scholarly journals ELECTROPHORETIC STUDIES ON HUMAN RED BLOOD CELLS

1941 ◽  
Vol 24 (4) ◽  
pp. 447-457 ◽  
Author(s):  
Robert F. Furchgott ◽  
Eric Ponder
Keyword(s):  
Ionic Strength ◽  
Phosphate Buffer Solution ◽  
Strong Acid ◽  
Red Cells ◽  
Radius Of Curvature ◽  
Large Radius ◽  
Buffer Solution ◽  
Mobility Data ◽  
Human Red Blood Cells ◽  
Human Red Cells

1. The electrophoretic mobility of unhemolyzed human red cells has been determined as a function of ionic strength at approximately constant pH in isotonic mixtures of glucose solution and saline-phosphate buffer solution. 2. Above an ionic strength of about 0.02 the cells behave as particles with a smooth surface of large radius of curvature. Below an ionic strength of about 0.02, changes of the surface occur, probably involving a decrease of charge density and perhaps connected with injury of the surface. 3. The mobility as a function of pH at an ionic strength of 0.172 has been determined for human red cells, for the lipid extract of the cells, and for the stroma protein of the cells. The isoelectric points of cells, lipid, and protein have been found to be about 1.7, 2.6, and 4.7 respectively. 4. The pH-mobility data lead to the conclusion that a red cell surface is composed largely of lipid and dominated by strong acid groups, possibly the phosphoric acid groups of cephalin molecules.

Influence of Red Blood Cell Concentrations on the Measurement of Turbulence Using Hot-Film Anemometer

1983 ◽  
Vol 105 (4) ◽  
pp. 406-410 ◽  
Author(s):  
A. M. Sallam ◽  
N. H. C. Hwang
Keyword(s):  
Phosphate Buffer Solution ◽  
Experimental Information ◽  
Characteristic Change ◽  
Density Functions ◽  
Buffer Solution ◽  
Number Range ◽  
Flow Phenomenon ◽  
Human Red Blood Cells ◽  
Spectral Density Functions ◽  
Hot Film

Measurement of local velocity fluctuations was made with an L-shaped conical hot-film probe in a submerged circular jet. The experiment was carried out in solutions of washed human red blood cells (RBC) in a phosphate buffer solution (PBS), at hematocrit concentrations (Ht percent) of 10, 19, 29, and 38 percent. The viscosity of the testing solutions was kept at 3.2 c.p. by adding proper amount of dextran. The experiment was conducted at Reynolds numbers (NR) 674, 963, 1255 and 1410, based on the jet exit velocity and exit diameter. Statistical analyses were performed on the recorded instantaneous velocity signals to obtain the root-mean-square (rms) values, the probability density functions (PDF) and the power spectral density functions (PSDF) of the signals. Within the range tested, we noticed an incidental rise in rms values at 19 to 29 Ht percent for NR = 963 similar to those reported earlier in the literature. Further analyses using PDF and PSDF, however, showed neither a trend nor any physical significance of this rise. Based on the analyses of both the PDF and the PSDF, we believe that the incidental rise in rms value can be partially attributed to the high spikes registered by the probe in a high RBC concentrations fluid flow. The bombardment of RBC on the probe thermal boundary layer may cause a characteristic change in the probe response to certain flow phenomenon, at least within the Reynolds number range used in this study. Additional theoretical and experimental information is needed to pin point the nature of this response. We thus suggest that the second and higher moments of the HFA signals obtained in a fluctuating flow field involving a liquid with relatively high contaminant concentrations cannot be interpreted as a simple flow phenomenon.

