Chapter 4 Effect of Biodegradation and Water Washing on Crude oil Composition Susan e. palmer Amoco Production Company Tulsa, Oklahoma, U.s.A INTRODUCTION Eganhouse and Calder, 1976; May et al, 1978a, b; Lafargue and Barker, 1988). Extensive review articles covering The study of crude oil geochemistry becomes difficult work done prior to 1985 on biodegradation and water when crude oils are altered by microbial action(biodegra washing have been prepared by Milner et al.(1977)and dation)and/or water washing. These processes can alter Connan( 1984). These two reviews give an overview of parameters used to compare oils when determining the effects of biodegradation and water washing gleaned genetic relationships (oil-oil correlation), depositional from the many exemplary papers found in the literature environments, and time of oil generation (i.e. thermal Also, Lafargue and Barker(1988)have reviewed and maturity of the source rock at the time of oil generation discussed data obtained from laboratory water washing and expulsion ). Much of this knowledge comes from experiments and present their own observations and histories of such occurrences through the years(e.g chapter is to compile and present Winters and Williams, 1969; Bailey et al., 1973a, b, the results of these studies and review articles in an abbre- Rubinstein et al., 1977; Connan et al, 1975, 1980; Seifert viated form to aid the explorationist in understanding the and moldowan, 1979: Rullkotter and Wendish 1982 Volkman et al, 1983: Momper and williams, 1984; Palmer, regard, the reader is directed to the literature for details of In addition, microbiologists and petroleum individual studies. Results of more recent studies geochemists have studied the action of bacteria on (1985-1988) are also included here. Examples of para ters and hydrocarbon distributions demonstrating the petroleum in the laboratory (e. g McKenna, 1972, effects of biodegradation and water washing are given to Horowitz et al., 1975; Jobson et al, 1979: Connan, 1981; aid the reader in recognizing altered oils. Because many Goodwin et al., 1983). Some workers have isolated products of bacterial metabolism of crude oils or classes of geochemical laboratory groups have developed their own hydrocarbons(e.g, Gibson, 1976 Higgens and gilbert, ways to portray data, some of these techniques will be 1978; Cripps and Watkinson, 1978; Cain, 1980; Mackenzie referenced in this discussion. Organic geochemical et al., 1983). Connan(1984) illustrates the metabolic parameters and types of hydrocarbon classes referred to products recognized by some of these workers and others. here are defined in the Glossary at the back of this volt Products of aerobic degradation are most often organic cids and CO. Anaerobic bacteria can live on the metabo- GEOLOGICAL CONSTRAINTS AND lites of the aerobes but do not grow on hydrocarbons. PHYSICOCHEMICAl CONdItIoNS tion is only sligh FOR BIODEGRADATION AND Alteration of crude oil by water washing has been indi- ctly studied in the laboratory through determination of WATER WASHING yater solubilities of individual hydrocarbons and The processes of microbial degradation and water mixtures of several hydrocarbons and by studying washing of crude oils occur when certain conditions are compositional changes of whole crude oils( e.g., met. Milner et al. (1977) and Connan(1984 )have outlined McAuliffe, 1966, 1980; Bailey et al., 1973b, Price, 1976; the requirements for both processes in their review
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 (1985-1988) 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
