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 Table of Contents  
ORIGINAL ARTICLE
Year : 2021  |  Volume : 10  |  Issue : 2  |  Page : 231-239

Phytochemical screening, antioxidant, antibacterial, and antidandruff activities of leaf extract of Artemisia indica


1 Department of Biochemistry, Universal Engineering and Science College, Pokhara University, Lalitpur, Kathmandu, Nepal; Department of Medical Microbiology, B.P. Koirala Institute of Health Sciences, Dharan, Nepal
2 Department of Biochemistry, Universal Engineering and Science College, Pokhara University, Lalitpur, Kathmandu, Nepal
3 Department of Pharmacy, Purbanchal University College of Medical and Allied Sciences, Gothgaun, Morang, Nepal

Date of Submission15-Sep-2020
Date of Acceptance14-Aug-2021
Date of Web Publication17-Dec-2021

Correspondence Address:
Mr. Pragyan Dahal
Universal Engineering and Science College, Pokhara University, Lalitpur, Kathmandu.
Nepal
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/jrptps.JRPTPS_110_20

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  Abstract 

Background: Artemisia indica is a traditionally used medicinal plant in the treatment of various conditions such as loss of appetite, abdominal discomfort, antimalarial infection, dermal wound infection, etc. Objectives: This study aims to determine the presence of phytochemical content, antioxidant, antibacterial, and antidandruff activity of leaf extract of A. indica. Materials and Methods: Dried ground leaves were subjected to a cold extraction method using an absolute concentration of methanol, ethanol, and water. Total phenolic, flavonoid, and proanthocyanidin content was estimated by using a linear regression equation from the calibration curve and expressed in terms of gallic acid equivalent (GAE) and rutin equivalent (RE). Antioxidant properties were determined using DPPH (1,1-diphenyl-2-picryl-hydrazyl), nitric oxide, and hydrogen peroxide assay, and their IC50 values were calculated. The antibacterial activity was tested using the agar well diffusion method against the common five pathogenic strains, and the zone of inhibition is compared with gentamicin (1 mg/mL) as a positive control. The minimum inhibitory concentration (MIC) value was obtained by the microbroth dilution method. The antidandruff assay was performed on Malassezia furfur by the disk diffusion method into Sabouraud dextrose agar overlaid with 1 mL of olive oil, and the MIC value was determined by the microtiter plate method. Results: The result showed that the Artemisia methanolic extract represents ample content of phenolics (248±3.29 mg/g of GAE), flavonoids (222.33±4.41 mg/g of RE), and proanthocyanidin (222.83±1.62 mg/g of RE equivalent). The antioxidant assay revealed that methanolic extract has the highest radical scavenging activity followed by aqueous extract and then ethanolic extract. The antibacterial activity of leaf extract shows MIC value ranging from 6 to 25 μg/mL against various human pathogenic bacteria. The antidandruff assay showed that MIC value of methanolic extract is lesser than that of ethanolic extract (350<400) mg/mL. Conclusion: The results concluded that leaf extract of A. indica contains phenolics, flavonoids, and proanthocyanidin and exhibits adequate antibacterial, antidandruff, and antioxidant activity.

Keywords: Antibacterial, antidandruff, antioxidant, Artemisia indica, phytochemical screening


How to cite this article:
Dahal P, Bista G, Dahal P. Phytochemical screening, antioxidant, antibacterial, and antidandruff activities of leaf extract of Artemisia indica. J Rep Pharma Sci 2021;10:231-9

How to cite this URL:
Dahal P, Bista G, Dahal P. Phytochemical screening, antioxidant, antibacterial, and antidandruff activities of leaf extract of Artemisia indica. J Rep Pharma Sci [serial online] 2021 [cited 2022 Jun 29];10:231-9. Available from: https://www.jrpsjournal.com/text.asp?2021/10/2/231/332777




