Anal. Chem. 1990, 62,1069-1074
spectable at about 1.2 Fg/mL. The smallest volumes examined were 25 l L , about 450 fmol of HSA at the detection limit (signal-to-noise ratio = 3). On the down side, the sensor has a finite lifetime and cross reactivity is increased compared to bulk solution results. Apparently, surface immobilization adversely affects the binding affinity of the immobilized antibody fragment. After repeated regenerations the surface begins to inactivate also. ESCA experiments indicate that protein is still immobilized, but its activity is completely eliminated after about 75 chaotropic cycles. After 50 cycles the response is still 50% of the initial value. In the future, we plan to explore the suitability of this new sensor scheme for other antigens and smaller haptens. Also, in an effort to minimize photodecomposition problems, we have begun to investigate fluorescent labels other than dansyl. We also are exploring more mild regeneration conditions, and other spacers besides GOPS are being examined.
ACKNOWLEDGMENT We thank T. G. Vargo and J. A. Gardella, Jr., for helping us to interpret the early ESCA spectra and for commenting on the reaction protocols. We also thank Gary Sagerman and Steve Palistrant for constructing the special fiber-optic mounts, etc., used in this work. LITERATURE CITED (1) Turner, A. P. F.; Swain, A. Biotech. Lab. 1888, 8 , 10. (2) Tromberg, B. J.; Sepaniak, M. J.; Vo-Dinh, T.; Griffin, G. D. Anal. Chem. 1887, 59, 1226.
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(3) Vo-Dinh, T.; Tromberg, B. J.; Griffin, G. D.; Ambrose, K. R.; Sepaniak, M. J.; Gardenhire, E. M. App. Spectrosc. 1887, 4 7 , 735. (4) Miller, W. G.; Anderson, F. P. Clin. Chem. 1888, 34, 1417. (5) Seitz, W. R. CRC Crit. Rev. Anal. Chem. 1888, 19, 135. (6) Angel, S. M. Spectroscopy 1887, 2 , 38. (7) Arnold, M. A.; Meyerhoff, M. E. CRC Crit. Rev. Anal. Chem. 1888, 20, 149. (8) Wolfbeis, 0. S.Pure Appl. Chem. 1887, 59, 663. (9) Andrade, J. D.; Vanwagen, R. A.; Gregonis. D. E.; Newby, K.; Lin, J. N. IEEE Trans. Electron Devices 1985, 32, 1175. (IO) Loeb, G. E.: McHardy, Kelliher, E. M.; Brummer, S. B. I n Biocompatability in Clinical Practice, Vol. II; Williams, D. F., Ed.; CRC Press: Boca Raton, FL, 1982. (11) Reginer, F. E.; Noel, R. J. Chromafogr. Sci. 1876, 74, 316. (12) Nilsson. K.; Mosbach, K. Methods Enzymol. 1884, 704, 56. (13) de Alwis, U.; Wilson, G. S.Anal. Chem. 1887, 59, 2786. (14) Nebesny, K. W.; Maschoff, B. L.; Armstrong, N. R. Anal. Chem. 1888, 6 7 , 469A. (15) Gardelia. J. A., Jr. Anal. Chem. 1888, 67, 589A.
RECEIVED for review December 15, 1989. Accepted February 26,1990. This work was supported by BRSG SO7 RR 07066 awarded by the biomedical Research Support Grant Program, Division of Resources, National Institutes of Health, the donors of the Petroleum Research Fund, administered by the American Chemical Society, a Non-Tenured Faculty Grant from 3M, Inc., a grant from the Health Care Instruments and Devices Institute at SUNY-Buffalo, the National Institute of Mental Health, and the National Science Foundation. Finally, K.S.L. wishes to acknowledge support from an ACS Analytical Division Summer Fellowship sponsored by the Pittsburgh Conference.
