1.

1 Overview

Human communities have always had a close relationship with their surroundings
and have used natural resources for both healing and nutrition. Through trial and
error, the knowledge of and use of plants for food and medicine developed. As time
goes by, understanding of medicinal herbs have been passed down through the
generations, aiding in the advancement of civilizations. Although many
civilizations have long used medicinal herbs, there is growing concern worldwide
about the safety and effectiveness of these plants (Rao et al., 2007). Healthcare may
enter a new phase with the standardization and evaluation of plant-derived
chemicals' health benefits. It is essential to preserve traditional knowledge about
medicinal plants in order to appropriately utilize natural resources and identify
possible advantages. Herbal remedies have an important role in both conventional
medicine and trade, meeting the demands of distant markets engaged in the
development of novel drugs. It takes training researchers and practitioners in both
modern and traditional therapeutic techniques connected to plant chemicals to
successfully integrate plants into healthcare (Ahad B et al., 2021). Throughout
human history, these herbal medicines have developed into complex sciences in a
variety of societies. Since ancient times, people have used medicinal herbs as a
natural form of healing. Even though more than 50,000 plant species are used to
make medications and cosmetics, their distribution around the world varies.
Medicinal plants are essential for the production of drugs because of their
therapeutic qualities, which may be found in their seeds, roots, and leaves, among
other sections (Abd et al., 2016). Certain diseases can be prevented and severe
illnesses like cancer can be treated with substances obtained from plants. Concerns
over the toxicity and negative consequences of conventional drugs are the reason
behind the rising demand for herbal therapies. It is essential to understand the
historical background of medicinal plants in order to appreciate and advance this
file, solve present issues, and influence future advancements (Kia et al., 2017).

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1.2 Medicinal plants

The most ancient kind of medicine is derived from medicinal plants, which have
been utilized for thousands of years in traditional medicine throughout many
nations. Over the years, empirical information on their positive benefits has been
passed down among human groups. Any plant with one or more chemicals or
organs that can be used medicinally is considered medicinal. Chemical components
are present in every part of the plant, including the leaves, roots, rhizomes, stamps,
and barks. These bioactive substances that are present in plants but are not nutrients
are sometimes called "phytochemicals" or non-nutritive plant compounds.
Depending on a number of variables, including the degree of accumulation,
location, manner of collection, extraction method, and amount of plant material
utilized, these bioactive components may be present or missing (Hamburger et al.,
1991). Phytochemicals found in many plants have medicinal or disease prevention
properties. Plants have been shown to be a great source of natural compounds for
maintaining human health as research on natural medicines has grown. Many of the
more than 150,000 plant species that have been researched have been found to have
therapeutic chemicals. Historically, sources of pharmacological lead compounds
have been mostly derived from medicinal plants. Because early people were guided
by sensation, taste, and learning, they relied on plants to alleviate their ailments
(Yuan H et al., 2016).

1.3 History of Medicinal Plants

Throughout history, humans have examined nature's gifts for possible treatments
against diseases, depending first on instinct when employing medical plants and
animals. Due to a lack of understanding about plant characteristics and disease
causation, early techniques relied heavily on practical learning. As knowledge about
the unique advantages of certain plants for healing ailments grew, empirical
methods gave way to evidence-based treatments. While iatrochemistry brought

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about a change in the 16th century that favored synthetic medications, the problems of
declining effectiveness and expanding contraindications in the present have
brought about a resurgence of interest in natural medicines (Petrovska, 2012).

1.3.1 Prehistoric Medicine

Prior to the development of reading and writing by humans, medicine was practiced
throughout history and varied greatly depending on the area and culture.
Anthropologists use the analysis of artifacts and human remains to generate
informed assumptions about the medical practices of the ancient past.
Understanding historical medicine involved combining natural and supernatural
explanations for various ailments; there was no comparable point of reference,
unlike the placebo effect in modern clinical studies. Trial and error may have been
included, but factors like coincidence, lifestyle, family history, and the placebo
effect that are taken into account in contemporary research are probably not taken
into consideration. It's still unclear exactly how ancient cultures understood how the
human body functions (Neil, 2020).

1.3.2 Ancient Egyptian

A diverse array of plants was used by the ancient Egyptians for therapeutic reasons.
They used their extensive knowledge of the characteristics of numerous plants and
herbs to cure a wide range of illnesses. Historical medical practices and herbal
medicines are described in great detail in medical papyri, such as the Edwin Smith
Papyrus and Ebers papyrus. Ancient Egyptians were well-versed in the botanical
characteristics of plants. They used plants in a variety of preparations, including as
ointments, poultices, and potions, after realizing the advantages of certain herbs. For
their therapeutic qualities, plants including opium poppies, garlic, and aloe vera
were often used (Petrovska, 2012).

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1.3.3 Ancient Roman

Greek and Egyptian medicine had a great effect on the Romans. Many herbal
medicines and information about medicinal plants were taken up and modified by
them from these civilizations. Particularly important were the writings of classical
Greek doctors like Hippocrates and Galen. Romans were renowned for their vast
gardens, which included gardens filled with therapeutic herbs. They recognized the
value of having a ready supply of herbs, so they cultivated a variety of plants for
medical uses. These gardens were often connected to temples, which used medicinal
plants in ceremonial and therapeutic contexts (Srivastava, 2018). The extensive
trade channels of the Roman Empire enabled the movement of products, including
medicinal plants, across various locations. Roman pharmacopeia included exotic
plants from the Mediterranean, the Middle East, and even Asia. Romans created a
wide range of medicinal concoctions and ointments, such as herbal infusions and
decoctions. They improved the processes for removing plant-based medicinal
substances, and they often blended many components together to create treatments.
Other Roman doctors and academics, including Pliny and Galen, wrote medical
treatises and manuscripts that included knowledge on therapeutic plants. These
writings included instructions on how to identify, gather, and use different plants for
medicinal reasons (Halberstein, 2005).

1.3.4 Ancient Greece

Herbalism was greatly influenced by the works of ancient Greek doctors such as
Hippocrates, Dioscorides, and Theophrastus. They recorded the uses of plants,
categorized them according to their therapeutic qualities, and developed the idea of
Materia medica, a catalog of materials used in medicine. The exploration and
colonization of several locations by the ancient Greeks is well known, and this
contributed to the interchange of botanical knowledge. They integrated information
about medicinal plants from several civilizations into their own customs. Greek

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doctors abandoned magical and superstitious ideas in favor of a more methodical
and logical approach to treatment. They contributed to a more scientific knowledge
of medicinal herbs by emphasizing observation, experimentation, and empirical data
(Carod, 2013).

1.3.5 Chinese Traditional Medicine

Traditional Chinese medicine is one of the oldest medical specialties. Chinese

herbal formulas are made up of carefully chosen ingredients that complement one
another. Herbs and herbal preparations are used in traditional Chinese medicine to
enhance organ function and promote health. A traditional Chinese medicine
practitioner uses diagnostic techniques such as smell, hearing, voice vibration,
touch, and heartbeat to determine the source of an unbalanced health condition, the
organ linked to it, and the meridians affected. By observing the functioning of
natural law, ancient healers recognized the five essential components of the
universe: wood (mu), fire (huo), earth (tu), metal (jin), and water (shui). They also
determined that there are several visible and hidden correspondences between these
components (Cheng J.T et al., 2000).

1.4 Characteristics of Medicinal Plants

Medicinal plants are valuable due to their medicinal and therapeutic qualities
because of a variety of attributes. The following are some essential qualities of
therapeutic plants:

 Bioactive Compounds: Alkaloids, flavonoids, terpenes, and phenolic

compounds are examples of bioactive substances found in medicinal plants
that may be good for human health. These substances frequently have
particular pharmacological effects.

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  Therapeutic Properties: The unique medicinal qualities of medicinal plants
are well known, and they can have effects ranging from antibacterial and
antioxidant to analgesic and anti-inflammatory. Different plants have
varying therapeutic qualities.
 Traditional Use: Many ancient medical systems, such as Ayurveda, ancient
Chinese Medicine and indigenous medicine, have traditionally used a
variety of therapeutic plants. Generation after generation has passed down
this customary wisdom (Rasool, 2012).

