Development and Evaluation of a Nanoemulsion-Based Hydrogel from Moringa Leaves Extract: Standardization, Biological Activity, and Formulation

Betty Ekawati Suryaningsih¹*, Oktavia Indrati², Mustofa Mustofa³,

Viviane Annisa², Asih Triastuti²*

¹Department of Dermatology and Venereology, Faculty of Medicine,

Universitas Islam Indonesia, Yogyakarta, Indonesia.

²Department of Pharmacy, Universitas Islam Indonesia, Yogyakarta, Indonesia.

³Department of Pharmacology, Faculty of Medicine, Universitas Gadjah Mada, Yogyakarta, Indonesia.

*Corresponding Author E-mail: betty.e.s@uii.ac.id, asih.triastuti@uii.ac.id

ABSTRACT:

Moringa oleifera, known for its rich phytochemical profile, has garnered attention for its potential in skin health applications, particularly concerning antihyperpigmentation and anti-acne effects. The objective of this study investigated the antioxidant, antityrosinase, and antibacterial activities of standardized Moringa leaf extract (MLE) formulated into a nanoemulsion-based hydrogel for topical application. The ethanolic extract was prepared and standardized following the Indonesian Herbal Pharmacopoeia. The antioxidant, antityrosinase, and antibacterial properties of the extracts were assessed to confirm their biological efficacy. Ultra-high-performance liquid chromatography-high-resolution mass spectrometry (UHPLC-HRMS) was used to characterize the chemical composition of the nanoemulsion, which was formulated using six different oil preparations. The nanoemulsion was characterized using several parameters, including percentage of transmittance, particle size, polydispersity index, and zeta potential. The stability of the nanoemulsion-based hydrogel was tested while paired and unpaired student t-tests were used for data analysis of hedonic test, with p <0.05 considered statistically significant. The MLE met the Indonesian standardization parameters and demonstrated antioxidant activity, with an IC₅₀value of 0.21 mg/ml, which is more than twice the IC₅₀ value of vitamin C (0.096 mg/ml). The IC₅₀ value of MLE in the inhibitory tyrosinase assay was 9.394 mg/ml, which was higher than the IC₅₀ of kojic acid (0.096 mg/ml). MLE was also active against S. epidermidis and S. aureus but not against E. coli. The particle sizes of the nanoemulgel preparations ranged from 30.3 ± 0.47 nm to 74.4 ± 4.8 nm. Our study demonstrated that MLE, which contains flavonoids and polyphenols, possesses antioxidants, antityrosinase, and antibacterial activities. Furthermore, MLE has been successfully incorporated into a stable nanoemulsion-based hydrogel, potentially offering a novel therapeutic approach for skin care.

KEYWORDS: Antityrosinase, Antibacterial, Antioxidant, Moringa oleifera, Nanoemulsion-based hydrogel.

INTRODUCTION:

Herbal medicines play a vital role in the foundation of contemporary medicine. Despite their in vivo efficacy, they are underutilized in clinical practice due to several challenges, such as solubility issues, bioavailability, and the need for high doses. Designing dosage forms for natural chemicals is essential for their recognition in modern medicine while ensuring their safety and efficacy. Several herbal medication formulations, such as herbal nano-tablets, nano-capsules, nanoemulsions, nanopastes, and nanogels, have been developed in recent years ¹. Moringa oleifera is a medicinal plant with a wide range of nutritional and therapeutic benefits ^2,3. It is widely used as a nutraceutical in several countries and is known as a superfood. All plant parts have been reported to exhibit medicinal properties, although the leaves are the most widely used⁴. Moringa leaves are rich in phenolic acids, flavonoids, glucosinolates, and isothiocyanates^5–7. In addition to its typical potential medicinal properties as an antibacterial ⁸, antifungal ⁹, antioxidant ¹⁰, immunomodulator ¹¹, and painkiller, Moringa oleifera also exhibits anti-inflammatory ¹², anti-cancer, anti-arthritic, antidiabetic, and anti-infertility properties ^13–15.

