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(17) Langer, S. H.; Patton, J. E. I n "New Developments In Gas Chromatography"; Purnell, J. H., Ed.; Wiley: New York, 1973; pp 293-373. (18) Langer, S. H.; Melton, H. R.; Griffith, T. D.; Coca, J. J. Chromatogr. 1976, 722,487-503. (19) Phllllps, C. S. G. I n "Gas Chromatography 1970" Stock, R., Ed.; Institute of Petroleum: London, 1971; pp 1-19. (20) Katsanos, N. A.; Tsiatsios, A. J . Chromatogr. 1981, 273, 15-24. (21) Bolme, M. W.; Langer, S. H. J . Phys. Chem. 1983, 87,3363-3366. (22) Sindorf, D. W.; Maciel, G. E. J. Am. Chem. SOC. 1083, 705, 1848-1851. (23) Bayer, E.; Albert, K.; Relners, J.; Nleder, M.; Muller, D. J. Chromatogr. 1983, 264, 197-213. (24) Marshall, D. B.; McKenna, W. P. Anal. Chem. 1984, 56,2090-2093. (25) Gilpin, R. K.; Gangoda, M. E. J. Chromafogr. Scl. 1983, 27, 352-361. (26) Lochmuller, C. H.; Marshall, D. B.; Wilder, D. R. Anal. Chlm. Acta 1981, 730,31-43. (27) Bogar, R. G.; Thomas, J. C.; Callis, J. 6 . Anal. Chem. 1984, 56, 1080-1084. (28) Sander, L. C.; Callis, J. B.; Field, L. R. Anal. Chem. 1983, 55, 1068-1075. (29) Bush, S.G.; Jorgenson, J. W.; Miller, M. L.; Linton, R. W. J. Chromafogr. 1983, 260, 1-12. (30) Lochmuller, C. H.; Wilder, D. R. Anal. Chim. Acta 1080, 778, 101- 108. (31) Wirth, M. J.; Hahn, D. A.; Holland, R. A. Anal. Chem. 1983, 55, 787-790. (32) Langer, S. H.; Chu, A. H.; Bolme, M. W.; Turner, M.; Quinting, G. R. J. Chem. Res ., in press. (33) Majors, R. E.; Barth, H. G.; Lochmuller, C. H. Anal. Chem. 1982, 54, 323R-363R. (34) Majors, R. E.; Barth, H. G.; Lochmuller, C. H. Anal. Chem. 1984, 58, 300R-349R. (35) Colin. H.; Gulochon, G. J. Chromatogr. 1077, 747, 289-312. (36) Colin, H.; Guiochon, G. J . Chromatogr. 1978, 758,183-205. (37) Melander, W. R.; Horvath, C "High Performance Liquld Chromatography: Advances and Perspectives"; Horvath, C., Ed.; Academlc Press: New York, 1980; Vol. 2, "Reversed-Phase Chromatography", pp 114-3 19. (38) Gllpin, R. K.; Sisco, W. R. J. Chromatogr. 1980, 794,285-295.
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RECEIVED for review February 13, 1985. Accepted June 3, 1985. We thank the Army Research Office, the National Science Foundation, and the University of Wisconsin for support of this work.
High-Performance Liquid Chromatography of Vanadyl Porphyrins Padmanabhan Sundararaman Chevron Oil Field Research Company, P.O. Box 1627, Richmond, California 94802-0627
A rapid, hlghly reproduclble hlgh-performance llquld chromatographic method Is descrlbed for the analysis of vanadyl porphyrlns from fossll fuels. The ablllty to separate structural as well as posltlonal Isomers Is also demonstrated. This method ellminates the problems and amblgultles assoclated wlth the demetalation of porphyrin mixtures whlch Is currently belng used by several workers In this fleld.
The original discovery of petroporphyrins in fossil fuels by Treibs ( I , 2 )indicated a biological source for petroleum. In spite of their discovery 50 years ago, porphyrins have found very little application in geochemical correlations. Moreover, until recently very little was known about their structure. This slow progress in petroporphyrin chemistry has mainly been due to the difficulties encountered in their isolation and analysis. Porphyrins in fossil fuels occur as a complex mixture and complete structures of a few of the compounds have been determined by nuclear magnetic resonance spectrometry and are listed in Figure 1. The major problem in determining porphyrin composition is that under normal conditions they are too involatile to be analyzed by gas chromatography (GC). 0003-2700/85/0357-2204$0 1.50/0
Probe mass spectrometry (3) and in recent years high-performance liquid chromatography ( 4 , 5 )and gas chromatography/mass spectrometry (GC/MS) of suitable volatile derivatives (6-9) have been applied to the analysis of petroporphyrins. Most of these methods involve demetalation as a principal step. Demetalation of metalloporphyrins is usually carried out by heating with a strong acid such as methanesulfonic acid and the yield, especially in the case of vanadyl porphyrins, is seldom greater than 75%. The basic assumption is made that every compound in this complex mixture decomposes to the same extent, e.g., the relative amounts of the various compounds present are unchanged during demetalation. This assumption has never been proven beyond doubt. Moreover, demetalation is one additional step in the analysis, thereby increasing the time required for the analysis: an especially precious commodity in an industrial laboratory. Therefore, because of the inconvenience and ambiguities surrounding the demetalation, attempts are being made to analyze metalloporphyrins directly. Recently Barwise ( I O ) introduced a method for purifying vanadyl porphyrins using propanesulfonic acid bonded silica, and the MS data on vanadyl porphyrins can be obtained directly without demetalation. Roberts et al. (11) and Gallegos 0 1985 American Chemical Soclety
R = Et R=Me
Q @ \' ,/
/
R'R=Me R = -CH2-COlH
/
RiEt R=Me R i H
t0,H R. =R,= Rl= Me: R. = n
Figure 2. E1 mass spectrum of vanadyl pwphyrins isolated from BOScan 011. Venezuela.
