EFFECT OF METHANOLIC Gmelina arborea LEAF EXTRACT ON

RUMEN FERMENTATION PARAMETERS AND MICROBIAL COUNTS

OF WEST AFRICAN DWARF SHEEP

BY

LOWEN, ROBERT OLUWAFERANMI NDAH/23/6031

ONWUKWE, JAMES MIRACLE NDAH/23/6082

A PROJECT REPORT SUBMITTED TO THE DEPARTMENT OF ANIMAL

HEALTH AND PRODUCTION TECHNOLOGY IN PARTIAL
FULFILMENT OF THE REQUIREMENTS FOR THE AWARD OF
NATIONAL DIPLOMA IN ANIMAL HEALTH AND PRODUCTION
TECHNOLOGY

OF THE

FEDERAL COLLEGE OF ANIMAL HEALTH AND PRODUCTION

TECHNOLOGY, MOOR PLANTATION, IBADAN

MAY, 2025.
 CEERTIFICATION
We hereby certify that this project was carried out by LOWEN ROBERT
OLUWAFERANMI, with matriculation number HNDAPT/23/6031 under our
supervision in the Department of animal Production Technology of the Federal
College of Animal Health and Production Technology, MOOR Plantation Ibadan.

……………………….............. …..………………………
Dr. (Mrs.) Ogunleke Date
Supervisor

……………………….............. …..………………………
Dr. A.A. Saka Date
Co-Supervisor
 ABSTRACT
A 56 days dietary trial was conducted to investigate the influence of methanolic gmelina arborea
leaf extract (MGALE) on rumen fermentation parameters of west african dwarf (WAD) sheep. A
total of twenty (20) WAD sheep were allotted into 4 dietary treatments,consisting of 5 replicates
per treatment in a completely randomised design. MGALE was administered at different varying
levels of 0mL,10mL,20mL and 30mL for the 4treatments T1,T2,T3 and T4 respectively. Same
basal diet and concentrate was fed across the treatment groups. Data was collected on rumen
fermentation parameters and microbial counts. Results revealed that MGALE had significant
influence on the rumen fermentation parameters investigated except in pH,temperature and
Ammonia nitrogen. Sheep administered 1pmL had the highest values of rumen fermentation
parameters. Administration of MGALE had significant effect on microbial count observed,sheep
administered 10mL had the highest total bacteria count (11.50 × 10_⁷cfu/mL) and lowest Total
colony count (1.80 × 10_²cfu/mL) . It was therefore concluded that MGALE can be administered
10mL in WAD sheep as it influenced the rumen fermentation parameters values and the total
bacteria count.
 ACKNOLEDGEMENT
My profound gratitude goes to Almighty God for giving me the capability, wisdom,
understanding and knowledge during the course of my program at the Federal College of Animal
Health and Production Technology, Moor Plantation, Ibadan, Oyo State and the successful
completion of my project
My utmost appreciation goes to my supervisor Dr. (Mrs.) Ogunleke for her guidance, support
and assistance during the period of the research.
My sincere gratitude goes to Dr. Saka A.A for his dedication, assistance and encouragement
during this research.
I appreciate the Head of Department and the entire staff of the department both teaching and non
teaching.
To my parents, Mr & Mrs Lowen, I am very grateful for the love and support you showed me
and the gift of education you gave unto me.
My appreciation is not complete without mentioning the efforts of my project mates Sulaimon
Olamide, Anthony Taiwo, Temitayo Fafowora, Abubakir Sekinat, Olaoye Zainab, Adeshina
David, Adebayo Agbolahan, Adeoye Rhoda, Benedict Barnabas, James Miracle, Alakufo Zainab,
David Ruth, Taiwo Abiodun, Aderibigbe Addul, Adigun ikimot, Adepoju Abdulafeez, Omotosho
kemi, Okunola Segun.
See you at the top.
 TABLE OF CONTENT
Title Page
Title page
Certification
Abstract
Dedication
Acknowledge
Table of content
List of tables

CHAPTER ONE
1.0 Introduction
1.1 Justification
1.2 Objectives of Study

CHAPTER TWO
2.0 Literature review
2.1 Overview of the Rumen
2.1.1 Ruman pH
2.1.2 Ammonia Nitrogen (NH3-N)
2.1.3 Volatile Fatty Acids (VFAs)
2.2 West African dwarf (WAD) sheep
2.3 Botanical and Nutritional profile of Gmelina arborea
2.3.1 Traditional and Ethnomedicinal Uses of Gmelina arborea
2.3.2 Phytochemical Constituents and Their Potential Effects On Rumen Microbes
2.3.2.1 Tannins
2.3.2.2 Saponins
2.4 Plant Extracts In Ruminant Nutrition
2.4.1 Merits And Potential Demerits of Using Methanolic Extracts In Ruminant Nutrition
3.0 Materials And Method
3.1 Experimental Site And Duration
3.2 Source and Preparation of Test Ingredient
3.2.1 Extraction
3.3 Experimental Animals And Management
3.4 Experimental Diet
3.5 Experimental Design
3.6 Data Collection
3.7 Chemical Analysis
3.8 Statistical Analysis
REFERENCES
 LIST OF TABLES
Tables Pages
Table 1: Gross and Chemical Composition of the Experimental Diet
 CHAPTER ONE
1.0 INTRODUCTION

