1. Introduction

Bangladesh, a country blessed with lush landscapes and rich biodiversity, is home to an extraordinary variety of indigenous medicinal plants. For centuries, these plants have played a central role in traditional healthcare practices, particularly in rural communities where access to modern medical facilities is often limited. Far from being simply remnants of the past, these traditional remedies represent living systems of knowledge that continue to guide daily healthcare decisions for millions of people. Beyond cultural heritage, these plants carry untapped scientific promise, offering bioactive compounds that may lead to novel therapeutics in the modern pharmaceutical world (Faruque et al., 2018; Ahmad et al., 2016; Bilal et al., 2016).

The use of indigenous medicinal plants in Bangladesh is deeply interwoven with cultural practices and local traditions. Folk healers, often referred to as kabiraj or traditional practitioners, rely heavily on plants for the treatment of a broad range of health conditions, from digestive issues and infections to chronic inflammatory disorders. These healers not only provide remedies but also act as custodians of ethnobotanical knowledge, safeguarding centuries of empirical wisdom. For instance, communities living along the Ghaghut, Bengali, and Padma rivers have been documented as possessing extensive ethnomedicinal knowledge, passing down practices orally across generations (Faruque et al., 2018; Dirir et al., 2017). Such practices reveal a profound and practical understanding of the local flora, positioning traditional medicine as a valuable complement to modern healthcare systems.

Scientific exploration of these plants highlights their rich phytochemical diversity. Many species are endowed with alkaloids, flavonoids, tannins, glycosides, and saponins—compounds known for their therapeutic roles. These phytochemicals are not mere chemical curiosities; they form the basis for pharmacological activities such as antimicrobial, anti-inflammatory, antioxidant, and even anticancer effects. For example, flavonoids are celebrated for their ability to neutralize oxidative stress and mitigate inflammation, while saponins are widely recognized for antimicrobial properties (Akhtar, 2022; Grochowski et al., 2019; Kähkönen et al., 1999). These findings lend scientific credibility to traditional claims, bridging the gap between ancient wisdom and modern validation.

One of the most pressing health concerns in Bangladesh is the persistent burden of infectious diseases. Rural populations, in particular, face heightened vulnerability due to limited access to antibiotics and rising concerns about antimicrobial resistance. Medicinal plants present a promising avenue for addressing these challenges. Several studies have demonstrated that extracts from Bangladeshi plants show notable antimicrobial activity against a spectrum of pathogens (Siddique et al., 2021; Gemeda et al., 2018; Pawar & Khan, 2007). Such discoveries are timely, given the global urgency to discover alternatives in the face of antibiotic resistance.

The antimicrobial effects of these plants often stem from their ability to disrupt vital microbial processes. Phytochemicals can interfere with cell wall synthesis, impair protein production, and destabilize microbial membranes, making them highly versatile defense tools (Kadir et al., 2012; Khurm et al., 2016). This pharmacological potential suggests that Bangladeshi medicinal plants should not be viewed merely as adjuncts to conventional medicine but rather as strong candidates for the development of entirely new classes of therapeutic agents.

In addition to fighting infections, Bangladeshi medicinal plants have long been valued for their wound-healing properties. Traditional healers frequently apply plant extracts, pastes, or poultices to cuts, burns, and chronic sores, and such practices are now being validated by scientific research (Sidhu et al., 1999; Varoglu et al., 2010; Murthy et al., 2013; Permata et al., 2024). Phytochemicals like flavonoids and tannins have been shown to promote tissue regeneration, reduce inflammation, and accelerate the closure of wounds (Sairam et al., 2001; Sharath et al., 2010). For instance, Bacopa monnieri has demonstrated significant wound-healing activity in animal models, improving both wound contraction and epithelialization time (Murthy et al., 2013; Sairam et al., 2001). Similarly, Calotropis gigantea, traditionally used for treating skin ailments, has been reported to enhance wound repair through mechanisms involving anti-inflammatory action and collagen synthesis (Permata et al., 2024; Argal & Pathak, 2006). These findings not only validate indigenous practices but also highlight the importance of preserving traditional knowledge as a reservoir of biomedical innovation.

