Experimental evaluation of methods for assessing the carboxyhemoglobin (COHb) content in blood using gas chromatography, UV-visible spectrophotometry (three methods), and magnetic circular dichroism (MCD) has been made. From the point of view of measurements where the COHb content is low (less than 15%), elevated simply by the smoking population, the spectrophotometric method by Rodkey, et al. (7) emerges as the most satisfactory. MCD analysis, however, is potentially considerably simpler, not requiring chemical pretreatment of the blood, and provided that analysis time can be improved, is likely to replace the spectrophotometric method as the method of choice.
A systematic study of the effect of procedural variables on the ALA-D assay has been carried out. Because of metal content in vacutainers, artifactual results are obtained on blood samples drawn by this method and the method of choice is the use of plastic syringes and tubes. The use of isopropyl alcohol as a skin sterilizing agent, although it gives measurable blood alcohol readings, does not affect the ALA-D assay. The handling of the whole blood has been investigated and the two methods of hemolysis compared. Trichloroacetic acid, a reagent used in the final assay of porphobilinogen, has been found to be a serious interference in the method, making it essential for this reagent to be kept in a securely sealed container. The deterioration of the trichloroacetic acid--mercuric chloride reagent is also a potential procedural problem that must be considered.
The effect of blood alcohol on the pH-activity profile of ALA-D in humans and rabbits has been measured. The shape of the pH-activity profile remains unchanged, although enzyme activities are significantly affected (up to 40%). In order for the blood lead assay using ALA-D to remain independent of the presence of alcohol, an alternative methodology, such as a ratio of activities at two pH values, and not a measurement at a single pH value should be used. The effect of carbon monoxide on the pH-activity profile of ALA-D, on the other hand, is minimal.
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