The various solutes travel through the column at different rates. The fastest moving solute exits (elutes) the column first then is followed by the remaining solutes in corresponding order. After the compounds have been eluted from the end of the column, they must be detected, one at a time. As each solute elutes from the column, it enters the heated detector. An electronic signal is generated upon interaction of the solute with the detector. The size of the signal is recorded by a data system and is plotted against elapsed time to produce a chromatogram.
The plot created by this process is called a chromatogram. The size of the signal is proportional to the amount of a compound in the sample. In conjunction with the data recorder, an integrator is frequently used to provide measurements of the height of the compound’s signal and the time that it took for the compound to elute through the column and reach the detector. While many types of detectors exist, the most common detector used for the blood alcohol analysis is the flame ionization detector. The flame ionization detector simply burns each organic compound as it elutes from the column. As the compound burns, it produces an electrical signal. The electrical signal is displayed as a “peak” with a known retention time. The period of time between injection and reaching the detector is referred to as the retention time.
The retention time of a compound alone is insufficient to identity of compound. The retention time of a known compound must be compared to the retention time of the unknown compound to determine if the known compound also exists in the testing sample. By comparing the retention time of the compound to a known standard, the compound can be identified. Identifying the compound is the qualitative portion of the analysis.
The concentration of alcohol in a sample is measured by comparing the size of the peak produced by the testing sample to the size of the peak produced by different known concentrations of alcohol. Measurement of the amount of the compound in the sample is the quantitative portion of the analysis. Each compound has a different retention time with a particular mobile phase, stationary phase, and temperature. If testing conditions change, identifying the compound is not possible. Therefore, the instrument must be periodically tested with a known specimen to ensure the testing conditions have not changed and the calibration of instrument remains intact.
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