Potential of a methanolic extract of Lawsonia inermis (L.) leaf as an alternative sanitiser in the time of COVID-19 Pandemic and beyond

Abstract

To harness the antimicrobial properties of a crude methanolic extract of Henna (Lawsonia inermis) leaf as a potential alternative sanitiser, there is the need to test its performance in different solutions. In this work, the effects of distilled water (dH20), Acetate-HCL (AH) Buffer (pH 4.6), Phosphate Buffer Saline (PBS) (pH 7.2) and Tris-HCL (TBH) Buffer (pH 8.6) on the antibacterial and antiviral activity of the extract were assessed. Through standard phytochemical screening and HPLC-MS (LCMS STANDARD 7.M), it was found that the extract consisted of about 30 different compounds including flavonoids. The extent of the antimicrobial activity of the extract in solutions was in the increasing order of AH > dH2O >>>> TBH > PBS. Under the same conditions, reduced antibacterial activity and complete cessation of the antiviral activity of the extract in TBH and PBS was observed. However, in AH and dH20, within 1–5 min, 1 mg ml⁻¹, 0.125 mg ml⁻¹ and 0.0625 mg ml⁻¹ of the extract caused complete inactivation of E.coli (reductions of 8.2 log CFU ml⁻¹), B. subtilis (reductions of 8.2 log CFU ml⁻¹) and MS2 (reductions of 9.7 log PFU ml⁻¹) respectively. The fluorescence microscopy images of the live/dead staining of the inactivated bacterial samples validated the extent of the inactivation. The broad spectrum and high antimicrobial activity of the extract, coupled with the plant not a staple food, has long history of safe use by humans as a medicine and cosmetic, cheaply available in abundance in many regions of the world, thus making the extract a potential candidate as an alternative sanitiser in the time of COVID-19 Pandemic and beyond.

1. Introduction

Sanitisers are antimicrobial agents with a broad spectrum against microorganisms including pathogenic bacteria and viruses. Sanitisation of hands and surfaces are required routine practices in the laboratories, health care settings, food, water and drug processing, production and packaging, toilets, and others. to achieve the public heath standard of zero to minimal risk of transmitting or contracting contagious pathogens and diseases.

In addition to the social distancing measures, good hand hygiene practices have been strongly recommended by the WHO and other health organisations, scientists and health professionals to be effective in curtailing the COVID-19 Pandemic (World Health Organization, 2020a, World Health Organization, 2020b, World Health Organization, 2020c). Frequent hand washing with soap under running water is the most preferred hand hygiene practice (World Health Organization (WHO), 2020b). In the absence of soap and water, hand sanitisers can be used to rub over all the surface of hands and fingers until the hands are dry (World Health Organization, 2015, World Health Organization, 2020b). Although there are several sanitisers with different compositions on the market, alcohol-based sanitisers with at least 60 % alcohol are the ones recommended as the most effective (World Health Organization (WHO), 2020b; 2015). However, in the times of a global health crisis like COVID-19 Pandemic which has led to intense and increased use of hand sanitisers universally, there are huge demands which sanitiser supply chains cannot presently satisfy. Sanitisers became very scarce, highly costly or not available at all in most rural areas of developing countries. There are also reports of harm due to the use of adulterated conventional hand sanitisers during the COVID-19 Pandemic in developing countries. Thus there is the need for alternative sanitisers that are cheap, easy/simple to produce, eco-friendly, food grade and available to the remotest part of the world which could be used in the time of pandemics and beyond to sanitise hands, surfaces and food including fresh produce.

Food grade antimicrobial plant extracts could be used to produce alternative sanitisers. However, such plants must not be a) staple foods to avoid food security risks, b) has broad spectrum antimicrobial activity, c) long history of safe use by humans as medicine, d) cosmetics e) for making drinks and beverages and available in abundance in many regions of the world. One of such plants that satisfied all these conditions is Lawsonia inermis (Henna) (Chaudhary et al., 2010, Hsouna et al., 2011, Ekwealor and Oyeka, 2015, Sharma and Goel, 2018).