R, = 4 R 2 = Et: A, =Ma R, = R t = E t : R J = Me R. =RI= R.= Et
Flgure 1. Smctures of geoporphyrins Isolated and Conflnmd by specbal data (ref 15-30).
et al. (12)have f d their attention on the GC/MS analysis of metalloporphyrins with limited success. Recently, the increasingly powerful technique of high-performance liquid chromatography has been applied to the analysis of vanadyl porphyrins. Both Fetzer e t 81. (13)and Fish e t al. (14) concentrated on developing an element specific detector for vanadyl complexes, but no attempts were made to improve the resolution, which was generally poor. The major objective of our work is to develop an efficient and reproducible method for analyzing metalloporphyrins directly, thereby eliminating the demetalation step and problems associated with it, and to apply it to exploration problems. We have focused our attention on vanadyl porphyrins and the results of our efforts are summarized in this paper. EXPERIMENTAL SECTION Reagents. Organic solvents were LC grade, Distilled-in-Glass from Burdick and Jackson, and the water was double deionized with a Millipore instrument. The propanesulfonic acid bonded silica was obtained from Analytichem International and the alumina (Activity Grade 11) was obtained from E. Merck & Co. The Zorhar Si1column was obtained from Du Pont Instruments as 5-pm spherical silica particles in 25 cm X 9.6 mm prepacked column. The Hypenil CI column was obtained from Shandon Instruments as 3-pm particles in 250 mm X 4.6 mm prepacked columns. Instrumentation. The HPLC was oerformed on a HewIetLPackard 1090 system with three solveit capability, equipped with a photodiode array detector.
0
10
20
30
40
IO
60
70
10
o(I
Tine. Mh.
Figure 3. Ulmmatogram of vanadyl porphyrins isolated from &scan oil. Venezuela: column, 25 cm X 0.46 cm, 3 Mm Hypersil C,,, Shandon Southern; detectlon, UV at 406 nm; solvent. 4 5 % CH,OH. 4 5 % CH&N, and 10% H 2 0 flow rate, 1 mL/mln.
Procedures Isolation. One gram of crude oil or bitumen was adsorbed onto 10 g of active alumina and introduced on top of 100 g of alumina in a glass column. The saturated and aromatic hydrocarhons were eluted with 10% ether in hexane. Themetalloporphyrins were eluted with methylene chloride. The eluent was comtantly monitod hy UV-vis spectrometry for quantitative isolation of porphyrins. Separation into Nickel and Vanadyl Porphyrins. By use of a semipreparative Zorhax Si1 column (9.6 mm x 25 em) and a gradient elution starting from 10090 hexane to 10090 methylene chloride over 120 min followed by 100% methylene chloride for 60 min a t a flow rate of 5 mL/min, the metalloporphyrins can he separated into nickel and vanadyl porphyrins. The less polar nickel porphyrins elute first. The eluents were monitored constantly with a UV-vis spectrophotometer. Purification of Vanadyl Porphyrins for MS and HF'LC Analysis. The vanadyl porphyrins were further purified by the procedure described by Barwise (IO)using silica functionalized with propanesulfonic acid. H P U : Analysis of Vanadyl Porphyrins. The vanadyl porphyrins were analyzed hy using a 3-pm Hypersil C,, column (4.6 mm X 25 cm). The mobile phase was 45% CH,CN, 45% methanol, and 10% H20. The flow rate was 1mL/min and the column back pressure was approximately 160 bar. The eluent was monitored with the diode array detector set at 406 nm. RESULTS AND DISCUSSION The mass spectrum and the chromatogram of vanadyl porphryins from Boscan crude oil, Venezuela, are shown in Figures 2 and 3, respectively. The mass spectrum is able to separate the homologous aeries and the types (ET10 and DPEP) of porphyrins present in this mixture. Unfortunately, it is unable to separate structural isomers, since all the
2206
ANALYTICAL CHEMISTRY, VOL. 57, NO. 12, OCTOBER 1985
C32D
,
- L L L
I, I
C31E
C29E DPEP
Figure 4. Elution profiles showing the resolution of structural isomers within each carbon number and porphyrin type. Conditions were as given In Figure 2 (D, DPEP; E, ETIO).
structural isomers of single-carbonnumber and type have the same molecular weight and thus appear as a single peak. On the other hand, inspection of Figure 3 shows that HPLC not only can separate the homologous series and type but can also separate structural isomers. The ability to separate these structural isomers with the 3-pm Hypersil C18 column is further demonstrated in Figure 4. The individual peaks from Boscan oil were separated by using a combination of reverse-phase and silica semipreparative columns and their molecular weights were determined by MS. The peaks with the same molecular weight were recombined and analyzed using the Hypersil C18column. The results of a series of such analyses demonstrating the abiltiy to separate structural isomers are shown in Figure 4. For example, in Figure 4 the Cz8E chromatogram was obtained by combining individual peaks with a molecular weight of mlz 487 and analyzing the resultant mixture of structural isomers thus obtained by using the Hypersil C18 column. Each of the chromatograms (e.g., C&, C&, etc.) in Figure 4 was obtained in a similar manner.
ACKNOWLEDGMENT I thank M. M. Pens for carrying out experimental work, J. C. Fetzer for isolating vanadyl porphyrins from Boscan
crude oil, E. J. Gallegos for MS data, and J. M. Moldowan and W. K. Seifert for helpful discussion.
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RECEIVED for review March 18,1985. Accepted June 13,1985. I wish to thank Chevron Oil Field Research Company for permission to publish this work.
