The effect of plasma on the reaction of cyclophosphamide with 4(-p-nitrobenzyl)-pyridine.

Cyclophosphamide is not structurally modified by blood plasma and may be recovered quantitatively from it. The apparent loss of alkylating ability of cyclophosphamide, as measured by its ability to react with 4-(p-nitrobenzyl)-pyridine (NBP), following treatement with plasma is not, as has been reported, due to metabolism of the drug but rather to an inhibition of the colorimetric reaction by a constituent of the plasma. This inhibition is strongly pH dependent, reaching 100% when the pH of the solution of cyclophosphamide in plasma is above 8, and falling to less than 20% when the pH is below 6, but extraction with chloroform at pH 7 separates cyclophosphamide from the inhibitor. Although the nature of the inhibitor has not been elucidated, its presence in plasma is of great importance in the quantitative determination of cyclophosphamide, and may also be of significance in the biological effects of cyclophosphamide and other alkylating agents in vivo.