The increasing global prevalence of multidrug-resistant (MDR) bacterial infections has intensified the search for new antibacterial agents from underexplored natural sources. Endophytic bacteria associated with medicinal plants are recognized as potential producers of bioactive secondary metabolites. This study aims to isolate and characterize antibacterial endophytic bacteria from Smilax zeylanica collected in Bukit Duabelas National Park, Jambi, Indonesia. Endophytic bacteria were isolated from roots, stems, leaves and rhizomes using a surface sterilization method, which was validated by a final rinse test which showed no microbial growth. Antibacterial activity was screened using a dual culture assay on Nutrient Agar against methicillin-resistant Staphylococcus aureus (MRSA) WS 29 and Escherichia coli RES 22-2020, followed by phenotypic and genomic characterization. A total of 45 endophytic bacterial strains were obtained, of which seven exhibited inhibitory activity against both pathogens. Strain ELSZ6 showed the strongest inhibition and was characterized as Gram-positive, rod-shaped, and endospore-forming. Whole genome sequencing identified the strain as Bacillus cereus (6.3 Mbp; 50× coverage). Genome mining using antiSMASH revealed several biosynthetic gene clusters associated with secondary metabolites, including bacillibactin, zwittermicin A and petrobactin. The findings indicate that strain ELSZ6 represents a potential candidate for further investigation as a source of antibacterial compounds for use against MDR bacteria.
Many areas in Indonesia face severe pollution from copper and antibiotics, impacting microbial diversity and underscoring the need for effective remediation strategies. Harnessing of copper- and antibiotic-resistant bacteria has become a promising approach. This study evaluates the antimicrobial resistance (AMR) profile of copper-accumulating Klebsiella pneumoniae strain CJK 500 2.1.2 as a bioremediation agent candidate. Whole-genome analysis of K. pneumoniae CJK 500 2.1.2 revealed multiple AMR genes, primarily encoding efflux pumps, with one fieF gene facilitating resistance to heavy metals. The disk-diffusion assay highlighted multidrug-resistance traits, regardless of CuSO4. Notably, increased cefoxitin resistance in the presence of CuSO4 and intermediate susceptibility to tetracycline were observed, which was corroborated by a broth microdilution assay. The genome analysis identified 22 virulence factors (VFs) and 16 insertion sequences (ISs), with some ISs located near AMR and VF genes, suggesting possible horizontal gene transfer (HGT). Overall, K. pneumoniae CJK 500 2.1.2 showed a potential as a model study for bioremediation research on genetically engineered microorganism (GEM). Through risk assessment, GEMs have the possibility to be applied to copper and antibiotics bioremediation.
Leaf fall disease of Hevea brasiliensis in Indonesia and Southeast Asia is an emerging threat to rubber production. While tentatively associated with Pestalotiopsis, Colletotrichum, and Neopestalotiopsis species, the disease etiology remains poorly characterized. This study aimed to identify the fungal community associated with symptomatic rubber leaves using culture-independent ITS amplicon sequencing. Leaf samples representing three symptom severity levels were collected from Central Java and South Sumatra. Fungal community composition analysis revealed that geographic origin, rather than symptom severity, was the primary driver of community clustering. Phyllosticta and Colletotrichum were consistently dominant across all samples, with Phyllosticta displaying notably higher relative abundance. Phyllosticta was identified as a newly reported pathogen of H. brasiliensis. Minor proportions of Pestalotiopsis, Neopestalotiopsis, Corynespora, and Pestalotia were also detected. Multigene phylogenetic analysis (ITS and β-tubulin) revealed three major clades corresponding to Neopestalotiopsis, Pestalotiopsis, and Colletotrichum. Pathogenicity assays confirmed that representative isolates reproduced field symptoms. This first metagenomic assessment of the phyllosphere mycobiota in diseased rubber leaves identifies Phyllosticta as a critical yet overlooked component of the leaf fall disease complex, providing insights into fungal ecology and pathogenic interactions underlying this emerging production threat.
