INTRODUCTION:

The skin consists of three main layers known as the hypodermis, dermis, and epidermis. First, the epidermis is a non-vascular tissue primarily composed of keratinocytes (90-95%), along with minor populations of melanocytes, Langerhans cells, and Merkel cells.

Second, the intermediate layer of the skin, consists of connective tissue primarily made up of collagen and elastin, which provide structural strength and elasticity. These fibers are supported by hyaluronic acid, which helps maintain moisture and skin integrity. Additionally, the dermis layer contains various specialized structures, such as sweat and sebaceous glands, capillaries, sensory nerves, and immune-related cells like mast cells, macrophages, and fibroblasts. Third, the hypodermis, which is the innermost skin layer, mainly comprises adipose tissue and performs essential activities including energy storage, thermal insulation, and anchoring the skin to the underlying muscle.^1,2 The skin, as the largest organ of the human body, experiences aging from both internal and external influences. Intrinsic aging is a natural process influenced by genetic predisposition, leading to gradual structural and functional changes over time. Conversely, extrinsic aging is predominantly influenced by environmental factors such as ultraviolet (UV) radiation, air pollutants, and elevated temperatures. Continual exposure to these factors hastens skin aging, resulting in wrinkles, dryness, reduced elasticity, sagging, and an uneven texture. These effects are largely attributed to the breakdown of essential structural components, such as collagen, elastin, and hyaluronic acid. UV radiation triggers molecular reactions within the skin, primarily by promoting the formation of reactive oxygen species (ROS) via photochemical reactions and cellular oxidative metabolism. This sequence of events results in DNA damage, protein degradation, and lipid peroxidation, while simultaneously depleting the skin's antioxidant defenses.^3,4

Natural bioactive compounds with the ability to counteract skin deterioration are gaining popularity due to modern consumers' preference for ingredients that are safe, environmentally friendly, and scientifically validated.⁵ S. trilobatum, commonly known as Ma Waeng Kruea in Thailand, is a climbing shrub. It belongs to the Solanaceae family and is acknowledged for its notable therapeutic use in traditional medicine. It is also listed in Thailand's National List of Essential Medicines. The leaves of S. trilobatum are rich in iron, fiber, calcium, carbohydrates, minerals, fats, and phosphorus. Traditionally, S. trilobatum has traditionally served in the management of multiple ailments, including hypoglycemia, asthma, coughs, and skin diseases. The plant contains phytochemical constituents such as alkaloids, phenols, and specific bioactive compounds like solanine and tomatine, which contribute to its diverse medicinal properties, including anti-inflammatory, antifungal, and antibacterial activities.^6,7,
8,9

Current literature reveals a lack of comparative studies on the antioxidant activity, total phenolic content, and the inhibitory effects on elastase, collagenase, and hyaluronidase, as well as the cytotoxicity of ethanolic extracts from S. trilobatum unripe and ripe fruits. These parameters are crucial for assessing their potential use as anti-aging agents. Accordingly, this research was designed to examine and contrast the antioxidant capacity and enzyme-inhibitory effects related to skin aging of ethanolic extracts from both unripe and ripe S. trilobatum fruits.

MATERIALS AND METHODS:

Materials:

The following substances were obtained from Sigma-Aldrich: L-ascorbic acid, tannic acid, α-tocopherol, epigallocatechin gallate, butylated hydroxytoluene, Folin and Ciocalteu′s phenol reagent, DPPH, ABTS, TPTZ. Additionally, collagenase type I, elastase type I, FALGPA, AAAPVN, sodium hyaluronic, p-dimethylaminobenzaldehyde, and Trizma^®base were used. All experiments were conducted using analytical-grade chemicals and solvents.

Cell cultures: Human skin fibroblast cells (HFF-1) obtained from ATCC (USA) were maintained under standard culture conditions.

