J. Phys. Chem. 1996, 100, 19603-19610
19603
Preparative-Scale Liquid Chromatography and Characterization of Large Fullerenes Generated in Low-Pressure Benzene Flames Henning Richter,† Koli Taghizadeh,‡ William J. Grieco,† Arthur L. Lafleur,‡ and Jack B. Howard*,† Department of Chemical Engineering and Center for EnVironmental Health Sciences, Massachusetts Institute of Technology, 77 Massachusetts AVenue, Cambridge, Massachusetts 02139 ReceiVed: August 5, 1996; In Final Form: October 1, 1996X
Soot generated in a premixed benzene/oxygen/argon low-pressure flame was extracted sequentially with dichloromethane, toluene, CS2 and 1,2-dichlorobenzene. Quantifications by HPLC were performed for C60, C70, C76, C78, and C84 by means of external standards using a monomeric octadecylsilica (ODS) bonded stationary phase and acetonitrile/toluene (55:45) as a mobile phase. Toluene was found to be the most suitable solvent for all the cases studied here. After the partial removal of smaller fullerenes with preparative HPLC, the presence of fullerenes up to C104 was shown by means of HPLC-APCI-MS using a heated nebulizer interface. The macroscopic isolation of C86, C90, C92, C94, and C96 was performed using a two-step HPLC separation with a 2-(1-pyrenyl)ethylsilica stationary phase and toluene as eluent preceded by the partial removal of C60 and C70 with a poly(divinylbenzene) HPLC column. UV-vis spectra were measured, and the presence of a C90 adduct could be discerned. Based on the results of this work, the estimated ratios C60:C70:C76:C78: C84 in the investigated flame-generated soot are 100:100:4:3:6 while the corresponding values for C84:C86: C90:C92:C94:C96 are 100:0.5:5:0.5:1.25:1.
1. Introduction 19851
Since the discovery of fullerenes by Kroto et al. in and their first macroscopic synthesis by Kra¨tschmer et al. in 1990,2,3 the existence of C60 and C70 fullerenes has been well established, and their physical and chemical properties have been investigated. The potential for applications in material sciences4,5 as superconductor,6 batteries, catalysts, or novel optical devices7 in organic or pharmaceutical chemistry8-10 is enhanced by the possibility of producing fullerenes of different sizes and shapes. Besides the relatively well-known C60 and C70 fullerenes, a large class of so-called fullerenic compounds like nanotubes11 or onionlike nanostructures12 have been identified and usually investigated by different microscopic methods. The different fullerenes and fullerenic structures were first obtained using high-intensity laser,1 electric-arc,2,3,11 or electron-bombardment12 heating, but sooting hydrocarbon flames represent a competitive alternative.13 The presence of charged fullerenes in premixed low-pressure acetylene/oxygen/argon and benzene/oxygen/argon flames was described by the Homann group in 1987.14 The macroscopic isolation of C60 and C70 fullerenes from flamegenerated soot by Howard et al. followed in 199115-17 and has been confirmed and extended.18 Other fullerene molecules including oxygen- and hydrogen-containing compounds19,20 and fullerenic nanostructures21-23 have been found in samples condensed from sooting flames. A significant advantage of fullerene synthesis in flames would be the relative ease of scaling up for industrial purposes. The purpose of this work is the investigation of higher fullerenes beyond C60 and C70 generated in premixed lowpressure benzene/oxygen/argon flames. In a previous work the presence of fullerenes containing up to 250 carbon atoms in benzene/oxygen low-pressure flames was suggested using online mass spectrometry.24 Nevertheless, the lack of knowledge of the behavior of such species during condensation with and †
Department of Chemical Engineering. Center for Environmental Health Sciences. X Abstract published in AdVance ACS Abstracts, November 15, 1996. ‡
S0022-3654(96)02356-8 CCC: $12.00
separation from the solid phase does not allow direct conclusions concerning the presence of these compounds in samples condensed from sooting flames. Direct mass spectrometric measurements of species released from flame-generated soot using matrix-assisted laser desorption ionization (MALDI) showed mass peaks corresponding to fullerenes containing more than 150 carbon atoms.18 Despite their great interest, these mass spectrometric results do not reveal the quantity of the detected compounds, and the impact of the ionization process on the resulting spectrum cannot be neglected25 but is only poorly known. Another motivation for this work was the identification of about 5% of the flame-generated material observable by transmission electron microscopy (TEM) as nanostructures, including nested tubes and particles. Examples of multishelled fullerenic nanostructures with dimensions corresponding to C330 to C4200 have been shown.22 These results raise the question as to whether fullerenic structures with carbon numbers between the usually known fullerenes (C60, C70, C76, C78, C84) and