Dengue remains a major global health concern, with current licensed vaccines offering variable protection across the four virus serotypes and posing safety considerations related to antibody-dependent enhancement (ADE). To contribute toward improved vaccine design, we developed a bivalent virus-like particle (VLP) targeting DENV-1 and DENV-2, the two most prevalent serotypes in Indonesia. Consensus envelope (CE) protein sequences from Indonesian DENV-1 and DENV-2 strains were codon-optimized and co-expressed in Pichia pastoris GS115 using the AOX1 promoter. The expressed proteins were solubilized from membrane fractions under denaturing conditions, purified via Ni2+-affinity chromatography, and subjected to urea gradient dialysis for VLP assembly. Transmission electron microscopy confirmed the formation of spherical VLPs. Western blot analysis confirmed expression and purification via anti-His detection, and supported antigenic integrity of the envelope proteins as recognized by anti-dengue E antibodies. The study demonstrates the feasibility of using P. pastoris for chimeric dengue VLP production based on regionally relevant strains. The findings support the development of serotype-focused VLP platforms as modular components in the development of future tetravalent dengue vaccines.
Erythritol, a sugar alcohol, is commercially produced through fermentation. Yarrowia lipolytica and Moniliella pollinis serve as efficient erythritol producers due to their osmotic tolerance. Optimizing the cost-effectiveness of erythritol production involves harnessing strains capable of utilizing low-cost carbon sources. This study examines the impact of two different carbon sources (glucose and molasses) on the growth and erythritol production of native and UV-induced mutant strains of Y. lipolytica and M. pollinis. In the glucose-based medium (GYM), the M. pollinis SP5 mutant strain achieved the highest erythritol biosynthesis, with a yield mass of 0.275 g/g and a volumetric productivity of 0.052 g/L/h, while in the molasses-based medium (MYM), it attained a yield mass of 0.120 g/g and a volumetric productivity of 0.065 g/L/h. Random mutagenesis might improve erythritol biosynthesis by increasing enzyme activity and altering carbon flux. Molasses emerged as a potential carbon source for M. pollinis, which favors sucrose hydrolysis and glucose metabolism for erythritol production. To further enhance erythritol yield with M. pollinis, it is advisable to increase the concentration of molasses, adjust the carbon-to-nitrogen (C/N) ratio in the MYM medium, and optimize the fermentation systems.
Triclosan, a common antimicrobial compound, persists in wastewater and sludge because conventional treatment systems do not completely remove it, leading to ecological and health risks. This study investigates the mechanisms of triclosan phytoremediation by Glycine max and its rhizobacterium Ochrobactrum sp. MC35 under controlled hydroponic conditions. A laboratory hydroponic experiment was conducted in which soybean seedlings were exposed to 50–200 µg L–1 TCS, with and without bacterial inoculation. TCS concentrations and metabolites were quantified using LC–MS/MS, while enzymatic activities, including peroxidase, glutathione-S-transferase, laccase and esterase, were measured spectrophotometrically. Degradation kinetics were evaluated using first-order models, and principal component analysis (PCA) was applied to assess correlations among the variables. The combined plant–microbe system achieved 85.4 ± 3.2% triclosan removal within 10 days, nearly twice that of plant-only treatments. Chemical analysis confirmed transient formation of methylated and hydroxylated intermediates, followed by complete degradation through sequential demethylation, hydroxylation and oxidative cleavage pathways. Kinetic modeling exhibited first-order behavior (R2 > 0.96) with an apparent rate constant of 0.312 d–1 in the inoculated system, indicating enhanced microbial degradation. Enzymatic assays showed significant increases in peroxidase, glutathione-S-transferase, laccase and esterase activities, supporting cooperative detoxification between plant and microbe. PCA indicated a close association between enzymatic activity and degradation efficiency, suggesting coordinated rhizospheric interactions. Overall, Glycine max not only absorbed triclosan, but also enhanced its microbial breakdown, likely through exudate-mediated enzyme induction. The findings provide mechanistic insight into bioaugmented phytoremediation and highlight its potential as a sustainable, nature-based strategy for removing persistent antimicrobial pollutants from aquatic environments.