Development and Validation of HPLC Determination of related Substances in A Novel Anticonvulsant agent Epimidin

2021 ◽  
pp. 3223-3231
Author(s):  
Hanna I. Severina ◽  
Svitlana M. Gubar ◽  
Ivan V. Bezruk ◽  
Anna S. Materiienko ◽  
Liudas Ivanauskas ◽  
...  
Keyword(s):  
Mobile Phase ◽  
Phosphate Buffer Solution ◽  
Anticonvulsant Drug ◽  
Strong Acid ◽  
Gradient Elution ◽  
Stress Conditions ◽  
Drug Candidate ◽  
Buffer Solution ◽  
Related Substances

1-(4-methoxyphenyl)-5-[2-[4-(4-methoxyphenyl)piperazin-1-yl]-2-oxo-ethyl]pyrazolo[3,4-d]pyrimidin-4-one has been reported as a promising new anticonvulsant drug candidate with a code name “Epimidin”. A new HPLC method for the related substances determination of potential active pharmaceutical ingredient has been developed and validated. The method uses ACE C18 column (250x4.6mm, 5µm) and gradient elution. Mobile phase consisted of a mixture of methanol R (mobile phase A) and phosphate buffer solution with triethanolamine, adjusted to pH 7.0 (mobile phase B). During the analysis, the ratio of mobile phases was changing according to a gradient mode at a flow rate of 1ml/min. The DAD detection was set at 240nm. The method was validated according to the ICH guidelines and requirements of State Pharmacopoeia of Ukraine. Drug substance was thoroughly explored for stability assessments under various stress conditions such as high temperature, as well as the influence of strong acid and base and oxidizing agents. The obtained solutions were analyzed by HPLC and LC/MS. It has been shown that the substance Epimidin was not resistant to the action of peroxide, alkali and acid decomposition – the mentioned stress conditions lead to the formation of unidentified impurities.

scholarly journals Kinetics and stoichiometry of Na-dependent Li transport in human red blood cells.

1978 ◽  
Vol 72 (2) ◽  
pp. 249-265 ◽  
Author(s):  
B Sarkadi ◽  
J K Alifimoff ◽  
R B Gunn ◽  
D C Tosteson
Keyword(s):  
Transport System ◽  
Red Cells ◽  
Normal Male ◽  
Red Cell ◽  
Red Cell Membrane ◽  
Human Red Blood Cells ◽  
Na Efflux ◽  
Tightly Coupled ◽  
Male Subjects ◽  
Human Red Cells

This paper describes the kinetics and stoichiometry of a tightly coupled Na-Li exchange transport system in human red cells. The system is inhibited by phloretin and furosemide but not by ouabain. Li influx by this system increases and saturates with increasing concentrations of external Li and internal Na and is inhibited competitively by external Na. Comparable functions relate Li efflux and Na efflux to internal and external Li and Na concentrations. Analysis of these relations yields the following values for the ion concentrations required to half-maximally activate the transport system: internal Na and Li 9.0 and 0.5 mM, respectively, external Na and Li 25 and 1.5 mM, respectively. The system performs a 1:1 exchange of Na and Li moving in opposite directions across the red cell membrane. We found no evidence for a simultaneous transport of more than one Na and Li by the system. The maximum transport rate of Na-dependent Li transport varied between 0.1 and 0.37 mmol/(liter of cells X h) in the red cells of the five normal male subjects studied. No significant variations between individual subjects were observed for bicarbonate-stimulated Li transport and for the residual Li fluxes which occur in the absence of bicarbonate and in the presence of ouabain plus phloretin.

scholarly journals THE PROPERTIES OF MAMMALIAN STRIATED MYOFIBRILS ISOLATED BY AN ENZYMATIC METHOD

1950 ◽  
Vol 91 (6) ◽  
pp. 655-664 ◽  
Author(s):  
Armin F. Schick ◽  
George M. Hass
Keyword(s):  
Ionic Strength ◽  
Potassium Chloride ◽  
Phosphate Buffer ◽  
Phosphate Buffer Solution ◽  
Potassium Phosphate ◽  
Microscopic Structure ◽  
Buffer Solution ◽  
Buffer Solutions ◽  
Alkaline Side ◽  
Fresh Frozen

