Stabilization of blood cyanide.

The measurement of blood cyanide is of clinical importance because, aside from the relatively few cases of accidental or selfinduced poisoning, raised levels are found in patients receMng the hypotensive agent sodium nitroprusside (1), in victims of smoke inhalation injury (2), and in lx~pulations at risk of ataxic mydopathy in areas of high cassava consumption C,). However, very few hospital laboratories have the means of measuring cyanide, and blood samples may need to be transported to one offering the service. As losses of cyanide may occur (/4,5) in transit, resulting in underestimation, a stabilizing technique would improve analytical accuracy. In one procedure, the blood is added to acidified silver sulfate. By subsequently increasing the acid concentration, the cyanide can be liberated from the silver cyanide and transferred to a small amount of sodium hydroxide sohttion with a stream of nitrogen (6). In another method, a small volume of sodium nitrite solution is added to the blood so that cyanide is retained as cyanmethemoglobin. It is possible then to separate the red cells, which contain all of the cyanide, and measure the cyanide in a protein-free extract without recourse to nitrogen aeration (7). A modification of the second procedure was adopted for a study of cyanide levels in patients with smoke inhalation injury admitted to casualty departments throughout the LJ.t;,. because blood samples were sent to the laboratory by mail. In order to determine the best anticoagulant/nitrite combination, 20 pL of a 50% aqueous solution of sodium nitrite was added to 5mL tubes containing four different anticoagulants (L.I.P. Shipley, West Yorkshire, tl.l(.) and dried in vacuo over anhydrous calcium chloride. ]\venty-five microliters of an isotonic saline solution of KCN (approximately 10raM) was added to 5 mL of blood in each of the 12 tubes (3 for each anticoagulant) and to two plain tubes containing 5 mL of 0.161 NaOH and stored at room temperature. The amount of hemolysis was checked, and the recovmy of cyanide was measured daily for three days. Aliquots (0.5 mL) of blood fi'om each tube were transferred to 2-mL graduated microcentrifuge tubes (Camlab, Cambridge, U.K.) and made up to 2 mL with isotonic saline. Following centrifugation at 12,000 rpm for5 rain, the cells were washed three more times with 2 mL of saline. The volume was then restored to 0.5 mL with saline and 5% trichloroacetic (TCA) layered on top, up to the 2-mL mark. Similarly, trichloroacetic acid (TCA) was added to 0.5-mL aliquots of the saline KCN diluted in NaOH. Following vortex mixing and centrifuging, the cyanide content of the TCA extracts was determined by means of an automated version of the Aldridge technique (8). The microcentrifuge tubes remained closed until immediately before analysis. It was apparent from the initial saline wash on the third day that a small amount of hemolysis had occurred in the blood samples anticoagulated with heparin and with Is whereas much more hemolysis had taken place with fluoride-oxalate. Because the citrate/nitrite combination produced no visible hemolysis and gave a reliable retrieval of cyanide (Figure 1 ), it was adopted for out study. 40