Breast cancer stem cells (bCSCs) exhibit high plasticity, therapeutic resistance and immune evasion, making them critical targets for effective cancer immunotherapy. Pentagamavunone-1 (PGV-1), a curcumin analog, and its derivative, Chemoprevention Curcumin Analog-1.1 (CCA-1.1), possess cytotoxic and reactive oxygen species (ROS)-modulating properties. However, their potential role in modulating immune-related phenotypes in bCSCs remains unclear. In the study, bCSCs derived from the MDA-MB-231 cell line were treated with PGV-1 or CCA-1.1 to evaluate cytotoxicity, apoptosis, cell cycle distribution, stemness-associated markers and immune checkpoint expression. Cell viability was assessed using the CCK-8 assay. Apoptotic, stemness and immune-checkpoint profiles were analyzed using flow cytometry. PGV-1 exhibited stronger cytotoxicity against bCSCs than CCA-1.1, characterized by lower IC50 values (10.70 and 28.78 µM for PGV-1 and CCA-1.1 respectively at 24 h) and greater induction of apoptosis in a dose-dependent manner. Both compounds at concentrations of 2.67 and 7.19 µM significantly (p < 0.001) induced G2/M cell cycle arrest and reduced the CD44+CD24– stem-like population, indicating loss of stemness and enhanced differentiation. Importantly, PGV-1 significantly downregulated PD-L1 while upregulating CD80, suggesting a shift toward immunophenotypic alterations associated with a potentially more immunogenic profile. These dual effects of cytotoxic and preliminary immunophenotypic modulation highlight the ability of PGV-1 to overcome immune resistance in bCSCs. Further investigation using immune cell–based functional assays or in vivo models is required to validate whether the observed immunophenotypic modulation translates into functional immune activation and to assess the potential of PGV-1 in a co-immunotherapy setting.
Anti‐miR molecules can suppress specific microRNA (miRNA) functions within critical signaling pathways. Chitosan acts as a delivery system for miRNAs; therefore, encapsulating miR‐203a‐3p is essential for targeted delivery and biological activity. This study investigates the impact of chitosan‐encapsulated anti‐miR‐203a‐3p nanoparticles (CS‐NPs) on the viability, proliferation, and migration of triple‐negative breast cancer (TNBC) 4T1 cells. The nanoparticles were synthe‐ sized using the ionic gelation method in a 5:1 ratio of chitosan to anti‐miR‐203a‐3p, incorporating sodium tripolyphosphate (STPP) as a crosslinker. Characterization was conducted using gel electrophoresis and particle size analysis. Cytotoxicity and cell viability were assessed through the MTT assay, while colony formation and wound healing assays evaluated cell proliferation and migration. The nanoparticles demonstrated an encapsulation efficiency of 89.47% and showed significant inhibitory effects on 4T1 cell proliferation and migration. The MTT results indicate an IC50 value of 2.454 µM, while colony formation analysis revealed that both ½ and IC50 doses significantly reduced colony numbers compared to the control. Similarly, wound healing assays showed notable inhibition of cell migration at ¼, ½, and IC50 concentrations. These findings suggest that anti‐miR‐203a‐3p‐loaded CS‐NPs may offer a promising therapeutic approach to managing aggressive breast cancer subtypes, particularly TNBC.
