Buffers in Biology PDF

## Buffers Buffers are solutions which can resist changes in pH when acid or alkali is added. ### Composition of a Buffer - Weak acid and its salt with strong base. This is the more important and common in human body e.g (CH3COOH/ CH3COONa) and H2CO3/ NaHCO3 - Or weak base and its salt with stron...

## Buffers Buffers are solutions which can resist changes in pH when acid or alkali is added. ### Composition of a Buffer - Weak acid and its salt with strong base. This is the more important and common in human body e.g (CH3COOH/ CH3COONa) and H2CO3/ NaHCO3 - Or weak base and its salt with strong acid e.g (NH3/NH4CI). - The pH of buffers are determined by Henderson-Haselbalch equation, which represents the relationship between pH and pKa (the dissociation constant of weak acid). This equation is used to prepare buffer solution with a certain pH. Let us consider a weak acid that ionizes as follows: HA -> H+ + A- Then equilibrium constant K will be: Ka = [H+][A-]/[HA] Rearranging the equation, we get: Ka[HA] = [H+][A-] [H+] = Ka[HA]/[A] Taking log on both the sides: log [H+] = log Ka + log [HA]/[A] Multiplying by -1, we get: -log [H+] = -log Ka + (-log [HA]/[A]) pH = pKa + log [A]/[HA] or pH = pKa + log [Salt]/[Acid] pH = pKa + log [Cojugate base]/[Conjugate acid] - Calculate the pH of mixture of acetic acid of 0.1 M and sodium acetate of 0.2 M, where Ka = 1.8 x 10^-5 pH = pKa + log [Salt]/[Acid] = -log 1.8 x 10^-5 + log 0.2/0.1 = 4.74 + 0.3 = 5.04 - Calculate the pH of mixture of acetic acid of 0.6 M and potassium acetate of 0.6 M, where Ka = 1.8 x 10^-5 pH = pKa + log [Salt]/[Acid] = -log 1.8 x 10^-5 + log 0.6/0.6 = 4.74 + 0 = 4.74 ### Factors Affecting pH of a Buffer The pH of a buffer solution is determined by two factors: a. The value of pK: ↓ pK → ↓ pH of the solution. b. The ratio of salt to acid concentrations **Buffer capacity** is the number of grams of strong acid or base that must be added to change the pH of one liter of solution by one pH unit. - The buffering capacity of a buffer is defined as the ability of the buffer to resist changes in pH when an acid or base is added. - The buffer capacity is only affected by the ratio of salt to acid concentrations.. **Effective range of buffer:** A buffer is most effective when: [acid] = [base] or pH = pKa ### Blood buffers: (Buffers Act Quickly, But Not Permanently) - Buffers are the first line of defense against acid load. Buffers can respond immediately to addition of acid or base, but they do not serve to eliminate the acid from the body. - For the final elimination of acids, the respiratory and renal regulations are very essential. The blood contains three important buffer systems: I- Bicarbonate buffer (H_2CO_3/HCO_3¯) II- Phosphate buffer (H_2PO_4¯/HPO_4^(2-)) III- Protein buffer e.g (H+-albumin/albumin) ### Bicarbonate buffer system (ECF): 1) It is the most important buffer system in plasma. 2) It is present in very high concentration than other buffer systems (26 to 28 millimole per liter). 3) It consists of carbonic acid H_2CO_3 and bicarbonate salt HCO_3. 4) The carbonic acid is the respiratory component because it is regulated by the respiratory system. So this buffer is directly linked up with respiration. 5) The bicarbonate salt HCO_3 is the metabolic component because it is regulated by kidney. Normally, all acids except H_2CO_3 react with bicarbonate to liberate CO_2. 7) In plasma the concentration of [HCO_3¯] = 24 mmol/l (alkali reserve) and the concentration of [H_2CO_3] = 1.2 m mol/l. 8) The pH of bicarbonate buffer in plasma is calculated from Henderson-Hasselbalch equation: pH = pKa + log [Salt]/[Acid] = 6.1 + log 24/1.2 = 6.1 + 1.3 = 7.4 ### Advantages of bicarbonate buffer system: Bicarbonate buffer system is efficient as compared to other buffer systems as: 1. It is present in very high concentration than other buffer systems. (26 to 28 millimole per liter). 2. Produces H_2CO_3, which is a weak acid and volatile and CO_2 is exhaled out. 3. It is a very good physiological buffer and acts as a front line defence. ### Disadvantage: As a chemical buffer, it is rather weak, pKa is further away from the physiological pH. ### Phosphate buffer system (ICF): 1) The concentration of phosphate in the blood is very low thus it is unimportant buffer in blood. 2) Phosphate are important buffer intracellularly and in urine where their concentration is higher. 3) It consists of H_2PO_4¯/HPO_4^(2-) in plasma in ratio 4:1. This ratio is kept constant with the help of the kidneys. Thus, phosphate buffer system is directly linked up with the kidneys. 4) KH_2PO_4 acts as acid by donating proton (H+), and is converted to its conjugate base K_2HPO_4. The pH of phosphate buffer in plasma is calculated by Henderson-Hasselbalch equation: pH = pKa + log [Salt]/[Acid] = 6.8 + log 4/1 = 6.8 + 0.6 = 7.4 ### Advantage of phosphate buffer: As a chemical buffer it is very effective and better, as pka approaches physiological pH. ### Disadvantage: Concentration in blood is low (1.0 millimole/liter), As a physiological buffer it is less efficient. **Note:** - In urine, the concentration of NaH_2PO_4 exceeds that of Na_2HPO_4 and the ratio is 9:1. ### Protein buffer: - Buffering capacity of plasma proteins is much less than Hb.. - Intracellular proteins and plasma proteins form a large pool of protein molecules, which act as buffer molecules. - Hemoglobin (in RBCs) is one of the most important intracellular proteins. - Other intracellular molecules like histone proteins associated with nucleic acids also act as buffers. - The most effective amino acid in protein buffer is histidine which contains imidazole (pKa 6.8) and binds H+.. - Hb molecule contains (38) histidine residues. - Albumin contains (16) histidine residues. - At the tissue level, hemoglobin binds to H⁺ ions and helps to transport CO₂ as HCO_3 with a minimum change in pH (referred to as isohydric transport). - As the concentration of HCO_3 increases in the RBC, it diffuses into plasma with the concentration gradient, in exchange for Cl⁻ ions, to maintain electrical neutrality. This phenomenon, referred to as chloride shift, helps to generate HCO_3¯. - The term "isohydric transport" refers to the actual chemical mechanism whereby CO_2 is transported from the tissues to the lungs. ### Respiratory regulation of pH: (The Second Line of Defense) i. Respiratory system provides a rapid mechanism for the maintenance of acid-base balance. This is achieved by regulating the concentration of carbonic acid (H_2CO_3) in the blood ii. The rate of respiration (rate of elimination of CO_2) is controlled by the respiratory center (in brain) which is sensitive to changes in the pH of blood. iii. pH of plasma (acidosis) → Stimulation of the respiratory center → respiratory rate → hyperventilation →↑ elimination of CO_2 → ↓ H_2CO_3 level.. iv. Respiratory control of blood pH is rapid but only a short term regulatory process/ since hyperventilation cannot proceed for long. **Figure:** Diagram comparing the function of RBCs in peripheral tissue versus RBCs in lungs.