Arch Toxicol DOI 10.1007/s00204-015-1526-5

ANALYTICAL TOXICOLOGY

Concentrations in human blood of petroleum hydrocarbons associated with the BP/Deepwater Horizon oil spill, Gulf of Mexico Paul W. Sammarco1   · Stephan R. Kolian2 · Richard A. F. Warby3 · Jennifer L. Bouldin4 · Wilma A. Subra5 · Scott A. Porter1,2,6 

Received: 4 March 2015 / Accepted: 5 May 2015 © Springer-Verlag Berlin Heidelberg 2015

Abstract  During/after the BP/Deepwater Horizon oil spill, cleanup workers, fisherpersons, SCUBA divers, and coastal residents were exposed to crude oil and dispersants. These people experienced acute physiological and behavioral symptoms and consulted a physician. They were diagnosed with petroleum hydrocarbon poisoning and had blood analyses analyzed for volatile organic compounds; samples were drawn 5–19 months after the spill had been capped. We examined the petroleum hydrocarbon concentrations in the blood. The aromatic compounds m,p-xylene, toluene, ethylbenzene, benzene, o-xylene, and styrene, and the alkanes hexane, 3-methylpentane, 2-methylpentane, and iso-octane were detected. Concentrations of the first four aromatics were not significantly different from US National Health and Nutritional Examination Survey/US National Institute of Standards and Technology 95th percentiles, indicating high concentrations of contaminants.

* Paul W. Sammarco [email protected] 1

Louisiana Universities Marine Consortium (LUMCON), 8124 Hwy. 56, Chauvin, LA 70344, USA

2

EcoRigs Non-Profit Organization, 6765 Corporate Blvd., Suite 1207, Baton Rouge, LA 70809, USA

3

The Warby Group LLC, 244 Park St B14, North Attleborough, MA 02760, USA

4

Department of Biological Sciences, Ecotoxicology Research Facility, Arkansas State University, PO Box 599, State University, AR 72467, USA

5

Subra Company, Louisiana Environmental Action Network (LEAN), and Lower Mississippi Riverkeeper, PO Box 9813, New Iberia, LA 70562, USA

6

EcoLogic Environmental, Inc., PO Box 886, Houma, LA 70361, USA







The other two aromatics and the alkanes yielded equivocal results or significantly low concentrations. The data suggest that single-ring aromatic compounds are more persistent in the blood than alkanes and may be responsible for the observed symptoms. People should avoid exposure to crude oil through avoidance of the affected region, or utilizing hazardous materials suits if involved in cleanup, or wearing hazardous waste operations and emergency response suits if SCUBA diving. Concentrations of alkanes and PAHs in the blood of coastal residents and workers should be monitored through time well after the spill has been controlled. Keywords  BP–Deepwater Horizon · Oil spill · PAH · Alkanes · Blood · VOC

Introduction The 2010 BP/Deepwater Horizon oil spill was the largest marine spill in the USA (Joye et al. 2011; Bolt 2014). It lasted for almost 3 months and leaked ~0.8–1.1 billion L of crude oil into the northern Gulf of Mexico (GOM). The spill was treated with >6.0 million L of dispersant (Corexit 9527 and 9500A; Judson et al. 2010). The spill covered up to 62,159 km2 (Norse and Amos 2010). The water column (Sammarco et al. 2013), benthos (Joye et al. 2011), commercial seafood (Tunnell 2011), and human health (Solomon and Janssen 2010) were all affected by the spill. A spill of this magnitude has important impacts on human health and implications for oil spills on a global basis. This is particularly true, since recent evidence indicates that this well may still be leaking and releasing crude oil (Kolian et al. 2013, 2015). Crude oil is composed of 1000s of organic compounds (Bjorlykke 2011) with varying volatilities (including

