Effect of Biodegradation and Water Washing on Crude Oil Composition

Transcription

1 Chapter 4 Effect of Biodegradation and Water Washing on Crude Oil Composition Susan E. Palmer Amoco Production Company Tulsa, Oklahoma, U.S.A. INTRODUCTION The study of crude oil geochemistry becomes difficult when crude oils are altered by microbial action (biodegradation) and/or water washing. These processes can alter parameters used to compare oils when determining genetic relationships (oil-oil correlation), depositional environments, and time of oil generation (i.e., thermal maturity of the source rock at the time of oil generation and expulsion). Much of this knowledge comes from petroleum geochemists who have been documenting case histories of such occurrences through the years (e.g., Winters and Williams, 1969; Bailey et al., 1973a, b; Rubinstein et al., 1977; Connan et al., 1975, 1980; Seifert and Moldowan, 1979; Rullkotter and Wendish, 1982; Volkman et al., 1983; Momper and Williams, 1984; Palmer, 1984; Williams etal., 1986) In addition, microbiologists and petroleum geochemists have studied the action of bacteria on petroleum in the laboratory (e.g., McKenna, 1972; Horowitz et al., 1975; Jobson et al., 1979; Connan, 1981; Goodwin et al., 1983). Some workers have isolated products of bacterial metabolism of crude oils or classes of hydrocarbons (e.g., Gibson, 1976; Higgens and Gilbert, 1978; Cripps and Watkinson, 1978; Cain, 1980; Mackenzie et al., 1983). Connan (1984) illustrates the metabolic products recognized by some of these workers and others. Products of aerobic degradation are most often organic acids and CQ2. Anaerobic bacteria can live on the metabolites of the aerobes but do not grow on hydrocarbons. Thus, the impact of anaerobic bacteria on oil biodegradation is only slight. Alteration of crude oil by water washing has been indirectly studied in the laboratory through determination of water solubilities of individual hydrocarbons and mixtures of several hydrocarbons and by studying compositional changes of whole crude oils (e.g., McAuliffe, 1966, 1980; Bailey et al., 1973b, Price, 1976; Eganhouse and Calder, 1976; May et al., 1978a, b; Lafargue and Barker, 1988). Extensive review articles covering work done prior to 1985 on biodegradation and water washing have been prepared by Milner et al. (1977) and Connan (1984). These two reviews give an overview of the effects of biodegradation and water washing gleaned from the many exemplary papers found in the literature. Also, Lafargue and Barker (1988) have reviewed and discussed data obtained from laboratory water washing experiments and present their own observations and conclusions. The purpose of this chapter is to compile and present the results of these studies and review articles in an abbreviated form to aid the explorationist in understanding the effects of biodegradation and water washing. In this regard, the reader is directed to the literature for details of individual studies. Results of more recent studies ( ) are also included here. Examples of parameters and hydrocarbon distributions demonstrating the effects of biodegradation and water washing are given to aid the reader in recognizing altered oils. Because many geochemical laboratory groups have developed their own ways to portray data, some of these techniques will be referenced in this discussion. Organic geochemical parameters and types of hydrocarbon classes referred to here are defined in the Glossary at the back of this volume. GEOLOGICAL CONSTRAINTS AND PHYSICOCHEMICAL CONDITIONS FOR BIODEGRADATION AND WATER WASHING The processes of microbial degradation and water washing of crude oils occur when certain conditions are met. Milner et al. (1977) and Connan (1984) have outlined the requirements for both processes in their review 47