the much larger nanostructures observed by electron microscopy might be present in macroscopic amounts. Macroscopic separation of such compounds from the soot and other condensed material is a necessary step for unambiguous identification of the compounds and an eventual determination of their structures and properties. Separation usually involves extraction with appropriate organic solvents such as toluene, followed by HPLC using various stationary and mobile phases.26 For C60 and C70 also selective complexation27,28 or flash chromatography on activated carbon or carbon fibers has been successful.29 A recent approach uses the reversible addition of fullerenes to silicasupported dienes.30 In the present work condensable material including soot was generated in benzene/oxygen/argon flames. Sequential extractions using different solvents were performed in order to check whether extraction may be the limiting step in the isolation of larger fullerenes. The quantities of C60, C70, C76, C78, and C84 obtained in the different extraction steps were determined by HPLC using external calibration standards. For the investigation © 1996 American Chemical Society
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TABLE 1: Sequential Extraction of 8.38 g of Sample of Flame-Generated Condensable Material Including Soot solvent
mass of extracted material (g)
yield (%)
% of total
dichloromethane toluene CS2 1,2-dichlorobenzene total
0.0975 0.21825 0.0090 0.0084 0.33315
1.16 2.60 0.11 0.10 3.97
29.27 65.51 2.70 2.52 100
of higher fullerenes, most of the C60 and C70 were removed by means of preparative HPLC because of the limited solubility of fullerenes. The resulting solution containing relatively more fullerenes beyond C70 was analyzed with liquid chromatography coupled to a mass spectrometer using atmospheric pressure chemical ionization (HPLC-APCI-MS). Compounds with molecular masses corresponding to fullerenes up to C104 were found to be present. After the preparative removal of all fullerenes Cn with n e 84 by means of HPLC, the macroscopic isolation of different fullerenes beyond C84 was performed in a final HPLC step. 2. Experimental Section 2.1. Extraction. The basic sample for this work was 8.38 g of condensable material including soot generated in 2 h operation time in a flat premixed benzene/oxygen/argon flame at a pressure of 5.3 kPa (40 Torr), an atomic C/O ratio of 0.96, 10% argon, and an initial velocity of the fresh gas mixture of 25 cm s-1 (at 25 °C) using equipment described previously.16,17 Although this flame does not give the highest fullerene yields among the flames investigated in the past,16,17 it was selected because a relatively important set of data concerning its properties is available.31 The soot was extracted sequentially with dichloromethane, toluene, CS2, and 1,2-dichlorobenzene, in this order. According to Ruoff et al.,32 C60 solubility in different solvents increases in the order dichloromethane (0.26 mg mL-1), toluene (2.8 mg mL-1), CS2 (7.9 mg mL-1), and 1,2-dichlorobenzene (27 mg mL-1). Assuming the solubilities of higher fullerenes follow the same order, the sequential extraction allows assessment of the influence of solvent strength on the extractability of higher fullerenes which are eventually not dissolved if only toluene is used for extraction. Soxhlet extraction with approximately 600 mL of solvent was used in the cases of dichloromethane and toluene, with extraction times of 16 h, 15 min and 90 h, respectively. Soxhlet extraction has been reported as more efficient than simple reflux;33 former work at MIT34 has shown that when toluene is used as solvent, extraction by sonication is nearly as efficient as Soxhlet extraction. The extraction step with 1,2-dichlorobenzene was performed by sonication in a volume of 75 mL during 90 min due to the high boiling point (179-180 °C) of this solvent. Neither of these extractions methods could be used with CS2 because of the low flash point. The extraction was performed by means of shaking in 100 mL of cold CS2 for some minutes the material remaining after extraction with dichloromethane and toluene; the solvent was removed after about 45 min. The mass of the extracted material was determined for each extraction. Table 1 shows for each extraction the mass of the dissolved material or extract and the yield relative both to the initial sample and to the total extract. The concentrations of C60, C70, C76, C78, and C84 in each extract were measured with a Hewlett-Packard Series 1050 HPLC using a Vydac 201HS54 analytical column (4.6 mm i.d. × 250 mm) with a monomeric octadecylsilica (ODS) bonded stationary phase and an acetonitrile/toluene (55:45) mixture35 as mobile phase at a flow rate of 1.0 mL min-1 with detection at 330 nm. The injected volume
Figure 1. HPLC chromatogram of toluene extract of initial flame sample: Vydac 201HS54 analytical column, 1 mL min-1 toluene/ acetonitrile (55:45), detection at 330 nm, injection of 20 µL.
