Comprehensive Biochemistry for Dentistry Textbook for Dental Students

480 18  Metabolism of Minerals

Characteristic Features
• Lethargy (weakness)
• Nausea, vomiting, and constipation
• Polyuria and renal calculi
• Increase in cardiac contractility
• Decrease in deep tendon reflex (normal reflex is knee jerk on striking with rubber

hammer)
• Confusion (lack of clarity)

H ypocalcemia
It is a clinical condition characterized by a decrease in plasma calcium level
lower than reference value (<8.5 mg/100 ml).

Etiology
• Decrease plasma albumin level
• Hypoparathyroidism
• Renal disease
• Acute pancreatitis
• Iatrogenic (glucocorticoids)

Characteristic Features
• Convulsions
• Cardiac arrhythmia

–– Irregular heart beat either very slow or high, palpitation, angina pectoris, and
decrease in cardiac contractility

• Tetany
• It is a sudden, forceful, and involuntary contraction of muscles (muscle

spasm). The following events occur in tetany:
–– Low calcium in plasma and ECF.
–– Increase in sensitivity of voltage-gated sodium channels.
–– Low-threshold stimulus can open sodium channels across neuronal

membrane.
–– Influx of sodium ions.
–– Rapid and progressive depolarization of neurons.
–– Muscle spasm.
–– Carpopedal spasm

It is the involuntary contraction of muscles of the hand and feet.
It may be elicited by applying a blood pressure cuff over the upper arm for 3 min.
It is called as Trousseau’s sign.
–– Facial Spasm
It is the involuntary contraction of facial muscles.
It may be elicited by tapping over bones of the cheek.
It is called as Chvostek’s sign.
–– Laryngospasm
Sudden and involuntary contraction of vocal cords.
Difficulty in breathing, inhalation is difficult.

18.2 Phosphorus 481

• Hyperactive deep tendon reflex
• Irritability of muscles and nerves

18.2 Phosphorus

Phosphorus is important mineral of hard tissues of the body. It interacts with the
calcium mineral for the calcification of bones and teeth. Phosphorus has equally
important role in synthesis of nucleic acids in the body. It is essential structural
constituent of all biological membranes. It is necessary in the oxidative phosphory-
lation to produce energy.

Distribution of Phosphorus in Body
The human body contains nearly 700 g of phosphorus. About 80% of total phos-
phorus is found in association with calcium ions in hard tissues of the body.
Muscles and blood contain about 10% phosphorus which exists in bound state
with proteins (phosphoproteins), carbohydrates, and lipids (phospholipids). The
residual 10% phosphorus is disseminated in the form of organic compounds in
tissues.

Sources
Animal Sources
• Animal sources are milk, cheese, and eggs.

Plant Sources
• Plant sources are vegetables and cereals.

Recommended Dietary Allowance
Adults
• RDA for adults is 800 mg per day.

Children
• RDA for children varies between 500 mg and 1.2 g per day.

Absorption of Phosphorus
Dietary phosphorus is absorbed through intestinal mucosa (jejunum).

Factors Regulating Absorption of Phosphorus
Promoters
• Parathyroid hormone and calcitriol promote absorption of phosphorus.
• Low pH in intestinal lumen promotes absorption of phosphorus.
• Calcium: Phosphorus ratio governs absorption of phosphorus in the intestine.

Inhibitors
• Organic compound like phytic acid in diet retards absorption of phosphorus.

482 18  Metabolism of Minerals

Excretion
• Phosphorus is excreted by kidneys. Phosphate ions (80%) are reabsorbed in

proximal convolute tubules. Excretion of phosphate is regulated by dietary intake
of phosphorus. Nearly 500 mg of phosphorus is excreted in urine daily.

Biochemical Functions
• Phosphorus is essential for calcification of teeth and bones.
• It is necessary for synthesis of phospholipids, phosphoproteins, and nucleic

acids.
• Phosphorus is essential for synthesis of high-energy phosphate compounds like

ATP and GTP.
• Phosphorus is a structural component of coenzymes like NAD+, FAD, FMN, and

NADP+.
• Phosphate is a component in the phosphate buffer system.

Clinical Significance
• In renal failure, serum phosphate level is raised.
• In serum phosphate level is affected by disease of parathyroid gland. In hyper-

parathyroidism, serum phosphate level is declined, while in hypoparathyroidism,
it is elevated.
• In renal rickets, serum phosphate level is reduced.

18.3 S elenium

Selenium is a trace mineral. It is an antioxidant element in the human body. In 1957,
Schwartz and Flotz observed that selenium was helpful in the prevention of liver
cell necrosis.

Distribution of Selenium
• Selenium is extensively distributed in body tissues. In the liver and kidneys, con-

centration of selenium is the highest, while less vascular organs like the muscle
and adipose tissues contain low concentration of selenium.
• Selenium exists in association with amino acids as selenomethionine and
selenocysteine.

Dietary Sources
• Cereals are average source of selenium (0.1–0.7 μg).
• Vegetables, fruits, and milk products are poor source of selenium.

Absorption and Transport
• Dietary selenium is absorbed in the duodenum. It enters blood circulation in

selenomethionine.
• It complexes with lipoprotein in blood circulation and is distributed to

tissues.

18.4 Copper 483

• Selenomethionine is taken up by body tissues like adipose tissues, myosin, and
myoglobin.

Biochemical Functions
• Selenium (selenocysteine) is a prosthetic group in glutathione peroxidase

enzyme. This enzyme is located in mitochondria and cytosol. Glutathione per-
oxidase catalyzes reduction of H2O2 into water and oxygen. It is an important
antioxidant enzyme in cells.
• Selenium, ascorbic acid, and vitamin E are antioxidants in cells. Selenium pro-
tects tissues against oxidative damage.
• Selenium has affinity to heavy metals like cadmium, mercury, and silver.
Selenium protects tissues against toxic effects of heavy metals.

Selenium Toxicity
• Excessive intake of selenium results into clinical condition called as selenosis. It

is manifested as:

–– GIT disturbances like nausea, vomiting, and loss of weight

–– Change in behavior

–– Garlic odor from the mouth owing to formation of dimethyl selenide

18.4 C opper

Copper is a trace element. The human body contains around 100 mg of copper in
tissues.

Dietary Sources
• Copper is found in cereals, nuts, egg yolk, and green leafy vegetables.
• Milk is a poor source of copper.

Recommended Dietary Allowance
• RDA for adults is 2 mg per day.
• RDA for children is 0.5 mg per day to 1.5 mg per day.

Absorption
• Dietary copper is absorbed in duodenum.
• Metallothionein is a conjugated protein. It helps in the absorption of copper.
• Zinc, molybdenum, and phytic acid retard absorption of copper in

duodenum.

Transport in Blood Circulation
• About 95% of copper exists as ceruloplasmin in blood circulation. The remain-

ing 5% of copper is bound to albumin in circulation.
–– Ceruloplasmin

484 18  Metabolism of Minerals

Normal serum ceruloplasmin concentration is 25–50 mg/100 ml. It contains
6–8 copper atoms which are present in cuprous state and cupric state. It is a
storage protein.
• Normal serum copper concentration is 100–200 μg/100 ml.

Biochemical Functions
• Copper is a structural component of ALA synthase enzyme. It is necessary for

synthesis of hemoglobin.
• Copper is a component of superoxide dismutase, catalase, cytochrome oxidase

and other copper-dependent enzymes.
• Ceruloplasmin is a copper containing protein. It is essential for oxidation of iron

in blood circulation. Oxidized iron (Fe3+) is distributed in form of transferring in
circulation.
• Copper is necessary for myelination of nerves in the brain.

Clinical Significance
Menkes’ Disease
• Disease is characterized by impaired absorption of copper in intestinal mucosa.

Probably, copper remains chelated by metallothionein within enterocytes.
• Serum copper concentration is decreased.
• Copper deficiency leads to anemia.

Wilson’s Disease
• It is a disorder of copper metabolism. Excessive deposition of copper occurs in

the liver and brain leading to liver necrosis and brain necrosis.
• Serum copper concentration is reduced.
• Serum ceruloplasmin concentration is decreased.
• Copper is deposited in renal tubules leading to renal necrosis.

18.5 Z inc

Zinc is a trace mineral. The human body contains around 2 g of zinc.

Dietary Sources
• Vegetables, cereals, beans, and milk are good sources of zinc.

Recommended Dietary Allowance
• RDA for adults is 10 mg per day.
• RDA in pregnancy and lactation is about 15 mg per day.

Absorption
• It is absorbed in duodenal mucosa.
• Conjugated protein metallothionein helps in the absorption of zinc through intes-

tinal mucosa.
• Divalent metal ions like copper, iron, and calcium interfere in absorption of zinc.

18.6 Fluorine 485

• Peptides promote zinc absorption.

Biochemical Functions
• Zinc is a structural component of enzymes like alcohol dehydrogenase, carbonic

anhydrase, alkaline phosphatase, and carboxypeptidase.
• Zinc is essential for storage of insulin in the pancreas.
• Zinc has a role in normal reproductive function.

18.6 Fluorine

Fluorine is an electronegative element. It is widely distributed in its oxidized form
(fluoride) in nature. It is found in drinking water in varying proportions.

Dietary Sources
• Drinking water is the exclusive, easily accessible, and most probable source of

fluorides for consumption to humans.
• Alternate sources of fluorides can be salmon, sardine, and tea leaves.

Recommended Dietary Allowance
• According to WHO recommendation, safe dose of fluoride in water is between

0.5 and 1.0 mg/L.
• Lethal dose of fluoride is 2.5 g.

Absorption and Transport
• Dietary fluoride enters the stomach. It combines with HCl to form hydrogen

fluoride (HF). It is absorbed from the mucosa of the stomach in two
forms.
• At low pH, hydrofluoric acid remains in undissociated form. It is absorbed by
diffusion through the mucosa.
• At high pH (>2.5), hydrogen fluoride is ionized into hydrogen and fluoride ions.
Diffusibility of fluoride from the stomach is pH-dependent and carrier-­
independent. It is owing to the sensitivity of phospholipid layer of plasma mem-
brane hydrogen fluoride. Phospholipid layer is highly permeable to hydrogen
fluoride than fluoride ions. Therefore, HF can rapidly pass through plasma
membrane in favor of pH gradient in the body.
• Large proportion of fluorides is absorbed through intestinal mucosa. Its
absorption in the small intestine is pH-independent and carrier-dependent.

Transport
• Absorbed fluoride rapidly enters blood circulation. It exists in ionic fluoride

and organic fluorocompounds with lipids. From plasma, fluorides are distrib-
uted to hard tissues of the body.
• In bones, fluorides are deposited in bone matrix, as well as fluorides are
incorporated in hydroxyapatite crystal structure.

