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U.S. GEOLOGICAL SURVEY
Scientific Investigations Report 2007–5144

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Sample Collection and Analytical Methods

Sample Collection

Water

Water samples for gasoline-related volatile organic compounds (VOCs) were collected using a stainless-steel hand sampler (fig. 2A) that was lowered to the desired depth (1 meter) on either a stainless-steel cable or nylon rope (Shelton, 1997). The sampler holds four 40-mL glass vials that are flushed with approximately seven volumes of water before being filled by the final 40 mL. Immediately upon retrieval of the sampler, the sampler was opened and each vial was removed, preserved with 1:1 hydrochloric acid, capped, placed on ice, and shipped overnight to the U.S. Geological Survey (USGS) National Water Quality Laboratory (NWQL) in Lakewood, Colo.

Detection of PAHs in water samples is problematic because of their low solubility in water and their transient nature, leading to low concentrations. Semipermeable membrane sampling devices (SPMDs) were used to sample these organic compounds in the water column. SPMDs consist of a flat low-density polyethylene tube filled with triolein (lipid) and are deployed in stainless-steel canisters (fig. 2B) suspended in the water column (Huckins and others, 1990 and 1993). These devices are effective in sequestering hydrophobic organic compounds from water and are useful in assessing the bioavailability of these compounds to the biota (Bevans and others, 1996).

During May–June 2004, SPMDs were placed at three locations in Lake Mead, two locations in Lake Mohave, and one location in Las Vegas Wash. During May–June 2006, SPMDs were placed at four locations in Lake Mead and two locations in Lake Mohave. The SPMDs were placed by suspending the canisters 3-meters below the water surface from a platform or buoy chain. When the SPMDs were retrieved, they were placed on ice and shipped overnight to Environmental Sampling Technologies (sole vendor of SPMDs) in St. Joseph, Missouri, for processing.

Bottom Sediment

Lake-bottom sediment samples were collected from a boat using a small Ponar dredge (fig. 2C), which was lowered to the bottom on a nylon rope. The dredge typically sampled the upper few centimeters of lake-bottom sediment. The dredge was retrieved and then placed in a cleaned stainless steel tray where the sediment was exposed by opening the dredge. Sediment was placed into a pre-cleaned (washed with detergent, rinsed with deionized water, and baked at 450 ^oC) 1- liter glass jar with a Teflon® spatula and the jar was sealed and placed on ice. The samples were frozen as soon as possible after collection, packed on ice, and sent to the USGS NWQL.

Laboratory Analyses of Samples

Water

Water samples were analyzed for volatile organic compounds (VOCs) at the USGS NWQL. Compounds specifically targeted were BTEX (benzene, toluene, ethylbenzene, and xylene isomers), oxygenates (diisopropyl ether, ethyl tert-butyl ether, methyl tert-butyl ether, and methyl tert-pentyl ether), and degradation products of oxygenates (acetone, methyl acetate, tert-amyl alcohol, and tert-butyl alcohol). The individual compounds were quantified using a heated purge and trap followed by gas chromatography and mass spectrometry (Rose and Sandstrom, 2003). Laboratory reporting limits for BTEX and other VOC compounds analyzed are shown in table 2.

Semipermeable Membrane Devices (SPMDs)

Compounds were recovered from the SPMDs by dialysis and gel-permeation chromatography, extracted into hexane, and sealed in glass ampoules. These samples were sent to the USGS NWQL for analysis. The hexane extracts were analyzed for PAHs using gas chromatography and mass spectrometry (Furlong and others, 1996). Individual PAH compounds that were analyzed in SPMD extracts and their laboratory reporting limits are shown in table 3.

Samples collected from five sites (three from Lake Mead and two from Lake Mohave) were dialyzed with hexane, transferred to DMSO (dimethylsulfoxide) and were shipped under ice to the Microbiology Laboratory at the Columbia Environmental Research Center (CERC) for analyses. All samples were handled under subdued lighting pursuant to CERC Standards. A hand-held Picofluor® dual channel fluorometer (Turner Designs, Sunnyvale, CA) monitored SPMD extracts for bioavailable PAHs. PAHs with three or more rings are known to fluoresce. The environmental contaminant pyrene was used as the standard to estimate the concentrations of fluorescent PAHs in unknown samples. A pyrene index was developed to estimate the presence and concentrations of PAHs detected in each SPMD dialysate. The fluorescence of individual PAHs varies widely, so this method serves as an approximation of the relative concentrations and presence or absence of PAHs in the sample. The index used the linear range where the concentration range in the readout of the Picofluor® is directly proportional to the concentration range for the fluorophore. Samples were recorded as µg PAH per SPMD dialysate.

The DMSO extracts also were used to determine relative toxicity of the PAHs in the samples. Microtox bioassays were conducted according to the standard protocol for the basic test described in the Microtox Manual, Volume III (Microbics, 1992) to determine the acute toxicity of compounds detected in water from the sites. Validation of the Microtox Toxicity Assay in single and complex mixtures of pesticides, polychlorinated biphenyls (PCBs), petroleum products, and PAH has been previously reported by Johnson and Long (1998) and Kaiser and Palabrica (1991). Suspensions of a selected strain of luminescent bacteria (Vibrio fischeri, Azur Environmental, Inc. Carlsbad, CA) were exposed to each test substance in a standard four-tube plus controls 1:2 dilution series. Samples were incubated at 17^oC in a temperature-controlled incubator; light emissions were measured after 5 minutes with a luminometer (Azur Analyzer 500). Phenol and DMSO were the assay’s standard positive and negative controls, respectively; carrier solvent (DMSO) did not exceed 5 percent of the sample volume. The standard dose-response curve method was used to determine the concentration that caused a 50-percent loss of light production in the bacteria. To calculate the effective concentration (EC50) values of the test samples, supporting computer software used a standard log-linear model and expressed the mean value of three replicates with confidence intervals (mg SPMD per mL carrier solvent). Samples were designated “toxic” when their EC50 values were significantly less than the EC50 value of the trip blank (TB). The lower the EC50 value the higher the acute toxicity. A toxicity index (TI) was calculated by dividing the mean phenol control EC50 value by the exposed SPMDs EC50 value (TI = TB/unknown sample).

Bottom Sediment

The PAH compounds were extracted from the sediment samples by solvent extraction followed by partial separation using gel-permeation chromatography. The compounds were identified and quantified using capillary-column gas chromatography and mass spectrometry (Olson and others, 2004). The individual PAH compounds analyzed for in sediment samples and their laboratory reporting limits are shown in table 4.

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