was in all cases 20 µL. Calibration curves using external standards were established by preparing three solutions of C60 and C70 (MER, Tucson, AZ) and three solutions of C76, C78, and C84 (TechnoCarbo, Plan de Grasse, France) at different concentrations. Each solution was injected three times. The concentration of the dichloromethane and the toluene extracts was increased in order to improve the precision of the measurement. The solvent of the CS2 and 1,2-dichlorobenzene extracts was evaporated and replaced by toluene. The HPLC chromatogram of the toluene extract is shown in Figure 1, and the amounts of C60, C70, C76, C78, and C84 found in the extracts are given in Table 2 for each extraction and the sum of all extractions in terms of masses as well as yields relative to both the initial sample and the extract. These results show that most of the extractable material and most of each fullerene included here are extracted with dichloromethane and toluene. The C70/C60 ratio in the different extracts increases in the sequential extractions from dichloromethane to CS2 and then decreases slightly for 1,2-dichlorobenzene. The results of the dichloromethane extraction might be low due to the relatively short extraction time of 16 h, 15 min compared to 90 h in the case of toluene; an increase of the dichloromethane extraction time could lead to higher values. The fraction of C60-C84 fullerenes as part of the material dissolved by a given solvent exhibits a maximum of 79.60% of the extract for toluene and a minimum of 41.31% for 1,2-dichlorobenzene. As shown in Table 2, 96.55% of the extracted C60-C84 fullerenes are extracted with dichloromethane or toluene. Taking into account that the solubility of C6032 and probably all other fullerenes is much higher in toluene than in dichloromethane, the direct extraction with toluene without preliminary extraction with dichloromethane should dissolve about the same amount of fullerenes as shown here for both solvents. As can be seen in Table 2, the extracted amount of C76, C78, and C84 fullerenes relative to that of C60 and C70 increases only slightly using solvents stronger than toluene. Despite an increase in solubility of C60 by a factor of about 10 between toluene and 1,2dichlorobenzene, more than 90% C76, C78, and C84 are extracted with dichloromethane or toluene. This yield could probably be improved by increasing the extraction time. Also, similar results could be expected in a single-step extraction with CS2 or 1,2-dichlorobenzene, but use of these solvents would not be justified considering the explosion risk of CS2, the high boiling point of 1,2-dichlorobenzene, and the health effects of both solvents. Therefore, with long-time Soxhlet extraction or with sonication for smaller samples, toluene remains the solvent of first choice also for C76, C78, and C84. Despite the poor
Large Fullerenes Generated in Benzene Flames
J. Phys. Chem., Vol. 100, No. 50, 1996 19605
TABLE 2: Amounts of Fullerenes C60, C70, C76, C78, and C84 Found in Each Sequential Extraction of 8.38 g of Flame Sample with Four Different Solvents solvent fullerene C60
C70
C76
C78
C84
C60-C84
quantity
dichloromethane
toluene
carbon disulfide
1,2-dichlorobenzene
sum for all extractions
mass (mg) % of sample % of extract % of C60 total mass (mg) % of sample % of extract % of C70 total mass (mg) % of sample % of extract % of C76 total mass (mg) % of sample % of extract % of C78 total mass (mg) % of sample % of extract % of C84 total mass (mg) % of sample % of extract % of (C60-C84) total
34.056 0.41 34.93 33.28 14.914 0.18 15.30 12.75 0.6474 0.0077 0.66 17.34 0.3897 0.0047 0.40 13.61 0.6150 0.0073 0.63 9.47 50.622 0.61 51.92 21.79
66.51 0.79 30.47 64.98 96.76 1.15 44.34 82.76 2.86 0.034 1.31 76.60 2.27 0.027 1.04 79.26 5.32 0.063 2.44 81.96 173.72 2.06 79.60 74.76
