Flavonoids are naturally occurring compounds found in fruits, vegetables, and other plant-based foods, and they are known for their health benefits, such as UV protection, antioxidant, anti-inflammatory, and antiproliferative properties. This study investigates whether flavonoids, such as quercetin and 2,3-dehydrosilybin, can act as photoactivatable carbon monoxide (CO)-releasing molecules under physiological conditions. CO has been recently recognized as an important signaling molecule. Here, we show that upon direct irradiation, CO was released from both flavonoids in PBS with chemical yields of up to 0.23 equiv, which increased to almost unity by sensitized photooxygenation involving singlet oxygen. Photoreleased CO reduced cellular toxicity caused by high flavonol concentrations, partially restored mitochondrial respiration, reduced superoxide production induced by rotenone and high flavonol levels, and influenced the G0/G1 and G2/M phases of the cell cycle, showing antiproliferative effects. The findings highlight the potential of quercetin and 2,3-dehydrosilybin as CO-photoreleasing molecules with chemopreventive and therapeutic implications in human pathology and suggest their possible roles in plant biology.

Department of Chemistry Faculty of Science Masaryk University Kamenice 5 Brno 62500 Czech Republic

Institute of Medical Biochemistry and Laboratory Diagnostics and 4th Department of Internal Medicine General University Hospital Prague and 1st Faculty of Medicine Charles University Na Bojišti 3 Prague 2 12108 Czech Republic

Laboratory of Biotransformation Institute of Microbiology of the Czech Academy of Sciences Vídeňská 1083 Prague 4 CZ 14200 Czech Republic

RECETOX Faculty of Science Masaryk University Kamenice 5 Brno 62500 Czech Republic

Zobrazit více v PubMed

Kumar S.; Pandey A. K.; Lu K. P.; Sastre J. Chemistry and biological activities of flavonoids: An overview. Sci. World J. 2013, 2013, 16275010.1155/2013/162750. PubMed DOI PMC

Li Q. L.; Wang Y. H.; Mai Y. X.; Li H. Y.; Wang Z.; Xu J. W.; He X. J. Health Benefits of the flavonoids from onion: Constituents and their pronounced antioxidant and anti-neuroinflammatory capacities. J. Agric. Food Chem. 2020, 68 (3), 799–807. 10.1021/acs.jafc.9b07418. PubMed DOI

Dinu M.; Abbate R.; Gensini G. F.; Casini A.; Sofi F. Vegetarian, vegan diets and multiple health outcomes: A systematic review with meta-analysis of observational studies. Crit. Rev. Food Sci. Nutr. 2017, 57 (17), 3640–3649. 10.1080/10408398.2016.1138447. PubMed DOI

Tapas A. R.; Sakarkar D. M.; Kakde R. B. Flavonoids as nutraceuticals: A review. Trop. J. Pharm. Res. 2008, 7 (3), 1089–1099. 10.4314/tjpr.v7i3.14693. DOI

Pradhan S. C.; Girish C. Hepatoprotective herbal drug, silymarin from experimental pharmacology to clinical medicine. Indian J. Med. Res. 2006, 124 (5), 491–504. PubMed

Křen V.; Valentová K. Silybin and its congeners: From traditional medicine to molecular effects. Nat. Prod. Rep. 2022, 39 (6), 1264–1281. 10.1039/D2NP00013J. PubMed DOI

Burda S.; Oleszek W. Antioxidant and antiradical activities of flavonoids. J. Agric. Food Chem. 2001, 49 (6), 2774–2779. 10.1021/jf001413m. PubMed DOI

Viktorová J.; Stránská-Zachariášová M.; Fenclová M.; Vítek L.; Hajšlová J.; Křen V.; Ruml T. Complex evaluation of antioxidant capacity of milk thistle dietary supplements. Antioxidants 2019, 8, 317.10.3390/antiox8080317. PubMed DOI PMC

Viktorová J.; Dobiášová S.; Řehořová K.; Biedermann D.; Káňová K.; Šeborová K.; Václavíková R.; Valentová K.; Ruml T.; Křen V.; et al. Antioxidant, anti-inflammatory, and multidrug resistance modulation activity of silychristin derivatives. Antioxidants 2019, 8 (8), 303.10.3390/antiox8080303. PubMed DOI PMC

