O2-) which was trapped by 5, 5-dimethyl-pyrroline-N-oxide or phenyl-tert-butyl-nitrone. The formation of .O2- and hydroxyl radical (.OH) by paraquat during microsomal incubation was also confirmed by the inhibition of the EPR spectra of DMPO-O2- and DMPO-OH spin adducts by superoxide dismutase and dimethyl sulfoxide, respectively. Our results provide direct evidence for the generation of reactive oxygen species during redox cycling of paraquat in microsomes. The formation of DMPO-H spin adduct during incubation of paraquat in the microsomal system is a strong indication of the involvement of hydrogen atom transfer mechanisms in paraquat-induced generations of reactive oxygen species in the lung.

Animals↗
PubMed1995

The specificity and product of quenching singlet oxygen by 2,2,6,6-tetramethylpiperidine.

The specificity of 2,2,6,6-tetramethylpiperidine to singlet oxygen was shown using Rose Bengal as a singlet oxygen generator, and Xanthine-Xanthine Oxidase and KO2 as the sources for the superoxide radical. The highest concentration of produced-singlet oxygen occurred at 25% of O2 by Rose Bengal photosensitization. The linewidth of the EPR signal for photosensitized nitroxyl radical, increasing solvent polarity. Deuterated solvents enlarge the EPR signal intensity in a dose-dependent manner. No EPR signal increase was observed in xanthine-xanthine oxidase reaction or KO2 systems, indicating that TEMP does not react with the superoxide anion. Thus, reaction of TEMP with 1O2 is highly specific.

Cyclic N-Oxides↗
PubMed1995

EPR studies on the kinetics of quenching singlet oxygen.

The specific reaction between singlet oxygen (1O2) and 2,2,6,6-tetra-methyl-piperidine (TEMP) was utilized to investigate the kinetics of singlet oxygen quenching. Rose bengal was used for the generation of singlet oxygen (1O2). The deuterium isotope effect (rho) was found to be 22.1. The rate constant (kT) of the reaction of TEMP with 1O2 in H2O, D2O and ethanol were found to be 9.0 x 10(5) M-1s-1, 1.5 x 10(6) M-1s-1 and 7.3 x 10(5) M-1s-1, respectively. The intercept of the plot of [TEMPO]-1 at different H2O/D2O ratios was found to be constant, indicating the lifetime of 3Sens is independent of solvent deuteration. The quenching of 1O2 by azide and the sensitizer itself were confirmed by the constant intercept on the kinetic plots. Thus, the quenching rate constants for azide anion in ethanol solution (96%) and for ground state rose bengal in 0.05 M phosphate buffer (pH 7.4), were found to be 8.36 x 10(8) M-1s-1 and 1.72 x 10(4) M-1s-1, respectively. These studies not only verify the specificity of the reaction between TEMP and 1O2 but also provide a sensitive and specific assay for 1O2.

Cyclic N-Oxides↗
PubMed1994

Uric acid and glutathione levels during short-term whole body cold exposure.

Ten healthy subjects who swim regularly in ice-cold water during the winter (winter swimming), were evaluated before and after this short-term whole body exposure. A drastic decrease in plasma uric acid concentration was observed during and following the exposure to the cold stimulus. We hypothesize that the uric acid decrease can be caused by its consumption after formation of oxygen radicals. In addition, the erythrocytic level of oxidized glutathione and the ratio of oxidized glutathione/total glutathione also increased following cold exposure, which supports this hypothesis. Furthermore, the baseline concentration of reduced glutathione was increased and the concentration of oxidized glutathione was decreased in the erythrocytes of winter swimmers as compared to those of nonwinter swimmers. This can be viewed as an adaptation to repeated oxidative stress, and is postulated as mechanism for body hardening. Hardening is the exposure to a natural, e.g., thermal stimulus, resulting in an increased tolerance to stress, e.g., diseases. Exposure to repeated intensive short-term cold stimuli is often applied in hydrotherapy, which is used in physical medicine for hardening.

Acclimatization↗
PubMed1994

Vitreal insulin-like growth factor binding proteins (IGFBPs) are increased in human and animal diabetics.

Although patients with diabetic retinopathy have been reported to have elevated vitreal IGF-I levels, it is not known whether diabetes also affects the levels of vitreal IGF binding proteins (IGFBPs) which control IGF's bioavailability. To address this issue, vitreal IGFBP levels were assayed in human diabetics, rats with streptozotocin-induced diabetes and galactose-fed dogs with diabetic-like retinopathy. Using 125I-IGF-II ligand blots, it was found that human diabetics have a 4-fold increase in vitreal IGFBP levels. Also, western blots on human diabetic vitreous reveal increased levels of IGFBP-2 and proteolytic fragments of IGFBP-3. IGF binding assays on vitreous from streptozotocin-treated rats (three months in duration) also indicate a 5-fold increase in IGF binding activity. IGF ligand blots using vitreous from rats with a shorter duration of diabetes (one month) show a 63% increase in IGFBP binding and a marked decrease in serum IGFBP binding. IGF ligand blots and IGFBP-2 and -4 western blots using vitreous from galactose-fed dogs with diabetic-like retinopathy exhibit a 6-fold increase in vitreal IGFBPs. The observation that vitreal IGFBPs are elevated in diabetic humans and rats without overt retinopathy suggests that these increases are not the result of a preexisting end-stage retinopathy but rather are an early ocular event in the diabetic process. Increases in vitreal IGFBPs thus could participate in the proliferative aspects of diabetic retinopathy by virtue of their putative intrinsic bioactivity or their capacity to alter IGF bioavailability.