Extraction:

S. trilobatum unripe (white) and ripe (red) fruits were collected from a Thai herbal garden in Nakhon Pathom province and verified by the Bangkok Herbarium, Thailand (voucher specimen no. BK085694). After washing and air-drying, the fruits were oven-dried at 50°C until fully dehydrated, then pulverized using an herbal grinder. The powdered fruit was extracted with ethanol at a 1:10 ratio. The extraction process involved ultrasound-assisted maceration at 40 kHz and 40°C for two cycles of 30 minutes each, followed by a 7-day maceration period. After being macerated, the extract solution was filtered and concentrated using a rotary vacuum evaporator at low pressure. This made a crude extract that was then kept at -20°C until it was analyzed. The extraction yield (w/w) was determined following the methodology outlined by Khuanekkaphan et al.¹⁰

Total Phenolic Content:

A modified Folin-Ciocalteu procedure was employed to quantify the total phenolic content (TPC), as outlined by Khuanekkaphan et al.¹⁰ Briefly, 100 µL of sample was diluted with distilled water and reacted with Folin-Ciocalteu reagent. After incubation, sodium carbonate solution was added to initiate color development. Following a 90minute incubation, absorbance was recorded at 725nm, and the total phenolic content was determined in terms of gallic acid equivalents using a standard calibration curve (2.5 to 40µg/mL).

GC-MS Analysis:

The ethanolic extracts of S. trilobatum unripe and ripe fruits were analyzed using GC-MS with an HP5MS capillary column (30m × 250µm × 0.25µm) to identify phytochemical constituents. Ultra-pure helium was used to inject a 2µL hexane-diluted sample in splitless mode at a rate of 1.0mL/min. First, the oven was preheated to 70°C for a duration of 2 minutes. Then heated it to 250°C at a rate of 10°C per minute for 5 minutes and then to 300°C at the same rate for an additional 5 minutes. The mass spectrometry was conducted with a 280°C transfer line temperature and a 3-minute solvent delay, scanning 40-1000amu. To identify compounds, mass spectra were compared to the NIST spectral database.

Evaluation of Antioxidant Activities:

a) DPPH assay:

The DPPH assay was employed to assess antioxidant activity. In summary, 100 µL of each sample was mixed with an equal volume of freshly prepared 0.2mM DPPH solution and dispensed into a 96-well microplate. The reaction mixture was incubated in the absence of light for 30minutes. Absorbance was subsequently recorded at 517nm using a microplate reader. L-ascorbic acid was utilized as the standard reference antioxidant. The percentage of inhibition and IC₅₀ values were calculated based on the procedure described by Noysang et al.¹¹

b) ABTS assay:

The ABTS assay was conducted following the method described by Khuanekkaphan et al.¹² with slight alterations. The ABTS radical cation (ABTS•⁺) was generated and diluted with ethanol to achieve an optical density of approximately 0.70 at 734nm. Samples (20 µL) were combined with the radical solution (2mL), and the resultant mixture was subjected to brief incubation. The absorbance was then recorded at 734 nm, and the radical scavenging capacity was expressed relative to L-ascorbic acid. IC₅₀ values were calculated based on the percentage inhibition.

c) FRAP assay:

Ferric-reducing antioxidant power was assessed following a modified procedure of Janmin et al.¹³ A freshly mixed FRAP reagent was used, and each sample (20µL) was reacted with the reagent (1.8mL) and incubated for 4minutes at room temperature. The resulting absorbance was then measured at 595 nm, and values were determined based on a calibration curve prepared from FeSO_4·7H₂O.