2 48 Palmer articles. Their findings are summarized here. Biodegradation occurs in surface seeps and in relatively shallow reservoirs, e.g., ft ( m) or less. Although anaerobic bacteria can survive on partially degraded oils, both case histories and microbiological studies show that aerobic bacteria are the major agents of crude oil degradation. Aerobic bacteria can grow in relatively cool reservoirs (i.e., below 80 C, or 176 F) that are invaded by oxygen-charged waters. In addition to dissolved oxygen, nutrients such as nitrate and phosphate must be present and the salinity of the water must be less than %o. Also, unless special cases such as oxygenated microenvironments exist, the amount of H2S in the oil must be very low, as it is toxic to aerobic bacteria. Thus, cool, shallow reservoirs that are flushed by oxygenated, nutrient-rich fresh water can be expected to contain oil that is being actively biodegraded. However, biodegraded oils are also present in deeper reservoirs. In areas where tectonic activity causes subsidence of reservoirs, biodegraded oils are found preserved far below the arbitrary 6000-ft cutoff for bacterial activity. Thus, cases where biodegraded oils occur in reservoirs of, e.g., ,000 feet ( m) have been reported. Also, with regard to maximum depth of reservoirs and ongoing degradation, lower thermal gradients can permit deeper occurrences. Connan (1984) points out that aerobic bacteria degrade oil at the oil-water interface. In such cases, the lower part of an oil accumulation will be degraded rather than the upper portions, unless more than one oil-water contact is present. Some reservoirs have multiple oil-water contacts and different hydrological regimes that could lead to a complex and perhaps confusing array of degraded and undegraded oils. Lafargue and Barker (1988) mention that hydrodynamically tilted oil-water contacts are indicators of actively flowing waters and delineate areas where degradation is occurring. Physical and chemical processes other than biodegradation and water washing (e.g., fractionation of light and heavy ends during migration and in-reservoir maturation) can add to the difficulty of understanding the transformation of oil from its initial state to the time it is recovered in a discovery well or surface seep. Some of these other processes could be mistaken as biodegradation or water washing. Thus, a good understanding of the postgeneration history of an oil is of major importance for establishing cause and effect relationships. Because water is a necessary ingredient for biodegradation, the process of water washing generally accompanies biodegradation. In spite of many studies that have attempted to isolate the effects of water washing from microbial alteration, questions concerning the actual cause of alteration of a crude oil's geochemistry, especially specific parameters, are still open for discussion. Conditions favorable for water washing exist during oil migration if oil is passing through a water-wet carrier bed and reservoir system. However, Lafargue and Barker (1988) have suggested that water washing during migration must be minimal because highly water soluble molecules, namely, benzene and toluene, are present in most oils. They suggest that most water washing occurs after accumulation where, given the proper conditions, biodegradation is also occurring. Water washing can, however, take place outside the temperature, oxygen, and salinity constraints of biodegradation. Price (1976) and Lafargue and Barker (1988) have noted that the solubility of crude oil components increases markedly at higher temperatures. These results demonstrate that water washing can occur in zones where microbial activity is precluded by high temperature. The work of Price (1976) shows that high salinities (over 270%o) cause exsolution of hydrocarbons; thus, salinity may control the occurrence of water washing. Price's work is supported by the results of Lafargue and Barker (1988) EFFECTS OF BIODEGRADATION AND WATER WASHING ON CRUDE OIL COMPOSITION Biodegradation Biodegradation produces heavy, low API gravity oils depleted in hydrocarbons and enriched in the nonhydrocarbon nitrogen-, sulfur-, oxygen-bearing (NSO) compounds and asphaltenes (see Glossary for definition of terms). Water washing usually accompanies biodegradation, removing the more water-soluble hydrocarbons, especially the lower molecular weight aromatics such as benzene and toluene. It also aids in concentration of the heavier molecules in the residual oil. Studies demonstrating the selective loss of the gasoline-range hydrocarbons (i.e., the less-than-cis fraction) by water washing and mild biodegradation have been reviewed by Milner et al. (1977). It is not always clear whether the composition (i.e., relative amounts of saturates, aromatics, and naphthenes) of light hydrocarbons is altered by biodegradation, water washing, or evaporative loss. For example, in microbiological experiments using these volatile compounds, unless one collects and identifies