Lawsonia inermis is a shrub with greyish-brown bark with Leaves which are opposite, sub-sessile, elliptic or broadly lanceolate, entire, acute or obtuse, 2–3 cm long and 1–2 cm wide (Hsouna et al., 2011). It is frequently cultivated in India, Middle east and Africa (Hsouna et al., 2011, Ekwealor and Oyeka, 2015, Sharma and Goel, 2018). Aside from its cosmetic use for staining hands, nails and hair, the leaves of henna are also used as a prophylactic agent against skin diseases (Hsouna et al., 2011, Ekwealor and Oyeka, 2015). Phytochemical compositions of the Lawsonia inermis predominantly include coumarins, flavonoids, naphthalene and gallic acid derivatives; these phenolic compounds could be glycosylated. Other compounds such as triterpenoids, steroids and aliphatic hydrocarbons have also been isolated from the plant (Siddiqui et al., 2003, Hsouna et al., 2011, Gallo et al., 2008, Sharma and Goel, 2018). Extracts (Leaves, flowers, seeds, stem barks and roots) of Lawsonia inermis have been shown to have antibacterial, antioxidant, anti-inflammatory, antiviral, antifungal, anthelminthic, anticarcinogenic, wound healing, immunomodulatory, analgesic, antipyretic, molluscicidal and hepatoprotective properties (Saadabi, 2007, Nayak et al., 2007, Ekwealor and Oyeka, 2015, Hsouna et al., 2011, Raja et al., 2013, Sharma and Goel, 2018). If the antimicrobial properties of Henna extracts are to be used in the making of sanitisers, there is a need to test the antimicrobial activity performance of the extract in different solutions including water.

In this work, the antibacterial and antiviral activity of crude methanolic extract of Lawsonia inermis leaf in water was investigated. The effects of three buffers solutions (Acetate-HCL Buffer (pH 4.6), Phosphate Buffer Saline (pH 7.2) and Tris-HCL Buffer (pH 8.6)) on the antimicrobial activity of the extracts were also assessed.

2. Materials and methods

2.1. Lawsonia inermis leaf (Henna)

The Henna leaves and its dried fine powder were purchased from the Kure Market, Minna, Niger State, Nigeria. The leaves and its powder were identified and authenticated by a Botanist in the Department of Biology, Ibrahim Badamasi Babangida University Lapai, Niger state, Nigeria and a voucher specimen of the leaf was deposited at their herbarium with a voucher/reference number IBBU210510. The Henna leaves were dried and powdered and used with little prior preparation and processing for the extraction of its phytochemicals.

2.2. Solvents, buffers and other consumables

All the chemicals, solvents and other consumable used were of analytical grade. All inorganic chemicals and salts were purchased from Sigma-Aldrich (Germany). Methanol was purchased from Acros Organics (Belgium). LB broth and agar for culturing bacteria and double layer plaque assay for bacteriophage MS2 were purchased from Fisher Scientific: Janssen Pharmaceuticalaan (Belgium). The distilled water was provided by the Thermo Scientific (UK). Composition details of the buffers used are as follows; Phosphate Buffer Saline (PBS) (pH 7.2; 10 mM Na₂HPO4, 1.8 mM KH₂PO4, 137 mM NaCl, and 2.7 mM KCl), Acetate-HCl (AH) Buffer (pH 4.6; 0.1 M CH₃COOH, 0.1 M CH₃COOK) and Tris Base-HCl (TBH) Buffer (pH 8.6; 0.1 M Tris, 0.1 M HCL).

2.3. Bacteria and virus strains

The bacteria strains – E. coli Nissle 1917 (a model for Gram negative bacteria) and B. subtilis DB 104 (a model for Gram positive bacteria) were from the laboratory of Prof. Ulrich Dobrindt, University of Munster, Germany. The bacteriophage MS2 DSM 13767 (a model for human viruses) (Kohn and Nelson, 2007, Zhong et al., 2016, Majiya et al., 2018) and its E. coli DSM-5695 host cell stocks were purchased from DSMZ, Germany.

2.4. Extraction and screening of phytochemicals of Lawsonia inermis leaf (Henna)

Methanol was used as a solvent for the extraction of phytochemicals of the dried powdered leaves of Henna under cold (room temperature 20 °C) conditions. 10 g each of the dried powdered leaves was dispensed into a 250 ml round bottom flask and 50 ml of methanol was added. The flask was then placed on the shaker set at 500 rpm for 24 h. The phytochemical extract was then decanted into a separate Falcon bottle (50 ml). The extract was then centrifuged at 1871 × g for 5 min. After the centrifugation, the supernatant was decanted into a fresh clean round bottom flask (100 ml). The methanol extract was then concentrated and dried using Rotavapor (BUCHI, Switzerland). The dried sample was weighed, and the yield of the extract was determined. The sample was then dispensed into a smaller labelled bottle and stored in the dark under refrigeration for subsequent use. Phytochemical screening of the extract was carried out using the standard methods (Sharma and Goel, 2018, Debela, 2002). Hager’s test, Dragendorff’s test and Mayer’s test were performed to identify alkaloids in the extract. Legal’s test was carried out to identify glycosides while ferric chloride test was used to determine the presence of tannins and polyphenolic compounds in the extract. Ninhydrin and Biuret tests were used to detect proteins while flavonoids were tested through alkaline tests. Steroids were identified through the Salkowski test and the presence of carbohydrates in the extract was determined through Biuret and Fehling’s test (Sharma and Goel, 2018, Debela, 2002).

