DECLARATION

I, Papias NTEZIYAREMYE, a student in University of Rwanda, Department of Chemistry, Option of Environmental

Chemistry, hereby declare that this internship conducted in University of Rwanda, Huye campus Chemistry
Laboratories from 2nd July to 31st August is the best of my knowledge and belief, original except as acknowledged
in internship report and had never been submitted elsewhere for an award of academic qualification.
I, Papias NTEZIYAREMYE
Signature......................................................
Date…………………………………………

i
 DEDICATION

I dedicate this report to Almighty God for his guidance and protection in my daily life.
This report is also dedicated to my family who ever love and support me in whatever I do.
To My brothers and sisters, my classmates and Best Friends, is this internship report dedicated. I sincerely dedicate
this work globally to everyone who helped me; morally, financially, or by counseling, for successfully achievement of
this work.

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 ACKOWLEDGEMENTS

Firstly, My greatest gratitude goes to God Almighty, the infinity of goodness from whom every good gift comes
from, for the gifts of life and knowledge and who permitted the success and completion of this report.
I also want to express my gratitude to The Head of Department Dr. Theoneste MUHIZI, all Lecturers and staff in the
Department of Chemistry, whose efforts in producing great leaders of tomorrow will never be in vain.
Particularly, I express my gratitude to my facilitators Lab-technicians include Mr. Emmanuel NKUNDIMANA in
water quality analysis and Mr. Hubert RUDAKEMWA in bio-organic chemistry, who made this work successful
through their encouragement, patience and support. I deeply express my gratitude to my father Mr. B. Deogratias, my
mother Mrs. N. Verdiane and my brothers and sisters whose prayers, materials and moral supports went a long way in
the success of this work and during my course of study in University of Rwanda.

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Table of content
DECLARATION............................................................................................................................................................. i
DEDICATION ...............................................................................................................................................................ii
ACKOWLEDGEMENTS ............................................................................................................................................ iii
Table of content............................................................................................................................................................. iv
LIST OF ABBREVIATIONS AND ACRONYMS ......................................................................................................vii
List of tables ............................................................................................................................................................... viii
List of figures and images ............................................................................................................................................. ix
ABSTRACT ................................................................................................................................................................... x
CHAPTER I. GENERAL INTRODUCTION ............................................................................................................... 1
I.1. Literature Review .......................................................................................................................................... 2
I.2. GENERAL OBJECTIVES ............................................................................................................................ 3
II. 1. OILS EXTRACTION FROM NATURAL PRODUCTS ............................................................................ 3
1.1. Introduction ......................................................................................................................................... 3
II.1.2. Physical and chemical properties of oils and fats ............................................................................ 4
II.1.3. Objectives ........................................................................................................................................ 4
II.1.4. MATERIALS AND REAGENTS .................................................................................................... 4
II.1.5. METHODOLOGY .......................................................................................................................... 4
II.1.6. PROCEDURES ............................................................................................................................... 6
II.1.7. CALCULATIONS AND RESULTS ................................................................................................ 7
II.2. EXTRACTION OF ESSENTIAL OILS FROM NATURAL PRODUCTS ................................................. 9
II.2.1. Introduction ...................................................................................................................................... 9
II.2.2. OBJECTIVES ................................................................................................................................ 10
II.2.5. MATERIALS AND REAGENTS .................................................................................................. 12
II.2.6. METHODOLOGY ........................................................................................................................ 12
II.2.6. EXTRACTION USING HYDRO DISTILLATION METHOD ................................................... 13
II.2.7. PROCEDURES ............................................................................................................................. 17

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 II.2.8. CALCULATIONS, RESULTS AND DISCUSSIONS .................................................................. 18
II.2.9. GENERAL USES OF EUCALYPTUS ESSENTIAL OILS........................................................... 20
II.3. WATER ANALYSIS ................................................................................................................................. 20
II.3.1. Introduction .................................................................................................................................... 20
II.3.2. Objectives ...................................................................................................................................... 21
II.3.3. Physical tests .................................................................................................................................. 21
II.3.4.Chemical tests ................................................................................................................................. 22
II.3.5. Bacteriological tests ....................................................................................................................... 22
II.3.6. Sampling ........................................................................................................................................ 23
II.3.7. TRANSPORT AND CONSERVATION OF SAMPLES ............................................................... 24
II.3.8. MATERIALS AND APPARATUS ................................................................................................ 24
II.3.9. ANALYSIS OF SOME IONS IN WATER .................................................................................... 24
CHAPTER IV: CONCLUSION AND RECOMMENDATIONS ................................................................................ 32
1. CONCLUSION .............................................................................................................................................. 32
2. RECOMMENDATION .................................................................................................................................. 32
REFERENCES ............................................................................................................................................................ 34
APPENDICES ............................................................................................................................................................. 35
APPENDICES 1: preparation of sample :Natural products : soybeans and groundnuts ................................... 35
APPENDICES 2: preparation of solution for analysis of ammonium ions. ...................................................... 35
Procedures for analyzing Ammonium ...................................................................................................... 36

v
vi
LIST OF ABBREVIATIONS AND ACRONYMS

UR: University of Rwanda

UV: Ultra-violet
o
C: Celsius degree
%: percentage
mg: milligram
L: liter
S1: sample1
St1: standard1
WHO: World Health Organization
VT: total volume
AAS: Atomic Absorption Spectrometer
λ: Wavelength
DO: Dissolved Oxygen
TDS: Total Dissolved Solid
pH: Hydrogen potential.
LADAMET: Laboratoire d’Analyse des Denrées Alimentaires, Médicaments, Eaux et Toxiques.
B.O.D: Biological Oxygen Demand.

vii
 List of tables

Table 1: Summary of Results for groundnut and soybeans. ........................................................................................ 8

Table 2: Characteristics difference between essential and vegetables oils................................................................ 10
Table 3: Results summary for essential oils extraction............................................................................................... 19
Table 4: physico-chemical parameters to analyze for different water applications ................................................ 23
Table 5: Results of standard solutions obtained using UV- spectrophotometer....................................................... 25
Table 6: The concentrations of ammonium ions in samples ...................................................................................... 27
Table 7Summary table of results found about concentrations of ions in water samples ........................................ 30
Table 8: standard solution preparation ............................................................................................................................. 36

viii
 List of figures and images

Figure 1soxhlet extractor connected with condenser and round-bottomed flask. ............................................................. 5
Figure 2: The Soxhlet apparatus are set for the extraction ................................................................................................ 6
Figure 3 : Rotary evaporator (Rotavapor) .......................................................................................................................... 7
Figure 4: Rotary evaporator in use..................................................................................................................................... 7
Figure 5: Comparison chart ............................................................................................................................................ 9
Figure 6:Dean-stark setup for extraction. ........................................................................................................................ 14
Figure 7: Dean stark with condenser and flask ................................................................................................................ 14
Figure 8Essential oils extraction set up ........................................................................................................................ 17
Figure 9Graph present absorbance vs concentration of standard solutions ............................................................ 26

