Anal. Chem. 1994,66, 1884-1889
Surface Ionization Organic Mass Spectrometry of Imipramine, Desipramine, Clomipramine, and Lidocaine Toshihiro Fujii' and Yoshiyuki Kurihara
National Institute for Environmental Studies, 16-2 Onogawa, Tsukuba, Ibaraki 305, Japan Hiromi Arimoto
Shimadzu Corporation, Nakagyo-ku, Kyoto 604, Japan Yoshihiro Mitsutsuka
Meisei University, Hodakubo, Hino, Tokyo 186, Japan
Surface ionization organic mass spectrometry (SIOMS) has been performedon some clinically important drngs (imipramine, desipramine, clomipramine, lidocaine) by using quadrupolemass spectroscopy in which the thermal ion source has a rhenium oxide emitter. The mass spectra were presented, interpreted in a purely empirical way, by means of evidence from previous investigations, and then compared to conventionalE1 techniques. Sensitivityand selectivity have also been studied,demonstrating that (a) these drug compounds are efficiently surface-ionized, (b) experimental results rationalize the high sensitivity of the surface ionization detector (SID) of gas chmtography (CC) for the examined drugs and, (c) the GC/SIOMS coupling can be used for sensitive and selective detection of the drugs in the serum. An approach to detection of these drugs in serum by GC/SIOMS and GC with SID isdescribed. The characteristics of both methods provide a reliable, sensitive, and selective method, which is needed for low concentration level measurements in complex mixtures. Drugs are continuously causing severe poisonings. Antidepressant substances are often misused and abused. The incidence of these cases is high, and many types of drugs are involved. Drug analysis is often needed for low-level concentrations.' A number of drug analyses have been developed using thinlayer chromatography,2 fluorescence spectr~photometry,~ radioisotope derivatization method^,^ and radioimmun~assay.~ Gas chromatography (GC)6with flame ionization or nitrogenspecific7 detectors has been used, as has high-performance liquid chromatography,8 but these methods are prone to problems of sensitivity and/or lack of specificity, mainly due to interference from coeluting compounds. (1) Florey, K., Ed. Analytical Profiles ofDrugSubstances;Academic Press: San Diego, CA, 1988; Vol. 17. (2) Harper, J. D.; Martel, P. A.; O'Donnell, C. M. J. Anal. Toxicol. 1989, 13, 31. (3) Moody, J. P.; Whyte, S. F.; Naylor, G. J. Clin. Cfiim. Acta 1973, 43, 355. (4) Wallace, J. E.; Hamilton, H. E.; Goggin, L. K.; Blum, K.Anal. Cfiem.1975, 47, 1516. ( 5 ) Cleeland, R.; Christenson, J.; Usategui-Gomez, M.; Heveran, J.; Davis, R.; Grunberg, E. Clin. Cfiem. 1976, 22, 712. (6) de Silva, J. A. F.; Puglisi, C. V. Anal. Cfiem. 1970, 42, 1725. (7) Monca, D.; Ferron, L.; Weber, J. P. Clin. Cfiem. 1989, 35, 601. (8) Overzet, F.; Rurak, A.; van der Voet, H.; Drenth, B. F. H.; Ghijsen, R. T.; de Zeeuw, R. A. J. Cfiromatogr. 1983, 267, 329.