1.4.1 Utilization Parts of the Plants in Medicine

Plants have long been used in traditional medicine, with various portions providing
unique medicinal properties. Certain roots, such as valerian and ginger, have
calming and anti-inflammatory qualities, respectively. Essential oils and other
substances found in leaves, such as those of eucalyptus and Ginkgo biloba, support
respiratory health and cognitive function. Elderflower and chamomile are two
popular flowers for their immune-stimulating and relaxing properties. Flaxseed and
fennel seeds, for example, provide omega-3 fatty acids that are beneficial to
cardiovascular health and assist with digestion. Willow and cinnamon bark have
anti-inflammatory and analgesic qualities. Aloe vera and Fo-ti are two examples of
stems that provide skin healing and possible anti-aging benefits. Resins with
antibacterial and anti-inflammatory properties include frankincense and myrrh.
Whole plants with anti-inflammatory and perhaps depressive qualities include St.
John's Wort and turmeric. The many ways that plant components are used highlight
the wide range of pharmacological diversity that exists in nature (Purwanti, E et al.,
2020).

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 Figure 1.1: Plant parts Use in Medicine

1.4.2 Importance of Medicinal Plants

A number of phytochemicals included in plant-based diets may be able to delay the

beginning of diseases including cancer and heart disease, according to recent
studies. Over 3000 plant species have been shown to have anticancer
characteristics, and over 25% of all recommended therapies worldwide are plant-
based. Conventional plant-based medications provide around 80% of the majority of
people's fundamental medical requirements in developing nations. The idea or
foundation for many pharmacological treatments comes from compounds that may
be found in medicinal plants. Natural substances have served as the basis for the
development of several modern pharmaceuticals, including painkillers, antibiotics,
and anticancer drugs. Since they function as dietary supplements to prevent
diseases in the human body and have fewer side effects when used to treat
illnesses, herbal drugs are natural items that have been demonstrated to be safe
(Harriet U et al., 2020).

Herbal products are now available as pills, capsules, powder, dried or fresh herbs,
and tea extracts in response to customer demand. People take herbal treatments
without a prescription because they are usually believed to be safe. However, some
may cause health problems, while others are useless and conflict with other
treatments. To find the concentration of their active ingredients in the present state
of affairs, standardizing herbal products and assessing the quality of medicines
seem to be crucial. Even after many legislative attempts to curb the practice in an
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attempt to boost profits, large-scale illegal medicinal plant collection continues.
Because the species in question is considered endangered, collecting it is prohibited
(Aslam et al., 2016).

Due to their elite populations, availability of propagules and high-quality

production, value addition, and guaranteed markets, Aconitum heterophyllum and
Picrorhiza kurrooa received particular attention in the technical sessions of a
workshop on Himachal Pradesh's endangered medicinal plant species. Even though
Swertia chirata is one of the finest plants at high altitudes, its extended gestation
time and meticulous field management precluded it from being planted right away
(Raina, H et al., 2016).

1.4.3 Drug Development Process

The approval of new medications is declining, and the prices of new medications
are rising, which is making new drug innovation very challenging. The advent of
combinatorial chemistry gave rise to expectations of higher national chemical entity
success rates; however, this scientific progress hasn’t affected the success rate of
new medication development, however. This circumstance has prompted us to
create a novel integrated drug discovery approach in which the creation of natural
drugs may coexist peacefully with Ayurvedic expertise. One of the initial steps in
the creation of new medications is the identification of the national chemical entity,
which may be generated by chemical synthesis or isolated from natural materials
via biological activity-guided fractionation. Numerous innovative drugs and the
active ingredients in them originate from diverse sources (Kayser, O et al., 2018).

The steps involved in developing drugs from plants are outlined below:

 Selecting and correctly recognizing the appropriate medicinal herb.

 Crude extraction with the right solvent.
 Developing an assay that permits the identification of the bioactive elements and
the removal of such inactive components.

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  Determining the bioactivity of crude extract.
 All crude extract fractions were physiologically evaluated and active separation
was used, using the finest chromatographic techniques.
 The bioactive components are separated repeatedly in order to isolate pure
molecules.
 Using spectroscopic methods to make pure substances' chemical structure visible.
 Examining how pure chemicals affect biology.
 Testing for toxicity using pure chemicals.

1.5 List of Key Medicinal Plants Species and Its Uses

Table 1.1: List of Key Medical Plant Species and Its Uses (Rahman et al., 2014)

Plant Scientific Name Local/Tribal Parts Medical Uses

and Family Name
Used

Adansonia digitata Baobab/ Leaf, Antimalarial

L. root,
Monkey-bread
Family: flower
Tree
Bombacaceae

Adhatoda vasica Basak, Basak Root, Expectorant and

Pata
Family: Acanthaceae leaves bronchodilator

Holorheena Kurai (S) Bark, Scabies,

Antipyretic,
antidysentrica Seed,
Amoebic
Family: Flower,
dysentery
Apocynaceae Leaves

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Phyllanthous Bhumi Amla Whole Anemic,
Jaundice,
amarus Plant
Dropsy.
Family:

Euphorbiaceace

Bidens Shumophul Flowers To treat acne

sulphurea
and leaves
Family:
Asteraceae

Peeper longum Long peeper / Fruit, Appetizer,

Family: Pippali Root enlarged spleen ,
Piperaceae Bronchitis, Cold,
antidote.

Gymnema Gudmar / Leaves Diabetes,

sylvestre Madhunashini hydrocil, Asthma.
Family:
Asclepiadaceae

Tinospora Guluchi Stem Gout, Pile,

cordifolia general debility,
Family: fever,
Menispermaceae Jaundice.

Swertia chiraita Chirata Whole Skin Disease,

Family: plant Burning,
Gentianaceae censation, fever.

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1.6 Phytochemistry in Medicinal Plants

Derived from the Greek term phyto, which means "plant," phytochemicals are
naturally occurring, physiologically active compounds found in plants that provide
health advantages for humans that go beyond those of macronutrients. and smaller
nutritional components. They enhance the plant's color, flavor, and taste while
shielding it from harm and illness. Plant substances known as phytochemicals shield
plant cells against environmental threats such pollution, stress, drought, UV ray
exposure, and viral attack. It has been publicly accepted recently that they have a
responsibility to protect human health when it comes to nutrition. Over 4,000
phytochemicals have been classed based on their chemical, physical, and defensive
qualities, and about 150 phytochemicals have received extensive investigation.
Among the many phytochemicals found in food, while it's well accepted that plants
make these compounds to defend themselves, recent research has shown that a
number of phytochemicals may also shield humans from disease. Despite not
seeming to be essential nutrients or anything the body needs to sustain life,
phytochemicals do possess certain significant properties that may aid in the
treatment of a number of common illnesses. Because of this characteristic, some of
those advantages suggest that phytochemicals may play a part in the prevention and
treatment of illness; several researches are being carried out to determine the
positive effects of phytochemicals on health (Kurmukov, 2012).

1.6.1 Alkaloids

Alkaloids are a broad class of bitter-tasting, possibly dangerous compounds that are
present in many medicinal plants. They have different mechanisms of action and
are employed as recreational and therapeutic medications. Several well-known
alkaloids and their sources of plants include (Rehman, 2021):
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  Atropine, hyoscyamine, and scopolanide, which are derived from
certain nightshade species.
 Berberine, which comes from plants such as Mahonia and Berberis.
 Coffea arabica produces caffeine.
 Erythroxylum coca cocaine
 Ephedra sinica produces ephedrine.
 Papaver somniferum morphine
 Nicotiana tabacum produces nicotine.

1.6.2 Glycosides
Anthraquinone glycosides are found in a number of medicinal plants (MPs),
including cascara, rhubarb, senna, and Alexandrian. Plant-based laxatives like aloe
vera, senna, and rhubarb may be derived from MPs. Furthermore, digoxin and
digitoxin are cardiac glycosides, powerful medications made from MPs like foxglove
and lily. These substances function as diuretics and support normal cardiac rhythm
maintenance (Rehman, 2021).

1.6.3 Terpenes
In addition to resinous plants like conifers, a wide variety of therapeutic plants also
contain terpenes and terpenoids in different forms. These substances have an
overpowering scent that naturally deters herbivores. Certain plant extracts, including
those from lavender and rose, are useful for use in perfumery and aromatherapy
because of their pleasant scent. Additionally, several terpenes and terpenoids have
therapeutic qualities, such as the thymol's antiseptic properties have long been used
as a vermifuge and an anti-worm medication (Rehman, 2021).