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ABSTRACT

The Chemistry Department planed the internship of students for the main objective of allowing them to be
familiar with practical knowledge based on the theory learned in class and make appropriate application of what they
studied during the previous academic years. The period of internship was one month, that is from 2nd to 31st July, 2015
and it is mainly made with two activities: Extraction of oils and Water analysis. Firstly, we focused on extraction of
fat and oil from natural products as soybeans and groundnuts using soxhlet extraction method, here the yield were
45.128% and 21.961% of groundnuts and soybeans respectively. This implies that , the groundnuts are good for
production of a large amount of oil than soybeans. Secondly we worked on extraction of essential oil from the fresh
leaves of two types of eucalyptus; eucalyptus globulus and eucalyptus macurata by using hydrodistillation method.
As results for this session, we found that many essential oils were produced by eucalyptus macurata with a yield of
1.39%, and 0.984% for eucalyptus globulus. Essential oils are very useful in different domains due to their chemical
constituents such as terpenes, alcohols, flavonoids, fatty acids, steroids, etc. In third session of this internship, I
worked on analysis of water. We know that Water is one of the most important and abundant compounds of the
ecosystem and all living organisms on the earth need water for their survival and growth. In this session, different ions
like chloride, ammonium, fluoride, zinc, manganese and iron were analyzed in water samples. Based on analysis done
in laboratory, these water samples do not contain any chloride, zinc, manganese and iron ions . The only ions present
were ammonium and fluoride ions in which their concentration were also below the standards recommended by WHO
in 2014.These water samples collected from different sites in MUSANZE and NYABIHU Districts, should be
accepted as drinking-water after manual addition of some important ions like chloride to kill micro-organisms and
others needed by our body to improve their quality.
This report is made up of four chapters:
Chapter one comprises of the general introduction, objectives, and literature review.
Chapter two comprises equipment used, sampling and methodology.
Chapter three comprises all works performed and results interpretation.
Chapter four comprises the conclusion and recommendations.

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 CHAPTER I. GENERAL INTRODUCTION

The internship provides a great opportunity for prospective students, to gain experience in a
particular field or industry. it may also help the students to practice many theories learned in the
class. This is the reason why the Department of chemistry planned the internship of four weeks
for chemistry students, this internship is carried out in different industries, institutions and
Laboratories of related field. The internship carried out in laboratories of University of Rwanda/
Huye Campus has three parts to be dealt with in a whole month, these are: water analysis, oil
extraction and essential oil extraction from natural products. For water analysis, It has the target
of allowing students to analyze different physico-chemical parameters (such as pH, alkalinity,
conductivity, total dissolved solids, Hardness, color, turbidity, etc ) and determine the
concentration of different ions in given samples of water . The session of water analysis also
allows us to become familiar with the use or/and run of machines like UV-Spectrophotometer,
DR-Spectrophotometer and other instruments available in the Laboratory. The second session
was the extraction of essential oils. We know that Essential oils are highly concentrated, volatile
substances extracted from flowers, leaves, stems, roots, seeds, barks, or fruit rinds of plants and
they are used for their characteristic smell and their therapeutic or odoriferous properties, in a
wide selection of products such as foods, medicines, and cosmetics. The world production and
consumption of essential oils are increasing very fast. So, during internship, we were interested
in knowing the amount of essential oil that are produced by different plant species and compare
them in overall yield and quality of essential oil they produce. The other covered activity is the
extraction of oil and fats from natural products such as soybeans and groundnuts. Oils and fats
are lipids extracted from vegetables, nuts, seeds, milk, avocadoes, etc. These Fats and oils are
extensively used as food and also in cosmetics, pharmaceuticals and other industries. They
constitute an important resource in the food and cosmetic fields. In the internship, we found out
the amount of oil in two lipid- containing seeds which are soybean and groundnut. The
extraction of oil and fat was done using soxhlet extractor apparatus. A Soxhlet extractor is a
piece of laboratory apparatus invented in 1879 by Franz von Soxhlet and it was originally
designed for the extraction of a lipid from solid materials as ours (soybean and groundnut).

1
Our internship carried out in laboratories of University of Rwanda/ Huye Campus commenced
on 13rd to 31st July, 2015 and it took place in two laboratories: One is LADAMET( Laboratoire
d’Analyse des Denrées Alimentaires, Médicaments, Eaux et Toxiques.) at MAMBA; this modern
laboratory is well-equipped with different materials and apparatus that were used in Water
analysis and distillation of oil by rotary vapor. The other laboratory locates at RUHANDE
(former NUR) and it is equipped with materials that we used in fat and oil extraction and
essential oil extraction.

I.1. Literature Review

Water is one of the most important and abundant compounds of the ecosystem, as of now only
earth is the planet having about 71 % of water. But due to the increased human population,
industrialization, use of fertilizers in the agriculture and man-made activity it is highly polluted
with different harmful contaminants. This contamination causes water born-diseases which have
led to the death of millions of people. ( Adefemi and Awokunmi, 2010). It is very essential and
important to test the water before it is used for drinking, domestic, agricultural or industrial
purpose. Water must be tested with different physic-chemical parameters and the parameters to
test in water are solely depend upon for what purpose we going to use that water and what
extent we need its quality and purity.
Lipids (fats and oils ) are a group of substances that, in general, are soluble in organic solvents
but are relatively insoluble in water.
Lipids (Fats and oils ) have became an integral part of human diet. The per capita fat
consumption has increased significantly, from an average of only 53 grams in 1967- 1969 to as
much as 73 g/capita/day in 1997-1999 around the world and contributes 30% of total energy
supply of mankind (FAO, 2003) . It is also projected to continue growing. The increase in oil
production which began with the application of solvent extraction method, made the access to
these commodities easier. Because of commercial regulations, it is important for food producers
and consumers to be able to report fat content in a serving size of a food item. The Soxhlet
procedure allows for the calculation of total lipids(fat and oils) content in seeds like soybeans
and groundnuts or other natural products. In many of its published methods, the U.S.
Environmental Protection Agency requires Soxhlet extraction of components from a variety of
sample matrices including foods and soils.

2
Essential oils are volatile substances extracted from flowers, leaves, stems, roots, seeds, or fruit
rinds of plants. In this regard, the International Organization for Standardization (ISO) defines
an essential oil as a: “product obtained from natural raw material, either by distillation with
water and steam, or from the epicarp of citrus fruits by mechanical processing, or by dry
distillation”.

I.2. GENERAL OBJECTIVES

The ambition of internship was to enhance practical skills, to facilitate students in gaining new
knowledge and practical experiences of the theories learned and make appropriate application of
what studied during the previous years of study. The other purpose is to improve the personal
ability for using laboratory materials and apparatus. It also experiences us about working in
team and being familiar with working environment.

CHAPTER II. PERFOMED ACTIVITIES

II. 1. OILS EXTRACTION FROM NATURAL PRODUCTS

1.1. Introduction

Seed oils, Nuts oils and oils of fruits and vegetables are receiving growing interest due to their
high concentration of bioactive lipid components, such as polyunsaturated fatty acids and
phytosterols, which have shown various health benefits. Fats and oils, and their several lipid
components are extensively used in the food and also in cosmetics, pharmaceuticals,
oleochemicals and other industries. They constitute an important resource in the food and
cosmetic fields. Vegetable lipids are usually obtained by solvent extraction and/or by pressing
seeds and fruits of many plants.
The conventional extraction methods are time consuming, have low selectivity and/or low
extraction yields, and possible solvent contamination of final products. Supercritical fluid
extraction allows obtaining extracts free of toxic residues that can be directly used, without any
further treatment, and with excellent features and ultra pure composition of final product.

