WOUND HEALING POTENTIAL OF BACOPA MONNIERIIN ACID BURN & BOILING WATER INDUCED WOUNDS

Abstract

The present invention relates to a wound healing composition comprising Bacopa monnieri (BM) leaf extract in varying concentrations of 20%, 50%, and 100%. The composition is formulated for topical application to promote wound healing in burn-induced wounds. Bacopa monnieri, a medicinal plant known for its therapeutic properties, has been shown to enhance collagen synthesis and accelerate wound healing through the increase of hydroxyproline content, wound contraction, and reduction in epithelization time. The composition is prepared by extracting BM leaves with a hydroalcoholic solution, followed by concentration and formulation into a cream. The wound healing potential of the BM leaf extract was evaluated in animal models using both acid burn and thermal burn (boiling water) wound models. This invention provides a promising natural remedy for effective wound healing, offering an alternative to conventional wound care treatments.

Specification

Description:FIELD OF INVENTION
The present invention relates to wound healing and therapeutic formulations, specifically exploring the potential of Bacopa monnieri extracts for promoting tissue repair in acid burn and boiling water-induced wounds.
BACKGROUND OF THE INVENTION
Burn wounds, particularly those caused by acid exposure and boiling water, present unique challenges in wound care due to their potential for deep tissue damage, infection risk, and delayed healing. Effective treatment options for these injuries are essential to prevent complications, such as infection, scarring, and prolonged inflammation. While several synthetic treatments exist, there is growing interest in natural therapies due to their lower side effects, cost-effectiveness, and often potent healing properties. Traditional herbal medicines have shown promise in promoting wound healing by leveraging bioactive compounds found in plants known for their regenerative effects.
Bacopa monnieri, commonly known as Brahmi, is a medicinal herb renowned in traditional Ayurvedic medicine for its cognitive and anti-inflammatory benefits. Recent research has highlighted its potential beyond neurological applications, showing it to possess strong antioxidant, antimicrobial, and anti-inflammatory properties, which are crucial in wound healing. The bioactive compounds, such as bacosides, present in Bacopa monnieri can enhance collagen synthesis, reduce oxidative stress, and aid in tissue repair, making it an ideal candidate for wound treatment applications. Its efficacy in the healing process, especially in challenging wound types like acid burns, holds promise for accelerating tissue regeneration and improving outcomes.
This invention explores the application of Bacopa monnieri extracts in treating acid and boiling water-induced wounds by developing a formulation specifically targeted at promoting rapid and effective healing. Through controlled trials and assessments, this formulation aims to validate the effectiveness of Bacopa monnieri in wound care, offering a natural alternative to synthetic treatments that supports skin recovery, minimizes scarring, and leverages the plant's bioactive compounds for therapeutic benefits. This research builds on traditional knowledge with modern scientific validation, aiming to expand the scope of Bacopa monnieri in dermatological and wound healing applications.
OBJECTS OF THE INVENTION
Some of the objects of the present disclosure, which at least one embodiment herein satisfies, are as follows.
It is an object of the present disclosure to ameliorate one or more problems of the prior art or to at least provide a useful alternative
An object of the present disclosure is to promotes faster wound healing through enhanced collagen synthesis.
Another object of the present disclosure is to reduce inflammation, minimizing pain and swelling at the wound site.
Still another object of the present disclosure is providing strong antioxidant protection, reducing oxidative stress on skin tissue.
Another object of the present disclosure is to possess antimicrobial properties, lowering infection risks in wounds.
Still another object of the present disclosure is to improve tissue regeneration, aiding recovery of both superficial and deep wounds.
Still another object of the present disclosure is to offer a natural, side-effect-free alternative to synthetic wound treatments.
Yet another object of the present disclosure is to minimize scar formation, promoting smoother skin recovery.
Yet another object of the present disclosure is cost-effective and accessible, a widely available herbal remedy.
Yet another object of the present disclosure is to accelerates wound healing and tissue regeneration by boosting collagen synthesis and reducing inflammation.
Yet another object of the present disclosure is to provides antioxidant and antimicrobial protection, lowering infection risk and oxidative stress on healing tissues.

