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Chemistry 201

Lecture 15

Practical aspects of buffers

NC State University
The everyday pH scale
To review what pH means in
practice, we consider the pH
of everyday substances that
we know from experience.

Remember that [H+] = 10-pH.

pH + pOH = 14

Therefore that [OH-] = 10pH-14.


Two ways to make a buffer
Method 1 Method 2

Add the acid and conjugate Add a strong acid to the weak base
base to the solution in a (or vice versa) until the desired
defined proportion. proportion [A-]/[HA] is obtained.
Buffer strength
The ratio [A-]/[HA] should be as close as possible 1:1,
but the amounts may vary. To make a stronger buffer
you simply need to increase the amount of each
component. Lets investigate.

Suppose we add 1 mL of 1 M HCl to 1 liter of solution.


The final concentration of HCl is 0.001 M.
Buffer strength
The ratio [A-]/[HA] should be as close as possible 1:1,
but the amounts may vary. To make a stronger buffer
you simply need to increase the amount of each
component. Lets investigate.

Suppose we add 1 mL of 1 M HCl to 1 liter of solution.


The final concentration of HCl is 0.001 M. pH = 3
Buffer strength
The ratio [A-]/[HA] should be as close as possible 1:1, but the
amounts may vary. To make a stronger buffer you simply
need to increase the amount of each component. Lets
investigate.

Suppose we add 1 mL of 1 M HCl to 1 liter of solution. The


final concentration of HCl is 0.001 M. pH = 3

Suppose we add 1 mL of 1 M HCl to 1 liter of 10 mM optimal


phosphate buffer solution (pKa = 7.2) solution. The final
concentration of HCl is 0.001 M. pH = pKa + log10([A-]/[HA])
Buffer strength
Keeping in mind that the unbuffered solution in this example
([HCl] = 0.001 M) would be pH = 3

Suppose we add 1 mL of 1 M HCl to 1 liter of 10 mM


optimal phosphate buffer solution (pKa = 7.2) solution. The
final concentration of HCl is 0.001 M.
pH = pKa + log10([A-]/[HA])
[A-] = 0.005 0.001 = 0.004 [HA] = 0.005 + 0.001 = 0.006
pH = 7.2 + log10(0.004/0.006) = 7.02

If the target pH = 7.2 (i.e. pH = pKa) then this buffer is too


weak. An error of 0.2 pH units could be significant.
Buffer strength
Keeping in mind that the unbuffered solution in this example
([HCl] = 0.001 M) would be pH = 3

Suppose we add 1 mL of 1 M HCl to 1 liter of 100 mM


optimal phosphate buffer solution (pKa = 7.2) solution. The
final concentration of HCl is 0.001 M.
pH = pKa + log10([A-]/[HA])
[A-] = 0.05 0.001 = 0.049 [HA] = 0.05 + 0.001 = 0.051
pH = 7.2 + log10(0.049/0.051) = 7.18

If the target pH = 7.2 (i.e. pH = pKa) then this buffer is


reasonable. The difference is only -0.02.
Buffer strength
Keeping in mind that the unbuffered solution in this example
([HCl] = 0.001 M) would be pH = 3

Suppose we add 1 mL of 1 M HCl to 1 liter of 300 mM


optimal phosphate buffer solution (pKa = 7.2) solution. The
final concentration of HCl is 0.001 M.
pH = pKa + log10([A-]/[HA])
[A-] = 0.150 0.001 = 0.149 [HA] = 0.150 + 0.001 = 0.151
pH = 7.2 + log10(0.149/0.151) = 7.194

If the target pH = 7.2 (i.e. pH = pKa) then we would say that


this buffer is definitely strong enough, difference = -0.006
Titrating to make a buffer
You can create a buffer either by adding the acid and
Its conjugate base to a solution or by titrating in strong
base to acid (or vice versa). Remember, regardless of
the method used to prepare it:

The buffering strength is maximum when [HA] = [A-]

The buffering range is considered to extend from


[HA] / [A-] = 0.1 to [HA] / [A-] = 10. This is subjective.
Wertz suggests 0.01 to 100 is an acceptable range.
Understanding the titration curve
Suppose we want to make a buffer by titrating [OH-].
We cannot use the H-H equation initially. We do not know
the concentration of [A-]. Instead at this initial point we will
use the other form of the
equilibrium constant and
make an ICE table.

Starting point [HA] = [HA]0

Added [OH-] = 0
Understanding the titration curve
Suppose we want to make a buffer by titrating [OH-].
We cannot use the H-H equation initially. We do not know
the concentration of [A-]. Instead at this initial point we will
use the other form of the
equilibrium constant and
make an ICE table.

Starting point [HA] = [HA]0

Added [OH-] = 0
Understanding the titration curve

We can calculate x = [H+]


and therefore the pH from
the equilibrium constant.

