Comprehensive Biochemistry for Dentistry Textbook for Dental Students

534 21 Nutrition

21.4 Measurement of Calorific Value

Carbohydrates, proteins, and lipids constitute important foodstuffs. Different food-
stuffs release variable amount of energy by complete combustion in an apparatus
called as bomb calorimeter.

Procedure
• Bomb calorimeter is a thick-walled steel apparatus. It contains sufficient amount

of oxygen to burn foodstuff completely.
• 1 g of foodstuff is burnt in the presence of oxygen within sealed apparatus

(bomb). Air inside the apparatus becomes hot and it escapes via a copper tube. It
is surrounded by water column. Hot air raises water temperature.
• Thermometer displays rise in temperature.

Calculation

Calorific value of food (H) = W (T2-T1)

M

W mass of water
(T2-T1) change in temperature
M mass of foodstuff

Calorie
Calorie is defined as energy required to raise the temperature of 1 g of water
by 1° C at a given pressure.

Calorie is food energy and is expressed as (Cal). It is also expressed in Kcal in

routine practice, where 1 Kcal is equal to 1000 cal. It should be noteworthy that heat
is expressed in joules.

Calorie and joule are related as shown in:

1cal = 4.2 J (SI unit of energy)

1Kcal = 1000 cal

Calorific Value of Foodstuffs
Within body tissues, dietary carbohydrates and lipids are completely oxidized to
form carbon dioxide and water and release energy. However, dietary proteins are
incompletely oxidized in body tissues. Proteins undergo deamination and trans-
amination to form ammonia and urea which are excreted. It leads to loss of energy.

Similarly, in carbohydrates and fats, some amount of energy is lost in the diges-
tion and excretion of dietary foodstuff.

21.5  Respiratory Quotient of Foods (RQ) 535

Table 21.1  Showing calorific values of foods Calorific value in calorimeter
4.3 cal/g
Dietary foodstuff Calorific value in the body 9.4 cal/g
Carbohydrates 5.4 cal/g
Lipids 4.1 cal/g
Proteins 9.3 cal/g
4.1 cal/g

Therefore, calorific value of carbohydrates and lipids and proteins differ in bomb
calorimeter and body tissues. However, difference in calorific values of carbohydrates
and lipids is minimum, and it is maximum in proteins. It is due to loss of ammonia
and urea from metabolism of proteins in body (Table 21.1).

21.5 R espiratory Quotient of Foods (RQ)

Definition
Respiratory quotient of foods is the ratio of amount of CO2 produced to amount
of O2 consumed in complete combustion of foods.

Respiratory Quotient of Carbohydrates
• RQ of carbohydrates is nearly 1. It is due to complete combustion of carbohy-

drates into carbon dioxide and water.

C6H12O6 + 6O2 6CO2 + 6H2O

RQ of carbohydrates = 6CO2/6O2 = 1 114CO2 + 110H2O

Respiratory Quotient of Lipids
• RQ of lipids is 0.7.

2 C15H110O6 + 163O2
(Tristerin)

• In the above example, the number of oxygen atoms in comparison to carbon and
hydrogen atoms per molecule of tristearin is considerably reduced. Lipids inher-
ently contain lesser number of oxygen atoms in their structure. Lipids require
more amount of oxygen in combustion.

RQ of Lipids = 114/163 = 0.7

Respiratory Quotient of Proteins
• RQ of proteins is 0.8.

536 21 Nutrition

C72H112N18O22S + 72O2 63CO2 + 38H2O + SO3 + 9CO(NH2)2
RQ of Proteins (Alanine) = 63/72= 0.8

• Proteins represent variable chemical structure. So it is difficult to ascertain num-
ber of oxygen atoms in protein molecule.

Respiratory Quotient of Mixed Diet
• RQ of mixed diet depends on relative proportion of dietary components. In routine

practice, RQ is nearly 0.85. However, it can be higher in carbohydrate-rich diet.

Significance of Respiratory Quotient
• RQ is a numerical value. It represents source of energy in human body. For

example, RQ of 0.7 shows that lipids are mainly utilized for energy production
in the body.
• RQ is a convenient way of determination of energy released by oxidation of
foodstuff.

21.6 B asal Metabolic Rate (BMR)

Calorie requirement of individuals is highly variable. It is dependent on various fac-
tors like age, gender, environmental conditions, and occupation. However, mini-
mum calorie requirement to sustain life under standard conditions is constant in all
individuals.

Definition
Basal metabolic rate is defined as the minimum quantity of energy necessary to
sustain physiological functions of the body under basal conditions in postab-
sorptive state.

Characteristics of Basal Conditions
• Individual should be in complete physical and mental.
• Individual should be in postabsorptive state (12  h without intake of solid or

liquid).
• Individual should be placed in reclining position in bed and should be awake and

alert.

In basal conditions, energy is utilized by body to perform basic physiological
functions like respiration, blood circulation, muscle tone, and brain activity. Cellular
metabolism utilizes energy.

21.6.1  Estimation of Basal Metabolic Rate

Depending on the procedure employed, basal metabolic rate can be estimated by the
following two methods:

21.6  Basal Metabolic Rate (BMR) 537

O pen-Circuit Method
This method employs measurement of oxygen consumption and production of car-

bon dioxide.

• Advantage
–– BMR can be estimated with accuracy with this method.

• Disadvantage
–– Apparatus used in this method is sophisticated and costly.
–– Procedure requires high technical efficiency and trained lab technicians.

Examples of Open-Circuit Method

1. Douglas method
2. Tissot method

Closed-Circuit Method
This method employs closed-circuit environment. The consumption of oxygen by

patient is measured for 2–6 min in basal conditions.

Examples of Closed-Circuit Method
Benedict-Roth Apparatus

• It is a cylindrical apparatus which is filled with oxygen. Patient under test is
advised to keep nostrils closed and breathe by mouth. Patient utilizes oxygen
from spirometer during inhalation, and carbon dioxide released during exhala-
tion is absorbed by lime water. Respiratory movements are transmitted to respi-
rometer and in turn are recorded on a drum. Consumption of oxygen is obtained
from recordings on drum.

Calculation of BMR
Standard heat production for 1 L of oxygen consumption = 4.825 °C
Oxygen consumed in 6 min = A
Oxygen consumed in 1 h = 10A
Standard heat production for 1 L of oxygen consumption in 1 h = 4.825 × 10A
Since BMR = cal/h/m2, therefore,

BMR = Square 4.8250 C ´ 10A area
meter of body surface


• In a state of health, BMR is constant for any individual.
• BMR variation between −15% and +20% is accepted normal.
• BMR requires 60–70% of total calorie requirement of the body.

Normal Value of BMR
In Adult Males

538 21 Nutrition

• BMR is 40 cal/h/m2.
• BMR in adult males per day is nearly 1600 cal/day.
• BMR represents 50–60% of total calorie requirement of individual per day.

In Adult Females

• BMR is 37 cal/h/m2.
• BMR per day is 1400 cal/day.
• BMR represents about 50–60% of total calorie requirement of individual per

day.

21.6.2  Factors Regulating BMR

Age
Infants and children have larger body surface area in relation to body weight. They
have higher BMR than adults. BMR at age of 6 years is 57 cal and at age of 16 years
is 50 cal. It is estimated that BMR declines nearly by 12% for every 10 years of life.

Exception is BMR of newborn babies. It is 20–25 cal/h/m2.

Gender
Males have greater mass of muscles than females. Males have nearly 5% higher
BMR than females.

Body Surface Area
Body surface area is directly proportional to BMR. Since surface area of body is
dependent on weight and height, therefore, thin and tall persons have greater surface
area and higher BMR in comparison to obese and short stature individuals.

Exercise
Regular physical exercise as in sportsmen and athletes increases lean muscle
mass and hence increases body surface area. It leads to increase in BMR (lean
muscle mass is metabolically more active and energy demanding than adipose
tissues).

Starvation
In period of starvation, dietary intake is reduced. Consequently, BMR is decreased.
Probably, it is an adaptation to starvation.

Fever
BMR is increased in fever. It is estimated that about 10% BMR rises with every
increase in 1 °C of body temperature.

Diseases
In diseased state of body, BMR is altered.

21.6  Basal Metabolic Rate (BMR) 539

Increase in BMR
In following diseases, BMR is increased as:

• Infectious diseases (sore throat, cellulitis)
• Polycythemia
• Leukemia
• Cushing syndrome
• Acromegaly
• Thyrotoxicosis
• Hypertension
• Chronic obstructive pulmonary disease

Decrease in BMR
In the following diseases, BMR is decreased:

• Myxedema
• Addison’s disease (adrenal insufficiency)
• Parkinson’s disease

In thyrotoxicosis, BMR increases 50–70% higher than normal. However, respi-
ratory quotient does not change owing to rise in oxygen consumption proportionate
to carbon dioxide production.

• In myxedema, BMR declines by 30–40% below normal.

Climate
Environmental conditions determine BMR of an individual. It increases in cold cli-
mate than hot climate.

Hormones

Hormones from thyroid gland (T3 and T4), hormones from adrenal medulla (adren-
aline), and hormones from anterior pituitary gland are responsible for rise in BMR.

Pregnancy
BMR in pregnancy increases after sixth month of pregnancy. Basal metabolic rate
of pregnant women is decided as:

• BMR of women as in nonpregnant state.
• BMR of fetus.
• Conclusively, BMR in pregnancy rises.

Racial Variations
Human race differs in physique and composition of body in different continents.
Racial variations influence BMR.  Eskimos have 33% higher BMR than normal.

540 21 Nutrition

Women belonging to Eastern countries living in the USA have 10% lesser BMR
than women of America in the same age group.

Drugs
Alcohol, caffeine, nicotine, and adrenaline elevate BMR.

21.6.3  Clinical Significance of BMR

Planning of Diet
BMR helps in estimation of calorie requirement of an individual. It in turn is neces-
sary for selection of nutrients from basic food groups and planning a balanced diet.

Research Based-Activity
Effect of drugs and nutrients on basal metabolic rate can be ascertained.

Diagnosis
BMR estimation is a useful tool in diagnosis of diseases.

