routinely to ensure no contamination. Second, simultaneous quantitative multielement analysis of potassium through strontium is achieved in an irradiation time of 10-20 minutes. The analysis is nondestructive, so that the sample may be used to determine concentration of other elements, which are not determined by this method. Finally, the type of instrumentation used, lends the method to routine analyses (e.g.7 quality. control type analysis), on the order of 3 to 4 per hour. Thus, the analysis of various pharmaceuticals or analysis of repetitive environmental samples (i.e,, sediment, plants, fish) can be greatly simplified.
LITERATURE CITED (1) K. A. Hardy, R. Akseisson, J. W. Nelson, and J. W. Winchester, Environ. Sci. Techno/.,IO, 176 (1976). (2) R. G. Flocchini et al., Environ. Sci. Techno/., io, 76 (1976). (3) "X-Ray Energy Spectrometry", Rolf Woldseth, Kevex Corporation, Burlin-
game, Calif. (1973).
(4) R. D. Giauque, F. S. Goulding, J. M.Jaklevic, and R. H. Pehl, Anal. Chern., 45, 671 (1973). (5) PierreMarmierand Eric Sheldon, "Physics of Nucleiand Particles", demic Press, New York, 1971.
RECEIVEDfor review September 16, 1976. Accepted November 29,1976.
Identification of Barbiturates by Chemical Ionization and MassAnalyzed Ion Kinetic Energy Spectrometry E. Soltero-Rigau, T. L. Kruger, and R. G. Cooks* Department of Chemistry, Purdue University, West Lafayette, Ind. 47907
The electron impact (El) and chemlcal Ionization (CI) mass spectra of a number of barbiturates and the ion kinetic energy spectra of some ions present in these mass spectra have been recorded. The El mass and MIKE spectra are not distinctive for all the barbiturates. These compounds can, however, be distinguishedby the MIKE spectra of the protonated or ethylated molecules generated by CI.
Numerous techniques have been employed for the identification of barbiturates (1-3).Recent reviews ( 4 , 5 )and current applications of some of these techniques are reported in the literature: thin-layer chromatography (6-9), color profile test (IO),microphase extraction ( 1 1 ), luminescence (12,13), high speed ion-exchange chromatography (14), nuclear magnetic resonance (15),and high pressure liquid chromatography (16,17).Preeminent among methods of barbiturate analysis has been gas-liquid chromatography (GC). This technique is a valuable tool for drug analysis but extraction of barbiturates from biological samples presents a problem in GC since other biological compounds also extracted interfere in the analysis (18-21). To avoid detecting these interferences, specific detectors (22-24) such as the Hall electrolytic conductivity detector have been used. Identification of the barbiturates directly as free acids or sodium salts is further complicated by the fact that they are strongly adsorbed on GC columns (25).To overcome this problem the solid support may be deactivated (26).Alternatively, derivatization may be accomplished either prior to GC analyses (27) or by the "oncolumn" derivatization technique (28,29).Alkyl (20,27-34) and silyl derivatives (29,35)have been prepared. The occurrence of side reactions in derivative formation has been a further problem (28, 36, 37). In analyses by mass spectrometry (MS), sample extracts or the pure compounds can be introduced directly, thus avoiding derivatization and other problems associated with GC columns and detectors. Different ionization sources have been employed: E1 (38,39),CI (40,41),FI ( 4 2 ) ,FD ( 4 2 ) ,and API (43).The E1 mass spectra of barbiturates do not always show molecular ions but the CI spectra show the presence of protonated molecular ions from which the molecular weight can be determined. However, neither methane nor isobutane reagent gas allows isomers or other compounds with the same
molecular weight to be distinguished. Hunt (41)used CIMS with an argon/water mixture as reagent gas for distinguishing the isomers pentobarbital and amobarbital. Although gas chromatography and mass spectrometry are by themselves often insufficient to provide a reliable identification of barbiturate mixtures, the combination of gaschromatography mass spectrometry (GCMS) has proved to be a more valuable tool for the identification and quantification of these compounds (34,44:i.Applications of GCMS to barbiturate analyses have used EII(33,44),CI (45),and API. The GC retention time is particularly useful when a CI source is used so that isomers may be distinguished. The aim of the present work is to apply the technique of mass-analyzed ion kinetic energy spectrometry (MIKES) (46, 47) to the identification of barbiturates. The MIKES technique and its application to mixture analysis have been described previously (48-51). Reference 51 summarizes the state of the art including limits of detection, accuracy, and methodology. Structural identification of the mass-selected ion is best made by collision-induced dissociation (CID) although spontaneous (metastable) ion dissociations also give useful results (52). The fundamental question is whether this method, in which an energy spectrum is taken on a selected ion(s), gives results which uniquely characterize each individual barbiturate. To this end the set of barbiturates chosen for analysis (Table I) includes many isomers and homologues which are usually difficult to identify. EXPERIMENTAL Two barbiturate kits containing commercial analytical standards were obtained from Applied Science. 'The samples were used without further purification. However, no significant impurities were found as judged from the mass spectra. All data, except the E1 mass spectra, were recorded using a reverse sector mass spectrometer (MIKES) (46, 4 7 ) . The E1 mass spectra were taken in a Hitachi RMU-6 single focusing mass spectrometer operated a t an ionizing electron energy of 70 eV. Introduction of samples was via a solid probe inlet system and volatilization was accomplished by slowly heating the source. These E1 spectra were in good agreement with the E1 spectra obtained with the MIKE spectrometer. The MIKES was operated at 70 eV, 125-150 O C and an accelerating voltage of 7 kV. Operating conditions for the CI source were: an electron-ionizing energy of 500 eV, 100-125 "C, an electron emission current of 0.080.14 mA and an accelerating voltage of 7 kV (except secobarbital which was run at 6 kV). Methane was used as reagent gas a t an indiANALYTICAL CHEMISTRY, VOL. 49, NO. 3, MARCH 1977
435
_ _ _ _ _ _ _ ~
Table 1. Structures and Molecular Weights of Barbiturates Studied
Barbiturate Hexobarbital Alphenal Phenobarbital Mephobarbital Allobarbital Aprobarbital Butalbital
Rl
Other name
R2
Hexenal Methyl Propenal Allyl Luminal Ethyl Mebaral Ethyl Dial Allyl Alurate Allyl Sandoptal Allyl Itobarbital Secobarbital Seconal Allyl Barbital Verona1 Ethyl Butabarbital Butisol Ethyl Butethal Neonal Ethyl Pentobarbital Nembutal Ethyl Amobarbital Amytal Ethyl aValues obtained from The Merck Index (1968).