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1.6.4 Tannins

One kind of polyphenol that has antiseptic and antibacterial properties is tannins.
They were identified in fifteen distinct plant species and were the most common
chemical in a wide variety of plants in Wadi Yalamlam, a major Saudi Arabian
area. Aerva javanica, Spergula fallax, Cleome hanburyana, and other plants are
examples of plants that contain tannin (Al Shehri et al., 2022).

1.6.5 Resins

Plant byproducts called resins, which are not soluble in water, have a tendency to
harden when exposed to air. Woody plants like Commiphora myrrha, whose water-
based extract has antibacterial activity against a variety of pathogenic
microorganisms, are widely used to make them. Furthermore, according to Al
Shehri et al. (2022) additional plants that also generate resins include Zingiber
officinale and Alpinia galanga.

1.6.6 Flavonoids

Flavonoids are low molecular weight secondary metabolic chemicals that are
extensively distributed throughout the kingdom of plants. They build up in the plant
cellular vacuoles and are responsible for the fruit and flower color and scent. These
substances function as both antibacterial defenses and detoxifiers. They are also
widely distributed in many plant-based meals and drinks, including tea, chocolate,
wine, fruits, and vegetables. For these reasons, they are known as dietary
flavonoids.
Essential components found in onions, especially quercetin and its derivatives, a
subclass of flavonoids, play a major role in the advantageous bioactive
characteristics of onions and their derivatives. Similarly, broccoli (Brassica

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oleracea), a vegetable well-known for its health advantages, has a large number of
secondary metabolites, such as glucosinolates and flavonoids, with the blossom
head being the main edible portion (Al Shiehri et al., 2022).

1.6.7 Saponins

Saponins are heterosides made up of a liposoluble triterpene or steroid structure and

a water-soluble carbohydrate chain. They are widespread in several plant groups,
such as Saponaria officinalis and Glycyrrhizia glabra of the Fabaceae family.
Flavonoids, saponins, mucilaginous materials, and resinous components are all
present in the whole Saponaria officinalis plant. Alfalfa and Yucca schidigera are
two examples of plants that contain saponins and have been utilized as feed
additives in ruminant production to improve growth, milk, or wool production (Al
Shehri et al., 2022).

1.6.8 Volatile Oils

The main terpenoids, low molecular weight aliphatic hydrocarbons, acids, alcohols,
aldehydes, acyclic esters or lactones, nitrogen, sulfur-containing chemicals,
coumarins, and homologs of phenylpropanoid are all found in complex
combinations known as volatile oils. These oils' terpene content supports the
culinary, medicinal, and aromatic applications of many plants. Different sections of
different plants contain these essential oils. For example, Nigella sativa is extracted
from the seeds of Cinnamomum verum, Salvia officinalis is extracted from the
leaves of the plant, and Caryocar villosu is extracted from the stems. The mature
leaves, fruits, immature flowers, and stems of Eucalyptus oleosa produce volatile
oils. Beyond being used in food flavorings, hygiene, and fragrances, volatile oils
have several uses. They may be applied as herbicides or insecticides as well.
Studies have indicated that volatile oils extracted from plants like hairy basil
(Ocimum americanum), citronella grass (Cymbopogon winterianus), kaffir lime
(Citrus hystrix), and turmeric (Curcuma longa) have been shown to have notable

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effects on three mosquito vectors: Aedes aegypti, Anopheles dirus, and Culex
quinquefasciatus (Al Shehri et al., 2022).

1.6.9 Fats and Lipids

Triglycerides and high fatty acids are among the important categories of fats and
lipids. Unlike volatile oils, these compounds are usually present in plants as fatty
droplets and cannot be distilled without first decomposing. They also neither
evaporate nor volatilize. Saturated fats may raise blood HDL-cholesterol levels in
an efficient manner without significantly altering the total cholesterol/HDL-
cholesterol ratio, according to scientific study. One important ingredient, palmitic
acid, which is made from palm tree oil, lowers the risk of cardiovascular disease by
performing crucial structural and functional tasks throughout fetal development and
infancy. Human breast milk is the only source of this particular acid. Furthermore,
a wide range of industrial and medical uses employ castor oil, which is derived from
the seeds of Ricinus communis (Al Shehri et al., 2022).

1.7 Medicinal Plants in Bangladesh

The traditional medicine of Bangladesh, a biologically diverse nation, has

traditionally relied on medicinal plants to treat a wide range of illnesses and provide
other health advantages. Bangladesh may include a large number of naturally
occurring medicinal plants that are essential to traditional medicine. Currently, the
most popular kind of traditional medicine, both domestically and globally, is herbal
therapy. They are respected since they have no bad effects. Furthermore, a vast
variety of medicinal plants are grown by farmers in the Mymensingh, Tangail,
Sylhet, Madhupur, Kushtia, and Chittagong Hill Tracts. The indigenous people of
Bangladesh have traditionally depended on traditional healers to treat a variety of

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ailments. The main source of healing for these healers is medicinal herbs.There are
many different kinds of medicinal plants in Bangladesh. Currently, this country is
home to over a thousand distinct plant species that are known to have therapeutic
value (Khan, 2016). Many of these plants have been the subject of chemical,
pharmacological, and toxicological experiments to identify the components or
compounds that may be bioactive; these investigations are currently ongoing. Of
these, 546 have recently been shown to be helpful in therapy, despite the fact that
over a thousand have been linked to being poisonous when taken medicinally
(Polash et al., 2017).
1.7.1 Classification of Medicinal Plants According to The Habitats in
Bangladesh
Table 1.2: List of medicinal plants grow in different habitats (Sadi, 2012)

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 SL Plant Type Habitats Type Example
:
No.

1. Forest and Deciduous Nallota, Nimaada, Nota,

Hill tracts forest, Hill
Bormasri
plants track forest

2. Coastal areas Coastal area Koromcha, Sadormala,

plants and
Sadakolmi
Mangrove forest

3. Plain land and Plain land and Kalomegh, Shiakata,

crop land plants Ghagra, Nallota
cultivate land

4. Fresh Water Side of the Sadapata, Hizor, Halencha,

and water canal, hill Jolkolmi, Sapla, Saluk,
logging land Boure, Doba Padma
plant

1.7.2 Bangladesh Market for Medicinal Herbs, Their Production

Bangladesh has a long history of employing plants for medical purposes in
Ayurvedic and Unani medicine systems. The market's demand for medicinal plants

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is influenced by the cultural importance of herbal treatments. Bangladesh's varied
topography and climate encourage a large range of plant species with possible
therapeutic uses. The biodiversity of the nation offers a base for the production and
gathering of therapeutic plants. In Bangladesh, traditional medical practices are
common, and a large number of people use herbal treatments to treat a variety of
illnesses. This generates a steady market for therapeutic plants. It is crucial to
guarantee the uniformity and quality of medicinal plants for both local and foreign
markets. Observing quality control procedures and good manufacturing practices
(GMP) is essential. Working together, the public, commercial, and academic sectors
may promote innovation and result in the discovery of novel therapeutic plants as
well as enhanced growing methods (Jagaran., 2023).

Figure 1.2: Herbal Medicine Market in Bangladesh (Jagran, 2023)

1.7.3 Future Possibilities for Medicinal Plants

Ongoing investigation of therapeutic plants in search of novel bioactive substances

and potential medications. Technological developments in areas like metabolomics,
genomics, and artificial intelligence to speed up the discovery and creation of
medicinal substances. Using medicinal plants into personalized medicine strategies
to customize care according to a patient's genetic, lifestyle, and environmental
characteristics. Creation of specialized medications made from certain plant

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chemicals. synthesis of biopharmaceuticals, such as vaccinations, antibodies, and
therapeutic proteins, using plant-based systems. Application of plant biotechnology
for the manufacture of complicated medicinal molecules. Expansion of functional
foods and nutraceuticals utilizing medicinal plant extracts for preventative health
and wellbeing. Creation of dietary supplements made of plants to target certain
health issues. Integration of traditional medical methods into modern healthcare
systems, with a focus on evidence-based treatments. Cooperation for all
encompassing patient care between contemporary healthcare providers and
traditional healers. Developments in bioengineering to produce pharmaceuticals
using synthetic biology techniques. Genetic alteration of plants to promote the
production of certain medicinal chemicals (Mohammad, 2018).

