3. Determining the Fraction Freely Dissolved ( ffd). As illustrated by Equation 5 10, the fraction of the nonionic organic chemical that is freely dissolved in the study water is required for calculating a baseline BAF R f d from a field measured BAF tT . The freely dissolved fraction is the portion of the nonionic organic chemical that is not bound to 5 26 ( Equation 5 12) particulate organic carbon or dissolved organic carbon. Together, the concentration of a nonionic organic chemical that is freely dissolved, bound to dissolved organic carbon, and bound to particulate organic carbon constitute its total concentration in water. As discussed further in the Bioaccumulation TSD, the freely dissolved fraction of a chemical is considered to be the best expression of the bioavailable form of nonionic organic chemicals to aquatic organisms ( e. g., Suffet et al., 1994; USEPA, 1995b). Because the fraction of a nonionic organic chemical that is freely dissolved may vary among different bodies of water as a result of differences in dissolved and particulate organic carbon in the water, the bioavailability of the total chemical concentration in water is expected to vary from one body of water to another. Therefore, BAFs which are based on the freely dissolved concentration in water ( rather than the total concentration in water) are considered to be more reliable for extrapolating and aggregating BAFs among different bodies of water. Currently, availability of BAFs based on measured freely dissolved concentrations is very limited, partly because of difficulties in analytically measuring the freely dissolved concentration. Thus, if a BAF based on the total water concentration is reported in a given study, the fraction of the chemical that is freely dissolved should be predicted using information on the organic carbon content in the study water.

a. Equation for Determining the Freely Dissolved Fraction. If reliable measured data are unavailable to directly determine the freely dissolved fraction of the chemical in water, the freely dissolved fraction should be estimated using the following equation.

where:

POC = concentration of particulate organic carbon ( kg/ L) DOC = concentration of dissolved organic carbon ( kg/ L) K ow = n octanol water partition coefficient for the chemical In Equation 5 12, K ow is being used to estimate the partition coefficient to POC ( i. e., K POC in L/ kg) and 0.08 @ K ow is being used to estimate the partition coefficient to DOC ( i. e., the K DOC in L/ kg). A discussion of the technical basis, assumptions, and uncertainty associated with the derivation and application of Equation 5 12 is provided in the Bioaccumulation TSD.

b. POC and DOC Values. When converting from the total concentration of a chemical to a freely dissolved concentration using Equation 5 12 above, the POC and DOC concentrations should be obtained from the original study from which the field measured BAF is determined. If POC and DOC concentrations are not reported in the BAF study, reliable estimates of POC and DOC might be obtained 5 27 from other studies of the same site used in the BAF study or closely related site( s) within the same water body. When using POC/ DOC data from other studies of the same water body, care should be taken to ensure that environmental and hydrological conditions that might affect POC or DOC concentrations ( i. e., runoff events, proximity to ground water or surface water inputs, sampling season) are reasonably similar to those in the BAF study. Additional information related to selecting POC and DOC values is provided in the Bioaccumulation TSD.

In some cases, BAFs are reported using the concentration of the chemical in filtered or centrifuged water. When converting these BAFs to a freely dissolved basis, the concentration of POC should be set equal to zero when using Equation 5 12. Particulates are removed from water samples by filtering or centrifuging the sample.

c. Selecting Kow Values. A variety of techniques are available to measure or predict K ow values. The reliability of these techniques depends to a large extent on the K ow of the chemical. Because K ow is an important input parameter for calculating the freely dissolved concentration of nonionic organic chemicals and for deriving BAFs using the other three methods of Procedure # 1, care should be taken in selecting the most reliable K ow value. The value of K ow for use in estimating the freely dissolved fraction and other procedures used to derive national BAFs should be selected based on the guidance presented in the Bioaccumulation TSD.

4. Determining the Fraction Lipid ( f R ) . Calculating a baseline BAF R f d for a nonionic organic chemical using Equation 5 10 also requires that the total chemical concentration measured in the tissue used to determine the field measured BAF tT be normalized by the lipid fraction ( f R ) in that same tissue. Lipid normalization of tissue concentrations reflects the assumption that BAFs ( and BCFs) for nonionic organic chemicals are directly proportional to the percent lipid in the tissue upon which they are based. This assumption means that an organism with a two percent lipid content would be expected to accumulate twice the amount of a chemical at steady state compared with an organism with one percent lipid content, all else being equal. The assumption that aquatic organisms accumulate nonionic organic chemicals in proportion to their lipid content has been extensively evaluated in the literature ( Mackay, 1982; Connell, 1988; Barron, 1990) and is generally accepted. Because the lipid content in aquatic organisms can vary both within and across species, BAFs that are expressed using the lipid normalized concentration ( rather than the total concentration in tissue) are considered to be the most reliable for aggregating multiple BAF values for a given species. Additional discussion of technical basis, assumptions, and uncertainties involved in lipid normalization is provided in the Bioaccumulation TSD.

a. The lipid fraction f R , is routinely reported in bioaccumulation studies involving nonionic organic chemicals. If the lipid fraction is not reported in the BAF study, 5 28 ( Equation 5 13) it can be calculated using the following equation if the appropriate data are reported:

where:

M R = Mass of lipid in specified tissue M t = Mass of specified tissue ( wet weight)

b. Because lipid content can vary within an aquatic organism ( and among tissues within that organism) due to several factors including the age and sex of the organism, changes in dietary composition, season of sampling and reproductive status, the lipid fraction used to calculate a baseline BAF R f d should be measured in the same tissue and organisms used to determine the field measured BAF tT , unless comparability is demonstrated across organisms.

c. Experience has shown that different solvent systems used to extract lipids for analytical measurement can result in different quantities of lipids being extracted and measured in aquatic organisms ( e. g., Randall et al., 1991, 1998). As a result, lipid measurements determined using different solvent systems might lead to apparent differences in lipid normalized concentrations and lipid normalized BAFs. The extent to which different solvent systems might affect lipid extractions ( and lipid normalized concentrations) is thought to vary depending on the solvent, chemical of concern, and lipid composition of the tissue being extracted. Guidance on measurement of lipid content, including the choice of solvent system and how different solvent systems may affect lipid content, is provided in the Bioaccumulation TSD.

B. Baseline BAF R f d Derived from BSAFs The second method of determining a baseline BAF R f d for the chemical of concern in Procedure # 1 involves the use of BSAFs. Although BSAFs may be used for measuring and predicting bioaccumulation directly from concentrations of chemicals in surface sediment, they may also be used to estimate BAFs ( USEPA, 1995b; Cook and Burkhard, 1998). Since BSAFs are based on field data and incorporate effects of chemical bioavailability, food web structure, metabolism, biomagnification, growth, and other factors, BAFs estimated from BSAFs will incorporate the net effect of all these factors. The BSAF approach is particularly beneficial for developing water quality criteria for chemicals which are detectable in fish tissues and sediments, but are difficult to detect or measure precisely in the water column. 5 29 As shown by Equation 5 14 below, predicting baseline BAF R f ds using BSAFs requires that certain types of data be used for the chemicals of interest ( for which BAFs are to be determined) and reference chemicals ( for which BAFs are measured) from a common sediment waterorganism data set. Differences between BSAFs for different organic chemicals are good measures of the relative bioaccumulation potentials of the chemicals. When calculated from a common organism sediment sample set, chemical specific differences in BSAFs reflect the net effect of biomagnification, metabolism, food chain, bioenergetics, and bioavailability factors on the degree of each chemical's equilibrium/ disequilibrium between sediment and biota. At equilibrium, BSAFs are expected to be approximately 1.0. However, deviations from 1.0 ( reflecting disequilibrium) are common due to: conditions where water is not at equilibrium with surface sediment; differences in organic carbon content of water and sediment; kinetic limitations for chemical transfer between sediments and water associated with specific biota; biomagnification; or biological processes such as growth or biotransformation. BSAFs are most useful ( i. e., most predictable from one site to another) when measured under steady state ( or near steady state) conditions. The use of non steady state BSAFs, such as found with new chemical loadings or rapid increases in loadings, increases uncertainty in this method for the relative degree of disequilibrium between the reference chemicals and the chemicals of interest. In general, the fact that concentrations of hydrophobic chemicals in sediment are less sensitive than concentrations in water to fluctuations in chemical loading and distribution makes the BSAF method robust for estimating BAFs. Results from validation of the BAF procedure in Lake Ontario, the Fox River and Green Bay, Wisconsin, and the Hudson River, New York, demonstrate good agreement between observed and BSAF predicted BAFs in the vast majority of comparisons made. Detailed results of the validation studies for the BSAF procedure are provided in the Bioaccumulation TSD.

Baseline BAF R f ds should be calculated using acceptable BSAFs for chemicals of interest and appropriate sediment to water fugacity ( disequilibrium) ratios ( J socw) r /( K ow) r for reference chemicals under the following guidelines.

1. Baseline BAF R f d Equation. For each species with an acceptable field measured ( BSAF) I, a baseline BAF R f d for the chemical of interest may be calculated using the following equation with an appropriate value of ( J socw) r /( K ow) r:

( Equation 5 14)

where:

( Baseline BAF R f d) I = BAF expressed on a freely dissolved and lipidnormalized basis for chemical of interest " I" ( BSAF) I = Biota sediment accumulation factor for chemical of interest " I" 5 30 ( Equation 5 15)

( Equation 5 16)

( Equation 5 17) ( J socw) r = sediment organic carbon to water freely dissolved concentration ratio of reference chemical " r" ( K ow) I = octanol water partition coefficient for chemical of interest " I" ( K ow) r = octanol water partition coefficient for the reference chemical " r" D i/ r = ratio between J socw / K ow for chemicals " I" and " r" ( normally chosen so that D i/ r = 1)

The technical basis, assumptions, and uncertainties associated with Equation 5 14 are provided in the Bioaccumulation TSD. Guidance for determining each component of Equation 5 14 is provided below.

2. Determining Field Measured BSAFs. BSAFs should be determined by relating lipidnormalized concentrations of chemicals in an organism ( C R ) to organic carbon normalized concentrations of the chemicals in surface sediment samples ( C soc) using the following equation:

a. Lipid Normalized Concentration. The lipid normalized concentration of a chemical in an organism should be determined by:

where:

C t = Concentration of the chemical in the wet tissue ( either whole organism or specified tissue) ( µ g/ g) f R = Fraction lipid content in the tissue b. Organic Carbon Normalized Concentration. The organic carbon normalized concentration of a chemical in sediment should be determined by: 5 31 where:

C s = Concentration of chemical in sediment ( µ g/ g sediment) f oc = Fraction organic carbon in sediment The organic carbon normalized concentrations of the chemicals in surface sediment samples should be associated with the average exposure environment of the organism.