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volatile organic compounds; VOCs, BTEX—benzene, toluene, ethylbenzenes, etc.; USGS 2011) and levels of toxicity for marine biota (Ryerson et al. 2011) and humans (Baars 2002). Polycyclic aromatic hydrocarbons (PAHs) are highly toxic compounds of light crude and can bio-concentrate in marine organisms (Meador 2003) and humans (Knap et al. 2002; Fleming et al. 2006). In addition, oil combined with dispersants, as was the case in this spill, are known to be more toxic than either oil or dispersant alone (National Research Council 2005; Zhang et al. 2013; Polli et al. 2014). Up to 10 % of crude oil can be made up of PAHs. They are often used as an indicator of the general distribution of petroleum hydrocarbons in a spill environment (Vinas et al. 2010). VOCs can have negative effects on human health. Human exposure pathways include dermal contact, inhalation, and ingestion (Fingas 2000). These organic compounds are lipophilic and easily taken up by human tissues (Cheng et al. 2010) (e.g., fat, kidneys, liver, blood, etc.) and are known to be toxic to the human nervous and immune systems. Cancer and leukemia are known risks of exposure to some of these aromatic compounds. Symptoms in residents, tourists, and cleanup workers in the coastal areas affected by the BP/DWH spill have experienced respiratory distress, headaches, and skin problems (McCauley 2010). The latter group suffered from shortness of breath, nausea (Gardner 2010), and gross hematuria (blood in the urine; Rivera et al. 2012). The symptoms associated with this study were more severe and acute. In a spill situation, and particularly in this spill, people who came into contact with crude oil included workers in cleanup operations, fishers in the GOM, SCUBA divers in the region, and individuals living in the coastal communities. In the case of fishers, they became exposed to hazardous materials not only via inhalation but also by contact with their fishing gear and also via contaminated clothes or direct dermal contact with oil. People with exposure to high concentrations of crude oil, either through inhalation or direct dermal contact, can exhibit a variety of tell-tale physiological and behavioral signs and symptoms. For example, hydrogen sulfide, associated with crude oil, can cause acute or chronic central nervous system effects (Solomon and Janssen 2010). Kerosene mid-range compounds are known to cause neurotoxicity, central nervous system depression, ataxia, hypoactivity, and prostration, through both inhalation and dermal exposure (Koschier 1999). Experiments performed exposing of Pembina Cardium crude oil to cattle caused tremors, nystagmus, vomiting, and pulmonary stress (Khan et al. 1996). Gohlke et al. (2011) have reviewed and analyzed previous large oil spills in terms of their protocols and risks associated with PAHs and metals. One of their conclusions was that the duration of the post-spill analysis needed to be extended.

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Arch Toxicol

Here, we analyzed data on concentrations of certain petroleum hydrocarbons—particularly VOCs, alkanes and PAHs—found in human blood within subjects exposed to crude oil, as an indicator of human health. We focused on those organic ring compounds/PAHs and alkane carbon chain compounds which were reported by the analytical laboratory. PAHs within crude oil are known to be toxic and carcinogenic, in turn making them a health concern for responsible federal agencies (Bolger and Carrington 1999; Ylitalo et al. 2012). The objectives of this study were to examine the concentrations of petroleum hydrocarbons in human blood. Samples were drawn from patients who exhibited symptoms or signs of exposure to toxic petroleum hydrocarbons. This study was a response to a given environmental situation, i.e., it was an a posteriori study, not a pre-planned a priori one. Ideally, it would have been best to have preplanned the study and collected samples from equal numbers of patients—one group exhibiting symptoms and the other not, or samples from one group prior to the spill and one afterward. This, however, was not possible. Thus, we compared our “treatment” data with “control” data derived from the US human population overall. These “control” data were available through NHANES/NIST and provided information on concentrations of specific toxins in human blood on a percentile basis. Here, we will compare our results to US NIST standards.

