Larger sample volumes are helpful in increasing the sensitivity in the headspace measurement. However, because the headspace equilibration is operated in a closed vial, it is impossible to achieve a truly full evaporation even if the temperature is high. Some of the analyte could be trapped in the condensate phase. Fig. 3 shows the effect of the sample size on the ethanol full evaporation from a fermentation liquor sample in a 20-mL sample vial. It can be seen that when the sample size is less than 50 μL, the GC signal response in the headspace ethanol measurement is linearly proportional to the amount of sample introduced, indicating that a near-complete mass transfer of ethanol from the liquid phase to vapor phase is consistently achieved. When the sample size is greater than 50 μL, less than the expected amount of ethanol was detected. The data show that the effect of the sample size is observable when a larger volume larger that 50 μL was used. Thus, under the conditions used in this experiment, 50 μL is the maximum sample size that can be used for an accurate determination of ethanol in the fermentation liquor. It was noticed that the maximum sample size (directly affecting the volume ratio of the headspace and liquid sample) permitted in FE headspace analysis varies [23], [24] and [25], and depending on both the analyte and the nature of the sample matrix. Therefore, it is necessary to conduct a sample size optimization study for a new FE headspace GC application (Fig. 3).