Principle

Of

Food
Analysis
 Topic
1
Afzal sir Spectroscopy
• Introduction
• Energy state of matter Itroduction
• Electronic Energy Spectroscopy is a scientific technique that
Levels involves the study of the interaction
• Nuclear Energy between matter and electromagnetic
Levels radiation (light) as a function of
• Nature of electromagnetic wavelength or frequency. It is used to
radiation obtain information about the composition,
• Theory of Spectroscopy structure, and properties of materials, as
• Energy levels transitions well as to study various physical processes.
in spectroscopy Energy state of matter
• Absorption of radiation Atoms and molecules can only exist in a
• Emission of radiation limited number of discrete energy levels
• Modes of measurement they cannot have energies between these
• Spectroscopic Analysis levels, i.e., their energy levels are
• Quantitative Analysis quantized. Each molecular species has a
• Qualitative Analysis unique set of energy levels that depends on
• Spectroscopic techniques its unique atomic structure (electrons,
• γ- Ray and X-Ray protons, neutrons) and molecular structure
Spectroscopy (type and arrangement of atoms and
• Ultraviolet and bonds).
Visible spectroscopy • Imagine a ladder with several steps, and
• Applications you are a tiny ant trying to climb the
• Infrared Spectroscopy ladder. However, you can only step on
• Application specific steps and not on the spaces
• Raman spectroscopy between the steps. These steps represent
• Spectrofluorometry the allowed energy levels that atoms
• Nuclear Magnetic and molecules can have.
Resonance (NMR)
Spectroscopy
• Mass Spectroscopy
• Summary
• Now, let's consider that each ladder represents a different type of
material with its own unique set of energy levels. For example, a
wooden ladder might have six steps, while a metal ladder could have
eight steps. Each ladder's steps are specific to that material's atomic and
molecular structure.
The relative potential energy of an atom or molecule is made-up of
contributions from a number of different sources: electronic, vibrational,
rotational, translation and nuclear.

1. Electronic Energy Levels: Electronic energy levels refer to the

specific quantized energy states that electrons can occupy within
an atom or a molecule.

2. Nuclear Energy Levels: The nuclei of certain atoms have a

property known as spin. A (charged) spinning nucleus generates a
small magnetic field and can be thought of as being a small
magnet. Normally, this magnet can be orientated in any direction,
but in the presence of an external magnetic field it can only align
itself either with or against the field, i.e., it is quantized.
• Spinning Compass Trees: The compass needles (nuclei) can spin, just
like a top spinning on its axis. When they spin, they generate a small
magnetic field, much like how a spinning magnet creates a magnetic
field around it.
• Free Orientation: Normally, in the absence of any external influence,
these compass needles (nuclei) can point in any direction. They have
the freedom to orient themselves in any way they like, just like a
compass needle that can point in any direction.
• External Magnetic Field: Now, imagine that someone outside the
forest brings in a large magnetic bar. This bar represents an external
magnetic field applied to the forest.
• Quantization of Alignment: When the external magnetic bar is
brought close to the compass trees, something interesting happens.
The compass needles (nuclei) can only align themselves either with
the magnetic bar's direction or against it. They can no longer point in
any random direction but have to choose between two specific
orientations—aligned or anti-aligned with the external magnetic
field.
Nature of electromagnetic radiation
Electromagnetic radiation is a form of energy that propagates through space
in the form of waves or particles. It consists of both electric and magnetic
fields oscillating perpendicular to each other and also perpendicular to the
direction of wave propagation. This type of radiation doesn't require a
medium to travel through and can propagate through a vacuum, which sets it
apart from other forms of waves, like sound waves.
• Here are some key characteristics and properties of electromagnetic
radiation:
• Wide Spectrum: Electromagnetic radiation spans a broad spectrum of
wavelengths and frequencies, from radio waves to gamma rays.
• Speed of Light: Electromagnetic radiation travels at the speed of
light in a vacuum, which is approximately kilometres per second.
The speed of light is the maximum possible speed for anything in the
universe.
• Dual Nature: Electromagnetic radiation exhibits both wave-like and
particle-like behaviour. At high energies, electromagnetic radiation
behaves more like particles, called photons, while at lower energies,
it behaves more like waves.
• Quantization of Energy: The energy of electromagnetic radiation is
quantized into discrete packets called photons. Each photon carries a
specific amount of energy, which is directly related to the frequency
(or inversely related to the wavelength) of the radiation.

• Interaction with Matter: It interacts with matter through absorption,

reflection, transmission, or scattering.
• Types of Radiation: The spectrum includes radio waves, microwaves,
infrared, visible light, ultraviolet, X-rays, and gamma rays.
Theory of Spectroscopy
• Energy Levels and Transitions: At the atomic and molecular level,
electrons and nuclei have quantized energy levels. Interactions with light
can cause transitions between these levels, resulting in absorption,
emission, or scattering of light.
• Quantization of Energy: Light energy is quantized into discrete packets
called photons. The energy of photons determines the frequency or
wavelength of light.
• Spectral Patterns: Interactions between light and matter produce
characteristic spectral patterns, like fingerprints, specific to each material.
These patterns reveal information about the material's composition and
structure.
• Types of Spectroscopy: Different spectroscopic techniques exist, such as
visible and ultraviolet spectroscopy, infrared spectroscopy, NMR
spectroscopy, mass spectrometry, Raman spectroscopy, and X-ray
spectroscopy, each suited to different applications.