A new method for the isolation of large numbers of individual myofibrils from fresh mammalian skeletal and cardiac muscle has been described. Purification of isolated myofibrils was accomplished by differential centrifugation of fresh frozen sections of muscle which had been mechanically agitated after exposure for 30 to 45 minutes at 0°C. to the action of a dilute solution of trypsin in a phosphate buffer solution with a pH of 7.0 and an ionic strength of 0.25. Isolated skeletal myofibrils of the rabbit and man have similar constant solubility properties. They dissolve in an aqueous mixture of 0.5 N potassium chloride and 0.03 N sodium bicarbonate, giving viscous solutions which exhibit conspicuous birefringence of flow. They are soluble in buffer solutions (ionic strength 0.15) on the acid side of pH 4 and alkaline side of pH 10. If the ionic strength of potassium phosphate buffer solutions is increased to 0.5 or if the ionic strength of phosphate-borate buffer solutions is increased to a similar value by addition of potassium chloride, the isolated myofibrils become soluble at neutrality. Hence, it is possible, first to isolate the myofibrils and then dissolve them without deviating appreciably from physiologic ranges of pH. The extent to which myofibrils are modified by the conditions imposed by the method of isolation is unknown. There is no significant change in microscopic structure or optical birefringence. Furthermore, there is retention of a form of physiological reactivity, for when the isolated skeletal myofibrils are immersed in solutions of adenosinetriphosphate, they promptly and irreversibly change from elongated fibrils with distinct structural detail into dense spherical masses without recognizable microscopic structure.

scholarly journals Lithium transport pathways in human red blood cells.

1978 ◽  
Vol 72 (2) ◽  
pp. 233-247 ◽  
Author(s):  
G N Pandey ◽  
B Sarkadi ◽  
M Haas ◽  
R B Gunn ◽  
J M Davis ◽  
...  
Keyword(s):  
External Medium ◽  
Red Cells ◽  
Red Cell Membrane ◽  
Transport Pathways ◽  
Human Red Blood Cells ◽  
Lithium Transport ◽  
Ca Ions ◽  
Dominant Cation ◽  
Human Red Cells ◽  
In Cells

In human red cells, Li is extruded against its own concentration gradient if the external medium contains Na as a dominant cation. This uphill net Li extrusion occurs in the presence of external Na but not K, Rb, Cs, choline, Mg, or Ca, is ouabain-insensitive, inhibited by phloretin, and does not require the presence of cellular ATP. Li influx into human red cells has a ouabain-sensitive and a ouabain-insensitive but phloretin-sensitive component. Ouabain-sensitive Li influx is competitively inhibited by external K and Na and probably involves the site on which the Na-K pump normally transports K into red cells. Ouabain does not inhibit Li efflux from red cells containing Li concentrations below 10 mM in the presence of high internal Na or K, whereas a ouabain-sensitive Li efflux can be measured in cells loaded to contain 140 mM Li in the presence of little or no internal Na or K. Ouabain-insensitive Li efflux is stimulated by external Na and not by K, Rb, Cs, choline, Mg, or Ca ions. Na-dependent Li efflux does not require the presence of cellular ATP and is inhibited by phloretin, furosemide, quinine, and quinidine. Experiments carried out in cells loaded in the presence of nystatin to contain either only K or only Na show that the ouabain-insensitive, phloretin-inhibited Li movements into or out of human red cells are stimulated by Na on the trans side and inhibited by Na on the cis side of the red cell membrane. The characteristics of the Na-dependent unidirectional Li fluxes and uphill Li extrusion are similar, suggesting that they are mediated by the same Na-Li countertransport system.