3. Sediment to Water Partition Coefficient ( J socw) r. Sediment to water partition coefficients for reference chemicals should be determined by:

( Equation 5 18)

where:

( C soc) r = Concentration of a reference chemical in sediment normalized to sediment organic carbon ( C w f d) r = Concentration of the reference chemical freely dissolved in water 4. Selecting Reference Chemicals. Reference chemicals with ( J socw) / ( K ow) similar to that of the chemical of interest are preferred for this method. Theoretically, knowledge of the difference between sediment to water fugacity ratios for two chemicals, " I" and " r" ( D i/ r), could be used when reliable reference chemicals that meet the fugacity equivalence condition are not available. Similarity of ( J socw) / ( K ow) for two chemicals can be indicated on the basis of similar physical chemical behavior in water ( persistence, volatilization), similar mass loading histories, and similar concentration profiles in sediment cores.

Validation studies have demonstrated that choosing reference chemicals with well quantified concentrations in water is important because the uncertainty associated with measurement of barely detected chemicals is large ( see the Bioaccumulation TSD). Similarity between K ow values of the reference and target chemicals is generally desirable, although recent validation studies indicate that the accuracy of the method is not substantially decreased through use of reference chemicals with large differences in K ow , as long as the chemicals are structurally similar and have similar persistence behavior in water and sediments.

5. The following data, procedural, and quality assurance requirements should be met for predicting baseline BAF R f ds using field measured BSAFs: 5 32 a. Data on the reference chemicals and chemicals of interest should come from a common organism water sediment data set at a particular site.

b. The chemicals of interest and reference chemicals should have similar physicochemical properties and persistence in water and sediment.

c. The loadings history of the reference chemicals and chemicals of interest should be similar such that their expected sediment water disequilibrium ratios ( J socw/ K ow) would not be expected to be substantially different ( i. e., D i/ r ~ 1).

d. The use of multiple reference chemicals is generally preferred for determining the value of ( J socw) r so long as the concentrations are well quantified and the aforementioned conditions for selecting reference chemicals are met. In some cases, use of a single reference chemical may be necessary because of limited data.

e. Samples of surface sediments ( 0 1 cm is ideal) should be from locations in which sediment is regularly deposited and is representative of average surface sediment in the vicinity of the organism.

f. The K ow value for the target and reference chemicals should be selected as described in the Bioaccumulation TSD.

g. All other data quality and procedural guidelines described earlier for determining field measured BAFs in Section <IP_ADDRESS>( A) should be met.

Further details on the requirements for predicting BAFs from BSAF measurements, including the data, assumptions, and limitations of this approach are provided in the Bioaccumulation TSD.

C. Baseline BAF R f d from a Laboratory Measured BCF tT and FCM The third method in Procedure # 1 consists of using a laboratory measured BCF tT ( i. e., a BCF based on total concentrations in tissue and water) and FCMs to predict a baseline BAF R f d for the chemical of concern. The BCF tT is used in conjunction with an FCM because non aqueous routes of exposure and subsequent biomagnification is of concern for the types of chemicals applicable to Procedure # 1. A laboratory measured BCF inherently accounts for the effects of chemical metabolism that occurs in the organism used to calculate the BCF, but does not account for metabolism which may occur in other organisms of the aquatic food web.

1. Baseline BAF R f d Equation. For each acceptable laboratory measured BCF tT , calculate a baseline BAF R f d using the following equation: 5 33 ( Equation 5 20) ( Equation 5 19)

where:

Baseline BAF R f d = BAF expressed on a freely dissolved and lipidnormalized basis Measured BCFtT = BCF based on total concentration in tissue and water f R = Fraction of the tissue that is lipid f fd = Fraction of the total chemical in the test water that is freely dissolved FCM = The food chain multiplier either obtained from Table 5 1 by linear interpolation for the appropriate trophic level, or from appropriate field data The technical basis for Equation 5 19 is provided in the Bioaccumulation TSD. Guidance for determining each component of Equation 5 19 is provided below.

2. Determining the Measured BCFt T. The laboratory measured BCF tT shown in Equation 5 19 should be calculated using information on the total concentration of the chemical in the tissue of the organism and the total concentration of the chemical in the laboratory test water. The equation to derive a measured BCF tT is:

where:

C t = Total concentration of the chemical in the specified wet tissue C w = Total concentration of chemical in the laboratory test water The data used to calculate a laboratory measured BCF tT should be reviewed thoroughly to assess the quality of the data and the overall uncertainty in the BCF value. The following general criteria apply in determining the acceptability of laboratory measured BCF tT .

a. The test organism should not be diseased, unhealthy, or adversely affected by the concentration of the chemical because these attributes may alter accumulation of chemicals compared with healthy organisms.

b. The total concentration of the chemical in the water should be measured and should be relatively constant during the exposure period. 5 34 c. The organisms should be exposed to the chemical using a flow through or renewal procedure.

d. The percent lipid of the tissue used to normalize the BCF tT should be either measured or reliably estimated to permit lipid normalization of chemical concentrations.

e. The concentrations of particulate organic carbon and dissolved organic carbon in the study water should be measured or reliably estimated.

f. Aquatic organisms used to calculate a laboratory measured BCF tT should be representative of those aquatic organisms that are commonly consumed in the United States. An aquatic organism which is not commonly consumed in the United States can be used to calculate an acceptable laboratory measured BCF tT provided that the organism is considered to be a reasonable surrogate for a commonly consumed organism. Information on the ecology, physiology, and biology of the organism should be reviewed when assessing whether an organism is a reasonable surrogate of a commonly consumed organism.

g. BCFs may be based on measurement of radioactivity from radiolabeled parent compounds only when the BCF is intended to include metabolites, when there is confidence that there is no interference due to metabolites of the parent compounds, or when studies are conducted to determine the extent of metabolism, thus allowing for a proper correction.

h. The calculation of the BCF tT should appropriately address growth dilution, which can be particularly important in affecting BCF tT determinations for poorly depurated chemicals.

I. Other aspects of the methodology used should be similar to those described by the American Society of Testing and Materials ( ASTM, 1999) and USEPA Ecological Effects Test Guidelines ( USEPA, 1996).

j. In addition, the magnitude of the K ow and the availability of corroborating BCF data should be considered. For example, if the steady state method is used for the BCF tT determination, exposure periods longer than 28 days will generally be required for highly hydrophobic chemicals to reach steady state between the water and the organism.

k. If a baseline BCF R f d derived from a laboratory measured BCFtT consistently increases or decreases as the chemical concentration increases in the test solutions for the test organisms, the BCF tT should be selected from the test concentration( s) that would most closely correspond to the 304( a) criterion. Note: a BCF tT should not be calculated from a control treatment. 5 35 3. Selecting Food Chain Multipliers. An FCM reflects a chemical's tendency to biomagnify in the aquatic food web. Values of FCMs greater than 1.0 are indicative of biomagnification and typically apply to organic chemicals with log K ow values between 4.0 and 9.0. For a given chemical, FCMs tend to be greater at higher trophic levels, although FCMs for trophic level three can be higher than those for trophic level four.

Food chain multipliers used to derive baseline BAF R f ds using Procedure # 1 can be selected from model derived or field derived estimates.

a. Model Derived FCMs. For nonionic organic chemicals appropriate for Procedure # 1, EPA has calculated FCMs for various K ow values and trophic levels using the bioaccumulation model of Gobas ( 1993). The FCMs shown in Table 5 1 were calculated using the Gobas model as the ratio of the baseline BAF R f ds for trophic levels 2, 3, and 4 to the baseline BCF R f d.

EPA recommends using the biomagnification model by Gobas ( 1993) to derive FCMs for nonionic organic chemicals for several reasons. First, the Gobas model includes both benthic and pelagic food chains, thereby incorporating exposure of organisms to chemicals from both the sediment and the water column. Second, the input data needed to run the model can be readily defined. Third, the predicted BAFs using the model are in agreement with field measured BAFs for chemicals, even those with very high log K ow s. Finally, the model predicts chemical residues in benthic organisms using equilibrium partitioning theory, which is consistent with EPA's equilibrium partitioning sediment guidelines ( USEPA, 2000d).

The Gobas model requires input of specific data on the structure of the food chain and the water quality characteristics of the water body of interest. For calculating national BAFs, a mixed pelagic/ benthic food web structure consisting of four trophic levels is assumed. Trophic level 1 is phytoplankton, trophic level 2 is zooplankton, trophic level 3 is forage fish ( e. g., sculpin and smelt), and trophic level 4 are predatory fish ( e. g., salmonids). Additional assumptions are made regarding the composition of the aquatic species' diets ( e. g., salmonids consume 10 percent sculpin, 50 percent alewives, and 40 percent smelt), the physical parameters of the aquatic species ( e. g., lipid values), and the water quality characteristics ( e. g., water temperature, sediment organic carbon).

A mixed pelagic/ benthic food web structure has been assumed for the purpose of calculating FCMs because it is considered to be most representative of the types of food webs that occur in aquatic ecosystems. FCMs derived using the mixed pelagic/ benthic structure are also about mid range in magnitude between a 100% pelagic and 100% benthic driven food web ( see the Bioaccumulation TSD). The validity of FCMs derived using the mixed pelagic/ benthic food web structure has 5 36 Table 5 1 Food Chain Multipliers for Trophic Levels 2, 3 and 4 ( Mixed Pelagic and Benthic Food Web Structure and J socw / KOW = 23)

