{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,21]],"date-time":"2026-09-21T06:02:07Z","timestamp":1789970527949,"version":"4.0.1"},"reference-count":42,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2026,9,21]],"date-time":"2026-09-21T00:00:00Z","timestamp":1789948800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Microbiol."],"abstract":"<jats:sec>\n                    <jats:title>Introduction<\/jats:title>\n                    <jats:p>\n                      <jats:italic>Clerodendrum cyrtophyllum<\/jats:italic>\n                      Turcz., a traditional Chinese medicinal plant, exhibits broad bioactivity. Nevertheless, the flavonoid constituents conferring anti-\n                      <jats:italic>Staphylococcus aureus<\/jats:italic>\n                      (\n                      <jats:italic>S. aureus<\/jats:italic>\n                      ) activity and their mechanistic basis remain to be elucidated.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>\n                      Total flavonoid extraction from\n                      <jats:italic>Clerodendrum cyrtophyllum<\/jats:italic>\n                      Turcz. (TFC) was first evaluated by single-factor experiments to screen key variables and subsequently optimized by a Box\u2013Behnken design (BBD) within response surface methodology (RSM). followed by purification with D101 macroporous resin. Liquid chromatography\u2013tandem mass spectrometry (LC\u2013MS\/MS) characterized the chemical profile. Antibacterial activity against\n                      <jats:italic>S. aureus<\/jats:italic>\n                      was assessed via inhibition zones, minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), and growth curves. Mechanisms were explored by evaluating cell surface hydrophobicity (CSH), extracellular alkaline phosphatase (AKP), leakage of nucleic acids\/proteins, membrane potential, ATP levels, reactive oxygen species (ROS) content, and morphological alterations.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      The optimal extraction conditions for TFC were determined as 70% ethanol as the solvent, an extraction time of 102 min, a temperature of 64 \u00b0C, and a solid\u2013liquid ratio of 1:25 (g: mL), resulting in a yield of 6.48%\u202f\u00b1\u202f0.58. After purification, the purity of TFC was increased to 76%, and preliminary analysis identified 85 flavonoid constituents. TFC exhibited inhibition zone diameters of 13\u202f\u00b1\u202f0.45\u202fmm, 16\u202f\u00b1\u202f0.38\u202fmm, and 20\u202f\u00b1\u202f0.45\u202fmm at concentrations of 50, 100, and 150\u202fmg\/mL, respectively, with MIC and MBC values of 0.781\u202fmg\/mL and 1.562\u202fmg\/mL. Notably, TFC effectively inhibited the growth of\n                      <jats:italic>S. aureus<\/jats:italic>\n                      within 24\u202fh. Mechanistic investigations revealed that TFC treatment increased\n                      <jats:italic>S. aureus<\/jats:italic>\n                      CSH and decreased the membrane potential, leading to the leakage of intracellular nucleic acids and proteins, indicating damage to\n                      <jats:italic>S. aureus<\/jats:italic>\n                      cells, which was further confirmed by SEM. Furthermore, TFC induced significant intracellular ATP depletion, decreased intracellular nucleic acids and protein contents, and triggered excessive ROS production.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusion<\/jats:title>\n                    <jats:p>\n                      This study optimized the extraction conditions of TFC and successfully obtained TFC rich in multiple flavonoid constituents. TFC exerts potent inhibitory effects on\n                      <jats:italic>S. aureus<\/jats:italic>\n                      by disrupting the integrity of cell walls and membranes, depleting intracellular energy reserves, decreasing the contents of intracellular nucleic acids and proteins, and inducing intracellular oxidative stress.