CHAPTER 2

Water: The Solvent

for Biochemical
Reactions
 Chapter Outline
2-1 Water and Polarity

2-2 Hydrogen Bonds

2-3 Acids, Bases and pH

2-4 Buffers
 Biologically important elements
O 1. Organic compounds on most living:
O Carbon (C)
O Nitrogen (N)
O Hydrogen (H)
O Oxygen (O)
processes depend.
O 2. An inorganic ions , which represent only
about 0.5%
O Sodium (Na)
O Potassium (K)
O Chlorine (Cl)
O Manganese (Mg)
3
4
Chemical Evolution.
By sparking CH4, NH3, H2O and H2 these are formed:
 Glycine
 Glycolic acid
 Sarcosine
 Alanine
 Lactic acid
 N-Methalanine
 a-Amino-n-butyric acid
 a - Aminoisobutyric acid
 b- Alanine
 Succinic Acid
 Glutamic acid
 etc 5
Interactions between Biomolecules —
Chemical Bonds
OInteractions between
biomolecules depend on the
forming and breaking of
chemical bonds.

6
NON-COVALENT INTERACTIONS

7
 Water
O The medium in which biological systems
operate is water
O Water is the solvent of life
O provides a medium for movement of
molecules
O separates charged molecules, dissipates
heat, and participates in chemical reactions.

8
Body composition of Water

9
 What makes water polar

What is a polar bond:

• Electrons are unequally shared,more negative charge
found closer to one atom.
• Due to difference in electronegativity of atoms involved in
bond.
 Electronegativity
O Electronegativity: a measure of the force of an
atom’s attraction for electrons it shares in a chemical
bond with another atom
O Oxygen and Nitrogen,
more electronegative
than carbon and
hydrogen
O Fluorine is most
electronegative (4)
 Polar Bonds & Molecules
O Molecules such as CO2 have polar bonds
but, given their geometry, are nonpolar
molecules; that is, they have a zero dipole
moments
 Solvent Properties of H2O
O Ionic Bonds: Held together by positive and negative
ions
O Salt Bridge: Attraction that occurs when oppositely
charged molecules are in close proximity.
O Ion-dipole interactions: When ions in solution intreract
with molecules with dipoles, e.g., KCl dissolved in H 2O
O van der Waals Forces: bonds that do not involve
electrostatic interactions
O Dipole-dipole interactions: Forces that occur between
molecules with dipoles, one positive and one negative
O Dipole induced-dipole interactions: weak and
generally do not lead to solubility in water
Hydration Shells Surrounding Ions in
Water
Ion-dipole and Dipole-dipole Interactions

• Ion-dipole and dipole-dipole interactions help ionic and polar compounds

dissolve in water
Dipole-Induced Dipole Interaction
 Induced Dipole-Induced Dipole
Interactions or London Dispersion
Forces
Biochemically-Relevant Bond
Energies
 Solvent Properties of H2O
O Hydrophilic: water-loving
O tend to dissolve in water

O Hydrophobic: water-fearing
O tend not to dissolve in water

O Amphipathic: has characteristics of both properties

O molecules that contain one or more hydrophobic and
one or more hydrophilic regions, e.g., sodium palmitate
Hydrophobic and Hydrophilic
Substances
 Amphipathic molecules
• both polar and nonpolar character
• Interaction between nonpolar molecules is
very weak
– called van der Waals interactions
 Micelle formation by amphipathic molecules

O Micelle: a spherical arrangement of

organic molecules in water solution
clustered so that
O their hydrophobic parts are buried inside the
sphere
O their hydrophilic parts are on the surface of
the sphere and in contact with the water
environment
O formation depends on the
attraction between temporary
induced dipoles
24
 Hydrogen Bond
 The hydrogen atoms have a partially positive charge (δ+);
the oxygen atom has a partially negative charge (δ–).

 This charge distribution is due to the electronegativity

difference between hydrogen and oxygen atoms

 such partial charges result in a intermolecular force known

as a dipole-dipole force, in which the positive end of one
molecule attracts the negative end of another molecule.

25
 Hydrogen bond
 This leads to stronger-than-expected intermolecular forces.

 These unexpectedly strong intermolecular forces have a

special name: hydrogen bonds.

 The interaction of a
 hydrogen atom bonded to either oxygen, nitrogen,
 fluorine atoms with an oxygen, nitrogen or
 fluorine on another molecule (intermolecular) or the same
molecule (intramolecular).

