1442
Anal. Chem. 1909, 61,1442-1447
Fast Atom Bombardment Induced Reduction of Aromatic Oximes M. Graca 0. Santana-Marques a n d A n t h i o J. V. Ferrer-Correia Department of Chemistry, University of Aveiro, 3800 Aveiro, Portugal Michael L. Gross* Midwest Center for Mass Spectrometry, Department of Chemistry, University of Nebraska, Lincoln, Nebraska 68588
Aromatic oximes and P-hydroxyoximes are reduced to their corresponding imines by interaction of their solutions in giyceroi solvent with a 7-keV argon atom beam. Evidence that the reduction occurs for the protonated oximes comes from fast atom bombardment (FAB) mass spectrometry, tandem mass spectrometry, and time-resolved FAB studies. The requirement that the oxime be in solution was established by comparing the FAB results with those obtained by electron ionization, chemical Ionization, and laser desorption. The timedependent studies of the concentration of the protonated oxime and the reduced product, a protonated imine, were made with and without added anionic surfactant. The abundance of the reduced species depends on the concentration of the protonated oxime at the matrix surface, thus showing that the first step of the reduction mechanism is the protonation of the analyte, followed by reduction and eventual desorption of the reduced species. Surface reduction occurs for ail the compounds Containing an oxime group that were studied and Is a useful characteristic for identtfylng oximes by fast atom bombardment mass spectrometry.
INTRODUCTION Fast atom bombardment mass spectrometry is a useful technique especially for the analysis of polar and nonvolatile compounds. Mechanisms of desorption that occur upon fast atom bombardment are still the object of some controversy, and several interpretations of the ionization phenomena under fast atom bombardment conditions were advanced by different authors (1-6). A better understanding and a better utilization of this technique can be achieved through the investigation of the many facets of desorption ionization from a liquid matrix. Reduction of the analyte under desorption ionization conditions has received considerable attention and occurs for different types of compounds (7-10). Evidence exists that reduction takes place for the same type of compound both in the absence (11) and in the presence of a liquid matrix (12, 13). Suggestions that radicals produced in the liquid matrix play a role in reduction phenomena were advanced by Field (14) and Ligon (15),whereas Cerny and Gross (16) proposed multiple protonation followed by one and two electron reductions. Williams et al. (17) attempted to generalize the mechanism for reduction under fast atom bombardment and secondary ion mass spectrometry by postulating that the electrons produced by the impact of kiloelectronvolt ionizing particles are the effective reducing agents. Reduction involves in part the capture of thermal or near thermal energy electrons by sample molecules. Under fast atom bombardment conditions, reductions involving dehalogenation were observed (18, 19). For some N-hydroxy nitrogen compounds (benzohydroxamic acid, 1-
Table I. Aromatic Oximes of the General Formula CGH&(R)NOH
I I1 I11 IV V
VI VI1
R
oxime
mol wt
H CH3 C,H6 C3H7 C6HI3 CllH23 C,H,
benzaldoxime acetophenone oxime propiophenone oxime butyrophenone oxime heptanophenone oxime dodecanophenone oxime benzoDhenone oxime
121 135
149 163 205
275 197
hydroxybenzotriazole, and N-hydroxysuccinimide), reduction to their deoxygenated counterparts was also reported (19). Deoxygenation was observed for sulforhodamine B, a xanthane type dye (10). In all these studies, the mechanism and generality of the reduction processes were not established. A substance that was extensively used for studying reduction is methylene blue (11-13, 20). A correlation of the extent of the reduction occurring in fast atom bombardment mass spectrometry and secondary ion mass spectrometry with its electrochemical potential in solution was established (12, 13),and it was concluded that the substance must be dissolved in a liquid matrix for reduction to occur (12). On the other hand, the same reduction processes were shown to occur in the absence of any protic solvent, due to self-hydrogenation by the analyte molecules, and the correlation mentioned above was suggested to be fortuitous (11). Thus, the requirements of fast atom bombardment mass spectrometry and the presence of a matrix for inducing reduction remain open questions. We will address these questions for oximes by studying their reduction to the corresponding imines under fast atom bombardment conditions. Oximes are widely used in industry (21,22)and in chemical analysis (23-26). Methods for structure proof and analysis of oximes are therefore needed. The reduction that we observe is common to all the compounds containing an oxime function group and is useful for establishing more clearly the mechanism of reduction induced by fast atom bombardment. Furthermore, reduction is a general feature of the fast atom bombardment (FAB) mass spectrometry of oximes and, as such, can be used in the identification and structure proof of this class of compounds, in either the pure or mixed state. The reader is directed to a recent review for a more detailed discussion of chemical reactions that accompany desorption ionization (27). EXPERIMENTAL SECTION All the oximes (Table I) and some of the hydroxyoximes (I to VI, Table 11) were synthesized from their corresponding aldehydes and ketones by using standard procedures (28). Industrial hydroxyoximes (VI1 to X, Table 11) were purified first by complexation with Cu(I1) (29) and further by high-pressure liquid chromatography (30). Benzophenone imine was purchased from Aldrich (Milwaukee, WI).