Moringa leaves are also used topically in dermatology to promote several advantages, such as accelerating wound healing¹⁶, providing antioxidant benefits^17,18, preventing wrinkles², and reducing the appearance of dark spots^19,20. Recent studies have shown that topical formulations containing Moringa oleifera extract can revitalize human skin¹³, facilitate atopic dermatitis in vivo^21,22, and act as a potential skin whitening agent²³, which has led to the increased demand for this ingredient in cosmetics. Herbal cosmetics can successfully shield the skin against allergies, infections, and other skin disorders. Because of their improved safety profile, efficacy, high quality, affordability, and reduced side effects, herbal cosmetics are favoured over synthetic ones. Herbal extracts from different plant sections can be made and used in a variety of skin care cosmetic creams, lotions, and ointments ²⁴.

The formulation of Moringa has been studied in previous research into oral administration, such as lozenge ²⁵ and chewable gummy ²⁶. However, the research of Moringa formulation as a topical dosage form is still lacking. The objective of this study is to formulate moringa leaf extract into nanoemulsion. In prior work, Pakan et al. developed topical gel preparations that contained ethanol-based extracts of Moringa oleifera leaves at doses of 5%, 10%, and 15% for use against Staphylococcus epidermidis ²⁴. This study aims to develop a nanoemulgel formulation from Moringa leaves, standardize the process, and evaluate the dosage form.

MATERIALS AND METHODS:

Materials:

Ethanol and rose oil were purchased from Brataco Chemical (Indonesia), while potassium dihydrogen phosphate (KH₂PO₄₎, NaOH, L-tyrosin, tyrosinase from mushroom, and kojic acid were provided by Sigma Aldrich (Singapore). Other chemicals were purchased for nanogel formulation, including propylene glycol, aquadest (Dow Chemical Pacific, Indonesia), carbopol 934 (PT. Dunia Kimia Jaya Indonesia), cremophor EL (PT. KAO Indonesia Chemicals), dimethylol-5-5-dimethy hydantoin (Beijing Biotech).

Methods:

Figure 1 illustrates the various stages of research for this study. Moringa leaf was processed into Moringa leaf extract (MLE) and subsequently standardized. The MLE was evaluated for its phytochemical composition, antibacterial properties, tyrosinase inhibition, and antioxidant activities. MLE was transformed into nanoemulsions and subsequently assessed through the measurement of particle size, zeta potential, and polydispersion index. The nanoemulsion was subsequently transformed into a hydrogel form, followed by an evaluation of the sensory attributes of the nanoemulsion-based hydrogel.

Figure 1. The phases of investigation in this research. The process involves preparation, standardization, activity evaluation, and formulation into a hydrogel nanoemulsion.

Plant extract preparation and standardization:

Moringa leaf powder was obtained from PT. Moringa Organik Indonesia (Lombok, Indonesia). A voucher for powder specimen no. BF-UII-pm-004 was deposited at the Biopharmaceutical Laboratory, Department of Pharmacy, Universitas Islam Indonesia and the microscopical analysis was performed. The extract was prepared by macerating 2 kg of Moringa leaf powder in 20 L of ethanol for 24 hours, followed by twice re-maceration. The filtrates were evaporated using a rotary evaporator (Heidolph).

Detection of chemical constituents by UHPLC-HRMS:

