Effects of polyoxyethylene detergent (Brij 58) on platelet aggregation,release and clotting activity

Low concentrations of a polyoxyethylene detergent, Brij 58, inhibited the secondary phase of platelet aggregation induced by ADP in human citrated platelet-rich but had no effect on primary aggregation. Thrombin-induced aggregation of washed human platelets suspended in Tyrode's buffer was inhibited after incubation of cells with 4 · 10^?6 M detergent. Efflux of [¹⁴C]serotonin, ⁴⁵Ca²⁺ and labile aorta contracting substance (thromboxane A₂) and development of prothrombin-converting activity (platelet factor 3) were abolished concomitantly. Aggregation of washed platelets either by sodium arachidonate or by collagen was also inhibited by the same concentration of Brij 58 which inhibited thrombin aggregation. This concentration did not itself produce any release of a cytoplasmic marker, lactate dehydrogenase, from platelets. Higher concentrations of Brij 58, exceeding 4 · 10^?5 M, lysed the cells liberating lactate dehydrogenase, serotonin and Ca²⁺. When albumin was included as a platelet stabilizer in the suspending medium the concentration of detergent required for the inhibitory effects was increased ten-fold. This could be attributed to competitive binding of the detergent to albumin, demonstrated with [¹⁴C]acetylated Brij 58. A variety of other polyoxyethylene detergents, at concentrations from 8 · 10^?4 to 5 · 10^?3 M, also inhibited platelet aggregation induced by thrombin. It is concluded that low concentrations of Brij 58 stabilize the platelets against the action of aggregation agents, while higher concentrations produce membrane destabilization and cell lysis.     

Effects of α,β-methylene-adenosine-5′-diphosphate on blood platelet aggregation

The effects on platelet aggregation of α,β-methylene-adenosine-5′-diphosphate (Ado-PCP) have been investigated. Using human citrated platelet-rich plasma it has been shown that: (i) at concentrations of 10^?3 M or higher Ado-PCP is able to induce platelet aggregation; (ii) the rate of Ado-PCP-induced aggregation increases on raising the pH of platelet-rich plasma above the pK_a for the secondary phosphonyl dissociation of Ado-PCP; (iii) at concentrations from 1 · 10^?4 to 5 · 10^?4 M Ado-PCP does not cause platelet aggregation itself, but it inhibits ADP-induced aggregation. This inhibition is also observed in washed platelet suspensions. The data suggest that Ado-PCP acts at the same site on the platelet membrane as does ADP and that ADP to AMP transformation is not a prerequisite for the process of aggregation. The observed effect of pH on the rate of Ado-PCP induced aggregation suggests that the ionization state of a nucleotide terminal acid group is important in the process of aggregation.     

Spectroscopic properties of protochlorophyll forms in Triton X-100 detergent micelles

Protochlorophyll (PChl) forms were performed in Triton X-100 detergent micelles. The concentration of Triton X-100 was 7·10^?4 M (above the critical micellar concentration); the concentration of PChl varied between 1.6·10^?5 and 1.8·10^?4 M. Absorption, fluorescence and circular dichroism (CD) spectra were registered. The absorption spectra were resolved into Gaussian components by computer analysis. PChl forms with absorption bands at 632–634, 638, 652–654, 663–664, 668 and 676 nm and with fluorescence emission bands at 634–636, 640–644, 652–655, 677–678, 686 and 694–696 nm were observed in micellar solutions of different PChl concentrations. The CD spectra showed a strong dependence on the concentration of PChl: positive CD signals or positive Cotton effects were observed in the vicinity of 650 nm. The intensity of these signals increased in parallel with increasing concentration of PChl. No CD signals were found in the region of the longer wavelength absorption bands. These data show that the PChl exists in many different forms in this system, and the spectroscopic properties of these forms are determined by different molecular interactions viz., interactions of PChl with Triton X-100 or water molecules and/or by the aggregation of PChl.     

