Separation and identification of higher molecular weight fullerenes by

faster than C76. Monomeric ODS cannot resolve two of the C7g isomers, although a lower column temperature of 10 °C gives its partial separation. UV-v...
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Anal. Chem. 1993, 65, 2650-2654

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Separation and Identification of Higher Molecular Weight Fullerenes by High-Performance Liquid Chromatography with Monomeric and Polymeric Octadecylsilica Bonded Phases Kiyokatsu Jinno,' Takashi Uemura, Hatsuichi Ohta, Hideo Nagashima, and Kenji Itoh School of Materials Science, Toyohashi University of Technology, Toyohashi 441, Japan

The separation of fullerenes with monomeric and polymeric octadecylsilica (ODS) bonded phases has been studied and the existence of three C78 isomers confirmed. It has been found that the elution order of higher fullerenes is controlled by their molecular size and shape and that polymeric ODS resolves three C78isomers, one being eluted faster than C76. Monomeric ODS cannot resolve two of the C78 isomers, although a lower column temperature of 10 OC gives its partial separation. UV-visible spectral assignments were confirmed by FAB-MS measurements of the fractionated samples. INTRODUCTION Separation of fullerenes in the soot extract is an important analytical problem, considering the increasing attention paid to these new compounds of potential importance in materials science, superconductivity, and other related technologies. Liquid chromatographic approaches have been reported to separate typical fullerenes such as Cm and C70 and other higher molecular weight f~llerenes.l-~ Stationary phases which have had some success in separating fullerenes, such as Pirkletype,6 multilegged phenyl bonded phase,7 and multi-methoxyphenylpropyl phases8 indicate the need to study in more detail the retention mechanism of fullerene LC separations as based on molecular recognition. However, those approaches still need development before they could be useful for industrial or large laboratory-scale separations. Therefore, the main approach a t present for separation of various sizes of fullerenes is to use octadecylsilica (ODS) bonded phases with toluene-based mobile-phase systems, which increase the solubility of fullerenes in the mobile phase and hence the amount injectable into the separation system. Although several investigators have reported the use of ODS stationary phases for this analysis,9-l8 basic discussions on whether (1)Ajie, H.; Alvarez, M. M.; Anz, S. J.; Beck, R. D.; Diederich, F.; Fostiropoulos, K.; Huffman, D. R.; Kraetachmer, W.; Rubin, Y.; Schriver, K. E.; Sensharma,D.; Whetten,R. L. J.Phys. Chem. 1990,94,8630-8633. (2) Taylor, R.; Hare, J. P.; Abdul-Sada, A. K.; Kroto, H. W. J . Chem. SOC., Chem. Commun. 1990, 1423-1425. (3) Hawkins, J. M.; Lewis, T. A.; Loren, S. D.; Meyer, A.; Heath, J. R.; Shibato, Y.; Smalley, R. J. J. Org. Chem. 1990, 55, 6250-6252. (4) Kikuchi, K.; Nakahara, N.; Honda, M.; Suzuki, S.; Saito, K.; Shiromaru, H.;Yamauchi,K.; Ikemoto,I.; Kuromochi,T.; Hino, S.;Achiba, Y. Chem. L e t t . 1991, 1607-1610. (5) Bethune, D. S.; Meijer, G.; Tang, W. C.; Rosen, T. H.; Golden, W. G.; Seki, H.; Brown, C. A.; de Vries, M. S. Chem. Phys. L e t t . 1991, 179, 181-186. (6) Welch, C. J.; Pirkle, W. H. J. Chromatogr. 1992, 609, 89-102. (7) Jinno, K.; Yamamoto, K.; Ueda, T.; Nagashima, H.; Itoh, K. J . Chromatogr. 1992, 594, 105-109. (8) Jinno, K.; Saito, Y.; Chen, Y.-L.; Luehr, G.; Archer, J.; Fetzer, J. C.; Biggs, W. R. J . Microcolumn Sep. 1993, 5, 135-139. (9) Fetzer, J. C.; Biggs, W. R. Polycyclic Aromat. Compd. 1992, 2, 245-251. 0003-2700/93/0365-2650$04.00/0

polymeric-type or monomeric-type phases are a better choice have not appeared in the literature. In this paper, we investigate which is the better stationary phase for fullerene separation, monomeric or polymeric, focusing on isomer resolution. It has been found that monomeric ODS phase is the more powerful stationary phase for the isomer separations, based on the difference in their molecular weights, but polymeric ODS is better for the separation based on molecular shape difference.