Tauco, a traditional Indonesian fermented soybean paste, is a promising yet under‐characterized source of dietary bioactives. To systematically evaluate its functional potential, we first identified ultrasonic‐assisted extraction (UAE) as the most efficient method, yielding the highest total phenolic content (28.70 mg GAE/g) and the strongest in vitro antioxidant activity (ABTS IC50 = 0.238 mg/mL). Untargeted LC–HRMS‐based metabolite profiling of this optimal extract revealed a diverse phytochemical profile rich in isoflavone aglycones, sterols, and specialized lipids. To link this chemical inventory to a specific health‐related mechanism, we performed molecular docking against HMG‐CoA reductase (HMGR), the rate‐limiting enzyme in cholesterol biosynthesis. This in silico screening prioritized the triterpenoid uvaol and the phytosterol stigmasterol as top‐binding candidates, with predicted binding affinities stronger than the reference ligand (−8.2 and −7.6 kcal/mol respectively). Overall, our integrated approach shows that UAE efficiently recovers a complex mixture of antioxidants from tauco and identifies specific metabolites with high predicted affinity for a key cardiometabolic target, providing a mechanistic hypothesis for its functional benefits and prioritizing lead compounds for future validation.
Type 2 diabetes mellitus (T2DM) leads to the non‐enzymatic glycation of proteins, resulting in the formation of advanced glycation end products (AGEs), which contribute to diabetic complications. Human serum albumin (HSA), a major plasma protein, undergoes structural alterations upon glycation (gHSA), reducing its stability and biological functions. Astaxanthin (ASX), a potent antioxidant, is limited by its instability and moderate binding affinity. In this study, we explore the use of copper (Cu2+) to form a stable ASX‐Cu2+ complex, enhancing the antioxidant properties of ASX and improving its interaction with HSA and gHSA. Utilizing computational approaches such as molecular docking, molecular dynamics (MD) simulations, and free energy landscape (FEL) mapping, we analyze the stability and conformational changes of HSA and gHSA upon binding with ASX and ASX‐Cu2+. The residue interaction network (RIN) analysis reveals that ASX‐Cu2+ complexes create a more robust and interconnected network of non‐covalent interactions, particularly enhancing hydrogen bonding, π‐stacking, and ionic interactions. The ASX‐Cu2+ complex at a 1:2 molar ratio significantly improved the binding affinity and structural stability of both native and glycated HSA, reducing protein fluctuations and promoting a more compact conformation. These findings suggest that ASX‐Cu2+ complexes offer therapeutic potential for stabilizing albumin under glycation‐induced stress, with implications for managing oxidative stress and diabetes‐related complications.
The marine environment represents a promising source of diverse enzymes produced by marine microorganisms, including L‐asparaginase. This has been widely studied due to its ability to hydrolyze extracellular L‐asparagine, an amino acid required for the growth of certain cancer cells. In this study, a synthetic L‐asparaginase gene derived from the marine bacterium Pseudoalteromonas tetraodonis GFC was recombinantly expressed in Escherichia coli BL21 (DE3). The gene was cloned into the pD861‐SR expression vector and transformed into E. coli BL21 (DE3). Positive transformants were confirmed by restriction digestion using SapI and Sanger sequencing. Recombinant protein expression was induced with L‐rhamnose under the control of the rhaBAD promoter. The expressed L‐asparaginase was then purified using Ni‐Sepharose affinity chromatography followed by membrane dialysis, yielding a protein purity of 73.6%. SDS‐PAGE analysis revealed a prominent protein band at approximately 37 kDa, corresponding to the expected molecular weight of recombinant L‐asparaginase. Enzymatic activity was evaluated using the Nessler method, and the purified enzyme exhibited a specific activity of 5.863 U/mg. These results demonstrate the successful recombinant expression and preliminary functional validation of L‐ asparaginase from P. tetraodonis GFC in E. coli, providing a basis for further optimization and characterization in future studies.