21.7 Specific Dynamic Action (SDA)

Definition
Specific dynamic action is the surplus heat production in the body, which is
above the estimated calorific value, after a given quantity of food is metabo-
lized in the body.

Specific dynamic action is also termed as food-induced thermogenesis.

Specific Dynamic Action of Foods
Dietary foodstuff involves digestion, absorption, and storage of food in body tis-
sues. These cellular activities release heat which is above the heat of cellular metab-
olism during basal conditions. Therefore, basal metabolism and food processing
together produce heat in body tissues, which together constitute food-induced
thermogenesis or SDA.

SDA is dependent on the nature of nutrient in diet; hence, proteins, carbohy-
drates, and lipids have different specific dynamic action.

SDA for Carbohydrates
• Specific dynamic action for carbohydrates is 5% higher than calculated calorific

value.

SDA for Proteins
• Specific dynamic action for proteins is 30% higher than calculated calorific

value.

21.8  Balance Diet 541

SDA for Lipids
• SDA for lipids varies between 5 and 13% higher than calculated calorific value.

SDA for Mixed Diet
• SDA is between 6 and 10%.
• Carbohydrates and lipids in diet reduce SDA of proteins. In mixed diet, SDA is

lowered. It is not the sum of individual SDA of food components. Addition of
carbohydrates in protein reduces SDA by 12%. Addition of lipids in protein diet
reduces SDA by 54%.
• Addition of lipids comparatively reduces SDA more than addition of carbohy-
drates in mixed diet.

Explanation

Suppose a 20  g of proteins are oxidized in bomb calorimeter in laboratory. This

results in heat production.
Heat produced in burning of proteins in bomb calorimeter

= 20 × 4 = 80 cal (calorific value of protein 4 cal/g)
Heat produced in metabolism of 20 g of proteins in body tissues

= calculated calorific value + food induce thermogenesis (SDA)

= 80 cal + 30% higher than calculated calorific value for proteins

= 80 cal + 24 cal
Total heat production in body tissues = 104 cal
Specific dynamic action or food-induced thermogenesis is always higher
than basal metabolic rate.

Mechanism of Specific Dynamic Action
Food processing involves conversion of nondiffusible foodstuff into diffusible form,
absorption of micronutrients into circulation, and storage of food.

In case of dietary proteins, SDA is the highest among all macronutrients. In
amino acids, nitrogenous fraction undergoes oxidative deamination and carbon
skeleton enters intermediate metabolism. The body spends energy in these physio-
logical activities.

In the case of dietary carbohydrates, the body spends energy when glucose is con-
verted into lipids as well as when glucose is metabolized into carbon dioxide and water.

In the case of dietary lipids, oxidation of fats releases energy.
Therefore, food-induced physiological activities generate heat in body. It is
called SDA.

21.8 B alance Diet

Definition
Balanced diet is defined as a diet which comprises proportional amounts of
food stuffs drawn from basic food groups to fulfill energy and nutrient require-
ments of body.

542 21 Nutrition

Characteristics of Balanced Diet

• Balanced diet should contain one part protein, one part lipids, and four parts
carbohydrates (1:1:4).

• It should contain vitamin and minerals in appropriate quantity.
• It should be enriched with appropriate amount of foods selected from basic food

groups so as to provide phytonutrients, vitamins, and trace elements.
• Balanced diet should be prepared keeping in mind the physiological needs of an

individual. The quantity of its ingredients should be adjusted to fulfill increased
calorie requirement in pregnancy, lactation, childhood, and convalescent
periods.
• Balanced diet should be economical and contains local food groups.
• Balanced diet should have adequate amount of fiber foods (salad).

21.8.1  Basic Food Groups

Basic food groups are fundamental groups of foods that deliver wide range of
macro- and micronutrients. They are essential in maintaining health. Each food
group has characteristic nutrients that serve particular function. It is recommended
to select food from each basic food group so as to provide necessary calories and
minerals to match energy requirement of an individual.

21.8.2  T ypes of Basic Food Groups

In 1943, the US Department of Agriculture introduced a guide for healthy nutri-
tion. USDA promulgated seven basic food groups for general health.

1. Yellow green vegetables
2. Oranges, grapefruit, and tomatoes
3 . Fruits, other vegetables, and potatoes
4 . Milk and milk products
5 . Meat, fish, poultry, or eggs
6 . Bread, flour and cereals
7. Vitamin A fortified butter and fortified margarine

The Australian government Department of Health recommended five basic
food groups as:

1 . Grains (breads, cereals, rice, pasta, noodles, and other grains) (40% of daily diet)
2. Vegetables (30% of daily diet)
3. Fruits (10% of daily diet)
4 . Milk (milk, yoghurt, cheese, and/or alternatives) (10% of daily diet)
5 . Meat (lean meat, fish, poultry, eggs, and beans) (10% of daily diet)

21.8  Balance Diet 543

In 1992, US Department of Agriculture again recommended four basic food
groups as:

1 . Green leafy vegetables and fruits
• This food group delivers vitamins A and C. Vegetables and fruits are good
sources of dietary fibers and carbohydrates and minerals. It is recommended
to have 2–3 servings per day from this group.

2 . Milk
• Milk is an excellent source of calcium, phosphorous, lipids, and proteins.
Cheese constitutes as good casein protein. Milk should be served 2–3 times a
day in preschool children, adolescents, and adults.

3 . Meat
• Meat group represents good source of animal and plant proteins. Meat group
includes meat, fish, eggs, pulses, beans, peas, and nuts. Two servings are rec-
ommended per day.

4 . Cereals
• Whole grains as wheat, barley, maize, and rice are good source of carbohy-
drates and proteins. They are rich in minerals and vitamin B complex. Whole
grains also provided fibers. Two to three servings of cereals are recommended
per day.

Food Guide Pyramid
In 1992, the US Department of Agriculture (USDA) recommended the number of
adequate servings which should be consumed from each basic food group per day.
It was called as Food Guide Pyramid.

Food Pyramid
Food pyramid is also called as diet pyramid.

It is a diagrammatic representation indicating the type of nutrient and its
optimum servings which should be selected from each basic food group and
should be consumed per day to meet calorie and mineral requirement of an
individual.

Cereals and bread constitute the base of food pyramid with 6–11 servings per
day to be consumed depending on age, gender, and dietary needs. Further, oils, fats,
and sweets are placed at the top of food pyramid which should be used judiciously
per day as in Fig. 21.1.

MyPyramid
In 2005, the USDA Center for Nutrition Policy and Promotion introduced
updated version of nutrition guidelines. It replaced earlier Food Guide Pyramid.

MyPyramid is a diagrammatic representation in the form of colorful verti-
cal bars without images of foods.

In left side of pyramid, image of stairs and climber represent a need for physical
activity. In extreme left-hand side, orange vertical bar represents proportion of
cereals per day. Vegetables and milk groups are represented by green and blue bars,

OIL544 21 Nutrition
CARBO-
HYDRATES BREAD, CEREALS, RICE
6–11 SERVINGS
MILK
GROUP DEPENDING ON AGE,
2 SERVINGS GENDER HEALTH
MEAT
GROUPFig. 21.1  Showing food pyramid

FRUIT GROUPrespectively. Both have equal proportions. Red bar represents fruit group which is
2–4 SERVINGSsmaller than vegetables and milk groups. Another yellow narrow band represents
2–4 SERVINGSprotein group and thin silver bar gives amount of oils per day to be consumed.
MyPlate
VEGETABLEIn 2011, the USDA Center for Nutrition Policy and Promotion introduced a lat-
GROUPest version of nutrition guidelines. It replaced MyPyramid.

2–4 SERVINGSMyPlate is a diagrammatic representation in the form of a pie chart depict-
ing a plate and glass representing five food groups.

Plate is divisible into four zones indicating 30% cereals, 40% vegetables,
10% fruits, and 20% proteins to be consumed per day. Plate is shown to be asso-
ciated with a small circle representing a dairy (glass of milk) as in Fig. 21.2
(Table 21.2).

21.8.3  Types of Balanced Diet
Quantity of ingredients of balanced diet varies according to age group, gender, and
nature of work. Few examples of balanced diet have been provided from the report
of nutrition expert group, ICMR (1968) as in Tables 21.3, 21.4, 21.5 and 21.6.

21.8  Balance Diet 545
Fig. 21.2  My Plate

DAIRY

10% 20%
FRUITS PROTEINS

40%
VEGETABLES

30%
GRAINS

Table 21.2 Calorie Adult male 2400 cal
requirement per day  Sedentary work 2600 cal
 Light work 3000 cal
 Moderate work 4000 cal
 Heavy work
Adult female 1800 cal
 Sedentary work 2000 cal
 Light work 2400 cal
 Household women

Table 21.3  Balanced diet Sedentary lifestyle
recommended for adult men
Food groups Vegetarians (g) Non-v­ egetarians (g)
with sedentary lifestyle
Cereals 400 400

Pulses 70 55

Green leafy vegetables 100 100

Other vegetables 75 75

Roots and tubers 75 75

Fruits 30 30

Milk 300 100

Fats/oils 400 35

Meat/fish Nil 30

Eggs Nil 30

Sugars 30 30

Source: Report of nutrition expert group, ICMR (2007, 2010a,
2010b, 2012); Deb 2004

546 21 Nutrition

Table 21.4  Balanced diet Food groups Moderate work Non-v­ egetarians (g)
recommended for adult men Vegetarians (g) 475
with moderate work Cereals 475
Pulses 65
Green leafy 80 125
vegetables 125
Other vegetables 75
Roots and tubers 75 100
Fruits 100
Milk 30
Fats/oils 30 100
Meat/fish 300
Eggs 40
Sugars 45 50
Nil 30
Nil 40

40

Source: Report of nutrition expert group, ICMR (2007, 2010a,
2010b, 2012); Deb 2004

Table 21.5  Balanced diet Food groups Moderate work
recommended for adult Vegetarian (g) Non-vegetarian (g)
Cereals
women with moderate work Pulses 350 350
Green leafy vegetables 70 55
Other vegetables 125 125
Roots and tubers 75 75
Fruits 75 75
Milk 30 30
Fats/oils 200 100
Meat/fish 40 35
Eggs
Sugars Nil 30
Nil 40

30 30

Source: Report of nutrition expert group, ICMR (2007, 2010a,
2010b, 2012); Deb (2004)

Table 21.6  Balanced diet Food groups Age 1–3 years Age 4–6 years
recommended for preschool
Cereals 150 200
children Pulses 50 60
Green leafy vegetables 50 75
Roots and tubers 30 30
Fruits 50 50
Milk 500 400
Fats/oils 20 25
Sugars 30 40

Source: Report of nutrition expert group, ICMR (2007, 2010a,
2010b, 2012); Deb 2004

21.9  Nutritional Value of Dietary Carbohydrates 547

21.9 N utritional Value of Dietary Carbohydrates

Carbohydrates are important component of human diet. A healthy adult person
requires around 2800 cal per day. Calorific value of carbohydrates is 4 cal/g. It is
recommended that around 60% of calories should be obtained from carbohydrates.
Ideally, an individual should consume around 450 g of carbohydrates per day.

