CHAPTER 1: INTRODUCTION

1.1. Overview of Essential Oils

An essential oil is a concentrated hydrophobic liquid containing volatile
(easily evaporated at normal temperatures) chemical compounds from
plants. Essential oils are also known as volatile oils, ethereal oils,
aetheroleum, or simply as the oil of the plant from which they were
extracted, such as oil of clove.
They are commonly used in aromatherapy, perfumes, cosmetics, and as
flavourings in food and beverages. Essential oils are made up of a
complex mixture of natural chemicals, which can vary depending on the
plant species, growing conditions, and extraction method.
Essential oils have a wide range of chemical compositions and physical
properties. They can be colourless or have a pale yellow or green colour.
The viscosity of essential oils can range from watery to thick and syrupy.
Essential oils have strong, characteristic Odors that can vary from floral
and fruity to spicy and woody.
Essential oils have many potential health benefits, such as reducing stress
and anxiety, improving sleep quality, and boosting the immune system.
However, it is important to note that essential oils can also have potential
risks and side effects, particularly if used improperly or in large amounts.
The Essential Oil extraction method generally depends on the botanical
material used. It is one of the main factors determining Essential Oils
quality. An inappropriate extraction procedure can damage or alter the
chemical composition of EO. This results in a loss of bio-activity and
natural characteristics.
The principle of EO extraction is relatively straightforward. However, the
process chosen can have a significant effect on the yield and quality of
the distillate obtained. Various processes have therefore been developed

1
 to obtain maximum yields of EO with a chemical state as close as
possible to their native structure.

1.2. Importance of Extraction methods

The extraction of the EOs is the first important step in preparing these
compounds. Modern extraction methods are effective in the efficient
development of these compounds. Moreover, the compounds extracted
from plants have natural antimicrobial activity against many spoilage and
disease-causing bacteria.
The choice of extraction method profoundly shapes the quality and
characteristics of essential oils. Each method, whether hydro distillation,
cold-pressing, solvent extraction, or CO2 extraction, offers unique
advantages and considerations. From preserving delicate aromas to
ensuring purity and safety, the extraction process significantly impacts the
final product. Moreover, environmental sustainability and cost-
effectiveness are essential factors to consider in the selection of an
extraction method. Ultimately, understanding the intricacies of each
method and its suitability for different plant materials is key to producing
high-quality essential oils with desired therapeutic properties.

1.3. Objectives of the study

The objective of extracting volatile essential oils from plant materials
using hydro-distillation and solvent extraction methods is to isolate and
capture the aromatic compounds present in the plants and compare both
the methods on various factors such as effectiveness, versatility etc.
These essential oils often contain complex mixtures of volatile organic
compounds that contribute to the characteristic aroma and potential
therapeutic properties of the plants.

2
CHAPTER 2: EXTRACTION TECHNIQUES

2.1 Hydrodistillation
In order to isolate essential oils by hydrodistillation, the aromatic plant
material is packed in a still and a sufficient quantity of water is added and
brought to a boil; alternatively, live steam is injected into the plant
charge. Due to the influence of hot water and steam, the essential oil is
freed from the oil glands in the plant tissue. The vapor mixture of water
and oil is condensed by indirect cooling with water. From the condenser,
distillate flows into a separator, where oil separates automatically from
the distillate water.

3
2.1.1 Mechanism of Distillation
Hydrodistillation of plant material involves the following main
physicochemical processes:
2.1.1.1. Hydrodiffusion
Diffusion of essential oils and hot water through plant
membranes is known as hydrodiffusion. In steam distillation,
the steam does not actually penetrate the dry cell
membranes. Therefore, dry plant material can be exhausted
with dry steam only when all the volatile oil has been freed
from the oil-bearing cells by first thorough comminution of
the plant material.
2.1.1.2. Hydrolysis
Hydrolysis in the present context is defined as a chemical
reaction between water and certain constituents of essential
oils. Esters are constituents of essential oils and, in the
presence of water, especially at high temperatures, they tend
to react with water to form acids and alcohols.

( alcohol ) ×(acid)
K=
( ester ) ×(water )

where K is the equilibrium constant.

Therefore, if the amount of water is large, the amounts of
alcohol and acid will also be large, resulting in a decreased
yield of essential oil.

2.1.1.3. Decomposition by heat

Almost all constituents of essential oils are unstable at high
temperature. To obtain the best quality oil, distillation must
be done at low temperatures. The temperature in steam

4
 distillation is determined entirely by the operating pressure,
whereas in water distillation and in water and steam
distillation the operating pressure is usually atmospheric.