Importantly, the integration of herbal medicine into modern healthcare is no longer seen as optional but rather as essential. The World Health Organization has long advocated for recognizing traditional medicine as part of national healthcare strategies, particularly in countries like Bangladesh where plant-based remedies remain widely used. However, successful integration requires careful balancing of cultural respect with scientific rigor. Traditional remedies must undergo stringent pharmacological evaluation to ensure efficacy and safety before they can be formally adopted into mainstream medicine (Nahar et al., 2024; Anders et al., 2010).

The study of indigenous medicinal plants is not only about discovering new drugs but also about honoring cultural heritage and addressing unmet health needs. In Bangladesh, the reliance on medicinal plants is both a matter of tradition and necessity, particularly in remote areas where conventional healthcare may be scarce. As scientific evidence continues to mount, it becomes increasingly clear that these plants represent a vital bridge between the wisdom of the past and the innovations of the future. Exploring their therapeutic potential aligns with global health priorities, particularly in the fight against infectious diseases, chronic inflammation, and wound management (Bilal et al., 2016; Gemeda et al., 2018).

Therefore, the present study seeks to evaluate the wound-healing efficacy of methanolic extracts from selected indigenous plants of Bangladesh using in vivo excision wound models. By focusing on Alpinia nigra, Bacopa monnieri, Calotropis gigantea, and Cynodon dactylon, this research not only tests traditional claims under controlled conditions but also contributes to the growing body of scientific evidence supporting the pharmacological importance of Bangladeshi flora (Annapurna et al., 2013; Dande & Khan, 2012). Ultimately, such efforts may pave the way for developing affordable, accessible, and culturally relevant medicines, bringing together the best of tradition and science.

2. Materials and Methods

2.1 Study Design

This study was designed as an experimental investigation to explore the wound-healing efficacy and antimicrobial activities of methanolic extracts obtained from four indigenous Bangladeshi medicinal plants: Alpinia nigra, Bacopa monnieri, Calotropis gigantea, and Cynodon dactylon. These plants were selected based on ethnobotanical reports of their therapeutic applications in wound care and infection management (Faruque et al., 2018; Kadir et al., 2012). The study integrated both in vitro assays to assess antimicrobial potential and in vivo animal models to evaluate wound-healing properties, ensuring a comprehensive pharmacological evaluation.

The methodology followed internationally accepted experimental pharmacology guidelines, with an emphasis on scientific reproducibility, ethical standards, and alignment with the principles of the 3Rs—Replacement, Reduction, and Refinement (Murthy et al., 2013).

2.2 Plant Collection and Authentication

Fresh plant materials were collected during the dry season (December–February) from different districts of Bangladesh known for their rich medicinal flora, including Sylhet, Rangamati, and Rajshahi. Local folk healers, with extensive ethnomedicinal knowledge, assisted in plant identification, which ensured that culturally validated species were prioritized (Siddique et al., 2021).

The plants were authenticated by a taxonomist from the Department of Botany, University of Dhaka. Voucher specimens were deposited in the university herbarium for future reference, and each was assigned an accession number for traceability.

2.3 Preparation of Plant Extracts

Collected plant materials—leaves of Bacopa monnieri, rhizomes of Alpinia nigra, latex of Calotropis gigantea, and aerial parts of Cynodon dactylon—were thoroughly washed with distilled water to remove soil and debris. They were shade-dried at room temperature (25–28 °C) to preserve sensitive phytoconstituents and then powdered using a mechanical grinder.

For each plant, 500 g of powdered material was subjected to Soxhlet extraction using methanol as the solvent, chosen for its ability to extract a wide spectrum of polar and non-polar bioactive compounds (Akhtar, 2022). Extraction continued until the solvent became colorless, indicating exhaustive extraction. The filtrates were concentrated under reduced pressure using a rotary evaporator at 40 °C and subsequently stored at 4 °C in amber-colored vials to prevent light-induced degradation.

2.4 Preliminary Phytochemical Screening

Qualitative phytochemical screening was carried out following standard pharmacognostic protocols. Tests for alkaloids (Mayer’s and Dragendorff’s reagents), flavonoids (alkaline reagent test), tannins (ferric chloride test), saponins (froth test), and glycosides (Keller-Kiliani test) were conducted. These phytochemicals are widely recognized as contributors to wound healing and antimicrobial activity (Siriwatanametanon et al., 2010).

