Evaluation of wound healing activity of herbal gel formulation of Tridax procumbens and Aloe vera in goats

Introduction

A wound is defined as the disruption of the normal structure and continuity of the skin due to physical, chemical, or biological injury. The skin acts as a protective barrier, and any damage initiates a complex healing process involving haemostasis, inflammation, proliferation, and remodelling. Proper progression of these phases is essential for effective wound repair (Shivhare et al., 2019;Guo and DiPietro, 2010). Goats play an important role in rural economy, and wounds are common due to their active nature and exposure to environmental hazards. Such injuries can lead to infection, delayed healing, and economic losses, highlighting the need for effective wound management strategies (Department of Animal Husbandry and Dairying, 2019; Farghali et al., 2020). Herbal medicines have gained attention as alternative therapeutic agents due to their antimicrobial, anti-inflammatory, and tissue-regenerating properties. Among them, Tridax procumbens and Aloe vera are widely recognized for their wound healing potential and are commonly used in traditional medicine (Ghosh et al., 2019;Manvitha and Bidya, 2014). Therefore, the present study was undertaken to evaluate the wound healing efficacy and safety of a herbal gel formulation containing Tridax procumbens and Aloe vera in goats [20, 12, 8, 10, 11, 18] .

Materials and methods

Collection and authentication of plant material

Fresh leaves of Tridax procumbens were collected from the Parbhani region and authenticated by a botanist at VNMKV, Parbhani. The leaves were washed thoroughly, shade-dried at room temperature (25-30 °C) for 7-10 days, and ground into fine powder after achieving constant weight. Fresh Aloe vera leaves were collected on the day of formulation and authenticated similarly. The leaves were washed, outer rind removed, and inner mucilaginous pulp was separated. The pulp was homogenized, filtered to remove fibrous material, and used in fresh form for preparation of herbal gel formulations.

Preparation of Herbal Formulation

The gel base was prepared by dispersing Carbopol-940 (2% w/v) in distilled water with continuous stirring and allowing it to hydrate for 8-10 hours. Triethanolamine (0.5% v/v) was then added gradually to obtain a uniform gel base. For preparation of the herbal gel, powdered Tridax procumbens leaves and fresh Aloe vera pulp were incorporated into the gel base at concentrations of 10% and 5%. The mixture was stirred continuously to achieve uniform consistency, and the final volume was adjusted with distilled water.

Acute Dermal Toxicity Study of herbal formulation

Result and discussion

Dermal reaction of herbal formulation

The dermal toxicity was conducted in female Wistar rats to assess dermal tolerability of the herbal gel formulation. Healthy animals with intact skin were selected, and the dorsal area was shaved approximately 24 hours prior to application. The formulation was applied topically at a limit dose of 2000 mg per kilogram body weight over approximately 10% of the body surface area, and the site was covered with gauze and secured with a non-irritant dressing for 24 hours and animals were observed at 24, 48, and 72 hours for dermal reactions as per OECD guideline 402.No dermal irritation, clinical abnormalities, or systemic toxicity was observed in any group throughout the observation period, indicating that both formulations were safe for topical application.

Weekly feed consumption: Weekly feed intake of rats in both groups showed a gradual increase over the study period (table 2). In group B, mean feed intake increased from 91.36 ± 0.38 g in the first week to 93.50 ± 0.47 g in the second week. Similarly, in group C, feed intake increased from 91.14 ± 0.32 g to 92.71 ± 0.47 g. Statistical analysis indicated that the differences between the groups were non-significant.

Weekly body weight of rats in group B and C

Body weight of rats in both groups increased progressively over the study period (table 3). In group B, mean body weight increased from 216.8 ± 1.82 g on day 0 to 231.8 ± 1.15 g on day 7 and 246.2 ± 0.58 g on day 14. Similarly, in group C, body weight increased from 217.0 ± 1.78 g to 231.0 ± 1.44 g and 246.0 ± 0.54 g on days 0, 7, and 14, respectively. The increase in body weight over time was significant within groups, while differences between the two groups were nonsignificant.

