Addis Ababa Institute of Technology (AAiT)

School of Graduate Studies

Department of Chemical Engineering

EFFECT OF PROCESSING ON PHYTOCHEMICALS AND NUTRIENTS COMPOSITION OF

FENUGREEK (TRIGONELLA FOENUM-GRAECUM L.), AND DEVELOPMENT OF VALUE
ADDED PRODUCTS

A Thesis Submitted to the School of Graduate Studies of Addis Ababa University,

Institute of Technology, in Partial Fulfillment of the Requirements for the Degree
of Masters of Science in Chemical Engineering (Food Engineering)

By: Birhane G/Silassie

Advisor: Dr.Eng. Shimelis Admassu (Asso. Prof.)

Addis Ababa, Ethiopia

October, 2012
 ADDIS ABABA INSTITUTE OF TECHNOLOGY (AAiT)
ADDIS ABABA UNIVERSITY

SCHOOL OF GRADUATE STUDIES

DEPARTMENT OF CHEMICAL ENGINEERING

EFFECT OF PROCESSING ON PHYTOCHEMICALS AND NUTRIENTS COMPOSITION OF

FENUGREEK (TRIGONELLA FOENUM-GRAECUM L.), AND DEVELOPMENT OF VALUE
ADDED PRODUCTS

A Thesis Submitted to the School of Graduate Studies of Addis Ababa

University, Institute of Technology, in Partial Fulfillment of the
Requirements for the Degree of Masters of Science in Chemical
Engineering (Food Engineering)

By
Birhane G/Silassie

Approved by the Examining Board

______________________________________ __________________________
(Chairman, Department’s Graduate Committee)

Dr.Eng. Shimelis Admassu (Asso. Prof.) __________________________

(Advisor)

_________________ __________________________
(Internal Examiner)

_________________ __________________________
(External Examiner)

i
 Acknowledgements

First and foremost I would like to thank my advisor Dr.Eng. Shimelis Admassu (Asso. Prof.) for
his advice, guidance, support and encouragement throughout my thesis work. My gratitude also
extends to Eng. Gizachew Shiferaw, Ato Adamu Zegeye, Prof. Kuman Jha., and Ato Mulugeta
Teamir for their valuable advices and suggestions.

I am extremely happy to acknowledge my sincere gratitude to FAFFA Foods Share Company for
its financial support and Ethiopian Spice Extraction Factory for letting me to use their laboratory
facilities and chemicals. I also express my sincere thanks to Debrezeith Agricultural Research
Center which provides me fenugreek seed variety, Addis Ababa Institute of Technology
Department of Chemical Engineering and its staffs for their cooperation during this thesis work,
and, Ethiopian Health and Nutrition Research Institute (EHNRI) and JIJE laboglass PLC for letting
me to use their food analysis laboratory.

I also take this opportunity to express my overwhelming gratitude to my parents for their blessing
and unstinted co-operation in the present venture. This work would not have seen the light of the
day but for the patience, forbearance, constant encouragement and support of my father Mr.Atsbha
Reda and my special heart-felt thanks to my Brother G/Egiziabher in completing this work
successfully.

Finally, the study was joyful and successful because of the brotherhood cooperation of my friends
Habtamu Asmare, Simo Hailu, Derese Mokennen, Tuamelsam Shumuye, Shewarega Habtamu,
Yared Tesfaye, G/Hana Ashine, Eskindir Getachew, Abulkerim Mohammed, Lemesa Etana,
Addisu Kussie and Endiris hassen.

ii
 Abstract

The primary purpose of this research was to study the effects of germination, fermentation,
autoclaving, extraction and their combinations on the phytochemicals and nutrients composition of
fenugreek seed varieties (Challa, Ada’a and Wello). The chemical compositions, physico-chemical
and functional properties, mineral bioavailability, and phytate and non phytate phosphorus contents
of fenugreek seeds were investigated. Furthermore, during polar solvent extraction process, the
influences of rinsing stages, solvent type, and proportion of solvent to water on the rate of
extraction were also examined. The average mean value on dry weight basis of fenugreek’s
moisture content was 7.5%, crude protein 29.3%, crude fat 7.9%, crude, fiber 7%, total ash 3% and
total carbohydrates 45.2%. The major mineral constituents of fenugreek seeds were K (10351.7
mg/kg), P (5651.3 mg/kg), Mg (1257.3 mg/kg) and Na (121.5 mg/kg), Ca (553.7 mg/kg), Fe (81.3
mg/kg) and Zn (43.4 mg/kg). Saponins, phytates and tannins concentrations ranged from 4581.3 to
6320.8, 76.11to 111.08 and 235.24 to 269.13mg/100g; respectively. Water and oil absorption
capacities were 7.5, 10.2 & 7.2 g/g of water and 1.04, 1.12 & 2.3g/g of oil absorption capacity for
Challa, Ada’a and Wello; respectively. Crude protein, fat, fiber, mineral bioavailability and non
phytate phosphorus compositions were improved during processing methods. Significant reduction
of phytochemicals was achieved by employing conventional, heat and chemical processing
methods. Germination reduced phytates, tannins, and saponins by 47.9 – 66.4%, 33.7 – 56.8%, &
46.3 – 69.9%, fermentation 68.7 – 100%, 40.7 – 87.9% & 67.8 – 81.0%, autoclaving 32.5 – 36.4%,
66.4 – 74.7%, & 88.6 – 94.6% and extraction 6.7 – 7%, 100%, & 100%; respectively. Extraction
was less effective in reducing phytates as compared to other processing methods due to insolubility
of phytates in alcohol (ethanol &methanol). The combination of germination and fermentation
resulted in a significant and successive reduction in phytates, tannins and saponins levels. However,
complete removal of all phytochemicals was achieved by the combination of fermentation followed
by extraction process. During extraction, as the proportion of water increases the percolation rate
decreases due to water binding property of fenugreek. Methanol extraction was faster and powerful
than ethanol extraction. The extraction process also removed the astringent smell and bitterness of
fenugreek. Thus, debitterized fenugreek powder was produced. In addition, to preserve the health
benefits and to maintain dose of fenugreek required for different age groups, sex and health
conditions, fenugreek capsules were also produced.

Key words: Chemical composition, extraction rate, Fenugreek, phytochemical, processing methods,
solvent

iii
 Table of Contents

Chapter Title Pages

Title page i
Acknowledgement ii
Abstract iii
Table of contents iv
List of tables viii
List of figures x
List of abbreviations xi

1 Introduction 1

1.1. Background 1

1.2. Statement of the problem 2

1.3. Objectives 3

1.4. Scope of the study 4

1.5. Significance of the project 4

2 Literature Review 5

2.1. Overview of fenugreek 5

2.1.1. Origin, distribution, production and different species of

fenugreek in the world 6

2.1.2. Production and distribution of fenugreek in Ethiopia 8

2.1.3. Uses of fenugreek 8

2.2. Nutraceutical properties of fenugreek 9

2.3. Functional food aspects of fenugreek 11

2.4. Major phytochemicals of fenugreek 12

2.4.1. Phytic Acid 13

iv
 2.4.2. Phenolic compounds 14

2.4.3. Saponins 15

2.5. Processing methods to reduce phytochemicals 16

3 Materials and Methods 19

3.1. Experiment Location, materials, chemicals and reagents 19

3.2. Processing methods and their combination 20

3.2.1. Germination 20

3.2.2. Autoclaving 20

3.2.3. Natural fermentation 21

3.2.4. Extraction 21

3.2.5. Combination of processes 21

3.3. Analysis methods 21

3.3.1. Proximate chemical composition 21

3.3.2. Physico-chemical properties of fenugreek 26

3.3.3. Functional properties of fenugreek flour 27

3.3.4. Phytochemicals composition in fenugreek 28

3.4. Development of value added products from fenugreek 29

3.5. Sensory analysis 30

3.6. Experimental setup of the thesis 30

3.7. Experimental design and statistical data analysis 32

4 Results and Discussion 33

4.1. Proximate, mineral and phytochemicals composition of fenugreek seeds 33

4.1.1. Proximate composition of fenugreek 33

v
 4.1.2. Mineral composition of fenugreek 34

4.1.3. Phytochemicals composition of fenugreek 36

4.2. Physicochemical properties of fenugreek 37

4.3. Functional properties of fenugreek 39

4.4. Effect of processing on the proximate and mineral compositions of

fenugreek 41

4.4.1. Effect of processing methods on the proximate compositions of

fenugreek 41

4.4.2. Effect of processing methods on mineral contents of fenugreek 44

4.4.3. Effect of processing methods on phytate phosphorus and non

phytate phosphorous contents of fenugreek 46

4.4.4. Effect of processing methods on mineral bioavailability of

fenugreek 48

4.5. Effect of processing methods on phytochemicals composition of fenugreek 49

4.5.1. Effect of germination 50

4.5.2. Effect of natural fermentation 51

4.5.3. Effect of autoclaving 52

4.5.4. Effect of extraction 54

4.5.5. Effect of combination of processes 54

4.6. Effect of rinsing stage, solvent type and water to solvent proportion on
extraction rate of fenugreek flour 55

4.7. Comparison of effect of various processing methods on phytochemicals 57

4.8. Sensory evaluation of value added products 59

5 Techno-economic Feasibility for Fenugreek Products 60

5.1. Process description on production of debitterized fenugreek powder 60

vi
 5.2. Process description on fenugreek capsule production 61

5.3. Material and energy balance 64

5.4. Financial and economic analysis 79

5.4.1. Plant capacity and production programming 79

5.4.2. Project cost estimation 80

5.4.3. Financial Evaluation and project appraisal 84

6 Conclusions and Recommendation 89

6.1. Conclusions 89

6.2. Recommendations 90

References 91

Appendices 100

vii
 List of Tables

Table Title Page

2.1 Fenugreek and its associated medicinal properties 10

2.2 Summary of research progress with respect to the functional food aspect of
fenugreek 11

3.1 AOAC (2000) official methods used for proximate chemical composition analysis 22

4.1 Proximate composition of fenugreek varieties (g/100 g DMd)ã of whole seed

samples 34

4.2 Mineral composition of raw fenugreek varieties (on dry matter basis) 35

4.3 Phytochemicals composition of raw fenugreek varieties 37

4.4 General characteristics of fenugreek seed varieties 38

4.5 Physicochemical properties of Fenugreek seed varieties 38

4.6 Water and oil absorption capacity of flours of raw fenugreek varieties 39

4.7 Effect of processing on proximate composition of Challa variety (g/100 g DMd) ã 41

4.8 Effect of processing on proximate composition of Ada’a cultivar (g/100 g DMd) ã 42

4.9 Effect of processing methods on mineral compositions of Challa 45

4.10 Effect of processing methods on mineral compositions of Ada’a 45

4.11 Effect of processing methods on phytate phosphorous and non phytate

phosphorus of Challa 46

4.12 Effect of processing methods on phytate phosphorous and non phytate

phosphorus of Ada'a 47

4.13 Effect of processing on molar ratios of phytate: Fe, phytate: zinc, Phytate: Ca, and
[calcium*phytate]: [zinc] for Challa 48

4.14 Effect of processing on molar ratios of phytate: Fe, phytate: zinc, Phytate: Ca, and
[calcium*phytate]: [zinc] for Ada’a 49

viii
4.15 Effect of processing methods on phytochemical compositions of Challa variety 52

4.16 Effect of processing methods on phytochemical compositions of Ada’a cultivar 53

4.17 Effect of rinsing stage and alcohol to water ratio on percolation rate 56

4.18 Sensory evaluation of extracted fenugreek flours and capsules 59

5.1 Plant capacity and production programming 79

5.2 Purchased equipment cost 80

5.3 Total investment cost 81

5.4 Cost of manpower 83

5.5 Total product cost estimation 83

5.6 Projected profit and loss 85

5.7 Data required for break-even analysis calculation 86

ix
 List of Figures

Figure Title Page

2.1 Fenugreek leaves and seeds 5

2.2 World map showing distribution of fenugreek (Trigonella foenumgraecum L.) 7

3.1 Fenugreek seed varieties used in this thesis study 19

3.2 Framework of experiments carried out in this thesis 31

4.1 Comparison of effect of processing methods on phytochemicals of Challa

variety 58

4.2 Comparison of effect of processing methods on phytochemicals of Ada’a

variety 58

5.1 Flow sheet diagram for the production of debitterized fenugreek powder 60

5.2 Flow sheet diagram for the production of fenugreek capsule 61

5.3 Equipment layout for debitterized fenugreek powder and fenugreek capsule
production 62

5.4 Break-Even analysis for production of debitterized fenugreek powder 88

x
 List of Abbreviations

AAFRD Alberta Agriculture, Food and Rural Development

AOAC Association of Official Analytical Chemistry

CSA Central Statistical Agency

CF Control Fermentation

EHNRI Ethiopian Health and Nutrition Research Institute

FAO Food and Agriculture Organization

IBC Institute of Biodiversity Conservation

NF Natural Fermentation

PA Phytic acid

PGRFA Plant Genetic Resources for Food and Agriculture

Qtl Quintal

USDA United States Department of Agriculture

W/V Weight/Volume

W/W Weight/Weight

xi
 CHAPTER ONE

1. Introduction

1.1. Background

Spices are natural food adjuncts that have been in use for thousands of years to enhance the sensory
quality of foods. Spices impart characteristic flavour, aroma, taste and colour to foods. Some
spices, like fenugreek, can also modify the texture of food (Srinivasan, 2006).

Fenugreek (Trigonella foenum-graecum L.) is an erect annual herb native to Southern Europe and
Asia, belonging to the family Leguminosae. Over 80% of the total world's production of this seed is
contributed by India, one of the major producers and exporters of fenugreek legume in the world.
The seeds are used as spices worldwide, whereas the leaves are used as green leafy vegetables in
the diet. Fenugreek seeds are bitter to taste and are known for a long time for their medicinal
qualities (Saikat, 2006). The seeds of fenugreek have high protein content (25%), lysins (5.7g/16g
of nitrogen), soluble (20%) and insoluble (28%) dietary fibre, L-tryptophan and trigonelline besides
being rich in calcium, iron and beta-carotene (Sharma, 1986).

Fenugreek is regarded as the oldest known medicinal plant in recorded history. Its seed and leaves
have medicinal value, and have been used to reduce blood sugar and lower blood cholesterol in
humans and animals (Dahanukar et al., 2000). Several health beneficial attributes of the spice,
fenugreek, have been experimentally evidenced in recent decades, which have the potential of
possible therapeutic application. In view of these promising beneficial physiological effects,
fenugreek is understood to exert galctagogue, cholesterol-lowering, antidiabetic, digestive
stimulant, antioxidant, gastroprotective and hepatoprotective effects (Srinivasan, 2006). The
therapeutic uses of fenugreek also include its use as hypoglycemic, antiulcerogen
hypocholesterolemic and antihypertensive agent (Sharma et al., 1996 & Tayyaba et al., 2001). It is
also widely used as a galctagogue (milk producing agent) by nursing mothers to increase
inadequate breast milk supply. Studies have shown that fenugreek is a potent stimulator of breast
milk production (Chantry et al., 2004).

Commercially, fenugreek has been utilized in the food industry as stabilizer, emulsifier, thickening
agent, spice, supplement of wheat and rice flour, beverage, coffee substitute, and tea. In Ethiopia, it
is known as “Abish” and has been considered as medicinal plant to lower blood glucose level, treat
gastric disease and abdominal discomfort, boost milk production in lactating mothers, and to

1
increase body weight. It is also used, for softening and preserving of “Ingera”, infant food
preparation, food blend, as beverage, spice and cosmetics as jelling agent for hair.

However, the seeds are bitter in taste due to the presence of bitter saponins, which limit their
acceptability in foods (Sharma 1986; Udayasekhera et al., 1987). Fenugreek has been found to
contain relatively large quantities of saponins (Valette et al., 1984). On the other hand, it has been
possible to debitterize fenugreek seeds by employing various processing methods such as soaking,
germination, roasting, etc. (Sharma, 1986; Shashi, 1997). As fenugreek seeds are rich in
mucilaginous fiber and other dietary essentials, their use can be exploited as functional and
nutritional foods as well as therapeutic agent. By keeping these facts in view various value-added
baked and extruded products from wheat-fenugreek flour blends have been developed (Hooda and
Jood 2003b).

Therefore, in this project the effects of processing methods include germination, autoclaving,
fermentation, extraction and their combinations on nutrients and phytochemicals composition of
fenugreek seeds were examined and value added products were developed.

1.2. Statement of the Problem

Fenugreek has been used as food spice and medicinal plant in many countries for centuries. It is a
good source of essential amino acids, soluble and insoluble dietary fiber, and minerals. Now it is
considered as one of the most important nutraceutical food especially due to its ability to reduce
cholesterol level and its antidiabetic, antioxidant, anticarcinogenic, galctagogue, and antibiotic
effects.

However, it contains phytochemicals such as phytates which binds minerals and interferes with
their availability, and polyphenols and tannin that hinder the absorption of nutrients and the
bioactive compound, bitter saponin that limits acceptability of fenugreek. Hence, the presence of
these phytochemicals hinders the versatile application and exploitation of fenugreek as:

 The phytates as an antinutrient interfere with the availability of some important minerals
e.g. calcium, iron and zinc.

 The tannins interact with the protein to form complexes. This interaction increases the
degree of cross linking, decreasing the solubility of protein and making protein complexes
less susceptible to proteolytic attacks, thereby adversely affecting the protein digestibility
and protein quality.

2
  The presence of bitter saponins, alkaloids and volatile acids in fenugreek seeds and,

 The astringent smell, like maple syrup, of sweat and urine limits their acceptability for
consumption.

In addition, despite of some researches on its medical value, fenugreek has not been well studied
with respect to nutritional composition, phytochemicals concentration, processing characteristics
and value added products development. Especially in Ethiopia, the historical and traditional
utilization of fenugreek has not been supported and improved by academic researches regarding its
agro-climatic preference, productivity, disease resistance, variety distribution, storage and handling
conditions, chemical and nutritional composition, processing quality, therapeutic and functional
values, product diversification, and process technology developments. Therefore, this study was
conducted at the expectation that it contributes its part to pursuit the stated problems and to enhance
the awareness and exploitation of the functional and nutritional values of fenugreek.

1.3. Objectives

General objectives

The overall objective of this study was to investigate the effects of processing methods on the
nutrients and phytochemicals composition, and development of value added products from
fenugreek seed varieties grown in Ethiopia.

Specific objective

Specific objectives of this work were to:

1. Study proximate and mineral composition, physico-chemical, and functional characteristics

of three selected fenugreek seed varieties.
2. Quantify the levels of saponins, phytic acid and tannins in raw and processed fenugreek.
3. Evaluate effects of processing methods (germination, autoclaving, fermentation, extraction
and their combinations) on nutrients, minerals and phytochemicals concentration, and
mineral bioavailability of fenugreek.
4. Study the effects of solvent type, rinsing stage and water to solvent proportion on extraction
rate of fenugreek flour.
5. Develop value added products and conduct sensory quality evaluation.
6. Conduct techno-economic feasibility study on fenugreek products.

3
 1.4. Scope of the study

In this research physicochemical, proximate and phytochemicals composition of three fenugreek

seed varieties were studied and two varieties were selected for processes such as germination,
autoclaving, fermentation, extraction and their combinations. Different experiments were carried
out to evaluate the composition of processed samples. Then, value added products were produced
and analyzed for their compositions.

1.5. Significance of the project

Most Ethiopians are aware of the advantages and health benefits of consuming fenugreek but they
use it only when they have abdominal discomfort, feel weakness, and in general when they are
forced to take at during sickness and other situations due to the bitter test and astringent smell of
their sweat after consuming of fenugreek. In addition, there is high mortality rate of children under
the age of five and mothers due to insufficient supply of milk to children, and nutrient and
complementary food to mothers.

On the other hand, if the side effects of phytochemicals are reduced, fenugreek has high protein
content, lysine which is not found in most cereals, soluble and insoluble dietary fibre, fat and
essential amino acids besides being rich in magnesium, calcium, iron, zinc and beta-carotene. It has
also health benefits as it prevents diabetics, reduces cholesterol level, and its antioxidant,
anticarcinogenic and antibiotic effects, and increases milk production in lactating mothers.

In addition, Ethiopia has suitable agroclimatic conditions and land for the growing of all fenugreek
varieties. Therefore, this project:

 Creates awareness and contributes additional knowledge regarding to the composition,

processing quality, product diversification and utilization of fenugreek.
 Enables Ethiopia to export value added products.
 Enable to exploit the therapeutic and functional values of fenugreek.
 Use to combat the hidden hunger, shortage of micronutrients, through improving the
bioavailability of minerals.
 Alleviate the health of the society.
 Supports the prevention based health policy of the country.

4
 CHAPTER TWO

2. Literature Review

This literature review covers an overview of fenugreek, nutraceutical and functional food aspects of
fenugreek, phytochemical components in fenugreek seeds, food processing methods including
conventional, heat, chemical methods and their combinations that reduce phytochemicals and
enhance nutritional value of fenugreek are briefly discussed.

2.1. Overview of fenugreek

Fenugreek (Trigonella foenum-graecum L.) is an annual crop belonging to the legume family.
Although grown as a spice in most parts of the world, the species name "foenum-graecum" means
"Greek hay" indicating its use as a forage crop in the past (Petropoulos 2002).

The seed sown on a prepared soil sprouts in 3 days, grows inherently resistant to most infestations
and diseases to a robust, erect plant to a height of 30–60 cm, with compound pinnate trifoliate
leaves, auxiliary white to yellowish flowers, and 3–15 cm long thin pointed beaked pods, which
contain 10–20 oblong greenish-brown seeds with unique hooplike groove. As a leguminous plant, it
fixes atmospheric nitrogen, thus enriching the soil. Greens are harvested at the 3 or 4 leaved stage,
after which saponin content increases, making them bitterer. The seeds are harvested 30–35 days
after flowering or 155–165 days after sowing (Chatterjee and prakashi, 1995).

Fenugreek plant Fenugreek seeds

Source: Srinivasan (2006)
Figure 2.1: Fenugreek leaves and seeds

Fenugreek (Trigonella foenum-graecum L.) is an annual, self-pollinated plant from Leguminosae

family with small seeds and since ancient times has always been known as a medicinal herb
(Slinkard, 2006). Its leaves are consumed as leafy green vegetables in India and are rich in calcium,
iron, carotene B and other vitamins (Sharma, 1986). Fenugreek seeds are rich in protein, fixed oils
and minerals; thus, it is quite nutritious. Fenugreek protein is also rich in lysine amino acid and in

5
terms of supplying the protein requirements of the human body is similar to soybean protein
(Hidvegi et al., 1984).

2.1.1. Origin, distribution, production and different species of fenugreek in the

world

Different authors have widely divergent opinions about the probable ancestry of T. foenum-
graecum. Vavilov (1951) has suggested that fenugreek is native to the Mediterranean region, while
Fazli and Hardman (1968) proposed an Asian origin for the crop. Indigenous species have been
reported by Petropoulos (2002) on the continents of Asia, Europe, Africa and Australia. Fenugreek
also is reported as a cultivated crop in parts of Europe, Africa, west and south Asia, North and
South America and Australia (Edison, 1995; Ethiopia: Country Report, 1996; Petropoulos 2002).

The exact number of species of fenugreek also has been debated. Petropoulos (2002) indicated that
older taxonomists like Linnaeus have suggested that as many as 260 species of fenugreek exist. In
contrast, about 128 species of fenugreek were reported by Vasil'chenko (1953), 97 by Fazli (1967)
and 70 by Rouk and Mangesha (1963).

Cultivation climate and soil

Fenugreek is suitable for areas with moderate or low rainfall. A temperate and cool growing season
without extreme temperatures is favourable for the best development of fenugreek. It can tolerate
10 –15 oC of frost (Duke, 1986). Fenugreek is fairly drought resistant and fairly frost sensitive
(Talelis, 1967). According to Del’ Gaudio (1952), fenugreek adapts well during summer droughts
and the wet and raining winter, while it does not like severe winter and raining summer, but it is
resistant to winter cold, especially when it is covered with snow. Also, Rouk and Mangesha (1963)
noticed that in Ethiopia fenugreek is grown primarily in regions where the climatic conditions
approach those of the Mediterranean area. The climate of these regions is mostly subtropical and is
characterized by a wet followed by a dry season. Duke (1986) reports also that fenugreek grows
fairly well on gravely or sandy soils and it is not adaptable to heavy clay or soil that becomes hard
and it is fairly tolerant to salt. Bunting (1972) also reports that heavy and wet soils are unsuitable
for cultivation of fenugreek and mentions as an optimum pH 8–8.5.

Distribution, cultivated area and production of fenugreek in the world

The species T. foenum-graecum, wild or cultivated, is widely distributed throughout the world, as is
indicated by the great number of names it possesses with Arabic, Indian (Sanskrit) and European
(Greek and Latin) roots. Fenugreek has been reported as a cultivated crop in Portugal, Spain,

6
United Kingdom, Germany, Austria, Switzerland, Greece, Turkey, Egypt, Sudan, Ethiopia, Kenya,
Tanzania, Israel, Lebanon, Morocco, Tunisia, India, Pakistan, China, Japan, Russia, Argentine and
the United States of America (Rouk and Mangesha, 1963; Fazli and Hardman, 1968). At the
present time fenugreek is an important cash crop in India (the leading fenugreek producing
country), Morocco, China, Pakistan, Spain, Tunisia, Turkey, Lebanon, Israel, Egypt, Ethiopia,
Kenya, Tanzania etc. (Smith, 1982; Edison, 1995).

Figure 2.2: World map showing distribution of fenugreek (Trigonella foenumgraecum L.)

As far as the world cultivated area of fenugreek and the annual production of seed are concerned,
statistics are very limited and scattered, as the area seeded with fenugreek is relatively small and
not recorded by the agricultural statistics of different countries. In spite of this, the following
analysis based on the exported quantities of the principal producing countries, the domestic use of
fenugreek and the existing statistics of the cultivated area for some countries represent a reasonably
accurate assessment of the world production and cultivated area of fenugreek.

1. The cultivated area of fenugreek in India, an average for the last twenty years (1975–95),
accounts for 34,534 ha with a production of 41,530 tons and an export of 4203 tons, that is
domestic use accounts for 90 per cent of the production (Anonymous, 1996).
2. Recently, there has been an increase in the export of fenugreek from India: in 1994–95 it
accounted for 7,956 tons (Anonymous, 1996). According to Edison (1995) India claims 70–
80 per cent of the world export in fenugreek. This means that the world export of fenugreek
until 1995 fluctuated around 10,500 tons, and export from the other countries mentioned
above can be estimated as approaching 2,700 tons. According to the forementioned
considerations, the cultivated area from these countries accounts for about 22,000 ha with a
production of 26,700 tons.

7
 3. These considerations permit us to estimate that in the world, the annually cultivated area of
fenugreek amounts to roughly 57,000 ha with a seed production of 68,000 tons.

2.1.2. Production and distribution of fenugreek in Ethiopia

Fenugreek is locally used as a pulse, spice and medicinal plant, and has a long history in Ethiopia.
Even though the hectarage is limited, the species has a considerable genetic diversity in seed
colour, maturity and other morpho-agronomic characters (IBC, 2008).

Characterization is essential for effective utilization of conserved germplasm. Characterization and

preliminary evaluation on basic morpho-agronomic characteristics have been undertaken on about
70 percent of the crop germplasm accessions, including 34648 cereals, 8037 oil crops, 5355 pulses,
424 coffee and 360 accessions of fenugreek (IBC, 2008).

Currently, 271,220.46 Quintals of fenugreek seeds were produced and about 51.09 per cent of
fenugreek was used for household consumption, 15.19 per cent for seed and about 32.88 per cent
for sale. The remaining 0.84 per cent of fenugreek was used for wages, animal feed and others
(CSA, 2009/10).

2.1.3. Uses of fenugreek

Human food

Fenugreek is a chemurgical cash crop, usually cultivated as a break crop for cereal, as it is
considered a good soil renovator. The whole plant is used as forage and vegetable, while the seeds
(whole, powdered, in flour, or roasted) are used as human and animal food, spice, dyeing,
flavouring, as well as for medicinal and industrial purposes (Petropoulos, 2002).

Young plants and fresh tips of fenugreek are succulent and eaten as a salad, or cooked and
generally served as a condiment in India and Egypt, as the fresh plant is very rich in vitamin C (207
mg per cent) (Saleh et al., 1977).

The fenugreek seed is rich in protein, fixed oils and minerals and so it is nutritive and a tonic
(Anonymous, 1994). In Sudan and Egypt the seeds are used in making beverages and in some
countries the roasted seeds are used as a coffee substitute, probably because of the alkaloid
trigonelline content, which is a basic constituent of the coffee seed. While in Ethiopia the seeds are
prepared for infant feeding by boiling the whole seed (Fazli and Hardman, 1968), in North Africa it
is mixed with breadstuff (Manniche, 1989); in Egypt also the seeds of the fenugreek are added to

8
bread as a supplement of wheat and maize (Hidvegi et al., 1984). In Yemen it is widely used every
day by the general population (Manniche, 1989).

Spice

As a spice, fenugreek seeds add nutritive value to food, as well as flavouring and are used in soups
and curries (Duke, 1986). Fenugreek seed is commonly used for seasoning purposes and as an
ingredient of curry powder and sauces (Fazli and Hardman, 1968).

