TITLE PAGE

EVALUATION OF CAFFEINE CONTENT OF COCOA BEAN FROM ETUNG LOCAL

GOVERNMENT AREA OF CROSS RIVER STATE, NIGERIA.

DEDICATION
I dedicate this work to God who has blessed me with the opportunity to acquire knowledge and
grow intellectually. I also dedicate it to my parents whose love, encouragement and sacrifices
have been the foundation of my educational pursuits.

ACKNOWLEDGEMENT
I would like to express my heartfelt appreciation to my supervisor Prof. N. E. Abu for her
invaluable guidance, expertise and unwavering support throughout the completion of this work.
Her knowledge and insights have greatly enriched my understanding of the subject matter.
Additionally I would like to extend my gratitude to the department of Plant Science and
Biotechnology for providing the necessary resources and creating a conducive environment for
academic growth. Their dedication to fostering intellectual development is truly commendable.

TABLE OF CONTENTS
CHAPTER ONE
Introduction

CHAPTER TWO
Literature review
2.1 Botanical description
2.2 Taxonomy
2.3 History
2.4 Cultivation and processing
2.5 Different Cacao varieties
2.6 Different Cocoa products
2.7 Caffeine and cocoa

CHAPTER THREE
Materials and methods
3.1 Sample collection and identification
3.2 Chemicals and instruments
3.3 Sample preparation
3.4 Caliberation solutions preparation
3.5 Caffeine extraction
3.6 Statistical analysis

CHAPTER FOUR
Results

CHAPTER FIVE

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5.1 Discussion
5.2 Conclusion

List of tables

Table 1: Caffeine values of samples gotten from Abia community of Etung LGA.
Table 2: Caffeine values of samples gotten from Bendegehe community in Etung LGA.
Table 3: Caffeine values of samples gotten from Bendegehe farm in Etung LGA
Table 4: Caffeine values of samples gotten from the Cocoa Research Institute of Nigeria.

List of plates

Plate 1: Dried cocoa bean

Plate 2: Centrifuge 800 D
Plate 3: DK-420 Water bath
Plate 4: 721 visible spectrophotometer
Plate 5: Commercial dehydrator
Plate 6: Sample being weighed by a weighing balance.

ABSTRACT
Beans of Theobroma cacao is used widely as a source of beverage when grinded and processed.
It is the only plant with commercial use in the production of chocolate. It is also an important
ingredient in the confectionery and pharmacy industries. Theobroma cacao bean contains
caffeine, a white crystalline xanthine alkaloid that acts as a psychoactive stimulant drug and a
mild diuretic. This study was carried out to investigate and evaluate the caffeine content of cocoa
beans gotten from Etung Local Government Area of Cross River state in Nigeria. Four Cacao
samples were prepared by drying sample seeds at 500°C in a commercial dehydrator and grinded
to fine powder with an electrical blender. The caffeine levels for grounded cocoa samples were
taken using a visible spectrophotometer. The results revealed the presence of caffeine at varying
levels. These variations could be greatly attributed to cocoa bean variety and external conditions.
The caffeine levels of all the samples ranged from 2.65 mg/g through 4.74 mg/g. A dose above
300 mg can cause caffeine intoxication whose symptoms includes restlessness, nervousness,
insomia, irritability etc. However, the values obtained in this study are way below 300mg,
therefore, these cocoa samples are safe for human consumption.

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 CHAPTER ONE

INTRODUCTION

Cocoa is a major agricultural export crop of Nigeria and a major employer of labour in the crop
producing areas. Nigeria is the fourth largest world producer of cocoa after Ivory Coast,
Indonesia and Ghana and the third largest exporter. It is the only plant with commercial use in
the production of chocolate. Despite having a pulp with a pleasant flavour, cocoa seed is the part
of cocoa mostly used in the food industry, generating several products with the greatest focus on
chocolate. In Nigeria, Cross River State is the second largest producer of cocoa after Ondo.
Other producing states are Ogun, Akwa Ibom, Ekiti, Delta, Osun, and Oyo. Ikom, Boki and
Etung local government areas are the highest producers of cocoa in Cross River State (Michael
and Utetiang, 2020).