The spectroscopic techniques can thus divide into two categories: absorption
and emission spectroscopy.
1. Absorption: The sample absorbs specific wavelengths of light, causing
electrons or molecules to transition to higher energy levels. The absorbed
energy is characteristic of the sample's chemical composition and can be
used to identify and quantify substances.
2. Emission: Excited electrons or molecules in the sample can return to lower
energy levels, releasing energy in the form of emitted light. The emitted
light can be analysed to provide information about the sample's properties.
Energy levels transitions in spectroscopy
Spectroscopic techniques utilize the fact that atoms and molecules have a
discrete set of energy levels and that transitions can only occur between them.
• The Energy Staircase:
• Imagine a building with multiple floors, each floor representing a
different energy level of an atom or molecule. These energy levels are
quantized, like discrete steps in a staircase.
• At the ground floor, an atom or molecule is in its lowest energy state,
with electrons occupying specific energy levels called electron shells.
• When light (energy balls) shines on the building, electrons can absorb
the energy and move up to higher floors (higher energy levels) in a
process called absorption.
• Conversely, electrons on higher floors can release excess energy and
move back down to the ground floor, emitting light in the form of new
energy balls, in a process called emission.
• Energy level transitions are quantized, meaning electrons can only
move between specific floors, not staying in intermediate positions.
Absorption of radiation
Absorption is the process by which energy is transferred from an
electromagnetic wave to an atom or molecule and causes it to move to an
excited state. Absorption can only occur when an atom or molecule absorbs a
photon of light that has an energy which exactly corresponds to the difference
between two energy levels, i.e., it must be quantized.
Emission of radiation
Emission of radiation refers to the process by which an atom or molecule
releases energy in the form of electromagnetic radiation, such as light, after
being in an excited state. When atoms or molecules absorb energy and
become excited, they can later return to lower energy states by emitting this
excess energy in the form of photons (particles of light).
• Fireworks in Excitement: Initially, the fireworks are at rest,
representing atoms or molecules in their ground state (lowest energy
level).
• Adding Energy: To start the show, a group of performers (external
energy source) launches firecrackers towards the fireworks. These
firecrackers represent the energy input, which can come from various
sources like heat, light, or electricity.
• Excited Fireworks: When the firecrackers reach the fireworks, they
explode, releasing a burst of energy. This represents the absorption of
energy by the atoms or molecules, causing them to move to higher
energy levels (excited state).
• Colourful Emission: After reaching the excited state, the fireworks
cannot stay there for long. They are ready to put on a spectacular
show by emitting bursts of colorful lights (emission of radiation) in
all directions.
• Specific Colours: Each firework (atom or molecule) emits light of
specific colours. The colours depend on the material and its unique
energy levels. Just as different fireworks produce different colours,
different atoms or molecules emit radiation at specific wavelengths
based on their energy level differences.
• Returning to Rest: Once the fireworks have released their energy and
dazzled the audience, they return to their original state (ground state).
This represents atoms or molecules returning to lower energy levels
by emitting radiation.
• Non-radiative decay
• Non-radiative emission, also known as non-radiative decay or non-
radiative relaxation, is a process by which an excited atom or
molecule releases its excess energy without emitting a photon of light
(electromagnetic radiation). Instead of emitting light, the energy is
transferred to another form, such as heat or vibrations within the
material.
• Radiative decay
• Radiative decay, also known as radiative emission or spontaneous
emission, is a process in which an excited atom or molecule releases
its excess energy in the form of electromagnetic radiation (photons of
light).
Modes of measurement
The design of an analytical instrument based on spectroscopy depends on the
nature of the energetic transitions involved (e.g., electronic, vibration,
rotation, translation, nuclear), the nature of the radiative process involved
(e.g., absorption, emission, fluorescence) and the nature of the food matrix
(e.g., absorbing or non-absorbing).
• Emission: Measures the intensity and wavelengths of light emitted by
a sample in an excited state.
• Absorption/Transmission: Measures the amount of light absorbed by a
sample at different wavelengths.
• Reflection: An electromagnetic wave generated by the analytical
instrument is reflected from the surface of the sample and the
reduction in its amplitude due to interaction with the sample is
measured at different wavelengths
Spectroscopic Analysis
Spectroscopic analysis is a powerful and versatile technique used to study the
interaction between matter and electromagnetic radiation.
I. Quantitative Analysis
II. Qualitative Analysis
Quantitative Analysis
Quantitative analysis in spectroscopy aims to determine the concentration or
amount of specific substances in a sample. It involves measuring the intensity
of light absorbed or emitted by the sample and relating it to the concentration
of the analyte of interest using calibration curves or mathematical models.
A spectrometer is an optical instrument used to analyse light and other forms
of electromagnetic radiation.
• Light Source: A spectrometer starts with a light source that emits
electromagnetic radiation. This can be a lamp, a laser, or another type
of light emitter.
• Collimation: The emitted light is collimated, meaning it is converted
into a parallel beam by passing through a collimator. This ensures the
light travels in a straight line with consistent direction.
• Dispersion: The collimated light interacts with a dispersive element,
such as a prism or a diffraction grating. This element causes the
different wavelengths or frequencies of light to spread out or disperse
in different directions.
• Separation of Wavelengths: As the light is dispersed, the various
wavelengths or frequencies become spatially separated. Each
wavelength corresponds to a specific colour or frequency of light.
• Detection: The dispersed light is detected by a detector, such as a
photodiode or a charge-coupled device (CCD). The detector measures
the intensity of light at different wavelengths and converts it into an
electrical signal.

Calibration Curves: These curves are generated by measuring the absorption

or emission intensities of standard solutions with known concentrations of the
analyte. The resulting curve serves as a reference to quantify the
concentration of the analyte in an unknown sample.
The quantitative determination of compounds by spectrometric techniques
relies on two fundamental laws: Beer's law and Lambert's law.
• Beer's Law: According to Beer’s Law the amount of light absorbed is
directly proportional to the concentration of absorbing solute in the
solution. Thus

• Lambert's Law: The Lambert’s law states that light absorbed by a

solution is directly proportional to the length of light path through the
solution Hence,

• Combining the above equations, we get

Qualitative analysis: Qualitative analysis using spectrometry involves

identifying the presence of specific substances or chemical components in a
sample based on their unique spectral patterns.
• Atomic or Molecular Type: Determining the atomic or molecular type
using spectroscopy involves identifying the specific building blocks
(atoms or molecules) present in a sample by analysing the unique
patterns of light they absorb or emit.
• Molecular structure: Determining molecular structure using
spectrometry involves analysing the unique patterns of light
absorption or emission to understand how atoms are connected within
a molecule.
• Molecular environment: Using spectrometry to understand the
molecular environment involves analysing how the surroundings of
atoms or molecules in a sample affect the way they absorb or emit
light.
Spectroscopic technique
1. UV-Vis Spectroscopy:
• Principle: Measures the absorption and transmission of UV and
visible light by molecules. The energy difference between electronic
energy levels causes absorption.
• Applications: Determines the concentration of coloured compounds in
solutions, analyses chemical reactions, and studies electronic
transitions in molecules. (Identification of compounds, Measurement
of enzyme activities)
2. Infrared (IR) Spectroscopy:
• Principle: Analyses molecular vibrations by measuring the absorption
of infrared light. Bonds between atoms vibrate at characteristic
frequencies.
• Applications: Identifies functional groups in organic compounds,
studies molecular structure, and analyses complex mixtures. (NIS is
widely used in the meat, dairy and seafood industries for proximate
analysis. This is aimed predominantly at determining fat / oil, protein
and moisture levels in the associated products, serving a grading and
quality assurance function.)
3. Raman Spectroscopy:
• Principle: Measures the scattering of light by molecules. Provides
information about molecular vibrations and energy levels.
• Applications: Identifies chemical bonds, functional groups, and
molecular structures, and is used in materials characterization.
4. Fluorescence Spectroscopy:
• Principle: Measures the emission of light by molecules after
excitation. Excited molecules emit light as they return to lower
energy states.
• Applications: Studies fluorescent molecules in biological systems,
detects trace amounts of substances, and probes molecular
interactions.
5. Nuclear Magnetic Resonance (NMR) Spectroscopy:
• Principle: Exploits the magnetic properties of atomic nuclei. Nuclei
absorb radiofrequency radiation in the presence of a magnetic field.
• Applications: Determines molecular connectivity, identifies
chemical environments, and elucidates molecular structures. (NMR
spectroscopy is employed in a wide range of food safety areas
broadly aimed at averting significant chemical, biological or
microbiological threats to the food chain.)
6. Mass Spectrometry:
• Principle: Measures the mass-to-charge ratio of ions generated from
molecules. Provides information about molecular composition and
fragmentation patterns.
• Applications: Identifies unknown compounds, quantifies isotopes,
studies molecular fragmentation, and analyses biomolecules.
Summery
1. Spectroscopy Overview:
- Spectroscopy is the science of analyzing how matter interacts with light.
- Different types of spectroscopy techniques exist, each with its own
principles and applications.

2. Energy States of Matter:

- Atoms and molecules have discrete energy levels, and their energy
transitions are quantized.
- Energy levels depend on atomic and molecular structures, leading to unique
spectral patterns.

3. Electronic Energy Levels:

- Electrons in atoms and molecules can jump between energy levels by
absorbing or emitting specific amounts of energy.
- These transitions create distinct spectral lines in absorption and emission
spectra.

4. Nuclear Energy Levels:

- Nuclei possess spin and magnetic properties, resulting in quantized
orientations in external magnetic fields.
- These properties give rise to nuclear magnetic resonance (NMR)
spectroscopy, used to study molecular environments.

5. Nature of Electromagnetic Radiation:

- Electromagnetic radiation encompasses a wide range of energies, from
radio waves to gamma rays.
- It travels in waves and exhibits particle-like behavior (photons).

6. Theory of Spectroscopy:
- Spectroscopy involves studying how atoms and molecules interact with
light, leading to absorption or emission of energy.
- Absorption occurs when energy matches the difference between energy
levels, and emission happens when excited states return to lower states.
7. Energy Level Transitions:
- Spectroscopy focuses on energy transitions between discrete levels in
atoms and molecules.
- These transitions lead to absorption and emission of specific wavelengths,
resulting in unique spectral patterns.