0003-2700/89/0361-1442$01,50/0 1989 American Chemical Society
ANALYTICAL CHEMISTRY, VOL. 61, NO. 13, JULY 1, 1989
Table 11. Aromatic &Hydroxyoximes of the General Formula RC6HaOHC(R’)=NOH R
hydroxy oxime
R
I
H
H
I1 I11 IV V
H H CH3 CH3
CH3 C6H6 H CH3
VI
CHB
C6H6
2-hydroxybenzaldoxime (salicylaldoxime) 2-hydroxyacetophenone oxime 2-hydroxybenzophenone oxime 2-hydroxy-5-methylbenzaldoxime
2-hydroxy-5-methylacetophenone oxime 2-hydroxy-5-methylbenzophenone
oxime VI1 CBH17 H VIII C ~ H I ~ ( C H ~H) ~ C
2-hydroxy-5-octylbenzaldoxime
IX
C5H13(CH3)2CCH3
2-hydroxy-5-(1’,l”-dimethyl-
X
C5H,3(CH3)2CC6H6
2-hydroxy-5-(l’,l”-dimethyl-
2-hydroxy-5-(l’,l/’-dimethylheptyl)benzaldoxime hepty1)acetophenone oxime hepty1)benzophenone oxime
Fast atom bombardment mass spectra were obtained with a Kratos MS-50 triple analyzer mass spectrometer (Manchester, U.K.), which was described elsewhere (31). A standard Kratos FAB source equipped with an Ion Tech gun was used. The samples were dissolved in glycerol and bombarded with 7-keV argon atoms. For the time dependence studies, 0.01 M solutions of the oximes in glycerol and a 0.01 M solution of lithium dodecyl sulfate in glycerol were used. Ion abundances were measured by using the same calibration file (mass range from 99 to 282 amu) and doing the same number of successive scans, with the same instrumental parameters, for the selected mass range. Normal spectra were acquired at low mass resolution (ca. 3000). Metastable ion and collisionallyactivated decomposition (CAD) mass analyzed ion kinetic energy spectra (MIKES) were obtained by scanning the second electrostatic analyzer. CAD spectra were obtained by using helium as collision gas (collision cell located between the magnetic sector and the second electrostatic analyzer), with 50% suppression of the intensity of the signal. The data were processed with a standard DS-55 data system by using software written at the Midwest Center for Mass Spectrometry. Accurate mass measurements under FAB conditions were made by peak-matching with a Kelvin-Varley voltage divider, using selected ions from a desorption of a cesium iodide/glycerol sample as references. Full scan, high resolution ( R = lOO00) electron ionization (EI) mass spectra were obtained with a Kratos MS-50 double focusing mass spectrometer, interfaced to a Nova-4X computer and operating with DS-55 software. The source temperature was 250 “C, and the solids probe was heated sufficiently so that an ion beam was produced. Laser desorption spectra were obtained with a Fourier transform mass spectrometer, constructed in the Midwest Center for Mass Spectrometry (32) and controlled with a Nicolet FTMS-1000 computer and data acquisition system. A Quanta-Ray DCR-2 Nd:YAG laser was used at two wavelengths and pulse intensities: 266 nm, 5 mJ and 1064 nm, 140 mJ.