  Introduction Top


Traditionally used herbal plant genus Artemisia exceeds more than 400 species found in nature and it is named in honor of “Artemis” known as the Greek goddess. The common name includes “Wormwood,” which is named due to wide application in the past for the treatment of worms (intestinal helminth). Geographically, Artemisia family is worldwide distributed, mainly abundant across the temperate zones of the Northern hemisphere, whereas few species are also found in the Southern hemisphere. Artemisia species are pungent in odor and bitter in taste due to the presence of essential oil named terpenoids. Phytochemically, about 839 compounds have been reported from 14 species of wormwood.[1]

Artemisia indica is considered a valuable medicinal plant due to its antihelminthic, antipromastigote, antioxidants, cytotoxic, antimicrobial, antimalarial, and antidiabetic activity.[1],[2],[3],[4],[5] The leaf infusion is also considered to enhance the appetite. The juice of the plant is used to treat diarrhea, dysentery, and abdominal discomfort associated with helminth infection.[1] Studies have shown that A. indica contains phytochemicals such as saponins, tannins, flavonoids, alkaloids, sterol, phenol, proanthocyanidin, and glycosides with various bioactivities.[6],[7] Javid et al.[8] in their study have shown the significant antibacterial activity of A. indica against Escherichia coli, Pseudomonas aeruginosa, Salmonella typhi, and Staphylococcus aureus and antifungal activity against Aspergillus flavus, Aspergillus niger, Aspergillus fumigatus, and Fusarium species. However, evidence for the antidandruff activity of this plant species in literature till date is scarce. This study aims for phytochemical screening and investigates in vitro antioxidant, antibacterial, and antidandruff activity of this plant.


  Materials and Method Top


Specimen collection and preparation of extracts

The A. indica plant was collected from UN Park, Lalitpur district of Nepal. The identification of plant sample was confirmed by the National Herbarium and Botanical Laboratories, Department of Plant Resources located at Godavari, Lalitpur (Reg no.: 051-072/073, Ref no.: 101). Fresh leaf material was washed thoroughly under running tap water, shade dried, and used for extraction. The dried leaf was powdered to a fine powder and stored in airtight bottles. Twenty grams of powdered leaves were weighed and soaked in 200 mL of different solvents for 72 h with intermittent stirring for 30 min every 16–24 h using a magnetic stirrer. The solvents used to obtain extracts were methanol, ethanol, and water. The extracts were then filtered in a Whatman filter paper. The filtrate was taken and dried in rotatory evaporator at 70°C.[9]

Qualitative phytochemical screening

In order to identify the phytochemical derivatives present in a leaf extract of A. indica, a standard procedure of phytochemical screening was performed. Presence of alkaloids was tested using Dragendorff’s and Meyer’s tests, amino acids by ninhydrin test, carbohydrates by Barfoed’s and Fehling’s tests, flavonoids by FeCl3 and Shinoda’s tests, glycosides by Libermann’s test, saponin by froth test, tannins by FeCl3, and terpenoids by Salkowski’s test.[6],[9]

Quantitative phytochemical screening

Determination of total phenolic contents

Total phenol contents in the extracts were estimated using the Folin–Ciocalteu method. Aliquots of 50 μL of 12.5, 25, 50, 100, 200, and 400 μg/mL gallic acid solutions were mixed with 0.1 mL Folin–Ciocalteu reagent (diluted 10-fold) and 0.1 mL (75 g/L) sodium carbonate. Incubation of mixture was done for 30 min and observed in a spectrophotometer at an absorbance of 765 nm. The calibration curve was made by plotting absorbance against concentration. The total phenolic content was expressed as milligrams of gallic acid equivalent (GAE) per gram of extract.[10]

Determination of total flavonoid contents

The total flavonoid content was determined by the Kumaran and Karunakaran method.[11] The rutin calibration curve was prepared by mixing 2 mL of various concentrations of ethanolic solutions of rutin with 2 mL (20 mg/mL) of aluminum trichloride and 6 mL (50 mg/mL) of sodium acetate. The absorbance at 440 nm was read after 2.5 h. The same procedure was used for 2 mL of plant extract (10 mg/mL) instead of rutin solution. All determinations were carried out in duplicates. The content of flavonols was calculated using a standard curve obtained from various concentrations of rutin.