486 18  Metabolism of Minerals

Excretion
• Dietary fluorides are chiefly excreted by kidneys. Excretion of fluoride in

urine is again pH-dependent. Alkaline urine (owing to intake of fruits and salad)
suppresses excretion of fluorides, while acidic urine (owing to intake of protein
enriched diet) promotes urinary excretion of fluorides.
• Dietary fluorides which are not absorbed from the small intestine are excreted in
feces.
• Dietary fluorides are also secreted in saliva in trace amounts (0.01–0.03 ppm).
Salivary fluorides have biological significance as they are involved in excreting
anticariogenic effect on the teeth.

Biochemical Functions
Tooth Development
• Fluorine has a positive role in normal tooth development. Additionally, fluorine

has anticariogenic effect on teeth.
–– During tooth development stage, fluorine is incorporated into crystal structure

and forms fluoroapatite crystals. These crystals are less soluble in oral acidic
environment than hydroxyapatite crystals. Fluoroapatite crystals have higher
resistance to attack of acids than hydroxyapatite crystals. It also maintains
normal crystal structure of teeth.

Bone Development
• Fluorine has healthy effect on normal bone development.

–– In 1 PPM dose, fluorine promotes deposition of calcium and phosphates in
organic bone matrix. Fluorine helps in retention of Ca++ in the bone, thereby
preventing demineralization of skeletal tissues.

–– Fluorine retards age-dependent osteoporosis in bones.

18.6.1 F luoride Toxicity

Fluoride toxicity is a clinical condition characterized by excessive intake of
fluoride in water or food leading to a variety of clinical manifestations in the
body.

Fluoride toxicity can be grouped into two categories depending on its manifesta-
tions as:

Dental Fluorosis
Dental fluorosis is a disorder of tooth mineralization owing to intake of exces-
sive amount of fluoride in drinking water during tooth development stage.

Etiology
• High fluorine content in drinking water is the prime cause of dental fluorosis. A

dose of fluorine exceeding 1 PPM in drinking water during stage of tooth
development is detrimental to normal development of teeth.

18.6 Fluorine 487

Pathogenesis
• Ingested fluorides exhibit harmful effects on tooth bud through in situ mode.

High serum fluoride concentration alters normal tooth development via the fol-
lowing probable mechanisms:
–– Inhibitory effect on proteases in maturing enamel

In normal tooth development, ameloblasts lay down enamel organic matrix
rich in amelogenin proteins. These proteins provide nucleation for the growth
of hydroxyapatite crystals.
During enamel maturation stage, enamel matrix proteins are hydrolyzed
and removed from maturing enamel. Proteolysis of matrix proteins provides a
large space for the growth of hydroxyapatite crystals as enamel of the tooth is
95% inorganic by weight.
Enamel matrix proteins proteolytic enzymes
Matrix metalloproteinase-20 (MMP-20) also called as enamelysin.
Kallikrein 4 (KLK-4) also called as enamel matrix serine protease-1.
These enzymes catalyze hydrolysis of amelogenins along with enamel matrix
proteins.

High level of serum fluoride has an inhibitory effect in situ on proteases. As a
result, degradation of amelogenin and enamel matrix proteins does not take place. It
impairs deposition of calcium and phosphates in maturation stage of the tooth and
results into hypomineralization.

• Effect on Hydroxyapatite Crystals
In normal tooth mineralization, calcium and phosphates are mineralized into
hydroxyapatite crystals.
In high level of serum fluoride, excess fluoride ions are incorporated into crystal
structure. Fluoride ions replace hydroxyl ions from hydroxyapatite crystals to form
fluoroapatite crystals. Accordingly, high amount of hydroxyl ions are released in
extracellular matrix space of enamel which maintain alkaline pH.
High pH converts amelogenin into insoluble mass and impairs its removal from
extracellular matrix space. Alkaline pH additionally favors growth of apatite
crystals along the diameter than length which retains enamel proteins.
Overall effect is hypomineralization of tooth enamel.

Clinical Manifestations
Critical period in which toxic effects of fluoride intake are manifested is
between 1st year and 6th year of life.

Depending upon the severity of dental fluorosis, hypomineralized enamel exhib-

its structural deformities as follows:

1 . Subsurface hypomineralization
• Mineralization of surface enamel is normal. It appears translucent, glossy,
and smooth. However, subsurface enamel shows hypomineralization and
porosity which may extend to dentinoenamel junction.

488 18  Metabolism of Minerals

2. White opaque spots
• With higher level of hypomineralization, enamel shows white opaque areas
over cusps of teeth.

3. White spots with discoloration
• White opaque spots are widespread over enamel surfaces of tooth. Appearance
of brown discoloration is also manifested on tooth surfaces.

4. Enamel discoloration with attrition
• With further rise in severity of hypomineralization, enamel surface shows
brown to black discoloration. Enamel undergoes attrition over cusps and inci-
sal edges of teeth.
• There is formation of pits over enamel surfaces.

Dental fluorosis is also called as enamel mottling.

Skeletal Fluorosis
Excessive fluoride intake is manifested into skeletal fluorosis. It has the follow-
ing manifestations:

• Initial lesion is marked by bone pain, stiffness of joints, and osteosclerosis of the
pelvis and vertebral column.

• Later stages of skeletal fluorosis are marked by chronic joint pain, arthritis, and
ligament calcification.

• Fluorosis causes increase in bone density.
• Terminal stages of skeletal fluorosis are marked by limited joint movement,

deformity of the large joints and spine, wasting of muscles, and neurological
manifestations.

18.7 I ron Metabolism

Iron is the most essential trace element in the human body. It is needed in a dose
(<100 mg) per day. Total body iron represents a small fraction (0.01%) of total body
weight in healthy person. It is around 3.5 g in adult males and 2.5 g in adult females.
Iron has multiple functions in the body. Iron is necessary for gaseous transport in
pulmonary alveoli and body tissues.

Dietary Sources of Iron
Animal Sources
• Rich sources of iron are liver, meat, spleen, and eggs.
• Liver is the richest source containing around 5 mg/100 mg of iron.

Plant Sources
• The richest source is green leafy vegetables containing around 20 mg of iron per

100 mg serving of vegetables.
• Good sources are cereals, pulses, lentils, spinach, molasses, and nuts.
• Jaggery is another good source of iron.

18.7  Iron Metabolism 489

Recommended Dietary Allowance
Adults

• RDA of iron for adults is nearly 10 mg per day.

Children
• RDA of iron for children varies between 10 and 15 mg per day.

Pregnancy and lactation
• RDA of iron in pregnancy and lactation varies between 20 and 25 mg per day.

Forms of Iron in Body Tissues

Iron is present in two forms in tissues as:
• Functional Iron

Functional form of iron is essential for iron-dependent enzymatic activity, meta-

bolic functions, transport proteins, and hemopoietic functions in the body.
Functional iron is found in the following compounds:
–– Heme-conjugated proteins

Hemoglobin
Myoglobin
Catalases
Peroxidases
–– Cytochromes
–– Other enzymes
Succinate dehydrogenase
Aconitase
• Storage Iron

• Storage form of iron is the iron reservoir of the body. Iron is stored in two forms as:
–– Ferritin

Ferritin form of iron is made up of ferric form of iron which is coplexed with

apoferritin protein. Ferritin is stored in the liver, spleen, bone marrow, blood,

and intestinal mucosa.
–– Hemosiderin

It is another form of iron storage in the body. Hemosiderin contains larger

amount of iron than ferritin. In the condition of iron overload, iron is stored in

hemosiderin. It is deposited in the liver, pancreas, and spleen.

Absorption of Dietary Iron
In a routine diet, about 10–20 mg of iron is consumed with diet. However, only 2 g
(10% of intake) of iron is absorbed in circulation. About 80–90% of total iron con-
sumed is excreted in stools.

Iron in blood circulation is in dynamic equilibrium with iron store of the body, bone
marrow uptake, uptake for synthesis of iron-dependent enzymes, and iron loss from body.

Garnick postulated mucosal block theory for absorption of dietary iron.

• Dietary iron enters the stomach. Hydrochloric acid liberates ferric form of iron
from dietary non-heme iron.

490 18  Metabolism of Minerals

• Ferric form of iron is reduced into ferrous form in the gastrointestinal tract.
Ascorbic acid in diet helps in reduction of iron and forms iron-ascorbate com-
plex. This complex is highly soluble in intestinal juices. Dietary amino acids also
help in chelation of iron as iron amino acid complex. Heme iron is absorbed
without reduction in GIT.

• Iron from non-heme iron and heme iron is absorbed in ferrous form (Fe2+)
through duodenal mucosa.

• Ferrous iron is transported across the plasma membrane of enterocytes (luminal
surface of enterocytes).

• Within cytoplasm of enterocytes, ferrous iron is oxidized into ferric form.
Reaction is catalyzed by ferroxidase I enzyme. The intracytoplasmic transport of
ferric iron is carried by intracellular iron carrier. It transports ferric iron to
apoferritin within enterocytes to form ferritin. Iron is stored in the form of fer-
ritin in enterocytes.

• Ferritin is a transient iron stored in intestinal mucosa. A fraction of ferric iron
from intracellular carrier is converted into ferrous form. It is exported through
serosal surface of enterocytes. Ferroportin helps in the export of iron.

Regulation of Absorption of Iron
• Mucosal block regulates absorption of dietary iron. In case iron store of the

body is exhausted, mucosal cells enhances absorption of dietary iron. Otherwise,
in condition of adequate iron store of the body, mucosal cells limit the absorption
of iron.
• Total body iron store regulates iron absorption. Whenever body iron store is
depleted, absorption of iron is increased through intestinal mucosal cells.
• Erythropoietin secretion also regulates iron absorption. In case of iron defi-
ciency in the body, juxtaglomerular apparatus of kidneys secretes erythropoietin.
It stimulates mucosal cells to enhance absorption of iron.

Factors Promoting Absorption of Iron
• Presence of ascorbic acid in diet promotes iron absorption.
• Low pH in stomach favors iron absorption.
• Protein-rich diet favors iron absorption.
• Gastroferrin in the stomach favors iron absorption.

Factors Retarding Absorption of Iron
• Presence of phytic acid in diet retards iron absorption.
• Diet rich in phosphates retards iron absorption.
• Achlorhydria retards iron absorption.

Transport of Iron
• In blood circulation, ferrous iron is converted into ferric state. It combines with

apotransferrin to form transferrin.
• Iron is chiefly transported by transferrin from GIT to the bone marrow and other

body tissues.

18.7  Iron Metabolism 491

Excretion of Iron
• Iron is an exclusive element that is primarily stored in the body. A negligible

fraction of iron is excreted from the body in the following ways:
–– Desquamated mucosal cells of GIT containing ferritin
–– Desquamated skin cells
–– Menstrual bleeding
• Iron loss in urine is non-traceable. Appearance of blood in urine is considered a
pathological condition.
• Undigested iron is lost in feces.

18.7.1 Clinical Significance

Iron metabolism is clinically oriented toward two conditions that arise either due to
iron deficiency or overload of iron in the body. These are described as follows:

Iron Deficiency
• This condition is characterized by deficiency of iron in body stores. Iron defi-

ciency is associated with the following biochemical parameters:
–– Concentration of hemoglobin is reduced.
–– RBC count is reduced.
–– Serum ferritin level is declined.
–– Erythropoiesis is retarded.