3
II.1.2. Physical and chemical properties of oils and fats

 Fats and oils (lipids) are soluble in non-polar solvents (Ether, benzene, liquid alkanes,...)
it means they are non-polar compounds, are insoluble in water(polar solvent).
 Mainly oils are liquid but fats are solid at room temperature.
 They have relatively high-boiling point than water.
 Both fats and oils have relatively low density.

II.1.3. Objectives

The objective of oil and fats extraction is to determine lipid contents of various oil-seeds by
weight in a sample. This session helped me to calculate the yield of oils from two different seed
species and compared them in oil production. It also helped in gaining some skills about
operation of a Soxhlet extractor.

II.1.4. MATERIALS AND REAGENTS

Materials Reagents
Extraction thimble Soybean
Ring stand Groundnut (peanut) Round flask
Soxhlet extractor petroleum Ether (as solvent)
Analytical balance
Heater and Funnel

II.1.5. METHODOLOGY

There are many methods that can be used for extracting oils from natural products, those
include; mechanical methods, chemical methods, and soxhlet method. The later method was
used during this session.

II.1.5.1. Mechanical methods

Mechanical extraction of the oil is accomplished by exerting sufficient force on confined seed.
Under this condition pressure is high enough to rupture the cells and force oil from the seed to
“escape.” Extraction is accomplished by compressing the material in a container that has small
perforations, either round or slotted, that allow the liquid component to leave.

4
Mechanical pressing alone does not remove all the oil from the seed, it's followed by chemical
extraction to remove the remaining oils.

II.1.5.2. Chemical methods

Those methods are applicable to almost of fats-containing products. They consist of using the
chemical solvents to remove oils from seeds. Chemical method is commonly used at small scale
and for research purposes.
Solvent extraction method belongs in chemical methods, and is the most popular method for
separation of oil from soybeans, mainly because of the method’s high extraction efficiency (over
99%) as well as its capability to handle large quantities.

II.1.5.3. Soxhlet method

It is a method of extraction in which the soxhlet extractor is used. A Soxhlet extractor is a piece
of laboratory apparatus invented in 1879 by Franz von Soxhlet. It was originally designed for
the extraction of a lipid from a solid material. However, a Soxhlet extractor is not limited to the
extraction of lipids. Typically, a Soxhlet extraction is only required where the desired compound
has a limited solubility in a solvent, and the impurity is insoluble in that solvent. If the desired
compound has a significant solubility in a solvent then a simple filtration can be used to separate
the compound from them insoluble substance.

Figure 1soxhlet extractor connected with condenser and round-bottomed flask.

5
II.1.6. PROCEDURES

During our internship, we preferred to use the well-known natural products such as: Soybeans
and Groundnut
Firstly, we need to choose the best quality of our sample (soybeans or groundnut) and separate
them with other materials like stones, sand,...that can be in mixture with. Both products were
harvested from GISAGARA District.
We weighed 0.25kg of sample (soybeans or groundnut)
We crashed them in mortar using the pestle, so that we get the powder of sample.
Note: It is better to use the powder of sample in order to facilitate its dissolution in solvent.
From the obtained powder, we weighed in 35.0g and 48.60g of groundnut and soybeans
respectively put them gently in thimble, then the thimble is carefully placed into soxhlet
extractor.
About 150 ml of petroleum Ether is poured into 250 ml round-bottomed flask with boiling
chips. The solvent is boiled ; the vapors rise to the condenser where they condense and drip
down through the sample back into the boiling solvent below. The extraction process is
continuous and was completed between3 and 4 hours.

Figure 2: The Soxhlet apparatus are set for the extraction

6
 In a Soxhlet extractor, the solvent is heated in a boiler; the pure vapor rises up through a by-pass
and into the top part of the Soxhlet container where the sample to extract is contained. In the
condenser, the vapors are condensed and drip into the sample-containing thimble. When the level
of liquid reaches the same level as the top of the siphon, the liquid containing the extracted
material is siphoned back into the boiler.

Figure 3 : Rotary evaporator (Rotavapor) Figure 4: Rotary evaporator in use

Once the extraction is finished, the solvent is removed from the extracted oil by distillation
using a Rotavapor, also called Rotary evaporator, Which is the instrument used in distillation
to separate solvents and oils at low pressure.
Note: At low pressure the boiling point of a compound becomes slightly low, this is important
for some organic compounds that can be destroyed or volatilized at high pressure( high
temperature).
At the end of distillation , we got separately both , oils and solvents . Solvent should also be
used again and again since it's regained at the end of distillation.

II.1.7. CALCULATIONS AND RESULTS

1. From sample of Groundnut (peanut).

Mass of Sample (Groundnut powder ): 35.0g
Mass of Beaker : 105.083g
Total mass of oil and beaker : 121.284g
Mass of oil only : (121.284 – 2105.083 )g =16.201g
The oils obtained by extraction is equal to 16.201g as mass, and we calculated the yield as
follow:

7
 mass of obtained oil
Yield =
mass of sample taken x100,

16.201g
Yield =
35.90g x100 = 45.128%

2. From sample of Soybeans.

Mass of Sample (Soybean powder ): 48.60g
Mass of Beaker: 105.526g , and the Total mass of oil and beaker : 116.199g
Mass of oil only: (116.199 – 105.526 )g =10.673g
The oils obtained by extraction is equal to 10.673g as mass, and we calculated the yield as
follow:
10.673g
Yield =
48.60g x100 = 21.961%

Table 1: Summary of Results for groundnut and soybeans.

Scientific Common Mass of Mass of oil yield conclusion

name name sample obtained

Glycine Max Soybeans 48.60g 10.673g 21.96% The groundnuts yields

arachis Groundnut 35.0g 16.201g 45.13% more oils than soybeans

hypogaea

8
 60

48.6
50

40 35

30 mass of sample
massof oil
20 16.201
10.673
10

0
soybeans groundnuts

Figure 5: Comparison chart

By comparing above results of calculated yields, we found that the yield of groundnut is about
2.055 times greater than that of soybeans. This means that, the groundnut is a higher lipid-
containing species than soybeans.

II.2. EXTRACTION OF ESSENTIAL OILS FROM NATURAL PRODUCTS

II.2.1. Introduction

Essential oils are highly concentrated, volatile substances extracted from flowers, leaves, stems,
roots, seeds, barks, resins, or fruit rinds of plants . Not all plants produce essential oils. These
oils are present in natural products at small amount because the amount of essential oils found in
these plants can be anywhere from 0.01 percent to 10 percent of the total. That's why tons of
plant material are required for just a few hundred pounds of oil. they are often used for their
characteristic smell and their therapeutic or odoriferous properties, in a wide selection of
products such as foods, medicines, and cosmetics.
Extraction of essential oils is one of the most time- and effort- consuming processes.
There are a wide number of ways to extract the Essential oil but the quality never remains the
same, it means that the way in which oils are extracted from plants is important because some
processes use solvents that can destroy the therapeutic properties.