26
 Hydrogen Bonds
O Hydrogen bond: the attractive interaction between dipoles
when:
O positive end of one dipole is a hydrogen atom bonded to an
atom of high electronegativity, most commonly O or N, and
O the negative end of the other dipole is an atom with a lone
pair of electrons, most commonly O or N
O Hydrogen bond is non-covalent
Hydrogen Binding Sites
 Hydrogen bonds

Physical properties of ice and water are a

result of intermolecular hydrogen bonding
Heat of sublimation at 0 oC is 46 kJ/mol yet
only 6 kJ/mol is gaseous kinetic energy and
the heat of fusion of ice is 6 kJ/mol which is
only 15% of the energy needed to melt ice.
Liquid water is only 15% less hydrogen
bonded than ice
CH4 boils at -164 oC but water is much higher29.
Interesting and Unique Properties of
Water
• Each water molecule can be involved in 4
hydrogen bonds: 2 as donor, and 2 as
acceptor
• Due to the tetrahedral arrangement of the
water molecule.
 Hydrogen Bonding
O Even though hydrogen bonds are weaker
than covalent bonds, they have a
significant effect on the physical properties
of hydrogen-bonded compounds
Hydrogen Bonding Between Polar
Groups and Water
Other Biologically Important Hydrogen
bonds
• Hydrogen bonding is important in
stabilization of 3-D structures of biological
molecules such as: DNA, RNA, proteins.
Hydrogen bonds

Physical properties of ice and water are a result

of intermolecular hydrogen bonding
Heat of sublimation at 0 oC is 46 kJ/mol yet
only 6 kJ/mol is gaseous kinetic energy and the
heat of fusion of ice is 6 kJ/mol which is only
15% of the energy needed to melt ice. Liquid
water is only 15% less hydrogen bonded than
ice
CH4 boils at -164 oC but water is much higher.34
Hydrogen bonds
O-H N N-H O
2.88 Å 3.04 Å

H bond donor or an H bond acceptor

N H O C

3-7 kcal/mole or 12-28 kJ/mole

very strong angle dependence

35
A hydrogen bond between two water molecules

36
 Specific heat
O is the amount of heat required to change the
temperature of a gram of water 1° Celsius.

O A high specific heat means it isn’t easy to change

the temperature of water.

37
 Acids, Bases and pH
O Acid: a molecule that behaves as a proton
donor

O Strong base: a molecule that behaves as a

proton acceptor
 Weak Acids and Bases Equilibria
•Strong acids / bases – disassociate completely
•Weak acids / bases – disassociate only partially
•Enzyme activity sensitive to pH
• weak acid/bases play important role in
protein structure/function

39
 Equilibrium
 When the concentrations of hydrogen ion (H+) and
hydroxide ion (OH–) are the same, the solution is neutral.
 If the hydrogen ion concentration exceeds
the hydroxide ion concentration, the solution is acidic.
 If the hydroxide ion concentration is greater, the solution
is basic.
 Water is a contributor to the hydrogen ion concentration
because it undergoes autoionization,

40
 Ionization of water
O The ability of water to ionize, while slight, is of central
importance for life.
O H2O <-> H+ + OH-

O Since water can act both as an acid and as a base, its

ionization may be represented as an intermolecular proton
transfer that forms a hydronium ion (H3O+) and a
hydroxide ion (OH–):

41
H2O <-> H+ + OH-

42
43
O The concentration of H2O remains almost unchanged
especially in dilute acid solutions.
O What is the concentration of H2O?

O Remember the definition: Moles per liter

O 1 mole of H2O = 18 g = 18 ml

1g  1ml
O 1000 g/liter
1000 g
 55.5M
18 g / mol 44
 The concentration of H2O
 1 mole (mol) of water weighs 18 g (Periodic Table of elements)
 1 liter (L) (1000 g) of water contains
 1000 ÷ 18 = 55.56 mol. (1gm=1ml)
 Pure water thus is 55.56 Molar.{ [H2O]=55.56M}
 Keq H2O= 1.8 x 10–16 M
 the molar concentration of H+ ions in pure water is the product
of the probability, 1.8 x 10–16, times the molar concentration of
water, 55.56 mol/L.
 the probability that a hydrogen in pure water will exist as a [H+]
= [OH-] which:
 (1.8 x 10–16 M) (55.56 M) = 1.0 X 10–14 M2
 Kw = [H+] [OH-] = 1.0 X 10–14 M2
 The result is 1.0 x 10–7 mol/L. 45
 Equilibrium expression
O Dissociation of water is described by:
K = [H+][OH-]
[H2O]
O brackets represent molar concentrations
O K is the dissociation constant
O Considering [H2O] constant yields
Kw = [H+][OH-]