The crop responds well to regions with coordinates not more than 20°, North or South of the
equator. This area is characterized by high temperatures and rainfall. However, the performance
of Nigeria’s cocoa economy has not been stable. The factors that are responsible for the yields
include climate change, impact on pests and diseases incidence, poor soil management practices
and use of old planting materials. Soil fertility is one of the essential components for a
sustainable cocoa plantation especially in tropical climates (Ozpalas and Ozer, 2017).

Cocoa as a cash crop is of considerable importance to both producing and consuming countries,
because it generates revenue from export, provides income and offers job opportunities. Cocoa is
an important ingredient in the beverage, confectionery and pharmaceutical industries. Therefore,
enhancing the sustainability of cocoa production and improving on its value chain is imperative,
particularly at a time when young people do not regard farming (including cocoa farming) as a
veritable business choice (Effiom and Odey, 2019).

As cocoa bean hull is an underutilized component of cocoa beans, they are receiving increased
attention. It has been reported that cocoa bean hull contained various functional compounds such
as phenolic phytochemicals, fibre, active compound packaging material, and significant fat
content that are similar to cocoa butter (Cai et al, 2021).

The rapid growth of the chocolate industry is driven by increased consumption, also increases
the amount of cocoa bean hulls generated annually. Cocoa bean hulls are the outer shell of cocoa
beans and weigh about 10-17% of the weight of a cocoa bean. It is the by-product obtained
during the production of cocoa products and usually disposed of and considered as waste in the
cocoa industry but yet to be discovered as a potential new ingredient for a high value product.
Cocoa bean hulls were left to rot on the cocoa plantation as fertilizer but leaching of excess
nutrients from decomposed hull contributes to pollution and eutrophication. Utilization of cocoa
bean hulls would reduce waste by circularizing the supply chain, turning it into a resource.
(Bdullahi et al, 2019).
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Caffeine on the other hand is a bitter, white crystalline xanthine alkaloid that acts as a
psychoactive stimulant drug and a mild diuretic. Variety of plants contain caffeine in their seeds,
fruits, and leaves (Rob et al, 2020). Besides coffee and tea, these plants include cacao beans (an
ingredient of chocolate), yerba matte leaves (used to make an herbal tea), and guarana berries
(used in various beverages and supplements) (Rob et al, 2020). Caffeine can also be synthesized
and is added to foods and beverages, including soft drinks, energy drinks, and energy shots, and
to tablets marketed for reducing fatigue. In addition, caffeine is widely used as a treatment for
apnoea of prematurity in infants, and caffeine and analgesic agents are used together in pain
medications (Rob et al, 2020).

Though caffeine content is relatively low in many cocoa species (Pavithra, 2021) this research
work aims to evaluate the caffeine content of cocoa bean gotten from Etung Local Government
Area in Cross River State, Nigeria.

CHAPTER TWO
LITERATURE REVIEW

2.1 BOTANICAL DESCRIPTION

Theobroma cacao is cauliflorous and semi-deciduous. The tree is low, reaching an average
height of 5-10 m. The main trunk is short; branches in whorls of 5, dimorphic; vertical chupons
growing from the trunk have leaves arranged in 5/8 phyllotaxy (agreement of leaves on stem).
The lateral branches (fans) have 1/2 phyllotaxy.
Petiole with 2 joined pulvini, one at the base and the other at the point of insertion of the leaf.
Stipules 2, deciduous. Lamina elliptical-oblong or obovate-oblong, simple, 10-45 cm long;
generally smooth, sometimes hairy, rounded and obtuse at the base, pointed apex.
Inflorescence dichasial; primary peduncle very short, often thick and lignified. Flower peduncle
1-4 cm long. Sepals 5, triangular, whitish or reddish in colour. Petals 5, joined at the base into a
cuplike structure, whitish-yellow with dark purple bands adaxially; ligules spathulate, yellowish.

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Stamens 5, fertile, alternating with 5 staminodes, the 2 whorls uniting to form a tube. Anthers 2,
stamens fused. Ovary superior with a single style terminating in 5 sticky stigmatic surfaces.
Fruit variable in shape, ovoid, oblong; sometimes pointed and constricted at the base or almost
spherical, with 10 furrows of which 5 are prominent.
Axial placentation, seeds embedded in mucilage, flat or round with white or purple cotyledons.
The generic name comes from the Greek ‘Theos’ (god), and ‘broma’ (food) and means the ‘food
of the gods’ (Orwa et al, 2009).

