BCH2301 GENERAL BIOCHEMISTRY I

Acids and Bases, pH, and Buffers

A. Acids and Bases

Electrolytes when dissolved in water split up into two or more electrically charged
particles. The charged particles, which are formed when the electrolyte is dissolved in
water, are called ions, and this process is called ionization. The process of ionization is
reversible. These ions may have positive or negative charge. The total number of
positive charges is equal to the total number of negative charges in the solution. Since
they are charged, they are responsible for carrying the electric current through the
solution.
In 1887, Svante Arrhenius explained this property of electrolytes, and hence, it is known
as Arrhenius theory of electrolyte dissociation. The extent of ionization is given by the
ratio of the total number of dissociated molecules to the number of molecules
dissolved. The degree of ionization can be increased by diluting the solution in case of
weak electrolytes.

What are Acids and Bases?

According to the Arrhenius theory of dissociation, an acid is a substance that can release
hydrogen ion (H+) when dissolved in water. For example; Hydrochloric acid

–
HCl H+ + Cl

While a base is a substance that can release a hydroxyl ion (OH-) when dissolved in
water. For example; Sodium Hydroxide

–
NaOH Na+ + OH

However, the Arrhenius definitions have some limitations of shortcomings. Some of

which are;
 1. Water is essential to determine whether a substance is an acid or a base.
2. It does not explain the acidity of basicity of non-aqueous solvents such as
Benzene
3. It does not explain the acidity or basicity of gases such as ammonia (NH3) and
carbondioxide (CO2).
However, in 1923 Danish chemist Johannes Nicolaus Brønsted and English chemist
Thomas Martin Lowry proposed the theory that defined acids as substances which are
able to donate protons, and bases as substances which are able to accept protons.

In the above example, HA is an acid because it donates a proton to water to form A–

and a hydronium ion. A– is an anion liberated by the deprotonation of the acid and
behaves like a base, so it is called conjugate base. Similarly, B is a base because it
accepted a proton from water to form BH+ and a hydroxide ion. BH+ is created b
protonation of the base and behaves like an acid, so it is called conjugate acid.
Interestingly, water acts both as an acid (proton donor) and a base (proton acceptor).
Such substances are referred to as Amphoteric.
Note: When the conjugate base of an acid is weak i.e. it has less affinity for a proton, it
gets dissociated easily and hence that acid is considered a strong acid. However, when
the conjugate base of an acid is strong i.e. it has high affinity for a proton, it does not get
dissociated easily and hence that acid is considered a weak acid.
This implies that the degree of dissociation of an acid, determines its strength.
Therefore it is important to identify an index that measures the strength of an acid. To
do that, we look at the equilibrium constant for the dissociation reaction of an acid (HA),
which can be written as follows;
–
HA ⇌ H+ + A
By applying the law of mass action at equilibrium,

𝐻 + [𝐴− ]
𝐾𝑎 = , Ka is the dissociation constant
[𝐻𝐴]

The equilibrium constants for ionization reactions are commonly called dissociation
constants (Ka).
The reaction above can be rearranged thus;

𝐾𝑎 × [𝐻𝐴]
[𝐻 +] =
[𝐴−]

Multiplying the above equation by -1 and taking logarithm of both sides will give;

𝐾𝑎 × [𝐻𝐴]
−[𝐻 +] = −
[𝐴− ]

[𝐻𝐴]
− log[𝐻 +] = − log 𝐾𝑎 − log
[𝐴−]

Let’s refer to the expression, − log[𝐻 +] as pH and − log 𝐾𝑎, as pKa;

As the [𝐻 +] increases, the pH of the solution decreases and as the [𝐻 +] decreases, the
pH increases. Thus, [𝐻 + ] and pH are reciprocally related.

[𝐴−]
𝑝𝐻 = 𝑝𝐾𝑎 + log
[𝐻𝐴]

[𝐶𝑜𝑛𝑗𝑢𝑔𝑎𝑡𝑒 𝑏𝑎𝑠𝑒]
𝑝𝐻 = 𝑝𝐾𝑎 + log
[𝐴𝑐𝑖𝑑]