Iron deficiency is clinically manifested as iron-deficiency anemia.

I ron Overload
Iron overload is the excessive accumulation of iron in body tissues. It is generally
caused by frequent blood transfusions and or a genetic disorder.

Iron overload can be grouped into two categories as:

Hemosiderosis
Hemosiderosis is a localized excessive accumulation of iron without any injury
to body tissues. Hemosiderin is deposited in tissues.

It is of two types as follows:

• Hemosiderosis is associated with repeated blood transfusions. Hemosiderin is
deposited in the liver. It is called as transfusional hemosiderosis. It is commonly
observed in patients suffering from thalassemia, sickle cell anemia, and leukemia.

• Hemosiderosis can be idiopathic. In this condition, hemosiderin is deposited in
lungs. It is called as idiopathic pulmonary hemosiderosis.

Hemochromatosis
Hemochromatosis is generalized excessive accumulation of iron in body tissues
accompanied by irreversible cell injury.

492 18  Metabolism of Minerals

It is of two types as follows:
Primary Hemochromatosis
• It is a hereditary disorder of iron metabolism. P. hemochromatosis is caused by
autosomal recessive genes.
• It is manifested by the following signs:
–– Cirrhosis of the liver is caused by deposition of iron in hepatocytes. Liver

cell undergo irreversible injury.
–– Diabetes mellitus is caused by accumulation of iron in islets of Langerhans

in the pancreas. Beta cells are damaged.
–– Pigmentation of the skin

Skin appears bronze colored. Skin pigmentation is associated with insulin
deficiency and collectively termed as bronze diabetes.
–– Cardiomyopathy
Iron is deposited in cardia muscle fibers. It results in hypertension, arrhyth-
mia, and valvular defects.

Secondary Hemochromatosis
S. hemochromatosis is a consequence of another disorder. It is frequently fol-
lowed by the following conditions:

• Severe hemolysis
• Frequent blood transfusion
• Beta-thalassemia major
• Parental iron therapy

Muscle Spasm
• It is the sudden, forceful, and involuntary contraction of muscles.
Muscle Cramp
• Muscle cramp persisted for prolonged period.
• Quadriceps, hamstrings, and gastrocnemius muscles in thigh, back

thigh, and calve regions may undergo cramp.
Tetany
• It is caused by hypocalcemia.
• Increased neuronal membrane permeability to sodium ions.
Tetanus
• It is a bacterial disease. It is caused by invasion of Clostridium tetani.
• Bacteria release tetanospasmin (toxin) which inhibit the release of

inhibitory neurotransmitter (glycine) from Renshaw cells in the spinal
cord.
• Rapid and progressive depolarization of motor neurons.
• Persistence of skeletal muscle contraction

Suggested Readings 493

Suggested Readings

Alberti KGMN (ed) (1978) Recent advances in clinical biochemistry. Churchill Livingstone,
London

Baron DN (1982) A short textbook of chemical pathology, 4th edn. Wiley, New York
Conn EE, Stump PK (1969) Outline of biochemistry, 2nd edn. Wiley, New Delhi
Gupta A (2017) Iron metabolism in human body. In: Nutritional anemia in preschool children.

Springer-Nature, Singapore
Harper HA (1979) Review of physiological chemistry, 17th edn. Lange Medical Publisher,

New York
Kleiner IS, Orten JM (1966) Biochemistry, 7th edn. Mosby, St Louis
Latner AL (1975) Cantarow and Trumper. Clinical biochemistry, 7th edn. Saunders, Philadelphia
Murray RK, Granner DK, Mayes PA, Rodwell VW (1999) Harper’s biochemistry. Lange Medical

Publisher, New York
Murray RK, Granner DK, Mayes PA, Rodwell VW (2003) Harper’s illustrated biochemistry, 26th

edn. Lange Medical Books, New York
Oser BL (ed) (1965) Hawk’s: physiological chemistry, 14th edn. Mc-Graw Hill, New York
Whitford GM (1994) Effects of plasma fluoride and dietary calcium concentrations on GI absorp-

tion and secretion of fluoride in the rat. Calcif Tissue Int 54:421–425

Biological Oxidation 19

19.1 Definition

Biological oxidation is an enzyme-controlled biochemical reaction in which
organic molecules of food are oxidized in tissues to release energy.

Dietary carbohydrates and lipids are major sources of energy through oxida-
tion. Carbohydrates undergo oxidation in tissues to yield energy along with car-
bon dioxide and water. Fatty acids are oxidized in tissues to yield energy as well
as formation of acetyl CoA which subsequently enter TCA cycle to generate
energy and carbon dioxide. Dietary proteins are minimally utilized for energy
production. They are involved in the synthesis and repair of tissues. However,
protein also undergoes oxidative deamination to form ammonia and α-keto acids
whose carbon skeleton is subjected to oxidation (TCA cycle) to yield energy and
produce CO2 and H2O.

19.2 M ethods of Biological Oxidation

Addition of Oxygen to Substrate
• Oxygen is added to substrate. It is catalyzed by oxidase enzyme as shown in the

following reaction:

Oxidase

2AH2 + O2 2H2O + 2A
(Addition of O2) (Oxidized)

© Springer Nature Singapore Pte Ltd. 2019 495
A. Gupta, Comprehensive Biochemistry for Dentistry,
https://doi.org/10.1007/978-981-13-1035-5_19

496 19  Biological Oxidation

Removal of Hydrogen from Substrate

• In this reaction, hydrogen atoms are removed from a substrate. Hydrogen atoms
must be accepted by a molecule, called as hydrogen acceptor. Reaction is cata-
lyzed by dehydrogenase enzyme as shown in the following reaction:

Dehydrogenase 2B + AH2
BH2 + AB (Oxidized)
(H-acceptor)

Transfer of Electrons
In living tissues, biological oxidation can be better explained in terms of trans-
fer of electrons, wherein a molecule releases electron (electron donor) and it

becomes oxidized. Other molecule accepts electron (electron acceptor) and becomes

reduced.
This type of biological oxidation is called as oxidation-reduction or redox

reaction.

Electron donor in redox reaction is called as reducing agent or reductant, while

electron acceptor is called as oxidizing agent or oxidant.

In general, a redox reaction can be written as:

Donor electron e– + Acceptor electron

Fe++ e– + Fe+++
(reduced form) (oxidized form)

Therefore, biological oxidation is characterized by simultaneous occurrence
of oxidation and reduction reaction.

In redox reactions, a substance exists into two forms:

• Oxidized form
• Reduced form

Example: (NAD+/NADH), (FMN/FMNH2)
These two forms in redox reaction are called as redox couple or conjugate
redox pair.
Oxidation-reduction reaction is dependent on two factors:

• Standard reduction potential (standard redox potential)
• Oxidoreductase enzymes

19.3  Standard Reduction Potential (Redox Potential) 497

19.3 S tandard Reduction Potential (Redox Potential)

Redox potential is a measure of the ability of a redox couple to accept or donate
electrons under standard conditions (0.1 M conc, 25 °C, pH 7.0).

Redox potential is expressed in volts or millivolts and is symbolized by (E0).

• Reduction potential of hydrogen is taken as (–0.41 V) which is the lowest as in
Table 19.1.

• Oxygen atom has a reduction potential (+0.82 volt) which is the highest.
• Reduction potentials of other substances range between (−0.42 and +0.82 volt)

as in Table 19.1.
• A substance with lower reduction potential donates electrons, and a sub-

stance with higher reduction potential accepts electrons. In ETS, electrons
move from hydrogen atom to molecular oxygen through a series of com-
plexes placed with increasing reduction potential.
• Therefore, electron donors have lower reduction potential than electron
acceptor.
• Transfer of electrons from lower reduction potential to higher reduction potential
liberates energy and is called as free energy change.
• The amount of free energy change is dependent on the difference of redox
potential.
• Free energy change marked positive indicates energy consumption, while
energy change marked negative indicates release of energy.

Table 19.1 Showing Conjugate pair Redox
redox potential of conjugate potential (V)

pairs 2H+ + 2e− → H2 −0.41

NAD+ + H+ + 2e− → NADH −0.32

FMN + 2H+ + 2e− → FMNH2 −0.30
Ubiquinone + 2H+ + 2e− → ubiquinol (CoQ.H2) +0.04
Cytochrome b (Fe+3) + e → cytochrome b (Fe+2) +0.07
Cytochrome c1 (Fe+3) + e− → cytochrome c1 (Fe+2)− +0.23
Cytochrome c (Fe+3) + e− → cytochrome c (Fe+2) +0.25
Cytochrome a (Fe+3) + e− → cytochrome a (Fe+2) +0.29
Cytochrome a3 (Fe+3) + e− → cytochrome a3 (Fe+2) +0.55
+0.82
½O2 + 2H+ + 2e− → H2O

At standard conditions (25  °C temp, pH  =  0.7,
concentration = 1 M)

498 19  Biological Oxidation

19.4 Oxidoreductase Enzymes

These enzymes catalyze electron transfer from an electron donor to electron accep-
tor molecule. Oxidoreductases belong to EC-1 as per enzyme classification system.
They are further divided into 22 subclasses. Important groups of oxidoreductases
are mentioned as follows:

Oxidases
Oxidase is a subclass of oxidoreductase enzyme that catalyzes removal of
hydrogen atoms from a substrate by utilizing molecular oxygen as an acceptor
of hydrogen.

Oxidases transfer electrons from substrate to oxygen atom (oxygen atom is
direct acceptor of electrons).

Oxidases catalyze formation of either H2O or H2O2 as in following reactions:

Oxidase

2AH2 + O2 2A + 2H2O

Oxidase

AH2 + O2 A + H2O2

Example:

• Cytochrome oxidase produces H2O in the reaction.
• Xanthine oxidase and L-amino acid oxidase produce H2O2 in the reaction.

Oxygenases
Oxygenase is a subclass of oxidoreductase enzyme that catalyzes direct addi-
tion of oxygen to substrate.

Depending upon the number of oxygen atom, these enzymes are subdivided
into monooxygenase and dioxygenase enzymes.

• Monooxygenase
Catalyzes addition of single oxygen atom to substrate in the form of hydroxyl
group, while second oxygen atom is reduced to a molecule of water
Example: dopamine β-hydroxylase (monooxygenase).

Monooxygenase

BH2 + O2 + XH2 BOH + H2O + X

• Dioxygenase

Catalyzes addition of two oxygen atoms into substrate
Example: homogentisic acid 1,2-dioxygenase

Dioxygenase

A + O2 AO2

19.4  Oxidoreductase Enzymes 499

Dehydrogenases
Dehydrogenase is a subclass of oxidoreductase enzyme that catalyzes the removal
of hydrogen atoms from a substrate and brings about oxidation of substrate.