8. Absorption of Radiation:
- Absorption involves transferring energy from light to an atom or molecule,
causing it to enter an excited state.
- Energy absorbed matches the energy difference between energy levels.

9. Emission of Radiation:
- Emission occurs when excited atoms or molecules release energy in the
form of light as they return to lower energy states.
- This emitted light carries information about the substance's composition
and structure.

10. Non-Radiative Emission and Radiative Decay:

- Non-radiative emission involves energy release through processes other
than light emission.
- Radiative decay, on the other hand, involves the emission of photons.

11. Spectrometry Modes and Measurement:

- Spectroscopy provides various modes of measurement, including
transmission, absorption, and emission measurements.
- These modes offer insights into sample properties and composition.

12. Spectroscopic Analysis - Qualitative and Quantitative:

- Spectroscopy is used for both qualitative (identifying components) and
quantitative (measuring concentrations) analysis.
- Beer's law and Lambert's law are fundamental principles for quantitative
analysis.
13. Molecular Identification and Structure Determination:
- Spectroscopy helps identify atomic or molecular types based on unique
spectral patterns.
- Molecular structure determination involves analyzing patterns of
absorption and emission to understand how atoms are connected.

14. Molecular Environment Analysis:

- Spectroscopy reveals changes in the molecular environment by detecting
alterations in absorption or emission patterns.
- This can be used to monitor changes in complex systems like protein
folding or aggregation.

15. Spectroscopic Techniques:

- Various spectroscopic techniques include UV-Vis, IR, NMR, mass
spectrometry, fluorescence, Raman, X-ray, CD, and emission spectroscopy.
- Each technique has distinct principles and applications for studying
different aspects of substances.

Spectroscopy plays a crucial role in understanding the microscopic world and

is applied across scientific disciplines for a wide range of research, analysis,
and exploration.
 Topic
1 Spectroscopy and structural
Afzal sir
• Isolation and purification
• Elemental analysis
• Molecular formula
• Mass spectroscopy
• Functional groups
• Absorption spectroscopy
• Ultraviolet-visible spectroscopy
• Infrared spectroscopy
• NMR spectroscopy
• Summary
determination
Introduction
Spectroscopy is a powerful set of
techniques used in the field of chemistry
to study the interaction between matter
and electromagnetic radiation. It provides
valuable information about the structure,
composition, and properties of molecules
and materials. Spectroscopy plays a
crucial role in structural determination,
helping researchers identify the
arrangement of atoms in a molecule and
gain insights into its chemical and physical
characteristics.
Isolation and purification
Isolation and Purification in Chemistry:
Isolation and purification are vital
processes in chemistry for obtaining pure
substances from complex mixtures. These
processes are essential in various
applications, from drug discovery to
material science. Different techniques are
employed:
1. Filtration: Separates solids from
liquids or gases using a porous
medium.
2. Distillation: Separates components
based on boiling points through
vaporization and condensation.
3. Crystallization: Precipitates compounds by controlled evaporation of a
solution.
4. Chromatography: Separates compounds using differences in interactions
between phases.
5. Extraction: Transfers substances between phases based on solubility
differences.
6. Sublimation: Converts solid to vapor without a liquid phase.
7. Recrystallization: Dissolves and reprecipitates a compound for
purification.
8. Electrophoresis: Separates charged molecules in an electric field.
9. Centrifugation: Separates particles based on density or size through high-
speed spinning.

Elemental analysis
The empirical formula of a chemical compound represents the simplest
whole-number ratio of the atoms of different elements present in the
compound. It provides information about the relative proportions of the
elements but does not give the exact number of atoms or their arrangement
within the molecule.
1. Determine Mass or Percent Composition: You need information about the
mass or percent composition of each element in the compound.
2. Convert to Moles: Convert the masses or percentages of the elements to
moles. To do this, divide each mass by the molar mass of the respective
element.
3. Find the Simplest Ratio: Divide each element's moles by the smallest
number of moles obtained. This step ensures that you're working with
whole-number ratios.
4. Write the Empirical Formula: Write the subscripts for each element based
on the ratio of moles obtained in the previous step. The subscripts should
be in their simplest whole-number form.

• A compound returns the following analysis: What is its empirical formula?

Assume
This gives a ratio of C:H:O is 2:4:1 the emperical formula C 2H4O1

Molecular formula
• Molecular Formula vs. Empirical Formula: Molecular formula shows the
actual number of atoms of each element in a compound, whereas the
empirical formula represents the simplest whole-number ratio of elements.
• Example: Benzene (C6H6) and acetylene (C2H2) share the empirical
formula CH but have different molecular masses and molecular formulas.
• Mass Spectrometry: This technique determines the molecular mass of a
compound. It involves:
• Vaporizing a small sample of the compound under low pressure and
high temperature.
• Irradiating the vapor with high-energy electrons (e.g., 4000-6000
kJ/mol).
• Resulting in electron ejection from molecules, creating positively
charged cations (ions).
• The molecular ion (parent ion) represents the compound's molecular
mass.

• The molecular ions (M+.) may break down into fragment ions or
daughter ions (m+).

The cations are accelerated through an electric field into a magnetic field
where they are deflected.
The ions are characterised by their mass (m) to charge (z) ratio (m/z). The
charge is always = 1.
• Molecular Ion Peak: Represents the intact molecular ion, allowing
determination of the compound's molecular weight.
• Fragmentation Patterns: In EI and some other ionization methods, the
molecule can fragment into smaller ions. The resulting fragments reveal
structural information about the compound.
• Isotope Patterns: Different isotopes of elements can be distinguished based
on their mass differences. This helps confirm the presence of specific
elements in a compound.

Funtional group
Absorption spectroscopy
The energy (E) of electromagnetic radiation is directly proportional to its
frequency (), given by the equation , where h is Planck's constant. Higher
frequency radiation has higher energy.
The frequency () and wavelength () of light are related by the speed of light
(c), where .
1. Ultraviolet-Visible (UV-Vis) Spectroscopy:
• Principle: UV-Vis spectroscopy measures the absorption of UV and
visible light by molecules, providing information about electronic
transitions. Electron that are most easily promoted are those in
conjugated π bond.
• Applications: It's used to identify the presence of conjugated systems
in organic compounds, such as aromatic rings and double bonds.
Organic compound that contain multiple conjugated bond strongly
absorb ultraviolet-visible radiation.
• Example:
2. Infrared (IR) Spectroscopy
• Principle: IR spectroscopy measures the absorption of IR radiation by
molecular vibrations, revealing information about functional groups
and bond types. Electromagnetic radiation in IR region of the
spectrum has the correct energy to cause bond in a molecule to stretch
and bend.
• Applications: It's widely used to identify and characterize functional
groups in organic compounds, making it valuable for structural
analysis.
3. Nuclear Magnetic Resonance (NMR) Spectroscopy:
• Principle: NMR spectroscopy involves the interaction of atomic
nuclei with magnetic fields and radiofrequency radiation. The nuclei
of some isotopes of many elements (H) absorb electro magnetic
radiation in radio frequency region when placed in a strong magnetic
field. The number of signals in NMR spectrum corresponds to the
distinct types of hydrogen atom in a molecule.
• Applications: NMR provides information about the arrangement of
atoms within molecules, making it essential for structural
determination and identifying functional groups
• Example:
Chemical shift: The frequncy axis of the spectrum is callibrated with a scale
called 𝛿 scale. The 𝛿 scale is callibrated with respect to a reference
compound, tetramethylsilane (). The friquency difference between the signal
arises from the sample and the TMS signal is called the chemical shift.
Splitting: The signals in 1H NMR frequently show fine structure (more than
one peak) which termed as splitting. The splitting pattern of 1H NMR arises
from the hydrogen atoms attached to neighbouring carbon atoms. If the
hydrogen atoms number is “n” then splitting pattern will be “n+1”
 Topic
2 Refractometry and polarimetry
Afzal sir

• Basics of Refraction Basic of refraction

• Index of Refraction
• Snell's Law
Index of Refraction
• Critical Angle The index of refraction (n) is a fundamental
• Refractometers optical property of a material that describes
• Principle of Operation
how much the speed of light is reduced when it
• The Abbe
refractometer enters the material. The index of refraction (n)
is calculated using the following formula:

• "n" is the index of refraction.