The phytochemical contents of the extract were analyzed using ultra-high-performance liquid chromatography-high resolution mass spectrometry (UHPLC-HRMS) with a Vanquish Tandem Q Exactive Plus Orbitrap HRMS instrument (Thermo Scientific, USA), following a previously modified method²⁷. The UHPLC separation was performed using the column Accucore C18 (Thermo Scientific) with a particle size of 1.5 μm and dimensions of 10 mm × 2.1 mm at a flow rate of 0.2 ml/min. The eluent contained H₂O + 0.1 % formic acid (A) and acetonitrile + 0.1 % formic acid (B) with a gradient of 0–1 minute (5 % B), 1–25 minutes, and 0–1 minute (5 % A) (5%–95% B), 25–28 minutes (95 % B), 28–30 minutes (5 % B). The column temperature was set to 30°C. The injection volume was 2.5 µl, and the mass range was 100–1500 m/z in both positive and negative ionization modes. Mass detection was performed using heated-electrospray ionization (H-ESI). Full MS was performed at 70,000 resolutions, and data-dependent MS2 was performed at 17,500 full widths at half maximum (FWHM). The capillary temperature of the H-ESI source was set to 320°C, and the aux gas heater temperature was 39°C. Thermo Scientific™ Compound Discoverer Software was used to identify the compounds.

Determination of the antioxidant activity by the DPPH assay:

The 2,2′-Diphenyl-1-picrylhydrazyl (DPPH) assay was performed using a modified version of the procedure reported by Braham et al ²⁸ . A total of 5 µl of Moringa leaf extract (MLE) at concentrations of 1000, 800, 400, 200, and 100 ppm were added to each well of a 96-well clear polystyrene microplate containing 150 µl of 30 µg/mL DPPH solution using ascorbic acid as a standard. All experimental procedures were performed in triplicate. After 30 minutes of incubation at room temperature in the dark, the absorbance at 510 nm was measured using a microplate reader (Thermo Scientific™ Multiscan™ GO Microplate Spectrophotometer). Radical scavenging activity was calculated according to the following formula:

Ab - As

Inhibition rate % = ------------ X 100 ………….(1)

Where Ab is the absorbance of the blank and As is the absorbance of the sample, the 50% inhibitory concentration (IC₅₀) value was defined as the extract concentration required to scavenge 50% DPPH radicals.

Tyrosinase inhibition assay:

The tyrosinase inhibition assay was performed following the previous study ²⁹ with minor modifications. The sample solution was prepared by dissolving Moringa extract in 0.8% DMSO at a concentration of 100,000 ppm. The test solutions were prepared at a series of concentrations: 10,000 ppm, 8,000 ppm, 4000 ppm, 2000 ppm, and 1000 ppm. The assay medium was prepared by combining 30 µl of 1 mM mushroom tyrosinase solution (250 U/mL, Sigma), 100 µl phosphate buffer solution (pH 6.8), and 70 µL of sample. The mixture was preincubated at 25 °C for 10 minutes, and then 100 µL of an L-tyrosine solution (1.7 mM, Sigma) was added. After incubating the reaction mixture for 20 minutes, the absorbance was measured at 510 nm. As a control, the absorbance of a similar mixture without the extract was also measured using kojic acid as a standard. All experimental procedures were performed in triplicate. The percentage of inhibition of tyrosinase activity was calculated using the following equation:

Ac - As

% Inhibition = ------------ X 100 ………….(2)

where Ac is the absorbance of the control, and As is the absorbance of the sample. The IC₅₀ values were calculated using nonlinear regression analysis of concentration-response data.

Antibacterial assay:

The antimicrobial activity of MLE was assessed against S. aureus, S. epidermidis, and E. coli using the disk diffusion technique described in the Manual of Antimicrobial Susceptibility Testing²⁰. S. aureus (ATCC 25923), S. epidermidis (ATCC 12228), and E. coli (ATCC 35218) were purchased from Fisher Scientific. Sterile filter paper discs (6 mm in diameter, Whatman, Maidstone, England) were impregnated with 20 µL of extract dissolved in 0.8% DMSO and placed on TSA agar plates inoculated with bacteria at 10⁸ CFU/mL. The plates were then incubated at 37°C for 24 h. The microbial activity of the samples was determined by measuring the diameter of the zone of inhibition of microbial growth using the Scan 500 inhibition zone reader (Interscience). The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were determined following the Clinical and Laboratory Standards Institute microdilution method in 96-well microtiter plates²⁰.