The Loch Eil project: Planktonic pigments in sediments from Loch Eil and the firth of Lorne

Photosynthetic pigments and their derivatives were measured in sediments in the fjordic Loch Eil and the Firth of Lome, Scotland, between November 1975 and November 1976. After acetone extraction from the top 10 mm of sediment cores, pigments were crudely separated, by fluorescence change on acidification, into (chlorophyll a + chlorophyllide a) and phaeopigments. The greatest pigment concentrations (mean 73 μg · g sediment dry wt^?1) were found in the most reducing sediments which also had a high average proportion (23%) of chlorophyll. The least mean pigment concentration (23 μg · g^?1) and proportion of chlorophyll (17%) were found in the most oxidizing sediments in the Firth of Lorne where there was a clear seasonal cycle, with a peak in sediment pigment concentration and chlorophyll proportion in May and June, just after the planktonic spring increase. The Loch Eil stations showed a less clear or no seasonal cycle; the station most affected by organic input was the most variable from month to month. It was concluded that redox status was the most obvious control of sediment pigment content, whereas the effect of sedimentation of phytoplankton was complex.     

Influence of carbon-dioxide on the growth of Spirulina sp. (MCRC-A0003) isolated from Muttukadu backwaters,South India

Growth of Spirulina sp. (MCRC-A0003), a cyanobacterium, was evaluated under different concentrations of carbon-dioxide (CO₂) (4–50 %) in a closed glass photobioreactor. Although significant CO₂ utilization by the cyanobacterial strain was observed up to 50 % concentration, complete utilization was observed only at 4, 10 and 20 % concentrations on 3rd, 6th and 8th day respectively. However, considerable reduction was witnessed in reactors containing 30–50 % CO₂ only between 6th and 9th day. A corresponding increase in the biomass and primary metabolites like chlorophyll-a, carbohydrate and protein were observed. Biomass productivity of Spirulina in reactors sparged with 4, 10 and 20 % CO₂ were 13.7, 43 and 44 % more than that in control reactor without CO₂. While CO₂ increased the levels of primary metabolites in the cyanobacterial cells, it was quite prominent in 10 % CO₂ concentration with the chlorophyll-a, carbohydrate and protein contents were 64, 183 and 626 mg g^?1 respectively. While 10 and 6.6 % increase were noticed in chlorophyll-a and protein, 17 % increase in carbohydrate levels was observed in Spirulina cells, which could be attributed to the conversion of CO₂ to carbohydrate by the cyanobacterium.     

Remote sensing of phytoplankton-macrophyte coexistence in shallow hypereutrophic fluvial lakes

We investigated with remote sensing (APEX images) the coexistence of phytoplankton and macrophytes in three interconnected shallow and hypereutrophic fluvial lakes (Mantua Lakes, Northern Italy). High concentrations of chlorophyll-a, up to 60 mg m^?3, were determined in the open water between well-developed stands of floating-leaved, submerged, and emergent macrophytes. Our data suggest a general inhibition of phytoplankton by macrophytes, evidenced by decreasing chlorophyll-a concentrations in proximity of macrophyte stands. Chlorophyll-a concentrations halved in the proximity of emergent stands (~6 mg m^?3 within 21 m from the stand border) when compared to the outer zones (~13 mg m^?3). Contrasting trends were observed for submerged stands, where concentrations decreased inwards from ~8 to ~3 mg m^?3. Floating leaved stands had a neutral effect, chlorophyll-a being nearly constant in both inner and outer zones. Overall, remotely-sensed data allow evaluation of quantitative and spatially defined interactions of macrophytes and phytoplankton at the whole ecosystem scale.     

Effect of detergent on aggregation of the light-harvesting chlorophyll a/b complex of photosystem 2 and its impact for carotenoid function and fluorescence quenching

