Fabrication and Evaluation of Polyherbal Handwash Utilizing Tulsi (Ocimum sanctum), Bhringraj (Eclipta alba) and Harad (Terminalia chebula)

Pallavi, Deepak Prashar*, Shilpa, Priya Kumari, Akhil Tomar

Department of Pharmacy, LR Institute of Pharmacy, Jabli-Kyar, Solan HP-India

*Correspondence: prashardeepak99@yahoo.in;

DOI: https://doi.org/10.71431/IJRPAS.2026.5408

INTRODUCTION

Hand hygiene is widely recognized as one of the most critical measures in preventing healthcare associated infections (HAIs) and community acquired diseases. The World Health Organization (WHO) has consistently emphasized the role of handwashing with soap or antiseptic handwash in curbing the spread of pathogens including bacteria, viruses, and fungi [1]. The COVID-19 pandemic further underscored the indispensable nature of hand hygiene as a frontline defence against infectious disease transmission [2]. Conventional synthetic handwash products contain a range of chemical components including triclosan, parabens, sodium lauryl sulphate (SLS), and artificial fragrances. While effective, these compounds have raised concerns regarding skin sensitization, antimicrobial resistance, hormonal disruption, and environmental toxicity [3]. Triclosan, a widely used antibacterial agent, has been associated with the development of cross-resistance in bacteria and has demonstrated endocrine disrupting activity in animal studies [4]. Similarly, parabens used as preservatives have been detected in human breast tissue, raising safety concerns over their long term use [5]. The global resurgence of interest in phytotherapy has led researchers to explore herbal alternatives to synthetic antimicrobial agents. Herbal handwash formulations represent a compelling solution, offering antimicrobial activity through multicomponent mechanisms that are less prone to resistance development, combined with moisturizing and skin-conditioning benefits [6].

Ocimum sanctum Linn. (Tulsi), a sacred plant in Indian tradition, belongs to the family Lamiaceae. It is rich in volatile oils, flavonoids (luteolin, apigenin), phenolic acids, and terpenoids including eugenol, linalool, and ursolic acid. Tulsi exhibits potent antibacterial, antifungal, antiviral, anti-inflammatory, and adaptogenic properties and has been extensively documented in Ayurvedic texts for its purifying and health-promoting attributes [7]. Eclipta alba Hassk. (Bhringraj), belonging to the family Asteraceae, contains wedelolactone, ecliptine, nicotine, wedelic acid, and coumestan derivatives. It is traditionally employed in hair care formulations and as a hepatoprotective agent. Recent pharmacological studies have confirmed its significant antimicrobial and antioxidant activities, alongside its role in wound healing and dermal health [8-9]. Terminalia chebula Retz. (Harad), a member of the family Combretaceae, is one of the three constituents of the classical Ayurvedic formulation Triphala. It is rich in hydrolysable tannins (chebulinic acid, chebulagic acid, gallic acid), triterpenoids, and flavonoids. It demonstrates broad-spectrum antimicrobial activity, strong antioxidant properties, and wound-healing potential, making it an excellent candidate for topical formulations [10].

The combination of these three herbs leverages their complementary phytochemical profiles to produce a synergistic antimicrobial and skin-protective effect. The present study was therefore undertaken to formulate and evaluate a polyherbal handwash gel incorporating standardized extracts of Tulsi, Bhringraj, and Harad, and to compare the optimized formulation against a marketed standard.

MATERIALS AND METHODS

Plant Material Collection and Authentication

Dried leaves of Ocimum sanctum and Eclipta alba, and dried fruits of Terminalia chebula, were procured from a certified herbal supplier (Zandu Pharmaceutical Works Ltd., Mumbai, India) and from local herbal markets of Panipat, Haryana.

Chemicals and Reagents

Carbopol 940 (Lubrizol Corporation, USA), triethanolamine (TEA; Loba Chemie, India), glycerin (Merck, India), sodium lauryl sulphate (SDS-grade; Himedia, India), EDTA disodium salt, methyl paraben, propyl paraben, citric acid, rose water, and purified water were used. All solvents used for extraction (ethanol 70%, distilled water) were of analytical grade.