2.5. Mass spectroscopy of methanolic extract of Lawsonia inermis leaf

Mass spectroscopy of the methanolic extract of Henna was determined in methanol using MicroTof-ESI (Bruker Daltonics, Germany). Additionally, the fractionization and mass spectroscopy of the extract was carried out using 6110 HPLC-MS-ESI (Agilent Technologies, USA). In all cases, both positive and negative ionisation modes of the mass spectroscopy were acquired.

2.6. Inactivation of model bacteria and virus using the methanolic extract of Lawsonia inermis leaf

Inactivation of the model bacteria (E. coli Nissle 1917 and B. subtilis DB 104) and model virus (bacteriophage MS2 DSM 13767) were investigated with different concentrations of the methanol leaf extract of Lawsonia inermis (LM) in distilled water (dH₂O), Acetate-HCl (AH) Buffer (pH 4.6), PBS (pH 7.2) and Tris Base-HCl (TBH) Buffer (pH 8.6). Actively growing log phase cultures of the model bacteria with Optical Density (OD) 0.5 at 600 nm (about 8 log CFU ml⁻¹) were used for the inactivation experiments. The concentrations of the extract-LM used were 0.03125 mg ml⁻¹ to 1 mg ml⁻¹ and inactivation times were 1–5 min. The stock solution (100 mg ml⁻¹) of the extract-LM was made in DMSO. All experiments were repeated three times. After the inactivation experiments, the solutions were plated for bacterial enumeration to determine the rate and extent of inactivation. Also, live/dead staining (BacLight Bacterial Viability Kit, Invitrogen) was used according to the manufacturer’s instructions to determine the viability of the bacteria after the inactivation experiments. Fluorescence images were captured using 40 × magnification objective with a Nikon Eclipse Ci microscope (Nikon Corporation Tokyo, Japan). The double layer agar plaque assay was used to determine the infectivity and titre of bacteriophage MS2 (starting titre of about 9.7 log PFU ml⁻¹) after the inactivation experiments to determine the rate and extent of inactivation. To prepare 100 ml of soft/top agar, 0.6 g of agar was added to 100 ml of LB broth which was then autoclaved. After sterilisation, 4 ml each of the top agar was dispensed into 15 ml falcon tubes and placed in a water bath set at 45 °C. Tenfold serial dilutions of the MS2 from 10² to 10¹⁴ were carried out in LB broth. Microcentrifuge tubes (1.5 ml) were set up and labelled with dilution factors for the initial incubation of the MS2 and host E. coli cells. To each of these, 0.1 ml of the corresponding MS2 dilution and 0.3 ml log phase E. coli (DSM-5695) culture (0.5 OD at 600 nm) were added and incubated for 20 min at 37 °C. After the incubation, they were added to their corresponding top agars in a water bath and were vortexed before plating them out on their respective corresponding agar plates. After the top agar had solidified, the plates were incubated at 37 °C for 24 h. After incubation, the plates were observed for plaques and counted (Majiya, 2017, Kropinski et al., 2009).

3. Results

3.1. Physico-chemical characteristics of methanolic extract of Lawsonia inermis leaf

The physico-chemical characteristics including the yield, phytochemical constituents, mass spectroscopy and HPLC fractions of methanolic extract of Lawsonia inermis leaf are shown in Tables 1 and S1. The mass spectroscopy and HPLC fractionization of the extract showed it is a complex compound with about 30 different compounds/fractions (Table S1).

Table 1.

Physico-chemical characteristics of methanolic extract of Lawsonia inermis leaf.