Evaluation of Anti-Aging Potential via Enzymatic Inhibition:

a) Collagenase inhibition:

An enzymatic assay using FALGPA substrate was employed, following a modified version of the collagenase enzymatic assay (EC 3.4.24.3). The test sample (20µL) was mixed with collagenase solution (20 µL) and incubated at 25°C for 10minutes. Then, 200µL of 1mM FALGPA substrate was introduced. Absorbance at 340nm was recorded at 30second intervals for a duration of 10minutes. EGCG served as a positive control. Enzyme inhibition was assessed by measuring the decrease in absorbance over time.

b) Elastase inhibition:

The elastase inhibitory activity was assessed via a modified AAAPVN-based enzymatic test, derived from the enzymatic test of elastase (EC 3.4.21.36). A test sample of 20µL was mixed with 180µL of elastase enzyme solution (0.5U/mL in 1 M Tris-HCl buffer). The mixture was pre-incubated at 25°C for 10minutes prior to substrate addition. Subsequently, 40µL of 2.2mM AAAPVN substrate was introduced. Absorbance at 410 nm was recorded at 30-second intervals for a duration of 10minutes. EGCG served as the positive control. The decrease in kinetic absorbance was quantified to determine the percentage of elastase inhibition.

c) Hyaluronidase inhibition:

The inhibitory effect on hyaluronidase was assessed following an adapted protocol from Jiratchayamaethasakul et al.¹⁴ To initiate the reaction, a sodium hyaluronate solution (1.2mg/mL) prepared in acetate buffer at pH 3.5 was mixed with 50µL of the test sample and incubated at 37°C. Following the sequential addition of CaCl₂, sodium hyaluronate substrate, sodium borate, and DMAB reagent, by absorbance detection at 585nm. The assay employed tannic acid as a standard positive control.

Cytotoxicity assay in HFF-1 cells:

The cytotoxic effects of ethanolic extracts from unripe and mature S. trilobatum fruit were evaluated in HFF-1 cells with the MTT test, based on a modified protocol outlined by Pressi et al. and Taofiq et al.^15,16 Cells were seeded in 96-well plates (2 × 10⁴ cells/well) and incubated for 24hours prior to treatment with 100 µL of extract at various concentrations. After 24 hours of treatment, MTT solution (5.0mg/mL) was introduced and incubated for 3hours. Formazan crystals formed were dissolved in DMSO, and absorbance was recorded at 570nm. Cell viability (%) was determined using the following equation:

Cell viability (%) =	OD _sample	× 100
	OD _control

Statistical analysis:

The results were reported as mean±SD based on three independent replicates. Statistical comparisons were analyzed using one-way ANOVA in GraphPad Prism 5.01, with significance defined at p<0.05.

RESULT:

Extraction Yield:

The ethanolic extract of S. trilobatum ripe and unripe fruits yielded differing amounts. The ripe fruits produced a higher yield (10.73%±0.74%) than the unripe fruits (9.32%±1.12%) (Table 1).

Total Phenolic Content:

The TPC of the ripe fruit extract was significantly higher (12.91±0.97mg GAE/g crude extract) than that of the unripe fruit extract (8.33±1.84 mg GAE/g crude extract) (Table 1).

GC-MS Analysis:

GC-MS profiling showed distinct phytochemical compositions between the unripe and ripe fruit extracts (Figure 1 and Figure 2). The major components of the unripe extract were 1,4-dioxaspiro[4.4]nonane, 7-butyl-(R,R)-2,3-dimethyl- (34.71%), glycerol tricaprylate (26.10%), sarcosine, N-(1-naphthoyl)-, octyl ester (15.87%), oleic acid (7.02%), and hexadecanoic acid, 2-hydroxy-1-(hydroxymethyl)ethyl ester (4.78%) (Table 2). The ripe extract was rich in hexadecanoic acid, 2-hydroxy-1-(hydroxymethyl)ethyl ester (47.98%), octadecanoic acid, 2,3-dihydroxypropyl ester (22.03%), hydroxylamine, O-methyl- (9.53%), silane, triethyl-1-pentenyl- (6.98%), and glycerol tricaprylate (6.57%) (Table 3).


Figure 1. Presents the *GC-MS chromatogram of ethanolic extract from unripe S. trilobatum* fruits.**	Figure 2. Presents the *GC-MS chromatogram of ethanolic extract from ripe S. trilobatum* fruits.**

Table 2. Five major phytochemical components of ethanolic extract from unripe S. trilobatum fruits and their reported biological activities.