metabolites, the loss of these compounds could have causes other than biodegradation (i.e., evaporation and water washing). It is clear, however, that biodegraded and water-washed oils generally are depleted in less-than-cis hydrocarbons; are enriched in sulfur, NSOs, and asphaltenes; and have low API gravities. Enrichment in NSOs, asphaltenes, and cyclic hydrocarbons relative to n-paraffins causes an increase in the optical rotation of an oil. Winters and Williams (1969) and Momper and Williams (1984) demonstrate that optical activity is a useful parameter for indicating degrees of biodegradation In addition to these gross compositional changes, biodegraded oils are most readily recognized by low concentrations of n-paraffins relative to branched (e.g., the

3 4. Effect of Biodegradation and Water Washing on Crude Oil Composition 49 C19 and C20 isoprenoids, pristane and phytane) and cyclic hydrocarbons (naphthenes and aromatic hydrocarbons). For example, the weight percent of n-paraffins relative to naphthenes and aromatics is approximately 2-15 wt. % in the C15+ fraction of biodegraded oils (Figure 1). Gas chromatograms of whole crude oils show that low molecular weight components, e.g., C10 to C14 n-paraffins, are depleted first; the C15+ n-paraffins are then attacked (e.g., Williams et al., 1986). Gas chromatographic patterns of C15+ saturated hydrocarbon fractions (Figure 2) of biodegraded oils contain low amounts of n-paraffins relative to pristane, phytane, and naphthenes. Thus, the loss of n- paraffins relative to branched and cyclic hydrocarbons (in conjunction with low API gravity and enrichment in percent sulfur, NSOs, and asphaltenes) is the most common parameter alluded to as an indicator of biodegradation. Perhaps the focus on the use of the saturated hydrocarbon fraction (e.g., n-paraffins, branched paraffins, and naphthenes such as steranes and terpanes) in the application of oil geochemistry has led to a better understanding of the effects of bacterial action on this fraction. However, aromatic hydrocarbons can also be degraded by bacteria. AromawcB 100% Paraffins Figure 1. Gross ds* hydrocarbon composition of crude oils in terms of percent abundance of paraffins, naphthenes, and aromatic hydrocarbons. Biodegradation removes paraffins leaving an oil enriched in aromatic and naphthenic hydrocarbons. ( a > NONDEGRADED OIL (») SEVERELY BIODEGRADED OIL Numbered peaks = n-paraffins Pr and Ph = isoprenoids, pristane and phytane f lit naphthenes r IB Retention Time, Minutes i i i i i i i. i i i i i IB Retention Time, Minutes Figure 2. Effect of biodegradation on the saturated hydrocarbon fraction of crude oils, (a) Gas chromatogram of Ci-*. saturated fraction of a nondegraded oil contains prominent n-paraffin and branched paraffins, (b) Chromatogram of $5+ saturated fraction of a severely biodegraded oil contains primarily iiaprithenes; the paraffiiis have been removed.

4 50 Palmer Connan (1984) lists examples where aromatic fractions are altered and concludes that more in-depth studies, such as laboratory cultures of aromatic hydrocarbon-metabolizing bacteria, are needed. More documented field examples are needed of biodegradation of the aromatic fractions of reservoired crude oils and the types of bacteria that attacked these oils. Aromatic Hydrocarbons The major classes of aromatic hydrocarbons that are altered by bacteria are those with paraffin side chains, such as the alkylbenzenes (single-ring aromatics). Twoand three-ring aromatics are more resistant to bacterial attack than are the single-ring aromatics (Connan, 1984). Thus, alkylbenzenes are depleted in moderately biodegraded oils. In a study of a sequence of biodegraded oils from south Texas, Williams et al. (1986) showed mat some dimethylnaphthalenes (two-ring aromatics not to be confused with the class of saturated ring compounds, the naphthenes) are removed prior to others. In line with earlier findings of Volkman et al. (1984), this study showed the selective removal of specific dimethylnaphthalenes (2,6-, 2,7-, 1,3-, 1,7-, and 1,6-) relative to other homologs. In contrast, removal of ethylnaphthalenes prior to dimethylnaphthalenes is indicative of water washing (Eganhouse and Calder, 1976). Wardroper et al. (1984) showed a loss of the C20 and C21 triaromatic steranes (i.e., four-ring compounds with three aromatized rings) relative to C26 to C28 homologs during degradation. These