EXPERIMENTAL SECTION Instrumentation. The mass spectrometer (MS) used in this work was a JEOL JMS-SXlO2 A (Akishima, Japan). The ionizationwas performed by fast atom bombardment (FAB)using Xe (5 kV) as the primary beam and m-nitrobenzyl alcohol (mNBA) plus 1% toluene as the matrix. The samples were introduced into the MS system using a flow injection method. Basic studies on LC separations were performed using a 880 PU LC pump (Jasco, Hachioji, Japan) combined with a UV detector (Shodex M315, Showa Denko, Tokyo, Japan) set at 320 nm or a Jasco MD-920 UV-visible photodiode array detector. Mobile phases were toluene-methanol and toluene-acetonitrile mixtures. A 4.6 mm i.d. X 250 mm Develosil ODS-5 column (monomeric ODs, 5 pm, Nomura Chemicals, Seto, Japan) and the same size Wakosil I1 5C18AR column (polymeric ODs, 5 pm, Wako Chemicals, Tokyo, Japan) were used in the evaluation. The flow rate of the mobile phases was always 1 mL/min. The same Develosil ODs-5 stationary phase packed into a 20 mm i.d. X 250 mm column was used for the semipreparative-scale separations together with a Jasco 880 PU pump connected to a Jasco 880 UV detector. The mobile phase for the sample fraction collection process was a toluene-methanol mixture which was optimized using an analytical-scale column with a flow rate of 10 mL/min. Column temperature was controlled by a LABThermo Model LH-1000E (Toyo Seisakusho, Tokyo, Japan) and Tosoh RE-8000 oven (Tokyo, Japan). Sample Preparation. Carbon soot was produced by an arc discharge in an inert gas environment. The soot was first extracted with toluene (fraction A in Figure 1,where the sample preparation process is schematically summarized),and then the residue was extracted with trichlorobenzene in order to obtain (10) Diack, M.; Hettich, R. L.; Compton, R. N.; Guiochon, G. Anal. Chem. 1992, 64, 2143-2148. (11)Klute, R. C.; Dorn, H. C.; McNair, H. M. J . Chromatogr. Sci. 1992, 30, 438-442. (12) Jinno, K.; Uemura, T.; Nagashima, H.; Itoh, K. Chromatographia 1993, 35, 38-44. (13) Jinno, K.; Ohta, H.; Saito, Y.; Uemura, T.; Nagashima, H.; Itoh, K.; Chen, Y. L.; Luehr, G.; Archer, J.; Fetzer, J. C.; Rings, _ _ W. R. J . Chromatogr., in press. (14) Jinno, K.: Uemura, T.; Nagashima. H.; Itoh, K. J . Hinh Resolut. Chromatogr. 1992, 15, 627428. (15) Hirsch, A,; Soi, A.; Karfunkel, H. R. Angew. Chem. 1992, 104, 808-810.

(16) Elemes, Y.; Silverman, S. K.; Sheu, C.; Kao, M.; Foote, C. S.; Alvarez, M. M.; Whetten, R. L. Angew. Chem. 1992, 104, 364-366. (17) Mittelbach, A,;Honle, W.;vonSchnering, H. G.; Carlsen, J.;Janiak, R.; Quast, H. Angew. Chem. 1992, 104, 1681-1683. (18)Cui, Y.; Lee, S. T.; Olesik, S. V.; Flory, W.; Mearini, M. J . Chromatogr. 1992, 625, 131-140. 1993 American Chemical Society

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Flgure 2. Chromatograms of fraction A with toluene-acetonltrik and toluene-methanol mobile-phase systems: (A) toluene/acetonltrile = 50150; (B) toluenelmethanol = 50150. Column, Develosll 008-5; temperature, room temperature (ca.15-20 OC), mobllephase Row rate, 1 mL1mln; detector, UV at 325 nm.

