Buffers
Introduction
Buffers are important in biochemical processes.
Whether they occur naturally in plasma or in the
cytosol of cells, buffers assure biological reactions
occur under conditions of optimal pH. The word
“buffer” is so common in biochemistry its replaces the
word “water” in experimental protocols.
Buffers
3
What are buffers?
 Buffers are solutions in which the pH remains
relatively constant, even when small amounts of acid
or base are added
Example:
 Contain a weak acid (CH3COOH) and its
salt(CH3COONa); H2CO3 & NaHCO3
 Or a weak base (B) and its (BHCl); (NH3 and
NH4Cl)
Importance of Buffers
Buffers
resist changes in pH from the addition of acid or
base.
in the body, absorb H3O+ or OH- from foods and
cellular processes to maintain pH.
are important in the proper functioning of cells
and blood.
in blood maintain a pH close to 7.4. A change in
the pH of the blood affects the uptake of oxygen
and cellular processes.
5
Acidic buffer solution
 An acidic buffer solution is simply one which has a pH
less than 7.
 Acidic buffer solutions are commonly made from a
weak acid and one of its salts - often a sodium salt.
 Example: acetic acid and sodium acetate
CH3COOH and CH3COONa
Alkaline(Basic) buffer solutions
 An alkaline buffer solution has a pH greater than 7.
 Alkaline buffer solutions are commonly made from
a weak base and one of its salts.
 A frequently used example is a mixture of
ammonia solution and
solution. NH3 and NH4Cl
ammonium chloride
How buffers work
 Equilibrium between acid and base.
 Example: Acetate buffer
 CH3COOH  CH3COO- + H+
 If more H+ is added to this solution, it simply shifts
the equilibrium to the left, absorbing H+, so the
[H+] remains unchanged.
 If H+ is removed (e.g. by adding OH-) then the
equilibrium shifts to the right, releasing H+ to keep
the pH constant
Buffer Capacity
 The buffer capacity is the amount of acid or base
that can be added before a significant change in
pH
 This depends on the amounts of HA and Apresent in the buffer
 Most efficient buffer is when
[A-] = 1
[HA]
Henderson-Hasselbach Equation
 Derived from the equilibrium expression of a weak
acid and the pH equation.
[A-]
pH = pKa + log[HA]
 This equation allows you to determine the pH of a
buffer solution
 pKa = -log Ka
 Example 1: What is the pH of a mixture of 5 ml of 0.1
M sodium acetate and 4 ml of 0.1 M acetic acid (pKa =
4.76 at 250C)?Concentration of CH3COOH=4/9 × 0.1 M
Concentration of CH3COONa=5/9 × 0.1 M
[NA]
pH = pKa + log
[HA]
pH = 4.76+ log (5/4)
pH = 4.76 + ( + 0.097) = 4.86
Learning Check
Which combination(s) make a buffer solution?
A. HCl and KCl
B. H2CO3 and NaHCO3
C. H3PO4 and NaCl
D. CH3COOH and CH3COOK
E. NaOH and NaCl
F. NH3 and NH4Cl
12
Solution
B. H2CO3 + NaHCO3
A weak acid and its salt
D. CH3COOH + CH3COOK
A weak acid and its salt.
F. NH3and NH4Cl
A weak base and its salt
13
PREPARATION OF BUFFER SOLUTIONS
Three practical methods to prepare a buffer:
1- First Method : By the Titration, in the presence of one of
the two buffer forms with strong base or acid:
 Prepare a buffer composed of an acid and its salt by
adding a strong base(e.g. NaOH) to a weak acid (e.g.
Acetic acid) until the required pH is obtained
 If the other form of buffer is available (in this case
sodium acetate), a strong acid is added (e.g. HCl) until
the required pH is obtained.
CH3COONa+HClCH3COOH+NaCl
 So acetate buffer is formed(CH3COOH/CH3COONa)
2- Second Method: Using the buffer pKa , calculate the
amounts (in moles) of acid/salt or base/salt present in
the buffer at the desired pH.
 If both forms (i.e., the acid and the salt) are available,
convert the amount required from moles to grams
,using the molecular weight of that component, and
then weigh out the correct amounts of both forms. Or
convert moles to volume if the stock is available in the
liquid form.
.3- The Thired Method: Using table
 Find a table of the correct amounts of acid/salt or
base/salt required for different pH's
 Dissolve the components in slightly less water
than is required for the final solution volume.
 Check that the pH and correct if necessary.
 Add water to the final volume.