phenoxy-, dithiocarbamate, arylsulfonate-, phenate-, or cyclopentadienyl-, etc. groups may survive engine service temperatures to yield discrete organometal or metallo-organic complexes. Many of these, though highly polar, would form hydrocarbon-soluble molecules possessing strongly UV-absorbing moieties [28]. Among immediate objectives for future work should be improvements in the HPLC-GFAA speciation scheme, coupled with additional supportive compound-specific detectors operating coincidently with the GFAA detector. Of special benefit to identification of complexing ligands, particularly as a basis for comparison between new and re-refined oils, is addition of a spectrofluorimetric detector operating in either continuous or stopped-flow modes. This approach will accelerate our progress. The potential for precise characterization of many classes of chromophoric organic structures is well documented in another presentation at this conference [29]. Applications of spectrofluorimetry to speciation of trace biogenic organometallic species was shown to be highly successful at the NBS [30] in an earlier study. Future work must also exploit another feature of the HPLC-GFAA speciation method. The availability of lead-containing molecules in oils for leaching into fresh or salt water media can be assessed by use of reverse-bonded-phase HPLC columns operated with acetonitrile-water or methanol-water solvent-gradient programs [31]. By coordinating these studies with the oil-based ligand characterization studies above, a partition chemistry relating forms and rates of interchange of lead-containing bioactive molecules from oils to waters can be established. Finally, studies aimed to evaluate bioavailability of lead-containing (or other metal-containing) species in waste, re-refined, or "virgin" oils are required to provide comparisons for environmental impact/assessment from these different technological sources. In particular, the biotransformations of characterized lead-containing molecules leached into ground waters from such oils must form the focus of such research. Thereby, presence or absence of certain desirable "telltale" chemical features can be established or standardized and the results applied to redesigning additive packages or refining processes supporting production of such necessary commodities. Acknowledgements We The authors gratefully acknowledge partial financial support from the NBS Office of Air and Water Measurement and the NBS Recycled Oil Program Office. thank Mr. D. A. Becker and his associates for ultracentrifuged samples of oils; the assistance of Dr. W. P. Iverson was invaluable for microscopic characterization of the ultracentrifuged oils. We thank Mr. E. J. Parks for pointing out several valuable separation procedures for trace metal ions in oils. References [1] Golightly, D. W., and Weber, J. L., Studies of calibration standards used in Department of Defense Equipment 0.1 Analysis Program, NBS Technical Note 751 (Jan. 1973). on [2] Coyle, T. D., and Siedle, A. R., A survey of metals in oil: occurrence and significance for reuse of spent automotive lubricating oils (Proc. Conf. Measurements and Standards for Recycled Oil, NBS, Gaithersburg, Maryland, Nov. 29 and 30, 1977, NBS Special Publication 556. [3] Crosby, E. S., Rudolfs, W., and Heukelekian, H., Biological growths in petroleum refinery waste waters, Ind. Eng. Chem 46, 296 (1952). [4] Walker, J. D., and Colwell, R. R., Mercury-resistant bacteria and petroleum degradation, Appl. Microbiol. 27, 285 (1974). [ 5] [ 6] Gish, C. D., and Christensen, R. E., Cadmium, nickel, lead, and zinc in GURC Report No. 123, The Federal Register 38 [6], Part III, 9 (Jan. 10, 1973). [7] Robinson, J. W., and Wolcott, D. K., Simultaneous determination of particulate and molecular lead in the atmosphere, Environ. Letters 6, 321 (1974). [8] Roslow, E. E., Smith, W. H., and Staskawisz, B. J., Lead-containing particles on urban leaf surfaces, Environ. Sci. Technol. 11, 1019 (1977). [9] Zimdahl, R. L., and Skogerboe, R. K., Behavior of lead in soil, Environ. Sci. Technol. 11, 1202 (1977). [10] Gaskill, A., Jr., Byrd, J. T., and Shuman, M. S., Fractionation and trace metal content of a commercial humic acid, J. Environ. Sci. Health A12, 95 (1977). [11] Furr, A. K., et al., Multielement and chlorinated hydrocarbon analysis of municipal sewage sludges of American cities, Environ. Sci. Technol. 10, 685 (1976). [12] Bloomfield, C., and Pruden, G., The effects of aerobic and anaerobic_incubation on the extractabilities of heavy metals in digested sewage sludge, Environ. Pollut. 