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RESULTS AND DISCUSSION Mobile-Phase Selection. In order to separate higher fullerenes by LC, the first task is to find the optimum mobilephase composition. n-Hexane is common in several previous publications with ODS stationary phases,Q-l4 but the low solubility of higher fullerenes limits its use. A toluene-based mobile phase should be useful in this context and toluenemethanol and toluene-acetonitrile were, therefore, evaluated as candidates. With Develosil ODs-5, the monomeric ODs, two mobile phases were examined and the results are shown in Figure 2 [toluene-acetonitrile (A) and toluene-methanol (B)].Although solvent strength of both solvents does not explain the elution if the retention is induced by a typical reversed-phase mechanism, both give relatively good separationsfor fullerenes higher than C70. One can choose the countersolvent in toluene on the following bases: (1)if a better separation for higher fullerenes is desired, acetonitrile is the choice; (2) if faster analysis with a high-resolution column is required, methanol should be used. As this inveatigation was using conventionalsize columns for analytical separation, toluene-acetonitrile was preferred. Experimentsto find the optimum composition of toluene and acetonitrile in the mobile phase showed 4060% acetonitrile in toluene to give the best compromise in resolution, analysis time, and solubility of the samples. Comparison of Monomeric and Polymeric O D s Phases for Higher Fullerene Separation. Figure 3 illustrates the chromatogramof fraction A with the monomeric ODs, several peaks later than C70 retention time being seen. The UVvisible spectra of these peaks are summarized in Figure 4. The peak assignments are tentatively made by reference to published The results are as follows: (A) CW, (B)c70, (C)c78, (D) C78 CW,(E)C78 CZV,(F) no reference data (19)

Diederich, F.; Whetten, R. L. Ace. Chem. Res. 1992,25,119-126.

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mobile phase, toluene1acetonltrlk = 45/55; flow rate, 1 mumin; detection, UV at 325 nm; temperature, room temperature.

found but probably C82, and (G)CU. More accurate identification is made later by FAB-MS measurements. For the comparative study of monomeric and polymeric ODS phases, similar experiments were performed with the Wakosil ODS polymericphase and the chromatagramis shown in Figure 5. The UV-visible spectra of the peaks are summarized in Figure 6. It was confirmed that peaks J, K, L, and N are assigned to the same solutes of peaks D,C, E, and G, respectively, in Figure 3. This means that the elution order of c78 and C78 CW for the monomeric ODS is reversed with the polymeric ODS phase using the same mobile phase. It is also noted that the solute in peak M in Figure 5 cannot be seen in Figure 3, and ita spectrum is very similar to that (20)Diederich,F.;Whe~,R.L.;Thilgen,C.;Etti,R.;Chao,L;Al~z, M. M. Science 1991,254,1768-1770. (21) Diederich, F.; Ettl,R.; Rubin, Y .;Whetten,R. L.; Beck, R.; Alvarez, M. M.; Ans,S.;Senaharma,D.;Wudl,F.; Khemani,K. C.;Koch,A. Science 1991,252, 548-549. (22) Kikuchi,K.;Nakahara,Y.;Suzuki,S.;Saito,K.;Ikemoto,I.;Achi~ Y .Proceedinga of the 3rd Cw Symposium, July 14-16, Tokyo,1992; p 23.

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UV-vielbk spectra of the peaks In the chromawarn In w e 3. Aphabetkal order coneaponds totheso paako In the chrometogrsm.