palmer articles. Their findings are summarized here. (1988) have suggested that water washing during Biodegradation occurs in surface seeps and in relatively migration must be minimal because highly water soluble shallow reservoirs, e.g, 4000-6000 ft(1220-1830 m)or less. molecules, namely, benzene and toluene, are present in though anaerobic bacteria can survive on partially most oils. They suggest that most water washing occurs degraded oils, both case histories and microbiological after accumulation where, given the proper conditions, studies show that aerobic bacteria are the major agents of biodegradation is also occurring. Water washing can, crude oil degradation. Aerobic bacteria can grow in rela- however take place outside the temperature, oxygen, and tively cool reservoirs (i.e, below 80C, or 176F)that are salinity constraints of biodegradation. Price(1976)and invaded by oxygen-charged waters. In addition to Lafargue and Barker(1988)have noted that the solubility dissolved oxygen, nutrients such as nitrate and phosphate of crude oil components increases markedly at higher must be present and the salinity of the water must be less temperatures. These results demonstrate that water than 100-150%o. Also, unless special cases such as washing can occur in zones where microbial activity is oxygenated microenvironments exist, the amount of Hs precluded by high temperature. The work of Price(1976) in the oil must be very low, as it is toxic to aerobic bacteria. shows that high salinities(over 270%o)cause exsolution of Thus, cool, shallow reservoirs that are flushed by hydrocarbons; thus, salinity may control the occurrence of oxygenated, nutrient-rich fresh water can be expected to water washing. Price's work is supported by the results of contain oil that is being actively biodegraded. Lafargue and Barker(1988) However, biodegraded oils are also present in deeper reservoir dence of reservoirs, biodegraded oils are found preserved EFFECTS OF BIODEGRADATION far below the arbitrary 6000-ft cutoff for bacterial activity. AND WATER WASHING ON CRUDE Thus, cases where biodegraded oils occur in reservoirs of e.g,8000-10,000 feet(2440-3050 m) have been reported OIL COMPOSITION Also, with regard to maximum depth of reservoirs and gr Biodegradation deeper occurrences. Connan(1984) points out that aerobic Biodegradation produces heavy, low API gravity oils bacteria degrade oil at the oil-water int In such depleted in hydrocarbons and enriched in the nonhydro- cases, the lower part of an oil accumulation will be carbon nitrogen-, sulfur-, oxygen-bearing(NSO) degraded rather than the upper portions, unless more compounds and asphaltenes(see Glossary for definition than one oil-water contact is present. Some reservoirs of terms). Water washing usually accompanies biodegra have multiple oil-water contacts and different hydrolog- dation, removing the more water-soluble hydrocarbons, ical regimes that could lead to a complex and perhaps especially the lower molecular weight aromatics such as onfusing array of degraded and undegraded oils. benzene and toluene. It also aids in concentration of the Lafargue and Barker (1988)mention that hydrodynami- heavier molecules in the residual oil. Studies demon- ally tilted oil-water contacts are indicators of actively strating the selective loss of the gasoline-range hydrocar- flowing waters and delineate areas where degradation is bons(ie, the less-than-- Ci5 fraction )by water washing and mild biodegradation have been reviewed by Milner et al. Physical and chemical processes other than biodegrad- (1977). It is not always clear whether the composition(ie ation and water washing (e.g fractionation of light and relative amounts of saturates, aromatics, and naphthenes) heavy ends during migration and in-reservoir maturation) of light hydrocarbons is altered by biodegradation, water can add to the difficulty of understanding the transforma- washing, or evaporative loss. For example, in microbiolog- tion of oil from its initial state to the time it is recovered in ical experiments using these volatile compounds, unless a discovery well or surface seep. Some of these other one collects and identifies metabolites, the loss of these processes could be mistaken as biodegradation or water compounds could have causes other than biodegradation tion bi g. Thus, a good understanding of the postgenera- (ie, evaporation and water washing).It is clear, however istory of an oil is of major importance for estab- that biodegraded and water-washed oils generally are lishing cause and effect relationships depleted in less-than-C15 hydrocarbons; are enriched in Because water is a necessary ingredient for biodegrada- sulfur, NSOs, and asphaltenes; and have low API tion, the process of water washing generally accompanies gravities. Enrichment in NSOs, asphaltenes, and