STANDARD OPERATING PROCEDURE (SOP) FOR ANALYSIS OF

ALCOHOL TRACERS USED AS PARTITIONING TRACERS

(SOP-UF-HiII-95-07-OO1O-v.2)

1.0 SCOPE AND APPLICATION

1.0.1. This SOP describes the analytical procedures utilized by University of Florida (UF) for analysis of alcohols used as partitioning tracers, in both laboratory and field studies, to quantify the amount and distribution of residual non-aqueous phase liquids ~APLs) present in the saturated zone.

1.0.2. This SOP was written by D.P. Dai, H.K. Kim, and P.S.C. Rao at the Universitv of Florida. This version (v.2) was updated on July 19, 1995. The method described here was modified from a protocol provided by Professor Gary Pope at the University of Texas-Austin.

1.0.3. The alcohol tracers used in the UF laboratory and field studies were: methanol; ethanol. n-butanol; n-pentanol; n-hexanol: 2,2-dimethyl-3-pentanol; and 6-methvl-2-heptanol

1.0.4. The method involves gas chromatography (GC) techniques for estimation of the concentrations of alcohol(s) in water samples; a flame-ionization detector (FID) is used to quantify the analyte concentrations in the sample. The method has been found to provide reliable and reproducible quantitation of alcohol tracers for concentrations > 1 mg/ml; this value is then considered to be the Method Detection Level (MDL). The standards calibration curve for FID response was found to be linear up to 3,000 mg/ml for ethanol and methanol, and it may be linear even beyond this value, but was not tested. (In laboratory and field experiments, the input concentrations of the alcohol tracers did not exceed this upper limit.)

1.0.5. Samples selected for GC-FID analysis may be chosen on the basis of preliminary screening to determine approximate concentration ranges. and select appropriate GC parameters (e.g., sample injection volumes; concentration range for standard curves, etc.) However, the strategies for sample screening themselves are outside the scope of this SOP.

1.0.6. Water samples from laboratory and field experiments are sub-sampled into 2-ml GC vials for analysis.

2.0 PURPOSE

2.0.1. The purpose of this SOP is to ensure reliable and reproducible analytical results of alcohols in water samples for laboratory-based or on-site (field-based) OC-FID analyses, and to permit traceability of possible causes of error in analytical results.

3.0 PROCEDURES

3.1 Sample Containers, Collection, Transportation and Storage

3.1.1. Sample Containers. Water samples are contained in 5-ml glass sample vials (Fisher Catalog # 06406-19F) with Teflon-faced septa caps. The glass vials or the caps are not reused.

3.1.2. Sample Collection. Each sample vial is completely filled with aqueous samples, such that no headspace of air exists, and capped. The vials are not opened until the time of sub-sampling or analysis.

3.1.3. Transportation and Storage. For field studies, the samples are stored in coolers containing "blue ice", and later stored in refrigerators in a trailer located on the site. Samples may be subjected to on-site GC analysis, and/or shipped back to UF laboratories; samples are packed in coolers and shipped via overnight air express (e.g., FedEx). The samples are stored in the cold storage room or refrigerator at 4 ºC, until they are ready for GC analysis. After sub-sampling. the samples are returned to cold storage. For laboratory studies, the samples are stored in a refrigerator if the period prior to analysis is expected to exceed eight hours.

3.2. Sub-Sampling and Dilution

3.2.1. Disposable, Pasteur glass pipettes (Fisher Catalog # 13-678-20B) are used to transfer samples from 5-ml sample vials to the 2-ml GC vials. Samples may need to be diluted with HPLC water prior to GC analysis. The dilution (usually 10x to 100x) necessary is determined from preliminary screening analysis.

4.0 APPARATUS AND MATERIALS

4.0.1. Glassware. Disposable micro-pipettes (10, 100 ~L; Fisher Catalog # 21-175B: 21-175F) and Class A volumetric pipettes (1 or 2 ml) are required for sample dilution. Disposable Pasteur glass pipettes (Fisher Catalog # 13-678-20B) are required for sub-sampling. GC vials (2-ml) with Teflon-faced caps (Fisher Catalog * 03-375-16A) are required for GC analysis. Volumetric Class A pipettes (0.5, 1, 2, 5, 10 ml) are required for preparations of the calibration standards.

4.0.2. Gas Chromatograph System. An analytical GC system is required, complete with a temperature-programmable oven, an auto-injector suitable for on-column injection, and either an integrator or a PC-based data acquisition/analysis software. Also required are other accessories, including analytical columns and the gases required for GC-FID operation.

4.0.3. Two types of GC systems were used at UF; a Shimadzu 14-A system equipped with an autosampler and FID was used to analyze water samples in laboratory and on site; while a Perkin Elmer Autosystem with an HD and an integrated autosampler was used for some laboratory analyses. The Shimadzu GC was linked to Shimadzu CR-601 Chromatopac integrator, while the Perkin Elmer system was linked to a IBM-compatible PC loaded with Turbochrom (version 4) software.

•Column: The capillary column used was a Restek RTX 624 column (75-m long, 0.53-mm i.d.) with a 3.0-mm film thickness (Restek Catalog # 10974).

•Gases: Zero-grade air, and ultra-high purity hydrogen are used for the FID, and ultra-high purity helium is used as the carrier gas.

4.0.4. Reagents. Reagent water is defined here as the water in which an interferant is not observed at the MDL of the parameters of interest. Laboratory reagent water used is HPLC-grade or OPTTMA-grade water (Fisher Catalog #W54 and W74).

4.0.5. Standard Solutions. Analytical standard solutions are prepared from pure materials in the laboratory. Stock standard solutions, at 1000 mg/ml of each analyte, are prepared in HPLC water and kept in 22-ml glass vials (Fisher Catalog # 03-393-D) with Teflon-lined caps; minimal headspace ensures no volatile losses. These stock solutions are stored at 4 ºC.