Animal feed

Originally, it was grown in the ancient world and especially in Europe and was recognized as good
forage, hence the name ‘Greek hay’ or foenum-graecum (Rouk and Mangesha, 1963). Hidvegi et
al. (1984) report that fenugreek seeds are used for feeding cattle. Ground fine and mixed with
cotton seed it is fed to cows to increase the flow of milk.

Others

Pharmaceuticals and cosmetics, flavouring agent, dyeing, remedy, herbal medicine and industrial
material (Fazli and Hardman, 1968). For Repellent flavouring perfume (Anonymous, 1994).

2.2. Nutraceutical properties of fenugreek

Fenugreek has been referred to as a medicinal herb both in Indian Ayurvedic and traditional
Chinese medicines (Tiran 2003). Ancient literature, religious scripture, travel records and anecdotes
from different continents and from different periods of human history, record a wide variety of
medicinal properties associated with fenugreek (Lust 1986). Medicinal uses vary from wound-
healing to bust enhancement and, from promotion of lactation in weaning mothers, to its use as a
sex stimulant or aphrodisiac (Petropoulos 2002; Tiran 2003).

Fenugreek is reported to have anti-diabetic, anti-fertility, anticancer, anti-microbial, anti-parasitic

and hypocholesterolaemic effects (Al-Habori and Raman, 2002) and protective effect on ethanol
toxicity (Thirunavukkarasu et al., 2003). Some medicinal properties attributed to fenugreek are
presented in Table 2.1.

Fenugreek contains three important chemical constituents with medicinal value; i.e., steroidal
sapogenins, galactomannans and isoleucine. These constituents have placed fenugreek among the
most commonly recognized "nutraceutical" or health food products (Srichamroen et al., 2005).

9
  Steroidal sapogenins

Fenugreek seed is an important source of steroidal sapogenins such as diosgenin which are used
extensively by both pharmaceutical and nutraceutical industries (Srichamroen et al. 2005).
Diosgenin is often used as a raw precursor for the production of steroidal drugs and hormones such
as testosterone, glucocorticoids and progesterone (Fazli and Hardman, 1968; Raghuram et al. 1994;
Srichamroen et al. 2005). McAnuff et al. (2002) reported that steroidal sapogenins are effective
agents for the treatment of hypocholesterolemia, a disorder often associated with diabetes.

 Galactomannans

Fenugreek galactomannans appear to be beneficial for control of type 2 diabetes in both animals
and humans (Sharma et al., 1996) by reducing hyperglycemia in these individuals. They have a
high water-binding capacity and are able to form highly viscous solutions at relatively low
concentration, which appears to reduce glucose absorption in the digestive tract (Raghuram et al.,
1994).

Table 2.1: Fenugreek and its associated medicinal properties.

Medicinal properties reported References

Anti-diabetic and cholesterol lowering properties Hannan et al., (2003)
Anti- hyperthyroidism Tahiliani and Kar (2003a)
Against thyroxine- induced hyperglycaemia Tahiliani and Kar (2003b)
Anti-cancer effects Devasena and Menon (2003)
Gastro-protective and hepatoprotective effect Srinivasan (2006)
Antioxidant property Raskin et al., (2002)
Antinociceptive property Javan et al., (1997)
Antimicrobial property Bhatti et al., (1996)
Anthelmintic property Ghafgazi et al., (1980)
Anti-sterility and anti-androgenic effects Kamal et al., (1993)
Wound healing property Taranalli and Kuppast (1996)
Anti-inflammatory and antipyretic actions Ahmadiani et al. (2001)
Galctagogue Srinivasan (2006)
Digestive stimulant Srinivasan (2006)
 Isoleucine

The amino acid isoleucine is a precursor of 4-hydroxyisoleucine which is known to regulate the
secretion of insulin in animals (Broca et al., 2000). Most hypoglycaemic and anti-hyperglycaemic

10
effects of fenugreek are attributed to the gastrointestinal effect of dietary fiber and systemic effects
of amino acids like 4-hydroxyisoleucine present in the seed (Petropoulos, 2002).

2.3. Functional food aspects of fenugreek

A summary showing studies associated with functional foods developed with fenugreek additives is
presented in Table 2.2.

Table 2.2: Summary of research progress with respect to the functional food aspect of fenugreek

Functional food aspect of fenugreek References

Fenugreek seed has protein, lipid, cellulose starch and ash. It is also Sauvaire et al., (1976)
reported to be rich in calcium, iron and β-carotene.
Fenugreek seed endosperm is characterized by soluble fiber. Aspinall (1980)
Approximately half the dry weight of fenugreek seed has Soluble
Dietary Fibers (SDF), an edible dietary fiber
Boiling of fenugreek seed results in loss of crude proteins, total sugars, Ismail (1996)
ash but subsequently increases the level of crude fibers; without any
significant effect on lipid levels.
Good quality wheat breads with high nutritional characteristics and (Bakr 1997)
higher acceptability have been produced in Egypt by supplementation
of 4 % fenugreek flour.
Cooking fenugreek leaves in a pressure cooker improve retention of Yadav and Sehgal (1997)
essential chemical nutrients such as ascorbic acid and beta-carotene
Fenugreek galactomannans as food emulsifiers Gatri et al., (1997)
Fenugreek seed contain sufficient ascorbic acid Riddoch et al., (1998)
Nutritional quality of lactic acid fermented fenugreek leaves Gupta et al., (1998)
Fenugreek fibers, psyllium husk and wheat bran could be used as Al-Khalidi et al., (2001)
dietary supplements to increase roughage in the human diet
Supplementation of wheat flour with ground debittered fenugreek Sharma and Chauhan
improves the physicochemical, nutritional and rheological properties (2000)
Suitability of the development of food products based on millets, Pathak et al., (2000)
legumes and fenugreek seed in diabetic diet
Use of corn bread mixed with a small amount of fenugreek (3 %) or Galal (2001)
with wheat flour (30 %) is used as staple food in Egypt
Fenugreek contains 23-26 % protein, 6-7 % fat and 58 % carbohydrate USDA (2001)
and 25 % dietary fiber.
Properties of fenugreek mucilage and its use in ice cream as a stabilizer Balyan et al., (2001)
Nutritional value and physiological properties of fenugreek. Billaud and Adrian (2001)
Improvement of oxidative stability of raw food products like eggs by Armitage et al., (2002)

11
fenugreek
Fenugreek seed mucilages can be used as viscosity builders Seghal et al., (2002)
Supplementation of fenugreek flour in rice bran improved the physical Sharma and Chauhan
and sensory properties and qualities of breads and cookies (2002).
Reduction in phytic acid levels and simultaneous in vitro increase with Bhatia and Khetrapaul
respect to calcium and iron with higher temperature and longer (2002)
fermentation duration in case of fenugreek supplemented Indian bread
Analysed the composition of raw, soaked and germinated seed and Hooda and Jood (2003a.)
found that nutritional quality of fenugreek seed could be improved with
subsequent reduction of bitterness through processing
Physiological, rheological and organoleptic characteristics of wheat Hooda and Jood (2003b.)
fenugreek supplemented blends showed increase in protein and fat
contents

2.4. Major Phytochemicals of fenugreek

Phytochemical comes from the Greek word “Phyto” for plant. It refers to every naturally occurring
chemical presents in plants. In plants, phytochemicals act as a natural defense system for host
plants and provide colour, aroma and flavour (Okwu, 2004). There is a wide distribution of
biologically-active constituents throughout the plant kingdom, particularly in plants used as animal
feeding stuff and in human nutrition (Igile, 1996). Phytochemicals are present in a variety of plants
utilized as important components of both human and animal diets. These include fruits, seeds, herbs
and vegetables. Herbs and spices are harmless sources for obtaining natural antioxidants (Okwu,
2004).

Phytochemicals found in plant foods have both adverse effects and health benefits. For example,
phytic acid, lectins, phenolic compounds (tannins), saponins and enzyme (amylase and protease)
inhibitors have been shown to reduce the availability of nutrients and cause growth inhibition,
while phytoestrogens and lignans have been linked with infertility problems. However, phytic acid,
lectins, phenolic compounds, amylase inhibitors and saponins have also been shown to reduce the
blood glucose and insulin responses to starchy foods and/or the plasma cholesterol and
triglycerides. In addition, phytic acid, phenolics, saponins, protease inhibitors, phytoestrogens and
lignans have been related to reduce cancer risks. Because antinutrients can also be mitigating
agents, they need re-evaluation and perhaps a change in name in the future (Sharma, 1984; Sharma,
1986; Sidhu & Oakenful, 1986).

12
 2.4.1. Phytic Acid

Phytic acid (myoinositol 1,2,3,4,5,6, hexakis-dihydrogen phosphate; PA) is present in foods in

concentrations ranging from 0.1 to 6.0%. It is found as crystalline globoid inside protein bodies in
the cotyledon of legumes or oilseeds or in the bran region of the cereal grains (Reddy et al., 1982).
Salts of phytic acid, designated as phytates, are found in plants, animals and soil. Phytate is
ubiquitous among plant seeds and grains, comprising 0.5 to 5 percent (w/w). It is primarily present
as a salt of the mono- and divalent cations K+, Mg 2+, and Ca 2+ and accumulates in the seeds during
the ripening period. Phytate is regarded as the primary storage form of both phosphate and inositol
in plant seeds and grains (Loewus, 2002).

Because phytate is a naturally occurring compound formed during maturation of plant seeds and
grains, it is a common constituent of plant-derived foods. Depending on the amount of plant-
derived foods in the diet and the grade of food processing, the daily intake of phytate can be as high
as 4500 mg (Reddy, 2002).

Figure 2.2: Proposed structures of phytic acid.

Source: Reddy (2002)

Health benefits of phytate

A significant negative relationship was observed between the blood glucose response to different
starchy foods, expressed as glycemic index, and the intake of phytic acid (Yoon et al., 1983). The
addition of PA (0.2 - 9%) to the diet of rats significantly reduced the plasma cholesterol and
triglyceride levels (Sharma, 1984). This was suggested to be related to the ability of PA to bind to
Zn and thus lower the plasma Zn to copper (Cu) ratio; lower ratios tend to predispose humans to
cardiovascular disease.

Certain minerals such as iron and copper catalyze oxidative enzymes that generate free radicals,
resulting in undesirable oxidative damage such as cell membrane damage (produces leaky cells).
Because phytates have the ability to chelate minerals that participate in undesirable oxidative

13
reactions, phytates have been suggested to have protective effects. The ability of phytates to chelate
the divalent minerals makes them a natural antioxidant.

Phytate as an antinutrient

The major concern about the presence of phytate in the diet is its negative effect on mineral uptake.
2+ 2+ / 3+ 2+ 2+ 2+ 2+
Minerals of concern in this regard would include Zn , Fe , Ca , Mg , Mn , and Cu
(Vucenik, 2003). Its highly negatively charged structure at a wide range of pH values makes it very
reactive with other positively charged ions such as minerals, forming insoluble complexes which
are less available for digestion and absorption in the small intestine. This is the main reason why
PA has traditionally been considered as an antinutrient. The adverse effect of PA in mineral
availability depends on a number of factors including the concentration of PA and the strength of
its binding with different minerals. For example, zinc (Zn) forms one of the strongest mineral
complexes with PA (Evans & Martin, 1988).

PA can also react directly with the positively charged group or indirectly with the negatively
charged group of the proteins mediated by a positively charged mineral ion such as calcium. It can
bind with starch either directly by hydrogen bonding with the phosphate group or indirectly through
the proteins to which it is associated with. The formation of these complexes is likewise thought to
reduce the solubility and digestibility of the proteins or starch and several in-vitro (Carnovale et al.,
1988) studies have indeed shown reductions in protein digestibility by PA and in-vivo (Atwal et al.,
1980).

2.4.2. Phenolic compounds

Phenolic compounds encompass a wide variety of compounds characterized by the presence of an

aromatic ring with one or more hydroxyl groups and a variety of substituents. Flavonoids have a
basic C6C3-C6 structure and include the anthocyanin pigments, flavonols, flavanoids and
isoflavones. They occur mostly as glycosides except the flavanols which tend to polymerise to
condensed tannins. The tannins could be classified either as condensed or hydrolysable. Most
condensed tannins are polymers of flavan-3-ols (catechins) or flavan-3, 4-diols
(leucoanthocyanidins) while most hydrolysable tannins are glucose or polyhydric alcohol esterified
with gallic acid (gallotannins) or hexahydrodiphenic acid (ellagitannins). The stable dilactone of the
latter is ellagic acid (Deshpande et al., 1984). Condensed tannins occur widely in cereals and
legumes. These compounds are concentrated in the bran fraction of cereals. Tannins may be
classified as polyphenolic substances (Shimelis and Rakshit, 2007).

14
Health benefits of Phenolic compounds

Several phenolics (e.g. chlorogenic acid, gallic acid, caffeic acid, tannit acid, catechin) have been
shown to inhibit the mutagenic effects of both direct-acting carcinogens (e.g. benzo(a)pyrene diol
epoxide) and carcinogens that require metabolic activation (e.g. aflatoxin B) and to trap nitrite, thus
reducing the nitrosating species and preventing the endogenous formation of carcinogenic
nitroseamines (Stich & Rosin, 1984).

Antinutritional effect of Phenolic compounds

The antinutritional and toxic effects of phenolic compounds, particularly the tannins, have been
categorised as depression in food/feed intake, formation of the less digestible tannin-dietary protein
complexes, inhibition of digestive enzymes, increased excretion of endogenous protein,
malfunctions in digestive tract, and toxicity of absorbed tannin or its metabolites (Salunkhe et al.,
1990). Increased risks of cancer of the mouth and esophagus have been linked to dietary tannins in
some epidemiological studies (Kapadin et al., 1983).

Although the depressed food intake has been related in part to the astringent taste of the phenolic
compounds, the adverse effects of tannins have traditionally been attributed to their ability to bind
with proteins and hence inhibit their digestion and absorption (Butler, 1989).

Tannins decreased feed consumption in animals, bind dietary protein and digestive enzymes to
form complexes that are not readily digestible. They also cause decreased palatability and reduced
growth rate (Soetan and Oyewole, 2009).

2.4.3. Saponins

Saponins are a diverse group of compounds commonly found in legumes, e.g. chick peas, soya
beans, lentils, peanuts, lentils, phaseolus beans and alfalfa sprouts; and in some plants commonly
used as flavourings, herbs or spices, e.g. ginseng, fenugreek, sage, quillaja bark, thyme, sarsaparilla
and nutmeg (Oakenful & Sidhu, 1990). Their structures are characterised by the presence of a
steroid or triterpene group, referred to as the aglycone, linked to one or more sugar molecules. The
presence of both polar (sugar) and non-polar (steroid or triterpene) groups provide saponins with
strong surface-active properties which then are responsible for many of its adverse and beneficial
biological effects.

Fenugreek contains approximately 4 to 8% saponins and about 1% alkaloids, contributing to

bitterness, gastric stimulation, increased acidity, and increased appetite (Ambasta, 2000).

15
Uses of saponins

Of the phytochemicals, saponins have been studied the most extensively for the
hypocholesterolaemic effect. Decreases in cholesterol have been observed when rats were fed diets
containing fenugreek (Sharma, 1986).

The biological properties of saponins suggest that they may also have some anticarcinogenic effect
(Messina & Barnes, 1991) through a number of mechanisms. Saponins are able to bind cholesterol
and bile acids (Sidhu & Oakenful, 1986). The binding of the primary bile acids by saponins would
reduce their bacterial conversion to secondary bile acids and consequently reduce their ability to
promote tumourigenesis. Saponins have been shown to enhance the immune response and thus
have been used as adjuvants for oral vaccines (Heath et al., 1991).

Problems associated with saponin

A well-known toxic effect of saponin is its ability to lyse erythrocytes, this, in general, being due to
its interaction with the cholesterol in the erythrocyte membrane (Birk & Peri, 1980). If provided
intravenously to mammals, saponins can cause local inflammation, and in large doses, can result in
death due to massive release of erythrocyte debris and reduction in the oxygen-carrying capacity of
the blood (Scott et al., 1985). Saponins can also lyse other cells such as those found in the intestinal
mucosa and consequently affect nutrient absorption. Decreased weight gain has been observed with
high saponin intake due to a number of reasons including reduced food intake attributable to the
bitter taste of saponin (Birk & Petri, 1980), or decreased absorption and utilization of nutrients
caused either by (a) the inhibition of metabolic and digestive enzymes, e.g. protease, amylase,
lipase and cholinesterase inhibition by soya saponin (Cheeke, 1971), and chymotrypsin, protease
and succinoxidase inhibition by alfalfa saponins (Birk & Petri, 1980); or (b) binding with nutrients,
e.g. Zn binding by alfalfa saponins (West & Greger, 1978).

2.5. Processing methods to reduce phytochemicals

 Soaking

Long soaking (16 h) in water or bicarbonate solution was required to cause a considerable reduction
in the antinutrients of cowpeas. Also, the data indicated that pressure cooking was more effective
than ordinary cooking in decreasing phytic acid and trypsin inhibitor, while for tannins the results
were comparable. Moreover, cooking the presoaked seeds further reduced the studied antinutrients.
Germination also caused a significant reduction in the antinutrients and it was better than soaking
when these pretreatments (soaking or germination) were combined with cooking, i. e. cooking

16
germinated cowpeas was more effective in reducing the antinutrients than cooking soaked seeds.
Fermentation completely eliminated trypsin inhibitor, raffinose and stachyose from the seeds. It
also caused the highest reduction (47.36%) in phytic acid; however, more research is needed to
optimize fermentation conditions to modify cowpea composition without increasing tannins
(Ibrahim et al., 2001).

Soaking of seeds in plain water and mineral salt solution for 12 h decreased phytic acid to the
maximum 46–50% (Khokhar and Chauhan, 1997).

 Germination

El-Beltagy (1996) reported that the hemagglutinin activity of mung bean seeds decreased by about
84.4% after three days of germination. Mubarak (2004) also reported that tannins and phytic acid in
mung bean seeds were significantly (p < 0.05) reduced by germination and cooking processes.
Dehulling and soaking processes were less effective than germination and cooking processes in
reducing phytic acid and tannins. The higher reduction of phytate could have been due to the
phytase activity during germination. Sprouting for 60 h had the most pronounced saponin lowering
effect (46%) (Khokhar and Chauhan, 1997).

 Fermentation

Lactic acid fermentation (by Lactic acid bacteria, LAB) performs a number of essential roles
including the preservation and production of wholesome foods. They are generally inexpensive and
often little or no heat is required in their preparation thus making it fuel efficient. In human
nutrition such fermentations can be used as a means of partial and/or complete elimination of
antinutritional and flatulence producing factors in raw substances (Reddy & Pierson, 1994).

The fermentation processes (NF and CF) increased the protein digestibility to about 90% in all
samples. This result was consistent for dry bean with an initial protein digestibility value of 66%
and was increased to around 85% after the fermentation process. Dry bean fermentation resulted in
the reduction of phytates, TIA, saponins, tannins and raffinose oligosaccharides. Fermentation
offers unique nutritional advantages for making the protein of coarse-grained dry beans more
digestible by reducing its antinutrients. Natural fermentation had a better effect on the breakdown
of the oligosaccharides, raffinose and stachyose. However, CF significantly reduced the saponins,
phytate and tannin contents of the dry bean samples (Shimelis and Rakshit, 2008).

17
  Autoclaving

Effects of hydration, autoclaving, germination, cooking and their combinations, on the

reduction/elimination of antinutrients, flatus-producing compounds and the improvement of in vitro
protein digestibility of three selected phaseolus vulgaris varieties were investigated. Due to their
heat-sensitive nature, saponins, trypsin inhibitors and phytohaemagglutins, diminished drastically
to undetectable amounts when heating processes (cooking and autoclaving) were employed.
Hydration and germination processes were less effective in reducing trypsin inhibitors, saponins
and phytohaemagglutinins as compared with cooking/autoclaving processes. Germination process
reduced stachyose, raffinose, phytic acid and tannins which was due to metabolic activity. The
combination of germination followed by autoclaving processes yielded the most promising result in
this study (Shimelis and Rakshit, 2007).

18
 CHAPTER THREE

3. Materials and Methods

3.1. Experiment location, materials, chemicals and reagents

 Experiment Location

The experiment on the processing methods such as germination, autoclaving, fermentation and their
combinations, physico-chemical and functional properties were conducted in the food processing,
chemical reaction, size reduction and environmental engineering Laboratories of the Chemical
Engineering Department of Addis Ababa Institute of Technology (AAiT), Addis Ababa University.
The experiment on extraction process was done in the Laboratory of Ethiopian Spice Extraction
Factory. Laboratory analysis for proximate composition was conducted at Ethiopian Health and
Nutrition Research Institute (EHNRI) and JIJE Laboglass. Phytochemicals analysis was carried out
at EHNRI, Ethiopian spice extraction factory and Food Engineering Laboratories, AAiT.

 Sample collection, Transportation and Preparation

Three fenugreek varieties, namely, Challa, Ada’a and Wello were selected for experimental studies
in this thesis. The Challa variety was collected from Debrezeith Agricultural Research Center and,
both Ada’a and Wello varieties were purchased from a market (Ehil Berenda, Addis Ababa). All
varieties were transported, and cleaned manually by winnowing and hand sorting to remove husks,
damaged grains, stones, dust, light materials, glumes, stalks, undersized and immature seeds and
other extraneous materials then after the seeds were packed in airtight polythene plastic bags for
further processing.

Challa Ada’a Wello

Figure 3.1: Fenugreek seed varieties used in this thesis study

19
The cleaned seeds were divided into two sub-portions. The first sub-portion, not subjected to any
treatment, was used as a control and the other sub-portion was subjected to germination,
autoclaving, fermentation, extraction and their combinations. The raw and processed samples were
ground to pass a 0.71 mm aperture size of a laboratory test sieve (Endecotts LTD., London
England) to obtain fine powder and then packed in airtight polythene plastic bags for further
analysis of nutrients and phytochemicals composition, and physicochemical functional properties.

 Chemicals and Equipments

There were a number of equipments such as incubator, autoclave, centrifuge, percolator, tray drier,
digital balance, oven, mills, etc used during the experimental work. All the chemicals and reagents
used for analysis were analytical grade.

3.2. Processing methods and their combination

The processing methods used for the reduction of phytochemicals were germination, autoclaving,
fermentation, extraction and their combinations.

3.2.1. Germination

Each fenugreek variety seeds of 350g weight were washed three times using distilled water and
soaked in 99 % ethanol for one minute to prevent microbial growth during the germination period
(Huang et al., 2003). Then, the seeds were washed two times with distilled water to remove the
ethanol. The cleaned and washed fenugreek seeds were soaked in distilled water (seed to water ratio
of 1:8 w/v) for 16 h, 27°C in an incubator (Einrichtungen, Germany) in plastic containers. The
steeping water was drained off and the soaked fenugreek seeds were washed twice using distilled
water to protect the growth of microorganisms during germination. The soaked and washed seeds
were again divided into two sub-portions. The first sub-portion was germinated for 24 hours and
the second sub-portion was germinated for 48 hours. The soaked seeds were covered with wet clean
cloth and placed in plastic sieve. The content was left at 37°C in an incubator and watered 2-3 times
a day to enhance the germination process. Finally, the germinated seeds were dried in tray drier.

3.2.2. Autoclaving

Each fenugreek seed varieties of 350g weight were washed two times using distilled water and then
autoclaved at 15 psi pressure with a temperature of 121 oC in distilled water (1:8 w/v) for 15 min
according to the method of Vijayakumari et al., (1996). Then, the seeds were rinsed with distilled
water and dried in tray drier.

20
 3.2.3. Natural fermentation

Suspensions of fenugreek flour in distilled water were prepared in plastic containers at a

concentration of 1:18 w/v flour/ water, covered with aluminum foil and allowed to ferment
naturally with the endogenous microflora of the flour at 36°C incubator for 24, 48 & 72h. Aliquots
of the fermenting slurry for analysis were drawn at 0, 24, 48 and 72h under aseptic conditions. All
samples were analyzed for pH and titratable acidity. Slurry samples were dried in tray drier. The
dried sample was flaky and therefore reground mildly to homogenize the flour.

3.2.4. Extraction

Cleaned fenugreek seeds from each variety were coarsely ground to 1mm to 1.5mm size and then
50g of fenugreek flour were used for extraction at a time. 100%, 95%, 90%, 85%, 80% methanol
and ethanol was prepared and 500 ml of solvent was used for each washing stages of extraction.
The extraction was done eight times (3 times rinsing in the first and second stage, and single fresh
wash in the third and fourth stages) in a percolator with 1 mm sieve size. Percolation time was
recorded in each rinsing stages and the extracts were dried and reground mildly to homogenize the
flour, and the alcohol was recovered by distillation.

3.2.5. Combination of processes

Suspensions of fenugreek flours from 24h germination and autoclaving processes in distilled water
were prepared in plastic containers at a concentration of 1:18 w/v flour/ water, covered with
aluminum foil and allowed to ferment naturally with the endogenous microflora on the flour at
36°C incubator for 24h. All samples were analyzed for pH and titratable acidity. Fenugreek flour
samples from three days fermentation were extracted with polar solvents to study the effect of
combination of extraction and fermentation.

3.3. Analysis methods

Official standard methods of analysis of Association of Official Analytical Chemistry (AOAC,

2000) were used for proximate chemical analysis of fenugreek.

3.3.1. Proximate chemical composition

Proximate chemical composition analysis such as moisture content, total ash, crude protein
(N*6.25), crude fiber, crude fat, and minerals of the raw, processed fenugreek and products were
performed according to table 3.1.

21
Table 3.1: AOAC (2000) official methods used for proximate chemical composition analysis

No. Parameters Test Method

1 Ash AOAC Official Method 923.03
2 Crude fat AOAC Official Method 920.39
3 Crude protein AOAC Official Method 920.87
4 Crude fiber AOAC Official Method 962.09
5 Moisture content AOAC Official Method 925.10
6 Total calcium AOAC Official Method 923.03-Flame AAS
7 Total magnesium AOAC Official Method 923.03-Flame AAS
8 Total iron AOAC Official Method 923.03-Flame AAS
9 Total potassium AOAC Official Method 923.03-Flame Photometry
10 Total sodium AOAC Official Method 923.03-Flame Photometry
11 Total phosphorus Vanadomolybdophosphoric Acid Method- Colorimetric
 Determination of moisture content

A crucible was dried in an oven at 105°C for 5h and placed in a desiccator to cool. The weight of
the crucible (W1) was determined. Samples were weighted in this dry crucible (W2), dried at 105°C
for 24 hour and reweighted again (W3). All weights are measured in grams.

The moisture content was determined by the following relationship:

W2– W3
Moisture content (MC) = × 100 Eq. (3.1)
W2 − 𝑊𝑊1

 Moisture is reported as grams of water per hundred grams of wet sample.

 Determination of total ash

The apparatus used for total ash analysis were silica dish and muffle furnace. An empty silica dish
of capacity 45 mL was weighed and about 2 g samples added in it. This was put into a muffle
furnace where the temperature was raised to about 200oC for 1 to 2 h. Then, further increased the
temperature to 550°C and incinerated until a white or grayish ash is formed. The dish was placed in
a desiccator, cooled and weighed until constant weight reached.

The ash content was calculated on dry mass basis by the following relationship:

(W2– W)
% ash = × 100 Eq. (3.2)
(W1 − W)

Where: W: Weight in grams of the empty dish.

22
 W1: Weight in grams of the dish plus the dried test material.
W2: Weight in grams of the dish plus ash

 Determination of crude protein

Protein content was determined based on AOAC (2000) 920.87 of Kjeldahl method. About 2 g of
the test material was transferred to the Kjeldahl flasks. About 10 g of the catalyst was added and 25
mL or more, if necessary, of the concentrated sulphuric acid. The flask was placed in an inclined
position and heated below the boiling point of the acid until the frothing ceases. The heat increased
and boiled to digest until the mixture is clear. Then after the contents of the flask was cooled. A few
pieces of pumice stone were added to prevent bumping. About 120 mL of sodium hydroxide was
added to the side of the flask so that it forms a layer below the acid level.

The contents of the flask was mixed by shaking and distil until all the ammonia has passed over
into the boric acid solution to which 0.5mL of the indicator solution has been added. The solution
was titrated with the standard sulphuric acid solution until the color just changes from green to
pink.

14.007 × (T − B) × N × 100
% Nitrogen = Eq. (3.3)
W

Where:

T: volume in mL of the standard sulphuric acid solution used in the titration for the test.
B: volume in mL of the standard sulphuric acid solution used in the titration for the blank
determination
N: Normality of standard sulphuric acid.
W: Weight in grams of the test material.
14.007: Molecular weight of nitrogen
Percent nitrogen was multiplyed by factor 6.25 to calculate percent protein.
The calculated protein was on a total nitrogen basis.

 Determination of crude fat

Fat content was determined by AOAC (2000) 920.39 modified Mojonnier petroleum Ether
Extraction method. About 5 g to 10 g of the samples were weighed and extracted with 150 mL
petroleum ether for a minimum period of 8 hours in the Soxhlet extractor. The solvent is then

23
removed using a steam bath. The flask containing the extracted fat was dried for about one hour at
103 ± 2°C. Cooled in a desiccator’s and weighed.

 Crude fat was determined dry mass basis by the following relationship

(W2 – W1 ) × 100
Fat % = Eq. (3.4)
W

Where: W: Weight in grams of the dried test material.