cy biodegradation. In spite of many studies that have hydrocarbons relative to n-paraffins causes an increase in attempted to isolate the effects of water washing from the optical rotation of an oil. winters and williams( 1969) microbial alteration, questions concerning the actual cause and Momper and williams(1984)demonstrate that of alteration of a crude oil's geochemistry, especially optical activity is a useful parameter for indicating degrees pecific parameters, are still open for discussion. of biodegradation. Conditions favorable for water washing exist during oil In addition to these gross compositional changes, migration if oil is passing through a water-wet carrier bed biodegraded oils are most readily recognized by low and reservoir system. However, Lafargue and Barker concentrations of n-paraffins relative to branched (e.g, the
48 Palmer articles. Their findings are summarized here. Biodegradation occurs in surface seeps and in relatively shallow reservoirs, e.g., 4000-6000 ft (1220-1830 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 100-150 %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., 8000-10,000 feet (2440-3050 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
4. Effect of Biodegradation and Water Washing on Crude Oil Composition C19 and C2o isoprenoids, pristane and phytane)and cyclic 100 Parasit d 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 chro- matograms of whole crude oils show that low molecular weight components,e.g, C1o to C14n-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 biode- graded oils contain low amounts of n-paraffins relative to pristane, phytane, and naphthenes. Thus, the loss of n- araffins relative to branched and cyclic hydro conjunction with low API gravity and enrichment in percent sulfur, NSOs, and asphaltenes)is the most er alluded to as an indicator of biod dation Perhaps the focus on the use of the saturated hydro- rbon fraction(e-g n-paraffins, branched paraffins,and naphthenes such as steranes and terpanes)in the applica- Figure 1. Gross Cis, hydrocarbon compositon of tion of oil geochemistry has led to a better understanding and aromatic hydrocarbons. Biodegradation remo aromatic hydrocarbons can also be degraded by bacteria (a)NONDEGRADED OIL (b)SEVERELY BIODEGRADED OIL Pr and Ph= isoprenoids, pristane naphthenes _2557.511256 25}5mzs5h,5aa5如 Retention Time, Minutes Retention TIme, MInutes Figure 2. Effect of biodegradation on the saturated hydrocarbon fraction of crude fraction of a nondegraded oll contains prominent paraffin and branched paraffins, chromatogram of C15+ saturated fraction of a severely biodegraded oil contains primarily naphthenes; the paraffins have been removed
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 nparaffins 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. 100% Paraffins AromawcB 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 f lit Numbered peaks = n-paraffins Pr and Ph = isoprenoids, pristane and phytane naphthenes 2.5 5 -1 1 1 1 1 1 1 r— 1 1 1 — 7.5 IB 12.5 15 17.5 28 22.5 25 27.5 38 32.5 (») SEVERELY BIODEGRADED OIL —i—i—i—i—i—i—i—.—i—i—i—i—i — 2.5 57. 5 IB 12.5 15 17.5 28 22.5 25 27.5 38 32.5 Retention Time, Minutes - 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
Palmer Connan(1984)lists examples where aromatic fractions are effects of degradation on distributions of compounds used altered and concludes that more in-depth studies, such as in determining genetic relationships among oils(oil-oil laboratory cultures of aromatic hydrocarbon-metabolizing correlation) and thermal maturities must be understood bacteria, are needed. More documented field examples Other parameters used in correlation, such as stable are needed of biodegradation of the aromatic fractions of carbon isotopic composition, can also be influenced by reservoired crude oils and the types of bacteria that degradative processes attacked these oils Removal of saturated hydrocarbon compound classes Aromatic Hydrocarbons in order of their increasing resistance to biodegradation The major classes of aromatic hydrocarbons that and a scale of degrees of biodegradation are presented in altered by bacteria are those with paraffin side