RESULTS AND DISCUSSION Fast Atom Bombardment and Fast Atom Bombardment Mass Analyzed Ion Kinetic Energy Spectra. Fast atom bombardment of all the compounds studied (aromatic oximes and P-hydroxyoximes) produces [(M + H) - O]+ ions in the gas phase. The abundances of [(M + H) - 0]+ions increase with increasing time, to the point where this ion is the most abundant. In Table 111, the fast atom bombardment mass spectra of the oximes with the general formula C6H5C(R)=NOH, listed in Table I, are presented. The data were obtained from spectra recorded at the same elapsed time after sample introduction in all cases. Because the relative abundances of the [M + HIt and [(M + H) - O]+ ions vary with time, the choice of the spectra was made after the time-dependent studies were analyzed. Spectra are reported at the point when the ratios of both the [M + H]+ and [(M + H)
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Table 111. Relative Ion Abundances of Fast Atom Bombardment Mass Spectra of Oximes I to VI1 (% of Base peak)a
I I1 I11 IV V
VI VI1
28
25
34 27 70 18 30
100 100 55 100 100 74
19
35
50 49
49 13
26
19 30
42 15 11 13
54 17 11
“Plateau” values.
- O]+ ion abundances to their sum became constant within experimental error. Besides the [ (M + H) - O]+ ions, the ions formed by a loss of one H20molecule are also significant for oximes. Formation of the molecular radical cation is only found in the case of the benzaldoxime. The ion of m f z = 104, probably having the formula C6H5CNH+,is a common fragment for all the oximes. To confirm that the [(M + H) - O]+ ions are not the product of a gas-phase decomposition of the [M + H]+ ions, but the result of an independent reduction process, metastable ion and collisionally activated MIKE spectra were obtained of the [M + H]+ ions of all the oximes. The three most abundant ions for each of the oximes listed in Table I, are shown in Tables IV and V. The principal decompositions as registered in the metastable ion spectra are the loss of HzO, the formation of ions of m f z = 104, m / z = 91, and m f z = 77, and alkene elimination in the case of oximes with larger alkyl groups (from C3H7 to CllH23). Upon collisional activation, the principal decompositions are the losses of H, ROH, and CRNHOH, the last two giving the ions of mlz = 104 and mf z = 77. Alkane elimination is only important for the heptanophenone oxime and dodecanophenone oxime. Loss of HzO gives the most abundant fragment in the mass spectrum of benzophenone oxime. From these results, the complete absence of a decomposition, either metastable or collisionally activated, giving rise to the loss of an oxygen atom from the protonated quasimolecular action, is ascertained. Similar results were found in the case of the 0-hydroxyoximes, so we can conclude that the [(M + H) - O]+ ions are not formed by gas-phase decomposition of the [M H]+ ions but are probably produced in the matrix and then desorbed. Identification of the Reduction Products. For the phydroxyoximes with the general formula RC6H30HC(R’)= NOH, listed in Table I1 from VI1 to X, the composition of the ions formed by the loss of 16 m u from the [M H]+ ions was established on the basis of accurate mass measurements. These are presented on Table VI. The correct composition is [(M + H) - O]+, and other possible compositions such as [(M + H) - NHz]+ and [(M + H) - CH,]+ are ruled out because they give deviations in the range 67-97 and 106-151 ppm, respectively. In addition, the iminium ion structure of the [(M + H) O]+ ions had to be checked. It is well-known that imines of the general formula RR’C=NH are often very unstable and highly reactive (33). Nevertheless, some aromatic imines such as the benzophenone imine are stable enough to be synthesized and stored. Therefore, we sought to compare the CAD spectrum of the protonated ion of the benzophenone imine and that of the [ (M + H) - O]+ ion of the benzophenone oxime (see Figure 1). On the basis of their near identity, the proposed iminium structure for the reduced form is confirmed. It is interesting to note that an iminium type intermediate has been postulated for the electrochemical reduction of ox-
+
+
ANALYTICAL CHEMISTRY, VOL. 61, NO. 13, JULY 1, 1989
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Table IV. Metastable Ion MIKE Spectra of [M
+ H]+ Given as Decreasing Abundances"
oxime
fragments
I
ion
C6H5+
(mlz)
(77) 34 %
re1 abund ion
I1
C6H5f
biz)
(77) 4 YG
re1 abund ion
111
[(M + H) - ROH]+
(mlz)
(104) 24%
VI
re1 abund ion (mlt) re1 abund ion (mlz) re1 abund ion 31 %
VI1
rel'abund ion
(mlz)
(180) 62%
IV
V
(mlz) [(M + H) - HzO]+
rel'abund a
9%
Ion abundances were normalized to 100.