Other objects and advantages of the present disclosure will be more apparent from the following description, which is not intended to limit the scope of the present disclosure.

SUMMARY OF THE INVENTION
The following presents a simplified summary of the invention in order to provide a basic understanding of some aspects of the invention. This summary is not an extensive overview of the present invention. It is not intended to identify the key/critical elements of the invention or to delineate the scope of the invention. Its sole purpose is to present some concept of the invention in a simplified form as a prelude to a more detailed description of the invention presented later.

The present invention is generally directed to the wound healing potential of Bacopa monnieri (BM) leaf extract in two wound models: acid burns and boiling water-induced burns. The study aimed to assess the efficacy of various concentrations (20%, 50%, and 100%) of BM leaf extract cream in promoting wound healing. BM leaves were collected, authenticated, and extracted using a hydroalcoholic solution. The resulting extract was formulated into a topical cream and applied to Wistar albino rats to evaluate its wound healing effects over 21 days.
An embodiment of the present invention involved a comparison of BM leaf extract with a standard wound healer (Lobate Gm) and a cream base in five groups of animals. Key parameters such as hydroxyproline content, wound contraction, and epithelization period were measured to evaluate the wound healing activity. The findings demonstrated a dose-dependent effect, with higher concentrations of BM leaf extract significantly improving the healing process compared to the control groups.
An embodiment of the present invention highlights Bacopa monnieri as a promising natural wound healing agent. The results suggest that BM leaf extract possesses significant potential in promoting collagen synthesis, wound contraction, and reducing the epithelization period, making it a valuable candidate for the development of wound healing formulations.

DETAILED DESCRIPTION OF THE INVENTION
The following description is of exemplary embodiments only and is not intended to limit the scope, applicability or configuration of the invention in any way. Rather, the following description provides a convenient illustration for implementing exemplary embodiments of the invention. Various changes to the described embodiments may be made in the function and arrangement of the elements described without departing from the scope of the invention.
The present invention relates to the wound healing potential of Bacopa monnieri (BM) leaf extract, particularly in acid burn and boiling water-induced wounds. The study involved the extraction of BM leaves using a hydroalcoholic solution, which was then formulated into a cream at varying concentrations (20%, 50%, and 100%) and tested on Wistar albino rats. The efficacy of BM leaf extract was evaluated by measuring key parameters such as hydroxyproline content, wound contraction, and epithelization period over a 21-day period.

EXAMPLE 1: Collection, Authentication, and Extraction
Fresh leaves of Bacopa monnieri (BM) were collected and authenticated by a botanist at MJPRU, Bareilly. The leaves were thoroughly washed to remove any soil residues, finely chopped, and weighed (20g). The chopped leaves were then soaked in a hydroalcoholic solution (1:1) for 15 days, with frequent stirring to enhance extraction. After this period, the solution was initially filtered through a cotton plug, followed by filtration using Whatman filter paper to obtain a clear extract. The extract was then concentrated using a rotary evaporator at 40ºC, dried, and stored in a desiccator until further use.
The % yield was obtained as 69% for BM leaves when calculated with theoretical and practical.
EXAMPLE 2: Preparation of animals
Wistar albino rats of either sex weighing 150- 200g were obtained from the animal House, RBCP Bareilly. The animals are kept in good health, with room temperatures of 25°C and a 12-hour light/dark cycle. The relative humidity is kept at 44-56 percent, and the rats are provided a regular rodent pellet chow diet and free access to water.
To evaluate the wound healing potential of BM leaves extract, 6 rats were taken in each group.
Animals were divided into 5 following groups-
Group I: applied base used in cream.
Group II: applied Lobate Gm once daily, for 21 days.
Group III: applied BM leavesextract cream (20%) once daily, for 21 days.
Group IV: applied BM leavesextract cream (50%) once daily, for 21 days.
Group V: applied BM leavesextract cream (100%) once daily, for 21 days.
EXAMPLE 3: Evaluation of Wound Healing Potential of Bacopa monnieri (BM) Leaf Extract
To assess the wound healing effects of BM leaf extract, 30 rats were divided into five groups, with six rats per group, as follows:
• Group I: Applied with the base cream only.
• Group II: Treated with Lobate GM cream once daily for 21 days.
• Group III: Treated with BM leaf extract cream (20%) once daily for 21 days.
• Group IV: Treated with BM leaf extract cream (50%) once daily for 21 days.
• Group V: Treated with BM leaf extract cream (100%) once daily for 21 days.
On the day of the experiment, the animals were anesthetized via intraperitoneal injection of ketamine and xylazine (5:1). The lateral dorsal area was shaved using a hair removal ointment and disinfected with spirit. To induce acid burns, 0.5 ml of a 50% v/v HCl solution was applied dropwise to the shaved area, with contact maintained for 15 seconds to ensure consistent burn creation. Key metrics-percentage of wound contraction, epithelialization period, and hydroxyproline content-were measured over a 21-day observation period to evaluate the extract's wound healing efficacy.