Determination of total proanthocyanidins

In this method, 1 mL of standard solution of concentration 0.5 mL (1 mg/mL) was mixed with 3 mL of 4% vanillin methanol solution and 1.5 mL of HCl: the mixture was allowed to stand for 15 min at room temperature and the absorbance was measured at 500 nm. A standard curve was constructed using rutin as standard. All the measurements were done in triplicates, and the total proanthocyanidin content was expressed as rutin equivalent (RE).[12]

Antioxidant assay

DPPH (1,1-diphenyl-2-picryl-hydrazyl)

The DPPH assay is performed for the assessment of free radical scavenging activity of an antioxidant molecule which is considered as the standard and easy colorimetric process for the scientific evaluation of antioxidant activity of the pure compounds. Five different concentrations of 100, 200, 300, 400, and 500 μg/mL are taken in a test tube. From each concentration, 1 mL of plant extract is mixed with 1 mL aliquot of 0.5 mM DPPH and incubated in the dark at 37°C for 30 min, and the absorbance was noted at 517 nm.[12] Ascorbic acid was taken as positive control.

The percentage of inhibition was calculated using the below formula:



where Ac is the absorbance of DPPH solution and As is the absorbance of the sample after 30 min of incubation in the dark.

IC50 values were determined using GraphPad Prism software after obtaining percentage of inhibition.

Nitric oxide scavenging activity

The nitric oxide scavenging assay was performed according to the procedure done by Awah and Verla.[13] An aliquot of 5 mM of sodium nitroprusside was prepared in phosphate-buffered saline adjusting pH 7.4. At 1 mL of different concentrations (25, 50, 100, 200, 400 μg/mL) of methanolic, ethanolic, and aqueous extract of Artemisia, 0.3 mL of previously prepared 5 mM sodium nitroprusside solution was added. The test tubes containing all the above mixtures were incubated at 25°C for 5 h, and then 500 μL of Griess reagent was used. The absorbance was noted at 546 nm. Quercetin was used as the standard solution to which effect of each extract (methanolic, ethanolic, and aqueous extracts) was compared. NO scavenging activity in terms of percentage was calculated as:



where AC is the absorbance of the control (without extract or standard) and AS is the absorbance of extract or standard.

Hydrogen peroxide scavenging assay

The hydrogen peroxide scavenging assay was carried out following the procedure of Ruch et al.[14] To perform this assay, 43 mM of H2O2was prepared in phosphate-buffered saline (0.1 M, pH 7.4). Artemisia extract (100, 200, 300, 400, and 500 μg/mL) concentration in 3.4 mL of phosphate-buffered saline was added to 600 μL of H2O2 solution (43 mM). The absorbance value of the reaction mixture was noted at 230 nm. Sodium phosphate buffer without H2O2 served as blank, and quercetin was used as a standard for this assay.

The percentage of H2O2 scavenging and standard compounds was calculated using the following equation:



where AC is the absorbance of the control (without extract or standard) and AS is the absorbance of the extract or standard.

Collection of test organisms

The pure cultures of the test organisms of bacteria were collected in a sterile nutrient agar slant from National Public Health Laboratory, Kathmandu, Nepal. The test bacterial organisms used were four Gram-negative bacteria E. coli (ATCC 25922), Proteus mirabilis (ATCC 15028), P. aeruginosa (ATCC 27853), and Salmonella typhi (ATCC 14028), and one Gram-positive bacteria S. aureus (ATCC 14028) for antibacterial assay. The bacterial turbidity was adjusted to 0.5 MacFarland, and pure culture of one of the dandruff-causing fungi, i.e., Malassezia furfur, was inoculated from the Dixon agar and collected in a sterile Sabouraud dextrose agar (SDA) plate.