9
II.2.2. OBJECTIVES

The essential oils are used in a wide variety of consumer goods such as detergents, soaps, toilet
products, cosmetics, pharmaceuticals, perfumes, confectionery food products, soft drinks,
distilled alcoholic beverages (hard drinks) and insecticides. So, based on their uses, the objective
of extracting essential oils from different plant species is to know the amount and yield of
essential oils produced, so that we make a comparison of plant species in production of essential
oils.

Table 2: Characteristics difference between essential and vegetables oils

essential oils vegetables oils

 They are typically non-oily,  They are called fixed oils because they are
 They are volatile and evaporate at or non-volatile and do not evaporate like
above room temperature. essential oils.
 They normally dissolve in alcohol.  They are also not soluble in alcohol.
 They are found in plants at small amount  They are found in plants at large amount
compare to essential oils.

II.2.4. Chemical Constituents of Essential Oils

The chemical composition of both vegetable oils and essential oils is complex and the exact
chemical composition of each oil depends on the plant species from which it was extracted
from. Pure essential oils are mixtures of more than 200 components, normally mixtures of
terpenes or phenylpropanic derivatives, in which the chemical and structural differences
between compounds are minimal. They can be essentially classified into two groups:
Volatile fraction, such as aliphatic aldehydes, alcohols, and esters.
Nonvolatile residues, such as fatty acids, sterols, carotenoids, waxes, and flavonoids.
1. Hydrocarbons:
Essential Oils consist of Chemical Compounds that have hydrogen and carbon as their
building blocks. Basic Hydrocarbon found in plants are isoprene having the following
structure
(Isoprene)

10
 2. Ketones
Essential oils containing ketones are beneficial for promoting wound healing and encouraging
the formation of scar tissue.
Ketones are usually (not always) very toxic. The most toxic ketone is Thujone found in
mugwort and wormwood oils. Other toxic ketones found in essential oils are pulegone in
pennyroyal, and pinocamphone in hyssops. But there are Some non-toxic ketones like jasmone
in jasmine oil, fenchone in fennel oil, and menthone in peppermint oil.
3. Alcohols
Alcohols exist naturally, either as a free compound, or combined with a terpenes or ester. They
improve the quality of essential oils for their anti-septic, anti-viral, bactericidal and germicidal
Properties. Here are some alcohols found in typical essential oils: linalool found in ylang-ylang
and lavender, Geraniol in geranium and rose.
4. Terpenes:
Generally this class of compounds include: Limonene, Pinene, Piperene, Camphene,...
Terpenes are anti- inflammatory, antiseptic, antiviral, and bactericidal. Terpenes can be further
categorized in monoterpenes, sesquiterpenes and diterpenes. Referring back to isoprene units
under
the Hydrocarbon heading, when two of these isoprene
units join head to tail, the result is a monoterpene, when three join, it’s a sesquiterpene
and four linked isoprene units are diterpenes.

(Limonene) (Menthol)
5.Aldehydes
Medicinally, essential oils containing aldehydes are effective in treating Candida and
other fungal infections because of their anti-fungal, anti-inflammatory, and anti-viral Properties.
Example of Citral in lemon and citrus eucalyptus .
6. Acids

11
Organic acids with their anti-inflammatory Properties in free state are generally found in very
small quantities within Essential oils. Plant acids act as components or buffer systems to control
acidity.
Example of Cinnamic and benzoic acid in benzoin.
8. Ester
Essential oils containing esters are used for their soothing, balancing effects. Because of the
presence of alcohol, they are effective antimicrobial agents. Medicinally, esters are characterized
as antifungal
and sedative, with a balancing action on the nervous system.

II.2.5. MATERIALS AND REAGENTS

Materials Reagents
Ring stand and its accessory Leaves of eucalyptus macurata
Dean-stark Leaves of eucalyptus globulus spp
Heater Water(H2O)
Plastic tube

II.2.6. METHODOLOGY

The essential oils have many methods of extraction but the method in which oils are extracted
from plants is important because some methods use solvents that can destroy their therapeutic
properties. Some methods that are used for extractions of essential oils are given below:

a. Hydro distillation
This method is often used in primitive countries. The risk is that the still can run dry, or be
overheated, burning the aromatics and resulting in an Essential Oil with a burnt smell. Hydro
distillation seems to work best for powders (spice powders, ground wood, etc.) and very tough
materials like roots, wood, or nuts. This method is cheaper and was preferred during our
internship.

b. Solvent Extraction
An hydrocarbon solvent is added to the plant material to help dissolve the essential oil. When
the solution is filtered and concentrated by distillation, a substance containing resin (resinoid),

12
or a combination of wax and essential oil (known as concrete) remains. From the concentrate,
pure alcohol is used to extract the oil. When the alcohol evaporates, the oil is left behind.

c. Steam Distillation
The most essential oils are commonly extracted from the plants using this technique.
One type of distillation places the plants or flowers on a screen. Steam is passed through the
area and becomes "charged" with the essence.
The steam then passes through an area where it cools and condenses. This mixture of water and
essential oil is separated and bottled.

e. Cold Pressing
This method involves the simple pressing of the rind at about 120 degrees F to extract the oil.
The rinds are separated from the fruit, are ground or chopped and are then pressed. The result is
a watery mixture of essential oil and liquid which will separate given time. It is important to
note that oils extracted using this method have a relatively short shelf life, so make or purchase
only what you will be using within the next six months. Cold pressing is used to extract the
essential oils from citrus rinds such as orange, lemon, grapefruit and bergamot.

II.2.6. EXTRACTION USING HYDRO DISTILLATION METHOD

Essential oils can be extracted using a variety of methods, although some are not commonly
used today.
Hydrodistillation is the most common method used for volatile oil extraction. This method is
described in the European Pharmacopoeia 2005 (Duran A., 2010).
Many researchers think that this method is not so good, because it is not certain that the
compounds cannot be destroyed by heat during the process. Hydrodistillation needs a large
amount of plant material and the time for extraction is quite long (around 3 hours).
Because of the long time for extraction, the energy consumption is quite high. In isolation of
essential oils using hydro distillation method, the aromatic plant material is packed in a Dean-
stark 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 oils are
freed from the oil glands in the plant tissues (leaves). The vapor mixture of water and oils is
condensed by indirect cooling with water. From the condenser, distillate flows into a separator,

13
where oils separate automatically from the distillate water.
As disadvantages: The process is slow and the distillation time is much longer thereby
consuming more firewood / fuel making process uneconomical. Sometimes, the Prolong action
of hot water can cause hydrolysis of some
constituents of the essential oils such as esters etc which reacts with the water at high
temperatures to form acids & alcohols . This method is not suitable for large scale distillations .

Figure 6:Dean-stark setup for

Figure 7: Dean stark with Hydro extraction.
condenser and flask
distillation involves the following main physicochemical
processes:
Hydro diffusion, Hydrolysis, Decomposition by heat .
Hydro diffusion is a diffusion of essential oils and hot water through plant material membranes
Hydrolysis 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.
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 distillation is determined entirely by the operating pressure, whereas
in water distillation and in water and steam distillation the operating pressure is usually
atmospheric.
It is possible to obtain better yield and quality of oils by:
Maintaining the temperature as low as possible,

14
Using as little water as possible, in the case of steam distillation, and
Thoroughly comminuting the plant material and packing it uniformly before distillation.
Here there are three types of hydro distillation for isolating essential oils from plant materials:
Water distillation , Water and steam distillation , and Direct steam distillation.

