Biological Detergents
Biological Detergents - the basics
Anatrace - commercial site with booklet, tables, etc.
Useful new detergent
Essential concepts
1- Amphiphile or surfactant - a molecule with distinct hydrophilic and hydrophobic surfaces. Any substance which causes a solution to foam is likely an amphiphile.
2- Detergent - an amphiphile which disrupts membranes
3- Micelle - a detergent aggregate
4- Critical micelle concentration (cmc) - concentration above which detergent forms micelles. Above the cmc detergent monomer concentration is independent of total detergent concentration (at least for "discrete" amphiphiles). For ionic detergents cmc depends on counterion identity and salt concentration.
5- Critical micelle temperature (Krafft point) - temperature below which micelles are insoluble. The Krafft point of potassium dodecyl sulfate is notorious for being above room temperature.
6- Micelle structure - is size and concentration independent for detergents with distinct hydrophilic and hydrophobic domains (e.g. SDS). For detergents (e.g. bile salt) with less defined hydrophilic and hydrophobic domains, micelle structure varies with and size increases with detergent concentration.
7- Aggregation number - number of detergent monomers in a micelle. Detergents which form large micelles (e.g. Triton X-100) are hard to remove from proteins. Detergents which form small micelles (e.g. deoxycholate) are easier to get rid of.
8- Cloud point - temperature above nonionic detergents separate into two phases
9- Head group types - anionic, cationic, nonionic, zwitterionic
10- HLB number (hydrophile-lyophile balance) - an empirical measure of emulsifying power (devised by cosmetic chemists). The HLB number is the sum of plus numbers assigned for hydrophilic groups and minus numbers assigned for hydrophobic groups. Detergents with HLBs below 10 are water insoluble. Membranes are commonly solubilized with detergents of HLBs about 15. McCutcheon's contains a table of HLB values. SDS, with an HLB number of 40, is the best solubilizing agent among common detergents.
11- Detergent binding - Detergents bind strongly to lipids (and proteins). Unless detergent is in considerable excess, ratio of detergent to lipid (or protein) is more important than absolute detergent concentration. If, for example residual phospholipid is essential to enzyme activity, then activity will be especially sensitive to detergent protein ratio. Most proteins do not bind nonionic detergents.
12- Dialysis of detergents - Micelles dialyze slowly. To dialyze choose a detergent with a high cmc. Incomplete dialysis invalidated early studies of SDS binding to proteins.
13- Detergent "matching" - detergents with similar structures and similar HLB numbers are likely interchangeable.
14- UV absorbance - Where UV absorbance can interfere, choose nonionic detergents with alkyl groups (e.g Triton DF series); avoid those with alkylphenoxy groups (e.g. Triton X series).
Important Detergent Properties
1- Chemical nature and size of hydrophobic tail
2- Chemical nature and size of head group
3- Cmc and aggregation number.
4- Effect of solution variables (e.g. salt concentration, temperature, etc.) on cmc and aggregation number
5- Solubilizing power - The HLB number is a useful guide
6- Polyionic detergent micelles can precipitate oppositely charged polyions (e.g. CTAB precipitation of nucleic acids)
7- Detergent head groups dictate surface interactions. Headgroups of cationic detergents bind to anionic glass surfaces leaving the surface "greasy". For similar reasons nonionic detergents tend to make plastics "greasy".
8- Binding - Don't forget the influence of protein and lipid binding on free detergent concentration. Proteins bind twice their weight of SDS and lipids can "consume" 10x their weight of SDS.
9- Purity - Most commercial detergents are mixtures. The detergent can be a minor component. CAUTION - 99% pure usually means 99% detergent which can still be an ill-defined mixture. Polyoxyethylene detergents often contain iron and peroxides which can inactivate enzymes. Two batches of a single commercial detergent can differ more than two different, but similar detergents.
Detergent Bibliography
Furth, Bolton, Potter, Priddle (1984) Separating detergent from proteins. Methods in Enzymology 104, 318-328. (QP601.C133)
Helenius, McCaslin, Fries, Tanford (1979) Properties of detergents. Methods in Enzymology 56, 734-749.
(QP601.C133)
Helenius, Simons (1975) Solubilization of membranes by detergents. BBA 415, 29-79.
Hildebrand* (1979) Is there a "hydrophobic effect"?. PNAS 76, 194.
Hjelmeland (1986) The design and synthesis of detergents for membrane biochemistry. Methods in Enzymology 124, 134-164. (QP601.C133)
Hjelmeland, Chrambach (1984) Solubilization of functional membrane proteins. Methods in Enzymology 104, 305-318. (QP601.C133)
Lichtenberg, Robson, Dennis (1983) Solubilization of phospholipids by detergents: structural and kinetic aspects. BBA 737, 285-304.
Lindman, Wennerstrom (1980) Micelles, amphiphile aggregation in aqueous solution. Topics in Current Chemistry 87 ,1-87 (Chem - QD1.F58).
McCutcheon's Detergents and Emulsifiers - annual (TP992.5M24) (A guide to commercial detergents - published annually)
Myers, Drew (1992) Surfactant Science and Technology, 2nd ed. VCH (Chem - QD506.M94)
Rosen, Milton J. (1989) Surfactants and Interfacial Phenomena, 2nd ed. Wiley, a little pricey, I got a used copy for $15 (Chem TP994.S77)
Selinger, Ben (1998) Chemistry in the Marketplace, 5th ed. - a great book with a very useful chapter on detergents
Shinoda, Friberg (1986) Emulsions and Solubilization. Wiley (Chem - TP156.E6S55)
Tanford** (1980) The Hydrophobic Effect 2nd ed., Wiley (QP521.T73)
Weber, Osborn (1975) Proteins and sodium dodecylsulfate: molecular weight determinations on polyacrylamide gels and related procedures. The Proteins, I, 179-223 (QD431.N453)
* - Hildebrand is a former American Chemical Society president. He coached the US olympic ski team in 1936 and wrote this article when he was 98. He died in 1983 at 101.
**Tanford coined the term "hydrophobic effect" - to which Hildebrand objects, but which remains in common use.