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Discriminatory Analysis of Marine-Engine Lubricants Using Hopane Biomarkers Masato Taki*,† and Masatoshi Nagai‡ Coast Guard Research Center, Japan Coast Guard, 1156, Tachikawa-city, Tokyo 190-0015, Japan, and Graduate School of Bio-applications and Systems Engineering, Tokyo UniVersity of Agriculture and Technology, 2-24, Nakamachi, Koganei-shi, Tokyo 184-8588, Japan
Twenty marine-engine lubricants from five Japanese oil companies were discriminated on the basis on a hopane analysis using a gas chromatograph-mass spectrometer (GC-MS). The hopane ratios of 18R-22,29,30trisnorneohopane (1) to 17R-22,29,30-trisnorhopane (2) and of 17R-21β(H)-norhopane (3) to 17R-21β(H)hopane (5) were determined. Each lubricant can be discriminated by combining the figures represented by the 1 to 2 and 3 to 5 ratios and principal component analysis from 1 through (22R)-17R-21β(H)-29pentakishomohopane (15). Oleanane (4), one of the biomarkers inherent in the crude oils from Southeast Asia, has been detected in the lubricants of eight out of 20 types, and 4 represents a significant feature for the discrimination. The discrimination of the lubricants in the spilled oils was also investigated. Introduction Approximately half of lubricants used in engine crankcases is released into the environment, both through engine exhaust and through leakage and spillage. Ships sometimes illegally dump overboard mixtures of their lubricants, fuel oil, and water which are accumulated in ship bottoms. The released oils cause extensive damage to marine life, natural resources, and human health. To prevent such pollution, contaminants and the source of the spilled oils need to be analyzed and identified in a short time as well as the spilled oils being cleaned up. The lubricants spilled in the sea are usually identified based on their hydrocarbon composition using a chromatograph flame ionization detector (GC-FID)1 and a chromatograph combined with a mass spectrometer (GC-MS) scan mode analysis,1-3 and the distribution of their molecular weights is determined by gel permeation chromatography (GPC).4,5 The GC-FID and GC-MS scan mode analyses can detect with broad and unresolved peaks, but they can hardly classify several specific types of compounds, especially hydrocarbons contained in the lubricants without long accumulated results. The GPC has similar concerns regarding the distribution analysis of the molecular weights of the lubricants. Lubricants are manufactured during the purification process of crude oils, such as solvent refining, hydrorefining, and solvent dewaxing after the distillation of the crude oils under normal or reduced pressure during the separation. Furthermore, a manufacturer adds its own additives to the lubricant to create a unique blend. For that reason, lubricants certainly contain substances derived from crude oil. For the lubricants spilled in the sea, these conventional methods analyze the lubricants in order to compare the isolated additives with authentic samples,6-8 but they require a lot of time to extract the additives. Therefore, these methods are unsuitable for the required prompt analysis of spilled oils in the sea. Furthermore, it is extremely difficult to extract and analyze small amounts of additives floating on the sea. In addition, there are only a few studies on the separation and analysis of individual pure additives or those contained in the lubricants. Crude oils have their origins from the organic debris of plankton, algae, bacteria, and plants that inhabited the * To whom correspondence should be addressed. Tel.: +81-42-5265634. Fax +81-42-526-5636. E-mail:
[email protected]. † Japan Coast Guard. ‡ Tokyo University of Agriculture and Technology.