However, carbohydrates constitute major ingredient of daily diet in economi-
cally weaker section of society. Carbohydrates furnish 90% of total calorie require-
ment per day to poor population.

Carbohydrates can be categorized into two groups depending on their
nutritional value:

Digestible Carbohydrates
These carbohydrates are easily digested and metabolized in the body, for example,
glucose, starch, glycogen, fructose, and sucrose.

Nondigestible Carbohydrates
These carbohydrates are not digested in the body, for example, cellulose, inulin, and
pectin.

Chief Source of Energy
• Carbohydrates fulfill 60–70% of total calorie need of the body per day. Glucose

is an instant source of energy during hypoglycemia. They fulfill 90% of energy
requirement in poor population owing to major proportion of carbohydrates in
daily diet.

Exclusive Source of Energy
• Brain tissues and erythrocytes are exclusively dependent on glucose metabolism

for their total energy demand.

Source of Energy for Muscles
• Skeletal muscles store about 500  g of glycogen. It is an important source of

energy for skeletal muscles. It is converted into glucose which in turn is metabo-
lized to produce energy for muscular contractions.

Roughage Value
• Salads, fruits, and whole grains are rich in cellulose. It is a nondigestible carbo-

hydrate among humans owing to the absence of cellulose enzyme in alimentary
canal. Cellulose accumulates in the intestine and provides bulk to volume to the
intestine. It promotes intestinal peristalsis. Cellulose relieves constipation, and
this effect is termed as roughage effect.

Protein-Sparing Effect
• Body tissues fulfill energy demand from dietary carbohydrates and proteins that

are marginally utilized for energy production. This is protein-sparing effect.
• Dietary proteins utilized for growth and catalysis of metabolic reactions.

548 21 Nutrition

Formation of Pentose
• Dietary carbohydrates are metabolized in hexose monophosphate shunt. This

results in formation of pentoses like ribose and deoxyribose. Pentoses are con-
stituents of nucleic acids.

Role in Lipogenesis
• Dietary carbohydrates are converted into triglycerides. They are stored in adi-

pose tissues.

Role in Fat Oxidation
• Fatty acid oxidation produces acetyl CoA. It is condensed with oxaloacetate to

form citric acid in TCA cycle. Therefore, carbohydrates are essential in proper
utilization of end products of fatty acid oxidation.

21.10 N utritional Value of Dietary Proteins

Dietary proteins are building elements of living body. Proteins are primarily neces-
sary for generation and regeneration of body tissues. Proteins have higher impact on
growth than lipids and carbohydrates. 1 g of protein provides 4 cal of energy.

A healthy adult person should consume around 1 g of protein per kg of weight of
the body per day. It is around 70 g in adult males and 60 g in adult women. Its daily
requirement increases during pregnancy and lactation. Children in age group between
1 and 5 years require 30–40 g of protein daily. Old-aged persons, convalescents, and
persons suffering from chronic diseases like cirrhosis and renal failure require higher
quantity of protein. The carbohydrates are important component of human diet.

A healthy adult person requires around 2800  cal per day. It has been recom-
mended to consume 0.75 g/kg body weight of proteins for adults per day.

Dietary proteins primarily serve as a source of amino acids to the body pool of
amino acids. They are utilized for synthesis of structural proteins as well as func-
tional proteins. Dietary proteins are necessary for synthesis of hormones. Dietary
proteins help to maintain positive nitrogen balance. They are necessary for repair of
body tissues.

Nutritional value of dietary proteins can be assessed from two stand-
points as:

21.10.1  Quality of Dietary Proteins

There are many procedures that grade dietary proteins on the basis of quality.

B iological Value of Dietary Protein
Definition
Biological value is the percentage of absorbed protein from diet which becomes
part of tissue proteins.

Or

21.10  Nutritional Value of Dietary Proteins 549

Biological value can also be defined as percentage of absorbed nitrogen which
is retained in body.

It is a scale which determines what percentage of a protein is utilized by the
body. Biological value describes how rapidly the body can utilize the dietary
protein.

Biological value is considerable in the case of dietary proteins owing to lack of
storage of amino acids in the body. Biological value is of little importance in carbo-
hydrates and lipids because sufficient quantity of carbohydrates and lipids is stored
in body tissues. However, dietary proteins should have high biological value to ful-
fill daily requirement of amino acids.

Limiting amino acid in dietary protein determines biological value of protein.
For example, suppose the body requires daily 1 g of methionine. Routine diet sup-
plies 300  g of protein with 500  mg of methionine. It means dietary proteins are
deficient of 500 mg of methionine and proteins have low biological value. It can be
increased by mixing the protein with other dietary protein which has high histidine
content. In this way, biological value of mixture of proteins is higher than individual
dietary proteins.

Estimation of Biological Value (BV)
It is estimated in terms of percentage of absorbed nitrogen which is retained in tis-
sues of the body.

Procedure

• An experiment is performed on weaning albino rats.
• Rats are fed upon protein-free diet for 10 days. Amount of nitrogen is calculated

in urine and feces samples of rats.
• Later on, rats were fed upon 10% protein diet. Amount of nitrogen is estimated

in urine, feces, and diet.

Calculation (BV) = Amount of N2 retained × 100
Amount of N2 absorbed

BV = ((IN – (FN – FC) – (UN – UC)(

IN IN – (FN – FC)
FN Ingested nitrogen
FC
UN Nitrogen content in feces with protein diet
UC
Nitrogen content in feces without protein in diet

Nitrogen content in urine with protein diet

Nitrogen content in urine without protein in diet

550 21 Nutrition

Significance of Biological Value
• It is a measure of usefulness of proteins.
• It is an index of nutritive importance of proteins.
• Biological value is a useful index for vegans and vegetarians to select a good

source of dietary protein. Animal proteins, in general, have high biological value
as in Table 21.7.

Egg has the highest biological vale. It is considered to have the best amino
acid composition for humans. In plant-based proteins, soya bean has a compara-
tive composition of amino acids, but it has low biological value in comparison
to egg.

Limitation
• This method does not consider digestibility criteria of proteins.

Net Protein Utilization (NPU)
Definition
Net protein utilization is defined as a ratio between the proportion of protein
retained in body to the proportion of protein ingested.

This method is designed to assess quality of proteins. It is a better procedure. It

involves digestibility criteria of dietary proteins.

Calculation (NPU) = Amount of nitrogen retained × 100
Amount of nitrogen consumed

Significance
• It is an index of quality of dietary protein for human consumption.
• The value of net protein utilization varies from 0 to 100. A value of 100 indicates

that dietary protein is completely utilized in the body, whereas a value of “0”
indicates that ingested protein is not utilized in the body.
• Egg has NPU score of 100 and is considered as reference.

Limitation
• Net protein utilization is affected by limiting amino acids in dietary proteins.

Table 21.7 Showing Biological value Food source
biological values of food
94 Egg
sources 84 Milk
75 Beef
65 Soya bean
67 Whole cereals
60 Corn

21.10  Nutritional Value of Dietary Proteins 551

C hemical Score
Definition
Chemical score is defined as a ratio between quantity of the most limiting
essential amino acid in protein under test to quantity of similar essential amino
acid in egg protein.

This method determines chemical composition of proteins. It analyzes composi-

tion of amino acids of a given protein. It is compared with a reference composition

of amino acids, usually egg.

Chemical Score = quantity(mg) of limiting essential amino acid per gram of test protein
quantity (mg) of limiting essential amino acid per gram of egg protein ×100

Chemical score of egg protein is considered as 100. Its value is taken as reference
for comparison with same amino acid.

Egg protein contains adequate proportion of essential amino acids. Therefore,
chemical score of egg protein is used as a mark to determine chemical score of other
dietary proteins as in Table 21.8.

P rotein Efficiency Ratio
Definition
Protein efficiency ratio is defined as a ratio between the weight gain of a subject
under test and quantity of protein consumed by the subject.

PER had been a widely used method for evaluating the quality of protein in food.

Calculation

Protein efficiency ratio = quantity weight gain (g) (g )

of protein consumed

Supplementation of Dietary Proteins
• Egg is the source of all essential amino acids to humans.
• Animal proteins are better in quality than plant proteins. Wheat and rice proteins

are limiting in lysine and threonine amino acids. Therefore, intake of wheat or
rice as staple food results in deficiency of amino acids. It can be overcome by
mixing of one type of food with another food. It helps to supplement deficient
amino acids in one food with other food. Human race is consuming mixed diet.
This activity helps to improve nutritive value of proteins.

Table 21.8  Showing nutritive values of amino acids

Protein source BV NPU Chemical score PER Limiting amino acid

Egg 94 91 100 4.5 None

Milk 84 75 65 3.0 Sulfur amino acids

Meat 75 76 70 2.7 Sulfur amino acids

Wheat 67 47 42 1.5 Lysine, threonine

Soya bean 65 55 55 2.1 Sulfur amino acids

552 21 Nutrition

Example:
Rice (deficient in lysine, low in threonine) is supplemented with a preparation of
red grams.
Wheat (deficient in lysine, high in methionine) is supplemented with potatoes
(sufficient methionine, high in lysine).
Rice is supplemented with meat.

Limiting Amino Acid
• It is the most deficient essential amino acid in a dietary protein. This amino

acid becomes limiting for protein, for example, sulfur-containing amino
acid in wheat.