2.5 Experimental Animals

Adult Wistar albino rats (Rattus norvegicus) weighing 150–200 g were procured from the Bangladesh Council of Scientific and Industrial Research (BCSIR) animal facility. The rats were housed under standard laboratory conditions: temperature maintained at 25 ± 2 °C, relative humidity at 50–60%, and a 12-hour light/dark cycle. Standard pellet diet and water were provided ad libitum.

Before initiating experiments, animals were acclimatized for seven days. All procedures strictly followed the guidelines of the Committee for the Purpose of Control and Supervision of Experiments on Animals (CPCSEA) and were approved by the Institutional Animal Ethics Committee (Ethics approval: DGDA/CRO-4/2017/9170).

2.6 Experimental Grouping and Sample Size

Thirty-six rats were randomly divided into six groups, with six animals per group (n = 6). The grouping was as follows:

Control Group – treated with ointment base only.

Standard Group – treated with 5% povidone-iodine ointment (reference drug).

Test Group 1 – treated with 5% w/w ointment of Alpinia nigra extract.

Test Group 2 – treated with 5% w/w ointment of Bacopa monnieri extract.

Test Group 3 – treated with 5% w/w ointment of Calotropis gigantea extract.

Test Group 4 – treated with 5% w/w ointment of Cynodon dactylon extract.

The sample size was selected in accordance with prior wound-healing studies using rodent models, which typically employ 5–6 rats per group to achieve statistical significance (Permata et al., 2024).

2.7 Wound Healing Activity (Excision Model)

The excision wound model was selected as it closely mimics clinical wound healing (Murthy et al., 2013). Animals were anesthetized using ketamine (50 mg/kg) and xylazine (10 mg/kg) intraperitoneally. After shaving and sterilizing the dorsal thoracic region, a full-thickness circular wound (8 mm in diameter) was created using sterile surgical scissors. Hemostasis was achieved via blotting with sterile cotton swabs.

Topical treatments (ointment base, standard, or extract formulations) were applied once daily until complete healing.

Wound contraction was measured on days 4, 8, 12, and 16 post-wounding using a transparent graph sheet to trace wound margins. The wound area was then quantified by planimetry. Wound contraction percentage was calculated as:

Epithelialization time, the number of days required for complete wound closure without residual raw surface, was also recorded. Researchers conducting measurements were blinded to treatment groups to minimize bias.

2.8  Antimicrobial Activity

The antimicrobial potential of extracts was tested using the agar disc diffusion method (Kadir et al., 2012). Clinical isolates of Staphylococcus aureus (Gram-positive), Escherichia coli (Gram-negative), Candida albicans (yeast), and Aspergillus niger (filamentous fungus) were selected to represent common wound pathogens.

Sterile discs (6 mm) were impregnated with 100 µL of plant extract solutions at concentrations of 100, 200, and 400 µg/disc and placed on Mueller-Hinton agar plates inoculated with bacterial or fungal suspensions standardized to 0.5 McFarland units. Plates were incubated at 37 °C for 24 hours (bacteria) or 28 °C for 48 hours (fungi). Zones of inhibition were measured in millimeters. Methanol served as a negative control, while ciprofloxacin (10 µg/disc) and ketoconazole (10 µg/disc) served as positive controls for antibacterial and antifungal assays, respectively.

2.9 Data Collection and Analysis

For wound healing studies, wound contraction percentages and epithelialization times were recorded as mean ± standard deviation (SD). Normality of data distribution was confirmed using the Shapiro–Wilk test. One-way ANOVA followed by Tukey’s post hoc test was employed to identify significant differences among treatment groups.

For antimicrobial assays, zones of inhibition were expressed as mean ± SD. Data were analyzed by two-way ANOVA, assessing the effect of both extract concentration and plant type. Post hoc analysis was performed with Tukey’s test.

All analyses were conducted using SPSS version 26.0, and p-values < 0.05 were considered statistically significant. This approach ensured methodological rigor and minimized type I and II errors (Saraf et al., 2011).

2.10 Ethical Considerations

Animal welfare was prioritized throughout the experiment. Surgical wounds were created under anesthesia to minimize pain, and animals were monitored daily for signs of distress. Humane endpoints were predefined, including excessive weight loss or infection, though none occurred.

