SECTIONS

Acids, Bases, and pH

1. Introduction

2. Acids and Bases

3. The pH Scale

4. Strong and Weak Acids

5. Acid-Conjugate Base
Pairs
6. The HendersonHasselbalch Equation
7. Buffers
8. Review

INTRODUCTION
Acids and bases in our lives
Acids and bases are two broad classes of compounds that have a great deal of importance in both
chemistry and biochemistry. In industry, acids and bases are used in various reactions. Sulfuric acid,
one of the most important industrial chemicals, is used to manufacture fertilizers for agriculture, to
make man-made fibers, paints and dyes, and to purify petroleum products. The base sodium
hydroxide (sometimes called caustic soda, or lye) is used for the production of fabrics, paper, and
cleaning agents.
Acids and bases are also common in our everyday lives. Acids
have a sour taste, and many of the sour-tasting foods with which
we are familiar are acidic. Vinegar, for example, is diluted acetic
acid (normal household vinegar is a 3% solution of acetic acid), and
gives salad dressings and pickled vegetables their tart tastes. Other
familiar foods with sour flavors get their tartness from acids:
oranges and lemons contain citric acid, wine contains tartaric acid,
and aspirin contains acetylsalicylic acid.
While the tart taste of some acids can be a pleasant addition to
many kinds of foods, bases have a bitter flavor, and therefore are
not typically preferred for human consumption. However, many
people have acquired tastes for caffeine and nicotine, both of which
are alkaloids, a class of nitrogen-containing bases. Quinine, the
ingredient that gives tonic water its bitter taste, is also an alkaloid.
Antacids, including sodium bicarbonate (baking soda) and calcium
Acids are sour or tart: vinegar,
carbonate (Tums), are basic and work by neutralizing stomach
lemon and orange juice, wine,
acids to water and carbon dioxide (CO2) gas. Bases feel slippery
aspirin.
because they are soapy in nature, which is why they are used in
cleaners. Lye (sodium hydroxide), a strong base, can dissolve
grease and protein, and is used in oven cleaners, products for unclogging drains, and in hair-

removal lotions.
Acids and bases are also essential for life. For example,
without the strong acid present in our stomachs, we would not
be able to digest food. More importantly, an organism needs to
be able to control the level of acid within its cells for life to be
possible. The worlds oceans are able to sustain life in part
because the level of acid (or pH) of the water is kept constant
through acidbase chemistry. In the same way, the pH of living
cells is tightly regulated to allow cells to maintain their
structural integrity, and for cellular processes to function.

Bases are bitter: coffee, cigarettes,

tonic water, baking soda, antacid
tablets, soap.

To keep living cells or body fluids at the proper pH, special

compounds called buffers are used. A solution containing a
buffering agent tends to resist changes in pH, so that biochemical reactions, which often produce or
use up acids, can proceed without producing wild fluctuations in cellular pH. In this way, the whole
cell is protected from high concentrations of acid, but critical acidbase reactions can still proceed in
the cell. Since pH is so critical to biological molecules, any biochemical experiment that involves
studying proteins or cellular systems needs to be performed in a buffered solution to keep the
molecules in their natural, or native, conformation. Thus, an understanding of how buffers can help
regulate pH is essential for any biochemist.
Learning objectives

Learn the definitions of acid, base, and pH.

Know how to convert from [H+] to pH and vice versa.

Understand the meaning of Ka and pKa.

Know how to use the HendersonHasselbalch equation.

Learn how buffers work.

ACIDS AND BASES

Chemical definitions of acid and base
Acids and bases can be defined by their chemical properties in several different ways. However, because acid
base reactions important for biochemistry occur in aqueous solution (that is, in water), the BronstedLowry
definitions of acids and bases are generally used. In the BronstedLowry scheme:
An acid is a proton (H+) donor
A base is a proton (H+) acceptor

A proton is a hydrogen atom that has lost its electron and is designated H +. Thus, acids are chemical compounds
that tend to give up H+ ions when placed in water. For example, when hydrochloric acid (HCl) is placed in water,
the reaction that occurs is
HCl

H3 O +

H2 O

proton
donor
(acid)

proton
acceptor
(base)