EXAMPLE 4: Wound Healing Activity
Hydroxyproline Content
To measure hydroxyproline content, wound tissue samples from all rats in both wound models were collected. These tissues were initially dried at 60°C to a constant weight and then hydrolyzed in 6N HCl at 130°C for 4 hours. After hydrolysis, the pH was adjusted to 7.0, and 0.4M perchloric acid was added to halt further reactions. Color development was achieved by adding Ehrlich's reagent to the solution, followed by incubation at 60°C with continuous stirring. The absorbance of the resulting solution was measured using a UV spectrophotometer, and hydroxyproline content was calculated by comparing results to a standard curve prepared with pure L-hydroxyproline.

Table 1. Effect of BM leaves extracton hydroxyproline content in wound tissues

Group
Treatment Hydroxyproline content (mg/100g tissue)
Acid burn model Boiling water model
Group I Cream base 1.25±1.45** 1.57±1.20**
Group II Lobate Gm 5.10±2.57*** 4.99±2.31***
Group III BM leaves extract (20%) 1.83±1.51* 2.30±1.62*
Group IV BM leaves extract (50%) 2.12±1.33** 3.51±1.71**
Group V BM leaves extract (100%) 3.81±2.24** 4.10±2.41***

Results shows that in the control group, hydroxyproline content was minimal, while the standard treatment group exhibited the highest levels. Treatment with BM leaf extract demonstrated significant wound healing potential by increasing hydroxyproline levels. The hydroxyproline content observed in animals treated with BM leaf extract was 1.83 ± 1.51* for 20% concentration, 2.12 ± 1.33** for 50% concentration, and 3.81 ± 2.24** for 100% concentration. These results indicate a dose-dependent effect, with higher doses of BM extract contributing to increased collagen synthesis and enhanced wound healing.

Wound Contraction Method
In both models, burn wound areas were recorded over a 21-day period. Burn boundaries were traced on transparent sheets and measured against graph paper. Wound contraction was assessed every third day using the formula:

Table 2. Effect of BM leaves extract on wound contraction in wound tissues
Group Treatment % of wound contraction
Acid burn model Boiling water model
Group I Cream base 52.21±1.15* 53.37±1.30***
Group II Lobate Gm 78.11±2.27** 81.29±2.11**
Group III BM leaves extract (20%) 63.43±1.61*** 64.20±1.32***
Group IV BM leaves extract (50%) 67.11±1.43** 68.31±1.41**
Group V BM leaves extract (100%) 75.41±2.34** 77.10±2.11**
Significance Level: *
Values are expressed in Mean± SEM; (n=6)

Results shows that in the acid burn model, the percentage of wound contraction was 63.43 ± 1.61* for the 20% BM leaf extract, 67.11 ± 1.43** for the 50% dose, and 75.41 ± 2.34** for the 100% dose. In the boiling water burn model, the wound contraction percentages were 64.20 ± 1.32*** for the 20% BM leaf extract, 68.31 ± 1.41** for the 50% dose, and 77.10 ± 2.11** for the 100% dose, showing a highly significant effect. The following table summarizes the wound contraction observed in both the acid burn and boiling water models.

