P. Rispens

The apparent first dissociation constant of carbonic acid in plasma between 16 and 42.5

Abstract The variation with pH and temperature of the apparent first dissociation constant of carbonic acid in plasma has been studied using normal human plasma. A new tonometry set-up was used for equilibrating plasma with mixtures of carbon dioxide and oxygen, CO2 concentration 2–16%, at 42.5, 37.5, 35.0, 32.5, 30.0, 26.0, 20.0 and 16.0°. Total carbon dioxide concentration in plasma was measured by means of the cediometer, plasma pH and CO2 concentration of tonometry gas was measured using standard methods. The dissociation constant calculated from these measurements is denoted by the symbol p K ′ 1 . A total of 555 p K ′ 1 determinations have been performed in duplicate, the reproducibility being 0.008 (s.d.). p K /t 1 can be described as a function of pH and temperature by means of the equation p K ′ 1 = −47416+ 1840.141 T +0.15906T−log(1+ 0.020682 10 −pH+7 ) The agreement between experimentally determined and calculated p K ′ 1 values is within 0.01379 (s.d.). The abnormal values of p K ′ 1 recently reported for seriously ill patients, found when using CO2 electrodes for measuring blood PCO2, are discussed and supposed to be caused by errors in the determination of pH, PCO2 or [HCO3−].

INHIBITION OF CARBONIC-ANHYDRASE IN DOG PLASMA

Using a pH stat method, we measured the activity of carbonic anhydrase (CA) from dog erythrocytes in the presence of various amounts of dog plasma. A plasma factor appeared to be able to inhibit about 86% of the total CA activity, corresponding to the relative activity of CA II. Naiodoacetate was shown to inhibit the total CA activity up to about 13%, corresponding to the relative activity of CA I. Cl− inhibited the total CA activity up to about 20%, presumably mainly through its strong influence on Ca I. It is concluded that with a degree of haemolysis of up to 3%, no appreciable plasma CA activity will occur.

Determination of extracellular fluid volume in the dog with ferrocyanide

SummaryThe suitability of ferrocyanide as an indicator for the measurement of extracellular fluid volume was tested. Added ferrocyanide could be recovered completely from urine, plasma and blood. Inin vitro experiments ferrocyanide did not penetrate into erythrocytes, nor did it adhere to the red cell membrane. In gel filtration and electrophoresis experiments binding of ferrocyanide to plasma proteins could not be demonstrated. Inin vivo experiments on dogs, the urinary recovery of intravenously administered ferrocyanide was 98.9±2.1% (n=14). The partition ratio of ferrocyanide between lymph water and plasma water was 0.99±0.02 (n=20). Ferrocyanide could not be detected in cerebrospinal fluid or red cells of dogs after administration by intravenous infusion. No untoward effects of the infused ferrocyanide were observed during or after the experiments.In nephrectomized dogs ferrocyanide reached its ultimate distribution volume 2 hrs after intravenous administration of a single dose and remained constant for up to 10 hrs. The average distribution volume was 224±17 ml· kg−1 body mass (n=6). In intact dogs continuously infused with indicator, ferrocyanide also reached its ultimate distribution volume in 2 hrs and remained constant thereafter for up to 7 hrs after the start of the infusion. The average distribution volume was 237±27 ml· kg−1 body mass (n=14).It is concluded that ferrocyanide fulfils the requirements to be met by an indicator for the measurement of the extracellular volume, and is well suited for repeated determinations of the extracellular fluid volume in one and the same experiment.

Determination of total carbon dioxide in blood and plasma by means of the cediometer. Theory and experimental verification

Abstract The cediometer, an instrument for the determination of total carbon dioxide in blood and plasma samples, consists of a closed system incorporating an electrode chamber filled with a NaHCO 3 solution, a sample chamber in which the CO 2 of injected blood or plasma is set free with an acid reagent and a roller pump which circulates the CO 2 -containing gas until the P CO 2 , has become uniform throughout the system and a stable pH in the electrode chamber solution has been reached. The relationship between this pH and the total carbon dioxide content of the sample has been formulated exactly. For an arbitrarily chosen standard system with constant volumes of the three parts of the circuit, constant volume of the sample injected and constant temperature as well as constant pre-injection pH of the electrode chamber solution, the relationship of final pH and total carbon dioxide of the samples was calculated and presented in a table. This table can be used with any cediometer to convert the final pH into total carbon dioxide content. A simple calibration procedure using Na 2 CO 3 reference solutions can be used. The method is accurate within ± 1%, when the volumes of the various parts of the circuit do not differ by more than 10% from those of the standard system. The apparatus needs no special care and performs well under ordinary clinical conditions.