Spectroscopy was used to investigate the fluorescence quenching mechanism in light-harvesting complex 2 (LHC2). The 77 K fluorescence excitation spectroscopy was performed for detection of aggregation state of LHC2 treated with different concentrations of octylphenol poly(ethyleneglycol ether)₁₀ (TX-100). Resonance Raman (RR) spectra excited with 488, 496, and 514 nm provided molecular configuration of neoxanthin, lutein 1, and lutein 2, respectively. At increased concentration of TX-100, the RR signals of xanthophylls were enhanced in the four frequency regions, which was accompanied with increase of fluorescence of chlorophyll (Chl) a. Thus the absorption of the three xanthophyll molecules was inclined to excitation wavelength, which proved that functional configurations of xanthophyll molecules in LHC2 were vital for fast transfer of excitation energy to Chl a molecules. Changes in the v4 region (C-H out-of-plane bending modes, at ∼960 cm⁻¹ in RR spectra) demonstrated that the twist feature of neoxanthin, lutein 1, and lutein 2 molecules existed in LHC2 trimers, however, it was lost in the LHC2 macro-aggregates. In the second derivative absorption spectra of LHC2, neoxanthin absorption was not detected in LHC2 macro-aggregates, while evident absorption was found in LHC2 trimers and this absorption decreased obviously when TX-100 concentration was higher than 1 mM. Hence the neoxanthin molecule had a structural role in formation of LHC2 trimers. The RR and absorption spectra also implied that carotenoid molecules constructed the functional LHC2 trimers via their intrinsic configuration features, which enabled energy transfer to Chl a efficiently and led to lower fluorescence quenching efficiency. In contrast, these intrinsic twist configurations were lost in LHC2 macro-aggregates and led to lower energy transfer efficiency and higher fluorescence quenching efficiency.     

Energy transfer by chlorophyll a in detergent micelles

The process of energy transfer was studied in the chlorophyll a-containing detergent micelle, serving as a possible model of the photosynthetic unit. Chlorophyll a was added to aqueous solutions of the detergent Triton X-100 and incorporated into the micelles. The energy transfer process was studied by investigating the concentration depolarization of fluorescence of chlorophyll a. On the basis of the experimental depolarization curves as well as the value of the Förster parameter $\text{R}{}_0\text{= 56}\text{A}\text{?}$ calculated from the overlap of absorption and fluorescence spectra it was concluded that energy transfer between chlorophyll a molecules in this model follows the Förstertype mechanism of inductive resonance. Furthermore it was found that the local concentration of chlorophyll a in the micelles is higher by 1–3 orders of magnitude than its overall concentration in the solution and by choosing the appropriate ratio between the concentration of chlorophyll a and the detergent it is possible to reach the in vivo chlorophyll concentration of 0.1 M within the micelles. Thus the chlorophyll-detergent micelle model may be applied as a model of the separate package-type photosynthetic unit.     

PHYCOBILIN CONTENT OF THE CONCHOCELIS PHASE OF ALASKAN PORPHYRA (BANGIALES,RHODOPHYTA) SPECIES: RESPONSES TO ENVIRONMENTAL VARIABLES1

Variations of pigment content in the microscopic conchocelis stage of four Alaskan Porphyra species were investigated in response to environmental variables. Conchocelis filaments were cultured under varying conditions of irradiance and nutrient concentrations for up to 60 d at 11°C and 30 psu salinity. Results indicate that conchocelis filaments contain relatively high concentrations of phycobilins under optimal culture conditions. Phycobilin pigment production was significantly affected by irradiance, nutrient concentration, and culture duration. For Porphyra abbottiae V. Krishnam., Porphyra sp., and Porphyra torta V. Krishnam., maximal phycoerythrin (63.2–95.1 mg · g dwt^?1) and phycocyanin (28.8–64.8 mg · g dwt^?1) content generally occurred at 10 μmol photons · m^?2 · s^?1, f/4–f/2 nutrient concentration after 10–20 d of culture. Whereas for Porphyra hiberna S. C. Lindstrom et K. M. Cole, the highest phycoerythrin (73.3 mg · g dwt^?1) and phycocyanin (70.2 mg · g dwt^?1) content occurred at 10 μmol photons · m^?2 · s^?1, f nutrient concentration after 60 d in culture. Under similar conditions, the different species showed significant differences in pigment content. P. abbottiae had higher phycoerythrin content than the other three species, and P. hiberna had the highest phycocyanin content. P. torta had the lowest phycobilin content.     

Low-temperature time-resolved spectroscopic study of the major light-harvesting complex of Amphidinium carterae

The major light-harvesting complex of Amphidinium (A.) carterae, chlorophyll-a–chlorophyll-c ₂–peridinin–protein complex (acpPC), was studied using ultrafast pump-probe spectroscopy at low temperature (60 K). An efficient peridinin–chlorophyll-a energy transfer was observed. The stimulated emission signal monitored in the near-infrared spectral region was stronger when redder part of peridinin pool was excited, indicating that these peridinins have the S₁/ICT (intramolecular charge-transfer) state with significant charge-transfer character. This may lead to enhanced energy transfer efficiency from “red” peridinins to chlorophyll-a. Contrary to the water-soluble antenna of A. carterae, peridinin–chlorophyll-a protein, the energy transfer rates in acpPC were slower under low-temperature conditions. This fact underscores the influence of the protein environment on the excited-state dynamics of pigments and/or the specificity of organization of the two pigment–protein complexes.     