\n                    <\/jats:p>\n                  <\/jats:sec>","DOI":"10.3389\/fmicb.2026.1961127","type":"journal-article","created":{"date-parts":[[2026,9,21]],"date-time":"2026-09-21T05:35:08Z","timestamp":1789968908000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":0,"title":["Optimization of extraction conditions for total flavonoids from Clerodendrum cyrtophyllum Turcz. and its antibacterial activity and mechanism against Staphylococcus aureus"],"prefix":"10.3389","volume":"17","author":[{"given":"Yue","family":"Hu","sequence":"first","affiliation":[{"name":"School of Ethnic Medicine, Guizhou Minzu University","place":["Guiyang, Guizhou, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wenwen","family":"Luo","sequence":"additional","affiliation":[{"name":"School of Ethnic Medicine, Guizhou Minzu University","place":["Guiyang, Guizhou, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chengjian","family":"Tan","sequence":"additional","affiliation":[{"name":"School of Ethnic Medicine, Guizhou Minzu University","place":["Guiyang, Guizhou, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Guoshun","family":"Pei","sequence":"additional","affiliation":[{"name":"School of Ethnic Medicine, Guizhou Minzu University","place":["Guiyang, Guizhou, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2026,9,21]]},"reference":[{"key":"ref1","doi-asserted-by":"publisher","first-page":"88","DOI":"10.1093\/ajcp\/71.1.88","article-title":"In-vitro methods for determining minimal lethal concentrations of antimicrobial agents","volume":"71","author":"Barry","year":"1979","journal-title":"Am. J. Clin. Pathol."},{"key":"ref2","doi-asserted-by":"publisher","first-page":"209","DOI":"10.1080\/20477724.2023.2285187","article-title":"Staphylococcus aureus epidemiology, pathophysiology, clinical manifestations and application of nano-therapeutics as a promising approach to combat methicillin resistant Staphylococcus aureus","volume":"118","author":"Bashabsheh","year":"2024","journal-title":"Pathog. Glob. Health"},{"key":"ref3","doi-asserted-by":"publisher","first-page":"547","DOI":"10.1080\/21505594.2021.1878688","article-title":"Pathogenicity and virulence of Staphylococcus aureus","volume":"12","author":"Cheung","year":"2021","journal-title":"Virulence"},{"key":"ref4","doi-asserted-by":"publisher","first-page":"10303","DOI":"10.2147\/dddt.S555032","article-title":"Momordicae semen: a review of Phytochemistry, pharmacology, toxicology, herbal processing, clinical applications, and Q-markers prediction","volume":"19","author":"Deng","year":"2025","journal-title":"Drug Des. Devel. Ther."},{"key":"ref5","doi-asserted-by":"publisher","first-page":"551","DOI":"10.3390\/antibiotics11050551","article-title":"Multifractal analysis to determine the effect of surface topography on the distribution, density, dispersion and clustering of differently organised coccal-shaped bacteria","volume":"11","author":"Evans","year":"2022","journal-title":"Antibiotics"},{"key":"ref6","doi-asserted-by":"publisher","first-page":"1163","DOI":"10.3390\/antibiotics14111163","article-title":"Antimicrobial and anti-infective activity of natural products: unveiling mechanisms, synergies, and translational applications","volume":"14","author":"Fernandes","year":"2025","journal-title":"Antibiotics"},{"key":"ref7","doi-asserted-by":"publisher","first-page":"2669","DOI":"10.3390\/molecules27092669","article-title":"Cannabis sativa CBD extract shows promising antibacterial activity against Salmonella typhimurium and S. Newington","volume":"27","author":"Gildea","year":"2022","journal-title":"Molecules"},{"key":"ref8","doi-asserted-by":"publisher","first-page":"1208617","DOI":"10.1155\/2020\/1208617","article-title":"Optimization of extraction of shikonin using ultrasound-assisted response surface methodology and antibacterial studies","author":"Huang","year":"2020","journal-title":"Evid. Based Complement. Alternat. Med."},{"key":"ref9","doi-asserted-by":"publisher","first-page":"190","DOI":"10.3969\/j.issn.1672-2779.2025.10.060","article-title":"Modern research progress of textual research of medicinal herb on Clerodendrum cyrtophyllum Turcz. and the clinical application of phenylpropanoid components","volume":"23","author":"Huang","year":"2025","journal-title":"Chin. Med. Modern Dist. Educ. China"},{"key":"ref10","doi-asserted-by":"publisher","first-page":"220","DOI":"10.3109\/10520299509108199","article-title":"DAPI: a DNA-specific fluorescent probe","volume":"70","author":"Kapuscinski","year":"1995","journal-title":"Biotech. Histochem."},{"key":"ref11","doi-asserted-by":"publisher","first-page":"1138","DOI":"10.3390\/antibiotics14111138","article-title":"Anti-infective-associated AKI: a narrative review of the epidemiology, mechanisms, risk factors, biomarkers, clinical course, monitoring, prevention, and therapeutic strategies","volume":"14","author":"Karimzadeh","year":"2025","journal-title":"Antibiotics (Basel)"},{"key":"ref12","doi-asserted-by":"publisher","first-page":"34056","DOI":"10.1038\/s41598-025-14396-z","article-title":"Murraya koenigii methanolic extract inhibits bacterial growth and biofilm of Staphylococcus aureus and Enterococcus faecalis","volume":"15","author":"Kashyap","year":"2025","journal-title":"Sci. Rep."