21.10.2  Q uantity of Dietary Proteins

Quantity of dietary proteins signifies the quantity of proteins which is to be
consumed daily to fulfill calorie need of an individual.

It may be called as recommended dietary allowance of proteins.
RDA of dietary proteins is dependent on total calorie requirement of a person,
age of a person, health and disease state of a person, and job profile of a
person.
A healthy adult individual must consume around 0.8–1 g/kg body weight of pro-
tein daily. This value amounts to nearly 560 g in adult with 70 kg of body weight.
However, amount of protein intake is raised during physiological conditions like
pregnancy and lactation. Additionally, preschool children in age group between 1
and 5 years need 30–40 g of protein daily. Aged individuals, convalescents, and
patients affected with diseases like liver cirrhosis and kidney failure need addi-
tional quantity of proteins per day. This group of individuals exhibit negative
nitrogen balance. Additional amount of proteins help to replenish nitrogen
balance.

21.11 Nutritional Value of Dietary Lipids

Lipids are important source of energy for living organisms.
It is recommended that around 20–30% of total calories should be obtained from

dietary lipids. Appropriate calculation of calories from lipids depends on total calo-
ries demand per day for the person. A healthy adult person requires around 2800 cal
per day. Calorific value of fat is (9 cal/g). Therefore, a quantity of lipids between 60
and 90 g per day is recommended.

Triglycerides constitute 80–90% of total dietary lipids consumed per day.
Triglycerides of polyunsaturated, short-chain, and medium-chain fatty acids are
digested more rapidly and easily than saturated fatty acids. Oils from plant seeds
like mustard, sunflower, soya bean, and groundnut are beneficial to saturated and
trans-fatty acids.

21.12  Protein Energy Malnutrition 553

According to American Heart Association, calories from saturated fatty acids
should not be more than 10% of total calories for healthy persons. For individuals
who are at risk of coronary artery disease, it should be less than 7% of total calories
per day. Further, proportion of trans-fatty acids in diet should be minimal to provide
<2% of total calories.

Source of Energy

• Fats are source of energy. Calorific value of 1 g of fat is 9 cal, and it is the highest

among all macronutrients. Oxidation of palmitic acid (C15H31COOH) produces
129 ATP molecules, whereas oxidation of glucose (C6H12O6) produces 38 ATP
molecules.

Transport Fat-Soluble Vitamins
• Dietary lipids help to carry fat-soluble vitamins A, D, E, and K.

Protein-Sparing Effect
• Dietary lipids are good source of energy and fulfill energy demand of body tis-

sues. They exert protein-sparing effect similar to carbohydrates.

Palatability of Food
• Palatability is a pleasure gained through eating a favorable food. Fats in diet

enhance palatable value. Fats also provide a feeling of satiety.

Source of Essential Fatty Acids
• Dietary lipids contain essential fatty acids. Linoleic acid is abundant in oils from

plant seeds. Arachidonic acid is found in animal fats. Proportion of EFA in bal-
anced diet should be 30% of total fat intake per day. Essential fatty acids have
high beneficial effects on health. They are useful in lowering low-density lipo-
protein. They are necessary for health of the liver and transport of cholesterol in
blood circulation. They are essential for growth and cognitive development in
preschool children.

21.12 Protein Energy Malnutrition

Definition
Protein energy malnutrition (PEM) is a type of malnutrition which is associ-
ated with inadequate intake of calories and/or insufficient intake of proteins in
diet.

Causes of Protein Energy Malnutrition
Primary Causes
• Inadequate intake of calories
• Intake of poor quality and inadequate quantity of proteins

554 21 Nutrition

Secondary Causes
• Malabsorption syndrome
• Celiac disease
• Environmental enteropathy

Types of Protein Energy Malnutrition
PEM is categorized into three types as:
• Kwashiorkor
• Marasmus
• Marasmic-kwashiorkor

21.12.1  Kwashiorkor

Definition
Kwashiorkor is a type of severe protein deficiency disorder.

Etiology
• Kwashiorkor is caused by severe inadequacy of proteins in diet. Protein-­

deficient diet supplies adequate calories to children for survival but impairs phys-
iological functions.

Predisposing Factors
• Natural calamity leading to food crisis
• Poverty
• Improper weaning
• Persistent diarrhea
• Acute respiratory infections
• Poor sanitation and hygiene
• Geophagia

Age Predilection
• It is commonly seen in children in 2–3 years age group.

Clinical Manifestations
• Retarded growth of children.
• Pitting edema of ankles and feet is the characteristic sign of kwashiorkor.
• Distension of the abdomen.
• Hairs dry and sparse. Brown discoloration of hairs.
• Dermatitis and loss of skin pigmentation.
• Anorexia (loss of appetite).
• Liver enlargement (fatty liver).
• Muscle wasting.
• Physical fatigue.
• Anemia.

21.12  Protein Energy Malnutrition 555

Prognosis
• Malnutrition (weight to height <−3SD) in kwashiorkor is associated with poor

prognosis despite the start of proper treatment.

21.12.2  M arasmus

Definition
Marasmus is primarily an energy (calorie) deficiency disorder.

Etiology
• Inadequate intake of diet (including protein, carbohydrates, lipids, and

minerals) for prolonged period

Predisposing Factors
• Natural calamity leading to food crisis
• Poverty
• Improper weaning
• Persistent diarrhea
• Acute respiratory infections
• Poor sanitation and hygiene
• Geophagia
• Helminthic infestations
• Preterm infants
• Prolonged breast feeding

Age Predilection
• Marasmus is observed in infants (<1 year).

Clinical Manifestations
• Retarded growth, pronounced emaciation, and underweight are common

features.
• Persistent diarrhea is another important sign of marasmus.
• Edema is absent.
• The skin becomes flaccid, wrinkled, and pale in color.
• Hairs are dry and thin and have loss of luster.
• Acute dehydration.
• Infant is highly irritable and restless.
• Anemia.

Prognosis
• Marasmus has good prognosis after the start of proper treatment.

556 21 Nutrition

Suggested Readings

Deb AC (2004) Fundamentals of biochemistry, 8th edn. New Central Book Agency, Kolkata
Food Act (2014) New Zealand Legislation, no. 32, 2014, New Zealand. http://www.legislation.

govt.nz/act/public/2014/0032/latest/whole.html#DLM2996074
Gopalan C, Ramasastri BV (1990) Nutritive value of Indian food. National Institute of Nutrition,

ICMR, Hyderabad
Gupta A (2015) Effect of geophagy on the nutritional status of children under five years of age.

Ind Stream Res J 4(12):5859
Gupta A (2017) Assessing stunting and predisposing factors among children. Asian J Pharm Clin

Res 10(10):1–8
ICMR (2007) Diet and diabetes. NIN, ICMR, Hyderabad
ICMR (2010a) Nutrient requirements and recommended dietary allowances for Indians. NIN,

ICMR, Hyderabad
ICMR (2010b) Nutritive value of Indian foods. NIN, ICMR, Hyderabad
ICMR (2011) Dietary guidelines for Indians – a manual. NIN, ICMR, Hyderabad
Khanna K et al (2003) Textbook of nutrition and dietetics. Phoenix Publishing House Pvt. Ltd.,

New Delhi
Passmore R (1986) Eastwood. Human nutrition and dietetics, 8th edn. Churchill Livingstone,

London
Planning Commission (2005) 10th plan (2002–2007) volume II, nutrition. Planning Commission,

GOI, New Delhi
Robinson CH, Lawler MN (1986) Normal and therapeutic nutrition, 17th edn. Macmillan

Publishing Company, New York
Swaminathan M (1990) Essentials of food and nutrition, vol vol 1/vol 2. Bangalore Printing and

Publishing Co Ltd., Bengaluru
Wadhwa A, Sharma S (2003) Nutrition in the community. Elite Publishing House Pvt. Ltd., Darya

Ganj

Serum Enzymes and Organ Function 22
Tests

22.1 Definition

Clinical enzymology is the branch of medical science which deals with study of
enzyme activity for diagnosis and prognosis of diseases.

In 1956, Wroblewski and his colleagues published their work on SGOT and
LDH. It was a new venture in medical science that paved a way to emergence of
clinical enzymology.

22.2 Types of Plasma Enzymes

Plasma enzymes are two types depending on their source and function.

Functional Plasma Enzymes
• Concentration of these enzymes is very high in plasma.
• These enzymes are synthesized by the liver.
• These enzymes have high catalytic activity in plasma.
• They may be called as plasma-derived enzymes.
• Examples are fibrinogen, prothrombin, and lipoprotein lipase.

Nonfunctional Plasma Enzymes
• The concentration of these enzymes has very low plasma.
• These enzymes are synthesized by rough endoplasmic reticulum in cells.
• These enzymes have no specific function in plasma.
• They may be called as cell-derived enzymes.
• Examples are SGPT, SGOT, and creatine kinase.

© Springer Nature Singapore Pte Ltd. 2019 557
A. Gupta, Comprehensive Biochemistry for Dentistry,
https://doi.org/10.1007/978-981-13-1035-5_22

558 22  Serum Enzymes and Organ Function Tests

Nonfunctional enzymes are continuously discharged into plasma by the follow-
ing activities:

1 . Normal diffusion through membrane of cells
2 . Normal cell death

There exists a balance between rate of influx of an enzyme in plasma and its
catabolism and clearance from plasma.

A disease causes imbalance by either increasing release or decreasing clear-
ance of nonfunctional enzyme from plasma.

Clinical enzymology is based on the estimation and interpretation of plasma
enzymes in disease.

22.3 S ignificance of Clinical Enzymology

Diagnosis of Disease
• Estimation of plasma enzymes is a noninvasive method. It helps in the detection

of organ dysfunction and diagnosis of disease.

Pattern of Disease
• Plasma enzyme estimation helps to understand the pattern of a disease. For

example, SGPT and SGOT estimations aid in diagnosis of acute viral hepatitis,
alcoholic cirrhosis, fatty liver, and obstructive liver disease.

Prediction of Severity of Disease
• Plasma enzyme is helpful in assessing the severity of a disease. For example,

rising value of SGPT helps to predict severity of acute viral hepatitis.

Prognosis of Disease
• Plasma enzyme estimation helps to assess the prognosis of a disease. Their study

predicts the morbidity and mortality.

