Introduction to Carbohydrates
Page: 1-7 (7)
Author: Mrittika Sarkar, Sinchan Hait* and Sai Joshi
DOI: 10.2174/9789815165852124010004
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Abstract
Carbohydrates are one of the four major classes of biologically essential
organic molecules in living organisms. They are the most abundant class of
biomolecules in nature based on their mass. And they make up most of the organic
bodies on earth due to their extensive role in every life form. Carbohydrates are
polyhydroxy aldehydes or ketones or substances that result in such compounds upon
hydrolysis. These macromolecules are composed of carbon (C), hydrogen (H), and
oxygen (O). They are also referred to as saccharides (sakcharon = sugar or sweetness)
since most of them have a sweet taste. Apart from that, carbohydrate serves as a
primary energy source, a structural framework for nucleotides, and provides structural
support to organisms. They also play a crucial role as mediators in cellular interaction.
Carbohydrates are classified into monosaccharides, oligosaccharides, and
polysaccharides. Monosaccharides are the simplest form of carbohydrates, and a few
monosaccharides hydrolyse to form oligosaccharide, and many monosaccharides
together form polysaccharides. They also act as a precursor for glycoproteins and
glycolipids. In this section, we are discussing some basic concepts about carbohydrates.
Qualitative Tests for Carbohydrate Detection
Page: 8-31 (24)
Author: Mrittika Sarkar, Sinchan Hait* and Sai Joshi
DOI: 10.2174/9789815165852124010005
PDF Price: $15
Abstract
Qualitative tests for the determination of carbohydrates in the given sample
are a crucial part of before getting into major analytical procedures. Every mono-,
oligo-, and poly-saccharide vary depending upon the number of carbohydrate molecule
present in it as well as changes in the side chains. Thus, every carbohydrate molecule
has distinct functions and properties. Depending upon these physio-chemical properties
of the carbohydrates, they respond to certain specific chemical reactions under certain
conditions. Only mono- and dis-saccharides respond to the solubility test as they are
soluble in water at room temperature. The Molisch test is only for the determination of
the presence of carbohydrate, not depending on the types of it. The iodine test gives a
result for polysaccharides. Whereas Fehling, Benedict and Osazone tests distinguish
between reducing and non-reducing sugars. The Bradford test differentiates between
mono- and di-saccharide-reducing sugars. Seliwanoff test is only for sucrose which is a
non-reducing sugar. Bial’s test is to determine the presence of pentose sugars.
Quantification Tests for Carbohydrate Detection
Page: 32-54 (23)
Author: Mrittika Sarkar, Anshika Sah, Payal Agarwal, Sai Joshi and Sinchan Hait*
DOI: 10.2174/9789815165852124010006
PDF Price: $15
Abstract
Quantification of carbohydrates by the anthrone method is a colourimetric
assay. In this method, first, the complex carbohydrates are hydrolysed in a highly
acidic medium, resulting in monosaccharides. Thereafter, these monosaccharides are
dehydrated to form 5-hydroxymethylfurfural (5-HMF) or 2-furfural (2-F), followed by
reacting with anthrone reagent. This reaction produces a blue-green complex which is
colourimetrically determined. In DNSA method 3,5-dinitrosalicylic acid (DNSA), a
yellow colour reagent reacts with reducing sugar’s carbonyl group and forms orangered colour compound 3-Amino-5-nitrosalicylic acid (ANSA). The quantity of
carbohydrate present in a sample is determined by Beer-Lambert’s law. Apart from
these techniques Folin-Wu, Hexokinase, Phenol-Sulphuric acid, Somogvi-Nelson, and
GOPOD method are discussed in this chapter in detail.