2.1.2 Three Types of Hydrodistillation

Three are three types of hydrodistillation for isolating essential oils
from plant materials:
2.1.2.1.1. Water distillation
In this method, the material is completely immersed in
water, which is boiled by applying heat by direct fire,
steam jacket, closed steam jacket, closed steam coil or
open steam coil. The main characteristic of this
process is that there is direct contact between boiling
water and plant material
2.1.2.1.2. Water and steam distillation
In water and steam distillation, the steam can be
generated either in a satellite boiler or within the still,
although separated from the plant material. Like water
distillation, water and steam distillation is widely used
in rural areas. Moreover, it does not require a great
deal more capital expenditure than water distillation.
Also, the equipment used is generally similar to that
used in water distillation, but the plant material is
supported above the boiling water on a perforated
grid.
2.1.2.1.3. Direct steam distillation
direct steam distillation is the process of distilling
plant material with steam generated outside the still in
a satellite steam generator generally referred to as a

5
 boiler. As in water and steam distillation, the plant
material is supported on a perforated grid above the
steam inlet. A real advantage of satellite steam
generation is that the amount of steam can be readily
controlled.

2.2. Soxhlet Extraction

Soxhlet extraction is one of the most popular techniques for extraction of

analytes from solid materials. Since its discovery in 1879, the standard
Soxhlet technique has been routinely applied in almost every analytical
laboratory. Up to this day, Soxhlet extraction technique remains a
standard technique to which the performance of modern extraction
techniques is compared.
SE is the most conventional method used to extract phenolics and other
secondary metabolites from plants. In SE, phenolics are extracted from
the plant biomass by means of solubilization in the solvent. The SE
technique is very simple and requires a Soxhlet unit, a thimble (normally
made from cellulose) to contain the sample, and a percolator to circulate
solvent through the sample (a circulating water bath connected to the
Soxhlet unit and a heating mantle). An advantage of using SE is that a
filtration step to recover the solvent containing extracted phenolics is not
required because the plant residue is contained in the thimble and not
mixed in directly with the solvent. Normally the SE process is carried
over a 2–12 h period, commonly using a 40%–60% ethanol or methanol
in water solution as solvent at reflux. Other solvents such as acetone,
acetonitrile, or pure water have been used as solvent, but a solution made
using ethanol or methanol in water is more common.

6
2.3. Eutectic Solvent Extraction

Extraction using deep eutectic solvents (DES) is an innovative approach

in green chemistry. These solvents are composed of a eutectic mixture of
two or more components, typically a hydrogen bond acceptor (HBA) and
a hydrogen bond donor (HBD), which form a low-melting-point liquid.
DES offer several advantages over traditional organic solvents, such as
being environmentally friendly, biodegradable, and non-toxic.

7
the extraction process using DES:
2.3.1. Selection of Components: The first step is to select appropriate
components to form the deep eutectic solvent. These
components should have suitable properties for the intended
extraction process.
2.3.2. Preparation of DES: The HBA and HBD are mixed in a specific
ratio and heated until they form a homogeneous liquid. This
mixture typically forms a eutectic point, where the melting point
of the resulting mixture is lower than that of the individual
components.
2.3.3. Extraction Process: The DES is then used as a solvent to extract
desired compounds from a solid or liquid matrix. This can be
done through techniques such as liquid-liquid extraction, solid-
phase extraction, or other extraction methods.
2.3.4. Separation: After extraction, the DES containing the desired
compounds is separated from the matrix. This can be achieved
through methods like filtration or centrifugation.
2.3.5. Recovery of Compounds: The extracted compounds are then
recovered from the DES. This can be done by evaporating the
solvent under vacuum or through other separation techniques.
2.3.6. Reuse of DES: One of the advantages of DES is that they can
often be reused multiple times, making the process more
sustainable.
Extraction using DES has been applied in various fields such as
pharmaceuticals, food processing, and environmental remediation,
offering a promising alternative to traditional solvent extraction methods.
Its green credentials make it particularly attractive in industries where
sustainability is a priority.

8
9
CHAPTER 3: EXPERIMENTAL WORK

3.1. Methods and materials

3.1.1. Chemicals and reagents: cinnamon powder, Folin-Denis

reagent, sodium carbonate, methanol, hexane, distilled water,
silica gel and visible chromatography.
3.1.2. Plant material: Bark of cinnamon was collected from the local
market at Mumbai. The bark was powdered and used as a raw
material and stored in an air tight container.
3.1.3. Visible spectrophotometry: In visible spectrophotometry,
Solutions of different compositions were prepared using a stock
solution. A wavelength of 375 nm was selected to check the
absorbance of solutions with different compositions. Methanol
was used for the baseline correction using a quartz cuvette.