Dehydrogenases are of two types as follows:

1 . Aerobic dehydrogenases
• These enzymes can transfer hydrogen directly to molecular oxygen.

• Aerobic dehydrogenases contain FMN or FAD as prosthetic groups which are

reduced into FMNH2 and FADH2 after accepting (2H+ + 2e−) two hydrogens.
These reduced coenzymes are reoxidized by transferring hydrogens to molec-

ular oxygen.

• There is always formation of H2O2 which in turn is decomposed by catalase
enzyme.

• Synthesis of ATP is always absent.

2 . Anaerobic dehydrogenases
• These enzymes cannot directly transfer hydrogen to molecular oxygen.

• They transfer hydrogens (electrons) to intermediate electron acceptors in

ETC which ultimately are accepted by molecular oxygen.

• There is always formation of H2O.
• Synthesis of ATP is always a feature of anaerobic dehydrogenases.

Flavin coenzymes (FMN/FAD) and nicotinamide coenzymes (NAD+/NADP+)

serve as electron acceptors.
Examples:

Flavin coenzyme-dependent dehydrogenases

Example: FMN-dependent L-amino acid oxidase, FAD-dependent D-amino
acid oxidase, and succinate dehydrogenase

Nicotinamide coenzyme-dependent dehydrogenases

Example: NAD+-dependent isocitrate dehydrogenase, malate dehydrogenase,
and NADP+-dependent glucose-6-P dehydrogenase

Cytochromes (Cyt b, Cyt c, Cyt P450) are dehydrogenase enzymes.
Cytochrome oxidase (Cyt a, a3) oxidase enzyme.
Cytochrome P450 (monooxygenase enzyme).

Hydroperoxidases
Hydroperoxidase is a subclass of oxidoreductase enzyme that utilizes H2O2as a
substrate and catalyzes oxidation of another reduced substrate.

Hydroperoxidases are of two types as follows:

1. Peroxidase
Glutathione peroxidase requires reduced glutathione for activity of enzyme.

500 19  Biological Oxidation

Peroxidase

2 G-SH + H2O2 G-s-s-G + H2O
Reduced Oxidized

Glutathione Glutathione

Glutathione reductase enzyme brings about reduction of glutathione disulfide
(oxidized) as:

G-s-s-G +NADPH+ H+ → 2 GSH+ NADP+.

2. Catalase
Catalase decomposes two molecules of H2O2 into a molecule of water and
oxygen molecule.

Catalase 2 H2O + O2
2 H2O2

19.5 Electron Transport System (ETS)

Definition
Electron transport system is defined as a series of complexes which are involved
in the transfer of electrons from a donor to an acceptor molecule coupled with
synthesis of ATP.

19.5.1 Structural Components of Electron Transport
System (ETS)

ETS is comprised of two components as follows:

1 . Electron carriers (Acceptors)
2 . Enzyme complexes in electron transport system

19.5.2 Electron Carriers (Acceptors)

• Electron carriers are complex organic molecules which transfer electrons from
one molecule to another molecule. Electron carriers act as prosthetic groups
(coenzymes) with enzymes.

• Dehydrogenase enzymes catalyze the dehydrogenation reactions in which
electron are transferred from substrates to electron carriers. Substrate mol-
ecules are oxidized and electron carriers are reduced. Subsequently, electron
carriers transfer electrons to successive molecules and are oxidized for
reuse.

19.5  Electron Transport System (ETS) 501

T ypes of Electron Acceptors
NAD+

• Nicotinamide adenine dinucleotide is a coenzyme.
• Structurally, it contains two nucleotides. Its one nucleotide has adenine as base,

ribose sugar, and phosphate group. Other nucleotide has nicotinamide as base,
ribose sugar, and phosphate group. Nicotinamide ring is found in highly reactive
group of molecules.
• NAD+ acts as electron acceptor.
• Enzyme catalyzed dehydrogenation of substrate releases two hydrogen atoms
and two electrons. NAD+ accepts one hydrogen atom and two electrons. It is
reduced into NADH. Other hydrogen atom enters in aqueous medium.
• Two electrons are added to carbon atom opposite to N+ atom in nicotinamide ring.

Flavoproteins (Fp)
FAD

• Flavin adenine dinucleotide is a coenzyme. It contains adenine, ribose sugar,
riboflavin ring, and two phosphate groups. Riboflavin ring is highly reactive
group in FAD molecule.

• FAD acts as electron acceptor.
• FAD accepts two hydrogen atoms and two electrons. It is reduced into FADH2.

FMN

• Flavin mononucleotide is a coenzyme. It contains riboflavin, ribose sugar, and
phosphate group. It acts as electron acceptor.

• It accepts two hydrogen atoms and two electrons. It is reduced into FMNH2.

Coenzyme Q (Ubiquinone)

• Coenzyme Q is a ubiquitous coenzyme in living tissues. It is composed of qui-

nine ring and isoprenoid units in side chain. The number of isoprenoid units is

variable. Generally, coenzyme Q10 has ten isoprenoid units in molecule.
• It acts as electron acceptor.

• It is not covalently bound to protein and serves as mobile electron carrier.
• Coenzyme Q can accept two hydrogen atoms and is reduced into ubiquinol.

Otherwise, acceptance of one hydrogen atom reduces coenzyme Q into
semiquinone.

Cytochromes

• Cytochromes are red- or brown-colored conjugated proteins called as hemopro-
teins (“heme”-conjugated proteins) or chromoproteins (pigmented proteins) or
metalloproteins (iron-coordinated proteins).

502 19  Biological Oxidation

• In 1925, David Keilin coined the name cytochrome.
• Cytochromes contain heme (iron-porphyrin complex) as a prosthetic

group. Heme is an iron-chelated tetrapyrrole compound. Iron atom in
cytochromes exists in ferrous state (Fe++) and ferric state (Fe+++). Its inter-
convertibility into ferrous and ferric states is helpful in the transfer of
electrons in ETS.
• Types of Cytochromes
• Depending upon the nature of “heme” and its binding to apoprotein
molecule,
• cytochromes are designated as cytochrome a, cytochrome b, and cyto-
chrome c.
• Three groups of cytochromes have been described as follows:
–– Cytochrome a

This cytochrome contains heme as prosthetic group. Heme is a complex of
iron and porphyrin. It is called as heme A.
Heme A contains copper atoms in addition to iron atoms.
Heme moiety is linked to protein through a coordinate bond between iron of
heme and amino acid residue.
Cytochrome a has further two members which are designated by writing sub-
script as cytochrome a and cytochrome a3.
Cytochrome a and cytochrome a3 are terminal component of ETS.  They
contain copper atoms and referred as cytochrome oxidase.
Heme in cytochrome a differs from protoheme in cytochrome b and cyto-
chrome c as it contains:
Formyl group that replaces a methyl group
Isoprenoid side chain that replaces vinyl side chain
–– Cytochrome b
This cytochrome contains protoheme as a prosthetic group. Protoheme is a
complex of iron and protoporphyrin IX. It is called as heme B.
Heme moiety is linked to protein through a coordinate bond between iron of
heme and amino acid residue.
–– Cytochrome c
This cytochrome also contains protoheme as a prosthetic group. Protoheme
is a complex of iron and protoporphyrin IX. It is called as heme C.
Heme moiety is linked to protein through a covalent bond between iron of
heme and amino acid residue.
Cytochrome c has further two members which are designated by writing sub-
script as cytochrome c and cytochrome c1.
Cytochrome C is a mobile electron carrier.

Cytochromes can also be classified depending upon their absorbance at a
particular wavelength of light band in reduced state as:

• Cytochrome a
Absorbance at wavelength 605 nm

19.5  Electron Transport System (ETS) 503

• Cytochrome b

Absorbance at wavelength 560 nm
• Cytochrome c

Absorbance at wavelength 550 nm
Heme A is a prosthetic group of cytochrome a and cytochrome a3.
Heme B is a prosthetic group of cytochrome b.
Heme C is a prosthetic group of cytochrome c and c1.
Heme moiety in cytochromes is attached to protein molecule by two thio-
ether linkages (covalent bonding).

Iron-Sulfur Complexes

• They are designated as Fe-S complexes. In these complexes, iron is not a
component of heme group. Therefore, Fe-S complex is also called as non-
heme iron complex. These complexes are associated as prosthetic groups in

various metalloproteins like NADH dehydrogenase, ferrodoxin, and coen-

zyme Q.

• Fe-S complexes contain chelated iron and sulfur atoms. The iron atoms are cova-

lently linked to sulfur atoms. Sulfur atoms are provided by cysteine residues of

protein and inorganic sulfides.
• Types of Fe-S Complexes

–– In Fe4S4 complex
This complex has four iron atoms, four cysteine residues, and four inor-
ganic sulfide ions. Each iron atom is linked to two sulfur atoms from cysteine

residues and two sulfur atoms from inorganic sulfides.
–– In Fe2S2 complex

This complex has two iron atoms, two sulfur atoms, and four cysteine
residues. Each iron atom is linked to two sulfur atoms from cysteine residues

and two sulfur atoms from inorganic sulfides.
–– Fe-S complex

This complex has one iron atom and four cysteine residues. Inorganic sulfide

is absent. Single iron atom is linked to four sulfur atoms of cysteine

residues.
• Ferrodoxin (Fe2S2) was the first recognized Fe-S complex. It is involved in

nitrogen fixation in plants.
• In electron transport chain, complex I and complex II contain complexes of

Fe-S.
• Fe-S complexes are involved in electron transfer in ETS. The iron atom in Fe-S

complex undergoes oxidation and reduction, hence catalyzing the transfer
of electrons.

Cytochrome c and coenzyme Q constitute mobile electron carriers in elec-
tron transport chain.

Cytochromes and Fe-S complexes carry one electron in ETS.
NADH, FAD, FMN, and coenzyme Q carry two electrons in ETS.

504 19  Biological Oxidation

19.5.3 Enzyme Complexes in Electron Transport System

• Enzyme complexes are highly organized transmembrane proteins. These
complexes are located in the inner mitochondrial membrane.

• Enzyme complexes are interlinked by electron carrier molecules.

Enzyme complexes are described as follows:
Complex I

• Complex I is NADH dehydrogenase (NADH-CoQ reductase).
• NADH dehydrogenase is associated with coenzyme FMN and Fe-S complex.
• Complex I catalyzes electron transfer from NADH to CoQ as in Fig. 19.1.

Complex II

• Complex II is succinate dehydrogenase (succinate-CoQ reductase).
• Succinate dehydrogenase is associated with coenzyme FAD and Fe-S complex.
• Complex II catalyzes electron transfer from succinate to CoQ.

Complex III

• Complex III is ubiquinol-dehydrogenase (CoQ-Cytochrome c reductase).
• This enzyme is associated with cytochrome b and cytochrome c1 as coenzymes

and iron-sulfur clusters (Fe-S) as in Fig. 19.1.
• This complex catalyzes electron transfer from CoQ.H2 to cytochrome c.

