A. Buursma

The oxygen binding capacity of human haemoglobin

SummaryThe internationally standardized HiCN method for measuring the total haemoglobin concentration of the blood (cHb*) is based on the chemical composition of the haemoglobin molecule. If all haemoglobin measured in the blood by this method were active, i.e., capable of reversibly binding O2, the O2 binding capacity (β) would be 1.39 ml · g−1 The experimental values of β to be found in the literature vary over a wide range, values as low as 1.24 ml · g−1 being no exception. We determined β for 36 blood samples of healthy men, using for the determination of the O2 content 3 independent methods, i.e., a manometric, a polarographic and a chemical one. Before the measurements the blood was equilibrated with humidified pure O2 for 3 h. Appropriate corrections for dissolved O2 were made. The mean β was 1.368 ml · g−1, with a s.d. of 0.017 ml · g−1 (median 1.372 ml · g−1; range 1.322–1.387 ml · g−1). This mean β corresponds to 98.4% active haemoglobin. The inactive haemoglobin consisted of 0.7% HbCO, Hi and SHb (“dyshaemoglobin”) and 0.9% unidentified inactive haemoglobin. In 4 samples a high fraction of unidentified inactive haemoglobin was found (2.8–4.3%). Taking into account that in addition to this fraction a considerable amount of dyshaemoglobin (especially HbCO) may be present in the blood of normal men, it must be concluded that calculating the O2 carrying capacity of the blood by multiplyingcHb* by a constant value of β, may be subject to an appreciable error. If β=1.39 ml · g−1 is used, the O2 carrying capacity of the blood may easily be found 10–15% too high.The internationally standardized HiCN method for measuring the total haemoglobin concentration of the blood (c Hb * ) is based on the chemical composition of the haemoglobin molecule. If all haemoglobin measured in the blood by this method were active, i.e., capable of reversibly binding O2, the O2 binding capacity (β) would be 1.39 ml · g−1 The experimental values of β to be found in the literature vary over a wide range, values as low as 1.24 ml · g−1 being no exception. We determined β for 36 blood samples of healthy men, using for the determination of the O2 content 3 independent methods, i.e., a manometric, a polarographic and a chemical one. Before the measurements the blood was equilibrated with humidified pure O2 for 3 h. Appropriate corrections for dissolved O2 were made. The mean β was 1.368 ml · g−1, with a s.d. of 0.017 ml · g−1 (median 1.372 ml · g−1; range 1.322–1.387 ml · g−1). This mean β corresponds to 98.4% active haemoglobin. The inactive haemoglobin consisted of 0.7% HbCO, Hi and SHb (“dyshaemoglobin”) and 0.9% unidentified inactive haemoglobin. In 4 samples a high fraction of unidentified inactive haemoglobin was found (2.8–4.3%). Taking into account that in addition to this fraction a considerable amount of dyshaemoglobin (especially HbCO) may be present in the blood of normal men, it must be concluded that calculating the O2 carrying capacity of the blood by multiplyingc Hb * by a constant value of β, may be subject to an appreciable error. If β=1.39 ml · g−1 is used, the O2 carrying capacity of the blood may easily be found 10–15% too high.

A Multi-Wavelength Spectrophotometric Method for the Simultaneous Determination of Five Haemoglobin Derivatives

A method is described by which the concentration of deoxyhaemoglobin, oxyhaemoglobin, carboxyhaemoglobin, haemoglobin and sulfhaemoglobin in a human blood sample is determined by passing the haemolysate without air contact through a coarse filter and subsequently measuring the absorbance at lambda = 500, 569, 577, 620 and 760 nm. The ensuing set of equations is solved by matrix calculation with the aid of a simple computer program. The method has been tested by comparing it with conventional methods for the determination of the various haemoglobin derivatives separately.

Carboxyhaemoglobin. spectrophotometric determination tested and calibrated using a new reference method for measuring carbon monoxide in blood

The spectrophotometric determination of HbCO at lambda = 562 and 540 nm in the system HbCO/HbO2 was reinvestigated using a new reference method for measuring CO in blood. This reference method is based on the conversion of CO from HbCO into CO2 which is determined by titration. Plotting the absorbance ratio A562/A540 against the titrimetrically determined HbCO fractions of 46 blood samples demonstrated a linear relationship up to 90% HbCO and yielded more accurate values for the constants in the equation for calculating the HbCO fraction from A562/A540. The standard deviation of the differences between the spectrophotometric and the titrimetric method was 1.2% HbCO. It is shown that the influence of other haemoglobin derivatives and other possible sources of error is either negligible or can be prevented by simple precautions.

