3.3.1 Toxicity Evaluation Requirements
For Toxicity evaluation requirements, ASTM D40542 refers to the Department of Defense Aerospace Fuels Certification Handbook58, Appendix E. Appendix E lists seven (7) minimum toxicity tests, Five (5) additional tests, and an optional physiologically-based pharmacokinetic model development effort. The minimum tests include in vitro and in vivo genotoxicity, acute inhalation, skin irritation, and inhalation studies of 2-week and 90-day duration, a typical initial toxicity screen. The present analysis must diverge from the Appendix E process because there are currently no toxicology test data for Bio-SPK. Appendix E does not address a process that could be used in the absence of data on the fuel of interest. An alternative approach is necessary due to the lack of toxicity testing on Bio-SPK. This review discusses available alternative methods, applies appropriate methods to Bio-SPK, and reviews the toxicology literature to support the resulting evaluation.
3.3.2 Reciprocal Calculation Procedure
Occupational Exposure Limits (OEL) have generally been developed for specific chemicals or for defined mixtures based on toxicological data for the chemical or mixture of interest. Because exposure to mixtures of chemicals is typical in occupational settings, the American Conference of Governmental Industrial Hygienists (ACGIH) developed and has recommended a Reciprocal Calculation Procedure (RCP) for mixtures (since 1940, according to ECETOC59). The RCP is a calculation of a mixture exposure limit based on the weighted exposure limits of the individual components. It can be applied to an airborne sample or to the composition of a liquid mixture. Application of the RCP assumes that the components of a mixture have similar toxicity and act in an additive manner. The RCP approach is limited in its application to Bio-SPK or similar hydrocarbon mixtures because of the number of chemically distinct components, and the lack of chemical specific OELs for the components. The need for a more generalizable approach was addressed in the petroleum refining industry, where many different mixtures of varying composition are produced, with their composition depending of the starting material, the processing steps, and the finishing steps used. Although petroleum derived hydrocarbon mixtures have numerous components, they are primarily aliphatics (paraffins) and aromatics. In view of the similarity of the toxicity of many individual chemicals within these classes, and lack of toxicity data on most, the UK Health and Safety Executive developed OELs for the hydrocarbon classes (aliphatics, cycloaliphatics, aromatics) that could be used in the RCP calculation to develop a mixture OEL for a hydrocarbon solvent, fuel, or refinery stream6061. The chemical categories and their respective group OELs are in Table 3-12.
Chemical group OEL (mg/m3)
C5-6 aliphatics* 1800
C7-15 aliphatics 1200
C5-6 cycloaliphatics* 1800
C7-16 cycloaliphatics 800
Aromatics* 500
*excluding those with specific OELs; from HSE60
Table 3-12. Hydrocarbon Class OELs
These OEL values are used in the RCP calculation along with the OELs for specific chemical components of the mixture such as hexane, cyclohexane, and several aromatics. The chemicals with specific OELs were often those for which the assumption of additivity was not justified due to different toxic endpoints, potency, or mode of toxic action (common examples cited in this context are n-hexane, benzene, cyclohexane, and naphthalene), and for this reason they often had lower OELs.
A critical review of the available methods and data for hydrocarbon mixture assessment59 concluded that the group OEL values assigned by HSE were reasonable, and that the RCP approach was the best available. McKee et al.62 presented an updated assessment of the applicability of the RCP and group OELs for hydrocarbon mixtures.
They developed chemical group guidance values (GGV, e.g., OELs for chemical groups analogous to the hydrocarbon class OELs used by HSE) based on central nervous system effects. To do this, they incorporated new toxicological information and recent changes in established OELs, and also performed new research on acute exposures in animals and humans to representative chemicals from each group. The group classifications and guidance values recommended by McKee et al.62 are shown in Table 3-13.
