Evaluation of Anti-inflammatory activity of Jacquemontia caerulea Leaf Methanolic Extract in Wistar rats

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Adiba Afreen¹ , Shaik Mohd Khasim¹ , Sayeda Umme Bathool² , Fouzia Tehseen³

¹Department of Pharmacology, Shadan College of Pharmacy, Peerancheruvu, Hyderabad, Telangana, 500091, India

²Shadan College of Pharmacy, Peerancheruvu, Hyderabad, Telangana, 500091, India

³Department of Pharmaceutical Chemistry, Anwarul Uloom College of Pharmacy, Mallepally, Hyderabad, 500001, Telangana, India

Corresponding Author Email: adibafreen12@gmail.com

DOI : https://doi.org/10.51470/AMSR.2024.03.01.01

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INTRODUCTION
In recent times, detailed studies on Pharmacological and Phytochemical investigations provide extensive knowledge and information for the medicinal use of herbal plants by demonstrating the presence of active principles present in them and their therapeutic effect on living beings. Hence, it has become crucial for screening and collecting plants for the exploration of new and safe medicinal agents[1]. Inflammation is the body’s mechanism in which white blood cells and chemicals defend against infections caused by foreign substances such as bacteria and viruses [2]. It is regarded as an intricate process. The inflammatory process involves various mediators, potent chemical substances found in the body, including prostaglandins, leukotrienes, lymphokines, and chemokines like interferon-α, γ, interleukin (IL)-1, IL-8, histamine, 5-hydroxytryptamine (5-HT), and tumor necrosis factor-α. These substances initiate chemical pathways and events to provoke a coordinated response against external stimuli.[3]Inflammation is of two types:
 Acute inflammation
 Chronic inflammation
Acute inflammation: is of short duration, which could be anything from a few minutes to a few days. Such inflammation is caused by the foreign substances entering the body, or by physical damage.[4]
The vascular events of the acute inflammatory response involve three main processes:

1. Changes in vessel caliber and consequently blood flow
2. Increased vascular permeability
3. Formation of fluid exudates
   Chronic inflammation: is of longer duration. lasts for an extended period, enduring for weeks, months, or even years.[5]
   Chronic inflammation tends to occur under the following conditions:
    Infections by organisms that are resistant to killing and clearing by the body tend to cause chronic inflammation.
    Repeated bouts of acute inflammation can result in a chronic inflammation.
    Chronic inflammation is a prevalent element in numerous autoimmune disorders.
    Prolonged exposure to toxins.
   The five fundamentals of acute inflammation- “PRISH”
    Pain-
   the puffed-up area is likely to be throbbing, chiefly when touched.
   Chemicals vitalizing nerve endings are allowed to leave, making the area much more subtle.
    Redness-
   this is because the capillaries are charged with more vital fluid than usual
    Immobility-
   There may be somemisplacement of outcome
    Swelling-
   results from edema, the assemblage of solution in the extravascular space as a portion of inflammatory fluid exudates and to a lesser extent, from the physical mass of the inflammatory cells migrating into the area.
    Heat-
   increase in temperature is readily determined. As with the objective for the reddishness, blood in the pretentious area makes it sense hot to touch.[6]

Jacquemontia caerulea is a member of the morning glory family Convolvulacea, which includes 55 genera and 1650 species, widely cultivated for their colourful flowers and heart-form leaves.[7] This family is distributed in both topical and temperate regions of the world. Convolvulacea includes many commercial uses, edible crops, ornamentals, medicinal herbs, and significant weeds.[8] Jacquemontia caerulea is an attractive ornamental, flourishing twining climber. Chemically strains of this family include phytoconstituents as; alkaloids, flavonoids, terpenoids, and coumarins.[9]. Also known as cluster vine, this perennial plant possesses a woody base and has the potential to adopt a shrubby growth form. It primarily thrives in sandy littoral beachfront and maritime hammock environments [10]. The plant blooms from September to June, with vining upper stems adorned with relatively tomentose leaves. [11] Its flowers are bell or wheel-shaped, ranging from blue to white, with a relatively short corolla tube. [12]