Production of biomass and lutein by Chlorella protothecoides at various glucose concentrations in heterotrophic cultures

The influence of initial glucose concentrations on the production of biomass and lutein by Chlorella protothecoides CS-41 was investigated in batch cultures using both shake flasks and fermentors. The maximum biomass concentration increased from 4·9 to 31·2 g litre⁻¹ dry cells with an increase in initial glucose concentration from 10 to 80 g litre⁻¹. An even higher initial glucose concentration (100 g litre⁻¹) resulted in a lower biomass concentration, a lower specific growth rate, a lower growth yield coefficient and a considerably longer lag phase, which might be due to substrate inhibition. The initial glucose level also had a significant effect on the production of lutein. In a 3·7-litre fermentor an increase in lutein production from 19·39 to 76·56 mg litre⁻¹ was obtained with an increase in initial glucose concentration from 10 to 40 g litre⁻¹, within which range, lutein yield coefficient was constant (Y_Itn=1·90±0·02 mg g⁻¹). A simple substrate inhibition model was developed, which fitted the experimental data better than the classical Haldane model. A group of time-dependent kinetic models for algal cultivation in fermentors were also constructed, which were in good agreement with the experimental results and could be employed to predict the production of biomass and lutein, and the consumption of glucose in fermentor cultures.     

Kinetics of interaction of some α- and β-D-monosaccharides with concanavalin A

The rates of formation and dissociation of concanavalin A with some 4-methylumbelliferyl and p-nitrophenyl derivatives of α- and β-D-mannopyranosides and glucopyranosides were measured by fluorescence and spectral stopped-flow methods. All process examined were uniphasic. The second-order formation rate constants varied only from 6.8 · 10⁴ to 12.8 · 10⁴ M^?. s^?1, whereas the first-order dissociation rate constants ranged from 4.1. to 220 s^?1, all at ph 5.0, I = 0.3 M, and 25°C. Dissociation rates thus controlled the value of binding constant. The effect of temperature on these reactions was examined, from which enthalpies and entropies of activation and of reaction could be calculated. The effects of pH at 25°C on the reaction rates of 4-methylumbelliferyl α-D-mannopyranoside and 4-methylumbelliferyl α-D-glucopyranoside with concanavalin A were examined. The value of the binding constant K_ap (derived from the kinetics) at any pH could be related to the intrinsic binding constant K by the expression K_ap = K_aK(K_a + [H⁺])^?1. The values of K_a, the ionization constant of the protein segment responsive to sugar binding, were 3 · 10^?4 M and 1 · 10^?4 M for 4-methylumbelliferyl α-D-mannopyranoside and 4-methylumbelliferyl α-D-glucopyranoside, respectively. The binding constant of p-nitrophenyl α-D-mannopyranoside is surprisingly much less sensitive to a pH change from 5.0 to 2.7. Ionic strength had little effect on the binding characteristics of 4-methylumbelliferyl α-D-mannopyranoside to concanavalin A at pH 5.2 and 25°C.     

Effect of adrenalectomy on cellular calcium metabolism and on the response to adrenergic stimulation of hepatocytes isolated from male and female rats

The effects of adrenalectomy on cell calcium metabolism and on the effects of epinephrine on cAMP, phosphorylase a activity, and calcium efflux were studied in hepatocytes isolated from adult male and female rats. Adrenalectomy increased the total calcium of hepatocytes, all exchangeable calcium pools, and all calcium fluxes between the cellular pools in both sexes. After adrenalectomy, basal cAMP was elevated, phosphorylase a + b was decreased, but basal phosphorylase a activity was not changed. In adrenalectomized males and at all concentrations of epinephrine studied (1·10^?8?1·10^?5M) stimulation of calcium efflux was decreased and cAMP accumulation was enhanced, while the resulting phosphorylase a activation was depressed. In hepatocytes from adrenalectomized females there was a similar increase in cAMP accumulation induced by epinephrine, and a decrease in the stimulation of calcium efflux; however, the depression in phosphorylase a activation was much less and was significant only at 1·10^?8 and 1·10^?5M epinephrine. In the male, while activation of phosphorylase a shifted from a pure α-adrenergic response mediated by calcium to one also involving a cAMP-mediated β-adrenergic response, the contribution of the attenuated calcium signal was still significant. Hepatocytes from female rats did not show a comparable α- to β-shift, since the relative contribution of calcium and cAMP to phosphorylase activation was similar in sham-operated and adrenalectomized animals.     

