Polynuclear Aromatic Compounds - American Chemical Society

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Downloaded by UNIV LAVAL on March 11, 2013 | http://pubs.acs.org Publication Date: December 22, 1987 | doi: 10.1021/ba-1988-0217.ch018

Correlations Between the Spatial Configuration and Behavior of Large Polynuclear Aromatic Hydrocarbons J. C. Fetzer Chevron Research Company, Richmond, CA 94802-0627

Twenty previously unreported polycyclic aromatic hydrocarbons (PAHs) were synthesized by condensing mixtures of 1-phenalenone -type ketones. These PAHs were isolated and identified by using field -ionization mass spectrometry, spectrofluorometry, and UV-visible absorbance spectrometry. The spectrometry and chromatography of these and other PAHs were compared. Correlations between the steric strain and the spectral valley depths between maximums, or chromatographic retention, were found. Steric effects were not the only factors affecting retention; resonance stability also appeared to increase retention.

O F M A T E R I A L S for polycyclic aromatic hydrocarbons (PAHs) have become more important in the past few years. The possible etiological effects and ubiquity of these compounds make them interesting. PAHs are found in shale oil, coal liquids, petroleum, asphalt, and many other hydrocarbonbased materials (I). When these materials are combusted, fly ash, chimney soot, and engine-derived air particulates are produced. These products have higher levels of PAHs than the original materials. Discharging of PAHs into the environment has resulted in their universal occurrence in air and water. PAHs do not necessarily present a problem when they are found in a sample. A problem only occurs if certain P A H isomers are found. The mu tagenic and carcinogenic levels of a P A H are structurally dependent. Certain isomers can be very active, whereas other similar ones are not active. For ANALYSES

0065-2393/88/0217-0309$06.75/0 © 1988 American Chemical Society

In Polynuclear Aromatic Compounds; Ebert, L.; Advances in Chemistry; American Chemical Society: Washington, DC, 1987.

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example, of the five PAHs that contain four rings and have a molecular weight of 228, three (chrysene, benz[a] anthracene, and benzo[c]phenanthrene) are mutagenic, and two (naphthacene and triphen ylene) are not (2). Analytical methods for P A H analysis must be able to differentiate specific P A H isomers to accurately assess the potential health effects.


Limitations in PAH Analysis The possible use of most techniques for P A H analysis is limited by the need to differentiate isomers. Mass spectrometry, for example, yields only P A H molecular weights because only the molecular ion gives useful information. The fragment ion spectra of most isomers are almost identical; thus, they cannot be used to identify specific isomers. P A H physical properties also limit the use of other analytical methods. Gas chromatography (GC), for example, requires volatile compounds. This limitation allows G C to be used for P A H analyses of only up to six or seven rings. The higher temperatures needed to elute larger PAHs would degrade the column stationary phases and some PAHs. High-performance liquid chromatography ( H P L C ) is not limited in its application to P A H analysis. The wide variations possible in mobile and stationary phases can separate P A H mixtures with large numbers of isomers or a large number of rings. For example, several PAHs of up to 12 rings were found in a dichloromethane extract of carbon black (3). Ten compounds with a molecular weight of 424 and six compounds with a molecular weight of 448 were found by using nonaqueous reverse-phase H P L C . This analysis, however, was tedious because individual peaks had to be collected and subsequently identified by fluorescence and mass spectrometries. The difficulties inherent in off-line analysis of H P L C peaks are becoming less of an unavoidable problem because H P L C also has another advantage over other techniques. The most commonly used H P L C detectors are ideal for P A H analysis. Fluorescence excitation-emission and U V absorbance de tection are both highly sensitive and very selective for PAHs. These methods detect electronic transitions in the P A H molecules. The transition energies are determined by the P A H size and shape. Therefore, isomeric species that differ in ring configuration also differ in spectral character. The locations of absorbance maximums and the intermediate minimums, as well as the rel ative intensities of each, form a unique pattern characteristic of a particular P A H (4). The development of photodiode array detectors has made it possible to acquire on-line UV-visible absorbance spectra of an H P L C eluent. In con trast with older detectors, which monitored U V absorbance at only one wavelength, the photodiode array detectors monitor all wavelengths within