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of C,8 Da aa observed by Kikuchi et al.22 The right ehoulder of peak N (Z in the figure) gives a spectrum eimilar to peak F in Figure 3, although mme overlapping of their elution distorts ita epectrum. The reaulta indicate that the polymeric ODS phaae givee a different elutiqn order compared to that with the moaomeric ODS phrye; such differences have also been found in the &udy of elution cbaractmbtica of planar and nonplanar polycyclic aromatic hydrocarbons (PAHe) by Wise and Sander- and by Jiaao et al.*n The remita can be explained by the eupmitiollthat the polymeric phaee haa higher molecular planarity recognition capability than the monomeric ODs phaae. Consideration of the shape of the isomers of the higher fullerenee can explain the retentkn differenma. For the isomers of C,& C-78 CW haa the lo est of the ehorter diameter (most bulky), C~llhaa ,%%le different eymmetry, and DSha a narrower shape than the other two isomers. The polymeric ODs elutea bulkier CW fmt and the more cylindricalDa late by virtue of ita planarity recognition capability. The reversed elution order of CM and Cm with the polymeric ODS [aa tentatively aaaigned, peak F (23)Sander, L. C.;Wise, S.A. AM^. C k m . 1987,59,2309-2313. (24) Sander, L. C.;WW, 8. A. LC-CC 1998,8,378490. (25) JIplr0,IC;l h k i , T.;T.nrL., N.; Ohmoto, M.;Fetza, J. C;Bigp, W.R;Gnffih, P.R;Ohger, J. M.J. Cluamotogr. 1980,461,Mg!22'7. (28) J h , K; W,Y.;Mnlhan nm chopn,R.; P e d , J. J.; Fetzer, J. C.; B m , W. R J. Chromatugr. 1891,667,464(6& (27) J m o , K.;Yeummoto, K.; Kuwamoto, T.; NagH.;U d a , T.; Tejima, S.;.Kimura,E; Itoh, K.; Fcttaer, J. C.; Bigga, W. R Chromotolpophro l%Ot,34,381-386.

in Figure 3 and the right ehoulder of peak N (Zin Figure 611 can be ale0 explaiaed the aame way. The Ca dupe is co~d~tobelongerlndnnrrowerthnC~(although~ are pmibilities for at leaut three isomers of thb fullerene to e d t q , and this difference can induce such an elution d e r with the polymeric phase becauaeof ita planarity recognition capability. However, the problem remained aa to why the4 isomer of C,could not be found with the monomeric ODs phaee. The following inveatigation of temperature &ecta addreseee this queation. Temperature Effectr in separation ef Higher Fullerenes. The temperature effect on the aeparation of higher fullerener has been investigabd, the reaulta being eummaried in pigure 'I and 8 for the monomeric and the polymeric phaaea, respectively. In Figure 7, four chromatognunswith the monomeric ODs ere illustrated at column temperaturee from 10 (D) to 70 OC (A). High temperature doea aot give good resolution, and dbueasing the temperature incrsasee the re4ution. A t 30 O C , five clear peake are awn in the chromatogram (C in Figure 7) andthe W-visible spectra of each peak wign themlube in the order of C,8, CW,C,8 CZU,Cg,and CM,respectively. the existence of which haa been However, the peak for &Ds, confirmed by separation with the polymeric phaae m shown in Figures 6 and 6, is not found. When the temperature is decreased to 10 O C with the monomeric pham, th6fifth peak in chromatogram D in Figure 7 appeara to include at least two components, m indicated by the eeteriak in the f w e . W-visible epectral matching for the left aide and the right aide of thie peak aneigna the former aa (&a CZ~, and the latter aa c 7 8 Ds.It is also intereating that thie peak wema to have other componenta coeluted (aa a h o m by the two aeterielrs in the chromatogram), and t h i a might be assignedBB the m n d isomer of CM; further c o n f i i t i o n is required by ke.tuing the column efficiency or decreasing the temperature to 0 OC or below. In Figure 8, four chromatagramewith the poly.raetiCODS are ehown with changing column temperature from 10 to 70 OC at 20 O C intervals. Focueing on the elution profilm for the &e, (278, Cg,and C, isomers,reeolution for those ieomen ia ale0 improved upon decreasing the temperature. At 70 O C , only two major peaks are found and the higher fullerenee em eluted et the ehoulder of the eecond @, however, at 50 OC, fourmajorpeakeareeeen. At 3OoC,clearpealreforthehigher (28) Waknbyaohi, T.;Kikuchi, IC; Nabjima, Y.; Shiromnm, H.; Suzuki, 9.; Achiba, Y.F'roceedinge of the 4th & Sympoeium, January 26-27,ToyohYhi, 1993,p 144.