The HCl has donated its H+ ion, which was accepted by the water molecule. Notice how the H + ion, once
donated by the HCl molecule, does not simply float around in the water freely, but instead associates strongly
with water (the proton acceptor) to form a hydronium ion (H3O+). Thus, using the BronstedLowry convention, the
acid proton H+ is often written as H3O+ to reflect this fact.
Similarly, ammonia (NH3) is a base because it acts as a proton acceptor:
NH3

proton
acceptor
(base)

NH4+

H2 O

proton
donor
(acid)

As you can see in the examples above, water can act not only as a proton acceptor, but also as a proton donor.
In fact, water actually reacts with itself. The result is that even in a sample of pure water, not every molecule is in
the form of H2O. A very small proportion of molecules undergo this reaction, called autoionization:
H2 O

H3 O +

H2 O

proton
donor
(acid)

proton
acceptor
(base)

Autoionization of water
Just how common is this reaction? Is a cup of water out of the tap really just a collection of charged molecules
rather than H2O?
Let's look closer. The equilibrium constant for the reaction of water molecules is
[H3O+][OH]
Keq

=
[H2O]

(the square brackets indicate molar concentrations)

By convention, when a reaction involves water, the concentration of water is factored into the value of K eq
the reaction of water molecules, the resulting equilibrium constant is known as K w, the ion product constant of
water, and its value is 1014.
Kw = Keq[H2O] = [OH][H3O+] = 1014

Scientists determined experimentally that the equilibrium constant for this reaction of water molecules is
Kw = [OH][H3O+] = 1014

The value of Kw is known as the ion product constant of water. Note that by convention, water molecules are not
included in the equilibrium constant expression, because the reaction is happening in water, and the
concentration of water is already factored in to the Kw constant. Therefore Kw is just the Keq for the ionization of
water, with the concentration of water factored into the constant (Kw = Keq/[H2O]).
Pure water is neutral, having neither an overall positive nor an overall negative charge. This is because each
time a water molecule donates a proton, another accepts it, so that the concentrations of OH and H3O+ are
always equal:
[OH] = [H3O+] in pure water

Since the Kw expression for pure water states that the product of these two must equal 10 14:
[H3O+] = 107 M and [OH] = 107

for pure water ONLY

These are very minute concentrations, especially since pure water is 55.5 M. In other words, the fraction of
+

3O and OH ions is very tiny compared to the overall H 2O concentration. In fact, there are over 500 million
times more H2O molecules than either H3O+ or OH ions in water!
The Kw value is still very important for biochemists, however, since it aids in the understanding of acidbase
chemistry. The Kw value is useful because it says that in any aqueous solution, the product of [OH] and
[H3O+] ions is a constant value. Thus, an acid solution contains not only H3O+ ions, but also a lesser amount of
OH ions.
Example 1

If a 101 M solution of HCl was prepared, what would be the equilibrium concentrations of H3O+and OH
Solution
Returning to the hydrochloric acid dissociation reaction from above, we can describe the reaction of the acid with
water as:
HCl

proton
donor

H2 O

H3 O +

proton
acceptor

Because HCl is a strong acid, essentially all of the acid will dissociate, and the concentration of H 3O+ ions would
be equal to 101 M. Plugging this into the Kw expression gives the following:
Kw = [OH][H3O+
[OH](101
[OH
Similarly, a basic solution contains not only OH ions, but also some H3O+ ions. To further grasp this
concept,interact with the graph below. Note that while the ratio of hydronium ion to hydroxide ion changes, the
w (the product of these two) does not change.

Move the slider left to add more acid or right to add more
base to an aqueous solution

THE pH SCALE
In the previous section, we learned that the Kw expression for water is
Kw = [OH][H3O+] = 1014

and that pure water, being neutral, has

[OH] = [H3O+] = 107 M

By convention, acidic solutions are those where the [H3O+] is greater than the [OH], and basic
solutions are those where the [OH] is greater than the [H3O+]. In other words:

Acidic solutions have [H3O+] greater than 107 M

(what follows from the Kw expression is that [OH] is less than 107 M).