chains, Volkman et al. ( 1984). Oil biodegradation in general aa+amm时 observed. Thus, other wo如hpN attack than are the single-ring aromatics( connan, 1984). slightly different scales based on their own suite of Thus, alkylbenzenes are depleted in moderately biode- samples. These deviations remind us that oil transforma- graded oils. In a study of a sequence of biodegraded oil tion is the result of a complex process and that some from south Texas, Williams et al. (1986) showed that some factors might not be known for a given case. Mild to dimethylnaphthalenes( two-ring aromatics not to be moderate effects of biodegradation can be readily detected confused with the class of saturated ring compounds, the in gas chromatograms of the saturated hydrocarbons,but naphthenes)are removed prior to others. In line with more extensive degradation(i.e, of the naphthenes) requires gas chromatographic-mass spectrometric earlier findings of Volkman et al. (1984), this study analysis(GCMS); these data are usually displayed as showed the selective removal of specific dimethylnaph- thalenes(2, 62, 7,1 3-, 1, 7,and 1 6-)relative to other gle ion mass chromatograms Volkman et al. ( 1984)indicate initial or mild biodegrad homologs. In contrast, removal of ethylnaphthalenes tion as the removal of low molecular weight n-paraffins prior to dimethylnaphthalenes is indicative of water ashing(Eganhouse and Calder, 1976). (e. g, gasoline-range n-paraffins), which is most readily Wardroper et al. (1984)showed a loss of the Cx and,, observed on whole-oil gas chromatograms. Moderate triaromatic steranes(ie, four-ring compounds with three biodegradation is marked by a nearly total loss of n- aromatized rings)relative to C2 to C2s homologs during (moderate to extensive), branched paraffins(pristane and degradation. These authors suggested that the Cz and C2l phytane)and single-ring naphthenes are removed(Figure triaromatic steranes are depleted because of water washing rather than biodegradation. However the lower 2). In the aromatic fraction, alky benzenes are dep solubility of C20+ hydrocarbons may preclude water and selective removal of dimethylnaphthalenes occurs In the same study, C20 and C2l monoaromatic during moderate biodegradation. Extensive biodegrade- tion is indicated by removal of two-ring naphthenes(C14 water soluble than triaromatics (McAuliffe, 1966). to C16 bicyclics), detected by changes in mass chro- tion is denoted as loss of a group of four-ring naphthene relative to C2 through C28 is important because the ratio the Cr to Ca"normal"steranes(Figure 3). Of particular is used to assess relative oil maturity (ie, timing of oil importance is the selective removal o the 20(R)-sa(H- generation).Connan(1981)showed that even the sulfur- steranes to assess the maturity level of an oil (ie, timing of graded oils(e.g, asphalts from the Aquitaine basin). It oil generation and expulsion of an oil from its source rock) biodeg ion is indicated by demethylation of was suggested that anaerobic sulfate-reducing bacteria the five-ring naphthenes, the Cz to C35 hopanes(Figure 4) (rather than aerobic bacteria)attacked these usually A methyl group is removed from the"A"ring(ring number 1 out of 5), producing a new series of compounds This brief discussion of the effects of biodegradation of the C-10 demethylated hopanes detected by the m/z 177 aromatic hydrocarbons shows that much remains to be ion(Seifert and Moldowan, 1979: Rulkotter and Wendish, leamed about alteration of oils in the subsurface. Indeed, 1982). The C3o to Cas hopanes appear to be altered before the causes of alteration and the distribution of the affected oils in the subsurface are not always straightforward the Czz to C2g hopanes, Demethylated hopanes predomi ate in cases of extreme biodegradation, and the Cz to C29 Saturated Hydrocarbons normal"steranes are completely absent. Philp (1985a) As previously mentioned, the biodegradation of the added an additional biodegradation step, the alteration of saturated hydrocarbon fraction has been more extensively the "rearranged"steranes, which is considered to be uery studied than that of the aromatics. In more detailed extreme degradation discussions of biodegradation, changes within classes of An alternative degradation path for hopan saturated hydrocarbon compounds are considered. The appears to exist. A series of C2 to C3o(and possibly C3)