Materials and methods General Samples of human blood were collected in the post-spill period between December 2010 and February 2012. They were collected from 69 adults in the GOM, male and female, ranging between 20 and 78 years of age, as well as from eight children, 2–13 years of age. Since it was not the object of this study to compare hydrocarbon concentrations between age groups or genders, but rather determine overall blood concentrations, data were pooled for all subjects. All subjects were GOM residents who had either inhalation or dermal exposure or both to crude oil and/or dispersant and reported medical problems assumed to be derived from exposure to these materials (see Diaz 2011; D’Andrea and Reddy 2013). Signs included uncontrollable shaking in the extremities and lack of muscular control or tenor in the facial muscles (Michael Robichaux, M.D., pers. obs., pers. comm., 2010–2012; also see Suarez et al. 2005; Sim et al. 2010; Levy and Nassetta 2011; Ha et al. 2012). They were exposed to the following contaminated environments prior to sampling: seawater, air, wetlands, or a sandy beach. Whole blood samples were analyzed for VOCs by gas

Arch Toxicol

chromatography/mass spectrometry, following the procedures of current procedural terminology (CPT) code 84600 (test #0762) (Blount et al. 2006). Blood analyses were performed by Genova Diagnostics/Metametrix Clinical Laboratory in Duluth, GA, USA, which is licensed/accredited by Clinical Laboratory Improvement Amendments (CLIA), the Georgia Department of Community Health, and the National Environmental Laboratory Accreditation Conference (NELAC; see Chirinos et al. 2008; Lord and Bralley 2008; McDaniel et al. 2008, for descriptions of similar blood analyses performed at Genova/Metametrix). Genova Diagnostics/Metametrix also participates in proficiency tests with the College of American Pathologists (CAP), New York State, Wisconsin State Laboratory of Hygeine (WSLH), State of Pennsylvania, Quebec Multielement External Quality Assessment Scheme (QMEQAS), and Innovation Beyond Limits (IBL) International. It is known that inhalation exposure to PAHs or other contaminants (Lewtas et al. 1997; Abraham et al. 2005; Jakubowski and Czerczak 2009) results in temporary contamination of the blood in humans and rats, including DNA adducts, and lasts a short period of time. Dermal exposure, however, results in higher concentrations of these contaminants (Van Rooij et al. 1993; Semple 2004; Cirla et al. 2005; Alikhan and Maibach 2011; Augusto et al. 2012), reaching higher concentrations and most likely being released over a longer period of time, potentially resulting in serious effects, including death (Alikhan and Maibach 2011). Statistical analyses Petroleum hydrocarbon concentration data were compared to the NHANES 95th percentile value for the distribution of a given compound known for the general human population. The test used was “Comparison of a Single Observation with the Mean of a Sample” (Sokal and Rohlf 1981). In a few cases, multiple values were reported for the NHANES value. In those cases, we used the mean of those values to test our samples against. Percent data were transformed by square root of (Y + 0.5) for normalization purposes as part of the analysis, if required (Sokal and Rohlf 1981). In addition to the mean of the human blood data, additional descriptive statistics were calculated, including the standard deviation, 95 % confidence limits, range, and minimum and maximum values for petroleum concentrations. Average concentrations were reported for all compounds observed here. Raw means, standard deviations, sample sizes, range, and 95 % confidence limits are reported. Concentration data were reported for all samples. Concentrations below detectable limits were considered to be, conservatively, zero. Zero concentration data were not included in the

percentile analyses because explicit percentile information for these levels was not afforded to us by the reporting laboratory. Percentile NHANES data falling above the 95th percentile were (conservatively) considered to be 95, as precise percentile data above this level were also not reported to us. Whether a datum or average contaminant level reported here falls within the 95 % confidence limits of the US NHANES data or not, does not necessarily imply that it represents an acceptable level of exposure. It merely demonstrates whether that point falls within a certain range characteristic of the US population. Data regarding “acceptable” or “unacceptable” levels of exposure or concentrations in the blood of a contaminant are generally derived from specific toxicity tests not pursued in this study.