Complex IV

• Complex IV is cytochrome c oxidase.
• This enzyme is associated with cytochrome a and cytochrome a3 as coenzymes.
• This complex catalyzes electron transfer from cytochrome c to oxygen mol-

ecule as in Fig. 19.1.

19.5.4 P hysiology of Electron Transport System

Hydrogen atoms are produced in the body through oxidation and dehydrogenation
reactions. Hydrogen atoms are accepted by NAD+ and FAD which in turn furnish
hydrogen atoms to molecular oxygen through a series of complexes in ETC. This
exchange is coupled with synthesis of ATP.

The NADH2 and FADH2 are oxidized for reuse.

Sources of NADH2
1. Carbohydrate metabolism

• Oxidative decarboxylation of pyruvate into acetyl CoA-SH by pyruvate
dehydrogenase

• Oxidative decarboxylation of isocitrate by isocitrate dehydrogenase

19.5  Electron Transport System (ETS) 505
SUCCINATE
ATP

ADP + PI

NADH FMN SITE FAD
Fe-S I Fe-S

COMPLEX I COMPLEX II

COQ ADP + PI ATP

Cytb - CytC1 SITE
Fe-S II

COMPLEX III

• MOBILE Cytc
ELECTRON CARRIER ADP + PI

• CONNECTING LINK
Between Complex III &
Complex IV

ATP

Cyta, Cyta3 SITE
Cutt III

COMPLEX IV

Molecular (½O2) 2Ht
oxygen H2O

Fig. 19.1  Diagram showing Electron Transport Chain

• Oxidative decarboxylation of alpha-ketoglutarate by alpha-ketoglutarate
dehydrogenase

• Dehydrogenation of malate into oxaloacetate by malate dehydrogenase
2 . Ketogenesis

• Dehydrogenation of beta-hydroxybutyrate into acetoacetate by beta-­
hydroxybutyrate dehydrogenase

506 19  Biological Oxidation

3. Lipid metabolism
• Dehydrogenation of beta-hydroxyacyl CoA into beta-ketoacyl COA by beta-­
hydroxyacyl CoA dehydrogenase

4. Amino acid metabolism
• Oxidative deamination of L-glutamate into alpha-ketoglutarate by L-g­ lutamate
dehydrogenase

Sources of FADH2
1. Lipid metabolism

• Dehydrogenation of acyl CoA into alpha-beta unsaturated acyl COA by acyl
COA dehydrogenase

2 . Carbohydrate metabolism
• Dehydrogenation of succinate into fumarate by succinate dehydrogenase

Sequence of steps in electron transport chain is described as follows:

(Complex I)
• NADH2 furnishes two electrons and two H+ ions and it is oxidized into NAD+.

It is reutilized in dehydrogenation reactions in metabolic pathways as in
Fig. 19.1.
• Complex I (NADH-CoQ reductase) catalyzes transport of electrons from NADH
to CoQ (lipid-soluble electron carrier) in ETC through a series of reactions as
follows:
–– Two electrons are accepted by flavoproteins in complex I to form reduced

flavoprotein.

NADH + H+ + FMN FMNH2 + NAD+

–– Reduced flavoproteins transfer electrons to iron clusters in complex I which in
turn transfer electrons to CoQ.

CoQ + e_ CoQ.H (semi-quinone)

e_

CoQ.H2 (quinol)

Transfer of electrons from NADH to CoQ is associated with the release of free
energy (redox potential +0.42 V) which is sufficient to pump protons from mito-
chondrial matrix to intermembrane space.

Transfer of electron from NADH2 to CoQ synthesizes 1 ATP molecule.
It is the Site I.

19.5  Electron Transport System (ETS) 507

(Complex II)
• Succinate molecule furnishes two electrons and two hydrogen ions which

are transferred to CoQ through FAD and iron clusters in complex II as in
Fig. 19.1.

Succinate + CoQ Fumarate + CoQ.H2

Transfer of electron from succinate to CoQ is associated with reduction potential
of +0.11 V. The small energy change is unable to pump protons into intermembrane
space.

Complex II is an electron transport pathway parallel to complex 1.
However, complex II is not associated with proton pumping into intermem-
brane space. ETS through complex II generates 2 ATP than complex I which
produces 3 ATP molecules.

Electron transfer from succinate to CoQ lacks synthesis of ATP.

(Complex III)
• CoQ is a mobile electron carrier in ETC. It is lipid soluble. It acts as an acceptor

of electrons from NADH2 and succinate molecules. The CoQ.H2 transfers elec-
trons to complex III.
• In complex III, electrons are initially accepted by Cyt b which transfers electrons
to Cyt c1, and ultimately electrons are passed on to Cyt c.
• CoQ.H2 is oxidized into CoQ for its reuse.

Transfer of electrons from CoQ.H2 to Cyt c is associated with high-energy
change which is sufficient to pump four protons to intermembrane space.

CoQ.H2 + 2 Cyt c [Fe3+) CoQ + 2 Cyt c [Fe2+) + 2H+

• Cytochrome c is a mobile electron carrier.

Electron transfer from CoQ.H2 to Cyt c through Cyt b and Cyto c1 synthe-
sizes 1 ATP molecule.

It is the Site II.

(Complex IV)
• Reduced Cyt c transfers electrons to complex IV.
• In complex IV, electrons are transferred to Cyt a, which in turn transfers elec-

trons to Cu2+ and finally to Cyt a3.

508 19  Biological Oxidation

• Cyt a3 transfers electrons to molecular oxygen (terminal electron acceptor).
• Cyt a and Cyt a3 together are called cytochrome oxidase.

4 Cyt c [Fe2+) + 4H+ + O2 4 Cyt c [Fe3+) + 2H2O

Transfer of electrons from Cyt a3 to O2 is associated with high-energy change which
is sufficient to pump two protons into intermembrane space.

Electron transfer from Cyt a3 to O2 synthesizes 1 ATP molecule.
It is the Site III.

19.6 Oxidative Phosphorylation

Definition
Oxidative phosphorylation is a biochemical process in which ATP is formed by
phosphorylation of ADP-utilizing energy generated in oxidation of reduced
coenzymes (NADH/FADH2).

Oxidative phosphorylation converts free energy released during biological oxi-
dation into chemical energy in the form of ATP.

Oxidative phosphorylation is closely coupled with electron transport system in
mitochondria.

Coenzymes like NAD+ and FAD participate in biological oxidation and are
reduced into NADH and FADH2. These reduced forms of coenzymes are oxidized
through the release of electrons and protons in electron transport chain.

Electrons are transported across a series of complexes in ETC. The electrons are
transferred from one redox couple to another in increasing order of redox potential.
The redox potential difference between two redox couples is associated with release
of free energy change.

Free energy during electron flow drives protons from mitochondrial matrix into
intermembrane space through inner membrane.

Proton pump generates proton gradient across inner membrane that drives pro-
tons back into matrix, and it is coupled with phosphorylation of ADP catalyzed by
ATP synthase.

Free Energy Change in Oxidative Phosphorylation
• Redox potential of redox couple NADH/NAD+ is (Eo = −0.32 V).
• Redox potential of terminal redox couple ½ O/H2O is (Eo = +0.82 V).
• Redox potential difference between two redox couples is (Eo = 1.14 V). This

difference in redox potential is equivalent to 52.6  K  cal/mol of free energy
change.
• One ATP formation requires 7.3 K cal/mol of free energy. Oxidation of NADH
in ETC is associated with synthesis of 3 ATP molecules which requires 21.9 K cal
of free energy.

19.6  Oxidative Phosphorylation 509

Percentage of Energy Conservation
21.9 × 100
52.6

Percentage of Energy Conservation = 42%

• Oxidation of NADH helps to conserve 42% of energy in the form of 3 ATP
molecules. The efficiency of energy conservation is 42%, while 58% of the
biological energy is dissipated as heat.

Sites of Oxidative Phosphorylation
Oxidative phosphorylation occurs in the inner mitochondrial membrane at three
sites as:

Site-I

• Electron transfer from NADH2 to CoQ synthesizes 1 ATP molecule at
Site I.

Site-II

• Electron transfer from CoQ.H2 to Cyt c through Cyt b synthesizes 1 ATP
molecule at Site II.

Site-III

• Electron transfer from Cyt a3 to O2 synthesizes 1 ATP molecule at Site
III.

19.6.1 T heories of Oxidative Phosphorylation

C hemiosmotic Hypothesis
This theory was proposed by Peter Mitchell in 1961. It is a widely accepted theory
for understanding oxidative phosphorylation. The theory is also named as Mitchell’s
hypothesis.

Assumptions of Chemiosmotic Theory
1 . Electron transport

• Electrons are translocated by enzyme complexes present in ETS. It occurs at
Site I, Site II, and Site III in electron transport chain. Redox couples are
arranged in order of higher redox potential in ETC.  The flow of electrons
from a redox couple with lower redox potential to a redox couple with higher
redox potential is exergonic and liberates free energy.

510 19  Biological Oxidation

2 . Proton Pump
• Electron transport chain carries proton pumps.
• Proton pump is an intrinsic membrane protein located in biological
membrane. It serves to translocate protons across biological membrane.
–– Proton pumps in ETC are as follows:
NADH-CoQ reductase (Site I)
CoQ-cytochrome c reductase (Site II)
Cytochrome c oxidase (Site III)
Proton pump utilizes released energy associated with electron flow in
ETC. Proton pump undergoes conformational change and actively trans-
ports protons.
• Protons are translocated from mitochondrial matrix to intermembrane space
across inner mitochondrial membrane. Protons accumulate in intermembrane
space. It leads to an increase in concentration of protons toward the outer side
of inner membrane than the inner side of inner membrane. Furthermore, pro-
ton translocation produces proton gradient across the inner membrane.

3 . Proton Gradient
• Proton gradient is responsible for two conditions as:
–– Low pH on outer side of inner membrane than the inner side.
–– H igh electrical potential on the outer side of inner membrane than the inner
side.
–– T herefore, proton gradient generates electrochemical gradient across
inner membrane.

4. ATP Synthesis
• In the inner mitochondrial membrane, proton gradient or electrochemical
gradient is neutralized by backflow of electrons from intermembrane
space into mitochondrial matrix across the inner membrane.

ATP synthase (ATP phosphorylase) is also called as ATPase as it can catalyze
hydrolysis of ATP into ADP and Pi.

ATP synthase is a large-sized asymmetrical protein molecule with a shape resem-
bling mushroom. ATP synthase is a molecular motor (molecule that rotates upon
input of energy).

ATP synthase is composed of total eight subunits which are arranged into
two main fractions as:

• F1 fraction
–– It is named as Fraction 1.
–– F1 fraction is composed of alpha, beta, gamma, delta, and epsilon subunits.
–– F1 fraction is hydrophilic.
–– F1 fraction has catalytic activity.

• Fo fraction
–– It is named with subscript “o” owing to the ability of Fo fraction to bind with
oligomycin antibiotic.
–– Fo fraction is composed of a, b, and c subunits.