46
 Kw
Kw is used to calculate the pH of acidic and basic solutions
Kw = [H+][OH-]

 Where Kw is the ionization constant of water

 For pure water ionization constant is
Kw = [H+] [OH–] = 1.0 × 10–14 (at 25°C)
 For pure water
[H+] = [OH-] = (Kw)1/2 = 10-7 M
 In the human body, where T = 37°C

Kw = 2.4 × 10–14
47
O In pure water, at 25°C, [H+] = 1.0 × 10–7 M (1.6 × 10–7 M
at 37°C)
O (M is a concentration term, the molarity. Molarity
is the number of moles of solute per liter of
solution)

48
 Acids and bases

O Since there is equal amounts of [H+] and [OH-] for pure

water (neutral)

[H+] = [OH-] = (Kw)1/2 = 10-7 M

O Acidic if [H+] > 10-7 M

O Basic if [H+] < 10-7 M

49
 pH
O pH is way of simplifying the representation of the
hydrogen ion concentration: 10-7
O the pH for any solution by using the following equation:
pH = –log [H+]
O For instance, in the case of a solution with a hydrogen ion
concentration of 1.0 × 10–7 M

pH = –log (1.0 × 10–7) = 7.0

50
[H+] pH

10-7 = 7
pH = -Log[H+]
10-3 = 3

10-2 = 2 It is easier to think in

10-10 = 10 log of concentrations
5x10-4 = 3.3 but it takes practice!!
7x10-6 = 5.15

3.3x10-8 = 7.48

51
 The pH in your
stomach is close to 1

The pH of pure water

is 7

52
Relationship between pH and [H+] / [OH-]
concentration

53
 Calculating pOH
 You can calculate pOH in a similar manner to the pH
calculation.
pH = –log [H+]
pOH = –log [OH–]
 You can calculate the hydroxide ion concentration from
the hydrogen ion concentration and the Kw (equilibrium
constant) relationship:
Kw = [H+][OH-]

[OH–] = Kw ÷ [H+]

54
 Exercise for pH and pOH
calculation
O example, if a solution has a [H+] = 6.2 × 10–6
its pH would be
pH = –log [H+]
pH = –log [6.2 × 10–6]
pH = 5.21
O pH + pOH = 14.00 (for any aqueous
solution)
O pOH= 14.00 – pH = 14.00 – 5.21 = 8.79
55
 Acids and Bases
 Many biochemicals possess functional groups that are
weak acids or bases.
 Carboxyl groups, amino groups, and phosphate esters,
whose second dissociation falls within the physiologic
range, are present in proteins and nucleic acids, most
coenzymes, and most intermediary metabolites.
 Knowledge of the dissociation of weak acids and bases
thus is basic to understanding the influence of
intracellular pH on structure and biologic activity.

56
 Acids and Bases
O the protonated species (eg, HA or R—NH3+) =the acid
O the unprotonated species (eg, A– or R—NH2) =its
conjugate base
O Weak acid Conjugate base pKa

57
 Acids and Bases
O pKa ;dissociation constants is used to express the
relative strengths of both acids and bases.

O For any weak acid, its conjugate is a strong base.

O the conjugate of a strong base is a weak acid.

O The relative strengths of bases are expressed in

terms of the pKa of their conjugate acids
58
 Acid Strength
O One can derive a numerical value for the strength of an
acid (amount of hydrogen ion released when a given
amount of acid is dissolved in water).
O Describe by Ka:

O Written correctly,
 Ionization of H2O and pH
O Lets quantitatively examine the dissociation of water:

• Molar concentration of water (55M)

• Kw is called the ion product constant for water.

• Must define a quantity to express hydrogen ion concentrations…pH

Henderson-Hasselbalch
equation
 Henderson-Hasselbalch
O Equation to connect Ka to pH of solution
containing both acid and base.
O We can calculate the ratio of
weak acid, HA, to its conjugate
base, A-, in the following way
 Henderson-Hasselbalch (Cont’d)
O Henderson-Hasselbalch equation

O From this equation, we see that

O when the concentrations of weak acid and its
conjugate base are equal, the pH of the
solution equals the pKa of the weak acid
O when pH < pKa, the weak acid predominates
O when pH > pKa, the conjugate base
predominates
 -
[A ]
pH  pK  log
[HA]

[A ] 1 10
ratio varies from 
[ HA ] 10 1

Above and below this range there is

insufficient amount of conjugate acid or base
to combine with the base or acid to prevent
the change in pH. 64
 For weak acids
HA A- + H+
This equilibrium depends on concentrations of each component
.
If [HA] = [A-] or 1/2 dissociated


[A ]
Then log  log 1  0 : pH = pK
[ HA]
By definition the pK is the pH where [HA] = [A-] :
50% dissociated 65
1: Example -the equilibrium from the
acetate

66
Example -the equilibrium from the acetate

67
Example -the equilibrium from the
acetate

68
 The buffer effect can be seen in a titration curve.
To a weak acid salt, CH3C00-, add HC1 while monitoring pH vs. the
number of equivalents of acid added.
or
do the opposite with base.