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 nparaffins. 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)
4. Effect of Biodegradation and Water Washing on Crude Oil Composition 51 (a NONDEGRADED OIL 一,2、a() SEVERELY BIODEGRADED OIL e’·之52 √“NM 28A8 MM 253-.-20-1220130 car to Czs distributions(m/z =21n)of (a) a nondegraded oil and (b a severely biodegraded oil. Normal sterar -11 and 15-22)are consumed by bacteria in(b), leaving an abundance of rearranged steranes(peaks 1-7 and 12-13) re 5 for names of individual steranes. tetracyclic compounds, the 8, 14-seco-hopanes, are formed Water Washin by opening the C ring(ring number 3 out of 5) ( Rullkotter and Wendish, 1982). In such cases, demethylated hopanes in Water washing is most readily recognized by cha the composition of the gasoline-range hydrocarbons can also be present and the steranes may be only slightly because these compounds are more water soluble than th processes can operate to produce severely biodegraded given carbon number, ring formation, unsaturation, and required to allow specific bacteria to grow on oils. would expect that when water washing occurs, aromatic The C1 to C2 three-ring naphthenes(tricyclic terpanes) hydrocarbons of a given carbon number would decrease survive extreme biodegradation, although demethylated first, followed by naphthenes, branched paraffins, and n tricyclic terpanes have been tentatively identified(e.g, paraffins. Generally, the loss of benzene and toluene is a Howell et al., 1984; Philp, 1985a). Because of their resis- good indicator that water washing has occurred.These tance to biodegradation, tricyclic terpanes have been used low molecular weight aromatics are also biodegradable, for oil-oil correlation in severely biodegraded oils. Their however, their high water solubilities make them useful distributions also supply information concerning deposi- indicators of water washing. Other indicators of water onal environments(e. g, Zumberge, 1987) washing are the loss of ethy naphthalenes relative to As previously mentioned, the stable carbon isoto dimethylnaphthalenes(Eganhouse and Calder, 1976)and composition of crude oils can also be altered by biodegra- possibly(as discussed in the previous section)the loss of dation, although not in a consistent manner. For example, C2o and C2 triaromatic steranes wardroper et al., 1984) in a 42-day simulated oil biodegradation study, Stahl Experimental water washing studies by Lafargue and (1980)observed that the saturated hydrocarbon fraction Barker(1988)do support the loss of gasoline-range (less was enriched inC(ie, more positive 8 C values), but than C1s)aromatic hydrocarbons relative to naphthenes he isot ition of the aromatic hydrocarbon and paraffins in line with the solubility studies previously fraction remained unchanged. Sofer(1984)and Momper noted. and williams(1984)showed that the saturated fraction of An example of the effect of water washing on the Cl naturally biodegraded oils is also enriched in c. hydrocarbon composition involved a field study of However, field examples showing no or little change in Philippine oils having abundant sulfur-containing isotopic composition or changes in both the saturated and aromatic hydrocarbons( dibenzothiophenes). Based on the aromatic fractions have also been reported(Sofer, 1984). idea that heteroatomic compounds are more water soluble Connan(1984)has reviewed other studies in which the than aromatic, cyclic, branched, and straight-chain hydro isotopic composition of crude oil fractions other than the carbons(e-g, Price, 1976), water washing was thought by saturated hydrocarbons also become enriched Palmer (1984)to cause the loss of dibenzothiophene 12H8S)and methyldibenzothiophene(C13H1oS)relative
4. Effect of Biodegradation and Water Washing on Crude Oil Composition 51 # ' i l l 3 ,!' w^ miwy 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 resistance to biodegradation, 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 13C (i.e., more positive #3C 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 13C. 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 in1 13C 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 nparaffins. 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