Characteristics		Methanol extract
Colour		Dark green
Texture		Gummy oily
Yield (mg) per 10 g of dried Henna leaf		2009.2 mg
Percentage Yield (%)		20.1 %
Alkaloids		+
Carbohydrates		+
Flavonoids		+
Glycosides		+
Resins		+
Saponins		+
Sterols		+
Tannins		+
Quinones		+
Phenol		+
Proteins		−
MS Peaks m/z (Positive)		1st	2nd	3rd
		203.0528	365.1056	101.0007
HPLC-MS	Positive ionisation mode	30 compounds (fractions)
	Negative ionisation mode	15 compounds (fractions)

Key: + = present, − = absent.

3.2. Inactivation of Escherichia coli with the methanolic extract of Lawsonia inermis leaf in solutions

The inactivation of E.coli using 0.0625 mg ml⁻¹ to 1 mg ml⁻¹ methanolic extract of Lawsonia inermis leaf (LM) was determined in distilled water (dH2O), Acetate-HCl (AH) Buffer (pH 4.6), Phosphate Buffer Saline (PBS; pH 7.2), and Tris Base-HCl (TBH) Buffer (pH 8.6). The results showed that E.coli is stable in all the solutions used for the inactivation ( Fig. 1, Fig. 2). Also, the final dilution of the DMSO used to make the stock solution of the LM extract did not have any effect on the bacterium (result not shown). The results showed that 1 mg ml⁻¹ of the LM extract in AH and dH2O and one (1) minute of inactivation time led to complete inactivation (limit of detection) equivalent to reductions of about 8.2 log CFU ml⁻¹ of E. coli (Fig. 1 A). Under the same conditions, the same results were not observed for the inactivation that took place in PBS and TBH was observed; a reduction of the antibacterial activity of the LM in PBS and TBH and only reductions of 2 log CFU ml⁻¹ and 3.3 log CFU ml⁻¹ of E.coli were observed respectively (Fig. 1 A). The LM concentration dependent inactivation in AH and dH2O showed that the minimal concentration of 0.5 mg ml⁻¹ of LM caused reductions of 3.3 log CFU ml⁻¹ and 1.6 log CFU ml⁻¹ of E.coli respectively (Fig. 1 B and C). The fluorescence microscopy images of the live/dead staining of the E.coli samples inactivated with the 1 mg ml⁻¹ LM in dH20 and AH validated the extent of inactivation (Fig. 2).

Fig. 1.

Inactivation of E. coli using different concentrations of the LM extract in different solutions. (A) Inactivation of E.coli in different solution; (B) LM concentration dependent inactivation of E.coli in AH buffer; (C) LM concentration dependent inactivation of dH20. AH, acetate-HCl buffer; dH2O, distilled water; PBS, phosphate buffer saline; TBH, Tris base-HCl buffer; S, solution; LM, methanolic extract of Lawsonia inermis leaf. Data are mean ± SD (n = 3). Error bars show ± SD.

Fig. 2.

Fluorescence microscopy images of live/dead staining of the E. coli samples inactivated with 1 mg ml⁻¹ LM in dH20 and AH buffer. AH, acetate-HCl buffer; dH2O, distilled water; LM, methanolic extract of Lawsonia inermis leaf.

3.3. Inactivation of Bacillus subtilis with the methanolic extract of Lawsonia inermis leaf in solutions

The inactivation of B. subtilis was investigated using 0.0625 mg ml⁻¹ to 1 mg ml⁻¹ methanolic extract of Lawsonia inermis leaf (LM) in distilled water (dH2O), Acetate-HCl (AH) Buffer (pH 4.6), Phosphate Buffer Saline (PBS; pH 7.2), and Tris Base-HCl (TBH) Buffer (pH 8.6). The final dilution of the DMSO used to make the stock solution of the LM extract does not have any effect on the B. subtilis (result not shown). It was observed that B. subtilis is not stable in AH; ordinarily and without the LM, the buffer-AH caused the reductions of about 5 log CFU ml⁻¹ of B. subtilis ( Fig. 3, Fig. 4). Also, about 0.8 log CFU ml⁻¹ reductions of the bacterium was observed in dH2O without LM (Fig. 3). The results showed that 1 mg ml⁻¹ of the LM extract in all of the solutions and one (1) minute of inactivation time caused complete inactivation (limit of detection) equivalent to reductions of about 8.3 log CFU ml⁻¹ of B. subtilis (Fig. 3 A). The concentration dependent inactivation with the LM in AH showed that even the minimal concentration of 0.0625 mg ml⁻¹ of LM used caused complete inactivation of the bacterium (Fig. 3 B). However, the minimal concentration of LM that showed activity against B. subtilis in water was 0.0625 mg ml⁻¹ and caused reductions of 2.4 log CFU ml⁻¹ of the bacterium (Fig. 3 C). The fluorescence microscopy images of the live/dead staining of the B. subtilis samples inactivated with the 1 mg ml⁻¹ LM in dH20 and AH validated the effect of the AH buffer and extent of inactivation (Fig. 4).