AMMONIUM AND UV RADIATION STIMULATE THE ACCUMULATION OF MYCOSPORINE‐LIKE AMINO ACIDS IN PORPHYRA COLUMBINA (RHODOPHYTA) FROM PATAGONIA,ARGENTINA1

The combined effects of ammonium concentration and UV radiation on the red alga Porphyra columbina (Montagne) from the Patagonian coast (Chubut, Argentina) was determined using short‐term (less than a week) experimentation. Discs of P. columbina were incubated with three ammonium concentrations (0, 50, and 300 μM NH₄Cl) in anilluminated chamber (PAR=300 μmol photons·m^?2·s^?1, UVA=15 W·m^?2, UVB=0.7 W·m^?2) at 15°C. Algae incubated at 300 μM ammonium showed a significant increase (P<0.05) in the concentration of mycosporine‐like amino acids (MAAs) compared with the initial value or with the other ammonium treatments. The increase of MAAs was, however, a function of the quality of irradiance received, with a higher increase in samples exposed to UVA compared with UVB (29% and 5% increase, respectively). However, UVB radiation was more effective in inducing MAA synthesis per unit energy received by the algae. Samples exposed to PAR only had an intermediate increase in MAA concentration of 16%. Chl a concentration decreased through the incubation with the greatest decrease at high ammonium concentration. Phycobiliprotein (BP) decreased through time with the smallest decrease occurring at high ammonium concentration. Photoinhibition (as a decrease of optimal quantum yield) was significantly greater under nitrogen‐deprived conditions than that under replete ammonium levels. Maximal gross photosynthesis (GP_max), as oxygen evolution, and maximal electron transport rate (ETR_max), as chl fluorescence, increased with the ammonium concentration. Positive relationships between maximal GP or ETR and pigment ratios (BP/chl a and MAAs/chl a) and negative relationships with chl a concentration were found.     

SEASONAL PHOTOSYNTHETIC PERFORMANCE OF MACROCYSTIS INTEGRIFOLIA (PHAEOPHYCEAE)1

The seasonal photosynthetic performances of three age classes of blades of Macrocystis integrifolia Bory were estimated by studying their photosynthetic rate vs. irradiance curves and pigment contents for 15 months. All blade types were irradiance-saturated between 25 and 70 μE · m^?2· S^?1. Young and mature blade tissues had higher photosynthetic maxima and initial slopes on an area basis than older blade tissue. The latter, however, had pigment concentrations similar to those in mature blade tissues. All these parameters varied on a seasonal basis. The photosynthetic maxima ranged from 0.1–0.8 μmol · C · cm^?2· h^?t and showed two peaks, one in late summer-early fall and the other in late winter. Changes in initial slope and pigment concentrations in the blade tissues suggest that, changes in the size or efficiency of electron transfer in the photosynthetic unit occur. These data are discussed in relation to changes in seawater temperature and nitrate concentrations.     

A diffusional analysis of the temperature sensitivity of the Mg2+-induced rise of chlorophyll fluorescence from pea thylakoid membranes

The kinetics of the chlorophyll fluorescence rise induced by adding 20 mM MgCl₂ to a suspension of isolated pea chloroplasts treated with 3-(3,4-dichlorophenyl)-1, 1-dimethylurea (DCMU) have been examined experimentally and theoretically as a function of temperature. The application of similarity arguments and particle aggregation theory to the experimental results suggests that at the first approximation, the salt-induced time-dependent fluorescence changes may be described by the diffusion-controlled lateral movement of Photosystem II pigment-protein complexes. From an analysis of the temperature dependence of the fluorescence changes, estimates obtained for the lateral diffusion coefficients were 1.85 · 10^?12–3.08 · 10^?11 cm²/s over the temperature range 10°C ? T?30°C.     