Differential Diagnosis
• The study of plasma enzymes helps in differential diagnosis of diseases with

similar clinical manifestations. For example, pulmonary embolism can exhibit
the same clinical manifestation as acute myocardial infarction. Assessment of
LDH, SGOT, and CK is helpful in differential diagnosis.

Interpretation Unit of Serum Enzymes (Plasma Enzymes)
Plasma enzyme activity is expressed in international unit.

It is defined as enzyme activity that converts 1 μmol of substrate per minute
per liter under optimum conditions.

22.4  Serum Enzymes in Heart Diseases 559

22.4 Serum Enzymes in Heart Diseases

22.4.1 C reatine Kinase (CK or CPK)

Creatine kinase is an important enzyme that catalyzes phosphorylation of cre-
atine into creatine phosphate (phosphocreatine, a mobile source of energy in
skeletal muscles and brain tissues which provides high energy phosphate to
ADP to form ATP).

Creatine
• Nitrogenous compound is formed from glycine and arginine in the liver and kid-

neys and released into blood circulation and enters skeletal muscles (95%) and
brain tissues (small fraction).

Occurrence
It is present in high amount in cardiac muscle fibers, skeletal muscle fibers, brain
tissues, and the retina. It is absent in liver cells, erythrocytes, and the kidneys.

Normal Value
Adult males: 10–100 IU/L
Adult females: 10–80 IU/L

Creatine
• It is a nitrogenous compound. It is synthesized from glycine and arginine amino

acids in the liver and kidneys. Creatine enters circulation and transported to the
skeletal muscles.
• It is phosphorylated into phosphocreatine by creatine kinase in the presence of
ATP. Phosphocreatine is energy currency for the muscles and brain. It delivers
high-energy phosphate to ADP to form ATP.

Creatinine
• Creatinine is formed by a nonenzymatic breakdown of phosphocreatine in skel-

etal muscles. It is produced at uniform rate. It is not reabsorbed and excreted
freely by kidneys. Serum creatinine level is an indicator of renal function and is
inversely related to renal function.

Interpretation in Heart Disease
In Acute Myocardial Infarction (AMI)
• In acute myocardial infarction, cardiac muscle fibers are damaged. The enzyme is

released in plasma in large amount. Its detection is helpful in the diagnosis of AMI.
• Value of creatine kinase is elevated in acute myocardial infarction. Its value tends to

rise within 3–6 h, and it attains a maximum value in 24 h. Its value normalizes in 72 h.

560 22  Serum Enzymes and Organ Function Tests

Isozyme of creatine kinase (CK)
Creatine kinase is a dimer and its subunits are called as B (brain) and M (muscles).
On the basis of two subunits, creatine kinase has three isoenzymes:

• CK-MM (CK-3)
It is present in high concentration in skeletal muscle fibers (98%). Skeletal
muscles have low concentration (1%) of CK-MB. Its serum concentration is
80%.

• CK-MB (CK-2)
Cardiac fibers have (30%) concentration of CK-MB. Its serum concentration is
5%. Its value rises in serum after onset of AMI owing to its release from dam-
aged cardiac fibers.

• CK-BB (CK-1)
It is present in high concentration in the brain and smooth muscle fibers. Its
serum concentration is 1%.

Clinical Significance in AMI
• Serum creatine kinase is helpful to detect early AMI cases when changes in ECG

do not correlate with onset of AMI.
• Creatine kinase is not able to diagnose congestive cardiac failure and coronary

ischemia.
• Creatine kinase is not increased in hemolysis as in lactate dehydrogenase. Its

estimation is more advantageous in comparison to LDH.

22.4.2 Serum Glutamate Oxaloacetate Transaminase (SGOT)

It is also called as aspartate transaminase (AST). It catalyzes reversible transamina-
tion in between aspartate and glutamate.

Occurrence
This enzyme is present in high concentration in liver cells, cardiac muscle fibers, the
kidneys, and the brain.

Normal Value
In adult males: 5–40 IU/L
In adult female: 5–35 IU/L

Interpretation in Heart Disease
In Acute Myocardial Infarction

Its value increases in 12 h and attains peak value in within 24 h. It normalizes
after 3–5 days.

22.5  Serum Enzymes in Liver Diseases 561

Significance of SGOT in AMI
• Quantity of rise (SGOT value) is related to size of infarct.
• SGOT value is helpful in prognosis of AMI as:

–– SGOT value <50 IU/L indicates good prognosis and low mortality.
–– SGOT value >150 IU/L indicates poor prognosis and high mortality.
–– SGOT value >350 IU/L indicates very high mortality.

22.4.3 L actate Dehydrogenase (LDH)

It catalyzes reversible conversion in between pyruvate and lactate.

Occurrence
Lactate dehydrogenase is widely distributed in body tissues. It is present in high
concentration in liver cells, cardiac fibers, skeletal fibers, erythrocytes, brain tissues,
and the kidneys. Erythrocytes contain 100 times higher concentration of LDH than
plasma. LDH is liable to false-positive test in hemolysis. Therefore, it is a non-­
specific biomarker.

Normal Value
Its normal value varies between 120 and 360 IU/L.

Interpretation in Acute Myocardial Infarction
Its serum value increases 12 h after AMI and attains peak value in 48 h. It normal-
izes between the 8th and 12th day.
Significance in Acute Myocardial Infarction (AMI)

• LDH-2 is the predominant isoenzyme in plasma.
• In normal condition, concentration of LDH-2 in plasma is higher than concen-

tration of LDH-1.
• In acute myocardial infarction, LDH-1 concentration becomes higher than

LDH-2 concentration in plasma. This reversal of concentration of between two
LDH isoenzymes is called flipped pattern.
• Concentration of LDH.
• LDH is not a specific biomarker of AMI.

22.5 S erum Enzymes in Liver Diseases

The liver is the master organ to control metabolism of proteins, carbohydrates, and
lipids. Liver cells contain various enzymes. After injury to hepatocytes, enzymes are
released into plasma. Therefore, estimation of serum enzymes is a valuable clinical
tool that serves multiple functions.

562 22  Serum Enzymes and Organ Function Tests

22.5.1 S erum Transaminases

Serum Glutamate Pyruvate Transaminase (SGPT)
It is also called as alanine transaminase (ALT). It catalyzes reversible transamina-
tion between alanine and glutamate.

Occurrence
SGPT is primarily and abundantly found in liver cells.

Normal Value
Its normal value ranges between 5 and 45 IU/L.

Interpretation
• Elevation of SGPT >500 IU/L is suggestive of viral hepatitis, toxin-induced liver

disease, and ischemic liver disease.
• Value of SGPT rises in biliary cirrhosis (50–350 IU/L).
• Elevation of SGPT between 300 and 500 IU/L is found in Laennec’s cirrhosis.
• Elevation of SGPT between 150 and 300 IU/L is suggestive of obstructive jaun-

dice (posthepatic jaundice).

S erum Glutamate Oxaloacetate Transaminase (SGOT)
It is also called as aspartate transaminase (AST). It catalyzes reversible transamina-
tion between aspartate and glutamate.

Occurrence
SGOT is predominantly found in cardiac muscle fibers. It is also present in liver
cells.

Normal Value
Its normal value ranges between 5 and 40 IU/L.

Interpretation
• Elevation of SGOT is seen in cirrhosis patients.

Ratio of SGOT/SGPT has better diagnostic and prognostic significance than
either test performed alone.

Significance
• Elevation in SGPT level starts before clinical appearance of jaundice. Its peak

value is attained between 7 and 10 days. SGPT normalizes within 4 weeks of
onset of acute viral hepatitis.
• SGOT/SGPT ratio is <1in hepatitis excluding viral hepatitis.
• SGOT/SGPT ratio is >1in advanced cirrhosis of the liver and chronic hepatitis C.
• SGOT/SGPT ratio >2 is suggestive of chronic alcoholic cirrhosis.

22.5  Serum Enzymes in Liver Diseases 563

Gamma-Glutamyl Transpeptidase (GGT)
It is a transferase enzyme. It catalyzes the transfer of gamma-glutamyl group from

donor molecule like glutathione to an amino acid.

Occurrence
Gamma-glutamyl transpeptidase is mainly found in the liver. In the liver, it is
necessary for metabolism of drugs. It is also present in the pancreas, kidneys, and
spleen.

Normal Value
Its normal value ranges between 5 and 50 IU/L.

Interpretation
• GGT value rises in chronic viral hepatitis, alcoholic cirrhosis, and drug-induced

hepatitis.

Alkaline Phosphatase
It is orthophosphoric-monoester phosphohydrolase (hydrolase) enzyme.
It catalyzes breakdown of phosphate monoesters through addition of water at
alkaline pH.  Its exact physiological functions are still obscure. In bones, it is
helpful in bone mineralization. ALP is a biomarker for osteoblastic activity in
bones.

Occurrence
Alkaline phosphatase is ubiquitous in distribution. It is widely found in body tis-
sues. It is abundantly found in the liver, bone, intestine mucosa, kidneys, and
placenta.

Normal Value
Its normal value is 20–140 IU/L.

Interpretation
• Elevation in alkaline phosphatase value is two times its normal value in hepato-

cellular jaundice.
• Elevation in alkaline phosphatase value is nearly ten times its normal in obstruc-

tive jaundice (posthepatic jaundice).

Significance
• GGT to ALP ratio has better diagnostic and prognostic value than either value of

test alone.
• GGT/ALP >1.4 is highly suggestive of alcoholic cirrhosis than other liver

diseases. Therefore, ratio has a value in differential diagnosis of liver
diseases.

564 22  Serum Enzymes and Organ Function Tests

22.6 S erum Enzymes in Gastrointestinal Tract Diseases

22.6.1 Serum Amylase

Amylase is a hydrolase. It splits dietary starch into maltose under normal physiolog-
ical condition. In diseases of pancreas (acute pancreatitis), salivary glands (acute
parotitis), kidneys (renal failure) and diabetes mellitus, serum value of amylase is
elevated.

Occurrence
• Amylase is found in the saliva and pancreatic juice.

Normal Value
Its normal value ranges between 40 and 120 IU/L.