2.2 TAXONOMY

Kingdom: Plantae
Super division: Spermatophyta (Seed plants)
Division: Magnoliophyta (Flowering plants)
Class: Magnoliopsida (Dicotyledons)
Sub class: Dilleniidae
Order: Malvalves
Family: Sterculiaceae
Genus: Theobroma
Specie: T. cacao
Binomial name: Theobroma cacao L.
(Kokou and Ngo-sammick, 2014).

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2.3 History

The cacao tree was cultivated more than 3,000 years ago by the Maya, Toltec and Aztec peoples,
who prepared a beverage from cocoa beans (sometimes using it as a ceremonial drink) and also
used the beans as currency. The Maya considered chocolate to be the food of the gods and held
the cacao tree to be sacred. For the Aztec people, the drinks made from cocoa beans were
reserved for the emperor and aristocracy. After destroying the Aztec empire, the colonizing
Spanish began to experiment with cocoa; mole poblano, the rich chili-cocoa sauce, is probably
an early example of this. When cocoa reached Europe, the Italians used the new spice in savoury
dishes and created some of the earliest chocolate sweets with sugar. Cocoa and other chocolate
products are enjoyed by billions of people around the globe, but surprisingly few people know
the history of the confection. In fact, cocoa has appeared in different cultures worldwide for
hundreds of years. Cocoa was first developed as a crop in many ancient South American
cultures, with the Aztecs and Mayans being the most well-known of these indigenous
populations (Avendano-Arrazatech et al, 2018)

Researchers have found evidence of cocoa-based food dating back several thousand years. The
modern word chocolate stems from two words in Nahuatl, the language spoken by many native
groups: chocolatl, which translated literally means hot water, and cacahuatl, which referred to a
bitter beverage made with cocoa that was shared during religious ceremonies. The cacao bean
was so significant to the local cultures that it was used as a currency in trade, given to warriors as
a post-battle reward, and served at royal feasts. When the Spanish Conquistadors arrived in the
New World and began the process of invading, colonizing, and ultimately destroying the native
cultures, they also discovered the value of the local cacao crop. However, they brought their own
innovation to the appropriated drink the addition of sugar and spices to sweeten the bitter cocoa.
After that point, chocolate became wildly popular amongst the Spanish, who kept the production
method a secret from other Europeans for almost 100 years after their discovery. The Spanish
could not hold onto their secret forever, and chocolate quickly spread across the rest of western
Europe. Chocolate then still exclusively in the form of a drink appeared in France, and then
England, in royal courts and special chocolate houses that served the social elite. Hot chocolate
was hailed by the upper classes as both delicious and healthy, and cocoa ultimately gained the
reputation of being an aphrodisiac (Gatew and Mengistu, 2019).

The exclusivity of chocolate was ultimately diminished by the onset of the Industrial Revolution,
when steam-powered machines made the production of cocoa powder significantly quicker and
more affordable. Solid chocolate hit the market and found wild success by 1850, due to the
discovery by Joseph Fry that adding cacao butter to the cocoa powder formed a solid mass. Sixty
years later, the art of creating chocolate confections with flavored filling referred to as pralines
by their Belgian inventor, Jean Neuhaus II went public. From there, the chocolate and cocoa
industry exploded in popularity and quickly spread around the world. Throughout its centuries-
long evolution, one factor has remained consistent and cocoa has attracted devotees worldwide.
Today, over 4.5 million tons of cocoa beans are consumed annually around the globe, in
everything from drinks to candy bars. It is safe to say that the ancient Mesoamericans who
4
pioneered the crop could never have imagined the popularity cocoa would someday experience.
To secure the future of chocolate and ensure that it is available for generations to come, it is
essential that sustainable farming practices and ethical means of production are implemented in
the cocoa supply chain (Russel and Martin, 2023).