• "c" is the speed of light in a vacuum,
approximately meters per second.
• "v" is the speed of light in the given
material.

Snell's Law
Snell's Law, also known as Snell–Descartes law or the
law of refraction, is a fundamental principle in optics
that describes how light rays change direction when
they pass from one medium into another with a
different refractive index.

• n₁ is the refractive index of the initial medium (the

medium the light is coming from).
• is the angle of incidence, which is the angle between the
incident light ray and the normal (a line perpendicular to
the surface) at the point of incidence in the initial
medium.
• n₂ is the refractive index of the second medium (the
medium the light is entering).
• is the angle of refraction, which is the angle between the
refracted light ray and the normal at the point of
refraction in the second medium.
Critical Angle
The critical angle (usually denoted as ) is the angle of incidence at which the
angle of refraction () becomes degrees. It is a specific angle for a given pair
of media with different refractive indices.
Refractometers
Principles of Interferometry(Measurement of RI)
Interference: Interferometers rely on the principle of interference, where two
or more coherent waves overlap. Interference can result in constructive
interference (waves reinforce each other, creating bright regions) or
destructive interference (waves cancel each other out, creating dark regions).
Path Difference: The key parameter measured in interferometry is the path
difference () between the reference and sample beams. This path difference
determines how the waves interfere when they recombine. It is often adjusted
or controlled to investigate different aspects of the sample.
Fringe Pattern: The interference pattern produced by the recombined beams is
called a fringe pattern or interferogram. The pattern can be visualized as
alternating bright and dark regions, and its characteristics provide information
about the sample's properties.
The Abbe refractometer
1. Prism: The Abbe refractometer typically consists of a glass prism with a
flat, polished surface. The sample to be measured is placed on this surface.
2. Illumination System: An illumination system, often consisting of a light
source, a condenser lens, and a collimator, is used to project a well-
defined beam of light through the prism and onto the sample.
The operating principle of the Abbe refractometer involves measuring the
critical angle of total internal reflection (TIR) at the prism-sample interface.
Here's how it works:
• A beam of monochromatic light (usually sodium or lithium light) is
directed through the prism and onto the sample.
• The angle of incidence is gradually increased until the critical angle of
TIR is reached at the prism-sample interface.
• At the critical angle, the light is internally reflected within the prism and
does not penetrate the sample. This critical angle is related to the
refractive index of the sample.
• The operator observes the interference pattern (usually a dark field)
created at the critical angle. The position of this pattern is used to
determine the refractive index of the sample.
• The refractive index reading can be converted to various scales, including
the Brix scale for sugar solutions or specific scales for specific
applications.
Afzal sir

4
Rheological Properties of Foods

Introduction
• Introduction to Rheology Rheology is the study of how materials
• Flow of Material
deform and flow in response to applied
• Newton’s Law of Viscosity
• Viscous Fluids forces. In the context of food science,
• Newtonian Fluids rheological properties are essential for
• Non-Newtonian understanding the texture, consistency, and
Fluids overall sensory attributes of food products.
• Plastic Fluids Flow of Material
• Bingham Plastic 1. Newton’s Law of viscosity
Fluids Newton's Law of viscosity describes
• Non-Bingham
the relationship between shear stress and
Plastic Fluids
• Time Dependency velocity gradient in a fluid. It states:
• Viscosity Measurement
• Capillary Flow • The rate of deformation (and thus the
Viscometers velocity gradient) is directly
• Rotational Viscometers proportional to the shear stress 𝜏,
• Viscoelastic Behaviours Viscous Fluids
• Stress Relaxation Test Viscous fluids tend to deform
• Creep Test
continuously under the effect of an applied
• Mechanical Models
• Maxwell Model stress. They can be categorized as
• Kelvin-Voigt Model Newtonian or non-Newtonian fluids.
• Burger Model 1. Newtonian fluid
• Newtonian fluids are those that follow
Newton's law of viscosity.
• According to Newton's law of viscosity,
the relationship between shear stress
and shear rate is linear, and the
viscosity remains constant and
independent of the shear rate.
• Examples of Newtonian fluids include gases, oils, water, and most liquids
that contain over 90% water, such as tea, coffee, beer, carbonated
beverages, fruit juices, and milk.

1. Non-Newtonian fluids
Fluids that do not follow Newton’s law of viscosity are known as non-
Newtonian fluids. Shear thinning or shear thickening fluids obey the power
law model (Ostwald-de Waele equation):

k = the consistency coefficient (Pa · sn), n = flow behaviour index.

I. Shear Thinning (Pseudoplastic) Fluids.
• Shear thinning fluids are characterized by a decrease in viscosity as
the shear rate (rate of deformation) increases. This behaviour occurs
when friction between layers of the fluid decreases due to shearing
forces.
• Long-chain molecules in these fluids become unentangled and align
with the direction of flow under shear stress, reducing the fluid's
resistance to flow.
• Pseudoplastic behaviour is common in food systems. Products like
applesauce, banana puree, and concentrated fruit juices exhibit shear
thinning behaviour.
• For shear thinning (pseudoplastic) fluids n < 1,
II. Shear Thickening Fluids
• Shear thickening fluids, also known as dilatant fluids when
accompanied by volume expansion, are characterized by an increase
in internal friction and apparent viscosity as the shear rate (rate of
deformation) increases.
• As the shear rate rises, these fluids resist flow more strongly,
requiring greater force for movement.
• In food systems, corn starch suspension is provided as an example
of shear thickening fluids.
• For shear thickening fluids n > 1
Plastic Fluids
1. Bingham Plastic Fluids
A plastic fluid, also known as a yield-stress fluid, is a type of non-Newtonian
fluid that behaves as a solid below a certain critical shear stress (yield stress)
but flows like a liquid when the shear stress exceeds that critical value.
• Bingham plastic fluids are characterized by the fact that they remain
rigid (do not flow) when the applied shear stress is smaller than a
specific yield stress ().
• However, when the shear stress exceeds the yield stress (), these
fluids behave like Newtonian fluids and flow normally.
• In food systems, mayonnaise, tomato paste, and ketchup are
examples of Bingham plastic fluids.
II. Non-Bingham Plastic Fluids
• Non-Bingham plastic fluids also require a minimum shear stress
(yield stress) to be exceeded before they begin to flow, similar to
Bingham plastic fluids.
• However, the relationship between shear stress and shear rate is not
linear for these fluids. They may exhibit either shear thinning or shear
thickening behaviour in addition to having a yield stress.