Starting point [HA] = [HA]0

Added [OH-] = 0
Understanding the titration curve
When we have a buffer we can use the Hendersen-Hasselbach
equation. This is nice since it is the simplest treatment of the
acid-base equilibrium. In the case shown we have pH = pKa.

Maximum buffer capacity


[HA] = [A-].

Added [OH-] = 1/2 [HA]0


Understanding the titration curve
The buffer range is defined as approximately from:
pH = pKa 1 R = [A-]/[HA] = 0.1 [OH-] ~ 0.091 [HA]0
to
pH = pKa + 1 R = [A-]/[HA] = 10 [OH-] ~ 0.91 [HA]0

Maximum buffer capacity


[HA] = [A-] when Buffer region

[OH-] ~ 0.5 [HA]0


Understanding the titration curve
Once the solution moves outside the buffer range the pH shoots
up. The equivalence point is reached when the added is equal
to the original acid concentration, i.e. [OH-] ~ [HA]0
At this point one can no longer
use the H-H equation. Instead,
we assume that [A-] ~ [HA]0
Then we use the base equil-
-ibrium:

Buffer region

Note that pKb = 14 - pKa


Types of buffers
There are inorganic buffers, e.g. phosphate, but there are
many more organic buffers. In fact, the number of buffers
is staggering.

Organic buffers: Tris, HEPES, MOPS, MES

Biological buffers: citrate, acetate, carbonate, malonate,

Proteins themselves are polyelectrolytes and therefore


tend to the buffer the solution they are in. This can have
important physiological impact (e.g. hemoglobin).
Tris buffer
Tris(hydroxymethyl)aminomethane), is
an organic buffer with the formula
(HOCH2)3CNH2. Tris has a pKa = 8.07.
The buffer range is 7.07 9.07.
It is is widely used as a component for
solutions of nucleic acids, proteins and for
any application in which phosphate is not a
good choice.
For example, calcium phosphate has a low solubility product,
Which means that phosphate buffers are a poor choice in
any application where calcium is present.

Important point: Tris can react with aldehydes since it is a


primary amine. Choice of buffer should done by consulation
of the chemical interactions in your application.
Other organic buffers

HEPES: pKa = 7.5

MOPS: pKa = 7.2


Citrate buffer
Citric acid is found in abundance
in citrus fruits. It is also part of the
citric acid cycle in biochemistry.
It is a triprotic acid, with three
carboxylic acid groups as seen
by its structure (on the right).

pKa1 = 3.14

pKa2 = 4.75

pKa3 = 6.40 Citric acid crystals


under polarized light
Citrate buffer: species in solution

The middle H atom has the lowest


pKa. This is because the neighbor-
ing OH group has an electron with-
drawing effect that stabilizes the
negative charge created.
Amino acids are amphipathic

All amino acids contain the carboxylic acid and amino


group. These have very different pKa values so the amino
acids have a doubly charged form (zwitterion) at pH 7.
In proteins only the N- and C-terminus have these pKas.
Amino acids can be classified in part according to the pKa
of their side chains.
Amino acid side
chain pKa
Proteins are made of up of a
number of titratable amino
acids. At pH 7 the terminal
carboxyl, aspartate and
glutamate have a negative
charge. Terminal amino,
lysine and arginine are
positively charged. Others
are neutral, but can be
charged due to interactions
within the protein. The pKa
of any of these groups may
be altered by the protein.
The isoelectric point of a protein pI
Proteins have many titratable groups on their surface. It is
not possible to define a single pH since all of the groups
have different pKa values. However, we can define the point
at which protein is neutral in charge: the isoelectric point.
At the isoelectric point the protein has 0 net charge, which
means that there as many positive as negative groups on
the surface.

The isoelectric point concept applies to polymers,


nanoparticles etc. Any macromolecule can be described in
terms of its overall charge.

IMPORTANT: When pH = pI a macromolecule has a neutral


surface. This is the minimum stability point. Macromolecules
tend to precipitate at this poin.
Hemoglobin as a buffer
Hemoglobin is the most abundant protein inside of red blood cells
and accounts for one-third of the mass of the cell. During the
conversion of CO2 into HCO3-, H+ liberated in the reaction are
buffered by hemoglobin, which is reduced by the dissociation of
oxygen. This buffering helps maintain normal pH. The process is
reversed in the pulmonary capillaries to re-form CO2, which then
can diffuse into the air sacs to be exhaled into the atmosphere.
Bicarbonate (hydrogen carbonate):
an important regulator in the body