region in ancient times. Carbon skeletons can be related to a precursor molecule from a specific type of organism, which is called a biomarker. A biomarker is an analytical method superior to refractory substances and compounds resistant to biodegradability which does not change by evaporation or organisms. A discriminative analysis based on the biomarker is required for the discriminative analysis of lubricants spilled in the sea. The hopane homologous series are derived from their components in living organisms.11 Hopanes of the biomarker, contained in lubricants, characteristically have a m/z 191 fragment ion, and are a suitable method for the GC-MS selected ion monitoring mode. This technique identifies the lubricants faster and easier than conventional methods. In a previous paper,9 the analysis of the hopane, norhopane, and oleanane series in crude oil made it possible to discriminate 41 crude oils and identify their oil stratum wells in a region. These biomarkers are also contained in the lubricants. The analysis of the crude oils applies to the discrimination of the lubricants. The study focused on the rapid determination of lubricants without isolation and knowledge of special additives added during their production. This new analyzing method is original and has not been previously reported. Furthermore, one example is reported for the identification of spilled and discharged oil (bilge) from a ship. Experimental Section Lubricants and Reagents. Twenty types of marine-engine lubricants for ships were purchased from five domestic producers. The 20 types of lubricants used for the experiments are shown in Table 1. In the code, such as A1 and B4, the letters A-E are designated for each company, and the following numbers from 1 to 4 are the reference numbers. One lubricant, which had been used for 15 638 h, was obtained from a Japan Coast Guard patrol ship. The five companies are large domestic distributors of lubricants that possess their own oil refineries. The 20 types of lubricants were the standard ones from the five companies along with four types of motor oils per company based on Japanese Industrial Standards (JIS). The authentic standards of 1, 2, 3, 4, 5, and 17β-21β(H)-hopane were purchased from Chiron, Norway. The solvents, such as hexane, were purchased from Wako Pure Chemical, Ltd. (Tokyo, Japan).
10.1021/ie100904b 2010 American Chemical Society Published on Web 10/26/2010
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Table 1. Classification of the Lubricants kinematic viscosity (mm2 /s) company abbreviation
lubricant
40 °C
100 °C
flash point (°C)
pour point (°C)
base number (mg of KOH/g)
A
A1 A2 A3 A4 B1 B2 B3 B4 C1 C2 C3 C4 D1 D2 D3 D4 E1 E2 E3 E4
146 146 136 92 103 142 105 152 124 130 108 95 145 110 130 110 106 140 104 140
14.4 14.4 14.3 11.2 11.5 14.1 11.5 14.7 13.9 13.8 14.6 11.2 14.2 12.0 13.5 12.0 12.0 14.5 11.8 14.4
258 275 278 246 264 278 252 266 254 254 232 242 230 230 230 220 260 260 260 262
-9 -12 -18 -21 -18 -15 -13 -13 -13 -13 -35 -13 -10 -10 -10 -10 -18 -18 -20 -20
15 30 10 10 12 12 11 11 20 10 17 10 10 30 30 10 11 11 20 20
B
C
D
E
Preparation and Analysis of Hopanes in Lubricants by GC-MS. The marine-engine lubricants are discriminated by GC-MS based on biomarkers, such as 18R-22,29,30-trisnorneohopane (1), 17R-22,29,30-trisnorhopane (2), 17R-21β(H)norhopane (3), oleananes (18R- and 18β-oleananes) (4), 17R21β(H)-hopane (5), and (22S)-17R-21β(H)-29-homohopane (6) to (22R)-17R-21β(H)-29-pentakishomohopane (15). Each lubricant sample (0.100 g) was dissolved in 20 mL of hexane and then separated at 7000 rpm for 10 min using a centrifugal separator. The same separation procedure for the hexane-soluble material and insoluble residue in the lubricants was used instead of the method of extraction and fractionation. The soluble materials contained 1 mL of a hexane solution and 1 mL of 17β-21β(H)-hopane, which were added as the internal standard substances for the analytical sample. The test lubricant samples contain no or little 17β-21β(H)-hopane. The internal standard material has been reported in various articles including that of
Wang et al.1 and Zakaria10 et al. As lubricants can be analyzed with good accuracy using 17β-21β(H)-hopane, it was used as the internal standard substance. The 20 types of marine-engine lubricants and the used lubricant were analyzed using Agilent Technologies equipment (Models HP5972 and HP5890) along with a J&W Scientific Durabond fused-silica capillary DB-1 column having an inner diameter (i.d.) of 0.25 mm, a film thickness of 0.25 µm, and a length of 60 m with a helium carrier gas (1.1 mL/min). The GC-MS operating conditions in the electron impact (EI) mode included an ionization potential of 70 eV with the ion source at 180 °C and the electron multiplier voltage at ∼1.2 kV. The sample (1 µL) was injected into the injection port maintained at 320 °C in the splitless mode, followed by a purge for 1 min after the injection. The column temperature was increased from 200 to 310 °C at the rate of 4 °C/min. The selected ion monitoring method was applied after a delay of 7 min, and the hopanes were quantified at m/z )
Figure 1. m/z 191 mass chromatograms of lubricants (A1, B1, C1, and D1). The notations 1-15 are illustrated in Figure 2.