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a spectral range. Conventional detectors dispersed white light from a source and passed only one wavelength through the flow cell. Photodiode array detectors pass white light through the flow cell and then disperse it. The dispersed light is focused on a row of photosensitive elements (the photodiode array). These elements each measure the intensity at a particular wavelength. The total output of this array is a series of absorbance spectra of the H P L C eluent. Another factor limiting P A H analysis has been the few compounds that are available as standards for spectral and chromatographic comparisons. A collection of commercially available standards has consisted of a few com pounds having six or fewer rings (those having 24 or fewer ring carbons). The many other isomers of this size, or any larger ones, have not been easily obtained. The problem is being slowly overcome by new P A H syntheses. Even the products of these syntheses are of limited availability, and many other structures have not yet been produced. Thus, most studies have fo cused on the small PAHs for which standards are available; few studies have investigated the larger PAHs. Therefore, only a few PAHs have been un ambiguously identified in most samples. At best, the remaining components can be tentatively identified by matching their spectra to those i n the lit erature. This approach, however, has inherent chances of error because of different solvents and conditions. Quantitation would be impossible. Overall, analyses for PAHs have not been complete or definitive.

Previous Findings As part of a program to analyze various materials for large PAHs, analytical standards had to be acquired. One of the ways to gain standards was through the synthesis of several large PAHs (5-7). These PAHs included five previously unreported compounds. Some were peropyrenes (dibenzo[cd,lm]pery\ene and its benzo and dibenzo analogues). These com pounds resulted by condensing a mixture of 1-phenalenone and 7Hbenz[cfe]anthracen-7-one and isolating the resulting PAHs. When these compounds and many other PAHs were chromatographed to see if retention time could be used as another characterizing tool, an unexpected result was found. Even though the P A H structures were osten sibly similar (if conventional graphical representations were compared), their true spatial configurations must be very different. These conformational dif ferences resulted in vastly different spectral and chromatographic charac teristics. Some PAHs had high degrees of intramolecular steric strain because of ring locations. These PAHs assumed nonplanar configurations. Other iso mers, which only differed in the location of one ring, were not significantly strained and were more planar. Their higher degree of planarity increased



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interaction with the chromatographic stationary phase. Thus, the strained PAHs eluted much earlier than other, more planar, isomers. The most ex treme example was the nonplanar nine-ring tetrabenzo[a,cd,j,ira]perylene, which eluted before the planar four-ring pyrene. The amount of elution earlier than expected could be estimated from UV-visible spectra. The earliest eluting PAHs had the shallowest minimums between absorbance maximums. As retention increased, the valley between the absorbance maximums also increased for an isomeric series. Some PAHs even showed changes in the depth of their spectral valleys as solvent com position changed. These spectral characteristics paralleled the chromato graphic retention behaviors. This finding was assumed to indicate changes in a molecule's planarity with changes in solvent composition. The broad ening in spectral bands can be ascribed to the greater possibility of bending and twisting motions in nonplanar structures. These effects superimpose themselves on the electronic transitions (8). Three distinct types of PAHs were found: (1) PAHs that were planar in all of the solvents (eluting predictably and having very deep spectral valleys, (2) those that were nonplanar in all of the solvents (eluting much earlier than the planar PAHs of the same carbon number and having very shallow spectral valleys, and (3) those that had varying planarity that was dependent on solvent composition (always eluting in a varying fashion in comparison with the planar PAHs and having spectral valleys that changed depth). For ex ample, two nine-ring isomers eluted after the seven-ring dibenzo[cd,im]perylene at low dichloromethane concentrations in reverse-phase H P L C . At higher concentrations, they eluted before it, and at the same time their spectra had increasingly broader absorbance bands with shallower minimums. Size-exclusion chromatography indicated a relative change in their sizes. Although this variation in behavior was initially found for only a small set of peropyrene isomers, it has since been seen for many other PAHs of various structural types by using both normal- and reverse-phase H P L C (7, 9). The parallel trends in both the chromatographic and spectral behaviors were also seen. This research continues the previous work to further determine the structural features that control chromatographic retention and to see if these features can be measured by spectrometric behavior. Approximately 30 other peropyrene-type PAHs have been synthesized, isolated, and characterized. These PAHs were the products of other condensation reactions of 1-phenalenone-type ketones. 1-Phenalenone, 7f7-benz[cfe]anthracen-7-one, and a pair of five-ring ketones were used as reactants. PAHs with 9-12 rings resulted. Their chromatography and spectrometry were studied. The new results agree with previous correlations between steric effects and chro matographic and spectral behaviors.