Ruminant livestock play a crucial role in Nigeria's agricultural sector, contributing significantly
to food security, income generation, and socio-cultural practices. Among these, small ruminants
particularly sheep and goats are indispensable assets for rural households due to their
adaptability, low maintenance requirements, and multifaceted utility. They serve as sources of
meat, milk, manure, and income, and often function as financial reserves or assets that can be
liquidated in times of need (Bayer, 1985, Abiola-Olagunju et al., 2024).

Nigeria boasts the largest small ruminant population in Africa, with approximately 42.1 million
sheep and 73.8 million goats, predominantly indigenous breeds (CSIRO, 2016). These animals
are primarily reared by smallholder farmers in low-input, extensive systems, relying on natural
pastures and crop residues. Despite the challenges of seasonal feed shortages, diseases, and
limited veterinary services, small ruminant production remains a vital component of rural
livelihoods (CSIRO, 2016).

The West African Dwarf (WAD) sheep, in particular, holds significant importance in the southern
regions of Nigeria. This breed is renowned for its adaptability to the humid tropical environment,
resistance to trypanosomiasis, and ability to thrive on low-quality forages (Adjibode et al.,
2017). WAD sheep are predominantly managed by smallholder farmers, often under traditional
systems that involve free-range grazing and minimal supplemental feeding (Ahaotu, 2011). Their
small size, early maturity, and prolific breeding make them well-suited for the resource-
constrained conditions typical of rural communities.

Beyond their economic value, WAD sheep also play a role in the social and cultural fabric of
rural Nigeria. They are commonly used in traditional ceremonies, religious rites, and as gifts
during social events. Moreover, their manure contributes to soil fertility, supporting crop
production and sustainable farming practices (Bayer, 1985). Given the integral role of WAD
sheep in rural economies, enhancing their productivity through improved nutrition, health
management, and breeding practices is essential (Whannou et al., 2021). Research into
alternative feed resources, such as the use of plant extracts like methanolic Gmelina arborea leaf
extract, offers promising avenues for improving rumen fermentation and microbial efficiency,
thereby boosting the overall performance and sustainability of WAD sheep production systems
(Adjibode et al., 2017). In many parts of Nigeria, the dry season brings about a serious decline in
both the availability and quality of forage, which poses a major challenge to ruminant livestock
production. During the rainy season, ruminants especially sheep and goats have access to lush,
green pastures that are rich in nutrients (Jiwuba et al., 2017). However, once the rains stop and
the dry season sets in, these pastures dry up and lose their nutritional value.

One of the main issues is that the crude protein content of forage drops significantly during the
dry season, while the fiber content increases. This makes the feed less digestible, which in turn
reduces how much the animals are willing or able to eat (Ojo et al., 2020). As a result, ruminants
often suffer from weight loss, reduced milk production, slower growth, and poor reproductive
performance. These effects are even more pronounced in smallholder farming systems where
animals depend entirely on natural grazing.

To cope with the scarcity of feed, many herders resort to feeding their animals with crop residues
or agro-industrial by-products like cassava peels and maize husks. Unfortunately, these
alternatives are usually low in essential nutrients and are only suitable as fillers rather than
complete rations. The situation is worsened by the fact that many farmers do not have the
resources or knowledge to process or supplement these feeds to make them more nutritious
(Lamidi & Ologbose, 2014). In some cases, pastoralists are forced to migrate with their herds in
search of greener pastures. While this may help in the short term, it often leads to overgrazing in
certain areas and sometimes results in land disputes with crop farmers. To address these
challenges, experts have recommended strategies like hay and silage production, planting
drought-tolerant fodder species, and supplementing animal diets with nutrient-rich additives.
However, the adoption of these solutions remains limited, mostly due to a lack of training,
infrastructure, and financial support among small-scale farmers (The Junction, 2021; Ojo et al.,
2020).