Polyak S. J.; Morishima C.; Lohmann V.; Pal S.; Lee D. Y.; Liu Y.; Graf T. N.; Oberlies N. H. Identification of hepatoprotective flavonolignans from silymarin. Proc. Natl. Acad. Sci. U.S.A. 2010, 107 (13), 5995–5999. 10.1073/pnas.0914009107. PubMed DOI PMC

Wagoner J.; Negash A.; Kane O. J.; Martinez L. E.; Nahmias Y.; Bourne N.; Owen D. M.; Grove J.; Brimacombe C.; McKeating J. A.; et al. Multiple effects of silymarin on the hepatitis C virus lifecycle. Hepatology 2010, 51 (6), 1912–1921. 10.1002/hep.23587. PubMed DOI PMC

Rutter K.; Scherzer T. M.; Beinhardt S.; Kerschner H.; Stättermayer A. F.; Hofer H.; Popow-Kraupp T.; Steindl-Munda P.; Ferenci P. Intravenous silibinin as ‘rescue treatment’ for on-treatment non-responders to pegylated interferon/ribavirin combination therapy. Antivir. Ther. 2011, 16 (8), 1327–1333. 10.3851/IMP1942. PubMed DOI

Agarwal C.; Wadhwa R.; Deep G.; Biedermann D.; Gažák R.; Křen V.; Agarwal R.; Polyak S. J. Anti-cancer efficacy of silybin derivatives - A structure-activity relationship. PLoS One 2013, 8 (3), e6007410.1371/journal.pone.0060074. PubMed DOI PMC

Chambers C. S.; Holečková V.; Petrásková L.; Biedermann D.; Valentová K.; Buchta M.; Křen V. The silymarin composition··· and why does it matter???. Food Res. Int. 2017, 100, 339–353. 10.1016/j.foodres.2017.07.017. PubMed DOI

Flaig T. W.; Gustafson D. L.; Su L. J.; Zirrolli J. A.; Crighton F.; Harrison G. S.; Pierson A. S.; Agarwal R.; Glodé L. M. A phase I and pharmacokinetic study of silybin-phytosome in prostate cancer patients. Invest. New Drugs 2007, 25 (2), 139–146. 10.1007/s10637-006-9019-2. PubMed DOI

Darband S. G.; Kaviani M.; Yousefi B.; Sadighparvar S.; Pakdel F. G.; Attari J. A.; Mohebbi I.; Naderi S.; Majidinia M. Quercetin: A functional dietary flavonoid with potential chemo-preventive properties in colorectal cancer. J. Cell. Physiol. 2018, 233 (9), 6544–6560. 10.1002/jcp.26595. PubMed DOI

Asgharian P.; Tazekand A. P.; Hosseini K.; Forouhandeh H.; Ghasemnejad T.; Ranjbar M.; Hasan M.; Kumar M.; Beirami S. M.; Tarhriz V.; et al. Potential mechanisms of quercetin in cancer prevention: Focus on cellular and molecular targets. Cancer Cell Int. 2022, 22 (1), 257.10.1186/s12935-022-02677-w. PubMed DOI PMC

Chen L.; Cao H.; Huang Q.; Xiao J. B.; Teng H. Absorption, metabolism and bioavailability of flavonoids: A review. Crit. Rev. Food Sci. Nutr. 2022, 62 (28), 7730–7742. 10.1080/10408398.2021.1917508. PubMed DOI

Enjalbert F.; Rapior S.; Nouguier-Soulé J.; Guillon S.; Amouroux N.; Cabot C. Treatment of amatoxin poisoning: 20-year retrospective analysis. J. Toxicol. Clin. Toxicol. 2002, 40 (6), 715–757. 10.1081/CLT-120014646. PubMed DOI

Li H. W.; Li M. Z.; Fu J. X.; Ao H.; Wang W. H.; Wang X. T. Enhancement of oral bioavailability of quercetin by metabolic inhibitory nanosuspensions compared to conventional nanosuspensions. Drug Delivery 2021, 28 (1), 1226–1236. 10.1080/10717544.2021.1927244. PubMed DOI PMC