Results Average concentrations for petroleum hydrocarbons in the blood are reported in Table 1. Two sets of compounds were found in the blood of the human subjects. The aromatic compounds found were m,p-xylene, toluene, ethylbenzene, benzene, o-xylene, and styrene (Fig. 1). The alkanes were hexane, 3-methylpentane, 2-methylpentane, and iso-octane. Blood samples exhibited aromatic petroleum hydrocarbon concentrations averaging 0.71 for m,p-xylene (range 0–3.48 ppb), 0.55 ppb for toluene (range 0–28.69 ppb), 0.25 ppb for ethyl benzene (range 0–1.18 ppb), and 0.06 ppb for benzene (range 0–3.0 ppb) (Table 1). Levels found for other aromatics were, for o-xylene, an average of 0.02 ppb (range 0–1.08 ppb), and for styrene, 0.005 ppb (range 0.0–0.12 ppb). The mean concentrations for the alkanes hexane, 2- and 3-methylpentane, and iso-octane, ranged between 8.34 ppb for iso-octane and 175.2088 ppb for hexane. Average concentrations for all compounds varied between individuals. With respect to individual compounds, average concentrations of various VOCs in human blood also varied to different degrees from the NHANES 95th percentiles, used for comparison. When the observed aromatic petroleum hydrocarbon concentrations and their 95 % confidence limits were graphed against the 95 % NHANES/NIST limits, variation in each suggested which compounds were within and outside of those limits (Fig. 2). Some compounds appeared to be significantly higher than those concentrations considered to be high in the US population. These were toluene, m,p-xylene and ethyl benzene-ring compounds. Concentrations of the other aromatic compounds detected all appeared, and we emphasize appeared, to fall below the 95 % NHANES levels. These were toluene, benzene, o-xylene, and styrene. The four alkanes detected in the samples all appeared to be below the NHANES 95th percentile levels (Fig. 3).

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13 n.s 77.1 65.71 16.417 15

n.s.

77.1 68.93 22.740

15

80.44 57.42

 Significance Percentile (arcsine)  95th percentile  Mean  SD

 n

 95th % upper  95th % lower

16

74.02 57.40

90.47 68.90 98 25

62

95.67 89.36 98 25

95 79.7 22.01

95 92.5 12.69

 n

1.2869 −0.1906 28.69 0 n.s.

0.8340 0.5865 3.48 0 n.s.

 95th % upper  95th % lower  Maximum  Minimum  Significance Percentile (raw)  95th percentile  Mean  SD

 95th % upper  95th % lower  Maximum  Minimum

3.3074 77

0.5540

77

 SD

0.68 0.548

Toluene

 n

0.34 0.710

Concentration  95th percentile  Mean

m,p-Xylene

Aromatics

72.08 51.18

14

77.1 61.63 19.946

n.s.

93.89 87.16 98 40

76

95 90.5 14.96

0.2965 0.2009 1.18 0 n.s.

77

0.2139

0.11 0.249

Ethylbenzene

66.92 49.99

15

77.1 58.46 16.734

n.s.

82.61 58.64 98 40

16

95 70.6 24.46

0.1421 −0.0125 3 0 n.s

77

0.3462

0.26 0.065

Benzene

54.86 44.11

15

77.1 49.49 10.618

***

64.84 48.76 97 40

15

95 56.8 15.90

0.0534 −0.0030 1.08 0 n.s.

77

0.1262

0.09 0.025

o-Xylene

56.00 40.29

15

77.1 48.14 15.527

n.s.

64.28 41.85 98 40

15

95 53.1 22.16

0.0098 0.0001 0.12 0 ***

77

0.0218

0.12 0.005

Styrene

39.90 30.97

70

77.1 35.43 19.070

*

41.79 29.71 98 2

77

95 35.8 27.05

188.1857 162.2319 400.000 0.000 n.s.

77

58.09899

239.0 175.2088

Hexane

Alkanes

41.93 33.22

77

77.1 37.57 19.515

n.s.