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A number of papersg-" have appeared describing the use of gas chromatography/massspectrometry(GC/MS) for drug analysis. The use of combined GC/MS made it possible to obtain the required specificity by monitoring the identity of the peaks as they eluted from thegas chromatograph. Selected ion monitoring brought the analytical sensitivity of the drug in blood serum into the 5-10 ng/mL range, using the internal standard for quantitation.12 Among many MS schemes, electron ionization (EI) has been widely used as an ionization technique. With a quadrupole MS for the detection of ions, E1 has been demonstrated to be a versatile and sensitive technique with a powerful means of molecular identification and structural analysis. However, despite the number of mass spectrometric procedures available, determination of serum concentrations of drugs encountered in patients receiving small dosages remains difficult in some ~ a s e s . 1 3 , ~ ~ Surface ionization (SI)has proven to be a useful technique for mass spe~trometryl~ and gas chromatography.'6 With SI,some organic molecules can be ionized with high efficiency. Since the SI mass spectral data are lacking for most organics, the so-called surface ionization organic mass spectrometry (SIOMS) has been performed on more than 500 compounds over several years in our laboratory.15J7 In general, soft ionization can be obtained, where only the molecular ion or the molecular ion with very few fragments is generated. In addition, high selectivity can be achieved by SI on the basis of thermal characteristics and ionization energy of chemical species. An area where S I can be successfullyapplied to complement conventional techniques represents the characterization of moderately polar and relatively small biomolecules.I8 Hence, (9) Soo,V. A.; Bergert, R. J.; Deutsch, D. G. Clin. Cfiem. 1986, 32, 325. (10) Neill, G. P.; Davies, N. W.; McLean, S. J . Cfiromafogr.1991, 565, 207. (11) Lillsunde, P.; Korte, T. J. Anal. Toxicol. 1991, 15, 71. (12) Belvedere, G.; Burti, L.; Friegerio, A,; Pantarotto, C. J. Cfiromatogr. 1975, 111, 313. (13) Brooks, K. E.;Smith, N. B. Clin.Cfiem. 1991, 37, 1975. (14) Alkalay, D.; Volk, J.; Carlsen, S . Biomed. Mass Spectrom. 1979, 6, 200. ( I 5) Fujii, T.; Arimoto, H. Am. Lab. 1987, (Aug), 54, and references cited therein. (16) Fujii, T.; Arimoto, H. A d . Chem. 1985, 57, 2625. (17) Fujii, T.; Kakizaki, K.;Ishii, H. Cfiem.Phys. 1992,163,413, and references cited therein. (18) Fujii, T.; Inagaki, Y.; Mitsutsuka, Y. Int. J . Mass Spectrom. Ion Processes 1993, 124, 45.
0003-2700/94/036&1004$04.50/0
0 1994 American Chemical Society
ion gauge
* r - l
EXPERIMENTAL SECTION Instrumentation. The experimental setup for GC/SIOMS, shown in Figure 1, has been reported in detail elsewhere.20 Briefly, a dual EI/SI Finnigan Model 3300 mass spectrometer coupled to a Model 9500 gas chromatograph was used. A SI source assembly is home-made so that the rhenium oxide emitter can be fitted into the center of the E1 ion source chamber when the assembly is inserted. Analysis was carried out in the SI mode with 02 added continuously through the
gas flow line to allow preparation of rhenium oxide and an increase in work function at the emitter surface. The mass resolution of the quadrupole mass spectrometer (QMS), R = M / A M ( A M , width at half height), is adjusted to the unit resolution ( M / A M = 2M). As a result, the sensitivity, to some extent, is unavoidably smaller at the higher mass. The GC(S1D) system21 consisted of a gas chromatograph (Shimadzu, GC-15APF) fitted with a van der Berg type solventless injector and a surface ionization detector with an electrically heated Pt emitter (Shimadzu, SID- 14/ 15). Analytical Conditions. For all gas chromatographic separations (both GC(S1D) and GC/SIOMS), a 25 m X 0.2 mm i.d. X 0.25 pm CBP-5 (chemically bonded 5% phenyl poly(dimethylsi1oxane)) fused silica capillary column was used. The column temperature was maintained at 260 and 250 OC for antidepressants and lidocaine, respectively. The flow rates of the helium carrier gas were 0.9 (antidepressants) and 1.4 mL/min (lidocaine). The other instrumental parameters used were as follows: for GC(SID), air flow rate 150 mL/min, detector temperature 300 "C; for GC/SIOMS, separator-room temperature (260 "C); for electron ionization MS (EIMS), electron energy 55 eV, electron emission current 0.7 mA; for SIOMS, surface