A common mathematical model may be used to express the non-Newtonian

characteristics. This model is referred to as the Herschel–Bulkley model
(Herschel and Bulkley, ):
Time Dependency
1. Thixotropic Fluids
• Thixotropic fluids are characterized by a decrease in shear stress and
apparent viscosity with respect to time when subjected to a constant
shear rate. In other words, these fluids become thinner as they are
sheared over time.
• This behaviour is often attributed to the breakdown of the material's
internal structure as shearing continues.
• Examples of thixotropic fluids include gelatine, egg white, and
shortening.
• Thixotropic behaviour can exhibit varying degrees of reversibility. It
may be fully reversible, partially reversible, or irreversible (termed
rheomalaxis or rheodestruction) when the applied shear is removed,
and the fluid is allowed to rest.
Hysteresis Loop:
• Thixotropic behaviour can be studied by measuring shear stress as a
function of shear rate while increasing and then decreasing the shear
rate. This typically results in the observation of a hysteresis loop in
the shear stress versus shear rate curve.
• The hysteresis loop represents the difference in behaviour between
increasing and decreasing shear rates, highlighting the thixotropic
nature of the fluid.
2. Rheopectic Fluids
• Rheopectic fluids exhibit an increase in shear stress and apparent
viscosity over time when subjected to a constant shear rate. In other
words, their internal structure builds up and thickens as shearing
continues.
• This behaviours is relatively rare in food systems but can be observed
in materials like bentonite-clay suspensions.
Viscoelastic Behaviour
When a force is applied to a material, it can exhibit different responses based
on its rheological properties.
• Viscous fluids, when subjected to a force, deform continuously and
their deformation is proportional to the magnitude of the applied
force. They do not return to their original position when the force is
removed. This behaviour is characteristic of viscosity.
• Solids, on the other hand, exhibit elasticity, meaning they deform
under a force but return to their original shape when the force is
removed.
• Some materials, including certain foods, exhibit both viscous and
elastic properties, and these materials are known as viscoelastic
materials.
1. Stress Relaxation Test
If food materials are deformed to a fixed strain and the strain is held
constant, the stress required to maintain this strain decreases with time. This
is called stress relaxation. In this test, stress is measured as a function of time
as the material is subjected to a constant strain.
2. Creep Test
If a constant load is applied to biological materials and if stresses are
relatively large, the material will continue to deform with time. This is known
as creep. In a creep test, an instantaneous constant stress is applied to the
material and the resulting strain is measured as a function of time. There is a
possibility of some recovery of the material when the stress is released as the
material tries to return to its original shape.
Mechanical model
To understand the rheological behaviour of viscoelastic foods, some
mechanical models are used. These models consist of springs and dashpots.
Elastic (Spring) Model

Viscous (Dashpot) Model

1. Maxwell Model
The Maxwell model has been used to interpret stress relaxation of viscoelastic
liquids, especially polymeric liquid. Since the arrangement is a series
arrangement in Maxwell model

2. Kelvin-Voigt Model
Creep behaviour can be described by the Kelvin-Voigt model. This model
contains a spring and a dashpot connected in parallel

3. Burger Model
It is the series combination of the Kelvin and Maxwell models
Afzal sir

•
5
Texture of food
• Compression
Texture of foods
Introduction
The texture of foods is a crucial aspect
of the overall eating experience and can
• Snapping-Bending greatly influence our perception of flavour
• Cutting Shear and enjoyment. Food texture refers to the
• Penetration physical properties of a food item as it is
• Penetration perceived through touch and mouthfeel.
• Texture Profile Analysis 1. Compression
Compression (deformation) test measures
the distance that a food is compressed
under a standard compression force or the
force required to compress a food a
standard distance. This test can be
compared to the squeezing of bread by the
consumers to be sure that bread is fresh.
The sensory description of this test is
softness or firmness.
• In this test, a Universal Testing machine
fitted with 36 mm diameter of
aluminium plunger with cross-head
speed of 100 mm/min and a chart speed
of 50 mm/min is used.
2. Snapping-Bending
This test measures the force required to
bend or snap brittle foods such as biscuits
or crackers. The sample is laid across two
vertical rails that support it in horizontal
position.
A third bar mounted above the sample and equidistant between the supporting
rails is lowered until the sample breaks and the force is measured. The
deflection at the centre of the beam produced by a given force is expressed as:

3. Cutting Shear
The Pea Tenderometer, which was introduced in , works via the principle of
cutting shear. It consists of a grid of blades rotated at constant speed through a
second grid of blades. As the peas are cut by the blades, the maximum force is
measured. This instrument is still used to determine the maturity of peas at
harvest.
4. Puncture
Puncture test measures the force required to push the probe into the food and
expressed as firmness or hardness of the product. It is used mostly for fruits,
gels, vegetables, and some dairy or meat products.
The puncture test is not widely used on cereal products since hard baked
products are susceptible to fracture when subjected to this test.
5. Penetration
Penetrometers were originally designed to measure the distance that a cone or
a needle sinks into a food such as margarine or mayonnaise under the force of
gravity for a standard time. This is a simple and relatively inexpensive
apparatus used for determination of spreadability of butter
Texture Profile Analysis
Texture Profile Analysis (TPA) compresses a bite-sized piece of food
(usually 1 cm cube) twice to simulate the chewing action of the teeth.
Compression is usually of the original length of the
sample. As a result of TPA, sensory properties such as gumminess,
cohesiveness, and so forth can be determined objectively. Texture analysers
are used to obtain texture profile analysis.
• Fracturability (brittleness) is defined as the force at the first
significant break in the first positive bite area.
• Hardness is defined as the peak force during the first compression
cycle.
• Cohesiveness is defined as the ratio of the second positive bite area to
the first positive bite area.
• Adhesiveness is defined as the negative force area for the first bite
representing the work required to pull the plunger away from the
food.
• Springiness (elasticity) is defined as the height to which the food
recovers during the time that elapses between the end of the first bite
and start of the second bite (distance or length of the compression
cycle during the second bite).
• Gumminess is the product of hardness and cohesiveness. In sensory
terms, it is the energy required to disintegrate a semisolid food so that
it is ready for swallowing.
• Chewiness is the product of gumminess and springiness. In sensory
terms, it is known as the energy required for chewing a solid food
until it is ready for swallowing.
Afzal sir Properties of Granular Foods,

6
Powder & Solid Foods

Density
Bulk density
• Density Bulk density is the mass of a material per
• Compression
• Snapping-Bending unit of volume, including the interior pores
• Cutting Shear that are filled with air.
• Penetration 1. Loose bulk density: measured after
• Penetration placing the product in the constant
• Texture Profile Analysis volume container without vibration.
2. Packed bulk density: measured after the
sample placed in the constant-volume
container has been vibrated until the
volume seems constant
3. Apparent Particle Density: This density
represents the density of the solid
particles within the granular product,
excluding the void spaces.
4. Void: Void is the ratio of the volume of
the empty spaces or gaps between
particles in a granular material to the
total volume occupied by the material.
5. Porosity: Porosity is the measure of how
much empty space is present in a
material, expressed as a ratio of the
volume of voids to the total volume of
the material. P. Where, ρt is the true
density of the air free solids making up
the product.
Particle size and size distribution:

Problem: Compare the arithmetic, surface and volume-surface mean diameter

for particles in a dry food product with the following distribution of sizes:

Flow of Food Powders

The manner in which granular foods or powders may flow into or out of
containers is of concern in processing plants.
Two common parameters used for this purpose are the angle of repose and the
angle of slide.
• The angle of repose is defined in the following way:

where H' is the height of a mound of a powder which forms as it flows from a
container directly above a horizontal surface, and S is the circumference of
the mound at its base.
• parameter determined by placing the powder on a horizontal plate and the
angle of the plate is changed until the powder slides from the plate.
Angle of internal friction

flow parameter for a powder is a function of the shear stress (τ) and the
normal stress (σn)
Flow rate of powder through an orifice in the bottom of a storage container:

mass flow rate of powder (w) is a function of the orifice diameter (D) and the
angle of repose (β), discharge coefficient (Cc), bulk density(ρB). The discharge
coefficient (Co) usually varies between 0.5 and 0.7.
Compressibility index and Hausner ratio:

Major problems in the flow of powders

Arching or bridging of the material above the opening. The minimum distance
between the two walls of the opening (DL) which will not result in bridging is
as follows:

Angle of internal friction (φ)