8, 217 (1975). [13] Wong, P. T. S., Chau, Y. K., and Luxon, P. L., Methylation of lead in the environment, Nature 253, 263 (1975). [14] Schmidt, U., and Huber, F., Methylation of organolead and lead (II) compounds to (CH3) Pb by microorganisms, Nature 259, 157 (1976). 4 [15] Jernelöv, A., Lann, H., Wennergren, G., Fagerström, T., Åsell, B., and Andersson, R., Analysis of methylmercury concentrations in sediments from the St. Clair System, Unpublished Report of the Swedish Water and Air Pollution Research Laboratory, Stockholm, Sewden, (in English) (1972). [16] Jensen, J., and Jernelöv, A., Biological methylation of mercury in aquatic organisms, Nature 223, 753 (1969). [17] Brinckman, F. E., and Iverson, W. P., Chemical and bacterial cycling of heavy metals in the estuarine system, Book, Marine Chemistry in the Coastal Environment, T. Church, ed., ACS Symposium Series No. 18, Amer. Chem. Soc., Washington, D.C. (1975) and reference 18 cited therein. [18] Jewett, K. L., Brinckman, F. E., and Bellama, J. M., Chemical factors influencing metal alkylation in water, Book, Marine Chemistry in the Coastal Environment. T. Church, ed., ACS Symposium Series No. 18, Amer. Chem. Soc., Washington, D.C. (1975). [19] Hofstader, R. A., Milner, O. L., and Runnels, J. H., eds., Book, Analysis of Petroleum for Trace Metals, Advances in Chemistry Series No. 156, Amer. Chem. Soc., Washington, D.C. (1976). [20] Office of Standard Reference Materials Catalog, NBS Special Publication 260, 1975-6 Ed. (June 1975). [21] Parris, G. E., Blair, W. R., and Brinckman, F. E., Chemical and physical considerations in the use of atomic absorption detectors coupled with a gas chromatograph for determination of trace organometallic gases, Anal. Chem 49, 378 (1977). [22] Chau, Y. K., Wong, P. T. S., and Saitoh, H., Determination of tetraalkyl lead compounds in the atmosphere, J. Chromatogr. Sci. 14, 162 (1976) and references 1-4 cited therein. [23] Blair, W., Iverson, W. P., and Brinckman, F. E., Application of a gas chromatograph-atomic absorption detection system to a survey of mercury transformations by Chesapeake Bay microorganisms, Chemosphere 3, 167 (1974). [24] Bauman, F., and Hadden, N., Eds., Basic Liquid Chromatography (Varian Aerograph, Palo Alto, California, 1971). [25] [26] [27] [28] [29] [30] [31] Brinckman, F. E., Blair, W. R., Jewett, K. L., and Iverson, W. P., Application Turina, N., and Turina, S., Trace analysis of lead in oil by TLC, Chromatographia 10, 97 (1977). Issaq, H. J., and Barr, E. W., Combined thin-layer chromatography/flameless Sandell, E. B., Colorimetric Determination of Traces of Metals, 3rd Ed. May, W. E., and Brown, J. M., The analysis of some residual fuel and waste Komae, H., and Hayashi, N., Separation of essential oils by liquid chromatography, THE ANALYSIS OF SOME RESIDUAL FUEL AND WASTE LUBRICATING OILS BY A HIGH-PERFORMANCE LIQUID CHROMATOGRAPHIC PROCEDURE W. E. May and J. M. Brown Organic Analytical Research Division Introduction In recent years there has been increasing concern about the possible harmful effects caused by some of the organic compounds that are found in the environment. The polynuclear aromatic hydrocarbons (PAH) are a class of pollutants that are of great concern because some of them, such as benzo(a)pyrene, benz (a) anthracene, and dibenz (ah) anthracene, have been shown to possess carcinogenic [1-3] and mutagenic [4,5] properties. PAH have been found in air, food, and water from such sources as engine exhaust; the burning of fossil fuels; tobacco smoke; and natural sources, such as petroleum-based oils. Most schemes for the analysis of PAH involve an extraction, separation (sample cleanup using either open-column or thin-layer chromatography), and qualitative and quantitative analyses steps. Most early investigators used ultraviolet absorption (UV) or fluorescence spectroscopy for quantitative analysis of such extracts [6]. Later, gas chromatography (GC) became the analytical technique of choice because it allowed further isolation of the analyte and gave some qualitative information in the form of the chromatographic retention time. Recently, the use of capillary GC has become very popular and has been used in the analysis of PAH in engine exhausts [17], anthracene oil [8], cigarette smoke [9], soot samples [10], and sediments [11]. Janini, et al. [12] have discussed some of the difficulties associated with separating complex mixtures of PAH by GC and suggested the use of a