1-Cyclohexenyl Phenyl Phenyl Phenyl Allyl Isopropyl Isobutyl sec-Pentyl Ethyl sec-Butyl Butyl sec-Pentyl Isopentyl
'b
R,
Molecular weight
Methyl H H Methyl H H H
236 244 232 246 208 210 224
H H H H H H
238 184 212 212 226 226
Class
i i i i ii ii 11 11 ...
111
iii iii iii ...
111
50
41 I
504
20
60
100
140
180
220
T I
I
'i'
I,
180
~
,%. ; 2"
220
dl 01)
2
5 1 L 180
161)
100-
220
-31
20
Mass Figure 1. Electron impact mass spectra of the allyl-type barbiturates: (a) allobarbital, (b) aprobarbital, (c) butalbital, and (d) secobarbital
cated source pressure of 5 X Torr which corresponds to an ion chamber pressure of approximately 1 Torr. Sample was introduced on a direct insertion probe and the magnet was set to the mle value corresponding to (M + 1)+or (M 29)+. Then the source pressure was varied slowly to obtain the methane-to-sample ratio where maximum protonation or ethylation occurs.
+
436
ANALYTICAL CHEMISTRY, VOL. 49, NO. 3, MARCH 1977
The low-pressure MIKE spectra of selected ions were obtained by scanning the electric sector voltage. Introduction of collision gas (air) into the second field-free region at an indicated pressure of 5 X IO-b Torr allowed high pressure MIKE spectra to be recorded. The energy axis in these spectra can be converted to mass units by the simple equation: E2 = (mz/ml)E1.High resolution ion kinetic energy scans
Table 11. EI/MIKES Results of Barbiturates Barbiturate
MIKES of
Hexobarbital Alphenal Phenobarbital Mephobarbital Allobarbital Aprobarbital
236 244 232 246 208 210b
Butalbital Secobarbital Barbital Butabarbital Butethal Pentobarbital Amobarbital
181
Fragment loss and relative abundanceu 15 15 15 15 15 15 43 43 15 15 42 17 42 17
197 156 183 184 197 198
(0.86) (0.06); 17 (0.05); 29 (1.8) (0.02); 28 (0.40) (0.15); 28 (2.2) (0.52); 29 (0.28); 43 (0.54) (7.7); 28 (0.04); 29 (0.06); 4 1 (1.6); 42 (0.42) (0.33); 56 (0.15) (0.05) (0.15); 29 (0.03); 42 (0.20); 56 (0.30) (0.67) (0.19) (1.88); 29 (0.08); 42 (0.24); 43 (0.05) (0.19); 56 (0.20) (0.44); 43 (0.04); 56 (0.38)
aPercent of metastable ion relative to the ion scanned, bMolecular ion.
IO(
a1 41
100
1,
50
I
60
.
100
140
1;
:
I80
"[
226
!
2 20
212
180
220
7-
156
-3
IOC
29
I?
50
Iel2
20
Mass Figure 2.
60
100
I
Mass
Electron impact mass spectra of the ethyl-type barbiturates: (a)butabarbital, (b) butethal, (c)pentobarbital,and (d) amobarbital
were taken to verify the fragment ion masses and to allow calculation of the abundances of the metastable and CID peaks relative to the main ion beam. The relative abundances were measured in terms of
peak heights.
RESULTS A N D DISCUSSION The results are conveniently discussed in terms of three classes of barbiturates (Table I):
(i) hexobarbital, alphenal, phenobarbital, and mephobarbital (ii) allyl-type barbiturates (iii) ethyl-type barbiturates In this way barbiturates will be discussed in order of the increasing difficulty encountered in their analyses. The types of scans used for the analysis include E1 and CI mass spectra and MIKE spectra on ions generated by E1 and by CI. The ANALYTICAL CHEMISTRY, VOL. 49, NO. 3, MARCH 1977
437
Table 111. Low and High Pressure CI/MIKE Spectra of the (M
+ H)+ Ions"
Butalbital (M = 224) Low
42 (0.04)
High 42 (0.16); Secobarbital (M = 238) Low
44 (0.10);
54 (0.06); 54 (0.19);
56 (0.03); 56 (0.26);
57 (0.02) 57 (0.27);
58 (0.18);
71 (0.07);
86 (0.04)
29 (
