September18, Unitedkingdom  2021 

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Abstract Volume: 2 Issue: 2 ISSN:

Extraction of Active Components from Crude Ocimum Sanctum and Maintenance of C. Elegans

Amrita Mishra*, Dr. Rakhi Agarwal

 

1.Associate Professor, IFS, GFSU Worked at laboratory of analytical and molecular toxicology Institute of forensic science gujarat forensic sciences university, Near DFS Head Quarter, Sector 9, Gandhinagar, Gujarat, India.

 

*Corresponding Author: Amrita Mishra, M.Sc. Forensic Science, Specialization in Forensic Chemistry Toxicology.


Received Date:  February 5, 2021

Publication Date: March 01, 2021


Abstract
Ayurveda is the traditional Indian system of medicine that is meant for curing diseases and also in preventing the occurrence of diseases. It focuses on healthy lifestyle practices and regular consumption of herbs especially mediational plants for curing various diseases. In the traditional system of medicine, different parts of the Tulsi plant (Ocimum sanctum Linn.) possess various therapeutic effects. Eugenol (1-hydroxy-2-methoxy-4-allylbenzene), the active constituents present in O. sanctum L. is largely responsible for the therapeutic potentials.

Caenorhabditis elegans is a free-living, transparent nematode, about 1mm in length, that lives in the temperate soil environment. This has been widely used as a model organism for various studies. The effect of crude extract of Ocimum sanctum on C. elegans has been studied here. Firstly, it includes extraction process from Tulsi leaves, then the maintenance of C. elegans which involves Preparation of NGM agar plates, M9 buffer solution, MEM for E. coli; seeding and Chunking of plates; age synchronization.

The study is basically on the effect of crude extract of Ocimum on C. elegans. It shows the antioxidant capacity, resistance against thermal stress, protective and life span extending properties of the medicinal plant (Tulsi) in C. elegans.

Extraction of Active Components from Crude Ocimum Sanctum and Maintenance of C. Elegans

Introduction

Ayurveda

The word “Ayurveda” comes from the Sanskrit language which means “life-knowledge”. Ayurveda is a system of medicine that has been used for several millennia. Its therapies are based on complex herbal compounds, minerals, and metallic substances. While Ayurveda is considered a complementary and alternative form of medication in the modern world, it is the only medicine and therapeutic therapy in many rural villages.

Ayurveda names three elemental and constitutional substances or “energies” called doshas which are Vata, pitta, and Kapha, which loosely translates to air, fire, and water, respectively. Doshas and its balance in the human body is the emphasis on maintenance of health and treatment of disease in the Ayurvedic system of medicine. Medicinal plants that have been focused on Ayurveda include – Neem (Azadirachta indica), Guggul (Commiphora weight), Tulsi (Ocimum uniform), Amla- Indian gooseberry (Emblica Officinalis), Turmeric/ Haldi (Curcuma longa). (Rupani & Chavez, 2018)

Figure 1: Ayurvedic medication – Herbal extracts and products

The ethno pharmacological relevance of Ayurveda entails a scientific tradition of harmonious living and its origin is contained in Rigveda and Atharvaveda. Ayurveda is a traditional healthcare system of Indian medicine since time immemorial. Several Ayurvedic medicines have been exploiting for the management and treatment of various diseases in human beings. Hence, Ayurveda has now become a modern practice and can be referred to as “Tradition to Trend”. The potential of Ayurvedic medicine needs to be explored and validation approaches for better therapeutic leads. (Mukherjee et al., 2017)

Ayurveda uses the natural elements or components to eliminate the root cause of a disease by restoring balance and also creates a healthy lifestyle to prevent the reoccurrence of imbalance. Herbal medicines have existed worldwide with a long record history and were used in ancient Chinese, Greek, Egyptian, and Indian medicine for various therapeutic purposes. World Health Organization estimated that 80% of the world’s inhabitants still rely mainly on traditional medicines for their healthcare.