In recent years, there has been increasing interest in the use of plant-based supplements to
improve livestock nutrition and reduce the environmental footprint of ruminant production
systems. One such plant with promising potential is Gmelina arborea, a fast-growing deciduous
tree commonly found in tropical regions, including Nigeria. The leaves of Gmelina arborea are
rich in phytochemicals such as tannins, flavonoids, and saponins, which have been shown to
influence rumen microbial activity and fermentation processes (Oskoueian et al., 2013).

Methane production in the rumen, a natural by-product of microbial fermentation, represents

both an energy loss to the animal and a significant contributor to greenhouse gas emissions.
Research suggests that around 2–12% of the gross energy consumed by ruminants is lost as
methane, depending on the diet and animal type (Patra & Saxena, 2010). Reducing methane
output is not only beneficial for the environment but can also enhance the animal’s productivity
by redirecting energy toward growth and milk production.

Several studies have shown that plant secondary metabolites, especially tannins and saponins,
can help suppress methane production in the rumen by inhibiting methanogenic archaea and
modifying the fermentation pathway (Jayanegara et al., 2011). This action shifts the microbial
balance in favor of more efficient digestion and energy utilization.

Although Gmelina arborea is less studied in this context compared to other tannin-rich plants, its
phytochemical profile positions it as a strong candidate for improving rumen fermentation and
mitigating methane emissions. Its use as a dietary supplement particularly in the form of
methanolic extracts could provide a natural, locally available solution to some of the challenges
associated with dry season feeding and enteric methane production. Further investigation into its
specific effects on rumen parameters and microbial populations in animals such as West African
Dwarf sheep is therefore both timely and important.

1.1 JUSTIFICATION OF THE STUDY

Methanolic Gmelina arborea leaf extract could be a potential alternative to antibiotics in

modifying rumen fermentation parameters and microbial count in West African Dwarf sheep,
addressing concerns over antibiotics use and its impact on animal (Chessan, 2006).

The increasing pressure on conventional feed resources and the rising cost of high-quality
additives have underscored the need for sustainable and cost-effective solutions in ruminant
nutrition. In Nigeria, where smallholder farmers depend on low-input systems, enhancing
ruminant productivity is critical for improving livelihoods and food security. Sustainable feed
additives offer a practical pathway to achieve this goal by helping to optimize feed utilization,
enhance nutrient absorption, and ultimately boost animal performance. Recent research has
demonstrated that natural plant extracts can serve as viable alternatives to synthetic additives,
reducing feed costs and supporting environmental sustainability (Silva et al., 2019).

Gmelina arborea, commonly known as beechwood, is a fast-growing tree species widely

distributed across Nigeria. Its leaves are rich in bioactive compounds such as tannins, flavonoids,
and saponins, which have been shown to positively influence rumen microbial populations and
fermentation processes (Patra & Saxena, 2010). These compounds can enhance feed digestibility,
reduce methane emissions, and improve overall animal health (Oskoueian et al., 2013).

Research indicates that incorporating Gmelina arborea leaf meal into ruminant diets can serve as
a cheap protein supplement, improving voluntary feed intake, digestibility, and general
performance of animals fed low-quality feeds. Additionally, the plant's high crude protein content
and favorable fiber composition make it a valuable feed resource, especially during periods of
forage scarcity (Adewale & Olaniyi, 2020).

1.2 OBJECTIVES

 To determine the effect of methanolic Gmelina arborea leaf extract on rumen

fermentation parameters such as pH, volatile fatty acid concentration and ammonia
nitrogen levels.
 To evaluate the effect of methanolic Gmelina arborea leaf extract on the microbial
population in the rumen including total bacteria, protozoa and fungi counts.
 CHAPTER TWO

2.0 LITERATURE REVIEW

2.1 THE OVERVIEW OF THE RUMEN

The rumen, which is the largest compartment of the ruminant stomach, plays a central role in
digesting fibrous feeds that monogastric animals cannot efficiently utilize. It acts as a
fermentation chamber where a complex community of microorganisms including bacteria,
protozoa, fungi, and archaea break down ingested plant material into simpler, usable forms of
energy for the host animal (Yáñez-Ruiz, Abecia, & Newbold, 2015).

Among these microbes, bacteria are the most dominant, accounting for up to 10¹¹ cells per
milliliter of rumen content. These bacteria primarily degrade carbohydrates like cellulose and
hemicellulose into volatile fatty acids (VFAs), such as acetate, propionate, and butyrate, which
serve as major energy sources for ruminants (Dai et al., 2023). Protozoa, though smaller in
number, also contribute to fiber digestion and play a key role in starch breakdown and microbial
regulation through predation (Cholewińska et al., 2020).