Introduction to Proteins
Page: 55-61 (7)
Author: Sinchan Hait* and Sai Joshi
DOI: 10.2174/9789815165852124010007
PDF Price: $15
Abstract
Proteins are the ambidexterity macromolecules which play a crucial role in
biological processes. Through proteins, genetic information is expressed. They exhibit
enormous functions which are crucial in various pathways. Some proteins act as an
enzyme which is vital for catalyzing various metabolic reactions in living cells. All
proteins are made up of amino acids which join together via a peptide bond to form a
polypeptide chain which is a precursor for the further development of protein. A
nascent protein undergoes several post-translational modifications to achieve its true
function. Based on the amino acids present in the protein, its function determines its
role as hormones, enzymes, antibodies, transporters, etc. In this chapter, a brief
introduction to protein’s structure and function is discussed.
Qualitative Tests for Protein Detection
Page: 62-83 (22)
Author: Sinchan Hait*
DOI: 10.2174/9789815165852124010008
PDF Price: $15
Abstract
Qualitative tests are the primary assays for detecting the presence of any
protein in a given sample. Qualitative detection of proteins is based on the chemical
interactions between amino acids and certain chemical reagents added to them. These
chemical reactions yield colour formation, which corresponds to specific amino acid or
a group of amino acids present in the sample. In this chapter, some of such most
important assays for qualitative analysis are discussed in detail.
Protein Determination by SDS-PAGE
Page: 84-97 (14)
Author: Sai Joshi*
DOI: 10.2174/9789815165852124010009
PDF Price: $15
Abstract
SDS-PAGE is the abbreviated form of sodium-dodecyl sulphatepolyacrylamide gel electrophoresis. It is a commonly used gel electrophoresis for the
qualitative estimation of proteins. It is based on the principle that treatment of protein
with SDS, an anionic detergent, causes a constant negative charge throughout the
protein surface. It also breaks the protein chain into smaller peptide chains. Hence, the
separation of proteins is solely based on the size and shape of the fragments generated
and is independent of the charge of the protein. This, combined with polyacrylamide
gel electrophoresis, allows for visualization of these fragments where the smaller
fragments retain at the bottom of the gel. This method has been discussed in detail in
this chapter.
Protein Determination by Western Blotting or Immunoblot
Page: 98-105 (8)
Author: Sinchan Hait*
DOI: 10.2174/9789815165852124010010
PDF Price: $15
Abstract
Western blotting, also known as immunoblot, is a chromatography-based
technique in which quantitative detection of proteins can be performed based on their
molecular weight. This is often performed after the SDS-PAGE technique. It is used to
identify, analyze and quantify proteins from a protein mixture. The major steps include
1. Separation of proteins (e.g., SDS-PAGE), 2. Transfer onto a solid support, 3.
Utilizing primary and secondary antibodies for visualization. Western blotting can be
used qualitatively and quantitatively. In this chapter, both of these techniques are
explained in detail.
Quantitative Tests for Protein Detection
Page: 106-127 (22)
Author: Sinchan Hait, Sai Joshi* and Ankita Nitin Padhye
DOI: 10.2174/9789815165852124010011
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Abstract
Quantitative tests for protein are essential for understanding the amount of
protein present in a sample. Various tests have been developed, such as BCA, biuret,
Bradford, and Lowry tests. These are the most common and standardised procedures
for the determination of protein. The mentioned techniques are colourimetric assays.
However, over a while, several more advanced techniques are also being developed. In
this chapter, quantitative methods for protein are discussed.
Protein Purification
Page: 128-152 (25)
Author: Sinchan Hait, Sai Joshi* and Ankita Nitin Padhye
DOI: 10.2174/9789815165852124010012
PDF Price: $15
Abstract
Protein purification is an essential step after protein isolation. Protein
purification ensures the removal of all contaminants from the sample pool. Purification
depends on the chemical and physical interactions of protein molecules and chemical
moieties. These techniques are performed with tags and without tags. Tag purification
involves ammonium sulfate precipitation and dialysis. Purification of proteins with tags
utilizes affinity and size exclusion chromatography. There are various methods
available for this, however, we will discuss some most common methods for protein
purification.