0.8558 0.010 9.51 0.84 3.1227 0.037 34.70 2.67 0.1344 0.0016 1.49 3.60 0.1057 0.0013 1.17 3.69 0.3229 0.0039 3.59 4.97 4.5415 0.054 50.46 1.96
0.9240 0.011 11.00 0.90 2.1231 0.025 25.27 1.82 0.0917 0.0011 1.09 2.46 0.0985 0.0012 1.17 3.44 0.2333 0.0028 2.78 3.60 3.4706 0.0411 41.31 1.49
102.35 1.22 30.72 100 116.92 1.39 35.10 100 3.7335 0.0444 1.12 100 2.8639 0.0342 0.86 100 6.4912 0.0770 1.95 100 232.36 2.77 69.75 100
knowledge of the solubility behavior of fullerenes larger than C84, there is no reason to assume a sudden change from the trend seen in the C60-C84 range. Continuation of the decrease of solubility with increasing molecular weight seems to be more likely. The results in Table 2 show that the reported32 increase of C60 solubility by a factor of about 10 in going from toluene to 1,2-dichlorobenzene has only a very small effect on the extraction of all fullerenes up to C84. Therefore, at least the first fullerenes beyond C84 condensed from sooting flames can be assumed to be extracted efficiently using toluene as solvent. 2.2. Mass Spectrometric Measurements. Mass spectrometry was used to check qualitatively the fullerenes present in the toluene extract. The measurements were based on detection of positive ions with a API-I single quadrapole mass spectrometer (Sciex, Thornhill, Ontario, Canada) equipped with a heated nebulizer interface and using chemical ionization.20,36 Two spectra were measured: one in the mass range 700-1050 amu (Figure 2a) and one in the mass range 1000-1350 amu (Figure 2b). The split into two spectra allows measurement of a smaller mass range and higher sensitivity of the instrument. Mass peaks corresponding to C60, C60O, C70, C70O, C76, C78, C84, C86, C88, C90, C92, C94, C96, and C98 could be identified. The C60 and C70 peaks are by far the most prominent, in contrast to those of fullerenes larger than C84 as is revealed by the C84 peak in both spectra. Although the exact concentrations cannot be determined from mass spectrometric peak heights alone, in the present case the peak heights can serve as a first indication of relative abundances. The ionization efficiencies of at least C60, C70, C76, C78, and C84 are very similar, as it has been shown in Figure 3 by results from the simultaneous measurement of these fullerenes in a standard solution containing 0.0049 mg mL-1 of each compound. The evolution of the peak height follows in a first approximation the molar concentrations. Sequential extractions of condensed material, referred to as soot, from carbon vaporization under fullerene-forming conditions have been previously performed using other solvents such as 1,2,3,5-tetramethylbenzene,33,37 N-methyl-2-pyrrolidinone,33 1,2,3-trichlorobenzene,33 quinoline,38 and toluene at high pressure/high temperature.33,39 Shinohara et al.38 report fast atom bombardment mass spectra containing peaks equivalent in mass
Figure 2. Mass spectra of toluene extract of initial flame sample, positive ion mode: (a) 700-1050 amu and (b) 1000-1350 amu.
to fullerenes up to C300. Parker et al.33 claim on the basis of the successive extractions with benzene, pyridine, and 1,2,3,5tetramethylbenzene which dissolved 44% of the initial soot and the extraction yield of 94% for N-methyl-2-pyrrolidinone (NMP) using raw soot that a large portion of the soot has a molecular fullerene-type structure. Using laser desorption Fourier transform mass spectrometry (LD-FTMS), they detected species equivalent in mass to fullerenes up to C466. However, the
19606 J. Phys. Chem., Vol. 100, No. 50, 1996
Richter et al. TABLE 3: Recovery of Fullerenes C60, C70, C76, C78, and C84 from Different Columns after Preparative HPLC Separation column no. 1 2 3
Figure 3. Mass spectrum of standard solution containing 6.81 × 10-9 mol mL-1 (or 6.81 µM) C60, 5.83 µM C70, 5.37 µM C76, 5.24 µM C78, and 4.86 µM C84.