Log KOW Trophic Level 2 Trophic Level 3 Trophic Level 4 Log KOW Trophic Level 2 Trophic Level 3 Trophic Level 4 4.0 1.00 1.23 1.07 6.6 1.00 12.9 23.8 4.1 1.00 1.29 1.09 6.7 1.00 13.2 24.4 4.2 1.00 1.36 1.13 6.8 1.00 13.3 24.7 4.3 1.00 1.45 1.17 6.9 1.00 13.3 24.7 4.4 1.00 1.56 1.23 7.0 1.00 13.2 24.3 4.5 1.00 1.70 1.32 7.1 1.00 13.1 23.6 4.6 1.00 1.87 1.44 7.2 1.00 12.8 22.5 4.7 1.00 2.08 1.60 7.3 1.00 12.5 21.2 4.8 1.00 2.33 1.82 7.4 1.00 12.0 19.5 4.9 1.00 2.64 2.12 7.5 1.00 11.5 17.6 5.0 1.00 3.00 2.51 7.6 1.00 10.8 15.5 5.1 1.00 3.43 3.02 7.7 1.00 10.1 13.3 5.2 1.00 3.93 3.68 7.8 1.00 9.31 11.2 5.3 1.00 4.50 4.49 7.9 1.00 8.46 9.11 5.4 1.00 5.14 5.48 8.0 1.00 7.60 7.23 5.5 1.00 5.85 6.65 8.1 1.00 6.73 5.58 5.6 1.00 6.60 8.01 8.2 1.00 5.88 4.19 5.7 1.00 7.40 9.54 8.3 1.00 5.07 3.07 5.8 1.00 8.21 11.2 8.4 1.00 4.33 2.20 5.9 1.00 9.01 13.0 8.5 1.00 3.65 1.54 6.0 1.00 9.79 14.9 8.6 1.00 3.05 1.06 6.1 1.00 10.5 16.7 8.7 1.00 2.52 0.721 6.2 1.00 11.2 18.5 8.8 1.00 2.08 0.483 6.3 1.00 11.7 20.1 8.9 1.00 1.70 0.320 6.4 1.00 12.2 21.6 9.0 1.00 1.38 0.210 6.5 1.00 12.6 22.8 been evaluated in several different ecosystems including Lake Ontario, the tidally influenced Bayou D'Inde in Louisiana, the Fox River and Green Bay, Wisconsin, and the Hudson River in New York. Additional details of the validation of EPA's national default FCMs and the assumptions, uncertainties, and input parameters for the model are provided in the Bioaccumulation TSD. 5 37 Although EPA uses the FCMs in Table 5 1 to derive its national 304( a) criteria, EPA recognizes that food webs of other waterbodies might differ from the assumptions used to calculate national BAFs. In these situations, States and authorized Tribes may wish to use alternate food web structures for calculating FCMs for use in setting State or Tribal water quality criteria. Additional guidance on the use of alternate food web structures for calculating State, Tribal, or sitespecific criteria is provided in the Bioaccumulation TSD.

b. Field Derived FCMs. In addition to model derived estimates of FCMs, field data may also be used to derive FCMs. Currently, the use of field derived FCMs is the only method recommended for estimating FCMs for inorganic and organometalic chemicals because appropriate model derived estimates are not yet available ( see Section 5.6). In contrast to the model based FCMs described previously, fieldderived FCMs account for any metabolism of the chemical of concern by the aquatic organisms used to calculate the FCM.

Field derived FCMs should be calculated using lipid normalized concentrations of the nonionic organic chemical in appropriate predator and prey species using the following equations.

FCM TL2 = BMF TL2 ( Equation 5 21)

FCM TL3 = ( BMF TL3) ( BMF TL2) ( Equation 5 22)

FCM TL4 = ( BMF TL4) ( BMF TL3) ( BMF TL2) ( Equation 5 23)

where:

FCM = Food chain multiplier for designated trophic level ( TL2, TL3, or TL4) BMF = Biomagnification factor for designated trophic level ( TL2, TL3, or TL4)

The basic difference between FCMs and BMFs is that FCMs relate back to trophic level one ( or trophic level two as assumed by the Gobas ( 1993) model), whereas BMFs always relate back to the next lowest trophic level. For nonionic organic chemicals, BMFs can be calculated from tissue residue concentrations determined in biota at a site according to the following equations.

BMF TL2 = ( C R , TL2) / ( C R , TL1) ( Equation 5 24)

BMF TL3 = ( C R , TL3) / ( C R , TL2) ( Equation 5 25)

BMF TL4 = ( C R , TL4) / ( C R , TL3) ( Equation 5 26) 5 38 where: C R = Lipid normalized concentration of chemical in tissue of appropriate biota that occupy the specified trophic level ( TL2, TL3, or TL4)

In addition to the acceptability guidelines pertaining to field measured BAFs, the following procedural and quality assurance requirements apply to field measured FCMs.

( 1) Information should be available to identify the appropriate trophic levels for the aquatic organisms and appropriate predator prey relationships for the site from which FCMs are being determined. General information on determining trophic levels of aquatic organisms can be found in USEPA 2000a, b, c.

( 2) The aquatic organisms sampled from each trophic level should reflect the most important exposure pathways leading to human exposure via consumption of aquatic organisms. For higher trophic levels ( e. g., 3 and 4), aquatic species should also reflect those that are commonly consumed by humans.

( 3) The studies from which the FCMs are derived should contain sufficient supporting information from which to determine that tissue samples were collected and analyzed using appropriate, sensitive, accurate, and precise methods.

( 4) The percent lipid should be either measured or reliably estimated for the tissue used to determine the FCM.

( 5) The tissue concentrations should reflect average exposure over the approximate time required to achieve steady state in the target species.

D. Baseline BAF R f d from a Kow and FCM The fourth method in Procedure # 1 consists of using a K ow and an appropriate FCM for estimating the baseline BAF R f d. In this method, the K ow is assumed to be equal to the baseline BCF R f d. Numerous investigations have demonstrated a linear relationship between the logarithm of the BCF and the logarithm of the octanol water partition coefficient ( K ow ) for organic chemicals for fish and other aquatic organisms. Isnard and Lambert ( 1988) list various regression equations that illustrate this linear relationship. When the regression equations are constructed using lipidnormalized BCFs, the slopes and intercepts are not significantly different from one and zero, respectively ( e. g., de Wolf, et al., 1992). The underlying assumption for the linear relationship between the BCF and K ow is that the bioconcentration process can be viewed as the partitioning of a chemical between the lipid of the aquatic organisms and water and that the K ow is a useful 5 39 ( Equation 5 27) surrogate for this partitioning process ( Mackay, 1982). To account for biomagnification, Procedure # 1 requires the K ow value be used in conjunction with an appropriate FCM.

1. Baseline BAF R f d Equation. For each acceptable K ow value and FCM for the chemical of concern, calculate a baseline BAF R f d using the following equation.

where:

Baseline BAF R f d = BAF expressed on a freely dissolved and lipid normalized basis for a given trophic level FCM = The food chain multiplier for the appropriate trophic level obtained from Table 5 1 by linear interpolation or from appropriate field data ( used with Procedure # 1 only) K ow = Octanol water partition coefficient The BCF K ow relationship has been developed primarily for nonionic organic chemicals that are not readily metabolized by aquatic organisms and thus is most appropriate for poorly metabolized nonionic organic chemicals ( i. e., Procedures # 1 and # 3 as depicted in Figure 5 1). For poorly metabolized nonionic organic chemicals with large log K ow s ( i. e., > 6), reported log BCFs are often not equal to log K ow. EPA believes that this nonlinearity is primarily due to not accounting for several factors which affect the BCF determination. These factors include not basing BCFs on the freely dissolved concentration in water, not accounting for growth dilution, not assessing BCFs at steady state, inaccuracies in measurements of uptake and elimination rate constants, and complications from the use of solvent carriers in the exposure. Application of Equation 5 27 for predicting BAFs has been conducted in several different ecosystems including Lake Ontario, the tidally influenced Bayou D'Inde in Louisiana, the Fox River and Green Bay, Wisconsin, and the Hudson River in New York. Additional detail on the validation, technical basis, assumptions, and uncertainty associated with Equation 5 27 and is provided in the Bioaccumulation TSD.

2. FCMs and Kows. Food chain multipliers and K ow values should be selected as described previously in Procedure # 1.

<IP_ADDRESS> Selecting Final Baseline BAF R f ds After calculating individual baseline BAF R f ds using as many of the methods in Procedure # 1 as possible, the next step is to determine a final baseline BAF R f d for each trophic level from the individual baseline BAF R f ds ( see Figures 5 1 and 5 2). The final baseline BAF R f d will be used in the 5 40 last step to determine the national BAF for each trophic level. The final baseline BAF R f d for each trophic level should be determined from the individual baseline BAF R f ds by considering the data preference hierarchy defined by Procedure # 1 and uncertainty in the data. The data preference hierarchy for Procedure # 1 is ( in order of preference):

1. a baseline BAF R f d from an acceptable field measured BAF ( method 1) 2. a baseline BAF R f d predicted from an acceptable field measured BSAF ( method 2), 3. a baseline BAF R f d predicted from an acceptable BCF and FCM ( method 3), or 4. a baseline BAF R f d predicted from an acceptable K ow and FCM ( method 4).

This data preference hierarchy reflects EPA's preference for BAFs based on field measurements of bioaccumulation ( methods 1 and 2) over those based on laboratory measurements and/ or predictions of bioaccumulation ( methods 3 and 4). However, this data preference hierarchy should not be considered inflexible. Rather, it should be used as a guide for selecting the final baseline BAF R f ds when the uncertainty is similar among two or more baseline BAF R f ds derived using different methods. The following steps and guidelines should be followed for selecting the final baseline BAF R f ds using Procedure # 1.

1. Calculate Species Mean Baseline BAF R f ds. For each BAF method where more than one acceptable baseline BAF R f d is available for a given species, calculate a species mean baseline BAF R f d as the geometric mean of all available individual baseline BAF R f ds. When calculating a species mean baseline BAF R f d, individual baseline BAF R f ds should be reviewed carefully to assess the uncertainty in the BAF values. For highly hydrophobic chemicals applicable to Procedure # 1, particular attention should be paid to whether sufficient spatial and temporal averaging of water and tissue concentrations was likely achieved in the BAF, BSAF, or BCF study. Highly uncertain baseline BAF R f ds should not be used. Large differences in individual baseline BAF R f ds for a given species ( e. g., greater than a factor of 10) should be investigated further. In such cases, some or all of the baseline BAF R f ds for a given species might not be used. Additional discussion on evaluating acceptability of BAF values is provided in the Bioaccumulation TSD.

2. Calculate Trophic Level Mean Baseline BAF R f ds. For each BAF method where more than one acceptable species mean baseline BAF R f d is available within a given trophic level, calculate a trophic level mean baseline BAF R f d as the geometric mean of acceptable species mean baseline BAF R f ds in that trophic level. Trophic level mean baseline BAF R f ds should be calculated for trophic levels two, three, and four because available data on U. S. consumers of fish and shellfish indicate significant consumption of organisms in these trophic levels.

