Difference Between Reverse Phase and Hydrophobic Interaction Chromatography

The main difference between reverse phase and hydrophobic interaction chromatography is that the reverse phase chromatography (RPC) uses a more hydrophobic medium, which leads to more stronger interactions whereas the hydrophobic interaction chromatography (HIC) uses a less hydrophobic medium when compared to the ...

1. What is hydrophobic interaction chromatography?
2. What is the difference between normal phase and reverse phase chromatography?
3. What is reverse phase chromatography used for?
4. What is the difference between HPLC and RP HPLC?
5. What is meant by hydrophobic interactions?
6. What does the word hydrophobic mean?
7. Is C18 polar or nonpolar?
8. Is silica polar or nonpolar?
9. Why is reverse phase HPLC more common?
10. What are the advantages of reverse phase chromatography?
11. Which column is more polar C8 or C18?
12. Is the mobile phase polar or nonpolar?

What is hydrophobic interaction chromatography?

Hydrophobic interaction chromatography (HIC) separates molecules based on their hydrophobicity. HIC is a useful separation technique for purifying proteins while maintaining biological activity due to the use of conditions and matrices that operate under less denaturing conditions.

What is the difference between normal phase and reverse phase chromatography?

In normal-phase chromatography, the stationary phase is polar and the mobile phase is nonpolar. In reversed phase we have just the opposite; the stationary phase is nonpolar and the mobile phase is polar. ... In normal-phase chromatography, the least polar compounds elute first and the most polar compounds elute last.

What is reverse phase chromatography used for?

Reversed-phase chromatography is the most common HPLC separation technique and is used for separating compounds that have hydrophobic moieties and do not have a dominant polar character (although polarity of a compound does not exclude the use of RP-HPLC).

What is the difference between HPLC and RP HPLC?

Reversed phase HPLC (RP-HPLC) has a non-polar stationary phase and an aqueous, moderately polar mobile phase. ... With such stationary phases, retention time is longer for molecules which are less polar, while polar molecules elute more readily (early in the analysis).

What is meant by hydrophobic interactions?

Hydrophobic interactions describe the relations between water and hydrophobes (low water-soluble molecules). Hydrophobes are nonpolar molecules and usually have a long chain of carbons that do not interact with water molecules. The mixing of fat and water is a good example of this particular interaction.

What does the word hydrophobic mean?

Meaning " water fearing". Hydrophobic compounds do not dissolve easily in water, and are usually non-polar. Oils and other long hydrocarbons are hydrophobic.

Is C18 polar or nonpolar?

A C18 column is an example of a "reverse phase" column. Reverse phase columns are often used with more polar solvents such as water, methanol or acetonitrile. The stationary phase is a nonpolar hydrocarbon, whereas the mobile phase is a polar liquid.

Is silica polar or nonpolar?

Silica gel is a polar adsorbent. This allows it to preferentially adsorb other polar materials. When it comes to polarity, materials interact more with like materials. This principle is particularly important to many laboratories, which use silica gel as the stationary phase for column chromatography separations.

Why is reverse phase HPLC more common?

Reverse phase columns have a hydrophobic stationary phase which works well for retention of most organic analytes. ... Reverse phase chromatography also has the advantage of being able to use pH selectivity to improve separations. There are also many more choices in stationary phases for reverse phase vs. normal phase.

What are the advantages of reverse phase chromatography?

Reversed-phase chromatography provides better solubility for polar analytes, uses nontoxic solvents, offers a method for removal of contaminants and mobile phase additives, and gives timely sample recovery with little solvent evaporation.

Which column is more polar C8 or C18?

C18 has 18 carbon atoms while C8 has only 8 carbon atoms. C18 has a longer carbon chain, but C8 has a shorter one. C18 has higher retention while C8 has shorter retention. C18 has higher hydrophobicity, but C8 has a lower hydrophobicity.
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Is the mobile phase polar or nonpolar?

Stationary phases are usually very polar, while mobile phases vary widely in polarity, but are less polar than the stationary phase. This is called normal phase (NP) chromatography. The exception is reverse phase (RP) chromatography, in which a polar mobile phase, and a less polar stationary phase are used.