Chemical group OEL (mg/m3)
C5-8 aliphatics/cycloaliphatics 1500 C9-15 aliphatics/cycloaliphatics 1200
C7-8 aromatics 200
C9-15 aromatics 100
From McKee et al.62
Table 3-13. Group Guidance Values
There are several differences between the McKee et al.62 and the UK-HSE60 recommendations. McKee et al.62 recommended slightly different category definitions (carbon number ranges), slightly lower OEL values for the same aliphatics, and considerably lower OEL values for the aromatics. McKee et al.62 also compared different aliphatic chemical classes and concluded that there was no clear difference between iso-, normal, and cyclic aliphatics, so they do not derive different values for cycloaliphatics and aliphatics. They also note that some data supports a general increase in toxicity with increase in carbon number, but that there is limited data for 9-carbon compounds or larger. ACGIH63 added a new appendix to the TLV documentation describing the group guidance value/RCP (GGV/RCP) approach for hydrocarbon solvent mixtures. ACGIH63
“considers this method to be applicable” when there is no OEL for the specific mixture of interest or OELS for all components, and they cite the UK-HSE60 and McKee et al.62 recommendations as examples.
3.3.3 Applicability of GGV/RCP Approach to Bio-SPK
Although the GGV/RCP approach was developed to facilitate evaluation of petroleum refinery streams, generally hydrocarbons with 5-15 carbon atoms that boil at temperatures from <50 to >300oC, it is useful for refined hydrocarbon mixtures regardless of source. A primary limitation of the method is its application to mixtures which contain components that do not meet the assumptions of additivity and similar mode-of action. Its applicability to Bio-SPK is more straightforward and less uncertain than for petroleum derived mixtures in this regard. Chemicals whose presence in a mixture make the GGV/RCP method inapplicable, or at least suspect, are low molecular
weight compounds (hexane, cyclohexane) or aromatic compounds (benzene, naphthalene, methylnaphthalene), and are not present in Bio-SPKs. Based on the conclusions of McKee et al.62, ACGIH 200963, and ECETOC 199759, the assumption of additivity for the components of Bio-SPKs is reasonable. In Table 3-14, the GGV/RCP approach is applied to SPK and jet fuels using both sources of GGVs discussed above. The estimates are similar for Bio-SPK based on the GGVs from two sources, but the values differ for the jet fuels due to their different GGV for aromatics. Detailed isomer composition data were not available for the jet fuels, so these calculations assume all hydrocarbons in the fuels are C9 or greater. These methods predict much lower toxicity for Bio-SPK than for jet fuel. The ACGIH TLV for jet fuel is 200 mg/m364.
Fuel Composition Mixture OEL (mg/m3) GGV Source Composition Source
SPK 100% aliphatics 1200 McKee et al.62 Figure 2-1
Bio-SPKs 1-4, Figure 2-1
90-99% iso- or n-aliphatics 1-10% cycloaliphatics
1142-1194 HSE60 Figure 2-1
Jet A 81% aliphatics 18.1% aromatics
402 McKee et al.62 CRC39 JP-8 50% n-, iso-paraffin,
30% cyclic, 20% aromatic
375 McKee et al.62 NRC65 JP-8 50% n-, iso-paraffin,
30% cyclic, 20% aromatic 839 HSE60 NRC65 Table 3-14. Occupational Exposure Limit Based on GGV/RCP Method.
The application of this method to Bio-SPK is uncertain due to the lack of toxicity test data for many of the chemicals in the mixture, including compounds with C-9 or greater. The assumption of common toxicological mode of action and additivity needs to be verified as new toxicological research is done. Specific questions of ongoing interest are whether new information supports combining normal-, iso- and cyclo-paraffins as was done by McKee et al.62, and whether the group OEL values for C5-8 and C9-15 compounds are supported. To investigate these questions, toxicology literature was searched for 73 C7-15 paraffins (search included CCOHS, RTECS, HSDB, TOXLINE, US EPA High Production Volume Information System, European Chemical Substances Information System and other public sources). Several recent studies have evaluated individual components of jet fuel with regard to properties related to inducing inflammation or inflammatory mediators in cultured cells or animal skin. These studies do not provide a sufficient basis for assessment of the overall toxicity of related mixtures.
No new data were found that would challenge the assumptions used in the analysis of McKee et al.62, namely that iso-, normal-, and cyclo-paraffins in the C9-15 range are similar and additive. No data were found to suggest that any chemicals in these groups have unique toxic effects, such that they should be treated separately. However, there remains limited data, especially on the longer chain hydrocarbons, so this approach to analysis of Bio-SPK is inherently uncertain.