Piscinibacter sakaiensis, first isolated in Japan, is the only well‐characterized bacterium known to possess both PETase and MHETase, enabling complete polyethylene terephthalate (PET) degradation. To date, no additional habitats for the species have been reported. This study aims to identify homologous PETase and MHETase DNA from plastic‐contaminated landfill soils in Indonesia. Enrichment cultures were established from soil samples collected at Galuga (Bogor) and Cipeucang (South Tangerang). PCR amplification and sequencing revealed a full‐length MHETase homolog (G2MHETase, 1,812 bp) from Galuga, showing 99.4% and 99.3% nucleotide identity to MHETase from P. sakaiensis and Delftia sp. respectively. The deduced amino acid sequence shared 98.5% identity with both. In contrast, a partial PETase homolog (502 bp of 873 bp) was amplified from the Cipeucang sample, displaying 96 and 93% amino acid identity to PETase from P. sakaiensis and P. gummiphilus respectively. Nanophore NGS analysis of bacterial diversity indicated distinct microbial community profiles between the two sites. Rare taxa potentially associated with the detected genes included P. gummiphilus, Delftia sp., Delftia tsuruhatensis and Xenophilus aerolatus from Galuga, and Piscinibacter and Acidovorax from Cipeucang. These findings demonstrate the feasibility of detecting homologous PET degrading enzyme genes from plastic‐contaminated soils using PCR‐based approaches.
Tannic acid (TA) has been shown in a previous study to expedite cutaneous wound healing in rats; however, the precise mechanism by which it operates remains poorly understood. This research evaluates the effects of TA on wound healing using both in vitro and in silico methods. In vitro, its influence on the inflammatory cytokine interleukin‐1β (IL‐1β) and the growth factor fibroblast growth factor (FGF) throughout the healing process were assessed. In silico molecular docking was employed to predict direct ligand–protein interactions and to provide a mechanistic insight into whether these proteins represent primary molecular targets or downstream effects. Parameters evaluated included cell viability and proliferation, scratch assays, and the activity of pro‐inflammatory cytokines in the lipopolysaccharide (LPS)‐stimulated RAW 264.7 macrophage cell line, together with growth factors in the NIH 3T3 fibroblast cell line; all were evaluated using enzyme‐linked immunosorbent assay (ELISA). The results indicate that TA significantly facilitates wound closure by promoting NIH 3T3 fibroblast cell proliferation, enhancing FGF expression, and suppressing IL‐1ß synthesis in both in vitro and in silico approaches. These findings suggest that TA may hold considerable promise for wound‐healing management.
Industrial batik wastewater containing naphthol dyes is persistent, dark in color, and contains complex organic compounds that have the potential to pollute aquatic environments. Conventional physicochemical treatment methods are often ineffective in degrading resistant aromatic compounds and involve high operational costs, thus necessitating alternative approaches that are more environmentally friendly and sustainable. This study evaluates the effectiveness of extracellular enzymes from Aspergillus sp. immobilized in a chitosan matrix, combined with a zeolite‐activated carbon adsorbent medium, in the decolorization of naphthol dye wastewater in a treatment system using a wastewater treatment plant. The enzymes were obtained from liquid cultures and subsequently immobilized using an encapsulation method in chitosan beads. The treatment process was conducted over 72 hours of incubation, with observations of the percentage of dye decolorization. The results show that the combination of immobilized enzymes and zeolite‐activated carbon provides a significant increase in decolorization efficiency compared to single treatments. The highest decolorization was obtained in black‐blue naphthol waste, with an efficiency of 86.94% after 72 hours of incubation. These findings indicate that a combination system of immobilized biocatalysts and adsorbents has the potential to become a more stable and effective alternative technology for batik waste treatment. The approach opens up opportunities for the development of more applicable and sustainable enzyme‐based waste treatment systems on an industrial scale.
