Phosphate buffer and protein buffers

Phosphate buffer

The pKa for phosphate, H₂PO₄ , is 6.8, close to the desired blood pH of 7.4, so phosphate is a good buffer.

 In the ECF, phosphate is present as inorganic phosphate in a very low concentration  (about 1 – 2 mmol/L), so it plays a minor role in extracellular buffering.

Phosphate is an important intracellular buffer  for the reasons:

1.    Cells contain large amounts of phosphate in such organic compounds as adenosine triphosphate (ATP), adenosine diphosphate (ADP), and creatine phosphate. Although these compounds primarily function in energy metabolism, they also act as pH buffers.

2.    Intracellular pH is generally lower than the pH of ECF and is closer to the pKa of phosphate which is 6.8 (e.g.The cytosol of skeletal muscle has a pH of 6.9.).

Phosphoric acid is triprotic weak acid and has a pKa value for each of the three dissociations:

	pKa1 = 2		pKa2 = 6.8		pKa3 = 12
H3PO4	<= = = >	H+ + H2PO4-	<= = =>	H+ + HPO4-2	< = = = >	PO4-3 + H+

The three pKa values are sufficiently different so that at any one pH only the members of a single conjugate pair are present in significant concentrations.

The pKa2 is 6.8 and this makes the closed phosphate buffer system a good buffer intracellularly and in urine. The pH of glomerular ultrafiltrate is 7.4 and therefore phosphate is predominantly in the monohydrogen form so as to  combine with more H⁺ in the renal tubules efficiently. This makes the phosphate buffer more effective in buffering against a drop in pH than a rise in pH.

Protein buffer

Proteins act as efficient buffers because of their amino acid structure which  have a central carbon with four groups off of it.

The carboxyl and amino groups are what enable proteins to act as buffers.

Protein buffers in blood include haemoglobin (150g/l) and plasma proteins (70g/l). Buffering is by the imidazole group of the histidine residues which has a pKa of about 6.8. This is suitable for effective buffering at physiological pH (7.4).

Haemoglobin is quantitatively about 6 times more important than the plasma proteins as it is present in about twice the concentration and contains about three times the number of histidine residues per molecule. For example if blood pH changed from 7.5 to 6.5, haemoglobin would buffer 27.5 mmol/l of H⁺ and total plasma protein buffering would account for only 4.2 mmol/l of H⁺.

Deoxyhaemoglobin is a more effective buffer than oxyhaemoglobin and this change in buffer capacity contributes about 30% of the Haldane effect. The major factor accounting for the Haldane effect in CO₂ transport is the much greater ability of deoxyhaemoglobin to form carbamino compounds.

Haemoglobin has a special place in the pHstabilizing mechanisms of blood because

(a)  Haemoglobin and oxyhaemoglobin have different iso-electric points and different ionization constants so that at the pH of blood, absorption of about 0.7 g. of hydrogen ion [from carbonic acid mainly] (and the release of an equivalent amount of potassium ion in exchange) takes place during liberation of oxygen from 1 g molecule of oxyhemoglobin.

(b)  (b) Haemoglobin, more easily than other proteins, reacts reversibly with carbonic acid to form a still weaker carbaminoacid; and an increasing accumulation of carbamino-haemoglobin as the blood passes through the capillaries and a sharp reversal when, in the lungs, the equilibrium is disturbed, carbon dioxide being excreted, carbonic acid converted to carbon dioxide under the influence of carbonic anhydrase, and, consequently, carbaminohaemoglobin  is decomposed.

(Frazer &Stewart, J. clin. Path. (1959), 12, 195-206)

For more details readers are advised to go through the above mentioned paper and then directed to the link http://www.madsci.org/posts