When operating the gas chromatograph, if the column is normal, the sensitivity of the sample is sufficient, the analytical method is appropriate, and the peak shape should be symmetrical and sharp under the condition that the peak time is short. However, when the degree of understanding of the sample is insufficient, the method is not appropriate, the sample processing method and the injection method are unreasonable, some unexpected problems may occur, and it is difficult to make a reasonable explanation for the chromatographic peak. Below we, Tengzhou Lu Chuang chromatograph based on their own experience, put forward some views, to advise colleagues.
A chromatographic bimodal refers to a substance, but if a double peak appears in the chromatogram, it indicates that it contains two substances. We can divide this situation into four reasons.
1, the column
If you analyze the sample and find that each peak has a double peak, especially when using a single pure substance, you can be sure that the column is out of order - the stigma is damaged or the stigma stationary phase is dirty or lost. If the injection volume is small, the original chromatographic column is normal, and the shape of the chromatographic peak is mostly a large peak with a small peak, not necessarily tailing. This should generally be the plugging of the stigma, the column is reversed, and the mobile phase is washed or pickled. Or other solvents, wash away the residue stuck in the stigma, and vice versa, under normal circumstances. Of course, there is no recoil, and the positive impulse will sometimes be normal. If the peak tails, the peak intensity is not much difference, the stigma stationary phase becomes dirty or the possibility of loss is greater. This is to unscrew the injection head, ultrasonically filter the microporous filter, scrape off a part of the packing, and re Fill in the new packing and tighten, but this kind of work requires a certain skill, and at the same time can not do the same thing, otherwise it will not be used several times, the column will be scrapped due to low efficiency.
2, solvent polarity and injection volume
Many HPLC analysts may not agree with this, and the general HPLC books and literature will not mention this aspect, and this is indeed a very important reason for the double peak. At present, HPLC analysis is mostly reversed phase chromatography. The mobile phase is mostly methanol, acetonitrile, water, and various additives are added to improve the separation performance. The sample is typically dissolved in a solvent that is compatible with the mobile phase. The best way to dissolve is to dissolve with the mobile phase, but in many cases it is inconsistent. When using solvents with high polar polarity, such as pure methanol, pure acetonitrile, pure ethanol, and the analysis system is mainly water, the sample injection volume is large, such as 20ul of quantitative tube, under this condition, it is completely certain, single The pure material has a double peak, the second peak is smaller than the first peak, and the tailing time, the retention time will be advanced, the injection volume will be reduced by more than half, and the peak shape will become normal. This is because the solvent and the mobile phase polarity of the sample are too different, and the mobile phase is too late to dilute it to equilibrium. The above mentioned one reason for the double peak caused by the injection volume. Another reason is that the injection volume is not necessarily large, but the absolute amount is very large. The double peaks on the chromatogram are close together, basically high, no tailing. . Dilute the sample and re-inject it. This is due to excessive injection volume and column overload. ,
3, the characteristics of the sample
Some samples have tautomerism due to their chemical structure, and such tautomers cannot be separated, but exist in a dynamic equilibrium. In chromatographic analysis, under a specific condition, a substance will appear double peaks, the double peaks are close, basically high, no tailing, the conditions change slightly, especially pH, the double peak phenomenon will disappear, such as Erythromycin, etc. Some samples do not see double peaks on the UV chromatogram, but under LC-MS, the total ion chromatogram of the mass spectrum is more obvious with a mass spectrometer detector, such as the acetamiprid, which I analyzed.
4, parameters
The recorded parameters are generally defined and do not need to be modified, but the GC and HPLC parameters are not completely consistent. For example, the general recording time interval GC on the C-R3A data logger is 2 ms, HPLC is 5 ms, if the recording interval is shortened. , one peak will become two peaks or more.
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