CALCULATED CHANGES IN PH AND PCO2 IN ARTERIAL BLOOD-PLASMA ASSUMING ABSENCE OF ION AND WATER EXCHANGE BETWEEN PLASMA AND ERYTHROCYTES DURING THEIR EQUILIBRATION WITH ALVEOLAR GAS

As a contribution to solving the problem of pH disequilibrium in arterial blood, the results of two modes of gas exchange in the lung have been calculated using an equilibrium state model of the blood. In both cases the HCO3−/Cl− exchange was assumed to occur after the gas exchange in the pulmonary capillaries. When the gas exchange was assumed to be dependent on intra-erythrocytic carbonic anhydrase. the plasma pH in the arterial blood increased. Whe plasma and erythrocytes were assumed to equilibrate separately with the alveolar gas due to presence of extra-erythrocytic carbonic anhydrase, plasma pH in the arterial blood decreased. In each case there was a slight increase inpCO2 after the blood had left the pulmonary capillaries.

A direct method for the determination of the HCO3-concentration as total carbon dioxide in blood and plasma

A method is described to determine the [HCO3−] as total CO2 in blood or plasma samples rapidly and accurately. By the addition of acid to the sample all CO2 is released into the gaseous phase. The released gas is led through a bicarbonate solution in a closed circuit and the change in pH of this solution is recorded. The CO2(tot) content is then read directly from a table relating ΔpH to [CO2(tot)], for which an equation is given. The accuracy of the method is shown to be approximately 1%, the time necessary for a determination is but 3 min. The method easily lends itself to semi-automation.

PITFALLS IN ACID/BASE EXPERIMENTS WITH CONSCIOUS DOGS

Arterial pH and blood gases were measured at intervals in conscious dogs after their first human contact of the day. Blood was sampled through an indwelling catheter in the aorta without disturbing the animals. It appeared that in the first 90 min arterial PO2, oxygen saturation and haemoglobin concentration significantly declined. PCO2 and pH changed less consistently when the acid/base status of the dogs was normal, but when a non-respiratory acidosis was present there was a significant decrease in pH and a significant increase in PCO2. Arterial pH and blood gases were also measured before and after feeding the animals. It appeared that an appreciable metabolic alkalosis developed within 2 h after a meal. The “alkaline tide” was accompanied by a trend to higher values for PCO2. It is concluded that, after a period of seclusion, renewed human contact causes behavioural changes in a dog, which may result in appreciable transitory changes in arterial pH and blood gas values. Blood sampling from conscious dogs should therefore take place after a proper period of habituation; preferably, a few samples should be taken at intervals to check that a steady state has been reached. If possible, blood should be collected before feeding; in any case the relationship in time of blood sampling to feeding should be constant throughout.

Horizontal rotating tonometers for the equilibration of blood or plasma with gasmixtures at constant temperatures

SummaryThe construction of a thermostatted tonometry unit allowing the simultaneous equilibration of six blood or plasma samples with different gas mixtures is described. The tonometers rotate along their horizontal axis, insuring a large equilibration surface, resulting in equilibration with CO2 in the gasphase being reached within 10 min. Samples may be introduced into or withdrawn from the rotating tonometers at any time. The gas mixtures leaving the tonometer may also be analyzed at any moment.

A new reference method for the determination of the total CO2 concentration in biological fluids.

It appeared that a part of a measuring system recently developed for the determination of the oxygen content of blood (Dijkhuizen, P., Kwant, G. and Zijlstra, W.G. (1976) Clin. Chim. Acta 68, 79), was perfectly suitable for measuring the total CO2 content of blood, plasma or other fluids. CO2-free room air is pumped through an extraction vessel in which all the CO2 of the sample is set free by an acid reagent, and swept by the carrier gas to a titration vessel containing a BaCl2 solution. CO2 is bound as BaCO3 and the ensuing H+ titrated with an NaOH solution. The method was tested by measuring a series of Na2CO3 reference solutions. The values measured by titration amounted to 99.4 +/- 0.8% of the concentration of the reference solutions (range 10--50 mmol 1(-1). The coefficient of variation was 1.8% for 5 mmol 1(-1) solutions and 0.2% for 50 mmol 1(-1) solutions. In measuring a series of 60 blood samples the coefficient of variation as calculated from duplicate determinations was 1%.

Influence of changes in cardiac output on the acid-base status of arterial and mixed venous blood

While maintaining the arterial CO2 tension constant near the normal level of the dog (4.3 kPa), we studied the influence of decreasing cardiac output on both the arterial and mixed-venous blood acid-base status in anaesthetized, artificially ventilated dogs. Cardiac output was manipulated by applying positive end-expiratory pressure (PEEP), and by β-adrenergic blockade to suppress a compensatory heart rate response. The systemic vascular response was attenuated by α-adrenergic blockade. Metabolic rate remained virtually unchanged when cardiac output decreased. Under these conditions a fall in cardiac output led to a shift of the arterial acid-base status in the direction of a metabolic acidosis. The changes occurring in the mixed-venous blood resembled those of an in-vivo CO2 bufferline, with the shift being such as if a respiratory acidosis was developing.