W1: Weight in grams of the extraction flask.
W2: Weight in grams of the extraction flask plus the dried crude fat.

 Determination of Crude Fiber

About 2 g of the sample was weighed and the fat extracted for about 6h with petroleum ether using
a Soxhlet extractor. The fat-free dry residue (the residue obtained from crude fat determination)
was transferred to a conical flask and then 200 mL of 0.255N sulphuric acid pre-heated for one
minute was added to the flask containing the fat-free material. Immediately the flask was connected
to a reflux condenser and heated for one minute. The flask frequently was rotated to keep the
material from remaining on the sides of the flask out of contact with the acid. Boiling was
continued in a reflux condenser for 30 min. The reflux was filtered through a filter-paper and
washed with boiling water until the washing is no longer acid to litmus. The residue was boiled
with 0.313N sodium hydroxide solution under a reflux condenser for 30 min. The residue was
washed thoroughly with boiling water and transfer to a sintered glass crucible. Subsequently, the
residue was washed first with hot water and then with about 15 mL of ethanol. The sintered glass
crucible and contents was dried at 103 ± 2°C in the oven to constant weight. Finally the crucible
was cooled and weighed. The content of the sintered glass crucible was incinerated at 550°C in a
muffle furnace. Made to cool the sintered glass crucible containing the ash in a desiccator’s and
weighed.

 Crude fiber content of the flour was analyzed dry mass basis according to:

𝑊𝑊1 − 𝑊𝑊2
Crude fiber % = 𝐸𝐸𝐸𝐸. (3.5)
𝑊𝑊
Where: W= Weight in grams of the dried test material.
W1 = Weight in grams of the sintered glass crucible and contents before ashing.
W2 = Weight in grams of the sintered glass crucible after ashing.

24
  Determination of Total Carbohydrates

The total carbohydrates excluding the crude fiber was calculated on dry mass basis by difference.

𝐶𝐶 = 100 − (𝑀𝑀 + 𝐴𝐴 + 𝐹𝐹 + 𝑃𝑃) 𝐸𝐸𝐸𝐸. (3.6)

Where: M, A, F and P are the mass % of moisture, total ash, fat and total protein; respectively.
Soluble or utilizable carbohydrates shall be calculated after estimation of total carbohydrates using
the recommended method of Sumati and Rajagopal (1996).

 Energy value estimation

Energy value (calorific value) was quantified using an indirect calculation method. The three
groups of nutrients, which provide the body with energy, are carbohydrates, fats and proteins
(Gaman and Sherrington, 1986). One gram of carbohydrate (C) was assumed to give 15.71 KJ
energy; one gram of fat (F) 37.71 KJ energy and one gram of protein (P) 16.76 KJ. Therefore,
determination of calorific value (KJ/100 g) of fenugreek seeds was determined according to
Osborne and Voogt (1978). The energy values for one gram of the three groups of nutrients which
provide the body energy were calculated by using specific values of water factors for protein, fat
and total carbohydrate as recommended by Birch et al., (1980)

𝐸𝐸𝐸𝐸 = (𝑃𝑃 × 16.76) + (𝐹𝐹 × 37.71) + (𝐶𝐶 × 15.71) 𝐸𝐸𝐸𝐸. (3.7)

Where P: Protein content (%)

F: Fat content (%)
C: Total carbohydrates (%)
EV: Energy value (KJ)
The result was in KJ/ 100 g of the sample.

 Minerals Analysis

The mineral contents were determined by the procedure of AOAC (2000). Calcium, magnesium,
potassium, sodium, iron, and zinc were determined using an atomic absorption spectrophotometer
while phosphorous was determined by colorimetric method using vanadomolybdophosphoric acid
method. After removal of organic materials by dry ashing, the residue was dissolved in dilute acid.
The solution was sprayed into the flame of atomic absorption spectrophotometer and the absorption
of the metal to be analyzed was measured at a specific wavelength.

[(𝑎𝑎 − 𝑏𝑏) ∗ 𝑉𝑉]

𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐 (𝑚𝑚𝑚𝑚/100𝑔𝑔) = 𝐸𝐸𝐸𝐸. (3.8)
10𝑊𝑊

25
Where: W = weight (g) of samples;
V= volume (L) of extract;
a = concentration (μg/ml) of sample solution
b = Concentration (μg/ml) of blank solution

3.3.2. Physico-chemical properties of fenugreek

Unprocessed seeds of the given varieties were analyzed for seed weight, bulk density, volume,
hydration capacity, hydration index, swelling capacity and swelling index.

 Thousand seed mass

One thousand seed mass was determined by counting 1000 seeds manually and weighing. Results
were expressed as the mean of triplicate determinations.

 Density

After accurately weighing of hundred grams of fenugreek seeds and transferred to a measuring
cylinder, 100 ml of distilled water at room temperature (19oC) were added. Seed volume (ml/100 g
seeds) was obtained after subtracting 100 ml from the total volume (ml). Volume increase was
measured immediately, so that swelling character can not be a problem. The density of fenugreek
seeds was calculated and recorded as g/ ml (Shimelis and Rakshit 2005).

 Hydration and swelling coefficient

The hydration coefficient of raw fenugreek seeds soaked in distilled water for 18 h was calculated
as the percentage increase in mass of seeds. The volume of raw fenugreek seeds before and after
soaking in distilled water for 18 h was estimated by determination of displaced water. Swelling
coefficient was calculated as a percentage of the ratio of volume of fenugreek seeds after soaking to
before soaking (Shimelis and Rakshit 2005).

 Hydration, swelling capacity and indices

About 100 g of fenugreek seeds were counted and transferred to plastic cup and 500 ml distilled
water was added. The plastic cups covered with aluminum foil and left overnight at room
temperature. The next day all seeds were drained, superfluous water removed with filter paper and
swollen seeds reweighed. Hydration capacity expressed as hydration capacity per seed is
determined by dividing the mass gained by the number of seeds present in the sample.

Seeds mass of 100 g were counted, their volume noted and soaked overnight. The volume of the
soaked seeds was noted in a graduated cylinder (Shimelis and Rakshit 2005). Swelling capacity per

26
seed calculated as the volume gained by the seeds divided by the number of seeds. The swelling
index can be calculated as the ratio of swelling capacity per seed to volume of one seed (ml).
Hydration index then can be calculated using the formula (Bishnoi & Khetarpaul, 1993) as the ratio
of the average hydration capacity per seed and the mass of one seed.

 Water uptake

Fenugreek seed samples with known mass were soaked in distilled water at room temperature
overnight. The water left was removed from the soaking tank and the mass of soaked fenugreek
seed was measured. The water uptake was measured as percentage of soaked seeds per seeds
(Shimelis and Rakshit 2005).

𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚 𝑜𝑜𝑜𝑜 𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠 𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠 − 𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚 𝑜𝑜𝑜𝑜 𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠

𝑤𝑤𝑤𝑤𝑤𝑤𝑤𝑤𝑤𝑤 𝑢𝑢𝑢𝑢𝑢𝑢𝑢𝑢𝑢𝑢𝑢𝑢 (𝑊𝑊𝑊𝑊) = × 100 𝐸𝐸𝐸𝐸. (3.9)
𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚 𝑜𝑜𝑜𝑜 𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠

3.3.3. Functional properties of fenugreek flour

 Water absorption capacity

Since one gram of fenugreek flour absorbed more than 10ml distilled water, the Beuchat (1977)
procedure in which 1g flour was mixed with 10 ml of distilled water and centrifuged at 500 rpm for
20 min was modified to determine the water absorption capacity of fenugreek where about two
gram of fenugreek flour was mixed with 40 ml distilled water in a pre-weighed 50 ml centrifuge
tube. The slurry was agitated for 2 min, allowed to stand at room temperature for 30 min and then
centrifuged at 500 rpm for 20 min. The clear supernatant was decanted and discarded. The adhering
drops of water in the centrifuge tube were removed with cotton wool and the tube was weighed, the
weight of water absorbed by 1g of flour or protein was calculated and expressed as water
absorption capacity.

 Oil absorption capacity

The oil absorption capacity was determined according to Beuchat (1977) where one gram of
fenugreek flour was mixed with 10 ml refined palm oil in a pre-weighed 50 ml centrifuge tube. The
slurry was agitated for 2 min, allowed to stand at room temperature for 30 min and then centrifuged
at 500 rpm for 20 min. The clear supernatant was decanted and discarded. The adhering drops of oil
in the centrifuge tube were removed with cotton wool and the tube was weighed, the weight of oil
absorbed by 1 g of flour or protein was calculated and expressed as water or fat absorption capacity.

27
 3.3.4. Phytochemicals composition in fenugreek

 Phytic acid

Phytic acid was determined by using Vaintraub and Lapteva (1988). About 1g of sample was
extracted with 10 ml 2.4% HCl for 1h at an ambient temperature and centrifuged (3000 rpm/30min)
(Nüve, bench-top centrifuge, NF 800R, 2001, Ankara, Turk). The supernatant was used for phytate
estimation. About 2 ml of wade reagent (0.03% solution of FeCl3.6H2O containing 0.3%
sulfosalicylic acid in water) was added to 3 ml of the sample solution and centrifuged. The
absorbance at 500 nm was read using spectrophotometer (BECKMAN, DU-64, Japan).

The phytate concentration was calculated from the difference between the absorbance of the control
(3ml of water + 1ml of Wade reagent) and that of assayed sample. The concentration of phytate
was calculated using phytic acid standard curve (by preparing a series of standard solution
containing 0, 5, 10, 20, and 40 µg/ml μg/ml phytic acid in 0.2 N HCl) using water to zero the
spectrophotometer, and results were expressed as of phytic acids in mg/100gm dry weight by the
following formula.

𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎 – 𝐵𝐵𝐵𝐵𝐵𝐵𝐵𝐵𝐵𝐵 − 𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼 10

𝑃𝑃𝑃𝑃 = � �× 𝐸𝐸𝐸𝐸. (3.10)
𝑆𝑆𝑆𝑆𝑆𝑆𝑆𝑆𝑆𝑆 × 𝑆𝑆𝑆𝑆𝑆𝑆𝑆𝑆𝑆𝑆𝑆𝑆 𝑤𝑤𝑤𝑤𝑤𝑤𝑤𝑤ℎ𝑡𝑡 3

 Determination of phytate and non-phytate phosphorus

Phytate and phosphorous were determined by the above methods. Phytate phosphours was
calculated with the following formula (Duhan et al., 2002).

𝑃𝑃𝑃𝑃 × 28.18
𝑃𝑃ℎ𝑦𝑦𝑦𝑦𝑦𝑦𝑦𝑦𝑦𝑦 𝑝𝑝ℎ𝑜𝑜𝑜𝑜𝑜𝑜ℎ𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜 = 𝐸𝐸𝐸𝐸. (3.11)
100

Where: PA = phytate content (mg/100g), 28.18 = molecular weight of phytate

The result is expressed as mg/100g.

Non-phytate phosphorus was calculated as a difference between the total phosphorus and phytate
phosphorus.

 Tannins

Tannin was determined by the modified vanillin assay (Butter et al., 1982). About 200 mg
fenugreek flour samples were weighed and then extracted with 10 ml absolute methanol for 20 min
rotating screw cap culture tubes (130*100mm). The mixture was then centrifuged for 10 min at
3000*G and the supernatant were used in the analysis. About 0.0 – 1.0 ml aliquot of catechin

28
standard was dispended into two sets of culture tubes and each sample was brought to 1.0 ml by the
addition of absolute methanol. Incubate the tubes in water bath. 5 ml of the working vanillin
reagent was added at 1 min interval to one set of standards, and 5ml of the 4% HCl solution is
added at 1 min interval to the second set of standards. After 20 minutes, the absorbance of sample
solution and the standard solution were measured at 500 nm by using spectrophotometer. The
absorbance of the blank is subtracted from the absorbance of the corresponding vanillin- contain
sample. A standard curve has been constructed (Absorbance vs catechin) and the linear portion of
the curve will be extrapolated to produce the standard curve. Finally, the tannin contents were
calculated. Values of tannins were expressed in mg/100g sample as follow:

𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 – 𝐵𝐵𝐵𝐵𝐵𝐵𝐵𝐵𝐵𝐵 − 𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼

𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇 = � � × 10 𝐸𝐸𝐸𝐸. (3.12)
𝑆𝑆𝑆𝑆𝑆𝑆𝑆𝑆𝑆𝑆 × 𝑆𝑆𝑆𝑆𝑆𝑆𝑆𝑆𝑆𝑆𝑆𝑆 𝑤𝑤𝑤𝑤𝑤𝑤𝑤𝑤ℎ𝑡𝑡 × 0.791

 Saponin

The saponins concentration was determined by modifying the Okwu and Josiah (2006) procedure
where they extract 20 g of each sample by dispersing in 200 ml of 20% ethanol and the suspension
was heated over a hot water bath for 4 h with continuous stirring at about 55°C. The mixture was
filtered and the residue re-extracted with another 200 ml of 20% ethanol. However, in this study,
the extraction procedure was modified in which fenugreek seeds were coarsely ground to 1mm to
1.5mm size and then 20g of flour were extracted ten times (3 times rinsing in the first and second
stage, 2 times rinsing in the third stage and single fresh wash in the fourth and fifth stages) in a
percolator with 500 ml 80% methanol for each washing stages of extraction. The extract was
reduced to 40 ml over water bath at about 90°C. The concentrate was transferred into a 250 ml
separator funnel and 20 ml of diethyl ether was added and shaken vigorously. The aqueous layer
was recovered while the ether layer was discarded. The purification process was repeated. 60 ml of
n-butanol was added. The combined n-butanol extracts were washed twice with 10 ml of 5%
aqueous sodium chloride. The remaining solution was heated in a water bath. After evaporation, the
samples were dried in the oven to a constant weight. The results were expressed in mg/100g.

3.4. Development of value added products from fenugreek

 Debitterized fenugreek powder

The seeds of fenugreek from each variety were cleaned and coarsely ground to 1mm to 1.5mm size
and then 50g of fenugreek flour were used at a time for extraction. 90% ethanol was prepared and
500 ml of solvent was used for each washing stages of extraction. The extraction process was done

29
six times (3 times rinsing in both the first and second stages) in a percolator with 1 mm sieve size.
The extracted flours were dried, reground and packed in airtight polythene plastic bags.

 Fenugreek capsule

Fenugreek flours were weighed on a weighing balance and mixed with supplementary ingredients.
The ingredients were magnesium stearate as lubricant to prevent stickiness, gladiant to improve
flowability and starch as disintegrant. The flour blend was filled in to capsules with different sizes
by using manual capsule filling machine. The capsules were polished by passing through a polisher
to remove powders from the exterior part of the capsules and to give shinny color and then packed
in jars.

3.5. Sensory analysis

Fenugreek flours extracted by polar solvents (90% methanol and 90% ethanol) and fenugreek
capsules were evaluated for their taste and palatability by a panel of judges.

Debitterized fenugreek powder mix and fenugreek capsule were presented to 7 panelists. Just
before each test session, panelists were given orientation about the procedure of sensory evaluation.
The health status of the panelist was also considered during panelist selection (not suffering from
colds and allergies that affect their sensitivity for the product). Panelists were asked to rinse their
mouth with tap water that was provided to them, before the next serving.

The panelist were asked to rank the flours on the basis of visual color, flavor, texture, aroma and
overall acceptability using a nine point hedonic scale rated from 1(dislike extremely), 5 (neither
like nor dislike) to 9 (like extremely).

3.6. Experimental setup of the thesis

In this thesis, initially three varieties of fenugreek were studied for their proximate composition,
physico-chemical and functional properties, and phytochemical compositions. Two varieties were
then selected on the basis of phytochemical and nutrient composition for further processes such as
germination, autoclaving, fermentation, extraction and their combinations. The overall framework
of experiments of the thesis is summarized in figure 3.2.

30
 Three varieties of Fenugreek seeds

Physicochemical and Proximate chemical Phytochemicals

Functional Properties Composition analysis

 1000-Seed mass  Crude protein  Phytic acid

 Density, Volume  Crude fat  Tannins
 Hydration and swelling  Crude fiber  Saponins
coefficient  Minerals
 Hydration, swelling  Mineral bioavailability
capacity and indices  Moisture
 Water absorption  Total ash
capacity  Total Carbohydrates
 Oil absorption capacity

Two varieties of fenugreek

Processing methods and their combination

 Sensory
analysis
Extraction Germination Fermentation Autoclaving  Process
technology
Extraction development
rate study  Techno-
Proximate composition,
Product
mineral bioavailability & economic
phytochemicals Analysis Development
feasibility

Figure 3.2: Framework of experiments carried out in this thesis

31
 3.7. Experimental design and statistical data analysis

All the experiments were conducted in complete randomize design (CRD). Samples in triplicate
were taken from each of the three unprocessed fenugreek varieties. Accordingly, the Physico-
chemical and functional properties, proximate, mineral and phytochemical composition of seed
flours were recorded.

The data obtained from the chemical and functional determination were statistically analyzed by
using JMP software version 5.0. Comparisons between sample treatments and the indices were
done using analysis of variance (ANOVA) with a probability p< 0.05. “Student’s t” significant
difference test was used to identify significant differences among mean main effects for processing
methods. The analysis of physical and functional properties was performed in triplicate while
proximate and chemical measurements were performed in duplicate.

32
 CHAPTER FOUR

4. Results and Discussion

The influences of germination, fermentation, autoclaving, extraction, and their combinations on the
levels of certain phytochemicals (phytates, tannins and saponins), proximate and mineral
compositions of fenugreek seeds varieties were studied. The effects of solvent type, rinsing stage
and solvent to water proportion on the extraction rate of fenugreek flours were also investigated.

4.1. Proximate, mineral and phytochemicals composition of fenugreek seeds

The proximate chemical compositions (moisture, ash, crude protein, crude fat, crude fiber, total
carbohydrates and energy), minerals (Na, K, Ca, Mg, Fe and Zn) and phytochemicals (Tannins,
phytates and saponins) of whole fenugreek samples were presented on dry mater basis in tables 4.1,
4.2 and 4.3; respectively.

4.1.1. Proximate composition of fenugreek

The proximate compositions of three fenugreek varieties before any treatment are given in Table
4.1. Concentrations of moisture, protein, fat, ash, fiber, total carbohydrates, and energy value of
three varieties of fenugreek studied on dry mater basis are shown in table 4.1. For all varieties,
significant differences (P<0.05) were observed in the nutrient contents.

Moisture content of fenugreek varieties ranged from 6.88 % for Wello variety to 8.06 % for Ada’a
variety (Table 4.1). Crude protein was found to be 30.07, 29.21, and 28.65 % in Challa, Wello and
Ada’a; respectively. These values were within the range 21.7 % to 29.0% that were reported by
Nazar (2006) and similar to 30.0% obtained by Srinivasan (2006) for fenugreek seeds but higher
than 25.9% as reported by Hooda and Jood (2003a) and 24.7% by Ahmed (1982).

Oil content of the Ada’a variety was 8.84% which was significantly (P<0.05) higher than 7.55%
(Challa) and 7.57% (Wello) while the difference between Challa and Wello was not significant
(P>0.05). The oil compositions were greater than the range 5.8%, 6.9% and 7.2% that were
reported by Nazar (2006), EHNRI (1997), and Hooda and Jood (2003a); respectively. The results
for Challa and Wello were similar with the results reported by Srinivasan (2006) where he reported
7.5% of oil for fenugreek while Ada’a is still higher.

Crude fiber composition of 6.72 for Challa was found to be the highest followed by 7.22% for
wello and the least was 7.06% for Ada’a. The results for the three varieties were less than the 9.3%

33
that of reported by Nazar (2006) while Hooda and Jood (2003a) reported 6.88% crude fiber which
was similar to the Challa variety.

Table 4.1: Proximate composition of fenugreek varieties (g/100 g DMd)ã of whole seed samples

Variety % % Ash % Crude % Crude % Crude % Total Energy (KJ /

Moisture protein fat fiber Carbohydrates 100g)

Challa 7.70±0.10b 2.96±0.3a 30.07±0.07a 7.55±0.25b 6.72±0.03c 44.99±0.26b 1495.85±1.11c

Ada’a 8.06±0.01a 3.15±0.2a 28.65±0.63c 8.64±0.05a 7.06±0.05b 44.44±0.36b 1504.14±0.91a

Wello 6.88±0.02c 2.9±0.09a 29.21±.18b 7.57±0.27b 7.22±0.1a 46.22±0.02a 1501.14±0.58b

ã Values are means of triplicate determinations ± SD.
d DM, each values expressed on dry matter basis.
a-c Means with the same superscript letters within a column are not significantly different (p>0.05)

The seeds of fenugreek varieties contain 2.9 to 3.15% ash content. The difference in the ash
compositions between the varieties was not significant (P>0.05). The ash compositions are in the
range with 2.6 to 3.6% that of reported by EHNRI (1997), higher than 2.88% that was obtained by
Hooda and Jood (2003a), and less than 3.28% reported by Nazar (2006).

Total carbohydrates value of 46.22% for Wello was found to be the highest followed by 44.99% for
Challa and 44.44% for Ada’a was the least. The carbohydrates content of wello was significantly
(P<0.05) higher than the other two varieties. The results for carbohydrates contents were slightly
lower than the findings of Nazar (2006) where he reported 47.4% of carbohydrates and higher than
Hooda and Jood (2003a) in which they reported 43.7% for fenugreek seeds.

The energy values for Ada’a, Wello and Challa were 1504.14, 1501.14, and 1495.85KJ / 100g;
respectively in their descending order. For all varieties, significant differences (P<0.05) were
observed in the energy values.

4.1.2. Mineral composition of fenugreek

The mineral compositions of fenugreek seeds given in table 4.2 comprise the concentrations of
sodium, potassium, calcium, magnesium, iron and zinc on dry weight basis. For all varieties,
significant differences (P<0.05) were observed in the mineral contents. The data obtained showed
that Mg and K were the major mineral constituents of fenugreek seeds.

Sodium (Na) content varied from 102.07 mg/kg (Wello) to 154.17 mg/kg (Ada’a). The values for
sodium were lower than 350mg/kg that of reported by EHNRI (1997) and 190 mg/kg Srinivasan
(2006). As the result indicates Ada’a cultivar is superior in sodium content.

34
Table 4.2: Mineral composition of raw fenugreek varieties (on dry matter basis)

Variety Minerals (mg/kg)

Na K Ca Mg P Fe Zn

Challa 108.33±0.45b 9750.4±0.35c 502.9±0.32c 1237.8±0.16b 4879.0±1c 69.6±0.31c 39.47±0.31b

Ada’a 154.17±0.31a 10967.3±0.3a 532.5±0.15b 1443.8±0.46a 6750.0±2a 99.03±0.15a 52.45±0.21a

Wello 102.07±0.08c 10337.3±0.35b 625.6±0.2a 1090.2±0.32c 5325.0±1b 74.9±0.06b 38.3±0.2c

All values are means of triplicate ± standard deviation
a-c Means with the same superscript letters within a column are not significantly different (p>0.05)

Potassium ranged from 9750.4 mg/kg (Challa) to 10337 mg/kg (Wello) noticeably according to the
seed varieties. Wello has the highest (10337 mg/kg) potassium content followed by 10967 mg/kg of
Ada’a and the least 9750.4 mg/kg of Challa variety. The values obtained in this study are greater
than the result, 8805 mg/kg, reported by EHNRI (1997) and 5300 mg/kg by Ahmed (1982) while
lower than 16000 mg/kg that of expressed by Srinivasan (2006). Calcium content also varied
according to seed types from 502.9 to 625.6 mg/kg. The results are lower than 720.5 mg/kg that of
reported by Hooda and Jood (2003a), 1550.9 & 1730 mg/kg (EHNRI, 1997), and 1600 mg/kg
(Srinivasan, 2006).

Magnesium content of 1443.8 mg/kg for Ada’a was found to be the highest followed by 1237.8
mg/kg for Challa and the least 1090.2 mg/kg for Wello. The content of phosphorus in whole
fenugreek seed flours obtained in this study were higher than values recorded in a previous reported
by Abdel-Nabey and Damir (1990).

Zinc content of Ada’a was 52.45mg/kg, 39.47 mg/kg for Challa and 38.3 mg/kg for Wello. The
results for Challa and wello are similar with the value 38.0 mg/kg given by EHNRI (1997). Ada’a
was found to have the highest iron as well as zinc compositions among the fenugreek seed varieties
studied in this thesis. Zinc fights diarrhea and infections and promotes growth. It also promotes
immunity, resistance to infection, and growth and development of the nervous system. It also
enhances the production of antibodies against intestinal pathogens as described by Lazzerini and
Ronfani (2008).

Iron composition also varied conspicuously with the variety of seeds, Ada’a cultivar scored the
highest iron value of 99.03 mg/kg then Wello with a value of 74.9mg/kg and Challa was the least
by 69.6 mg/kg among the fenugreek seed varieties studied. The results were less than 141 mg/kg
for fenugreek reported by Ahmed (1982). As compared to the value 97.0 mg/kg obtained by

35
EHNRI (1997) Ada’a was higher where as Wello and Challa were lower. Iron is essential for
maternal and fetal health, learning, and productivity. It is an essential mineral for human
development and function. It helps to produce haemoglobin, the oxygen-carrying component of red
blood cells. As these cells carry oxygen to the muscles and brain, iron is critical for motor and
cognitive development in childhood, and for physical activity in all humans. If iron levels are too
low, the body makes too few red blood cells, and individuals develop anaemia. Iron is also critical
to the health of a pregnant mother and her unborn child. According to the report by UNICEF
(2009), a woman needs iron during pregnancy because the fetus and placenta both need additional
iron.

As the result of mineral compositions in this study indicates, fenugreek is rich in micronutrients,
calcium, iron and zinc. Therefore, fenugreek can be an important food ingredient to be used in food
fortifications enhancing micronutrient supplementations to alleviate the hidden hunger,
micronutrient deficiency, of the African children and the poor people as whole.

4.1.3. Phytochemicals composition of fenugreek

The major phytochemicals composition of fenugreek seed varieties are presented in table 4.3.
Saponins ranging from 4581.3 to 6320.8 mg/100g were found to be the highest phytochemical
content in fenugreek seeds. Tannins and phytates were varied from 199.86 to 269.13 mg/100g and
76.11 to 111.08 mg/100g; respectively. Significant differences (P<0.05) between varieties were
observed in the phytochemicals content of fenugreek seed samples before any processing treatment.
The differences in phytates, tannins and saponins contents for the three varieties could be due to
both genotypic and environmental conditions.

Tannin content of 269.13 mg/100g for Challa was the highest among the fenugreek varieties
studied followed by 235.24 mg/100g for Wello and the least was 199.86 mg/100g for Ada’a
cultivar (Table 4.3). The results are higher than that of reported by Hooda and Jood (2003a) where
they obtained 148.26 mg/100g of tannin content in a fenugreek variety grown in India. The tannin
compositions of the studied fenugreek varieties were lower than other legumes such as 390
mg/100g described by Vidal-Valverde et al. (1994) for Lentil, 537 to 1755 mg/100g for dry bean
reported by Shimelis and Rakshit (2007) and 1928 to 1085 mg/100g Barampama and Simard
(1993).

36
Table 4.3: Phytochemicals composition of raw fenugreek varieties

Variety Tannin Saponin Phytate

(mg/100g) (mg/100g) (mg/100g)

Challa 269.13±0.035 a 4581.3±0.8c 88.46±0.008 b

Ada’a 199.86±0.02c 6320.8±1a 111.08±0.24 a

Wello 235.24±0.11b 5008.1±0.5b 76.11±0.06 c

All values are means of triplicate ± standard deviation
a-c Means with the same superscript letters within a column are not significantly different (p>0.05)

As shown in table 4.3, saponin content of 6320.8 mg/100g for Ada’a was the highest among the
fenugreek varieties investigated followed by 5008.1 mg/100g for Wello and the least 4581.3
mg/100g for Challa variety (Table 4.6). The results are within the range of 4 to 8% reported by
Ambasta (2000).

The phytic acid content of fenugreek varied from 76.11 to 111.08 mg/100g. Ada’a has the highest
phytates content followed by Challa while Wello score the least. Similarly, as their tannin content,
the studied fenugreek varieties have lower phytates composition than other legumes such as 620 to
810 mg/100g for Lentil as reported by Vidal-Valverde et al. (1994), 2351 to 2406 mg/100g which
were reported by Shimelis and Rakshit (2007) and 1304 to 2111 mg/100g reported by Barampama
and Simard (1993) for dry bean.

Phytates potentially form complexes with minerals and dietary proteins and decreases their
bioavailability (Reddy, 2002). Similarly, The tannins cause decreased feed consumption in animals,
bind dietary protein and digestive enzymes to form complexes that are not readily digestible
(Soetan and Oyewole, 2009). The fenugreek varieties studied in this thesis, which are grown in
Ethiopia, are found to have lower phytates and tannins content as compared to the fenugreek
varieties grown in India and other legumes. Therefore, with respect to the antinutritional effect of
phytates and tannins the fenugreek varieties studied in this research can be advantageous.

4.2. Physicochemical properties of fenugreek

General characteristics and physicochemical properties such as 1000 seed mass, seed density,
hydration capacity, hydration index, swelling capacity, swelling index, hydration coefficient and
swelling coefficient of the three varieties of fenugreek seeds are presented in tables 4.4 and 4.5.
The moisture content of the fenugreek seed varieties varied significantly between 8.03% (Challa)

37
and 10.85% (Ada’a) with Challa having the least and Ada’a the highest followed by Wello(Table
4.4).