Interpretation
In Acute Pancreatitis
• Elevation of serum amylase is found in acute pancreatitis. Serum amylase

>1000  IU/L is seen in first 24  h of onset of acute pancreatitis. It normalizes
within 2–3 days of onset.

In Salivary Gland Diseases
• Serum amylase is elevated in mumps, bacterial parotitis, and sialolithiasis (sali-

vary stones). Its value ranges between 500 and1000  IU/L in salivary gland
diseases.

22.6.2 Serum Lipase

Lipase is a hydrolase enzyme. It digests lipids by hydrolyzing the ester linkage in
triglycerides.

Occurrence
Lipase is secreted by Ebner’s gland, gastric glands, pancreatic glands, and intestinal
glands in alimentary canal.

Normal Value
Its normal value ranges between 5 and 160 IU/L.

Interpretation
• Serum lipase is elevated to 1000 times the normal value in acute pancreatitis. It

normalizes within 10–14 days after onset of disease.
• Serum lipase level is also elevated in duodenal ulcer, carcinoma of the pancreas,

and liver cirrhosis.

22.8  Liver Function Tests 565

22.7 Serum Enzymes in Bone Diseases

Serum alkaline phosphatase
Alkaline phosphatase activity is associated with bone mineralization. Its activity is
a biomarker for osteoblastic activity in bones.

Occurrence
It is found in tissues like liver, intestine, bone and placenta.

Normal Value
In males: 45–110U/L, In females: 40–100 U/L.

Clinical Significance
• This enzyme has high clinical significance in detection of bone diseases.
• Serum alkaline phosphatase is elevated in Pagets’ disease, rickets, and

osteomalacia.
• Serum alkaline phosphatase is decreased in hypophosphatasia (rare, hereditary

disorder where bone mineralization is impaired).
• Its value is elevated in malignancy of bones.

Serum Acid Phosphatase in Prostate Cancer
Acid phosphatase is hydrolase enzyme. It catalyzes breakdown of phosphate mono-
ester at low pH.

Occurrence
It is found in the prostate gland, kidneys, spleen, liver, and erythrocytes.

Normal Value
Its normal value is <2 ng/ml.

Clinical Significance
• Its value is elevated in prostate cancer.

22.8 L iver Function Tests

Liver is a major organ of human body that performs multiple functions. It is a mas-
ter organ that controls metabolism of carbohydrates, lipids, and proteins. It detoxi-
fies ammonia into urea. It metabolizes drugs. It synthesizes physiologically
important compounds. The liver excretes biles.

Definition
Liver function tests comprise a batter of biochemical tests that help in assess-
ment and management of liver diseases.

566 22  Serum Enzymes and Organ Function Tests

Indications
Liver function tests (LFT) serve following functions as:
Diagnosis of Liver Disease
Liver function tests are noninvasive biochemical tests. They help to assess function-
ing of the liver. They are helpful in diagnosis of liver disease.
Differential Diagnosis of Liver Diseases
Liver function tests help to differentiate liver diseases. For example, the ratio of
SGOT/SGPT describes pattern of liver diseases and helps in differential diagnosis
between viral hepatitis and alcoholic hepatitis.
Prognosis of Liver Diseases
Liver function tests are useful in detection of severity of liver diseases. They are also
needful in determining outcome of liver diseases.
Long-term Follow-Up
Liver function tests are helpful in long-term management of liver diseases. For
example, chronic hepatitis B and C and alcoholic cirrhosis are associated with com-
plications. It requires periodic evaluation of liver functioning and evaluation of
therapeutic response to disease.

Limitations
Lack sensitivity: Test sensitivity is the ability to detect persons actually suffer-
ing from a disease (true positives).

Liver function tests may have compromised sensitivity in certain liver diseases.
Example:
Normal LFT in non-cirrhotic portal hypertension.
Lack specificity: Test specificity is its ability to detect persons who are actu-
ally healthy (without disease) (true negatives).
Liver function tests have poor specificity in liver diseases.
Example:
Hypoalbuminemia is found in chronic liver diseases and renal failure.
Serum aminotransferases are elevated in liver diseases and acute myocardial
infarction.

22.8.1 C lassification of Liver Function Tests

Group I (Tests for Excretory Function of the Liver)
• Serum bilirubin
• Urine bilirubin
• Urine urobilinogen

Group II (Tests for Enzymes of the Liver)
• Serum aminotransferases
• Serum alkaline phosphatase
• Serum γ-glutamyl transpeptidase

22.8  Liver Function Tests 567

Group III (Tests for Synthesizing Function of the Liver)
• Serum total protein, serum albumin, and serum globulin
• Prothrombin time

Group IV (Test for Carbohydrate Metabolism)
• Glucose tolerance test

Group V (Test for Lipid Metabolism)
• Serum cholesterol level

Group VI (Test for Amino acid Metabolism)
• Serum ammonia level test
• Ammonia tolerance test

Group VII (Test for Detoxification Function)
• Hippuric acid synthesis test

22.8.2 Group I (Tests for Excretory Function of the Liver)

• Serum bilirubin
• Urine bilirubin
• Urine urobilinogen

Serum Bilirubin Test
Bilirubin is a metabolic product of heme catabolism in the spleen. It is insoluble in
water and is called as unconjugated bilirubin or indirect bilirubin.

It undergoes conjugation with glucuronic acid in hepatocytes and forms bilirubin
diglucuronide. It becomes water soluble and called as conjugated bilirubin or
direct bilirubin.

Principle of Estimation
Its estimation is based on van den Bergh reaction. Serum bilirubin reacts with
diazo reagent to form a purple-colored compound.

Normal Value
Serum total bilirubin varies between 0.2 and 1.2 mg/100 ml.

Serum direct bilirubin is 0.2  mg/100  ml (represents 15% of total serum
bilirubin).

Serum indirect bilirubin is <1.2  mg/dl (represents 85% of total serum
bilirubin).

Interpretation
In Prehepatic Jaundice (Hemolytic Jaundice)
• Indirect bilirubin level is elevated. VD Bergh reaction is indirect positive (pur-

ple color appears only after addition of alcohol).

568 22  Serum Enzymes and Organ Function Tests

In Hepatocellular Jaundice
• Direct and indirect bilirubins are elevated. VD Bergh reaction is biphasic (purple

color appears with diazo reagent, its intensity ↑ with alcohol).

In Posthepatic Jaundice (Obstructive Jaundice)
• Direct bilirubin is elevated. VD Bergh reaction is direct positive (purple color

appears on addition of diazo reagent in sample).

Unconjugated Hyperbilirubinemia
Conditions like hemolytic jaundice and Gilbert syndrome are associated with
increased bilirubin formation and impaired conjugation of bilirubin, respectively.

It results into elevation of unconjugated fraction of bilirubin in body (>85%) and
is called as unconjugated hyperbilirubinemia. Serum unit may rise to <5 times
the normal value of serum unconjugated bilirubin (<6 mg/dl).

Conjugated Hyperbilirubinemia
Condition like obstructive jaundice or cholestasis is associated with obstruction
inflow of the bile from the liver to the duodenum. As a result, conjugated bilirubin
regurgitates into systemic circulation leading to rise in fraction of direct bilirubin in
serum (>50%) and is called as conjugated hyperbilirubinemia.

U rine Bilirubin Test
Principle of Test
Urine bilirubin is estimated by Gmelin’s test. Concentrated nitric acid oxidizes bili-
rubin into biliverdin to form greenish color in test tube.

Direct bilirubin is water soluble. It can pass the glomerular filtration membrane.
It appears in urine and the condition is called bilirubinuria. It is associated with
increase in serum direct bilirubin.

Normal Value
Urine bilirubin is 0.02 mg/100 ml. It is not traceable under normal condition.

Interpretation
In Prehepatic Jaundice (Hemolytic Jaundice)
• Serum indirect bilirubin is elevated which is insoluble in water. Therefore, bili-

rubin is absent in urine. The condition is called as acholuric jaundice.

In Hepatocellular Jaundice
• Direct and indirect bilirubins are elevated.

In Posthepatic Jaundice (Obstructive Jaundice)
• Direct bilirubin is formed in the liver. However, its excretion is obstructed and it

regurgitates into blood circulation. It is excreted by kidneys and appears in urine.
The condition is called as choluric jaundice.

22.8  Liver Function Tests 569

Urine and Fecal Urobilinogen
Bilirubin is reduced into urobilinogen by colon bacteria. It is a colorless com-

pound. A small amount of urobilinogen enters hepatic portal vein and reaches

the liver. It is delivered to circulation and excreted by kidneys. On exposure to
air, urobilinogen is oxidized into urobilin which provides yellow color to
urine.

Intestinal urobilinogen is reduced into stercobilin. It is excreted in feces. On
exposure to air, stercobilinogen is oxidized into stercobilin which provides
yellowish brown color to feces.

Principle of Test
Urine urobilinogen is detected by addition of Ehrlich reagent (mixture of paradi-
methyl amino benzaldehyde + Conc HCL) in 2 ml of urine sample. Red color of
solution appears in the presence of urobilinogen.

Normal Value
Urine Urobilinogen

Its value is 0.6 mg/per 24 h. In normal condition, it is untraceable in urine.

Interpretation
In Prehepatic Jaundice (Hemolytic Jaundice)

• Urine urobilinogen is increased. Urine appears dark yellow colored.

In Posthepatic Jaundice (Obstructive Jaundice) and Hepatocellular Jaundice
• Urobilinogen in urine is not detected.

Fecal Urobilinogen
Principle of Test
Fecal urobilinogen is detected by Edelman’s reagent (alcoholic mercuric chlo-
ride + alcoholic zinc chloride + amyl alcohol) with feces. Appearance of greenish
fluorescence indicates urobilinogen.

Normal Value
Its value ranges between 50 and 200 mg per day.

Interpretation
In Prehepatic Jaundice (Hemolytic Jaundice)
• Fecal urobilinogen concentration is increased. Feces appear dark brown

colored.

In Hepatocellular Jaundice
• Fecal urobilinogen is decreased. Feces appear pale colored.

In Posthepatic Jaundice (Obstructive Jaundice)
• Urobilinogen in feces becomes absent. Feces appear clay colored.