RT (min)	Chemical Name	Area (%)	Biological Activity	Reference
18.957	Oleic Acid	7.02	Anti-inflammatory	18
22.684	Hexadecanoic acid, 2-hydroxy-1-(hydroxymethyl)ethyl ester	4.78	Antioxidant, pesticidal, nematicidal.	19
30.066	Glycerol tricaprylate	26.10	Skin conditioning and antihistaminic	20
32.887	1,4-Dioxaspiro[4.4]nonane, 7-butyl-(R,R)-2,3-dimethyl-	34.71	NI	NI
35.415	Sarcosine, N-(1-naphthoyl)-, octyl ester	15.87	NI	NI

NI = No information available in current literature sources.

Table 3. Five major phytochemical components of ethanol extract from ripe S. trilobatum fruits and their reported biological activities.

RT (min)	Chemical Name	Area (%)	Biological Activity	Reference
1.267	Hydroxylamine, O-methyl-	9.53	NI	NI
22.701	Hexadecanoic acid, 2-hydroxy-1-(hydroxymethyl)ethyl ester	47.98	Antioxidant, pesticidal, nematicidal, anti-microbial	19, 21
26.094	Octadecanoic acid, 2,3-dihydroxypropyl ester	22.03	Antioxidant Anti-microbial	21
30.051	Glycerol tricaprylate	6.57	Skin conditioning and antihistaminic	20
32.866	Silane, triethyl-1-pentenyl-	6.98	NI	NI

NI = No information available in current literature sources.

Evaluation of Antioxidant Activities:

a) DPPH assay:

The antioxidant potential of ethanolic extracts from S. trilobatum unripe and ripe fruits was assessed using the DPPH assay. The IC₅₀ values were 0.47±0.01mg/mL for the unripe extract and 0.37±0.02mg/mL for the ripe extract (Table 1). The lower IC₅₀value of the ripe extract compared to the unripe one suggests stronger free radical scavenging activity. L-ascorbic acid exhibited significantly higher activity, with an IC₅₀ of 0.02±0.00 mg/mL.

b) ABTS assay:

In the ABTS assay, which evaluates the ability of antioxidants to quench the ABTS⁺• radical, the unripe and ripe extracts exhibited IC₅₀ values of 19.54±0.06 mg/mL and 18.34 ± 0.04mg/mL, respectively (Table 1). L-ascorbic acid exhibited significantly higher activity, with an IC₅₀ of 0.22± 0.00mg/mL.

c) FRAP assay:

The FRAP assay, which measures electron-donating capacity, showed that the unripe and ripe fruit extracts had FRAP values of 248.76±9.14 and 290.89±8.96 μmol Fe(II)/g extract, respectively. L-ascorbic acid exhibited a markedly higher reducing power (13,683.14±33.85μmol Fe(II)/g extract).

Evaluation of Anti-Aging Potential via Enzymatic Inhibition:

a) Collagenase inhibition:

The collagenase inhibitory activity of ethanolic extracts from S. trilobatum unripe and ripe fruits (1mg/mL) was evaluated and compared with epigallocatechin gallate (EGCG, 0.1mg/mL) as a positive control (Figure 3). The ripe fruit extract exhibited significantly higher collagenase inhibition (70.00% ±3.54%) than the unripe fruit extract (56.25% ±1.77%) (p<0.05). EGCG showed the highest inhibition at 86.25%±5.30%.

Figure 3. Presents the anti-collagenase activity of S. trilobatum fruit ethanolic extracts compared with EGCG. The values are presented as the mean±SD and displayed statistical significance (*p<0.05 and **p<0.01).

b) Elastase inhibition:

The elastase inhibitory activity of ethanolic extracts from S. trilobatum unripe and ripe fruits (1mg/mL) was also determined, using EGCG (0.1mg/mL) as a positive control (Figure 4). The unripe extract showed 51.37%± 2.91% inhibition, whereas the ripe extract exhibited significantly higher inhibition at 62.63%±2.86% (p< 0.01). EGCG presented the highest inhibition at 91.19% ±2.06%.