Afzal sir

•
6
Definition
• Optical Properties
Optical properties
1. Optical Properties: Optical properties refer
to material characteristics that result from
interactions with light. In the context of
foods, the primary optical property
• Snapping-Bending considered by consumers is colour, followed
• Cutting Shear by gloss, translucency or turbidity, and other
• Penetration related properties.
• Penetration 2. Gloss: Gloss refers to the specular reflection
• Texture Profile Analysis of light from a smooth, plain surface. It can
be quantitatively described by a gonio
photometric curve, which represents how the
intensity of reflected light changes with
different angles of incidence and viewing.
3. Colour Variability: The colour of a material
can vary depending on the type of
illumination it is exposed to. The same
material may appear differently under
different lighting conditions due to
variations in the wavelengths of light
emitted.
4. Translucency in Foods: Translucency is an
optical property used to describe how light
interacts with a material, especially in the
context of foods. In liquid foods, light that
passes through the material may scatter
randomly when interacting with suspended
particles. Translucency is distinct from
colour and is perceived as a sensory
attribute. Some food products, like cloudy
fruit and vegetable juices, exhibit
translucency, falling between opacity and
transparency on a scale
Factors Affecting Perceived Food Colour:
1. Spectral Composition of Light Source: The type of light illuminating the
food affects its perceived colour.
2. Chemical and Physical Characteristics of the Food: The food's inherent
properties, such as pigments and surface texture, impact its colour.
3. Spectral Sensitivity Properties of the Eye: Human vision responds
differently to various wavelengths of light.
Colour Analysis Systems:
Various colour analysis systems have been developed to standardize and
quantify colour measurement. Some commonly used systems include:
4. CIE (Commission Internationale de l'Eclairage): An international standard
that defines colour spaces and measurement techniques.
• The CIE system is based on the principle that any colour can be
matched by combining suitable proportions of three primary colours:
red, green, and blue.
• These primary colour combinations are known as tristimulus values
and are used to quantify colours.
• To define a specific colour in the CIE system, chromaticity
coordinates, typically denoted as x and y, are used.
• These coordinates represent the position of the colour on a
chromaticity diagram, which is a two-dimensional space describing
the hue and saturation of the colour.
5. Munsell: In the Munsell system, all colours are described using three
attributes: hue, lightness, and saturation (or purity).
• Hue: Hue represents the type of colour and is based on a scale of ten
hues distributed on a circumference, numbered from 1 to 10.
• Lightness: Lightness is a measure of how light or dark a colour
appears relative to the proportion of light emitted. It ranges from
black (0) to white (10) and is distributed on a perpendicular line.
• Saturation or Purity: Saturation or purity is associated with the
clearness or darkness of a colour's perception. It is measured on an
irregular scale, starting from 0 for central gray and extending to the
limit of purity achievable with available pigments in the Munsell
book of colour.
Topic
2
Mobarak sir Food analysis
• Food analysis 1. Food analysis
• Defination Introduction
• Importance/Reason of Food analysis is the process of evaluating
food analysis and examining food products to determine
• Types of sample that
their composition, quality, safety, and other
are analyzed
• Food properties that relevant characteristics.
are typically analyzed.
• Sample and sampling Importance of food analysis
• Defination • Food Safety: Perhaps the most critical
• Steps in sampling aspect of food analysis is ensuring the
• Selection of sampling safety of food products.
procedure • Quality Assurance: Food analysis helps
• Sampling plan
• Factors Affecting
maintain and improve the quality of
the Choice of food products. It ensures that food
Sampling Plans meets established quality standards,
• Sampling by adheres to industry regulations, and
Attributes and provides consistent sensory attributes
Sampling by (taste, texture, appearance, aroma) that
Variables consumers expect.
• Developing a • Nutritional Information: Food analysis
sampling plan***
• Sampling Techniques
provides accurate information about the
• Probablity sampling nutritional content of food
• Non probablity • Labelling Compliance: To inform
sampling. consumers accurately, food products
• Preparation of sample must have correct labelling, including
• Performing assay ingredient lists, nutritional facts, and
• Choice and Validity of allergen information.
A Method
• Criteria for Choice of
Food Analysis Methods
• Official/ References
Methods
• Product Development: Food analysis plays a vital role in developing new
food products by helping manufacturers adjust formulations to meet
desired nutritional profiles, taste preferences, and shelf life requirements.
• Shelf Life Assessment: Understanding the shelf life of food products
through stability testing and monitoring helps reduce food waste by
ensuring that products remain safe and of high quality for as long as
possible.
• Regulatory Compliance: Food analysis is essential for food producers and
manufacturers to meet the regulatory requirements and quality standards
set by government agencies and industry organizations.
• Consumer Confidence: Consumers expect safe, high-quality, and
accurately labelled food products. Food analysis helps build trust among
consumers by providing assurance that these expectations are met.
• Global Trade: In international food trade, food analysis is crucial for
compliance with import and export regulations, including the
determination of sanitary and phytosanitary measures.
• Health and Wellness: As dietary preferences and health trends evolve, food
analysis assists in developing and marketing healthier food products that
meet specific dietary requirements, such as low-sugar, low-sodium, or
organic options.

Types of sample that are analysed

• Raw materials: Do they meet your specifications? Do they meet required
legal specifications? Are they safe and authentic? Will a processing
parameter have to be modified because of any change in the composition
of raw materials? Are the quality and composition the same as for previous
deliveries? How does the material from a potential new supplier compare
to that from the current supplier?
• Process control samples:
• Finished product: Does it meet the legal requirements? What is the
nutritive value, so that label information can be developed? Or is the
nutritive value as specified on an existing label? Does it meet product
claim requirements?
• Competitor’s sample: What are its composition and characteristics? How
can we use this information to develop new products?
• Complaint sample: How do the composition and characteristics of a
complaint sample submitted by a customer differ from a sample with no
problems?

2. Sample and sampling

Defination
• Population: The population in food analysis refers to the entire group or
collection of similar food products that you are interested in studying or
evaluating.
• Sample:
1. A sample is a subset of the population that is selected for testing,
analysis, or evaluation.
2. In the case of food analysis, a sample would be a representative
portion of the food products (e.g., a few cans of the brand of canned
tomatoes) chosen from the entire population.
3. The goal of selecting a sample is to obtain enough information about
the population's characteristics without having to analyse every
single item, which could be impractical or expensive.
• Sampling: Sampling is the process of selecting and collecting a sample
from the population.
Steps in sampling
1. Clearly define the population.
2. Selection and obtaining of the gross sample.
3. Reduction to a laboratory size sample.

Types of sample/Sampling
• Zero sampling
• 100% sampling
• Spot cheking
• Statistical Samples
Selection of sampling procedure
Once sampling is conducted, a series of stepwise procedures – from sample
preparation, laboratory analysis, data processing, and interpretation – is
needed to obtain data from the samples.

Sampling plan
“A predetermined procedure for the selection, withdrawal, preservation,
transportation, and preparation of the portions to be removed from a lot as
samples”
• A sampling plan should be a well-organized document that establishes the
goals of the sampling plan, the factors to be measured, sampling point,
sampling procedure, frequency, size, personnel, preservation of the
samples, etc.

Factors Affecting the Choice of Sampling Plans

1. Purpose of the inspection
2. Nature of the product
3. Nature of the test method
4. Nature of the population being investigated

Sampling by Attributes and Sampling by Variables

Sampling plans are designed for examination of either attributes or variables
5. Atribute sampling: In attribute sampling, sampling is performed to decide
on the acceptability of a population based on whether the sample
possesses a certain characteristic or not. The result has a binary outcome
of either conforming or nonconforming.
2. Variable sampling: In variable sampling, sampling is performed to
estimate quantitatively the amount of a substance (e.g., protein content,
moisture content, etc.) or a characteristic (e.g., colour) on a continuous
scale.