Preparation of Herbal Extracts

Aqueous Extract: Coarsely powdered plant material (50 g each) was macerated in 500 mL of distilled water for 72 hours with occasional stirring. The extract was filtered through Whatman No. 1 filter paper, concentrated in a rotary evaporator at 60°C, and dried to yield a semisolid extract. Percentage yield was calculated.

Hydroalcoholic Extract: Soxhlet extraction was performed using 70% ethanol as solvent for 6 hours. The extract was filtered, concentrated, and dried. The dried extracts were stored at 4°C until use. Preliminary phytochemical screening was performed for all extracts using standard methods as described by Harborne and Kokate et al. [11-12].

2.4 Formulation of Polyherbal Handwash

Six formulations (F1–F6) were prepared with varying concentrations of herbal extracts using carbopol 940 as the primary gelling agent. Carbopol 940 was dispersed in 60 mL of purified water and allowed to swell for 24 hours. Glycerin was added with stirring to improve skin conditioning. SLS was dissolved separately in warm water and added to the gel base. Methyl paraben and propyl paraben were dissolved in a small quantity of propylene glycol and added as preservatives. The herbal extracts (dissolved in a minimum volume of 70% ethanol) were incorporated into the gel base with gentle stirring. EDTA disodium salt was added as a chelating agent. The pH was adjusted to 6.0–6.8 by gradual addition of TEA. Rose water was added for fragrance and volume was made up to 100 mL with purified water. The gel was homogenized using a mechanical stirrer at 500 rpm for 15 minutes and deaerated by standing at room temperature.

Table 1: Formulation Composition of Polyherbal Handwash (F1–F6) per 100 mL

q.s. = quantum sufficit (sufficient quantity); pH adj. = added to adjust pH to 6.0–6.8

Evaluation Parameters

Organoleptic Properties: All formulations were evaluated for colour, odour, appearance, texture, and phase separation by visual observation.

pH Determination: The pH of each formulation was measured using a calibrated digital pH meter (Systronics, India) at 25°C. Triplicate measurements were recorded [13].

Viscosity: Viscosity was determined using a Brookfield viscometer (LV-DV II+, spindle No. 64, speed: 50 rpm) at 25 ± 1°C. Results were expressed in centipoise (cP) [14].

Spreadability: Spreadability was determined by the parallel plate method. One gram of gel was placed on a glass plate and a second plate applied with a 100 g weight for 5 minutes. The spread diameter was measured. Spreadability (S) = W × L / T, where W = weight (g), L = spread diameter (cm), T = time (seconds) [15].

Foam Height and Stability: 2 mL of each formulation was placed in a graduated 50 mL cylinder containing 8 mL of distilled water. The cylinder was stoppered and shaken 20 times. Initial foam height was recorded immediately and again after 5 minutes. Foam stability was expressed as the percentage of foam remaining after 5 minutes relative to the initial foam height [16].

Washing Efficacy Test: Ten volunteers with artificially contaminated hands (charcoal suspension) washed with 2 mL of each formulation for 30 seconds. Efficacy was rated on a scale of 1–5 based on the degree of cleansing [17].

Antimicrobial Activity: Antimicrobial activity was assessed by the agar well diffusion method on Mueller-Hinton agar (bacteria) and Sabouraud Dextrose agar (fungi). Test organisms included Staphylococcus aureus ATCC 25923, Escherichia coli ATCC 25922, and Candida albicans ATCC 10231. Wells (6 mm diameter) were made using a sterile cork borer. 100 µL of each formulation was added per well. Ciprofloxacin (5 µg/disc) and fluconazole (25 µg/disc) were used as positive controls. Plates were incubated at 37°C for 24 hours. Zones of inhibition (mm) were measured. Marketed handwash (Dettol® Original) served as the reference standard [18].

Skin Irritation Study (Patch Test): A patch test was conducted on the inner forearm of ten healthy volunteers. 1 g of each formulation was applied under occlusion for 24 hours. Skin reactions were graded (0 = no reaction, 1 = mild erythema, 2 = erythema + oedema, 3 = vesiculation) [19].

Stability Studies: Stability studies were conducted as per ICH Q1A(R2) guidelines at accelerated conditions (40 ± 2°C / 75 ± 5% RH) and intermediate conditions (30 ± 2°C / 65 ± 5% RH) over three months. Physical appearance, pH, viscosity, and antimicrobial activity were evaluated at 0, 1, 2, and 3 months [20].