Palmer to phenanthrene(C14H1o)and methylphenanthrene C15H12) from the C15+ aromatic hydrocarbon fraction. Also, because the C1s+ 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 Iteration In support of this idea, sulfur-bearing aromatics are as readily degraded by bacteria as are aromatic hydrocar bons(e.g, Connan, 1981), especially in comparison with n- threne in oils not depleted in n-paraffins may be a useful indicator of water washing. Unfortunately, not all oils contain sufficient quantities of dibenzothiophene to allow monitoring of such changes; therefore, this parameter cannot be used universally. Also, Connan(1981)postur- lated that anaerobic sulfate-reducing bacteria caused the preferential removal of aromatic hydrocarbons(and even r mz 191-titerpane 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 hydro- carbons 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 The isotopic composition of crude oils can be altered by water washing(Sofer, 1984). In the case of the Philippin oils, the saturated fraction(especially the naphthenes) became depleted inC, resulting in an isotopically light mz 177-demetylated oil(with hopanes negative values). The aromatic fraction, although ically altered, showed little change in 〔 Palmer,1984) SUMMARY Processes that alter the composition of a crude oil can MWwNWLA have a direct bearing on the commercial value of an oil field. Therefore, the petroleum geochemistry community as made a major effort to understand the occurre and causes of biodegradation and water washing Time, Minutes Biodegradation generally occurs in relatively shallow, cool conditions allow the growth of aerobic bacteria. Both case Figure 4. Example of a crude oll containing demethylated history studies of biodegraded crude oils and laboratory- panes(m/z=17. The presence of demethylated panes is indicative of severe blodegra In the case induced changes in oil composition using bacteria have shown here, an earty oll became severely biodegraded shown that aerobic bacteria are the major agents of oil migration of a nondegraded oll later filled the same degradation. Special cases exist in which anaerobic servoir, mixing with the severely degraded oil. (a)The bacteria are the suspected agents of degradation chromatogram of the C15+ saturated fraction thus repre- However, the present understanding is that anaerobic sents hydrocarbons from the nondegraded oll(bThe demethylated hopanes are initially detected in the penta cyclic triterpane fraction by the large DMC3o peak, the demethylated Cao hopane. c) For verfication, the complete Figure 5.(Facing page)(Top)List of Identified triterpanes. series of demethylate scan ( Bottom)ust of indentified steranes. Assumes the mz177 ion.(From Soferet al, 1906 )See Figure 5 for 8e)9dH),14d H), 17d) unless otherwise stated; names of individual triterpanes. dia s rear
52 Palmer C B 7 IOMCBB I <OMCJ, 7 J r 1, ^Xjfc^JvJkX, m/z 177 - demethylated hopanes C M C M 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 C M hopane. (c) For verification, the complete series of demethylated hopanes are identified by scanning therrvz177ion. (FromSoferetal., 1986.) SeeRgure5for names of individual triterpanes. 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 nparaffins. 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 13C, resulting in an isotopically light oil (with more negative values). The aromatic fraction, although chemically altered, showed little change in isotopic composition (Palmer, 1984) 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
