Stopped-flow measurements of CO and O<sub>2</sub> uptake by hemoglobin in sheep erythrocytes

Sirs, John A.; Roughton, F. J. W.

doi:10.1152/jappl.1963.18.1.158

Cited by 53 publications

(18 citation statements)

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“…This in turn was adapted from the system used by Sirs & Roughton (1963). It consists of a simple optical system, the stopped-flow and rapid-mixing unit, and a photomultiplier, amplifier and recorder.…”

Section: Methodsmentioning

confidence: 99%

The rate of osmotic influx of water by flexible and inflexible erythrocytes

Sirs¹

1969

The Journal of Physiology

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Section: Methodsmentioning

confidence: 99%

The rate of osmotic influx of water by flexible and inflexible erythrocytes

Sirs¹

1969

The Journal of Physiology

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“…For studies of the effect of butylurea on the rate of red cell oxygenation, 0.1 M butylurea was added to the buffers in which Aliquots from the two syringes,.one containing the red cell suspension, the second containing the oxygenation or deoxygenation buffer, were driven simultaneously into a mixing chamber, of eight 1-mm diameter tangential jets (12,20) leading to a 2-mm diameter, 2-cm long observation tube. The apparatus was contained in a thermally regulated water bath.…”

Section: Methodsmentioning

confidence: 99%

“…Improvements in the detections system by Legge and Roughton (8) and by Forster et al (9) were followed by contributions from the laboratories of Gibson (10), Roughton (11), and Forster (9). Subsequently, Sirs and Roughton (12) constructed an apparatus for the study of red blood cells, combining the split-beam detection system (9) with the stopped-flow principle. This apparatus or modifications of it have been used by later workers.…”

Section: Abstracimentioning

confidence: 99%

Ligand kinetics of hemoglobin S containing erythrocytes.

Harrington¹,

Elbaum²,

Bookchin³

et al. 1977

Proc. Natl. Acad. Sci. U.S.A.

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ABSTRACIOxygen uptake of fully deoxygenated sickle (SS) erythrocytes is slower than that of normal (AA) erythrocytes, as demonstrated by the half-times of the overall oxygenation reactions: at 250 in an isotonic phosphate buffer the normal red cells have a tl/2 = 82 ± 4.7 msec, as compared to sickle red cells where t1/2 = 135 i 17.6 msec. The effects of temperature, extracellular osmolality, and the presence of an antisickling agent (n-butylurea) on the rate of red cell oxygenation strongly suggest that the differences in oxygenation rates encountered with sickle red cells is directly related to the intracellular polymerization of deoxyhemoglobin S. Erythrocytes from persons with sickle cell anemia have an abnormal oxygen equilibrium curve, reflecting an increase in the partial pressure of oxygen needed to half-saturate the available binding sites (1-3). The mechanism of this shift is not clearly established, but it is connected with the intraerythrocytic polymerization of the deoxy form of hemoglobin (Hb) S (4, 5).Rates of oxygen and carbon monoxide uptake by red blood cells were first measured 50 years ago by Hartridge and Roughton (6, 7), using the reversion spectroscope and their continuous flow, rapid reaction technique. Improvements in the detections system by Legge and Roughton (8) and by Forster et al. (9) were followed by contributions from the laboratories of Gibson (10), Roughton (11), and Forster (9). Subsequently, Sirs and Roughton (12) constructed an apparatus for the study of red blood cells, combining the split-beam detection system (9) with the stopped-flow principle. This apparatus or modifications of it have been used by later workers.The findings of Hartridge, Roughton, Gibson, Forster and their colleagues have been summarized by Roughton (13,14) and by Forster (15). The pure chemical kinetics of the reactions of Hb in solution and in the red cell appear to be the same. Nonetheless, the rates of reaction of Hb in the red cell are slower than in solution. It is noted that the discrepancy between the rate of a given process in Hb solutions and in the red cell suspension increases pari passu with the speed of the reaction in solution. The rate of the fast process, combination of Hb with oxygen, is 30-fold slower in the red cell at 370 than in solution.Since the transit time for the erythrocytes in the pulmonary capillaries is about 700-800 msec, several questions arise regarding oxygen exchange in the Hb S-containing red cells that could have physiological significance. To approach these questions we have utilized a custom-made stopped-flow apparatus specially designed for the study of red cell ligand kinetics. Our investigations explored the effects of variations in temperature and in extracellular osmolality, and the effects of an antisickling agent (n-butylurea) on the rate of red cell oxygenation. In addition, a high-density fraction of sickle red cells rich in irreversibly sickled forms (ISC), which possess rigid membranes (17), were compared to the low density fraction to help assess the ...

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“…The rate of the reaction of carbon monoxide with fully oxygenated haemoglobin was measured with the stopped-flow rapid-mixing method of Sirs & Roughton (1963). While being mixed the haemoglobin solution was simultaneously diluted 1:5 with solutions equilibrated with carbon monoxide (Sirs, 1966).…”

Section: Methodsmentioning

confidence: 99%

The effect of temperature on the reaction of carbon monoxide with oxygenated haemoglobin.

Sirs¹

1976

The Journal of Physiology

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SUMMARY1. The rate at which carbon monoxide displaces oxygen from its combination with haemoglobin in solution, has been measured spectrophotometrically, using a rapid-mixing stopped-flow technique.2. In the presence of carbon dioxide, the reaction proceeds by a unimolecular dissociation, with a rate constant r.3. The relationship of the reciprocal of r to the ratio Po2IPco is non-linear, and a different curve is obtained at each carbon monoxide concentration.4. From measurements of the rate constant at temperatures between 5 and 350 C, it is concluded that the non-linearity is due to the formation at a finite rate of an intermediate complex, CO2Hb4O6, by the reaction of carbon dioxide with Hb4O6.

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Stopped-flow measurements of CO and O₂ uptake by hemoglobin in sheep erythrocytes

Cited by 53 publications

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The rate of osmotic influx of water by flexible and inflexible erythrocytes

The rate of osmotic influx of water by flexible and inflexible erythrocytes

Ligand kinetics of hemoglobin S containing erythrocytes.

The effect of temperature on the reaction of carbon monoxide with oxygenated haemoglobin.

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Stopped-flow measurements of CO and O2 uptake by hemoglobin in sheep erythrocytes

Cited by 53 publications

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The rate of osmotic influx of water by flexible and inflexible erythrocytes

The rate of osmotic influx of water by flexible and inflexible erythrocytes

Ligand kinetics of hemoglobin S containing erythrocytes.

The effect of temperature on the reaction of carbon monoxide with oxygenated haemoglobin.

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Stopped-flow measurements of CO and O₂ uptake by hemoglobin in sheep erythrocytes