3.3.4 Similar Mixture Approach
The EPA 198666, risk assessment guidelines for mixtures proposed a 3-tier approach to assessing the human health hazard of a chemical mixture. Toxicological data on the mixture of interest is preferred. If not available, data for a toxicologically similar mixture could be used. If no “sufficiently similar” mixture is available, data on individual components could be used. EPA 200067 discusses the data requirements to determine if a
mixture is sufficiently similar. Since there is no toxicological data on Bio-SPK, and inadequate data on its components, the best option, as described by EPA is to evaluate a similar mixture. Several paraffinic hydrocarbon mixtures have been tested for toxicity and the results of those tests may be applicable to Bio-SPK if the mixtures are sufficiently similar. Similarity of a mixture is assessed in terms of chemical composition and toxicological action. McKee et al.62 concluded that iso-, normal-, and cyclo-paraffins in the C9-15 range are all toxicologically similar and additive. Saturated hydrocarbons have no metabolically active or chemically reactive chemical sites and are relatively stable under physiological conditions. As described above, no data has been found that contradicts the conclusion that iso-, normal-, and cyclo-paraffins in the C9-15 range are similar and additive. Therefore, to be sufficiently similar toxicologically to serve as a surrogate for Bio-SPK, another mixture would need to be mostly C9-16 paraffins.
3.3.5 Similar Mixtures
Purified paraffinic solvent products have been available commercially for many years and toxicity testing has been done on some of these mixtures. In general, these products contain defined distillation cuts of paraffinic hydrocarbon mixtures.
Composition and toxicity data on several were reviewed by Mullin et al.68, and Table 3-15 shows their identity and properties.
Table 3-15. Isoparaffinic Hydrocarbon Solvents Reviewed by Mullin et al.68
The data discussed above and by McKee et al.62 does not suggest differences between subsets of C9-15 paraffins, so further chemical characterization of these products in terms of iso-, normal-, and cyclic paraffins, and distribution of carbon chain length are not needed. All of the products listed in Table 3-15 are similar to Bio-SPKs based on the discussion above, so data on any of the products could be used as a surrogate for Bio-SPKs. The toxicity data for the paraffinic solvents are summarized in Table 3-16 along with similar data for jet fuels.
Manufacturer
hydrotreated heavy Hydrotreated petroleum fraction with C6-13;
boiling in the range of approximately 65-230°C
Isopar M C12-15 64742-47-8 191 205-254 Distillates (petroleum),
hydrotreated light Hydrotreated petroleum fraction with C9-16, boiling in the range of approximately 150-290°C Phillips
Soltrol 100 C9-11 68551-16-6 142 157-173 C9-11 Isoalkanes Soltrol 130 C10-13 68551-17-7 158 176-208 C10-13 Isoalkanes Soltrol 170 C10-14 68551-19-9 185 218-238 C12-14 Isoalkanes
Shell
Shell Sol 71 C9-12 64741-65-7 158 179-202 Naphtha (petroleum), heavy alkylate
Predominantly branched chain saturated hydrocarbons; C9-12, boiling range of ~ 150-220°C, from reaction products of isobutane with monoolefins
Shell Sol TS C8-C12 isoparaffin Texaco
Texsolve S-2 C9-10 64742-88-7 135 156-157 Solvent naphtha (petroleum), medium aliph.
Predominantly saturated hydrocarbons; C9-12, boiling range of ~140-220°C, from distillation of crude oil
Test Isoparaffinic solventsa,b Jet Ac JP-8d Oral LD50, g/kg, rat >10 (G, L, M); >25 (100, 130, 71); >8000 (71, Shell69). >20 >5 Dermal LD50, g/kg, (rabbit) >3.2 (G, L, M); >15 (100, 130); >5 (71); >4000 (71, Shell69) >4 >2 Inhalation LC50, 4 hr (rat) >2000 ppm (G); >715 ppm (L); >3684 ppm (100); >1227 ppm
(130); >592 ppm (71); >290 ppm (vapor); >5991 mg/m3 (vapor + aerosol) (M).