Fungi in the rumen, particularly anaerobic types, assist in disrupting lignin structures in plant cell
walls, thereby improving the accessibility of fibrous content to enzymatic digestion (Dai et al.,
2023). Meanwhile, archaea are primarily involved in methane production through the reduction
of carbon dioxide using hydrogen produced during fermentation. While this process helps
maintain a stable rumen environment, it also leads to energy loss and contributes to greenhouse
gas emissions (Denman et al., 2015).

The efficiency of rumen fermentation is influenced by several factors, including feed type,
quality, and composition, as well as animal-specific variables such as age and physiological
status. Improving this efficiency has become a major goal in ruminant nutrition, not just for
enhancing animal productivity, but also for minimizing environmental impacts, especially
methane emissions (Mizrahi & Jami, 2018).

The rumen hosts a complex microbial ecosystem comprising bacteria, protozoa, fungi, and
archaea. These microorganisms are integral to the fermentation process, breaking down feed
components into absorbable nutrients. The balance and diversity of these microbial populations
are influenced by various factors, including diet composition and environmental conditions.
Alterations in microbial communities can significantly impact fermentation efficiency and
overall animal health.

2.1.1 RUMEN pH

Rumen pH is perhaps the most fundamental parameter influencing the entire fermentation
process. An optimal pH range of 6.0 to 7.0 supports the activity of cellulolytic bacteria that
digest fibrous plant material (Chen et al., 2023). When pH falls below this range, particularly
under acidic conditions near pH 5.0, the growth and activity of fiber-digesting microbes decline,
leading to reduced fermentation efficiency. Zhang et al. (2022) found that changes in pH could
significantly shift the composition and activity of microbial communities, affecting both VFA
production and fiber breakdown.

2.1.2 AMMONIA NITROGEN (NH₃-N)

Ammonia nitrogen in the rumen results from the breakdown of dietary proteins and non-protein
nitrogen compounds. It serves as a nitrogen source for microbial protein synthesis. However, its
concentration must remain within an optimal range generally 5 to 25 mg/dL. Levels above this
can be harmful, especially under low-pH conditions. According to Chen et al. (2023), high
ammonia concentrations inhibited rumen microbial fermentation in vitro, but the degree of
inhibition was strongly dependent on the surrounding pH.

2.1.3 VOLATILE FATTY ACIDS (VFAs)

VFAs are the major end-products of microbial fermentation of carbohydrates in the rumen. They
include acetate, propionate, and butyrate, all of which are absorbed across the rumen wall and
used as energy by the host animal. The proportions of these VFAs vary with diet: forage-based
diets tend to produce more acetate, while concentrate-rich diets promote propionate production
(Zhang et al., 2022). Maintaining a balanced VFA profile is essential not just for energy supply,
but also for regulating rumen pH and microbial health.

2.2 WEST AFRICAN DWARF (WAD) SHEEP

The West African Dwarf or Djallonké is an African breed or group of breeds of domestic sheep.
It is the dominant breed of West and Central Africa (Ojo et al., 2019). This breed is primarily
raised for meat. The Cameroon or Cameroon Dwarf is a breed within this group (WAD/Benin,
2009; Jiwuba et al., 2017).

The West African Dwarf is generally white or piebald, the front half being black and the back
half white. However, skewbald (tan on white) and the black belly pattern are found, and the Kirdi
type are specially selected to be entirely black (Ofori and Hagan, 2020). Rams weigh
approximately 37 kg, have a well-developed throat ruff and are usually horned. The horns are
wide at the base, curve backwards, outwards and then forwards again, with a maximum of one
and a half coils. Ewes weigh about 25 kg and are usually polled (hornless), but may have slender
short horns (Bemji et al., 2018). The ears are short and pendulous, the neck is long and slender,
the chest is deep, the legs are short, the back is long and dished, higher at the withers than at the
tail-head, and the tail reaches the hocks (Whannou et al., 2021).

On average, ewes produce 1.15 to 1.50 lambs per lambing (Ojo et al., 2019). This breed grows
slowly as evaluated in Nigeria in the last 1970s. The overall growth rates from 0–90, 91–150 and
151–350 days old were 74, 49 and 34 g/day, respectively. This breed is also highly tolerant of
trypanosome. This breed is thought to go into oestrus throughout the year (Whannou et al.,
2021).

The West African Dwarf sheep is found in West Africa, its range extending from Senegal to
Chad, Gabon, Cameroon and the Republic of the Congo. It is adapted for life in humid forested
areas, sub-humid areas and savannahs. The Kirdi or Poulfouli is a wholly black variant found in
northern Cameroon and southwestern Chad (Bemji et al., 2018).