Human coronaviruses (HCoVs) are responsible for mild common cold to severe pneumonia‐like symptoms in infected individuals. The first HCoV was HCoV‐229E, discovered in 1962 in the US, which causes moderate symptoms. Since then, HCoVs have evolved, leading to epidemics or the recent SARS‐CoV‐2 pandemic. The main objective of this study was to understand the modifications occurred and what led to the transition from mild to pandemic form. Of the viral proteins, the RNA‐dependent RNA polymerase (RdRp) plays a crucial role in viral evolution, mutation, pathogenesis and transmission; this protein was therefore analyzed using in silico tools. We observed that RdRp has shown many mutations during its transition from mild to severe forms in HCoVs, which may have affected its enzymatic activity. The RdRp of HoV‐229E and HCoV‐NL63 showed 171 mutations, while SARS‐CoV‐2 showed the presence of 312. SARS‐CoV‐2 also showed a reduction in hydrophobic amino acid compared to the other HCoVs, consequently contributing to faster replication. Although mutations in the RdRp subdomains were found, yet five conserved regions was also presence among all the seven HCoVs; the finger and thumb subdomains had one conserved region, while the palm subdomains had three. Therefore, it can be inferred that on one hand the mutations reported in RdRp appeared to be the major cause of increased virulence leading to sporadic disease outbreaks, while on the other hand the presence of five conserved regions might prove to be potential targets for the development of new antiviral drugs.
The application of Fusarium‐antagonistic endophytic bacteria with plant growth‐promoting traits offers an effective method to enhance the success of banana plantlet tissue culture while combating Fusarium wilt disease caused by Fusarium oxysporum f.sp. cubense Tropical Race 4 (FocTR4) (VCG 01213). This study evaluates the endophytic bacterium AP3311, isolated from healthy banana roots in direct association with orchid roots. AP3311 exhibited strong antagonism toward FocTR4, hyphal colonization ability, and multiple growth‐promoting activities, including phosphate solubilization, nitrogen fixation and auxin production. 16S rRNA gene sequencing identified that AP3311 belongs to the genus Bacillus, while metabarcoding analysis revealed that Bacillus species dominate the root microbiomes of both bananas and orchids. The bacterial supernatants stimulated root development and leaf growth in vitro. Metabolomic profiling indicated that antimicrobial compounds, together with plant growth regulators, promoted both root and shoot growth. Overall, the research demonstrates that Bacillus sp. AP3311 and its supernatants are valuable components in banana tissue culture, providing the dual benefits of plant growth promotion and effective disease control.
Phosphorus (P) is an essential element for oil palm growth and development. Acid phosphatase (Apase) and Pti‐interacting serine/threonine kinase are two enzymes which enzymes confirmed to be related to P‐uptake in oil palm, therefore their activities in oil palm treated with P‐limitation need to be quantified. Acid phosphatase is believed to be induced by P‐deprivation. Conversely, the Pto‐interaction (Pti) serine/threonine kinase activity is associated with abiotic stress. The aim of this study was to quantify of activities of two selected enzymes and phytohormone content in oil palm‐clones in the P‐limitation condition. Two oil palms genotypes were treated with three P dosages i.e. 0% (v/v), 4.67% (v/v), and 14.02% (v/v) represented as starvation, deficiency, and optimum condition, respectively. The activity of these two enzymes was quantified in mitochondria and cytoplasm using spectrophotometry and modified dot‐blot methods, while abscisic acid, indole acetic acid and gibberellic acid content was quantified using ultra performance liquid chromatography (UPLC). The result showed that the Apase activity in P‐optimum was higher than starvation and deficiency in leaf and root tissues in both genotypes, whereas Pti serine/threonine kinase activity was higher in prolific than non‐prolific genotypes in P‐deficient dosage. Furthermore, abscisic acid content was higher in prolific than non‐prolific genotypes in starvation and deficient, whereas other hormone contents were similar. Association study showed that prolific was separated with non‐prolific ones at different doses of P. Finally, the prolific genotype is more adaptable with P deficiency.
Leptospirosis presents with nonspecific clinical features and requires time‐consuming laboratory tests for gold standard diagnosis. This study aims to design and characterize the recombinant LipL32 from synthetic gene and assess its performance as an antigen for detecting leptospirosis. The antigen was developed by cloning the LipL32 gene conserved portion of Leptospira interrogans serovar Icterohaemorrhagiae strain Langkawi. The immunoinformatic was used to characterize the developed rLipL32. Western blot results using anti‐histidine revealed a band of rLipL32 protein at ~40 kDa. Subsequently, it was used to examine the IgM antibody on human sera by using ELISA. The IgM‐LipL32 ELISA was evaluated using 67‐positive and 25‐negative sera and compared with a commercial ELISA. With a cut‐off value of 0.8, it showed 85.7% sensitivity, 83.3% specificity, a 48% positive prediction value (PPV), and 97% negative prediction value (NPV), indicating modest performance compared to existing commercial kits. The rLipL32 is a potential antigen for detecting IgM using ELISA; however, for use in low incidence areas, a confirmation test is crucial.