1.8 Increased Demand of Medicinal Plants

Natural and traditional medicines are becoming more and more popular than
manufactured medications. There is a growing market for medicinal plants as a
result of individuals looking for alternatives with less negative consequences.
People are becoming more interested in holistic and preventative healthcare
practices as they place a greater emphasis on wellbeing and health. Many people
believe that using medicinal plants to treat a variety of health issues is a more all-
natural and comprehensive approach to maintaining health. Globally, traditional
medical systems like Ayurveda, Traditional Chinese Medicine (TCM), and native
healing methods have become more and more well-liked. The demand for certain
plant species is driven by these systems, which often depend on the usage of
medicinal herbs. In order to produce new drugs, the pharmaceutical industry is
looking more and more into natural substances originating from plants. The demand
for certain plant extracts is rising as a result of several pharmaceutical firms funding
research and development aimed at creating plant-based medications. Estimates of
the size of the herbal medicine business in five years are shown in Figure, assuming

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that the current simple growth rate for herbal practitioners, self-treatment, and the
Ayurvedic and Unani sectors would continue. It is projected that the market for
herbal medicines will increase from Tk 3,700 million in 2003 to Tk 5,850 million
in 2008 at current market values. These growth rate conjectures make sense.
Because of the robust government assistance, market liberalization, and corporate
dynamism, these growth rates ought to be exceeded (Mohammad, 2018).

1.9 WHO's Policy on Herbal Medicines

The World Health Organization promotes the use of medicinal plants and their
byproducts because it recognizes the importance of herbal treatments to many of its
Member States. Early in 1978, the World Health Assembly, the highest authority of
the World Health Organization, recognized the role of medicinal plants in the
healthcare system when making decisions on drug administration and policy. The
World Health Assembly recommended establishing protocols and guidelines for
proving the safety and efficacy of therapeutic herbs, compiling a list of beneficial
plants, and distributing relevant information in order to organize efforts. Three
further resolutions on the categorization, assessment, preparation, cultivation,
application, management, and preservation of medicinal plants were enacted in
1987, 1988, and 1989. In light of those resolutions, the WHO's position on herbal
medicine might be summed up as follows:
 The World Health Organization (WHO) recognizes the importance of herbal
treatments for the well-being of a considerable global population. It is
recommended to utilize herbal medicines sparingly since they are valuable
and conveniently available resources.
 To encourage the proper use of medicinal herbs, a comprehensive program
for their identification, evaluation, preparation, development, recognition as
significant and readily available resources, and their right use is advised.

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  It is crucial to carry out a comprehensive stockpile and evaluation
(preclinical as well as clinical) of medicinal herbs in order to ensure quality
control of herbal supplements by using contemporary methodologies,
applying relevant standards, and adhering to good manufacturing practices.
Herbal remedies must also be included in federal standards or pharmacopeia
(WHO, 2015).

1.10 New Drugs Derived from Medicinal Sources

Nowadays, almost every pharmacopeia in the world suggests plant-based

medications that have real therapeutic value. Certain countries, including the UK,
Russia, and Germany, have their own plant pharmacopeias. On the other hand, the
use of illicit drugs is virtually always far more common. Their use is predicated on
the findings of current scientific research, conventional medicine, popular
medicine, or both. Numerous plant species are used internally or under the
guidance of a physician or pharmacist (Shakya et al., 2016). They may be used
independently of synthetic treatments or in combination with them (complementary
medicine). To achieve an effective and well-executed treatment, it is essential to
comprehend not only the physiological influence of medicinal plants but also the
correct diagnosis of the ailment. The most common types of plant drugs and phyto
preparations utilized as therapeutic approaches are those having established active
components, shown efficacy, and, on rare occasions, clinical consequences (Wang,
2016).

1.10.1 Medicinal Plants as Pharmaceuticals

According to a 2003 WHO survey, 30% of pharmaceuticals marketed worldwide

included substances derived from plants, and in 2002, sales of herbal cures were
estimated to have brought in USD 600 million. About 80% of the people in

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developing countries rely largely on plant-based medicines to meet their medical needs
(Tungmunnithum et al., 2018). Such as:
• Artemisinin (Medication for Malaria): Artemisinin and its derivatives, which
are derived from the Artemisia annua plant, are essential to the creation of
antimalarial medications. Nowadays, ACTs, or artemisinin-based combination
treatments, are often used to treat malaria.
• Paclitaxel (Cancer Treatment): Paclitaxel is a significant chemotherapy
medication that was first made from the Pacific yew tree (Taxus brevifolia). It is
used to treat a number of malignancies, including lung, ovarian, and breast
cancer.
• Morphine (Pain Management): Morphine, which is derived from the opium
poppy plant (Papaver somniferum), has long been a staple in the treatment of
pain. It functions as a model for opioid analgesics, which are prescribed to treat
pain.
• Vincristine and Vinblastine (Cancer Treatment): These alkaloids, which were
extracted from the Catharanthus roseus plant in Madagascar, have been used in
the creation of anticancer medications. Leukemia and Hodgkin's lymphoma are
among the malignancies that may be treated with vinblastine and vincristine.
• Aspirin (Inflammation and Pain): Although salicylic acid, the active component
of aspirin, is not obtained directly from a plant, it was first extracted from the
willow tree's (Salix spp.) bark. Because of its analgesic and anti-inflammatory
qualities, aspirin is often utilized.
• Anticancer Derivatives of Camptothecin: Originally obtained from the bark and
stem of the Chinese tree Camptotheca acuminata, camptothecin provided the
foundation for the creation of medications such as topotecan and irinotecan,
which are used to treat certain malignancies.

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 1.11 Scientific Classification

Artocarpus lacucha, commonly known as Monkey Jack or Monkey Jackfruit, is a

tropical tree belonging to the family Moraceae. Here is its scientific classification:
Kingdom: Plantae

Phylum: Angiosperms (Flowering plants)

Class: Eudicots

Order: Rosales

Family: Moraceae

Genus: Artocarpus Figure 2.1: Artocarpus lacucha

Species: Artocarpus lacucha

1.12 Botanical Description

Artocarpus lacucha, commonly known as Monkey Jack or Monkey Jackfruit, is a
tropical tree that belongs to the Moraceae family. Here are some details regarding
the botanical description of Artocarpus lacucha:
1. Morphology:

• Tree Form: It is a medium-sized to large evergreen tree with a straight

trunk and a rounded or spreading crown.

• Height: The tree can reach heights of 25 to 30 meters.

• Bark: The bark is rough and dark brown.

2. Leaves:

• Type: The leaves are alternate, simple, and broadly ovate to elliptic in
shape.

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 • Margin: The leaf margins are entire (smooth).

• Venation: The venation is pinnate with prominent midribs.

3. Flowers:

• Inflorescence: The flowers are arranged in densely packed, axillary, or

cauliflorous (borne on the trunk) catkins.
• Sex: The species is monoecious, meaning individual trees bear both male
and female flowers.
• Male Flowers: These are small, cylindrical, and borne on long spikes.
• Female Flowers: They are larger and occur on shorter spikes.
4. Fruits:

• Type: The fruit is a syncarp, which is a multiple fruit composed of

numerous individual drupelets.

• Size: The individual drupelets are small, usually about 1 to 2 cm in

diameter.

• Color: The mature fruits are greenish to yellowish in color.

• Texture: The fruit has a fibrous and somewhat rough texture.

5. Seeds:

• Number: Each drupelet contains one seed.

• Shape: The seeds are ovoid or ellipsoid.

7. Ecological and Cultural Significance: The tree is valued for its timber, and the
wood is used for various purposes. The fruits are consumed locally, and the latex from the
tree may have certain uses (Adhikary et al., 2023).

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 Figure 2.2: Parts of Artocarpus lacucha (Ganeshaiah et al., 2012)

1.13 Geographical Distribution

Artocarpus lacucha is endemic to South and Southeast Asia. India, Bangladesh,

Myanmar, Thailand, Cambodia, Laos, Vietnam, Malaysia, Indonesia, and the
Philippines are among the nations that make up its natural distribution. Evergreen
and tropical rainforests are ideal habitats for monkey jack trees. Although they may
be found in submontane regions up to a height of around 1,200 meters (3,937 feet)
above sea level, they are more often seen in lowland locations. These trees are often
found next to rivers and streams and require well-drained soils (Vanajakshi et
al.,2020). In addition to its natural habitat, Artocarpus lacucha is grown in other
tropical climates worldwide. This is because of its edible fruit as well as other
applications, such as conventional medicine. Areas with appropriate tropical
conditions, such as portions of Africa, the Caribbean, Central America, and South
America, may be subject to cultivation. A medium- to large-sized evergreen tree
with a straight trunk and a thick crown is the monkey jack tree. Its glossy green
leaves are elliptic and oblong in shape and alternating. The fruit on the tree is
unique; it is knobby and has a prickly outer layer. Although it is not as popular as
some other Artocarpus species, such as jackfruit (Artocarpus heterophyllus), the
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fruit of Artocarpus lacucha is edible. The fruit is green and prickly on the outside
and white and fibrous inside. It may be consumed raw or cooked. Different portions
of the tree are used in traditional medicine in addition to its culinary purposes
(Aneklaphakij et al.,2020).