3.2. Hydrodistillation

100g of dried cinnamon was crushed and placed in the 1000 ml round
bottom flask with 500-600 ml of distilled water. The temperature was
adjusted to 100 ֯ C. The condensate was collected in a funnel. As there
was a mixture of water and different types of oils in the condensate, 1,2-
dichloroethane was added to the condensate to separate the oil and water.
The oils present in the condensate varies in density thus forms layers in
the mixture. A syringe was used to suck the water from the 1,2
dichloroethane and condensate mixture. the 1,2-dichloroethane and oil
was separated using simple distillation.

10
11
3.3. Soxhlet extraction

10g of cinnamon was crushed and placed in a Soxhlet extraction

apparatus with hexane.
(Precaution: at least 2/3 of the volume of solvent should be used to the
capacity of the round bottom flask, this is to prevent the solvent from
drying up before extraction is achieved.)
The solution was gently refluxed for 2-4 hours and allowed to cool to
room temperature. The extract was mixed with ethanol for further
separation of solvents. The content was allowed to rest for 48 hours for
liquid -liquid separation to take place. The liquid was then taken to a
water bath where the temperature was adjusted to 65 ֯ C. the remaining
oily liquid in the flask was the essential oil which was later used to plot
absorbance vs concentration graph.

12
3.4. Extraction using eutectic solvents

3.4.1. Preparation of DES (Deep eutectic solvents): The preparation of

DES was carried out by mixing choline chloride and glycerol at
ratios of 1:2. The required amount of the individual components
according to their molar ratio and molecular weight was
measured using and analytical balance. The two components

13
 were heated on a hot plate with magnetic stirrer at 80 ֯C until a
homogeneous transparent liquid was formed.

3.4.2. 1g of crushed cinnamon was mixed with 10 ml of DES. It was

allowed to rest for 1-2 hours. The mixture was then centrifuged
for 4 hours.

3.5. Preparation of absorbance vs concentration graph of extracted oil

(Assumption: oil extracted from Soxhlet apparatus is pure)

Two solutions were prepared. 0.1g of oil in 10 ml of methanol (stock
solution) and 20% w/v of sodium carbonate in water. The sodium

14
carbonate solution was prepared by heating 100 ml of water on hot plate
and adding sodium carbonate intermittently to avoid forming lumps.
Preparation of 5 test tubes with following compositions were
implemented.
3.5.1. 100 µl of stock solution with 900 µl of methanol
3.5.2. 200 µl of stock solution with 800 µl of methanol
3.5.3. 300 µl of stock solution with 700 µl of methanol
3.5.4. 400 µl of stock solution with 600 µl of methanol
3.5.5. 500 µl of stock solution with 500 µl of methanol

5 µl of Folin reagent and 2 ml of sodium carbonate solution was added in

each test tube.

The spectrophotometer was set with a wavelength of 375 nm to check and

note the absorbance of solutions with above compositions.

15
CHAPTER 4: COMPARITIVE ANALYSIS OF EXTRACTION

METHODS.

4.1. Yield Comparison:

4.1.1. Hydro Distillation: Hydro distillation can provide moderate to
high yields of essential oils, depending on factors such as the
plant material, duration of distillation, and equipment used.
However, it may not be as efficient in extracting all volatile
compounds compared to other methods.
Yield from hydrodistillation was found to be 2.81g
4.1.2. Soxhlet Extraction: Soxhlet extraction typically yields higher
amounts of essential oils compared to hydro distillation due to
the continuous extraction process and the ability to continuously
replenish the solvent, leading to better extraction efficiency.
Yield from Soxhlet extraction was found to be 0.13g
4.1.3. Deep Eutectic Solvent Extraction: Deep eutectic solvent
extraction can offer competitive yields similar to Soxhlet
extraction due to its ability to dissolve a wide range of
compounds and its mild extraction conditions.
Yield from deep eutectic solvent extraction couldn’t be
calculated as the research of separation of oil from eutectic
solvent is still ongoing.

4.2. Quality of Extracts:

4.2.1. Hydro Distillation: Hydro distillation may lead to the loss or
degradation of certain delicate compounds due to prolonged
exposure to heat and water. However, it can preserve certain
heat-stable compounds better than other methods.