3. Select a Final Baseline BAF R f d for Each Trophic Level. For each trophic level, select the final baseline BAF R f d using best professional judgment by considering: ( 1) the data preference hierarchy shown previously, ( 2) the relative uncertainty in the trophic levelmean baseline BAF R f ds derived using different methods, and ( 3) the weight of evidence among the four methods. 5 41 ( Equation 5 28) a. In general, when more than one trophic level mean baseline BAF R f d is available for a given trophic level, the final trophic level mean baseline BAF R f d should be selected from the most preferred BAF method defined by the data preference hierarchy for Procedure # 1.

b. If uncertainty in a trophic level mean baseline BAF based on a higher tier ( more preferred) method is judged to be substantially greater than a trophic level mean baseline BAF from a lower tier method, and the weight of evidence among the various methods suggests that a BAF value from lower tier method is likely to be more accurate, then the final baseline BAF R f d should be selected using a trophic level mean baseline BAF R f d from a lower tier method.

c. When considering the weight of evidence among the various BAF methods, greater confidence in the final baseline BAF R f d is generally assigned when BAFs from a greater number of methods are in agreement for a given trophic level. However, lack of agreement among methods does not necessarily indicate less confidence if such disagreements can be adequately explained. For example, if the chemical of concern is metabolized by aquatic organisms represented by a BAF value, one would expect disagreement between a field measured BAF ( the highest priority data) and a predicted BAF using a K ow and model derived FCM. Thus, field measured BAFs should generally be given the greatest weight among methods because they reflect direct measures of bioaccumulation and incorporate any metabolism which might occur in the organism and its food web.

d. The above steps should be performed for each trophic level until a final baseline BAF R f d is selected for trophic levels two, three, and four.

<IP_ADDRESS> Calculating National BAFs The last step in deriving a national BAF for each trophic level is to convert the final baseline BAF R f d determined in the previous step to a BAF that reflects conditions to which the national 304( a) criteria will apply ( Figure 5 2). Since a baseline BAF R f d is by definition normalized by lipid content and expressed on a freely dissolved basis, it needs to be adjusted to reflect the lipid fraction of aquatic organisms commonly consumed in the U. S. and the freely dissolved fraction expected in U. S. bodies of water. Converting a final baseline BAF R f d to a national BAF requires information on: ( 1) the percent lipid of the aquatic organisms commonly consumed by humans, and ( 2) the freely dissolved fraction of the chemical of concern that would be expected in the ambient waters of interest. For each trophic level, a national BAF should be determined from a final baseline BAF R f d according to the following guidelines.

1. National BAF Equation. For each trophic level, calculate a national BAF using the following equation. 5 42 where:

Final Baseline BAF R f d = Final trophic level mean baseline BAF expressed on a freely dissolved and lipid normalized basis for trophic level " n" f R ( TL n) = Lipid fraction of aquatic species consumed at trophic level " n" f fd = Fraction of the total chemical in water that is freely dissolved The technical basis of Equation 5 28 is provided in the Bioaccumulation TSD. Guidance for determining each component of Equation 5 28 is provided below.

2. Determining the Final Baseline BAF R f d. The final trophic level mean baseline BAF R f ds used in this equation are those which have been determined using the guidance presented in Section <IP_ADDRESS> for selecting the final baseline BAF R f ds.

3. Lipid Content of Commonly Consumed Aquatic Species. As illustrated by Equation 5 28, the percent lipid of the aquatic species consumed by humans is needed to accurately characterize the potential exposure to a chemical from ingestion of aquatic organisms.

a. National Default Lipid Values. For the purposes of calculating a national 304( a) criterion, the following national default values for lipid fraction should be used: 1.9% ( for trophic level two organisms), 2.6% ( for trophic level three organisms), and 3.0% ( for trophic level four organisms).

These national default values for lipid content reflect national per capita average patterns of fish consumption in the United States. Specifically, they were calculated using the consumption weighted mean lipid content of commonly consumed fish and shellfish as identified by the USDA Continuing Survey of Food Intake by Individuals ( CSFII) for 1994 through 1996. This same national survey data was used to derive national default values of fish consumption. To maintain consistency with the fish consumption assumptions, only freshwater and estuarine organisms were included in the derivation of the national default lipid values. Additional details on the technical basis, assumptions, and uncertainty in the national default values of lipid fraction are provided in the Bioaccumulation TSD.

Although national default lipid values are used by EPA to set national 304( a) criteria, EPA encourages States and authorized Tribes to use local or regional data on lipid content of consumed aquatic species when adopting criteria into their water quality standards because local or regional consumption patterns ( and lipid content) can differ from national consumption patterns. Additional guidance on 5 43 ( Equation 5 29) developing site specific values of lipid content, including a database of lipid content for many commonly consumed aquatic organisms, is found in the Bioaccumulation TSD.

4. Freely Dissolved Fraction. The third piece of information required for deriving a national BAF is the freely dissolved fraction of the chemical of concern that is expected in waters of the United States. As noted previously, expressing BAFs on the freely dissolved concentration in water allows a common basis for averaging BAFs from several studies. However, for use in criteria development, these BAFs should be converted back to values based on the total concentration in the water to be consistent with monitored water column and effluent concentrations, which are typically based on total concentrations of chemicals in the water. This should be done by multiplying the freely dissolved baseline BAF R f d by the fraction of the freely dissolved chemical expected in water bodies of the United States where criteria are to be applied, as shown in Equation 5 29.

where:

POC = national default value for the particulate organic carbon concentration ( kg/ L) DOC = national default value for the dissolved organic carbon concentration ( kg/ L) K ow = n octanol water partition coefficient for the chemical Equation 5 29 is identical to Equation 5 12, which was used to determine the freely dissolved fraction for deriving baseline BAF R f ds from field measured BAFs. However, the POC and DOC concentrations used in Equation 5 29 reflect those values that are expected in U. S. bodies of water, not the POC and DOC values in the study water used to derive the BAF. Guidance for determining each component of Equation 5 29 follows.

a. National Default Values of POC and DOC. For estimating the freely dissolved fraction of the chemical of concern that is expected in U. S. water bodies, national default values of 0.5 mg/ L ( 5 × 10 7 kg/ L) for POC and 2.9 mg/ L ( 2.9 × 10 6 kg/ L) for DOC should be used. These values are 50th percentile values ( medians) based on an analysis of over 110,000 DOC values and 85,000 POC values contained in EPA's STORET database from 1980 through 1999. These default values reflect a combination of values for streams, lakes and estuaries across the United States. Additional details on the technical basis, assumptions, and uncertainty in the 5 44 derivation and application of the national default values of POC and DOC are provided in the Bioaccumulation TSD.

Although national default values of POC and DOC concentrations are used by EPA to set national 304( a) criteria as described by this document, EPA encourages States and authorized Tribes to use local or regional data on POC and DOC when adopting criteria into their water quality standards. EPA encourages States and Tribes to consider local or regional data on POC and DOC because local or regional conditions may result in differences in POC or DOC concentrations compared with the values used as national defaults. Additional guidance on developing local or regional values of POC and DOC, including a database of POC and DOC values segregated by waterbody type, is found in the Bioaccumulation TSD.

b. KowValue. The value selected for the K ow of the chemical of concern should be the same value used in earlier calculations ( e. g., for calculating baseline BAF R f ds and FCMs). Guidance for selecting the K ow value is found in the Bioaccumulation TSD.

5.4.4 Deriving National BAFs Using Procedure # 2 This section provides guidance for calculating national BAFs for nonionic organic chemicals using Procedure # 2 shown in Figure 5 1. The types of nonionic organic chemicals for which Procedure # 2 is most appropriate are those that are classified as moderately to highly hydrophobic and subject to high rates of metabolism by aquatic biota ( see Section 5.4.2 above). Non aqueous contaminant exposure and subsequent biomagnification in aquatic food webs are not generally of concern for chemicals that are classified in this category. As a result, FCMs are not used in this procedure. In addition, K ow  based predictions of bioconcentration are not used in this procedure since the K ow / BCF relationship is primarily based on poorly metabolized chemicals. Some nonionic organic chemicals for which Procedure # 2 is probably appropriate include certain PAHs which are believed to be metabolized substantially by fish ( e. g., benzo[ a] pyrene, phenanthrene, fluoranthene, pyrene, benzo[ a] anthracene and chrysene/ triphenylene; USEPA, 1980; Burkhard and Lukasewycz, 2000).

According to Procedure # 2, the following three methods can be used in deriving a national BAF:

C using a BAF from an acceptable field study ( i. e., a field measured BAF) ( method 1), C predicting a BAF from an acceptable BSAF ( method 2), and C predicting a BAF from an acceptable BCF ( method 3).

Each of these three methods relies on measured data for assessing bioaccumulation and therefore, includes the effects of chemical metabolism by the study organism in the BAF estimate. 5 45 The field measured BAF and BSAF methods also incorporate any metabolism which occurs in the aquatic food web.

As shown in Figure 5 2, the next steps in deriving a national BAF after selecting the derivation procedure are: ( 1) calculating individual baseline BAF R f ds, ( 2) selecting the final baseline BAF R f ds, and ( 3) calculating the national BAFs. Each of these three steps is discussed separately below.

<IP_ADDRESS> Calculating Individual Baseline BAF R f ds As described previously in Procedure # 1, calculating individual baseline BAF R f ds involves normalizing the measured BAFtT or BCF tT ( which are based on the total chemical in water and tissue) by the lipid content of the study organisms and the freely dissolved fraction of the chemical in the study water. Converting measured BAF tT ( or BCF tT ) values to baseline BAF R f d ( or BCF R f d) values is designed to account for variation in measured BAF tT s that is caused by differences in lipid content of study organisms and differences in the freely dissolved fraction of chemical in study waters. Therefore, baseline BAF R f ds are considered more amenable for extrapolating and averaging BAFs across different species and different study waters compared with total BAF tT s.

1. For each species where acceptable data are available, calculate all possible baseline BAF R f ds using each of the three methods shown above for Procedure # 2.

2. Individual baseline BAF R f ds should be calculated from field measured BAF tT s, fieldmeasured BSAFs, and laboratory BCF tT s according to the following procedures.

A. Baseline BAF R f d from Field Measured BAFs 1. Except where noted below, a baseline BAF R f d should be calculated from a field measured BAFtT using the guidance and equations outlined in Section <IP_ADDRESS>( A) for determining baseline BAF R f ds from field measured BAFs in Procedure # 1.

2. Because nonionic organic chemicals applicable to Procedure # 2 have relatively high rates of metabolism in aquatic organisms, they will tend to reach steady state more quickly than nonionic organic chemicals with similar K ow values but which undergo little or no metabolism. Therefore, less temporal averaging of chemical concentrations would generally be required for determining field measured BAF tT s with highly metabolizable chemicals compared with chemicals that are poorly metabolized by aquatic biota. 5 46 B. Baseline BAF R f d Derived from Field measured BSAFs 1. A baseline BAF R f d should be calculated from a field measured BSAF using the guidance and equations outlined in Section <IP_ADDRESS>( B) for determining baseline BAF R f ds from fieldmeasured BSAFs in Procedure # 1.