19.6  Oxidative Phosphorylation 511

–– Fo fraction is hydrophobic in nature.
–– Fo fraction acts a proton-translocating channel or proton pore in inner mito-

chondrial membrane.
• Protons pass through proton pore (Fo fraction) in inner membrane and induces

conformational change in F1 fraction which has catalytic activity.
• F1 fraction utilizes energy and catalyzes phosphorylation of ADP into ATP.

The overall electron chain transport reaction is:

2 H + +2 e + +1 / 2 O2 ® H2O + energy
Two hydrogen ions, two electrons, and an oxygen molecule react to form as a

product water with energy released in an exothermic reaction.

Chemical Coupling Hypothesis
Chemical coupling hypothesis was proposed by Edward Slater in 1953.

Its assumptions are as follows:

• The flow of electrons in ETC is associated with the formation of high-energy
intermediates. They are cleavaged to release energy that is utilized for phos-
phorylation of ADP.

• This hypothesis relies on chemical coupling to synthesize ATP.
• However, high-energy intermediates have not been isolated from

mitochondria.

C onformational Coupling Hypothesis
Conformational coupling hypothesis was proposed by Paul Boyer in 1964.

Its assumptions are as follows:

• Flow of electrons in electron transport chain brings about release of free energy.
• This free energy induces conformation change in membrane protein which

assumes high-energy conformational state.
• As membrane protein regains low-energy state, it promotes synthesis of ATP by

union of ADP with Pi.
• However, no concrete evidence has been put forward for high-energy con-

formation state of inner membrane.

19.6.2 Inhibitors of Electron Transport System and Oxidative
Phosphorylation

Inhibitors may be classified into two groups as follows:

Inhibitors of ETC
Inhibitor substances combine with complexes of ETC and interfere in the transfer of
electrons at different sites. These inhibitors are subclassified depending on the site
of action as follows:

512 19  Biological Oxidation

• Inhibitors of Site 1 (complex I)
–– These inhibitors prevent transfer of electrons from FMNH2 to CoQ via
complex I.
Rotenone (insecticide and fish poisoning)
Chloropromazine (anxiolytic)
Barbiturates (hypnotic).

• Inhibitors of Site II (complex III)
–– These inhibitors prevent electron transfer from reduced Cytb to Cyt c1
through complex III.
Antimycin A (antibiotic)
BAL (British anti-Lewisite)
Dimercaprol
Phenformin (hypoglycemic drug)
Napthoquinone

• Inhibitors of Site III (complex IV)
–– These inhibitors prevent electron transfer from reduced Cytcto molecu-
lar oxygen through complex IV.
Cyanide
Carbon monoxide (CO)
Hydrogen sulfide (H2S)

Inhibitors of ETC and Oxidative Phosphorylation
Inhibitor substances combine with complexes in ETC and enzymes involved in
oxidative phosphorylation. However, these inhibitors do not inhibit ETC which
are not associated with phosphorylation reactions.

• Rutamycin (inhibits ETC and ATP synthase enzyme)
• Oligomycin (inhibits ETC and ATP synthase enzyme)
• Atractyloside (toxic glycoside, herbicide)

Suggested Readings

Alberti KGMN (ed) (1978) Recent advances in clinical biochemistry. Churchill Livingstone,
London

Baron DN (1982) A short textbook of chemical pathology, 4th edn. Wiley, New York
Conn EE, Stump PK (1969) Outline of biochemistry, 2nd edn. Wiley, New Delhi
Harper HA (1979) Review of physiological chemistry, 17th edn. Lange Medical Publisher,

New York
Kleiner IS, Orten JM (1966) Biochemistry, 7th edn. Mosby, St Louis
Latner AL (1975) Cantarow and Trumper. Clinical biochemistry, 7th edn. Saunders, Philadelphia
Mazur A, Harrow B (1971) Textbook of biochemistry, 10th edn. Saunders, Philadelphia
McGilvery RW (1983) Biochemistry-a functional approach, 3rd edn. Saunders, Philadelphia
Murray RK, Granner DK, Mayes PA, Rodwell VW (1999) Harper’s biochemistry. Lange Medical

Publisher, New York
Murray RK, Granner DK, Mayes PA, Rodwell VW (2003) Harper’s illustrated biochemistry, 26th

edn. Lange Medical Books, New York

Suggested Readings 513

Oser BL (ed) (1965) Hawk’s: physiological chemistry, 14th edn. Mc-Graw Hill, New York
Rawn JD (1989) Biochemistry. Neil Patterson Publsihers, Burlington, NC
Streyer L (1975) Biochemistry, 3rd edn. Freeman WH, New York
Swaminathan M (1981) Biochemistry for medical students, 1st edn. Geetha Publishers, Mysore
Thorpe WB, Bray HG, James HP (1970) Biochemistry for medical students, 9th edn. Churchill,

London
Varley H (1969) Practical clinical biochemistry. WH Medical Books, London
Yudkin M, Offord K (1973) Comprehensive biochemistry. Longman, London

Part IV
Medical Biochemistry

Acid-Base Balance 20

20.1 Definition

Acid-base balance is a regulatory mechanism by which pH of extracellular
fluid is stringently maintained.

Intracellular fluid (ICF) has normal pH 6.7 and it fluctuates between pH 6.7 and
7.4. This high variation in pH of intracellular fluid is attributed to metabolic produc-
tion of carbon dioxide in body tissues.

Extracellular fluid (interstitial fluid and plasma) has normal pH of 7.4, and it has
very slight variation between pH 7.35 and 7.45 (7.4 ± 0.05). Despite the intake of
variety of foods per day, acid-base level of plasma and interstitial fluid is thoroughly
maintained within narrow limit of fluctuation. Human body has homeostatic mecha-
nism of constancy of pH.

A sharp change in pH of ECF leads to wide fluctuation in pH of ICF. It in turn
has high detrimental consequences on metabolic functions of cells. Plasma proteins
and enzymes are extremely sensitive to change in pH beyond the acceptable limits.
They undergo denaturation. There is malfunctioning of ion channels and receptors
across the cell membrane of cells.

20.2 Sources of Acids in Body

Routine diet especially protein-enriched food is a source of enormous amount of
acids in human body. Foods like meat, eggs, milk, cheese, alcohol, and animal
proteins liberate high quantity of acids in body tissues.

Two types of acids are produced in body: volatile acid and nonvolatile acids.

Volatile Acid
It is so called as it is excreted by lungs through exhalation. Carbonic acid is the
single volatile acid which is produced in body tissues. Carbon diaoxide is produced

© Springer Nature Singapore Pte Ltd. 2019 517
A. Gupta, Comprehensive Biochemistry for Dentistry,
https://doi.org/10.1007/978-981-13-1035-5_20

518 20  Acid-Base Balance

in tissues due to metabolic reactions of various food products. Carbon dioxide is
produced through complete oxidation of carbon chain of nutrients in carbohydrates,
lipids, and proteins as in Fig.  20.1. It becomes the major source of volatile acid
(carbonic acid) in tissues.

Carbonic acid is produced by dissolution of CO2 into H2O. It occurs in plasma
non-enzymatically and within RBC by action of carbonic anhydrase enzyme.

Metabolic activity of body tissues generates nearly 15,000–20,000 mmol of
carbon oxide. Similar quantity of carbonic acid is produced in plasma as in
Fig. 20.1.

Nonvolatile Acids (Metabolic Acids)
They are not excreted by lungs. They are fixed acids. Nonvolatile acids are further
grouped into organic and inorganic acids.

Organic and Inorganic Nonvolatile Acids
These acids are produced by incomplete oxidation of carbohydrates, lipids, and
proteins in body tissues. Important organic nonvolatile acids are pyruvate, lac-
tate, alpha-keto acids, acetoacetate, and beta-OH-butyric acid.

Important inorganic nonvolatile acids are sulfuric acid and phosphoric acid.
Sulfuric acid is produced by oxidation of sulfhydryl group of methionine and cyste-
ine amino acids. Phosphoric acid is produced by oxidation of phospholipids, phos-
phoprotein, and nucleic acids.

Daily production of nonvolatile acids is between 1 and 1.5  mmol H+/kg
weight and a total amount of 70–80 mmol H+/day as in Figs. 20.2, 20.3, 20.4,
and 20.5.

Fig. 20.1 Diagram Complete Oxidation of Nutrients
showing oxidation of Carbon Chain → CO2 + H2O

carbon chain from foods H2CO3

Fig. 20.2 Diagram Incomplete Oxidation of Carbohydrates
showing incomplete Carbohydrates → Glucose

oxidation of carbohydrates Pyruvate

Lactate

H+

20.3  Sources of Bases in Body 519

Fig. 20.3 Showing Incomplete Oxidation of Trigycerides
incomplete oxidation of Triglycerides
triglycerides

Acetoacetate β-OH-Butyric acid

Fig. 20.4 Showing H+
oxidation of lipids and Oxidation of Phospholipid, Phosphoprotein and Nucleic acid

nucleic acids H3PO4
H+

Oxidation of Sulfhydryl Group of Methionine and Cysteine Amino Acids

H2SO4

H+

Fig. 20.5  Showing oxidation of sulfur containing amino acids

20.3 Sources of Bases in Body

Routine diet delivers a negligible quantity of basic compounds in body. Fruits, veg-
etables, leguminous foods, and nuts constitute alkaline foods and provide alkaline
compounds in the body. These foods are rich in bicarbonates and potassium com-
pounds. They help to raise the pH of plasma and ECF.

Foods like raisins, spinach, bananas, and apple have potential renal acid load
(PRAL) as −20, −13, −6, and −2.5, respectively. They have the highest alkalinizing
effect on plasma and ECF.

Vegetarian food provides sodium salts of citrates and lactates. These compounds
utilize hydrogen ions. Ammonia is also generated through deamination of amino
acids. It is transported to blood circulation. It has alkalinizing effect on plasma and
ECF. However, ammonia is converted into urea.

520 20  Acid-Base Balance

20.4 A cid-Base Homeostasis

Acid-base balance is maintained by three types of regulatory mechanisms in
body. Each one has its characteristic and role in acid-base homeostasis as
shown in Fig. 20.6.

20.4.1 B uffer Systems

• Bicarbonate buffer
• Phosphate buffer
• Protein buffers

20.4.2 Role of Buffer System in Acid-Base Homeostasis

Buffer
Buffer is an aqueous solution comprised of a weak acid and its salt with a
strong base or a weak base and its salt with strong acid.
Buffer can resist a change in pH of the solution after addition of an acid or a base.

Efficacy of buffer is determined by its pKa (dissociation constant of acid).
Efficacy of a buffer is good at its pKa close to pH ± 1 of solution.

20.4.3 Types of Buffer Systems

Three buffer systems are present in body. With mild variation, their compo-
nents are the same in blood, lymph, interstitial fluid, and intracellular fluid.