nematic liquid crystal column for such analyses. Such columns have high bleed rates and relatively short lifetimes. The high bleed rates also make their use in gas chromatography-mass spectrometry (GC-MS) impractical. GC-MS is generally recognized as being the most powerful tool available to the analyst for identifying trace components in complex mixtures. GC-MS has been used in the analysis of complex PAH mixtures [13-15] but does have certain limitations. Isomeric PAH are not readily distinguishable from their mass spectra alone and are often not resolved chromatographically. For example, 1,2-benzanthracene, chrysene, and naphthacene give essentially the same mass spectrum and are difficult to separate by GC. High-performance liquid chromatography (HPLC) offers the analyst another powerful tool for use in the analysis of PAH. Larger sample sizes can be accommodated, and less sample pretreatment is necessary. Columns are available that provide rapid and efficient resolution of complex mixtures of PAH. However, the sensitivity and selectivity of the detection systems that may be used in conjunction with HPLC are the major advantages afforded by the technique. Christensen and May [16] have recently shown that ultraviolet and fluorescence detectors are comparably sensitive and more selective for PAH than the mass spectrometer. In this presentation we will describe a HPLC method that we have developed for the analysis of benzo(a)pyrene in some waste lubricating and residual fuel oils. 1 Underlined numbers in brackets indicate the literature references at the end of this paper. Experimental Instrumentation. The liquid chromatographic unit used in this work consisted of two Model 6000A pumps and a solvent programmer (Waters Associates, Milford, MA); a Model 7120 sample injector (Rheodyne, Berkeley, CA); a Model 440, 254 nm filter photometric detector (Waters Associates, Milford, MA); and a Mark I spectrofluorometer, equipped with a 10 μL flow cell (Farrand Optical Company, Inc., Valhalla, NY). The columns used were μBondapa, NH2 and μBondapak C18. Data were recorded using a strip chart recorder. Methodology. The HPLC method described here utilizes two chromatographic columns for the isolation and separation of individual PAH. The data presented in table 1 show that chromatography on uBondapak NH2 provides a separation of PAH that is based on ring size. Alkylation has little effect on retention. On the other hand, chromatography on μBondapak C18 provides a separation that is related to the aqueous solubilities of the respective PAH. Parent PAH can be separated from both their alkylated homologs and their structural isomers. Efficient isolation of individual PAH from complex mixtures may be achieved by using both of these columns in an integrated procedure. The concentration of benzo(a)pyrene in some residual fuel and recycled oil samples was measured using the procedure described below. 1. The density of each sample was determined by weighing 10 mL of oil on a Mettler balance. The volume was measured in a class "A" volumetric flask. 2. Dilution and filtration of the samples were accomplished by filling a 1.33 mL stainless steel sample loop with the oil to be measured, and pumping the contents of the loop through a 2 um filter into a 10 mL volumetric flask. 3. Isolation of the five condensed ring PAH fraction was effected by injecting 200 μL of the diluted filtrate onto a 300 x 12 mm uBondapak NH2 column. The benzo(a)pyrene fraction was collected in a volume of 20 mL (see figure 1). 4. This 20 mL was reduced to a volume of between 200 and 500 uL using N2 purge at room temperature. The volume measurement was made by means of a Hamilton syringe, calibrated to the nearest 5 μL. 5. Separation of benzo(a)pyrene from other five condensed ring PAH and quantification was done by injecting 23 μL of the fractionated oil on a 300 x 4 mm Bondapak C18 column. Further selectivity was obtained by monitoring the chromatographic effluent fluorimetrically, with excitation and emission wavelengths optimized for detection of benzo(a)pyrene (ex-295nm, em-400nm). Benzo(a)pyrene was identified in each oil by chromatographic retention volume and fluorescence emission spectrus, as shown in figure 2. Results. The concentration of benzo(a)pyrene in each sample was calculated using the following equation: K3 = Unfractionated dilution factor; the ratio of the volume to which Fractionated dilution factor; the ratio of the volume to which = Fluorescence response factor for benzo(a)pyrene in mm/ug. |