A NEW TREND IN BLOOD-GAS CHEMISTRY - THE MEASUREMENT OF CLINICALLY RELEVANT HEMOGLOBIN DERIVATIVES - PERFORMANCE OF THE OSM3 HEMOXIMETER

Supplementing the determination of total hemoglobin (CHb*) and oxygen saturation (SO2) with measuring the fractions (F) of inactive hemoglobins (dyshemoglobins: carboxyhemoglobin, HbCO; methemoglobin, Hi; sulfhemoglobin, SHb) in human blood is becoming common practice in many hospitals. We tested the performance of a new instrument for this purpose, the hemoximeter OSM3 (Radiometer, Copenhagen, Denmark) by comparing the results with those of an established multiwavelength method (MWM). For 100 fresh blood samples from patients with SO2 ranging between 20 and 100%, the difference between the two methods (OSM3-MWM) was 0.90% SO2 +/- 1.14% (SD). A series of 214 consecutive blood samples from patients with FHbCO ranging from 0 to 12% showed for HbCO a difference (OSM3-MWM) of 0.03% FHbCO +/- 0.47% (SD). In the same blood samples FHi was between 0 and 1.4%. The OSM3 gave a mean value of 0.81%, the MWM 0.47%. In order to put the OSM3 to a more severe test we made from normal human blood several series of specimens with high concentrations of dyshemoglobins. The difference between OSM3 and MWM was for 37 specimens of blood with 1-60% HbCO 0.29% FHbCO +/- 0.96% (SD), and for 34 specimens of blood with 0-70% Hi -0.29% FHi +/- 2.29%. For specimens with high fractions of HbCO and Hi the agreement between the SO2 measurement made with the two methods remained good, even when the dyshemoglobins were present in combination. Hi fractions of up to 70% did not interfere with the measurement of HbCO; the same holds good for the measurement of Hi in the presence of HbCO.

Quality control in haemoglobinometry with special reference to the stability of haemiglobincyanide reference solutions

In haemoglobinometry grave errors are still being made even though an internationally accepted standardized method is available for the determination of the haemoglobin content of blood. Up to the present only haemiglobincyanide reference solutions have been available on a wide scale to check the measuring stage of the standardized haemiglobincyanide method. These reference solutions are shown to remain stable, under proper storage conditions, for more than 10 years. Concentrated haemoglobin solutions have become available recently, offering the possibility to control the dilution and conversion steps of the haemiglobincyanide method. Such a solution is shown to remain stable, under proper storage conditions, for at least a year. Using both haemiglobincyanide reference solutions and concentrated haemoglobin solutions, as well as having the possibility of checking the cyanide content of the reagent used, an acceptable intra-laboratory control program may now be set up.

The use of atomic absorption spectrophotometry for the measurement of haemoglobin-iron, with special reference to the determination oF εHiCN540

Abstract The possibility of using atomic absorption spectrophotometry for the determination of haemoglobin-iron has been investigated. Evidence is given that both the presence of inorganic substances and handling of blood samples can influence the results. The method thus may be prone to systematic errors. The technique is considered unsuitable for the absolute measurement of haemoglobin-iron, thus yielding no arguments requiring reconsideration of the internationally accepted value of 11.0 for e HICN 540 . A table summarizing the experimental evidence on which this value is based, is given.

Carboxyhemoglobin: Determination and significance in oxygen transport

Apart from being the direct cause of quite a few cases of near-fatal poisoning, carbon monoxide is a very common noxious agent functioning as an additional factor in human disease. The extent of the possible role of carbon monoxide in human pathology is demonstrated by the following data from the Diakonessenhuis Groningen, a 400-bed private hospital1. During a five-month period the carboxyhemoglobin fraction (FHbCO) was measured in the blood of all pre-operative patients. In 64.4% of the 1358 cases FHbCO was 10%. The highest value measured in this series was 15.5%. Thus it appears that in a considerable number of patients HbCO fractions are present that are of possible pathophysiological significance. This shows the practical importance of the pathophysiology of carboxyhemoglobin and the need for easy and reliable methods for the determination of FHbCO in blood.