Antibacterial assay

Bacterial growth inhibition was determined by using the agar well diffusion method. Test bacteria were spread on Muller-Hilton agar medium using a sterile cotton swab. About 6 mm diameter wells were made using a sterile cork borer. Dried methanolic, ethanolic, and aqueous extracts were dissolved in distilled water for making the concentration of 10 mg/mL, and then the required volume of each solvent was drawn with a micropipette and diluted in distilled water to prepare different volumes of each extract (30, 40, 50, and 60 μL) and loaded into the well. Pure distilled water was used as solvent control in the antimicrobial susceptibility test for all three different extracts, and broad-spectrum antibiotic gentamicin (1 mg/mL) was used as a positive control. The plates were incubated for 24 h at 37oC. After incubation, the mean zone of inhibition of four different volumes of same concentration (10 mg/mL) of extracts was measured (in mm). The final data were presented in form of mean ± standard deviation.[15]

Determination of minimum inhibitory concentration

The minimum inhibitory concentration (MIC) of the methanolic, ethanolic, and aqueous extracts was determined by the microbroth dilution method using peptone water. For finding MIC value, 2-fold serial dilutions of plant extract were carried out (200, 100, 50, 25, 12.5, and 6.25 μg/mL). Tubes were incubated at 37oC for 24 h and observed for any visible growth by observing broth turbidity. The bacterial suspensions in peptone water were used as positive control, and uninoculated extracts in broth were used as negative control in a test tube. The MIC was interpreted as the lowest concentration of the extract that inhibited bacterial growth when compared with control tubes that contained only the extracts.[16]

Antidandruff assay

Test organism

The fungal strain was tested with 10% H2O2 to investigate the presence of catalase which is an indicator of Malassezia spp. The catalase test was performed by using 10% H2O2 in a clean test tube in which inoculation of Malassezia spp. Results in the formation of oxygen bubbles. Inoculation was done on SDA plates incorporated with 1 mL of olive oil for providing source of lipid, and SDA was added with penicillin and streptomycin combination (PenStrep) in order to inhibit bacterial growth. The plates were incubated for 48 h at 32°C after which the formation of yellowish-creamy colonies which are typically smooth to slightly wrinkled were examined and studied.[17]

Evaluation of antidandruff assay

Methanolic, ethanolic, and aqueous extracts of A. indica were subjected to antifungal screening by the agar disk diffusion method. SDA plates which were overlaid with 1 mL of olive oil were inoculated with the test isolate by spreading the standardized inoculum on the surface of the agar plate with sterile swab. Five-millimeter diameter plates were punched onto the sterile filter paper and soaked with 20 μL of each of the extract solutions at 100 mg/mL concentration. Fluconazole disc (40 mg/mL) is used as positive control and 10% dimethyl sulfoxide (DMSO)-containing disk alone was used as a negative control for antifungal assay. The disks were dried for 7 h and placed onto the agar plates. The incubation was done at 32°C for 48 h due to optimum growth temperature for Malassezia spp. The antimicrobial activity was observed by measurement of zone of inhibition, and the assay was performed in triplicates to calculate mean and standard deviation.[17]

Determination of MIC

The methanolic, ethanolic, and aqueous extracts from the leaves of A. indica were performed using broth microdilution method for determination of MIC values.[18] The 96-well microtiter plate was used and prepared by pipetting 95 μL of SDA broth with 1 mL of olive oil and was left for 15 min before addition of 5 μL of the yeast suspension into each well. The stock solution of A. indica extracts at 100% was added into the first well, and then followed by 100 μL of 75%, 50%, and 25% of plant extracts added to the next three wells consecutively. The plate was covered with a sterile plate shaker at 300 rpm for 20 s and then incubated at 32°C for 48 h. At the end of the incubation period, the plates were evaluated for the presence or absence of growth. MIC is the lowest concentration of the antifungal agent showing no turbidity after 48 h, where the turbidity is interpreted as visible growth of the fungi. The bioassay was performed in triplicates in order to calculate the mean value.[17]

Statistical analysis

Values of quantitative phytochemical screening, antioxidant assay, and antidandruff assays were performed in triplicates and expressed in mean ± standard deviation using regression equation in Microsoft Excel 2013. IC50 values were calculated by plotting inhibition % vs. concentration using GraphPad prism.