2.4 CULTIVACULTIVATION AND PROCESSING

After four years a mature cacao tree produces fruit in the form of elongated pods that range in
colour from bright yellow to deep purple. A single tree may yield up to 70 such fruits annually,
and the fruits ripen in less than six months. Each has numerous ridges running along its length
and holds 20 to 60 seeds, the cocoa beans, arranged around the long axis of the pod. The oval
seeds are about 2.5 cm (1 inch) long and are covered with a sweet sticky white pulp. Harvesting
of cocoa beans can proceed all year, but the bulk of the crop is gathered in two flush periods
occurring from October to February and from May to August. The ripe seed pods are cut from
the trees and split open with a machete. The beans, removed from the pods with their
surrounding pulp, are accumulated in leaf-covered heaps, in leaf-lined holes dug in the ground,
or in large shallow boxes perforated at the bottom to provide for drainage (Michael, 2021).

The beans with their pulp are fermented for one to seven days, depending on the type and grade.
Frequent turnings dissipate excess heat and provide uniformity. During fermentation the juicy
sweatings of the pulp are drained away, the germ in the seed is killed by the increased heat, and
flavour development begins. The beans become plump and full of moisture, and the interior
develops a reddish brown colour and a heavy sharp fragrance. The fermented beans are sun-dried
or kiln-dried to reduce moisture content to 6-7 percent and are bagged for shipment.

In the case of cocoa butter, the fermented and dried cocoa beans are then typically roasted.
Roasting develops flavour, reduces acidity and astringency, lowers moisture content, deepens
colour, and facilitates shell removal. After roasting comes a cracking and fanning (winnowing)
process, in which machines crack the shells and then separate them from the heavier nibs by
means of blowers. Nibs can be sold or further processed. Grinding releases the fat, or cocoa
butter, and forms a paste, called chocolate liquor or cocoa mass. The chocolate liquor is pressed
and pulverized into cocoa powder to make chocolate. Cocoa bean shells, the major by-products
of cocoa processing, represent 8-10 percent of raw cocoa bean weight and are blown off in the
cracking and fanning, or winnowing, operation. They are used for fertilizer, mulch, and fuel.
(Stradler and Guilaume, 2014).

2.5 DIFFERENT CACAO VARIETIES

There are three varieties of Theobroma cacao, as it was officially named in 1753 by the Swedish
scientist Carl von Linné. They are: Criollo, Forastero and Trinitario. There are also multiple
hybrids of each strain. Criollo and Trinitario are the fine varieties prized for making chocolates
with the most interesting and complex flavours. Forastero is typically the hardier, more reliable
strain and has a higher yield of cacao pods. As a result, it’s easier and cheaper to grow, which is

5
why it is generally used to make mass-produced chocolate. It’s consistent and reliable, but can be
rather bland and somewhat acidic. The reason all this matters is that fine varieties of cacao all
taste very different, infact there are said to me more than 400 different flavour notes in cacao.
Think of cacao like wine or coffee (Lam et al, 2016).

In Maya and Aztec times, lower-grade cacao was mixed with maize and other seeds, while high-
quality chocolate was drunk in a purer form. It is the same now. Manufacturers use cheap, less
interesting Forastero cacaos to make confectionery bars and save the fine, rare beans for making
rich, flavoursome dark chocolate. Because there is such a huge quality range in cacao, the
chocolate industry has always attracted counterfeiters and swindlers. Aztec market traders, when
selling cacao beans, often used to make their wares go further by mixing them up with avocado
stones, bits of dough, wax, clay and anything else that could pass for the real thing. In Europe in
the eighteenth and nineteenth centuries, chocolate was frequently diluted with brick dust or red
lead, and lard was added to cacao butter to bulk it out. These days, ingredients list show
everyone how much fat, sugar and lecithin goes into your average chocolate bar. It is amazingly
people still call it chocolate (Suratam et al, 2019].

The Criollo and Forastero varieties came first, but although they both evolved in the Amazon
Basin, they are quite different. Criollo beans are what the Maya and Aztecs went wild about, the
strain that later seduced and enchanted Europeans in the seventeenth and eighteenth centuries.
Today, Criollo tends to be the preferred bean of chocolate connoisseurs. The beans are
tantalizingly rare, accounting for only 3% of the world crop, and unusually they are often white
in colour.