Introduction to Nucleic Acids
Page: 153-161 (9)
Author: Sai Joshi*, Sinchan Hait and Ahiree Ghosal
DOI: 10.2174/9789815165852124010013
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Abstract
Nucleic acids are biomolecules which carry all the genetic information in the
form of DNA and RNA. The nucleic acids are composed of primarily a 5-carbon sugar,
a phosphate and a nitrogenous basis. These nitrogenous bases can be adenine, guanine,
thymine, cytosine and uracil. Nucleic acids can be measured qualitatively by UV
absorbance, diphenylamine method/orcinol method, fluorescence dyes and capillary
electrophoresis. It can be measured qualitatively by agarose gel electrophoresis.
Qualitative Tests for Nucleic Acid Detection
Page: 162-175 (14)
Author: Anshika Sah, Mrittika Sarkar, Sinchan Hait, Sai Joshi* and Ahiree Ghosal
DOI: 10.2174/9789815165852124010014
PDF Price: $15
Abstract
Qualitative tests of nucleic acids are important to detect the presence or
absence of nucleic acid in the given sample. It can be done by performing agarose gel
electrophoresis. Nucleic acids are negatively charged and hence can be attracted
towards a positive charge in an electric field. This phenomenon is used in agarose gel
electrophoresis to detect nucleic acids. Based on the mass of nucleic acids, different
bands are formed in the gel, which can be visualized using fluorescent dyes.
Quantitative Tests for Nucleic Acid Detection
Page: 176-193 (18)
Author: Mrittika Sarkar, Sinchan Hait, Sai Joshi* and Revati Supekar
DOI: 10.2174/9789815165852124010015
PDF Price: $15
Abstract
Nucleic acid quantification is essential in experiments associated with
genetic engineering. Over periods, several techniques have been developed to quantify
nucleic acids. UV-Vis spectrophotometer is the simplest technique where it is
associated with absorbance, which is proportional to the concentration of the sample.
There are some colourimetric methods, such as the diphenylamine (DPA) method, and
the orcinol method, where nucleic acid reacts with diphenylamine and forms a bright
blue colour complex, and orcinol reacts and forms a green colour complex. Apart from
that, some advanced techniques, such as fluorescence techniques, are used for
quantification. In this chapter, each of these methods is explained in detail.
Purification of Nucleic Acids
Page: 194-206 (13)
Author: Sai Joshi*, Sinchan Hait and Niriksha Patel
DOI: 10.2174/9789815165852124010016
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Abstract
Besides qualitative and quantification of nucleic acids, purification is an
integral part of any genetic engineering experiments. In genetic engineering
experiments, the purity of nucleic acid matters the most. Nucleic acid purification can
be achieved by the phenol-chloroform method, alcohol-based purification, agarose gel
electrophoresis, and chromatographic purification. These purifications are based on the
interactions between bases of nucleic acids and chemical reagents. In this chapter,
nucleic acid purification is explained in detail.
Introduction to Lipids
Page: 207-216 (10)
Author: Sai Joshi, Sinchan Hait* and Pranali Garde
DOI: 10.2174/9789815165852124010017
PDF Price: $15
Abstract
Lipids are water-insoluble hydrocarbon derivatives with many physiological
functions. They are the structural components of membranes. The long chain fatty acid
chains of lipids can be broken down to serve as energy fuels, as well as they play a role
in intracellular signaling. The entire composition of lipid molecules in a cell constitutes
its lipidome. In this chapter, we will discuss the different roles played by lipids by their
chemical properties in detail.
Qualitative Tests for Lipid Detection
Page: 217-233 (17)
Author: Sai Joshi*, Sinchan Hait and Pranali Bhosale
DOI: 10.2174/9789815165852124010018
PDF Price: $15
Abstract
Qualitative tests are the primary assays for detecting the presence of any
lipids in a given sample. Qualitative detection of lipids is based on the chemical
interactions between components of lipids and certain chemical reagents added to
them. These chemical reagents allow for the properties of lipids like hydrophobicity,
translucency, or colourimetric analysis due to colour changes. These tests have been
explained in detail in this chapter.