Mechanism of Acid-Base balance (Homeostasis)
Acid-base homeostasis is maintained by following systems as:

Buffer Systems First-line Defence Mechanism
Respiratory System Short Term Homeostasis

Renal System Second-line Defence Mechanism
Long Term Homeostasis

Fig. 20.6  Showing systems of acid-base homeostasis

20.4  Acid-Base Homeostasis 521

Based on components of a buffer system, they are grouped into the following
categories:

1. Bicarbonate Buffer
2. Phosphate Buffer
3. Protein Buffer

B icarbonate Buffer
Composition
• Bicarbonate buffer is composed of a weak acid (carbonic acid) and its salt with

strong base (sodium bicarbonate). It is represented as:

NaHCO3 / H2CO3

• Bicarbonate/carbonic acid represents the most important buffer system of blood
and ECF.

• Sodium bicarbonate and carbonic acid are present in a fixed ratio (20:1) in
blood.

• Plasma concentration of bicarbonates is 20 times higher than plasma concentra-
tion of carbonic acid. Therefore, bicarbonates constitute alkali reserve.

Mechanism of Action
Neutralization of Nonvolatile Acids
• Bicarbonate buffer system neutralizes nonvolatile acids like lactic acids, hydro-

chloric acid, and sulfuric acid.
• After an acid enters extracellular fluid, it furnishes H+ ions. The bicarbonate buf-

fer becomes active. Sodium bicarbonate (NaHCO3) dissociates into Na+ ion and
HCO3− ion. The bicarbonate ions react with H+ ions to form carbonic acid as in
Fig. 20.7.
• Carbonic acid is a weak acid. It further dissociates into carbon dioxide and
a molecule of water. Carbon dioxide is removed by lungs through
expiration.
• Therefore, a strong nonvolatile acid is buffered by bicarbonate component
into a weak volatile acid.

Neutralization of Alkali
• After an alkali enters blood, carbonic acid dissociates into hydrogen ions and

bicarbonate ions. Hydrogen ions react with hydroxyl ions to form water as in
Fig. 20.8.

Significance
• Bicarbonate buffer is significant buffer of blood and ECF. It helps to neutralize

effectively hydrogen ions produced in plasma and ECF.

522 20  Acid-Base Balance
Fig. 20.7  Acid buffered
by bicarbonate buffer Addition of NaHCO3 | H2CO3
system Acid in Body

Fig. 20.8  Bicarbonate HA Na+ + HCO3–
Buffer System H+

Liberation of Dissociation of
H+ ions Sodium Bicorbonate

By Acid HCO3–

H+

H2CO3
Carbonic acid

Addition of NaHCO3 | H2CO3
Bicarbonate Buffer System
Alkaline Substance
[NaOH]

Na+ + OH– H+ + HCO3

Dissociation of Dissociation of
Alkaline Substance Carbonic acid

OH– + H+ H2O
Na+ + HCO3–

NaHCO3

• This buffer system generates weak and volatile carbonic acid. It rapidly dissoci-
ates into carbon dioxide which is easily removed by lungs. This is an advantage
of bicarbonate buffer system over other buffer system in body.

• Bicarbonate buffer system is an indicator of acid-base balance of body.
• Bicarbonate buffer system is directly associated with lungs.

Phosphate Buffer
• Phosphate buffer system is comprised of weak acid (monosodium phosphate)

(NaH2PO4) and its salt (disodium phosphate) (Na2HPO4). It is represented as:

20.4  Acid-Base Homeostasis 523

Na2HPO4 / NaH2PO4

• NaH2PO4 acts as weak acid and Na2HPO4 acts as salt of weak acid with strong
base.

• Disodium phosphate and monosodium phosphate are present in fixed ratio (4:1).
• Phosphate buffer system is an important buffer of intracellular fluid (ICF).
• Concentration of disodium phosphate (alkali phosphate) is four times higher

than concentration of monosodium phosphate (acid phosphate) in blood.

Mechanism of Action
Neutralization of Acid
• After an acid enters blood, it furnishes hydrogen ions.
• Disodium phosphate Na2HPO4 becomes active, and it dissociates into Na+ ion

and NaHPO4− ion as in Fig. 20.9.
• Hydrogen ions (H+) from acid are neutralized by NaHPO4− ions. There is forma-

tion of monosodium phosphate. It is excreted by kidneys.

Neutralization of Alkali
• After an alkali enters blood, phosphate buffer system becomes active. Its

monosodium phosphate component participates to neutralize addition of
alkali.
• Monosodium phosphate dissociates into H+ ions and NaHPO4− ions. Latter ions
react with OH group of alkali and neutralize it. NaHPO4− ions are converted into
disodium phosphate, and it is excreted by kidneys as in Fig. 20.10.

Fig. 20.9  Acid buffered Addition of Na2HPO4 | NaH2PO4
by phosphate buffer Phosphate Buffer
Acid in Body
[HA]

H+ Na HPO4– + Na+
Dissociation of Buffer
Acid liberates
Hydrogen ions

H+ + NaHPO4–

Na H2PO4 Excreted in
Urine

524 20  Acid-Base Balance
Fig. 20.10  Alkali
buffered by phosphate Addition of
buffer
Na2HPO4 | NaH2PO4

Alkaline Compound Phosphate Buffer

[NaOH]

Na+ + OH– Na HPO4– + H+

Dissociation of Dissociation of
alkali Phosphate Buffer

Na+ + NaHPO4–

Na2 HPO4 Excreted in
Urine

Significance
• Dissociation constant of acid (pKa) of phosphate buffer system is 6.8 and it is

close to pH of blood (7.4). Therefore, physiologically, it is a better buffer than
bicarbonate buffer.
• Phosphate buffer is present in low concentration in blood than bicarbonate buf-
fer; therefore, it is a weak buffer system than bicarbonate buffer system.
• It is directly associated with kidneys.

Protein Buffers
• Plasma Protein Buffer

• Hemoglobin Buffer

Plasma Protein Buffer

• Plasma proteins and hemoglobin constitute protein buffer system.

• Plasma protein represents a better and more effective buffer of blood.

• Plasma proteins particularly albumin represent 95% of protein buffer system of

blood and plasma.

• Buffering efficacy of protein is dependent on dissociable group in plasma pro-

teins. These groups are carboxylic group (COOH), amino group (NH2), guani-
dine group (C=NH), and imidazole group (C-NH).

• In acidic medium, amino group (NH2) accepts hydrogen ions from acidic
medium. It is converted into NH3− and acts as base. Plasma protein gets posi-
tively charged.

• In alkaline medium, carboxylic group (COOH) dissociates to release hydrogen

ions. It is converted into COO− ion and acts as acid. Plasma protein becomes

positively charged.

20.4  Acid-Base Homeostasis 525

Hemoglobin Buffer
• Hemoglobin is a conjugated protein and is an effective protein buffer.
• Buffering capability of hemoglobin is due to the presence of imidazole (C-NH)

in histidine amino acid.
• Hemoglobin has 38 histidine residues. Imidazole of histidine is the most effec-

tive buffering group of hemoglobin. Its pKa (7.3) is almost similar to pH of
plasma (7.4).
• Imidazole (C-NH) is closely linked with ferrous iron of heme. In oxygenated
state, imidazole is highly dissociable. It releases hydrogen ion and acts as
acid.
• In deoxygenated state, imidazole is least dissociable. It acts as proton acceptor
and base.

Mechanism of Hemoglobin as Buffer
At Body Tissue Level

• Cellular respiration produces carbon dioxide in tissues. Due to high pCO2 at tis-
sues level, it rapidly diffuses into blood. Inside RBCs, CO2 combines with water
to form H2CO3 under the influence of carbonic anhydrase. Carbonic acid is
unstable and dissociates into H+ ions and HCO3 ions.

• Deoxyhemoglobin accepts H+ ions (H.Hb) and acts as base. Therefore,

hemoglobin buffers hydrogen ions and prevents change in pH of blood and

ECF.

• Bicarbonates ions are transported to level of pulmonary alveoli with a least
change in pH of blood. This transport is called isohydric transport.

At Pulmonary Alveoli Level

• Due to high pO2, deoxyhemoglobin (H.Hb) releases hydrogen ions, and they
combine with bicarbonate ions to form carbonic acid. It dissociates into carbon

dioxide and water.

20.4.4 R ole of Respiratory System in Acid-Base Homeostasis

Respiratory system acts as second line of defense in maintaining pH of blood and
ECF. Pulmonary alveoli have rich blood supply and help to remove tissue-gener-
ated carbon dioxide from blood circulation and constancy of pH within a narrow
range. Hemoglobin plays an integral role in the transport of CO2 from tissues to
the lung.

Mechanism of Homeostasis
• Respiration is regulated by respiratory center located in medulla oblongata in

the brain. Respiratory center is a cluster of neurons which are sensitive to change
in pH and pCO2 of blood and ECF.  These neurons are called central
chemoreceptors.

526 20  Acid-Base Balance

• At elevated pCO2, carbon dioxide can rapidly cross blood-brain barrier. It dif-
fuses in cerebrospinal fluid and combines with water to form carbonic acid. It
splits immediately to deliver H+ ions and HCO3− ions.

• H+ ions activate respiratory center in the brain. It stimulates rate and force of
respiration (ventilation). A slight rise (0.2%) in pCO2 (1.5 mm of Hg) in blood
causes 100% increase in pulmonary respiration.

• Therefore, respiratory system helps to wash out excess of carbon dioxide from
blood and preserves pH of blood and ECF.

20.4.5 Role of Renal System in Acid-Base Homeostasis

Kidneys are vital excretory organs of human body. They serve the following vital
functions:

• Removal of nitrogenous waste substances like creatinine, urea, uric acid, xan-
thine, and hypoxanthine from the body

• Secretion of erythropoietin and renin
• Maintenance of total body water
• Maintenance of acid-base homeostasis through the following:

–– Excretion of H+ ions from blood and body fluids
–– Reabsorption of Na+ and HCO3+ ions from glomerular filtrate and preserving

alkali reserve
–– Excretion of ammonium ions
–– Excretion of monosodium dihydrogen phosphate in urine
–– To preserve acidification of urine under normal health condition

Renal system operates through following mechanisms as:

R enal Mechanism of H+ Ion Excretion (Bicarbonate Mechanism)
Kidneys are importantly involved in removal of hydrogen ions from blood and
ECF.  Regulation of hydrogen ions is necessary for maintenance of pH of
blood.

Renal Mechanism of H+ Ion Excretion
• Renal mechanism of hydrogen ion excretion occurs in proximal convoluted

tubules (PCT) of nephrons.
• PCT are surrounded by peritubular capillaries. Carbon dioxide diffuses from

capillaries into the cytoplasm of proximal convoluted tubules. CO2 combines
with water in presence of carbonic anhydrase to form carbonic acid. It splits into
hydrogen ions and bicarbonate ions within PCT as in Fig. 20.11.
• Hydrogen ions are secreted into lumen of tubules. Secretion of one H+ ion is
exchanged with reabsorption of one Na+ ions by cells of PCT.
• Hydrogen ions are excreted in urine. Therefore, kidneys are significantly engaged
in acid-base balance.