temperature of the rhenium oxide emitter 850 "C. Samples. Thedrug samples investigatedin this study, along with the proposed thermal discussion patterns, are shown in Figure 2. All drug chemicals were purchased from Sigma Chemical Co., Tokyo, and used without further purification. To obtain reference data and to enable standardization of analysis, a series of drug solutions (concentration 1-1000 pg/ mL) was prepared in dichloromethane for GC(S1D) and GC/ SIOMS. The antidepressant drugs, as a free base, were obtained as follows. Drugs which were obtained as salts were first dissolved in water (1-2 mg in 1 mL), and then pH of the solution was adjusted with 1 mL of 1 N potassium hydroxide, pH 12. The free drug was extracted with 1 mL of a mixture of dichloromethane by vortex-mixing for 5 min, and after separation by centrifugation, the organic phase was transferred to a sample vial and concentrated under nitrogen. Serum Extraction Procedures. For the determination of IPM, DPM, and CPM in human serum, a simple and rapid extraction procedure was employed, which could be completed in less than 30 min. The procedures are as follows: (1) To 0.5 mL of serum, add 0.3 mL of 1 N KOH. (2) To each add 0.5 mL of dichloromethane. (3) Shake 1 min. Centrifuge at 12000 rpm for 2 min. (4) Aspirate off upper aqueous layer. (5) Filter dichloromethane layers into a single 5-mL conical tube. (6) Inject an extracted sample solution into a GC port with a syringe. Analytical Procedures. E1 and SI mass spectra were generated from the drug samples as a free base and in the hydrochloride form. They were analyzed individually. About 100 pg of solid sample was placed in a glass sample holder which was located in a GC oven about 20 cm away from the ion chamber of the E1 source. The desired amount of the sample was controlled by varying the GC oven temperature
(19) Fujii, T.; Jimba, H.; Arimoto, H. Anal. Chem. 1990, 62, 107. (20) Fujii, T.; Suzuki, H.; Obuchi, H. J. Phys. Chem. 1985, 89, 4687.
(21) Fujii, T.; Arimoto, H. Detectors for Capillary Chromatography; Hill, H. H., McMinn, D. G., Eds.; Wiley Interscience: New York, 1993; pp 169-191.
Figure 1. Schematics of the experimental apparatus for surface ionizationorganic massspectrometry. This combination-type ion source allows an easily interchangeableoperation both in SI mode and in E1 mode. When the SI assembly is withdrawn, the E1 source is exactly the same as delivered and, hence, provides the standard E1 sensitivity. The electron beam in the E1 mode is along the plane of this figure.
we are now in the process of evaluating the surface ionization techniques, incorporated with GC and MS, for the analysis of drugs. As examples, three tricyclic antidepressants, imipramine (IPM), desipramine (DPM), and clomipramine (CPM), and the antiarrhythmic local anaesthetic lidocaine are selected here because the relevant information was virtually obtained by preliminary surface ionization detector (SID) studies. Knowledge about the exact identification of charged species being formed from these drugs in the SID is limited, partly due to the lack of mass spectrometric study. Atmosphericpressure ionization mass spectrometric studies on the response mechanism of SID for GC were reported previou~ly.'~The results confirm that (1) positive surface ionization is the mechanism for the production of charge carriers in the SID, (2) the ion species produced are the same as those obtained with SIOMS in a vacuum condition, and (3) parallels exist between the sensitivity of GC(S1D) and SIOMS. Therefore, the SIOMS approach has allowed consistent rationalization of the SID results. This paper describes a highly selective and sensitive method for the analysis of drugs at serum level (as low as 1 ng/mL), utilizing GC with SID and GC/SIOMS. The analytical power of the SI technique is demonstrated by studying its sensitivity and selectivity. The basis of the present assay is rationalized through the SI mass spectrum obtained by SIOMS.
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imipramine
A - NH CI CH3
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clomipramine Flgure 2. Drug examples and proposed thermal dissociatlon patterns.
( Tc). Full mass spectra were acquired by scanning over the mass range m l z 500-40 approximately once per 10 s. The recovery was tested with serum samples prepared by adding the drugs to serum and giving a final concentration ranging from 1 to 4 pg/mL.