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Experimental Details


Synthesis. The ketones used in the syntheses were 1-phenalenone (Aldrich), 7i/-benz[cte]anthracen-7-one (Phaltz and Bauer), 6#-benzo[de]pyren-6-one (W. Schmidt, Biochemical Institute for Environmental Carcinogens, Ahrensburg, West Germany), and 6H-benzo[fg]naphthacen-6-one (W. Schmidt). The ketone structures are shown in Chart I.

Ο 1-phenalenone

Ο 6iî-benzo[ de]pyren-6-one

o 7 iï-benz [ de] anthacen-7-one

Ο 6iï-benzo[/g]naphthacen-6-one

Chart I. Structures of the starting ketones used.

All solvents for extraction and chromatography were from Burdick and Jackson Laboratories (American Scientific Products), except for the solvent for spectrofluorometry, which was a purer grade of 1,2,4-trichlorobenzene that was necessary so that spectra below 400 nm could be obtained (Omnisolv from E M Scientific). All solvents were used as received. The condensation reactions used either one ketone or a mixture of two ketones (a total of 10 separate reactions). A dry zinc dust melt caused the condensation (10). A mixture of ketone, zinc dust (J. T. Baker), sodium chloride (Leslie Salt), and zinc chloride (Mallinckrodt) (1:1:1:5 by weight, respectively) was ground, thoroughly mixed, and placed in a 50-mLflat-bottomedborosilicate glass (Pyrex) reaction vessel. The total amount of ketone used in each reaction was 0.5-1.0 g. For the reactions that used two ketones, the ratio was genereally 1:1. However, the proportion in reactions using 1-phenalenone was 1:3 (1-phenalenone to the other ketone) because 1-phenalenone was much more reactive than the other ketone. This smaller pro portioning of the more reactive ketone resulted in higher yields from the cross reactions of 1-phenalenone with the other ketone rather than from the reactions of 1-phenalenone with itself. The mixture was first heated on a high-wattage hot plate to 250-275 °C for 10


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min to activate the zinc dust (ridding it of any surface zinc oxide). The temperature was raised to 330-350 °C and kept there for 30 min. The liquefied mixture was constantly stirred to make sure that the reaction was complete and that the heating was uniform. The resulting melt both condensed the ketones and reduced the re sulting intermediates to the product PAHs. The melt changed from yellow to orange or red as the PAHs were made. Extraction and Separation. After cooling, the product mixture was an amor phous solid. The product mixture and the reaction vessel were ground and placed in a 250-mL Soxhlet thimble. The mixture could not be easily broken up, removed from the vessel, and extracted separately; thus, a small reaction vessel was sacrificed for each reaction to ensure that all product PAHs were collected. The material was exhaustively extracted in a Soxhlet apparatus with 500 mL of dichloromethane. The solution fluoresced an intense green when UV light was shown on it with a hand held light (Ultraviolet Products). This method was used to monitor the extent of extraction. The extraction typically lasted 75 h, and one Soxhlet cycle occurred every 10 min. After the dichloromethane extraction, the Soxhlet thimbles were immersed in 1,2,4-trichlorobenzene at 175 °C for 1 day to ensure that all the PAHs were removed. Synchronous scanning spectrofluorometry showed only traces of PAHs that were already seen in the dichloromethane extract. Each extract was concentrated to 50 mL in a Kuderna-Danish apparatus and then diluted to 500 mL with toluene. This solution was then separated on a 75- X 1-cm i.d. glass column of basic aluminum oxide (150 g, Brockman activity grade 1, 60-100 mesh). This solution and another solution eluted from another 500 mL of toluene were combined as the first fraction. Three other fractions were collected by sequential elution with 1 L each of ethyl acetate, dichloromethane, and a 1:1 mixture of dichloromethane and methanol. The first three fractions were usually intensely fluorescent; the first fraction was blue or violet, and the other fractions were various shades of green (from aquamarine to greenish yellow, depending on the reactants). The fluorescence was used to monitor the elution of each fraction from the column. Each fraction was dried under nitrogen on a steam table and redissolved in 50 mL of dichloromethane. H P L C analyses of the fractions were then performed. The H P L C was a Du Pont 8800 quaternary solvent system with a HewlettPackard (HP) 1040A photodiode array detector (5-8). Flexible disks were used for data storage, and postrun data evaluation was performed by the detector's computer (HP 85). Samples were injected with a loop injector; 10 was used for analytical scale, and 200 μL· was used for preparative scale. Spectra were run on a spectro photometer (Perkin-Elmer Lambda 3) for static absorbance and on a spectrofluorometer (Perkin-Elmer MPF-66) for fluorescence. Field-ionization mass spectra were obtained at a resolution of 45,000 (corresponding to a 0.01-daltons error for a mass of 450). Normal-phase chromatography used an amino bonded-phase column (Du Pont Zorbax, 25- x 0.48-cm i.d., 5-μιη spherical particle size); n-hexane and dichloro methane mixtures were used as the mobile phases. Reverse-phase separations used an octadecyl bonded-phase column (Vydac 201TP5, 25- X 0.46-cm i.d. for analytical scale, and 25- X 0.94-cm i.d. for preparative scale, both with 5-μηι spherical par ticles, Separations Group); methanol and dichloromethane were the mobile phases. As in previous work (5-7, 9), the concentration of strong solvent in the mobile phase was varied from 0% to 100% for reverse-phase studies and from 0% to 30% for normal-phase studies. The peaks isolated from preparative-scale runs were further purified by separation on the analytical-scale octadecyl bonded-phase column. During a chromatographic run, spectra were acquired manually or through a software peak-detection subroutine. The software package was an upgraded version