Basic solutions have an [H3O+] less than 107 M

(what follows from the Kw expression is that [OH] is greater than 107 M).
Note: even in acidic solutions, OH is present, and in basic solutions H3O+ is also present.
We have learned that in aqueous solutions the actual concentration of [H 3O+] is often very small. For
instance, in pure water, which is neutral, the [H3O+] is 107 M. Since living cells keep their acid
concentrations close to neutral, working with these small physiological [H 3O+] values as exponents
can be very cumbersome. To circumvent this problem, a logarithmic measure of the [H 3O+] (or [H+]
for short) was devised by a Danish scientist in 1909. He called this measure pH, short for the power
of hydrogen. The relationship of pH to the concentration of H+ ions is given by the following equation:
pH = log[H+]

If pure water has an [H+] concentration of 107 M, then neutral water has a pH of log (107), or 7.
This value was designated the middle of the pH scale. The pH scale ranges from 0 to 14, reflecting
[H+] from 1 M (pH = 0) to 1014 M (pH = 14). Note how much easier it is to manipulate pH values from
0 to 14 than to deal with variations in H+ concentrations from 1 M to 0.00000000000001 M. Keep in
mind that since the log function removes the exponents from the [H +] expression, a one point
change in the pH is equivalent to a tenfold change in the [H+]!

Also keep in mind that in nonaqueous solution, it is possible to have a [H +] that lies outside the range
of the pH scale. However, such values of H+ are not found in biological systems, and are not
physiologically relevant.

Example 2
If the [H+] of a solution is 0.1 M, what is the [OH]? What is the pH?
Solution
From the Kw expression,
Kw = [OH][H3O+] = 1014
[OH](101) = 1014
(note 0.1 M = 101)

[OH] = 1013
From the pH equation,
pH = log [H+]
pH = log (0.1) or pH = log(101)
pH = (1)
pH = 1

STRONG AND WEAK ACIDS

Ionization
There are two classes of acids: strong acids and weak acids. Note that strong acids and bases are
rarely a concern in biochemistry (as they are in general chemistry). Strong acids ionize (break into
ions) completely when dissolved in water. In other words, a strong acid is a good proton donor. For
example, the strong acid HCl gives up all its protons to water:
HCl + H2O

Cl + H3O+

There are only a few strong acids, which are all listed below. HCl (hydrochloric acid, found in
stomach digestive juices) is a good example of a strong acid.
THE STRONG ACIDS
Chemical Formula

Name

HCl

Hydrochloric acid

HBr

Hydrobromic acid

HI

Hydroiodic acid

H2SO4

Sulfuric acid

HNO3

Nitric acid

HClO4

Perchloric acid

Because strong acids completely ionize, the [H+] of a solution made with a strong acid is easily
figured out, since it is equal to the molarity of the solution. Thus, a 0.1 M solution of HCl has a [H +] of
0.1. A 0.029 M solution of HCl has a [H+] of 0.029 and so on. (Remember that this only holds true as
long as the molarity of your solution is somewhat greater than 107 M. Otherwise, for solutions so
dilute that the H+ normally present in pure water is comparable or larger, you will need to take the
[H+] of pure water into consideration.)

Similar to the case with strong acids, there are just a few strong bases that ionize completely. For
example, when sodium hydroxide ionizes in water, the resulting hydroxide ion readily accepts a
proton:
NaOH

Na+ + OH

The strong bases are listed in the table below.

THE STRONG ACIDS
Chemical Formula

Name

NaOH

Sodium hydroxide

LiOH

Lithium hydroxide

KOH

Potassium hydroxide

RbOH

Rubidium hydroxide

CsOH

Cesium hydroxide

TlOH

Thallium hydroxide

Ca(OH)2

Calcium hydroxide

Sr(OH)2

Strontium hydroxide

Ba(OH)2

Barium hydroxide

Most acids of biological origin are weak. In part 1 of this tutorial, we saw that in pure water, a very
small percentage of the H2O molecules dissociate to form H+ and OH ions. Most of the molecules in
pure water remain intact as H2O. The same can be said of weak acids. When weak acids are
dissolved in water, only some of the acid molecules actually dissociate (break apart), while most
others remain intact.