C. Baseline BAF R f d from a Laboratory Measured BCF 1. Except where noted below, a baseline BAF R f d should be calculated from a laboratorymeasured BCFtT using the guidance and equations outlined in Section <IP_ADDRESS>( c) for determining baseline BAF R f ds from a laboratory measured BCF and FCM in Procedure # 1.

2. Because biomagnification is not an overriding concern for nonionic organic chemicals applicable to Procedure # 2, food chain multipliers are not used in the derivation of a baseline BAF R f d from a laboratory measured BCF tT .

<IP_ADDRESS> Selecting Final Baseline BAF R f ds After calculating individual, baseline BAF R f ds using as many of the methods in Procedure # 2 as possible, the next step is to determine a final baseline BAF R f d for each trophic level from the individual baseline BAF R f ds. The final baseline BAF R f d will be used in the last step to determine the national BAF for each trophic level. A final baseline BAF R f d for each trophic level should be determined from the individual baseline BAF R f ds by considering the data preference hierarchy defined by Procedure # 2 and uncertainty in the data. The data preference hierarchy for Procedure # 2 is ( in order of preference):

1. a baseline BAF R f d from an acceptable field measured BAF ( method 1), 2. a baseline BAF R f d from an acceptable field measured BSAF ( method 2), or 3. a baseline BAF R f d from an acceptable laboratory measured BCF ( method 3).

This data preference hierarchy reflects EPA's preference for BAFs based on fieldmeasurements of bioaccumulation ( methods 1 and 2) over those based on laboratorymeasurements ( method 3). However, as explained in Procedure # 1, this data preference hierarchy should not be considered inflexible. Rather, it should be used as a guide for selecting the final baseline BAF R f ds when the underlying uncertainty is similar among two or more baseline BAF R f ds derived using different methods. Although biomagnification is not generally a concern for chemicals subject to Procedure # 2, trophic level differences in bioaccumulation might be substantial to the extent that the rate of chemical metabolism by organisms in different trophic levels differs. For example, certain PAHs have been shown to be metabolized to a much greater extent by some fish compared with some invertebrate species ( James, 1989). Therefore, final baseline BAF R f ds for chemicals applicable to Procedure # 2 should be determined on a trophic levelspecific basis according to the following guidelines. 5 47 1. The final baseline BAF R f ds in Procedure # 2 should be selected according to the same steps described in Procedure # 1 but with the substitution of the data preference hierarchy described above for Procedure # 2. Specifically, the species mean baseline BAF R f ds, trophic level mean baseline BAF R f ds, and the final baseline BAF R f ds should be determined according to the guidelines presented in Procedure # 1 ( Section <IP_ADDRESS>, Steps 1, 2, and 3).

<IP_ADDRESS> Calculating the National BAFs As described in Procedure # 1, the last step in deriving national BAFs for nonionic organic chemicals is to convert the final baseline BAF R f ds determined in the previous step to BAFs which reflect conditions to which the national 304( a) criteria will apply ( Figure 5 2).

1. For trophic levels two, three, and four, national BAFs should be calculated from the final baseline BAF R f ds using the same equation and procedures described previously in Procedure # 1 ( see Section <IP_ADDRESS> entitled " Calculating the National BAFs").

5.4.5 Deriving National BAFs Using Procedure # 3 This section provides guidance for calculating national BAFs for nonionic organic chemicals using Procedure # 3 shown in Figure 5 1. The types of nonionic organic chemicals for which Procedure # 3 is most appropriate are those that are classified as low in hydrophobicity ( i. e., log K ow values less than 4.0) and subject to low ( or unknown) rates of metabolism by aquatic biota ( see Section 5.4.2 above). Non aqueous contaminant exposure and subsequent biomagnification in aquatic food webs are not generally of concern for chemicals that are classified in this category ( Fisk et al., 1998; Gobas et al., 1993; Connolly and Pedersen, 1988; Thomann, 1989). As a result, FCMs are not used in this procedure.

According to Procedure # 3, the following three methods can be used in deriving a national BAF:

C using a BAF from an acceptable field study ( i. e., a field measured BAF), C predicting a BAF from an acceptable laboratory measured BCF, and C predicting a BAF from an acceptable K ow.

After selecting the derivation procedure, the next steps in deriving a national BAF at a given trophic level for nonionic organic chemicals are: ( 1) calculating individual baseline BAF R f ds, ( 2) selecting the final baseline BAF R f d, and ( 3) calculating the national BAF ( Figure 5 2). Each of these three steps is discussed separately below.

<IP_ADDRESS> Calculating Individual Baseline BAF R f ds Calculating individual baseline BAF R f ds involves normalizing each measured BAF tT or BCF tT ( which are based on the total chemical in water and tissue) by the lipid content of the study organism and the freely dissolved fraction of the chemical in the study water. For additional 5 48 discussion of the technical basis for calculating baseline BAF R f ds, see Section <IP_ADDRESS> in Procedure # 1.

1. For each species where acceptable data are available, calculate all possible baseline BAF R f ds using each of the three methods shown above for Procedure # 3.

2. An individual baseline BAF R f d should be calculated from field measured BAF tT s, laboratorymeasured BCFtT s, and K ow values according to the following procedures.

A. Baseline BAF R f d from Field Measured BAFs 1. Except where noted below, a baseline BAF R f d should be calculated from a field measured BAFtT using the guidance and equations outlined in Section <IP_ADDRESS>( A) in Procedure # 1.

2. Freely Dissolved Fraction. Due to their low hydrophobicity ( i. e., log K ow < 4.0), nonionic organic chemicals applicable to Procedure # 3 are expected to remain almost entirely in the freely dissolved form in natural waters with dissolved and particulate organic carbon concentrations typical of most field BAF studies. Therefore, the freely dissolved fraction should be assumed to be equal to 1.0, unless the concentrations of DOC and POC are very high in the field BAF study. For studies with very high DOC or POC concentrations, ( e. g., about 100 mg/ L or higher for DOC or 10 mg/ L or higher for POC), the freely dissolved fraction may be substantially lower than 1.0 and therefore should be calculated using Equation 5 12.

3. Temporal Averaging of Concentrations. Also due to their low hydrophobicity, nonionic organic chemicals appropriate to Procedure # 3 will also tend to reach steady state quickly compared with those chemicals to which Procedure # 1 applies. Therefore, the extent of temporal averaging of tissue and water concentrations is typically much less than that required for highly hydrophobic chemicals to which Procedure # 1 is applied. In addition, field studies used to calculate BAFs for these chemicals should have sampled water and tissue at similar points in time because tissue concentrations respond more rapidly to changes in water concentrations. EPA will be providing additional guidance on appropriate BAF study designs for nonionic organic chemicals ( including those appropriate to Procedure # 3) in its forthcoming guidance document on conducting field BAF and BSAF studies.

B. Baseline BAF R f d from a Laboratory Measured BCF 1. Except where noted below, a baseline BAF R f d should be calculated from a laboratorymeasured BCFtT using the guidance and equations outlined in Section <IP_ADDRESS>( c) of Procedure # 1. 5 49 2. Food Chain Multipliers. Because biomagnification is not an overriding concern for the minimally hydrophobic chemicals applicable to Procedure # 3, FCMs are not used in the derivation of a baseline BAF R f d from a laboratory measured BCF tT .

3. Freely Dissolved Fraction. Due to their low hydrophobicity ( i. e., log K ow < 4.0), nonionic organic chemicals to which Procedure # 3 is applied are expected to remain almost entirely in the freely dissolved form in waters containing dissolved and particulate organic carbon concentrations typical of laboratory BCF studies. Therefore, the freely dissolved fraction should usually be assumed equal to 1.0. The freely dissolved fraction will be substantially less than 1.0 only in situations where unusually high concentrations of DOC and POC are present in the laboratory BCF study ( e. g., above about 100 mg/ L for DOC or about 10 mg/ L for POC). In this situation, the freely dissolved fraction should be calculated according to Equation 5 12.

C. Baseline BAF R f d from a Kow 1. Except where noted below, a baseline BAF R f d should be calculated from an acceptable K ow using the guidance and equations outlined in Section <IP_ADDRESS>( D) in Procedure # 1.

2. Because biomagnification is not an overriding concern for nonionic organic chemicals with low hydrophobicity ( i. e., log K ow < 4.0), food chain multipliers are not used in Procedure # 3 for deriving the baseline BAF R f d from a K ow.

<IP_ADDRESS> Selecting Final Baseline BAF R f ds After calculating individual baseline BAF R f ds using as many of the methods in Procedure # 3 as possible, the next step is to determine a final baseline BAF R f d for each trophic level from the individual baseline BAF R f ds ( Figure 5 2). The final baseline BAF R f d will be used in the last step to determine the national BAF for each trophic level. The final baseline BAF R f d for each trophic level should be determined from the individual baseline BAF R f ds by considering the data preference hierarchy defined by Procedure # 3 and uncertainty in the data. The data preference hierarchy for Procedure # 3 is ( in order of preference):

1. a baseline BAF R f d from an acceptable field measured BAF or laboratorymeasured BCF, or 2. a baseline BAF R f d predicted from an acceptable K ow value.

This data preference hierarchy reflects EPA's preference for BAFs that are based on measured data ( field measured BAFs and laboratory measured BCFs) over BAFs based on predictive methods ( K ow). This data preference hierarchy should be used as a guide for selecting the final baseline BAF R f ds when the uncertainty is similar among two or more baseline BAF R f ds derived using different methods. Since bioaccumulation via dietary uptake and subsequent biomagnification generally are not of concern for chemicals subject to Procedure # 3, field 5 50 measured BAFs and laboratory measured BCFs are considered equally in determining the national BAF.

Final baseline BAF R f ds should be selected for each trophic level using the following steps and guidelines.

1. Calculate Species Mean Baseline BAF R f ds. For each BAF method ( i. e., field measured BAF, BAF from a lab measured BCF, or BAF from a K ow) where more than one acceptable baseline BAF R f d is available for a given species, calculate a species mean baseline BAF R f d according to the guidance described previously in Procedure # 1.

2. Calculate Trophic Level Mean Baseline BAF R f ds. For each BAF method where more than one acceptable species mean baseline BAF R f d is available within a given trophic level, calculate the trophic level mean baseline BAF R f d as the geometric mean of acceptable species mean baseline BAF R f ds in that trophic level.