Examination of Ligand-Ligand Interactions by Fast Atom Bombardment Mass Spectrometry Dennis P. Michaud, J. N. Kyranos, T. F. Brennan, and Paul Vouros* Department of Chemistry and Barnett Institute of Chemical Analysis, Northeastern University, Boston, Massachusetts 02115
The fast atom bombardment mass spectra of mixtures of peptide ligands exhibit well-defined peaks corresponding to protonated molecular Ions of ligand complexes. Ouantltatlve selectivity In complex formation between llgands is indicated. A split-probe-tlp experlment has been designed In whlch lndlvldual ilgand solutions are placed one on each half of the probe tlp and exposed simultaneously to the atom beam. I n systems where complexatlon requires lnteractlon at several sites, no protonated complex formation is observed under these condltlons. Complexes formed by simple single-site lank Interactions, on the other hand, are readily formed In thls system. The data suggest that complexatlon Is, at least in part, the result of solution (or posslbly selvedge-Induced) lnteractions as opposed to a vapor-phase phenomenon.
INTRODUCTION In recent years fast atom bombardment mass spectrometry (FAB-MS) has emerged as a technique that permits the analysis of polar and nonvolatile substances by transferring them from a liquid matrix directly into the vapor phase (I). A prominent feature of FAB mass spectra is cluster ion for-
mation. For example, when glycerol is used as a matrix, protonated clusters are routinely observed with hydrogen bonding being the likely force holding these clusters together. Analyte-analyte clusters have also been detected by FAB-MS. In their use of FAB-MS for the sequence determination of peptides, Roepstorff et al. (2),observed dimerization of selected acetylated and underivatized tripeptides. Meijers et al. (31, showed that cluster formation is a common phenomenon of the FAB mass spectra of porphyrins and suggested that such cluster formation may be important in the tumor localization process. Ions reflecting intermolecular associations have also been reported by Williams et al. ( 4 ) in two examples of the FAB spectra of binary peptide mixtures. Some selectivity was apparent, but the authors caution against the quantitative interpretation of the FAB data generated from the condensed phase interaction of two ligands that differ widely in mass. Hydrogen bonding was invoked to explain the occurrence of dimers between imidazole and trimethyl phosphate systems (5) and between pyridinium ions and other electron donors (6). We have been interested in assessing the use of FAB-MS for the study of intermolecular associations in solutions (7). Although many of these associations may be examined by infrared or NMR, mass spectrometry alone provides greater
0003-2700/90/0362-1069$02.50/0@ 1990 American Chemical Society
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ANALYTICAL CHEMISTRY, VOL. 62, NO. 10, MAY 15, 1990
HpN-CNHCHpCHpCH, NH,*
-CHCOO I
NHCOCHCH2
I
NH3'
THYROTROPIN RELEASING HORMONE,
mA
5
KYOTORPHIN,
OH
ARGININE,
CH3
E
n
I
fYo "YN" 0
0
THYMINE,
ADENINE,
&
CYTOSINE, C n,
Figure 1.
GUANINE, G N
Structures of ligands examined.
sensitivity as well as unambiguous identity of cluster ion content. Conventional electron impact MS, which is typically preceded by a thermal vaporization step, is not suitable for a study of phenomena governed by the relatively weak ( < l o kcal/mol) noncovalent bonds. In contrast, FAB-MS is a low-energy process and there is a growing body of evidence that solution-phase chemistry can be studied by this technique (8). Although there is considerable conflict in the literature pertaining to the exact mechanism of proton attachment in FAB, there is significant evidence that the FAJ3 mass spectrum adequately represents the established equilibrium in the liquid state (8). It is not the intent in this work to advocate any one theory of protonation during the FAB process but, rather, to present evidence that may begin to clarify the extent of contribution of solution interactions as opposed to those of the gas phase in the cluster ion formation. This report focuses on the investigation by FAB-MS of ligand-ligand interactions involving several peptides and other biologically significant compounds. Intermolecular interactions are evidenced by the presence of peaks above the molecular ion peak of the monomeric species in the mass spectrum. For example, from the reaction 2A A, the FAB spectrum should contain not only the protonated molecular ion (AH') but also the protonated dimer (A2H+). The ratio of dimer to monomer is an indication of the extent of molecular self-association. In the case of two components A B -+A, AB + B2 there is the possibility for mixed-association as well as selfassociation. The extent of mixed-association, observed in FAB as ABH+, is that quantity which can be used to study the interaction of biologically important molecules. The possible involvement of a vapor phase as opposed to a solution derived attachment has also been examined by using a specially designed probe tip. Evidence for competitive interaction between ligands is also presented.
