Detergentes

Detergents - also termed as tensides or surfactants - are amphipathic compounds with both lipophilic (hydrophobic; non-polar) and hydrophilic (polar) sides within one molecule.

Therefore they are soluble in aqueous solution as well as in non-polar organic solvents - and they are able to influence the solubility of other molecules (like lipids or hydrophobic proteins in buffer solutions). Detergents are widely used in biochemistry, cell biology or molecular biology. Cell lysis, protein solubilization, protein crystallization or reduction of background staining in blotting experiments are just a few examples.

Detergents can be classified for instance according to their chemical structure stating their constituent polar and nonpolar group (glucosides, alkyl ionic detergents, polyoxyethylene alcohols, bile salts, sulphonates etc.), the charge character (anionic, cationic, zwitterionic = amphoteric; non-ionic) or simply whether they are mild or strong in terms of their ability to solubilize and / or to denature proteins. We decided to arrange our detergents according to their charge:

• anionic detergents
• cationic detergents
• zwitterionic detergents
• non-ionic detergents
• isomeric pure detergents for crystallography

But how to choose the right detergent?

There is a large number of different detergents available and choosing the best detergent for a special application is not trivial. In many cases, a set of detergents has to be tested to select the one with the best properties. If the goal is the isolation of a protein preserving its structural and functional state, one should consider the temperature, the pH and the ionic strength of the system as well as interference with assays. Several other parameters may influence the choice too. Assuming that the factors mentioned above are dictated by the protein and are held constant, the factors that are left to be optimised are the detergent's strength (in this case a mild, non-denaturating one), chemical structure, solubility and concentration.

The CMC value is specific to each detergent

Above a critical concentration, called CMC value (Critical Micellar Concentration), detergents start forming stable aggregates (micelles). The CMC value is specific to each detergent and different factors (chemical structure, pH value, temperature, ionic strenth) may influence it. In aqueous solutions, the hydrophilic site of the monomers will be on the outside of the micelle, and the lipohilic portion inside. In organic solvents, reversed micelles form, with the lipophilic site outside. The micelle size increases and the CMC decreases with increasing size of the lipophilic part of the detergent and, to a lesser extent, with decreasing size and polarity of the polar groups. That means, that detergents with a higher hydrophilic part are characterized by a higher CMC (start to form micelles at higher concentrations) and smaller micelles than higly lipophilic detergent. Above the CMC, free monomer molecules are in equilibrium with the micelles and the solubilizing ability increases. Detergents with high CMC values are more easily removed by dialysis.

Overview Detergents

Trademarks:
Brij (Atlas Chemicals Co.)
Nonidet (Shell)
Triton (Union Varbide Co.)
Pluronic, Tween (ICI America Inc.)

General properties of the main classes of detergents

Ionic detergents contain a negatively (anionic detergent) or positively (cationic detergent) charged hydrophilic head group. The hydrophobic part is either an alkyl chain (as for SDS, CTAB or alkyl sulfonic acids) or a more complicated steroidal structure as a bile acid salt (like cholate and deoxycholate). The latters are more rigid in their structure and carry additional hydrophilic groups (hydroxyl groups) within the hydrophobic steroide scaffold. Thus, bile salts only show a polar and an apolar face, not a well-defined polar head group.
The size of the micelles is affected by repulsion forces between the charged polar groups and hydrophobic attraction between the side chains. As a result, the micellar size of ionic detergents can be manipulated by the ion strength. Increasing the concentration of neutralizing counter ions leads to a larger micelle size. At the same time, the increased ion strength results in a lower CMC. Changing the temperature on the other hand only slightly influences the CMC. Ionic detergents generally have higher CMC values than their non-ionic analoga.
Alkyl-ionic detergents mostly act denaturating and separate protein complexes into their subunits.

Non-ionic detergents possess an uncharged, hydrophilic head group consisting of either polyoxyethylene units (e.g. Brij^® und Triton^®) or sugars (alkyl glucosides or maltosides). The CMC value and micellar size of this detergent group is mainly affected by temperature (the higher the temperature the higher the CMC), not by ion strength.
Non-ionic detergents generally are non-denaturating and therefore first choice for applications that require preservation of protein structure and activity. They are mild detergents that primarily break lipid-lipid and lipid-protein interactions, while protein-protein interactions stay unaffected. Especially alkyl glycosides and maltosides are suitable for isolation of biological active membrane proteins; advantages over polyoxyethylene detergents are e.g. homogenity in composition and structure (many polyoxyethylenes are composed of several homologs) und a lack of absorbance at 280 nm (detergents containing aromatic rings absorb in the ultraviolet region and may interfere with spectrophotometric monitoring of proteins at 280 nm).

Zwitterionic detergents, like CHAPS or sulfobetaine, combine the features of ionic and non-ionic detergents. Like non-ionic detergents they have no net charge. Consequently they show no electrophoretic mobility and do not bind to ion-exchange resins. Compared to ionic detergents, their CMC values are less sensitive to changes in ion concentration, but they have in common to break protein-protein interactions efficiently (denaturating effect).