Fig. 3.

Inactivation of B. subtilis using different concentrations of the LM extract in different solutions. (A) Inactivation of B. subtilis in different solutions; (B) LM concentration dependent inactivation of B. subtilis in AH buffer; (C) LM concentration dependent inactivation of B. subtilis in dH20. AH, acetate-HCl buffer; dH2O, distilled water; PBS, phosphate buffer saline; TBH, Tris base-HCl buffer; S, solution; LM, methanolic extract of Lawsonia inermis leaf. Data are mean ± SD (n = 3). Error bars show ± SD.

Fig. 4.

Fluorescence microscopy images of live/dead staining of the B. subtilis samples inactivated with 1 mg ml⁻¹ LM in dH20 and AH buffer. AH, acetate-HCl buffer; dH2O, distilled water; LM, methanolic extract of Lawsonia inermis leaf.

3.4. Inactivation of bacteriophage MS2 with the methanolic extract of Lawsonia inermis leaf in solutions

The inactivation of phage MS2 using 0.03125 mg ml⁻¹ to 1 mg ml⁻¹ methanolic extract of Lawsonia inermis leaf (LM) in distilled water (dH2O), Acetate-HCl (AH) Buffer (pH 4.6), Phosphate Buffer Saline (PBS; pH 7.2), and Tris Base-HCl (TBH) Buffer (pH 8.6) was investigated. The final dilution of the DMSO used to make the stock solution of the LM extract did not have any effect on the phage (result not shown). It was observed that phage MS2 is very stable in all the solutions used for the inactivation ( Fig. 5). The results showed that 1 mg ml⁻¹ of the LM extract in AH and dH2O and one (1) minute of inactivation time caused complete inactivation (limit of detection) equivalent to reductions of 9.7 log PFU ml⁻¹ of the virus (Fig. 5 A). However, with the same concentration of the LM and inactivation time, antiviral activity of the extract ceased completely in PBS and TBH (Fig. 5 A). The concentration dependent inactivation of the virus with the LM in AH showed that even the minimal concentration of 0.03125 mg ml⁻¹ of LM used caused complete inactivation of the phage MS2 (reductions of 9.7 log PFU ml⁻¹) (Fig. 5 B). However, the minimal concentration of LM that showed antiviral activity in water was 0.03125 mg ml⁻¹ and it caused reductions of 3 log CFU ml⁻¹ of the MS2 (Fig. 5 C).

Fig. 5.

Inactivation of phage MS2 using different concentrations of the LM extract in different solutions. (A) Inactivation of phage MS2 in different solutions; (B) LM concentration dependent inactivation of phage MS2 in AH buffer; (C) LM concentration dependent inactivation of phage MS2 in dH20. AH, acetate-HCl buffer; dH2O, distilled water; PBS, phosphate buffer saline; TBH, Tris base-HCl buffer; S, solution; LM, methanolic extract of Lawsonia inermis leaf. Data are mean ± SD (n = 3). Error bars show ± SD.

4. Discussion

In this work, methanol was used as a solvent for the extraction of phytochemicals from the dried powdered Lawsonia inermis (Henna) leaf. A yield of about 20 % was observed and the extract contained flavonoids as one of its phytochemical constituents (Table 1). These observations agreed with previous reports (Sharma and Goel, 2018). The HPLC-MS fractionalisation of the extract showed that it is made up of about 30 different compounds; the authors interest was on the whole crude extract’s antimicrobial activity performance in different solutions and not individual constituents of the extract (Tables 1 and S1).

The leaves and other parts (flowers, seeds, stem barks and roots) of Lawsonia inermis have been previously investigated for their antimicrobial activities especially antibacterial, antifungal and antiparasitic activities. However, most of the investigations used high concentrations of the extracts and could not quantitate the number of microorganisms inactivated by the extract; these maybe attributable to the methods used to test for the viability of microorganisms to determine the extents of inactivation with the extracts (Saadabi, 2007, Nayak et al., 2007, Ekwealor and Oyeka, 2015, Hsouna et al., 2011, Raja et al., 2013, Sharma and Goel, 2018). Presently, there is very little or no report of the antiviral activity of Lawsonia inermis extracts.