},{"key":"ref13","doi-asserted-by":"publisher","first-page":"32","DOI":"10.1038\/s44303-024-00036-0","article-title":"The role of [18F]FDG-PET\/CT in Staphylococcus aureus bacteremia: a clinical perspective","volume":"2","author":"Kouijzer","year":"2024","journal-title":"npj Imaging"},{"key":"ref14","doi-asserted-by":"publisher","first-page":"247","DOI":"10.1007\/s00284-025-04236-8","article-title":"Inhibiting Staphylococcus aureus virulence factors: advances in traditional Chinese medicines and active compounds","volume":"82","author":"Liang","year":"2025","journal-title":"Curr. Microbiol."},{"key":"ref15","doi-asserted-by":"publisher","first-page":"866","DOI":"10.3390\/antibiotics13090866","article-title":"Efficacy and safety of antibiotics in the treatment of methicillin-resistant Staphylococcus aureus (MRSA) infections: a systematic review and network meta-analysis","volume":"13","author":"Liu","year":"2024","journal-title":"Antibiotics (Basel)"},{"key":"ref16","doi-asserted-by":"publisher","first-page":"403","DOI":"10.3390\/foods11030403","article-title":"Antibacterial effect of oregano essential oil against Vibrio vulnificus and its mechanism","volume":"11","author":"Luo","year":"2022","journal-title":"Foods"},{"key":"ref17","doi-asserted-by":"publisher","first-page":"e202200076","DOI":"10.1002\/ijch.202200076","article-title":"Chemoproteomic approaches for unraveling prokaryotic biology","volume":"63","author":"Malarney","year":"2023","journal-title":"Isr. J. Chem."},{"key":"ref18","doi-asserted-by":"publisher","first-page":"5157","DOI":"10.1007\/s00210-025-04765-w","article-title":"Integrative network pharmacology and experimental validation of multi-target synergy against multidrug-resistant Klebsiella pneumoniae via PI3K\/AKT-MAPK pathway disruption","volume":"399","author":"Man","year":"2025","journal-title":"Naunyn Schmiedeberg's Arch. Pharmacol"},{"key":"ref19","doi-asserted-by":"publisher","first-page":"11803","DOI":"10.3390\/ijms262411803","article-title":"The pathogenicity mechanisms of Staphylococcus aureus","volume":"26","author":"Mlynarczyk-Bonikowska","year":"2025","journal-title":"Int. J. Mol. Sci."},{"key":"ref20","first-page":"e02456-20","article-title":"Bacterial cell wall quality control during environmental stress","volume-title":"mBio","author":"Mueller","year":"2020"},{"key":"ref21","doi-asserted-by":"publisher","first-page":"192","DOI":"10.3390\/antiox9030192","article-title":"Anti-inflammatory and antioxidant properties of the ethanol extract of Clerodendrum cyrtophyllum Turcz in copper sulfate-induced inflammation in zebrafish","volume":"9","author":"Nguyen","year":"2020","journal-title":"Antioxidants"},{"key":"ref22","doi-asserted-by":"publisher","first-page":"27404","DOI":"10.1038\/s41598-025-12616-0","article-title":"Optimisation of goose oil extraction process by response surface methodology","volume":"15","author":"Ning","year":"2025","journal-title":"Sci. Rep."},{"key":"ref23","doi-asserted-by":"publisher","first-page":"e2506007","DOI":"10.4014\/jmb.2506.06007","article-title":"Antibacterial and antibiofilm activity of Clerodendrum cyrtophyllum Turcz. 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Ethnopharmacol."},{"key":"ref25","volume-title":"Studies on Miao Folk Usage and Chemical Constituents of Clerodendrum cyrtophyllum in Qiandongnan [Master's thesis]","author":"Ran","year":"2017"},{"key":"ref26","doi-asserted-by":"publisher","first-page":"289","DOI":"10.1016\/j.jtemb.2014.08.001","article-title":"Diphenyl diselenide (PhSe)\u2082 inhibits biofilm formation by Candida albicans, increasing both ROS production and membrane permeability","volume":"29","author":"Rosseti","year":"2015","journal-title":"J. Trace Elem. Med. Biol."},{"key":"ref27","doi-asserted-by":"publisher","first-page":"128171","DOI":"10.1016\/j.foodchem.2020.128171","article-title":"In vitro and in situ abrogation of biofilm formation in E. coli by vitamin C through ROS generation, disruption of quorum sensing and exopolysaccharide production","volume":"341","author":"Shivaprasad","year":"2021","journal-title":"Food Chem."