1.14 Phytochemistry

1.14.1 Lacunin

Artocarpus lacucha has a special phytochemical called lacunin. It is classified as a

prenylated stilbenoid because it has prenyl side chains incorporated into the core
stilbene structure. The fruit's potential bioactivity has made lacunin a topic of
attention in phytochemical investigations. Antioxidant qualities in this substance
can aid in scavenging free radicals and lowering oxidative stress. Furthermore,
studies have indicated that lacunin could have anti-inflammatory properties, which
makes it potentially useful for reducing illnesses linked to inflammation (Sitorus et
al., 2022).

1.14.2 Flavonoids

Various flavonoids, a class of polyphenolic chemicals with antioxidant qualities, are

present in Artocarpus lacucha. Flavonoids are well recognized for their capacity to
lessen oxidative damage and neutralize dangerous free radicals. Flavones, flavonols,
and flavanones are a few of the particular flavonoids found in the fruit. These
substances could be involved in the fruit's possible health advantages, namely its
ability to shield cells from oxidative stress (Sitorus et al., 2022).

1.14.3 Tannins

Another group of polyphenolic substances discovered in Artocarpus lacucha are

called tannins, and they are mostly concentrated in the leaves and bark. Tannins are
known to have astringent qualities and to play a part in plant defense systems.
Tannins are well-known in phytochemistry for their capacity to precipitate proteins,

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which may have effects on the health of the digestive system and the suppression of
harmful microbes (Sitorus et al., 2022).

1.14.4 Phenolic Acids

The polyphenolic chemicals known as phenolic acids have also been found in
different regions of Artocarpus lacucha. Ferulic acid and p-coumaric acid are two
examples. Because of their antioxidant qualities, phenolic acids can aid in lowering
the body's oxidative stress. Additionally, they may have anti-inflammatory
properties (Kumar M et al., 2010).

1.14.5 Terpenoids and Triterpenes

According to certain research, Artocarpus lacucha's leaves and bark contain

terpenoids and triterpenes. These substances are varied and have a range of
bioactive characteristics. Terpenoids are abundant in the natural world and may
play a part in a plant's defensive mechanisms. Potential pharmacological effects of
triterpenes, such as those linked to inflammation, antioxidants, and other health
issues, have been studied (Sitorus et al., 2022).

1.14.6 Steroids

Various plant sections have been shown to contain steroids. A broad class of
chemicals with a range of physiological functions in plants are called steroids.
Because of their possible bioactivity and contribution to Artocarpus lacucha's
overall phytochemical composition, steroids may be of interest in phytochemistry
(Kumar M et al., 2010).

1.14.7 Alkaloids

The bark and leaves of Artocarpus lacucha contain a few different types of
alkaloids. Alkaloids are a family of chemicals with nitrogen that exhibit a wide
range of biological functions. They could play a part in plant defense, and more

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study to determine their possible medicinal qualities would be interesting (Sitorus et
al., 2022).

1.15 Pharmacological Actions

It has been reported that Artocarpus lacucha possesses antioxidant qualities.

Antioxidants aid in the body's defense against free radicals, which may be a factor
in a number of health problems, including chronic illnesses. According to some
research, Artocarpus lacucha extracts may have anti-inflammatory properties. Anti-
inflammatory chemicals may have therapeutic promise since inflammation is a
major contributing factor to a number of disorders. There has been evidence of
antibacterial activity of Artocarpus lacucha extracts against certain microbes. This
may suggest possible uses in the creation of antibacterial drugs or in conventional
therapy (Jagtap et al., 2010). There is little evidence to support the possibility that
Artocarpus lacucha has anti-diabetic qualities. Its effects on insulin sensitivity and
blood glucose levels have been the subject of much research. Wound care is one of
the traditional medical applications of Artocarpus lacucha. Although there hasn't
been much scientific study done in this field, it's plausible that some of the plant’s
constituents aid in the healing of wounds. Based on available data, Artocarpus
lacucha may possess hepatoprotective qualities, which might shield the liver from
harm. Its anti-inflammatory and antioxidant properties may be the cause of this
(Vanajakshi et al., 2016).

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Panal et al., (2022), conducted research on A. lacucha for focusing the
phytochemical constituents and pharmacological properties of the plant. One useful
fruit that may be included into a regular diet is the A. lacucha fruit. Fresh
counterparts are said to have a high water content (82%) and a high fiber content
(2%) along with a large vitamin A and C content. Numerous macro- and
microminerals, including calcium (66.6 mg), magnesium (23.6 mg), potassium (350
mg), phosphorus (22.1 mg), iron (778 μg), zinc (3981 μg), copper (7974 μg), and
manganese (2025 μg), are present in A. lacucha fruit, according to one source.
Here, It is possible to observe the DPPH radical scavenging process by measuring
the absorbance drop. Antioxidants (AH) can reduce radicals or react with other
radical chemicals to cause this. The wood, bark, and leaf aqueous extracts of A.
lacucha have antibacterial properties that may stop the development of fungus and
bacteria, with an average inhibition zone of ≥20 mm in the very strong category and
10–20 mm in the strong category. At the same time, the ethanol extract from A.
lacucha wood also suppressed, on average by 10–20 mm, the bacterial inhibition
zone in the strong category. It was discovered that A. lacucha leaves extracted in
methanol had dose-dependent and statistically significant anti inflammatory effects.
At 200 mg/kg (64.90%), A. lacucha was more effective than indomethacin at
reducing inflammation. The writhing reaction was inhibited by the A. lacucha leaf
methanol extract dosage by 29.63% and 57.41%, respectively (p < 0.05).

Additionally, Shanta et al., (2019), found out the antinociceptive activities of A.

lacucha and its isolated phenolic compound, catechin, in mice. The pathological
circumstances of pain can arise and persist as a result of nociception in the central
and peripheral nervous systems. Although there are now analgesic therapies on the
market, such as glucocorticoids, non-steroidal anti

29 | P a g e
inflammatory medications, and opioids, their applicability have been hindered by
their severe negative effects. It has been documented that Artocarpus species have
promise anti-arthritic, anti inflammatory, and antioxidant properties in both crude
extracts and isolated phenolic compounds. There have been reports that significant
levels of flavonoids and phenolics may be found in the methanol extract of A.
lacucha bark. Amyrin acetate, lupeol acetate, 5-(γ,γ -dimethylallyl)-
oxyresveratrol,3-(γ,γ-dimethylallyl)resveratrol,3-(2,3-dihydroxy-3-
methylbutyl)resveratrol, 3- (γ,γ-dimethylpropyl)moracin M, afzelechin-3-O-R-L-
rhamnopyranoside, oxyresveratrol, dihydromorin, (−)-epiafzelechin, and
epiafzelechin-(4β → 8)-epicatechin have also been reported to be present in the
bark of the tree. It has been shown that the phenolic component trans-2,4,3′,5′-
tetrahydroxystilbene (THS), which was extracted from an aqueous extract of the
plant, had anti-inflammatory, antioxidant, anti-HIV, and antiherpetic properties. The
plant's bark and twigs contain a phenolic chemical called catechin, which has been
shown to inhibit the cyclooxygenases COX-1 and COX-2 in vitro. It has already
been documented that catechin has encouraging anti arthritic properties. These
results led to the design of the current study, which assessed the antinociceptive
effects of a hydro-alcoholic extract of A. lacucha bark and its isolated component,
catechin, in mice models.