The percentage of the molecules that break apart depends on the weak acid. Some weak acids
dissociate very minimally, and as a consequence such an acid has little acid strength. Other weak
acids dissociate more, and as a consequence have more acid strength.
The degree of dissociation of a weak acid in water is described by the acid dissociation constant,
K a.
For the dissociation of acetic acid (the acid in vinegar) in water,
CH3COOH + H2O

CH3COO + H3O+

the equilibrium (or acid dissociation) constant expression for the reaction is given by
[CH3COO][H3O+]
Ka

1.74 x 105

[CH3COOH]

(Remember that by convention, [H2O] is left out of the expression because the reaction is taking
place in water). The a subscript on Ka is a reminder that this number represents an acid
dissociation constant. The larger the Ka, the more the acid dissociates (or reacts with water) to form
H3O+ ions, and the stronger the acid.
Note how Ka values for weak acids tend to be very small numbers. Because these numbers with
exponents tend to be cumbersome, Ka values are usually converted to pKa values, much in the same
way as [H+] values are converted to pH.
pKa = log Ka

For example, the Ka of acetic acid is 1.74 x 105.

The pKa for acetic acid is therefore pKa = log (1.74 x 105) = 4.76.
Below is a table listing some common weak acids and their Ka and pKa values.
SOME ACIDS AND pKa VALUES
Acid Name

Formula

Found in

Ka

pKa

4.5 x 107

6.35

1.74 x 105

4.76

CH3CH(OH)COOH Active muscles, milk, microorganisms

1.40 x 104

3.85

Formic acid
(weak acid)

HCOOH

Poison in stings from ants and nettle plants, leather

manufacture

1.78 x 104

3.75

Hydrofluoric acid
(weak acid)

HF

Semiconductor manufacture, glass etching

6.76 x 104

3.17

Carbonic acid
(weak acid)

H2CO3

Sea water, human blood, soft drinks

Acetic acid
(weak acid)

CH3COOH

Vinegar, spoiled wine

Lactic acid
(weak acid)

Hydronium ion
H3 O +
(the dividing line)

Water, aqueous solutions

Hydrochloric acid HCl

(strong acid)

Stomach, industrial processes

2.0 x 106 6.3

Note that the smaller the Ka, the larger the pKa. Thus, stronger acids are represented by larger
Ka values, but smaller pKa values.

Also, whether an acid is strong of weak can be readily identified by either its K a or pKa:
IDENTIFYING ACID STRENGTH
Type

Ka

pKa

Strong acid

>1

< 0 (negative)

Weak acid

<1

> 0 (positive)

Hydronium acid (H3O+), which is the protonated form of water, is the dividing line between a strong or
weak acid. In other words, a strong acid is defined as one that ionizes to a larger degree than the

H3O+ acid form of water does (Ka = 1, pKa = 0). Another way to put this is that if you put a strong acid
in water, most of it ionizes, while for a weak acid most stays in its conjugate acid (protonated) form.

ACIDS AND THEIR CONJUGATE BASES

Remember the BronstedLowry definition of an acid: an acid is a proton donor. Clearly, in our
previous example acetic acid donates its proton to H2O in the forward reaction, resulting in the
formation of the acetate ion. However, in the reverse reaction, the acetate ion accepts a proton from
H3O+ to revert to CH3COOH. From the BronstedLowry definition, a base is a proton acceptor.
CH3COOH

acetic
acid

H2 O

CH3COO

water

acetate
ion

H3 O +

hydronium
ion

Thus, the acetate ion acts as a base in the reverse reaction, and is called the conjugate base of
acetic acid. Acetic acid and the acetate ion are known as an acidbase conjugate pair. Interestingly
enough, there is a second acidbase conjugate pair in the reaction above. Notice how in the reverse
reaction, H3O+ donates a proton (making it an acid), while in the forward reaction, H 2O accepts a
proton (making it a base). H2O and H3O+ are a conjugate acidbase pair, with H3O+ being the
conjugate acid and H2O being the conjugate base.

In a conjugate acidbase pair, the conjugate acid has one more

proton than its corresponding conjugate base.