Ind. Eng. Chem. Res., Vol. 49, No. 23, 2010 Table 2. Compounds from GC-MS Analysis of Hopane Homology notation
compounds
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15
18R-22,29,30-trisnorneohopane 17R-22,29,30-trisnorhopane 17R-21β(H)-norhopane (C29 hopane) oleananes 17R-21β(H)-hopane (C30 hopane) (22S)-17R-21β(H)-29-homohopane (22R)-17R-21β(H)-29-homohopane (22S)-17R-21β(H)-29-bishomohopane (22R)-17R-21β(H)-29-bishomohopane (22S)-17R-21β(H)-29-trishomohopane (22R)-17R-21β(H)-29-trishomohopane (22S)-17R-21β(H)-29-tetrakishomohopane (22R)-17R-21β(H)-29-tetrakishomohopane (22S)-17R-21β(H)-29-pentakishomohopane (22R)-17R-21β(H)-29-pentakishomohopane
191. The peaks were identified by comparison of their retention times to those for the standards and their mass spectra, which were obtained from a different GC-MS run in the scan mode, with the retention times reported in the literature.10-13 The measuring limit of 17R-21β(H)-hopane was 1 mg/L for the scan mode and 0.1 mg/L for the selected ion monitoring (SIM) mode. Results and Discussion Hopanes in the Lubricant Samples. Four typical GC-MS chromatograms of the hopane homologous series (m/z ) 191) distributed in the 20 lubricant samples are shown in Figure 1. The names and structures of the hopane compounds (1, 2 3, 4,
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5, 6, 7, and 8-15), i.e., the pentacyclic triterpanes of the hopane series, are shown in Table 2 and Figure 2, respectively. Compounds 3 and 5 were detected in large amounts for 1-15 in the lubricants, while 1 and 2 were hardly observed. Figure 3 shows the relationship between the peak ratio 1/2 and the peak ratio 3/5 based on the average of five values analyzed by the SIM mode. Regarding A1, B1, C1, and E1, A1 and C1 contained compound 1 at a greater peak intensity than 2, while B1 and E1 contained 1 in a lower amount than 2. For compounds 3 and 5, the peak intensity of 5 was greater than that of 3 in A1, but it was lower in C1 and E1, and for B1, 5 had the same peak intensity as 3. Compound 4 was only contained in E1. The peak intensities from 6-15 exhibited a similar decreasing tendency for A1, B1, C1, and E1. E1 exhibited the highest decreasing trend, while A1 had the lowest. Thus, the discriminatory analysis of the 20 lubricants should be performed on the basis of the correlative combination of the various hopanes. Grouping All Lubricants Based on Hopane Analysis. In a previous paper,9 based on the analysis of the hopanes (1/2 and 3/5 ratios) in 41 crude oils, they were divided into three groups: the northern Middle East crude oils (classification a), the eastern part among the Middle East crude oils (Zagros suture zone and Oman) (classification b), and the southern part among the Middle East crude oils (classification c). The lubricants were made of crude oils and then determined using these combination ratios of the hopanes. The relationship between the 1/2 and 3/5 ratios is shown in Figure 3. The lubricants of A1-A4 exhibited the highest ratio of 1/2 (1.30-1.75) and the lowest ratio of 3/5
Figure 2. Structures of the hopane compounds: 1, 18R-22,29,30-trisnorneohopane; 2, 17R-22,29,30-trisnorhopane; 3, 17R-21β(H)-norhopane (C29 hopane); 4, oleananes (18R-oleanane, 18β-oleanane); 5, 17R-21β(H)-hopane (C30 hopane); 6, (22S)-17R-21β(H)-29-homohopane; 7, (22R)-17R-21β(H)-29-homohopane; 8-15, (22S)-17R-21β(H)-29-bishomohopane to (22R)-17R-21β(H)-29-pentakishomohopane.