In Nigeria, WAD sheep are primarily reared for meat production, contributing significantly to the
nation's animal protein supply. They are integral to the socio-economic fabric of rural
communities, serving as a source of income, food security, and cultural value (Jiwuba et al.,
2017). The breed's adaptability to local conditions and low maintenance requirements make it an
ideal choice for smallholder farmers (Ouattara et al., 2021).

2.3 BOTANICAL AND NUTRITIONAL PROFILE OF Gmelina arborea

Gmelina arborea, commonly referred to as white teak or beechwood, is a fast-growing,

deciduous tree species that holds substantial value both ecologically and economically. It belongs
to the family Lamiaceae, formerly classified under Verbenaceae, and is native to tropical and
subtropical parts of Asia. Its rapid growth, adaptability to different soil types, and multipurpose
applications have made it a desirable species for reforestation, agroforestry, traditional medicine,
and animal nutrition (Anandalakshmi et al., 2020).

The taxonomic classification of Gmelina arborea is outlined below:

 Kingdom: Plantae

 Clade: Angiosperms

 Clade: Eudicots

 Clade: Asterids

 Order: Lamiales

 Family: Lamiaceae

 Genus: Gmelina

 Species: Gmelina arborea (Thaw Maw Moe, 2021)

This species underwent a taxonomic revision based on molecular phylogenetic studies which
reclassified it from the Verbenaceae family to Lamiaceae, aligning it with species exhibiting
similar floral morphology and secondary metabolite profiles (Arora & Tamrakar, 2017).

Gmelina arborea is naturally distributed across a wide range of tropical and subtropical Asian
countries such as India, Nepal, Sri Lanka, Myanmar, Bangladesh, Thailand, Laos, Cambodia,
Vietnam, and parts of southern China. Due to its environmental resilience and fast-growth
characteristics, it has been widely introduced to other parts of the world, including Africa
(notably Nigeria, Ghana, and Tanzania), Central America, and South America for agroforestry,
timber plantations, and soil conservation purposes (Anandalakshmi et al., 2020).

In Nigeria, Gmelina arborea is commonly planted as a reforestation species and windbreak in

plantations. It thrives particularly well in the southern and middle belt zones where rainfall is
adequate. The species performs best in well-drained, loamy soils and is tolerant of drought once
established. Its ability to grow in degraded or moderately acidic soils makes it suitable for
marginal land rehabilitation a significant advantage in tropical agricultural economies (Sangeeta
et al., 2017).
2.3.1 TRADITIONAL AND ETHNOMEDICINAL USES OF Gmelina arborea

In traditional medicine, particularly in the Ayurvedic, Unani, and Siddha systems, Gmelina
arborea holds a long-standing reputation for its therapeutic properties. Almost every part of the
plant—root, stem bark, leaves, fruits, and flowers—is utilized for medicinal purposes, which
reflects its pharmacological versatility. This wide range of ethnomedicinal applications is
supported by the plant’s rich phytochemical content, including flavonoids, lignans, tannins, and
iridoid glycosides (Anandalakshmi et al., 2020).

 Roots: Traditionally used for their purgative, anthelmintic, and anti-inflammatory

properties. Decoctions are often administered for treating fever, arthritis, and digestive
disturbances. In some cultures, the roots are also believed to have lactogenic properties,
thus being given to nursing mothers to stimulate milk production (Sangeeta et al., 2017).

 Leaves: The leaves are known for their anti-inflammatory and wound-healing properties.
Poultices made from crushed leaves are applied externally on bruises, cuts, and skin
irritations. Additionally, in rural communities in West Africa, the leaves are occasionally
fed to livestock due to their moderate protein content and digestibility (Arora &
Tamrakar, 2017).

 Fruits: The fruits are edible and used in traditional medicine to treat ulcers, skin diseases,
and respiratory ailments. Their mucilaginous properties make them useful in soothing
sore throats and digestive tract inflammations. Some studies suggest aphrodisiac effects
attributed to their lignan content (Anandalakshmi et al., 2020).

 Bark and Wood: Though more commonly used in construction due to its lightness and
workability, the bark is sometimes processed into herbal extracts for its antioxidant and
antimicrobial properties (Jafari et al., 2019).

 Flowers: Traditionally used for skin conditions like leprosy and dermatitis. They are also
known to possess mild laxative properties (Sangeeta et al., 2017).
2.3.2 PHYTOCHEMICAL CONSTITUENTS AND THEIR POTENTIAL EFFECTS ON
RUMEN MICROBES

Gmelina arborea., commonly known as white teak, is recognized for its rich diversity of
bioactive phytochemicals. Recent research has increasingly highlighted its potential in animal
nutrition particularly ruminant feeding systems due to compounds such as tannins, saponins,
flavonoids, phenolics, alkaloids, and glycosides (Arora & Tamrakar, 2017; Thaw Maw Moe,
2021). Among these, tannins and saponins have attracted special attention for their ability to
influence rumen microbial populations and fermentation dynamics.