Methanolic extract of Lawsonia inermis leaf has been previously shown to have broad spectrum antimicrobial activity (Sharma and Goel, 2018). However, in this study, the antibacterial and antiviral activity of crude methanolic extract of Lawsonia inermis leaf in water was investigated. The effects of three buffers solutions (Acetate-HCL Buffer (pH 4.6), Phosphate Buffer Saline (pH 7.2) and Tris-HCL Buffer (pH 8.6)) on the antimicrobial activity of the extract was also investigated. Generally, the extent of antimicrobial activity of the extract in solutions was in the increasing order of AH > dH2O > >>> TBH > PBS. Under the same conditions, reduced antibacterial activity of the extract in TBH and PBS was observed and complete cessation of the antiviral activity with the same buffer solutions (TBH and PBS). The pH and salts contents of these buffer solutions might have negatively affected the antimicrobial constituents in the extract rendering it ineffective against the model organisms especially the phage MS2. Also, possibly, the constituents of the buffer solutions might have stabilised the model bacteria and virus, thereby making them resistant or to have recovered from the damage caused by the extract. However, the dH20 and constituents of AH might have sustained or even enhanced the antimicrobial activity of the extract. The authors believed that in dH2O and especially in AH, some components of the extract could be protonated thereby leading to a stronger interaction of the bacterial cells/MS2 and active agents which might have resulted to the enhanced antimicrobial activity observed (Galstyan et al., 2019). These possibilities may limit the use of the crude methanolic extract of Lawsonia inermis leaf mostly to the environmental applications including sanitisations of hands, objects, surfaces, topical applications and fresh produce where water and other buffer solutions like AH would be used as diluents of the extract to inactivate pathogenic and spoilage microorganisms including viruses. The crude extract may not be suitable for the inactivation of microorganisms in biological samples usually have a high salt contents like PBS. To the best of the authors knowledge, this will be the first study to show the effect of buffer solutions on the antimicrobial activity of crude methanolic extract of Lawsonia inermis leaf.

E. coli and B. subtilis were chosen as models for Gram-negative and Gram-positive bacteria, respectively. Under the same conditions, it was observed that more B. subtilis was inactivated compared with the E. coli. This is a general trend for most of the antibacterial agents. Gram-positive bacteria are easily inactivated agents compared with gram-negative bacteria due to differences in the structure of their cell membrane. Gram-negative bacteria have an additional outer membrane apart from the cytoplasmic (inner) membrane, giving them extra protection against antimicrobial agents (Bourré et al., 2010). It is important to note here that B. subtilis was sensitive to AH, possibly due to its low pH.

Bacteriophage MS2-a non-enveloped virus has been used as a viral model organism in several studies aimed at inactivation of human viruses because of its similarity in size and morphology to some human viruses such as noroviruses and picornaviruses (Kohn and Nelson, 2007, Zhong et al., 2016). Also, it safe to work with as it is non-toxic to human and easy to propagate. Generally, enveloped viruses are believed to be inactivated faster and higher by antiviral agents compared to the non-enveloped viruses. In this work, a high and fast inactivation of the MS2 with the crude methanolic extract of Lawsonia inermis leaf was observed. This may be an indication of high antiviral activity of the extract even at low concentrations.

5. Conclusions

In conclusion, buffer solutions affect antimicrobial activity of methanolic extract of Lawsonia inermis leaf; reduced antibacterial and complete cessation of the antiviral activity were observed in PBS and TBH. However, high and fast antibacterial and antiviral activities of the extract were retained and observed in AH and dH20; this means the extract would be suitable for making sanitisers for environmental and topical applications such as hands, objects and surfaces sanitisations in the time of COVID-19 Pandemic and beyond especially in places where conventional alcohol based sanitisers are lacking. This would of course be a reality with further standardisations and testing of the extract on more microorganisms and solutions. Also, cytotoxicity of the methanolic extract of Lawsonia inermis leaf should be performed to ascertain its safety on humans for topical biomedical applications.

CRediT authorship contribution statement

Hussaini Majiya: Conceptualization, Methodology, Formal analysis, Investigation, Writing – original draft, Funding acquisition. Anzhela Galstyan: Resources, Data curation, Writing – review & editing, Supervision, Visualization.

Ethical consideration

Ethics approval was not required for this research.

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Appendix A. Supplementary material

Supplementary material.