Moreover, Giash et al., (2020) , observed the evaluation of anthelmintic, antioxidant

and antiinflammatory potential of methanolic extract of Artocarpus lacucha leaves.
With a small modification, the technique of Ajaiyeoba et al. 14 was used to test the
anthelmintic activity of metahnolic plant extract. Perthima posthuma was utilized
in this test due to its physiological and anatomical similarities to human intestinal
worms. Earthworms were gathered from the Noakhali region's wet soil, which had
been cleaned of dirt and other dust particles using regular saline. The total phenolic
content of the extracts was calculated and contrasted with the gallic acid reference
solutions. The results indicated that methanolic extracts had a dose-dependent rise

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in anti-inflammatory action, whereas a 10 mg/ml concentration considerably
inhibited heat-induced hemolysis by 20.16% and a hypotonic solution by 15.06%.
The extract's capacity to scavenge free radicals or donate hydrogen is determined
by its DPPH radical scavenging activity. The stable free radical DPPH combines
with an antioxidant that has the ability to donate hydrogen. Artocarpus lacucha
crude extract's antioxidant effect may be attributed to its phenolic content.

Furthermore, Poppy et al., (2018), identified the antioxidant activity of n-Hexane,

Ethyl Acetate and Ethanol extract from lakoocha leaves using the DPPH method.
According to this study, the maximum antioxidant activity was found in the ethanol
extract of lakoocha leaves. The quantity and positioning of the hydroxyl and
methyl groups on the ring may have an impact on the higher antioxidant activity. A
compound with a large number of hydroxyl groups would be more effective in
absorbing free radicals due to its increased ability to donate hydrogen atoms. That
explanation clarified why, despite the presence of flavonoids in the phytochemical
screening findings, ethyl acetate showed inactive antioxidant properties. Because
ethyl acetate has more methyl groups than hydroxyl groups, the flavonoid it
contains tends to be semi-polar.

Tania et al., (2021) conducted a study on Phytochemical Screening And Antioxidant

Activities of Artocarpus lacucha leaves. Here the results of the phytochemical
screening test reveal the presence of phenolic substance, flavonoids, tannic acid,
triterpenoids, glycosides, and saponins. Certain serious chronic disorders including
diabetes mellitus, coronary heart disease, and cancer may be caused by free
radicals. The goal of the present investigation was to evaluate ethanol's anti-cancer
capabilities. Leaf extract from Artocarpus lacucha. The 1,1-diphenyl-2-
picrylhydrazyl (DPPH) free radical was used to measure the amount of free radical
scavenging activity. Ascorbic acid had an IC50 of 6.14μg/ml while the ethanol
extract of Artocarpus lacucha leaves was 49.89μg/ml. Figure Applying the equation

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revealed the total flavonoid concentration of the Artocarpus lacucha leaves extract
to be 811.9 mg QE/gm dry extract. Using the equation, the total phenolic content of
the Artocarpus lacucha leaves extract was calculated, and it was found to be 164.4
mg GAE/gm dry extract. Using the equation, the total tannin content of the
Artocarpus lacucha leaf extracts was calculated, and 400.87 mg GAE/gm dry
extract was found. According to the analysis, using Artocarpus lacucha leaves
might have some beneficial effects due to their strong antioxidant activity.
However, Prasit et al., (2012), determined anti-aging activity, non-toxic dose of
photo oxyresveratrol from Artocarpus lakucha. Here, glycation products and free
radicals play significant roles in the onset of cellular aging. Cellular aging is
associated with external radicals, particularly singlet oxygen, which produces
ultraviolet light. The phyto oxyresveratrol that was extracted from A. lakucha in
this investigation demonstrated anti-oxidant and anti-glycation properties. This
validates earlier research on the phytocompound. The results show that the phyto
oxyresveratrol isolate is bioequivalent to the reference standard, OxyResvenox, due
to its high degree of radical scavenging and antiglycation actions.

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Significance of The Study Comprehending the botanical traits, geographic range,
and ecological function of Artocarpus lacucha enhances our comprehension of local
biodiversity. Researching a tree's preferred environment, growth habits, and
relationships with other species may have an impact on managing and conserving
ecosystems. Evaluating Artocarpus lacucha's biodiversity and ecological
significance advances our knowledge of the surrounding environment. Conservation
plans for Artocarpus lacucha and its environment may benefit from an
understanding of the variables influencing the species' distribution and abundance.
Traditional usage of Artocarpus lacucha for therapeutic purposes is well-known. Its
different components' phytochemical content may be examined by researchers to
find possible bioactive substances. Pharmacological development or conventional
medicine may be impacted by an assessment of the pharmacological characteristics
and possible therapeutic uses of Artocarpus lacucha extracts.
New bioactive substances may be found by looking at the chemical makeup of
Artocarpus lacucha and its possible therapeutic uses. Comprehending the health
advantages linked to the ingestion or utilization of Artocarpus lacucha extracts
may have consequences for conventional treatment or the advancement of
pharmaceuticals.
For the sustainable management and conservation of Artocarpus lacucha, it is
essential to analyze the genetic diversity within these populations. Creating plans to
preserve genetic diversity helps to guarantee Artocarpus lacucha's long-term
survival and environmental adaptation. Understanding the nutritional makeup of
Artocarpus lacucha—especially if it is used as a food source—adds to our
understanding of regional cuisines and food security. Encouraging the production
and consumption of nutrient-dense species may benefit public health.

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Aim and Objective of The Work

The aims of the Study were to:

1. study the antioxidant activity by measuring total flavonoid content and

2. Assay of antimicrobial activity against different bacterial strains by disc

diffusion method

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3.1 Collection of Plant

Throughout the whole nation, Artocarpus lacucha is abundantly dispersed. The

specimen came from Dhaka in 2022. The plant was successfully recognized by a
skilled taxonomist. The specimen (stem) had gone through a preliminary
purification procedure to get rid of any unwanted materials or particulates. After
that, the plant samples were dried in the sun.

3.2 The Powdering Process

After the ingredients were dried, they were ground into a coarse powder using a
household blender. Through the process of cutting the pieces smaller, the inside
tissues and cells are made more accessible to the solvents, which may then enter the
cells and gather their contents. After that, the crushed product was placed into glass
containers that had been disinfected and sealed. There were 205 grams of powder in
total. The mixer was carefully cleaned before it was put to use to avoid
contamination from any leftover material from a previous crushing operation or any
other undesirable materials that could have gotten on the blender.

3.3 Extraction

Using 1.5 liters of methanol, the powder and solvent were properly combined by
shaking the container firmly. The combination was then kept for seven days in a
securely sealed glass container that was shaken often to help with the mixing of the
crushed ingredients and solvent. This procedure is referred to as maceration. The
glass container's lid was tightly sealed to prevent air from getting inside.

3.4 Filtration

The plant extracts were filtered in the following order: first through a cotton filter
that had been sterilized, then through Whatman No. 1 filter paper after the
extraction process. In a beaker, the filtrate gathered. Three times, cotton and filter

35 | P a g e
paper were used in the filtering process. After that, the filtrate was put in a conical
flask, covered with aluminum foil, and allowed to evaporate by rotation.

3.5 Preparation for Evaporation and Extraction

A revolving evaporator machine with a vacuum pump was used to evaporate the
solvent, lowering the pressure within the glass tube coil. Reduced pressure causes
evaporation to happen quickly. On the other hand, the condenser made it easier to
reuse the solvent. Seventy percent of these solvents return to their liquid form. The
extraction was taken out of the evaporating flask and the solvent was taken out of
the receiving flask. The extract was then put into a 50 ml beaker and tightly
covered with foil.

3.6 Anti-Microbial Activity Test

After major breakthroughs in the 1960s, the efficacy of essential antibiotics is

presently undermined due to the rise in bacterial resistance, raising concerns for
global public health. The search for new antibiotics necessitates concentrating on
investigating natural products produced by bacteria and plants as potential sources
of medicinal chemicals. The antibacterial qualities of synthetic substances, essential
oils, secondary metabolites, and plant and microbial extracts are currently being
studied. The in vitro antibacterial activity is assessed using a variety of laboratory
techniques, such as disk-diffusion and dilution processes (Balouiri et al., 2016).

3.6.1 Principle

Agar plates are inoculated with a standard inoculum of the microorganism being
studied. Filter paper discs with the test material at a predetermined concentration are
then placed on the agar’s surface. The Petri plates are put in an incubator and given
the best conditions possible—that is, 37°C for 24 hours. Usually, the target
microorganism's germination and growth are inhibited by an antibiotic drug that

36 | P a g e
 penetrates the agar. The areas where development is restricted are then measured for
size (Balouiri et al., 2016).

