Mobile Phase vs Stationary Phase: Key Concepts in Chromatography
To understand chromatography, you must first understand its two basic functions: the mobile and stationary phase. In this article, we will break down each phase into its components, the mediums and solvents used, and how they are used in the chromatography process.
What Is the Mobile Phase in Chromatography?
Simply put, the mobile phase is the phase in chromatography where the solution being tested or separated is injected before it is passed through the stationary phase.
The mobile phase is often called an eluent or solvent that is pumped through the column filled with the stationary phase at a specific flow rate. This can be done simply through gravity or by a pump to vary the flow rate through the column.
To work properly, the solvent used as the mobile phase must have a relatively low viscosity, low compressibility and be able to solubilize (or dissolve) the intended sample being tested or separated. The mobile phase must also have a favorable affinity for the stationary phase, meaning it allows compounds to separate effectively without sticking too much or passing through too quickly.
Solvents Used for Mobile Phase
In chromatography, several common solvents are used as the mobile phase, with some being more effective than others due to differences in how they interact with the stationary phase. Popular options include hydrocarbons like pentane, hexane, and petroleum ether and co-solvent mixtures like ether/hexane and methanol/dichloromethane.
The choice of solvent is critical and depends on the specific molecules being separated, as certain solvents work more efficiently with particular materials.
What Is the Stationary Phase In Chromatography?
The stationary phase is the material packed into a column through which the mobile phase will pass to perform separation. The materials used for the stationary phase are often:
- Gel
- Silica
- Alumina
- Sand
These materials allow the mobile phase, carrying the dissolved sample, to flow through them. The material can be monodisperse or polydisperse, meaning the particles are all the same size or vary in size, respectively. This variation can aid separation, as specific sample components may have a unique affinity for certain particle sizes.
Pore Diameter and Its Role in Separation
The pore diameter of the stationary phase plays a crucial role in how well separation can be performed during chromatography. The particles in the sample must be small enough to pass through the pores.
For example, cannabinoids like CBD and THC have very small particle sizes and need a small pore size for effective separation. In contrast, larger molecules, like plant proteins, cannot pass through smaller pores, allowing cannabinoids to be separated from them.
Selectivity of the Stationary Phase
Another key factor is the selectivity of the stationary phase material. This refers to how well the stationary phase interacts with the mobile phase on a molecular level.
The interaction between the media in the mobile and stationary phase in chromatography determines how well compounds are separated. For instance, some stationary phases may effectively separate compounds in one solvent but may not work as well with another solvent.
Polarity and Its Impact on Separation
Polarity also affects how well components separate during chromatography. Molecules can be polar (having a charge distribution, like water) or nonpolar (lacking a charge, like oil). Because cannabinoids are nonpolar, using a nonpolar solvent will be more efficient for their extraction and separation. While polar solvents can still dissolve nonpolar compounds, they are less effective than nonpolar solvents in these cases.
| Mobile Phase vs Stationary Phase | Mobile Phase | Stationary Phase |
|---|---|---|
| Composition | Liquid or gas | Solid material |
| Material | Solvent (pentane, hexane, petroleum ether, etc.) | Porous Solid (silica, alumina, gel) |
| Purpose | Dissolve and carry the mixture through the stationary phase | Separate the mixture into various components as they pass through it |
| Post-processing | Solvent removal needed | Remains in column |
How the Mobile and Stationary Phase Work in Chromatography
Both the mobile and stationary phases are essential in chromatography, and chromatography separation cannot be accomplished without them.
Let’s break down the chromatography process in simple steps:
- A column is prepared with a filter material, followed by a packing material, and then the stationary phase is added.
- The mixture intended to be separated is suspended in a small amount of solvent and added above the stationary phase.
- More packing material is added above the mixture solvent solution, followed by the remaining solvent, which is used as the mobile phase.
- Using simple gravity or some kind of vacuum pump, the mobile phase is passed through the mixture, pulling it through the stationary phase.
- As each component is separated by passing through the stationary phase, they are collected in separate vessels. The chromatography process is then complete.
Now that we’ve established the basic chromatography steps, let’s get more detailed:
The mobile phase is a solvent or fluid eluent that can move freely through a chromatography system using a pump. As a solvent, it is also important for the mobile phase to dissolve the solution being tested or separated so that the individual components can be identified and remediated. This is why some eluents or solvents work more efficiently than others, depending on the separated substance.
The material used as the stationary phase is always some sort of porous solid that remains fixed in a column that allows the eluent used as the mobile phase to pass through it. This is where the actual chromatography process takes place. As the mobile phase passes through the stationary phase, the various components of the solution will move through the material at various rates depending on their affinity for that material.
If a particular material passes through the stationary phase more quickly, it will be seen on a chromatograph or collected in a containment vessel. Then, the following fastest material will be seen or collected, and the process continues until each component is identified and separated. As they pass through the stationary phase, they can be identified in a chromatogram for testing purposes or collected into separate vessels.
Several factors can cause variations in the efficiency of a chromatography separation process, but the basic principles remain the same. The mobile and stationary phase in chromatography can be as simple as water and a coffee filter or as complicated as a co-solvent solution and a silica gel, but the separation process itself remains the same.
By adding a solution to a mobile phase and passing it through a stationary phase, you can effectively separate the individual components in a solution so that they can be identified, remediated, and isolated altogether.
Final Thoughts
Understanding the mobile and stationary phases of chromatography is crucial for efficient separations. These two components work together to ensure effective testing and purification, making them essential in the extraction industry.
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