3. Select a Final Baseline BAF R f d for Each Trophic Level. For each trophic level, select the final baseline BAF R f d using best professional judgment by considering: ( 1) the data preference hierarchy, ( 2) the relative uncertainties among trophic level mean baseline BAF R f ds derived using different methods, and ( 3) the weight of evidence among the three methods.

a. In general, when more than one trophic level mean baseline BAF R f d is available within a given trophic level, the final baseline BAF R f d should be selected from the most preferred BAF method defined by the data preference hierarchy for Procedure # 3. Within the first data preference tier, field measured BAFs and laboratory measured BCFs are considered equally desirable for deriving a final trophic level mean baseline BAF R f d using Procedure # 3. If a trophic level mean baseline BAF R f d is available from both a field measured BAF and a laboratorymeasured BCF, the final baseline BAF R f d should be selected using the trophic levelmean baseline BAF R f d or BCF R f d with the least overall uncertainty.

b. If uncertainty in a trophic level mean baseline BAF R f d based on a higher tier ( more preferred) method is judged to be substantially greater than a trophic level mean baseline BAF R f d from a lower tier method, then the final baseline BAF R f d should be selected using a trophic level mean baseline BAF R f d from a lower tier method.

c. The above steps should be performed for each trophic level until a final baseline BAF R f d is selected for trophic level two, three, and four.

<IP_ADDRESS> Calculating the National BAFs As described in Procedure # 1, the last step in deriving a national BAF for a given trophic level for nonionic organic chemicals is to convert the final baseline BAF R f d determined in the 5 51 previous step to a BAF that reflect conditions to which the national 304( a) criterion will apply ( Figure 5 2). Each national BAF should be determined from a final baseline BAF R f d according to the following guidelines.

1. National BAF Equation. Except where noted below, national BAFs for trophic levels two, three, and four should be calculated from the final, trophic level mean baseline BAF R f ds using Equation 5 28 and associated guidance described in Procedure # 1 ( see Section <IP_ADDRESS>).

2. Freely Dissolved Fraction. Due to their low hydrophobicity ( i. e., log K ow < 4.0), a freely dissolved fraction of 1.0 should be assumed for calculating national BAFs for nonionic organic chemicals using Procedure # 3. A freely dissolved fraction of 1.0 should be assumed because at a log K ow of less than 4.0, nonionic organic chemicals are expected to remain over 99 percent in the freely dissolved form at POC and DOC concentrations corresponding to national default values for U. S. bodies of water ( i. e., 0.5 mg/ L and 2.9 mg/ L, respectively).

5.4.6 Deriving National BAFs Using Procedure # 4 This section provides guidance for calculating national BAFs for nonionic organic chemicals using Procedure # 4 shown in Figure 5 1. The types of nonionic organic chemicals for which Procedure # 4 is most appropriate are those that are classified as having low hydrophobicity and subject to high rates of metabolism by aquatic biota ( see Section 5.4.2 above). Non aqueous contaminant exposure and subsequent biomagnification in aquatic food webs are not generally of concern for chemicals that are classified in this category. As a result, FCMs are not used in this procedure. In addition, K ow  based predictions of bioconcentration are not used in this procedure since the K ow / BCF relationship is primarily based on poorly metabolized chemicals. One example of a nonionic organic chemical for which Procedure # 4 appears appropriate is butyl benzyl phthalate in fish. Using radiolabeling techniques with confirmation by chromatographic analysis, Carr et al. ( 1997) present evidence that indicates butyl benzyl phthalate is extensively metabolized in sunfish. Carr et al. ( 1997) also report measured BCFs ( and subsequently lipid normalized BCFs) which are substantially below predicted BCFs based on log K ow. In a study of chlorinated anilines ( which would be essentially un ionized at ambient pH), de Wolf et al. ( 1992) reported measured BCFs substantially lower than those predicted based on K ow. The authors suggested that biotransformation ( metabolism) involving the amine ( NH 2) was responsible for the lower measured BCFs.

According to Procedure # 4, the following two methods can be used in deriving a national BAF:

C using a BAF from an acceptable field study ( i. e., a field measured BAF), and C predicting a BAF from an acceptable BCF. 5 52 After selecting the derivation procedure, the next steps in deriving a national BAF for a given trophic level for nonionic organic chemicals are: ( 1) calculating individual baseline BAF R f ds, ( 2) selecting the final baseline BAF R f d, and ( 3) calculating the national BAF ( Figure 5 2). Each of these three steps is discussed separately below.

<IP_ADDRESS> Calculating Individual Baseline BAF R f ds Calculating individual baseline BAF R f ds involves normalizing the measured BAF tT or BCF tT ( which are based on the total chemical in water and tissue) by the lipid content of the study organism and the freely dissolved fraction of the chemical in the study water. For additional discussion of the technical basis for calculating baseline BAF R f ds, see Section <IP_ADDRESS> in Procedure # 1.

1. For each species where acceptable data are available, calculate all possible baseline BAF R f ds using each of the two methods shown above for Procedure # 4.

2. Individual baseline BAF R f ds should be calculated from field measured BAF tT s and laboratory measured BCF tT s according to the following procedures.

A. Baseline BAF R f d from Field Measured BAFs 1. A baseline BAF R f d should be calculated from a field measured BAF tT using the guidance and equations outlined in Section <IP_ADDRESS>( A) in Procedure # 1.

2. Freely Dissolved Fraction. Due to their low hydrophobicity ( i. e., log K ow < 4.0), nonionic organic chemicals applicable to Procedure # 4 are expected to remain almost entirely in the freely dissolved form in natural waters with dissolved and particulate organic carbon concentrations typical of most field BAF studies. Therefore, the freely dissolved fraction should be assumed equal to 1.0 unless the concentrations of DOC and POC are very high in the field BAF study. For studies with very high DOC or POC concentrations, ( e. g., about 100 mg/ L or higher for DOC or 10 mg/ L or higher for POC), the freely dissolved fraction may be substantially lower than 1.0 and therefore should be calculated using Equation 5 12.

3. Temporal Averaging of Concentrations. Also due to their low hydrophobicity, nonionic organic chemicals appropriate to Procedure # 4 will also tend to reach steadystate quickly compared with those chemicals to which Procedure # 1 applies. Therefore, the extent of temporal averaging of tissue and water concentrations is typically much less than that required for highly hydrophobic chemicals to which Procedure # 1 is applied. In addition, field studies used to calculate BAFs for these chemicals should have sampled water and tissue at similar points in time because tissue concentrations should respond rapidly to changes in water concentrations. EPA will be providing additional guidance on appropriate BAF study designs for nonionic organic chemicals ( including those 5 53 appropriate to Procedure # 4) in its forthcoming guidance document on conducting field BAF and BSAF studies.

B. Baseline BAF R f d from a Laboratory Measured BCF 1. Except where noted below, a baseline BAF R f d should be calculated from a laboratorymeasured BCFtT using the guidance and equations outlined in Section <IP_ADDRESS>( c) of Procedure # 1.

2. Food Chain Multipliers. Because biomagnification is not an important concern for the minimally hydrophobic chemicals applicable to Procedure # 4, FCMs are not used in the derivation of a baseline BAF R f d from a laboratory measured BCF tT .

3. Freely Dissolved Fraction. Due to their low hydrophobicity ( i. e., log K ow < 4.0), nonionic organic chemicals to which Procedure # 4 is applied are expected to remain almost entirely in the freely dissolved form in waters containing dissolved and particulate organic carbon concentrations typical of laboratory BCF studies. Therefore, the freely dissolved fraction should usually be assumed to be equal to 1.0. The freely dissolved fraction will be substantially less than 1.0 only in situations where unusually high concentrations of DOC and POC are present in the lab BCF study ( e. g., above about 100 mg/ L for DOC or about 10 mg/ L for POC). In this situation, the freely dissolved fraction should be calculated according to Equation 5 12.

<IP_ADDRESS> Selecting the Final Baseline BAF R f ds After calculating individual baseline BAF R f ds using as many of the methods in Procedure # 4 as possible, the next step is to determine a final baseline BAF R f d for a given trophic level from the individual baseline BAF R f ds ( Figure 5 2). The final baseline BAF R f d will be used in the last step to determine the national BAF for each trophic level. A final baseline BAF R f d should be determined for each trophic level from the individual baseline BAF R f ds by considering the data preference hierarchy defined by Procedure # 4 and uncertainty in the data. The data preference hierarchy for Procedure # 4 is:

1. a baseline BAF R f d from an acceptable field measured BAF or predicted from an acceptable laboratory measured BCF.

Since bioaccumulation via dietary uptake and subsequent biomagnification generally are not of concern for chemicals subject to Procedure # 4, field measured BAFs and laboratorymeasured BCFs are considered equally in determining the national BAF.

Final baseline BAF R f ds should be selected for each trophic level using the following steps and guidelines. 5 54 1. Calculate Species Mean Baseline BAF R f ds. For each BAF method ( i. e., field measured BAF or a BAF from a lab measured BCF) where more than one acceptable baseline BAF R f d is available for a given species, calculate a species mean baseline BAF R f d according to the guidance described previously in Procedure # 1.

2. Calculate Trophic Level Mean Baseline BAF R f ds. For each BAF method where more than one acceptable species mean baseline BAF R f d is available within a given trophic level, calculate the trophic level mean baseline BAF R f d as the geometric mean of acceptable species mean baseline BAF R f ds for that trophic level.

3. Select a Final Baseline BAF R f d for Each Trophic Level. For each trophic level, select the final baseline BAF R f d using best professional judgment by considering: ( 1) the data preference hierarchy, and ( 2) the relative uncertainties among trophic level mean BAFs derived using different methods.

a. As discussed above, field measured BAFs and laboratory measured BCFs are considered equally desirable for deriving a final trophic level mean baseline BAF R f d using Procedure # 4. If a trophic level mean baseline BAF R f d is available from both a field measured BAF and a laboratory measured BCF, the final baseline BAF R f d should be selected using the trophic level mean baseline BAF R f d or BCF R f d with the least overall uncertainty.

b. The above steps should be performed for each trophic level until a final baseline BAF R f d is selected for trophic levels two, three, and four.

<IP_ADDRESS> Calculating National BAFs As described in Procedure # 1, the last step in deriving a national BAF for a given trophic level for nonionic organic chemicals is to convert the final baseline BAF R f d determined in the previous step to a BAF that reflects conditions to which the national 304( a) criterion will apply ( Figure 5 2). Each national BAF should be determined from a final baseline BAF R f d according to the following guidelines.

1. National BAF Equation. Except where noted below, national BAFs for trophic levels two, three, and four should be calculated from the final, trophic level mean baseline BAF R f ds using the same equation and procedures described previously in Procedure # 1 ( see Section <IP_ADDRESS> in Procedure # 1).