reported fullerene yields with N-methyl-2-pyrrolidinone contradicts the relatively poor solubility of C60 in this solvent (0.89 mg mL-1).32 Also, the possible formation of aggregated clusters during the ionization by laser desorption must be considered.40 In the present work a flame-generated sample of condensables preextracted with 1,2-dichlorobenzene was sequentially extracted with 1,2,3,5-tetramethylbenzene and N-methyl-2-pyrrolidinone, and the extracts were analyzed by mass spectrometry using the above-mentioned instrument in the positive ion mode with a mass resolution allowing the presence of individual fullerenes to be checked. Noisy spectra appearing to show regular maxima were obtained, but no prominent peaks corresponding to molecular masses of fullerenes were present. Interestingly, compounds were detected with molecular masses equivalent to C60 and C70 adducts containing up to 10 1,2,3,5-tetramethylbenzene units which could have been formed during either the extraction or ionization processes. Therefore, no special efficiency of 1,2,3,5-tetramethylbenzene or N-methyl-2-pyrrolidinone for the extraction of higher fullerenes is seen in the present study. Given that mass spectrometry alone is not sufficient for fullerene identification, a preparative isolation must be performed which allows further characterizations and accurate quantification of relatively pure compounds. In the present work all separations were performed by HPLC. 2.3. High-Performance Liquid Chromatography. The macroscopic separation of fullerenes beyond C84 by HPLC necessitates the use of a stationary phase with a sufficient separation capacity and of a mobile phase in which the fullerene solubility is high enough to dissolve the necessary amounts. Diederich et al.41 in early work used gravity chromatography on alumina with hexane-toluene gradient elution followed by fast atom bombardment (FAB) and laser desorption time-offlight (LD-TOF) mass spectrometry. Samples enriched in C90 and C94 were separated. The corresponding HPLC peaks were identified using SiO2 as support and hexane with addition of 2% of toluene for solubility reasons as mobile phase. Unfortunately, this approach is not suitable for the separation of larger amounts due to the very poor solubility of fullerenes in hexane (0.043 mg mL-1 for C6032). Kikuchi et al.42 report the analytical separation and identification of C90 and C96 by HPLC using CS2 as eluent in a time-consuming recycle method. HPLCMS measurements of Anacleto et al.43,44 show the presence of compounds with molecular masses corresponding to fullerenes up C9643 or C108.44 They used different monomeric or polymeric octadecylsilica (ODS) or 2-(1-pyrenyl)ethylsilica phases with a mobile phase gradient of initially acetonitrile/dichloromethane (50:50) programmed to 100% dichloromethane.43 This method
name
percentage of injected compound C60
C70
C76
C78
C84
Vydac 201TP510 10.98 15.91 5.22 2.80 3.36 Alltech Hypersil ODS5u 19.59 22.11 14.21 13.47 16.89 Jordi Gel-500 (PDVB) 80.35 75.92 33.29 58.99 61.98
also does not seem to be appropriate for preparative-scale separations due to the low solubility of fullerenes in the mobile phase. In the present work the first step of the separation by HPLC consists of a preparative-scale fractionation of the toluene extract in order to remove a substantial part of C60 and C70, the most prominent fullerenes in the sample (Table 2). This approach allows the relative abundance of higher fullerenes to be increased, a condition essential for preparative-scale separation considering the limited solubility of these compounds. For this purpose three different HPLC columns have been tested: (1) a polymeric octadecylsilica column (Vydac 201TP510, 250 × 10 mm) using a flow of 5.0 mL min-1 actetonitrile:toluene (55: 45); (2) a monomeric octadecylsilica column (Alltech Hypersil ODS5u, 250 × 22 mm) using a flow of 10.0 mL min-1 acetonitrile:toluene (50:50); and (3) a poly(divinylbenzene) (PDVB) column (Jordi Gel-500, 250 × 22 mm) with 4.0 mL min-1 dichloromethane:toluene (60:40) for elution. The injection volume was 5 mL, and all preparative sampling was performed with a Beckman System Gold HPLC (Solvent Module 116, Detector Module 166). Using columns 1 and 2, three fractions were collected: a fraction containing C60 and C70, a fraction containing Cn with 70 < n e 84, and a fraction including the tail of the C84 peak and the higher fullerenes. In the case of column 1, the separation capacity is close to that described for a similar analytical column by Jinno et al.,35 with an elution time sensitive to temperature. With column 2, a good separation of C60, C70, and C84 was observed while no base line resolution between C76 and C78 could be obtained. Using column 3, no base line resolution even between C60 and C70 could be observed so only two fractions were collected, one containing the major part of C60 and C70 and the tail of the spectra containing the higher fullerenes. Essential to the suitability of a column is sufficient recovery, expressed for each fullerene as the ratio of the amount of the fullerene collected to the amount of the fullerene injected. Recoveries of fullerenes C60, C70, C76, C78, and C84 for the three columns were determined by analyzing with the