The juice extracted from the leaves and roots is consumed as a remedy to treat bites of green mamba snakes and leaf infusion is used to wash wounds. Fruit decoction is utilized in the management of constipation, flatulence, liver complaints. Seeds of this family have carminative and purgative properties. Jacquemontia caerulea has been used topically in traditional medicine to address various skin conditions. It is often applied as a poultice or used in creams or ointments to treat rashes, insect bites, and skin irritations.[13] Extracts from Jacquemontia caerulea have shown antioxidant potential. Antioxidants play a crucial role in counteracting detrimental free radicals within the body, providing protection against oxidative stress, a condition linked to numerous diseases. The core foundation for the therapeutic effectiveness of plants lies in the phytochemicals they contain and their antioxidative capacities. Comprehensive phytochemical analyses on diverse components of the Jacquemontia caerulea plant have revealed the existence of a range of phytoconstituents and compounds, encompassing alkaloids, flavonoids, and saponins. These substances serve as secondary metabolites and also function as defense mechanisms. Flavonoids dissolve the extracellular proteins and form protein complexes. Alkaloidspossess antibacterial, antioxidants, antifungal, analgesic, anxiolytic, anticoagulant, and anticancer activities.[14]
MATERIALS & METHODS
COLLECTION, IDENTIFICATION, AND EXTRACTION OF PLANT MATERIAL:
The leaves of Jacquemontia caerulea were obtained from an authorized plant supplier and underwent authentication by the Department of Botany at Sri Venkateshwara University in Tirupati-517 502, Andhra Pradesh, India. The leaves were air-dried in the shade for 15 days and were coarsely powdered using a blender. The plant material was taken up for extraction using methanol and water of 80% respectively. The extraction was carried out by a soxhlet extraction process followed by distillation and subsequently stored in the refrigerator.[15]
PREPARATION OF EXTRACT:
300g of leaves of Jacquemontia caerulea were collected, washed thoroughly, and dried in the shade. Leaves are made into coarse powder. The powder of the leaves was extracted in a soxhlet extractor with successive methanol solvent. The extract was then subjected to distillation and heated in a water bath for semisolid consistency and then placed in the refrigerator.[16]
EXPERIMENTAL ANIMALS:
Adult Wistar rats, of both genders weighing between 200-250 g, were obtained from the animal facility at Shadan College of Pharmacy in Peerancheruvu, Hyderabad. The rats were acclimated to standard environmental conditions, including ambient temperature, relative humidity, and a 12/12-hour light-dark cycle, for one week before the commencement of the experiment. Throughout this period and the experiment, the animals had unrestricted access to a standard pellet diet and water. All procedures involving animals adhered to the guidelines set by the Committee for Control and Supervision of Experiments on Animals (CPCSEA). Additionally, the experimental protocol received approval from the Institutional Animal Ethical Committee.
Methods
Preliminary Phytochemical Screening
The preliminary Phytochemical Screening of Jacquemontia CaeruleaMethanolic Leaf Extract was assessed for the presence of steroids, flavonoids, saponins, glycosides, carbohydrates, oils, phenols, and fats. The phytochemical screening tests for active principles were carried out by using standard procedures[17-19].
Evaluation of Anti-inflammatory activity of JCLME

1. CARRAGEENAN INDUCED PAW EDEMA METHOD:
   30, Wistar rats of both genders weighing between 150-200 grams (divided into five groups containing 6 in each), extract of Jacquemontia caerulea, carrageenan, and normal saline.
   ANIMAL GROUPING:
   Procedure:
   The anti-inflammatory activity was evaluated following the method outlined by Winter et al. (1962). Thirty male rats were chosen and randomly divided into five groups, with six rats in each group. Rats in groups 2 and 3 were administered the methanolic extract of Jacquemontia caerulea orally, while those in group 1, serving as the control, received 1ml of distilled water (DW). The fourth group was given indomethacin (standard 1mg/kg P.O). The drugs were administered orally, and one hour later, 0.05ml of a 1% carrageenan suspension was injected into the sub-plantar surface of the left hind paw to induce edema. Paw volume was measured initially at 0, 1, 2, and 4 hours after carrageenan injection using a plethysmometer.[20]
2. COTTON PELLET GRANULOMA METHOD:
   30 Wistar rats of both genders weighing between 150-200 grams (divided into five groups containing 6 in each), extract of Jacquemontia caerulea, cotton pellets, and normal saline.
   ANIMAL GROUPING:
   Group 1: control (n=6)
   Group 2: Test I (n=6) treated with methanolic extract of Jacquemontia caerulea
   Group 3: Test II (n=6) treated with methanolic extract of Jacquemontia caerulea
   Group.4: standard group (indomethacin 10 mg/kg by p.o)
   Procedure:
   Thirty rats were randomly distributed into five groups, with six rats in each group. Under ether anesthesia and with appropriate precautions, autoclaved cotton pellets were subcutaneously implanted on both sides above the scapula region. The administration of either Jacquemontia caerulea extract or 1ml of distilled water (DW) orally was carried out for 7 days starting from the day of pellet injection. On the 8th day, the rats were euthanized, and the pellets, along with the granuloma, were extracted and subjected to drying in an oven at 60°C until a constant weight was achieved.[21].The percentage inhibition of inflammation was calculated as inhibition of edema volume [22] as follows:
   Percentage inhibition of inflammation = 1-Vt/Vcx 100
   Where Vtis the average paw edema volume of extract of a treated group or positive control group.
   Vcis the paw volume of the control that received only vehicle.
   STATISTICAL ANALYSIS
   The data were studied with the help of one-way ANOVA followed by Dunnet’s multiple comparison test. The mean ± standard error of the mean (S.E.M) values were computed for each group. Statistical significance was considered at a P< 0.05 using GraphPad Prism version 10.1.