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Figure 3. Diagram of 1/2 ratio plotted versus 3/5 ratio: A, company A (A1-A4); B, company B (B1-B4); C, company C (C1-C4); D, company D (D1-D4); E, company E (E1-E4).
Figure 4. Principal component analysis (PCA) of 1-15: B1-B4, C1-C4, D1-D4, E1-E4, spilled oil, and bilge oil.
(0.63-0.75). C1-C4 were in the middle range of the 1/2 ratio (1.00-1.17) with the highest 3/5 ratio (1.12-1.17). The 1/2 ratios for the lubricants of B, D, and E were significantly lower than those of A1-A4 and C1-C4. Based on these results, the 20 types of lubricants were classified on the basis of the ratios of 1/2 and 3/5 for each company. This may be caused by the fact that each manufacturer produces their lubricants from different crude oils. A1-A4 can be discriminated by the relationship between the 1/2 and 3/5 ratios. The lubricants of B1-B4, C1-C4, D1-D4, and E1-E4 cannot be clearly discriminated by the ratios of 1/2 and 3/5, but an additional factor using the other groups of hopanes, 1-15, leads to confirming the discrimination of the individual lubricants of B, C, D, and E. Figure 4 shows the principal component analysis (PCA) from 1-15. Regarding the rectus (22R) and sinister (22S) hopanes, Peters et al.11 reported that the isomerization at C-22 in the C31-C35 17R-hopanes occurs earlier than in many biomarker reactions used to assess the thermal maturity of oil and bitumen. During the biodegradation process of sterane, the 20S/(20R + 20S) ratio was used as the maturity index, because 20R is reported to originate from living organisms and is more selectively consumed than 20S. 22R was also more selectively consumed than 22S for hopane. For sterane, only the R
configuration at C-20 occurred in living organisms, and this was gradually converted during burial maturation to a mixture of the R and S sterane configuration. Suau et al.14 found that the homohopane degradation favored the formation of the R epimer over that of the S epimer. Furthermore, the sinister (22S) hopane compounds, 6, 8, 10, 12, and 14, likely exhibited a higher peak intensity than the corresponding rectus (22R) hopane compounds, 7, 9, 11, 13, and 15. Figure 4 shows the results of PCA from 1 to 15. The PCA results allowed the discrimination of B1 from B4, C1 from C4, D1 from D4, and E1 from E4. Five groups were classified: C1-C4; D1-D4; B1 and B4; B2, E1, E2, and E4; and B3 and E3. Lubricants Derived from Crude Oils in Regional Producing Area. The origin of the marine lubricants was investigated in order to determine from which region the crude oils originated. The result obtained by analyzing the hopanes (ratios of 1/2 and 3/5) in the crude oils is shown in Figure 5. In a previous paper,9 this analysis of the hopanes was separated into the following four groups: (1) a group of Southeast Asia, China, Gabon, and Russia crude oils with less than a 3/5 ratio of 0.8, and three groups of the (2) eastern, (3) northern, and (4) southern Middle East crude oils as shown in Figure 5. The separation of the lubricants of A-E was not completely determined by the
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Figure 5. Diagram of 1/2 ratio versus 3/5 ratio: A, company A (A1-A4); B, company B (B1-B4); C, company C (C1-C4); D, company D (D1-D4); E, company E (E1-E4). E4 was used for 15 638 h. The northern, eastern, and southern Middle East crude oils are divided by the lines. The spilled and bilge oils are denoted by the symbols + and 2, respectively.