The Trinitario bean has an interesting history. When virtually the entire crop of Trinidad’s
Criollo cacao was wiped out in the eighteenth century either by a hurricane or by a plant disease
of some kind. Forastero trees were introduced to the island, where they cross-pollinated with the
few remaining Criollo trees to produce a new strain of cacao. Trinitario is hardier than Criollo,
but tastier than Forastero, so it’s very versatile. It has classic nutty, coffee notes and layers of
complexity. Venezuela is home to many of the world’s finest cacaos, in particular Criollos. This
is in part because when they discovered oil, they lost interest for a while in the less lucrative
cacao and did none of the replanting with modern productive hybrids that so many countries did.

Venezuelan cacao used to be called Caracas wherever it was imported, but now it is known by
the region in which it grows, just like wine. Chuao, which is considered by some people to be
among the world’s finest Criollo cacao, comes from a village in Venezuela. Porcelana Blanca,
another highly prized variety, was originally called Maracaibo because it came from the area
around Lake Maracaibo (Lam et al, 2016).

2.6 DIFFERENT COCOA PRODUCTS.

This takes a look at some of the most popular types of chocolate and cocoa products. Baking
chocolate was traditionally an unsweetened bar of chocolate used for baking purposes However,
in recent times, it is possible to buy a wide range of sweetened baking chocolate products too.
These chocolate bars tend to be ‘cheap and cheerful’ options rather than premium chocolate

6
products, and their affordable pricing reflects this. Unsweetened, Bittersweet, Semisweet, Sweet
chocolates are the different kinds of baking chocolate that can be found in stores. Sweet baking
chocolate has the highest sugar content, while unsweetened, as the name suggests, contains no
added sugar (Thiago and Beatriz, 2018).

Brewed cocoa. Interestingly, it is possible to use cocoa beans in the same way as coffee. Brewed
cacao is an increasingly popular drink made from ground cocoa beans. Although it may sound
similar, there is a big difference between brewed cocoa products and regular cocoa powder. For
one thing, cocoa powder undergoes processing that removes a large proportion of the cacao
butter. In contrast, brewed cacao products are pure cacao beans that have been roasted and then
ground into a texture conducive for brewing. In other words, they are made in the same way as
ground coffee products. Although brewed cocoa is relatively low in caffeine, it contains large
amounts of theobromine. Brewed cocoa has a rich chocolatey taste, and it contains no calories.
(Karen et al, 2019).

Cocoa butter; is simply the pure fat removed from cocoa beans during their processing. Although
it is possible to buy pure cocoa butter, it is most commonly used for making chocolate bars.
However, cocoa butter is costly, and one of the most expensive isolated fats. For this reason,
many chocolate products use cheaper alternate fats such as palm oil. The primary fatty acid in
cocoa butter is stearic acid, which is a type of saturated fat. Interestingly, stearic acid does not
have a significant influence on lipids/cholesterol. As a result, cocoa butter has a smaller impact
on cholesterol levels compared to other rich sources of saturated fat. However, it does tend to
have slightly more of an effect than olive oil and other cooking oils because it also contains high
amounts of palmitic acid (Thiago and Beatriz, 2018).

2.7 CAFFEINE AND COCOA

Caffeine is a natural chemical stimulant that can also be created synthetically for consumption.
Natural caffeine is found in coffee beans, tea leaves, cacao beans, guarana berries, and yerba
maté leaves. Caffeine preparations can be added to drinks, food, tablets, or powdered
supplements. In the US, about 85% of adults consume caffeine daily, and average intake is 135
mg per day (equivalent to 12 oz of coffee). The most common source of caffeine is coffee for
adults and soft drinks and tea for teenagers (Walter, 2022).

A single dose consumption of 200 mg of caffeine, or less, by healthy people without

comorbidities and pharmacokinetic disturbances, is usually not associated with toxic effects.
However, a dose above 300 mg at once can cause caffeine intoxication, the symptoms of which
are mainly related to its stimulating effect. The most common ones are: restlessness,
nervousness, excitement, insomnia, facial flushing, increased urination, gastrointestinal
disorders, muscle tremors, chaotic flow of thoughts and speech, irritability, arrhythmia,
tachycardia, and psychomotor agitation. The severity of the undesirable effects of caffeine
consumption is dose dependent. The threshold of caffeine toxicity appears to be about 400
mg/day in healthy adults (19 years or older), 100 mg/day in healthy adolescents (12-18 years
old), and 2.5 mg/kg/day in healthy children (less than 12 years old) (Rodak et al, 2021).