3.6.2 Apparatus and Reagent

Table 3.1 Apparatus and reagents for antimicrobial test

Petri dishes Sterile Forceps

Autoclave Eppendorf Tube

Spirit Burner Nutrient agar medium

Ethanol Nose mask

Incubator Inoculating loop

Filter paper discs Sterile Cotton bud

Media Bottle Isotonic saline solution

Laminar Air Flow Hood Micropipette

3.6.3 Test Sample of Artocarpus lacucha

The test sample utilized in this study was a methanolic extract of Artocarpus
lacucha stems.

3.6.4 Test Organisms

The experiment utilized bacterial strains obtained as pure cultures from the microbiology
laboratory at East West University. The test included both gram-positive and gram negative
organisms (Table 3.2).

Table 3.2: List of micro-organisms

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 Type of Bacteria Name of Bacteria

Gram Positive Bacteria Bacillus cereus

Bacillus megaterium

Bacillus subtilis

Staphylococcus aureus

Gram Negative Bacteria Salmonella paratyphi

Salmonella typhi
Vibrio parahemolyticus
E. coli
Shigella dysenteriae
Pseudomonas aureus

Fungi Aspergillus niger

3.6.5 Procedure
3.6.5.1 Culture Medium Selection
Nutrient agar medium was used to create fresh cultures and evaluate the organisms'
susceptibility to the test ingredients.

3.6.5.2 Preparation of Medium

To create the necessary volume of this medium, exact amounts of deionized water
and nutrient agar were added to a media bottle. An autoclave (Figure 3.1) was used
to sterilize the medium, petri plates, and test tubes. It was set to 121°C for 20
minutes at a pressure of 15 psi. The slants were used to create new bacterial
cultures, which were then used to test for sensitivity.

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 Figure 3.1: Autoclave (Raypa, n.d.)

3.6.5.3 Sterilization Procedures

To avoid cross-contamination and contamination of any type by the test organisms,

the antimicrobial screening was carried out in a Laminar Hood. To guarantee the
integrity of the findings, stringent protocols were followed. One hour before the
commencement of work in the Laminar Hood, the UV light was turned on.
Sterilization of Petri dishes and other glassware was accomplished by autoclaving
them for 20 minutes at 121 degrees Celsius and 15 pounds of pressure. Sterilization
was also performed on cotton, forceps, blank discs, and micropipette tips.

3.6.5.4 Preparation of Subculture

The test organisms were moved from pure cultures to agar medium (for bacteria) in a
sterile setting within a laminar air cabinet in order to produce uncontaminated and
pure cultures. The strains were injected, and then they were grown for a whole day
at 37°C to find their ideal growth temperature.

3.6.5.5 Negative Control

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One petri dish was incubated without any bacterial inoculation in order to
guarantee that the medium had been thoroughly sanitized.

3.6.5.6 Preparation of Discs

Antimicrobial screening involved the utilization of three distinct types of discs

• Standard Discs: These were used as positive controls to confirm that the
standard antibiotic was effective against the test organisms and to
compare the test sample's response to that of the known antimicrobial
agent. A 30-μg-diameter ciprofloxacin disk served as the experiment's
reference.
• Blank discs: These were used as negative controls to ensure that neither
the filter paper nor the leftover solvent, which remained on the discs
even after air drying, had any activity of its own.
• Sample discs: These were dried and used to measure the anti-activity of
the samples after being submerged in solutions containing test samples
at predetermined concentrations. Antibacterial and antioxidant properties
are shown by Artocarpus lacucha cyclohexane extraction.

3.6.5.7 Preparation of Test Sample Discs

Gram positive and gram-negative bacteria at concentrations of 300 and 600 μg/disc
were used to test the extract fraction's antibacterial activity.

3.6.5.8 Preparation of The Test Plates

With the use of a sterile transfer loop, the tested isolates were extracted from the
culture. Under aseptic circumstances, the sample was moved to Eppendorf tubes
holding 1 ml of sterile saline water. The organisms were equally suspended as a
consequence of rotating and shaking the Eppendorf tubes. The bacterial solution

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quickly contaminated the aseptic petri plate. Using plates and cotton swabs,
uniformly distribute the test organisms throughout the medium.

3.6.5.9 Application of Test Samples

The pre-inoculated zones on the plates were meticulously overlaid with the sample
discs, the regular antibiotic discs, and the control discs.

3.6.5.10 Diffusion & Incubation

After that, the plates were kept in a refrigerator with a 4°C setting for about a day in
order to allow the ingredients from the discs to diffuse into the agar medium. After
that, the plates were turned upside down and kept in the incubator for twenty-four
hours at 37°C.

3.6.5.11 Determination of Antibacterial Activity by Measuring the Zone of

Inhibition

The antibacterial activities of the test materials were evaluated by quantifying the
diameters of the zones of inhibition in millimeters following incubation (Figure
3.2)

Figure 3.2: Determination of clear zone of inhibition (Shariar, 2020)

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3.7 Antioxidant Activity

3.7.1 Total Flavonoid Content

The colorimetric technique using aluminum chloride (AlCl3) is used to determine

the total flavonoid content of the crude plant extract. Assay methodology is
predicated on the idea that the C-4 keto group and the C-3 or C-5 hydroxyl groups
of the flavones and flavonols present in the crude extract may form stable
complexes with aluminum chloride. Additionally, aluminum chloride may combine
the ortho-hydroxyl groups present in the flavonoid A or B rings to form molecules
that are sensitive to acid. At a wavelength of 415 nm, the flavonoid-aluminum
combination formed by the flavonoid in the crude extract and aluminum chloride
reaches its maximum absorbance. Using a UV-visible spectrophotometer, the
quantity of flavonoid in the crude extract may be ascertained by comparing the
absorbance of the reaction mixture at 415 nm
with a blank that contains all of the reagents except the extracts. Quercetin was used
as the reference at various concentrations (Singatong et al., 2010).

Flavonoid (Extract) + AlCl3 (reagent) =Formation of flavonoid-aluminium

complex (λmax =415nm)

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 3.7.2 Apparatus & Reagents

Table 3.3: Apparatus and reagents for total flavonoid content test

Reagents Apparatus

Aluminium Chloride Spatula

Potassium Acetate Analytical balance

Methanol Pipette

Quercetin Test tubes and volumetric flask

3.7.3 Preparation of 10% Aluminium chloride (AlCl3) solution

A total of 2.5 grams of AlCl3 was added to a 25 ml volumetric flask

and the volume was then adjusted using distilled water.

3.7.4 Preparation of 1M Potassium acetate solution

2.4 g of potassium was taken into a 25 ml of a volumetric flask and the

volume was adjusted by distilled water.
3.7.5 Preparation of Standard solution

A stock solution of quercetin with a concentration of 5μg/μl was made by

dissolving 0.025 grams of quercetin in 5 milliliters of ethanol. The
experimental concentrations were made from the stock solution using the
following method:

Table 3.4: Preparation of standard in different concentrations

Concentration Solution taken from stock Volume adjusted by Final

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 solution (μl) methanol (ml)
of Quercetin volume
(μg/ml) (ml)

250 250 4.75 5

200 200 4.80 5

100 100 4.90 5

50 50 4.95 5

25 25 4.975 5

3.7.6 Preparation of Extract solution

The stock sample was prepared by diluting 4 mg of Artocarpus lacucha stem methanol
extract in 2 ml of methanol, yielding a concentration of 2000μg/ml. This was seen as a
stock answer. This stock solution's experimental concentration was made by diluting
100μl of the stock solution with 900μl of methanol in a test tube, yielding a 200μg/ml
concentration.
3.7.7 Experimental Procedure

1. 1 ml of plant extract (200μg/ml) and standard of different concentration solutions was taken
in a test tube.

2. 3 ml of methanol was added to the test tube.

3. Then 200 μl of 10% aluminum chloride solution was added into the same test tube.

4. Followed by the addition of 200μl of 1M potassium acetate solution into the test tube.
5. Finally, 5.6 ml of distilled water was mixed with the reaction mixture.
6. The reaction mixture is then incubated for 30 minutes at room temperature to
complete the reaction.

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7. Then the absorbance of the solution was measured at 415 nm using a
spectrophotometer against blank.
8. Methanol served as blank.