derived crude oils. It is possible that the lubricants of A1-A4 are Southeast Asia, China, Gabon, and Russia crude oils based on the 3/5 ratio (0.63-0.75, 0.8, and less). Compound 4 was detected in A2 and A3, which definitely proved its origin as Southeast Asian. The 3/5 ratios for B-E were greater than 0.8, indicating that the crude oils were from the Middle East and Mexico. Furthermore, the 3/5 ratio of B1-B4 was 0.87-0.90, but the 3/5 ratio of E1-E4 was 0.93-0.95. This result showed that the lubricants of B1-B4 and E1-E4 were derived from the eastern or a blend of eastern and southern Middle East crude oils. Since the lubricants of B2, B3, and E1-E4 contained 4, the lubricants of B2, B3, and E1-E4 had two possible crude oil origins. One was identified as a blend of eastern Middle East and Southeast Asian crude oils because of the detection of 4 in the Southeast Asian crude oil. The other possibility is the Iranian light (IR-IL) or Iranian heavy (IR-IH) which contains 4, although it is a rare case for the eastern Middle East crude oils. C1-C4 were derived from the crude oils produced from the southern or a blend of southern and northern Middle East crude oils based on a 1/2 ratio of 1.00-1.17 and a 3/5 ratio of 1.12-1.17. The lubricants D1-D4 were derived from the crude oil produced from the northern Middle East crude oils based on the following data: 0.48-0.57 for the peak 1/2 ratio and 1.06-1.08 for the 3/5 ratio. Consequently, the lubricants can be determined based on the type of crude oil by adopting the ratios of 1/2 and 3/5. The analysis of the hopanes allows one to determine the origin of the crude oils from plants or bacteria or by the degree of maturity. In the crude oil, 18R-22,29,30trisnorneohopane and 17R-22,29,30-trisnorhopane are not affected by biodegradation, but 18R-22,29,30-trisnorneohopane showed maturity with a greater thermal stability than 17R22,29,30-trisnorhopane.11,15 18R-22,29,30-Trisnorneohopane originated from green algae or bacteria.11 17R-21β(H)-Norhopane was contained in large amounts in the southern Middle East crude oils. 17R-21β(H)-Norhopane originated from bacteria residues in the evaporate and carbonate source rocks before the Cretaceous age. Also, oleanane is of plant origin which can be contained in large quantities in the Southeast Asia crude oils. 17R-21β(H)-Hopane originated from bacteria residues in mudstone rock.16 In the case of the crude oil from Southeast Asia, oleanane, a plant-derived biomarker, was detected. Oleanane, one of biomarkers inherent in the crude oil of Southeast Asia,
Figure 6. m/z 191 mass chromatograms of (A) spilled and (B) bilge oils.
has been detected in eight out of the 20 lubricants: A2, A3, B2, B3, and E1-E4. The eight marine-engine lubricants are derived from the Southeast Asia crude oils. As the used lubricants contained the hopanes, they can be identified on the basis of the hopane analysis. Spilled Oil. In October 2008, spilled oil was found near a certain city on the coast of the Sea of Japan. The chromatogram of the collected spilled oil on the sea is shown in Figure 6, and that of the discharged oil (bilge) collected from the bottom of a ship anchored nearby is also shown in Figure 6. These oils were analyzed on the basis of the hopane distribution. The hopane compositions of the spilled oil were similar to those of the discharged oil (bilge) in Figures 4 and 5. Both the 1/2 and 3/5 ratios and the PCA from 1-15 in the spilled oil were proved to conform to those of the discharged oil (bilge). The spilled
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oils were identified on the basis of the analysis of the homologous series of hopanes. This analytical method using the hopanes is sufficient for the determination of the lubricants, the lubricant-containing fuel oils, crude oils, and bunker C fuels (C-grade heavy oil). Conclusions Twenty types of lubricants were classified on the basis of the 1/2 and 3/5 ratios for each manufacturer. Each lubricant was identified by the combination of the ratios of 1/2 and 3/5. The PCA from 1-15 was performed. The analysis of the hopanes in the lubricant resulted in a specification for the corresponding crude oils. Compound 4 was characteristically contained in the crude oil from Southeast Asia and detected in eight of the 20 lubricants. This hopane analysis is an important index for the discrimination of lubricants and estimates the crude oil used for producing the lubricants. The hopanes can be detected in the used lubricants. This technique is useful for determining the origin of the lubricants, the crude oils, and bunker C fuels (C-grade heavy oil) which are illegally dumped into the sea from ships. This is a superior technique for the identification of the lubricants. Acknowledgment The authors acknowledge the many helpful suggestions of our co-workers at the Japan Coast Guard Research Center Chemical Analysis Division. Literature Cited (1) Wang, Z.; Fingas, M.; Lambert, P.; Zeng, G.; Yang, C.; Hollebone, B. Characterization and identification of the Detroit River mystery oil spill (2002). J. Chromatogr. 2004, A1038, 201–214. (2) No, M. H.; Kim, E.; Lee, J. S.; Jung, H. Hydrocarbon Compound Type Analysis of Lube Base Oil by GC-MSD: Advantages on Replacement of the AGHIS Magnetic Sector Type Mass Spectrometer. Energy Fuels 2007, 21, 3341–3345. (3) Hiltz, J. A.; Haggett, R. D. Identification and Quantitative Analysis of Contaminants in Lubricating and Hydraulic Fluids by Gas Chromatography/Mass Spectrometry. Lubr. Eng. 1990, 11 (47), 945–955.