2.3.2.1 TANNINS

Tannins are polyphenolic compounds known for their protein-binding properties. When included
in ruminant diets in moderate concentrations, they form complexes with dietary proteins,
reducing protein degradation in the rumen and allowing more undegraded protein to pass into the
small intestine for absorption (Adefalujo & Akiode, 2021). This improves nitrogen utilization
efficiency and may reduce ammonia nitrogen (NH₃-N) concentration in the rumen. However,
excessive tannin intake can have adverse effects, including reduced feed intake, nutrient
digestibility, and microbial activity (Ristasari, Natsir, & Dagong, 2019).

In a study conducted on West African Dwarf goats, supplementation with Gmelina arborea leaf
meal led to a significant reduction in rumen ammonia concentration, suggesting improved
microbial protein synthesis and reduced nitrogen loss (Adefalujo & Akiode, 2021). The study
further reported that digestibility parameters improved with Gmelina inclusion, emphasizing the
functional role of tannins when managed at beneficial levels.

2.3.2.2 SAPONINS

Saponins are glycosidic compounds that possess natural surfactant properties. In the rumen, they
are known to suppress protozoal populations, particularly the ciliate protozoa that harbor
methanogenic archaea responsible for methane production (Ristasari et al., 2019). By reducing
the protozoal load, saponins indirectly lower methane emissions while improving nitrogen
retention, as protozoa often consume valuable rumen bacteria involved in protein synthesis
(Jafari et al., 2019).
Research also supports the potential of saponins in Gmelina arborea to modify microbial balance
in favor of more efficient fermentation. Ristasari et al. (2019) demonstrated that goats fed diets
incorporating Gmelina leaves maintained stable rumen pH and volatile fatty acid (VFA)
concentrations while exhibiting reduced NH₃-N levels, implying improved fermentative
efficiency and less nitrogen wastage.

2.4 PLANT EXTRACTS IN RUMINANT NUTRITION

Methane emission from enteric fermentation in ruminants is a major concern, both for
environmental sustainability and livestock productivity. Methane represents an energy loss to the
animal (up to 12% of gross energy intake) and contributes significantly to greenhouse gas
emissions. In response, researchers have explored natural feed additives especially plant extracts
rich in secondary metabolites as eco-friendly alternatives to synthetic rumen modifiers (Patra &
Saxena, 2010).

Plant extracts modulate rumen fermentation and reduce methane production through several
mechanisms. Tannins, saponins, essential oils, and flavonoids can directly or indirectly influence
rumen microbial populations. Tannins, for example, form complexes with proteins and inhibit
methanogens and protozoa. Saponins disrupt protozoal membranes, indirectly reducing methane-
producing archaea due to their symbiotic relationships (Jafari, Ebrahimi, & Rajion, 2019).

Essential oils from plants such as oregano, thyme, and clove have also been shown to inhibit
methanogenic activity by altering the membrane integrity of microbes and interfering with
hydrogen metabolism in the rumen (Calsamiglia et al., 2007; Durmic et al., 2014).

Additionally, some phytochemicals shift rumen fermentation patterns towards greater propionate
production, which serves as a hydrogen sink and reduces the availability of hydrogen for
methane synthesis (Akanmu & Hassen, 2018).

Numerous studies have demonstrated the methane-reducing potential of plant extracts under both
in vitro and in vivo conditions:

Jafari et al. (2019) reviewed over a decade of studies and concluded that moderate inclusion of
tannin- and saponin-rich plant extracts significantly reduces methane production by up to 20–
30%, depending on dose and source.
Akanmu and Hassen (2018) investigated extracts from Azadirachta indica (neem), Moringa
oleifera, and Ocimum gratissimum and found significant reductions in methane production
during in vitro fermentation, along with stable pH and enhanced volatile fatty acid profiles.

Durmic et al. (2014) tested essential oils from oregano, garlic, and clove and reported up to 40%
reduction in methane output in vitro, while maintaining or improving rumen fermentation
profiles. Though not a terrestrial plant, Asparagopsis taxiformis, a red seaweed, has recently
gained attention. Kebreab et al. (2018) showed that its inclusion at less than 2% of dry matter
intake could cut methane emissions by up to 50% in dairy cattle trials.

2.4.1 MERITS AND POTENTIAL DEMERITS OF USING METHANOLIC EXTRACTS

IN RUMINANT NUTRITION

Methanolic extracts, derived through the use of methanol as a solvent, have garnered significant
attention in ruminant nutrition due to their efficacy in extracting a broad spectrum of
phytochemicals (Anandalakshmi et al., 2020). These extracts have been investigated for their
potential to modulate rumen fermentation, enhance feed efficiency, and mitigate methane
emissions.