Nanoemulsion and nanoemulsion-based hydrogel formulations:

The solubility of the oil, surfactant, and co-surfactant phases was first tested to optimize the nanoemulsion formulation. The solubility test was performed by weighing 0.5 g MLE leaf extract into multiple vials, each containing one of the three phases. Each vial containing the oil, surfactant, and co-surfactant phases was homogenized using an ultrasonic homogenizer to prepare the nanoemulsion-based hydrogel (Table 1). Nine nanoemulsion-based hydrogel formulas were prepared with different extract concentrations based on the quercetin content in MLE (0.07%–1.26%)³⁰, oils, and surfactants. The nanoemulsion was prepared by mixing the aqueous phase (mixture 1: aquadest and cremophor EL) and the oil phase (mixture 2: MLE, rose oil, and propylene glycol) in a beaker glass, followed by homogenization using an ultrasonic homogenizer (Biologic 300 VT). Mixture 1 was then added to mixture 2, and the final mixture was homogenized again using an ultrasonic homogenizer (Biologic 300 VT). To prepare the nanoemulsion-based hydrogel, carbopol 934 was combined with aquadest and homogenized. Next, TEA was gradually added to the mixture while stirring to form a gel base. The gel base was then homogenized at 2000 rpm for five minutes to create a uniform and stable mixture. The gel base was allowed to equilibrate 24 hours after formation. The MLE nanoemulsion was then gradually added to the base gel while homogenizing at 3000 rpm for 4 minutes until the nanoemulsion was completely incorporated. Finally, DMDM hydantoin was added as a preservative to the nanoemulsion-based hydrogel.

Nanoemulsion and nanoemulsion-based hydrogel evaluation:

Particle size, zeta potential, and polydispersion index of the nanoemulsion were measured with a Nanoparticle Analyzer SZ-100 instrument (Horiba Scientific) after 100x dilution³¹. Meanwhile, the characteristics of the nanoemulsion-based hydrogel were assessed, including organoleptic, viscosity, pH, and stability tests. The pH values were measured using a pH meter (Horiba Scientific), and the measurements were conducted in triplicate. The viscosity of the nanoemulsion-based hydrogel was determined using a cone and plate viscometer with spindle no 52 (Brookfield)³². Stability testing was carried out using the freeze-thaw method for four cycles. Each cycle consisted of storage at a temperature of 4°±2°C for 24 hours, then continued at a temperature of 40°±2°C for the next 24 hours. Observations were made for changes in colour, odour and phase separation of the preparation.

Sensory evaluation of MLE nanoemulsion-based hydrogel:

The sensory evaluation of participants in this study received ethical approval from The Ethics Committee of Universitas Islam Indonesia. All participants signed the informed consent form. Participants were over 18 years old, in good mental and physical health, free of skin blemishes, and without known allergies or sensitivities to the product being tested. After applying the gels or creams to the palms and hands, participants massaged the products until they were fully absorbed and let them dry for approximately 20 seconds. Participants rated their liking of the product on a 5-point hedonic scale, where one represented "dislike very much" and 5 "like very much."

Statistical analysis:

Statistical analysis was performed using Graph Pad Prism version 9 software. Paired and unpaired Student's t-tests were used to compare differences between and between groups, respectively. A p-value <0.05 was considered statistically significant.

Ethics statement:

This study was approved by the Faculty of Medicine UII (reference number 6/Ka. Kom. Et/70/KE/XII/2021) and conducted in compliance with the ethical standards of the institutional and national research committee and with the Helsinki Declaration of 1964 and its subsequent revisions or comparable ethical standards. All participants provided informed consent. Sixty untrained panellists were given a questionnaire to assess the sensory evaluation of the nanoemulsion-based hydrogel.

RESULT AND DISCUSSION:

Extract standardization:

The Indonesian Herbal Pharmacopeia defines specific and non-specific parameters for extract standardization³³. Specific parameters determine the efficacy of an extract, while non-specific parameters affect its safety/toxicity. Table 2 presents the result of MLE standardization.