Wetland soils in southern Iraq store large carbon pools and contain abundant phenolics that can modulate microbial decomposition. In this study, we investigate bacterial tyrosinases (TYRs), type III copper enzymes that oxidize mono‐ and diphenols, in Mesopotamian marsh soils using a combined metagenomic and biochemical approach. Degenerate primers targeting conserved CuA/CuB motifs recovered diverse partial tyr fragments affiliated with Proteobacteria and Actinobacteria. From one sample, we amplified the full melC operon from a Streptomyces lineage; expressed the tyrosinase in E. coli; and purified the enzyme (SZTYR). SZTYR displayed an alkaline pH optimum (~9); retained activity up to ~70 °C; and preferentially oxidized diphenols (e.g., L‐DOPA, dopamine) over monophenols. The enzyme also acted on phenolics relevant to peat/wetland matrices (e.g., caffeic, protocatechuic, p‐coumaric and gallic acids). The results document TYR genetic diversity in Iraqi marsh soils and establish the biochemical profile of an alkaline‐adapted Streptomyces tyrosinase. While ecosystem‐level impacts were not measured, our findings motivate field‐scale assessments of in situ TYR activity, phenolic pools and oxygen/pH dynamics to evaluate potential consequences for phenolic turnover and carbon cycling in aridifying wetlands.
Acute respiratory distress syndrome (ARDS) is a critical respiratory dysfunction triggered by intense in‐ flammation, microvascular damage, and increased epithelial and pulmonary vascular permeability. Human Wharton’s jelly mesenchymal stem cells (hWJMSCs) possess regenerative and anti‐inflammatory activities through the cytokines, chemokines, and growth factor secretion. The development of anti‐inflammatory agents derived from hWJMSCs has become one of the therapeutic solutions. Instead of direct cell use of hWJMSCs, their conditioned medium (CM) provides a cell‐free approach that delivers bioactive factors while minimizing the risks associated with stem cell transplantation. This study aims to measure the levels of vascular endothelial growth factor‐α (VEGF‐α), epidermal growth factor‐β (EGF‐β), interleukin‐10 (IL‐10), and hepatocyte growth factor (HGF) in CM‐hWJMSCs under non‐starvation and starvation conditions (24, 48 and 72 hours) using ELISA. The anti‐inflammatory potential of these factors was then analyzed through molecular docking with pro‐inflammatory cytokines. VEGF‐α, EGF‐β, IL‐10 and HGF levels were measured across all conditions. VEGF‐α ranged from 2590.37 to 3613.92 ng/mg protein; EGF‐β 347.01–504.43 ng/mg; IL‐10 302.59–729.28 pg/mg; and HGF 1747.20–2903.52 ng/mg. The molecular docking revealed strong binding between VEGF‐α, EGF‐β, IL‐10 and HGF with pro‐inflammatory cytokines, namely IL‐1β, IL‐6 and TNF‐α. VEGF‐α had the strongest bond with TNF‐α (–1162.3 kJ/mol), while EGF‐β formed the most hydrophobic and hydrogen interactions. The findings suggest that CM‐hWJMSCs, enriched with anti‐inflammatory and regenerative cytokines, may serve as a promising candidate for modulating the inflammatory pathways involved in ARDS pathogenesis. Longer starvation increased the secretion of VEGF‐α, EGF‐β, IL‐10 and HGF. These factors are known to promote angiogenesis, regulate immune responses, and protect against epithelial injury, thereby supporting the anti‐inflammatory and regenerative potential of hWJMSCs‐CM for ARDS therapy.