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of the original H P software (Infometrix). It stored a spectrum when a peak maximum was detected at a preset wavelength. The second derivative of the signal was mon itored for this purpose. A spectral range of 245-600 nm, with 2-nm resolution, was stored. Retention behavior was compared with a series of planar PAHs: pyrene, benzo[g/ii]perylene, coronene, benzo[pqr]naphtho[8,l,2-Z?cii]perylene, naph thoic, l,2-a&c]eoronene, and ovalene (obtained from Aldrich or by synthesis).


Results and Discussion New PAHs Found, H P L C analyses of the fractions showed that the toluene fractions contained only the small PAHs resulting when the ketones were reduced with no condensation. The other fractions contained the de sired PAHs (these fractions also had some partially reduced intermediates). The product PAHs were somewhat separated between the fractions. Thus, individual PAHs were easier to isolate by H P L C . The last fraction also contained the unreacted and intermediate ketones. Several previously unreported PAHs were found. Chart II shows these new structures, and the box on page 317 gives their I U P A C names. Chart III shows the structures of the previously known peropyrene-type PAHs produced, and the box on page 319 also gives their names. The structural assignments of the new PAHs are not absolute. They were based only on the known reactivities of the ketones (10-13), molecular weights from field-ionization mass spectrometry, and absorbance or fluo rescence spectra. The absorbance and fluorescence spectra were used to correlate isomeric structures by applying the rules of Clar's annellation the ory (4) and Aoki's resonance structure count theory (14). These two inde pendent approaches describe how the structure of a P A H determines its absorbance-band wavelengths. Both approaches rely on spectral trends ob served for numerous benzo-analogous P A H series. Important factors include the number of fully aromatic rings that can be drawn i n the structure (a fully aromatic ring is defined as one with three it bonds) and the location and number of additional rings on various core structures. The yields for each new P A H from the reactions varied. As mentioned earlier, 1-phenalenone was the most reactive starting material, and 7 H benz[cfe]anthracen-7-one, 6H-benzo[tfe]pyren-6-one, and 6H-benzo[fg]naphthacen-6-one were progressively less reactive. Not only did the yields of individual isomers vary because of the differences in the reactivities of the ketones, but the geometry of the ketones also determined the number of possible product isomers. Thus, the reaction of two ketones, 1-phenale none and 6f/-benzo[/g]naphthacen-6-one, gave two new compounds with impurities of two previously known PAHs. The yield of compound 7 was 23%, whereas that of the isomeric compound 8 was less than 1%. Each of the reacting ketones had only one reactive site, and the possible products were h i g h l y s y m m e t r i c a l . I n contrast, the reaction using 7 H - b e n z -




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Chart II. Structures of the new PAHs produced (1-20). (The corresponding names are given in the box on page 317.)


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Names of the New Product PAHs (see Chart H) 1. Benzo[7m]phenanthro[4,5,6-abcd]perylene 2. Dibenzo[; Zm]phenanthrof4,5,6-aZ?Cii]perylene 3. Dibenzo[rs,ut#x]naphtho[2,1,8,7-Wmn]hexaphene 4. Benzo[rsi]pyreno[3,4,5-c

Polynuclear Aromatic Compounds - American Chemical Society

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