Developing a sampling plan

• Selection of parameters, ranges and resolution
• Calculation of sample size
• Selection of sampling techniques/sampling scheme
• Data storage format
• Assign roles and responsibilities

Sample size
• Population Size (N): Total number of individuals or items in the population
you want to study.
• Confidence Interval (Margin of Error): Maximum acceptable difference
between the sample estimate and the true population value (often denoted
by "E"). Max deviation (error) from mean (±); If 90% likes sweet with ME
5%. , it means between 85 to 95% likes sweet.
• Confidence Level (Z): Desired level of confidence (e.g., 95% or 99%) that
the sample estimate will fall within a certain margin of error from the true
population value. How sure that the actual mean falls within confidence
interval? If CL 95%, liking of sweet within 85 to 95% will be 95% times.
• Standard Deviation: How much variance do you expect in your responses.
3. Sampling Techniques
4. Probablity sampling: Probability sampling plans prescribe the selection of
a sample from a population based on chance. It provides a statistically
sound basis for obtaining representative samples with elimination of
human bias.
• Simple Random Sampling: In simple random sampling, each member
of the population has an equal chance of being selected for the sample.
Simple random sampling requires that the number of units in the
population be known.
• Systematic Sampling: In systematic sampling, you select every nth
member from a list or sequence of the population. Systematic
sampling is used when a complete list of sample units is not available.
• Stratified Sampling: Stratified sampling involves dividing the
population into subgroups or strata based on certain characteristics or
attributes (stratification criteria). Samples are then independently
selected from each stratum using simple random sampling or another
method.
• Cluster sampling: In cluster sampling, the population is divided into
clusters or groups, often based on geographic proximity or some other
criterion. A random sample of clusters is selected, and then all
members within the selected clusters are included in the sample.
• Multistage Sampling: Multistage sampling combines two or more
sampling methods. It often starts with cluster sampling and proceeds
with additional stages of sampling within selected clusters. This
method is useful for large and diverse populations.
2. Non-Probability sampling: Non-probability sampling is a method of
selecting samples from a population in a way that does not ensure each
member of the population has a known and equal chance of being
included in the sample.
• Convenience Sampling: Convenience sampling involves selecting
samples that are readily available and convenient to the researcher.
May not be representative of the population.
• Judgmental or Purposive Sampling: In judgmental or purposive
sampling, the researcher selects specific individuals or items based on
their expertise or judgment about which samples are most relevant or
informative for the study.
• Quota Sampling: Quota sampling involves dividing the population
into subgroups or strata based on certain characteristics, and then the
researcher selects samples from each subgroup until a predetermined
quota is reached. Quota sampling is similar to stratified sampling but
does not involve random selection within each stratum, making it
non-probabilistic.
Sample preparation
The preparation of a sample in food analysis is a crucial step to ensure
accurate and reliable results.
• Consideration in sample preparation
• Samples type: dry, wet, fresh, processed
• Size reduction: based on dry or wet products
• Drying: Ovens, lyophilizes
• Sample size: Primary and secondary
• Concentration: Active materials
• Problems during sample preparation
• Preparing representative small samples from large samples
• Loss of plant materials
• Removal of extraneous material from plants without removal of plant
constituents
• Enzymatic changes before and during analyses
• Compositional changes during grinding
• Metal contamination during grinding
• Changes in unstable components
• Special preparation problems in analyses of oil seed materials
• Size Reduction Considerations (Grinding):
• Moist sample: To homogenize moist samples, bowl cutters, meat
mincers, tissue grinders, mortars and pestles, or blenders are used
• Dry sample: Mortars and pestles and mills are best for dry
samples.
• Enzymic and chemical treatment
• Enzyme inactivation
• Drying
• Storage
3. Perform the Assay
Choice and Validity of A Method
• Characteristics of the Method: Numerous methods often are available to
assay food samples for a specific characteristic or component.
• Objective of the Assay: Selection of a method depends largely on the
objective of the measurement.
• Consideration of Food Composition and Characteristics: The performance
of many analytical methods is affected by the food matrix
• Validity of the Method
Criteria for Choice of Food Analysis Methods
• Inherent properties Specificity/selectivity
• Precision
• Accuracy
• Applicability of method to laboratory Sample size
• Reagents
• Equipment
• Cost
• Usefulness
• Time required
• Reliability
• Need
• Personnel Safety
• Procedures
Official/ References Methods
• Official method: The choice of method for a specific characteristic or
component of a food sample is often made easier by the availability of
official methods.
• AOAC International (Association of the Official Analytical Chemists)
• American Oil Chemists’ Society (AOCS)
• American Association of Cereal Chemists (AACC)
• Other Endorsed Methods
• Standard Methods for the Examination of Dairy Products
• Standard Methods for the Examination of Water and Wastewater
• Food Chemical Codex
4. Calculation and Interpretation of Results
Estimating the Sample Size
Reliability of analysis(Precision calculation)
Precision and accuracy are tools to assess reliability of data. Precision is done
by controlling the confidence level. The confidence interval for a sample
mean is described by the following equation:

represents the maximum error (E) that is acceptable for a desired level of
confidence.
• A group of 10 foot surgery patients had a mean weight of 240 pounds. The
sample standard deviation was 25 pounds. Find a confidence interval for a
sample for the true mean weight of all foot surgery patients. Use a 95%
CL.

Calculation of SD
Problem: A sample of uncooked hamburger for percent moisture content four
times and obtained the following results: 64.53%, 64.45%, 65.10%, 64.78%.

Sample size

Deviation
Observed
from mean
moisture
Coefficient of variation
• How large is our SD compared to mean in percentage
• Small number indicates high precision or reproducibility
• Below 5% is acceptable.

Confidence Interval/Limit
• Desire degree of certainty or confidence have to select first. For large
sample(>30), z-value is used to calculate CI

• For small sample size (<30), t-value is used to calculate CI

• t-value has to be looked from t-Table.

Relative Deviation

Below 2% is acceptable
Useful for only two replications
Relative Average Deviation
• Given the following data on dry matter (88.62, 88.74, 89.20, 82.20),
determine the mean, standard deviation and coefficient of variation. Is the
precision for this set of data acceptable? Can you reject the value 82.20
since it seems to be different than the others? What is the 95% confidence
level you would expect your values to fall within if the test were repeated?
If the true value for dry matter is 89.40, what is the % relative error?

Mean and SD Observed

Deviation
from mean
moisture

Coefficient of variation

The precision for this set of data acceptable (Below 5% is acceptable)

Rejection of 82.20

Confidence interval
To calculate the 95% confidence interval for the mean, use the t-distribution
since the sample size is small (n = 4). The formula for the confidence interval
is:

The acceptable range is 85.52 to 88.86

Relative error
Mobarok sir

•
6
Introduction (Most
important 20%)
Chromatography
Introduction
Chromatography is a technique for
separating mixture to its component in
• Definition order to identify, purify, analyse and
• Terminology quantify the mixture or componenets.
• Principle of Terminology
chromatography 1. Mobile phase: It is a fluid that carries
• Types of the sample through the stationary
chromatography phase, allowing for the separation of
• Liquid chromatography compounds based on their interactions
• Ion exchange
with the stationary phase.
chromatography
• Basic Principles (Most 2. Stationary phase: It is a solid or liquid
important 20%) material that remain fixed in a column
• Ion Exchange Resins or a solid support. It serves as a surface
Overview onto which the sample compounds
• Structure of Ion interact during the chromatographic
Exchange Resins process.
• Capacity and
3. Partitioning: In liquid chromatography,
Selectivity
it refers to the distribution of
compounds between the mobile and
stationary phases. Compounds with
different partition coefficients will
separate differently.
Principle
Chromatography is a separation technique
based on the differential interaction of
compounds with stationary and mobile
phases.
Types of chromatography
Based on the mobile phase

Liquid chromatography
There are several liquid chromatography techniques applied in food analysis,
namely paper chromatography, thin layer chromatography (TLC) (both of
these techniques may be referred to as planar chromatography), and column
liquid chromatography, all of which involve a liquid mobile phase and either a
solid or a liquid stationary phase.
1. Paper chromatography: In paper chromatography the stationary phase and
the mobile phase are both liquid (partition chromatography). Paper
generally serves as a support for the liquid stationary phase. The dis-
solved sample is applied as a small spot or streak one half inch or more
from the edge of a strip. The dry strip is suspended in a closed container in
which the atmosphere is saturated with the developing solvent (mobile
phase), and the paper chromatogram is developed.
2. TLC: TLC is a chromatographic technique used to separate and analyse
non-volatile mixtures of compounds. Stationary Phase: In TLC, the
stationary phase is a thin layer of an adsorbent material, typically coated
onto a glass, plastic, or aluminium plate. The most commonly used
adsorbents include silica gel, alumina, or cellulose. Mobile Phase: The
mobile phase is a solvent or mixture of solvents that moves through the
stationary phase, carrying the sample mixture with it. The choice of the
mobile phase depends on the compounds being analysed and their
solubility characteristics.
• Separation and Visualization: During TLC, compounds in the
mixture are separated as spots on the TLC plate. These spots are
visualized, often using chemical staining or UV light.
Retention Factor (Rf): The retention factor (Rf) is used to quantify the
separation. It's calculated as the ratio of the distance travelled by the sample
spot to the distance travelled by the solvent front.