THE HENDERSONHASSELBALCH EQUATION

We have learned that the pH of a solution depends on the concentration of H + (more correctly called
H3O+) ions present in the solution. We have also learned that weak acids dont fully dissociate into
ions when placed in water. How much they ionize, or how acid they can make the water in which
they dissolve, is different for each weak acid. The pKa is a measure of the weak acids ability to

ionize. Thus, pH and pKa seem like very similar terms, and it is important not to get them confused.
Remember, pH is a direct measure of the H+ concentration, while pKa represents a particular acids
ability to ionize (dissociate into ions, including H+).
The HendersonHasselbalch equation
However, since the extent of ionization of a weak acid (the pK a) influences the final concentration of
H+ ions (the pH) of the solution, there must be a relationship between pH and the pK a of a weak acid.
This relationship is given by the HendersonHasselbalch equation:
[base]
pH

pKa

log
[acid]

Where base represents the conjugate base, and acid the conjugate acid, of a conjugate acid
base pair (for a review of conjugate acidbase pairs see section 5). Note that there is a way for the
pH and the pKa of a solution to be the same: the term log [base]/[acid] must be equal to zero! When
does that happen? If we solve the equation

log [base]/[acid] = 0
[base]/[acid] = 1
[base] = [acid]

Thus, only in the rare circumstance when the molar concentrations of the conjugate acid and
conjugate base in a solution are equal, the pH = pKa. Below is a graph of a weak acid titration, which
is also a graph of the Henderson-Hasselbalch equation, showing how pH changes as acid or base is
added. Note that when the pH = pKa, there are equivalent amounts of the conjugate acid and base. It
is also important to note that just because the amounts of conjugate acid and base are the same at
the pKa, the pH is not 7 (neutral) at this point. Remember, pH is a function of the the concentration of
H3O+, not the conjugate acid, and because weak acids do not ionize completely, these
concentrations will not be the same.

Buffer titration/HendersonHasselbalch graph: how the pH of a

buffered solution changes as acid or base is added.

It is important to remember that the pH of a weak acid solution can be calculated, using the K a value
of the weak acid.
Example 4
What is the pH when 10 mL of 1 M acetic acid is added to pure water so that the final volume
is 100 mL? The Ka for acetic acid is 1.74 x 105.
The reaction that occurs when acetic acid dissociates in water is
CH3COOH + H2O

CH3COO + H3O+

It is given that 10 mL of 1 M acetic acid is diluted to 100 mL in water. Thus, the initial acetic acid
concentration is
(0.01 liter)(1 mol/liter) / (0.100 liter total volume) = 0.1 M acetic acid.

Once the acetic acid is diluted in water, the dissociation reaction begins and continues until it
reaches equilibrium. We can use a table to organize what happens to all the species involved in the
reaction:
ACETIC ACID DISSOCIATION TABLE

Initial concentration
Equilibrium concentration

CH3COOH

CH3COO

H3 O +

0.1 M

0.1 M [CH3COOH]

[CH3COO]

[H3O+]

We also know that the Ka of acetic acid is 1.74 x 105 . Using the Ka expression for acetic acid:
[CH3COO][ H3O+]
Ka

=
[CH3COOH]

Plugging in the known Ka value, as well as the equilibrium concentrations of each species:
[CH3COO][ H3O+]
1.74 x 105

=
0.1 [CH3COO]

But since one H3O+ is produced for every CH3COO ion,

[CH3COO] = [ H3O+]

and the expression becomes

[ H3O+][ H3O+]
1.74 x 105

=
0.1 [ H3O+]

This is of course now just an algebra problem, where we have an quadratic equation in the form of
x2
1.74 x 105

=
0.1 x2

But dont drag out that quadratic formula just yet! Thankfully there is a shortcut that we can use.
Remember that weak acids dissociate very poorly in water, which means that the final CH 3COO
concentration will be very small compared to the 0.1 M acetic acid that we started with. This means
that the equilibrium concentration of CH3COOH will be very close to 0.1 M. This changes the
Ka expression to
[CH3COO][H3O+]
1.74 x 105

=
0.1

or
1.74 x 105

[H3O+][H3O+]

0.1
1.74 x 106 = [H3O+]2
[H3O+] = 1.32 x 103

Using the pHSECTIONS

equation:
pH = 1.logIntroduction
[H3O+]
pH = log (1.32 x 103)
pH = 2.88
2. Acids and Bases
3. The pH Scale
4. Strong and Weak Acids
5. Acid-Conjugate Base
Pairs
6. The HendersonHasselbalch Equation
7. Buffers
8. Review