(4) Philip, C. V.; Bullin, J. A.; Anthony, R. G. GPC Characterization for Assessing Compatibility Problems with Heavy Fuel Oils. Fuel Process. Technol. 1984, 9, 189–201. (5) Higashi, K.; Hagiwara, K. Characterization of crude oils by high speed gel permeation chromatography; Study of stationary phase. Bunseki Kagaku 1978, 27, 768–772. (6) Barnes, A. M.; Bartle, K. D.; Christopher, S.; Heathcote, C. Use of Capillary Supercritical Fluid Chromatography in the Analysis of Zinc Dialkyldithiophosphates. J. High Resolut. Chromatogr. 2000, 23, 389–392. (7) Habboush, A. E.; Farroha, S. M,; Khalaf, H. I. Extraction-gas chromatographic method for the determination of organophosphorus compounds as lubricating oil additives. J. Chromatogr. 1995, A696, 257– 263. (8) Rinco´n, J.; Can˜izares, P.; Garcia, M. T. Ind. Eng. Chem. Res. 2005, 44, 4373–4379. (9) Taki, M.; Asahara, T.; Mandai, Y.; Uno, T.; Nagai, M. Discriminatory Analysis of Crude Oils Using Biomakers. Energy Fuels 2009, 23, 5003– 5011. (10) Zakaria, M. P.; Horinouchi, A.; Tsutsumi, S.; Takada, H.; Tanabe, S.; Ismail, A. Oil Pollution in the Straits of Malacca, Malaysia: Application of Molecular Markers for Source Identification. EnViron. Sci. Technol. 2000, 34, 1189–1196. (11) Peters, K. E.; Walters, C. C.; Moldowan, J. M. The Biomaker Guide; Cambridge University Press: New York, 2005; pp 3-616. (12) Subroto, E. A.; Alexander, R.; Kagi, R. I. 30-Norhopanes: their occurrence in sediments and crude oils. Chem. Geol. 1991, 93, 179–192. (13) Chakhmakhchev, A.; Suzuki, M.; Waseda, A.; Takayama, K. Geochemical characteristics of Tertiary oils derived from siliceous sources in Japan, Russia and USA. Org. Geochem. 1997, 27, 523–536. (14) Suau, R. F.; Bost, F. D.; Mcdonald, T.; Morris, P. J. Aerobic Biodegradation of Hopanes and other Biomarkers by Crude Oil-Degrading Enrichment Cultures. EnViron. Sci. Technol. 2002, 36, 1189–1196. (15) Damste´, J. S. S.; Kuypers, M. M. M.; Pancost, R. D.; Schouten, S. The carbon isotopic response of algae, (cyano)bacteria, archaea and higher plants to the late Cenomanian perturbation of the global carbon cycle: Insights from biomarkers in black shales from the Cape Verde Basin (DSDP Site 367). Org. Geochem. 2008, 39, 1703–1718. (16) Waseda, A.; Nishita, H. Geochemical characteristics terrigenousand marine-sources oils in Hokkaido, Japan. Org. Geochem. 1998, 28, 27– 41.
ReceiVed for reView April 17, 2010 ReVised manuscript receiVed October 10, 2010 Accepted October 11, 2010 IE100904B