As with the phosphate buffer, a weak acid or weak base captures


the free ions, and a significant change in pH is prevented.
Bicarbonate ions and carbonic acid are present in the blood in a
20:1 ratio if the blood pH is within the normal range. With 20 times
more bicarbonate than carbonic acid, this capture system is most
efficient at buffering changes that would make the blood more
acidic. This is useful because most of the bodys metabolic wastes,
such as lactic acid and ketones, are acids. Carbonic acid levels in
the blood are controlled by the expiration of CO2 through the lungs.
The role of carbonic anhydrase

The enzyme does not change the equilibrium, but it accelerates the
Rate of reaching the equilibrium on each side of a membrane.
Acid/Base Mixtures : Reactions

How do you calculate pH after an


acid/base reaction occurs?
Text : Section 7.3
Examples: Strong acids and bases

What is the pH when:


a) 25 mL of 0.30 M HCl are added
to 35 mL of 0.20 M NaOH?

b) 15 mL of 0.25 M HClO4 are added


to 25 mL of 0.20 M NaOH?
Examples: Strong acids and bases
What is the pH when 25 mL of 0.30 M HCl are
added to 35 mL of 0.20 M NaOH?
Examples: Strong acids and bases
What is the pH when 25 mL of 0.30 M HCl are
added to 35 mL of 0.20 M NaOH?

Step 1. Calculate dilutions. First add the volumes

Total volume = 25 mL + 35 mL = 60 mL

Calculate concentrations in the solution


Examples: Strong acids and bases
What is the pH when 25 mL of 0.30 M HCl are
added to 35 mL of 0.20 M NaOH?
Step 2. Write a balanced chemical reaction for the limiting
reaction and the excess reaction.
Limiting reaction

Species HCl NaOH Na+ Cl-


Initial 0.125 0.117 0.0 0.0
Difference -x -x x x
Final 0.125-x 0.117-x x x
Examples: Strong acids and bases
What is the pH when 25 mL of 0.30 M HCl are
added to 35 mL of 0.20 M NaOH?
Step 2. Write a balanced chemical reaction for the limiting
reaction and the excess reaction.
Limiting reaction

Species HCl NaOH Na+ Cl-


Initial 0.125 0.117 0.0 0.0
Difference -0.117 -0.117 0.117 0.117
Final 0.008 0.0 0.117 0.117

Excess reaction
Species HCl H+ Cl-
Initial 0.008 0.0 0.0
Final 0.0 0.008 0.008
Examples: Strong acids and bases
What is the pH when 25 mL of 0.30 M HCl are
added to 35 mL of 0.20 M NaOH?
Recognize that both HCl and NaOH are strong acid/base,
respectively. Therefore, rather than find the equilibrium
constant, we assume that the reaction goes to completion.
In this case we find the limiting reagent which is NaOH.

In the general case we could include both H+ and OH- on


the right hand side. We may not know initially which one
is going to dominate, since we must first calculate the
limiting reagent.
Examples: Strong acids and bases
What is the pH when 25 mL of 0.30 M HCl are
added to 35 mL of 0.20 M NaOH?
Examples: Strong acids and bases
What is the pH when 25 mL of 0.30 M HCl are
added to 35 mL of 0.20 M NaOH?
Short cut method:
Step 1. calculate number of moles of each reagent
Examples: Strong acids and bases
What is the pH when 25 mL of 0.30 M HCl are
added to 35 mL of 0.20 M NaOH?
Short cut method:
Step 1. calculate number of moles of each reagent

Step 2. calculate the total volume (0.025 + 0.035 = 0.060 L)


Step 3. make a table considering only H+ and OH-
Species H+ OH- H2O
Initial 7.5 7.0 0.0
Difference -7.0 -7.0 +7.0
Final 0.5 0.0 7.0
Examples: Strong acids and bases
What is the pH when 25 mL of 0.30 M HCl are
added to 35 mL of 0.20 M NaOH?
Short cut method:
Step 4. calculate the final concentration of [H+ ]

Step 5. calculate the pH


Examples: Strong acids and bases
15 mL of 0.25 M HClO4 are added to 25 mL of
0.20 M NaOH?
Examples: Strong acids and bases
15 mL of 0.25 M HClO4 are added to 25 mL of
0.20 M NaOH?
Step 1. Calculate dilutions. First add the volumes

Total volume = 25 mL + 15 mL = 40 mL

Calculate concentrations in the solution


Examples: Strong acids and bases
15 mL of 0.25 M HClO4 are added to 25 mL of
0.20 M NaOH?
Step 2. Write a balanced chemical reaction for the limiting rxn.

Species HClO4 NaOH Na+ ClO4-


Initial 0.0937 0.125 0.0 0.0
Difference -0.0937 -0.0937 0.0937 0.0937
Final 0.0 0.0313 0.0937 0.0937

Excess rxn is
Species NaOH Na+ OH-
Initial 0.0313 0.0 0.0
Final 0.0 0.0313 0.0313
use rxn table
goes 100%
calc pH or pOH

Strong Strong
Acid Base

Weak Weak
Acid Base
Examples: One strong and one weak
What is the pH when 50. mL of 0.25 M NaOH are
added to 40. mL of 0.20 M HF?
Examples: One strong and one weak
What is the pH when 50.0 mL of 0.25 M NaOH are
added to 40.0 mL of 0.20 M HF?