7
Caffeine has both beneficial and harmful health consequences. It can be used to treat and prevent
premature new born respiratory issues like as bronchopulmonary dysplasia and apnea (Negeri,
2021). The WHO Model List of Essential Medicines includes caffeine citrate. It may have a
minor preventive effect against some disorders, such as Parkinson's disease. Caffeine
consumption can cause sleep disruption or anxiety in some people, although it has no effect on
others. Although there is no conclusive evidence of a danger during pregnancy, several
authorities advise pregnant women to limit their caffeine intake to the equivalent of two cups of
coffee per day or less (Negeri, 2021).

Caffeine is naturally found in the fruit, leaves, and beans of coffee, cacao, and guarana plants. It
is also added to beverages and supplements. There is a risk of drinking excess amounts of
caffeinated beverages like soda and energy drinks because they are taken chilled and are easy to
digest quickly in large quantities (Chan, 2020).

Several research efforts on cocoa have been focused on parameters for controlling the
transformation process to guarantee homogeneity and quality of cocoa beans, the main raw
material in the chocolate industry. The main changes that determine the final quality of cocoa—
and also the product’s homogeneity—occur during fermentation, given the great number of
factors that affect the process. Results have shown that temperature and pH profile are the key
factors to be monitored and controlled in order to achieve high-quality cocoa beans (Ana et al,
2021).

CHAAPTER THREE
MATERIALS AND METHODS

3.1 SAMPLE COLLECTION AND IDENTIFICATION.

Few cocoa pods samples were collected from Etung Local government area of Cross River state,
Nigeria. They were collected by Mr. Eugene Ojuah, a Research consultant of the University of
Nigeria Nsukka, Enugu state. The samples were authenticated my Mr. Felix Nwafor, a
Taxonomist of the University of Nigeria, Nsukka.

3.2 CHEMICALS AND INSRUMENTS.

The chemicals used in this study includes Chloroform (CHCL3), distilled water and Sodium
carbonate (Na2CO3). High performance liquid chromograph, Weighing balance, Laboratory
glass wares, filter paper, testube rack, water bath, syringe, aluminium foil, UV visible
spectrophotometer, dehydrator, corvette.

3.3 SAMPLE PREPARATIION

The cocoa fruit pods were cut open and the beans were separated from the pulp. The beans were
taken to the laboratory where they were dried in a dehydrator at 500c. After drying, the shells are
removed from the bean and ground to fine powder. Grinding is done using an electrical blender
(Vuletic et al, 2021).

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3.4 CALIBERATION SOLUTIONS PREPARATION.
A stock solution of caffeine, of 100ppm was made by dissolving 0.01g of recrystallized caffeine
in 100ml of chloroform in the volumetric flask. The following dilutions were prepared from the
caffeine stock solution: 1 ppm, 5 ppm, 10 ppm, 15 ppm, 20 ppm, 25 ppm. The absorbances of
these dilutions were measured in the spectrophotometer at wavelength of 274 nm in quartz
cuvettes (three times for each dilution). The absorbance values of the solutions were used to
make the calibration line for the caffeine content analysis. (Vuletic et al, 2021).

3.5 CAFFEINE EXTRACTION

Cocoa stock solution was prepared by dissolving 1g of cocoa powder in 10ml of water in a
testube. These solutions were further boiled in a water bath at 100 degrees Celsius for 60mins.
The solutions were filtered into a clean testube to eliminate the solid residue. 1g of Sodium
carbonate is then added to each testube containing the liquid. The liquid is further placed into a
High performance liquid chromograph and filtered after 10 minutes. 5ml of Chloroform is added
to each testube containing the filtered solution. The Chloroform is separated from the liquid and
put in a vial. The liquid is then transferred to a cuvette and placed in a UV visible
spectrophotometer and readings are taken. (Vuletic et al, 2021).

3.6 STATISTICAL ANALYSIS

The data obtained from this study was statistically analyzed using T-test and one-way analysis of
variance (ANOVA). The level of statistical significance was set at P≤0.05 (95% confidence
interval). The software used for this analysis is SPSS Version 23.

9
10
 CHAPTER FOUR
RESULTS

The mean caffeine values of samples obtained from Abia community ranged from 3.35 and 4.05.
The mean valus of caffeine content are presented in table 1.