2. Freely Dissolved Fraction. Due to their low hydrophobicity ( i. e., log K ow < 4.0), a freely dissolved fraction of 1.0 should be assumed for calculating national BAFs for nonionic organic chemicals using Procedure # 4. A freely dissolved fraction of 1.0 should be assumed because at a log K ow value of less than 4.0, nonionic organic chemicals are expected to remain over 99 percent in the freely dissolved form at POC and DOC 5 55 concentrations corresponding to national default values for U. S. bodies of water ( i. e., 0.5 mg/ L and 2.9 mg/ L, respectively).

5.5 NATIONAL BIOACCUMULATION FACTORS FOR IONIC ORGANIC CHEMICALS This section contains guidelines for deriving national BAFs for ionic organic chemicals ( i. e., organic chemicals which undergo significant ionization in water). As defined in Section 5.3.5, ionic organic chemicals contain functional groups which can either readily donate protons ( e. g., organic acids with hydroxyl, carboxylic, and sulfonic groups) or readily accept protons ( e. g., organic bases with amino and aromatic heterocyclic nitrogen groups). Some examples of ionic organic compounds include:

C chlorinated phenols ( e. g., 2,4,6 trichlorophenol, pentachlorophenol), C chlorinated phenoxyalkanoic acids ( e. g., 2,4 dichlorophenoxyacetic acid [ 2,4 D]), C nitrophenols ( e. g., 2 nitrophenol, 2,4,6 trinitrophenol), C cresols ( e. g., 2,4 dinitro o cresol [ DNOC]), C pyridines ( e. g., 2,4 dimethypyidine), C aliphatic and aromatic amines ( e. g., trimethylamine, aniline), and C linear alkylbenzenesulfonate ( LAS) surfactants.

Ionic organic chemicals are considered separately for deriving national BAFs because the anionic or cationic species of these chemicals behave much differently in the aquatic environment compared with their neutral ( un ionized) counterparts. The neutral species of ionic organic chemicals are thought to behave in a similar manner as nonionic organic compounds ( e. g., partitioning to lipids and organic carbon as a function of hydrophobicity). However, the ionized ( cationic, anionic) species exhibit a considerably more complex behavior involving multiple environmental partitioning mechanisms ( e. g., ion exchange, electrostatic, and hydrophobic interactions) and a dependency on pH and other factors including ionic strength and ionic composition ( Jafvert et al., 1990; Jafvert 1990; Schwarzenbach, et al., 1993). As a consequence, methods to predict the environmental partitioning of organic cations and anions are less developed and validated compared with methods for nonionic organic chemicals ( Spacie, 1994; Suffet et al., 1994).

Given the current limitations in the state of the science for predicting the partitioning and bioaccumulation of the ionized species of ionic organic chemicals, procedures for deriving national BAFs for these chemicals differ depending on the extent to which the fraction of the total chemical is likely to be represented by the ionized ( cationic, anionic) species in U. S. surface waters. When a significant fraction of the total chemical concentration is expected to be present as the ionized species in water, procedures for deriving the national BAF rely on empirical ( measured) methods ( i. e., Procedures # 5 and 6 in Section 5.6). When an insignificant fraction of the total chemical is expected to be present as the ionized species ( i. e., the chemical exists essentially in the neutral form), procedures for deriving the national BAF will follow those 5 56 established for nonionic organic chemicals ( e. g., Procedures # 1 through # 4 in Section 5.4). The following guidelines apply for assessing the occurrence of cationic and anionic forms at typical environmental pH ranges.

1. For the ionic organic chemical of concern, the dissociation constant, pK a, should be compared to the range of pH values expected in fresh and estuarine waters of the U. S. At pH equal to the pK a, 50% of the organic acid or base is expected to be present in the ionized species. The pH values for U. S. fresh and estuarine waters typically range between 6 and 9, although somewhat higher and lower values can occur in some bodies of water ( e. g., acidic bogs and lakes, highly alkaline and eutrophic systems, etc.).

2. For organic acids, the chemical will exist almost entirely in its un ionized form when pH is about 2 or more units below the pK a. For organic bases, the chemical will exist almost entirely in its un ionized form when pH is about 2 or more units above the pK a. In these cases, the aqueous behavior of the chemical would be expected to be similar to nonionic organic chemicals. Therefore, national BAF should usually be derived using Procedures # 1 through # 4 in Section 5.4.

3. When pH is greater than the pK a minus 2 for organic acids ( or less than the pKa plus 2 for organic bases), the fraction of the total chemical that is expected to exist in its ionized form can become significant ( i. e., $ 1% in the ionized). In these cases, the national BAF should usually be derived using Procedures # 5 and # 6 in Section 5.6.

4. In general, most organic acids ( e. g., pentachlorophenol and silvex), exist primarily in the ionized form in ambient waters because their pK a's ( 4.75 and 3.07, respectively) are much smaller than the pH of the ambient waters. Conversely, most organic bases, ( e. g., aniline) exist mostly in the un ionized form in ambient waters because their pK a's ( 4.63 for aniline) are much smaller than the pH of the ambient waters.

5. The above guidelines are intended to be a general guide for deriving national BAFs for ionic organic chemicals, not an inflexible rule. Modifications to these guidelines should be considered on a case by case basis, particularly when such modifications are strongly supported by measured bioaccumulation or bioconcentration data. For example, initial models have been developed for predicting the solid and organic phase partitioning of certain organic acids ( e. g., Jafvert 1990, Jafvert et al., 1990). As these or other models become more fully developed and appropriately validated in the future, they should be considered in the development of national BAFs. In addition, since pH is a controlling factor for dissociation and subsequent partitioning of ionic organic chemicals, consideration should be given to expressing BAFs or BCFs as a function of pH ( or other factors) where sufficient data exist to reliably establish such relationships. 5 57 5.6 NATIONAL BIOACCUMULATION FACTORS FOR INORGANIC AND ORGANOMETALLIC CHEMICALS This section contains guidelines for deriving national BAFs for inorganic and organometallic chemicals as defined in Section 5.3.5. The derivation of BAFs for inorganic and organometallic chemicals differs in several ways from procedures for nonionic organic chemicals. First, lipid normalization of chemical concentrations in tissues does not generally apply for inorganic and organometallic chemicals. Thus, BAFs and BCFs cannot be extrapolated from one tissue to another based on lipid normalized concentrations as is done for nonionic organic chemicals. Second, the bioavailability of inorganics and organometallics in water tends to be chemical specific and thus, the techniques for expressing concentrations of nonionic organic chemicals based on the freely dissolved form do not generally apply. Third, at the present time there are no generic bioaccumulation models that can be used to predict BAFs for inorganic and organometallic chemicals as a whole, unlike the existence of K ow based models for nonionic organic chemicals. While some chemical specific bioaccumulation models have been developed for inorganic and organometallic chemicals ( e. g., Mercury Cycling Model by Hudson et. al, 1994), those models currently tend to require site specific data for input to the model and are restricted to site specific applications. As the models become more fully developed and validated in the future, they should be considered on a case by case basis in conjunction with the following procedures for deriving national BAFs.

5.6.1 Selecting the BAF Derivation Procedure As shown in Figure 5 1, national BAFs can be derived using two procedures for inorganic and organometallic chemicals ( Procedures # 5 and # 6). The choice of the BAF derivation procedure depends on whether or not the chemical undergoes biomagnification in aquatic food webs.

1. For many inorganic and organometallic chemicals, biomagnification does not occur and the BCF will be equal to the BAF. For these types of chemicals, Procedure # 5 should be used to derive the national BAF. Procedure # 5 considers BAFs and BCFs to be of equal value in determining the national BAF and does not require the use of FCMs with BCF measurements. Guidance for deriving BAFs using Procedure # 5 is provided in Section 5.6.3.

2. For some inorganic and organometallic chemicals ( e. g., methylmercury), biomagnification does occur and Procedure # 6 should be used to determine the national BAF. Procedure # 6 gives general preference to the use of field measured BAFs over laboratory measured BCFs and requires FCMs to be used with BCF measurements for predicting BAFs. Guidance for deriving BAFs using Procedure # 6 is provided in Section 5.6.4.

3. Determining whether or not biomagnification occurs for inorganic and organometallic chemicals requires chemical specific data on measured concentrations of the chemical in aquatic organisms and their prey. Concentrations in aquatic organisms that increase 5 58 substantially at successive trophic levels of a food web suggest that biomagnification is occurring. Concentrations in aquatic organisms that remain about the same or decrease at successive trophic levels of a food web suggest that biomagnification is not occurring. When comparing tissue concentrations for assessing biomagnification, care should be taken to ensure that the aquatic organisms chosen actually represent functional predatorprey relationships and that all major prey species are considered in the comparisons.

5.6.2 Bioavailability The chemical specific nature of inorganic and organometallic bioavailability is likely due in part to chemical specific differences in several factors which affect bioavailability and bioaccumulation. These factors include differences in the mechanisms for chemical uptake by aquatic organisms ( e. g., passive diffusion, facilitated transport, active transport), differences in sorption affinities to biotic and abiotic ligands, and differences in chemical speciation in water. Some inorganic and organometallic chemicals exist in multiple forms and valence states in aquatic ecosystems that can differ in their bioavailability to aquatic organisms and undergo conversions between forms. For example, selenium can exist in various forms in aquatic ecosystems, including inorganic selenite(+ 4) and selenate(+ 6) oxyanions, elemental selenium ( 0) under reducing conditions ( primarily in sediments), and organoselenium compounds of selenide ( 2). Dominant forms of mercury in natural, oxic waters include inorganic (+ 2) mercury compounds and methylmercury; the latter is generally considered to be substantially more bioavailable than inorganic mercury compounds to higher trophic level organisms. Although a generic analogue to the " freely dissolved" conversion for nonionic organic chemicals does not presently exist for inorganic and organometallic chemicals as a whole, the occurrence and bioavailability of different forms of these chemicals should be carefully considered when deriving national BAFs.

1. If data indicate that: ( 1) a particular form ( or multiple forms) of the chemical of concern largely governs its bioavailability to target aquatic organisms, and ( 2) BAFs are more reliable when derived using the bioavailable form( s) compared with using other form( s) of the chemical of concern, then BAFs and BCFs should be based on the appropriate bioavailable form( s).

2. Because different forms of many inorganic and organometallic chemicals may interconvert once released to the aquatic environment, regulatory and mass balance considerations typically require an accounting of the total concentration in water. In these cases, sufficient data should be available to enable conversion between total concentrations and the other ( presumably more bioavailable) forms in water.