-
+
+
EXPERIMENTAL SECTION FAB-MS spectra were obtained by using a VG70-SE mass spectrometer. The target surface was bombarded by a beam of energetic xenon atoms produced by an Ion Tech FAB gun operated at 8 keV and 1 mA. Throughout all experiments, the ion source was maintained at room temperature. Peptide solutions (1 mg/mL) in 50% aqueous ammonium acetate (1M), 50% thioglycerol were combined and the mixture (2 pL) transferred to the probe tip of the FAB source. All other solutions (1 mg/mL) were prepared in methanol and used with the matrix indicated. Spectra were acquired at a rate of 5 s/decade, 1 s interscan time, and at nominal mass resolution. No significant variations in the spectral patterns were observed over a period of several minutes.
RESULTS AND DISCUSSION The following analyte systems were examined: (1) kyotorphin (K, Tyr-Arg) with leucine-enkephalin (L, Tyr-GlyGly-Phe-Leu); (2) a ternary mixture of thyrotropine releasing hormone (TRH),K, and L; (3) arginine (R) with L; (4) adenine (A) with thymine (T);(5) guanine (G) with cytosine (C). The structures of the compounds are given in Figure 1. The occurrence of an association between kyotorphin (MW 337) and L (MW 555) is shown in Figure 2. In this case an approximately 5:l molar mixture of kyotorphin to L was examined. The association between the two peptides is evidenced by the peak at m/z 893 (337 f 555 + l)corresponding to the protonated complex. Significantly, the intensity ratio of dimer to monomer for kyotorphin is 0.01, whereas the ratio of complex to L (the limiting reagent) is 0.04. These results suggest that mixed-association is preferred to self-association. The intermolecular associations defined by the K-L complex, as well as those between the other ligands described here, are characterized by interactions at multiple sites. For example, a structure such as the one indicated in Figure 3 may be envisioned for the neutral K-L complex prior to protonation. Here the L conformation is "U"shaped, exhibiting a type I P-bend as observed in several crystallographic deter-
ANALYTICAL CHEMISTRY, VOL. 62, NO. 10, MAY 15, 1990
Flgure 2. FAB spectrum of mixture of K and L.
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Flgure 4. FAB spectrum of ternary mixture containing TRH. K. and L. r n l i 173
m i l 205
N
N
L
K O
O
Flgure 3. Proposed structure of K-L complex minations (9-12). The conformation of L is stabilized by two intermolecular hydrogen bonds between the phenylalanine carboxyl groups. On the basis of related crystallographic data (12-14), it is further reasonable to propose an interaction of L and the complementary guanidinium cation of the K arginine residue, and (b) a tyrosine-tyrosine interaction consisting of hydrogen bonds between the ammonium and the tyrosyl hydroxyl groups of each peptide and a s-bonding interaction between the overlapping tyrosyl aromatic rings. In many respects, these parameters reflect the types of factors that may contribute to the eventual occurrence of an ionized dimer in the FAB analysis. The ternary mixture of equimolar TRH (MW 362), K (MW 3371, and L (MW 555) was examined in order to address the possible use of FAB-MS toward quantifying the selectivity of ligand competition. As shown in Figure 4, all poasible binary mixtures are present in the spectrum, hut considerably higher preference is apparent for the complex of TRH with L (mlz 918 (362 555 1)). I t is likely that this preference is due to an increased number of noncovalent binding sites. If the complex is in equilibrium with the individual components, then an increase in binding sites is expected to drive the equilibrium toward the observed complex formation. The question may be raised as to whether the occurrence of an intermolecular protonated complex of the type ABH+ in the spectra of Figures 2 and 4 might be the result of a vapor-phase ion/molecule reaction or a direct transfer to the vapor phase of a preformed ion in solution. According to the 'hot-gas" model proposed by Kebarle et al. (15),protonation of an analyte, An, during FAB in glycerol, occurs by a gasphase reaction between the sputtered analyte and a protonated glycerol ion, GlyH+
+
+
An
+ GlyH+
-
AnH+
+ Gly
(1)
In principle, it is unlikely that formation of the complex ABH+