1000 seed mass of the three varieties of fenugreek seed varied from 14.43 to 23.62 g/1000 seed,
Challa was the highest and the lowest was Wello. Challa had a significantly (P<0.05) larger size
than Ada’a and Wello. The results were within the range (1.7 to 2.9 g/100 seed) reported by
Petropoulos (2002). Density of fenugreek seed ranged from 1.21 to 1.19 g/ml, the highest being in
Ada’a and the lowest being in Wello. Ada’a had a significantly (P<0.05) higher density followed by
Challa. Hydration capacity (g/seed) varied from 0.026 to 0.046 among the different fenugreek
varieties (Table 4.5). Challa had the maximum hydration whereas, Wello had the minimum
capacity.

Table 4.4: General characteristics of fenugreek seed varieties

Varieties Type % Moisture Seed mass* Seed density Aspect

(g/1000 seed) (g/m L)
Challa Food 8.03 23.62±0.47a 1.18±0.000c Elongated
Ada’a Food 10.85 17.36±0.66b 1.19±0.001b Elongated
Wello Food 9.38 14.43±0.17c 1.21±0.001a Elongated
*
mass of 1000 Fenugreek seeds
Means within same column followed by the same letters are not significantly different, P> 0.05%, by students’ test.
All values are the mean ± SD of four independent determinations.

Table 4.5: Physicochemical properties of Fenugreek seed varieties

Varieties Hydration Swelling Hydration Swelling Hydration Swelling

ã v
capacity capacity index index coefficient coefficient
(g/seed) (mL/seed)

Challa 0.046±0.002a 0.048±0.002a 1.946±0.005a 2.388±0.005a 2.946±0.001a 3.338±0.004a

Ada’a 0.029±0.001b 0.030±0.002b 1.676±0.002c 2.095±0.002c 2.676±0.000c 3.095±0.002c

Wello 0.026±0.002b 0.027±0.001b 1.686±0.000b 2.168±0.002b 2.686±0.002b 3.168±0.002b

All values are the mean ± SD of three independent determinations.
ã
Mean increases in mass of seeds due to water uptake over 16 h divided by the number of seeds.
v
Mean increases in volume of seeds due to water uptake over 16 h divided by the number of seeds.
Means within same column followed by the same letters are not significantly different, P> 0.05, by students’ test.

Challa variety scored the highest hydration index followed by Ada’a. Wello, having the lowest
hydration capacity also had the minimum hydration index. Swelling capacity did vary significantly
among the varieties in a similar manner of hydration capacity. But, swelling index and hydration

38
index had a different trend in which Challa had the highest swelling index followed by Wello and
Ada’a had the lowest swelling index. Challa attain the highest hydration capacity, hydration index,
swelling capacity, swelling index, hydration coefficient and swelling coefficient.

4.3. Functional properties of fenugreek

Water absorption capacity was significantly (P<0.05) different between 7.23 and 10.18g/g. Ada’a
had the highest water absorption capacity followed by Challa while Wello was the least (Table 4.6).
The water absorption capacities were higher than the results (1.89 and 2.15 ml/g) reported by
Appiah et al., (2011) for cowpea flour.

Table 4.6: Water and oil absorption capacity of flours of raw fenugreek varieties

Variety Water absorption Oil absorption

capacity* capacity **

Challa 7.464±1.29b 1.04±0.18b

Ada’a 10.18±1.55a 1.12±0.12b

Wello 7.23±0.45b 2.27±0.51a

All values are means of triplicate ± standard deviation
a-c
Means with the same superscript letters within a column are not significantly different (p>0.05)
*grams of water absorbed per 1 g of solid matter and
**grams of oil absorbed per g of solid matter.

This high water absorption capacity of fenugreek can attributed to the high composition of
galactomannans. The most important property of galactomannans is their high water-binding
capacity and the formation of very viscous solutions at relatively high dilutions. The galactose side
chains cause extension of the very long mannan chains and keep them from forming hydrogen
bonded insoluble associates; pure linear polymannan is completely water insoluble; (Neukom,
1989). The galactomannans are important viscosity builders (Glicksman, 1982). Galactomannans,
especially fenugreek gum, have high water solubility and do not form insoluble associates in water
(at relatively dilute solutions < 8 g/kg). The fiber portion of fenugreek consists of both insoluble
(30%) and soluble fraction (20%), which is mostly galactomannan as described by Srinivasan
(2006). Similarly, Srichamroen et al., (2005) also explained that the high ratio of galactose
substitution contributes to the hydrophilic properties associated with the galactomannans in
fenugreek seed. The adsorbed gum shows an emulsifying capability for stabilizing oil-in-water
emulsions. The galactomannans are mainly used when an efficient thickening or stabilizing effect is
needed (Garti et al., 1997).

39
On the other hand, the high protein content, as shown in table 4.1, may also contribute to the high
water affinity of fenugreek. According to Appiah et al., (2011), protein has both hydrophilic and
hydrophobic properties, and so can interact with water in foods. Carbohydrates have also been
reported to influence water absorption capacity of foods. The ability of protein to bind water is
indicative of its water absorption capacity. The observed variation in water absorption among the
fenugreek flours may be due to different protein concentration, their degree of interaction with
water and their conformational characteristics.

The oil absorption capacity of the flours of the fenugreek varieties ranged between 1.04 and
2.27g/g. wello had scored the highest oil absorption capacity while the difference between Challa
and Ada’a was not significant (P>0.05) (Table 4.6). The oil absorption capacities of the flours of
the three varieties studied were higher than the results reported by Nazar (2006) for fenugreek
protein concentrate and Appiah et al., (2011) for cowpea flour. The mechanism of fat/oil absorption
capacity was explained by Kinsella (1979) as a physical entrapment of oil. Fat/oil absorption
capacity is a critical determinant of flavour retention, while fat emulsion capacity and stability are
important attributes of additives for the stabilization of fat emulsions. Chau and Cheung (1997)
reported that surface area and hydrophobicity improve oil absorption capacity. Results obtained in
this study indicate that fenugreek had good oil absorption capacity.

Thus, the observed extremely high water absorption capacity and high oil absorption of the
fenugreek flours could be attributable to the presence of Galactomannans and proteins. The high
water absorption capacity of the flours suggests that they would be useful functional ingredients in
bakery products.

The ability of these fenugreek varieties to bind oil makes them useful in food systems where oil
imbibition is desired. The flours could, therefore, have functional uses in foods such as sauce and
sausage production. Thus, with this respect, Challa could be superior to Ada’a and Wello varieties.
The high oil absorption capacity also makes the flours suitable in facilitating enhancement in flavor
and mouth feel when used in food preparations. Wello could, therefore, be superior to Challa and
Ada’a as flavor retainer since it had significantly higher oil absorption capacity. Therefore, from the
result, I can suggest that thick, stable and smooth solutions can be obtained in high dilution which
makes the fenugreek very economical to use. In many food applications the fenugreek can be also
used in the presence of oils or fats as stabilizing agents.

40
 4.4. Effect of processing on the proximate and mineral compositions of
fenugreek

4.4.1. Effect of processing methods on the proximate compositions of fenugreek

Table 4.7 and 4.8 shows the proximate compositions (%moisture, %crude protein, %crude %fat,
%ash, %crude fiber, %total carbohydrates, and energy value) of the two fenugreek varieties (Challa
and Ada’a) before and after different processing methods such as germination, autoclaving,
fermentation, extraction and, combination of germination and autoclaving with fermentation.

Table 4.7: Effect of processing on proximate composition of Challa variety (g/100 g DMd) ã

Treatment % % Ash % Crude % Crude % Crude % Total Energy

code Moisture Protein Fat Fiber CH value
(kcal/100g)
RC 7.70±0.01 2.96±0.36 30.07±0.15 7.56±0.41 6.72±0.23 44.99 1495.85
GC1 5.94±0.05 2.34±0.15 30.50±0.36 10.31±0.25 8.45±0.51 42.46 1567.02
GC2 6.02±0.90 1.60±0.68 33.16±0.11 11.42±0.44 11.28±0.1 36.52 1560.14
FC1 7.50±0.60 2.92±0.34 30.96±0.56 8.36±0.14 8.97±0.21 41.29 1482.81
FC2 9.06±0.52 2.58±0.01 31.17±0.23 9.60±0.25 10.06±0.9 37.53 1474.02
FC3 6.74±0.41 1.93±0.80 35.62±0.85 11.33±0.01 14.15±0.3 30.23 1499.16
AuC 7.36±0.33 2.00±0.59 33.00±0.60 10.83±0.39 7.12±0.12 39.69 1585.01
EC 6.06±0.02 2.52±0.36 34.91±0.00 10.50±0.11 9.69±0.24 36.32 1551.63
CoGFC 7.40±0.23 1.78±0.00 33.90±0.17 11.77±0.13 9.77±0.78 35.38 1567.83
CoAuFC 6.90±0.15 1.99±0.71 33.80±0.19 11.31±0.00 7.68±0.03 38.32 1595.00
CH- Total carbohydrates
ã
Values are means of triplicate determinations ± SD.
d
DM, each values expressed on dry matter basis.
RC- Raw (whole); GC1- Germination for 24hr; GC2- Germination for 48hr; FC1- Fermentation for 1day; FC2- Fermentation for
2days; FC3- Fermentation for 3days; AuC- Autoclaving; EC- Extraction; CoGFC- Combination of germination and fermentation ;
CoAuFC- Combination of autoclaving and fermentation for Challa variety.

Crude protein

Results from table 4.7 and 4.8 showed that protein content increased during all processing methods.
The protein content of Challa has increased from 30.07% to 35.62% while Ada’a has increased
from 28.65% to 35.22%. The best improvement of protein was observed during three days
fermentation followed by extraction, combination of germination and fermentation, combination of
autoclaving and fermentation, two days germination and autoclaving respectively for Challa variety
while two days germination followed by three days fermentation, extraction, combination of
germination and fermentation, combination of autoclaving and fermentation respectively for Ada’a.

41
Among the treatments, three days fermentation, extraction, two days germination and combination
of processes have been found to be better than one day germination and autoclaving which have
slightly increased protein content of both varieties of fenugreek.

Even though the increment level of protein varied according to the processing methods and
varieties, the crude protein contents of processed flours were significantly (p<0.05) higher than that
of raw fenugreek flours. Similar observations were made by Shimelis (2009); (Abdel-Nabey and
Damir, 1990).

Table 4.8: Effect of processing on proximate composition of Ada’a cultivar (g/100 g DMd) ã

Treatment % % Ash % Crude % Crude % Crude % Energy

code Moisture Protein Fat Fiber Total value
CH (kcal/100g)
RA 8.06±0.02 3.15±0.03 28.65±0.22 8.64±0.99 7.07±0.99 44.43 1503.98
GA1 6.92±0.57 2.90±0.55 30.57±0.37 8.77±0.07 10.10±0.58 40.74 1483.10
GA2 7.36±0.00 2.59±0.19 35.22±0.06 11.51±0.16 8.26±0.92 35.06 1575.12
FA1 8.84±0.28 2.91±0.12 29.25±0.89 8.56±0.84 9.38±0.43 41.06 1458.08
FA2 8.66±0.19 2.15±0.00 29.12±0.70 9.08±0.22 9.02±0.17 39.97 1458.39
FA3 6.90±0.03 2.42±0.35 33.66±0.44 12.32±0.76 13.80±0.25 30.9 1514.17
AuA 7.16±0.07 2.59±0.17 30.59±0.13 9.98±0.53 8.19±0.00 41.49 1540.84
EA 7.42±0.27 3.13±0.81 33.53±0.85 9.27±0.44 9.99±0.32 36.66 1487.46
CoGFA 8.12±0.02 2.12±0.14 31.96±0.11 10.48±0.10 10.56±0.48 36.76 1508.35
CoAuFA 6.80±0.11 2.36±0.33 30.82±0.71 10.48±0.66 7.62±0.44 41.92 1570.31
CH- Total carbohydrates
ã
Values are means of triplicate determinations ± SD.
d
DM, each values expressed on dry matter basis.
RA- Raw (whole); GA1- Germination for 24hr; GA2- Germination for 48hr; FA1- Fermentation for 1day; FA2- Fermentation for
2days; FA3- Fermentation of for 3days; AuA- Autoclaving; EA- Extraction; CoGFA- Combination of germination and fermentation;
CoAuFA-Combination of autoclaving and fermentation for Ada’a .

The increase in protein content of fenugreek seeds after fermentation may be due to the decrease in
carbon ratio of the total mass, resulting in redistribution of nutrients percentage. Microorganisms
utilize carbohydrates as an energy source and produced carbon dioxide as a by-product. Such a
process caused the nitrogen in the fermented slurry to be concentrated and thus the protein in the
total mass increases as reported by Nuha et al. (2010). The observed increase in protein content of
fenugreek flour resulting from germination might be due to dry matter loss during germination as
evidenced by decrease in ash content (Table 4.7 and 4.8). It might be also attributed to a net
synthesis of enzymatic protein by germinating seeds (Shimelis, 2009). Fermentation also believed
to increase total protein content. This is due to the decrease in starch and sugar as a result of

42
hydrolysis by bacterial enzymes with the formation of volatile products such as lactic acid, acetic
acid, carbon dioxide, and ethanol. This leads to changes in proportions of nutrient components. It is
believed that the increase in protein content at the expense of starch is beneficial to consumers who
need a higher protein intake (Mugocha, 2001).

Crude fat

Results from table 4.7 and 4.8 showed that fat content increased during all processing methods. The
improvement of fat varied according to the processing methods and varieties. The fat content of
Challa has increased from 7.56% to 11.77% while Ada’a has increased from 8.64% to 12.32%. The
maximum increase of fat was observed during combination of germination and fermentation
followed by two days germination for Challa while three days fermentation followed by two days
germination for Ada’a variety. Among the treatments, three days fermentation, two days
germination and combination of processes have been found to better than one day germination and
fermentation, autoclaving and extraction which have slightly increase fat content of both varieties
of fenugreek. However, fat content was not affected by extraction process due to the fact that fat is
insoluble in polar solvents (methanol and ethanol) used for extraction of phytochemicals from
fenugreek seed flours. The increase in fat content of fenugreek seeds may be due to the decrease in
carbon ratio of the total mass since soluble carbohydrates removed during processing which could
concentrate the fat content and resulting in redistribution of nutrients percentage.

Regarding the effect of natural fermentation and germination on fat contents, the results of this
study were similar with the results obtained by Meseret (2011) in which crude fat content increased
during fermentation of quality protein maize-soybean blend flours. Similarly, Soetan and Oyewole
(2009) reported that fermentation caused increases in lipids for both soaked and sprouted seeds due
to increased activity of lipolytic enzymes which produced more free fatty acids. However,
contradictory results were reported by Hellen (2011) where she stated that both natural
fermentation and germination decreased fat contents of finger millet. Similarly, Hooda and Jood
(2003a) reported germination of fenugreek decreased crude fat content.

Crude fiber

The changes of crude fiber during processing are presented in table 4.7 and 4.8. The results indicate
that fiber content increased during all processing methods. Crude fiber content of Challa has
increased from 6.72% to 14.15% while Ada’a has increased from 7.07% to 13.80%. The highest
increase of fiber was observed during three days fermentation for both varieties. The result of this

43
study regarding the effect of natural fermentation and germination on fiber contents are
contradictory with the report of Hellen (2011) which states that both natural fermentation and
germination decreased fiber contents of finger millet while similar with Amankwah et al. (2009)
where they stated that crude fiber contents increased during fermentation of corn meal, and Hooda
and Jood (2003a) in which germination of fenugreek increase crude fiber content.

Total carbohydrates

Total carbohydrates composition was decreased during all treatments as presented in table 4.7 and
4.8. This may be due to the increase of other nutrient ratio of the total mass, resulting in
redistribution of nutrients percentages and breakdown of carbohydrates. Similar observations have
been made by different scholars. An enzyme a-galactosidase from germinated fenugreek seeds
partially attacks galactomannan to yield galactose. The decrease in the polysaccharide and mucilage
content may be attributed to their breakdown and utilization by the growing sprouts (Hooda and
Jood, 2003a). Among the treatments, three days fermentation caused a significant (p<0.05)
reduction of total carbohydrates in both varieties. The decreased in total carbohydrates during
fermentation is due to the fact that starch and soluble sugars are principal substances for fermenting
microorganisms; therefore degradation and a subsequent decrease in starch content are expected to
occur as reported by Mugocha (2001).

Ash

Processing methods decreased the ash content in both varieties. The proximate composition of the
varieties showed that the whole flour samples had the highest values regarding the ash parameters
examined. The results were consistent with the reports of Abdel-Nabey et al., 1990; Hooda and
Jood (2003a); Amankwah et al.,(2009) and hellen (2011). This may be due to leaching out of macro
minerals as the reduction of the macro minerals, K and Mg, indicated in table 4.7 and 4.8.

4.4.2. Effect of processing methods on mineral contents of fenugreek

Data on the changes of mineral contents accompanied by processing methods of fenugreek seeds
are shown in table 4.9, 4.10, 4.11 and 4.12. Fenugreek seeds were generally found to be good
source of minerals.

Despite of the slight increase of Mg during polar solvent extraction, a reduction in K and Mg
contents was observed as a result of all processing methods. This might be due to the leaching out
of minerals into water during the processes.

44
Table 4.9: Effect of processing methods on mineral compositions of Challa

Treatment Minerals (mg/kg)

Code Na K Ca Mg P Fe Zn
RC 108.3±0.4 9750.8±0.54 162.7±0.06 1237.8±0.95 4879±0.48 69.5±0.27 39.4±0.4
GC1 212.63±0.06 6113.12±0.7 187.17±0.86 1055.52±0.1 6720±0.89 104.56±0.5 58.42±0.77
GC2 223.45±0.3 4788.25±0.88 219.30±0.19 1048.10±0.06 4932±0.08 80.71±0.19 43.84±0.5
FC1 200.00±0.3 8108.11±0.07 173.03±0.5 1132.43±0.23 3977±0.46 89.89±0.06 60.92±0.28
FC2 203.43±0.14 9346.82±0.9 173.80±0.73 1149.11±0.8 5568±0.37 64.49±0.48 38.98±0.75
FC3 250.35±0.44 6969.76±0.12 204.86±0.02 1147.33±0.01 6182±0.52 69.75±0.7 39.78±0.06
AuC 210.49±0.35 6476.68±0.61 159.87±0.25 1090.24±0.08 6258±0.85 84.74±0.1 40.05±0.24
EC 124.2±0.21 7185.4±0.17 214.6±0.81 1245.5±0.66 5834±0.2 92.2±0.91 40.3±0.44
CoGFC 145.79±0.01 8069.11±0.03 174.78±0.57 1198.70±0.51 68790±0.9 109.0±20.35 55.21±0.9
CoAuFC 209.45±0.22 9083.78±0.64 166.11±0.21 1034.59±0.11 6205±0.3 159.5±0.29 46.72±0.16
All values are means of duplicate ± standard deviation
RC- Raw (whole); GC1- Germination for 24hr; GC2- Germination for 48hr; FC1- Fermentation for 1day; FC2- Fermentation for
2days; FC3- Fermentation for 3days; AuC- Autoclaving; EC- Extraction; CoGFC- Combination of germination and fermentation ;
CoAuFC- Combination of autoclaving and fermentation for Challa variety.

Table 4.10: Effect of processing methods on mineral compositions of Ada’a

Treatment Minerals (mg/kg)

Code Na K Ca Mg P Fe Zn
RA 154.1±0.23 10967.3±0.15 156.7±0.7 1443.9±0.04 6750±0.8 99.2±0.32 52.7±0.09
GA1 247.10±0.7 6714.65±0.6 177.37±0.45 1415.45±0.35 9122±0.44 108.26±0.54 58.01±0.63
GA2 253.67±0.3 5397.24±0.28 208.23±0.88 1087.54±0.68 7637±0.56 100.18±0.7 61.83±0.52
FA1 131.64±0.49 9872.75±0.71 173.98±0.34 1198.44±0.5 6993±0.33 111.87±0.91 58.89±0.49
FA2 138.37±0.1 5534.65±0.03 227.63±0.76 1137.37±0.07 7788±0.48 137.92±0.5 62.67±0.27
FA3 177.23±0.08 10204.08±0.85 250.97±0.25 1334.59±0.56 8857±0.18 105.34±0.57 64.56±0.42
AuA 188.50±0.01 7001.29±0.41 140.56±0.85 1330.25±0.25 9281±0.42 112.13±0.23 57.36±0.18
EA 221.4±0.8 8416.1±0.3 189.2±0.1 1466.3±0.34 7372±0.78 120.3±0.87 52.7±0.2
CoGFA 146.93±0.55 8162.82±0.85 206.68±0.97 1325.10±0.8 7576±0.89 212.67±0.31 58.55±0.58
CoAuFA 241.42±0.39 1070.82±0.11 155.20±0.43 1349.25±0.78 8379±0.2 166.95±0.77 66.67±0.6
All values are means of duplicate ± standard deviation
RA- Raw (whole); GA1- Germination for 24hr; GA2- Germination for 48hr; FA1- Fermentation for 1day; FA2- Fermentation for
2days; FA3- Fermentation of for 3days; AuA- Autoclaving; EA- Extraction; CoGFA- Combination of germination and fermentation;
CoAuFA-Combination of autoclaving and fermentation for Ada’a .

Similarly, Ca content also decreases during autoclaving (table 4.9 and 4.10). The loss of Ca could
be due to the leaching out of Ca with water during autoclaving process since boiling in water

45
involves a complex reaction system, where the Ca which is concentrated in parenchyma cells,
migrates into the cell content complexing with phytate ions (Abdel and Damir, 1990). More soluble
Ca salts might be formed during these changes and are leached out. However, the levels of
phosphorus, iron and Zinc significantly increase during germination, fermentation, autoclaving,
extraction and their combinations (table 4.9 and 4.10). The observed increased in mineral contents
of fenugreek flour might be due to losses of water-soluble constituents and reduction of phytic acid
during processing. Similar observations were reported by Tizazu et al., (2010) for germination of
sorghum, Abdel-Nabey and Damir (1990) for boiling of fenugreek, and Nuha et al., (2010) for
fermentation and cooking of Sicklepod leaves.

4.4.3. Effect of processing methods on phytate phosphorus and non-phytate

phosphorous contents of fenugreek

Data on phytic acid and other phosphorus compounds during processing of fenugreek seeds are
presented in table 4.11 and 4.12. The phytic acid content of dry fenugreek seeds were 88.46
mg/100g (Challa) and 111.08 mg/100g (Ada’a) values which represent 18.13% and 16.46% of the
total phosphorus contents (487.9 and 675 mg/100 g); respectively.

Table 4.11: Effect of processing methods on phytate phosphorous and non phytate phosphorus of
Challa

Phytate Phosphorus* non-Phytate phosphorus**

Treatment Phytate Phosphorus Total % of total Total % of total
(mg/100g) (mg/100g) (mg/100 g) phosphorous (mg/100 g) phosphorous

RC 88.46 487.9 24.93 5.11 462.97 94.89

GC1 44.89 672 12.65 1.88 659.35 98.12
GC2 29.75 493.2 8.38 1.70 484.82 98.30
FC1 22.23 397.7 6.26 1.58 391.44 98.42
FC2 14.27 556.8 4.02 0.72 552.78 99.28
FC3 0.00 618.2 0.00 0.00 618.20 100.00
AuC 59.64 625.8 16.81 2.69 608.99 97.31
EC 82.53 583.4 23.26 3.99 560.14 96.01
CoGFC 12.40 687.9 3.49 0.51 684.41 99.49
CoAuFC 52.15 620.5 14.70 2.37 605.80 97.63
*Phytate phosphorous content was calculated by assuming that phytate contains 28.18% phosphorus; **non-phytate
phosphorous was the difference between total phosphorous and phytate phosphorous.
RC- Raw (whole); GC1- Germination for 24hr; GC2- Germination for 48hr; FC1- Fermentation for 1day; FC2- Fermentation for
2days; FC3- Fermentation for 3days; AuC- Autoclaving; EC- Extraction; CoGFC- Combination of germination and fermentation ;
CoAuFC- Combination of autoclaving and fermentation for Challa variety.

46
All processing techniques resulted in loss of phytate and phytate phosphorus. However, the greatest
reduction was observed during fermentation and its combination with germination, and the lowest
reduction was recorded during polar solvent extraction process. This is attributed to the water
soluble nature of fenugreek phytic acid phosphorus as described by El-Shimi et al., (1989).

Table 4.12: Effect of processing methods on phytate phosphorous and non phytate phosphorus of
Ada'a

Phytate Phosphorus* non-Phytate Phosphorus**

Treatment Phytate Phosphorus Total % of total Total % of total
(mg/100g) (mg/100g) (mg/100 g) phosphorous (mg/100 g) phosphorous

RA 111.08 675.00 31.30 4.64 643.70 95.36

GA1 57.93 912.20 16.32 1.79 895.88 98.21
GA2 45.05 763.70 12.70 1.66 751.00 98.34
FA1 34.75 699.30 9.79 1.40 689.51 98.60
FA2 11.84 778.80 3.34 0.43 775.46 99.57
FA3 2.17 885.70 0.61 0.07 885.09 99.93
AuA 70.68 928.10 19.92 2.15 908.18 97.85
EA 103.32 737.20 29.12 3.95 708.08 96.05
CoGFA 19.83 757.60 5.59 0.74 752.01 99.26
CoAuFA 62.56 837.90 17.63 2.10 820.27 97.90
*Phytate phosphorous content was calculated by assuming that phytate contains 28.18% phosphorus; **non-phytate phosphorous
was the difference between total phosphorous and phytate phosphorous.
RA- Raw (whole); GA1- Germination for 24hr; GA2- Germination for 48hr; FA1- Fermentation for 1day; FA2- Fermentation for
2days; FA3- Fermentation of for 3days; AuA- Autoclaving; EA- Extraction; CoGFA- Combination of germination and fermentation;
CoAuFA-Combination of autoclaving and fermentation for Ada’a .

The results in table 4.11 and 4.12 indicated that even though whole fenugreek seed varieties studied
in this thesis have high (94.89 and 95.36 %) non-Phytate phosphorus as compared to other legumes
and cereals, contents of non-phytate phosphorous (NPP) increased during all treatments. As the
total phosphorous (TP) and non-phytate phosphorous (NPP) contents of processed fenugreek flours
increased significantly while phytate phosphorus (PP) content changed inversely proportional to the
non-phytate phosphorous. Data in this respect are in general agreement with those reported by
Tizazu et al. (2010); Abdel-Nabey et al. (1990); Nuha et al. (2010) and hellen (2011). Generally,
diets with phosphorus as phytate (%) ≤ 60% are regarded as being adequate in bioavailable
phosphate (Hellen, 2011).Thus, the effect of phytate on the phosphorus bioavailability is trivial for
all fenugreek samples.

47
 4.4.4. Effect of processing methods on mineral bioavailability of fenugreek

The results in tables 4.13 and 4.14 indicated that all processing methods decreased the phytate:iron
and phytate:zinc and phytate:calcium molar ratios of fenugreek flours. Phytate: zinc molar ratio is
used to estimate the likely absorption of zinc from a diet. Tizazu et al., (2010) elucidated that diet
with a phytate: zinc molar ratio greater than 15 have relatively low zinc bioavailability, those with
phytate: zinc molar ratios between 5 and 15 have medium zinc bioavailability and those with a
phytate: zinc molar ratio less than 5 have relatively good zinc bio-availability. Low values
(phytate:zinc molar ratio <5) were found in all whole and processed fenugreek flours, indicative of
favorable zinc bio-availability.

Table 4.13: Effect of processing on molar ratios of phytate: Fe, phytate: zinc, Phytate: Ca, and
[calcium*phytate]: [zinc] for Challa

Phytate Phytate/Fea Phytate/Znb Phytate/Cac [Phytate x Ca]/Znd

Treatment (mg/100g) (molar ratio) (molar ratio) (molar ratio) (mol/Kg)
RC 88.46 1.08 1.97 0.33 0.009
GC1 44.89 0.36 0.68 0.15 0.004
GC2 29.75 0.31 0.60 0.08 0.004
FC1 22.23 0.21 0.32 0.08 0.002
FC2 14.27 0.19 0.32 0.05 0.002
FC3 ND 0.00 0.00 0.00 0.000
AuC 59.64 0.60 1.31 0.23 0.006
EC 82.53 0.76 1.80 0.23 0.011
CoGFC 12.4 0.10 0.31 0.04 0.002
CoAuFC 52.15 0.28 0.98 0.19 0.005
a
mg of Phytate/MW of Phytate: mg of iron/MW of iron
b
mg of Phytate/MW of Phytate: mg of Zinc/MW of Zinc
c
mg of phytate/MW of phytate: mg of Calcium/MW of Calcium
d
(mol/kg Phytate) x (mol/kg Calcium)/(mol/kg Zinc)
ND- non detectable
RC- Raw (whole); GC1- Germination for 24hr; GC2- Germination for 48hr; FC1- Fermentation for 1day; FC2- Fermentation for
2days; FC3- Fermentation for 3days; AuC- Autoclaving; EC- Extraction; CoGFC- Combination of germination and fermentation ;
CoAuFC- Combination of autoclaving and fermentation for Challa variety.