RESULTS AND DISCUSSION
(1) Mass SpectrometricConsiderations. We measured the mass spectra of all the drug compounds of interest for two ionization techniques, SI and EI. The sample was introduced directly from the glass holder in the GC oven. The mass spectra are presented in Table 1. The SI spectra were interpreted in a purely empirical way, characterized, and compared to conventional E1 spectra, in order to illustrate the potential and limitations of the SI technique in the analyses of drug molecules. These full mass spectra also make it possible to choose suitable ions for selected ion monitoring and to ensure maximum sensitivity for quantitative purposes. IPM, DPM, and CPM. Although the Tc (GC oven temperature) for these drug salts was ca. 35 OC higher than the Tc for the free base drugs, analysis of a given compound as a free base or in the hydrochloride form does not make a major difference in the resulting mass spectra for either SI or EI. The similarity is to be expected since the HCl salt should thermally decompose to the base and HCl. As an example, mass spectral data are given in Table 1 only for the IPM as a free base and in the salt form. 1886
Analytical Chemistry, Vol. 66,No. 11, June 1, 1994
lidocaine
All the E1 spectra of antidepressants exhibit a parent M+ ion of low intensity and many intense fragment ions. Some similarity exists, especially, between the IPM and DPM spectrum. CPM gives rise to an E1 spectrum showing the intense peaks only in the low-mass range. These spectra are comparable to those reported in literature^'^*^^ with a difference in intensity, but somehow different from those of ref 23. The SI spectrum of all the tricyclic drugs illustrates the features one expects from the SIOMS of biomolecules.'8 It is characterized by the extensive thermal dissociation on the hot emitter surface, which is followed by surface ionization of thermally dissociated products. The approach to structure interpretation of the signals in SIOMS is based on this mechanism. Dissociative surface reaction on a hot emitter results in loss of terminal amine groups so that major peaks were observed at m l z 44, 58, 70, 72, and 84. These can be reasonably assigned to (CH3)HNCH2+, (CH3)2NCH2+, (CH3)HC=NCH2+, and (CH&H5NCH2+, respectively. These abundant peaks of the alkylamine group fit with the evidence from other SI data of amine c o m p o ~ n d s . ' ~No J~ molecular ions are produced in these molecules and the high mass range peaks are not abundant. ~
~~~
(22) Claeys, M.; Muscettola, G.; Markey, S.P. Biomed. Mass. Spectrom. 1976, 3, 110. (23) Heller, S. R.; Milne, G. W. A. EPAINIH Mass Spectral Data Base; US. Dept. of CommerceINBS. Government Printing Office: Washington, DC, 1978.
accessible, and hence, the SI results are very useful for diagnostic analysis. mass spectra m/zd (% re1 intp (M%f&)C E1 vs SI. An important point common to all spectra S I 58 (loo),42 (8.6), 70 (8.2),84 (7.81, imipramine observed throughout this work is that the SI mass spectra 72 (3.8),193 (3.8),44 (3.0), 86 (2.2), (280,18.3) exhibit many fragment ions, which are basically different 194,208,206,204,143,236 (unfamiliar) from those of EI. It is also clear that E1 yields EI: 58 (loo), 85 (54), 35 (12),130 (8.4),193 (6.0), 194 (4.2),195 (3.9), 234 (2.2), 235 small M+ ions for all the examined drugs while SI does not (2.2), 178 (2.1), 208 (2.0), 220,179, 280 yield any ions in the molecular region. The preference of SI: 58 (loo),84 (18.8), 70 (7.4), 193 (7.0), imipramine 42 (4.4), 72 (4.0), 86 (2.6), 44 (2.1), 208, (HCl) (316, 20.7) different functional groups in a molecule is clearly indicated 143,206 in SI whereas the E1 mass spectrometry gives virtually all EI: 58 (loo),85 (56), 35 (23.0), 130 (16.3), 195 (9.0), 193 (7.4), 194 (6.1), 208 (2.6), diagnostic i n f ~ r m a t i o n .This ~ ~ result was not unexpected since 220 (2.1), 234, 235 previous investigations on other biomolecules have revealed S I 44 (loo),58 (13.0), 84 (8.0),70 (7.8), desipramaine 193(7.4), 72 (4.2), 30 (4.1), 42 (3.0),179, (266,7.2) this to be the case. 206,208,194 Next we considered the ionization efficiency of the SI EI: 44 Clod), 42 (75.4), 71 (51.3), 130 (22.7), 193(17.5), 194 (12.4), 195 (11.5), 91 (8.9), method. Since the present ion source is a combination type, 165 (7.4), 179 (7.5), 192 (7.0), 208 (6.1), it is very easy to compare the ionization efficiency in the SI 234 (6.0), 235 (6.0),178 (til),180 ( 4 3 , 220 (4.3). 206 (3.5). 204 (2.71. 228. 