Histopathological Observations: Gross examination of major organs including heart, liver, spleen, lungs, kidneys, and treated skin revealed normal morphology. Histopathological evaluation showed largely normal tissue architecture in all organs (fig 1). Cardiac tissue exhibited well-organized muscle fibers with no evidence of necrosis or inflammation. Renal sections showed intact glomeruli and normal tubular structure. Liver sections maintained normal hepatocellular architecture, although mild degenerative changes were observed in some animals treated with the 10% formulation. Lung tissue displayed normal alveolar and bronchial structures with minimal localized alterations. Splenic tissue showed normal distribution of red and white pulp. Skin at the site of application exhibited intact epidermal and dermal layers with mild inflammatory changes in a few sections. Overall, no significant histopathological alterations were observed in either treatment group, indicating dermal safety of the herbal gel formulation.

Image no. 1-6: Histopathological analysis of heart, kidney, liver, lung, skin and spleen Acute dermal toxicity study of herbal gel formulation revealed that the gel was safe to use topically with no unwanted effect or abnormalities in exposed rats was observed in terms of histopathology. [23, 21, 22, 13, 4, 19, 5] . The progressive increase in wound contraction indicated effective tissue repair and healing. The improved response in herbal gel-treated groups may be attributed to the phytoconstituents of Tridax procumbens and Aloe vera, which are known to enhance collagen synthesis, fibroblast proliferation, angiogenesis, and epithelialization. Their antiinflammatory and antioxidant properties further support faster wound closure. Overall, the findings confirm a timedependent increase in wound contraction, with better efficacy observed in the higher concentration herbal gel, indicating its potential as an effective topical wound healing agent.

Evaluation of clinical wound healing efficacy of herbal formulation in goats

Haematological and Biochemical analysis

Mean haematological parameters including haemoglobin, packed cell volume, red blood cell count, and total leukocyte count were evaluated in groups A, B and C on day 0 and day 14 of the study (table 5) and Biochemical parameters, including aspartate aminotransferase, alanine transaminase, total protein, blood urea nitrogen, and serum creatinine, were evaluated in groups A and B on day 0 and day 14, and the results are presented in table 6. Haematological findings over 14 days showed only slight changes in haemoglobin, packed cell volume, erythrocyte and leukocyte counts, and differential counts among the groups. All values remained within normal physiological limits, and the differences were not statistically significant. Biochemical parameters including AST, ALT, total protein, albumin, blood urea nitrogen, and serum creatinine also displayed only minor changes, all values remained within normal physiological limits, and the differences were not statistically significant. This indicates that the treatments did not harm liver or kidney functions. Aloe vera has been reported to support haemoglobin recovery and improve red cell parameters in experimental models, indicating its haematinic potential (Akhter et al., 2023;Archibong et al., 2018). In addition to their effects on erythropoiesis, both plants exhibit immunomodulatory properties. Tridax procumbens has been shown to increase total leukocyte count and enhance both humoral and cellmediated immunity (Tiwari et al., 2004), while Aloe vera has demonstrated anti-inflammatory activity and regulation of leukocyte responses (Ahmad et al., 2020;Kayode, 2017). Overall, these findings suggest that the maintenance of normal haematological parameters observed in the present study may be attributed to the protective, restorative, and immunomodulatory effects of the herbal formulation. In addition to liver protection, both plants exhibit proteinpreserving and tissue-repair properties. Extracts of Tridax procumbens have been reported to enhance collagen formation and improve biochemical parameters without adverse effects on protein status, while Aloe vera has shown protective effects against nephrotoxicity and restoration of total protein levels (Chatterjee et al., 2012). Furthermore, nephroprotective effects of both plants are supported by stabilization or reduction in serum urea and blood urea nitrogen levels. Aloe vera gel, particularly in diclofenac-induced toxicity models, has demonstrated notable renal protective potential, highlighting its therapeutic value (Iftikhar et al., 2015) [2, 3, 24, 1, 17, 7, 14 ] .