16
 4.2.2. Soxhlet Extraction: Soxhlet extraction may result in extracts
with a broad spectrum of compounds, but it could also lead to
the degradation of heat-sensitive compounds due to prolonged
exposure to high temperatures.
4.2.3. Deep Eutectic Solvent Extraction: DES extraction is generally
considered gentle and mild, making it suitable for preserving
delicate compounds and maintaining the quality of essential
oils.

4.3. Economic Considerations:

4.3.1. Hydro Distillation: Hydro distillation may have lower
equipment and operational costs compared to Soxhlet extraction
or DES extraction due to its simplicity and minimal need for
specialized equipment.
4.3.2. Soxhlet Extraction: Soxhlet extraction typically requires more
complex equipment and consumes larger amounts of solvents,
leading to higher operational costs. However, it may provide
higher yields, which can be economically beneficial for certain
applications.
4.3.3. Deep Eutectic Solvent Extraction: DES extraction may initially
involve higher costs due to the synthesis or purchase of DESs,
but it can offer advantages such as reduced solvent consumption
and potentially higher extraction efficiency, which could lead to
long-term cost savings.

4.4. Suitability of Raw Materials:

4.4.1. Hydro Distillation: Hydro distillation is suitable for a wide
range of plant materials, including flowers, leaves, seeds, and
roots. However, it may not be suitable for heat-sensitive or

17
 delicate materials that are prone to degradation at high
temperatures.
4.4.2. Soxhlet Extraction: Soxhlet extraction is versatile and can be
used for various types of plant materials, including both heat-
sensitive and heat-stable compounds. It is particularly effective
for extracting compounds with moderate to high solubility in
organic solvents.
4.4.3. Deep Eutectic Solvent Extraction: DES extraction can be
tailored to suit different types of raw materials by adjusting the
composition of the solvent mixture. It is suitable for both
hydrophilic and hydrophobic compounds and can be optimized
for specific extraction targets.

4.5. Scalability:
4.5.1. Hydro Distillation: Hydro distillation can be easily scaled up for
industrial production, as it involves relatively simple equipment
and processes. However, scaling up may require larger boilers
and condensers to accommodate higher volumes of plant
material and steam.
4.5.2. Soxhlet Extraction: Soxhlet extraction can also be scaled up for
industrial production, but it may require more complex and
expensive equipment compared to hydro distillation.
Additionally, larger quantities of solvents will be needed for
extraction, which can increase costs.
4.5.3. Deep Eutectic Solvent Extraction: DES extraction can be scaled
up for industrial production similar to Soxhlet extraction.
However, it may require optimization of DES formulations and
extraction conditions to maintain efficiency at larger scales.

18
4.6. Residue Production:
4.6.1. Hydro Distillation: Hydro distillation typically produces plant
residues or hydrosols, which may contain water-soluble
compounds and essential oil residues. These residues can be
further processed or utilized for other purposes, such as herbal
teas or animal feed additives.
4.6.2. Soxhlet Extraction: Soxhlet extraction generates a significant
amount of spent solvent, which may require proper disposal or
recycling to minimize environmental impact. Additionally, the
extracted plant material may still contain some residual solvent,
depending on the efficiency of the extraction process.
4.6.3. Deep Eutectic Solvent Extraction: DES extraction produces
minimal residue compared to Soxhlet extraction since DESs are
often biodegradable and can be easily separated from the
extracted compounds. However, the environmental impact of
residue disposal should still be considered.
4.7. Applications
4.7.1. Hydrodistillation:
4.7.1.1. Perfume and Aromatherapy Industries:
Hydrodistillation is widely used to extract essential oils
from aromatic plants such as lavender, rosemary, and
peppermint. These essential oils are then used in
perfumes, cosmetics, and aromatherapy products.
4.7.1.2. Food and Beverage Industry: Essential oils extracted
via hydrodistillation are used to flavor various food and
beverage products, including teas, candies, and baked
goods.
4.7.2. Soxhlet Extraction:

19
 4.7.2.1. Pharmaceutical Industry: Soxhlet extraction is
employed to extract medicinal compounds from plant
material for pharmaceutical purposes. For example,
active pharmaceutical ingredients (APIs) can be
extracted from medicinal herbs for use in drug
formulations.
4.7.2.2. Environmental Analysis: Soxhlet extraction is used in
environmental analysis to extract pollutants and
contaminants from solid samples such as soil,
sediment, and sludge for further analysis.
4.7.3. Extraction by Eutectic Solvents:
4.7.3.1. Green Chemistry and Sustainable Processes: Eutectic
solvents offer a sustainable alternative to traditional
organic solvents in various applications.
4.7.3.2. Phytochemical Extraction: Eutectic solvents can be
used to extract phytochemicals and bioactive
compounds from natural sources, including medicinal
plants, for use in pharmaceuticals, nutraceuticals, and
functional foods.
4.7.3.3. Extractive Metallurgy: Eutectic solvents are explored
for their potential in extractive metallurgy processes,
such as the extraction of metals from ores, providing a
more environmentally friendly alternative to
conventional methods.