same calibrated Vydac 201HS54 analytical column as were used in analyzing the initial toluene extract and maintaining similar experimental conditions. The results, summarized in Table 3, show a very small recovery for column 1 so that this column is not suitable for preparative separations. Such low recoveries can indicate that a part of the fullerenes sample either is irreversibly retained on the stationary phase or does not sufficiently interact with the stationary phase and is eluted with the solvent peak. The contribution of the second effect for column 1 was evaluated by collecting a fraction usually considered as the “solvent peak” and containing polycyclic aromatic hydrocarbons in flame samples. Analysis showed relative to the injected amounts 75.47% C60, 51.00% C70, and no detectable C76, C78, or C84. Therefore, for column 1, lack of interaction between C60, C70, and the stationary phase is clearly important, and retention on the stationary phase is also a considerable problem for C60 and C70 and especially for the other fullerenes. The observed retention may indicate irreversible adsorption, discussed by Gasper and Armstrong in the context of column deterioration.26
Large Fullerenes Generated in Benzene Flames Somewhat better but still low recoveries were observed with column 2 (Table 3), but the detailed reasons for the low recoveries were not studied in this case. Fifteen 5 mL injections of the toluene extract, representing 2.63 g of the initial flame sample, were made into this column, and three fractions were collected as described above. The fraction containing the tail of the C84 peak and the higher fullerenes was evaporated to 1.05 mL, and a 20 µL portion of this sample was injected into a Vydac 201HS54 analytical column coupled to the same mass spectrometer mentioned above via the heated nebulizer interface. The eluent was acetonitrile:toluene (55:45) with a flow of 0.5 mL min-1. The instrument is also equipped with a diode array detector (Groton Technology, Concord, MA). The mass spectrometer was started 60 min after the begin of the HPLC run, and the scan range covered 1100-1250 amu. As shown in Figure 4a-c, mass peaks corresponding to C84, C86, C88, C90, C92, C94, C96, C98, C100, C102, and C104 could be detected. UVvis spectra could be measured but were close to the detection limit. In a preliminary run using a sample containing all fullerenes larger than C70, C82 could be detected unambiguously as well as C84, C86, C88, C90, C92, C94, and C96. Unfortunately, the retention times for the different peaks attributed to the fullerenes C84-C104 overlap significantly for nearly all the species, so the present experimental conditions do not seem appropriate for collecting macroscopic amounts of pure compounds using a monomeric octadecylsilica stationary phase as in column 2. 2.4. Macroscopic Isolation of C86, C88, C90, C92, C94, and C96. Twenty milliliters of a 1,2-dichlorobenzene extract of 6.3 g of condensable material including soot, generated under the same conditions described above, was diluted with 80 mL of toluene. The resulting 100 mL solution was introduced as 20 5 mL injections into the poly(divinylbenzene) column 3, under the above-described conditions. Two fractions were collected, one containing C60 and C70 and the other the tail of the chromatogram. The recoveries shown in Table 3 are clearly higher than for columns 1 and 2, so that column 3 seems suitable for preparative work. Mass spectrometry of the tail fraction was performed using the same equipment and methods as described above. Peaks equivalent to fullerenes up to C98 were detected unambiguously, and relatively low peaks could be attributed to C102, C104, C108, and C110. For further separations a 2-(1-pyrenyl)ethylsilica stationary phase (Cosmosil BuckyPrep column, 250 × 4.6 mm) with a toluene flow of 1.0 mL min-1 for elution was used on a HewlettPackard Series 1050 instrument equipped with a HP 1040A diode array detector. The use of pure toluene for elution allows the loading of higher quantities of fullerenes; preliminary tests with standard solutions of C60, C70, C76, C78, and C84 show peak heights increased by a factor 5 for C60 and C70 and by 4.5-6.6 for C76, C78, and C84 compared to the Vydac 201HS54 analytical column. The peak areas increased by 1.25-1.42 and 1.592.78, respectively. These results show the occurrence of less adsorption of fullerenes on 2-(1-pyrenyl)ethylsilica stationary phase and thus the possibility of a better recovery for preparative separations. The volume of the dichloromethane/toluene solution containing the tail of the poly(divinylbenzene)-based separation was reduced to 22 mL and analyzed with the Vydac 201HS54 analytical column. The solution contained 0.9807 mg of C60, 3.0441 mg of C70, 0.4254 mg of C76, 0.2377 mg of C78, 1.2275 mg of C84, and an unknown quantity of higher fullerenes. In order to limit the number of injections, the solvent was replaced by 630 µL of 1,2-dichlorobenzene because of its greater solubility for fullerenes. A 550 µL aliquot of these solutions
J. Phys. Chem., Vol. 100, No. 50, 1996 19607
Figure 4. Mass spectrometric ion extract of HPLC measurement by direct HPLC/MS coupling: (a) C84, C86, C88; (b) C90, C92, C94, C96; (c) C98, C100, C102, C104.