Gas chromatography
Gas Chromatography (GC) is a powerful analytical technique used to separate
and analyse volatile compounds in a mixture. Its principles are rooted in the
differential partitioning of compounds between a stationary phase and a
mobile phase, which allows for the separation of individual components based
on their chemical properties.
Mobile Phase:
• In GC, the mobile phase is a pure, inert gas, such as helium, nitrogen,
or hydrogen. This gas carries the sample through the chromatographic
column. The choice of the mobile phase affects the efficiency and
speed of separation.
Stationary Phase
• The chromatographic column is a long, coiled tube typically made of
stainless steel or glass. It is packed with a stationary phase material or
coated with it. The stationary phase can be a liquid (liquid phase) or a
solid adsorbent (solid phase).
The separation in GC is primarily based on the differential partitioning or
distribution of compounds between the mobile phase (inert gas) and the
stationary phase. Compounds that have a higher affinity for the stationary
phase spend more time interacting with it, leading to longer retention times.
Ion exchange chromatography
Basic Principles
The core principle of IEC is the reversible exchange of ions between the
analyte molecule in solution and the charged site on the stationary phase (Ion
exchange resin). Three types of separation may be achieved: (1) ionic from
non- ionic, (2) cationic from anionic, and (3) mixtures of similarly charged
species. Ion exchange is an adsorption phenomenon. Electrostatic forces hold
ions to charged functional groups on the surface of the ion exchange resin.
The adsorbed ions replace ions that are on the resin surface on a 1:1 charge
basis.
1. Ion Exchange Resin: The stationary phase typically consists of resin beads
with functional groups that are positively (cation-exchange resin) or
negatively (anion-exchange resin) charged. Cation-exchange resins bind
negatively charged ions (acid), while anion-exchange resins bind
positively charged ions (base).
2. Selectivity: The selectivity in ion exchange chromatography is determined
by the type and density of the functional groups on the resin as well as the
pH and ionic strength of the mobile phase.
3. Mobile Phase: The mobile phase, also known as the eluent, is an aqueous
buffer solution with a specific pH and ionic strength. It is responsible for
carrying the sample through the column and competing with the analytes
for binding sites on the resin.
4. Elution: Elution is the process of washing out the bound analytes from the
resin using a gradient of increasing ionic strength or changing pH. This
elution process leads to the separation of analytes based on their affinity
for the stationary phase.
5. Eluate: The term "eluate" is commonly used in chromatography and refers
to the liquid or solution that emerges from the chromatography column
after the sample has undergone separation.
Resins
Ion Exchange Resins Overview:
• Ion exchange resins are organic or inorganic polymers used to exchange
cations or anions from a solution.
• They consist of a polymer matrix and functional groups for ion exchange.
• Natural sources of ion exchange materials include proteins, soils, lignin,
coal, metal oxides, aluminosilicates, and clay.
• Synthetic ion exchange materials include zeolite gels and polymeric resins
(e.g., macroreticular resins).
Structure of Ion Exchange Resins
• Polymer matrix materials commonly include polystyrene (85%),
polyacrylate (10%), and phenol-formaldehyde.
• Functional groups determine the type of ion exchange (cation, anion, or
chelating).
Capacity and Selectivity:
• Capacity is the amount of exchangeable ions per unit material.
• Proton exchange capacity (PEC) measures H+ exchange ability.
Factor affecting ion exchanger
• Affinity: The strength of attraction between the resin and target ions affects
selectivity.
• Ionic Valence: The charge on ions (valence) impacts their binding to the
resin. Na+ < Ca 2+ < Al 3+ < Th 4+
• Ionic Radius: The size of ions relative to resin binding sites can influence
selectivity. Li+ < H+ < Na+
• Ion Concentration: The concentration of ions in the sample can saturate
binding sites and affect separation.
• Ionic and Permeable Material: The resin must be both ionic and permeable
to allow ion exchange.
• Temperature: Temperature affects ion exchange kinetics and can impact
selectivity.
• Buffer: Buffers control pH and ionic strength, influencing selectivity in the
mobile phase.
Mobarok sir

•
7
Introduction (Most
important 20%)
Electrophoresis
Introduction
Electrophoresis is a widely used technique
in analytical chemistry and molecular
• Definition biology. It is a method for separating
• Theory molecules, such as DNA, RNA, proteins,
• Principle of and other charged particles, based on their
electrophoresis size and charge. The basic principle behind
• Zone electrophoresis electrophoresis is the movement of charged
• Liquid chromatography particles in an electric field.
• Ion exchange 1. Charged Particle Movement: Charged
chromatography
• Basic Principles (Most
particles move in response to an
important 20%) electric field. Cations (positively
charged) move towards the cathode
(negatively charged electrode), while
anions (negatively charged) move
towards the anode (positively charged
electrode).
2. Gel or Matrix: Electrophoresis is
conducted in a gel or matrix (e.g.,
agarose for DNA, polyacrylamide for
proteins). The gel acts as a sieve,
separating molecules based on size as
smaller molecules migrate faster.
3. Buffer Solution: A buffer maintains a
stable pH during electrophoresis,
preventing pH changes that could
affect molecular charge and mobility.
4. Controlled Voltage and Current: A
power supply applies an electric field
across the gel. Voltage and current are
controlled to optimize separation
without damaging molecules.
5. Sample Loading: Molecules to be separated (e.g., DNA or proteins) are
loaded into wells or slots at one end of the gel. When the electric field is
applied, molecules move through the gel.
6. Separation: Molecules separate based on size and charge as they migrate
through the gel. Smaller molecules travel further, while larger ones move
more slowly.
7. Visualization: After electrophoresis, molecules are visualized using
staining (e.g., ethidium bromide for DNA) or immunoblotting (for
proteins) to enable analysis and quantification.
Zone electrophoresis
Zone electrophoresis refers to a category of electrophoretic techniques where
molecules are separated based on their mobility (size, shape, charge) through
a supporting medium, typically a gel or a liquid, as they migrate in response
to an electric field.
• Zone electrophoresis is a technique used to separate components based on
differences in net charge, size, and shape.
• This separation occurs at a constant pH and ionic strength, ensuring
consistency in the electrophoretic conditions.
• Stabilizing media are used to provide support for the molecules being
separated. Common choices include polyacrylamide gel, cellulose, or
granulated gels, depending on the specific application.
• During the process, a narrow zone of the sample is applied on top of the
stabilizing gel.
• When an electric field is applied, the sample components migrate into the
gel. The molecules move through the gel at different rates due to
variations in their charge, size, or shape.
• The elution chamber typically has a funnel-shaped design to facilitate the
collection of separated components.
• Separated components are flushed out by a continuous stream of elution
buffer, allowing for their further analysis or downstream applications.