In summary, the choice between hydro distillation, Soxhlet extraction, and deep
eutectic solvent extraction for extracting essential oils depends on various
factors mentioned above. Each method has its advantages and limitations, and

20
the selection should be based on the specific needs and priorities of the
extraction process.

21
CHAPTER 5: CONCLUSION AND FUTURE PROSPECTS.

5.1. Conclusion:

5.1.1. Hydrodistillation: It's a traditional and widely used method for

extracting essential oils from aromatic plants. By heating water
and plant material, volatile compounds are released and
condensed to obtain the essential oil. Hydrodistillation offers
simplicity and effectiveness but may sometimes result in
degradation of heat-sensitive compounds due to prolonged
exposure to high temperatures.
5.1.2. Soxhlet Extraction: This method involves continuous extraction
using a solvent, typically ethanol or hexane. The solvent is
heated and continuously circulated through the plant material,
extracting essential oils and other compounds. Soxhlet
extraction is efficient in extracting a wide range of compounds,
but it requires a significant amount of time and solvent, and it
may lead to the extraction of non-volatile compounds as well.
5.1.3. Deep Eutectic Solvent (DES) Extraction: DES is a relatively
new and promising approach for extracting essential oils. DESs
are formed by mixing two or more components to create a
eutectic mixture with a lower melting point than any of the
individual components. DES extraction offers several
advantages, including milder extraction conditions, reduced
environmental impact, and potential selectivity for specific
compounds. However, further research is needed to optimize
DES formulations and extraction parameters for different plant
materials.
5.2. Future Prospects:

22
Future research should focus on optimizing extraction parameters such as
temperature, pressure, solvent composition, and extraction time to
improve the efficiency and selectivity of each method. This optimization
can enhance the yield and quality of essential oils while minimizing
energy consumption and environmental impact. There is a growing
interest in developing green extraction techniques that are more
sustainable and environmentally friendly. Methods such as microwave-
assisted extraction, ultrasound-assisted extraction, and supercritical fluid
extraction show potential for improving extraction efficiency and
reducing solvent usage and processing time. In-depth chemical
characterization of essential oil extracts is essential for understanding
their composition, bioactivity, and potential applications. Advanced
analytical techniques such as gas chromatography-mass spectrometry
(GC-MS), liquid chromatography-mass spectrometry (LC-MS), and
nuclear magnetic resonance (NMR) spectroscopy can provide valuable
insights into the chemical profile of essential oils and their functional
properties. Researchers can explore novel plant sources for essential oil
extraction, including underutilized or endemic plant species, agricultural
by-products, and waste materials. This exploration can not only diversify
the range of available essential oils but also contribute to sustainable
agriculture and waste valorisation efforts. Essential oils extracted using
different methods have numerous applications across industries such as
cosmetics, pharmaceuticals, food and beverages, and aromatherapy.
Future research should focus on exploring new applications and
formulations for essential oils to meet the evolving needs of consumers
and industries.

23
REFRENCES

1. Applications of Essential Oils and Plant Extracts in Different Industries.

Parisa Bolouri,1,2 Robab Salami,3 Shaghayegh Kouhi,4 Masoumeh Kordi,3
Behnam Asgari Lajayer,5,* Javad Hadian,6 and Tess Astatkie7,*
2. Soxhlet Extraction versus Hydrodistillation Using the Clevenger Apparatus:
A Comparative Study on the Extraction of a Volatile Compound from
Tamarindus indica Seeds. Kehinde Oluwakemi Fagbemi,corresponding
author Daniel Ayandiran Aina, and Olufunmiso Olusola Olajuyigbe
3. Deterpenation of Citrus Essential Oils Using Glycerol-Based Deep Eutectic
Solvents. Baranse Ozturk, Jesus Esteban, and Maria Gonzalez-Miquel
4. Extraction of Essential Oil from Cinnamon (Cinnamomum zeylanicum)
Y.C.Wong*, M. Y. Ahmad Mudzaqqir and W.A. Wan-Nurdiyana
5. Purification of Cinnamaldehyde from Cinnamon Species by Column
Chromatography Nandam Sree Satya, Surya Prakash D.V., Vangalapati
Meena

24