BUFFERS
In the previous section, the HendersonHasselbalch equation was introduced, which is
handy for dealing with pH calculations involving weak acids and their conjugate bases.
This equation is very important to biochemists, who are always concerned with keeping
proteins and other biological molecules in the lab at their proper pH. Indeed, a solution
that contains a weak acid and its conjugate base has the special property of being able
to resist changing its pH when either a base or an acid is added to it. Such a solution is
called a buffer because the solution is protected, or buffered, from pH changes even
when H3O+ or OH ions are added to the solution. Buffers help biochemists study
biomolecular reactions in the laboratory by stabilizing the pH of solutions used for

experiments.
To get a better sense of the utility of buffer solutions, lets compare the difference
between adding a strong base to a buffered solution, and adding the same amount of
strong base to water. This is done in the examples below.

Example 5
If 10 mL of 1M NaOH are added to one liter of a buffer that is 0.3 M acetic acid and 0.2
M sodium acetate (Na+CH3COO), how much does the pH change? The pKa for acetic
acid is 4.76.
Answer:
In the original solution, acetic acid is the weak acid, and acetate is the conjugate base.
Thus, [base] = 0.2 M, and [acid] = 0.3 M.
Using the HendersonHasselbalch equation,
[Base]
pH

pKa

log
[Acid]

pH = 4.72 + log (0.2/0.3)

pH = 4.54

NaOH is a strong base, and dissociates completely. Therefore, adding the strong base
results in
(0.01 liter)(1 M) = 0.01 moles of OH ions into the solution.
All of the OH ions will react with the acetic acid to form acetate ions:
CH3COOH + OH

CH3COO + H2O

Thus, when NaOH is added, the number of acetic acid molecules in the solution will
decrease, but the number of acetate molecules in the solution will increase. In this case,
0.01 moles of acetic acid will be used up to neutralize the 0.01 moles of OH, and this
will form 0.01 moles of acetate ion in the solution.
At the end of the reaction:

Moles of
acetic acid

Moles of
acetate

moles of acetic acid

before the reaction

moles of acetic acid used

up in the reaction

0.3 moles

0.01 moles

0.29
moles

0.21
moles

moles of acetate before

moles of acetate made in
+
the reaction
the reaction

0.2 moles

0.01 moles

Now we are almost ready to plug back into the HendersonHasselbalch equation to find
the new pH of the solution. We just need to remember that the volume has changed
slightly, from 1 liter (1000 mL) to 1 liter plus 10 mL (1010 mL). The Henderson
Hasselbalch equation requires that the [Base] and [Acid] be molar (M, or mol/L)
quantities. However, notice how that because both the acid and base species are in the
same volume, the volume correction cancels out:
[Base]
pH

pKa

log
[Acid]

(0.29 moles/1.01 liter)

pH

pKa

log
0.21 moles/ 1.01 liter)

pH = 4.72 + log 1.38

pH = 4.86
The pH change is 4.86 4.54 = 0.32 units.

Example 6
How much does the pH change when 10 mL of 1 M NaOH is added to 1 liter of water?
Answer: Pure water has a pH of 7.0. NaOH is a strong base, and once it has been
added to the water it will dissociate completely, contributing 0.01 moles OH ions, in a
total volume of 1010 mL ( 1 liter plus the 10 mL NaOH added). Thus,

[OH] = 0.01 moles / 1.01 liter = 0.0099 M

Plugging into the Kw expression: [OH][H3O+] = 1014:
(0.0099)[H3O+] = 1014
[H3O+] = 1.01 x 1012 M

Then plugging into the pH equation:

pH = log [H+]
pH = log 1.01 x 1012
pH = 11.99

The pH has shifted from the neutral pH of 7 for the pure water to a pH of almost 12 with
the NaOH added, a shift of 5 pH units!
Buffering agents stabilize the pH of aqueous solutions
In example 5, the pH of the buffer solution changed by less than 0.5 units when the
NaOH was added. Yet adding the same amount of NaOH to pure water resulted in a 5
unit pH shift in example 6. Clearly, the conjugate acid/base pair was able to protect the
solution from a large change in pH.
A buffer is a solution that resists a change in pH when acids or bases are added to it.
How is this possible? Buffers work by acting a little bit like a sponge, soaking up excess
H3O+ or OH ions when they are added to a solution. The dissociation reaction for
acetic acid shows that a solution of acetic acid contains acetate ions:
CH3COO + H3O+