5.6.3 Deriving BAFs Using Procedure # 5 This section contains guidance for calculating national BAFs for inorganic and organometallic chemicals using Procedure # 5 as shown in Figure 5 1. The types of inorganic and 5 59 organometallic chemicals for which Procedure # 5 is appropriate are those that are not likely to biomagnify in aquatic food webs ( see Section 5.1 above). In Procedure # 5, two methods are available to derive the national BAF for a given trophic level:

C using a BAF from an acceptable field study ( i. e., field measured BAF), or C predicting a BAF from an acceptable laboratory measured BCF.

Individual BAFs should be determined from field measured BAFs or laboratory measured BCFs according to the following guidelines.

<IP_ADDRESS> Determining Field Measured BAFs 1. Except where noted below, field measured BAFs should be determined using the guidance provided in Section <IP_ADDRESS>( A) of Procedure # 1.

2. As described previously, conversion of field measured BAFs to baseline BAF R f ds based on lipid normalized and freely dissolved concentrations does not apply for inorganic and organometallic chemicals. Therefore, the guidance and equations provided in Procedure # 1 which pertain to converting field measured BAFs to baseline BAF R f ds and subsequently to national BAFs do not generally apply to inorganic chemicals. As discussed in Section 5.6.2 above, an analogous procedure in concept might be required for converting total BAFs to BAFs based on the most bioavailable form( s) for some inorganic and organometallic chemicals of concern. Such procedures should be applied on a chemicalspecific basis.

3. BAFs should be expressed on a wet weight basis; BAFs reported on a dry weight basis can be used only if they are converted to a wet weight basis using a conversion factor that is measured or reliably estimated for the tissue used in the determination of the BAF.

4. BAFs should be based on concentrations in the edible tissue( s) of the biota unless it is demonstrated that whole body BAFs are similar to edible tissue BAFs. For some finfish and shellfish species, whole body is considered to be the edible tissue.

5. The concentrations of an inorganic or organometallic chemical in a bioaccumulation study should be greater than normal background levels and greater than levels required for normal nutrition of the test species if the chemical is a micronutrient, but below levels that adversely affect the species. Bioaccumulation of an inorganic or organometallic chemical that is essential to the nutrition of aquatic organisms might be overestimated if concentrations are at or below normal background levels due to selective accumulation by the organisms to meet their nutritional requirements. 5 60 <IP_ADDRESS> Determining Laboratory Measured BCFs 1. Except where noted below, BAFs should be predicted from laboratory measured BCFs using the guidance provided in Section <IP_ADDRESS>( c) of Procedure # 1.

2. As described previously, conversion of laboratory measured BCFs to baseline BCF R f ds based on lipid normalized and freely dissolved concentrations does not apply for inorganic and organometallic chemicals. Therefore, the guidance and equations provided in Procedure # 1 which pertain to converting laboratory measured BCFs to baseline BCF R f ds and subsequently to national BCFs do not generally apply to inorganic and organometallic chemicals. As discussed in Section 5.6.2 above, an analogous procedure in concept might be required for converting total BCFs to BCFs based on the most bioavailable form( s) of some inorganic and organometallic chemicals of concern. Such procedures should be applied on a chemical specific basis. In addition, the use of FCMs with BCFs does not apply to chemicals applicable to Procedure # 5.

3. BCFs should be expressed on a wet weight basis; BCFs reported on a dry weight basis can be used only if they are converted to a wet weight basis using a conversion factor that is measured or reliably estimated for the tissue used in the determination of the BCF.

4. BCFs should be based on concentrations in the edible tissue( s) of the biota unless it is demonstrated that whole body BCFs are similar to edible tissue BCFs. For some finfish and shellfish species, whole body is considered to be the edible tissue.

5. The concentrations of an inorganic or organometallic chemical in a bioconcentration test should be greater than normal background levels and greater than levels required for normal nutrition of the test species if the chemical is a micronutrient, but below levels that adversely affect the species. Bioaccumulation of an inorganic or organometallic chemical that is essential to the nutrition of aquatic organisms might be overestimated if concentrations are at or below normal background levels due to selective accumulation by the organisms to meet their nutritional requirements.

<IP_ADDRESS> Determining the National BAFs After calculating individual BAFs using as many of the methods in Procedure # 5 as possible, the next step is to determine national BAFs for each trophic level from the individual BAFs. The national BAFs will be used to determine the national 304( a) criteria. The national BAFs should be determined from the individual BAFs by considering the data preference hierarchy defined for Procedure # 5 and uncertainty in the data. The data preference hierarchy for Procedure # 5 is:

1. a BAF from an acceptable field measured BAF or predicted from an acceptable laboratory measured BCF. 5 61 Since bioaccumulation via dietary uptake and subsequent biomagnification are not of concern for chemicals subject to Procedure # 5, field measured BAFs and laboratory measured BCFs are considered equally in determining the national BAFs. The national BAFs should be selected for each trophic level using the following steps and guidelines.

1. Calculate Species Mean BAFs. For each BAF method where more than one acceptable field measured BAF ( or a BAF predicted from a BCF) is available for a given species, calculate the species mean BAF as the geometric mean of all acceptable individual measured or BCF predicted BAFs. When calculating species mean BAFs, individual measured or BCF predicted BAFs should be reviewed carefully to assess uncertainties in the BAF values. Highly uncertain BAFs should not be used. Large differences in individual BAFs for a given species ( e. g., greater than a factor of 10) should be investigated further and in such cases, some or all of the BAFs for a given species might not be used. Additional discussion on evaluating the acceptability of BAF and BCF values is provided in the Bioaccumulation TSD.

2. Calculate Trophic Level Mean BAFs. For each BAF method where more than one acceptable species mean BAF is available within a given trophic level, calculate the trophic level mean BAF as the geometric mean of acceptable species mean BAFs in that trophic level. Trophic level mean BAFs should be calculated for trophic levels two, three and four because available data on U. S. consumers of fish and shellfish indicate significant consumption of organisms in these trophic levels.

3. Select a Final National BAF for Each Trophic Level. For each trophic level, select the final national BAF using best professional judgment by considering: ( 1) the data preference hierarchy in Procedure # 5, and ( 2) the relative uncertainties among trophic level mean BAFs derived using different methods.

a. As discussed above, field measured BAFs and laboratory measured BCFs are considered equally desirable for deriving a final national BAF using Procedure # 5. If a trophic level mean BAF is available from both a field measured BAF and a laboratory measured BCF, the final national BAF should be selected using the trophic level mean BAF with the least overall uncertainty.

b. The above steps should be performed for each trophic level until a national BAF is selected for trophic levels two, three, and four.

5.6.4 Deriving BAFs Using Procedure # 6 This section contains guidance for calculating national BAFs for inorganic and organometallic chemicals using Procedure # 6 as shown in Figure 5 1. The types of inorganic and organometallic chemicals for which Procedure # 6 is appropriate are those that are considered likely to biomagnify in aquatic food webs ( see Section 5.6.1 above). Methylmercury is an 5 62 example of an organometallic chemical to which Procedure # 6 applies. In Procedure # 6, two methods are available to derive the national BAF:

C using a BAF from an acceptable field study ( i. e., field measured BAF), or C predicting a BAF from an acceptable laboratory measured BCF and a FCM.

Individual BAFs should be determined from field measured BAFs or laboratory measured BCFs and FCMs according to the following guidelines.

<IP_ADDRESS> Determining Field Measured BAFs 1. Field measured BAFs should be determined using the guidance provided in Section <IP_ADDRESS> of Procedure # 5.

<IP_ADDRESS> Determining Laboratory Measured BCFs 1. Except where noted below, BAFs should be predicted from laboratory measured BCFs using the guidance provided in Section <IP_ADDRESS> of Procedure # 5.

2. Because biomagnification is of concern for chemicals applicable to Procedure # 6, BAFs should be predicted from laboratory measured BCF using FCMs. Currently, there are no generic models from which to predict FCMs for inorganic or organometallic chemicals. Therefore, FCMs should be determined using field data as described in the section entitled: " Field Derived FCMs" in Section <IP_ADDRESS>( c) of Procedure # 1. Unlike nonionic organic chemicals, field derived FCMs for inorganic and organometallic chemicals are not based on lipid normalized concentrations in tissues. For calculating FCMs for inorganic and organometallic chemicals, concentrations in tissues should be based on the consistent use of either wet weight or dry weight concentrations in edible tissues. FCMs should be derived for trophic levels two, three, and four.

<IP_ADDRESS> Determining the National BAF After calculating individual BAFs using as many of the methods in Procedure # 6 as possible, the next step is to determine national BAFs for each trophic level from the individual BAFs. The national BAFs will be used to determine the national 304( a) criteria. The national BAFs should be determined from the individual BAFs by considering the data preference hierarchy defined for Procedure # 6 and uncertainty in the data. The data preference hierarchy for Procedure # 6 is ( in order of preference):

1. a BAF from an acceptable field measured BAF, or 2. a predicted BAF from an acceptable laboratory measured BCF and FCM.

This data preference hierarchy reflects EPA's preference for field measured BAFs over BAFs predicted from a laboratory measured BCF and FCM, because field measured BAFs are 5 63 direct measures of bioaccumulation and biomagnification in aquatic food webs. BAFs predicted from laboratory measured BCFs and FCMs indirectly account for biomagnification through the use of the FCM. For each trophic level, the national BAFs should be determined using the following steps and guidelines.

1. Calculate Species Mean BAFs. For each BAF method where more than one acceptable field measured BAF or BAF predicted using a BCF and FCM is available, calculate a species mean BAF according to the guidance described previously in Procedure # 5.

2. Calculate Trophic Level Mean BAFs. For each BAF method where more than one acceptable species mean BAF is available within a given trophic level, calculate the trophic level mean BAF according to guidance described previously in Procedure # 5.

3. Select a Final National BAF for Each Trophic Level. For each trophic level, select the final national BAF using best professional judgment by considering: ( 1) the data preference hierarchy in Procedure # 6, and ( 2) the relative uncertainties among trophic level mean BAFs derived using different methods.

a. When a trophic level mean BAF is available using both methods for a given trophic level ( i. e., a field measured BAF and a BAF predicted from a BCF and FCM), the national BAF should usually be selected using the field measured BAF which is the preferred BAF method in the data preference hierarchy in Procedure # 6.

b. If uncertainty in the trophic level mean BAF derived using field measured BAFs is considered to be substantially greater than a trophic level mean BAF derived using a BCF and FCM, the national BAF for that trophic level should be selected from the second tier ( BCF @ FCM) method.

c. The above steps should be performed for each trophic level until a national BAF is selected for trophic levels two, three, and four.

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