(5 × 20 µL and 9 × 50 µL) was injected into the Cosmosil BuckyPrep column. Figure 5 shows the chromatogram of a run with injection of 50 µL; the peak height of C70 is not represented correctly because the detector was saturated. C60, C70, C76, C78, and C84 could be attributed by means of the previous run of standard solutions. Several peaks are observed beyond C84. For each injection two fractions were collected:
19608 J. Phys. Chem., Vol. 100, No. 50, 1996
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Figure 5. HPLC chromatogram of toluene extract after partial removal of C60 and C70: Cosmosil Buckyprep column, 1 mL min-1 toluene, detection at 330 nm, injection of 50 µL.
Figure 7. Mass spectra of (a) fraction 3 and (b) fraction 4 as described in Table 4. Figure 6. HPLC chromatogram of toluene extract after removal of C60, C70, C76, C78, and partially C84: Cosmosil Buckyprep column, 1 mL min-1 toluene, detection at 330 nm, injection of 50 µL.
a fraction containing all peaks beyond C70 and including the major part of the C84 peak and another fraction with the tail of the C84 peak and beyond. The volume of the last fraction was reduced and the solvent replaced by 1,2-dichlorobenzene. Two 50 µL injections were done; after the first one the sample volume was reduced slightly. Figure 6 shows the chromatogram of the second injection. Ten fractions were taken for both injections, and the solution eluting between the fractions was also collected. All fractions collected were analyzed by mass spectrometry as described above. Figure 7a,b represents the mass spectra obtained for fractions 3 and 4. It can be concluded that nearly pure C90 was isolated (Figure 7a); the significance of the C90 peak in Figure 7b will be discussed below. Table 4 shows the collection times corresponding to Figure 6 and the assigned species in decreasing order of peak height. The mass spectra for fractions 8-10 are relatively noisy and include many peaks which cannot be assigned. A 20 µL sample of each fraction was reinjected into the HPLC using the same experimental conditions as for the initial separation represented in Figure 6. The volumes of each fraction were reduced to about 1 mL. The clearly different retention times confirm the successful separation. An approximate calibration of the amounts of collected fullerenes as shown in Table 4 was performed using C84 as external standard. The test of this procedure for C76 and C78 using a C76, C78, and C84 standard solution indicates only small errors. These results show that macroscopic amounts of C86, C88, C90, C92, C94, and C96 with different purities were isolated. Considering the total injection volume in each separation step,
TABLE 4: Fractionated Sampling of Cn Fullerenes with n g 84; Detected Species Are Listed in Order of Decreasing Peak Height fraction
collection time (min)
attributed species
1 2 3 4 5 6 7 8 9 10
28.5-31.5 32.5-35.3 41.2-44.3 47.1-50.7 53.3-58.8 62.3-66.2 68.5-74.9 80.1-91.3 99.4-103.5 104.4-147.5
C84 C84, C86, C70, C60 C90 C92, C90 C94, C92, C90, C84 C96 C84, C98, C92, C70 C84, C102, C102O C84, C102O C84, C102O
a
amount (µg) 60 150 56a 40 34 16 12
16 µg of C92 and 40 µg of C90 adduct.
