Biomedical Engineering Reference
In-Depth Information
So you can observe ion exchange and hydrophobic interactions in
GPC, and size-exclusion effects are not negligible if macroporous
ion exchange supports are used.
Substances destroying the water structure, the so-called chao-
tropic substances such as urea, guanidinium hydrochlorid, or some
organic solvents such as methanol, ethanol, or acetonitrile, sup-
press hydrophobic interactions. Ions also alter the water struc-
ture. The power of destruction of water structure is given by the
Hofmeister series:
increasing precipitation (“salting-out”) effect
NH
4
, Rb
+
, K
+
, Na
+
, Cs
+
>Li
+
>Mg
++
>Ca
++
>Ba
++
PO
4
>SO
4
>CH
3
COO
−
>Cl
−
>Br
−
>NO
3
>ClO
4
>
>F
3
CCOO
−
>I
−
> SCN
−
increasing chaotropic (“salting-in”) effect
←
→
Besides the application in chromatography, chaotropic substances
are used for protein extraction, e.g., of recombinant proteins from
inclusion bodies and for precipitation by salts or organic solvents
(cf. Protocols 3.7.2 and 3.7.3).
References
Deutscher MP (ed.) (1990) Guide to protein purification. Meth Enzymol
182
Gooding KM, Regnier FE (eds.) (1990) HPLC of biological macromolecules:
methods and applications. Chromatogr Sci Series, vol. 51. Marcel
Dekker, New York
Harris EL, Angal S (1990) Protein purification methods: a practical ap-
proach. Oxford Univ. Press, Oxford
Holtzhauer M (ed.) (1996) Methoden in der Proteinanalytik. Springer,
Berlin, p 7
Mikes O (1988) High-performance liquid chromatography of biopolymers
and biooligomers. Part A: principles, materials and techniques; Part B:
Separation of individual compound classes. J Chromat Library. Else-
vier, Amsterdam, vol. 41A and vol. 41B
Oliver RWA (ed.) (1989) HPLC of macromolecules: a practical approach.
IRL Press, Oxford
3.3 Gel Permeation Chromatography
(GPC; Gel Filtration, GF; Size-Exclusion
Chromatography, SEC)
Gel permeation chromatography is the mildest of all chromato-
graphic techniques. It allows to select the buffer conditions during
separation so that no alteration of the protein structure or function
occurs.