570 22  Serum Enzymes and Organ Function Tests

22.8.3 G roup II (Tests for Enzymes of the Liver)

• Serum aminotransferases
• Serum alkaline phosphatase
• Serum γ-glutamyl transpeptidase

Serum Transaminases

Serum Glutamate Pyruvate Transaminase (SGPT)
It is also called as alanine transaminase (ALT). It catalyzes reversible transamina-
tion between alanine and glutamate.

Occurrence
SGPT is primarily and abundantly found in liver cells.

Normal Value
Its normal value ranges between 5 and 45 IU/L.

Interpretation
• Elevation of SGPT >500 IU/L is suggestive of viral hepatitis, toxin-induced liver

disease, and ischemic liver disease.
• Value of SGPT rises in biliary cirrhosis (50–350 IU/L).
• Elevation of SGPT between 300 and 500 IU/L is found in Laennec’s cirrhosis.
• Elevation of SGPT between 150 and 300 IU/L is suggestive of obstructive jaun-

dice (posthepatic jaundice).

Serum Glutamate-Oxaloacetate Transaminase (SGOT)
It is also called as aspartate transaminase (AST). It catalyzes reversible transamina-
tion between aspartate and glutamate.

Occurrence
SGOT is predominantly found in cardiac muscle fibers. It is also present in liver cells.

Normal Value
Its normal value ranges between 5 and 40 IU/L.

Interpretation
• Elevation of SGOT is seen in cirrhosis patients.

Ratio of SGOT/SGPT has better diagnostic and prognostic significance than
either test performed alone.

Significance
• Elevation in SGPT level starts before clinical appearance of jaundice. Its peak

value is attained between 7 and 10 days. SGPT normalizes within 4 weeks of
onset of acute viral hepatitis.

22.8  Liver Function Tests 571

• SGOT/SGPT ratio is <1in hepatitis excluding viral hepatitis.
• SGOT/SGPT ratio is >1in advanced cirrhosis of liver and chronic hepatitis C.
• SGOT/SGPT ratio >2 is suggestive of chronic alcoholic cirrhosis.

Gamma-Glutamyl Transpeptidase (GGT)
It is a transferase enzyme. It catalyzes the transfer of gamma-glutamyl group from

donor molecule like glutathione to an amino acid.

Occurrence
Gamma-glutamyl transpeptidase is mainly found in the liver. In the liver, it is
necessary for metabolism of drugs. It is also present in the pancreas, kidneys, and
spleen.

Normal Value
Its normal value ranges between 5 and 50 IU/L.

Interpretation
• GGT value rises in chronic viral hepatitis, alcoholic cirrhosis, and drug-induced

hepatitis.

Alkaline Phosphatase
It is orthophosphoric-monoester phosphohydrolase (hydrolase) enzyme. It cata-

lyzes breakdown of phosphate monoesters through addition of water at alkaline

pH. Its exact physiological functions are still obscure. In bones, it is helpful in bone

mineralization. ALP is a biomarker for osteoblastic activity in bones.

Occurrence
Alkaline phosphatase is ubiquitous in distribution. It is widely found in body tis-
sues. It is abundantly found in the liver, bone, intestine mucosa, kidneys, and
placenta.

Normal Value
Its normal value is 20–140 IU/L.

Interpretation
• Elevation in alkaline phosphatase value is two times its normal value in hepato-

cellular jaundice.
• Elevation in alkaline phosphatase value is nearly ten times its normal in obstruc-

tive jaundice (posthepatic jaundice).

Significance
• GGT to ALP ratio has better diagnostic and prognostic value than either value of

test alone.
• GGT/ALP >1.4 is highly suggestive of alcoholic cirrhosis than other liver

diseases. Therefore, ratio has a value in differential diagnosis of liver
diseases.

572 22  Serum Enzymes and Organ Function Tests

22.8.4 G roup III (Tests for Synthesizing Function of the Liver)

• Serum total protein, serum albumin, and serum globulin
• Serum prothrombin

S erum Total Protein, Serum Albumin, Serum Globulin
The liver is the principal site of synthesis of albumin and globulin. Estimation of
serum albumin and globulin provide useful information regarding synthesizing
function of the liver.

Principle of Serum Total Proteins
Serum total protein can be estimated by biuret reaction. Protein in serum reacts
with copper sulfate in alkaline medium to form violet-colored biuret complex.
Serum protein concentration is proportional to intensity violet color.

Principle of Serum Albumin
Albumin in serum has the ability to interact with bromocresol green dye to form
BCG-albumin complex. This complex absorbs light at a specific wavelength.

Normal Value
• Serum albumin varies between 3.5 and 5.5 g/dl.
• Serum globulin varies between 2.5 and 3.5 g/dl.
• Serum total protein varies between 6 and 8 g/dl.
• Normal albumin/globulin ratio is 2:1.

Interpretation
In Prehepatic Jaundice (Hemolytic Jaundice)
• Serum albumin and globulin ratio remain normal in early stage of viral

hepatitis.
• In later stage, elevation of serum globulin has been found.

In Hepatocellular Disease
• Serum albumin is greatly reduced and serum globulin is elevated. A/G ratio is

reversed. It is highly suggestive of chronic cirrhosis liver.

In Posthepatic Jaundice (Obstructive Jaundice)
• Serum albumin and globulin ratio remain normal.

Prothrombin Time
Prothrombin is a clotting factor and plasma protein. It is synthesized by liver.
Prothrombin estimation is performed in the form of prothrombin time.

It is the time needed for clotting of a sample of citrated plasma which con-
tains thromboplastin and calcium.

22.8  Liver Function Tests 573

Normal Value
Normal value of prothrombin time is 14 s (PTcontrol).

Another method of expression of prothrombin time is international normalized
ratio (INR).

INR is the ratio of PT of patient to PT control as:

INR = PTpatient / PTcontrol
Normal value of INR is 1.0.

Interpretation
In Hepatocellular Diseases
• Prothrombin time is elevated depending upon severity of disease. It may rise to

ten times the normal value.

In Obstructive Jaundice
• Prothrombin time is elevated.

22.8.5 G roup IV (Test for Carbohydrate Metabolism)

• Glucose tolerance test

G lucose Tolerance Test
It determines the ability of body tissues to utilize carbohydrate after an intake of a
given amount of glucose.

22.8.6 T ypes of Glucose Tolerance Test

Oral Glucose Tolerance Test
Preparation of Patient
• Patient is advised to abstain from eating or drinking at least 8–12 h before test.
• Patient should be alert physically and mentally.

Procedure of Test
• A baseline fasting blood sample is collected.
• Patient is asked to drink a solution of 75 g of glucose (recommended by WHO

for adults) dissolved in 250 ml of water within 5 min.
• After every 30 min, five blood samples are collected.
• All six samples are estimated to determine blood glucose level. A graph is plot-

ted for six values of blood glucose concentration against time and it is called
glucose tolerance curve.

574 22  Serum Enzymes and Organ Function Tests

Interpretation
Normal Glucose Tolerance Curve
Characteristics
• Fasting blood glucose level should be <110 mg/dl.
• At 1 h period, blood glucose level should be <180 mg/dl. It is the highest peak

of blood glucose concentration. It should not exceed the renal threshold for glucose.
• At 21/2 h period, fasting blood glucose level (<110 mg/dl) should be obtained.

Diabetic Glucose Tolerance Curve
• Fasting blood glucose level is elevated (>110  mg/dl). Fasting glucose level

between 110 and 125 mg/dl indicates borderline impaired glucose tolerance.
• At 1 h period, blood glucose level rises >180 mg/dl. The highest peak of blood

glucose concentration is obtained after 1 h.
• At 21/2  h period, fasting blood glucose level is not obtained. It confirms

hyperglycemia.

Intravenous Glucose Tolerance Test
It is indicated in condition of malabsorption of glucose from alimentary canal.

Indications
• Coeliac disease
• Environmental enteropathy
• Hypothyroidism

Procedure
• A dose of 3 g/kg of weight of glucose is administered intravenously in 50% solu-

tion in 5 min.
• Baseline blood sample and half hourly five blood samples are taken.

Interpretation of glucose tolerance curve is same as in OGTT.

22.8.7 G roup V (Test for Lipid Metabolism)

• Serum Cholesterol

Liver regulates synthesis of cholesterol and helps in maintenance of serum cho-
lesterol level.

Normal Value
Normal total serum cholesterol varies between 150 and 250 mg/dl.

Interpretation
In Hepatocellular Diseases
• Serum cholesterol level is reduced.

22.8  Liver Function Tests 575

In Posthepatic Jaundice (Obstructive Jaundice)
• Serum cholesterol is elevated.

22.8.8 Group VI (Test for Amino Acid Metabolism)

Serum Ammonia Level
Liver is the chief organ for transamination and deamination of surplus amino acids.
These metabolic activities result into formation of ammonia.
Principle
• Serum ammonia level is determined by blood gas analysis.

Ammonia Tolerance Test
Test determines capability of liver to convert portal ammonia from intestine into urea.
Procedure
• Patient is advised to avoid any food overnight before the test.
• Blood sample in fasting state is procured.
• Patient is given 10 g of ammonium citrate in flavored syrup base to drink.
• Successive four blood samples are collected at intervals of ½ h, 1 h, 2 h, and

3 h.
• Ammonia gas is estimated by blood gas analysis.

Normal Value
Normal serum ammonia level varies between 40 and 70 μg/dl.

Interpretation
In Hepatocellular Disease
• Serum ammonia level is elevated. Its value may exceed 300 μg/dl. The liver func-

tions to convert ammonia coming from the intestine via portal vein into the urea.
This activity is impaired in hepatocellular diseases like cirrhosis, chronic hepati-
tis B and C, and alcoholic cirrhosis.
• Serum ammonia level is highly elevated (hyperammonemia) in hepatic
encephalopathy.

22.8.9 Group VII (Test for Detoxification Function)

Hippuric Acid Synthesis Test
Test determines two hepatic functions when benzoic acid in the form of sodium benzoate
is administered in body. Test determines synthesizing ability of liver along with
conjugation capability of the liver. Overall, test assesses detoxification ability of the liver.