Figure 4. Presents the anti-elastase activity of S. trilobatum fruit ethanolic extracts compared with EGCG. The values are presented as the mean±SD and displayed statistical significance (*p<0.05 and **p< 0.01).

c) Hyaluronidase inhibition:

The hyaluronidase inhibitory activity of the ethanolic extracts from S. trilobatum unripe and ripe fruits (1 mg/mL) was compared to tannic acid (0.1mg/mL) as a positive control (Figure 5). The unripe fruit extract inhibited hyaluronidase by 51.64%±2.19%, while the ripe extract showed slightly greater inhibition at 60.82% ±2.60%. Tannic acid exhibited the strongest inhibitory activity at 83.85%±2.81% (p< 0.01).

Figure 5. Presents the anti-hyaluronidase activity of S. trilobatum fruit ethanolic extracts compared with Tannic acid. The values are presented as the mean±SD and displayed statistical significance (*p < 0.05 and **p<0.01).

Cytotoxicity assay in HFF-1 cells:

The cytotoxic effects of ethanolic extracts from S. trilobatum unripe and ripe fruits were evaluated in HFF-1 cells. Both extracts exhibited no significant cytotoxicity at concentrations up to 1mg/mL, as cell viability remained above 90% (Figure 6). However, a dose-dependent reduction in cell viability was observed at concentrations exceeding 1mg/mL. At 2mg/mL, viability dropped below 70%, indicating potential cytotoxic effects at higher doses.

Figure 6. Presents cell viability of HFF-1 cells after 24-hour exposure to S. trilobatum fruit ethanolic extracts at different concentrations (mg/mL) compared with control or untreated cells. The values are presented as the mean±SD.

DISCUSSION:

The higher extraction yield observed in ripe fruits may be attributed to the progressive accumulation of bioactive constituents such as secondary metabolites, antioxidants, and other phytochemicals, which typically increase during fruit maturation. However, the concentration and composition of specific bioactive compounds at earlier developmental stages may also significantly influence both the yield and the bioactivity of the extracts.¹⁷ The significantly higher TPC in ripe fruits compared to unripe ones further supports this observation, suggesting that fruit maturation enhances phenolic biosynthesis in S. trilobatum.¹⁷

The GC-MS analysis revealed clear compositional differences between the S. trilobatum unripe and ripe fruit extracts, indicating potential variations in their biological activities. The unripe fruit extract was rich in diverse compounds, including spiroketals (e.g., 1,4-dioxaspiro[4.4]nonane) and sarcosine derivatives (e.g., sarcosine, N-(1-naphthoyl)-, octyl ester), along with medium-chain triglycerides such as glycerol tricaprylate, and unsaturated fatty acids like oleic acid, which may contribute to pharmacological properties such as anti-inflammatory, antioxidant, or skin conditioning effects.^18,19,20 In contrast, the ripe fruit extract contained higher levels of long-chain fatty acid esters, particularly hexadecanoic acid, 2-hydroxy-1-(hydroxymethyl)ethyl ester, and octadecanoic acid, 2,3-dihydroxypropyl ester, which are known for their antioxidant, emollient, and skin-conditioning properties.^19,20^,21 The notably higher proportion of hexadecanoic acid derivative in the ripe fruit extract suggests enhanced antioxidant potential, supporting its prospective use in pharmaceutical and cosmeceutical formulations.