Buffer capacity: the molar amount of acid which the buffer can
handle without significant changes in pH.

i.e

1 liter of a .01 M buffer can not buffer 1 liter of a 1 M solution of

HCl
but
1 liter of a 1 M buffer can buffer 1 liter of a .01 M solution of HCl
69
70
Distribution curves for acetate and acetic acid

71
Titration curve for phosphate

72
Dissociation constants and pK’s of Acids & buffers
Acid K pK

Oxalic 5.37x10-2 1.27

H3PO4 7.08x10-3 2.15
Succinic Acid 6.17x10-5 4.21 (pK1)
Succinate 2.29x10-6 5.65 (pK2)
H2PO4- 1.51x10-7 6.82
NH4+ 5.62x10-10 9.25
Glycine 1.66x10-10 9.78
73
74
 Exercise
You will be asked to solve Henderson -
Hasselbalch type problems:
-
[A ]
pH  pK  log
[HA]

You may be asked the pH, pK, the ratio of acid or base or solve for the final
concentrations of each.

75
 The 6 steps approach

1. Write the Henderson Hasselbalch equation.

2. Write the acid base equation

3. Make sure either an H+ or OH- is in the equation.

3. Find out what you are solving for

4. Write down all given values.

5. Set up equilibrium conditions.

6. Plug in H + H equation and solve. 76

 What is the pH of a solution of that contains 0.1M
CH3C00- and 0.9 M CH3C00H? (pK=4.76)
1) pH = pK + Log [A-]
[HA]

2) CH3C00H CH3C00- + H+

3) Find pH

4) pK = 4.76 A- = 0.1 M HA = 0.9 M

5) Already at equilibrium

6) X = 4.76 + Log 0.1

0.9

Log 0.111 = -.95 X = 4.76 + (-.95) X = 3.81

77
 What would the concentration of CH3C00- be at pH
5.3 if 0.1M CH3C00H was adjusted to that pH.
1) pH = pK + Log [A-]
[HA]

2) CH3C00H CH3C00- + H+
3) Find equilibrium value of [A-] i.e [CH3C00-]
4) pH = 5.3; pK = 4.76
5) Let X = amount of CH3C00H dissociated at equilibrium

[A-] = [X]
[HA] = [0.1 - X]

6) 5.3 = 4.76 + Log [X]

[0.1 - X]
78
Now solve.
 Blood Buffering System
O Bicarbonate most significant buffer
O Formed from gaseous CO2
CO2 + H2O <-> H2CO3
H2CO3 <-> H+ + HCO3-
O Normal value blood pH 7.4
O Deviations from normal pH value lead to acidosis

79
 Physiological buffer
O In the human body, the pH of various body fluids is
important.
O The pH of blood is 7.4,
O the pH of stomach acid is 1–2, and
O the pH in the intestinal tract is 8–9.
O If the pH of blood
0.2 pH units <normal=acidosis
0.2 > in pH than normal = alkalosis
O Acidosis and alkalosis may lead to serious health problems

80
Causes of acidosis and of alkalosis

81
 The bicarbonate buffer system
O is the main extracellular buffer system
O The presence of the conjugate acid-base pair (H2CO3 and
HCO3-) means this is a buffer system
O provides a means of eliminating carbon dioxide.
O The dissolution of carbon dioxide in aqueous systems sets
up the following equilibrium:
CO2 + H2O H2CO3 + H+ + HCO3-

82
 Buffers
O buffer: a solution whose pH
resists change upon addition
of small to moderate amounts
of a strong acid or base
O consists of a weak acid
and its conjugate base
O Examples of acid-base
buffers are solutions
containing
O CH3COOH and
CH3COONa
O H2CO3 and NaHCO3
O NaH2PO4 and Na2HPO4
 How Do We Choose a Buffer?
O By understanding the relationship between pH and
buffering capacity for the ionic environment needed for
the biological samples being studied
 Buffer Range
O A buffer is effective in a range of about +/- 1 pH unit of
the pKa of the weak acid
 Naturally Occurring Buffers
O H2PO4-/HPO42- is the principal buffer in cells
O H2CO3/HCO3- is an important (but not the only) buffer
in blood

O hyperventilation can result in increased blood pH

O hypoventilation can result in decreased blood pH
 Selecting a Buffer
O The following criteria are typical
O suitable pKa
O no interference with the reaction or
detection of the assay
O suitable ionic strength
O suitable solubility
O its non-biological nature