Antioxidant and Anti-Inflammatory Activities of Silene dichotoma Ehrh. subsp. dichotoma: Phytochemical Characterization and Metabolomic Insights into Oxidative Stress-Related Responses in HaCaT Cells
- 1Department of Pharmacognosy, Faculty of Pharmacy, Hacettepe University, Sihhiye, Ankara 06100, Türkiye
- 2Department of Analytical Chemistry, Faculty of Pharmacy, Hacettepe University, Sihhiye, Ankara 06100, Türkiye
Abstract
Background/Objectives: Silene dichotoma Ehrh. subsp. dichotoma is a relatively understudied member of a genus known for diverse secondary metabolites. This study integrated phytochemical characterization, biological activity assessment, and LC–MS-based cellular metabolomics to investigate its antioxidant, NO-inhibitory, and cytoprotective potential. Methods: An 80% ethanol extract of the aerial parts was partitioned into n-hexane, ethyl acetate, n-butanol, and aqueous fractions and evaluated for DPPH and superoxide radical-scavenging activities and total phenolic and flavonoid contents. The most active fraction was subjected to bioactivity-guided isolation, and fractions were assessed for cytotoxicity, LPS-induced nitric oxide inhibition, H2O2-induced cytoprotection, and untargeted LC–MS metabolomic profiling. Results: The ethyl acetate fraction exhibited the highest phenolic (51.84 mg GAE/g) and flavonoid (176.56 mg QE/g) contents and the strongest antioxidant activity, yielding isorhamnetin, isorhamnetin-3-O-β-D-galactopyranoside, and quercetin, reported for the first time from this species. None of the extract or fractions showed marked cytotoxicity toward RAW 264.7 macrophages or HaCaT keratinocytes. The ethyl acetate fraction most strongly inhibited LPS-induced nitric oxide production (IC50 = 118.34 µg/mL). In H2O2-challenged HaCaT cells, the n-hexane fraction modestly increased viability to 67.77% at 200 µg/mL; however, no statistically significant overall cytoprotective effect was demonstrated. Untargeted LC–MS profiling yielded 99 putatively annotated metabolites. After averaging analytical duplicates within biological replicates, 38 of 75 metabolites eligible for the overall seven-group analysis showed an FDR-adjusted difference. Exploratory pairwise comparisons suggested treatment-associated changes in energy-related, redox-associated, nucleotide, amino-acid, and lipid-associated features. Conclusions: These findings identify this species as a source of flavonoids with antioxidant and nitric oxide-inhibitory potential while demonstrating the value of metabolomics in characterizing cellular responses beyond viability endpoints.
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BibTeX
@article{bilal2026antioxidant,
title = {{Antioxidant and Anti-Inflammatory Activities of Silene dichotoma Ehrh. subsp. dichotoma: Phytochemical Characterization and Metabolomic Insights into Oxidative Stress-Related Responses in HaCaT Cells}},
author = {Melek Nur Bilal and Ozan Kaplan and Mustafa Çelebier and Vahap Murat Kutluay},
journal = {Metabolites},
year = {2026},
month = oct,
doi = {10.3390/metabo16100765},
url = {https://www.mdpi.com/2218-1989/16/10/765},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Antioxidant and Anti-Inflammatory Activities of Silene dichotoma Ehrh. subsp. dichotoma: Phytochemical Characterization and Metabolomic Insights into Oxidative Stress-Related Responses in HaCaT Cells
AU - Melek Nur Bilal
AU - Ozan Kaplan
AU - Mustafa Çelebier
AU - Vahap Murat Kutluay
PY - 2026
DA - 2026/10/01
JO - Metabolites
DO - 10.3390/metabo16100765
UR - https://www.mdpi.com/2218-1989/16/10/765
AB - Background/Objectives: Silene dichotoma Ehrh. subsp. dichotoma is a relatively understudied member of a genus known for diverse secondary metabolites. This study integrated phytochemical characterization, biological activity assessment, and LC–MS-based cellular metabolomics to investigate its antioxidant, NO-inhibitory, and cytoprotective potential. Methods: An 80% ethanol extract of the aerial parts was partitioned into n-hexane, ethyl acetate, n-butanol, and aqueous fractions and evaluated for DPPH and superoxide radical-scavenging activities and total phenolic and flavonoid contents. The most active fraction was subjected to bioactivity-guided isolation, and fractions were assessed for cytotoxicity, LPS-induced nitric oxide inhibition, H2O2-induced cytoprotection, and untargeted LC–MS metabolomic profiling. Results: The ethyl acetate fraction exhibited the highest phenolic (51.84 mg GAE/g) and flavonoid (176.56 mg QE/g) contents and the strongest antioxidant activity, yielding isorhamnetin, isorhamnetin-3-O-β-D-galactopyranoside, and quercetin, reported for the first time from this species. None of the extract or fractions showed marked cytotoxicity toward RAW 264.7 macrophages or HaCaT keratinocytes. The ethyl acetate fraction most strongly inhibited LPS-induced nitric oxide production (IC50 = 118.34 µg/mL). In H2O2-challenged HaCaT cells, the n-hexane fraction modestly increased viability to 67.77% at 200 µg/mL; however, no statistically significant overall cytoprotective effect was demonstrated. Untargeted LC–MS profiling yielded 99 putatively annotated metabolites. After averaging analytical duplicates within biological replicates, 38 of 75 metabolites eligible for the overall seven-group analysis showed an FDR-adjusted difference. Exploratory pairwise comparisons suggested treatment-associated changes in energy-related, redox-associated, nucleotide, amino-acid, and lipid-associated features. Conclusions: These findings identify this species as a source of flavonoids with antioxidant and nitric oxide-inhibitory potential while demonstrating the value of metabolomics in characterizing cellular responses beyond viability endpoints.
SN - 2218-1989
ER -