45.79 32.99 98 2

77

95 39.4 28.67

92.0840 79.9316 199.000 32.800 *

77

27.20382

146.0 86.0078

3-Methylpentane

34.65 27.01

77

77.1 30.83 17.107

**

34.61 23.60 98 2

77

95 29.1 24.64

47.3335 41.2795 94.000 16.200 **

77

13.55214

86.0 44.3065

2-Methylpentane

33.51 24.87

77

77.1 29.19 19.334

*

32.38 20.56 98 2

77

95 26.5 26.46

10.6637 6.0116 97.000 2.000 n.s

77

10.41398

20.4 8.3377

Iso-octane

Table 1  Mean concentrations of volatile organic compounds (VOCs) found in the blood of human subjects, sampled between December 2010 and February 2012; (the oil spill was capped in July 2010). Mean, standard deviation, sample size (n), range, and 95 % concentrations shown.  Mean percentile of concentrations also shown, in comparison to NHANES 95th percentile values for the US populations.  Same descriptive data also shown for the percentile data transformed by arcsine, for data normalization purposes

Arch Toxicol

**

*

When these data were subjected to statistical analyses; however, significant similarities and differences between measured compound averages and the NHANES 95th percentiles emerged (Table 1). Statistical analyses revealed similar results to the graphic analysis, but in greater detail, with an approximate doubling of the number of compounds being consistent with the NHANES 95th percentiles. The observed average blood concentrations for four aromatic compounds were not significantly different than the 95 % NHANES/ NIST levels (US Center for Disease Control and Protection 1999, 2010)—m,p-xylene, toluene, ethyl benzene, and benzene. This indicated that concentrations of these compounds in the blood were significantly higher than normal. These similarities were detected whether tested by concentration, percentile, or arcsine-transformed percentile. The only aromatic compound in the study significantly and consistently below the 95th percentile was o-xylene. Some statistical tests returned significant results for a given compound, but different tests yielded varying results, making the results equivocal. For example, this was the case with the aromatic compound styrene (Table 1). The results were also equivocal for the alkanes. Concentrations of hexane were high when calculated by concentration. Its percentile ranking, however, was low when considered either by raw or transformed percentage. Results were similar for 3-methylpentane and iso-octane. 2-Methylpentane consistently exhibited concentrations significantly lower than the NHANES 95th percentile.

* p < 0.05; ** p < 0.01; *** p < 0.001; n.s. not significant

Discussion * Significantly different from (lower than) the NHANES 95th percentile data

In that case, the sample mean would have a particularly high value

Lack of significance means no significant difference between the 95th percentile and the sample mean

Significance indicates significant difference between 95th percentile value and sample mean

Units = ng/mL (ppb)

n.s. mean of measured data nor significantly different than the 95th percentile data for NHANES data, i.e., high toxin concentrations

* n.s. n.s.  Significance

n.s.

n.s.

n.s.

*

*

81.87 8.13 81.87 8.13 81.87 8.13 81.87 8.13 81.87 39.23 81.87 0.00

81.87 30.00

81.87 39.23

81.87 39.23

80.03 39.23

Hexane Styrene o-Xylene m,p-Xylene

 Maximum  Minimum

Table 1  Continued

Aromatics

Toluene

Ethylbenzene

Benzene

Alkanes

3-Methylpentane

2-Methylpentane

Iso-octane

Arch Toxicol

The concentrations of a number of aromatic petroleum hydrocarbons considered in this study were well above average, as revealed by comparison with NHANES 95th percentile data (US Center for Disease Control and Protection 2010). Concentrations of a number of compounds were significantly below the NHANES 95th percentile levels—specifically, one to two aromatic compounds, and all four alkanes. This was indicated by either non-significant or equivocal statistical test results. Some average concentrations were well above average for the US population (two times >95th percentile; US Center for Disease Control and Protection 1999, 2010). The values were often an order of magnitude higher than the median (50th percentile) NHANES level in the US population (US National Institute for Occupational Safety and Health 2007). Exposure to petroleum hydrocarbons carries with it significant human health risks in the short and long term (White 1986; Wong and Raabe 2000; US National Institute for Occupational Safety and Health 2007; Ha et al. 2008). Exposure should be avoided or minimized through the use of personal protective equipment.