Table V. CA MIKE Spectra of [M
+ H]+ Given as Decreasing Abundances" fragments
oxime I
ion
[(M + H) - ROH]+
(mlz)
(104) 32%
re1 abund ion
I1
[(M + H) - ROH]+
(mlz)
(104) 25 %
re1 abund ion
111
[(M + H) - ROH]+
(mlz)
(104) 17YG
re1 abund ion
IV
[(M + H) - ROH]+ (104) 22 %
(mlz)
re1 abund ion
V
[(M + H) - ROH]+
(mlz)
(104) 18%
re1 abund ion
VI
[(M + H) - CgHzoI+
(mlz)
(148) 17%
re1 abund ion (rnlz) re1 abund
VI1
ROH]' 14%
[(M+ H) - H,O]+
M+'
(180) 39%
(197) 26 %
Sum of abundances normalized to 100.
Table VI. Accurate Mass Measurements Establishing the Composition of [(M H) - 161' Ions
+
hydroxyoxime VI1 VI11 IX
X
calculated mass for [(M + H) - O]+
found
error, ppm
234.185 789 248.201 439 262.217 089 324.232 739
234.184 79 248.200 66 262.216 14 324.230 74
-4.3 -3.1 -3.6 -6.2
imes in solution (34),where they are reduced, in some cases, to the corresponding amines via the imines in an overall two-step, four-electron process. These studies were mainly performed in an acidic or neutral media. Here we propose a two-electron process that will be discussed later. FAB in Other Matrices and CI. We attempted to evaluate whether reduction occurs in a matrix-less FAB experiment. Unfortunately, the signals for (M + H)+ were so
Figure 1. CAD MIKE spectra of ions (C,H,),C=NH,+ ( m I z = 182): (A) ion obtained in FAB spectrum of benzophenone oxime; (B) ion obtained in FAB spectrum of the benzophenone imine.
ANALYTICAL CHEMISTRY, VOL. 61, NO. 13, JULY 1, 1989
transient, even at the lowest atom dosages we could arrange with a saddle field gun, that no conclusion can be drawn from these experiments. Nevertheless, we are able to establish that the reduction is most facile in protic solvents. The fractions of the ion current of the [(M + H) - O]+ ion with respect to the sum of the currents of [(M H) - O]+and (M + H)+ for tetradecylaldoxime are given as follows for four test matrices: 3-nitrobenzyl alcohol, 7 % ; o-nitrophenyloctyl ether, 18% ; dithioerythritol/dithiothreitol,34% ; acidified glycerol, 52%. The trend is evidence that a protic solvent is required for reduction to occur. Under isobutane chemical ionization conditions, the relative abundance of the [(M + H) - 01 for the tetradecylaldoxime is 1% at best, also confirming the need for a protic matrix for the reduction. High-Resolution Electron Ionization and Laser Desorption Fourier Transform Mass Spectra. Having established that the [ (M + H) - O]+ ions are iminium ions and that their formation is not a result of gas-phase decompositions, we wish to demonstrate that the iminium ions are formed almost exclusively as the result of a fast atom bombardment induced reduction. The absence of imine traces in the oxime samples was established. High-resolution E1 mass spectra were obtained to ensure that even traces (