This expression is called the Handerson-Hasselbalch equation. If the conjugate base and
acid concentration is the same, the pH = pKa. The value of pKa is lower for strong acids
and higher for weak acids.
B. pH
To understand the concept of pH, let us take close look water. Being a weak electrolyte,
water dissociates to form H+ and OH– ions.
–
H2O ⇌ H+ + OH
Since the ionization of water is a reversible reaction, using the law of mass action at
equilibrium;
𝐻 + [𝑂𝐻 −]
𝐾𝑒𝑞 =
[𝐻2 𝑂]
For water, the molar mass is approximately 18 g/mol and 1 litre of water weighs 1000 g,
therefore, the concentration of H2O in pure water can be calculated to be 1000/18 or
55.56 moles/litre. The equilibrium constant Keq for the acid—base reaction between
H2O molecules is very small. At 25 °C, Keq = 1.8 x 10-16.
Since the concentration of H2O in dilute aqueous solution is essentially unchanged from
that in pure H2O, this figure may be taken as a constant. It is usually in fact incorporated
into the expression for the dissociation constant of water to give—
𝐻 + 𝑂𝐻 − = 𝐾𝑒𝑞 × [𝐻2 𝑂]
𝐻 + 𝑂𝐻 − = 1.8 × 10−16 × 55.56
𝐻 + 𝑂𝐻 − = 1 × 10−14
𝐾𝑤 = 𝐻 + 𝑂𝐻 − = 1 × 10−14
This Kw is called as ionic product of water and it expresses the relation between the
concentration of H+ and OH– ions in aqueous solutions. Thus, in water at 25 °C,
𝐻 + = 10−7 , and 𝑂𝐻 − = 10−7 . Now let’s proceed.
Since,
𝐻 + 𝑂𝐻 − = 1 × 10−14
Taking logarithm of both sides,
log 𝐻 + + log 𝑂𝐻 − = log( 1 × 10−14 ) = −14
Multiplying through by –1
−log 𝐻 + − log 𝑂𝐻 − = 14
Recall, pH is − log[𝐻 +], so − log 𝑂𝐻 − can be pOH. Then;
𝒑𝑯 + 𝒑𝑶𝑯 = 𝟏𝟒
The expression 𝒑𝑯 + 𝒑𝑶𝑯 = 𝟏𝟒 represents the pH scale. Therefore, if the [H+] of a
solution is greater than 10–7, the pH is less that 7 and the solution is acidic. Conversely, if
the [H+] concentration is less than 10–7, the pH is greater than 7 and the solution is
alkaline or basic.

C. Buffers
Buffers are solutions that resist changes in the pH of a when acid or alkali is added. The
buffer system has an acid component and salt component. Buffers are prepared by
mixing a weak acid and its salt with a strong base or by mixing weak base and its salt
with a strong acid. Buffers are used in a variety of ways. Some of the more important
ones are used in biological and biochemical research. In fact, it is often necessary for the
researcher to prepare a buffer of certain pH to support the chemical conditions the
system needs to maintain.
To understand how buffers work, let’s use the acetate buffer which is made up of acetic
14 Textbook of Biochemistry for Nurses
acid and sodium acetate i.e. CH3COOH/CH3COONa.
14 Acetate buffer
Textbook is made upforofNurses
of Biochemistry acetic acid and sodium acetate.
a. When an acid like HCl is added to the acetate buffer system, CH3COONa gets
Acetic acid / Sodium acetate
Acetate buffer is made up– of acetic acid and sodium acetate.
converted to CH3COO , which CH3combines
COOH / CH with the proton (H+) released by the HCl
3COONa
Acetic acid / Sodium acetate
+ to the acetate buffer system,
(a)to form
When CH acid
3
like
COOH. HCl is
The added
Na CHcombines with the Cl– to formCH3COONa (sodium
NaCl which is a acetate)
salt.
– 3COOH / CH3COONa
gets converted to CH3COO (acetate). Acetate combines with the protons released by
Thethe
(a) formed
When acidCH
strong 3COOH
like
acids HCl isisadded
to give aacetic
weak acid
toacid,
the with buffer
acetate
which a very
is small
acid,pH,
system,
a weak CH 3hence
andCOONa net pHin change
(sodium
the change pHacetate)
is very
–
gets
small.converted to CH 3 COO (acetate). Acetate combines with the protons released by
is very small.
the strong acids to give acetic acid, which is a weak acid, and the change in pH is very
small.

– +
b.(b)When
When a strongbase
a strong alkali
likelike NaOH
NaOH is added,CHOH
is added, combines with the H released by–the
3COOH dissociates to form CH3COO
+
CH3COOH to form water. Na combines with acetate to form sodium acetate.
(b) H+. aThe
andWhen H+combines
strong alkali like with
NaOHthe is added, OH+– combines
OH– released by with the H+ released by the
– NaOH to form H2O, while
NaOH Na + OH
CH3COOH
–
to form water. Na++ combines with acetate to form sodium acetate.
CH3COO combines with the Na to form CH3COONa
+ –
which is a salt.
CH3COOH NaOH Na –+ OH+
CH3COO H–