1. Water Distillation
In this method, the materials are completely immersed in water, which is boiled by applying heat
by direct fire. The main characteristic of this process is that there is direct contact between
boiling water and plant material.
When the still is heated by direct fire, adequate precautions are necessary to prevent the charge
from overheating. When a steam jacket or closed steam coil is used, there is less danger of
overheating; with open steam coils this danger is avoided. But with open steam, care must be
taken to prevent accumulation of condensed water within the still. Therefore, the still should be
well insulated.
The plant material in the still must be agitated as the water boils, otherwise agglomerations of
dense material will settle on the bottom and become thermally degraded. Consequently, before
any field distillation is done, a small-scale water distillation in glassware should be performed to
observe whether any changes take place during the distillation process.
During water distillation, all parts of the plant charge must be kept in motion by boiling water;
this is possible when the distillation material is charged loosely and remains loose in the boiling
water. For this reason only, water distillation possesses one distinct advantage, i.e. that it
permits processing of finely powdered material or plant parts that, by contact with live steam,
would otherwise form lumps through which the steam cannot penetrate.
Other practical advantages of water distillation are that the stills are inexpensive, easy to
construct and suitable for field operation.
The main disadvantage of water distillation is that complete extraction is not possible.
Besides, certain esters are partly hydrolyzed and sensitive substances like aldehydes tend to
polymerize.
Water distillation requires a greater number of stills, more space and more fuel. It demands
considerable experience and familiarity with the method. The high-boiling and somewhat water-
soluble oil constituents cannot be completely vaporized or they require large quantities of steam.
Thus, the process becomes uneconomical.

15
 For these reasons, water distillation is used only in cases in which the plant material by its very
nature cannot be processed by water and steam distillation or by direct steam distillation.

2. Water and Steam Distillation

For this type of 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.
Advantages of Water and Steam Distillation over Water Distillation include: Higher oil yield, If
refluxing is controlled, then the loss of polar compounds is minimized, Oil quality produced by
steam and water distillation is more reproducible, Steam and water distillation is faster than
water distillation, so it is more energy efficient. Many oils are currently produced by steam and
water distillation. Even if it's useful, some disadvantages of Water and Steam Distillation are
realized: Due to the low pressure of rising steam, oils of high-boiling range require a greater
quantity of steam for vaporization -hence longer hours of distillation, the plant material becomes
wet, which slows down distillation as the steam has to vaporize the water to allow it to condense
further up the still.

3. Direct steam distillation

It is the process of distilling plant material with steam generated outside the still in a satellite
steam generator generally referred to as a 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. Because steam is
generated in a satellite boiler, the plant material is heated no higher than 100° C and,
consequently, it should not undergo thermal degradation.
Steam distillation is the most widely accepted process for the production of essential oils on
large scale. There are some advantages of Direct Steam Distillation include: Amount of steam
can be readily controlled, No thermal decomposition of oil constituents can occur, it is the most
widely accepted process for large-scale oil production, superior to the other two processes.

16
The higher capital expenditure needed to establish this activity is like a disadvantage of Direct
Steam Distillation.

II.2.7. PROCEDURES

The fresh sample materials (leaves) were collected in the morning from arboretum, RUHANDE
parcel no 19. Fresh leaves of eucalyptus macurata or eucalyptus globulus spp were cut into
pieces less than 2X2cm within the morning after collection and conservation in fridge for last
one day.
A mass of 244.0g of eucalyptus globulus spp leaves and 252.0g of eucalyptus macurata leaves
boiled with 250 ml of distilled water in a Dean- Stark apparatus until oil distillation ceased after
2-3 h. The volume of essential oils was determined from a calibrated trap.

Figure 8Essential oils extraction set up

17
II.2.8. CALCULATIONS, RESULTS AND DISCUSSIONS

1. From sample1: Fresh leaves of eucalyptus macurata

mass of box(sample-containing box ):22.50g
Total mass of sample and box:274.50g
Mass of sample only :(274.50-22.50)g=252.0g
essential oils obtained after hydro distillation is equal to 4.2ml as volume , 3.50g as mass. So,
from the amount of oils obtained , we calculated the yield as follow:
mass of extracted oil
Yield =
mass of sample x100
3.50g
Yield =
252.0g x100=1.3888%

2. from sample2: Fresh leaves of eucalyptus globulus spp

mass of box(sample-containing box ):22.50g
Total mass of sample and box:266.50g
Mass of sample only :(266.50-22.50)g=244.0g
mass of small bottle( used in oil recuperation): 8.90g
mass of oils and bottle: 11.30g then, mass of oil only: (11.30-8.90)g=2.40g
2.40g
we calculated the yield as follow: Yield = 244.0g x100 = 0.984%

18
Table 3: Results summary for essential oils extraction

Scientific common Part of Mass of Mass of Duration

name of name plant sample oil of Yield Conclusion
species used used obtained heating

eucalyptus Tasmanian leaves 244.0g 2.40g 2-3h 0.98% Leaves of

globulus blue gum eucalyptus
spp macurata contain

eucalyptus Spotted leaves 252.0g 3.50g 2-3h 1.39% more essential

macurata gum oils than the
leaves of
eucalyptus
globulus spp

300
244 252
250
200
150 Mass of sample used (in g)
100
Mass of E.oil obtained
50
2.4 3.5
0
eucalyptus globulus eucalyptus macurata
spp

Figure 8: comparison chart

Based on the yields above, we need to conclude that, the leaves of eucalyptus macurata contain
more essential oils than the leaves of eucalyptus globulus spp . The ratio of yields shows that it's
1.4times.

19
II.2.9. GENERAL USES OF EUCALYPTUS ESSENTIAL OILS

1. Medicinal and antiseptic

Eucalyptus oil has antibacterial effects on pathogenic bacteria in the respiratory tract. Inhaled
eucalyptus oil vapor is a decongestant and treatment for bronchitis. Cineole controls airway
mucus hypersecretion and asthma via anti-inflammatory cytokine inhibition.
Eucalyptus oil also stimulates immune system response by effects on the phagocytic ability of
human monocyte derived macrophages.
Eucalyptus oil is also used in personal hygiene products for antimicrobial properties in dental
care and soaps. It can also be applied to wounds to prevent infection.

2. Repellent and biopesticide

Cineole-based eucalyptus oil is used as an insect repellent and biopesticide. In the U.S.,
eucalyptus oil was first registered in 1948 as an insecticide and miticide.

3. Flavoring
Eucalyptus oil is used in flavouring. Cineole-based eucalyptus oil is used as a flavouring at low
levels (0.002%) in various products, including baked goods, confectionery, meat products and
beverages. Eucalyptus oil has antimicrobial activity against a broad range of foodborne human
pathogens and food spoilage microorganisms.

4. Fragrance
Eucalyptus oil is also used as a fragrance component to impart a fresh and clean aroma in soaps,
detergents, lotions and perfumes. It is known for its pungent, intoxicating scent.

