Aim of the study: We aim to evaluate the antiplasmodial, antimicrobial, anti-Trypanosoma brucei

rhodesiense and cytotoxicity activities of the seven medicinal plants of Bhutan selected using an
ethno-directed bio-rational approach. This study creates a scientific basis for their use in the BTM
and gives foundation for further phytochemical and biological evaluations which can result in the
discovery of new drug lead compounds.

Materials and methods: A three stage process was conducted which consisted of: (1) an assessment
of a pharmacopoeia and a formulary book of the BTM for their mode of plant uses; (2) selecting 25
anti-infective medicinal plants based on the five established criteria, collecting them, and screening
for their major classes of phytochemicals using appropriate test protocols; and (3) finally analyzing
the crude extracts of the seven medicinal plants, using the standard test protocols, for their
antiplasmodial, antimicrobial, anti-Trypanosoma brucei rhodesiense and cytotoxicity activities as
directed by the ethnopharmacological uses of each plant.

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Results: Out of 25 medicinal plants screened for their major classes of phytochemicals, the majority
contained tannins, alkaloids and flavonoids. Out of the seven plant species investigated for their
biological activities, all seven of them exhibited mild antimicrobial properties, five plants gave
significant in vitro antiplasmodial activities, two plants gave moderate anti-Trypanosoma brucei
rhodesiense activity, and one plant showed mild cytotoxicity. Meconopsis simplicifolia showed the
highest antiplasmodial activity with IC(50) values of 0.40 μg/ml against TM4/8.2 strain (a wild type
chloroquine and antifolate sensitive strain) and 6.39 μg/ml against K1CB1 (multidrug resistant
strain) strain. Significantly the extracts from this plant did not show any cytotoxicity.

Conclusions: These findings provide the scientific basis for the use of seven medicinal plants in the
BTM for the treatment of malaria, microbial infections, infectious fevers, and the Trypanosoma
brucei rhodesiense infection. The results also form a good preliminary basis for the prioritization of
candidate plant species for further in-depth phytochemical and pharmacological investigations
toward our quest to unearth lead antiparasitic, anticancer and antimicrobial compounds.

The research interests of Dr. Kinghorn are on the isolation, characterization, and biological
evaluation of natural products of higher plant origin, and he has worked in particular on compounds
with potential antimicrobial, cancer chemotherapeutic, cancer chemopreventive, sweet-tasting, and
bitterness-blocking effects. Dr. Kinghorn is the former Editor in Chief (1994-2019) and current
Emeritus Editor (2020-present) of the "Journal of Natural Products" (co-published by the American
Chemical Society and the American Society of Pharmacognosy) and is Series Editor in Chief of the
book series "Progress in the Chemistry of Organic Natural Products" (published by Springer Nature,
Cham, Switzerland) (2008-present).

Abstract:Medicinal plants are the product of natural drug discoveries and have gained traction due
to their pharmacological activities. Pathogens are everywhere, and they thrive in ideal conditions
depending on the nutrients, moisture, temperature, and pH that increase the growth of harmful
pathogens on surfaces and textiles. Thus, antimicrobial agents and finishes may be the solution to
the destruction of pathogens. This review article presents an analysis of various aspects of
producing antimicrobial finishings, the microorganisms, their mechanism of attachment to natural
and synthetic fibre, the effect of microbial growth, and the principle and mechanism of the microbial
activity of the medicinal plants. Furthermore, the extraction methods, qualitative and quantitative
phytochemical evaluations of antimicrobial efficacy, and developments of antimicrobial treated
textiles using various agents are covered in this review.Keywords: antimicrobial agents; textile
finishings; extractions; solvents; phytochemical screening; qualitative analysis; quantitative analysis

Phenolic compounds are the most common type of phytochemical found in a wide range of plant-
based products, including phenolic acids (hydroxycinnamic acids, and hydroxybenzoic), polyphenols
(condensed tannins, and hydrolyzable), flavonoids (rutin, naringenin, quercetin, apigenin,
kaempferol)33, because of its importance in the pharmaceutical, and food industries, these
compounds have been extensively researched34. In this context, M. longifolia (Horsemint) is
considered a potential source of bioactive compounds as well as micronutrients, which can be
explored as a promising alternative for the formulation of nutraceutical products35. As previously
confirmed by our results, the phenolic content strongly depends on the selected extraction solvent.