Barzaghi N.; Crema F.; Gatti G.; Pifferi G.; Perucca E. Pharmacokinetic studies on IdB 1016, a silybin-phosphatidylcholine complex, in healthy human subjects. Eur. J. Drug Metab. Pharmacokinet. 1990, 15 (4), 333–338. 10.1007/BF03190223. PubMed DOI

Riva A.; Ronchi M.; Petrangolini G.; Bosisio S.; Allegrini P. Improved oral absorption of quercetin from quercetin Phytosome®, a new delivery system based on food grade lecithin. Eur. J. Drug Metab. Pharmacokinet. 2019, 44 (2), 169–177. 10.1007/s13318-018-0517-3. PubMed DOI PMC

Russo M.; Štacko P.; Nachtigallová D.; Klán P. Mechanisms of orthogonal photodecarbonylation reactions of 3-hydroxyflavone-based acid-base forms. J. Org. Chem. 2020, 85 (5), 3527–3537. 10.1021/acs.joc.9b03248. PubMed DOI

Jézéquel Y. A.; Svěrák F.; Ramundo A.; Orel V.; Martínek M.; Klán P. Structure-photoreactivity relationship study of substituted 3-hydroxyflavones and 3-hydroxyflavothiones for improving carbon monoxide photorelease. J. Org. Chem. 2024, 89 (7), 4888–4903. 10.1021/acs.joc.4c00070. PubMed DOI PMC

Anderson S. N.; Richards J. M.; Esquer H. J.; Benninghoff A. D.; Arif A. M.; Berreau L. M. A structurally-tunable 3-hydroxyflavone motif for visible light-induced carbon monoxide-releasing molecules (CORMs). ChemistryOpen 2015, 4 (5), 590–594. 10.1002/open.201500167. PubMed DOI PMC

Popova M.; Soboleva T.; Arif A. M.; Berreau L. M. Properties of a flavonol-based photoCORM in aqueous buffered solutions: Influence of metal ions, surfactants and proteins on visible light-induced CO release. RSC Adv. 2017, 7 (36), 21997–22007. 10.1039/C7RA02653F. DOI

Sánchez-Carnerero E. M.; Russo M.; Jakob A.; Muchová L.; Vítek L.; Klán P. Effects of substituents on photophysical and CO-photoreleasing properties of 2,6-substituted-carboxy BODIPY derivatives. Chemistry 2021, 3 (1), 238–255. 10.3390/chemistry3010018. DOI

Ramundo A.; Janoš J.; Muchová L.; Šranková M.; Dostál J.; Kloz M.; Vítek L.; Slavíček P.; Klán P. Visible-light-activated carbon monoxide release from porphyrin-flavonol hybrids. J. Am. Chem. Soc. 2024, 146 (1), 920–929. 10.1021/jacs.3c11426. PubMed DOI PMC

Ramundo A.; Hurtová M.; Bozek I.; Osifová Z.; Russo M.; Ngoy B. P.; Křen V.; Klán P. Multimodal carbon monoxide photorelease from flavonoids. Org. Lett. 2024, 26 (3), 708–712. 10.1021/acs.orglett.3c04141. PubMed DOI PMC

Matsuura T.; Matsushima H.; Nakashima R. Photoinduced reactions—XXXVI: Photosensitized oxygenation of 3-hydroxyflavones as a nonenzymatic model for quercetinase. Tetrahedron 1970, 26 (2), 435.10.1016/S0040-4020(01)97840-8. DOI

Kim H. P.; Ryter S. W.; Choi A. M. K. CO as a cellular signaling molecule. Annu. Rev. Pharmacol. Toxicol. 2006, 46, 411–449. 10.1146/annurev.pharmtox.46.120604.141053. PubMed DOI

Ahmad S.; Hewett P. W.; Fujisawa T.; Sissaoui S.; Cai M.; Gueron G.; Al-Ani B.; Cudmore M.; Ahmed S. F.; Wong M. K. K.; et al. Carbon monoxide inhibits sprouting angiogenesis and vascular endothelial growth factor receptor-2 phosphorylation. Thromb. Haemost. 2015, 113 (2), 329–337. 10.1160/TH14-01-0002. PubMed DOI