Quantitative Tests for Lipid Detection
Page: 234-266 (33)
Author: Sai Joshi, Sinchan Hait* and Pranali Garde
DOI: 10.2174/9789815165852124010019
PDF Price: $15
Abstract
Quantification of lipids can be broadly classified into the following methods:
gravimetric analysis, which employs the principle of phase separation, the acid value is
based on the property of rancidity of fats, saponification value which relies on alkali
required to saponify (hydrolyze) lipids containing mixture, iodine value depends on the
interaction of fats with halogen iodine, and blood cholesterol estimation determines the
cholesterol level by cholesterol esterase enzyme-based assay. These assays have been
explained in detail in this chapter. Advanced techniques used for lipid quantification
utilize the principles of chromatography (Thin-layer chromatography, gas
chromatography, and HPLC) and spectroscopy (infrared spectroscopy, Raman
spectroscopy, nuclear magnetic resistance spectroscopy, and fluorescence
spectroscopy), and different colourimetric assays are discussed in detail in this chapter.
Commonly Used Basic and Advanced Techniques
Page: 267-287 (21)
Author: Anshika Sah, Sinchan Hait and Sai Joshi*
DOI: 10.2174/9789815165852124010020
PDF Price: $15
Abstract
Different techniques are used in a biochemical laboratory to measure various
parameters. For example, a pH meter is used to measure the pH of the sample and
electrochemical cells are used to measure the redox potential. Zeta potential is another
property which can be measured using zeta cells. Chromatography is a technique used
to separate the components of a mixture. Two of the most common chromatography
techniques are HPLC and ion exchange chromatography. HPLC is based on the
interactions of the analytes in stationary as well as mobile phase. Whereas ion
exchange chromatography is based on the interaction of ions and charged site of the
stationary phase.
Significant Experimental Hazard: Safety Data Sheet (SDS)
Page: 288-298 (11)
Author: Sai Joshi*, Anshika Sah and Sinchan Hait
DOI: 10.2174/9789815165852124010021
PDF Price: $15
Abstract
A safety data sheet or SDS is a datasheet prepared by the manufacturer to
illuminate the user about any potential hazard and preserved by the owner. SDS also
provides information about the handling and working of the product. SDS is also
known as a material safety data sheet (MSDS) or product safety data sheet (PSDS). It
must include information about the physical, environmental health, and health hazards;
safety precautions, and protective measurements; transportation and storing. Guidelines
for the preparation of such SDS are laid down by the occupational safety and health
administration (OSHA). This chapter briefly explains the SDS of the significant
experimental hazards of those chemicals used in the previous chapters.
Introduction
Practical Biochemistry provides both foundational knowledge and advanced insights into biochemistry, including the basic compounds, and laboratory methods. The book is designed for students and academic professionals seeking a comprehensive understanding of the practical aspects of the subject. The book is systematically divided into five sections, each dedicated to a specific category of macromolecules and related biochemical techniques: 1) Carbohydrates, 2) Proteins, 3) Nucleic acids, 4) Lipids, 5) Supplementary Techniques and Safety Data Sheet (SDS). Each chapter within these sections is structured to provide a thorough understanding of the aim, principles, procedures, and practical applications of biochemical techniques. Key features: · Comprehensive Information: meticulously organized and structured chapters provide a thorough and methodical approach to learning · Additional Learning Tools: ‘Did You Know’ segments and ‘Viva Voice’ questions enrich the learning experience by offering interesting facts and stimulating critical thinking · Practical Focus: Step-by-step guides aid readers in understanding and applying the techniques in the lab · Safety and Accuracy: teaches how to conduct safe and accurate experiments with precautions · Accessible Language: simple and lucid language helps beginners to understand complex biochemical concepts