  Results Top


Phytochemical screening

The results of qualitative phytochemical screening are shown in [Table 1].
Table 1: Qualitative test of phytochemicals in methanol, ethanol, and aqueous extracts

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Estimation of total phenolic content of methanolic, ethanolic, and aqueous extracts of A. indica

The content of phenolic compounds in A. indica extracts (10 mg/mL) was determined from the regression equation of the calibration curve (y= 0.0003x−0.012, R2= 0.995). A calibration curve was drawn by using different concentrations of gallic acid and is as shown in [Figure 1]. The amount of phenolic compounds was expressed as GAE in mg/g dry weight of plant extract and is shown in [Table 2]. It was found that the total phenolic content was higher in Artemisia methanolic extract (AME) than Artemisia ethanolic extract (AEE) and Artemisia aqueous extract (AQE) with the increase in magnitude of the order of AME (248±3.29)>AEE (189.32±3.29)>AQE (114.17±0.82) mg/g dry extract GAE.
Figure 1: Standard calibration curve for quantification of total phenolic content

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Table 2: Summary table of total amount of phenol, flavonoid, and proanthocyanidin content in methanolic, ethanolic, and aqueous extracts of A. indica

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Total flavonoid content in methanolic, ethanolic, and aqueous extracts of A. indica

The flavonoid content of the A. indica extracts (10 mg/mL) was determined from the regression equation of the calibration curve (y= 0.002x−0.019, R2= 0.991). A calibration curve was drawn by using different concentrations of rutin and is shown in [Figure 2]. The amount of flavonoids was expressed as RE in mg/g dry weight of plant extract and is as shown in [Table 2]. Flavonoids content was also found to be higher in methanolic extract than in ethanolic and aqueous extracts with the corresponding increasing magnitude order of AME (222.33±4.41) > AEE (156.33±6.79) > AQE (153±7.21) mg/g dry extract of RE.
Figure 2: Standard calibration curve for quantification of total flavonoid content

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Total proanthocyanidin content in methanolic and ethanolic extracts of A. indica

The proanthocyanidin content of A. indica extracts (10 mg/mL) was determined from the regression equation of the calibration curve (y= 0.004x−0.041, R2=0.992). A calibration curve was drawn by using different concentrations of rutin and is shown in [Figure 3]. The amount of proanthocyanidins was expressed as RE in mg/g dry weight of plant extract and is shown in [Table 2]. The proanthocyanidin content was also found to be higher in the methanolic extract than in ethanolic and aqueous extracts of A. indica with the increase in magnitude of AME (222.83±1.6) > AEE (157.5±2.36) > AQE (198.66±1.8) mg/g dry extract of RE.
Figure 3: Standard calibration curve for quantification of total proanthocyanidin content

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Antioxidant activity

DPPH radical scavenging, NO scavenging, and hydrogen peroxide scavenging assay

In-vitro antioxidant activity of methanolic, ethanolic, and aqueous extracts of A. indica was measured on the basis of DPPH radical scavenging activity, NO scavenging activity, and hydrogen peroxide scavenging activity using ascorbic acid as standard in DPPH assay and quercetin as standard in NO and H2O2 scavenging assays. Using the results of all the three methods, the Artemisia methanolic extract had shown the highest antioxidant activity among all different solvent extracts, followed by aqueous extract and ethanolic extract, i.e., AME>AQE>AEE as shown in [Figure 4][Figure 5][Figure 6].
Figure 4: IC50 value of A. indica by DPPH assay with respect to percentage of inhibition measured in triplicates

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Figure 5: IC50 value of A. indica by NO scavenging assay with respect to percentage of inhibition measured in triplicates

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Figure 6: Bar diagram representing IC50 value of A. indica by H2O2 scavenging assay

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Antibacterial and antidandruff activity

The antibacterial activity leaf extract of A. indica was tested against five human pathogenic bacteria, i.e., S. typhi, S. aureus, P. mirabilis, P. aeruginosa, and E. coli. Antibacterial activity of leaf extract of A. indica against Gram-positive and Gram-negative bacteria are shown in [Table 3].
Table 3: Results of antibacterial activity of leaf extracts of A. indica

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Leaf extract of A. indica also showed the antidandruff potential against dandruff causing fungi. The antidandruff activity of A. indica against the test organism M. furfur is shown in [Table 4].
Table 4: Results of antidandruff activity of leaf extracts of A. indica

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[Table 3] shows that the significant zone of inhibition was observed in methanolic and ethanolic extracts of A. indica for P. aeruginosa and E. coli on comparison with other pathogens. The methanolic extracts of A. indica also showed high antimicrobial potency against S. aureus and P. mirabilis when compared with ethanolic extracts.