Table 1: Grouping and treatments of clinically wounded goats

Sr. No.

The wounds were cleaned with normal saline and a thin film of topical povidone iodine was applied twice daily till complete recovery 2 Group B (n=10)

The wounds were cleaned with normal saline and a thin film of topical herbal gel formulation (10% w/v) was applied twice daily till complete recovery

The wounds were cleaned with normal saline and a thin film of topical herbal gel formulation (5% w/v) was applied twice daily till complete recovery

Conclusion:

The present study demonstrated that the herbal gel formulation containing Tridax procumbens and Aloe vera was dermally safe and effective for wound management in goats. Acute dermal toxicity evaluation confirmed the absence of irritation, toxicity, or mortality in experimental animals, indicating its safety for topical application. Clinical evaluation revealed progressive wound healing in all groups, with reduction in swelling, exudation, pain, irritation, and improvement in wound colour over time. A significant increase in wound contraction was observed throughout the study period. The 10% herbal gel formulation showed better therapeutic response compared to the 5% formulation and exhibited healing effects comparable to the standard treatment. Haematological and biochemical parameters remained within normal physiological limits, indicating no systemic toxicity. Overall, the herbal gel formulation can be considered a safe, effective, and economical alternative for wound management in goats.

References

1. Ahmad Z, Hafeez A, Ullah Q, Naz S, Khan RU. Protective effect of Aloe vera on growth performance, leucocyte count and intestinal injury in broiler chicken infected with coccidiosis. Journal of Applied Animal Research. 2020;48(1):252-256.
2. Akhter S, Begum D, Jahan I, Sharna AH, Khan S, Azmin I. Effects of Aloe vera gel on red blood cell parameters in phenylhydrazine-induced anaemic Wistar albino rats. Ibrahim Cardiac Medical Journal. 2023:40-46.
3. Archibong AN, Udefa AL, Akwari AA, Leilei SA. Effect of Aloe vera gel on some haematological parameters and serum electrolytes in high salt loaded Wistar rats. MicroMed. 2018;6(2):69-77.
4. Atiba A, Ueno H, Uzuka Y. The effect of Aloe vera oral administration on cutaneous wound healing in type 2 diabetic rats. Journal of Veterinary Medical Science. 2011;73(5):583-588.
5. Brahma J, Saharia J, Sarma M, Boro P. Successful treatment of contagious ecthyma (ORF) in Assam hill goats by using turmeric powder and Aloe vera gel preparation. Journal of Entomology and Zoology Studies. 2020;8(4):1454-1456.
6. Can A, Akev N, Ozsoy N, Bolkent S, Arda BP, Yanardag R, et al. Effect of Aloe vera leaf gel and pulp extracts on the liver in type-II diabetic rat models. Biological and Pharmaceutical Bulletin. 2004;27(5):694-698.
7. Chatterjee P, Mukherjee A, Nandy S. Protective effects of the aqueous leaf extract of Aloe barbadensis on gentamicin and cisplatin-induced nephrotoxic rats. Asian Pacific Journal of Tropical Biomedicine. 2012;2(3):S1754-S1763.
8. Department of Animal Husbandry and Dairying. 20th livestock census. Government of India; 2019.
9. Enwa FO, Jemikalajah JD, Oghenejobo M, Oghenebrume VE. Effect of methanol extract of Tridax procumbens flower on hematological parameters and serum lipid profiles of Wistar albino rats. Journal of Pharmaceutical and Bioresources. 2020;17(1):24-28.