RESULTS

Effect Of Jacquemontia Caerulea On Cotton Pellet Wet Granuloma Method
The data were studied with the help of one-way ANOVA followed by Dunnet’s multiple comparison test. The mean ± S.E.M values were calculated for all groups. P< 0.05 was considered to be statistically significant.

Note: group 1: control, group 2: JCE-1, group 3: JCE-2, group 4: standard (Indomethacin)
JCE-Jacquemontia caerulea extract
WCPG-wet cotton pellet granuloma
Table no.3-Effect Of Jacquemontia Caerulea On Cotton Pellet Dry Granuloma Method

The data were studied with the help of one-way ANOVA followed by Dunnet’s multiple comparison test. The mean ± S.E.M values were calculated for all groups. P< 0.05 was considered to be statistically significant.

Note: group 1: control, group 2: JCE-1, group 3: JCE-2, group 4: standard (Indomethacin)
JCE-Jacquemontia caerulea extract
DCPG-dry cotton pellet granuloma
Table no.4-Effect Of Jacquemontia Caerulea On Carrageenan Induced Inflammation Method
The data were studied with the help of one-way ANOVA followed by Dunnet’s multiple comparison test. The mean ± S.E.M values were calculated for all groups. P< 0.05 was considered to be statistically significant.

Note: group 1: control, group 2: JCE-1, group 3: JCE-2, group 4: standard (Indomethacin) JCE-Jacquemontia caerulea extract
DISCUSSION
In this current research, samples from Jacquemontia caerulea were evaluated for their anti-inflammatory properties. In traditional drugs, Various natural substances have been employed for alleviating pain and reducing inflammation.
If the introduction of a foreign object into a host leads to tissue injury or harm, the host’s immune system responds by triggering an acute inflammatory reaction. This original response generally leads to minimum leukocyte accumulation and activation for the concurrence of the foreign body. Jacquemontia caerulea is a factory used for the treatment of painful and seditious conditions.
Methanol was selected as an extraction solvent to mimic the traditional mode of preparation of the plants. Phytochemical screening of the methanolic extract of Jacquemontia caerulea revealed the presence of considerable amounts of alkaloids, phenols, tannins, carbohydrates, glycosides, and amount of steroids. Several reports have shown the anti-inflammatory properties of flavonoids, tannins, and phenolic compounds in different experimental models, phytochemicals enhance the longevity of cells, prevent aging and minimizes predisposition to certain inflammatory disorders.
The methanolic extract of Jacquemontia caerulea showed a dose-dependent reduction in edema, similar to the standard indomethacin.
With the increasing need for alternatives to regulate the various forms of pro-oxidative and inflammatory processes, utilizing herbal products containing anti-inflammatory properties could represent a feasible choice.
Paw edema induced by carrageenan exhibits a two-phase reaction. The initial phase involves the release of histamine, serotonin, and kinins, while the second phase, occurring around 2-3 hours later, is attributed to the release of prostaglandin-like substances. Substances that impede carrageenan-induced paw edema may function by inhibiting both leukocyte migration and prostaglandin synthesis.

CONCLUSION:
In light of the current investigation, it can be deduced that the methanolic extract derived from the leaves of Jacquemontia caerulea possesses promising anti-inflammatory characteristics, introducing a novel aspect to pain management. Consequently, this study substantiates the traditional use of this plant in folklore for treating various painful conditions, offering pharmacological validation.These activities may be because of the existence of due to secondary metabolites such as alkaloids, flavonoids, tannins, steroids, and phenols. However, further study may be conducted to identify the compound that is responsible for possible anti-inflammatory effects and to fully elucidate the underlying mechanism of actions.
ACKNOWLEDGEMENT
I wish to acknowledge the assistance provided by my college management, colleagues, and students professionally and personally.
CONFLICT OF INTEREST
All authors approve the final manuscript and declare that there are no conflicts of interests.

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