CH3COO– + + H–
CH3COOH Na OH–

Na+ OH–

CH3COONa + H2O

CH3COONa + H2O
According to Henderson-Hasselbalch equation, when the concentration of the salt component
and the acid component
BuffersAccording
are most of the buffer system
effective in the pHequation,is equal,
range of pH of
± the the solution
1 concentration equals
unit of the ofdesired pKa. system.
At this pH,
to Henderson-Hasselbalch when
buffer can respond equally to both added acid as well as the alkali. the salt component
and the acid component
Furthermore, the closer ofthe
theconcentrations
buffer system is equal,
of thepH of theacid
weak solution
and equals pKa. At this
its conjugate weakpH,
buffer can respond
Buffering Capacity equally to both added acid as well as the alkali.
base salt or the weak base and its conjugate weak acid salt are to one another, the
Buffering
Capacity of theCapacity
buffer to resist the change in the pH of a solution when an acid or alkali is added
better thebuffering
is called chance of achieving
capacity. It isgreater buffering
estimated capacity.
by calculating the amount of acid or alkali required
Capacity of the buffer to resist the change in the pH of a solution when an acid or alkali is added
to change
What
is called isthe pH of one
thenbuffering
buffering litre of buffer by one unit.
capacity?
capacity. It is estimated by calculating the amount of acid or alkali required
Buffering
to change thecapacity depends
pH of one upon
litre of the by
buffer following factors:
one unit.
This is 1.amount of acid or base
The concentration
required
of the
to change the pH ofofone litre of a buffer by 1
Buffering capacity depends uponacid
the and base component
following factors: the buffer
unit. The As the concentration of acidupon
and the
basefollowing
component of the buffer increases, buffering
1. buffering capacity depends
The concentration of the acid and base factors:
component of the buffer
capacity of the buffer also increases.
As
1. 2.TheThe the concentration
concentration of theofacid
acidand
andbase
basecomponent
component of
of the
the buffer
buffer increases, buffering
pH of the Buffer
capacity of the buffer also increases.
Buffer
As the can act best atofpHacid
= pKa,
andand its buffering range
of isthe
about one increases,
pH unit above
its or
2. The concentration
pH of the Buffer base component buffer
below the pKa value.
Buffer can act best at pH = pKa, and its buffering range is about one pH unit above or
buffering capacity also increases.
below the pKa value.
 There will be no free OH– ions in solution and pH will remain same.
Similarly when HCl is added to buffer solution—
CH3COOH + CH3COONa + HCl ® NaCl + 2CH3COOH
2. The pH of the buffer
The salt (NaCl) is formed and no free H+ are in solution.
A buffer can actofbest
Addition acidwhenor pH
base= pKa
does andnot
its buffering
alter therange is about
pH of oneThe
buffer. pH unit
buffer acts almost
D:data/New-age~/Question/Third/III-1.pm5/Third proof/14-3-05
absorbing the added + –
above or below the pKafreevalue.H or OH ions.
Q. What are Physiological buffers? Or Describe the physiological importance of
Physiological Buffers
Ans. Buffers are of great physiological importance. The importance of buffers depe
6 Question Bank of Biochemistry
factors
In animal as molar
complex, concentration
bicarbonate of buffer
and carbonic components,
acid buffer ratiosystem
is the buffering of concentration
found in of conjugat
ofwill
There weakbe noacid.
free OH– ions in solution and pH will remain same.
circulating blood. This buffer neutralizes stronger dietary and metabolic acids (HA)
Similarly when HCl is added to buffer solution—
In animals complex and vital buffer system is bicarbonate and carbonic acid bu
converting them
CH3into
circulating weak
blood.
COOH + CHbases
This (A–+) with
buffer
3COONa ®the increase
HClneutralizes
NaCl in H2CO3dietary
stronger
+ 2CH 3COOH
. Stronger
andbases (B) are acids (HA) con
metabolic
alsoTheinto weak
salt (NaCl)
converted is bases
into formed and–no
(A
weak acids) (BH
with+ Hthe
free increase
+ are in solution.
) with the in H3–2. CO3. Stronger bases (B) are also convert
rise in HCO
Addition
acidsof(BH
acid+or base does
) with the not
risealter
in the
HCOpH of–.buffer. The buffer acts almost as if it were
absorbing the added free H+ or OH– ions. 3