The Indian subcontinent is well-known for its major biodiversity with about 45,000 plant species. About 15,000 medicinal plants have been recorded in India, out of which the communities use 7,000 to 7,500 plants for therapeutic purposes against various diseases. In Ayurveda, single or multiple herbs (polyherbal) are used for treatment purposes. The Ayurvedic literature “Sarangdhar Samhita” highlighted the concept of polyherbal to achieve greater therapeutic efficiency.

A combination of certain plant extracts may improve the therapeutic efficiency. The active phytochemical constituents of individual plants are sometimes insufficient to achieve the desired therapeutic effects, Hence, when multiple herbs are combined in a particular ratio, it may result in a better therapeutic effect and reduce the toxicity. This concept is known as polyherbal. (Parasuraman et al., 2014)

In this particular thesis, we are going to study one Ayurvedic herb i.e. Tulsi (Ocimum sanctum), and its effects on the living organism by using Caenorhabditis elegans as a model organism. C. elegans shows certain behavioral changes when exposed to an environment containing either a toxic substance or a medicinal substance. Hence, it is one of the best-preferred model organisms for such kind of studies that involves any toxin or a medicinal compound. Also, as it is microscopic, it is quite easy to grow and use for experimental purposes without violating ethics.


Tulsi (ocimum sanctum/ocimum tenuiflorum)

[queen of herbs]

Scientific classification

Kingdom: Plantae

Clade: Tracheophytes

Clade: Angiosperms

Clade: Eudicots

Clade: Asterids

Order: Lamiales

Family: Lamiaceae

Genus: Ocimum

Species: tenuiflorum/sanctum

Figure 2: Ocimum sanctum – Tulsi

Ocimum sanctum, commonly known as Holy basil or Tulsi is a perennial plant belonging to the family Lamiaceae. It is native to the Indian subcontinent and widespread as a cultivated plant throughout the Southeast Asian tropics.

Tulsi is cultivated for religious and traditional medicinal purposes and its essential oil. It is widely used as an herbal tea and is commonly used in Ayurveda medicine. It is also used by devotes (Hindu) for worship purposes.


Morphology

Tulsi is an erect, multibranched subshrub, 30-60 cm (12-24 inch) tall. Leaves are simple, opposite green or purple colored that are strongly scented and hairy stems. Leaves have petiole and are ovate, up to 5cm long. Flowers are purplish.


Ocimum Sanctum Genome

The draft assembly of Krishna Tulsi genome is about 386 Mb [1 Mb = 1,000,000 bp]. The plastid (Chloroplast) genome is about 1,42,524 base pairs. (Rastogi et al., 2015).

The pathway leading to the production of medicinally significant phytochemicals in O. sanctum is similar to that of Arabidopsis thaliana and certain other plants. Expression levels of anthocyanin biosynthesis-related genes in leaf samples of Krishna Tulsi were observed to be relatively high, explaining the purple coloration of the leaves. (Upadhyay et al., 2015)


Significance in Hinduism

Tulsi, the ‘Queen of herbs’, the legendary ‘Incomparable one’ of India, is one of the holiest and most cherished of the many healing and health giving herbs of the orient. The sacred basil is renowned for its religious and spiritual sanctity and has an important role in the traditional Ayurvedic and Unani system of holistic health and herbal medicine of the east.

Within Ayurveda, Tulsi is known as:

  • The Incomparable one
  • Mother Medicine of Nature
  • The Queen of Herbs

It is also referred to as the Elixir of life for both its medicinal and spiritual properties. The emerging science on Tulsi, which reinforces ancient Ayurvedic wisdom, suggests that Tulsi is a tonic for the body, mind, and spirit that offers solutions to many modern-day health problems.


Divine Tulsi

In Hinduism, Tulsi is worshiped as a goddess and every part of this plant is considered sacred. Even the surrounding soil, which has been found to harbor beneficial endophytic fungi, is considered as an aspect of the divine. Hindu households are considered incomplete without a Tulsi plant, which is typically ornated in an earthen pot situated in a courtyard where it serves both practical and ceremonial purposes.