RESULTS AND DISCUSSION

Phytochemical Screening

Preliminary phytochemical screening of all three extracts revealed the presence of key bioactive phytoconstituents. Ocimum sanctum extracts tested positive for tannins, flavonoids, terpenoids, saponins, and phenolic compounds. Eclipta alba extracts showed the presence of alkaloids, coumestans, flavonoids, and tannins. Terminalia chebula extracts demonstrated the presence of hydrolysable tannins (gallic acid, ellagic acid), triterpenoids, and flavonoids. These phytoconstituents are well-documented as responsible for the antimicrobial, anti-inflammatory, and antioxidant properties of the respective plants [7,8,10].

Organoleptic Evaluation

All six formulations were observed to be smooth, homogeneous gels free from any phase separation. F1 and F2 appeared light green in colour owing to the Bhringraj and Tulsi extracts, while F3–F6 presented as slightly deeper green gels with pleasant herbal aroma. No grittiness, stickiness, or separation was noted upon storage at room temperature. The formulations were found aesthetically acceptable by volunteer panellists.

Physicochemical Parameters

Table 2: Physicochemical Evaluation of Formulations F1–F6 (n=3, Mean ± SD)

Values expressed as Mean ± SD; n = 3 independent determinations

The pH values ranged from 6.2 to 6.8 across all formulations, which is compatible with human skin pH (4.5–6.5) and suitable for a handwash formulation to prevent microbial growth while maintaining skin integrity [13]. Higher carbopol concentrations in F4–F6 resulted in increased viscosity but slightly reduced spreadability. All formulations demonstrated acceptable foam height (>30 mL) and stability (>70%), with F3 and F6 showing the highest foam performance. The washing efficacy scores indicated satisfactory cleansing ability, with F3 rated highest (4.5/5).

Antimicrobial Activity

Table 3: Zone of Inhibition (mm) by Agar Well Diffusion Method (Mean ± SD, n=3)

*Control: Dettol® Original marketed handwash; Values as Mean ± SD of triplicate determinations

Formulation F3, containing the highest combined concentration of Ocimum sanctum (1.5 g) and Eclipta alba (1.0 g) extracts, demonstrated the greatest zones of inhibition: 22 mm against S. aureus, 19 mm against E. coli, and 17 mm against C. albicans. These values approached, though did not fully match, the marketed standard (Dettol® Original), which produced inhibition zones of 24 mm, 22 mm, and 21 mm respectively. The antimicrobial activity of Tulsi is primarily attributed to eugenol, a phenylpropanoid compound that disrupts bacterial cell membranes and inhibits ATP synthesis [7]. Gallic acid and chebulagic acid from Terminalia chebula contribute synergistic bacteriostatic activity by inhibiting DNA gyrase and membrane disruption [10]. Wedelolactone from Eclipta alba exerts antifungal effects via ergosterol biosynthesis inhibition [9].

Skin Irritation Study

No visible skin reactions (erythema, oedema, vesiculation) were observed in any of the ten volunteers during the 24-hour patch test study. All reactions were graded 0 (no reaction), confirming that all six formulations are non-irritant and suitable for topical application. The near-neutral pH (6.2–6.8) and the emollient properties of glycerin likely contributed to the good dermal tolerability profile [19].

3.6 Stability Studies

Table 4: Stability Study Results for Optimized Formulation F3 (ICH Q1A(R2) Accelerated Conditions: 40°C/75% RH)

No phase separation, syneresis, or microbial contamination detected throughout the study period

Stability studies conducted over three months at accelerated conditions (40°C/75% RH) indicated that formulation F3 remained physically and chemically stable throughout the study period. A marginal reduction in pH (6.5 to 6.3) and viscosity (5100 to 4890 cP) was observed, but these changes were within acceptable limits and not clinically significant. The slight colour darkening noted at month 3 may be attributed to oxidation of polyphenolic constituents, which can be minimized by incorporating antioxidants such as butylated hydroxytoluene (BHT) in future formulations. Antimicrobial activity was maintained throughout the study period, with zone of inhibition against S. aureus reducing marginally from 22 to 20 mm, confirming adequate preservative efficacy.