II.3. WATER ANALYSIS

II.3.1. Introduction

Water is life. No life can exist without water. Water is absolutely essential not only for survival
of human beings, but also for animals, plants and all other living beings. It has many beneficial
uses such as drinking, irrigation, navigation, propagation of wild life, fisheries, recreation;
aesthetics etc. water is one of the most valuable natural resources.

20
It is the basic element of social and economic infrastructure and is essential for healthy society
and sustainable development. Water, the matrix of life is exposed to pollution, unhealthy
environment, resulting in human affliction and diseases transmission due to rapid
industrialization and population.

II.3.2. Objectives

Firstly the objectives of water analysis is to get the knowledge about water analysis in physico-
chemical parameters such as turbidity, pH, alkalinity, conductivity, total dissolved solids,
Hardness, and conductance of a water sample to compare them with that of standards.
Secondly, the water analysis helps in quality improvement of water depends on what is going to
use for.
The parameters taken into account in water analysis are not universal, means it depends on the
use of analyzed water.
These parameters may be grouped into physical, chemical, bacteriological and microscopic
categories.
· Physical tests indicate properties detectable by the senses.
· Chemical tests determine the amounts of mineral and organic substances that affect water
quality.
· Bacteriological tests show the presence of bacteria, characteristic of faecal pollution.

II.3.3. Physical tests

Colour, turbidity, total solids, dissolved solids, suspended solids, odour and taste are recorded.
Colour in water may be caused by the presence of minerals such as iron and manganese or by
substances of vegetable origin such as algae and weeds.
Turbidity in water is because of suspended solids and colloidal matter. It may be due to eroded
soil caused by dredging or due to the growth of micro-organisms. High turbidity makes filtration
expensive. If sewage solids are present, pathogens may be encased in the particles and escape
the action of chlorine during disinfection.
Odour and taste are associated with the presence of living microscopic organisms; or decaying
organic matter including weeds, algae; or industrial wastes containing ammonia, phenols,
halogens, hydrocarbons.

21
II.3.4.Chemical tests

pH is a measure of hydrogen ion concentration. It is an indicator of relative acidity or alkalinity

of water. Drinking water should have a pH between 6.5 and 8.5. Harbor basin water can vary
between 6 and 9.
Total Hardness: Hardness is caused by divalent metallic ions
that are capable of reacting with sops to form precipitate. And with
certain anions present in the water to form scale.
There are two types of hardness-temporary hardness is also known as carbonate hardness and it
is mainly due to presence of carbonate and bicarbonates of Ca and Mg which is removed by
boiling or by adding Ca(OH)2 to it. The permanent hardness is also known as non-carbonate
hardness and is due to the sulphate, chlorides and nitrates of calcium and magnesium .
Total Dissolve Solids (TDS): The composition of solids present in a natural body of water
mainly depends upon the nature of the bedrocks and the soil developed from it .
Electrical Conductivity: The electrical conductivity is the capacity of waters to conduct
current, and is caused by the present salt, acids and bases, called electrolytes, capable of
producing cations and anions. As the conductivity is directly related to the presence of dissolve
salts, its magnitude can give the fair idea of the level of dissolved solids.
B.O.D.(Biological Oxygen Demand) It denotes the amount of oxygen needed by micro-
organisms for stabilization of decomposable organic matter under aerobic conditions. High
B.O.D. means that there is less of oxygen to support life and indicates organic pollution.

II.3.5. Bacteriological tests

The microbiological quality of water can be evaluated by the absence of pathogens in the water
sample. The main types of microorganisms found in water are bacteria, protozoa, viruses, and
helminthes.
Not all bacteria are pathogenic; many forms of bacteria are helpful such as bacteria in the human
gut. Examples of bacteria that are known to cause disease are Salmonella typhi and pathogenic
Escherichia coli, which cause typhoid fever and gastroenteritis, respectively.

22
Table 4: physico-chemical parameters to analyze for different water applications

Applications of water Parameters to analyze

Pathogen germs
Alkalinity & pH
Drinking water Elements in solutions( nutrients ions, heavy metals)
Appearance( color , turbidity)
organic chemicals

Alkalinity & Ph
Irrigation Salinity
heavy metals

Breeding(elevage) or animal husbandry heavy metals

pathogen germs

Elements in solutions( nutrients ions, heavy metals)

Temperatures
Aquaculture, also known as aqua-farming Dissolved oxygen(D.O)
Biological Oxygen Demand(B.O.D)
Alkalinity, pH and Total hardness

Industrial and domestic use Alkalinity, pH, Total hardness

heavy metals, pathogen germs

II.3.6. Sampling

During sampling, the materials must be washed and rinsed with distilled water to avoid
contamination. The most accurate measurements of water quality are made on-site, because
water exists in equilibrium with its surroundings. Measurements commonly made on-site and in
direct contact with the water source in question include temperature, pH, Dissolved oxygen,
Conductivity and turbidity.
During our Internship, we dealt with the parameters that were analyzed in laboratory, which
include: the quantity of different ions present in each water sample.

23
Even if we were not concerned in sampling, we got some information about collected samples.
After sampling, labeling must be done immediately on field, since it gives the necessary
information about identification of samples.
The water samples were collected from different sites in MUSANZE and NYABIHU Districts
and are named as S1:KINKWARE, S2:KIMBIKI1, S3:KIMBIKI2, S4:NYAKINAMA,
S5:BARIZO, S6:KABUGUNGU, S7:KAMAHEKE, S8:MUSEMBE, S9:KABAYA, AND
S10:MENGO.

II.3.7. TRANSPORT AND CONSERVATION OF SAMPLES

For water samples, some parameters must be analyzed in situ (on field) but other work takes
place in laboratory. So, the samples are gently transported to avoid contamination during
transport. In order to avoid deterioration and contamination, the samples must be kept in
specific storage at low temperature (40C) and dark to inhibit some bacteriological reactions that
can take place.

II.3.8. MATERIALS AND APPARATUS

There are many materials and apparatus that were used to analyze different ions in water
samples, some of those include: Atomic Absorption spectrometer(AAS, to analyze heavy metals
and other cations) , pH-meter,( to measure the alkalinity and acidity of samples) colorimeter,
UV-spectrophotometer, DR-Spectrophotometer,( both three , are used to analyze some ions in
samples), etc..... The later both apparatus were used in our internship.
Apart from above apparatus, there are other materials that were used as usual in
laboratory, they include: cooler box ( for samples transportation), beakers, washing bottles,
micropippettes, graduated cylinders, bottles, spatula, volumetric flasks and analytical balance.

II.3.9. ANALYSIS OF SOME IONS IN WATER

II.3.9.1. Analysis of ammonium ion in water by analytical method using UV-

spectrophotometer
Ammonia and ammonium compounds are excellent sources of plant nitrogen, and are present in
most fertilizers, whether natural or synthetic fertilizer formulations. Ammonium (NH4+) is the
ionized form of ammonia and will equilibrate with ammonia in solution.