authors suggested that the C20 and C21 triaromatic steranes are depleted because of water washing rather than biodegradation. However, the lower solubility of C20+ hydrocarbons may preclude water washing. In the same study, C20 and C21 monoaromatic steranes were not depleted possibly because they are less water soluble than triaromatics (McAuliffe, 1966). Recognition of the loss of the C20 and C21 triaromatics relative to C26 through C28 is important because the ratio of C20 and C21 versus C26 through C28 triaromatic steranes is used to assess relative oil maturity (i.e., timing of oil generation). Connan (1981) showed that even the sulfurcontaining aromatics can be removed from severely biodegraded oils (e.g., asphalts from the Aquitaine basin). It was suggested that anaerobic sulfate-reducing bacteria (rather than aerobic bacteria) attacked these usually resistant compounds. This brief discussion of the effects of biodegradation of aromatic hydrocarbons shows that much remains to be learned about alteration of oils in the subsurface. Indeed, the causes of alteration and the distribution of the affected oils in the subsurface are not always straightforward. Saturated Hydrocarbons As previously mentioned, the biodegradation of the saturated hydrocarbon fraction has been more extensively studied than that of the aromatics. In more detailed discussions of biodegradation, changes within classes of saturated hydrocarbon compounds are considered. The effects of degradation on distributions of compounds used in determining genetic relationships among oils (oil-oil correlation) and thermal maturities must be understood. Other parameters used in correlation, such as stable carbon isotopic composition, can also be influenced by degradative processes. Removal of saturated hydrocarbon compound classes in order of their increasing resistance to biodegradation and a scale of degrees of biodegradation are presented in Volkman et al. (1984). Oil biodegradation in general follows the path outlined as follows, but deviations are frequently observed. Thus, other workers have provided slightly different scales based on their own suite of samples. These deviations remind us that oil transformation is the result of a complex process and that some factors might not be known for a given case. Mild to moderate effects of biodegradation can be readily detected in gas chromatograms of the saturated hydrocarbons, but more extensive degradation (i.e., of the naphthenes) requires gas chromatographic-mass spectrometric analysis (GCMS); these data are usually displayed as single ion mass chromatograms. Volkman et al. (1984) indicate initial or mild biodegradation as the removal of low molecular weight n-paraffins (e.g., gasoline-range n-paraffins), which is most readily observed on whole-oil gas chromatograms. Moderate biodegradation is marked by a nearly total loss of n- paraffins. At slightly higher levels of biodegradation [moderate to extensive), branched paraffins (pristane and phytane) and single-ring naphthenes are removed (Figure 2). In the aromatic fraction, alkylbenzenes are depleted and selective removal of dimethylnaphthalenes occurs during moderate biodegradation. Extensive biodegradation is indicated by removal of two-ring naphthenes (C14 to Ci6 bicyclics), detected by changes in mass chromatograms of the m/z 123 ion. Very extensive biodegradation is denoted as loss of a group of four-ring naphthenes, the C27 to C29 "normal" steranes (Figure 3). Of particular importance is the selective removal of the 20(R)-5a(H)- steranes, which are ratioed against the 20(S)-5oc(H)- steranes to assess the maturity level of an oil (i.e., riming of oil generation and expulsion of an oil from its source rock). Severe biodegradation is indicated by demethylation of the five-ring naphthenes, the C27 to C35 hopanes (Figure 4). A methyl group is removed from the "A" ring (ring number 1 out of 5), producing a new series of compounds: the C-10 demethylated hopanes detected by the m/z 177 ion (Seifert and Moldowan, 1979; Rullkotter and Wendish, 1982). The C30 to C35 hopanes appear to be altered before the C27 to C29 hopanes. Demethylated hopanes predominate in cases of extreme biodegradation, and the C27 to C29 "normal" steranes are completely absent. Philp (1985a) added an additional biodegradation step, the alteration of the "rearranged" steranes, which is considered to be very extreme degradation. An alternative degradation path for hopanes also appears to exist. A series of C26 to C30 (and possibly C31)