[Figure 7] shows that the lowest inhibitory concentration was found to be in the positive control gentamicin, followed by methanolic and ethanolic extracts of A. indica. MIC values of the aqueous extract were not determined due to lack of antimicrobial activity of aqueous extract in the well diffusion assay. MIC values revealed that the minimum concentration to inhibit growth of pathogens was observed in the methanolic extract when compared with ethanolic extract.
Figure 7: Determination of MIC value of A. indica and gentamicin against test organisms

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[Table 4] shows that the methanolic extract was found to exhibit highest susceptibility (12±2) mm with MIC (300 mg/mL) over ethanolic extract having zone of inhibition (8±3) mm and MIC (400 mg/mL) on known standard strain of M. furfur, whereas the aqueous extract did not show any activity and hence MIC values were not determined. Here 10% DMSO4-containing disk was used as a negative control and fluconazole served as a positive control representing mean zone of inhibition and standard deviation, i.e., (16±2) mm with MIC values of 35 mg/mL.


  Discussion Top


The presence of secondary metabolites in A. indica extracts such as saponins, alkaloids, tannins, sterols, triterpenoids, phenolic, flavonoids, glycosides, and many enriched phytochemicals exhibits biological activity and serves as the preliminary evidence for the synthesis of plant-derived medicines. Qualitative phytochemical screening results in this study determined the presence of alkaloids, flavonoids, saponin, tannin, glycosides, phenol, and anthroquinonones. Similar findings have been observed in a study done in India on qualitative screening in A. indica extracts.[6] Quantitative phytochemical analysis in our study measured the total content of phenol, flavonoid, and proanthocyanidin. Our qualitative screening was further supported by work done by Pandey et al.[19] in Artemisia vulgaris. In our study, the methanolic extract of Artemisia (AME) exhibited the highest yield of phenol followed by AEE and least in the AQE per gram of GAE. Similarly, the results of total flavonoid content and proanthocyanidin content also revealed the highest yield in methanolic extract when compared with ethanol and aqueous solvent extracts. The study conducted by Ruwali et al.[6] in A. indica and Kumar et al.[20] in Artemisia pallens had also shown that majority of phytochemicals such as phenol, alkaloids, terpenoids, flavonoids, etc. were extracted using methanol solvent. This demonstrates methanol as excellent polar solvent for obtaining Artemisia extract. DPPH is commonly used as a substrate for the fast evaluation of antioxidant activity because of its stability in the radical form and simplicity of the assay. The principle behind this assay in the color change of DPPH solution from purple to yellow as the radical is quenched by the antioxidant. The color changes are measured quantitatively by UV–Vis spectrophotometer absorbance at 517 nm. DPPH inhibition percentage in this study finds AME>AEE>AQE with lower IC50 value (68.86±3) μg/mL. These findings correlate with another study conducted by Ruwali et al.[21] in A. indica, in which antioxidant activities in different concentrations show AME>AEE with comparatively lower IC50 value (83.8±1.25) μg/mL in concentration similar to the methanolic extract. Alternatively, antioxidant properties investigated in South Korea by Lee[22] in different Artemisia species such as Artemisia selengensis, Artemisia japonica, and Artemisia capillaries revealed to have high antioxidant potential (>90% inhibition) with lowest value ranging from 19 to 27 μg/mL. The degree of discoloration indicates the scavenging potential of the antioxidant extract, which is due to free radical scavenging activity. More reduction of the DPPH radical is related to the high scavenging activity of the extract and lower concentration of IC50 values. Similarly, nitric oxide is an unstable compound under the aerobic condition. It reacts with oxygen to produce the stable product nitrates and nitrite by means of intermediates such as NO2, N2O4, and N3O4. It is estimated by the application of Griess reagent. In the NO scavenging assay, NO acts as a scavenger where the level of nitrous acid decreases, whereas in the hydrogen peroxide scavenging assay, hydrogen peroxide itself is an unreactive compound that releases hydroxyl radical. According to our study, the nitric acid scavenging assay and hydrogen peroxide scavenging assay were used to evaluate antioxidant potential and found that the highest scavenging assay was shown by methanolic extract when compared with ethanolic and aqueous extracts. However, due to very limited data source for antioxidant activity on A. indica by these methods, we have compared our similar studies with another research work on antioxidant studies conducted by Ruwali et al. on A. indica,[21] in which methanolic extract was found to be superior by all these employed methodologies. Subsequently, a study conducted in India by Bora and Sharma[18] in another species of Artemisia (Artemisia absinthium) elucidated the significant hydrogen peroxide and nitrite scavenging activity in animal models.