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Arch Toxicol

Fig. 1  Chemical structure of volatile organic compounds (VOCs) found in human subjects’ blood. Upper line represents aromatic compounds, lower line represents alkanes– carbon chain compounds

Fig. 2  Bar diagram of petroleum hydrocarbon concentrations found in blood samples of up to 77 human subjects complaining of symptoms of exposure to crude oil during and after the BP/Deepwater Horizon spill. Blood samples collected between December 2010 and February 2012 (spill site capped in July 2010). Aromatic compounds shown here. Mean concentrations shown with 95 % confidence intervals and ranked by level. Dashed lines represent 95th percentile values for these VOCs in the US human population, as reported by NHANES (US Center for Disease Control and Protection 1999, 2010). For the first four compounds, means are above or equivalent to (not significantly different from) the 95th percentile NHANES values, indicating high toxicity (see Table 1). The last two compounds are equivocal in this regard. Data analyzed via “Comparison of a Single Observation with the Mean of a Sample” (Sokal and Rohlf 1981)

Fig. 3  Bar diagram of petroleum hydrocarbon concentrations found in blood samples of up to 77 human subjects complaining of symptoms of exposure to crude oil during and after the BP/Deepwater Horizon spill. Alkane compounds shown here. Mean concentrations shown with 95 % confidence intervals and ranked by level. Dashed lines represent 95th percentile values for these VOCs in the US human population, as reported by NHANES (US Center for Disease Control and Protection 1999, 2010). Mean concentrations are close to NHANES 95th percentiles, but results between statistical tests for any given compound are equivocal, except for 2-methylpentane which is significantly lower than the 95th percentile NHANES value in all tests (see Table 1). Data analyzed via “Comparison of a Single Observation with the Mean of a Sample” (Sokal and Rohlf 1981) (see legend of Fig. 2 for additional details)

One point which was evident from the data was that all of those compounds which were apparently retained in the body (m,p-xylene, toluene, ethylbenzene, and benzene, evident through particularly high concentrations) were aromatics, possessing a single-ring structure. Most of those

compounds which exhibited average concentrations lower than the 95th percentile NHANES values (hexane, 3-methylpentane 2-methylpentane, and iso-octane) were alkanes or long-chain hydrocarbons. It would appear that the aromatics represent a more persistent threat to human health.

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Arch Toxicol

Some of these subjects were not sampled until 1.5 years after the BP/Deepwater Horizon oil spill. The body may have a mechanism by which to clear or metabolize alkanes more readily than aromatic hydrocarbons. On the other hand, it is also possible that the alkanes may have been accumulated elsewhere in the body (liver, adipose tissue, etc.) and may no longer be evident in the blood. It is possible that uptake of these crude oil components was facilitated by Corexit® and one of its key components, a toxic solvent—2/BTE (2-butoxyethanol), which has been shown to facilitate this process (Burns and Harbut 2010). In addition, 2/BTE is known to be associated with human health problems, some of which overlap with symptoms observed here. For example, Corexit 9527A has been shown to rupture red blood cells, causing hemolysis and liver and kidney damage (Johanson and Boman 1991; Nalco 2010; see Earle et al. 2010 for further discussion). The people sampled represented a variety of ages, had different backgrounds, and may have lived different distances from the spill site. In addition, they may have been tobacco smokers or worked in industrial occupations which may have affected the levels of petroleum hydrocarbons in their blood. It is unlikely, however, that on the average, most of them would exhibit petroleum hydrocarbon concentrations above the national 95 % confidence limits under these conditions. What they all do have in common is some exposure to the BP/Deepwater Horizon oil spill. The patients whose blood data were considered here were exposed to crude oil through dermal contact or inhalation or both. Although it is likely that a certain components of the oil were dissolved in the water before the oil reached the surface after its 1500 m ascent (Ryerson et al. 2011), the high volumes of crude oil being released would have insured that even a small proportion of the spilled oil reaching the surface would still carry with it a substantial volume of benzene. BTEX in general are highly volatile, and any BTEX that successfully rise to the surface rather than dissolve in the water column could be expected to evaporate within 24 h of surfacing (Jordan and Payne 1980). With the constant high-volume renewal of surface oil for a period of 2 months, however, concentrations of BTEX could be expected to be almost continuously high in the atmosphere in the vicinity of the spill site. Some data suggest, however, that airborne BTEX concentrations measured for offshore workers were low and that the BTEX concentrations arising from the surfaced oil were negligible compared to other sources such as exhaust from boat engines (Avens et al. 2011). Under normal conditions, one might expect BTEX to evaporate within 1 day after capping. Another factor is that the dispersant Corexit was used continuously during the spill and afterward, drawing all petroleum hydrocarbons, including BTEX (benzene, toluene, ethylene, and xylene), down into the water column for months during and after the spill. It has been calculated that