20.4  Acid-Base Homeostasis 527

Blood Proximal tubule cell

CO2 CO2 Glomerular Lumen
filtrate of
Reabsorbtion Carbonic Na+ renel
HCO3 tubule
Anti porter

H2O anhydrase

H2 CO3 H+ H+
HCO3– Na+ HCO3–

NC+ Excreted in
urine
NaHCO3
Alkali reserve

Fig. 20.11  Renal bicarbonate mechanism of H+ excretion

R eabsorption of Bicarbonate Ions
Bicarbonates are important ions of blood and ECF.  They represent alkali
reserve of body.

Renal Mechanism of Reabsorption of Bicarbonate Ions
• Bicarbonate ions are reabsorbed by proximal convoluted tubules of nephrons.
• Bicarbonate ions are filtered into glomerular filtrate which is present in lumen of

tubules. HCO3 ions combine with hydrogen ions present in lumen of renal tubule
and form carbonic acid. It dissociates into carbon dioxide and water in tubular
lumen.
• Carbon dioxide diffuses inside the cells of PCT in favor of concentration gradi-
ent. Blood around PCT also delivers carbon dioxide to PCT cells.
• Inside PCT, carbon dioxide combines with water to form carbonic acid. It splits
in presence of carbonic anhydrase to form H+ ions and HCO−3 ions as in
Fig. 20.11.
• Bicarbonate ions and Na+ ions diffuse into capillaries from cytoplasm of PCT
cells.
• Therefore, renal system helps to maintain alkali reserve of blood and ECF in
addition to preserving pH.

Renal Mechanism of Phosphate Excretion
• Renal mechanism of phosphate excretion occurs in distal convoluted tubule of

nephron.
• Plasma contains alkaline phosphate, disodium monohydrogen phosphate

(Na2HPO2), and acidic phosphate, monosodium dihydrogen phosphate

528 20  Acid-Base Balance

Blood Renal tubule cell Glomerular
CO2 filtrate
CO2 +H2O
HCO–3 Na+
Na Carbonic HPO4–
anhydrase
H2CO3 H+

HCO–3 + H+ Na+
Na+ Anti porter

Na H2 Po4
Excreted in urine

Fig. 20.12  Renal phosphate mechanism of H+ excretion

(NaH2PO4), in a ratio (4:1). This ratio of phosphates is necessary to maintain
normal acidification of urine.
• Alkaline phosphate, disodium monohydrogen phosphate (Na2HPO2), is filtered
in glomerular filtrate.
• In lumen of tubules, hydrogen ions are exchanged with sodium ions from alka-
line phosphate, disodium monohydrogen phosphate. It is converted into monoso-
dium dihydrogen phosphate which is acidic, and it is excreted in urine as in
Fig. 20.12.
• Sodium ions are reabsorbed into DCT cells and enter blood circulation.

Renal Mechanism of Ammonium Ion Excretion
Ammonia is a toxic compound produced in blood and tissues as a result of deamina-
tion of amino acids. Kidneys are involved in excretion of hydrogen ions in the form
of ammonium ions to preserve pH of blood.

Mechanism of Excretion of Ammonium Ions
• This mechanism operates in distal convoluted tubules (DCT) of nephrons.
• Glutamine is deaminated inside the DCT cells. Reaction is catalyzed by gluta-

minase enzyme. It results in formation of ammonia and glutamic acid.
• Ammonia diffuses into tubular lumen. It combines with secreted H+ ions to form

ammonium ions (NH4+ ions). Ammonium ions are impermeable to cell mem-
brane of tubules, and hence, they cannot enter cells of DCT.
• Ammonium ions are excreted in urine. Therefore, renal system helps to remove
hydrogen ions in form of ammonium ions to preserve pH of blood and ECF. It

20.5  Disorders of Acid-Base Balance 529

has been estimated that nearly 75% of acid load of body is removed by excretion
of ammonium ions in urine.
• In acidosis, generation of ammonium ions is highly increased to counteract the
excess of hydrogen ions in blood and ECF. This declares ammonium excretion
mechanism as an important method of acid-base homeostasis.

Acidification of Urine
Under normal conditions, urine is acidic with a pH (6.5). Acidification of urine owes
to tubular secretion of H+ ions by proximal convoluted tubules of kidneys. Hydrogen
ions represent acid overload of body. Kidneys are the main organs to excrete
hydrogen ions in the form of acidic urine under normal physiological conditions. It
is surprising that acidic urine is formed through glomerular filtration of components
of alkaline blood (pH 7.4). It represents important excretory function of kidneys by
which hydrogen ions are eliminated and acid-base balance is maintained.

20.5 Disorders of Acid-Base Balance

Disturbance in acid-base homeostasis is manifested in the form of acidosis and
alkalosis.

20.5.1 A cidosis

It is a clinical condition characterized by a decrease in pH of blood.

Metabolic Acidosis
It is characterized by a decline in bicarbonate concentration of blood. It represents
a primary alkali deficit.

• Bicarbonate/carbonic acid ratio is decreased (↓20:1).
• Concentration of H+ ions in blood is increased (↓pH).
• As a compensatory mechanism, respiratory center is stimulated. Ventilation is

increased for rapid removal of CO2 from blood.
• The pCO2 is lowered leading to low blood carbonic acid level. It helps to restore

blood bicarbonate level.
• ↑NH3 and ↑H+ ion excretion by kidneys
• ↑ reabsorption of bicarbonates by kidneys.

Conditions Causing Metabolic Acidosis
Diabetes mellitus, lactic acidosis, starvation, hemorrhage, end-stage renal fail-
ure, and pyelonephritis

530 20  Acid-Base Balance

Respiratory Acidosis
It is characterized by an elevation of carbonic acid level. It represents primary car-
bonic acid overload.

• A decline in removal of CO2 by pulmonary alveoli. It results in ↑pCO2 in
blood.

• Concentration of carbonic acid in blood is increased (↑ carbonic acid).
• Bicarbonate/carbonic acid ratio is decreased (↓20:1).
• Concentration of H+ ions in blood is increased (↓pH).
• As a compensatory mechanism, kidneys are primarily active in respiratory aci-

dosis. Reabsorption of bicarbonates is increased by tubules. It helps to equalize
bicarbonate to carbonic acid ratio.

Conditions Causing Respiratory Acidosis
Asthma, pulmonary edema, emphysema, drug-induced (morphine, barbitu-
rates), and brain lesions

20.5.2 Alkalosis

It is a clinical condition characterized by an increase in pH of blood.

Metabolic Alkalosis
It is characterized by an elevation of bicarbonate concentration. It represents pri-
mary alkali overload.

• It is caused by increase in bicarbonate level in blood.
• Bicarbonate/carbonic acid ratio is increased (↑20:1) with (↑pH).
• As a compensatory mechanism, respiratory center is depressed. Rate of respi-

ration becomes decreased. Ventilation is reduced. It results in retention of carbon
dioxide and ↑pCO2 in blood. It generates more amount of carbonic acid and
helps to restore bicarbonate/carbonic acid ratio of blood.
• Excretion of bicarbonates is increased by kidneys.
• Hydrogen ion excretion is reduced and K+ ion excretion is increased by
kidneys.
• Decrease in NH3 synthesis by kidneys.

Conditions Causing Metabolic Alkalosis
↑ intake of bicarbonates and hypokalemia

Respiratory Alkalosis
It is characterized by a decline in carbonic acid level. It represents primary carbonic
acid deficit.

Suggested Readings 531

• The pCO2 in blood is decreased due to hyperventilation.
• Carbonic acid level in blood is increased.

• Bicarbonate/carbonic acid ratio is increased (↑20:1) with increase in (↑pH).
• As a compensatory mechanism, kidneys are primarily involved in restoration

respiratory alkalosis. Excretion of bicarbonates is decreased. Excretion of hydro-

gen ions is increased.

Diseases Causing Respiratory Alkalosis
Meningitis, salicylates poisoning, fever, and high altitude

Suggested Readings

Alberti KGMN (ed) (1978) Recent advances in clinical biochemistry. Churchill Livingstone,
London

Baron DN (1982) A short textbook of chemical pathology, 4th edn. Wiley, New York
Conn EE, Stump PK (1969) Outline of biochemistry, 2nd edn. Wiley, New Delhi
Harper HA (1979) Review of physiological chemistry, 17th edn. Lange Medical Publisher,

New York
Kleiner IS, Orten JM (1966) Biochemistry, 7th edn. Mosby, St Louis
Latner AL (1975) Cantarow and Trumper. Clinical biochemistry, 7th edn. Saunders, Philadelphia
Mazur A, Harrow B (1971) Textbook of biochemistry, 10th edn. Saunders, Philadelphia
McGilvery RW (1983) Biochemistry-a functional approach, 3rd edn. Saunders, Philadelphia
Murray RK, Granner DK, Mayes PA, Rodwell VW (1999) Harper’s biochemistry. Lange Medical

Publisher, New York
Murray RK, Granner DK, Mayes PA, Rodwell VW (2003) Harper’s illustrated biochemistry, 26th

edn. Lange Medical Books, New York
Oser BL (ed) (1965) Hawk’s: physiological chemistry, 14th edn. Mc-Graw Hill, New York
Rawn JD (1989) Biochemistry. Neil Patterson Publishers, Burlington, NC
Streyer L (1975) Biochemistry, 3rd edn. Freeman WH, New York
Swaminathan M (1981) Biochemistry for medical students, 1st edn. Geetha Publishers, Mysore
Thorpe WB, Bray HG, James HP (1970) Biochemsitry for medical students, 9th edn. Churchil,

London
Varley H (1969) Practical clinical biochemistry. WH medical books, London
Yudkin M, Offord K (1973) Comprehensive biochemistry. Longman, London

Nutrition 21

21.1 Definition

Nutrition is defined as biological process of consumption of food in proportion
to dietary requirement of living organism.

Nutrition is essential for growth and development of human body. Nutrition
comprises intake of macronutrients like proteins, carbohydrates, and lipids and
micronutrients. Determination of quantity of macronutrients per day for an indi-
vidual is most essential. It involves sound knowledge about the calorific value of
foods and calorie requirement of an individual. Intake of sufficient amount of
micronutrients is another dimension of human nutrition. Micronutrients help to sus-
tain normal endocrine and metabolic functions.

21.2 Food

Definition of Food
Food is defined as any edible substance that is used or intended for use by
humans either raw, cooked, or partly cooked excluding drugs.

21.3 Definition of Calorific Value of Food

Calorific value of food is defined as the amount of energy produced by total
combustion of 1 g of a particular food material in the presence of oxygen. It is
expressed in terms of caloric value.

© Springer Nature Singapore Pte Ltd. 2019 533
A. Gupta, Comprehensive Biochemistry for Dentistry,
https://doi.org/10.1007/978-981-13-1035-5_21