The critical phytate:calcium molar ratio is 1:6. The phytate:calcium molar ratio <6 is indicative of
favorable for calcium absorption (Tizazu et al., 2010). All the fenugreek flour samples analyzed in
this study exhibited phytate:calcium molar ratios less than 6, which indicated that calcium is
available for absorption from such diets. High calcium levels in foods can promote the phytate-
induced decrease in zinc bioavailability when the [calcium]x[phytate]:[zinc] millimolar ratio
exceeds 0.5 (Tizazu et al., 2010). However, values observed in all fenugreek flour samples studied

48
indicated that the possible contributions of calcium in exacerbate the low bioavailability of zinc and
iron due to phytate is minimal since all treatments are <0.5 (Table 4.13 and 4.14) .

When the phytate: iron molar ratio >0.15, indicates low iron bioavailability (Tizazu et al., 2010).
Among the treatments samples, only three days fermentation and combination of germination and
fermentation resulted in phytate: iron molar ratios <0.15, indicative of high iron bioavailability.
However, processing of fenugreek seed resulted in a reduction of the phytate: iron molar ratios,
especially during germination and fermentation.

In general, phytate mineral ratio was decreased significantly after each processing methods (Table
4.13 and 4.14). The lower phytate: mineral ratios from the processed fenugreek seed varieties may
be partly attributed to the decreased in content of phytic acid during the treatments which has a
significant negative correlation (p< 0.05) with the phytate: mineral ratio (bioavailability of
minerals).

Table 4.14: Effect of processing on molar ratios of phytate: Fe, phytate: zinc, Phytate: Ca, and
[calcium*phytate]: [zinc] for Ada’a

Phytate Phytate/Fe Phytate/Zn Phytate/Ca [Phytate x Ca]/Zn

Treatment (mg/100g) (molar ratio) (molar ratio) (molar ratio) (mol/Kg)
RA 111.08 0.95 1.85 0.43 0.008
GA1 57.93 0.56 0.88 0.20 0.004
GA2 45.05 0.64 0.95 0.13 0.006
FA1 34.75 0.32 0.79 0.12 0.004
FA2 11.84 0.07 0.24 0.01 0.004
FA3 2.17 0.02 0.03 0.01 0.000
AuA 70.68 0.53 1.08 0.30 0.004
EA 103.32 0.73 1.72 0.33 0.009
CoGFA 19.83 0.08 0.30 0.06 0.002
CoAuFA 62.56 0.32 1.18 0.24 0.005
a
mg of Phytate/MW of Phytate: mg of iron/MW of iron
b
mg of Phytate/MW of Phytate: mg of Zinc/MW of Zinc
c
mg of phytate/MW of phytate: mg of Calcium/MW of Calcium
d
(mol/kg Phytate) x (mol/kg Calcium)/(mol/kg Zinc)
RA- Raw (whole); GA1- Germination for 24hr; GA2- Germination for 48hr; FA1- Fermentation for 1day; FA2- Fermentation for
2days; FA3- Fermentation of for 3days; AuA- Autoclaving; EA- Extraction; CoGFA- Combination of germination and fermentation;
CoAuFA-Combination of autoclaving and fermentation for Ada’a .

4.5. Effect of processing methods on phytochemicals composition of fenugreek

The influences of germination, fermentation, autoclaving, extraction and combinations of

fermentation with the other processing methods on the levels of certain phytochemicals (phytic

49
acid, tannins and saponins) present in the seeds of fenugreek were studied. As indicated in table
4.15 and 4.16, the level of phytochemicals concentration were reduced by applying different
processing methods such as conventional (germination and fermentation), heat (autoclaving) and
chemical (polar solvent extraction).

4.5.1. Effect of germination

The levels of phytic acid, tannins and saponins in the samples fenugreek seed flours have been
affected by germination as shown in tables 4.15 and 4.16. Germination had significantly reduced
the phytochemicals content of the two fenugreek seed varieties. Higher reduction in phytic acid,
tannins and saponins was obtained at the end of the second day of germination than the first day of
germination. During the second day germination period, phytic acid contents of Challa and Ada’a
varieties decreased from 88.4 mg/100g to 29.75 mg/100g and 111.08mg/100g to 45.05mg/100g,
respectively. These 66.37% and 59.44% of reduction in phytic acid content for Challa and Ada’a;
varieties; respectively are comparable with previous reports on the effects of germination on phytic
acid. A reduction of 57.85% for fenugreek varieties grown in India (Hooda and Jood, 2003a) and
20 – 75% for different bean varieties (Shimelis and Rakshit, 2007) were observed. Tizazu et al.,
(2010) reported that reduction of phytic acid during germination is due to an increase in the phytase
activity, which degrades phytic acid in plant based foods. Degradation of phytate improves the
bioavailability of iron, zinc and phosphorus so that, germination enhance the nutritional quality of
fenugreek.

It also appears that germination is useful in reducing tannins. In this study, tannins were decreased
from 269.13 mg/100g to 116.21 mg/100g and 199.86 mg/100g to 88.82 mg/100g for Challa and
Ada’a varieties; respectively, at the end of second day of germination period. The observed
reduction in tannin content was attributed to the formation of hydrophobic association of tannins
with seed proteins and enzymes. Some loss of tannins during germination may be due to the
leaching of tannins into the water (Shimelis and Rakshit, 2007). Similarly, saponins were affected
significantly during germination process as shown in tables 4.15 and 4.16. The percentage decrease
in saponins content was 62 and 69.9% for Challa and Ada’a varieties; respectively. This reduction
of saponins during germination is attributed to the solubility of saponins in water.

50
4.5.2. Effect of natural fermentation

Data on the effect of fermentation on certain phytochemical (phytic acid, tannins and saponins)
contents, for Challa and Ada’a varieties, indicated that as the period of fermentation increased, a
significant decrease in phytochemical contents was observed (Table 4.15 and 4.16).

The phytate contents of Challa and Ada’a were reduced by 74.87 to 100% and 68.72 to 98.05%;
respectively. Complete removal of phytate was observed for Challa variety during three days of
fermentation period. Phytates reduction during fermentation has been reported in various foods
including beans (Shimelis1&Rakshit, 2008), sorghum (Osman, 2004) and finger millet (hellen,
2011). Phytate hydrolysis can occur during food preparation and production, either by phytase from
plants, yeasts, or other microorganisms. Biological processing techniques, which increase the
activity of native enzymes of cereals and legumes, are soaking, malting, hydrothermal processing,
and lactic fermentation. Lactic fermentation leads to lowering of pH as a consequence of bacterial
production of organic acids, mainly lactic acid, which is favorable for phytase activity (Reddy,
2002). Shimelis and Rakshit (2008) also explained that naturally occurring phytase is activated at
low pH and the hydrolysis of higher inositol phosphates to phosphate can lower phosphoric esters
of myo-inositol and in some cases can also free myoinositol. The reduction in phytic acid during
fermentation was attributed to the action of the microbial enzyme phytase. This reduction in phytic
acid may be useful in improving nutritional quality of fenugreek with respect to mineral
bioavailability.

Table 4.15 and 4.16 also shows that tannin contents significantly (p<0.05) decreased during
fermentation in the two fenugreek seed varieties. Reduction of 40.68 to 61.79% and 45.29 to
87.89% in tannins content of Challa and Ada’a varieties were obtained during the fermentation
processes. Similar observations on decreased tannins have been reported by other investigators.
Osman (2004) reported reduction of 52.7 to 92% in tannin contents during fermentation of sorghum
varieties. Shimelis &Rakshit (2008) also found that natural fermentation and control fermentation
of bean varieties decreased their tannins content by 26 to 47%. The reduction may be due to
amylolytic enzymes and microbial enzyme hydrolysis (Ogunsua, 1980; Osuntokun, 1981). Tannins
are known to be responsible for decreased feed intake, growth rate, feed efficiency and protein
digestibility in experimental animals. Therefore, reduction of tannin content of the fenugreek
varieties through fermentation would improve the nutritional value Osman (2004).

Natural fermentation also significantly (P < 0.05) reduced the saponins content of fenugreek flour
of the two varieties (Table 4.15 and 4.16). 67.8 to 79.13% and 73.96 to 81% decrease of saponins

51
for Challa and Ada’a were observed; respectively. The results in this study are comparable with
previous researchers. The decrease in saponin content of foods caused by fermentation has been
well documented for a variety of foods such as legumes and quinoa (Anderson and Wolf, 1995;
Zhou and Erdman, 1997; Ridout et al., 1991; Shimelis1&Rakshit, 2008).

4.5.3. Effect of autoclaving

During the autoclaving process, levels of phytic acid, tannins and saponins were significantly (P <
0.05) reduced (Table 4.15 and 4.16). However, autoclaving is found to be effective for reduction of
saponins in which 88 and 94.75% of reduction was observed for Challa and Ada’a varieties;
respectively. This is due to the heat-sensitive property of saponins. Degradation of saponins after
temperature of 100 0C was reported by Güçlü et al., (2007).

Table 4.15: Effect of processing methods on phytochemical compositions of Challa variety

Treatment Phytates Tannins Saponins

code
(mg/100g) Reduction (mg/100g) Reduction a (mg/100g) Reduction a
a
(d.m.) (%) (d.m.) (%) (d.m.) (%)
RC 88.4 269.13 4581.3
GC1 44.89 49.25 173.54 35.52 2458.33 46.34
GC2 29.75 66.37 116.21 56.82 1740.89 62.0
FC1 22.23 74.87 159.65 40.68 1475.18 67.8
FC2 14.27 83.87 126.11 53.14 1236.95 73.0
FC3 ND 100 102.83 61.79 956.12 79.13
AuC 59.64 32.58 90.32 66.44 524.10 88.56
EC 82.53 6.7 ND 100 ND 100
CoGFC ND 100 12.78 95.25 342.68 92.52
CoAuFC 52.15 41.05 52.43 80.52 ND 99.20
CoE&FC ND 100 ND 100 ND 100
ã
Reduction indicates % decrease over raw value
ND- non detectable
d.m., each values expressed on dry matter basis.
a-c
Means with the same superscript letters within a column are not significantly different (p>0.05)
RC- Raw (whole); GC1- Germination for 24hr; GC2- Germination for 48hr; FC1- Fermentation for 1day; FC2- Fermentation for
2days; FC3- Fermentation for 3days; AuC- Autoclaving; EC- Extraction; CoGFC- Combination of germination and fermentation ;
CoAuFC- Combination of autoclaving and fermentation; CoE&FC- Combination of extraction and fermentation for Challa variety

Autoclaving significantly (P < 0.05) lowered tannins and phytic acid concentration in fenugreek
seeds. Tannins content was reduced by 66.44 and 74.72% for Challa and Ada’a varieties;
respectively. The leaching of tannins into the water may cause the decrease of tannins

52
concentrations. The results of this study were consistent with those mentioned by previous
investigators that autoclaving process reduced tannins by 50–72% (Shimelis and Rakshit, 2007). As
compared to saponins and tannins, phytate lowered to a limited extent to 32.58 and 36.37%
reduction for Challa and Ada’a varieties; respectively. The reduction of phytic acid is less
significant than germination and fermentation processing methods. These could be during food
processing, such as soaking; germination, and fermentation, phytates are hydrolyzed because of the
activation of intrinsic plant phytases, extrinsic microbial phytases, or both. However, thermal
processing can lead to a partial non enzymatic hydrolysis of phytates (Sandberg et al., 1999).
Because phytate is heat stable, significant heat destruction of phytate is not expected to occur.
Prolonged times at elevated temperatures were lead to a progressive inactivation of the endogenous
enzymes (Greiner and Konietzny, 2006). Thus, it was observed that autoclaving was more
effective for the heat sensitive phytochemicals (saponins) than the heat stable (tannins and
phytates).

Table 4.16: Effect of processing methods on phytochemical compositions of Ada’a cultivar

Treatment Phytate Tannin Saponin

code
(mg/100g) Reduction a (mg/100g) Reduction a (mg/100g) Reduction a
(d.m.) (%) (d.m.) (%) (d.m.) (%)
RA 111.08 199.86 6320.8
GA1 57.93 47.85 132.51 33.7 2781.15 56
GA2 45.05 59.44 88.82 55.56 1902.56 69.9
FA1 34.75 68.72 109.34 45.29 1645.94 73.96
FA2 11.84 89.34 95.93 52 1364.66 78.41
FA3 2.17 98.05 24.20 87.89 1200.95 81.0
AuA 70.68 36.37 50.52 74.72 331.84 94.75
EA 103.32 6.99 ND 100 ND 100
CoGFA ND 100 12.37 93.81 189.62 97.0
CoAuFA 62.56 43.68 25.16 87.41 ND 100
CoE&FA ND 100 ND 100 ND 100
ã
Reduction indicates % decrease over raw value
ND- non detectable
d.m.- each values expressed on dry matter basis.
RA- Raw (whole); GA1- Germination for 24hr; GA2- Germination for 48hr; FA1- Fermentation for 1day; FA2- Fermentation for
2days; FA3- Fermentation of for 3days; AuA- Autoclaving; EA- Extraction; CoGFA- Combination of germination and fermentation;
CoAuFA-Combination of autoclaving and fermentation; CoE&FA- Combination of extraction and fermentation for Ada’a.

53
 4.5.4. Effect of extraction

As an alternative food processing method, polar solvent extraction by using low density alcohols
(methanol and ethanol) and water was attempted primarily to eliminate the bitterness of fenugreek
flour. As indicated in table 4.15 and 4.16, extraction of fenugreek flours with 90% alcohol leads to
elimination of tannins and saponins for both Challa and Ada’a varieties despite their genetic
difference. However, extraction slightly reduced phytates by only about 6% which was the least
reduction among the processing methods carried out in this study. This indicates that phytate is
insoluble in alcohol and the slight decrease of phytate content observed is due to the 10% water
used during the preparation of 90% alcohol solution.

The elimination of saponins and tannins during extraction is due to the fact that saponins and
tannins are soluble in polar solvents. Haslam (1966) mentioned that tannins readily dissolve in
water and alcohol to form colloidal solutions. Hostettmann and Marston (1995) reported that
solubility of saponins is also affected by the properties of the solvent. While water, alcohols
(methanol, ethanol) and aqueous alcohols are the most common extraction solvents for saponins,
solubility of some saponins in ether, chloroform, benzene, ethyl acetate, or glacial acetic acid has
also been reported. Water, lower alcohols (methanol and ethanol), or water: alcohol mixtures have
been widely used for extraction of saponins from plant matrices (Kitagawa, 1986; Bombardelli and
Gabetta, 2001).

4.5.5. Effect of combination of processes

Combination of food processing methods can reduce the level of phytochemicals in significant
(p<0.05) amount as compared to single processing methods. It was observed that fermentation leads
to a reduction in phytates. On the other side autoclaving and extraction led to a reduction in tannins
and saponins. Hence, combinations of autoclaving, germination, and extraction with fermentation
were attempted.

As the results indicated in tables 10.15 and 10.16, combination of fermentation and extraction
brought a complete removal of phytates, tannins and saponins. The combination of germination and
fermentation also resulted in a significant and successive reduction in the phytic acid, tannins and
saponins level than germination and fermentation. Similarly, combination of autoclaving and
fermentation was effective in reducing the concentrations of tannins and saponins than the
individual processes. However, combination of autoclaving and fermentation showed a reduction of
phytates less than the individual processes, autoclaving and fermentation. This might be due to

54
inactivation of the natural phytase enzymes found in the fenugreek seeds during autoclaving
process.

4.6. Effect of rinsing stage, solvent type and water to solvent proportion on
extraction rate of fenugreek flour

Extraction is an important separation process that is extensively used in various applications in food
engineering. Extraction is used either to recover important food components, being a main
processing stage for the production of certain food products (sugar, oils, proteins), or to isolate
desired components (antioxidants, flavors). It also removes contaminants and other undesirable
components from food sources (Coustantina and George, 2003). In this study, extraction process
(solid-liquid) was applied to reduce the contents of phytochemicals in fenugreek seeds.

As the results indicated in table 4.15 and 4.16, polar solvent extraction led to elimination of tannins
and saponins. However, the time taken to accomplish the extraction process is crucial for extraction
industries. The rate of extraction is affected by the property of solvent, type of extracted material,
particle size and quantity of material to be extracted, pH, length of extraction path, and temperature.
Therefore, the effect of solvent type, solvent to water ratio, genetic variety and washing stages on
the rate of extraction were studied and the results are presented in table 4.17.

Variation of extraction rates with variety, solvent type, and solvent to water proportion were
observed. Percolation time varied from 9 to 111 minutes during the extraction of fenugreek flour
samples (Table 4.17). Extraction of Challa variety with 100% methanol scored the least (9 minutes)
percolation time while extraction of Ada’a variety with 80% ethanol scored the longer time (111
minutes). Regarding to solvent type, methanol extraction was faster than ethanol because ethanol is
denser and higher solubility with water than methanol. As viscosity (density) increases mass
transfer decreases so that percolation (extraction) time increases. And concerning the solvent to
water ratio (proportion), as the proportion of water increased the percolation (extraction) rate was
also decreases. This might be due to:-

 The higher water binding property of fenugreek as explained earlier while discussing the
results obtained in this study (Table 4.6) for functional properties of fenugreek. Glicksman
(1982) also explained that galactomannans, especially fenugreek gum, have high water
solubility and galactomannans are important viscosity builders.
 The higher solubility of carbohydrates (gum, mucilage) in water. Gums and mucilages keep
(connect) the structure of outer and inner components (endosperm component) of the food

55
 material. Loss of structure and porosity of a material will occur when the gums and
musilages are extracted by water. Consequently, components of the food material become
fine (closed to each other) and impaired the movement of solvent which in turn decrease the
extraction rate.

Table 4.17: Effect of rinsing stage and alcohol to water ratio on percolation rate

Percolation time (min.)

Solvent to water Washing Rinsing Ethanol Methanol
ratio stage stage Challa Ada’a Challa Ada’a
First First 13.5 17 9 10.8
Second 15 20.3 10.5 13
Third 17 23.9 12 14.7
100 % alcohol Second First 20 28 11 17
Second 22 29.5 12 18.2
Third 24.8 33 13.5 19
Third 27 35 13 21.2
Fourth 29 36.7 12.5 22
First First 23 27.5 16 19.3
Second 26.5 29.6 18 21
Third 30.2 34 21 23.5
95 % alcohol Second First 32 38 18 26
/5% water Second 33.8 40 20 27
Third 36 44.5 23 30.5
Third 39 56 23 33
Fourth 40.7 58 24.5 34.2
First First 35 41.8 24 28
Second 39 46.2 26.7 31
Third 43 48 29 32.5
90 % alcohol Second First 48 54 31.5 34
/10% water Second 50.2 56.7 34 35.8
Third 52 59.2 35.8 37
Third 54 63 37 39
Fourth 59 66 38.2 40.7
First First 51.5 64.8 31 38
Second 55 67.5 35 41
Third 57.3 70.1 36.5 42.5
85 % alcohol Second First 60 72.6 40.5 44
/15% water Second 62.5 75 41 45.3
Third 65 78.2 41 47
Third 66.7 80 45 49
Fourth 68 81 47 49
First First 74 87 43 54
Second 78 90.5 45 57.5
Third 81.2 93 46.5 60.2
80 % alcohol Second First 85 96.5 49 65

56
 /20% water Second 87.7 90 51.5 66.5
Third 92 105 53 69
Third 96 110 58.5 77
Fourth 96 111 61 79.4

Extraction rate also showed inversely relationship with rinsing stage (washing step). As the
washing (rinsing) step increased the percolation rate decreased. This might be due to the loss of
structure and porosity of the fenugreek flour accompanied by solubility of carbohydrates,
musilages, and other materials that were maintained the structure or porosity through which the
solvent can easily passed.

Extraction rate of Ada’a variety was slower than Challa variety. This may be due to the higher
water absorption capacity of Ada’a cultivar (Table 4.6).

Even though extraction of saponins with methanol was higher and faster than ethanol, ethanol
extraction is recommended due to its cheaper price and toxicity of methanol. It was also observed
that as the water proportion increases solvent power to extract saponins increases and rate of
extraction decreases. Therefore, extraction with 90% ethanol (10% water) would be reasonable
(with respect to cost, safety, time, and efficiency) for extraction of fenugreek flour.

The byproduct (water + phytochemicals) from the extraction process was assessed for its
application in biopest management by applying on to roaches and insects. And it kills all of the trial
animals. Therefore, the byproduct, containing water and phytochemicals, can be used for pest
management. Similarly, the toxicity of saponins to insects (insecticidal activity), parasite worms
(anthelmintic activity), molluscs (molluscicidal), and fish (piscidal activity) and their antifungal,
antiviral, and antibacterial activity were documented by Güçlü-Üstündağ et al., (2007).

4.7. Comparison of effect of various processing methods on phytochemicals

The effect of germination, fermentation, autoclaving, extraction and their combinations on the
phytochemical contents of fenugreek seed varieties have been investigated in this study and
discussed in section 4.5. In this section a brief comparison of the processing methods based on their
effect on the reduction of phytochemicals level are made.

As Figures 4.1 to 4.3 shows, natural fermentation resulted in removal of phytic acid and increased
reduction in tannins and saponins because of the metabolic activities involving enzymes.
Extraction led to elimination of tannins and saponins. However, complete removal of all

57
phytochemicals (phytic acid, tannins and saponins) investigated in this thesis was obtained during
the combination process of fermentation followed by extraction.

% Reduction of phytochemicals
120
Processing methods
1. Raw
2. Germination (24h)
100 3. Germination (48h)
4. Fermentation (24h)
80 5. Fermentation (48h)
Percent (%)

6. Fermentation (72h)
7. Autoclaving
60 Phytates
8. Extraction
Tannins 9. Combination of
40 Saponins Germination&
fermentation
20 10. Combination of
Autoclaving&
fermentation
0
11. Combination of
1 2 3 4 5 6 7 8 9 10 11 Fermentation &
Processing methods extraction

Figure 4.1: Comparison of effect of processing methods on phytochemicals of Challa variety

% Reduction of phytochemicals
Processing methods
120
1. Raw
2. Germination (24h)
100 3. Germination (48h)
4. Fermentation (24h)
80
5. Fermentation (48h)
Percent (%)

6. Fermentation (72h)
7. Autoclaving
60 Phytates 8. Extraction
9. Combination of
Tannins
40 Germination&
Saponins fermentation
10. Combination of
20 Autoclaving&
fermentation
0 11. Combination of
1 2 3 4 5 6 7 8 9 10 11 Fermentation &
extraction
Processing methods

Figure 4.2: Comparison of effect of processing methods on phytochemicals of Ada’a variety

58
4.8. Sensory evaluation of value added products

The result obtained from panelists shows that fenugreek capsules were rated significantly (p<0.05)
higher in terms of overall acceptability and color compared to other samples. The overall
acceptability of raw Ada’a variety was higher than Challa variety (table 4.18).

According to the taste of the panelists, the aroma and taste of the ethanol extracted fenugreek flour
was preferable than that of methanol extracted flour. As compared to the control (raw), the odor and
taste of the extracted flours were preferred by the panelists. This is due to the fact that whole
fenugreek flour has bitter taste and unpleasant flavor while the extraction processes remove the
bitterness and odor from the raw fenugreek flours. However, the color of raw fenugreek flour
samples was preferred by the panelists than the extracted flours.

Concerning the therapeutic value of whole fenugreek seed, capsule filling of raw flour could be
advantageous. In addition, the amount or dose of fenugreek which is required for diabetic patients,
lactating mothers and infants can be managed by filling the required fenugreek in to the capsules.

Table 4.18: Sensory evaluation of extracted fenugreek flours and capsules

Sample (Challa variety) Color Aroma Taste Over all

acceptability
Unprocessed(control) 8.3±0.67a 4.5±0.10 a 3.9±0.32 a 5.0±0.21 a
Extracted flour (90% methanol) 6.7±0.24b 8.2±0.00 b 9.0±0.14 b 7.0±0.53 b
Extracted flour (90% ethanol) 7.0±0.63c 8.8±0.28 c 9.5±0.57 c 7.2±0.78 c
Capsule 9.8±0.65d - - 10.0±0.00d

Sample (Ada’a variety)

Unprocessed(control) 9.5±0.11 a 4.7±0.17 a 3.3±0.05 a 6.0±0.40 a
Extracted flour (90% methanol) 7.6±0.23 b 8.6±0.85 b 8.9±0.60 b 7.6±0.01 b
Extracted flour (90% ethanol) 8.1±0.20 c 8.9±0.34 c 9.4±0.47 c 7.7±0.19 c
Capsule 9.8±0.35 d - - 10.0±0.00 d
All values are means of triplicate ± standard deviation
a-d Means with the same superscript letters within a column are not significantly different (p>0.05)

59
 CHAPTER FIVE

5. Techno-economic Feasibility for Fenugreek products

The primary objective of this thesis was to study the influence of processing methods
(conventional, technological and chemical methods) on the reduction of phytochemical contents of
fenugreek seed varieties; however, developing of value added products to commercialize fenugreek
was also part of the objective of this study. Therefore, productions of debitterized fenugreek
powder and fenugreek capsule are selected for detail process technology development, techno-
economic feasibility and financial analysis.

5.1. Process description on production of debitterized fenugreek powder

Fenugreek
Cleaning Raw material drying Grinding
seed

Solvent
Water tank preparation
Vessel Extraction
Extract + solvent
Alcohol
storage vessel
Extract
+ Distillation Extracted flour
Water

Condenser Alcohol Desolventization

Debitterized Grinding Drying

fenugreek powder

Figure 5.1: Flow sheet diagram for the production of debitterized fenugreek powder

60
5.2. Process description on fenugreek capsule production

Fenugreek
Cleaning Raw material drying Grinding
seed

Capsule Capsule filling Mixing of fenugreek

Dispensing
machine with ingredients

Ingredient
Polishing Jar packing Preparation
unit

Figure 5.2: Flow sheet diagram for the production of fenugreek capsule

61
 Eq-22 Eq-24 Eq-28

Eq-23 Eq-25 Eq-26 Eq-27 Eq-29

Eq-32 Eq-31

Eq-30

Eq-5 Eq-2 Eq-1

Eq-4 Eq-3
Eq-6

Eq-12 Eq-8 Eq-7

Eq-9

Eq-13 Eq-11
Eq-10
Eq-14 Starch
Gladiant
Mg Stearate
Eq-16 Eq-15
Eq-21
Eq-17
Eq-18
Eq-19
Eq-33
Eq-20
Capsule Capsule
Fenugreek powder packaging unit filling
packaging unit

Figure 5.3: Equipment layout for debitterized fenugreek powder and fenugreek capsule production

62
Equipment List
Displayed Text Description
Eq-1 Fenugreek seed storage house
Eq-2 Conveyor
Eq-3 Screen Filter
Eq-4 Conveyor
Eq-5 Rotary Filter
Eq-6 Conveyor
Eq-7 Plate type Dryer
Eq-8 Conveyor
Eq-9 Hammer Crusher
Eq-10 Conveyor
Eq-11 Extractor feed Elevator
Eq-12 Extractor Hood fitted with perforated conveyor and spray nozzles
Eq-13 Extract discharge Elevator
Eq-14 Conveyor
Eq-15 Air blown heater
Eq-16 Conveyor
Eq-17 Dryer
Eq-18 Conveyor
Eq-19 Ball Mill
Eq-20 Conveyor
Eq-21 Mixer
Eq-22 Alcohol storage vessel
Eq-23 Centrifugal pump
Eq-24 Water storage tank
Eq-25 Centrifugal pump
Eq-26 Blender
Eq-27 Centrifugal pump
Eq-28 Solvent (Alcohol + water) Storage tank
Eq-29 Centrifugal pump
Eq-30 Evaporator
Eq-31 Water + Extract storage tank
Eq-32 Condenser
Eq-33 Conveyor

63
5.3. Material and energy balance

1000 kg/h of fenugreek seeds was taken as basis for calculations of material and energy balance.
The numerical values used for calculations are based on laboratory work results.

I. Material balance
A. Material balance for debitterized fenugreek powder
1) Mass balance on cleaning
• 5.6% of impurity was recorded during cleaning of fenugreek seeds bought from
local market.

