ABSTRACT
Fruit juices especially unpasteurized ones are well recognized by consumers for their fresh flavor, vitamin content and nutritive values. This study investigated a five-day microbial fermentation of the pulp of Theobroma cacao. The extracted juice samples were analyzed for total viable bacterial count (TVBC), total lactic acid bacterial count and total fungal (yeast and mould) count using the spread plating method with appropriate media. All isolates were characterized following standard methods. There was a microbial succession of a wide range of microorganisms including yeasts, lactic-acid, and acetic-acid bacteria during the fermentation period. Over-fermentation led to a rise in Bacilli and fungi that caused off-flavors in the samples. Results show that the mean viable bacterial count and lactic acid bacterial count of the juice samples ranged from 1.29 x 108cfu/ml to 2.41 x 108cfu/ml and 7.74 x 107cfu/ml to 1.04 x 108cfu/ml respectively. The microbial load decreased gradually during the study period. The pH recorded during the fermentation period ranged from 3.2 to 3.7. The pH decreased as the fermentation proceeded affirming the production of acidic products. The bacterial isolates were identified as Staphylococcus aureus, Bacillus spp., Acetobacter spp., and Lactobacillus spp. while the identities of the fungal isolates were Saccharomyces cerevisiae.
TABLE OF CONTENTS
Title
Page i
Certification
ii
Dedication
iii
Acknowledgements
iv
Table of
Contents v
Lists of
Tables vii
List of
Figures viii
Abstract ix
CHAPTER ONE
1.0 Introduction
1
1.1 Cocoa
Pulp 1
1.2 Aim
and Objectives 5
CHAPTER TWO: LITERATURE REVIEW
2.1 Background
of Cocoa Pulp 6
2.2 Fermentation
6
2.2.1 Phases
of cocoa fermentation 7
2.3 Microorganisms
involved in Fermentation of Cocoa Pulp 9
2.3.1 Yeasts 9
2.3.2 Lactic
acid bacteria 11
2.3.3 Acetic
acid bacteria 12
CHAPTER THREE
3.0 Materials
and Methods 15
3.2 Media
Preparation 15
3.3 Microbiological
Analysis 15
3.3.1 Sample
preparation 15
3.3.2 Inoculation 15
3.3.3 Isolation
and Characterization 16
3.4 Gram
Staining 16
3.5 Biochemical
tests 17
3.6 Determination
of pH Value 19
3.7 Identification
of Microbial Isolates 19
3.7.1 Bacterial
Isolates 19
3.7.2 Yeast
Isolates 19
CHAPTER FOUR:
Results 20
CHAPTER FIVE
5.0 Discussion 25
5.1 Conclusion 26
5.2 Recommendation
26
References 28
LIST OF TABLES
Table Title Page
1 Yeasts
isolated from Cocoa Fermentation in Four Countries 11
2 Lactic-acid
Bacteria isolated from Cocoa Fermentation in Four Countries 12
3 Acetic-acid
Bacteria isolated from Cocoa Fermentation in Four Countries 13
4 Total
viable count of Bacterial and fungi load obtained 21
5 Biochemical
characteristics of the isolates 22
6 pH
of the samples during the period of fermentation 23
7 Microorganisms isolated from the studied
samples 24
LIST OF FIGURES
Figure Title Page
1 Extraction
and fermentation of cocoa pulp juice 4
2 Microbial
activities during cocoa fermentation 14
CHAPTER ONE
1.0 Introduction
Theobroma
cacao belongs to the family Sterculiaceae and is
economically important due to its valuable seeds. The seeds, commonly known as
cocoa beans are the principal raw material for chocolate production. Cocoa
plants are cultivated on plantations in tropical regions throughout the world;
Ivory Coast, Ghana, Nigeria, Cameroon, Indonesia and Brazil are the major
producers. For many years, cocoa production and commercialization have been the
basis of the economy in some Brazilian states and particularly in Bahia. Brazil
is the world’s sixth largest cocoa producer while Nigeria is the world’s fourth
largest cocoa producer according to the United National Food and Agriculture
Organization (Freire, et al., 1999).
A specific type of bacteria Streptomyces camerooniansis was found to
be beneficial for Theobroma cacao by
helping plant growth by accelerating seed germination of T. cacao, inhibiting growth of various types of microorganisms
(Such as; different Oomycetes, fungi and bacteria) and preventing rotting by Phytophthora megakarya.
Extraction of pulp does not interface with
subsequent seed fermentation and reduction of pulp before fermentation may be
beneficial to cocoa quality (Camu, et al., 2008).
Cocoa is very important ingredient in
several kinds of foods, such as chocolate, cake, biscuit, child-food, ice
creams and sweet consumed in developed countries (Guehi et al., 2007).
1.1
Cocoa Pulp
Raw cocoa beans are sheathed in an
aromatic mucilaginous pulp, which constitutes about 40% of the seed fresh
weight. This mucilaginous pulp is spongy and contains the sap (Pulp juice)
known in the industry as “sweating”. It is made up of about 67% to 82% water,
it is rich in sugars (10 to 15%), salts (1 to 2%, protein (2 to 3%), organic
acids (1 to 2%) and proteins (0.6%) Cocoa sweating has relatively low pH (3.0
to 4.0) primarily due to high concentrations of citric acid (1 to 3%) (Schwan
and Wheals, 2004).