ADVANTAGES

 Broad-Spectrum Phytochemical Extraction: Methanol is a polar solvent capable of

extracting a wide range of bioactive compounds, including flavonoids, tannins,
saponins, and alkaloids. This comprehensive extraction ensures that multiple
phytochemicals with synergistic effects are available for modulating rumen
fermentation (Alem, 2024).
 Methane Mitigation: Studies have demonstrated that methanolic plant extracts can
significantly reduce methane production in the rumen. For instance, methanolic
extracts of Lawsonia inermis, Myristica fragrans, and Capsicum annuum have been
shown to decrease methane emissions during in vitro fermentation without adversely
affecting pH or protozoal populations (Open Vet J, 2012).
 Enhanced Fermentation Characteristics: The inclusion of methanolic extracts from
herbs and spices such as Azadirachta indica, Moringa oleifera, and Ocimum
gratissimum has been associated with improved ruminal fermentation profiles,
 including increased gas production and organic matter digestibility in West African
Dwarf sheep (PubMed, 2021).
 Antimicrobial Properties: Methanolic extracts possess antimicrobial activities that can
modulate the rumen microbial ecosystem. This modulation can lead to a reduction in
pathogenic microbes and an enhancement of beneficial microbial populations, thereby
improving overall rumen health and function (Alem, 2024).

POTENTIAL DEMERITS

 Variability in Phytochemical Composition: The concentration and efficacy of

bioactive compounds in methanolic extracts can vary based on factors such as plant
species, geographical location, harvesting time, and extraction methods. This
variability can lead to inconsistent results in rumen fermentation modulation (Alem,
2024).
 Potential Antinutritional Effects: While methanolic extracts can reduce methane
emissions, some compounds may also inhibit fiber digestion or reduce feed intake if
not properly dosed. For example, high concentrations of certain extracts have been
associated with decreased dry matter digestibility (Open Vet J, 2012).
 Residual Solvent Concerns: The use of methanol as an extraction solvent raises
concerns about residual solvent presence in the final extract. Methanol is toxic, and
its residues can pose health risks to animals if not adequately removed during the
extraction process (Alem, 2024).
 Limited In Vivo Studies: Most studies on methanolic extracts have been conducted in
vitro. There is a need for more in vivo research to confirm the efficacy and safety of
these extracts under practical feeding conditions (PubMed, 2021).
 CHAPTER THREE

3.0 MATERIALS AND METHOD

3.1 EXPERIMENTAL SITE AND DURATION

The experiment was conducted at the Teaching and Research Farm of the Federal College of
Animal Health and Production Technology, Ibadan, Nigeria. The study spanned a total of 56
days, equivalent to 8 weeks.

3.2 SOURCE AND PREPARATION OF TEST INGREDIENT

Fresh leaves of Gmelina arborea were harvested from mature trees within the college premises.
The leaves were air-dried under shade to preserve their phytochemical integrity, as recommended
by Abiola-Olagunju et al. (2024). Once adequately dried, the leaves were ground into a fine
powder using a laboratory mill.

3.2.1 EXTRACTION

Crude methanol (70%v/v) extraction

20 kg of the dried blended plant sample was transferred into glass container and 75L of 70%v/v
methanol was added, stirred at every 2hrs and allowed the extraction process to take place for
72hrs. The solvent (now containing the extract) was collected using muslin bag and the filtrate
was further filtered using Whatman filter paper 1mm. This process was repeated on the shaft
with another 75L of 70%v/v methanol. The combined filtrate was then concentrated with the aid
of rotary evaporator (Heidolph laborota 400 effiecient, made in Germany, model 517-01002-002)
set at 400C, after which the concentrate was further concentrated using a vacuum oven set at
400C with a pressure of 700mmHg.

3.3 EXPERIMENTAL ANIMALS AND MANAGEMENT

Twenty (20) healthy West African Dwarf (WAD) sheep, weighing up to 11-14kg, were procured
from a reputable sheep dealer at Akinyele market, Ibadan. Before the arrival of the animals, the
pen was washed, cleaned and disinfected to avoid the growth of microorganisms. Upon arrival,
the animals were neck tagged for easy identification and placed into individual pen. The animals
underwent a 14-day acclimatization period during which they were monitored for any health
issues and were given profilactic treatment. Post-acclimatization, the sheep were randomly
assigned to four treatment groups, each comprising five animals:

 Treatment 1 (Trt 1): Control group receiving no Gmelina arborea methanolic extract.