Váňová K.; Šuk J.; Petr T.; Černý D.; Slanař O.; Vreman H. J.; Wong R. J.; Zima T.; Vítek L.; Muchová L. Protective effects of inhaled carbon monoxide in endotoxin-induced cholestasis is dependent on its kinetics. Biochimie 2014, 97, 173–180. 10.1016/j.biochi.2013.10.009. PubMed DOI

Ling K.; Men F.; Wang W. C.; Zhou Y. Q.; Zhang H. W.; Ye D. W. Carbon monoxide and its controlled release: Therapeutic application, detection, and development of carbon monoxide releasing molecules (CORMs). J. Med. Chem. 2018, 61 (7), 2611–2635. 10.1021/acs.jmedchem.6b01153. PubMed DOI

Oliveira S. R.; Queiroga C. S. F.; Vieira H. L. A. Mitochondria and carbon monoxide: cytoprotection and control of cell metabolism - A role for Ca2+?. J. Physiol. 2016, 594 (15), 4131–4138. 10.1113/JP270955. PubMed DOI PMC

Lo Iacono L.; Boczkowski J.; Zini R.; Salouage I.; Berdeaux A.; Motterlini R.; Morin D. A carbon monoxide-releasing molecule (CORM-3) uncouples mitochondrial respiration and modulates the production of reactive oxygen species. Free Radic. Biol. Med. 2011, 50 (11), 1556–1564. 10.1016/j.freeradbiomed.2011.02.033. PubMed DOI

Bilban M.; Haschemi A.; Wegiel B.; Chin B. Y.; Wagner O.; Otterbein L. E. Heme oxygenase and carbon monoxide initiate homeostatic signaling. J. Mol. Med. 2008, 86 (3), 267–279. 10.1007/s00109-007-0276-0. PubMed DOI

Indo H. P.; Yen H. C.; Nakanishi I.; Matsumoto K.; Tamura M.; Nagano Y.; Matsui H.; Gusev O.; Cornette R.; Okuda T.; et al. A mitochondrial superoxide theory for oxidative stress diseases and aging. J. Clin. Biochem. Nutr. 2015, 56 (1), 1–7. 10.3164/jcbn.14-42. PubMed DOI PMC

Cadenas S. Mitochondrial uncoupling, ROS generation and cardioprotection. Biochim. Biophys. Acta Bioenerg 2018, 1859 (9), 940–950. 10.1016/j.bbabio.2018.05.019. PubMed DOI

Wang M.; Liao W. Carbon monoxide as a signaling molecule in plants. Front. Plant Sci. 2016, 7, 572.10.3389/fpls.2016.00572. PubMed DOI PMC

Feng L.; Wei L.; Liu Y.; Ren J.; Liao W. Carbon monoxide/heme oxygenase system in plant: Roles in abiotic stress response and crosstalk with other signals molecules. Nitric Oxide 2023, 138, 51–63. 10.1016/j.niox.2023.06.005. PubMed DOI

Biedermann D.; Hurtová M.; Benada O.; Valentová K.; Biedermannová L.; Křen V. Continuous diastereomeric kinetic resolution-Silybins A and B. Catalysts 2021, 11 (9), 1106.10.3390/catal11091106. DOI

Gažák R.; Trouillas P.; Biedermann D.; Fuksová K.; Marhol P.; Kuzma M.; Křen V. Base-catalyzed oxidation of silybin and isosilybin into 2,3-dehydro derivatives. Tetrahedron Lett. 2013, 54 (4), 315–317. 10.1016/j.tetlet.2012.11.049. DOI

Biedermann D.; Buchta M.; Holečková V.; Sedlák D.; Valentová K.; Cvačka J.; Bednárová L.; Křenková A.; Kuzma M.; Škuta C.; et al. Silychristin: Skeletal alterations and biological activities. J. Nat. Prod. 2016, 79 (12), 3086–3092. 10.1021/acs.jnatprod.6b00750. PubMed DOI

Martínek M.; Filipová L.; Galeta J.; Ludvíková L.; Klán P. Photochemical formation of fibenzosilacyclohept-4-yne for Cu-free click chemistry with azides and 1,2,4,5-tetrazines. Org. Lett. 2016, 18 (19), 4892–4895. 10.1021/acs.orglett.6b02367. PubMed DOI