Adult↗
PubMed1992

Quantitative determination of hydroxy fatty acids as an indicator of in vivo lipid peroxidation: oxidation products of arachidonic and docosapentaenoic acids in rat liver after exposure to carbon tetrachloride.

An improved gas chromatography-mass spectrometry method has been applied to the quantitation of both in vitro and in vivo products of lipid peroxidation in rat liver stimulated with carbon tetrachloride. The method avoids problems of autoxidation of unsaturated fatty acids during sample preparation, and the sensitivity permits assays on as little as 1 mg of tissue. This permits small samples of tissue to be obtained by biopsy from the same organ, thus making it possible to perform in vivo time studies on a single animal. Lipids from whole tissue or cell preparations are simultaneously extracted and reduced by catalytic hydrogenation and then saponified and derivatized to their pentafluorobenzyl esters and trimethylsilyl ethers. Quantitation is accomplished by negative ion chemical ionization gas chromatography-mass spectrometry, using either deuterated compounds or naturally occurring fatty acid metabolites as internal standards. Hydroxy fatty acids which result from reduction of the hydroperoxides of arachidonic and docosapentaenoic acids are found to increase within 20 min after exposure of liver or hepatocyte suspensions to carbon tetrachloride.

Animals↗
PubMed1992

2,6-Dichlorophenolindophenol is a competitive inhibitor for xanthine oxidase and is therefore not usable as an electron acceptor in the fluorometric assay.

Xanthine oxidase has been recognized as an important source of oxygen free radicals in ischemia-reperfusion injury. In order to study this enzyme in biological tissues, the conversion of pterin (2-amino-4-hydroxypteridine) to isoxanthopterin provides the basis for a very sensitive fluorometric assay. Xanthine oxidase is typically assayed in the presence of pterin only, while an electron acceptor which replaces NAD+ is used to determine the combined xanthine dehydrogenase plus xanthine oxidase activity. 2,6-Dichlorophenol-indophenol has been used as an electron acceptor in this assay. However, it was found in this study that it acts as an effective competitive inhibitor for xanthine oxidase. We concluded that methylene blue is the electron acceptor of choice in the fluorometric assays for xanthine oxidase.

2,6-Dichloroindophenol↗
PubMed1992

Fatty acid composition of the human macula and peripheral retina.

The fatty acids in the human retina and the macular region were measured quantitatively (mole percent) by gas chromatography. The major fatty acids of the human retina and macula were palmitic, stearic, oleic, arachidonic, and docosahexaenoic. Surprisingly, there was much less docosahexaenoic acid in the macular region (15.9% of total) than in the peripheral retina (22.3% of total). There was a group of "other fatty acids," not any of the five major fatty acids, that were relatively more abundant in the macula (21.0% of total) than in the peripheral retina (10.7% of total). These results indicate that the human macula has a unique biochemical composition, which differs substantially from the peripheral retina. Establishment of the biochemical composition of the macula may be important for helping recognize possible changes associated with diseases such as age-related macular degeneration.

Aged↗
PubMed1991

Photolysis intermediates of human rhodopsin.

Photochemical studies were conducted on human rhodopsin at 20 degrees C to characterize the intermediates which precede the formation of metarhodopsin II, the trigger for the enzyme cascade mechanism of visual transduction. Human rhodopsin was prepared from eyes which had previously been used for corneal donations. Time resolved absorption spectra collected from 10(-8) to 10(-6) s after photolysis of human rhodopsin in detergent suspensions displayed biexponential decay kinetics. The apparent lifetimes obtained from the data are 65 +/- 20 and 292 +/- 25 ns, almost a factor of 2 slower than the corresponding rates in bovine rhodopsin. The spectra can be fit well using a model in which human bathorhodopsin decays toward equilibrium with a blue-shifted intermediate (BSI) which then decays to lumirhodopsin. Spectra and kinetic rate constants were determined for all these intermediates using a global analysis which showed that the spectra of the human intermediates are remarkably similar to bovine intermediates. Microscopic rate constants derived from this model are 7.4 x 10(6) s-1 for bathorhodopsin decay and 7.5 x 10(6) s-1 and 4.6 x 10(6) s-1 for the forward and reverse reactions of BSI, respectively. Decay of lumirhodopsin to later intermediates was studied from 10(-6) to 10(-1) s after photolysis of rhodopsin in human disk membrane suspensions. The human metarhodopsin I in equilibrium metarhodopsin II equilibrium appears to be more forward shifted than in comparable bovine studies.

Humans↗
PubMed1991

Quantitative determination of hydroxy fatty acids as an indicator of in vivo lipid peroxidation: gas chromatography-mass spectrometry methods.