F, Fenugreek Fp, Pure fenugreek

Cleaning

I, Impurity (5.6%)
𝐹𝐹 = 𝐹𝐹𝐹𝐹 + 𝐼𝐼 𝐸𝐸𝐸𝐸. (5.1)

𝐹𝐹𝐹𝐹 = 𝐹𝐹 − 𝐹𝐹 × 0.056

𝐹𝐹 = 944 𝑘𝑘𝑘𝑘/ℎ 𝑜𝑜𝑜𝑜 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐 𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓 𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠

2) Mass balance on raw material drying

Fp, mass of Mw, mass of

cleaned Fenugreek Raw material moisture removed
944 kg drying

Fd, Mass of dried fenugreek

𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀 𝑜𝑜𝑜𝑜 𝑖𝑖𝑖𝑖𝑖𝑖𝑖𝑖𝑖𝑖 = 𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚 𝑜𝑜𝑜𝑜 𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜

𝐹𝐹𝐹𝐹 = 𝐹𝐹𝐹𝐹 + 𝑀𝑀𝑀𝑀 𝐸𝐸𝑞𝑞. (5.2)

Component balance on moisture loss

Moisture content of cleaned, whole, fenugreek seeds were 12% and dried to 5% moisture content.
Thus,

64
 0.12𝐹𝐹𝐹𝐹 = 0.05𝐹𝐹𝐹𝐹 + 𝑀𝑀𝑀𝑀 𝐸𝐸𝐸𝐸. (5.3)

0.12𝐹𝐹𝐹𝐹 = 0.05𝐹𝐹𝐹𝐹 + 𝑀𝑀𝑀𝑀 , 𝑤𝑤ℎ𝑒𝑒𝑒𝑒𝑒𝑒 𝐹𝐹𝐹𝐹 = 944 𝑘𝑘𝑘𝑘

0.12𝐹𝐹𝐹𝐹 = 0.05(𝐹𝐹𝐹𝐹 − 𝑀𝑀𝑀𝑀) + 𝑀𝑀𝑀𝑀

0.12𝐹𝐹𝐹𝐹 − 0.05𝐹𝐹𝐹𝐹 = 𝑀𝑀𝑀𝑀 − 0.05𝑀𝑀𝑀𝑀

0.07𝐹𝐹𝐹𝐹 = 0.95𝑀𝑀𝑀𝑀, 𝑤𝑤ℎ𝑒𝑒𝑒𝑒𝑒𝑒 𝐹𝐹𝐹𝐹 = 944 𝑘𝑘𝑘𝑘

𝑀𝑀𝑀𝑀 = 69.56 𝑘𝑘𝑘𝑘 𝑜𝑜𝑜𝑜 𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚 𝑤𝑤𝑤𝑤𝑤𝑤 𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒 𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑 𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑

𝐹𝐹𝐹𝐹 = 874.44 𝑘𝑘𝑘𝑘/ℎ 𝑜𝑜𝑜𝑜 𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑 𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓 𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠 𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟 𝑓𝑓𝑓𝑓𝑓𝑓 𝑔𝑔𝑔𝑔𝑔𝑔𝑔𝑔𝑔𝑔𝑔𝑔𝑔𝑔𝑔𝑔

3) Mass balance on grinding

There was around 7% loss of fenugreek flour during grinding in laboratory, however 4% loss is
assumed at industrial scale level.

Fd, Mass of dried Lf, lost fenugreek

fenugreek flour
Grinding
874.44 kg

F (flour), Fenugreek flour

𝐹𝐹𝐹𝐹 = 𝐹𝐹 (𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓) + 𝐿𝐿𝐿𝐿 𝐸𝐸𝐸𝐸. (5.4)

𝐹𝐹 (𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓) = 𝐹𝐹𝐹𝐹 − 𝐹𝐹𝐹𝐹 × 0.04

𝐹𝐹 (𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓) = 874.44 − 874.44 × 0.04

𝐹𝐹 (𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓) = 839.46 𝑘𝑘𝑘𝑘 𝑜𝑜𝑜𝑜 𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓 𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓

65
 4) Mass balance on extraction

S, Solvent (90% ethanol)

F (flour),
fenugreek flour E, Extract
Extraction

Wdf, wet debitterized

fenugreek

Component balance on debitterized wet fenugreek

To study the effect of extraction on phytochemicals, 50g of fenugreek flour was used at a time and
the following data were obtained:

 About 1L of solvent (90% ethanol) was used to extract the 50g of fenugreek flour. Since
ethanol has a density less than one (0.789g/ml), the weight of solvent was 810.1g using
the density formula;
𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚
𝐷𝐷𝐷𝐷𝐷𝐷𝐷𝐷𝐷𝐷𝐷𝐷𝐷𝐷 = 𝐸𝐸𝐸𝐸. (5.5)
𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣

 772.1g of extract that contains solvent and leached out materials, and
 88g of wet debitterized fenugreek flour were obtained.

Therefore, according to these results, the mass balance on the extraction process is summarized in
the following chart by scaling up the data obtained from extraction of 50g of fenugreek flour to the
basis, 839.46kg of fenugreek flour, from grinding unit operation.

S = 13600.93kg

F (flour) = 839.46 kg E = 12962.94kg

Extraction

Wdf =1477.45

66
 𝑊𝑊𝑊𝑊𝑊𝑊 = 𝟏𝟏𝟏𝟏𝟏𝟏𝟏𝟏. 𝟒𝟒𝟒𝟒𝟒𝟒𝟒𝟒 𝑜𝑜𝑜𝑜 𝑤𝑤𝑤𝑤𝑤𝑤 𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑 𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓 𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓

5) Mass balance on distillation

Ethanol, MET,

E = 12962.94kg
Distillation MW + MPHY
MW + MET +MPHY
tower

Where:-

E – Extract that contains extracted matter (phytochemicals and solvent (ethanol & water)

MW, MET, and MPHY - mass of water, ethanol and phytochemicals; respectively

 Mass of phytochemicals (MPHY) = 169.24 kg; this was calculated from laboratory result in
where 10.08g of phytochemicals was extracted from 50g of fenugreek flour. Therefore,
169.24 kg phytochemicals will be extracted from 839.46 kg of fenugreek flour.

Mass of solvent (ethanol and water) = 12962.94 - 169.24 = 12793.7 kg and assuming that the
proportion of ethanol to water is not affected by the extraction process;

 Mass water (MW) = 10% of solvent = 1,279.37 kg and,

 Mass of ethanol (MET) = 90% of solvent = 11,512.33 kg

 MW + MPHY = 169.24 kg + 1279.37kg = 1448.61 kg. This was the end byproduct from the
extraction process and it was assessed for its application in biopest management by applying
on to roaches and insects. And it kills all of the trial animals. Therefore, the byproduct,
containing water and phytochemicals, can be used for pest management. Similarly, the
toxicity of saponins to insects (insecticidal activity), parasite worms (anthelmintic activity),
molluscs (molluscicidal), and fish (piscidal activity) and their antifungal, antiviral, and
antibacterial activity were documented by Güçlü-Üstündağ et al., 2007.

67
 6) Mass balance on desolventization

Ethanol, METd,

Wdf =1477.45kg Wf
Desolventization
(MWd + METd + Df) (MWd + Df)

Where:

Wdf – wet (ethanol + water) debitterized fenugreek after extraction

MWd, METd, and Df - mass of water, ethanol and dry solid of debitterized fenugreek;
respectively in the desolventizer.

Wf – wet (water) debitterized fenugreek after desolventization

 Mass of dry solid of debitterized fenugreek (Df ) = 670.22 kg; this is calculated from
laboratory result in which 39.92g of dry solid of debitterized fenugreek obtained from 50g
of raw fenugreek flour. Therefore, 670.22 kg phytochemicals will be extracted from
839.46kg of fenugreek flour.

Mass of solvent (ethanol and water, MWd + METd) = 1477.45 – 670.22 = 807.23 kg and assuming
that the proportion of ethanol to water is not affected by the extraction process;

 Mass water (MWd) = 10% of solvent = 80.723 kg and,

 Mass of ethanol (METd) = 90% of solvent = 726.507 kg

Wf (MWd + Df) = 80.723 kg + 670.22 kg = 750.943 kg transferred to dryer

7) Mass balance on product drying

Wf, Product
M, Moisture
750.943 kg drying (water)

Ff, Debitterized fenugreek flakes

On average 39.92g of debitterized fenugreek flakes were obtained from each 50g of flour or 88g of
wet debitterized fenugreek flour used in each runs of extraction process. Based on this result the

68
amount debitterized fenugreek flakes that could be obtained from 839.46 kg of flour or 1477.45kg
of debitterized wet fenugreek will be 670.22 kg.

𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀 𝑜𝑜𝑜𝑜 𝑖𝑖𝑖𝑖𝑖𝑖𝑖𝑖𝑖𝑖 = 𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚 𝑜𝑜𝑜𝑜 𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜

𝑊𝑊𝑊𝑊 = 𝐹𝐹𝐹𝐹 + 𝑀𝑀 𝐸𝐸𝐸𝐸. (5.6)

𝑀𝑀 = 80.723 𝑘𝑘𝑘𝑘 − 670.22 𝑘𝑘𝑘𝑘

𝑀𝑀 = 80.723𝑘𝑘𝑘𝑘 𝑜𝑜𝑜𝑜 𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚 𝑤𝑤𝑤𝑤𝑤𝑤 𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟 𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑 𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑, 𝑎𝑎𝑎𝑎𝑎𝑎

𝐹𝐹𝐹𝐹 = 670.22 𝑘𝑘𝑘𝑘 𝑜𝑜𝑜𝑜 𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑 𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓 𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓 𝑤𝑤𝑤𝑤𝑤𝑤𝑤𝑤 𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜

8) Mass balance on grinding/homogenizing & packing

There was only about 1% loss of fenugreek flour during grinding/homogenizing of dried flakes in
laboratory.

Ff, Debitterized
W, wasted flour
fenugreek flakes Grinding/homogenizing
670.22 kg
& packing

Dp, Debitterized fenugreek powder

𝐹𝐹𝐹𝐹 = 𝐷𝐷𝐷𝐷 + 𝑊𝑊 𝐸𝐸𝐸𝐸. (5.7)

𝐷𝐷𝐷𝐷 = 𝐹𝐹𝐹𝐹 − 0.01𝐹𝐹𝐹𝐹

𝐷𝐷𝐷𝐷 = 670.22 − 6.7

𝐷𝐷𝐷𝐷 = 663.52 𝑘𝑘𝑘𝑘 𝑜𝑜𝑜𝑜 𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑𝑑 𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓 𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝 𝑤𝑤𝑤𝑤𝑤𝑤 𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜

Therefore, 663.52 kg of debitterized fenugreek powder is obtained from 1000kg of whole

fenugreek seeds.

B. Material balance for fenugreek capsule

Until the grinding unit operation similar processes are carried out to produce fenugreek capsule and
debitterized fenugreek powder. After grinding, the ground fenugreek flour is subjected to extraction
during debitterized fenugreek powder production while it is directly filled in to capsules during
fenugreek capsule production.

69
Mass balance on capsule filling

According to the information collected from capsule manufacturers 0.2% of flour is lost during
filling in to capsules and 2% of ingredients are added to enhance flow ability.

Ing, Ingredients (2%)

Fg , ground flour Wf, wasted flour (0.2%)

Capsule filling
839.46 kg

Fc, Flour filled in capsule

𝐹𝐹𝐹𝐹 + 𝐼𝐼𝐼𝐼𝐼𝐼 = 𝐹𝐹𝐹𝐹 + 𝑊𝑊𝑊𝑊 𝐸𝐸𝐸𝐸. (5.8)

𝐹𝐹𝐹𝐹 + 0.02𝐹𝐹𝐹𝐹 = 𝐹𝐹𝐹𝐹 + 0.002𝐹𝐹𝐹𝐹

𝐹𝐹𝐹𝐹 = 854.57𝑘𝑘𝑘𝑘 𝑜𝑜𝑜𝑜 𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓 𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓 𝑖𝑖𝑖𝑖 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐

Summary of mass balance

1000kg/h = initial fenugreek seed used as a basis for calculation

944 kg/h = cleaned fenugreek (cleaning unit operation)
874.44 kg/h = dried fenugreek seeds (raw material tempering unit operation)
839.46 kg/h = ground fenugreek flour (grinding unit operation)
1477.45 kg/h = wet debitterized fenugreek flour (extraction process)
670.22 kg/h = debitterized and dried fenugreek flakes (product drying unit operation)
663.52 kg = packed debitterized fenugreek powder from 1000kg/h of raw fenugreek seeds
II. Energy balance

Energy balance for debitterized fenugreek powder

 Energy balance on raw material tempering

70
 TA2, WA2

Fenugreek Dried fenugreek

MS, W1, TS1, Ma1 Drier MS, W2, TS2, Ma2

TA1, WA1
Where:-

MS - dry solid matter of fenugreek seeds; respectively

TA1 and TA2 – temperature of air at the inlet and out let; respectively

TF1 and TF2 – temperature of fenugreek seed at the inlet and out let; respectively

WA1 and WA2 = moisture content of inlet and outlet air; respectively

Ma1, M a2 - mass flow rate of air at the inlet and outlet; respectively

HF and HP = enthalpy of feed and dried fenugreek seed; respectively

HA1 and HA2 = enthalpy of inlet and outlet air; respectively

Latent heat of vaporization at 0oc = 2507kJ/kg

Specific heat of air (CP, A) = 0.24+ 0.46 (enthalpy)

Specific heat of solid matter of dried fenugreek seed (CPfg, P) = 1.68553 kJ/kg

Specific heat of water (CW) = 4.2kJ/kg

q - thermal energy loss of the dryer

Assumption:-

• Hot air entering (TA1) into the drier = 60 OC at 10% RH, WA1= 0.014kg/kg dry and enthalpy
of inlet air is 90 kJ/kg dry air (from psychometric chart)
• Air temperature leaving the drier, TA2 = 300C, reference temperature, T0 = 00C
• Moisture content of fenugreek seed at inlet and out let are 12% and 5%; respectively

Data required:

 Solid matter of cleaned fenugreek seed (MS) = 944 – (944 * 0.12) = 830.72 kg

71
  Moisture content in feed (W1) = 12.0% = (12/ (100-12)) = 0.13 kg of water /kg of solid
 Moisture content in dried fenugreek seed (W2) = 5% = (5 / (100-5)) = 0.053 kg of water /kg
of solid
 Me - mass flow of evaporated water = Mw1 – Mw2
 TF1 = 20 0C, and 50% RH
 TF2 = 30 0C
 At pressure of 101,325 pa, latent heat of vaporization of water, λ = 2256.6 kJ/kg
 Specific heat of the fenugreek seed can be calculated from the proximate composition of
raw fenugreek flour (Table 4.7) using (Choi & Okos 1986) model using the formula:

𝐶𝐶𝐶𝐶 = � 𝐶𝐶𝐶𝐶𝐶𝐶 ∗ 𝑥𝑥𝑥𝑥 𝐸𝐸𝐸𝐸. (5.9)

Cprs = 1.424 * Xcfb + 1.549 * Xpfb + 1.675 * Xffb + 0.837 * Xafb + 4.187 * Xmfb

Where,

Cp = specific heat capacity of the material

Cpi = specific heat capacity of ith component

xi = mass fraction of ith component

Cpfg = specific heat capacity of fenugreek

Xcfb = mass fraction of total carbohydrates

Xpfb = mass fraction of protein

Xffb = mass fraction of fat

Xafb = mass fraction of ash

Xmfb = mass fraction of moisture

All values of these mass fractions are found in table 4.1 for Challa variety.

Cpfg = 1.424 * Xcfb + 1.549 * Xpfb + 1.675 * Xffb + 0.837 * Xafb + 4.187 * Xmfb

=1.424*0.517 + 1.549*0.307 + 1.675*0.0755 + 0.837*0.0296 + 4.187*0.077

= 1.68553

From mass balance equation:

MS (W1) + MA (WA1) = MS (W2) + MA (WA2)

72
830.72 (0.13) + MA (0.014) = 830.72 (0.053) + MA (WA2)

𝑘𝑘𝑘𝑘
𝑀𝑀𝑎𝑎 (𝑊𝑊𝑊𝑊2) = 63.97 + 𝑀𝑀𝑀𝑀 (0.014) 𝐸𝐸𝐸𝐸. (5.10)
ℎ

From energy balance equation:

𝑀𝑀𝑀𝑀 (𝐻𝐻𝐻𝐻) + 𝑀𝑀a(ℎ𝑎𝑎1) = 𝑀𝑀𝑀𝑀 (𝐻𝐻𝐻𝐻) + 𝑀𝑀𝑀𝑀 (ℎ𝑎𝑎2) + 𝑞𝑞 𝐸𝐸𝐸𝐸. (5.11)

Assuming an adiabatic system, the above equation can be rewritten as:

𝑀𝑀𝑀𝑀 (𝐻𝐻𝐻𝐻) + 𝑀𝑀a(ℎ𝑎𝑎1) = 𝑀𝑀𝑀𝑀 (𝐻𝐻𝐻𝐻) + 𝑀𝑀𝑀𝑀 (ℎ𝑎𝑎2) 𝐸𝐸𝐸𝐸. (5.12)

9) HF = Cpfg*T + W1* Cw* T

HF = 1.68553(20) +0.13*4.2*20

HF = 44.63 kJ/kg

10) HP = Cpfg*T + W2* Cw* T

HP = 1.68553(30) + 0.053*4.2*30

HP = 57.24 kJ/kg

Ha2 = CpaT + λ WA2

Ha2 = (0.24 + 0.46 WA2) 30 + 2256.6 * WA2

Ha2 = 7.2 + 2270.4WA2

𝐻𝐻𝐻𝐻2 = 7.2 + 2270.4𝑊𝑊𝑊𝑊2 𝐸𝐸𝐸𝐸. (5.13)

Substituting eq. 5.13 in eq. 5.12:

𝑀𝑀𝑀𝑀 (𝐻𝐻𝐻𝐻) + 𝑀𝑀a (ℎ𝑎𝑎1) = 𝑀𝑀𝑀𝑀 (𝐻𝐻𝐻𝐻) + 𝑀𝑀𝑀𝑀 (7.2 + 2270.4𝑊𝑊𝑊𝑊2) 𝐸𝐸𝑞𝑞. (5.14)

830.72 (44.63) + MA (90.0) = 830.72 (57.24) + MA (7.2 +2270.4WA2)

82.8 𝑀𝑀𝑀𝑀 = 10475.38 + 2270.4 𝑊𝑊𝑊𝑊2 (𝑀𝑀𝑀𝑀) 𝐸𝐸𝐸𝐸. (5.15)

Substituting eq. 5.10 it in to eq. 5.15:

𝑘𝑘𝑘𝑘
𝑀𝑀𝑎𝑎 (𝑊𝑊𝑊𝑊2) = 63.97 + 𝑀𝑀𝑀𝑀 (0.014)
ℎ

𝑘𝑘𝑘𝑘
82.8 𝑀𝑀𝑀𝑀 = 10475.38 + 2270.4 �63.97 + 𝑀𝑀𝑀𝑀 (0.014)�
ℎ

50.414 MA = 155712.87

73
MA =3088.68 kg/h of air flow rate

Substituting MA in eq. 5.10;

𝑘𝑘𝑘𝑘
𝑀𝑀𝑎𝑎 (𝑊𝑊𝑊𝑊2) = 63.97 + 𝑀𝑀𝑀𝑀 (0.014)
ℎ

3088.68 𝑘𝑘𝑘𝑘/ℎ (𝑊𝑊𝑊𝑊2) = 63.97𝑘𝑘𝑘𝑘/ℎ + 3088.68 𝑘𝑘𝑘𝑘/ℎ (0.014)

3088.68(𝑊𝑊𝑊𝑊2) = 107.21

WA2 = 0.035kg/kg dry air

HA2 = 7.2 + 2270.4 (0.035)

HA2 = 86.66 kJ/kg

Heat load calculations for raw material tempering

Heat leaving the drier with the water remaining in the dried fenugreek:

𝑄𝑄1 = 𝑀𝑀𝑀𝑀2 ∗ 𝐶𝐶𝐶𝐶𝐶𝐶 ∗ 𝛥𝛥𝛥𝛥𝛥𝛥 = 43.72 ∗ 4.187 ∗ (30 − 20) = 1,830.06𝐾𝐾𝐾𝐾/ℎ

Heat leaving the drier with water vapor in the drying air:

𝑄𝑄2 = 𝑀𝑀𝑀𝑀 ∗ 𝜆𝜆0 + 𝑀𝑀𝑀𝑀 ∗ 𝐶𝐶𝐶𝐶𝐶𝐶 ∗ 𝛥𝛥𝛥𝛥𝛥𝛥

𝑄𝑄2 = (69.56 ∗ 2256.9) + [69.56 ∗ 4.187 ∗ (60 − 30)] = 1,578,591.9𝐾𝐾𝐾𝐾/ℎ

Heat leaving the drier with the dried fenugreek seed:

𝑄𝑄3 = 𝑀𝑀𝑀𝑀 ∗ 𝐶𝐶𝐶𝐶𝐶𝐶𝐶𝐶 ∗ 𝛥𝛥𝛥𝛥𝛥𝛥 = 830.72 ∗ 1.68553 ∗ (30 − 20) = 14,000.2𝐾𝐾𝐾𝐾/ℎ

Total heat load of the drier:

𝑄𝑄𝑄𝑄 = 1,830.06 + 1,578,591.9 + 14,000.2 = 1581822.16𝐾𝐾𝐾𝐾/ℎ𝑟𝑟

𝑄𝑄𝑄𝑄 = 439.4 𝐾𝐾𝐾𝐾ℎ

74
  Energy balance on distillation

Ethanol, MET, CPET, TET2

Extract, E = 12962.94kg
MW, CPW, TW1 MW, CPW, TW2
Distillation MPHY, CPHY, TPHY2
MET, CPET, TET1 tower
MPHY, CPHY, TPHY1

Where:-

E – extract that contains extracted matter (phytochemicals and solvent (ethanol & water)

MW, MET, and MPHY - mass of water, ethanol and phytochemicals; respectively

CPW, CPET, and CPHY – specific heat capacities of water, ethanol and phytochemicals; respectively

TW1, TET1, and TPHY1 – inlet temperatures of water, ethanol and phytochemicals; respectively

TW2, TET2, and TPHY2 – outlet temperatures of water, ethanol and phytochemicals; respectively

Data required:

 Boiling point of ethanol = 78.5 0C

 Latent heat of vaporization (λe) of ethanol at 78.5 0C = 855kJ/kg
 Specific heat of water (CPW) = 4.2kJ/kg
 Specific heat of ethanol (CPET) = 2.845kJ/kg
 Specific heat of phytochemicals (CPHY) = 1.5kJ/kg; this was calculated from the proximate
composition results for raw and extracted fenugreek flours. Thus, considering that the
reduction of carbohydrates and mineral observed for extracted fenugreek flour, the specific
heat capacity was calculated using (Choi & Okos 1986) model by taking only the mass
fraction of carbohydrates and mineral.
 Inlet temperatures of extract (TW1, TET1, and TPHY1) = 22 0C and
 Distillation process is controlled at temperature of 80 0C so that the final temperature of
water, ethanol and phytochemicals (TW2, TET2, and TPHY2; respectively) = 80 0C and
 From material balance on distillation:-
• Mass of phytochemicals (MPHY) = 169.24 kg;

75
 • Mass water (MW) = 1279.37kg and,
• Mass of ethanol (MET) = 11512.33 kg

Heat load calculations during distillation

Heat leaving with the water in the extract:

𝑄𝑄𝑄𝑄 = 𝑀𝑀𝑀𝑀 ∗ 𝐶𝐶𝐶𝐶𝐶𝐶 ∗ 𝛥𝛥𝛥𝛥𝛥𝛥 = 1279.37 ∗ 4.187 ∗ (80 − 22) = 310,689.89𝐾𝐾𝐾𝐾/ℎ

Heat leaving with ethanol:

𝑄𝑄𝑄𝑄 = 𝑀𝑀𝑀𝑀𝑀𝑀 ∗ 𝜆𝜆𝜆𝜆 + 𝑀𝑀𝑀𝑀𝑀𝑀 ∗ 𝐶𝐶𝐶𝐶𝐶𝐶𝐶𝐶 ∗ 𝛥𝛥𝛥𝛥𝛥𝛥𝛥𝛥

𝑄𝑄𝑄𝑄 = (11512.33 ∗ 855) + [11512.33 ∗ 2.845 ∗ (80 − 22)] = 11,742,692𝐾𝐾𝐾𝐾/ℎ

Heat leaving with the phytochemicals:

𝑄𝑄𝑄𝑄ℎ𝑦𝑦 = 𝑀𝑀𝑀𝑀ℎ𝑦𝑦 ∗ 𝐶𝐶𝐶𝐶ℎ𝑦𝑦 ∗ 𝛥𝛥𝛥𝛥𝛥𝛥ℎ𝑦𝑦 = 169.24 ∗ 1.5 ∗ (80 − 22) = 14,723.88𝐾𝐾𝐾𝐾/ℎ

Total heat load during distillation:

𝑄𝑄𝑄𝑄 = 302,654.8 + 11,693,563 + 14,343.09 = 12,068,106𝐾𝐾𝐾𝐾/ℎ𝑟𝑟

𝑄𝑄𝑄𝑄 = 3,352.25𝐾𝐾𝐾𝐾ℎ

 Energy balance during desolventization

Ethanol, METd, CPET, TETd2

Wdf =1477.45kg MWd, CPW, TWd2

MWd, CPW, TWd1 Desolventization METd, CPET, TETd2
METd, CPET, TETd1
MDF, CPDF, TDF2
MDF, CPDF, TDF1

Where:-

Wdf – wet (ethanol + water) debitterized fenugreek after extraction

MWd, METd, and MDF - mass of water, ethanol and dry solid of debitterized fenugreek;
respectively in the desolventizer.

CPW, CPET, and CPDF – specific heat capacities of water, ethanol and dry solid of debitterized
fenugreek; respectively

76
 TWd1, TETd1, and TDF1 – inlet temperatures of water, ethanol and dry solid of debitterized
fenugreek; respectively

TWd2, TETd2, and TDF2 – final temperatures of water, ethanol and dry solid of debitterized
fenugreek; respectively

Data required:

 Boiling point of ethanol = 78.5 0C

 Latent heat of vaporization (λe) of ethanol at 78.5 0C = 855kJ/kg
 Specific heat of water (CPW) = 4.2kJ/kg
 Specific heat of ethanol (CPET) = 2.845kJ/kg
 Specific heat of the extracted fenugreek flour be calculated from its proximate composition
(Table 4.7) using (Choi & Okos 1986) model using the formula:𝐶𝐶𝐶𝐶 = ∑ 𝐶𝐶𝐶𝐶𝐶𝐶 ∗ 𝑥𝑥𝑥𝑥 = CPDF =
1.646638
 Inlet temperatures of extract (TWd1, TETd1, and TDF1) = 22 0C and
 Live (direct) steam is supplied during desolventization so that the final temperatures are
higher than distillation which is temperature controlled process at 80 0C of water. Thus,
outlet temperature of ethanol and dry solid of debitterized fenugreek (TWd2, TETd2, and TDF2;
respectively) = 85 0C and
 From bacterial balance on desolventization:-
• Mass of dry solid of debitterized fenugreek (MDF) = 670.22 kg;
• Mass water (MWd) = 80.723 kg and,
• Mass of ethanol (METd) = 726.507 kg

Heat load calculations during desolventization

Heat leaving with the water in the extract:

𝑄𝑄𝑄𝑄𝑄𝑄 = 𝑀𝑀𝑀𝑀𝑀𝑀 ∗ 𝐶𝐶𝐶𝐶𝐶𝐶 ∗ 𝛥𝛥𝛥𝛥𝛥𝛥 = 80.723 ∗ 4.187 ∗ (63) = 21,293.194𝐾𝐾𝐾𝐾/ℎ

Heat leaving with ethanol:

𝑄𝑄𝑄𝑄𝑄𝑄𝑄𝑄 = 𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀 ∗ 𝜆𝜆𝜆𝜆 + 𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀 ∗ 𝐶𝐶𝐶𝐶𝐶𝐶𝐶𝐶 ∗ 𝛥𝛥𝛥𝛥𝛥𝛥𝛥𝛥𝛥𝛥

𝑄𝑄𝑄𝑄𝑄𝑄𝑄𝑄 = (726.507 ∗ 855) + [726.507 ∗ 2.845 ∗ (63)] = 751,378.94𝐾𝐾𝐾𝐾/ℎ

Heat leaving with dry solid of debitterized fenugreek:

𝑄𝑄𝑄𝑄𝑄𝑄 = 𝑀𝑀𝑀𝑀𝑀𝑀 ∗ 𝐶𝐶𝐶𝐶𝐶𝐶𝐶𝐶 ∗ 𝛥𝛥𝛥𝛥𝛥𝛥𝛥𝛥 = 670.22 ∗ 1.646638 ∗ (63) = 69,527.411𝐾𝐾𝐾𝐾/ℎ

77
Total heat load during desolventization:

𝑄𝑄𝑄𝑄 = 21,293.194 + 751,378.94 + 69,527.411 = 842,199.54𝐾𝐾𝐾𝐾/ℎ𝑟𝑟

𝑄𝑄𝑄𝑄 = 233.94 𝐾𝐾𝐾𝐾ℎ

 Water required for condensation

𝑄𝑄 = 𝑀𝑀𝑀𝑀 ∗ 𝐶𝐶𝐶𝐶𝐶𝐶 ∗ 𝛥𝛥𝛥𝛥𝛥𝛥

Total heat required to evaporate ethanol ( 𝑄𝑄 = 𝑄𝑄𝑄𝑄 + 𝑄𝑄𝑄𝑄𝑄𝑄𝑄𝑄) = 11,742,692 + 751,378.94𝐾𝐾𝐾𝐾/ℎ

from distillation and desolventization; respectively and water enters the cooling tower at 22 0C and
leaves at 30 0C. Thus, 𝑄𝑄 = 12,494,071𝐾𝐾𝐾𝐾/ℎ

12,494,071 = 𝑀𝑀𝑀𝑀 ∗ 4.187 ∗ (30 − 22)

𝑀𝑀𝑀𝑀 = 373,001.88 𝑘𝑘𝑘𝑘(𝐿𝐿𝐿𝐿𝐿𝐿𝐿𝐿𝐿𝐿𝐿𝐿)

 Energy balance on product drying

Heat load calculations for product drying (Qpd)

Assumption:-

• Hot air entering (TA1) into the drier = 80 OC

• Air temperature leaving the drier, TA2 = 40 0C, reference temperature, T0 = 00C
• Moisture content of debitterized fenugreek flakes at out let 5%

Data required:

 Solid matter of cleaned fenugreek seed (MS) = 670.22 – (670.22 * 0.05) = 636.709 kg
 Me - mass flow of evaporated water = 80.72 kg; from mass balance of product drying
 Mw2 – moisture in debitterized fenugreek flakes = 670.22*0.05 = 33.511 kg
 Inlet temperature of debitterized fenugreek flakes (TF1) = 30 0C,
 outlet temperature of debitterized fenugreek flakes (TF2) = 40 0C
 At pressure of 101,325 pa, latent heat of vaporization of water, λ = 2256.6 kJ/kg

Heat leaving the drier with the water remaining in the dried fenugreek:

𝑄𝑄𝑄𝑄𝑄𝑄1 = 𝑀𝑀𝑀𝑀2 ∗ 𝐶𝐶𝐶𝐶𝐶𝐶 ∗ 𝛥𝛥𝛥𝛥𝛥𝛥 = 33.511 ∗ 4.187 ∗ (40 − 30) = 1,403.11𝐾𝐾𝐾𝐾/ℎ

Heat leaving the drier with water vapor in the drying air:

𝑄𝑄𝑄𝑄𝑄𝑄2 = 𝑀𝑀𝑀𝑀 ∗ 𝜆𝜆0 + 𝑀𝑀𝑀𝑀 ∗ 𝐶𝐶𝐶𝐶𝐶𝐶 ∗ 𝛥𝛥𝛥𝛥𝛥𝛥

78
𝑄𝑄𝑄𝑄𝑄𝑄2 = (80.72 ∗ 2256.6) + [80.72 ∗ 4.187 ∗ (80 − 40)] = 195,671.74𝐾𝐾𝐾𝐾/ℎ

Heat leaving the drier with the dried fenugreek seed:

𝑄𝑄𝑄𝑄𝑄𝑄3 = 𝑀𝑀𝑀𝑀 ∗ 𝐶𝐶𝐶𝐶𝐶𝐶𝐶𝐶 ∗ 𝛥𝛥𝛥𝛥𝛥𝛥 = 636.709 ∗ 1.646638 ∗ (40 − 30) = 10,484.292𝐾𝐾𝐾𝐾/ℎ

Total heat load during product drying:

𝑄𝑄𝑄𝑄 = 1,403.11 + 195,671.74 + 10,484.292 = 207,559.14𝐾𝐾𝐾𝐾/ℎ𝑟𝑟

𝑄𝑄𝑄𝑄 = 57.66 𝐾𝐾𝐾𝐾ℎ

Summary of energy balance:

Heat load of material tempering = QT =439.4Kwh

Heat load of distillation = QT =3,352.25Kwh

Heat load of desolventization = QT =233.94Kwh

Heat load of product drying = QT =57.66Kwh

Total heat load extraction (QT) = 4,083.25Kwh for 1000kg/h of raw fenugreek seeds

5.4. Financial and economic analysis

5.4.1. Plant capacity and production programming

Considering the availability of raw materials, machineries, skilled labor and future market, and
based on the material balance (663.22kg of debitterized fenugreek flour/ 1000kg of raw fenugreek
seed); the plant will have an installed processing capacity of 45,300Qtl of fenugreek seeds per year
with annual production of 30,000Qtl of debitterized fenugreek powder. The factory will initially
operate at 75% capacity in the first year, 85% in the second year and 100% after third year. The
plant is planned to work for 300 days per annum and in a double shift of 16h per day.