In our current study, we noticed that Gram-positive strains were more responsive to our extracts
compared to Gram-negative strains. The reason for the lower sensitivity of Gram-negative bacteria
could be attributed to the complexity of their double membrane, which includes a cell envelope
made up of a lipoprotein and a lipopolysaccharide layer (LPS). Unlike Gram-positive bacteria's single
membrane, this structure works as a biological barrier to antibacterial drugs48. The demonstrated
antibacterial activity of our Mentha longifolia extracts can be explained by its rich phytochemical
profile, which includes caftaric acid, rosmarinic acid, cryptochlorogenic acid, ρ-coumaric acid, m-
coumaric acid, chlorogenic acid, caffeic acid, gallic acid, luteolin, apigenin, quercetin, rutin,
coumarins, and isocoumarins49,50. Indeed, previous studies have reported greater antimicrobial
potency for caffeic, cryptochlorogenic, as well as chlorogenic acids49,51,52. Other phenolic
compounds found in various species of the genus Mentha such as ellagic acid, ferulic acid,
gallocatechin, epigallocatechin gallate, and catechins were reported to exhibit antibacterial or
bacteriostatic effects against multiple bacterial strains including Escherichia coli, Staphylococcus
aureus, Bacillus aureus, Bacillus pumilis, Bacillus subtilis, and Pseudomonas aeruginosa. This effect
is associated with their capability to penetrate the bacterial wall and reach the bacterial
cytoplasm52.

One of the important strategies and tactics in the search for new drugs still constists in the empirical
approach i.e. the systematic screening of pure compounds or plant extracts in order to find useful
lead compounds.

One of the key steps in the implementation of this rational serendipity methodology is the
development of screening bioassays, which are adaptable to the testing of plant extracts, which can
be highly colored, tarry, water insoluble and chemically very complex.

Phytochemicals are chemical compounds produced by plants, generally to help them resist fungi,
bacteria and plant virus infections, and also consumption by insects and other animals. The name
comes from Greek φυτόν (phyton) 'plant'. Some phytochemicals have been used as poisons and
others as traditional medicine.

As a term, phytochemicals is generally used to describe plant compounds that are under research
with unestablished effects on health, and are not essential nutrients. Regulatory agencies governing
food labeling in Europe and the United States have provided guidance for industry to limit or
prevent health claims about phytochemicals on food product or nutrition labels.

Phytochemicals are chemicals of plant origin.[1] Phytochemicals (from Greek phyto, meaning
"plant") are chemicals produced by plants through primary or secondary metabolism.[2][3] They
generally have biological activity in the plant host and play a role in plant growth or defense against
competitors, pathogens, or predators.[2]

Phytochemists study phytochemicals by first extracting and isolating compounds from the origin
plant, followed by defining their structure or testing in laboratory model systems, such as in vitro
studies using cell lines or in vivo studies using laboratory animals.[2] Challenges in that field include
isolating specific compounds and determining their structures, which are often complex, and
identifying what specific phytochemical is primarily responsible for any given biological
activity.[2][10][11]

Without specific knowledge of their cellular actions or mechanisms, phytochemicals have been used
as poison and in traditional medicine. For example, salicin, having anti-inflammatory and pain-
relieving properties, was originally extracted from the bark of the white willow tree and later
synthetically produced to become the common, over-the-counter drug aspirin.[12][13] The tropane
alkaloids of Atropa belladonna were used as poisons, and early humans made poisonous arrows from
the plant.[14] In Ancient Rome, it was used as a poison by Agrippina the Younger, wife of Emperor
Claudius on advice of Locusta, a lady specialized in poisons, and Livia, who is rumored to have used
it to kill her husband Emperor Augustus.[14][15] Other uses include perfumes, such as the
sequiterpene santolols, from sandalwood.[16]

As of 2017[update], the biological activities for most phytochemicals are unknown or poorly
understood, in isolation or as part of foods.[2][5] Phytochemicals with established roles in the body
are classified as essential nutrients.[4][17]

The phytochemical category includes compounds recognized as essential nutrients, which are
naturally contained in plants and are required for normal physiological functions, so must be
obtained from the diet in humans.[17][18]

Non-digestible dietary fibers from plant foods, often considered as a phytochemical,[24] are now
generally regarded as a nutrient group having approved health claims for reducing the risk of some
types of cancer[25] and coronary heart disease.[26]

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