Table 1. Composition of Moringa leaf extract (MLE) nanoemulsion-based hydrogel

Ingredients		Composition % (w/w)
Ingredients		F1	F2	F 3	F4	F5	F6	F7	F8	F9
Nanoemulsion	MLE	0.25	0.5	1	0.25	0.5	1	0.25	0.5	1
	Rose oil	1	1	1	-	-	-	-	-	-
	Lemon oil	-	-	-	1	2	2	-	-	-
	Capryol 90	-	-	-	-	-	-	1.4	1.4	1.4
	Cremophor EL	1	1	1	1	2	2	2	2	2
	Propylene Glycol	1	1	1	0.5	1	1	1	1	1
	Aquadest	Ad 100	Ad 100	Ad 100	Ad 100	Ad 100	Ad 100	Ad 100	Ad 100	Ad 100
Gel base	Carbopol	0.3	0.3	0.3	0.3	0.3	0.3	0.3	0.3	0.3
	DMDM Hydantoin	0.2	0.2	0.2	0.2	0.2	0.2	0.2	0.2	0.2
	Aquadest	49.5	49.5	49.5	49.5	49.5	49.5	49.5	49.5	49.5
	TEA	qs	qs	qs	qs	s	qs	qs	qs	qs

MLE phytochemical analysis using UHPLC-HRMS:

UHPLC-HRMS analysis of MLE metabolites was performed in negative and positive ionization modes (Figure 2). The metabolites detected in the extract are listed in Supplementary Table 1. Phytochemical analysis using UHPLC-HRMS revealed that MLE contains phenolic and flavonoid compounds, which may contribute to its antioxidant activity. Flavonoids interact with the reactive component of free radicals to stabilize them, as described by the following equation given by Panche et al.³⁴:

Figure 2. UHPLC-HRMS total ion chromatograms of Moringa oleifera leaf crude extract in negative ionization (NI) and positive ionization (PI) modes at retention times ranging from 0 to 30 minutes

Flavonoids (F-OH) scavenge free radicals (•R) by donating hydrogen atoms (H) to form flavonoid radicals (F-O•). These flavonoid radicals then react with a second free radical (•R) to form a stable quinone.

The antioxidant activity of flavonoids is influenced by the number of hydroxyl groups they contain: the more hydroxyl groups in the A or B rings, the more hydrogen atoms can be donated to free radicals ³⁵. For example, quercetin has five hydroxyl groups and stronger antioxidant activity than isorhamnetin 3-glucoside, which has three hydroxyl groups³⁵, as evidenced by the DPPH assay. This is because quercetin's hydroxyl groups can be easily oxidized by free radicals. Rutin is also a potent antioxidant, inhibiting xanthine oxidase, an enzyme that produces Reactive Oxygen Species (ROS)³⁶. Other compounds with previously reported antioxidant activity include 6,8-di-c-glucosyl apigenin (nicotiflorin)³⁷, quercetin, isoquercetin³⁸, and kaempferol and its derivates²⁸.

Antioxidant, antityrosinase, and antibacterial properties of MLE:

The DPPH radical assay was employed to assess the free radical scavenging potential of the test compounds. It measures the ability of a compound to donate a hydrogen atom to DPPH, converting it to a more stable DPPH free radical. Antioxidants interact with the DPPH radical by transferring an electron or hydrogen atom, converting it to 1-1,diphenyl-2-picrylhydrazyl. The decrease in DPPH radicals results in a colour change from purple to pale yellow, indicating scavenging activity^28,39. The IC₅₀ value for MLE was 0.21 mg/mL, while vitamin C, the standard antioxidant, had an IC₅₀ value of 0.096 mg/mL ppm (Table 3). Tyrosinase inhibitors are of high interest in the cosmetic industry due to their skin-whitening properties. Overproduction of tyrosinase in the skin can lead to excessive melanin production, which can cause melanoma and other dermatological disorders⁴⁰. MLE showed antityrosinase activity, primarily attributed to its flavonoid and polyphenol content, such as quercetin and kaempferol (Supplementary Table 1)^41–43. Flavonoid tyrosinase inhibitory activity is strongly influenced by the number and position of phenolic hydroxyls. Flavonoids with more phenolic hydroxyls exhibit greater antioxidant and antityrosinase activities³⁵.