Q. What are Physiological buffers?

HA + HCO –
Or Describe – + H CO importance of buffers.
theAphysiological
3 2 3
Ans. Buffers are of great physiological importance. The + importance of buffers depend on several
B + H CO3
factors as molar concentration of buffer2 components,
BH + HCO3 of conjugate base to that
ratio of concentration
of weak acid.
The buffer ratio in blood remain unchanged by the respiratory elimination of H2CO3
In animals complex and vital buffer system is bicarbonate and carbonic acid buffer found in
The buffer ratioelimination
urinary in blood remains
of HCO –
unchanged by the respiratory elimination of H2CO3 as
circulating blood. This buffer neutralizes 3 . dietary and metabolic acids (HA) converting them
stronger
into weak bases –
(A other
) with the increase present
in H CO3. Stronger bases (B) are also converted
CO2 or The
the urinary buffers
elimination inother
of HCO23–. The circulating blood
buffers present inare as into weak
Phosphate
circulating blood buffer, Haem
acids (BH+) with the rise in HCO3–.
Oxyhaemoglobin buffer.
are phosphateHA
buffer, haemoglobin
+ HCO –
and oxyhaemoglobin.
A– + H CO
3 2 3
Q. What
B+Hare
CO Indicators?
BH+ + HCO
2 3 3
Indicators
The bufferAns.
ratio inIndicators
blood remain are chemical
unchanged compounds,
by the respiratory which
elimination of Hchange their colour with change
2CO3 as CO 2 or the
urinary elimination of HCO –.
solution
Indicators in which
are chemical they arewhich
3 compounds, present. These
change theirindicators
colour withare dyes,
change which
in pH are weak acids or
of the
The other buffers present in circulating blood are as Phosphate buffer, Haemoglobin and
They have
Oxyhaemoglobin
solution one are
buffer.
in which they colour, but after
present. Theseionization
indicators change their
are dyes, original
which colour.
are weak acidsUniversal
or indicator i
indicators, which gives a wide variety of colours over a wide range of the pH. Indicators
Q. What are Indicators?
weak bases.(a)They have one colour,
Determination of thebutendafter ionization
point in acidchange their original colour.
base titrations.
Ans. Indicators are chemical compounds, which change their colour with change in pH of the
Universal
solution (b) Determination
indicator
in which they is
area present. ofofindicators,
mixtureThese pH of solutions.
indicators which gives
are dyes, a wide
which variety
are weak acidsoforcolours over a
weak bases.
They have one colour, but after ionization change their original colour. Universal indicator is a mixture of
wide range
indicators, of the
givespH.
Indicators
which Indicators
a wide variety ofare usedover
colours in (a)
PH Determination
arange
wide range of the pH.ofIndicators
the endare point
usedinin acid Colour
(a) Determination of the end point in acid base titrations.
base titrations (b) Determination of pH of solutions.
(b) Determination of pH of solutions. In acid solution In alkalin
Indicators
Thymol blue PH range 1.2 – 2.8 Colour Red Yellow
Methyl yellow 2.9In–acid
4.0solution In Red
alkaline solution Yellow
Methyl orange
Thymol blue 1.2 – 2.8 – 4.4
3.1Red Red
Yellow Orange yel
Methyl yellow – 4.0
Methyl red 2.9
Methyl orange 3.1 – 4.4
Red
– 6.1
4.3Red Yellow
Red
Orange yellow
Yellow
Phenol red 4.3 – 6.1
Methyl red – 8.3
6.7Red Yellow
Yellow Red
Phenol red 6.7 – 8.3
Phenolphthalein 8.2 – 10.0
Yellow RedColourless Violet red
Phenolphthalein 8.2 – 10.0 Colourless Violet red

Q.dialysis.
Q. Define Define dialysis.
Ans. Dialysis is a process by which the more diffusible materials can be separated from non
Ans. Dialysis is a process by which the more diffusible materials can be separat
diffusible materials. If we take water solution of egg albumin and sugar in the upper small container
diffusible materials. If we take water solution of egg albumin and sugar in the upper sm
Let’s solve some problems.
Q1. Given the under listed [H+], determine whether each of the molarities are acidic, basic or
neutral.
a. 1.0 x 10-7 M
b. 3.8 x 10-12 M
c. 7.3 x 10-4 M
d. 9.6 x 10-2 M
e. 8.4 x 10-10 M

Q2. Given the following set of information, calculate the requested quantities.
a. [H+] = 4.1 x 10-4 M, find pH and [OH-]
b. [OH-] = 6.6 x 10-9 M, find pOH and [H+]
c. pH = 9.0, find [H+], [OH-] and pOH
d. pOH = 12.5, find [H+], [OH-] and pH

Q3. a. If a solution was prepared by dissolving 0.02 moles of acetic acid in water to give 1 litre of
solution. What is the pH? (Pka = 4.8)
b. To this solution, 0.008 moles of concentrated sodium hydroxide was added. What is the new
pH?.

Thank you.