Table 5.1: Plant capacity and production programming

No. Description Production program

1St year 2nd year After 3rd year

1 Capacity utilization (%) 75 85 100

2 Production rate (Qtl/year) 22,500 25,500 30,000

79
 5.4.2. Project cost estimation

A. Purchased equipment cost

Table 5.2: Purchased equipment cost

No. Equipment Quantity Capacity Unit price (Birr) Total Price (Birr)
1 Raw material silo 1 250 m3 300, 000.00 300,000.00
2 Belt conveyer 2 Length 9m 190,353.00 380,706.00*
3 Cleaning machine 1 30Qtl/h 542,595.00 542,595.00*
4 Seed weighing scale 1 10Qtl/h 5,000.00 5,000.00
5 Tray dryer 2 15 m2 480,706.00 961,412.00 **
6 Hammer Mill 1 20Qtl/ h 622,650.00 622,650.00
7 Extractor 1 15 ton/day 7,900,000 6,010,000.00
8 Solvent storage vessel 3 5000 gallon 266,850.00 800,550.00*
9 Boiler 1 2500 lb/h 6,186,472.00 2,186,472.00*
10 Distillation tower 1 10000 kg/h 955,800.00 955,800.00**
11 Desolventization 1 1000 kg/h 553,700.00 553,700.00**
12 Vapor condenser 1 15000 kg/h 711,600.00 711,600.00**
13 Product silo 1 100 m3 190,000.00 190,000.00
14 Packaging machine 1 40Qtl/h 266,850.00 266,850.00
15 Miscellaneous 700,000.00
Total cost of purchased equipments 15,187,335.00
*Costs obtained from the internet (http://www.matche.com/EquipCost/Index.htm, retrieved on April 25, 2012.
**Costs obtained from the internet (http://www.alibaba.com/EquipCost/Index.htm, retrieved on May 02, 2012.
Other costs were taken from local company and personal contact.
B. Total capital investment cost

Total capital investment is estimated based on delivered equipment cost of solid-fluid processing
plant; according to Peters & Timmerhaus (1991). The capital investment is calculated as a factor of
the ratios of equipment purchase cost.

 Purchased equipment cost = 15,187,335.00 Birr

 Assuming delivery cost = 10% of purchased equipment cost
 Purchased-Delivered equipment cost = 16,706,068.50 Birr

80
Table 5.3: Total investment cost

Item Percentage (%) of PEC Price (Birr)

Direct Cost
Purchased & Delivered equipment cost (PEC) 100% 16,706,068.50
Purchased Equipment Installation 33% 5,513,002.61
Instrumentation and Controls 11% 1,837,667.54
Piping Installation 25% 4,176,517.13
Electrical Installation 10% 1,670,606.85
Buildings 29% 4,844,759.87
Yard Improvements 8% 1,336,485.48
Service Facilities 45% 7,517,730.83
Land 6% 1,002,364.11
Total Direct Cost (A) 44,605,202.90
Indirect Cost
Engineering and Supervision 26% 4,343,577.81
Construction Expenses 28% 4,677,699.18
Total Indirect Cost (B) 9,021,276.99
Total Direct and Indirect Cost (A+B) 53,626,479.89
Contractor's fee (C) = 5% (A+B) 2,681,323.99
Contingency (D) = 10% (A+B) 5,362,647.99
Fixed Capital Investment (FCI) = A+B+C+D 61,670,451.87
Working Capital (WC) = 15% FCI 9,250,567.78
Total Capital Investment (TCI) = FCI + WC 70,921,019.65

C. Total product cost

 Raw materials cost

The major raw materials used in the production of debitterized fenugreek powder are Fenugreek
seeds and ethanol. The annual raw fenugreek seed required in full plant capacity utilization is
45,300Qtl per year to produce 30,000Qtl of debitterized fenugreek powder. Since ethanol is
recovered by distillation, 10kg = 12.7 liters of ethanol/1000kg of raw material is consumed (lost
through leakages, washed with product and other wastages). Therefore, 45,300 kg (57,414 liters) of
ethanol is required annually.

81
 𝑈𝑈𝑈𝑈𝑈𝑈𝑈𝑈 𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝 𝑜𝑜𝑜𝑜 𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓 𝑝𝑝𝑝𝑝𝑝𝑝 𝑄𝑄𝑄𝑄𝑄𝑄 = 2000𝐵𝐵𝐵𝐵𝐵𝐵𝐵𝐵

𝑈𝑈𝑈𝑈𝑈𝑈𝑈𝑈 𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝 𝑜𝑜𝑜𝑜 𝑒𝑒𝑒𝑒ℎ𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎 𝑝𝑝𝑝𝑝𝑝𝑝 𝑙𝑙𝑙𝑙𝑙𝑙𝑙𝑙𝑙𝑙 = 13𝐵𝐵𝐵𝐵𝐵𝐵𝐵𝐵

𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇 𝑟𝑟𝑟𝑟𝑟𝑟 𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚𝑚 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐 𝑝𝑝𝑝𝑝𝑝𝑝 𝑦𝑦𝑦𝑦𝑦𝑦𝑦𝑦 = 45,300 ∗ 2000 + 57,414 ∗ 13

𝑻𝑻𝑻𝑻𝑻𝑻𝑻𝑻𝑻𝑻 𝒓𝒓𝒓𝒓𝒓𝒓 𝒎𝒎𝒎𝒎𝒎𝒎𝒎𝒎𝒎𝒎𝒎𝒎𝒎𝒎𝒎𝒎 𝒄𝒄𝒄𝒄𝒄𝒄𝒄𝒄 𝒑𝒑𝒑𝒑𝒑𝒑 𝒚𝒚𝒚𝒚𝒚𝒚𝒚𝒚 = 𝟗𝟗𝟗𝟗, 𝟑𝟑𝟑𝟑𝟑𝟑, 𝟑𝟑𝟑𝟑𝟑𝟑. 𝟎𝟎𝟎𝟎 𝑩𝑩𝑩𝑩𝑩𝑩𝑩𝑩

 Auxiliary materials cost

The debitterized fenugreek flour is packed in sacks with 1Qtl holding capacity. Since 30,000Qtl of
debitterized fenugreek flour is produce annually (from the material balance), 30,000 sacks are
required. The unit price sack is 4birr per piece.

𝑨𝑨𝑨𝑨𝑨𝑨𝑨𝑨𝑨𝑨𝑨𝑨𝑨𝑨𝑨𝑨𝑨𝑨 𝒎𝒎𝒎𝒎𝒎𝒎𝒎𝒎𝒎𝒎𝒎𝒎𝒎𝒎𝒎𝒎𝒎𝒎 𝒄𝒄𝒄𝒄𝒄𝒄𝒄𝒄 𝒑𝒑𝒑𝒑𝒑𝒑 𝒚𝒚𝒚𝒚𝒚𝒚𝒚𝒚 = 30,000 ∗ 4 = 𝟏𝟏𝟏𝟏𝟏𝟏, 𝟎𝟎𝟎𝟎𝟎𝟎 𝑩𝑩𝑩𝑩𝑩𝑩𝑩𝑩

 Cost of utilities

Electricity and water are the two major utilities used during production of debitterized fenugreek
flour.

From summery of energy balance, total heat load during extraction processes = 4,083.25Kwh for
1000kg/h of raw fenugreek seeds and assuming the electric consumption for other equipments and
lighting purpose of the company to be 50Kwh;

𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇 𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎 𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐 = (4,083.25 + 50)𝐾𝐾𝐾𝐾ℎ ∗ 4,530

𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇 𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎 𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑟𝑟𝑟𝑟𝑟𝑟 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐 = 18,723,623𝐾𝐾𝐾𝐾ℎ

𝑈𝑈𝑈𝑈𝑈𝑈𝑈𝑈 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐 𝑜𝑜𝑜𝑜 𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒 = 58𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐/𝐾𝐾𝐾𝐾ℎ

𝑨𝑨𝑨𝑨𝑨𝑨𝑨𝑨𝑨𝑨𝑨𝑨 𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆 𝒄𝒄𝒄𝒄𝒄𝒄𝒄𝒄 = 0.58𝐵𝐵𝐵𝐵𝐵𝐵𝐵𝐵 ∗ 18,723,623 = 𝟏𝟏𝟏𝟏, 𝟖𝟖𝟖𝟖𝟖𝟖, 𝟕𝟕𝟕𝟕𝟕𝟕. 𝟎𝟎𝟎𝟎𝟎𝟎𝟎𝟎𝟎𝟎𝟎𝟎

From the material balance13, 600.93kg of solvent (90% ethanol & 10% water) was used for
extraction of 839.46kg of fenugreek flour. Therefore, 1,360kg of water per 839.46kg of fenugreek
flour was consumed which is equivalent to annually 3,802,753.8kg (liters) of water per 45,300Qtl
of fenugreek flour. This will be 3,802.8m3 of water per year and adding 1000 m3 for other purposes,
the total water consumption per annum will be 4,802.8 m3.

𝑈𝑈𝑈𝑈𝑈𝑈𝑈𝑈 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐 𝑜𝑜𝑜𝑜 𝑤𝑤𝑤𝑤𝑤𝑤𝑤𝑤𝑤𝑤 = 8𝐵𝐵𝐵𝐵𝐵𝐵𝐵𝐵/𝑚𝑚3

8𝐵𝐵𝐵𝐵𝐵𝐵𝐵𝐵
𝑨𝑨𝑨𝑨𝑨𝑨𝑨𝑨𝑨𝑨𝑨𝑨 𝒘𝒘𝒘𝒘𝒘𝒘𝒘𝒘𝒘𝒘 𝒄𝒄𝒄𝒄𝒄𝒄𝒄𝒄 = ∗ 4,802.8 𝑚𝑚3 = 𝟑𝟑𝟖𝟖, 𝟒𝟒𝟒𝟒𝟒𝟒. 𝟒𝟒𝟒𝟒𝟒𝟒𝟒𝟒𝟒𝟒
𝑚𝑚3

𝑻𝑻𝑻𝑻𝑻𝑻𝑻𝑻𝑻𝑻 𝒖𝒖𝒖𝒖𝒖𝒖𝒖𝒖𝒖𝒖𝒖𝒖𝒖𝒖 𝒄𝒄𝒄𝒄𝒄𝒄𝒄𝒄 = 10,859,701.00 + 38,422.4 = 𝟏𝟏𝟏𝟏, 𝟖𝟖𝟖𝟖𝟖𝟖, 𝟏𝟏𝟏𝟏𝟏𝟏. 𝟒𝟒𝟒𝟒𝟒𝟒𝟒𝟒𝟒𝟒

82
  Manpower requirement

Table 5.4: Cost of manpower

No. Position Quantity Monthly salary per head Yearly salary

1 General Manager 1 6,000 72,000
2 Executive Secretary 1 2,000 24,000
3 Production and Technical Manager 1 4,500 54,000
3 Production Head 1 3,500 42,000
4 Secretary 1 1,500 18,000
5 Operators 8 2,500 240,000
6 Electrician 1 2,500 30,000
7 Mechanic 3 2,500 90,000
8 Quality control head 1 3,500 42,000
9 Laboratory technicians 2 2,500 60,000
10 Finance and Administration Head 1 4,500 54,000
11 Sales person 1 2,500 30,000
12 Purchaser 1 2,500 30,000
13 Accountant 1 2,500 30,000
14 Store keeper 1 1,500 18,000
15 Clerk 1 1,000 12,000
16 Personnel & General Service 1 3,000 36,000
17 Cashier 1 2,000 24,000
18 Guards 2 800 19,200
19 Cleaners 2 600 14,400
Total 32 939,600

Table 5.5: Total product cost estimation

Components Percentages Cost (Birr)/year

1.Manufacturing A. Direct production cost
cost
1.Raw material cost 91,346,382.00
2. Auxiliary materials cost 120,000.00
3.Operating labor (OL) 939,600.00
4.Direct supervision 10%OL 93,960.00
5. Utilities 10,898,123.40

83
 6.Maintenance and repair 3% FCI 1,850,113.60
7. Laboratory charges 10%OL 939,600.00
Total Direct production cost 106,187,779.00
B. Fixed charges
1.Depreciation 10%FCI 6,167,045.19
2.Capital charge 1%FCI 616,704.52
3.Insurance 0.5%FCI 308,352.26
Total Fixed charges 7,092,101.97
C. Plant overheads 50%OL 469,800.00
Total manufacturing cost 113,749,680.97
(A+B+C)
2.General expenses 1.Adminstrative cost 3%TPC 3,791,656.03
2.Distribution and selling cost 4%TPC 5,055,541.38
3. R&D 3%TPC 3,791,656.03
Total production cost (TPC) = Manufacturing cost + General 126,388,534.40
expenses
𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇 𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐 (𝑇𝑇𝑇𝑇𝑇𝑇) = 𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀𝑀 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐 + 𝐺𝐺𝐺𝐺𝐺𝐺𝐺𝐺𝐺𝐺𝐺𝐺𝐺𝐺 𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒𝑒

𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇 𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐 (𝑇𝑇𝑇𝑇𝑇𝑇) = 113,749,680.97 + 10%𝑇𝑇𝑇𝑇𝑇𝑇

𝑻𝑻𝑻𝑻𝑻𝑻𝑻𝑻𝑻𝑻 𝒑𝒑𝒑𝒑𝒑𝒑𝒑𝒑𝒑𝒑𝒑𝒑𝒑𝒑𝒑𝒑𝒑𝒑𝒑𝒑 𝒄𝒄𝒄𝒄𝒄𝒄𝒄𝒄 (𝑻𝑻𝑻𝑻𝑻𝑻) = 𝟏𝟏𝟏𝟏𝟏𝟏, 𝟑𝟑𝟑𝟑𝟑𝟑, 𝟓𝟓𝟓𝟓𝟓𝟓. 𝟒𝟒𝟒𝟒 𝒃𝒃𝒃𝒃𝒃𝒃𝒃𝒃/𝒚𝒚𝒚𝒚𝒚𝒚𝒚𝒚

5.4.3. Financial Evaluation and project appraisal

I. Financial analysis
A. Unit product cost determination

𝑈𝑈𝑈𝑈𝑈𝑈𝑈𝑈 𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐 = 𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇 𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐/𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎 𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝

𝑏𝑏𝑏𝑏𝑏𝑏𝑏𝑏
= 126,388,534.40 /3,000,000𝑘𝑘𝑘𝑘/ 𝑦𝑦𝑦𝑦𝑦𝑦𝑦𝑦
𝑦𝑦𝑦𝑦𝑦𝑦𝑦𝑦

𝟒𝟒𝟒𝟒. 𝟏𝟏𝟏𝟏𝟏𝟏𝟏𝟏𝟏𝟏𝟏𝟏
𝑼𝑼𝑼𝑼𝑼𝑼𝑼𝑼 𝒑𝒑𝒑𝒑𝒑𝒑𝒑𝒑𝒑𝒑𝒑𝒑𝒑𝒑 𝒄𝒄𝒄𝒄𝒄𝒄𝒄𝒄 =
𝒌𝒌𝒌𝒌

 Selling price of the debitterized fenugreek flour = unit product cost + 33% profit margin =
42.12 (1+0.33) = 56.00 birr/kg
B. Projected Profit and Loss

𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇 𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎 𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟 = 56 ∗ 3,000,000 𝑏𝑏𝑏𝑏𝑏𝑏𝑏𝑏

84
 𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇 𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎 𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟 = 168,000,000.00 𝑏𝑏𝑏𝑏𝑏𝑏𝑏𝑏

𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇 𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎 𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐 = 126,388,534.40 𝑏𝑏𝑏𝑏𝑏𝑏𝑏𝑏

Table 5.6: Projected profit and loss

Year 1 Year 2 Year 3

Description
(Annually) 75% 85% 100% Average

Total revenue 126,000,000.00 142,800,000.00 168,000,000.00 145,600,000.00

Total product cost 94,791,400.80 107,430,254.20 126,388,534.40 109,536,730.00

Profit before tax 31,208,599.20 35,369,745.80 41,611,465.60 36,063,270.20

Business tax (30%) 9362579.76 10610923.74 12483439.68 10818981.06

Net profit after tax 21,846,019.44 24,758,822.06 29,128,025.92 25,244,289.14

II. Project appraisal

A. The Return on Total Capital Invested (ROI)

𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝑛𝑛𝑛𝑛𝑛𝑛 𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝 (𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴)

𝑅𝑅𝑅𝑅𝑅𝑅 % =
𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇𝑇 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐 𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟

25,244,289.14
𝑅𝑅𝑅𝑅𝑅𝑅 % = × 100 = 35.6%
70,921,019.65

B. Pay-back period analysis

𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼𝐼 𝑖𝑖𝑖𝑖𝑖𝑖𝑖𝑖𝑖𝑖𝑖𝑖𝑖𝑖𝑖𝑖𝑖𝑖𝑖𝑖
𝑃𝑃𝑃𝑃𝑃𝑃 𝑏𝑏𝑏𝑏𝑏𝑏𝑏𝑏 𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝 =
𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝑛𝑛𝑛𝑛𝑛𝑛 𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝 (𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴)

70,921,019.65
𝑃𝑃𝑃𝑃𝑃𝑃 𝑏𝑏𝑏𝑏𝑏𝑏𝑏𝑏 𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝 = = 2.8 𝑦𝑦𝑦𝑦𝑦𝑦𝑦𝑦𝑦𝑦
25,244,289.14

85
 C. Break – even analysis (BEP)

Table 5.7: Data required for break-even analysis calculation

Components Value
Annual Sales 145,600,000.00
Annual Fixed costs:
Factory overhead costs (excluding depreciation) 469,800.00
Administrative overhead Costs 3,791,656.032
Annual factory depreciation 6,167,045.19
Research and development (RD) Costs 3,791,656.032
Marketing cost 5,055,541.376
Total fixed costs 19,275,698.63
Annual Variable Costs:
Raw materials 91,346,382.00
Direct labor 576,000.00
Total annual variable Costs 91,922,382.00

Break – Even sales

𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠 × 𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐
𝐵𝐵𝐵𝐵𝐵𝐵 (𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠) =
𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠 − 𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐
145,600,000 × 19,275,698.63
=
145,600,000 − 91,922,382.00

𝐵𝐵𝐵𝐵𝐵𝐵 (𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝑠𝑠𝑠𝑠𝑙𝑙𝑙𝑙𝑙𝑙) = 52,285,139.04 𝑏𝑏𝑏𝑏𝑏𝑏𝑏𝑏

 BES (Annual Sales) = Birr 52,285,139.04 of sales revenue is required reach the break-
Even point

Break – Even percentage

𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐 × 100
𝐵𝐵𝐵𝐵𝐵𝐵 (%) =
𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠 − 𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐
19,275,698.63 × 100
𝐵𝐵𝐵𝐵𝐵𝐵 (%) =
145,600,000 − 91,922,382.00

𝑩𝑩𝑩𝑩𝑩𝑩 (%) = 𝟑𝟑𝟑𝟑. 𝟗𝟗%

86
  BEP (Percentage) for the project is 35.9%. This indicates that 35.9% of capacity utilization
is enough to reach the break- Even point.
Break – Even quantity

 Amount annually produced ……………….... …..………………3,000,000kg

 Total variable Costs……………………………………… 91,922,382.00 birr
 Unit selling price ……….…………………………………………56.00birr/kg
 Unit variable cost…………………………………..……….……… 9.46 birr

𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐

𝑈𝑈𝑈𝑈𝑈𝑈𝑈𝑈 𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐 =
𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝

91,922,382
=
3,000,000

𝑈𝑈𝑈𝑈𝑈𝑈𝑈𝑈 𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐 = 30.64

𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓𝑓 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐

𝐵𝐵𝐵𝐵𝐵𝐵 (𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞) =
𝑈𝑈𝑈𝑈𝑈𝑈𝑈𝑈 𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠 𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝𝑝 − 𝑈𝑈𝑈𝑈𝑈𝑈𝑈𝑈 𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣𝑣 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐

19,275,698.63
𝐵𝐵𝐵𝐵𝐵𝐵 (𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞) =
56 − 30.64

𝐵𝐵𝐵𝐵𝐵𝐵 (𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞𝑞) = 760,082.75𝑘𝑘𝑘𝑘

 Therefore, 760,082.75kg of debitterized fenugreek powder must be produced to reach the

break- even point.
Based on the preliminary economic and project appraisal analysis, the suggested project has a
return on investment (ROI) of 35.6 %, payback period of 2.8 years and break-even percentage of
35.9%. Thus, the suggested project is financially viable and profitable.

87
 Fixed Cost Total Product cost Total Sales

180000,000.00

160000,000.00

140000,000.00

120000,000.00

100000,000.00

80000,000.00

60000,000.00

52,285,139.04
40000,000.00

20000,000.00

0.00
760,082.75kg
0 500,000 1000,000 1500,000 2000,000 2500,000 3000,000

Figure 5.4: Break-Even analysis for production of debitterized fenugreek powder

88
 Chapter six

5. Conclusion and Recommendation

6.1. Conclusions

The study was conducted with the objective of reducing the phytochemicals and improving
nutritional composition of fenugreek seed varieties by different processing methods such as
germination, fermentation, autoclaving, extraction, and their combinations. The proximate, mineral
and phytochemicals composition, physicochemical and functional properties of raw fenugreek
varieties, and mineral bioavailability, phytate and non phytate contents as affected by processing
methods were also studied. Moreover, during the extraction process, the effects of rinsing stages,
solvent type and solvent to water ratio were also examined.

The results obtained showed that fenugreek is rich in nutrients, especially protein (about 30%), and
mineral contents. The result on crude protein content for fenugreek seeds suggests their high
potential as a cheap source of alternative protein for human consumption. It can also be used as
supplement to enhance the low nitrogen content of traditional staples of cereals and tubers. The
water absorption (about 10g of water/1g of fenugreek flour) and oil absorption (2g of water/1g of
fenugreek flour) capacities were also greater than those of other legumes, which makes fenugreek
an important viscosity builder, emulsifier and thickening agent.

In the current study, processing methods increased protein, fat, fiber, iron, zinc, calcium and
phosphorous contents of fenugreek. Bioavailability of minerals and non phytate phosphorus also
improved during processing.

Fermentation was effective in reducing the heat-stable phytochemicals, phytate and tannins, while
autoclaving was effective for saponins. Extraction has eliminated saponins and tannins, whereas it
did not bring in significant effects on phytates. Combinations of processing methods were better
than single processes. Combination fermentation and extraction brought a complete removal of
phytates, tannins and saponins. However, combination of autoclaving and fermentation was less
effective in reducing of phytates as compared to the individual processes, autoclaving and
fermentation.

Variation of extraction rates with variety, solvent type, and solvent to water proportion were
observed. Methanol extraction was faster than ethanol and extraction rate decrease with increase of
water proportion. Extraction rate of Ada’a variety was slower than Challa variety due to higher
water absorption capacity of Ada’a.

89
The bitterness and astringent smell of fenugreek flour were removed by polar solvent extraction
and debitterized fenugreek powder was produced. In addition, to preserve the health benefits and to
maintain dose of fenugreek required for different age groups, sex and health conditions, fenugreek
capsules were produced.

Generally, the fenugreek varieties studied were rich in nutrients, micronutrients, bioactive
components and so that they can be used intensively in the food, pharmaceutical manufacturing,
and cosmetics industries.

6.2. Recommendations

Based on the gaps identified in this research work, the following recommendations are made for the
future researchers to bring integrated and valuable knowledge contributions to the country.

 It is evident that both adverse and health benefits may be attributed to phytochemicals in
foods. The physiological effects of phytochemicals are related to their level of intake and
the conditions they are taken such as presence of other dietary constituents, age, nutritional
and health status of individuals. Therefore, dose-response studies on fenugreek that will
produce health benefits without causing any adverse effect should be conducted in the
future.
 Synergistic effects of phytochemicals and interaction of phytochemicals with other
components of the diet and the mechanism of their action need to be further addressed if we
are to understand the role of phytochemicals in health and disease.
 Extraction parameters such as granular size, temperature and others shall be optimized in
detail.
 Studies on antimicrobial, antidiabetic, antioxidant, gastroprotective, preservative and
hypocholesterolemic effects of fenugreek varieties grown in Ethiopia are required.
 Researchers also shall focus on utilization and product development of fenugreek such as
health drinks, food fortification, supplementation with wheat and cereals and functional
foods.
 Fenugreek is versatile crop so that researches on its application for different foods,
pharmaceutical, and cosmetics industries are required.
 The byproduct of extraction which contains extracted bioactive components and water can
be used to produce high end products such as galactomnnans, gums, emulsifiers,
sapogennins, and biopest management chemicals.