Table 3. Antityrosinase and antioxidant activities of MLE

Sample	Antioxidant activity (mg/mL)	Antityrosinase activity (mg/mL)
Kojic acid	-	0.096
Vitamin C	0.096	-
Moringa leaves extract	0.21	9.394

Our study found that MLE exhibits activity against Gram-positive bacteria (S. aureus and S. epidermidis) but not against Gram-negative bacteria (E. coli), as revealed by the disk diffusion method (Table 4). This result differs from previous studies, which reported that MLE is active against both Gram-positive and Gram-negative bacteria, with MIC values of 400 µg/mL and 500 µg/mL against S. aureus and E. coli, respectively^44,45. Our findings also differ from those reporting that MLE exhibits minimal antibacterial activity against Gram-negative bacteria and no inhibition against Staphylococcus aureus. These inconsistencies may be due to the variability of the chemical compounds present in the extract⁴⁶.

Table 4. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of MLE against bacteria

Bacteria	MIC (mg/mL)	MBC (mg/mL)
S. aureus	2.5	5
S. epidermidis	2.5	5
E. coli	Not Active	Not Active

Nanoemulsion-based hydrogel formulation:

Nanoparticle preparations must have particle sizes in the 10–100 nm range and a narrow particle size distribution. The particle size, polydispersity index, and zeta potential of the nine (9) nanoemulsion formulas are shown in Table 5.

Table 5. Particle size, polydispersity index, and zeta potential of MLE nanoemulsions

Formula	Particle size (nm)	Polydispersity Index (PI)	Zeta potential (mV)
1	48,1 ± 5,1	0,531 ± 0,032	–11,27 ± 0,71
2	72,5 ± 0,7	0,553 ± 0,553	–12,57 ± 1,12
3	74,4 ± 4,8	0,505 ± 0,505	–12,93 ± 0,45
4	30,3 ± 0,47	0,295 ± 0,03	–60,8 ± 0,28
5	37,9 ± 0,61	0,396 ± 0,03	–68,2 ± 0,46
6	44,5 ± 0,39	0,474 ± 0,02	–60,9 ± 0,37
7	65,13 ± 1,52	0,31 ± 0,06	–5,27 ± 1,32
8	67,60 ± 2,34	0,26 ± 0,06	–8,63 ± 0,80
9	87,90 ± 6,67	0,16 ± 0,04	–9,53 ± 0,95

Nanoemulsions were formulated into nanoemulsion-based hydrogel by adding base gel. The nanoemulsion-based hydrogel products are shown in Figure 3.

Figure 3. Nanoemulgel formulations of the Moringa leaf extract (MLE)

The colour intensity of the nine formulas varied depending on the extract concentrations. All formulas met the pH requirement for cosmetics (4.5 – 6.5). The viscosity of the MLE nanoemulgel was also in the range of 230–1150 cPs (Table 6).

Evaluation of the sensory properties of MLE nanoemulgel:

A test was designed to assess the sensory properties of MLE formulations, including aroma, colour, texture, and skin irritation. The hedonic test results are presented in Table 7. Based on the results, formula 7 was the most preferred formulation.