ANALYTICAL CHEMISTRY, VOL. 65, NO. 19, OCTOBER 1, 1993

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Fbwr 8. Chromatograms of fractlon A with the polymeric ODs at dltferent cdumn temperatures: (A) 70, (E)50, (C) 30, and (D) 10 OC. Mobile phase, toluene/acetonblle = 45/55; flow rate, 1 mL/mln; detection, UV at 325 nm.

fullerene isomers are found, and eight peaks are resolved a t 10 OC. The elution order at 10 OC can be defined by the UV-visible spectral measurements and that is C78 CW,first, c76, C7a C2”, C78 D3,CU, and probably C82, over an elution range from 120to 240min. The peak in Figure 8 at a retention time of 220 min can be assigned to the same solute as in the right shoulder of peak N (Z) in Figure 5. The polymeric ODS phase is more rigid at a temperature range lower than its critical temperature, which is generally between 40 and 50 OC.26 The rigid situation of the alkyl chains at the lower temperature induces much higher molecular planarity recognition capability, and hence, the resolution among these higher fullerene isomers can be improved, based on the difference of the shape of those molecules. FAB-MSIdentificationof Higher Fullerenes. In order to confirm the tentative identification by UV-visible spectra, FAB-MS measurements have been performed for the samples fractionated by the semipreparative-scaleLC procedure. For this purpose, a highly concentrated fraction of higher fullerenes is required as shown in Figure 1,where fraction B

includes a greater concentration of higher fullerenes than fraction A because of their higher solubility in trichlorobenzene. This process has been established previously.*2 Three fractions, 1-3 for c76, C78, and CU isomers, respectively,were collected, since the resolution between the three C78 isomers and between the c82 and CMisomers was not adequate under the conditions used in the semipreparative scale LC separation. Those three fractions were analyzed by FAB-MS negative ion measurements, and the MS spectra obtained are illustrated in Figure 9. As can be seen, the existence of several isomers in these fractions is confirmed as mainly C78and CT8 in fraction 1, C,8 and small amounts of c76 in fraction 2, and c 8 2 and Cw in fraction 3. There are some signals caused by isomer oxides whose existence can be explained by two major reasons: (1)they may be produced in the ionization process of the FAB-MS measurements, and (2) it is well-known that if fullerenes are left in a solution which is exposed to air, oxidation of these materials occurs. The assignmentsby UVvisible spectral measurements are now confirmed. The UVvisible spectra of peak F in Figure 3 and the right shoulder

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F l g w 0. FAB-MS spectra for three fractkns: (A) fractlon 1, (B) fraction 2, and (C) fractkn 3. Mass number tentative assignments: (1) COS (2) GO, (3)c72, (4) C74i (5) c 7 4 0 , (6) (7) GOO*(8)C70v (9) C700. (10) CW, (11) C82, (12) C d , (13) CM, (14) h 0 , and (15) Cee.

of peak N (Z) were thus c o n f i i e d as Cm, because C ~signals Z are clearly seen in the ma88 spectrum of fraction 3.

The elution characteristics of higher fullerenes with monomeric and the polymeric ODS phases have been investigated, and the resulta clearly indicate that shape recognition capability controls the retention order of higher fullerenes. The polymeric phase has higher molecular shape recognition capability and elution order is determined by the shape of the mdecules, but the monomeric phase has a lower such capability and elutes higher fullerenes in the order of their molecular size (molecular weight) with the tolueneacetonitrile mobile phase. Three C78 isomers are resolved with the polymeric phase, and the lower resolution of thme solutes with the monomeric phase has been improved at a lower temperature. C ~ elutes Z later than (2%with the polymeric phase, but the order was reversed with the monomeric phase. These findings can be explained as previously propmed by the authorssn and Wise et al.B*ufor PAH separations. The UV-visible spectra obtained can be identified by FAB-MS measurements after a preparative-LC sample fractionation process.

ACKNOWLEDGMENT The authors show their sincere thanks to Mr. Matauura, JEOL, Akishima, Japan, for his help in FAB-MS measurementa. Useful discussions with Dr. J. C. Fetzer and Dr. W. R. Biggs of Chevron Research and Technology Co., Richmond, CA, are also acknowledged.

RECEIVED for review February 16, 1993. Accepted July 8, 1993.' 0

Abstract published in Advance ACS Abstracts, August 15,1993.