4.0.6. Old stock solutions are discarded and a fresh batch prepared every month. Any time a comparison with the check standards indicates a problem, a new batch of standards must be prepared.

•Working Calibration Standards (Secondary Dilution Standards): Working calibration standards are prepared by diluting stock standard solutions in reagent water.

•Calibration Check Standards: Calibration check standards are prepared by diluting stock standard solution in water.

5.0 CALIBRATION

5.0.1. Calibration Standards. Prepare five calibration levels as follows: Four standards are prepared by dilution of a stock solution (e.g., 2000 mg/m), while the fifth is an undiluted standard. As an example, add 0.1, 1, 2, and 20 ml of the calibration standard (200 mg/ml) solution into 20 ml of reagent water. This gives five calibration levels (mg/ml) as follows:

Level I Level 2 Level 3 Level 4 Level 5

Analyte Concentration 0.99 9.52 18.18 100.0 200.0

6.0 QUALITY CONTROL

6.0.1. Quality control procedures that will be followed are:

•CC injector septa must be changed every 60 to 80 injections, or sooner if any related problems occur.

•Injector liner must be cleaned or changed every 60-80 injections or sooner if any related problems occur.

•An acceptable method blank analysis must be performed once for each 12-hour time period.

•Standard calibration must be run every time the flame is started.

•Check standard and blank (reagent water) should be run every 10 to 12 samples.

7.0 INSTRUMENTAL PROCEDURES

7.0.1. Gas Chromatography Condition. Recommended operating conditions are as follows:

Injection port temperature 200 ºC

FID detector temperature 265 ºC

Initial column temperature 40 ºC

Initial hold time 1 min.

Final column temperature 200 C

Final hold time 1 min.

Ramp rate 30 ºC/min.

Linear flow velocity 100/sec at 40 ºC

A typical chromatogram for a mixture of several alcohol tracers in aqueous samples analyzed using the Shimadzu GC system is attached here as Figure 1.

8.0 SAMPLE PREPARATION

8.0.1. Sub-sampling. Samples are transferred from sample vials to GC vials and capped with open-op, Teflon-lined septa caps.

8.0.2. Dilution. Prepare a sample dilution if responses for any peaks in the sample exceed the calibration ranges. The dilution should allow the analyst to obtain the greatest response within the analytical range, or refer to screen results to determine if initial dilution is required. For on-site analyses, samples were diluted if screening analyses indicated that the alcohol tracer concentrations exceeded 200 mg/ml

9.0 SAMPLE ANALYSIS

9.0.1. Analysis. The samples are allowed to reach ambient temperature prior to GC analysis. The vials with sub-sampled (and diluted, if necessary) samples are loaded on the GC auto-injector; 1 -mL sample size is suggested for all samples and standards. (Note that on certain makes of GC, the FID flame may be extinguished if 1 mL aqueous sample is directly injected, thus requiring split-sample injector or dilution with a non-aqueous solvent. Both options may result in decreased levels of detection.)

9.0.2. Analyte Identification. Analyte identification is not based on absolute retention time. A relative retention time for each analyte of interest is established by comparison to a specific "standard' peak (e.g., the solvent peak; a major component peak; a spiked internal standard or reference). The analyte of interest must elute at the same relative retention time as the standard; small changes in the absolute retention times might be expected due to variations in flow rate, or column temperature, or other operational parameters. The primary criterion is that the relative retention time of the sample component must be within +/- 0.1 min. of the standard.

9.0.3. Analyte Quantitation. When an analyte has been identified, the concentration will be based on the peak area, which is converted to concentration using a linear standard calibration curve.

9.0.4. Interferences. Contamination by carryover can occur whenever high-level and low-level samples are sequentially analyzed.

•To reduce carryover, the injector syringe should be rinsed with reagent water between analyses.

•Whenever a sample with an unusually high concentration is encountered, it should be followed by an analysis of reagent water to check for cross-contamination.

10.0 SAFETY

10.0.1. The main safety issue concerning the use of the GC at a field site relates to the compressed gases in use. The FID gases (hydrogen and air) form explosive mixtures. It is important to keep this in mind at all times, and be aware of the hazard potential in the event of an undetected hydrogen leak. All gas connections must be properly leak tested at installation.

10.0.2. High-pressure compressed-gas cylinders must be secured to a firm mounting point, whether they are located internally or externally. Gas cylinders should preferably be located outside the trailer on a flat, level base, and the gas lines run inside through a duct or window opening. If the gases are located outside, then some form of weatherproofing for the gauges will be necessary. As a temporary measure, heavy-duty polyethylene bags, secured with tie-wraps, have been used successfully; this may not be very elegant but it is very effective for short-term use of the GC. A more permanent protective housing must be built if the GC is located at the trailer for an extended time period. If it is not possible to arrange external siting easily, the gas cylinders should be secured to a wall inside the trailer.

l0.0.3. The main operating drawback to locating the gas cylinders externally is that it is not easy to monitor the cylinder contents from the instruments; the gas which could be used up most quickly is air for the FID, particularly if two instruments are hooked up to the same supply and they are running continuously. It may be worth having a reserve cylinder of air, just in case.

10.0.4. Miscellaneous Safety Issues. It is good laboratory operating practice to make sure the flame is attended at all times. When it is necessary to change the injection liner on the GC, the detector gases should be shut off. The column must be connected to the detector before igniting the flame. The trailer should be kept well ventilated when using the GC. Refer to the Materials Safety Data Sheets (MSDS) for additional information on environmental toxicity data; and for safety information, procedures and regulations etc.