A method for gas chromatographic detection of codeine in serum is described. An application to a clinical drug evaluation study is given. A method for simultaneous detection of free and total codeine and its metabolites, norcodeine and morphine, in urine is also presented. SE OF GAS-LIQUID chromatography as a means of comparing serum levels in subjects receiving oral codeine dosages has been investigated. Although an efficient SEBO column enabled detection of nanogram quantities, codeine was not resolved from norcodeine on this nonpolar phase, as previously reported by Anders and Manneririg (I). Elliot et uZ. (2) used the acetates formed inside the column (1) to detect higher levels present in urine of narcotic addicts. The authors found small interfering peaks present in blanks which precluded the use of codeine acetate for reliable quantification of low levels present in serum. However, there is a great difference in reaction rate between codeine and norcodeine under mild acetylation conditions. This was used to remove norcodeine before injection on SE30 columns. The specificity and accuracy of the results were checked, using an XE6O column on which free codeine and norcodeine are well resolved.
,4 method for the estimation of codeine and its metabolites in urine is also described.
Methods
Codeine in Serum or Urine.-To 2.5 ml. of serum or urinc in a 15-ml. glass-stoppered conical centrifuge tuhe is added 9 ml. of 10% butanol in freshly distilled chloroform (3). The tuhe is hand shaken 5 sec. Then 0.12 rnl. of 16 N KOH is added (0.05 ml. for urine). Again mix by hand 5 sec., then mechanically shake in 10 rnin. After centrifuging, the aqueous scrum and jell layers are aspirated. If jell formation is excessive, the aqueous layer is aspirated and the jell broken by vigorous reshaking before recentrifuging. A 7-ml. aliquot of the organic layer is transferred to a clean centrifuge tube. Care must bc taken not to transfer any of the strongly basic aqueous layer. A quantity of 1.5 ml. of 0.01 N HnS04 is added. Shake 10 inin.