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Biomedical subjects

B Kok

Publications and source records attributed to B Kok.

At least 19 recordsLinked to original sources
PubMed1995

Nitrite and nitrate determinations in plasma: a critical evaluation.

Plasma nitrite and nitrate determinations are increasingly being used in clinical chemistry as markers for the activity of nitric oxide synthase and the production of nitric oxide radicals. However, a systematic evaluation of the determination of nitrite and nitrate in plasma has not been performed. In this study the recovery and stability of nitrite and nitrate in whole blood and in plasma, the relation between nitrite and nitrate concentrations in plasma, and possible sources of artifacts were investigated. The main conclusions are: (a) Recovery of nitrite and nitrate from plasma is near-quantitative (87%) and reproducible; (b) nitrite and nitrate are stable in (frozen) plasma for at least 1 year; (c) nitrite in whole blood is very rapidly (> 95% in 1 h) oxidized to nitrate, and therefore plasma nitrite determination alone is meaningless; (d) the ranges of nitrite and nitrate concentrations in plasma samples of 26 healthy persons are 1.3-13 mumol/L (mean 4.2 mumol/L) and 4.0-45.3 mumol/L (mean 19.7 mumol/L), respectively; (e) plasma nitrite and nitrate concentrations were not correlated (nitrite as % of total nitrite + nitrate varied from 3.9% to 88% in plasma samples); and (f) plasma samples should be deproteinized, and background controls for each sample should be included in the assay, to avoid measuring artifactually high nitrite and nitrate concentrations in plasma.

Drug Stability↗
PubMed1979

Redox titration of electron acceptor Q and the plastoquinone pool in photosystem II.

The primary photochemical quencher Q and the secondary electron acceptor pool in Photosystem II have been titrated. We used particles of Scenedesmus mutant No. 8 that lack System I and allowed the system to equilibrate with external redox mediators in darkness prior to measurement of the fluorescence rise curve. The titration of Q, as indicated by the dark level of Fi, occurs in two discrete steps. The high-potential component (Qh) has a midpoint potential of +68 mV (pH 7.2) and accounts for approximately 67% of Q. The pH sensitivity of the midpoint potential is -60 mV, indicating the involvement of 1 H+/e. The low-potential component (Q1) accounts for the remaining 33% of Q and shows a midpoint potential near--300 mV (pH 7.2). The plastoquinone pool, assayed as the half-time of the fluorescence rise curve, titrates as a single component with a midpoint potential 30--40 mV more oxidizing than that of Qh, i.e., at 106 mV (pH 7.2). The Em shows a pH sensitivity of -60 mV/pH unit, indicating the involvement of 1 H+/e. The observation that all 12--14 electron equivalents in the pool titrate as a single component indicates that the heterogeneity otherwise observed in the secondary acceptor system is a kinetic rather than a thermodynamic property. Illumination causes peculiar, and as yet unclarified, changes of both Q and the secondary pool under anaerobic conditions that are reversed by oxygen.

Chlorophyta↗
PubMed1979

Rate-temperature curves as an unambiguous indicator of biological activity in soil.

Experiments are described in which we used a mass spectrometer to monitor O(2) uptake of enclosed soil samples as a function of temperature. We found that an Arrhenius plot of the rate of O(2) uptake showed pronounced local maxima attributable to biological activity, whereas similar plots of rates obtained with abiotic soils yielded straight lines. This procedure thus provides a basis for distinguishing biological from chemical activity for reactions, such as O(2) uptake, that can occur via either biological or chemical pathways.

Journal Article↗
PubMed1978

Evidence that the intermediate electron acceptor, A2, in photosystem I is a bound iron-sulfur protein.

Absorption changes accompanying the formation of light-induced P-700+ were investigated in a highly enriched Photosystem I preparation where an intermediate electron acceptor preceding P-430 could be detected. In an enriched Photosystem I particle, light-induced reversible absorption changes observed at 700 nm in the presence of dithionite resembled those previously seen at 703 nm and 820 nm [9], thus indicating the presence of a backreaction between P700+ and A-2. After this same Photosystem I particle was treated to denature the bound iron-sulfur centers, the photochemical changes that could be attributed to P-700 in equilibrium A2 were completely lost. These results provide evidence that the intermediate electron acceptor, A2, is a bound iron-sulfur protein. Additional studies in the 400--500 nm region with Photosystem I particles prepared by sonication indicate that the spectrum of A2 is different from that of P-430.