Figure 5. Shematic of split-probe tip showing relative disposnion of profiles of K21+ and Na,l+ ion currents. is the outcome of an "all-vapor phase" proceas since that would require a three-body collision. Other alternative gas-phase processes may also be considered. For example, preformed AH+ (or BH+) may react with neutral B (or A) when sputtered off (reactions 2 and 3) or a protonated dimer A2H* (or B2H+) may react with B (or A) (reactions 4 and 5)
+ + + +B
ABH+
(2)
BH+ A
ABH+
(3)
+
AH+
A2H+ B2H+
B
ABH+ A
(4)
A
ABH+ + B
(5)
We addressed the possibility that such gas-phase processes might lead to ions ABH+ by designing a 'split probe tip" experiment. A FAB probe tip was cut across the middle with a laser drill creating a ea. 200-j~mgap between the two halves. Half was coated with KI and the other with Nd,and the direet insertion probe was moved laterally during spectral acquisition. The profiles of the ions corresponding to K21+( m / z 205) and Na21+ ( m / z 173) as a function of the position of beam impact on the probe tip surface are given in Figure 5. Ion peak maxima are observed at positions 1and 3 while position 2, in the shaded area, represents that of maximum xenon beam overlap with the two halves of the probe tip. This setting of the probe ensures concurrent sputtering and vapor-phase mixing of constituents placed separately on each half of the probe tip and was used in all subsequent experiments utilizing this system. Solutions of arginine and L in glycerol were placed one on each half of the split probe tip. We found that, even though the atom beam overlapped both solutions thus providing an opportunity for the sputtered substrates to mix together in
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ANALYTICAL CHEMISTRY, VOL. 62, NO. 10, MAY 15, 1990 L
'27
IB i
556
136
XlO
1
'
10.
da
II
556
;; b) No Cornolex
I I I
r-
I
I
I
/
I
L2 1111
Figure 6. (a) FAB spectrum of mixture of R and L. (b) FAB spectrum of solutions of R and L separated on split probe tip.
the vapor phase, no protonated complex of mass 730 (L + R 1) was observed (Figure 6b). However, when the two solutions were mixed together and analyzed in the same manner, a peak corresponding to the mass of the protonated complex was observed (Figure 6a). The results from the "split probe tip" experiment are not totally surprising since ligand-ligand interactions such as the ones examined above are likely to involve a multitude of sites (see, e.g., Figure 3). The orderly arrangement and orientation of the interacting sites are more easily formed in solution than in the vapor phase. In view of the stringent conformational requirements imposed by these peptides, we proceeded to investigate some more simplified systems in which the interaction is probably restricted to three or fewer sites. It was envisioned that in systems with a higher entropy there would be an enhanced probability for a vapor phase induced process, thereby providing a further measure of its contribution to intermolecular complexation observed in FAB-MS. In the adenine-thymine complexation a two-site interaction is expected. The protonated complex ( m / z 262) is clearly present in the spectrum obtained when the two components are dissolved together (Figure 7a), whereas no indication of its formation is evident in the split probe tip experiment (Figure 7b). A similar effect was observed for the guaninecytosine mixture. The fact that complexation is more likely to occur in solution is evident from the peak a t m / z 263 (Figure 8a). The same peak is absent when the two ligands are separated on the split probe tip (Figure 8b). In the final example we considered the interaction of potassium (K+)ions with an 18-crown-6 ether (CE, MW 264). In many ways this represents one of the most facile complexation processes leading to the formation of a very stable ionic species. While the potassium ion is bound to six different oxygen atoms, this metal chelation is a low-entropy process involving effectively only one level of interaction. In other words, it would appear that the rate-determining step leading
+
Figure 7. (a) FAB spectrum of mixture of adenine (A) and thymine (T). (b) FAB spectrum of solutions of A and T separated on spli probe tip. dh
!i.