266 and E1 modes. To provide an estimate of the efficiency of this SI 58 iioOj, 84 (6.4),'70 (3:6),'42 (2:8), 72, clomipramine method, the ion yield of base peak ions from SI was compared (314,26.6) 44,86,204,230,242 with that from E1 in terms of SI/EI. The SI/EI is the ratio EI: 58 (loo), 44 (17.2), 85 (15.2), 42 ( 1 3 8 , 72 (2.1), 268, 269, 227,226, 228, of the base peak intensity obtained in the SI mode to that in 242,314 the E1 mode. SI production proved to be much more abundant a Refer to Fi ure 4 for lidocaine. b Trivial name used in text. M, than that of E1 in all cases compared (Table 1). The garent nominafmass; SI/EI, ratio of the base peak intensity in the clomipramine result is striking. This compound yields a peak I mass s ectra to that in the E1 spectra. d The italic m/z appears in both tRe SI and E1 mass spectra. "The value in parantheses which is 26.6 times higher in the SI mode. This result indicatesthe percent relative intensity which represents monoisotopic demonstrates that GC/SIOMS and GC(S1D) can be useful intensity. Only peaks above 2 % relative to the base peak are listed, except for some small peaks in the high-mass range. Relative for a trace analysis of certain kinds of drug compounds. intensities are not given to these small peaks. However, it should be noted that our comparisons were limited to the SI and E1 sources we used and may have only a qualitative meaning. A few signals of higher mass range peaks can be reasonably (2) Determination of IPM, DPM, and CPM in Serum by assigned. For instance, the signal peaks at m / z 194,208, and GC(S1D). IPM, DPM, and CPM are widely used tricyclic 236 in the IPM spectrum are due to the products arising from antidepressants. Recent studies indicate a relationship a simple cleavage of the side chain of IPM in the thermal between blood levels of these drugs and therapeutic response.26 dissociation process (Figure 2). In other words, although the Blood levels are very low, even in fatal poisoning. Considerable amine bears the positive charge most intensively, the spectra interest has prompted the development of several analytical show that the skeletal tricyclic ring does show some affinity meth0ds.2~ for the positive charge. For some peaks, however, establishThe present SIOMS studies revealed that SI give better ment of a link with the structure becomes less accessible, output ionic currents for those drugs. Hence, the SID appears which is partly due to the tendency to undergo a complex to have greater potential use in the routine analysis of drugs thermal dissociation process with skeletal rearrangement. in clinical laboratories, which requires sensitive, selective, Lidocaine. The E1 method gives many fragment ions with rapid, and positive identification and determination. From only two small peaks at m / z 120 and 234 in the mass range the use of the SID, two advantages may be expected: First, above 120. The fragment at m / z 120 reaches only 1.5% of because of the high specificity, the possibility of interference the base peak at m / z 86 and the M+ are barely detectable. is greatly reduced and cleanup procedures are usually This result is in good agreement with Karlagnis's spectrum24 unnecessary, and second, owing to the detector's increased among many reported spectra. sensitivity, a better detection capability is achieved. The S I mass spectrum of lidocaine illustrates that, similar Figure 3 shows a typical SID chromatogram obtained from to the above-studied pharmaceuticals, there are no molecular the analysis of a dichloromethane extract of a serum sample ions but there are ionic species in the low-mass region. The to which three antidepressants in different amounts had been expected ionization of the thermally dissociated species is added. The major points of interest concern the relative clearly present; again, quite a few and abundant ionic species sensitivity, the detection limit, and the presence of interfering such as m / z 86 [(CzH&NCH2]+, m / z 58 [(C2H5)HNCH2]+, peaks. As can be seen from the Figure 3, the relative sensitivity m / z 42 (CH2NCH2)+, m / z 56 [(CzH,)HNCH2]+, m / z 72 between three drugs is a little different. No interfering peaks [ ( C ~ H S ) C H ~ N C H ~and ] + , m / z 84 [ ( C ~ H S ) C ~ H ~ N C H ~ ] + was observed in GC profiles of serum extracts, demonstrating appear. Also, signals at (M - 114)+, (M - 129)+, and (M the specificity of the SID. After confirming the linearity of 86)+ correspond to structure-specific losses of side chain from response, the overall detection limit was examined (Table 2). lidocaine (Figure 2). In this case, interpretation becomes quite straightforward. In this respect, the SIOMS results ( 2 5 ) Frigerio, A.; Belvedere, G.; De Nadai, F.; Fanelli, R.; Pantarotto, C.; Riva. certainly contain structural information which is fairly E.; Morselli, P. L. J . Chromatogr. 1972, 74, 201. Table 1. Drug Compounds Studkd and Maso Spectral Data.