Vreman H. J.; Stevenson D. K. Heme oxygenase activity as measured by carbon-monoxide production. Anal. Biochem. 1988, 168 (1), 31–38. 10.1016/0003-2697(88)90006-1. PubMed DOI

Šranková M.; Dvořák A.; Martínek M.; Šebej P.; Klán P.; Vítek L.; Muchová L. Antiproliferative and cytotoxic activities of fluorescein-A diagnostic angiography dye. Int. J. Mol. Sci. 2022, 23 (3), 1504.10.3390/ijms23031504. PubMed DOI PMC

Vaňková K.; Marková I.; Jašprová J.; Dvořák A.; Subhanová I.; Zelenka J.; Novosádová I.; Rasl J.; Vomastek T.; Sobotka R.; et al. Chlorophyll-mediated changes in the redox status of pancreatic cancer cells are associated with its anticancer effects. Oxid. Med. Cell Longevity 2018, 2018, 406916710.1155/2018/4069167. PubMed DOI PMC

Yim S. K.; Yun C. H.; Ahn T.; Jung H. C.; Pan J. G. A continuous spectrophotometric assay for NADPH-cytochrome P450 reductase activity using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide. J. Biochem. Mol. Biol. 2005, 38 (3), 366–369. 10.5483/BMBRep.2005.38.3.366. PubMed DOI

Smolková K.; Dvořák A.; Zelenka J.; Vítek L.; Ježek P. Reductive carboxylation and 2-hydroxyglutarate formation by wild-type IDH2 in breast carcinoma cells. Int. J. Biochem. Cell Biol. 2015, 65, 125–133. 10.1016/j.biocel.2015.05.012. PubMed DOI

Dávila Y. A.; Sancho M. I.; Almandoz M. C.; Blanco S. E. Solvent effects on the dissociation constants of hydroxyflavones in organic–water mixtures. Determination of the thermodynamic pKa values by UV–visible spectroscopy and DFT calculations. J. Chem. Eng. Data 2013, 58 (6), 1706–1716. 10.1021/je400153r. DOI

Herrero-Martínez J. M.; Sanmartin M.; Rosés M.; Bosch E.; Ráfols C. Determination of dissociation constants of flavonoids by capillary electrophoresis. Electrophoresis 2005, 26 (10), 1886–1895. 10.1002/elps.200410258. PubMed DOI

van Wenum E.; Jurczakowski R.; Litwinienko G. Media effects on the mechanism of antioxidant action of silybin and 2,3-dehydrosilybin: Role of the enol group. J. Org. Chem. 2013, 78 (18), 9102–9112. 10.1021/jo401296k. PubMed DOI

Li Y. C.; Zhang Y.; Wang J. X.; Xia D. D.; Zhuo M. M.; Zhu L.; Li D.; Ni S. F.; Zhu Y. P.; Zhang W. D. Visible-light-mediated three-Component strategy for the synthesis of isoxazolines and isoxazoles. Org. Lett. 2024, 26 (15), 3130–3134. 10.1021/acs.orglett.4c00671. PubMed DOI

Studer S. L.; Brewer W. E.; Martinez M. L.; Chou P. T. Time-resolved study of the photooxygenation of 3-hydroxyflavone. J. Am. Chem. Soc. 1989, 111 (19), 7643–7644. 10.1021/ja00201a071. DOI

Russo M.; Orel V.; Štacko P.; Šranková M.; Muchová L.; Vítek L.; Klán P. Structure-photoreactivity relationship of 3-hydroxyflavone-based CO-releasing molecules. J. Org. Chem. 2022, 87 (7), 4750–4763. 10.1021/acs.joc.2c00032. PubMed DOI

Zenkevich I. G.; Eshchenko A. Y.; Makarova S. V.; Vitenberg A. G.; Dobryakov Y. G.; Utsal V. A. Identification of the products of oxidation of quercetin by air oxygen at ambient temperature. Molecules 2007, 12 (3), 654–672. 10.3390/12030654. PubMed DOI PMC