 Treatment 2 (Trt 2): Received 10 ml/day of Gmelina arborea methanolic extract.

 Treatment 3 (Trt 3): Received 20 ml/day of Gmelina arborea methanolic extract.

 Treatment 4 (Trt 4): Received 30 ml/day of Gmelina arborea methanolic extract.

The extracts were administered orally using a dosing syringe once daily in the morning before
feeding. This dosing regimen was adapted from methodologies employed in studies assessing the
impact of plant extracts on ruminant performance (Akanmu & Hassen, 2021).

3.4 EXPERIMENTAL DIET

Same basal diet and concrete was used across all treatment

3.5 EXPERIMENTAL DESIGN

A Completely Randomized Design (CRD) was employed for this study. The 20 sheep were
randomly allocated to the four treatment groups, ensuring that each group had an equal number
of animals. This design facilitates the unbiased assessment of treatment effects on the measured
parameters.

3.6 DATA COLLECTION

Data will be collected on the following parameters rumen fermentation parameters and methane
emission:

Rumen fluid sample (40mL) was collected at the end of the feeding trial from three (3) randomly
selected animals per treatment. The method of collection was done through the use of suction
tube thrust into the rumen compartment as described by Babayemi and Bamikole (2006). As
soon as the sample was obtained, rumen pH was determined within two minutes of collection
with the use of digital pH meter. The digital pH meter was stabilized in distilled water with
specific pH recommendation before being used for the reading. 20mL of the rumen fluid sample
was stored in 40mL 10% formal saline prior to the direct microscopic counts of rumen bacteria.
While the other 20mL sample of rumen fluid was bulked for each animal before being made free
of coarse particles by filtration with cease cloth. Thereafter, 5mL sample of the filtrate was then
acidified with 1mL of alpha 5%(v/v) orthophosphoric acid solution and stored frozen in the air
tight plastic bottle container for determination of volatile fatty acid concentration and its fraction.
The other 15mL of the filtrate sample was added to 10% sulphuric acid solution before there
were stored for analysis of ammonia nitrogen concentration

Methane production was estimated using the in vitro gas production technique, as described by
Elghandour et al. (2012), to assess the impact of the methanolic extract on enteric methane
emissions.

3.7 Chemical Analysis

Proximate analysis of feed samples was determined according to AOAC, (2002), while fibre
fractions were determined according to Van Soest et al., (1999)

3.8 Statistical Analysis

Data collected will be subjected to analysis of variance (ANOVA) using the Statistical Package
for the Social Sciences (SPSS) version 25.0. Differences among treatment means were separated
using Duncan's Multiple Range Test at a 5% significance level. This statistical approach is
consistent with methodologies adopted in similar studies evaluating the effects of dietary
interventions on ruminant performance (Adebisi et al., 2016).
Table 1: Gross and Chemical Composition of the Experimental Diet

Ingredients %
Cassava peel 50.00
Soya bean 14.00
Wheat offal 22.00
Palm kernel cake 11.00
Premix 1.00
Limestone 1.00
Salt 1.00
Total 100.00
Calculated analysis
Metabolized energy (Kcal) 2274.05
Crude protein % 13.80
Crude fibre % 8.60
 CHAPTER FOUR
4.0 RESULTS AND DISCUSSION
4.1 RESULTS
4.1.1 The Effect of Methanolic Gmelina Arborea Leaf Extract on Rumen Fermentation
Parameters of West Africa Dwarf Sheep
The Effect of Methanolic Gmelina Arborea Leaf Extract on Rumen Fermentation Parameters of
West Africa Dwarf Sheep is shown in table 2. AGALE had significant (P<0.5) influence on the
rumen fermentation parameters of WAD sheep. Animal administered 10m1 of MGALE recorded
the highest values, for all parameters except rumen pH, temperature and Ammonia Nitrogen
(NH3N) while animal administered 20ml, 30ml of MGALE recorded same least values with the
exception of control respectively. The total volatile acids and acetic acid of this study ranged
from 62.78 to 102.20 mmole/100ml and 4.42 to 7.20 mmole/100ml respectively.
4.1.2 The Effect of Methanolic Gmelina Arborea Leaf Extract on Microbial Counts of West
Africa Dwarf Sheep
The Effect of Methanolic Gmelina Arborea Leaf Extract on Microbial Counts of West Africa
Dwarf Sheep is shown in figure 3. MGALE had significant (P<0.05) influence on the Microbial
count of WAD sheep. Animal administered 10ml of MGALE recorded the highest values of total
bacteria count and lowest values of total colony count except protozoan and total fungi count.
However, animal administered 20ml of MGALE recorded the highest values of total colony
counts respectively.
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