Amino acids consist of carboxylic acids (acidic) and amines(basic). Acids contain a hydrogen atom which is a proton with one electron,technically an H ion is just a proton, and hence an acid is a proton donor. Aswell as, bases are proton acceptors. Moreover, there are two options foramines, they can either be neutral (COOH) or lose a proton (COO-). The protoncount on each will be determined by the acidity of the solution and that aminoacids can only be neutral or zwitterion (overall neutral but have formalcharges across the molecule).
http://study.com/academy/lesson/zwitterion-structure-function.htmlA zwitterion is sometimes called dipolar ions. This is dueto having a negative end (anion) and a positive end (cation).
At a neutral PH,amino acids exist in their zwitterion form (dipolar form). This means that theamino acids contain a positive charge and a negative charge. a) At the low PH of0/1 the carboxylic group is protonated. At this acidic PH, amino acids arepositively charged species. b) As the PHincreased, the carboxylic acid group loses its hydrogen to form a zwitterion.
Ion exchange chromatography: c) The zwitterion formusually persists until around a PH of 9. At this basic PH, the low H+ causesthe amino acids to lose its hydrogen therefore forming a negatively chargedspecies. Do you know how to write the structure of an amino acid?Can you draw and explain the structure of a zwitter ion andhow is it affected by acidic and basic pHCan you explain two techniques which separate amino acids onbasis of their ionisation at different pH?Ion exchange chromatography: Thismethod can be used to purify mixture of proteins based on their net charge. Soion exchange chromatography separates charged molecules and polar molecules.Hydrophobic molecules cannot be separated by this method. There are twodifferent phases known as the stationary phase and the mobile phase.
The interactionbetween these two phases varies for different molecules. One again ion exchangechromatography separates ions or polar or charged molecules. In this case ofion exchange chromatography, we run a column, meaning it’s a structure madewith polymers which creates chamber. Inside that chamber we add the stationaryphase. An amino acid has different chargedproperties based on the PH of the solution it is in. PH dictates whether theamine and carboxyl group in an amino acid will be protonated or deprotonated. Overall,charge differences are affected by PH.
This is because increasing PH in generaltends to deprotonate functional groups and makes the net charge on an aminoacid more negative. However, if PH is decreased then it protonates functional groupsand make the net charge more positive.Ion exchange chromatography canseparate amino acids on the basis of their net charge. You have column filledwith resin beads that are either negatively charged or positively charged. As aresult, amino acids will stick to the column with certain timings based ontheir charges. On the diagram above, chargedmolecules attached to the beads hold onto the amino acids to be separated.
Theion exchange chromatography can be classified into two types which are theanion exchange chromatography and the cation exchange chromatography. Thismethod is based on the reversible electrostatic interactions of proteins withthe separation matrix. Mechanism of protein separation has two types which arePH based binding and salt based binding. The differences between the netsurface charges on the solute molecules, is what the PH based binding dependson. As you can see above, the value whichis the PH underneath pi, the protein will gain itself a net positive charge.
Itwill also bind reversibly to the width of the surface of a cation exchangeresin which is negatively charged group based on the PH. Note that At PH value below the pi, the proteinwill have a net positive charge and will tend to bind reversibly to the widthof the surface of a cation exchange resin that is one that has negativelycharged groups at the PH. (1)http://technologyinscience.blogspot.co.uk/2011/09/ion-exchange-chromatography-principle.html#.Wl-qIq5l_IU(1)