>5 mg/L for straight run
kerosene, vapor and aerosol >3.43 mg/L(vapor); >4.39 mg/L (vapor and aerosol)
Skin irritation (rabbit) Slight (G, L, 100); Mild – 0.5 ml occluded (M); Slight on intact
skin, severe on abraised skin (130); Moderate (71) Slight Nonirritating to slight irritation - 0.5 ml occluded
Skin sensitization No sensitization, phototoxicity, or photosensitization – 50% in
petrolatum (G, L, M); not sensitizing - 30% (M); human data. Negative Nonsensitizing to weakly sensitizing (guinea pig)
Eye irritation (rabbit) Slight (G, L, 100); Not irritating (M) Moderate to severe. Nonirritating - 0.1 ml (rabbit).
Respiratory irritation
(mouse) No irritation – ~420 ppm (G); slight - >1000 ppm vapor (C), no irritation – saturated vapor, some irritation – 1728-49919 mg/m3 (vapor + aerosol (M).
Dead. kerosene: RD50 - >0.1 mg/L (vapor); 6900 mg/m3 (vapor + aerosol (mouse)70
RD50 (30 min exposure) - 2876 mg/m3
Neurotoxicity No effect - 100 ppm x 6 hr (TS) 71 Systemic toxicity –
inhalation (rats) Mild liver effect, reduced body wt – 900 ppm x 6 hr/d x 5 d/wk x 12 wk; no effect – 300 ppm (G); Reduced body wt, increased liver, spleen wt – 1444 ppm x 6 hr/d x 5 d/wk x 13 wk; no adverse effect – 737 ppm (TD) 71
Decreased body wt – 500 and 1000 mg/m3 x 90 d (male rat), ~68 and 135 ppm, based on ave.
MW = 180.
Systemic toxicity - oral
(rats) Signs of anemia, increased rel liver, adrenal gland wt – 2500 and 5000 mg/kg/day x 13 wk, no effect – 500 or 1000 mg/kg/d (hydrotreated, light petroleum distillate)72.
Decreased body wt, incr. liver enzymes - 750, 1500 or 3000 mg/kg/d x 90 d (rat).
Subchronic dermal toxicity Severe skin irritation, organ
damage – 6.4 g/kg/d x 3 d/wk x 4 wk (rabbit).
Reversible skin lesions – 0.16 mL/day nonoccluded x 4 weeks (rat).
Reproductive toxicity in
rats: No fetotoxicity or developmental effects – 900 ppm x 6 hr/day on gestation days 6-15 (G); No maternal or fetal effects – 1200 ppm x 6 hr/d on gestation days 6-15 (C).
No maternal toxicity or developmental effects – 100 or 400 ppm on gestation days 6-15 (rat).
Maternal toxicity – 1000 mg/kg/d; fetal toxicity – 1500 mg/kg/d; no malformations – 2000 mg/kg/d, gestation days 6-15; No reproductive effects on males – 3000 mg/kg/d for 70 d (rat).
Genotoxicity Negative Ames test (G, L, M, 130); negative E. coli bacterial mutation assays (G); negative for mutagenicity in vivo in mouse micronucleus test and/or rat dominant lethal test (G, M, C);
negative mutations in mouse lymphoma cells, Sister chromatid exchanges in CHO cells (130) .
Negative - Ames test;
micronuclei in the bone marrow or peripheral blood; dominant lethal assay; Positive mouse lymphoma assay; chromosomal aberrations in bone marrow cells.
Neg - Ames test, mouse lymphoma assay, dominant lethal assay in rats and mice, mammalian micronucleus test; Positive in UDS Assay.