},{"key":"ref28","doi-asserted-by":"publisher","first-page":"92","DOI":"10.3390\/foods11010092","article-title":"Antibacterial mechanism of 3-carene against the meat spoilage bacterium Pseudomonas lundensis and its application in pork during refrigerated storage","volume":"11","author":"Tang","year":"2021","journal-title":"Foods"},{"key":"ref29","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1155\/2018\/3508506","article-title":"Extraction process, component analysis, and in vitro antioxidant, antibacterial, and anti-inflammatory activities of total flavonoid extracts from Abutilon theophrasti medic. Leaves","volume":"2018","author":"Tian","year":"2018","journal-title":"Mediat. Inflamm."},{"key":"ref30","doi-asserted-by":"publisher","first-page":"941","DOI":"10.1007\/s004240000390","article-title":"Assessment of mitochondrial polarization status in living cells based on analysis of the spatial heterogeneity of rhodamine 123 fluorescence staining","volume":"440","author":"Toescu","year":"2000","journal-title":"Pfl\u00fcgers Arch."},{"key":"ref31","doi-asserted-by":"publisher","first-page":"798","DOI":"10.1001\/jama.2025.4288","article-title":"Management of Staphylococcus aureus bacteremia: a review","volume":"334","author":"Tong","year":"2025","journal-title":"JAMA"},{"key":"ref32","doi-asserted-by":"publisher","first-page":"142","DOI":"10.4103\/2225-4110.114898","article-title":"Therapeutic potential of traditional Chinese medicine on inflammatory diseases","volume":"3","author":"Tsai","year":"2013","journal-title":"J. Tradit. Complement. Med."},{"key":"ref33","doi-asserted-by":"publisher","first-page":"2954","DOI":"10.3390\/foods10122954","article-title":"Control of foodborne Staphylococcus aureus by shikonin, a natural extract","volume":"10","author":"Wan","year":"2021","journal-title":"Foods"},{"key":"ref34","doi-asserted-by":"publisher","first-page":"668","DOI":"10.1186\/s12866-025-04354-0","article-title":"Antimicrobial activity and possible mechanisms of juglone against Escherichia coli, Staphylococcus aureus, and Salmonella pullorum","volume":"25","author":"Wang","year":"2025","journal-title":"BMC Microbiol."},{"key":"ref35","doi-asserted-by":"publisher","first-page":"115553","DOI":"10.1016\/j.ab.2024.115553","article-title":"Microwave-assisted and methanol\/acetic acid-free method for rapid staining of proteins in SDS-PAGE gels","volume":"691","author":"Wang","year":"2024","journal-title":"Anal. Biochem."},{"key":"ref36","doi-asserted-by":"publisher","first-page":"3589","DOI":"10.3390\/foods14213589","article-title":"Integrated phenotypic and transcriptomic analyses unveil the antibacterial mechanism of punicalagin against methicillin-resistant Staphylococcus aureus (MRSA)","volume":"14","author":"Wang","year":"2025","journal-title":"Foods"},{"key":"ref37","doi-asserted-by":"publisher","first-page":"116940","DOI":"10.1016\/j.ejmech.2024.116940","article-title":"Oleanolic acid derivatives against drug-resistant bacteria and fungi by multi-targets to avoid drug resistance","volume":"280","author":"Wei","year":"2024","journal-title":"Eur. J. Med. Chem."},{"key":"ref38","doi-asserted-by":"publisher","first-page":"e1231","DOI":"10.1002\/prp2.1231","article-title":"Determination of a vancomycin nephrotoxicity threshold and assessment of target attainment in hematology patients","volume":"12","author":"Wong","year":"2024","journal-title":"Pharmacol. Res. Perspect."},{"key":"ref39","doi-asserted-by":"publisher","first-page":"111268","DOI":"10.1016\/j.foodres.2022.111268","article-title":"A review of ultrasound-assisted extraction for plant bioactive compounds: Phenolics, flavonoids, thymols, saponins and proteins","volume":"157","author":"Yusoff","year":"2022","journal-title":"Food Res. Int."},{"key":"ref40","doi-asserted-by":"publisher","first-page":"116645","DOI":"10.1016\/j.foodres.2025.116645","article-title":"Foodborne outbreak of enterotoxigenic Staphylococcus aureus at a school in Hebei, China","volume":"214","author":"Zheng","year":"2025","journal-title":"Food Res. Int."},{"key":"ref41","doi-asserted-by":"publisher","first-page":"e202301308","DOI":"10.1002\/cbdv.202301308","article-title":"Optimization of flavonoid extraction from Eucommia ulmoides pollen using response surface methodology and its biological activities","volume":"21","author":"Zhu","year":"2024","journal-title":"Chem. Biodivers."},{"key":"ref42","doi-asserted-by":"publisher","first-page":"11226","DOI":"10.3390\/ijms241311226","article-title":"Eukaryotic cell membranes: structure, composition, research methods and computational modelling","volume":"24","author":"Zhukov","year":"2023","journal-title":"Int. J. Mol. 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