24
The source of ammonium in the environment mirrors that of ammonia at a pH of less 8, over
95% of the total ammonia is in the ammonium ion form at 15 degrees Celsius. Most vertebrates
including mammals will convert ammonium into urea where it can be excreted or
bioaccumulated.
Ammonium can be measured in situ with an ion selective electrode.
From the collected water sample, the ammonium ions were analyzed in UR-HUYE CAMPUS at
MAMBA Laboratories of science, and the obtained results were tabulated below.
The different standards of NH4+solutions of 5.0ml, their concentrations and measured
Absorbance are below:

Table 5: Results of standard solutions obtained using UV- spectrophotometer

Standards Conc.mgNH4/L Absorbance(Abs) at 655nm Abs.- Abs(sto)

0.000 0.341 0.000

Sto(blank)

st1 0.150 0.374 0.033

st2 0.300 0.432 0.091

st3 0.450 0.752 0.441

st4 0.600 0.793 0.452

st5 0.750 0.848 0.507

st6 1.500 1.302 0.961

st7 2.250 1.426 1.085

st8 3.000 2.075 1.734

From the standards solutions we need to find the regression line equation that is used to
calculate the concentrations of ammonium ions in sample water solutions. So, we have to plot
the Absorbance against Concentrations of standard solutions.

25
 Graph present absorbance vs concentration of standard
solutions y = 0.5473x + 0.0387
R² = 0.964
2
Absorbances(nm)

1.5

0.5

0
0 0.5 1 1.5 2 2.5 3 3.5
Concentration (mg/l)

Figure 9Graph present absorbance vs concentration of standard

solutions

Regression line Equation: Y=0.5473X+0.0387, Where Y=absorbance and X= concentration.

Based on the regression line equation obtained above, we have to calculate the concentrations of
ammonium ions in water samples at measured Absorbances.

26
Table 6: The concentrations of ammonium ions in samples

Samples Absorbance Abs.- conc.mgNH4+/L

(Abs) at Abs(sto) calculated from Conclusion
655nm samples

S1 1.616 1.275 2.259 Some drinking water treatment

processes add small amounts,
S2 0.394 0.053 0.026 generally less than
0.4 mg/l of ammonia to water to
S3 0.135 -0.206 N.D increase the disinfecting ability
(excluded) of chlorine. European nations have
S4 0.560 0.219 0.329 adopted a drinking water standard
of 0.5 mg/l. The higher
S5 0.162 -0.179 N.D concentration of ammonia in
(excluded) samples S1 and S7 can cause bad
smell of water. So, they are not
S6 0.313 -0.028 N.D
drinkable.
(excluded)

S7 2.150 1.809 3.235

S8 0.555 0.214 0.320

S9 0.416 0.075 0.066

S10 0.489 0.148 0.1997

Ammonium ion is less toxic than ammonia and because it is the most abundant form at the
typical pH of surface water. Ammonium ion will however become toxic at higher concentrations.
Negative effects of ammonia:
Ammonium and ammonia are naturally occurring organic compounds that are formed during
decomposition of proteins, manure wastes and urine wastes, and from other nitrogen-containing
compounds.

27
The ammonium ions are mainly derived from ammonia and their effects in environment are
similar. Human beings and higher animals are less sensitive to ammonia in water, but long-term
ingestion of water containing more than 1mg/l ammonia may be damaging to internal organ
systems. Solutions having concentrations greater than 1000 mg/l (ppm) can cause severe burns
and scarring of sensitive skin and mucous membranes.
As little as one teaspoonful of 10% ammonia solution can be lethal. Splashing into eyes can
cause temporary or permanent blindness.

II.3.9.2. Determination of some ions in water sample solutions using DR-

Spectrophotometer

1. Chloride
Chloride in surface and groundwater from both natural and anthropogenic sources, such as
run-off containing road de-icing salts, the use of inorganic fertilizers, landfill leachates, septic
tank effluents, animal feeds, industrial effluents, irrigation drainage, and seawater intrusion in
coastal areas. Large amounts of chlorine are manually added in water as disinfectants and , it is
widely used to disinfect drinking-water and swimming-pool water and to control bacteria and
odors in the food industry. Chlorine is present in most disinfected drinking-water at
concentrations of 0.2–1 mg/L

2. Fluoride
Fluoride is found in all natural waters at some concentrations. Seawater typically contains about
1.0mg/L while rivers and lakes generally exhibit concentrations of less than 0.5 mg/L.
In groundwaters, however, low or high concentrations of fluoride can occur, depending on the
nature of the rocks and the occurrence of fluoride-bearing minerals.
Potential health effects of long-term exposure to elevated fluoride concentrations include dental
and skeletal fluorosis (U.S. Environmental Protection Agency, 1994).

3. Manganese
Manganese is the abundant one of the metals of the ground surface,and is an essential element
for many living organisms, including humans. It is generally associated with iron .
In form of permanganate (MnO4-) is an energetic oxidant which sometimes used in water
treatment to oxidize the organic matters.

28
The studies suggest that infants exposure to manganese in drinking water should be avoided,
because o f its health effects , particularly the potential neurotoxic effects .

4. Iron
Iron is an essential trace element in living organisms.
Concentrations of iron in drinking-water are normally less than 0.3 mg/litre but may be higher in
regions where various iron salts are used as coagulating agents in water-treatment plants and
where cast iron, steel, and galvanized iron pipes are used for water distribution.
The total body iron in adult males and females is usually about 50 and 34-42 mg/kg of
body weight, respectively .The largest fraction is present as haemoglobin, myoglobin,
and haem-containing enzymes.
5. Zinc
Most zinc is introduced into water by artificial pathways such as by-products of steel production
or coal-fired power stations, or from the burning of waste materials. Zinc is also used in some
fertilizers that may leach into groundwater.
Although levels of zinc in surface water and groundwater normally do not exceed 0.01 and 0.05
mg/L, respectively, concentrations in tap water can be much higher as a result of dissolution of
zinc from pipes.

29
 Table 7Summary table of results found about concentrations of ions in water samples

Drinking
SAMPL water S1 S2 S3 S4 S5 S6 S7 S8 S9 S1 CONCLUSION
ES Standards 0
(WHO
Guidelines)
Chloride plays a great
role in water disinfection.
Chloride 250mg/L 0.0 0.0 0.0 0. 0.0 0. 0.0 0.0 0.0 0. But it is also needed by
0 0 0 our body. The addition of
chloride in these samples
improve their quality for
drinking
Fluoride is very important
for teeth and skeletal
Fluoride 2.0mg/L 0.0 0.0 0.0 0. 0.0 0. 0.0 0.0 0.0 0. growth. Its excess leads to
3 4 4 0 1 0 1 1 0 fluorosis and nervous
problems, but these
samples are under WHO
guidelines.
Zinc is essential in
metabolism. The manual
addition of zinc is
ZINC 5.0mg/L 0.0 0.0 0.0 0. 0.0 0. 0.0 0.0 0.0 0. necessary for quality
0 0 0 improvement. Its
deficiency in young
children or animals will
retard growth and
decreases body resistance
to disease.

30
 It is necessary for proper
functioning of some
enzymes. its deficiency
Mangan 0.05mg/L 0.0 0.0 0.0 0. 0.0 0. 0.0 0.0 0.0 0. leads to skeletal
ese 0 0 3 0 abnormalities,
reproductive deficits, but
the excess provoke
neurological effects
referred to as
“manganism.”