The antioxidant activities of S. trilobatum fruit extracts were evaluated using three standard assays, including DPPH, ABTS, and FRAP.^22,23^,2⁴ The DPPH assay is a widely used, cost-effective method that assesses antioxidant capacity through a colorimetric shift, where the stable DPPH radical changes from deep purple to pale yellow upon reduction.¹⁷ In this study, the ripe fruit extract exhibited a lower IC₅₀ value than the unripe fruit extract, indicating stronger radical scavenging activity. Similarly, the ABTS assay, which measures both hydrophilic and lipophilic antioxidant activity²⁵, showed enhanced performance in the ripe fruit extract, as reflected by a lower IC₅₀ value. However, both fruit extracts were less potent than the standard L-ascorbic acid. The FRAP assay, which measures electron-donating capacity, further confirmed the superior reducing power of the ripe fruit extract through higher FRAP values (μmol Fe(II)/g).²⁶ These antioxidant experiments demonstrated a clear correlation between total phenolic content and antioxidant activity in S. trilobatum fruit extracts. Extracts with higher phenolic content are likely to exhibit stronger radical scavenging potential.²⁷ However, despite these improvements, both ripe and unripe fruit extracts were consistently less effective than pure L-ascorbic acid across all three assays. This may be due to the crude nature of plant extracts, which contain a complex mixture of active and inactive constituents. Inactive components may dilute or interfere with the action of antioxidant molecules. In contrast, L-ascorbic acid is a pure compound with well-defined and highly efficient radical scavenging properties.²⁵^,²⁸ The antioxidant effects of S. trilobatum fruit extracts are primarily attributed to phenolic compounds such as tannins and flavonoids, which are known to donate electrons or hydrogen atoms, chelate pro-oxidant metal ions, and mitigate oxidative stress.²^{9, 30,31} GC-MS analysis further confirmed the presence of bioactive compounds in the ripe fruit extract, including hexadecanoic acid, 2-hydroxy-1-(hydroxymethyl)ethyl ester, and octadecanoic acid, 2,3-dihydroxypropyl ester, both of which are associated with antioxidant properties. These findings support the greater antioxidant potential of the ripe fruit extract and its relevance in pharmaceutical and cosmeceutical applications. Notably, herbal-based topical formulations, such as polyherbal anti-aging creams, have been developed to exploit similar antioxidant mechanisms for improving skin health and preventing photoaging.³²

Given that oxidative stress and enzymatic degradation of the extracellular matrix are critical drivers of skin aging. Consequently, phenolic compounds with antioxidant properties may also confer protective effects by inhibiting matrix-degrading enzymes. To further explore the anti-aging potential of S. trilobatum fruit extracts, the inhibitory activities of the ripe and unripe fruit extracts against collagenase, elastase, and hyaluronidase were investigated. Collagen is a primary structural protein responsible for maintaining dermal strength and firmness. Its enzymatic degradation by collagenase contributes to visible signs of aging, such as wrinkles and sagging.³³^,³⁴ In the present study, S. trilobatum ripe fruit extract demonstrated significantly stronger collagenase inhibitory activity than the unripe fruit extract. This enhanced effect is likely associated with the higher total phenolic content in the ripe fruit, as phenolic compounds have been shown to inhibit collagenase by binding to its active site and modulating enzymatic function.³⁵^,36 Elastin, another essential protein in the extracellular matrix, contributes to tissue flexibility and resilience. Elastase, a proteolytic enzyme, degrades elastin and leads to reduced elasticity, sagging, and wrinkle formation. Inhibition of elastase can help preserve elastic fibers and mitigate visible aging signs.³⁷ The ripe fruit extract exhibited significantly greater elastase inhibition than the unripe fruit extract (p<0.01), suggesting that ripening may enhance elastase-inhibitory activity. This effect also correlates with the higher phenolic content in the ripe fruit extract, consistent with reports that polyphenols act as potent elastase inhibitors.³⁸ Hyaluronic acid plays a critical role in maintaining skin hydration, while hyaluronidase breaks it down, leading to dryness and premature aging.³⁹ This degradation mechanism has been described in both dermatological and orthopedic contexts.⁴⁰ The ripe fruit extract showed moderately higher hyaluronidase inhibition compared to the unripe fruit extract. Although this effect was less pronounced than those observed with collagenase and elastase, the trend supports the role of increased bioactive content in ripe fruits. Additionally, the strong inhibition observed in the tannic acid control highlights the potential of phenolic-rich plant extracts to modulate this enzymatic pathway.³⁵ The collective findings from this study confirm that S. trilobatum ripe fruit extracts exhibit stronger inhibitory activity against collagenase, elastase, and hyaluronidase compared to unripe fruit extracts. This enhanced effect is likely attributed to a higher concentration of bioactive compounds, particularly phenolics, which have been previously associated with enzyme inhibition in anti-aging applications.³⁵^,41,42 These results support the hypothesis that fruit ripening promotes the accumulation of active phytochemicals, thereby enhancing skin-protective properties.