Ethical clearance was obtained from the Ethics Review Committee of Probe International Clinical Research Ltd. (ERC: DGDA/CRO-4/2017/9170). All experiments adhered to ARRIVE guidelines for animal research reporting, ensuring transparency and reproducibility.

3. Results

The present study explored the antimicrobial and wound healing potential of four indigenous medicinal plants of Bangladesh—Alpinia nigra, Bacopa monnieri, Calotropis gigantea, and Cynodon dactylon—through detailed phytochemical analysis, antimicrobial assays, and excision wound models in Wistar rats. The results demonstrated that these plants, long valued in traditional medicine, indeed harbor bioactive compounds with promising therapeutic effects.

3.1 Phytochemical Screening

Preliminary phytochemical analysis confirmed the presence of alkaloids, flavonoids, tannins, saponins, and glycosides in all four methanolic extracts. These secondary metabolites are widely recognized for their pharmacological relevance. For example, flavonoids act as powerful antioxidants and modulators of inflammation, while saponins are known for their antimicrobial and immunomodulatory properties (Table 1) (Akhtar, 2022). The consistent detection of these compounds across the tested species reinforces the ethnomedicinal claims that these plants promote healing, reduce infections, and restore tissue integrity.

Table 1. Phytochemical Constituents of Methanolic Extracts

Key: "+" indicates presence of the phytochemical.

3.2 Antimicrobial Activities

The antimicrobial assays revealed a concentration-dependent inhibition of microbial growth across all extracts. Among them, Alpinia nigra displayed the most notable antibacterial activity, producing inhibition zones up to 18 mm against Staphylococcus aureus and 16 mm against Escherichia coli at the highest concentration. Similarly, Bacopa monnieri showed pronounced antibacterial activity against E. coli, reaching an 18 mm inhibition zone (Table 2). On the other hand, Calotropis gigantea stood out for its antifungal efficacy, particularly against Candida albicans, producing an inhibition zone of 17 mm. In contrast, Cynodon dactylon demonstrated comparatively lower antimicrobial activity, though still exhibiting measurable effects against all tested pathogens.

Table 2. Zones of Inhibition (mm) of Plant Extracts Against Test Organisms

These findings echo earlier ethnopharmacological surveys documenting the antimicrobial use of Bangladeshi medicinal plants (Kadir et al., 2012; Siddique et al., 2021). The strong antibacterial activity of Alpinia nigra and Bacopa monnieri is particularly relevant in the context of Bangladesh, where infectious diseases remain a major public health challenge and drug-resistant strains of bacteria are increasing (Faruque et al., 2018). The antifungal efficacy of Calotropis gigantea aligns with recent evidence emphasizing its wound-healing and antimicrobial roles (Permata et al., 2024).

3.3 Wound Healing Activities

In the excision wound model, distinct differences emerged between treatment groups. The control group, treated with simple ointment, exhibited the slowest healing process, with only 60% wound contraction and a mean epithelialization time of 20 days. By contrast, the standard group treated with povidone-iodine showed nearly complete wound closure (95%) and the shortest healing time (14 days) (Table 3).

Table 3. Wound Healing Parameters of Methanolic Extracts

Among the plant extracts, Alpinia nigra performed remarkably well, achieving 92% wound contraction and full epithelialization in 14 days, nearly matching the standard treatment. Cynodon dactylon and Calotropis gigantea also demonstrated strong wound healing activity, with 90% and 88% contraction, respectively, and mean epithelialization times of 15 days. Bacopa monnieri, while effective, showed slightly slower healing, with 85% contraction and 16 days to epithelialization.

The improved wound healing observed in these groups can be attributed to the combined effects of phytochemicals such as flavonoids and tannins, which promote collagen synthesis, angiogenesis, and tissue regeneration, while also reducing oxidative stress at the wound site (Murthy et al., 2013; Siriwatanametanon et al., 2010). Importantly, the performance of Alpinia nigra nearly equaled that of povidone-iodine, suggesting its potential as a plant-based alternative for topical wound management.

3.4 Integrated Findings

Taken together, these results affirm that indigenous Bangladeshi plants possess both antimicrobial and wound healing properties, supporting their long-standing traditional use. The significant healing efficacy of Alpinia nigra, combined with the antifungal potency of Calotropis gigantea and the antibacterial activity of Bacopa monnieri, highlight the therapeutic diversity within this group. The findings underscore the importance of preserving and scientifically validating traditional knowledge, as these plants may provide novel, culturally relevant, and cost-effective solutions to common health challenges in Bangladesh (Nahar et al., 2024).