A new method has been developed for the quantitation of lipid peroxidation products by gas chromatography-mass spectrometry. An important advantage over existing gas chromatography-mass spectrometry methods is the elimination of autoxidation during sample preparation. The sensitivity is sufficient to permit measurement of lipid peroxidation products under normal physiological conditions on as little as 1 mg of tissue. Lipids from whole tissue samples or cell preparations are reduced by catalytic hydrogenation during extraction. The hydrogenation stabilizes the compounds by saturating the double bonds and reducing the hydroperoxides to hydroxy derivatives. The saturated lipids are then saponified and the resulting fatty acids are converted to pentafluorobenzyl esters. Hydroxy fatty acids are further converted to trimethylsilyl ether derivatives. Quantitation is accomplished by negative ion chemical ionization gas chromatography-mass spectrometry, using deuterated internal standards. Specific products from polyunsaturated fatty acids can be quantitated, and the method differentiates between products produced by free-radical and photooxidation mechanisms. Increased levels of lipid peroxidation products, above normal physiological levels, that result from prooxidant conditions, such as exposure of animals to carbon tetrachloride, can be measured.

Animals↗
PubMed1991

Characterization of products formed during the autoxidation of beta-carotene.

The anticarcinogenic action of carotenoids such as beta-carotene has been frequently ascribed to their antioxidant properties. However, very little is actually known about the nature of the antioxidant reaction or the products that are formed. beta-Carotene was exposed to either spontaneous autoxidation conditions or to radical-initiated autoxidation conditions. The products were separated by reverse-phase HPLC, and individual peaks were characterized with an on-line diode array detector. Carbonyl products were isolated and characterized by several procedures, including borohydride reduction to the corresponding alcohols, derivatization with O-ethyl-hydroxylamine to the corresponding O-ethyl-oximes of the carbonyls, and analysis by GC-MS. Under the conditions of the experiments, the formation of a homologous series of carbonyl products was demonstrated, including beta-apo-13-carotenone, retinal, beta-apo-14'-carotenal, beta-apo-12'-carotenal, and beta-apo-10'-carotenal. Several very hydrophobic compounds were formed, which have not been previously identified. In addition, the products of NaOCl-treatment of beta-carotene were analyzed, and shown to be significantly different from the autoxidation products. This type of product analysis should be useful in determining the nature of the oxidants reacting with beta-carotene in vivo.

Antioxidants↗
PubMed1991

Effects of ultraviolet light on the eye: role of protective glasses.

Global atmospheric changes such as depletion of ozone in the stratosphere are thought to lead to increased levels of ultraviolet radiation on earth. This can have adverse effects on human health, and long-term effects of ultraviolet light on the eye are of increasing concern. Ultraviolet light exposure to the eye has been associated with cataract formation and retinal degeneration. In both cases, it is hypothesized that ultraviolet light can initiate formation of free radicals, which can cause protein modification and lipid peroxidation. Several procedures can be recommended to prevent ultraviolet light damage to the eye, such as the use of suitable protective glasses when outdoors.

Adult↗
PubMed1991

Spectrophotometric quantitation of rhodopsin in the human retina.

The rhodopsin content of the human retina was determined spectrophotometrically. Retinas were removed completely by using an in vitro technique based on a simulation of retinal detachment. This method provides a total recovery of rod outer segments which contain the rhodopsin visual pigment or its apoprotein, opsin. Using this improved dissection method followed by a crude preparation of the retina, an opsin-rhodopsin content of 6.20 +/- 0.64 nmol/retina (n = 9) was found. This is larger than values previously reported. Regeneration with a threefold excess of 11-cis-retinal improved the recovery of rhodopsin dramatically because the corneas of these human donor eyes were removed for corneal transplant surgery under relatively bright light that bleaches the retina nearly completely. The amount of rhodopsin that can be isolated will be sufficient for studies on early events in visual transduction using laser photolysis.

Adult↗
PubMed1990

Preparative high-performance liquid chromatography purification of polyunsaturated phospholipids and characterization using ultraviolet derivative spectroscopy.

A preparative reversed-phase HPLC system utilizing an isocratic mobile phase to purify up to 10-mg quantities of phospholipids is described. The method was developed to separate oxidation products of polyunsaturated phospholipids from intact, parent lipids. The method is useful for phosphatidylcholine and phosphatidylethanolamine on a preparative scale and for phosphatidylserine and phosphatidic acid on an analytical scale. Both intact phospholipids and oxidized phospholipids were monitored by absorbance at 206 nm. The oxidation products were simultaneously monitored at 234 nm where the intact phospholipids have only a very slight end absorption. Second-derivative uv spectroscopy proved to be extremely useful to identify the presence or to verify the absence of oxidation products in phospholipid samples. For autoxidized docosahexaenoic acid containing phospholipids, the absorbance maximum of diene oxidation products is 237 nm for the trans,trans (t,t) isomer and 246 nm for the cis,trans (c,t) isomers. Similarly, five classes of triene oxidation product stereoisomers have distinct absorbance maxima detected by second-derivative spectroscopy ranging from 269 to 292 nm.

Chromatography, High Pressure Liquid↗