Table 1: Caffeine values of samples gotten from Abia community in Etung LGA of cross river
state.

Samples Values(mg/g)
ABI-ETU-TL 3.3500 ± 0.5939de
ABI-ETU-SC 4.050 ± 0.000bc
Values expressed as mean ±
standard deviation. Means with same letters as superscripts are not significantly different at p ≤
0.05

The mean caffeine values gotten from Bendegehe community ranged from 3.56 to 4.74. This
result shows minimal variation of mean values of caffeine from the cocoa samples gotten from
the community. Sample BE-ETU-SCB gotten from this community (Bendegehe) also had the
highest caffeine content compared with samples from all other communities.

Table 2: Caffeine values of samples gotten from Bendegehe community in Etung LGA of cross
river state.

Samples Values(mg/g)
BE-ETU-TLA 4.535 ± 0.2899ab
11
BE-ETU-SC 4.675 ± 0.2899bc
BE-ETU-TLES 3.560 ± 0.0989cd
BE-ETU-SCES 4.540 ± 0.0989ab
BE-ETU-SCB 4.740 ± 0.000a
BE-ETU-TLB 4.047 ± 0.1484bc
Values expressed as mean ± standard deviation. Means with same letters as superscripts are not
significantly different at p ≤ 0.05

The samples from the Bendegehe farm had the lowest mean values for caffeine content across all
samples examined from the various communities.

Table 3: Caffeine values of samples gotten from Bendegehe farm in Etung LGA of cross river
state
Samples Values(mg/g)
BEF-ETU-TL 3.42 ± 0.0989e
BEF-ETU-SC 2.65 ± 0.00d

Values expressed as mean ± standard deviation. Means with same letters as superscripts are not
significantly different at p≤ 0.05

The sample results gotten from the Cocoa Research Institute of Nigeria shows a relatively consistent
mean value.

Table 4: Caffeine values of samples gotten from Cocoa research institute of Nigeria.
Samples Values(mg/g)
CRIN-ABIL 3 ± 0.00de
CRIN-TRI 3 ± 0.00de
CRIN-SG 3 ± 0.00de
CRIN-EXT 3 ± 0.00de
CRIN-SGa 4.5 ± 0.707ab
CRIN-F3HY 4 ± 0.00bc

Values expressed as mean ± standard deviation. Means with same letters as superscripts are not
significantly different at p ≤ 0.05

CHAPTER FIVE
DISCUSSION AND CONCLUSION
The results obtained from this study shows that the different cocoa samples obtained from different
communities across Etung LGA contains varying quantities of caffeine. The mean caffeine values of
samples gotten from Abia community of the LGA varied slightly between 3.35 and 4.05. The examined
samples from the Bendegehe community have the highest level of caffeine while the samples from
Bendegehe farm had the lowest caffeine content in their beans.
The samples from the Cocoa Research Institute of Nigeria (CRIN) shows a consistent mean caffeine value
of 3.0 mg/g with the exception of two samples (CRIN-SGa and CRIN-F3HY) which had values of 4.0 mg/g
and 4.5 mg/g respectively.
12
 A single dose consumption of 200 mg of caffeine, or less, by healthy people without comorbidities and
pharmacokinetic disturbances, is usually not associated with toxic effects. However, a dose above 300
mg at once can cause caffeine intoxication, the symptoms of which are mainly related to its stimulating
effect. The most common ones are: restlessness, nervousness, excitement, insomnia, facial flushing,
increased urination, gastrointestinal disorders, muscle tremors, chaotic flow of thoughts and speech,
irritability, arrhythmia, tachycardia, and psychomotor agitation (Rodak et al, 2021).

It can be said that the caffeine levels determined from all the samples examined across the communities
in Etung Local Government Area are below the maximum required dosage of caffeine intake and is
therefore safe for consumption if consumed in the right quantity.

It is known that external conditions can have a modifying effect on the external and internal
characteristics of a plant. The external conditions are provided by climate and soil, which both
CONCLUSION
The evaluation of caffeine content of cocoa beans from Etung Local Government Area of Cross River
state shows varying level of caffeine across four accessions (Abia, Bendegehe, Bendegehe farm and the
Cocoa research institute of Nigeria) in the LGA.