Cocoa pulp is the raw material which
serves as the basis for fermentation, as it is a rich medium for microbial
growth which constitutes about 10% of the mass of the cocoa fruits. The
relatively high content of pectin (2 to 3%) and other polysaccharides makes the
pulp viscous limiting diffusion of air of the sugars present, about 60% to
sucrose and 39% is a mixture of glucose and fructose, the concentration of
sucrose, glucose and fructose is a function of cultivars and fruit age with
unripe pods containing mainly fructose and glucose (Thompson et al., 2001).
Cocoa fermentation occur largely on the
mucilaginous pulp, the reason is spontaneous (i. e. allowing naturally
occurring microorganism to ferment the pulp sugars) and involves succession of
microorganism. The microbial activity in
the cocoa pulp is a well defined temporal succession that is dominated by yeast during the first few hours, followed
by lactic acid bacteria (LAB) which
decline 48hrs of fermentation and
then they are overcome by acetic acid bacteria
(AAB) (Schwan and Wheals, 2004). The primary colonizers are yeasts, followed by
lactic acid bacteria (LAB), acetic acid bacteria (AAB) and Bacillus species.
During the early and mid-phase of the spontaneous fermentation of raw cocoa
beans, yeasts produce ethanol from carbohydrates and assimilate citric acid
under anaerobic and low pH conditions and cause depectinization of the pulp,
consequently the pulp liquefies and the juice drain off allowing air to diffuse
into the heap.
As air diffuses into the heap,
microaerophilic LAB, citrate-fermenting, acid tolerant and ethanol tolerant Lactobacillus
plantarum and Lactobacillus fermentum strains dominate this phase
(Galvez, et al., 2007)
The great majority of lactic acid bacteria
utilize glucose via the Embden-Meyerhof-Parnas pathway, yielding more than 85%
lactic acid. While some species utilize glucose through the hexose
monophosphate shunt, forming 50% lactic acid, as well as combinations of
ethanol, acetic acid, glycerol, mannitol, and carbon dioxide causing a slight
increase in pH of the pulp. Acetic acid bacteria are responsible for the
oxidation of ethanol to acetic acid and oxidation of acetic acid to carbon
dioxide and water.
Other pulp ingredients are presumably as
nutritious but nutrient declarations are hard to come by at this early stage in
the industry (Akinwale, 2000).
Cocoa pulp is the raw material on which
the fermentation proceeds and a key determinant of both quality and financial
viability of the process. Not only is the quantity of pulp crucial in affecting
the efficiency and nature of the fermentation but excess pulp can also be sold
as a high value commodity. Cocoa pulp can also be used in preparation of soft drinks, fresh
cocoa juice is collected, sterilized and bottled for the production of alcoholic
drinks such as brandy (Camu, et al.,2007).
Cocoa pulp is a very interesting
by-product of cocoa production and it forms the substrate for microbial growth
during the fermentation process. Most of the pulp goes to waste during the
processing of the cocoa beans and this has led to its utilization as a
potential raw material for development of soft drink to enhance the importance
of cocoa as a source of income for farmers.
A quick look at the historical use of
cocoa pulp reveals that cocoa farmer and their immediate communities have long
enjoyed the pulp as juice. This was discovered by Joseph Montgomery who owned
cocoa farms in Ecuador and launched Suavva, the first commercial scale juice
smoothie gotten from cocoa pulp.
Montgomery says that “Like most fruits,
the pulp is where the sweetness (fruit sugar) is found in cocoa”. The pulp can
be used to make jam, jelly, juice which can be used to do fermented beverage
like coca wine and other products (Fowler, 2009).
The fermentation of cocoa pulp is one of
the few remaining large scale spontaneous microbial processes in today’s food
industry. The microbiota involved in cocoa pulp fermentations is complex and
variable, which leads to inconsistent production efficiency and cocoa quality
(Michael, 2001)
Pulp
Extraction
The selection of cocoa pods is important
for the production of quality juice. The most suitable pods will be those three
quarters ripe and yellowish-green in colour (Adomako and Takrama, 1999).
Figure
1: Extraction and fermentation of cocoa pulp juice
1.2 Aim and Objectives
The aim of this work is to identify and
isolate the microorganisms that are involved in the fermentation of the cocoa
pulp.
1. To
evaluate the total heterotrophic bacterial and fungal load of the cocoa pulp
samples.
2. To
characterize and identify the various organisms involved in the fermentation of
the pulp.
Buyers has the right to create
dispute within seven (7) days of purchase for 100% refund request when
you experience issue with the file received.
Dispute can only be created when
you receive a corrupt file, a wrong file or irregularities in the table of
contents and content of the file you received.
ProjectShelve.com shall either
provide the appropriate file within 48hrs or
send refund excluding your bank transaction charges. Term and
Conditions are applied.
Buyers are expected to confirm
that the material you are paying for is available on our website
ProjectShelve.com and you have selected the right material, you have also gone
through the preliminary pages and it interests you before payment. DO NOT MAKE
BANK PAYMENT IF YOUR TOPIC IS NOT ON THE WEBSITE.
In case of payment for a
material not available on ProjectShelve.com, the management of
ProjectShelve.com has the right to keep your money until you send a topic that
is available on our website within 48 hours.
You cannot change topic after
receiving material of the topic you ordered and paid for.
Login To Comment