(24) Karlaganis, G.;Bircher, L. Biomed.Enuiron. MassSpecfrom. 1987,14, 513.
(26) Gram, L. F. Clin. Pharmacokinet. 1977.2.237, and references cited therein. (27) Borga, 0.; Garle, M. J . Chromatogr. 1971.68, 77.
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1
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(A)
(B) 3
I
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Flgure 3. Surface ionization detector gas chromatogram of a serum sample supplemented wlth three drugs: (A) human serum blank and (B)three antidepressants added Inhumanserum. Peaks: ( 1) imipramine (4 ng); (2) desipramine (17.6 ng); (3) clomipramine (4 ng). Table 2. Analytlcai Results for IPY, DPM, CPM, and Lldocalne, Determlned in Serum Samples wAh the Splked Drug Levels Ranglng from 1 to 4 pg/mL
drug IPM DPM CPM lidocaine
detn limit, ng/mL this worka literature 1.5 6.8 2.1 1.0
0.2* O.lb
5OC
accuracy and precision found added, mean, CV,d pg/mL % % 1.0 4.0 1.0 3.4
94.0 100.6 62.2 101.0
1.72 2.72 2.06 2.5
The detection limit for the overall method (a 2-pL in'ection of the extract from the spiked serum sample of 500 pL), which is given in drug concentration (ng) in serum (mL) at a S/Nratio of 3, provided that no other substance in the serum interferes with each peak. By usin a 4-mL serum sample and monitorin the specific ions of GC/ EI&. The in'ection size is &antitation of lidocaine up to this lever has been achieved with coefficients of variation less than 10% by GC/EIMS combined with mass fragmentogra~hy.~' d CV, coefficient of variation.
The routine determination of IPM in serum at concentrations as low as 1.5 ng/mL (at signal-to-noise ratio of 3) can be reliably made. A comparison of SID with another ionization detector of NTD, used in GC in terms of the overall detection limit of IPM in serum, reveals that the SID provides 3-times lower detection limit than the well-established NTD.28 The accuracy and precision of the assay procedure was determined by measuring five different plasma samples, each spiked with IPM, DPM, and CPM. The coefficients of variation for these assays were 1.72, 2.72, and 2.06%, respectively. The data for lidocaine are cited from the previous which had indicated that lidocaine is successfully applied by the same method as the present one. The good reproducibility of the method is due to the reliability of the SID and to the minimal handling of samples. The average recovery from five analyses ranges from 62.2 to 101.0%. Shinkuma and et aL30have recently used the above method to analyze IPM and DPM in the serum of patients receiving (28) Bailey, N.; Jatlow, P. I. Clin. Chem. 1976, 22, 1697. (29)Arimoto, H.; Shiomi, K.; Fujii, T. J . High Resolut. Chromarogr. 1991, 14, 672. (30) In private communication.