OEL 300 ppm (M, G; ~2000 mg/m3) 73,74 400 ppm (C) 75 based on product toxicity data; 1200 mg/m3, (G, M, H); 1400 mg/m3 (C) based on GGV/RCP method 76, 77
200 mg/m3 (~29 ppm, based on
ave mol wt = 170) 78 200 mg/m358
a Numbers in parentheses refer to the solvent product (Isopar G, L, M, C, Soltrol 100, 130, Shell Sol 71, TD, TS)
b Tox data on Isoparaffinic solvents from Refs 71, 72, 73, 79, 80
c Jet A data is from reference # 70; it cites several API studies for Jet A-+
d JP-8 data is from reference # 58, Appendix E
Table 3-16. Toxicity Data for Isoparaffinic Solvents and Jet Fuels
Using the data for similar isoparaffinic solvents, it can be concluded that Bio-SPK has very low acute oral, dermal, and inhalation toxicity. It is likely to be a slight skin irritant, but may be a moderate or severe irritant on damaged skin. It is not a skin sensitizer. It is expected to be a slight eye irritant and is not a respiratory irritant. Long term exposure is expected to cause mild liver effects and reduced body weight in animals exposed to 900 ppm or more for 12 weeks, with no effect at 300 ppm. Subchronic oral dose could cause liver, and blood effects at 2500 mg/kg/d or more, with no effect at 500-1000 mg/kg/d. Bio-SPK is not expected to cause genotoxicity, reproductive effects, or carcinogenicity. The available data for similar isoparaffinic solvents address all of the testing required in DOD, 2004, Appendix E 58. Direct comparison of toxicity test results for similar isoparaffinic solvents and current jet fuels allows only very general conclusions due to differences in experimental protocols. The acute toxicity data is not helpful because the lethal doses were higher than the tested doses for jet fuels and paraffinic solvents. Likewise, comparison of the irritation and sensitization data does not show clear differences because of different protocols and mild effects. There are data suggesting that the jet fuel may be a stronger eye irritant, and have some potential for skin sensitization, but the results are not consistent for Jet A and JP-8 tests. Sub-chronic inhalation and oral studies indicate that relatively mild effects occur at lower oral doses
or inhalation concentrations of jet fuel compared with paraffinic solvents. The genotoxicity data also indicates some positive results for jet fuels and consistently negative results for paraffinic solvents. The similar mixture approach would lead to the adoption of OELs based on data for similar mixtures to Bio-SPK. Exxon recommended occupational exposure limits of 300 ppm for Isopar M and G 73, 74 and 400 ppm for Isopar C75, based on the toxicological data for the specific mixtures shown in Table 3-16.
Similarly, a recommended OEL of 400 ppm was set for Soltrol 200, a C13-C17 isoalkane mixture81. These exposure limits and the data that they are based on indicate substantially lower toxicity of isoparaffinic mixtures compared with jet fuel. More recently, Exxon has elected to use the GGV/RCP approach as described by McKee et al.62, to develop OELs for and has adopted occupational exposure limits of 1,200 mg/m3 for the Isopar G, H, and M, and 1400 mg/m3 for the Isopar C.
3.3.6 FT-SPK
The US Air Force is in the process of completing toxicological studies on a natural gas derived SPK made using the Fischer Tropsch (FT) process82. When the results of these studies are available, they will provide data on another similar mixture and the toxicological data for FT-SPK will be applicable to Bio-SPK based on the similar mixture approach.
3.3.7 Summary
No toxicological data is yet available for SPK derived from biological sources.
Available methods for evaluation of complex mixtures, were reviewed and applied to Bio-SPK. A method based on addition of groups of chemically related components (GGV/RCP), and an approach that uses compositionally and toxicologically similar mixtures, are both applicable to the evaluation of Bio-SPKs. Occupational exposure limits developed from these methods were of a similar magnitude and were much higher than the current limits for jet fuel. The GGV/RCP approach, yielded occupational exposure limits of 1,100–1,200 mg/m3. The use of a similar mixture leads to use of an exposure limit recommendation of 300 ppm, equal to approximately 2,000 mg/m3, using data from paraffinic solvent products. This evaluation leads to the conclusion that Bio-SPKs would be expected to be considerably less toxic that current jet fuels. It is expected to have very low acute toxicity, to be mildly irritating to the skin, although exposure to damaged skin or for long durations might cause more severe irritation. Finally, long-term inhalation exposure is expected to be a very mildly toxic hazard with possible liver and CNS effects. These conclusions are supported by considerable data on similar mixtures, some data on components, and by the relatively simple nature of the mixture. The conclusion is limited by the fact that most of the data on similar mixtures are relatively old comparisons of results from different studies on different materials and is made difficult by differences in methods and technology used and limited toxicity testing is available for paraffins with nine or more carbons.