The antibacterial activity of A. indica is observed in five human pathogens, which include S. aureus, P. aeruginosa, S. typhi, E. coli, and P. mirabilis. A broad-spectrum antibiotic gentamicin (1 mg/mL) is used as standard drugs (control). Our study finds that 10 mg/mL of the ethanolic and methanolic extracts of A. indica for P. aeruginosa and E. coli with MIC of 25 μg/mL determined potent antimicrobial action. MIC is defined as the least concentration which inhibits bacterial growth. It varies based on morphology, physiology, and nature of bacteria. Surprisingly, the MIC value was low against Gram-positive bacteria (S. aureus) tested on methanolic extract. The aqueous extract has no action against all five pathogens in all four tested concentrations as aqueous extract had low phenolic yield (14.17±0.82) per gram of GAE. This finding is supported by Ouerghemmi et al.,[23] in which the authors have shown role of phenolic compounds in natural products for antimicrobial activities and yield of phenol from the extract can be correlated with antimicrobial activity. The MIC ranged from 6 to 25 μg/mL that designates the lower dose required for inhibiting bacterial growth. MIC plays a vital role in dose management to prevent side effects. Koul et al.[1] in their review had mentioned that lower dose intakes of all Artemisia species are considered relatively safe. However, few reports have been found on side effects specific to A. indica extracts, such as occurrence of hypoglycemia when consumed in high concentrations.[24] Another study conducted in China on Artemisia species depicts induction of allergic rhinitis and Artemisia hay fever among patients allergic to mugwort plants.[25]

Moreover, this forms the first antidandruff study of A. indica since no other report of the same is available in the public domain. However, Javid et al.[8] have shown antifungal activities of A. indica against filamentous fungi. In our study, methanolic extract was found to be superior over ethanolic extract, i.e., AME (12±2)>AEE (8±3) in mm on strain of M. furfur with MIC values for methanolic extract less than those of ethanolic extract (350 <400) mg/mL, whereas aqueous extract had no susceptibility against dandruff-causing fungus probably due to low yield of phytochemicals such as phenol. Fluconazole was used as a positive control and DMSO as a negative control. This finding shows beneficial effects of A. indica extract, which can be used as a herbal remedy for prevention and control of dandruff. Furthermore, different topical creams and soaps enriched with these phytochemicals can be introduced as one of the major ingredients.


  Conclusion Top


Phytochemically A. indica is a rich source of glycosides, phenols, sterols, flavonoids, alkaloids, and tanins. Malassezia zone of inhibition and lower MIC values shown by the leaf extract of A. indica in bacterial pathogen demonstrate effective antibacterial property. The free radical scavenging potential also enriches its medicinal values. The presence of antimicrobial and antioxidant properties in A. indica contributes in the synthesis of plant-derived drugs following their uses on the basis traditional medicines. The antidandruff potential against M. furfur also signifies that the leaf extract is also effective against dandruff caused by Malassezia species.

Acknowledgements

We would probably like to thank all faculty and technical staffs of Universal Science College, Lalitpur, Nepal. We also express our gratitude to hospital officials of National Public Health Laboratory, Teku for providing us microorganism.

Financial support and sponsorship

Nil.

Conflicts of interest

There is no conflict of interest.

Data availability

All data generated or analyzed during the study are included in this article.



 
  References Top

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    Figures

  [Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6], [Figure 7]
 
 
    Tables

  [Table 1], [Table 2], [Table 3], [Table 4]


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