5 4. Effect of Biodegradation and Water Washing on Crude Oil Composition 51 (a) NONDEGRADED OIL < 217. bl : c.6;s4 c-^:4=i?z:42 (b) SEVERELY BIODEGRADED OIL 1:0s r.4:rt.j ^^is4.^h': 72)42 7S iao-. = io;5s w> # ill 3,!' ' miwy w^ 16" 19 a,» is Du ieo.j ebb '2898 Figure 3. C27 to C» distributions (nvz = 217) of (a) a nondegraded oil and (b) a severely biodegraded oil. Normal steranes (peaks 8-11 and 15-22) are consumed by bacteria in (b), leaving an abundance of rearranged steranes (peaks 1-7 and 12-13). See Figure 5 for names of individual steranes. tetracyclic compounds, the 8,14-seco-hopanes, are formed by opening the C ring (ring number 3 out of 5) (Rullkotter and Wendish, 1982). In such cases, demefhylated hopanes can also be present and the steranes may be only slightly altered. These examples suggest that various degradative processes can operate to produce severely biodegraded oils. Perhaps certain environmental conditions are required to allow specific bacteria to grow on oils. The C19 to C26 three-ring naphthenes (tricyclic terpanes) survive extreme biodegradation, although demefhylated tricyclic terpanes have been tentatively identified (e.g., Howell et al., 1984; Philp, 1985a). Because of their resistancetobiodegradation, tricyclic terpanes have been used for oil-oil correlation in severely biodegraded oils. Their distributions also supply information concerning depositional environments (e.g., Zumberge, 1987). As previously mentioned, the stable carbon isotopic composition of crude oils can also be altered by biodegradation, although not in a consistent manner. For example, in a 42-day simulated oil biodegradation study, Stahl (1980) observed that the saturated hydrocarbon fraction was enriched in 13 C (i.e., more positive # 3 C values), but the isotopic composition of the aromatic hydrocarbon fraction remained unchanged. Sofer (1984) and Momper and Williams (1984) showed that the saturated fraction of naturally biodegraded oils is also enriched in 13 C. However, field examples showing no or little change in isotopic composition or changes in both the saturated and aromatic fractions have also been reported (Sofer, 1984). Connan (1984) has reviewed other studies in which the isotopic composition of crude oil fractions other than the saturated hydrocarbons also become enriched in 13 C Water Washing Water washing is most readily recognized by changes in the composition of the gasoline-range hydrocarbons because these compounds are more water soluble than the C15+ hydrocarbons (McAuliffe, 1966; Price, 1976). For a given carbon number, ring formation, unsaturation, and branching cause an increase in water solubility. Thus, one would expect that when water washing occurs, aromatic hydrocarbons of a given carbon number would decrease first, followed by naphthenes, branched paraffins, and n- paraffins. Generally, the loss of benzene and toluene is a good indicator that water washing has occurred. These low molecular weight aromatics are also biodegradable; however, their high water solubilities make them useful indicators of water washing. Other indicators of water washing are the loss of ethylnaphthalenes relative to dimethylnaphthalenes (Eganhouse and Calder, 1976) and possibly (as discussed in the previous section) the loss of C20 and C21 triaromatic steranes (Wardroper et al., 1984). Experimental water washing studies by Lafargue and Barker (1988) do support the loss of gasoline-range (kssthan-cis) aromatic hydrocarbons relative to naphthenes and paraffins in line with the solubility studies previously noted. An example of the effect of water washing on the Q54- hydrocarbon composition involved a field study of Philippine oils having abundant sulfur-containing aromatic hydrocarbons (dibenzothiophenes). Based on the idea that heteroatomic compounds are more water soluble than aromatic, cyclic, branched, and straight-chain hydrocarbons (e.g., Price, 1976), water washing was thought by Palmer (1984) to cause the loss of dibenzothiophene (G2H8S) and methyldibenzothiophene (C13H10S) relative