Disk-Sphere Transformation in Mammalian Red Cells

1940 ◽  
Vol 17 (2) ◽  
pp. 117-127
Author(s):  
ROBERT F. FURCHGOTT ◽  
ERIC PONDER
Keyword(s):  
Fatty Acid ◽  
Cell Surface ◽  
Serum Albumin ◽  
Blood Cells ◽  
Phosphate Buffer ◽  
Phosphate Buffer Solution ◽  
Red Cells ◽  
Red Cell ◽  
Buffer Solution ◽  
Ph Values

1. Experimental evidence has been presented showing that crystalbumin (the carbohydrate-poor fraction of serum albumin) is the factor which prevents mammalian red blood cells from becoming spherical at pH values over 9.2. 2. The amount of crystalbumin taken up from a solution of it by red cells previously freed of it is of the order of 800 mg. per 100 c.c. of red cells. If this amount is all taken up at the red cell surface, it would form a layer only a few molecules thick. 3. The electrophoretic mobility in phosphate buffer of pH 7.4 is the same for cells containing crystalbumin, cells free of crystalbumin, cells with anti-sphering activity counteracted by fatty acid, and ghosts which have been temporarily sphered by a rise in pH. The mobilities in a saline-glycine buffer solution of pH 10.1 for the first three classes of cells just mentioned are also the same. The mobility of cells sphered with lecithin in a saline-phosphate buffer solution is the same as that for untreated discoidal cells.

Effect of N-ethylmaleimide on K transport in density-separated human red blood cells

1987 ◽  
Vol 253 (1) ◽  
pp. C7-C12 ◽  
Author(s):  
L. R. Berkowitz ◽  
D. Walstad ◽  
E. P. Orringer
Keyword(s):  
Red Blood Cells ◽  
Blood Cells ◽  
Red Cells ◽  
Sheep Red Cells ◽  
Human Red Blood Cells ◽  
High K ◽  
K Transport ◽  
Human Red Cells

N-ethylmaleimide (NEM) is a sulfhydryl-reacting agent known to stimulate chloride-dependent K transport in a variety of red cells. In high K sheep red cells, NEM-induced K movements are greater in magnitude in young cells compared with old cells. We hypothesized that human red cells might respond to NEM like high K sheep red cells. To test this idea, cells of various age were exposed to 0.5 mM NEM. We found that, after a 4-h incubation, young cells lost 50% of cell K, compared with 10% K loss in older cells. K loss in all fractions was inhibited by chloride replacement or furosemide.

Conformational response of the glycocalyx to ionic strength and interaction with modified glass surfaces: study of live red cells by interferometry

1983 ◽  
Vol 63 (1) ◽  
pp. 101-112
Author(s):  
H. Wolf ◽  
D. Gingell
Keyword(s):  
Brownian Motion ◽  
Ionic Strength ◽  
Red Cells ◽  
Adherent Cells ◽  
Ionic Strength Dependence ◽  
Human Red Blood Cells ◽  
Absolute Requirement ◽  
Strength Dependence ◽  
Finite Aperture ◽  
Glass Surfaces

We have measured separation distances between live human red blood cells and simple or modified glass surfaces, using the finite aperture technique of microscope interferometry. In general, separation increases as the ionic strength falls, in isotonic solutions. Restriction on movement parallel to the glass in all except the most dilute salt solutions, coupled with the absence of Brownian motion, indicates direct molecular contact with the substratum. Thus increased separation must be due to swelling of the glycocalyx under electrostatic forces. However, at approximately less than to 2mM adherent cells show a separation greater than 100 nm, execute Brownian motion and the restriction on lateral motion is less evident. This suggests that secondary minimum adhesion by long-range forces with little or no direct molecular connection occurs at extreme dilution only. Treatment of cells with trypsin reduces separation by up to 40 nm, but the extent to which this reflects reduced double-layer repulsion due to loss of surface charge, as opposed to the reduced opportunity for swelling in a trimmed-down glycocalyx, is unclear. Adhesion at a separation approximately 100 nm in 1 mM buffer after trypsinization supports the view that adhesion can occur without very long glycoprotein connections, but does not prove it. Adhesion to unwettable methylated glass and completely wettable unmethylated glass, with an identical ionic strength dependence of the separation, shows that hydrophilicity is not an absolute requirement. Red cells interact closely at all ionic strengths with glass made polycationic with poly-L-lysine, owing to electrostatic attraction. The interference technique also shows that adherent cells can be spaced from the glass by an intervening layer of previously absorbed serum albumin.

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