'12
C
H
G
08
!I li
j
;I/
4 5 .'
(a. 15 ., 18.
~
li
H
Conplex
XlO
7
Yo corr3ex
Figure 8. (a) FAB spectrum of mixture of guanine (0) and cytosine (C).(b) FAB spectrum of solutions of G and C separated on split probe tip.
to complex formation is the initial potassium-oxygen inter-
action. Once that attraction is established, the reaction proceeds to form the stable complex. As a result, the mass spectrum of a mixture of KI and 18-crown-6ether in distilled
ANALYTICAL CHEMISTRY, VOL. 62, NO. 10, MAY 15, 1990
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[CE + K t l
[ C E + K']
303
18%.
35
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3r
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LCE + R b t l
45. 48
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397
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;; b)
[CE +]'H
[CE
+
K+l 85. 88 JI .
265
6% 15
b)
(CCE+Htl [CE + Na' 1 287
J8 .
JD .
65.
61.
68.
Ka.
55.
[CE+K+]
55. 58.
(I, 48.
5.
P R.
n
I/ yI
148
31
188
188
Figure 9. (a) FAB spectrum of mixture of KI and 18-crown-6 ether (CE). (b) FAB spectrum of solutions of K I and CE separated on split
probe tip. glycerol free of alkali-metal contamination is dominated by the peak at m / z 303 reflecting the formation of the complex (Figure 9a). Significantly, evidence for complexation is also apparent from the split-probe tip experiment. The spectrum, Figure 9b, shows an intense peak at m / z 265 corresponding to the protonated crown ether. An additional peak at mlz 303 confirms the propensity of this reaction to also occur in the vapor phase. Moreover, similar results were obtained when the above experiment was conducted by using any one of the other alkali iodides (i.e., LiI, NaI, RbI, CsI). These results are additional evidence in support of the hypothesis that the complexations described previously are the consequence of a solution-phase phenomenon rather than a gas-phase reaction. It would appear that, when steric constraints are an important consideration, only the most favorable conditions promote molecular associations in the gas phase. I t is well-known that the cavity that is formed by the 18crown-6 ether is especially well suited for the insertion of a potassium ion (16). In order to ascertain whether this specificity is maintained under the FAB conditions, an equimolar mixture of the alkali-metal iodides (MI) was added to an 18-crown-6 glycerol solution and the resulting solution subjected to FAB. The spectrum obtained is shown in Figure loa. Indeed, the most intense peak corresponds to the potassiumether complex ( m l z 303). The relative intensities of the [CE + M]+ ions are very much in line with the values reported by Johnstone et al. (17). When the solid alkali iodides were placed on one side of the split probe tip and the crown ether in glycerol on the opposite side, the spectrum obtained by bombardment of both sides of the tip is shown in Figure lob. Note that the protonated ether complex is the base peak indicating the solution phase separation of the ether from the metals. Moreover, the relative intensities of the different metal-ether complexes are now different. Notably, the sodium complex dominates over that of the more stable potassium
Figure 10. FAB mass spectra of CE in glycerol and equimolar mixture of alkali iodides (a) mixed together and (b) separated on split probe tip.