Figure 3: Indian women offering water to Tulsi Plant at courtyard


Types of Tulsi

(Jurges et al., 2018)

3 main types of Tulsi plant are:

1. Rama Tulsi

2. Krishna Tulsi

3. Vana Tulsi

Rama Tulsi

Rama Tulsi is also called Sri Tulsi. It is the most dominant form of all having green colored leaves with light purple flowers. It is used mostly for religious purposes and is usually termed as Ocimum tenuiflorum. It has a clove-like scent, due to the chemical Eugenol.


Krishna Tulsi

The purple-leafed Krishna Tulsi, named after it resembles the color of the Hindu deity Krishna, is preferred for medical use and also assigned as O. uniform L. It has a clove-like aroma and peppery flavor. Krishna Tulsi is the right medicinal Tulsi as it has more potency. It has also been used as a traditional contraceptive.


Vana Tulsi

Vana Tulsi, meaning ‘forest’ Tulsi may indicate that it is often found in the wild. The wild leaves of it are bright or light green. They grow in the wild and are indigenous to many areas of Asia and northeast Africa.

Figure 4: Three main types of Tulsi plant


Apart from there three main types, there is a fourth type of Tulsi known as Kapoor Tulsi. However, over 60 different species are belonging to the genus Ocimum tenuiflorum distributed all over the world.

Varieties of Tulsi mostly found in the Indian Subcontinent are Amrita Tulsi ( Amrita meaning immortality, it is called so because it is very difficult to kill this plant as it can grow in toughest and variety environment), sweet Basil, Lime Basil, Greek Basil, Thai Basil, Cinnamon Basil or Mexican Basil, Lettuce leaf Basil, Christmas Basil, Napoletano Basil, Summerlong Basil, Genovese Basil, Ararat Basil, etc.


Chemical composition of ocimum sanctum:

(Verma, 2016)(Cohen, 2014)

Phytochemical constituents of Tulsi includes:

-Eugenol

-Oleanolic Acid

-Ursolic Acid

-Rosmarinic Acid

-Carvacrol

-Linalool

-β- Caryophyllene

Essential oil consists mostly:

  • Eugenol (~70%)
  • β-Caryophyllene (~11.0%)
  • Germacrene (~2%)

With the balance being made up of various trace compounds, mostly terpenes.

Figure 5: Structures of Phytochemical constituents of Tulsi


Caenorhabditis Elegans: A Model Organism

Scientific Classification

Kingdom: Animalia

Phylum: Nematoda

Class: Chromadorea

Order: Rhabditida

Family: Rhabditida

Genus: Caenorhabditis

Species: Elegans


Life cycle of C. Elegans

(Organism, 2014)

Similar to other nematodes, the life cycle of C. elegans is comprised of:

  • The embryonic stage
  • Four larval stages (L1, L2, L3, L4), and
  • Adulthood


Review of Literature

Tulsi: A Potent Adaptogen

Ocimium Sanctum against Mercury

Tulsi- A Medicinal Plant for Prevention and Treatment of Cancer

Antiviral Activity of Ocimium Sanctum

Indian medicinal plants and their possible effect on covid-19

Antioxidant Activities of Ocimum Sanctum against Cadmium Induced Toxicity

Extraction Method (Tulsi Leaves)

Rotary Evaporator Conditions/Parameters:

Materials Required

Protocols

Preparation of M Buffer Solution

Preparation of Minimum Essential Media (mem) for E. Coli

Seeding of Plates

Culturing of Worms

Chunking involves the following steps:

Age Synchronization

Result and Discussion


Conclusion

The ever-increasing rise of debilitating chronic diseases and stress in the population has enforced the necessity for searching for novel therapeutics. The study regarding the medicinal properties of the Tulsi plant (Ocimum sanctum) has provided several health benefits of Tulsi including anti-oxidant, anti-aging, promoting healthy heart, treating kidney stone, relieves headaches, fights acne, relieves fever, improves eye health, cures respiratory disorders, etc. The extract also has been proved to be a strong free radical and increase resistance against thermal stress. It is also suggested that its antioxidant capacity results in a protective and life span extending action. Phytochemical constituents of Tulsi modulates several signaling pathways and increases stress tolerance and life span in C. elegans.


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Volume 2 Issue 2 March 2021

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