90
References
Abdel-Nabey, A.A., and Damir, A.A., (1990). Changes in some nutrients of fenugreek (Trigonella
Foenum graecum L.) seeds during water boiling. Plant Foods for Human Nutrition 40: 267-
274.
Ahmadiani, A., Javan., M., Semnanian, S., Bharat, E. and Kamalinejad, M. (2001).
Antiinflammatory and antipyretic effects of Trigonella foenum-graecum leaves extract in
the rat. Journal of ethnopharmacol. 75(2-3):283-286.
Ahmed Rafik EL-Mahdy (1982). Changes in phytate and minerals during germination and cooking
of fenugreek seeds. Food Chemistry 9 (1982) 149-158
Al-Habori, M., and Raman, A. (2002). Pharmacological Properties in Fenugreek - The genus
Trigonella (1st edition) by G.A. Petropoulos (ed.), Taylor and Francis, London and New
York, 10: 163- 182.
Al-Khalidi, S.F., Martin, S.A. and Prakash, L. (2001). Fermentation of fenugreek fribers, psyllium
husk and wheat bran by Bacteroides ovatus V975. Current Microbiology. 39(4): 231-232.
Amankwah, E.A., Barimah J., Acheampong R., Addai L.O. and Nnaji C.O., (2009). Effect of
fermentation and malting on the viscosity of maize-soyabean weaning blends. Pakistan
journal of nutrition, 8, 1671-1675.
Ambasta, S.P., (Ed.) (2000). Useful Plants of India; National Institute of Science Communication:
New Delhi.
Anderson, R.L., and Wolf, W.J. (1995). Compositional changes in trypsin inhibitors, phytic acid,
saponins and isoflavones related to soybean processing. Journal of Nutrition, 125: 581S–
588S.
Anonymous (1994). Plants and Their Constituents. Phytochemical Dictionary of the Leguminosae,
Vol.1, Cherman and Hall, London.
Anonymous (1996). Common catalogue of varieties of agricultural plant species. Official Journal
of European Communities, 39, C 272 A, 45.
AOAC (2000). Food composition; additives; natural contaminants. In: Official Methods of
Analysis of AOAC International, Vol. II, 17th (edited by H. William). Pp. 12–42.
Washington, DC: Association of Official Analytical Chemists (AOAC). Official method
942.15, 982.14.
Appiah F., Asibuo J. Y. and Kumah P. (2011). Physicochemical and functional properties of bean
flours of three cowpea (Vigna unguiculata L. Walp) varieties in Ghana. African Journal of
Food Science Vol. 5(2), pp. 100 - 104
Armitage, D.B., Hettiarachchy, N.S. and Monsoor, M.A. (2002). Natural antioxidants as a
component of an egg-albumen film in the reduction of lipid oxidation in cooked and
uncooked poultry. Journal of Food Science, 67(2): 631-634.
Aspinall, G.O. (1980). The biochemistry of plants. Academic publisher, New York. 3:473- 500.
Atwal, A., Eskin, N., McDonald, B. and Vaisey-Genser, M. (1980). The effects of phytase on
nitrogen utilization and zinc metabolism in young rats. Nutrition Reports International. 21,
257-67,
Bakr, A.A. (1997). Production of iron-fortified bread employing some selected natural iron sources.
Nahrung. 41(5):293-298.

91
Balyan, D.K., Tyagi, S.M., Dheer-Singh and Tanwar, V.K. (2001). Effects of extraction parameters
on the properties of fenugreek mucilage and its use in ice cream as stabilizer. Journal of
Food Science and Technology. 38(2):171-174.
Barampama, Z., and Simard, R.E. (1993). Nutrient composition, protein quality and antinutritional
factors of some varieties of dry beans (Phaseolus vulgaris, L.) grown in Burundi. Food
Chemistry, 47, 159–167.
Beuchart LR (1977). Functional and Electrophoretic Characteristics of Succinylated Peanut Flour
Protein. Journal of Agriculture and Food Chemistry, 25: 258- 261.
Bhatia, A. and Khetrapaul, N. (2002). Effect of fermentation on phytic acid and in vitro availability
of calcium and iron of 'doli-ki roti'-an indigenously fermented Indian bread. Ecological
Food Nutrition (41):243-253.
Bhatti, M.A., Khan, M.T.J., Ahmed, B., Jamshaid, M. and Ammad, W. (1996). Antibacterial
activity of Trigonella foenum-graecum seeds. Fitopterapia. LXVII:372-374.
Billaud, C. and Adrian, J. (2001). Fenugreek: composition, nutritional value and physiological
properties. Science Alim. 21(l):3-26.
Birch, G. G., Spencer, M., and Cameron, A. G.(1980). Food science (2nd ed., pp. 29–37). Oxford,
UK: Pergamon Press Ltd.
Birk, Y. and Peri, I. (1980). In Toxic Constituents of Plant Foodstuffs, ed. I. E. Liener. Academic
Press, New York, USA, pp. 161-82.
Bishnoi, S., & Khetarpaul, N. (1993). Variability in physico-chemical properties and nutrient
composition of different pea cultivars. Food Chemistry, 47, 371–373.
Bombardelli, E., and Gabetta, B. (2001). Soya extract, process for its preparation and
pharmaceutical composition. US Patent 6,280,777
Broca, C., Breil, V., Cruciani-Guglielmacci, C., Manteghetti, M., Rouault, C., Derouet, M.,
Rizkalla, S., Pau, B., Petit, P., Ribes, G., Ktorza, A., Gross, R., Reach, G. and Taouis, M.
(2004). The insulinotrophic agent 1D1101(4-hydroxyisoleucine) activates insulin signaling
in rat. American Journal of Physioogyl-Endocrinology and Metabolis., 287(3): E463-
E471.
Bunting, E.S. (1972). Cultivation of Fenugreek and Some existing its varieties, Feed Laboratory,
Oxford.
Butler, L. G. (1989). New perspectives on the antinutritive effects of tannins. In Food Proteins, ed.
Kinsella, J. E. & Soucie W. G. AOCS, Champaign, IL, USA, pp. 402-9.
Butter, L.D., Price, M.L. and Brotheton, J.E. (1982). Vanillin assay for proanthocaynidins
(Condensed Tannins): Modification of the solvent for estimation of the degree of
polymerization. Journal of agricultural food chemistry 30:1087-1089.
Carnovale, E., Lugaro, E. and Lombardi-Boccia, G. (1988). Phytic acid in fava beans and peas:
effect on protein availability. Cereal Chemistry, 65, 114-17.
Chantry, Caroline J., Howard, Cynthia R., Montgomery, Anne, Wight and Nancy (2004). Use of
galactogogues in initiating or augmenting maternal milk supply.
Chatterjee, A., and Prakashi, S.C. (1995). Treatise on Indian Medicinal Plants, Vol. 2; Council of
Scientific and Industrial Research: New Delhi.

92
Chau, C. F., and Cheung, P. C. K. (1997). Functional properties of flours prepared from three
Chinese indigenous legumes seed. Journal of Food Chemistry, 66, 429–433.
Cheeke, P. R. (1971). Nutritional and physiological implication of saponin. A review. Candian
Journal of Animal Science, 51, 621.
Choi, Y. and Okos, M. R. (1986). Effects of temperature and composition on the thermal properties
of foods, in Food Engineering and Process Applications, Vol. 1,Transport Phenomena,
Maguer, M. Le and Jelen, P., Eds., Elsevier, London, 93–101.
Coustantina Tzia and George Liadakis (2003). Extraction Optimization in food engineering.
National Technical University of Athens, Greece.
CSA (2009/10). Report on Crop and livestock product utilization (private peasant holdings, meher
season). P.17-27. The Federal Democratic Republic of Ethiopia Central statistical. Agency
Agricultural sample survey, volume VII.
Dahanukar, S.A., Kulkarni, R.A. and Rege, N.N. (2000). Pharmacology of medicinal plants and
natural products. Indian Journal of Pharmacology. 32:S81-S118.
Del’ Gaudio, S. (1952). II fieno greco, foraggera del colle et del monte. Italian Agriculture, 89,
127–36.
Deshpande, S. Sathe, S. K., and Salunkhe, D. K. (1984). Chemistry and safety of plant polyphenols.
In Nutritional and Toxicological Aspects of Food Safety, ed. M. Friedman. Plenum Press,
New York, USA, pp. 457-95.
Devasena, T. and Menon, V.P. (2003). Fenugreek affects the activity of beta-glucuronidase and
mucinase in the colon. Phytotherapy Research. 17(9): 1088-1091.
Duhan A., Khetarpaul N., and Bishnoi S. (2002). Changes in phytates and HCl extractability of
calcium, phosphorus, and iron of soaked, dehulled, cooked, and sprouted pigeon pea
cultivar (UPAS-120). Plant Foods for Human Nutrition 57: 275–284.
Duke, A.J. (1986). Handbook of Legumes of World Economic Importance, Plemus Press, New
York and London.
E1-Shimi, MM., Damir, AA. And Ragab, M. (1989). Changes in some nutrients of fenugreek seeds
during germination. Food Chemistry, 14:11-19
Edison, S. (1995). Spices-Research support to productivity. In N. Ravi (ed.) The Hindu Survey of
Indian Agriculture, Kasturi and Sons Ltd., National Press, Madras. Pp.101-105.
EHNRI, (1997). Ethiopian Health and Nutrition Research Institute: Food Composition Table for
use in Ethiopia Part III and IV , 4-5d
Ethiopia: Country Report. (1996). FAO Int. Tech. Conf. on Plant Genet. Res., Leipzig. Plant
Genetic Resource Center, Addis Ababa.
Evans, W.J. and Martin, C.J. (1988). Interaction of Mg(II), Co(II), Ni(II) and Zn(II) with phytic
acid, VIII. A calorimetric study. Journal of Inorganic Biochemistry, 32, 259 – 67.
Fazli, F. R. Y. (1967). Studies in the steroid yielding plant of the genus Trigonella PhD diss.,
University of Nottingham, UK.
Fazli, F.R.Y. and Hardman, R. (1968). The spice fenugreek (Trigonella foenum-graecum L.). Its
commercial varieties of seed as a source of diosgenin. Tropical Science, 10:66-78.

93
Galal, O.M. (2001). The nutrition transition in Egypt: obesity, undernutrition and the food
consumption context. Public Health and Nutrition 5(1A):141-148.
Gaman, P.M., and Sherrington, K.B. (1986). The science of food. An Introduction to Food Science,
Nutrition and Microbiology. 2nd Edition. Pergamon Press. pp.139-177.
Gatri, N., Madar, Z., Aserin, A., and Sternheim, B. (1997). Fenugreek galactomanans as food
emulsifiers. Lebensmittel-Wissenschafft and Technolog, 30(3):305-311.
Ghafgazi, T., Farid, H. and Pourafkari, A. (1980). In vitro study of the action of Trigonella foenum-
graecum grown in Iran. Iranian. Journal of Public Health. 9:21-26.
Glicksman, M. (1982). Comparative properties of hydrocolloids: Functional properties. In:
Glicksman, M. (Ed), Food Hydrocolloids, Vol. 1. Florida: CRC Press, pp. 47–59.
Greiner, R. and Konietzny, U.(2006). Phytase for food application. Food technology and
biotechnology 44(2),125-140.
Güçlü-Üstündağ, Özlem and Mazza, Giuseppe (2007). Saponins: Properties, Applications and
Processing', Critical Reviews in Food Science and Nutrition, 47: 3, 231 — 258
Gupta, U. Rudrama, Rati, E.R. and Joseph, R. (1998). The nutritional quality of lactic fermented
bitter gourd and fenugreek leaves. International Journal of Food Science and Nutrition
49(2): 101- 108.
Hannan, J.M., Rokeya, B., Faruque, O., Nahar, N., Mosihuzzaman, M., Azad Khan, A.K, and Ali,
L. (2003). Effect of soluble dietary fibre fraction of Trigonella foenum-graecum on
glycemic, insulenimic, lipidemic and platelet aggregation status of Type 2 diabetic model
rats. Journal of Ethnopharmacology, 88: 73-77.
Haslam, E. (1966). Chemistry of vegetable tannins. New York, NY, USA: Academic Press.
Heath, A.W., Nyan, O., Richards, C.E. and Playfair, J.H. (1991). Effects of interferon gamma and
saponin on lymphocyte traffic are inversely related to adjuvaticity and enhancement of
MHCII expression. Int. Immunol., 3, 285 - 92
Helen W/Michael (2011). Influence of processing on antinutrients and production of some value
added finger millet based products. Msc. Thesis. Department of chemical enginerring,
Addis Ababa University.
Hidvegi, M., El-Kady, A., Lòsztity, R., Bákás, F. and Simon-Sarkadi, L. (1984). Contribution to the
nutritional characterization of fenugreek (Trigonella foenum-graecum L.). Acta
Alimentaria, 13(4), 315–24.
Hooda, S. and Jood, S. (2003a). Effect of soaking and germination on nutrient and antinutrient
contents of fenugreek (Trigonella foenum-graecum L.). Journal of Food Biochemistry
27(2): 165- 176.
Hooda, S. and Jood, S. (2003b). Physicochemical, rheological, and organoleptic characteristics of
wheat-fenugreek supplemented blends. Nahrung, 47(4):265-268.
Hostettmann, K., and Marston, A. (1995). Saponins. Cambridge University Press, Cambridge, New
York.
Huang, Z., Haig, T., Wu, H., An, M. & Pratley, J. (2003). Correlation between phytotoxicity on
annual ryegrass (Lolium rigidum) and production dynamics of allelochemicals within root
exudates of an allelopathic wheat. Journal of Chemical Ecology, 29, 2263–2279.
IBC (2008). Ethiopia: Second Country Report on the State of PGRFA to FAO, January, 2008, pp.9

94
Ibrahim S. S., Habiba R. A., Shatta A. A. and Embaby H. E. (2002). Effect of soaking,
germination, cooking and fermentation on antinutritional factors in cowpeas. Nahrung, 46 ,
No. 2, pp. 92 - 95.
Igile GO (1996). Phytochemical and Biological studies on some constituents of Vernonia
amygdalina (compositae) leaves. Ph.D thesis, Department of Biochemistry, University of
Ibadan, Nigeria.
Ismail, I. A. (1996). Changes in some nutrients of fenugreek seeds after boiling. Nutrition Institute,
Egypt. 16(l):78-87.
Javan. M , Ahmadiani, A., Semnanian, S. and Kamalinejad, M. (1997). Antinociceptive effects of
Trigonella foenum-graecum leaves extract. Journal of Ethnopharmacology 58(2): 125-129.
Kamal, R., Yadav, R. and Sharma, J.D. (1993). Efficacy of the steroidal fraction of the fenugreek
seed extract on the fertility of male albino rats. Phytotherapy Research, 7:134- 138.
Kapadin, D.J., Subba Rao, G. and Morton, J.F. (1983). Herbal tea consumption and esophageal
cancer. In carcinogens and mutagens in the environmental (Vol 3 – Naturally occurring
compounds: Epidemiology and distribution), ed.H.F. Stich. CRC Press, Boca Raton, FL,
USA, Pp. 3 – 12.
Khokhar S, Chauhan BM, (1997). Antinutritional factors in Moth Bean (Vigna aconitifolia):
Varietal Differences and Effects of Methods of Domestic Processing and Cooking. Food
Chemistry Vol. 59(3): 367-371.
Kinsella, J. E. (1979). Functional properties of soy protein. Journal of American Oil Chemists
Society, 56, 242–249.
Kitagawa, I. (1986). Method of isolating soyasaponins. US Patent 4,594,412.
Lazzerini M. and Ronfani L. (2008). Oral zinc for treating diarrhea in children (review). p. 3.
Loewus FA. Biosynthesis of phytate in food grains and seeds (2002). In: Reddy NR, Sathe SK
(Eds.). Food Phytates. CRC Press, Boca Raton Florida ; 53–61.
Lust, J.B. 1986. The herb book. Bantam Books Inc. New York. Pp. 1-55.
Manniche, L. (1989). An Ancient Egyptian Herbal, British Museum Publ. Ltd., London.
McAnuff, M.A., Omoruyi, F.O., Morrison, E.Y.S.A., and Asemota, H.N. (2002). Plasma and liver
lipid distributions in streptozotocin-induced rats fed sapogenin extract of the Jamaican
bitter yam (Dioscorea polygonoides). Nutrition Research, 22:1427-1434.
Meseret Bekele (2011). Effect of fermentation on Quality Protein Maize-soybean blends for the
production of weaning food. Msc Thesis. Department of chemical enginerring, Addis
Ababa University.
Messina, M. & Barnes, S. (1991). The role of soy products in reducing risk of cancer, Journal of
National Cancer Institute, 83, 541-6.
Mubarak, A.E., (2004). Nutritional composition and antinutritional factors of mung bean seeds
(Phaseolus aureus) as affected by some home traditional processes. Food Chemistry 89
(2005): 489–495
Mugocha, P.T. (2001). Fermentation of a finger millet- dairy composite gruel.
Nazar A. El Nasri , A.H. El Tinay (2006). Functional properties of fenugreek (Trigonella foenum
graecum) protein concentrate. Journal of Food Chemistry 103 (2007) 582–589

95
Neukom, H. (1989). Galactomannan: Properties and applications. Lebensmittel-Wissenschaft und-
Technologie, 22, 41–45.
Nuha, M. O., Isam, A. M. A. and Elfadil, E. B. (2010). Chemical composition, antinutrients and
extractable minerals of Sicklepod (Cassia obtusifolia) leaves as influenced by fermentation
and cooking. International Food Research Journal 17: 775-785
Oakenful, D. & Sidhu, G. S. (1990). Could saponins be a useful treatment for
hypercholesterolemia? European Journal of clinical nutrition, 44, 79-88.
Ogunsua AO (1980). Changes in some chemical constituents during fermentation of cassava tubers
(Manihot esculenta (Crantz). Food Chemistry 5: 249-255
Okwu D. E. and Josiah C. (2006). Evaluation of the chemical composition of two Nigerian
medicinal plants. African Journal of Biotechnology Vol. 5 (4), pp. 357-361
Okwu, D.E., (2004). Phytochemicals and vitamins content of indigenous spices of South Easter
Nigeria. Journal of sustain Agriculture and Environment, 6: 30-34.
Osborne, D.R., and Voogt, P. (1978). The Analysis of Nutrients in Food.
Osman, A. (2004). Changes in sorghum enzyme inhibitors, phytic acid, tannins and in vitro protein
digestibility occurring during Khamir (local bread) fermentation. Food Chemistry 88
(2004) 129–134
Osuntokun BO (1981). Cassava diet chronic cyanide intoxication and neuropathy in Nigerian
Africans. World Review of Nutrition & Dietetics, 36: 141-173.
Pathak, P., Srivastava, S. and Grover, S. (2000). Development of food products based on millets,
legumes and fenugreek seeds and their suitability in diabetic diet. International Journal of
Food Science and Nutrition 51(5):409-414.
Petropoulos G. A. (2002). Fenugreek. The genus Trigonella. Taylor & Francis, London and New
York, p. 2-35.
Raghuram, T.C., Sharma, R.D., and Sivakumar, B. (1994). Effect of fenugreek seeds on
intravenous glucose disposition in non-insulin dependent diabetic patients. Phytother. Res.
8:83-86.
Raskin, I., Ribnicky, D.M., Komarnytsky, S., Llic, N., Poulev, A., Borisjuk, N., Brinker, A.,
Moreno, D.A., Ripoll, C, Yakoby, N., O'Neal, J.M., Cornwell, T., Pastor, I. and Fridlender,
B. (2002). Plants and human health in twenty-first century. Trends Biotechnology
20(12):522-531.
Reddy NR. (2002). Occurrence, distribution, content, and dietary intake of phytate. In: Reddy NR,
Sathe SK (Eds.). Food Phytates. CRC Press, Boca Raton Florida; 25–51.
Reddy, N. R., Sathe, S. K. & Salunkhe, D. K. (1982). Phytates in legumes and cereals. Advanced
Food Research, 28, l-92.
Reddy, N.R. & Pierson, M.D. (1994). Reduction in antinutritional and toxic components in plant
foods by fermentation. Food Research International, 27, 281–290.
Ridout, C. L., Price, K. R., DuPont, M. S., Parker, M. L., and Fenwick, G. R. (1991). Quinoa
saponins-Analysis and preliminary investigations into the effects of reduction by
processing. Journal of Food Science and Agriculture, 54:165–176.

96
Rouk, H.F., and Mangesha, H. (1963). Fenugreek (Trigonella foenum-graecum L.). Its relationship,
geography and economic importance. Exp. Stat. Bull. No. 20. Imperial Ethiopian College
of Agriculture and Mechanical Arts.
Saikat K. Basu (2006). Seed production technology for fenugreek (Trigonellafoenum-graecum L.)
in the Canadian Prairies. Department of Biological Sciences University of Lethbridge
Lethbridge, Alberta, Canada.
Saleh, N., El-Hawary, Z., El-Shobaki, F.A., Abbassy, M. and Morcos S.R. (1977). Vitamin content
of fruits and vegetables in common use in Egypt. Zeitschrift für Ernõhrungswiss, 16(3),
158–62.
Salunkhe, D. K., Chavan, J. K. & Kadam, S. S. (1990). Dietary Tannins: Consequences and
Remedies. CRC Press, Boca Raton, FL, USA.
Sandberg, A., Brune,M., Carlsson , N., Hallberg, L., Skoglund, E. and Rossander-Hulthen,
L.(1999). Inositol phosphates with different numbers of phytate groups influence iron
absorption in humans. America journal of clinical nutrition 70:240-246.
Sauvaire, Y., Baccou, J.C. and Besancon, P. (1976). Nutritional value of the properties of the
fenugreek (Trigonella foenum-graecum L.). Nutrition Reports International, 14(5):527-
537.
Scott, M. T., Gross-Sampson, M. & Bomford, R. (1985). Adjuvant activity of saponin: antigen
localization studies. Znt. Arch. Allergy. Applied Immunology, 77, 409-12.
Seghal, G., Chauhan, G.S. and Kumbhar, B.K. (2002). Physical and functional properties of
mucilage from yellow mustard (Sinapis alba L.) and different varieties of fenugreek
(Trigonella foenum-graecum L.) seeds. Journal of Food Science and Technology
39(4):367-370.
Sharma, H.R. and Chauhan, G.S. (2000). Physico-chemical and rheological quality characteristics
of fenugreek (Trigonella foenum-graecum L.) supplemented wheat flour. Journal of Food
Science and Technology 37(l):91-94.
Sharma, H.R. and Chauhan, G.S. (2002). Effects of stabilized rice bran-fenugreek blends on the
quality of breads and cookies. Journal of Food Science and Technology 39(3):225-233.
Sharma, R. D. (1984). Hypocholesterolemic effect of hydroxyl acid components of Bengal gram.
Nutrition Reports International, 29, 13, 15, 22.
Sharma, R.D. (1986). Effects of fenugrek seeds and leaves on blood glucose and serum insulin
response in human subjects. Nutrition Research, 6, 1353-1363.
Sharma, R.D., A. Sarkar and D.K. Hazra (1996). Hypolipidaemic effect of fenugreek seeds: a
chronic study in non-insulin dependent diabetic patient. Phytotherapy Research, 10: 332-4.
Shashi Kala (1997). Management of non insulin dependent diabetes mellitus by using traditional
medicinal plant products. Ph.D. Thesis. Haryana Agricultural University, Hisar, India.
Shimelis Admassu & Sudip Kumar Rakshit (2008). Influence of natural and controlled
fermentations on α-galactosides, antinutrients and protein digestibility of beans (Phaseolus
vulgaris L.). International Journal of Food Science and Technology 43, 658–665.
Shimelis Admassu, Sudip Kumar Rakshit (2005). Proximate composition and physico-chemical
properties of improved dry bean (Phaseolus vulgaris L.) varieties grown in Ethiopia.
Science direct 103, 331–338.

97
Shimelis Admassu, Sudip Kumar Rakshit (2007). Effect of processing on antinutrients and in vitro
protein digestibility of kidney bean (Phaseolus vulgaris L.) varieties grown in East Africa.
Food Chemistry 103, 161–172.
Shimelis Tizazu (2009). Improvement of energy and nutrients density of sorghum based weaning
food using germination. Msc Thesis. Department of Food Science and Nutrition, Addis
Ababa University.
Sidhu, G. S. & Oakenful, D. G. (1986). A mechanism for the hypocholesterolemic activity of
saponins. British Journal of Nutrition 55, 643-9.
Slinkard AE (2006). University of Saskatchewan (www.agr.gov.sk.ca).
Smith, A. (1982). Selected Markets for Turmeric, Coriander, Cumin and Fenugreek Seed and Curry
Powder. Pub. No. G165 .Tropical Product Institute, London.
Soetan K. O. and Oyewole O. E. (2009). The need for adequate processing to reduce the
antinutritional factors in plants used as human foods and animal feeds: A review. African
Journal of Food Science Vol. 3 (9), pp. 223-232.
Srichamroen, A., Ooraikul, B., Vasanthan, T., Chang, P., Acharya, S. and Basu, T. (2005).
Compositional differences among five fenugreek experimental lines and the effect of seed
fractionation on galactomannans extractability of a selected line. International Journal of
Food Science and Technology (in press).
Srinivasan K. (2006). ‘Fenugreek (Trigonella foenum-graecum): A Review of Health Beneficial
Physiological Effects', Food Reviews International, 22: 2, 203-224.
Stich, H. F. & Rosin, M. P. (1984). Naturally occurring phenolics as antimutagenic and
anticarcinogenic agents. In Nutritional and Toxicological Aspects of Food Safety, ed. M.
Friedman. Plenum Press, New York, USA. pp. l-29.
Sumati, R.M., and Rajagopal, M.V. (1996). Fundamentals of foods and nutrition. 3rd edition. New
Delhi, India.
Tahiliani, P. and Kar, A. (2003a). The combined effects of Trigonella and Allium extracts in the
regulation of hyperthyroidism in rats. Phytomedicine, 10(8): 665-668.
Tahiliani, P. and Kar, A. (2003b). Mitigation of thyroxineinduced hyperglycaemia by two plant
extracts. Phytotherapy Research, 17(3): 294-296.
Talelis, D. (1967). Cultivation of Legumes, Agricultural College of Athens, Athens (Greek).
Taranalli, A.D. and Kuppast, I.J. (1996). Study of wound healing activity of seeds of Trigonella
foenum-graecum in rats. Indian Journal of Pharmaceutical Sciences, 58(3): 117-119.
Tayyaba, Z., S.N. Hasnain and S.K. Hasan (2001). Evaluation of the oral hypoglycemic effects of
Trigonella foenum graecum L. (Methi) in normal mice. Journal of Ethnopharmacology, 75:
191-5.
Thirunavukkarasu, V., Anuradha, C.V. and Viswanathan, P. (2003). Protective effect of fenugreek
(Trigonella foenum-graecum) seeds in experimental ethanol toxicity. Phytotherapy
Research, 17(7): 737-743.
Tiran, D. 2003. The use of fenugreek for breast feeding women. Complement Ther Nurs
Midwifery, 9(3): 155-156.

98
Tizazu, S., Urga, K., Abuye, C. and Retta, N. (2010). Improvement of energy and nutrient density
of sorghumbasedcomplementary foods using germination. African journal of food
agriculture nutrition and development 10(8), 2928-2942.
Udayasekhera, P. and Sharma, R.D. (1987). An evaluation of protein quality of fenugreek seeds
and their supplementary effect. Food Chemistry 24, 1-5.
UNICEF. The State of World’s Children, 2009, pp. 5 and 54.
United States Department of Agriculture (USDA) (2001). Nutrient database for standard reference.
Release 14. Washington DC.
Vaintraub, IA. and Lapteva, N.A.(1988). Colorimetric determination of phytate in unpurified
extracts of seeds and products of their processing. Annual biochemistry 175: 227- 230.
Valette, G., Y. Sauvaire, J.C. Baccou, & C. Ribes (1984). Hypocholesterolemic effect of fenugreek
seeds in dogs. Atherosclerosis, 50: 105-1 11.
Vasil'chenko, I.T. (1953). Bericht uber die arten der Gattung. Trigonella Trudy Bot. Inst. Akad.
Nauk SSSR.L10.
Vavilov, N.I. (1951). The origin, variation, immunity and breeding of cultivated plants. Chronica
Botanica. 1:6.
Vijayakumari, K., Siddnuraju, K., & Janardhanan, K. (1996). Effect of soaking, cooking and
autoclaving on phytic acid and oligosaccharide contents of the tribal pulse, Mucuna
monosperma, Dc.ex.Wight. Food Chemistry 55, 173–177.
Vucenik I, Shamsuddin AM. (2003). Cancer inhibition by inositol hexaphosphate (IP6) and
inositol: From laboratory to clinic. Journal of Nutrition 133: 3778S–3784S.
West, L. G. & Greger, J. L. (1978). In-vitro studies of saponin-vitamin complexation. Journal of
Food Science 43, 1340-5.
Yadav, S.K. and Sehgal, S. (1997). Effect of home processing and storage on ascorbic acid and
beta-carotene content of bathua (Cheniopodium album) and fenugreek (Trigonella foenum-
graecum). Plant Food. Human Nutrition, 50(3):239-247.
Yoon, J. H., Thompson, L. U. & Jenkins, D. J. A. (1983). The effect of phytic acid on in-vitro rate
of starch digestibility and blood glucose response. American Journal of Clinical. Nutrition
38, 83542.
Zhou, J.-R., and Erdman, J.W.Jr. (1997). Chemical effects of processing and food preparation on
carotenoids and soy and garlic phytochemicals. In: Lachance P.A., Eds., Nutraceuticals:
Designer Foods III Garlic, Soy and Licorice. Food & Nutrition Press, Connecticut, pp. 23–
37.

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Appendices

Appendix I. Photos of finger millet processing and laboratory equipments used

Fenugreek plant Challa Ada’a Wello

Capsules

Debitterized fenugreek Debitterized fenugreek Extract (byproduct)

powder (Challa) powder (Ada’a)

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 Extraction of fenugreek flour Percolator Distillation column

Temperature controlled Tray drier Moisture analyzer

Incubator

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Appendix II. Sensory evaluation score card using nine point hedonic scale

Panelist code & name: _____________ sample code: _____________ date: _______

Sensory perception Sensory quality attributes Overall acceptability

(score) Appearance Aroma Taste hedonic scale

(Color)

1=dislike extremely 1=Extremely unacceptable

2=dislike very much 2=very much unacceptable

3=dislike moderately 3=moderately unacceptable

4=dislike slightly 4=Slightly unacceptable

5=neither like 5=neither acceptable nor

nor dislike Unacceptable

6=like slightly 6=Slightly acceptable

7=like moderately 7=moderately acceptable

8=like very much 8=highly acceptable

9=like extremely 9=Extremely acceptable

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