complex. This shift in relative intensities favoring the sodium complex may be explained in terms of the mechanism of formation of each complex. In solution the metals exist as solvated species which can attach themselves to the crown ether. During the split probe experiment, however, the alkali iodides are deposited on the tip as a solid and sputtered into the gas phase by the impact of the FAB beam. In addition to free ionic metal species present in the gas phase, there are also a number of different salt clusters that can interact with the crown ether and transfer the metal ion. The following are some examples:
MI
+ CE
+
MCE+ + I-
(6)
If any of these reactions contribute to the formation of the metal-ether complex, then the energy of bond formation must be greater than the energy released when the metal-halide bond is broken. The formation of the metal-ether complex is governed by both the ease with which the metal is released from the salt complex and the stability of the resulting metal-ether complex. Although the potassium adduct is more stable, the sodium-iodide bond is easier to break (18). Judging from the higher relative abundance of the sodium-ether complex in the split-probe spectrum (Figure lob), the ratedetermining step for gas-phase complexations of the type presented here may be the separation of the metal from the salt. Finally, the possibility of cation exchange occurring in the vapor phase was investigated by using the split probe experiment. On one side of the tip was placed a solution of metal (Li+,Na+, Rb+, or Cs+)-ether complex in glycerol. The opposite side of the tip was coated with solid KI. Although the potassium-ether complex is the most stable, and there is ample opportunity for gas phase collisions, no exchange of metal ions was observed. The activation energy needed for metal exchange is apparently not available in the gas phase.
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Anal. Chem. 1990, 62, 1074-1079
This further suggests that molecular associations may be solution-phase phenomena.
CONCLUSION The data presented above demonstrate that FAB-MS may be used not only to determine the presence of intermolecular complexes in solutions but also to probe the selectivity of ligand-ligand interactions. In selecting ligands that can best associate via multiple interacting sites, our split probe tip experiments have demonstrated that, at least for such ligand systems, gas-phase clustering is not favored. These results indicate that the normal FAB spectra of multiple-binding-site intermolecular associations, may reflect the solution equilibria. On the other hand, when intermolecular association requires a simpler interaction as in the crown ether-alkali metal complexes, even the wide gap separating the two halves of the split probe tip cannot suppress clustering. In a broad sense, intermolecular associations of the type described here may be influenced by structural parameters and solution conditions. Stabilization of the complex is enhanced through hydrogen bonding, R bonding, and electrostatic interactions. It is envisioned that by combining information from FAB-MS analysis with crystallographic data and molecular modeling techniques (4,5),it may be possible to evaluate the relative contributions of these parameters. As a result it should be possible to design modified ligands with enhanced selectivity and increased affinity in interactions with ligands, receptors, or enzymes. FAB-MS can provide a sensitive and rapid screening for such features.