Cytochromes↗
PubMed1978

Photosynthetic oxygen evolution from hydrogen peroxide.

A study was made of the interactions of flash-illuminated chloroplasts with hydrogen peroxide. We conclude: 1. The oxygen precursor system can be reduced beyond the S0 state to an S-1 state, which can be oxidized to S0 by a single flash. 2. In the dark, a two-electron donation by H2O2 takes place which reduces S2 to S0 and S1 to S-1. 3. At the same time, two-electron oxidations by H2O2 re-form, S2 from S0 and S1 from S-1. 4. The catalase-like activity due to this cyclic oxidation and reduction of the S enzyme is higher with the S2 in equilibrium S0 couple than with the S1 in equilibrium S-1 couple. Another process, however, is responsible for most of the O2 evolution from H2O2 in the light. Our evidence indicates that this process: (1) is independent of the S states and insensitive to Tris washing, (2) turns over rapidly in high concentrations of peroxide, (3) yields 1 O2 per electron passing through system II; (4) dismutates two H2O2 molecules, so that there is no net consumption of 'holes'.

Catalase↗
PubMed1978

Kinetics and Apparent K(m) of Oxygen Cycle under Conditions of Limiting Carbon Dioxide Fixation.

A mass spectrometer with a membrane inlet was used to monitor light-driven O(2) evolution, O(2) uptake, and CO(2) uptake in suspensions of algae (Scenedesmus obliquus). We observed the following. (a) The rate of O(2) uptake, which, in the presence of iodoacetamide, replaces the uptake of CO(2), showed a distinct plateau (V(max)) beyond approximately 30% O(2) and was half-maximal at approximately 8% O(2). We concluded that this light-driven O(2) uptake process, which does not involve carbon compounds, is saturated at lower O(2) concentrations than are photorespiration and glycolate formation. (b) In the absence of inhibitor, O(2) evolution was relatively unaffected by the presence or absence of CO(2). During the course of CO(2) depletion, electron flow to CO(2) was replaced by an equivalent flow to O(2). (c) There was a distinct delay between the cessation of CO(2) uptake and the increase in O(2) uptake. We ascribe this delay to the transient utilization of another electron acceptor-possibly bicarbonate or another bound form of CO(2).

Journal Article↗
PubMed1978

Comparison of photosynthetic activities of spinach chloroplasts with those of corn mesophyll and corn bundle sheath tissue.

Bundle sheath and mesophyll chloroplasts from Zea mays showed comparable rates of O(2) evolution, which amounted to about half of the rate observed in spinach (Spinacia oleracea) chloroplasts.Ratios of 4.5, 4.6, and 6.2 Mn(2+) atoms per 400 chlorophylls were observed in mesophyll, bundle sheath, and spinach chloroplasts, respectively. These ratios roughly correspond to the observed O(2) evolution rates.Rates of electron transport from water to methylviologen (photosystem I and II) in both types of corn chloroplasts were about one-third that in spinach. Compared to spinach, transport rates from reduced diaminodurene to methylviologen (photosystem I) were about one-third and greater than one-half in mesophyll and bundle sheath material, respectively.In both types of corn chloroplasts, electron flow from photosystem II to P700 was abnormal. This observation, together with the low rates of all activities, suggests that damage occurred during isolation. Such damage may limit the quantitative significance of observations made with these materials (including the following data).Measurements of flash yields of O(2) evolution or O(2) uptake showed that the size of the photosynthetic unit was the same in photosystems I and II and in all three types of chloroplasts (about 400 chlorophylls per equivalent).Similarity of the photochemical cross-section of the two photosystems in the three preparations was also found in optical experiments: that is the half-times of the fluorescence rise in the presence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) (photosystem II) and of the photooxidation of P700 (photosystem I).The ratio of P700 to chlorophyll appeared to be about 2-fold higher in bundle sheath chloroplasts than in the other materials (1/200 versus 1/400).

Journal Article↗
PubMed1977

Plastocyanin as the possible site of photosynthetic electron transport inhibition by glutaraldehyde.