10. Farghali HAMAA, Ali KK, Shamaa AAE. Incidence of the most common surgical affections among sheep and goats admitted to a referral hospital in the State of Kuwait. Advances in Animal and Veterinary Sciences. 2020;8(2):41-57.
11. Ghosh P, Biswas S, Biswas M, Dutta A, Sil S, Chatterjee S. Morphological, ethnobiological and phytopharmacological attributes of Tridax procumbens Linn. (Asteraceae): A review. International Journal of Scientific Research in Biological Sciences. 2019;6(2).
12. Guo SA, Dipietro LA. Factors affecting wound healing. Journal of Dental Research. 2010;89(3):219-229.
13. Haritha Yadav KC, Ravi Kumar J, Basha SI, Deshmukh GR, Gujjula R, Santhamma B. Wound healing activity of topical application of Aloe vera gel in experimental animal models. International Journal of Pharma and Bio Sciences. 2012;3(2):P63-P72.
14. Iftikhar A, Hasan IJM, Sarfraz L, Jafri, Ashraf MA. Nephroprotective effect of the leaves of Aloe barbadensis (Aloe vera) against toxicity induced by diclofenac sodium in albino rabbits. The West Indian Medical Journal. 2015;64(5):462.
15. Iji OT, Oyagbemi AA, Azeez OI. Assessment of chronic administration of Aloe vera gel on haematology, plasma biochemistry, lipid profiles and erythrocyte osmotic resistance in Wistar rats. Nigerian Journal of Physiological Sciences. 2010;25(2):107-113.
16. Ikewuchi JC. Alteration of plasma biochemical, haematological and ocular oxidative indices of alloxaninduced diabetic rats by aqueous extract of Tridax procumbens Linn. (Asteraceae). EXCLI Journal. 2012;11:291.
17. Kayode OA. Effects of Aloe vera gel application on epidermal wound healing in the domestic rabbit. International Journal of Research in Medical Sciences. 2017;5(1):101-105.
18. Manvitha K, Bidya B. Aloe vera: A wonder plant, its history, cultivation and medicinal uses. Journal of Pharmacognosy and Phytochemistry. 2014;2(5):85-88.
19. Roy KK, Rahman AME, Kar M, Hossen MI, Islam MS, Rima UK. Evaluation of topical application of Aloe vera gel and honey for acceleration of cutaneous wound healing in goat model. Journal of Bangladesh Agricultural University. 2022;20(4):413-424.
20. Shivhare RS, Awachat P, Mahapatra DK, Ingole AR, Borkar SS. Development of wound healing ointment formulation containing active extracts of Tridax procumbens, Calendula officinalis, Murraya koenigii, and Aloe barbadensis. International Journal of Pharmaceutical and Phytopharmacological Research. 2019;9(6):99-104.
21. Shrivastav A, Mishra AK, Abid M, Ahmad A, Fabuzinadah M, Khan NA. Extracts of Tridax procumbens Linn. leaves cause wound healing in diabetic and non-diabetic laboratory animals. Wound Medicine. 2020;29:100185.
22. Surjushe A, Vasani R, Saple DG. Aloe vera: A short review. Indian Journal of Dermatology. 2008;53(4):163166.
23. Talekar YP, Das B, Paul T, Talekar DY, Apte KG, Parab PB. Evaluation of wound healing potential of aqueous and ethanolic extracts of Tridax procumbens Linn. in Wistar rats. Asian Journal of Pharmaceutical and Clinical Research. 2012;5(4):141-145.
24. Tiwari U, Rastogi B, Singh P, Saraf DK, Vyas SP. Immunomodulatory effects of aqueous extract of Tridax procumbens in experimental animals. Journal of Ethnopharmacology. 2004;92(1):113-119.
25. Yahaya I, Ndatsu Y, Yunusa A, Musa AI, Uthman A, Amina M, et al. Proximate, electrolyte and biochemical profile of rabbits fed with Tridax procumbens residue meal in replacement of vitamins and minerals in Lapai, Niger State, Nigeria. Journal of Applied Sciences and Environmental Management. 2023;27(2):349-354.