7.1. BIOLOGICAL MATERIALS

Analytical methods exist for measuring heptachlor, heptachlor epoxide, and/or their metabolites in various tissues (including adipose tissue), blood, human milk, urine, and feces. The common method used is gas chromatography (GC) coupled with electron capture detection (ECD) followed by identification using GC/mass spectrometry (MS). Since evidence indicates that heptachlor is metabolized to heptachlor epoxide in mammals, exposure to heptachlor is usually measured by determining levels of heptachlor epoxide in biological media. A summary of the detection methods used for various biological media is presented in Table 7-1.

Heptachlor and heptachlor epoxide are measured in adipose tissue, blood, and serum using GC/ECD (Adeshina and Todd 1990; Burse et al. 1990; Polishuk et al. 1977a, 1977b; Radomski et al. 1971a, 1971b) and identified by GC/MS (LeBel and Williams 1986). Sample preparation steps for adipose tissue vary but, in general, involve a lipid extraction step followed by a clean-up procedure involving gel permeation chromatography (GPC) and/or Florisil column clean-up. Using GPC with methylene chloride and cyclohexane as solvents, individual organochlorine contaminants can be separated from adipose tissue to produce extracts clean enough for direct GC analysis. Clean-up efficiency using GPC is 99.9% (LeBel and Williams 1986). The sensitivity obtained using GC/ECD is in the low-ppb range. Recoveries for heptachlor are adequate (72–87%); recoveries for heptachlor epoxide are good (84–98%). Precision is good for both (Adeshina and Todd 1990; LeBel and Williams 1986). The preparation step used for measuring heptachlor epoxide in blood and serum involves lipid extraction, clean-up with column chromatography, and elution with acetonitrile, hexane, and methylene chloride (Burse et al. 1990; Polishuk et al. 1977a, 1977b). Recovery is adequate (80–96%). Precision is good (9–15%). Sensitivity was not reported (Burse et al. 1990).

Table 7-1

Analytical Methods for Determining Heptachlor and Heptachlor Epoxide in Biological Materials.

GC/ECD and GC equipped with a microcoulometric detector have been used to determine heptachlor and heptachlor epoxide in a variety of human tissues, including the liver, brain, adrenals, lungs, heart, kidneys, spleen, and pancreas (Curley et al. 1969; Klemmer et al. 1977; Radomski et al. 1968). Details of a sample preparation method were not reported for GC equipped with a microcoulometric detector (Curley et al. 1969). Sample preparation steps for GC/ECD include homogenization, extraction with petroleum ether or hexane, usually followed by a clean-up procedure (Klemmer et al. 1977; Radomski et al. 1968). Recovery, sensitivity, and precision data were not reported (Curley et al. 1969; Klemmer et al. 1977; Radomski et al. 1968).

Heptachlor and heptachlor epoxide have been measured in samples of human milk using GC/ECD and GC/MS (Mussalo-Rauhamaa et al. 1988; Polishuk et al. 1977b; Ritcey et al. 1972). Sample preparation steps for milk involve homogenization with chloroform/methanol, lipid extraction with petroleum ether, hexane or acetone-hexane, clean-up by column chromatography, and elution with acetonitrile, hexane, methylene chloride, or dichloromethane-petroleum ether. Precision, accuracy, and sensitivity were not reported for most of the studies; however, one study reported a sensitivity in the low-ppb range (Ritcey et al. 1972).

Heptachlor, heptachlor epoxide, and their metabolites have been measured in urine and feces using GC/ECD (Tashiro and Matsumura 1978). Sample preparation steps involve extraction with acetone and hexane, clean-up on Florisil and silicic acid columns, and extraction of the derivatized metabolites into hexane for GLC analysis. Precision, accuracy, and sensitivity were not reported (Tashiro and Matsumura 1978).

7.2. ENVIRONMENTAL SAMPLES

Methods exist for measuring heptachlor and heptachlor epoxide in air, water, soil, and food. The most common methods are GC/ECD and GC/MS. A summary of methods for detecting heptachlor and heptachlor epoxide in various environmental samples is presented in Table 7-2.

Heptachlor is measured in indoor and outdoor air samples using GC/ECD and GC/MS (Anderson and Hites 1989; Leone et al. 2000; Lewis et al. 1986; Savage 1989). Heptachlor has also been measured in house dust (Roberts and Camann 1989). Preparation methods involve the use of a variety of air trapping samplers. Examples of these include the Greenburg-Smith impinger, Chromosorb 102, low-volume samplers, and the Millipore miniature vacuum pump with a sampling tube. The next step includes extraction with diethyl ether, acetone-hexane, or toluene (Anderson and Hites 1989; Roberts and Camann 1989). For indoor air, sensitivity is in the sub-ppt range (Leone et al. 2000). For outdoor air, precision is good (13%) and recovery is excellent (99%). Sensitivity is in the sub-ppb range (Lewis et al. 1986).

