Spurious chromatographic peaks in chromatographic analysis

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Reasons for the formation of spurious peaks: Usually, there may be two types of interactions between the sample (X) and the mobile phase (M) – cooperative adsorption and competitive adsorption. Assume that the sample molecule X is undetectable (no UV absorption), while the mobile phase component M is detectable. M is often an impurity in the mobile phase, while X may be an impurity or component in the solvent used to dissolve the sample, or it may be a genuine component of the sample. Under cooperative adsorption, if tM is less than tX, the components of the mobile phase leave the column first; M leaves the column as an inverted peak, followed by X as a normal peak ； If the retention times of M and X are reversed, that is, tM is greater than tX, then X exits the column first as an inverted peak, followed by M, which emerges as a normal peak. In ion-pair chromatography, it operates on the principle of cooperative adsorption. In reverse or normal phase chromatography, it is competitive rather than synergistic, resulting in different forms of spurious peaks: the earlier appearing spurious peak is a positive peak, while the later appearing one is an inverted peak. Several approaches to eliminating spurious peaks: (1) Using pure reagents as the mobile phase. The mobile phase is prepared using high-quality ion-pair reagents and buffering agents, with the combination of these reagents resulting in minimal spurious peak effects. (2) Dissolve the sample in the mobile phase. Dissolving the sample in other solvents may produce spurious peaks or lead to their formation, while dissolving it in the mobile phase can reduce the likelihood of such spurious peaks. (3) Add the sample solution in the smallest volume possible. Spurious peaks are often proportional to the sample volume, and the injection volume for ion-pair chromatography should be below 50 microliters. (4) Pre-treat the sample. Impurities in the sample can cause the appearance of spurious peaks. If the spurious peaks still cannot be removed using the above methods, the chromatographic conditions can be changed to shift the position of these peaks, thereby avoiding the interference with the desired peaks.