Treatment of spinach chloroplasts with glutaraldehyde causes an inhibition in the electron transport chain between the two photosystems. Measurements of O(2) flash yields, pH exchange, and fluorescence induction show that the O(2) evolving apparatus, photosystem II and its electron acceptor pool are not affected. The behavior of P700 indicates that its reduction but not its oxidation, is severely inhibited. Cytochrome f is still reducible by photosystem II but also slowly oxidizable by photosystem I. The sensitivity of isolated plastocyanin to glutaraldehyde further supports the conclusion that glutaraldehyde inhibits at the plastocyanin level and thereby induces a break between P700 and cytochrome f.

Journal Article↗
PubMed1976

Stabilization by glutaraldehyde of high-rate electron transport in isolated chloroplasts.

Treatment of isolated chloroplasts with glutaraldehyde affects their ability to photoreduce artificial electron acceptors. The remaining rate of O2 evolution approaches zero with methyl viologen, is low with ferricyanide, but nearly normal with lipophilic Photosystem II acceptors, like oxidized p-phenylenediamine and oxidized diaminodurene. Since Photosystem I donor reactions are also affected, a specific site of inhibition of electron transport to Photosystem I is indicated. At the same time, glutaraldehyde prolongs the longevity of the chloroplasts stored in dark. In control samples the half-life of Photosystem II activity varied between 5 days at 4degreesC and 1 day at 25degreesC. Glutaraldehyde treatment increased these half times approx. 3-fold. The glutaraldehyde doses required to induce inhibition and stabilization were very similar.

Aldehydes↗
PubMed1976

Determination of H+/e- ratios in chloroplasts with flashing light.

Using a rapid pH electrode, measurements were made of the flash-induced proton transport in isolated spinach chloroplasts. To calibrate the system, we assumed that in the presence of ferricyanide and in steady-state flashing light, each flash liberates from water one proton per reaction chain. We concluded that with both ferricyanide and methylviologen as acceptors two protons per electron are translocated by the electron transport chain connecting Photosystem II and I. With methyl viologen but not with ferricyanide as an acceptor, two additional protons per electron are taken up due to Photosystem I activity. One of these latter protons is translocated to the inside of the thylakoid while the other is taken up in H2O2 formation. Assuming that the proton released during water splitting remains inside the thylakoid, we compute H+/e- ratios of 3 and 4 for ferricyanide and methylviologen, respectively. In continuous light of low intensity, we obtained the same H+/e- ratios. However, with higher intensities where electron transport becomes rate limited by the internal pH, the H+/e- ratio approached 2 as a limit for both acceptors. A working model is presented which includes two sites of proton translocation, one between the photoacts, the other connected to Photosystem I, each of which translocates two protons per electron. Each site presents a approximately 30 ms diffusion barrier to proton passage which can be lowered by uncouplers to 6-10 ms.

Chloroplasts↗
PubMed1976

Photoreduction of O(2) Primes and Replaces CO(2) Assimilation.

A mass spectrometer with a membrane inlet system was used to monitor directly gaseous components in a suspension of algae. Using labeled oxygen, we observed that during the first 20 seconds of illumination after a dark period, when no net O(2) evolution or CO(2) uptake was observed, O(2) evolution was normal but completely compensated by O(2) uptake. Similarly, when CO(2) uptake was totally or partially inhibited, O(2) evolution proceeded at a high (near maximal) rate. Under all conditions, O(2) uptake balanced that fraction of the O(2) evolution which could not be accounted for by CO(2) uptake.From these observations we concluded that O(2) and CO(2) are in direct competition for photosynthetically generated reducing power, with O(2) being the main electron acceptor during the induction process and under other conditions in which CO(2) reduction cannot keep pace with O(2) evolution. The high rate of the O(2) uptake reaction observed in the presence of iodoacetamide, KCN, or carbonyl cyanide p-trifluoromethyoxyphenylhydrazone, suggests that a special high capacity oxidase distinct from ribulose diphosphate oxygenase exists in whole cells. The rapid reduction of molecular O(2) after a period of darkness probably serves as a priming reaction for the photosynthetic apparatus. The high steady state rate of the O(2) cycle in the absence of CO(2) fixation suggests that the regulation of photosynthesis does not involve significant changes in the rate of photochemical electron transport.

Journal Article↗