Teflon. a sul~posedly inert surface: yields tailing peaks for the aliphatic amines. I t would appear, therefore, that the base must be altering the behavior of the liquid phase. I t is immaterial n-hether the base is dissolved in the 9 x 9 and the mixture added to the support or whether the base and the s ~ -are 9 on the cjupl,ort separately, so long as the requisite amount of base is present.
ACKNOWLEDGMENT
The writers thank the Dow Chemical for the ’Owfax 9F9.
cO*
LITERATURE CITED
(1) Decora, A. W.) Dineen, G . U., “Gas
Chromatography,” H. J. Soebels, R. F. Wall, S . Brenner, eds., p. 33, Academic Press, Sew York, 1961.
( 2 ) Landault, C., Guichon, G . J J . Chromatog. 9, 133 (1961J. (3) Smith, E. D.>liadford, It. I>., * \ S A L CHEM. 33, 1161 (1961).
(4) Sze, Y. L., Borke, 11. L., Ottenstein, D. Ibid., 35, 240 (1963).
RECEIVEDfor review April 9, 1964. Accepted August 5, 1964. This investigation was supported by research grant GlI-10064 from the Public Health Service of the U. S. Department of Health, Education, and Welfare.
Determination of Diazepam (Valium) in Blood by Gas Liquid Chromatography J. ARTHUR F. de SILVA, MORTON A. SCHWARTZ, V. STEFANOVIC, J. KAPLAN, and L. D’ARCONTE Department o f Pharmacology, Hoffmann-la Roche Inc., Nutley, N. J .
b A method for the! determination of diazepam (Valium) in whole blood of humans involves selei:tive extraction of diazepam into ether, acid hydrolysis, and analysis b y gas liquid chromatography. The methocl has high sensitivity (0.02 to 0.03 pg. per ml. of blood), i s specific for diazepam after it i s hydrolyzed to 2-methylamino-5chlorobenzophenone, and has a very acceptable recovery and reproducibility of the order of 94% + 5.0. The method has been successfully applied to the determination of diazepam in samples obtained from a variety of clinical studies including a blood level fall-off study and the placental transfer of diazepam in expectant mothers. Blood levels of diazepam and i t s N-demethylated metabolite can be determined in a single assay and the method i s also applicable to the determination of chlordiazepoxide-HCI (Librium) in blood.
D
i-chIoro-1,3-dihydro-lmethyl - 5 - phenyl - 2H - 1,4benzodiazepin-2-one (Valium) is a new psychot~herapeutic drug of the 1,4benzodiazepine class: of compounds (11). Its pharmacological and clinical properties have been reported by Randall et al. ( 8 ) . Although diazepam is chemically related to chlordiazepoxide (Librium) it’ differs from the latter in that’ it contains a meth!.lated-S at, position 1, and both the basic side chain a t ~jo>ition2 and the -Y-oxide function a t pmition 4 (Figure I ) . (Valium and Lihrium are registered trademarks of Hoffniann-La Roche Inc. for the phariiiawutical forinu1atio:ns of diazepam and chlordiaze;,ositie-HCl, respectively.) Because of these differences, diazepam IAZEPAM,
cannot be determined by the analytical procedures developed for chlorodiazepoxide. hydrolyhis to an aromatic primary amine which is measured by a Bratton-Marshall procedure ( 7 ) or coniersion to a lactam which is determined spectrofluoronietrically ( 5 ) . Studies by Schnartz, Koechlin, and I