4. Effect of Biodegradation and Water Washing on Crude Oil Composition LIST OF IDENT [F !ED TRI TERPANES Weight Triterpane Identification c27N46 370 18aH)-22,29 218(H)-28, 30-bfsnorhopane demetry lated hopane at A/B ring 412 cy052 178()2la(H)-° stand 17a(H),21B(H)-30-homohopane(22S) 17a(H), 21B(H)-30-homohopa ne(22R) 17B(H). 2la(H)-30-honomoretane C32H56 17c(H),21·(H)-30,31-b1 shomo hopane(225 c256 17a(H,218(H)-30,31-1 shonohopane(22R 178(H). 2la(H)-30, 31-bishomomoretane C33H58 a( H),21B(H]-30, 31, 32-trishomohopane(22S) C33H58 17a(H), 218(H)-30, 31, 32-trishomohopane(22R) 17a《H),2(H)-30,31,32, tetraktshomohopane (22R) 4B217a(N),2L昌(N}-30,31,32,33,34 penta ishomohopane(22R) LIST OF IDENTIFIED STERAES steran【 dentification 13B(H).17()diacholestane(20S) 37213H),17a(M) 13a(H). 17B(H)diacholestane(20s) 37213a(H),17a(N)dah 136()17a(H)diverge anged C28 sterane JB(H),17a(H)diaergostane 5a(M) cholestane·【205)+5s(H) cholestan 3725a(H)14B(M),1e《H) cholestane(20) 18(M),17aM)dast。msta№·(20s) 25a《M】,14(M),17H) cholistan·【20s 12c2952 400 arranged C2 steran會 c2952 5 C28H50 365a(N),148(M),178(M)· gostaM【20) 5(M】 ergostane(20R) 3055a(M),14B(H),17B(H)· rastan(20) C29k52 (H),14(),176(H)t1 gaston(20) 58(M】st1 gStaad(20R 21 CzgH52 4005(M),14B(M),176(")fn20s S(M)stigmstane(20R)
4. Effect of Biodegradation and Water Washing on Crude Oil Composition 53 Peak Oeslgnatl A B BP DM C D OL E F G H I J K L M N 0 Q R S T Peak 1 2 3 4 5 6 7 8 9 10 11 12 13 14 IS 16 17 ia 19 20 21 22 Molecular on Formula C27H46 C27>(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)
palmer bacteria can live on the metabolites of aerobic bacteria and Because water is present during cannot degrade crude oil directly washing and biodegradation can occur simultaneously. In Biodegradation produces a heavy, low API gravity oil such cases it is difficult to determine whether biodegrada depleted in hydrocarbons and enriched in nonhydrocar- tion and/or water washing has altered the oil. Water bons. The most commonly used indicator of biodegrade- washing can also occur in deeper reservoirs where lack of tion is the loss of the n-paraffins relative to the branched oxygen and high temperatures prohibit the growth of paraffins, naphthenes, and aromatic hydrocarbons. This is aerobic bacteria. Water washing is enhanced at higher because bacteria consume n-paraffins prior to branched temperatures but high salinities cause exsolution of hydro paraffins, naphthenes and aromatic hydrocarbons. The carbons that would reduce their removal by water loss of n-paraffins is readily observed in whole-oil gas Ithough other agents need to be considered,water chromatograms and in chromatograms of the C15+ washing is the suspected agent that degrades the aromatic saturated hydrocarbon fract fraction. This idea is based primarily on hydrocarbon Petroleum geochemists are also concemed with recog- solubility studies; aromatic hydrocarbons are more water nizing the effects of biodegradation on correlation parame- soluble than saturated hydrocarbons(n-paraffins and ters. Bacteria consume hydrocarbons used in correlation naphthenes). Because smaller molecules are more water paraffins, pristane, phytanes, steranes, and penta soluble, water washing is most readily recognized by cyclic terpanes)and alter the isotopic composition of an changes in the composition of the gasoline-range hydro- oil. These changes must be recognized in oil geochemical carbons(less-than C1s fraction). The loss of benzene and studies when one is determining genetic relationships toluene relative to less-than-C15 Paraffins and naphthenes among oils, proposing source rock types and depositional is the most useful parameter for this fraction. environments based on oil composition, and determining Loss of C1s. aromatic hydro the thermal maturity of the source rock at the time of oil more water-soluble sulfur-containing components, due to eneration water washing has also been proposed. In cases where Most co fraction. Therefore, a greater emphasis has been placed on is lacking, depletion of the C15+ aromatic fraction is understanding how biodegradation affects the saturated attributed to water washing. However, in cases of fraction than the aromatic fraction. Microbial studies, extensive biodegradation, as demonstrated by the destruc- owever, show that the aromatic hydrocarbons can also tion of the saturated hydrocarbons including naphthenic be biodegraded. More case studies need to be performed steranes and pentacyclic terpanes(hopanes), degradation to gain an appreciation for bacterial attack on these of the aromatic fraction may also be due to biodegrada compounds
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