Principle
Test is based on the following chemical reaction in hepatocytes:

Benzoic acid + aminoacetic acid (glycine) ® hippuric acid

576 22  Serum Enzymes and Organ Function Tests

Procedure
• Patient is provided breakfast.
• After 2 h, patient is instructed void the bladder.
• Provides a drink containing 6 g of sodium benzoate in 200 ml of water.
• All urine samples till next 4 h are collected and combined. Amount of hippuric

acid is estimated.

Interpretation
In Healthy Condition
• An amount of 3 g of hippuric acid is excreted.

In Hepatocellular Diseases
• Excretion of hippuric acid is decreased below 3 g.

Renal Function Tests
The kidneys are an indispensable excretory organ of the human body. The kidneys
perform multiple functions involving removal of waster substances, homeostasis,
and secretion of important hormones.
Functions of Kidneys
• Excretion of metabolic waste substances (urea, uric acid, xanthine, hypoxan-

thine, and creatinine)
• Excretion of exogenous substances from body (drugs, hormones)
• Homeostasis of acid-base balance
• Homeostasis of body fluid
• Secretion of hormones (renin, erythropoietin)

22.9 Renal Function Tests (RFT)

Renal function tests comprise a batter of biochemical tests that help in assess-
ment and management of renal diseases.

Indications
Renal function tests (LFT) serve the following functions.

Diagnosis of Renal Diseases
Renal function tests are noninvasive biochemical tests. They help to assess func-
tioning of kidneys. They are helpful in early detection of renal diseases.
Prognosis of Renal Diseases
Renal function tests are useful in detection of severity of renal diseases. They are
also needful in determining outcome of renal diseases.
Long-term Follow-Up
Renal function tests are helpful in long-term management of renal diseases.
These tests help to assess efficacy of drugs and need for kidney replacement.

22.9  Renal Function Tests (RFT) 577

Classification of Renal Function Tests
Group I (Tests for Glomerular Filtration)
• Urea clearance test
• Creatinine clearance test
• Inulin clearance test

Group II (Tests for Renal Plasma Flow)
• P-amino hippuric acid test
• Filtration fraction test

Group III (Tests for Tubular Functions)
• Concentration and dilution tests

Glomerular Filtration Tests
Glomerular filtration test estimates the capability of nephrons to form glomerular fil-
trate. In a healthy adult person, 180 L glomerular filtrate is formed in 24 h by kidneys.

Normal Value of Glomerular Filtration Rate (GFR)
Its normal value is 125 ml/min.

Glomerular filtration rate cannot be estimated directly. It is assessed by clear-
ance test.

Definition
Clearance (C) is the volume of plasma which is cleared from an indicator sub-
stance in 1 min.

Clearance of a substance can be calculated by the following formula:

Clearance of substance (Z) = U´V
P

where U = Concentration of substance in urine, V = Volume of urine, P = Concentration

of substance in plasma.

Since

GFR = clearance of an indicator substance subject to certain characteristics,
therefore

GFR = Uz ´ V / Pz

where Uz (concentration of substance in urine) and Pz (concentration of substance in
plasma).

Characteristics of Indicator Substance
• In case of endogenous substance, the rate of its formation should be constant.
• In case of exogenous substance, it should be inexpensive, easy to administer, and

physiologically inert.

578 22  Serum Enzymes and Organ Function Tests

• Indicator substance should be freely filtered by kidneys.
• It should not be reabsorbed by tubules.

Based on type of indicator substance, clearance tests are the following types:

Urea Clearance Test
Kidneys excrete urea from plasma.

Creatinine Clearance Test
Creatinine is endogenous substance. It is a metabolic waste product formed from
decomposition of creatine phosphate in skeletal muscles.
Characteristics of Creatinine as Indicator Substance
• Creatinine is synthesized at constant rate.
• Its synthesis is proportional to muscle mass.
• It is freely filtered by kidneys.
• It is not reabsorbed by tubules.
• It is secreted by renal tubules.

Procedure
• Urine sample is collected in 24 h.
• Blood sample is taken.
• Measure volume of urine, urine creatinine concentration, and serum creatinine

concentration.

Principle
• Serum creatinine is estimated by Jaffe’s reaction. Creatinine reacts with picric

acid in alkaline medium to form reddish orange color. Its intensity is proportional
to creatinine concentration.

Creatinine clearance = GFR

Creatine clearance = Ucr ´ V / Pcr

Normal Value
• Normal value of serum creatinine is 0.7–1.2 mg/dl in males and 0.6–1.1 mg/dl in

females.
• Serum creatinine is inversely proportional to renal function.
• Normal creatinine clearance is nearly 105 ml/min.

Interpretation
• Serum creatinine concentration is elevated in renal diseases. It is inversely pro-

portional to GFR.
• Creatinine clearance is decreased in renal diseases.

22.9  Renal Function Tests (RFT) 579

Remarks
• Creatinine is secreted by renal tubules. In renal failure, GFR decreases. However,

total creatinine clearance increases owing to tubular secretion. So it is not an
ideal and reliable biomarker for renal function. It leads to overestimation of
GFR.

Inulin Clearance Test
Inulin is a homopolysaccharide of fructose. It is employed as exogenous marker for
ascertaining renal dysfunction.
Characteristics of Inulin as Indicator Substance
• It is an exogenous substance.
• It is easy to administer.
• It is economical and chemically inert substance. It is not metabolized in the

body.
• It is freely filtered by the kidneys.
• It is neither reabsorbed nor secreted by renal tubules.

Procedure
• Test is performed after a light breakfast.
• Solution of 10 g of inulin in 100 ml water is prepared. It is administered through

IV route.
• Urine is collected after 2 and 4 h. Urine volume is measured.
• Blood sample is collected after 2  h of inulin administration. Serum inulin is

estimated.

Inulin clearance = Uinulin ´ V / Pinulin

Normal Value
Its normal value is 125 ml/min. Inulin clearance is almost comparable to glomerular
filtration rate.
Interpretation
• Inulin clearance is decreased in renal diseases. It indicates decline in glomerular

function of kidneys.

Test for Renal Blood Flow
This test is not performed in routine clinical practice.

Para-aminohippuric acid is utilized as a marker to determine for renal plasma
flow (RPF). RPF is estimated by PAH clearance, since it is filtered by glomeruli and
also secreted by tubules.

Normal Value of RPF
• In males

–– Its value is 650 ml/min/1.73 m2.
• In females

–– Its value is 600 mL/min/1.73 m2.

580 22  Serum Enzymes and Organ Function Tests

Tests for Tubular Function
Renal perfusion is important in determining normal tubular functioning. Renal
hypoperfusion due to hyperdynamic changes, drug- and/or toxin-induced, definitely
reduces blood flow to kidneys. It impairs tubular functions.

Concentration Test
Principle
Concentration test determines capability of kidneys to reabsorb water and concen-
trate urine. Test relies on estimation of specific gravity of urine by urinometer.
Procedure
• Patient is advised to have dinner 12 h before the test (8PM if test is scheduled on

next day morning 8AM). Thereafter, patient is advised to keep fasting till next
day morning.
• Any urine passed midnight is discarded.
• In the morning at 8AM, collect sample of urine. Thereafter, two more urine
samples are collected after an interval of 1 h (9AM–10AM).

Normal Value
• Normal specific gravity of urine is 1.025.

Interpretation
• In normal tubular functioning, specific gravity of at least one sample should be

≥1.025.
• In abnormal tubular functioning, specific gravity of urine sample is <1.020.
• In severe renal failure, specific gravity of all urine samples is fixed at 1.010.

Dilution Test
Principle
Test determines capability of the kidneys to remove water. Test relies on estimation
of urine output and its specific gravity after intake of a given volume of water.
Procedure
• Patient is advised to have dinner 12 h before the test (8PM if test is schedule on

next day morning 8AM). Thereafter, patient is advised to keep fasting till next
day morning.
• Patient is asked to empty bladder and discard urine.
• Patient is given 1200 ml of water to drink in 30 min.
• Four urine samples are collected after an interval of 1 h in between each sample.

Interpretation
• Normal Kidney Function

–– About 80% of total water intake (1000  ml) should be evacuated in urine
within 4 h.

–– Specific gravity of at least one urine sample should be ≤1.003.

Suggested Readings 581

• Kidney Dysfunction
–– Excretion of water intake is delayed.
–– Specific gravity of samples should not be ≤1.0031. It becomes almost con-
stant at 1.010.

Suggested Readings

Kleiner IS, Orten JM (1966) Biochemistry, 7th edn. Mosby, St Louis
Latner AL, Cantarow A (1975) 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, Phialdelphia
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. Churchill,

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

Part V
Immunochemistry

Immunoglobulins 23

Extrinsic substances invade the body. They stimulate the immune system of the
host. It produces endogenous substances which destroy extrinsic substances. The
former substance is the antigen, while the latter is called as antibody.

23.1 D efinition

Antigen
Antigen is exogenous organic molecule which can elicit either one or both types
of immune reactions in the body of host.

Generally, antigens can be polypeptides or polysaccharides in nature. For exam-
ple, pneumococcal capsule contains polysaccharide as antigen. The peptidoglycan
is the chief structural component of gram-positive bacteria. It is composed of poly-
mer of disaccharides cross-linked to peptides. Bacterial peptidoglycan is antigenic
in nature.

Lipids and nucleic acids are rarely antigen DNA fragments, or oligoribonucleo-
tides covalently linked to large protein molecule can act as antigen. Lipids or ste-
roids linked to large protein molecule are antigens and are called as “haptene,” for
example, aniline, dinitrophenol, and hydralazine.

Antibody
Antibody is an endogenous glycoprotein which exhibits specificity for a par-
ticular antigen.

In antibody, glycoprotein is made up of oligosaccharides covalently attached to

polypeptides. This is called as glycosylation. The polypeptides belong to immuno-

globulin superfamily of proteins. Immunoglobulin superfamily is constituted by cell
surface proteins and soluble proteins. Antibody and immunoglobulin are synony-
mously used terms.

Immunoglobulins are endogenous glycoproteins belonging to immunoglob-
ulin superfamily of proteins.

© Springer Nature Singapore Pte Ltd. 2019 585
A. Gupta, Comprehensive Biochemistry for Dentistry,
https://doi.org/10.1007/978-981-13-1035-5_23