9. The Total content of flavonoid compounds in plant methanol extracts in quercetin

equivalents was calculated by the following formula equation:

C = (c x V)/m

Where:

C = Total flavonoid contents, mg/g plant extract in QE,

c = Concentration of quercetin obtained from calibration curve

(mg/ml), V = The volume of the sample solution (ml),

m = Weight of the sample (g).

4.1 Antibacterial Activity

4.1.1 Result
The study utilized the cyclohexane extracts of Artocarpus lacucha to evaluate their
antibacterial properties against a range of Gram-positive and Gram-negative bacteria.

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 Table 4.1, 4.2, and 4.3 displays the results of measuring the zone of inhibition for
each bacterium in millimeters after a 24-hour incubation period. The standard
reference for this data was ciprofloxacin 30 μg/disc.
Table 4.1: Antibacterial Activities of A. lacucha on Gram positive bacteria

Gram Positive Bacteria Zone of Inhibition (mm)

CESAL CESAL Ciprofloxaci Blank

n
300 μg/disc 600 μg/disc
30 μg/disc

Sample Bacteria Disc1 Disc2 Disc1 Disc2 Disc 1 Disc

No. 1
Name

1 Bacillus 7 10 13 25 -

cereus

2 Bacillus - - - - 25 -

megaterium

3 Bacillus - - - - 30 -

subtilis

4 Sarcina lutea 10 - 10 - 25 -

5 Staphylococcus - - 10 20 29 -
aureus

CESAL= Cyclohexane Extract of Stem of Artocarpus lacucha

‘ - ’ Means no antimicrobial activity
Table 4.2: Antibacterial Activities of A. lacucha on Gram negativc bacteria

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Gram Negative Bacteria Zone of Inhibition (mm)

CESAL CESAL 600 Ciprofloxacin 30 Blank

μg/disc μg/disc
300 μg/disc

Sample Bacteria Name Disc Disc Disc Disc Disc 1 Disc

No. 1 2 1
1 2

1 Salmonella - - - - 20 -

paratyphi

2 Pseudomonas - - - - 22 -
aureus

3 Shigella - - - - 35 -

dysenteriae

4 E. coli - - - - 30 -

5 Vibrio - - - - 22 -

parahemolyticus

6 Salmonella - 15 10 11 35 -

typhi

CESAH= Cyclohexane Extract of Stem of Artocarpus lacucha

‘ - ’ Means no antimicrobial activity

Table 4.3: Antibacterial Activities of A. lacucha on fungi

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 Fungi Zone of Inhibition (mm)

CESAL CESAL Ciprofloxacin Blank

30 μg /disc
300 μg/disc 600 μg/disc

Sample Fungi Disc Disc Disc Disc Disc 1 Disc

No. 1 2 1 2 1
Name

1 Aspergillus - - - - 35 -
niger

MESAH= Methanolic Extract of Stem of Artocarpus heterophyllus

‘ - ’ Means no antimicrobial activity

4.1.2Discussion

Eleven bacterial species and one fungal species were used to assess the antibacterial
activity of the cyclohexane extract derived from the stem of Artocarpus lacucha.
Using the disc diffusion technique, the extract was administered at 300 μg/disc and
600 μg/disc. For comparison, 30 μg/disc of ciprofloxacin was used as the standard.
As shown by Tables 4.1, 4.2, and 4.3, it was discovered that the methanolic fruit
extract of Artocarpus lacucha lacked antibacterial activity.

The current investigation's absence of antibacterial growth might be attributed to

species or type heterogeneity. The biological system and soil environment in
Bangladesh may have varied, which might have resulted in a reduced concentration
or lack of the component causing the antimicrobial action, which would have
eliminated the antibacterial activity. The experiment was only going to be
conducted with a dose of 300 μg and 600 μg per disk. Changing the extract's dosage
48 | P a g e
could enhance its antibacterial properties. To assess the antibacterial efficacy
against different bacterial strains, further research is required.

4.2 Antioxidant Test by Total Flavonoid Content

4.2.1 Result
The total flavonoid content of the methanol fractions of the stem extracts of
Artocarpus lacucha were determined using the aluminium trichloride technique.
The total flavonoid content was determined by utilizing the standard curve of
quercetin (y = 0.003x + 0.155; R2 = 0.9883) and was reported as quercetin
equivalents (QE) per gram of the plant extract. The absorbance of each fraction is
determined by calculating the average of two readings.

4.2.2 Preparation of Standard Curve

Table 4.4: Total flavonoid content of quercetin

Concentration of Absorbance Regression line R value

2

Quercetin (μg/ml) (At 415 nm)

250 0.954 y=0.003x+0.155 0.9883

200 0.714

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 100 0.459

50 0.305

25 0.244

A linear correlation was detected when absorbance measurements were made for
various concentrations of quercetin, ranging from 25μg/ml to 250μg/ml. This
relationship is depicted in Figure 4.1. The linear curve was regarded as a benchm ark
curve.

Figure 4.1: Graphical representation of assay of flavonoid content of quercetin

4.2.3 Calculation of Total Flavonoid Content

After obtaining the equation for linear regression from the calibration curve of
Quercetin, the absorbance for methanol (average absorbance of duplicated value is
used here) was put in place of y of equation. And the obtained value of x will be
the value of concentration of Quercetin equivalent (QE). The calculation is as
follows:

y = 0.003x + 0.155

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Then for calculating total flavonoid content following equation
is used: C = (c x V)/m

Where:

C = total content of phenolic compounds, mg/g plant extract in QE

c = concentration of Quercetin obtained from standard curve

(mg/ml) V = the volume of the sample solution (ml) = 1 ml
m = weight of the sample (g) = 0.0002g

Table 4.5: Total Fiavonoid content of Cyclohexane extract of Artocarpus lacucha stem

Concentration Absorbance QE (mg/ml) Total Mean ±

(μg/ml) flavonoid SD

content (mg/gm)

(mg/gm)

51 | P a g e
 200 0.385 0.076 380 383.17±

12.066
200 0.393 0.0793 396.5

200 0.379 0.0746 373

4.2.4 Discussion

The standard curve was utilized to ascertain the overall flavonoid content of the test
samples. The average flavonoid concentration of the dried extract of Artocarpus
lacucha stem is 383.17 mg of QE per gram which can vary with a standard deviation of
12.066.

This suggests that the extract contains a significant quantity of flavonoids, which
support its antioxidant action. Optimizing antioxidant effectiveness might be
achieved by experimenting with different solvents and concentrations. To ascertain
the efficacy of the antioxidant activity, further research and analysis are necessary.

The natural world has been sufficiently generous to mankind to provide a wide range of
plants with medicinal properties. Plant screening with the purpose of identifying and
isolating naturally bioactive compounds enhances the therapeutic database and offers a
less expensive, safer, and more effective substitute for conventional illness treatment
methods. Together, physicians and botanists are attempting to utilize these medicinal
plants to research and create new medications to combat the dreadful ailments like
diabetes, cancer, stroke, myocardial infarction, and others that are becoming more and
more common. Currently, maintaining the therapeutic quality of botanical medicines is
a key challenge when using phytomedicines. As a result, plant materials may include
chemically and physiologically significant therapeutic candidates. This will allow for
more testing of the plant against various ailments to determine its potential usefulness

52 | P a g e
going forward, with high hopes from the Moraceae family's Artocarpus lacucha, which
is traditionally employed in many disease situations. This plant has been enhanced with
a variety of compounds that have unique medical properties. Therefore, it is crucial to
separate the chemical components. My study’s particular goal is to ascertain the
substance's antibacterial, antioxidant, and cytotoxic activities in order to demonstrate its
potential as a modern medication.

Conclusion

For a plant physiologist, working with medicinal plants offers a wide range of research
prospects, and plant physiology studies would undoubtedly play a big role in this
emerging sector. Many frequently used therapeutic plants have not yielded the full
physiological characterization gained by food plants or model plant systems, with just a
few exceptions. While many routes for the production of specific medicinal substances

53 | P a g e
and the factors (biotic and abiotic) influencing their development remain unclear, active
phytochemicals may have been identified. Currently, a major issue with the utilization
of phytomedicines in botanical medicine is maintaining constant medicinal quality.
Thus, plant materials may be sources of new, chemically and physiologically
fascinating pharmaceuticals. And for this reason, in order to determine the plant's
potential usefulness in the face of widespread sickness in the future, it may be further
tested against a variety of ailments.

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