Pharmaceutical Analytical Chemistry: Open Access
Open Access

Crucial Role of Mobile Phase Composition in Chromatography

Per Andren^{* *}Correspondence: Per Andren, Department of Pharmaceutical Analysis, Spatial Mass Spectrometry, Science for Life Laboratory, Uppsala University, Uppsala, Sweden, Email:

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Description

Chromatography is a powerful analytical technique widely employed in various industries, such as pharmaceuticals, environmental monitoring, food analysis, and more. Its effectiveness in separating and analyzing complex mixtures relies heavily on the careful selection of parameters, with mobile phase composition standing out as a critical factor. The mobile phase, a solvent mixture, plays a fundamental role in determining the separation efficiency and selectivity of chromatographic methods.

Understanding mobile phase composition

The mobile phase in chromatography is responsible for carrying the sample through the stationary phase, where separation occurs based on differences in the interactions between the sample components and the stationary phase. It consists of a solvent or a mixture of solvents that elute the sample components at different rates, resulting in their separation. The composition of the mobile phase significantly influences the chromatographic separation by affecting the retention times, resolution, and overall separation quality [1].

Role of solvent polarity

One of the key factors in determining the mobile phase composition is solvent polarity. In reversed-phase chromatography, a common technique where the stationary phase is nonpolar, the mobile phase typically consists of a mixture of polar and nonpolar solvents. The proportion of these solvents affects the interactions between the sample components and the stationary phase. A higher proportion of the polar solvent promotes stronger interactions with polar sample components, resulting in longer retention times. Conversely, a higher proportion of the nonpolar solvent favors weaker interactions, leading to shorter retention times. Balancing these interactions is essential for achieving optimal separation and resolution [2].

Influence on selectivity

Mobile phase composition also plays a crucial role in the selectivity of chromatographic separations. Selectivity refers to the ability of a chromatographic method to distinguish between closely related compounds in a mixture. By adjusting the mobile phase composition, chromatographers can fine-tune the selectivity to separate compounds that have similar properties. This is particularly important in complex samples where compounds with subtle structural differences need to be resolved effectively [3].

Optimizing resolution

Resolution, another essential parameter in chromatography, relies on the ability to separate adjacent peaks in a chromatogram. The mobile phase composition can significantly impact resolution. An appropriate composition can lead to wellseparated peaks, enhancing the accuracy of quantification and identification of sample components. Achieving high resolution often involves a delicate balance between solvent polarity, flow rate, and stationary phase properties [4].

Influence of pH and ionic strength

In certain chromatographic methods, such as ion exchange chromatography, the mobile phase composition includes buffers to control the pH and ionic strength. pH influences the ionization state of analytes, impacting their interactions with the stationary phase. Ionic strength affects the competition between the analytes and the ions present in the mobile phase. Adjusting pH and ionic strength allows chromatographers to optimize separation conditions for charged analytes, ensuring precise and reproducible results [5].

Mobile phase gradient elution

Gradient elution is a technique where the mobile phase composition changes over time during the chromatographic run. This method is particularly useful for separating complex mixtures, as it can improve resolution and decrease analysis time. By altering the composition of the mobile phase, chromatographers can achieve different separation mechanisms at various points in the analysis, leading to improved peak separation and reduced co-elution.

Conclusion

The composition of the mobile phase is an intricate and essential aspect of chromatographic analysis. It influences the interactions between sample components and the stationary phase, ultimately determining separation efficiency, selectivity, and resolution. Chromatographers must carefully select and optimize the mobile phase composition to achieve accurate and reliable results in their analyses. Through a thorough understanding of the principles behind mobile phase composition, scientists can harness the full potential of chromatography for a wide range of applications, from pharmaceutical development to environmental monitoring, ensuring the accurate analysis of complex mixtures.

References

1. Tsao R, Yang R. Optimization of a new mobile phase to know the complex and real polyphenolic composition: Towards a total phenolic index using high-performance liquid chromatography. J Chromatogr A. 2003;1018(1):29-40.

   [Crossref] [Google Scholar] [PubMed]

2. Valkó K, Snyder LR, Glajch JL. Retention in reversed-phase liquid chromatography as a function of mobile-phase composition. J Chromatogr A. 1993;656(1-2):501-20.

   [Crossref] [Google Scholar]

3. Heinisch S, Rocca JL. Effect of mobile phase composition, pH and buffer type on the retention of ionizable compounds in reversed-phase liquid chromatography: Application to method development. J Chromatogr A. 2004;1048(2):183-93.

   [Crossref] [Google Scholar] [PubMed]

4. Hao Z, Xiao B, Weng N. Impact of column temperature and mobile phase components on selectivity of hydrophilic interaction chromatography (HILIC). J Sep Sci. 2008;31(9):1449-1464.

   [Crossref] [Google Scholar] [PubMed]

5. Tanaka N, Goodell H, Karger BL. The role of organic modifiers on polar group selectivity in reversed-phase liquid chromatography. J Chromatogr A. 1978;158:233-48.

   [Crossref] [Google Scholar]