In addition to their bioactivity, the safety profile of the extracts is critical for potential applications. S. trilobatum is well known for its pharmacological properties, including antioxidant, anti-inflammatory, anticancer, antifungal, and antibacterial effects.⁴³ Its fruit has long been used in Thai traditional medicine, and recent studies have highlighted its potential as a natural anti-aging agent, primarily due to its antioxidant and enzyme-inhibitory activities. In this study, both unripe and ripe fruit extracts showed good biocompatibility in HFF-1 cells at concentrations up to 1 mg/mL. However, reduced cell viability at higher concentrations indicates dose-dependent cytotoxicity.

CONCLUSION:

The investigation of the antioxidant and anti-aging potential of ethanolic extracts from S. trilobatum fruit revealed distinctive properties between unripe and ripe fruit extracts. The ripe fruit extract exhibited higher total phenolic content and greater antioxidant activity compared to the unripe fruit extract. Additionally, it demonstrated stronger inhibition of collagenase, elastase, and hyaluronidase. These attributes support its potential for use in anti-aging formulations. Furthermore, cytotoxicity evaluation in HFF-1 cells confirmed the safety of both unripe and ripe fruit extracts at concentrations up to 1mg/mL, with dose-dependent effects observed at higher concentrations. Further investigation is warranted to isolate the active compounds involved and to clarify their underlying mechanisms. The findings contribute valuable insight into the cosmeceutical and pharmaceutical potential of S. trilobatum fruit extracts.

CONFLICT OF INTEREST:

The authors have no conflicts of interest regarding this investigation.

ACKNOWLEDGMENTS:

The authors would like to express their sincere gratitude to Miss Fangkao Larit for her kind guidance in identifying sources of herbal raw materials. Appreciation is also extended to the Bangkok Herbarium, Thailand for their assistance in the botanical authentication of the plant specimens used in this study. This research was financially supported by Grant No. FRB650070/0168 from the Science, Research and Innovation Promotion Funding (TSRI). The grant was administered by Rajamangala University of Technology Thanyaburi (Grant Code: FRB65E1208).

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Received on 22.04.2025 Revised on 12.08.2025

Accepted on 11.11.2025 Published on 20.05.2026

Available online from May 25, 2026

Research J. Pharmacy and Technology. 2026;19(5):2102-2110.

DOI: 10.52711/0974-360X.2026.00302

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

Test samples	Yield (%)	TPC (mg GEA/g crude extract)	DPPH IC₅₀ (mg/mL)	ABTS IC₅₀ (mg/mL)	FRAP Value μmole of Fe (II)/g sample
Unripe fruit extract of S. trilobatum	9.32 ± 1.12	8.33 ± 1.84	0.47 ±0.01	19.54 ± 0.06	248.76 ± 9.14
Ripe fruit extract of S. trilobatum	10.73 ± 0.74	12.91 ± 0.97	0.37 ± 0.02	18.34 ± 0.04	290.89 ± 8.96
L-ascorbic acid	-	-	0.02 ± 0.00	0.22 ± 0.00	13,683.14 ± 33.85