Vicia graminea lectin binding to human M and N erythrocytes

The mode of binding of Vicia graminea¹²⁵I-labelled lectin to human M and N erythrocytes at 4°C has been investigated. The labelled lectin retained the full activity of native lectin. Lectin association at 4°C was characterized by a $\text{t}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$ of 3 to 5 min, reaching steady-state within 15 min. Incubation of cells for 15 min at 4°C with increasing concentrations of Vicia graminea¹²⁵I-labelled lectin showed that saturation binding occurred. Scatchard analysis of equilibrium data determined over a wide range of lectin concentrations yielded a curvilinear plot with an upward concave slope; this representation indicated that there was not a single homogeneous class of noninteracting binding sites. This result could indicate two or more independent classes of binding sites or one class of interacting sites exhibiting negative cooperativity. Since unlabelled lectin, which at the concentration used, rapidly binds to available receptors, did not affect the dissociation rate of the labelled lectin and since identical Scatchard plots were found using native and formaldehyde-fixed erythrocytes we conclude that there are two classes of independent Vicia graminea binding sites on human erythrocytes. Computer analysis of the Scatchard plots gave high- and low-affinity constant (7.07±1.1) · 10⁷ M^?1 and (0.2±0.01) · 10⁷ M^?1, respectively, for N erythrocytes and (1.13±0.18) · 10⁷ M^?1 and (0.24±0.01) · 10⁷ M^?1, respectively for the M cells. N erythrocytes were estimated to have 0.085 · 10⁵ high-affinity and 2.1 · 10⁵ low-affinity sites and M erythrocytes, 0.011 · 10⁵ high affinity and 0.13 · 10⁵ low-affinity sites. N cells therefore have 10-times as many sites as M cells. Studies of the dissociation of ¹²⁵I-labelled lectin from N and M cells in the presence of unlabelled lectin gave dissociation rate constants of 51 · 10^?4 s^?1 and 1.97 · 10^?4 s^?1 for the high- and low-affinity sites of N cells and 13 · 10^?4 s^?1 and 1.6 · 10^?4 s^?1 for the high- and low-affinitym sites of M cells, indicating that the binding of Vicia graminea lectin to human erythrocytes is reversible.     

Determination of glucose,urea and penicillin using enzyme-pH-electrodes

Conventional hydrogen ion glas electrodes have been used for the preparation of enzyme-pH-electrodes by either entrapping the enzymes within polyacrylamide gels around the electrode or as liquid layer trapped within a cellophane membrane. The enzymes were glucose oxidase, urase and penicillinase.The pH response to glucose concentration was about linear within 10^?1–10^?3 M glucose and for urea linear within 5·10^t—–5·10^?5M. The pH response to penicillin was about linear in the range from 10^?3–10^?2 M resulting in a pH shift of 1.4 units; reproduceable pH response was obtained down to concentrations of 3·10^?5 M.Studies as to the effect of buffer using an urease–pH-electrode showed a buffer concentration of 10^?2 M a substantial shift of about one pH-unit in the range of 10^?4 to 10^?2 M urea. Both urease- and penicillinase–pH-electrodes were tested as to stability showing no decrease in pH response except at high substrate concentration (1·10^?2 M) over a period of 2–3 weeks kept at room temperature.     

Aqueous and salt solutions of quinine of low concentrations: Self-organization,physicochemical properties and actions on the electrical characteristics of neurons

Self-organization, the physicochemical properties of aqueous and salt solutions of quinine and the effects of salt quinine solutions in a wide range of concentrations (1 · 10^?22?1 · 10^?3 M) on the electrical characteristics of the edible snail’s identified neurons were studied. Similar non-monotonic concentration dependencies of physicochemical properties of aqueous and salt quinine solutions at low concentrations are obtained. This allows of predicting the occurrence of biological effects at low concentrations of quinine solutions. Intrinsic (within 5% of the interval) changes in membrane potential, the amplitude and duration of the neuron action potential under the influence of quinine salt solutions at concentrations of quinine of 1 · 10^?20, 1 · 10^?18, 1 · 10^?10 M are found. For these concentrations the extreme values of specific conductivity and pH are shown.