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two doses of these drugs in 1 day. They have encountered no difficulties with the method in these analyses and confirmed the validity of this method for routine analysis. Full details of the study will be described elsewhere. (3) Determination of Lidocaine in Serum by GC/SIOMS. Lidocaine is used extensively to treat ventricular cardiac arrhythmias, especially in patients who have had cardiac surgery or sustained an acute myocardial i n f a r ~ t i o n .Among ~~ many methods for its determination, the use of capillary columns with selected ion monitoring by E1 mass spectrometry may be the best32 and has been the basis of many clinical studies. Combining with a GC, SIOMS also seems to be a promising method for the detection of drugs. Work in this direction was planned for lidocaine. This drug was selected because, in a rough approximation, it can be expected that the amine radical has the potential to be surface-ionized efficiently and previous SID s t u d i e ~ l have ~ . ~ ~yielded promising results. Figure 4 shows the mass spectrum (scan mass range 30250 amu) of lidocaine obtained by SI and by E1 along with a total ion chromatogram (TIM). The sample was a 10-pL extract solution of drug-free serum samples supplemented with lidocaine in concentrations of 40 and 400 pg/mL, respectively. Interestingly, the TIM in the SI mode shows no interference from the compounds associated with the blood, in which a large variety of normal constituents can usually give interfering peaks. The lidocaine peak of both TIMs yields an easily identified mass spectrum, which is, as expected, almost identical to that obtained from the direct introduction of the pure lidocaine sample. Comparison with the E1 mode reveals that the GC/SIOMS does not give rise to peak-broadening, tailing, and baseline drift in the TIM profiles, owing to its fast-response characteristics. Thus, the SI is compatible with capillary column techniques. Another important aspect of using the SI technique for mass spectrometry is its high sensitivity, which is demonstrated by a comparison of TIM obtained with the SI and E1 of a spiked serum sample at different concentrations. If a spiked sample of 500 p L of serum was treated and a 1 0 - ~ L injection of the serum extract was made, a lidocaine concentration in the serum of 42 ng/mL was calculated, as the detection limit at a signal-to-noise ratio of 3, from TIM profiles obtained with the SI. Certainly, the detection procedure (to choose the ions such as the base peak m / z 86, in light of the SI mass spectrum) by selected ion monitoring allows the determination of much lower concentrations in serum, which is at or below those typically used in the clinically important sample. It was not done in this study because of instrumental limitation. The preliminary study on the accuracy and precision of this GC/SIOMS method showed that the results are comparable to those of GC(S1D) obtained for tricyclic drugs. CONCLUSION This is the first study on SIOMS for the characterization of some drugs as an alternative, at least complementary, to conventional E1 mass spectrometry. It has been shown that (31) Hignite, C.E.; Tschanz, C.; Steiner, J.; Huffman, D. H.; Azarnoff, D. L. J . Chromatogr. 1978, 161, 243. (32) Heusler, H. J. Chromarogr. 1985, 340, 273.
100
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Figure 4. Comparlson of total ion chromatograms and mass spectra of lidocaine in a spiked serum sample obtained by (A, bottom) using the SI mode and (B, top) using the conventlonat E1 mode. On the assumption that the extraction efficiency Is loo%, the lidocaine peak of the TIM profiles In the SI mode corresponds to 0.8 pg, while the peak In the E1 mode corresponds to 8 pg. Mass spectral Intensity Is plotted as the normalized percentage scale.
a better idea of the possibilities and limitations of the SI technique, which allows production of a few abundant specific ions, could be obtained, and in this respect, the methods of GC(S1D) and GC/SIOMS associated with the S I technique can result in new opportunities in the field of pharmacology. The principal advantages of these methods are as follows: (a) GC(S1D) is a valuable and important tool in drug analysis. The simplicity, specificity, and sensitivity of this method, provided by detecting abundant ions formed upon SI on a refractory metal surface, make it possible for sensitive and selective analysis of drug serum levels in patients undergoing clinical treatment; (b) GC/SIOMS allows identification of drugs with certainty as a complement to conventional E1 and offers a reliable basis to the GC(S1D) method; (c) GC/SIOMS may detect a large number of drug metabolites under the conditions that an extensive SI mass spectral library is
established; (d) to make full use of the advantages of GC(SID), an S I mass spectral library is also essential.
ACKNOWLEDGMENT This work was supported in part by the ministry of Education, Science, and Culture of Japan, Grant-in-Aid for General Scientific Research (No. 03804045). We thank Dr. Shinkuma and his associates at Hyougo College of Medicine for helpful discussions. The authors are grateful to Tom McMahon at the Alabama Language Academy for the manuscript preparation. Recehred for revlew October 6, 1993. Accepted February 18,
1994.' *Abstract published in Advance ACS Abstracts, April 1 , 1994.
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