6 52 Palmer C M m/z demethylated hopanes to phenanthrene (CuHio) and methylphenanthrene (C15H12) from the C15+ aromatic hydrocarbon fraction. Also, because the Q5+ aromatic hydrocarbon fraction was being altered more extensively than the C15+ saturated hydrocarbon fraction, water washing, rather than biodegradation, was proposed as the major agent of oil alteration. In support of this idea, sulfur-bearing aromatics are not as readily degraded by bacteria as are aromatic hydrocarbons (e.g., Connan, 1981), especially in comparison with n- paraffins. Loss of dibenzothiophene relative to phenanthrene in oils not depleted in n-paraffins may be a useful indicator of water washing. Unfortunately, not all oils contain sufficient quantities of dibenzothiophenes to allow monitoring of such changes; therefore, this parameter cannot be used universally. Also, Connan (1981) postulated that anaerobic sulfate-reducing bacteria caused the preferential removal of aromatic hydrocarbons (and even the sulfur-bearing aromatics) in asphalts of the south Aquitaine basin in France. Thus, the loss of aromatic hydrocarbons and dibenzothiophenes could be due to special cases of biodegradation rather than to water washing. Therefore, changes in the gasoline-range hydrocarbons remain best suited for studies of water washing. Loss of benzene and toluene from the gasoline-range hydrocarbon fraction appears to be the most useful parameter. The isotopic composition of crude oils can be altered by water washing (Sofer, 1984). In the case of the Philippine oils, the saturated fraction (especially the naphthenes) became depleted in 13 C, resulting in an isotopically light oil (with more negative values). The aromatic fraction, although chemically altered, showed little change in isotopic composition (Palmer, 1984) CB7 IOMCBBI <OMCJ,7J r 1, ^Xjfc^JvJkX, CM OMC31I C31 r COM C^l C M '. «-! lomca,! I I I, I c 1 1 I Time, Minutes Rgure4. Example of a crude oil containing demethylated hopanes (m/!z=177). The presence of demethylated hopanes is indicative of severe biodegradation. In the case shown here, an early oil became severely biodegraded; migration of a nondegraded oil later filled the same reservoir, mixing with the severely degraded oil. (a) The gas chromatogram of the Ct&. saturated fraction thus represents hydrocarbons from the nondegraded oil. (b)the demethylated hopanes are initially detected in the pentacyclic triterpane fraction by the large DMCn peak, the demethylated CM hopane. (c) For verification, the complete series of demethylated hopanes are identified by scanning therrvz177ion. (FromSoferetal., 1986.) SeeRgure5for names of individual triterpanes.» SUMMARY Processes that alter the composition of a crude oil can have a direct bearing on the commercial value of an oil field. Therefore, the petroleum geochemistry community has made a major effort to understand the occurrences and causes of biodegradation and water washing. Biodegradation generally occurs in relatively shallow, cool reservoirs that are charged with oxygenated water. Such conditions allow the growth of aerobic bacteria. Both case history studies of biodegraded crude oils and laboratoryinduced changes in oil composition using bacteria have shown that aerobic bacteria are the major agents of oil degradation. Special cases exist in which anaerobic bacteria are the suspected agents of degradation. However, the present understanding is that anaerobic Figure 5. (Facing page) (Top) List of identified triterpanes. (Bottom) List of indentified steranes. Assumes 8#H),9c<H),14a(H),17a(H) unless otherwise stated; dia = rearranged.

7 4. Effect of Biodegradation and Water Washing on Crude Oil Composition 53 LIST OF I0ENTIFIE0 TRITERPANES Peak Oeslgnatl on A B BP DM C D OL E F G H I J K L M N 0 Q R S T Molecular Formula C27H46 C27><46 C 28 M 48 C 29 H 50 C 29 M 50 C 29 H 50 C 30 H S2 C30 M 52 C30 H 52 C31«54 C31H54 C30H52 C31H54 C32"56 32*56 C32H56 C33HS8 C33HS8 C34H6O C34HSQ. C35"62 C35H62 Mo!ecu 1 ar Weight S Triterpane Identification 18a(H)-22, 29, 30-tr1snorhopane 17a(H)-22, 29, 30-tr1snorhopane 17o(H). 18a(H), 216(H)-28, 30-blsnorhopane demethylated hopane at A/B ring 17a(H), HB(H)-30-norhopane 178(H), 21a(H)-30-nornnretane Oleanane 17a(H), 218(H)-hopane 176(H), 21o(H)-moretane l7a(h). 21B(H)-30-honohopane (22S) 17a(H), 2U(H)-30-r,omohopane (22R) gamnacerane 176(H). 21a(H)-30-homomoretane 17a(H), 21B(H)-30. 