Measurement of Epithelization Period
The epithelization period for both acid burn and thermal burn models was determined by counting the days until dead tissue sloughed off, leaving fully closed wounds. This metric provided an estimate of healing time across the two burn models.

Table 3. Effect of BM leaves extract cream on epithelization period in wound tissues
Group Treatment epithelization period
Acid burn model Boiling water model
Group I Cream base 22.50±1.12* 23.27±1.17*
Group II Lobate Gm 15.17±2.21** 13.79±2.30***
Group III BM leaves extract (20%) 20.41±1.40* 21.10±1.41*
Group IV BM leaves extract (50%) 18.11±1.23** 17.21±1.61**
Group V BM leaves extract (100%) 17.41±2.52** 15.10±.32*

Significance Level: *
Values are expressed in Mean± SEM; (n=6).

A decrease in the epithelization period indicates faster wound healing. The longest epithelization period was observed in Group I, treated with the cream base, at 22.50 ± 1.12*, while the shortest period was seen in Group II, treated with the standard wound healer Lobate Gm, at 15.17 ± 2.21**. BM leaf extract demonstrated excellent wound healing properties, with epithelization periods of 20.41 ± 1.40*, 18.11 ± 1.23**, and 17.41 ± 2.52** for doses of 20%, 50%, and 100%, respectively, in the acid burn model. In both the acid burn and boiling water models, BM leaf extract exhibited a significant ability to promote wound healing.

Results clearly shows that the analysis of hydroxyproline content, wound contraction, and epithelization time revealed that BM leaf extract effectively accelerated the wound healing process across all parameters.

While considerable emphasis has been placed herein on the specific features of the preferred embodiment, it will be appreciated that many additional features can be added and that many changes can be made in the preferred embodiment without departing from the principles of the disclosure. These and other changes in the preferred embodiment of the disclosure will be apparent to those skilled in the art from the disclosure herein, whereby it is to be distinctly understood that the foregoing descriptive matter is to be interpreted merely as illustrative of the disclosure and not as a limitation.
, Claims:We Claim,
1. A wound healing composition comprising Bacopa monnieri leaf extract in varying concentrations of 20%, 50%, and 100%, for topical application, wherein the composition demonstrates enhanced wound healing effects by increasing hydroxyproline content, accelerating wound contraction, and reducing the epithelization period in burn-induced wounds.
2. The wound healing composition as claimed in claim 1, wherein the Bacopa monnieri leaf extract is prepared by soaking fresh leaves in a hydroalcoholic solution (1:1) for 15 days, followed by filtration and concentration to yield a 69% extract.
3. The wound healing composition as claimed in claim 1, wherein the Bacopa monnieri leaf extract is used in a cream formulation and applied topically once daily for 21 days to promote wound healing.
4. The wound healing composition as claimed in claim 1, wherein the wound healing potential of Bacopa monnieri leaf extract is demonstrated by an increase in hydroxyproline content, with values of 1.83 ± 1.51*, 2.12 ± 1.33**, and 3.81 ± 2.24** for 20%, 50%, and 100% concentrations, respectively, in an acid burn model.
5. The wound healing composition as claimed in claim 1, wherein the percentage of wound contraction in the acid burn model is 63.43 ± 1.61* for 20%, 67.11 ± 1.43** for 50%, and 75.41 ± 2.34** for 100% Bacopa monnieri leaf extract, and in the boiling water model is 64.20 ± 1.32***, 68.31 ± 1.41**, and 77.10 ± 2.11**, respectively.
Dated this 12 November 2024

Dr. Amrish Chandra
Agent of the applicant
IN/PA No: 2959

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