Retardation or retention factor (Rf) value is the ratio of distance traveled by the analyte to that of the solvent front on a chromatogram.
The chromatographic techniques in which the analytes are added to the stationary phases show a difference in the movement of analytes with mobile solvents (phases). This difference is due to the relative affinities of analytes with stationary and mobile solvents. The more the relative affinity with a stationary phase an analyte has, the more it will stay in place and the lesser the Rf value will get and vice versa.
|Chromatography and its types|
|Thin Layer Chromatography (TLC)|
Rf value is the characteristic identification value for analytes at given temperatures. It means that compounds can be analyzed and identified based on their Rf values. It is, however, not the case when a new compound is discovered.
Calculation of Rf values
A chromatogram is first to be developed with a suitable solvent (mobile phase), depending upon the nature of analytes, and the stationary phases. The developed chromatogram is then dried and the positions (migration values) are measured for analytes and the solvent front.
The Rf (retardation/retention factor) values can be calculated by using the given procedure using the above experiment.
A prepared sample solution (A+B) is applied on the chromatogram paper and run through a mobile phase. Analyte (A) and (B) separate out because of different affinities with mobile phase (solvent). The relative measurements are taken for the analytes, the solvent front, and the point where the mixture (A+B) was applied.
For the analyte (A)
Rf = Distance moved by analyte (A) / Distance moved by solvent front
Rf = 2.9 / 4.0
Rf = 0.725
For the analyte (B)
Rf = Distance moved by analyte (B) / Distance moved by solvent front
Rf = 1.3 / 4.0
Rf = 0.325
So, the Rf values for the analytes (A) and (B) are 0.725 and 0.325.
Factors affecting Rf values
There are several factors that affect the Rf values of a particular analyte:
- Stationary phase
- Concentration of stationary phase
- Mobile phase
- Concentration of mobile phase
Same substances (analytes ) have different Rf values if the nature of stationary and mobile phases is changed, i.e. the affinity factors are changed. Similar is the case when the concentration of these phases is changed. Temperature affects the rate of mobile action of solvent and also the solubilities of analytes in the solvent.
Rf values are independent of the concentration of analytes whatsoever.
How to calculate Rf?
Retention/retardation factor (Rf) can be calculated by the relative migration values of solute (analyte) and the solvent front.
Rf = Migration of analyte / Migration of solvent front
The calculation of the Rf value is basically the calculation of relative affinities of a solute with the stationary and mobile phases.
How to choose the solvent system?
The solvent system for a particular analyte separation via chromatography is chosen based on the polarity of solute (analyte), the stationary phases, and the temperature. The solvent system is actually the mobile phase which must relatively be either more or less polar than the stationary phase, which becomes the reason for the displacement of the analyte from the application point to a higher level in column chromatography.
A pure solvent can be used with particular polarity for a chromatographic procedure. Although if a series of continuously increasing or decreasing polarities is required, a mixture of two or more solvents may be used with a continuous manner of increase in the concentration of one solvent and decrease of other.
How to interpret the TLC?
Interpretation of the thin layer chromatography (TLC) is totally based on the Rf values. The Rf values evaluate the relative polarity, the relative affinities with stationary and mobile phases, the relative molecular weights, and even the concentration of analytes if sophisticated environments are used.
Why do we need the Rf value?
Rf values in chromatography are the basic requirement of the whole experiment. These values tell us whether the analyte (solute) is more affinitive with stationary or the mobile phase. Rf values evaluate the polarity, relative masses, and relative solubilities with stationary and mobile phases, etc.
How to find the Rf values of aspirin in specific solvents?
Rf values are found by chromatographic experiments. Aspirin is a polar molecule and will be more soluble in a polar solvent. As the general rule of solubility ‘like dissolves like’ suggests. The Rf values are always different for different stationary and mobile phases used. So, an aspirin solution is to be run against a polar solvent (mobile phase) when the stationary phase is less polar than the mobile one. This will leave aspirin at a specific height giving its Rf value between 0 and 1.
What are the units for Rf values?
Rf values are unitless as the ratio of similar quantities is always just a number. The solute migration value is compared by the solvent front migration value to calculate the Rf value which gives just a ratio number.
What are the advantages of calculating an Rf value in chromatography?
Chromatography is extremely useful only if there is an account for Rf values. These values give the necessary information required, for which the chromatography is basically performed. Rf values evaluate the polarities, the relative affinities with stationary and mobile phases, the relative molecular weights, and even the identification of analytes (solutes) are done by the Rf values unless there is no record for that particular solute in those conditions.
Which one is better in TLC, the higher or the lower Rf value?
The basic property of Rf values is to determine the relative properties of components of a mixture among each other and with the stationary and mobile phases. It doesn’t actually matter if their values are higher or lower as long as these are far apart from one another.
Rf values must not be either 0 or 1 because both of these conditions are not useful in the analytical procedure. An Rf value of zero means that the complete displacement (migration) of the analyte with the mobile phase (solvent), whereas the value one refers to no displacement at all.
Is chromatography paper polar?
In column chromatography, the chromatography paper is made of a cellulose network (like a normal paper) having capillary action to move solvent over it. The compound acting as the stationary phase is the adsorbed water on cellulose fibers which is a polar compound. So, the chromatographic paper in column chromatography is polar. This is also the reason why it is called liquid-liquid chromatography.
In thin layer chromatography, the TLC paper or sheet is having alumina (AL2O3) or silica (SiO2) pasted over glass or metal sheet, which are polar so a TLC paper is also polar.
What is the principle behind chromatography?
The main principle of chromatography is the relative solubilities of a single analyte in different solvents.
Retardation/retention factor (Rf) values drive the whole procedure because it is the Rf values that determine the relative polarities/solubilities of components in a mixture (solutes) based on the polarities of stationary and mobile solvents.
The principle of chromatography can thus be, the relative affinities of analytes (solutes) with stationary and mobile phases.
The principle of relative affinities is further described by the adsorption or dissolution of solute on stationary solvent or phase and then leaving the stationary solvent for a relatively higher solubility in mobile solvent/phase.
In chromatography, if solvent reaches the edge of the paper so how can we determine the Rf value?
In chromatographic techniques, the solvent front is usually not let to reach the end of the paper. The development of chromatogram is stopped before that happens and is placed to dry for further evaluation. But if somehow, the solvent front reaches the edge of paper during chromatography, the value of solvent front migration is taken to be the length of the paper.
What is a good Rf value?
Rf values are the retardation values in a specific environment for a particular analyte (solute). These values aren’t good or bad. These are just a ratio of two migration distances occurring simultaneously, the analyte and the solvent front. It would be better for the calculation and identification purposes if the Rf values of compounds of a mixture are not too close.
Although, if the Rf values of compounds in a mixture are too close, these would not be of much use.
Why is Rf less than 1?
Rf values are always less than one because they are the ratios of migration distances of solutes (analytes) and solvent fronts. As a general rule, the solvent front always travels more than that of solute, because the solutes have to have some attractive properties with stationary phases. It means that the denominator being higher in value, Rf values will always be between 0 and 1.
What is the significance Rf value?
Rf values are fairly significant because the outcome of most of the chromatographic procedures, especially TLC and column chromatography rest on these Rf values. It is Rf values that provide the relative properties of analytes like polarities, molecular weights, affinities with particular solvents, and even the identification of analytes (solutes) can only be achieved through the Rf values library.