DISCUSSION

The development of polyherbal formulations for topical use represents a scientifically grounded strategy to harness the synergistic phytochemical interactions among multiple medicinal plants. The present study successfully demonstrated that a combination of Ocimum sanctum, Eclipta alba, and Terminalia chebula can be incorporated into a stable, efficacious handwash gel using carbopol 940 as the gelling matrix. The selection of these three herbs was guided by their well-documented antimicrobial activity and complementary mechanisms of action. Ocimum sanctum, the cornerstone of Ayurvedic medicine, provides broad-spectrum bactericidal and antifungal activity primarily through eugenol-mediated membrane disruption, while simultaneously offering anti-inflammatory benefits via inhibition of cyclooxygenase and lipoxygenase pathways [7]. Eclipta alba contributes through wedelolactone's antifungal and hepatoprotective properties and through its coumestan content that enhances the formulation's UV-protective and skin-regenerative capacity [8]. Terminalia chebula, with its exceptionally high tannin content, provides astringent and bacteriostatic properties and has demonstrated synergistic interactions with conventional antibiotics, suggesting its utility in combination antimicrobial strategies [10]. The choice of carbopol 940 as the gelling agent was based on its excellent gelling properties at low concentrations, transparency, compatibility with a wide range of excipients, and pH-dependent viscosity that can be precisely controlled through neutralization with TEA [14]. SLS was included as a foaming surfactant at a concentration (1.5% w/v) low enough to minimize irritation potential while ensuring adequate cleansing efficacy. The inclusion of glycerin as a humectant is clinically important, as repeated handwashing is known to cause transepidermal water loss (TEWL) and skin barrier disruption; glycerin maintains skin hydration and supports the integrity of the stratum corneum [21].

Comparative analysis with the marketed control (Dettol® Original) revealed that while F3 did not fully match the marketed product in antimicrobial potency, the difference was modest and within 10–20% for bacteria. The somewhat lower antifungal activity against C. albicans warrants further investigation, potentially through optimization of extract concentrations or addition of neem (Azadirachta indica) or clove (Syzygium aromaticum) extracts known for superior antifungal properties. Future studies should also evaluate the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of individual and combined extracts to better characterize synergistic interactions [22].

The non-irritant profile of all formulations in the patch test is particularly significant from a safety standpoint. Chemical handwashes containing triclosan and SLS at higher concentrations have been associated with contact dermatitis and disruption of the skin microbiome. The herbal formulations in this study, with their near-neutral pH and low SLS content, preserved skin integrity as evidenced by the absence of erythema or oedema in all volunteers. These findings are consistent with previous reports on herbal handwash formulations employing neem, aloe vera, and lemon grass extracts [6,23]. The accelerated stability data over three months provides preliminary evidence of formulation stability, though longer-term stability studies (12 months) at real-time conditions (25°C/60% RH) as mandated by ICH Q1A(R2) guidelines are necessary before considering this formulation for commercial development. The marginal viscosity reduction observed may be mitigated by increasing the carbopol 940 concentration from 1.0% to 1.2% in future batches of F3 [20]. Additionally, microbiological challenge testing as per USP <51> is recommended to conclusively demonstrate preservative efficacy throughout the proposed shelf life.

CONCLUSION

The present investigation successfully demonstrates the feasibility of formulating a stable, safe, and efficacious polyherbal handwash gel incorporating the synergistic combination of Ocimum sanctum (Tulsi), Eclipta alba (Bhringraj), and Terminalia chebula (Harad). Among the six formulations developed, F3 (containing 1.5 g Tulsi, 1.0 g Bhringraj, and 0.5 g Harad extracts with 1.0% carbopol 940) emerged as the optimized formulation based on its superior antimicrobial activity, acceptable physicochemical parameters, satisfactory foam performance, and excellent skin tolerance profile. The polyherbal handwash formulation offers several advantages over conventional synthetic handwashes, including multi-target antimicrobial activity, reduced risk of resistance development, skin-conditioning properties, biodegradability, and consumer safety. The formulation demonstrated good stability over three months under accelerated conditions, supporting its commercial potential.

Future research should focus on clinical evaluation in healthcare settings, biofilm disruption studies, in vitro skin permeation assessment, stability studies at real-time conditions, and exploration of additional complementary herbs to further enhance antifungal activity. Scale-up studies and cost-benefit analysis should also be undertaken to evaluate the commercial viability of this polyherbal product.

CONFLICT OF INTEREST

The authors declare no conflict of interest.

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