LITERATURE CITED (1) Barber, M.; Bordoli, R. S.;Sedgwick, R. D.; Tyler, A. N. J . Chem. Soc., Chem. Commun. 1981, 325. (2) Roepstorff, P.; Hojrup, P.; Moller, J. Biomed. Mass Spectrom. 1985, 72,181. (3) Meijers, J. C. M.; Lim, C. K.; Lawson. A. M.; Peters, T. J. J . Chromatogr. 1988, 352,231. (4) Williams, D. H.; Bradley, C.; Bojesen, G.; Santikarn. S.:Taylor, L. C. E. J . Am. Chem. SOC. 1981, 703,5700. (5) Clark, J. H.; (;reen, M.; Madden, R.; Reynolds, C. D.; Dauter, 2.; Miller, J. M.; Jones, T. J . Am. Chem. SOC. 1984, 706, 4056. (6) Brown, S. J.; Miller, J. M. J . Chem. Soc., Perkin Trans. 2 1987, 1129. (7) Michaud, D. P.; Brennan, T. F.; Vouros, P. Presented at 36th Annual Meeting of the American Society for Mass Spectrometry, San Francisco, CA; 1988. (8) Caprioli, R. M. Anal. Chem. 1984, 55, 2387. (9) Johnstone. R. A. W.: Lewis, I. A. S.;Rose, M. E. Tetrahedron 1983, 39. 1597. (10) S&h,G. D.; Griffin, J. F. Science 1978, 799,1214. (11) Blundell, T. L.; Hearn, L.; Tickle, I.J.; Palmer, R. A,; Morgan, B. A,; Smith, G. D.: Griffin, J. F. Science 1979, 205, 220. (12) Ishida, T.; Kenmotsu, M.; Mino, Y.; Inoue, M.; Fujiwara, T.; Tomita, K.-I.; Kimura, T.; Sakaibara, S. Biochem. J . 1984, 278,677. Protein Res. 1983, 27,223. (13) Suresh, C. G.;Vijayan, M. Int. J . Pep* (14) Garofai, A. L.; Williams, D. A.; Brennan, T. F. Cryst. Struct. Commun. 1979, 8, 953. (15) Sunner, J. A.; Morales, A.; Kebarle, P. Anal. Chem. 1987, 59, 1378. (16) Cotton, F. A.; Wilkins, G. Advanced Inorganic Chemistry, 4th ed.: John Wiley and Sons: New York. 1980. (17) Johnstone, R. A. W.; Rose, M. E. J . Chem. SOC.,Chem. Commun. 1983, 1268. (16) CRC Handbook of Chemistry andPhysics, 66th ed.; Weast. R. C., Ed.; CRC Press, Inc.: Boca Raton, FL, 1986.
RECEIVED for review July 27, 1989. Revised manuscript received November 28, 1989. Accepted February 5, 1990.
Effect of Hydrogen Substitution on the Mass Spectrometry and Size-Exclusion Chromatography of Perfluorinated Polyether Fluids as Determined by Time-of-Flight Secondary Ion Mass Spectrometry Steven M. Hues, Jeffrey R. Wyatt, and Richard J. Colton* Chemistry Division, Naval Research Laboratory, Washington, D.C. 20375-5000 Bruce H. Black Ceo-Centers, Inc., 10903 Indian Head Highway, Suite 502, Ft. Washington, Maryland 20744 The secondary ion mass spectrometry (SIMS) of perfluorkrated polyether (PFPE) fkrids reveals an e x t d v e series of hlgh-mass Ions whose distribution depends on the molecular weight, structure, and fragmentation pattern of the oiigomers. Analysis by site-exclusion chromatography (SEC) showed the fluids to consist of elther single or double component systems. Allquots of the components were collected from the SEC column effluent and subsequently analyzed by time-of-flight SIMS. The oligomer distributlons of the PFPE fluids were found to dlffer both in their number average molecular weight and in the degree of hydrogen substitutlon. The observed SEC separation was dependent on the molecular weight and the effect of hydrogen substitution on the molecular volume.
INTRODUCTION Perfluorinated polyether (PFPE) fluids are finding widespread application as high-temperature lubricants ( I , 2 ) because of their thermal stability, good lubricity, viscosity index,
and general chemical inertness. However, the reaction of these fluids with ferric fluoride, aluminum chloride, and other Lewis acids and with iron and titanium alloys a t elevated temperatures has been noted (3, 4). The mechanism of fluid degradation or reactivity may be associated with the presence of a small number of substitutional hydrogen species ( 5 ) ,due to incomplete fluorination. The fluids used in this study are homopolymers of trifluoro(trifluoromethy1)oxirane derived from two different sources: Krytox (E. I. du Pont de Nemours & Co.) and Aflunox (PCR, Inc.). The structure of the polymer is CF,CF,(OCF,CF(CF,)),F, and its IUPAC name is a-perfluoroethyl-w-fluoropoly(oxyperfluoro-2-methylethylene). The specific perfluorinated fluids used in this study are commonly known as Krytox 143 AZ and Aflunox 606. These fluids are low vacuum grade P F P E s of relatively low viscosity and low molecular weight. Krytox 143 AZ and Aflunox 606 have average molecular weights of approximately 1850 (determined by NMR) and 2100 amu, respectively (6, 7). The third fluid used in this study, Krytox TLF-7076, was specially hydrogenated by Du Pont to give the molecular
0003-2700/90/0362-1074$02.50/0 0 1990 American Chemical Society