REFRENCES
Ana, M.C., Blanca, L. B., Maria, C. G., William, A. C., Andrea, C. M. and Jennifer, C. (2021).
Dynamics of cocoa fermentation and its effect on quality. PubMed Central. 1(11):146-167.

Avendano-Arrazatech, C. H., Martinez-Bolanos, M., Mendoza, L. A (2018). The cocoa

[Theobroma cacao L] native in Mexico. Biodiversity International Journal. 2(6):535-536.

Bdullahi, R., Lawal, A.M., Khalid, A., Ibrahim, M.S. and Muhammad, U.L (2019). Assessment
of the level of caffeine in some tea leaves marketed in Duste, Jigawa state. Korean Journal of
Food and Health convergence. 5(3):7-20.

Byrne, P. (2018). General overview on Theobroma cacao L. Colorado State University

Extension. 8(4):018-020

Cai, S. S., Eric, W. C., Chee, W. W. and Chee, W. N. (2021). Antioxidant and sensory evaluation
of Theobroma cacao tea formulated with bean hull of different origins. Future foods. 3(1):279-
284.

Chan, T. H. (2020). Caffeine. The nutrition force. 4(6):012-018.

13
Effiom, F. and Odey, B., (2019). Rainfall pattern and Nigerian agricultural value chain; the case
of cocoa production in Etung, Cross river state, Nigeria. 1(1):069-088

Gatew, H. and Mengistu, K. (2019). Discovery learning about cocoa. Journal of Agriculture and
Biomedicine. 9(6):157-163

Karen, H. N. and Nanay, V. M. (2019). Cocoa by products. Wiley online library. 1(2):142-151.

Kokou, E. A. and Ngo-Samnick, L. E. (2018). Cocoa production and processing. Technical

Centre for Agriculture and Rural Cooperation (CTA) Publishing, Cameroon. Pp 45.

Lam, T.V., Phung, T. H. and Helena, E (2016). Characterization of leaf, flower and pod
morphology among Vietnamese Cocoa varieties. 48(6):2375-2383

Michael, A. K. and Utetiang, L. U., (2020). Soil fertility evaluation of major cacao producing
areas of Cross River state, Nigeria. Journal of Agriculture and food sciences. 2(6):41-47.

Michael, J. (2021). 11 types of chocolate and cocoa products. Nutrition advance. 3(6):115-117

Negeri, Z. (2021). An overview on caffeine. Journal of clinical and experimental

pharmacology. 11[7]:100-289.

Orwa, C., Mutual, A., Kindly, R., Jamnadass, R. and Anthony, S. (2009). Agroforestry Database
referenceandselectionguideversion4.0(http://www.worldagroforestry.org/sites/treedbs/
treedatabases.asp)

Ozpalas, B. and Ozer, E. A. (2017). Effect of caffeine on human health. Journal of the American
college of cardiology.12(1):297-305.

Pavithra, V. (2021). Review article on caffeine activity. Journal of Nanotechnology research.

4(6):223-247

Rob, M., Frank, B., Walter, C. and Willet M.D. (2020). Coffee, Caffeine and health. The New
England Journal of Medicine. 283(4):369-378

Rodak, K., Kokot, I. and Kratz, E. M. (2021). Caffeine as a factor influencing the functioning of
the human body-- Friend or foe? Nutrients. 13(1):30-88
Russel, L. C. and Martin, C., (2023). Cocoa bean. Encyclopedia Britannica. 2(2):252-264.

Stradler, R. H. and Guil, L., (2014). Food materials, technologies and risks. Encyclopedia of food
safety. 5(2):098-102.

Suratman, S., Shubzam A. M. and Chykita, A. J. (2019). Exploration on the morphology of

Cacao plant in Bacan Island. Advances in Engineering research. 2(6):019-028.

14
Thiago, F. S. and Beartiz, M (2018). Cocoa by products: characterization of bioactive
compounds and beneficial health effect. 2(18):74-81

Vuletic, N., Bardic, L. and Ozdak, R. (2021). Spectrophotometric determining of caffeine

content in the selection of teas, soft and energy drinks available on the Croatian market. Food
Research. 5(2):325-330

Walter, K. (2022). Caffeine and health. Jama patient page. 327(7):693

15