4. Discussion

The present study examined the antimicrobial and wound healing potential of methanolic extracts from four indigenous medicinal plants of Bangladesh—Alpinia nigra, Bacopa monnieri, Calotropis gigantea, and Cynodon dactylon. The findings strongly support the traditional use of these plants in local healing practices, while also underscoring their potential as candidates for future drug development.

4.1 Phytochemical Significance

The phytochemical screening revealed that all extracts contained alkaloids, flavonoids, tannins, saponins, and glycosides. These bioactive compounds are well-documented for their pharmacological actions, including antimicrobial, antioxidant, and anti-inflammatory properties. Flavonoids, for example, play a crucial role in scavenging free radicals and reducing oxidative stress, which is essential in the wound-healing cascade (Akhtar, 2022). Similarly, saponins are associated with antimicrobial and immunomodulatory activities, which may contribute to their wound-healing benefits (Kadir et al., 2012). The consistent presence of these compounds across the selected plants highlights a shared therapeutic potential rooted in their chemical composition.

4.2 Antimicrobial Properties

The antimicrobial assays demonstrated concentration-dependent inhibition across bacterial and fungal pathogens. Alpinia nigra showed the strongest antibacterial action against Staphylococcus aureus and Escherichia coli, while Calotropis gigantea displayed significant antifungal activity against Candida albicans. These results resonate with earlier ethnopharmacological surveys that documented the antimicrobial potential of Bangladeshi medicinal plants in combating infectious diseases (Siddique et al., 2021). The activity of Bacopa monnieri against E. coli further reinforces its traditional application in treating gastrointestinal and infectious conditions (Murthy et al., 2013). In contrast, Cynodon dactylon exhibited relatively milder antimicrobial activity, yet its consistent effect suggests synergistic roles in broader therapeutic applications.

The antimicrobial actions observed can be attributed to multiple mechanisms, such as disruption of microbial cell walls, inhibition of protein synthesis, and interference with vital cellular pathways (Faruque et al., 2018). Considering the global rise in antimicrobial resistance, the efficacy of these extracts underscores their relevance as potential sources of new antimicrobial agents.

4.3 Wound Healing Potential

The wound healing study revealed that Alpinia nigra had the most pronounced effect, nearly matching the standard povidone-iodine treatment in terms of wound contraction and epithelialization time. Calotropis gigantea and Cynodon dactylon also promoted substantial wound closure, while Bacopa monnieri demonstrated moderate yet notable efficacy. These findings align with reports highlighting the wound-healing properties of Bacopa monnieri and Calotropis gigantea, both of which have been traditionally employed for skin-related ailments (Permata et al., 2024; Murthy et al., 2013). The observed healing activity is likely linked to the presence of flavonoids and tannins, which not only reduce inflammation but also facilitate collagen synthesis and tissue regeneration (Siriwatanametanon et al., 2010).

The study’s results carry practical implications for primary healthcare in rural Bangladesh, where access to modern medicine is often limited. By validating traditional practices through scientific methods, these findings create a bridge between indigenous knowledge and modern biomedical research (Nahar et al., 2024).

4.5 Implications and Future Directions

While the results are promising, several limitations remain. The study was restricted to in vivo animal models, and clinical trials are necessary to confirm efficacy and safety in humans. Moreover, further isolation and characterization of the active compounds would be valuable for identifying the exact mechanisms underlying their therapeutic actions. Future research could also explore synergistic effects of combining different plant extracts, potentially enhancing antimicrobial and wound-healing outcomes.

5. Conclusion

Overall, the findings provide compelling evidence that methanolic extracts of Alpinia nigra, Bacopa monnieri, Calotropis gigantea, and Cynodon dactylon possess significant antimicrobial and wound-healing activities. These results not only validate long-standing traditional uses but also highlight the potential of indigenous medicinal plants as a foundation for novel therapeutic agents in modern medicine.

Author Contributions

N. M. Badhon conceptualized the study, designed experiments, conducted phytochemical and antimicrobial analyses, performed in vivo wound-healing experiments, analyzed and interpreted the data, and wrote the manuscript.

Acknowledgement

None declared 

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