Iron is an essential
element in human
Iron 0.3mg/L 0.0 0.0 0.0 0. 0.0 0. 0.0 0.0 0.0 0. nutrition, and it is found in
0 0 0 bone marrow,
haemoglobin, and haem-
containing enzymes. The
excess of iron may cause
genetic disorder
(haemochromatosis)

31
CHAPTER IV: CONCLUSION AND RECOMMENDATIONS

1. CONCLUSION

Many activities were successfully covered during internship. I reached on my goals as I gained
skills and experience to work in different environments.
Through the internship, I really acquired many practical skills as to handle laboratory work with
different machines and apparatus such as UV-Spectrophotometer, and DR-Spectrophotometer,
etc. I learned how to collect, conserve, process and analyze the data, and perform statistical
analyses to interpret and present these data. Based on performed activities, now I am able to deal
with analysis of different ions and some physico-chemical parameters in water samples so that I
can improve the quality of water depend on their uses. The session of fat and oil extraction was
very helpful for us , since I acquired knowledge about different methods of extracting oil from
natural products, especially by use of soxhlet extraction method. Oils and fats, also called lipids
have the highest food-energy content than carbohydrates and proteins but their methods of
extraction are time consuming, laborious, have l low extraction yields, and possible solvent
contamination of final products. So , through this internship, we extracted oil from soybeans and
groundnuts, where we saw that the groundnuts yield more oil quantity than soybeans. And then,
we took an overview about the better methods of extracting those oil in order to improve their
quality and to reduce the negative impacts on users, that can results from their lower quality.
Nowadays, essential oils play a great role as medicinal and antiseptic, repellent and biopesticide,
and flavouring due to their chemical constituents. During internship, the eucalyptus macurata
and eucalyptus globulus were used in extraction of essential oil by hydrodistillation method, the
large amount of essential oil was found in eucalyptus macurata rather than eucalyptus globulus.
Through the skills that I gained from this internship, I hope, I would be equipped to work with a
range of companies , laboratories or other related fields, especially when they work on covered
sessions, so that I will improve my well-understanding.

2. RECOMMENDATION

Even if, I successfully achieved my aims during internship, they are many difficulties and
challenges that I had been faced with, that is why some recommendations were rose so that the
future internship will be better and more efficiency.

32
As Chemistry Department, It would be better to negotiate early with many industries,
laboratories and other companies where chemistry is applied to help students to acquire varied
knowledge through internship. Today it is very difficult for students themselves to find where
they can conduct an internship.
No laboratory protection, such as laboratory masks, grooves to protect the experimenter from
some chemical products that are harmful to health.
Sometimes, we worked in large group due to the lack of sufficient materials and apparatus . This
is a very big problem for us because everyone does not get an opportunity to carry out an
experiment her/himself. So, it will be better once the issue is handled by concerns .

33
REFERENCES

1. Manahan, Stanley E. "FRONTMATTER"Environmental Chemistry 7th ed. Boca Raton:

CRC Press LLC, 2000 (page 242-256, 268-272).
2. International Research Journal of Environment Sciences: ISSN 2319–1414 , Vol. 3(4),
74-81, April (2014) Int. Res. J. Environment Sci.:
3. Voudouris, K; Voutsa, D. (2012): Water quality monitoring and Assessment.
4. Environmental Chemistry selected Methods for water quality analysis, UNESCO-IHE
Page (26-28).
5. WHO Guidelines for Drinking-water Quality, WHO/SDE/WSH/03.04/17 /Analysis of
Zinc in Drinking-water
6. http:// pipeline.corante .com/ oil extraction by soxhlet method (visited on Sept,15th 2015)
7. http:/ / www.buchi.com/ soxhlet extraction method -: BUCHI-automated-extraction-sys.
17815. 0.html) .
8. http:/ /www.campbell.edu/ faculty/jung/soxhlet.ppt): soxhlet extractor working
explained ppt.
9. http://www.theherbsplace.com/index.html : A Project Report on extraction of essential
oil and its applications Submitted By Virendra P. S. Rao &Diwaker Pandey .
10. http://healingdeva.com/selena2.htm : Extraction of essential oil using steam
distillation”A thesis submitted by K.Satish Kumar.
11. http://healingdeva.com/selena3.htm: Chemical Constituents of Essential oils”.
12. Boniface GATARI (2011), Evaluation of the mineral water consumed in Rwanda: Water
Resources Management sub-sector strategic Plan (2011 – 2015)
13. http://water.epa.gov/action/advisories/drinking/upload/dwstandards2012.pdf
14. Physico-Chemical Parameters for Testing of Present Water Quality of Khan River
at Indore, India.
15. http://water.epa.gov/action/advisories/drinking/upload/dwstandards2012 Pdf.
16. http://www.stevenswater.com: Physicochemical Parameters of Natural Waters
Written by Keith Bellingham Stevens Water Monitoring Systems, Inc.

34
 APPENDICES

APPENDICES 1: preparation of sample :Natural products : soybeans and groundnuts

I chose the good quality of soybeans and groundnuts and weighed 0.25kg for each.
1.Crushing and Grinding the samples were ground into fine particles or powder by using
mortar and pestle. The samples to be ground were placed in the mortar and ground, crushed or
mixed using the pestle.
2.Sieving: the Sieving step separates materials according to size. From the ground sample,
Objects may pass an opening of sieve which is larger than their diameter while being retained by
an opening of smaller diameter
3. powder collection: the powder were collected using the watch glass and weighed.

APPENDICES 2: preparation of solution for analysis of ammonium ions.

The ammonium ion reacts with hypochlorite ions, formed by the alkaline hydrolysis of
dichloroisocyanurate, and with sodium salicylate in presence of sodium nitroprussiate to form a
blue dye which appears green due to the interference with the yellow color of reagent.
We weighed and mixed 3.9g of sodium salicylate, 3.9g of trisodic citrate, and 30.0mg of
sodium nitroprussiate in the same beaker.
We weighed aliquot of 1.0g of sodium hydroxide in closed cap.
For R1: we dissolved salicylate reagent aliquot in 30ml of distilled water .
For R2: we dissolved sodium hydroxide aliquot in 30ml of distilled water, and after cooling,
we added 60.0g of sodium dichloroisocyanurate.

Standard solutions:
we diluted 333x the stock solution to obtain a diluted solution at 3mgNH4+/L : add 0.75ml stock
solution into a well -washed 250m volumetric flask and add distilled water until the horizontal
line .
To prepare different standards, dilute the 3mgNH4/L standard solution in the next proportion :

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Table 8: standard solution preparation

Standards mgNH4/L Vol. mgNH4/L Vol. distilled water Total volume ( ml)
(ml) (ml)

St0 0 0 5 5

St1 0.15 0.25 4.75 5

St2 0.3 0.5 4.5 5

St3 0.45 0.75 4.25 5

St4 0.6 1 4 5

St5 0.75 1.25 3.75 5

St6 1.5 2.5 2.5 5

St7 2.25 3.75 1.25 5

St8 3 5 0 5

Procedures for analyzing Ammonium

Allow the spectrophotometer to warm up for at least 30min.

In 5ml of standards, Milli-Q and samples, add 0. 4ml R1 and mix,
Add 0.4ml R2, mix and wait 20-30min.
Set the spectrophotometer to 655nm wavelength and add distilled water into a 50mm cell set the
spectrophotometer to zero.
Measure absorbance in 50mm cell of standards, Milli-Q and samples after at least 30min.
Record the results.

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