the half-life of benzene in the blood is <1 day (Brugnone et al. 1992). Considering that the Macondo well was capped in July 2010 and that all blood samples were taken between December 2010 and February 2012 from divers and offshore workers, the question arises as to why strongly positive results were obtained for some of these VOCs. This raises the question of retention by the body, the influence of a potential continuing leak, or substantial seeps in the region. Evidence of the second of these has surfaced since the capping of the well (Kolian et al. 2013, 2015). It is important that during and after a crude oil spill, individuals avoid inhalation of fumes from, or direct dermal contact with, the oil, particularly if it has been treated with a dispersant. This can be done by avoidance of the affected region. In the case of industrial contact, this should primarily be done through the use of hazardous materials suits (HazMat suits). In the case of SCUBA diving, hazardous waste operations and emergency response (HazWoper) suits are recommended. Monitoring of VOC contamination in human blood derived from those working with, or in the vicinity of, a spill should be continued through time after a spill event. The presence of VOC in the blood of patients who have been associated with the spill well after the spill has ceased suggests that investigations should be made regarding the possibility of retention of the contaminants by those exposed to the crude oil, continuing leaks after capping, or seeps, natural or newly formed, associated with the spill site. Subjects The home institution of this study (LUMCON) does not possess an Institutional Research Board (IRB). For this reason, we consulted an external IRB system to determine whether the research required oversight by an IRB. We used guidelines used by Cornell University, Ithaca, NY, and followed advice received from its “…Research Involving Secondary or Existing Data, Documents, or Biological Specimens…” document (http://www.irb.cornell.edu/documents/IRB%20Decision%20Tree.pdf). This guide led us to the decision that this project was not considered to be Human Subjects Research and that IRB review or oversight was not necessary. This was primarily because (1) members of our research team had no contact with and were not involved with the subjects, (2) they were also not involved with collection of the samples, (3) only numerical data were provided to us and these could not be linked back to the identity of any of the subjects, and (4) the provider of the data (an M.D.) was not a collaborator/co-author on the study. For this type of study, formal consent is not required. Acknowledgments We recognize and thank Dr. Michael Robichaux, M.D. for his sincere dedication to the health of his patients in the Gulf of Mexico region during this challenging period.

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We also thank him for making blood data available for statistical analysis within the study. We also thank Ms. Mary Lee Orr of the Louisiana Environmental Action Network (LEAN) for her support of the sample processing and also making the resultant data available for analysis by our team. Many thanks to M. Genazzio and D. Beltz who assisted with data analyses and graphics. Thanks as well to Ms. B. Wiseman—The Earth Organization; Mr. David Fa-Kouri—Louisiana Economic Foundation (LLC), and Principal Strategic Economist/ Researcher, Strategic Consulting Group, Inc; and, Mr. Andrew Blanchard, Indian Ridge Shrimp Co., Chauvin, LA, USA for supplying the initial impetus for this and related studies of ours.

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