Table 6. Physicochemical assessment of the MLE nanoemulsion-based hydrogel

Formula	Colour	Homogeneity	Consistency	pH	Viscosity (cP)
1	Greenish yellow	Homogenous	Slightly thick like serum, light and transparent, fast absorption rate	4.89 ± 0.02	388.1 ± 18.3
2	Brownish green	Homogenous	Slightly thick like serum, light and transparent, fast absorption rate	4.86 ± 0.00	289.3 ± 13.4
3	Brown	Homogenous	Slightly thick like serum, light and transparent, fast absorption rate	5.33 ± 0.00	330.1 ± 13.7
4	Bright yellow	Homogenous	Viscous, moderate absorption rate	5.30 ± 0.08	1739 ± 16.78
5	Greenish yellow	Homogenous	Viscous, moderate absorption rate	5.08 ± 0.06	1033 ± 7.56
6	Dark green	Homogenous	Viscous, moderate absorption rate	5.44 ± 0.04	628 ± 6.24
7	Greenish yellow	Homogenous	Soft, fast absorption rate	4.81 ± 0.14	269.82 ±1.1
8	Brown	Homogenous	Soft, fast absorption rate	4.81 ± 0.01	134.46 ±0.3
9	Dark Brown	Homogenous	Soft, fast absorption rate	5.19 ± 0.15	91.93 ±0.2

Table 7. Sensory testing results

Parameter	Formula
Parameter	F1	F2	F3	F4	F5	F6	F7*	F8	F9
Smell	5.13	4.9	5.33	4.77	5.27	4.93	5.4	5.1	4.5
Colour	5.33	4.9	4.67	5.53	5.3	4.47	5.6	5.3	4.8
Texture	5.23	5.2	5.33	5.10	5.2	5.17	5.5	5.7	5.4
Total	15.69	15	15.33	15.4	15.77	14.57	16.5	16.1	14.7

*No irritation occurred during the observational procedure

All formulas met the requirements for nanoemulsion, with particle size in the nanometer range with uniform size (PI < 0.7). The zeta potential value varies because it is influenced by the type of oil, surfactant and co-surfactant used. A high zeta potential value indicates a strong repulsive force between negatively charged particles, resulting in a suitable dispersion system and a more stable preparation³¹. Based on the stability test data, there was no change in colour, odour, and phase separation. The colour, freshness, and phase of the preparation after the freeze-thaw test were relatively the same as at the beginning of the test. This indicates that hydrogel-based nanoemulsions have good stability or resistance.

CONCLUSION:

We found that standardized MLE has antioxidant, antityrosinase, and antibacterial properties. The metabolomic analysis of the extract revealed that polyphenols and flavonoids may be responsible for its biological activity. MLE can be formulated into a nanoemulsion-based hydrogel with favourable physicochemical characteristics and hedonic test results. The best compositions to formulate moringa leaf extract into nanoemulsion-hydrogel were Capryol 90 (1.4%), Cremophor EL (2%), Propylene Glycol (1%), and as gel base, Carbopol (0.3%) and DMDM hydantoin (0.2%) were used. The most preferred formulation should be evaluated further in in vivo models to assess drug release and long-term toxicity.

CONFLICT OF INTEREST:

The authors report no conflicts of interest in this work.

ACKNOWLEDGMENTS:

This work was supported by the Directorate of Research and Community Services, Universitas Islam Indonesia, with approval number 006/Dir/DPPM/70/Pen. Unggulan/XI/2021.

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Received on 15.01.2025 Revised on 03.05.2025

Accepted on 14.07.2025 Published on 13.01.2026

Available online from January 17, 2026

Research J. Pharmacy and Technology. 2026;19(1):105-112.

DOI: 10.52711/0974-360X.2026.00016

This work is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License. Creative Commons License.

Parameter		Result	Reference value	Note
Specific	Organoleptic	Thick extract, brownish-green colour, distinctive odour, bitter taste	Thick extract, brownish-green colour, distinctive odour, bitter taste	Suitable
	Yield	15.34%	>9.2%	Suitable
	Identity compound	Quercetin	Quercetin	Identified
Non-specific	Moisture content	8.71±0.64 %	< 10%	Suitable
	Pb content	0,51 ± 0,03 mg/L	< 10 ppm	Suitable
	Cd content	< 0,04 ± 0,00 mg/L	< 0.3 ppm	Suitable