Gas Chromatography

GC is the reference method for alcohol testing. Serum, plasma, whole blood, urine, or vitreous fluid may be used. This method is specific for ethanol and is also able to identify other monohydroxy alcohols at the same time (Fig.1). Sample may be introduced directly onto the column (direct injection method) or by headspace analysis where the airspace above the sample is allowed to equilibrate with ethanol in the sample, and the air sample is subsequently analyzed. The direct injection technique offers the advantage of more rapid analysis; however, fouling of the column and plugging of the inlet and syringe can occur. Ethanol and other alcohols are separated using chromatography, and each alcohol is identified by its retention time (Fig.1). Quantitation is accomplished by comparing the peak height ratio of the sample to an internal standard with the peak height ratio of the calibration standard to an internal

Fig. 1. Chromatogram of an alcohol standard containing 0.395% ethanol. N-propanol was used as the internal standard. The specimen injected in the column was 1.0 mL headspace sample of a blood alcohol mixture, with an isothermal oven temperature, 70°C equilibrium temp, and flow rate of 20 mL per minute. Retention time of each alcohol is on the horizontal axis.

Fig. 1. Chromatogram of an alcohol standard containing 0.395% ethanol. N-propanol was used as the internal standard. The specimen injected in the column was 1.0 mL headspace sample of a blood alcohol mixture, with an isothermal oven temperature, 70°C equilibrium temp, and flow rate of 20 mL per minute. Retention time of each alcohol is on the horizontal axis.

standard. The internal standard typically used is n-propanol (Fig.1). Another advantage of GC is the capability of this technique for simultaneous analysis of other volatiles such as diethylene glycol, ethylene glycol, methanol, acetone, and isopropyl alcohol along with ethanol. There are numerous methods reported in the literature. For example, Williams et al. (30) used a capillary column and GC methods for simultaneous quantitation of diethyl glycol, ethylene glycol, methanol, isopropanol, acetone along with ethanol. After removal of serum proteins from the specimen by ultrafiltration, 1 ml of ultrafiltrate was manually injected into the GC column. The authors used n-propanol as an internal standard for analysis of alcohols and acetone and 1,3-butanediol for analysis of glycols (30).

4.5. Gas Chromatography-Mass Spectrometry

GC/MS has also been utilized for determination of alcohol concentration in serum. Dean et al. (31) reported a method for simultaneous determination of ethanol and ethyl-d5 alcohol in serum using stable isotope GC/MS. Wasfi et al. (32) developed a sensitive and specific method using static headspace GC combined with MS for quantitative determination of ethanol in biological fluids using n-propanol as an internal standard. The GC was performed in an isothermal mode with a run time of only 2.6 min, and the quantification was performed using a mass spectrometer operated in scan mode abstracting a quantitative ion (m/z 31) and a qualifier ion (m/z 46) for ethanol and for the internal standard (m/z 31 and 60, respectively). The method was linear for a concentration range of 5-200 mg/dL, and no interference was observed from methanol, acetaldehyde, acetone, or endogenous materials (32). Maeda et al. (33) also used an automated headspace GC combined with mass spectrometry for analysis of postmortem ethanol concentrations in pericardial fluid and bone marrow aspirate.

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