Anderson S. N.; Dederich C. T.; Borowski T.; Berreau L. M. Thermal CO release reactivity of a π-extended flavonol anion. Org. Lett. 2024, 26, 10253.10.1021/acs.orglett.4c03651. PubMed DOI

Matsuura T.; Matsushima H. Photoinduced reactions—XXII: Photooxidative cyclization of 3-methoxyflavones. Tetrahedron 1968, 24 (22), 6615–6618. 10.1016/S0040-4020(01)96876-0. DOI

Schwartz B. J.; Peteanu L. A.; Harris C. B. Direct observation of fast proton-transfer - Femtosecond photophysics of 3-hydroxyflavone. J. Phys. Chem. 1992, 96 (9), 3591–3598. 10.1021/j100188a009. DOI

Han Y.; Jia Y.; Wang X.; Chen Z.; Jin P.; Jia M.; Pan H.; Sun Z.; Chen J. Ultrafast excited state dynamics of two non-emissive flavonoids that undergo excited state intramolecular proton transfer in solution. Chem. Phys. Lett. 2023, 811, 14018910.1016/j.cplett.2022.140189. DOI

Queiroga C. S. F.; Almeida A. S.; Vieira H. L. A. Carbon monoxide targeting mitochondria. Biochem. Res. Int. 2012, 2012, 74984510.1155/2012/749845. PubMed DOI PMC

Averilla J. N.; Oh J.; Kim J. S. Carbon monoxide partially mediates protective effect of resveratrol against UVB-Induced oxidative stress in human keratinocytes. Antioxidants 2019, 8 (10), 432.10.3390/antiox8100432. PubMed DOI PMC

Ferreyra M. L. F.; Serra P.; Casati P. Recent advances on the roles of flavonoids as plant protective molecules after UV and high light exposure. Physiol. Plant. 2021, 173 (3), 736–749. 10.1111/ppl.13543. PubMed DOI

Machlin L. J.; Bendich A. Free-Radical Tissue-Damage - Protective Role of Antioxidant Nutrients. Faseb J. 1987, 1 (6), 441–445. 10.1096/fasebj.1.6.3315807. PubMed DOI

Wätjen W.; Michels G.; Steffan B. R.; Niering P.; Chovolou Y.; Kampkötter A.; Tran-Thi Q. H.; Proksch P.; Kahl R. Low concentrations of flavonoids are protective in rat H4IIE cells whereas high concentrations cause DNA damage and apoptosis. J. Nutr. 2005, 135 (3), 525–531. 10.1093/jn/135.3.525. PubMed DOI

Boczkowski J.; Poderoso J. J.; Motterlini R. CO-metal interaction: Vital signaling from a lethal gas. Trends Biochem. Sci. 2006, 31 (11), 614–621. 10.1016/j.tibs.2006.09.001. PubMed DOI

Almeida A. S.; Queiroga C. S. F.; Sousa M. F. Q.; Alves P. M.; Vieira H. L. A. Carbon monoxide modulates apoptosis by reinforcing oxidative metabolism in astrocytes. J. Biol. Chem. 2012, 287 (14), 10761–10770. 10.1074/jbc.M111.306738. PubMed DOI PMC

Balaban R. S.; Nemoto S.; Finkel T. Mitochondria, oxidants, and aging. Cell 2005, 120 (4), 483–495. 10.1016/j.cell.2005.02.001. PubMed DOI

Song R. P.; Mahidhara R. S.; Zhou Z. H.; Hoffman R. A.; Seol D. W.; Flavell R. A.; Billiar T. R.; Otterbein L. E.; Choi A. M. K. Carbon monoxide inhibits T lymphocyte proliferation via caspase-dependent pathway. J. Immunol. 2004, 172 (2), 1220–1226. 10.4049/jimmunol.172.2.1220. PubMed DOI

Vítek L.; Gbelcová H.; Muchová L.; Váňová K.; Zelenka J.; Koníčková R.; Šuk J.; Zadinová M.; Knejzlík Z.; Ahmad S.; et al. Antiproliferative effects of carbon monoxide on pancreatic cancer. Dig. Liver Dis. 2014, 46 (4), 369–375. 10.1016/j.dld.2013.12.007. PubMed DOI