CH3COOH + H2O

How do acetic acid and the acetate ion work to buffer the solution? If a strong acid, such
as hydrochloric acid, is added to this buffer, the H3O+ ions generated will react with the
acetate ions, removing them from the solution:
CH3COO

acetate
ion

H3 O +

CH3COOH

hydronium
ion

acetic
acid

H2 O
water

Similarly, if a strong base, generating lots of OH ions, were added to the acetic acid
buffer solution
CH3COOH

OH

CH3COO

H2 O

acetic
acid

hydroxide
ion

acetate
acid

water

The OH ions are removed via a reaction with acetic acid. It is important to remember
that buffers cannot maintain their pH indefinitely as more acid or base is added. Imagine
slowly pouring a bucketful of water onto a small sponge on the kitchen floor. At first, as
the water pours out of the bucket it is absorbed by the sponge, keeping the kitchen floor
dry. But once the sponge is soaked through, it can hold no more and the water spills
onto the floor as fast as if the sponge werent there. In the same way, buffers will protect
the pH of the solution to some extent, but if they are inundated by large amounts of
H3O+ or OH ions, all the available conjugate acid or base molecules will have been
used up and pH changes will rapidly occur.

Thus, buffers only work as long as the amount of conjugate acid and base ions are
large compared to the H3O+amount of or OH ions to be removed. Remember from the
Henderson-Hasselbalch equation (section 5) that the numbers of conjugate acid
molecules is equal to the number of conjugate base molecules when the pH = pKa:
pH = pKa + log ( [Base] / [Acid] )
if [Base] = [Acid], then
pH = pKa + log 1
pH = pKa

Because the conjugate acid and conjugate base molecules are available in equal
amounts when the pH = pKa, the buffer solution has the strongest ability to protect
against pH changes caused by incoming H3O+ or OHions when the pH of the solution
is close to the pKa of the conjugate acid. If the pH of the solution strays too far from the
pKa point, the buffer uses up all its available conjugate acid or base molecules, and the
pH starts to fall (or rise) dramatically. At this point, the buffering capacity of the buffer
has been exceeded. this can be seen in the buffer (weak acid) titration graph below.
Buffers protect the pH of a solution best within one pH unit of the pKa.

Buffer titration: within one unit of the pKa, the solution resists
changes in pH.

Buffers are important for biochemistry because the structures (and therefore the
functions) of biological molecules are stable within a relatively narrow range of pH
values.The evolution of life, which is believed to have begun in water, was likely due in
part to the stable pH of seawater. Even today, the largest buffered systems in the world
are the Earth's oceans. Carbon dioxide (CO2) from the Earths atmosphere reacts with
H2O to produce carbonic acid, H2CO3. Carbonic acid in turn reacts with water to form
bicarbonate:
H2CO3

carbonic
acid

H2 O

HCO3

water

bicarbonate
ion

H3 O +

hydronium
ion

Furthermore, the bicarbonate ion can also react with water to form carbonate:
HCO3

bicarbonate
ion

H2 O

CO32

water

carbonate
ion

H3 O +

hydronium
ion

The carbonate and bicarbonate ions act together as conjugate acid/base pair to keep
the pH of the ocean at about 8.2.

In the living organisms, proteins (the molecules which do most of the work of a cell)
are very sensitive to acid concentration, because they only fold into their proper threedimensional shapes within a small pH range. Some of the individual amino acid building
blocks that make up a protein have ionizable side chains that change their ionization
state, or charge, at different pH. Thus, if the pH changes, some of the attractive forces
that hold the protein together will also change.
The ionization states of the side chain of the amino acid lysine are shown below. The
pKa of lysine's side chain amino group is approximately 10.8. Therefore, at a pH lower
than the pKa, the side chain is in the protonated (conjugate acid) state; whereas at a pH
higher than the pKa, the side chain is unprotonated (conjugate base).