these quantities result from about 4.5 g of flame sample. A more careful statement is justified for C98 and C102 (fractions 7 and 8) because C84 shows the most important mass peak; nevertheless, no C84 could be observed by HPLC after reinjection. This observation could be explained by the presence of a metastable C84-containing compound, decomposed in the ionization process before the mass spectrometric analysis. No fraction contains C88, but it could be detected by mass spectrometry in the eluent collected between the fractions. Its attribution to the small peaks between fraction 2 and 3 seems to be likely (Figure 6). UV-vis spectra were measured with a relatively high quality, but only beyond 240 nm due to the absorption properties of toluene, which has been used as eluent. It should be pointed out that all spectra shown in this work were measured near the maxima of the chromatogram of Figure 6 and not with the pure fractions after reinjection where the concentrations are much lower. Figure 8a represents the UV-vis spectra of C84, C86, and C88 while the spectra of C90 (fraction 3) and the two peaks of fraction IV are represented in Figure 8b. Both UV-vis
Large Fullerenes Generated in Benzene Flames
J. Phys. Chem., Vol. 100, No. 50, 1996 19609
Figure 9. HPLC chromatogram of fraction containing C70 to C84: Cosmosil Buckyprep column, 1 mL min-1 toluene/acetonitrile (80:20), detection at 330 nm, injection of 20 µL.
of a prominent C84 peak in mass spectrometry could eventually be attributed to an overlapping of the fullerene peaks with C84 adducts of unknown structure. An increase of resolution of the Cosmosil BuckyPrep column was achieved using a toluene:acetonitrile mixture (80:20) as eluent at a flow of 1.0 mL min-1; mass spectrometrically pure C90 (17 µg), C94 (2 µg), and C96 (4 µg) were isolated. The relative to the injected volume amount of collected fullerenes is smaller than in the case of elution with pure toluene (Table 4). This observation as well as the absence of all peaks after the elution of C96 could be related to adsorption on the stationary phase. Nevertheless, the analysis of the fraction containing all peaks beyond C70 and including the major part of the C84 peak (Figure 5) allowed the separation of C70O and of a C60 adduct, showing only a strong C60 peak in the mass spectrum despite its elution between C78 and C84 (Figure 9). Also, the UV-vis spectrum corresponds to previous results.34 3. Conclusions
Figure 8. UV-vis spectra of (a) C84, C86, C88; (b) C90, C92, C90 adduct; and (c) C94, C96.
spectra of fraction IV are different from C90 as isolated in fraction 3 despite the mass spectrometric results showing mass peaks corresponding to C90 and C92 (Figure 7b). This behavior could indicate the presence of metastable cyclopentadiene adducts, previously observed in flame-generated samples,45 which cannot be identified by mass spectrometry using ionization with a heated nebulizer interface because of their thermal instability. Characteristic for UV-vis spectra of the C60 and C70 cyclopentadiene adducts is the presence of a “hump” at about 300-350 nm.34 The assignment of the second chromatographic peak in fraction 4 to a cyclopentadiene adduct of C90 or alternatively another C90 derivative and of the first one to C92 seems justified. The UV-vis spectra of C94 and C96 are shown in Figure 8c. No assigment of UV-vis spectra to fullerenes beyond C96 has been made because of the relatively low concentrations and poor peak purity observed by mass spectrometry. The appearance of only one chromatographic peak after reinjection of fractions 7 and 8 despite the presence
The presence of fullerenes up to C104 in flame-generated material including soot has been shown by HPLC/MS analysis after extraction with toluene which has been demonstrated to be the appropriate solvent for this purpose. Due to adsorption of higher fullerenes on the stationary phase, the choice of the chromatographic column and of the eluent is essential for a macroscopic isolation of higher fullerenes by HPLC. A 2-(1pyrenyl)ethylsilica stationary phase has been identified as suitable. Macroscopic amounts of C86, C90, C92, C94, and C96 were isolated after the removal of most of the C60 and C70. High purity was obtained for C90, C94, and C96. A C90 adduct was collected with C92, and this observation will allow the isolation of pure C92 and of the C90 adduct. The overlapping of different adducts with higher fullerenes can be assumed so that the consequent use of Soxhlet extration or heating of the sample prior to separation would be convenient because of the thermal instability of fullerene adducts. Based on the results of this work, the ratio C60:C70:C76:C78:C84 can be estimated as 100: 100:4:3:6 while the corresponding values for C84:C86:C90:C92: C94:C96 are 100:0.5:5:0.5:1.25:1. The extraction of soot generated in premixed benzene/oxygen/argon low-pressure flames coupled to a multiple-step chromatographic separation allows the isolation of macroscopic amounts of fullerenes beyond C84. Using the existing combustion facility, the production of soot samples large enough for the isolation of these compounds in milligram quantities requires only some hours. Using the results of the present work, the isolation of 1 mg of C90 requires about 26 g of flame-generated soot (6 h of operation) and isolation of
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