Table 7-2

Analytical Methods for Determining Heptachlor and Heptachlor Epoxide in Environmental Samples.

Heptachlor and heptachlor epoxide are measured in water, drinking water, waste water, soil/sediment, and solid waste using GC/ECD, GC/MS, and liquid chromatography (LC)/MS/MS (Alford-Stevens et al. 1988; Canadian Ministry of the Environment 2003; Chandramouli et al. 2004; EPA 1994a, 1994b; Lopez-Avila et al. 1990; McDougall et al. 1987; Smith et al. 1987). Preparation of water, waste water, and drinking water samples involves extraction with methylene chloride, concentration, and solvent exchange to hexane or methyl tert-butyl ether. Mean recovery in water for heptachlor was low (52–68%) and precision was poor (48–57%) (Alford-Stevens et al. 1988). Poor recovery and precision data were thought to be attributable to chromatographic problems in some of the participating laboratories. For drinking water (EPA Method 508), recovery was excellent for heptachlor (99%) and heptachlor epoxide (95%). Precision was excellent for both compounds (<10%). Sensitivity was in the sub-ppb range (Lopez-Avila et al. 1990). Preparation of soil/sediment or solid waste samples involves extraction with methylene chloride, methylene chloride-acetone, methylene chloride-methanol, or acetone-hexane followed by clean-up with Florisil or GPC (Alford-Stevens et al. 1988; EPA 1994a). Overall precision was adequate to poor, ranging from 19 to 47% for heptachlor. Recovery and sensitivity were not reported (Alford-Stevens et al. 1988). EPA Test Methods 8080 and 8250 for evaluating waste water, soil sediment, and solid waste report sensitivity in the low-ppb range for both heptachlor and heptachlor epoxide (EPA 1994a, 1994b). Recovery for heptachlor is adequate (69–87%) and recovery for heptachlor epoxide is good (89–92%). Precision is adequate for both methods (EPA 1994a, 1994b).

GC/ECD is the method used to detect heptachlor and heptachlor epoxide in foods (butterfat, fruits, vegetables, milk, and animal feed) (Di Muccio et al. 1988; Hopper and Griffitt 1987; Korfmacher et al. 1987; Ober et al. 1987; Santa Maria et al. 1986). Preparation methods vary for the different types of foods. The sample preparation method for butterfat involves GPC. GPC is a rapid clean-up technique for separating pesticide residues from a lipid extract. It was developed into an automated clean-up apparatus for use on a wide variety of fats and oils. The automated GPC system is reproducible and reliable. After being cleaned on GPC, most samples can be analyzed by GC without additional clean-up (Hopper and Griffitt 1987). Recovery is complete (100%), and precision is very good (<3%). Sensitivity is in the sub-ppm range. The sample preparation for milk samples involves selective extraction on solid-matrix disposable columns by means of acetonitrile-saturated light petroleum, followed by Florisil column cleanup. Recovery is excellent (99%); precision is very good (<7%) (Di Muccio et al. 1988). Sample preparation for fruits, vegetables, and animal feed involves cyclic steam distillation extraction in hexane or isooctane with direct injection into the gas chromatograph. Recoveries for this method are very low (15–50%). This is an indication that heptachlor is not extracted quantitatively by steam distillation and is not a recommended preparation method (Ober et al. 1987; Santa Maria et al. 1986).

7.3. ADEQUACY OF THE DATABASE

Section 104(i)(5) of CERCLA, as amended, directs the Administrator of ATSDR (in consultation with the Administrator of EPA and agencies and programs of the Public Health Service) to assess whether adequate information on the health effects of heptachlor and heptachlor epoxide is available. Where adequate information is not available, ATSDR, in conjunction with NTP, is required to assure the initiation of a program of research designed to determine the health effects (and techniques for developing methods to determine such health effects) of heptachlor and heptachlor epoxide.

The following categories of possible data needs have been identified by a joint team of scientists from ATSDR, NTP, and EPA. They are defined as substance-specific informational needs that if met would reduce the uncertainties of human health assessment. This definition should not be interpreted to mean that all data needs discussed in this section must be filled. In the future, the identified data needs will be evaluated and prioritized, and a substance-specific research agenda will be proposed.