Step 1. Calculate dilutions. First add the volumes

Total volume = 50 mL + 40 mL = 90 mL

Calculate concentrations in the solution


Examples: One strong and one weak
What is the pH when 50.0 mL of 0.25 M NaOH are
added to 40.0 mL of 0.20 M HF?
Step 2. Write a balanced chemical reaction and determine
the form of the equilibrium constant. Make a reaction table.
Examples: One strong and one weak
What is the pH when 50.0 mL of 0.25 M NaOH are
added to 40.0 mL of 0.20 M HF?
Step 2. contd. We calculate 1/Kb from the data in the tables
Ka = 6.6 x 10-4.

Step 3. Make a reaction table.

Species HF OH- F-
Initial 0.0888 0.139 0.0
Difference -x -x x
Final 0.0888-x 0.139-x x
Examples: One strong and one weak
What is the pH when 50.0 mL of 0.25 M NaOH are
added to 40.0 mL of 0.20 M HF?
Step 4. Solve for x
Examples: One strong and one weak
What is the pH when 50.0 mL of 0.25 M NaOH are
added to 40.0 mL of 0.20 M HF?
Step 5. Calculate OH- and pOH.

We see from the table that [OH-] = 0.139 x = 0.0511

BIG PICTURE: This example is very high on the titration


Curve. We can get [OH-] approximately from
[OH-] = [OH-]0 [HF] 0 = 0.139 0.888 = 0.051
The additional information from the ICE is in the third
decimal place.
Strong base exceeds weak acid
The key point of the previous problem is that we are no longer
in the buffer range. We cannot use H-H in this case. Since:

[OH-]0 > [HA]0

While Kb still applies it is often


unnecessary since [OH-] is in
excess.

If you need to use Kb


then use:
Buffer region
use rxn table
goes 100%
calc pH or pOH

Strong Strong
Acid Base
use rxn table
goes 100% Weak Weak
buffer : H-H Acid Base
base : pOH
Examples: One strong and one weak
What is the pH when 25.0 mL of 0.40 M HCl are
added to 40.0 mL of 0.30 M NH3?
Examples: One strong and one weak
What is the pH when 25.0 mL of 0.40 M HCl are
added to 40.0 mL of 0.30 M NH3?
The total volume is 65 mL so the final concentrations
are:
[HCl] = 25/65(0.40 M) = 0.154 M
[NH3] = 40/65(0.30 M) = 0.184 M
In this case the [HCl] < [NH3] so this will make a buffer.
Assume that the strong acid reacts completely then at
equilibrium we have:
[NH3] = 0.184 0.154 M = 0.03 M and [NH4+] = 0.154 M
Examples: One strong and one weak
use rxn table
goes 100%
calc pH or pOH

Strong Strong
Acid Base
use rxn table use rxn table
goes 100% Weak Weak goes 100%
buffer : H-H Acid Base buffer : H-H
base : pOH acid : pH
Examples: Weak acid and weak base
For a reaction of a weak acid and a weak base
we need to calculate the equilibrium constant
from the known Kas. We take the example of
ammonium acetate.

We see that the overal reaction is composed


of two acid-base equilibria
for acetate

for ammonia
Examples: Weak acid and weak base
Therefore, the overall equilibrium constant for
the reaction is

Now, that we can see how to calculate the


Equilibrium constant, we can solve any acid-base
Reaction problem using the standard methods
That we have used.
Step. 1. determine dilutions
Step. 2. set up the reaction table
Step. 3. solve for the unknown and then calculate pH
Examples: Weak acid and weak base
20.0 mL of 0.30 M NaH2PO4 was added to
20.0 mL of 0.30 M NaHS. What are the concs.
of all species at equil.?
Examples: Weak acid and weak base
20.0 mL of 0.30 M NaH2PO4 was added to
20.0 mL of 0.30 M NaHS. What are the concs.
of all species at equil.?
Solution: Look up the Ka for each reaction involved
in this acid-base equilibrium.
Examples: Weak acid and weak base
Species
Initial 0.15 0.15 0.0 0.0
Difference -x -x x x
Final 0.15-x 0.15-x x x
Strong Strong
Acid Base

Weak Weak
Acid Base

if conjugates if not conjugates


use H-H calc. K
calc pH calc concs.
Goals
Calculate the pH of acid/base mixtures
Calculate the pH at any point in a titration
Calculate the concentration of all species in
acid/base mixtures at equilibrium