Amino acid charge changes: if the pH of the environment around

a basic amino acid shifts so that it is greater than the pKa, the
amino acid is deprotonated, no longer carries a charge, and is
unable to form ionic bonds.

Proteins consist of long chains of amino acids, many of which include acidic or basic
groups. These groups are chaged at physiological pH, and form ionic interactions with
each other that contribute to the stability of the folded protein. A change in pH of the
protein's aqueous environment may alter the charge of some of these charged groups,
disrupting the ionic interactions and the stability of the proteins structure.

Remember that the HendersonHasselbalch equation was used for calculating the pH
of a solution containing a weak acid and its conjugate base. Since such a solution is a
buffer, the HendersonHasselbalch equation is extremely useful for calculations

involving buffers, allowing the following:

Determination of the pH of buffered solutions.

Determination of the ratio of conjugate base to conjugate acid at a given pH.

Example 7
A solution of lactic acid has twice as many conjugate acid molecules as conjugate base
molecules. If the pKa of lactic acid is 3.85, what is the pH of such a solution?
Answer:
We are given that the concentration of conjugate acid is twice the concentration of
conjugate base. Thus,
[acid] = 2 [base]
Substituting into the Henderson-Hasselbalch equation:
pH = pKa + log ([base]/[acid])
pH = 3.85 + log
([base]/2[base])
SECTIONS
pH = 1.
3.85Introduction
+ log (0.5)
pH = 3.85 + (0.30)
pH = 2.
3.55Acids and Bases
3. The pH Scale
4. Strong and Weak Acids
5. Acid-Conjugate Base
Pairs
6. The HendersonHasselbalch Equation
7. Buffers
8. Review

REVIEW
Acids and bases are compounds that are important in industry, our everyday lives, and
most critically, for the functioning of our bodies. Because biochemistry is aqueous
chemistry, the most common definition of an acid is the BronstedLowry definition:
An acid is a proton (H+) donor
A base is a proton (H+) acceptor

Water can be both a proton donor and a proton acceptor, and a small proportion of
water molecules do react with one another:

H2 O

proton
donor

OH

H2 O

H3 O +

proton
acceptor

The ion product constant of this reaction, Kw, is

Kw = [OH][H3O+] = 1014

This expression is useful because it tells us that in water, the product of the OH and
H3O+ ion concentrations is always a constant value. It also indicates that in pure water,
the concentration of both OH ions and H3O+ions is 107 M. Because under
physiological conditions, the H3O+ concentrations tend to be small and the numbers
cumbersome to deal with, the pH scale was invented to make manipulation of these
numbers easier:
pH = log [H+]

Remember that H+ is written as H3O+ (the hydronium ion) in the BronstedLowry

definition of acids and bases to indicate that free protons (H+ ions) always strongly
associate with a water molecule.
In the pH scale, which ranges from 0 to 14, pH 7 is neutral, while lower pHs are acidic
and higher pHs are basic. What follows from the Kw expression is that
Acidic solutions have [H3O+] greater than 107.
Basic solutions have [H3O+] smaller than 107.

There are only a few strong acids, which dissociate completely (break apart into their
constituent ions) completely in water. Most acids are weak, meaning they only partially
dissociate in water. The degree of dissociation of a weak acid is given by its Ka, acid
dissociation constant. Because Ka values are small for weak acids, the Ka is often
converted to a pKa value.
pKa = log Ka

The larger the Ka value, and the smaller the pKa value, the stronger the acid.
When a weak acid dissociates in water, it results in the formation of a hydronium ion
and a basic molecule that is able to accept a proton. This basic molecule is called the
conjugate base of the weak acid, and the weak acid and its conjugate base are said to

be a conjugate acidbase pair.

HB

weak
acid

H2 O

H3 O +

water

hydronium
ion

conjugate
base

In a conjugate acidbase pair, the conjugate acid has one more proton than its
conjugate base.
A solution made with a weak acid and its conjugate base is called a buffer, because it
resists changes in pH when OH ions or H3O+ ions are added. The Henderson
Hasselbach equation is useful for buffer calculations:
[Base]
pH

pKa

log
[Acid]

The HendersonHasselbalch equation allows the following:

Determination of the pH of buffered solutions.

Determination of the ratio of conjugate base to conjugate acid at a given pH .

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