3l-b1shomohopane (22S) 17a(H), 2U(H)-30, 3l-b1shomohopane (22R) 171(H), 21«(H) blshomomoretane 17a(H), 218(H)-30, 31, 32-trishomohopane (22S) 17a(H), 2la(H) , 32-trishomohopane (22R) 17a(H), 216(H) , 32, 33- tetraktshomohopane (22S) 17o(H), 21B(H)-30, 31, 32, 33- tetraklshomohopane (22R) 170(H), 21B(H)-30, 31, 32, 33, 34- pentaklshoaohopane (22S) 170(H), 21B(H)-30, 31, , 34- pentaklshomohopane (22R) LIST OF IDENTIFIED STERANES Peak IS ia Molecular Formula Molecular Weight C27M4g C27 H 48 C27H48 c 28*50 C 28 H 50 C2B H 50 ^ Ho C 29 H 52 C 29 H 52 C29 H 52 C29 H 52 C2flH50 C 28 H 50 C 28 H 50 C 28 H 50 C29H52 C2«H52 C»H52 C 29 H S2 Sterant Identification 136(H). 17a(H) diacholestine (20S) 138(H), 17a(H) dlacholestane (20R) 13a(H). 17B(H) diacholestine (20S) 13a(H), 17a(H) dlacholesttne (20R) 136(H) 17a(H)d1e*rgostane (20S) Rearranged Cjg sterant Rearranged Cjg sterane 138(H), 17«(H) dlaergostane (20R) 5a(H) cholestane (20S) 5g(H) Cholestin* (20R) 5a(H), 14e(H), 17B(H) cholestan* (20R) 1311(H), 17 a(h) dlastlgnastan* (20S) Sa(H), 148(H), 178(H) cholestane (20S) Su(H) cholestane (20R) Rearranged Cyg sterane Rearranged Cjg sterane Rearranged Cjg sterane 5a(H) ergostane (20S) 50(H), 14B(H), 17e(H) ergostane (20R) 5B(H) ergostane (20R) Si>(H), 148(H), 178(H) ergostane (20S) 5a(H) ergostane (20R) 5a(H) stlgmstane (20S) 5o(H), 14s(H), 17e(H) stlgnastane (20R) 58(H) stlgmastane (20R) 5a(H), 14g(H), 176(H) stlgnastane (20S) 5a(H) stlgmastane (20R)

8 54 Palmer bacteria can live on the metabolites of aerobic bacteria and cannot degrade crude oil directly. Biodegradation produces a heavy, low API gravity oil depleted in hydrocarbons and enriched in nonhydrocarbons. The most commonly used indicator of biodegradation is the loss of the n-paraffins relative to the branched paraffins, naphthenes, and aromatic hydrocarbons. This is because bacteria consume n-paraffins prior to branched paraffins, naphthenes and aromatic hydrocarbons. The loss of n-paraffins is readily observed in whole-oil gas chromatograms and in chromatograms of the C15+ saturated hydrocarbon fraction. Petroleum geochemists are also concerned with recognizing the effects of biodegradation on correlation parameters. Bacteria consume hydrocarbons used in correlation (e.g., n-paraffins, pristanes, phytanes, steranes, and pentacyclic terpanes) and alter the isotopic composition of an oil. These changes must be recognized in oil geochemical studies when one is determining genetic relationships among oils, proposing source rock types and depositional environments based on oil composition, and determining the thermal maturity of the source rock at the time of oil generation. Most correlation parameters are based on the saturated fractioa Therefore, a greater emphasis has been placed on understanding how biodegradation affects the saturated fraction than the aromatic fraction. Microbial studies, however, show that the aromatic hydrocarbons can also be biodegraded. More case studies need to be performed to gain an appreciation for bacterial attack on these compounds. Because water is present during biodegradation, water washing and biodegradation can occur simultaneously. In such cases it is difficult to determine whether biodegradation and/or water washing has altered the oil. Water washing can also occur in deeper reservoirs where lack of oxygen and high temperatures prohibit the growth of aerobic bacteria. Water washing is enhanced at higher temperatures but high salinities cause exsolution of hydrocarbons that would reduce their removal by water. Although other agents need to be considered, water washing is the suspected agent that degrades the aromatic fraction. This idea is based primarily on hydrocarbon solubility studies; aromatic hydrocarbons are more water soluble than saturated hydrocarbons (n-paraffins and naphthenes). Because smaller molecules are more water soluble, water washing is most readily recognized by changes in the composition of the gasoline-range hydrocarbons (less-than-ci5 fraction). The loss of benzene and toluene relative to less-than-cis paraffins and naphthenes is the most useful parameter for this fraction. Loss of C15+ aromatic hydrocarbons, especially the more water-soluble sulfur-containing components, due to water washing has also been proposed. In cases where evidence for biodegradation of the C15+ saturated fraction is lacking, depletion of the C15+ aromatic fraction is attributed to water washing. However, in cases of extensive biodegradation, as demonstrated by the destruction of the saturated hydrocarbons including naphthenic steranes and pentacyclic terpanes (hopanes), degradation of the aromatic fraction may also be due to biodegradation.