pH and pOH

By definition, pH is the negative logarithm of the hydrogen ion concentration. Similarly, pOH is the negative logarithm of the hydroxyl ion concentration.

In an aqueous solution, the equilibrium for the ionization of water is

Taking the logarithms:

Sample problem:

What are the (a) H+ ion concentration, (b) pH, (c) OH- ion concentration, and (d) pOH of a 0.001 M solution of HCl?

Remember a strong acid dissociates 100% when diluted in an aqueous solution.

Ionization of weak acids

In an aqueous solution a weak acid ionizes to a limited extent:

The proton released from HA is accepted by water to form the hydronium ion H3O⁺. The reversible ionization reaction can be described by an equilibrium constant, Ki:

Because [H2O] is itself a constant, we can define a new constant, Ka, that combines Ki and [H2O].

Because [H3O⁺] is the same as the hydrogen ion concentration, [H⁺], the Ka expression is usually written as shown below:

When a weak acid, HA, is dissolved in water the following happens:

Where Ka is the equilibrium constant for the dissociation of the acid.

If we start with the conjugate base, A-, and dissolve it in water, it ionizes as a typical base; it accepts a proton from H2O to form OH- and the corresponding conjugate acid, HA. A Kb expression can be written for this ionization.

Solution Behavior of Proteins

Changes in protein solubility that do not destroy the molecule’s inner structural integrity can occur in several ways.

Isoelectric Precipitation. Proteins typically have charged amino acid side chains on their surfaces that make energetically favorable polar interaction with the surrounding water. The total charge on the protein is just the sum of the charges contributed by the individual side chains. However, the actual charge on the weakly acidic and basic groups characterizing ionic amino acid side chains also depends on the solution pH.

For each protein there will be a particular pH value where the sum of the positive side-chain charges exactly equals the sum of the negative charges, so that the net charge on the protein is zero. Since an uncharged molecules will not migrate in an imposed electric field, the pH value at which this occurs is termed the protein’s isoelectric point (pI). Proteins tend to be least soluble at their isoelectric pH. This decrease in solubility reflects the fact that the individual protein molecules, which would all have similar charges at pHs away from their isoelectric points, cease to electrostatically repel each other at their isoelectric points.

Salting In and Salting Out. Proteins also show a variation in solubility depending on their solution ionic environments. These frequently complex effects may involve either specific interactions between charged side chains and solution ions or, particularly at high salt concentration, reflect more comprehensive changes in the solvent properties. The effects of salts such as sodium chloride on increasing the solubility of proteins is often referred to as salting in. The salting in effect is related to the nonspecific effect the salt has on increasing the ionic strength. Increasing the ionic strength decrease the sphere of influence of each charged site on the protein.

Many globular proteins precipitate out at very low ionic strengths or in pure water. This happens because oppositely charged sites on different proteins are able to interact favorably leading to an electrostatic complex. When this complex formation is extended between many protein molecules, it can lead to protein precipitation.

Some salts, such as high concentrations of ammonium sulfate, have general effects on solvent structure that lead to decreased protein solubility and salting out. In this case, the protein molecules tend to associate with each other because protein-protein interactions become energetically more favorable than protein-solvent interaction. Proteins have characteristic salting out points, and these are used in protein separations in crude extracts. The ammonium sulfate is essentially having a dehydrating effect on the protein.

Protein separation based on protein charge

The surfaces of most globular proteins are populated by charged amino acid side chains. Although each ionizable side chain of a free amino acid has a well-defined pK, the specific environment of amino acid side chains on a protein’s surface causes slight alterations in their pK values. For example, a neutral histidine side chain that is partially buried at the protein’s surface may have a decreased tendency to accept an additional proton, since the formation of the resulting positively charged species is less favorable in a partially apolar environment than in pure water. As a result, the pK of the histidine can be lowered relative to its usual value in water. Similarly, interaction between ionic groups at the protein’s surface can influence individual pK values.

Owing to the sorts of environmental effect that perturb the pKs of protein amino acid side chains, even proteins that might on the basis of composition be expected to have identical formal charges are in fact slightly different in their charge properties.