{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,19]],"date-time":"2026-10-19T09:08:28Z","timestamp":1792400908288,"version":"4.4.0"},"reference-count":35,"publisher":"MDPI AG","issue":"1","license":[{"start":{"date-parts":[[2022,12,29]],"date-time":"2022-12-29T00:00:00Z","timestamp":1672272000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"National Research Foundation of Korea","award":["2016R1A2B1007540"],"award-info":[{"award-number":["2016R1A2B1007540"]}]},{"name":"National Research Foundation of Korea","award":["2019R1A2C1089617"],"award-info":[{"award-number":["2019R1A2C1089617"]}]},{"name":"National Research Foundation of Korea","award":["2021R1F1A1051466"],"award-info":[{"award-number":["2021R1F1A1051466"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>Porphyrin compounds are widely distributed in various natural products and biological systems. In this study, effects of porphyrin-related compounds including zinc protoporphyrin (ZnPP), protoporphyrin IX (PPIX), cyanocobalamin (CBL), hemin, and zinc phthalocyanine (ZnPC) were analyzed on color response of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) tetrazolium-based assay, a commonly-used method for analyzing cell viability. Color responses of MTT formazan formed in cells treated with ZnPP, PPIX, or ZnPC were significantly reduced even at submicromolar concentrations without affecting cell viability, whereas hemin and CBL did not. ZnPP, PPIX, and ZnPC rapidly induced degradation of MTT formazan already-produced by cells when exposed to light, but not under a dark condition. Photosensitizing properties of the three compounds were also verified through extensive generation of reactive oxygen species under light. The porphyrins did not affect the stability of water-soluble formazans including XTT, WST-1, WST-8, and MTS formazans. Several factors including different light sources and antioxidants modulated the degradation process of MTT formazan by the porphyrins. The results suggest that certain porphyrin compounds could cause a severe artifact in the MTT assay through rapid degradation of formazan dye due to their photosensitizing property, which needs to be considered carefully in the related assays.<\/jats:p>","DOI":"10.3390\/ijms24010562","type":"journal-article","created":{"date-parts":[[2022,12,29]],"date-time":"2022-12-29T02:56:50Z","timestamp":1672282610000},"page":"562","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":13,"title":["Interfering with Color Response by Porphyrin-Related Compounds in the MTT Tetrazolium-Based Colorimetric Assay"],"prefix":"10.3390","volume":"24","author":[{"given":"Bo Hee","family":"Choi","sequence":"first","affiliation":[{"name":"Division of Applied Food System, College of Natural Science, Seoul Women\u2019s University, 621 Hwarangro, Nowon-Gu, Seoul 01797, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mi-Ri","family":"Kim","sequence":"additional","affiliation":[{"name":"Division of Applied Food System, College of Natural Science, Seoul Women\u2019s University, 621 Hwarangro, Nowon-Gu, Seoul 01797, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yu Na","family":"Jung","sequence":"additional","affiliation":[{"name":"Division of Applied Food System, College of Natural Science, Seoul Women\u2019s University, 621 Hwarangro, Nowon-Gu, Seoul 01797, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Smee","family":"Kang","sequence":"additional","affiliation":[{"name":"Division of Applied Food System, College of Natural Science, Seoul Women\u2019s University, 621 Hwarangro, Nowon-Gu, Seoul 01797, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3878-6059","authenticated-orcid":false,"given":"Jungil","family":"Hong","sequence":"additional","affiliation":[{"name":"Division of Applied Food System, College of Natural Science, Seoul Women\u2019s University, 621 Hwarangro, Nowon-Gu, Seoul 01797, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2022,12,29]]},"reference":[{"key":"ref_1","first-page":"936","article-title":"Evaluation of a tetrazolium-based semiautomated colorimetric assay: Assessment of chemosensitivity testing","volume":"47","author":"Carmichael","year":"1987","journal-title":"Cancer Res."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"57","DOI":"10.1016\/0022-1759(86)90215-2","article-title":"Use of MTT colorimetric assay to measure cell activation","volume":"94","author":"Gerlier","year":"1986","journal-title":"J. Immunol. Method."},{"key":"ref_3","doi-asserted-by":"crossref","unstructured":"Kumar, P., Nagarajan, A., and Uchil, P.D. (2018). Analysis of Cell Viability by the MTT Assay. Cold Spring Harb. Protoc., 2018.","DOI":"10.1101\/pdb.prot095505"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"474","DOI":"10.1006\/abbi.1993.1311","article-title":"Characterization of the cellular reduction of 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT): Subcellular localization, substrate dependence, and involvement of mitochondrial electron transport in MTT reduction","volume":"303","author":"Berridge","year":"1993","journal-title":"Arch. Biochem. Biophys."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"279","DOI":"10.1038\/bjc.1987.190","article-title":"A study of some variables in a tetrazolium dye (MTT) based assay for cell growth and chemosensitivity","volume":"56","author":"Twentyman","year":"1987","journal-title":"Br. J. Cancer"},{"key":"ref_6","first-page":"4827","article-title":"Evaluation of a Soluble tetrazolium\/formazan Assay for Cell Growth and Drug Sensitivity in Culture using Human and Other Tumor Cell Lines","volume":"48","author":"Scudiero","year":"1988","journal-title":"Cancer Res."},{"key":"ref_7","doi-asserted-by":"crossref","unstructured":"Su\u0142ek, A., Pucelik, B., Kobielusz, M., Barzowska, A., and D\u0105browski, J.M. (2020). Photodynamic Inactivation of Bacteria with Porphyrin Derivatives: Effect of Charge, Lipophilicity, ROS Generation, and Cellular Uptake on Their Biological Activity In Vitro. Int. J. Mol. Sci., 21.","DOI":"10.3390\/ijms21228716"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"1801","DOI":"10.1021\/bi00885a017","article-title":"Spectra of molecular complexes of porphyrins in aqueous solution","volume":"4","author":"Mauzerall","year":"1965","journal-title":"Biochemistry"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"1369","DOI":"10.3109\/10715762.2012.715371","article-title":"In vitro antioxidant capacity and antigenotoxic properties of protoporphyrin and structurally related tetrapyrroles","volume":"46","author":"Huber","year":"2012","journal-title":"Free Radic. Res."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"2277","DOI":"10.1016\/j.freeradbiomed.2012.10.529","article-title":"Treatment with heme arginate alleviates adipose tissue inflammation and improves insulin sensitivity and glucose metabolism in a rat model of Human primary aldosteronism","volume":"53","author":"Jadhav","year":"2012","journal-title":"Free Radic. Biol. Med."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"17","DOI":"10.1016\/0190-9622(93)70002-B","article-title":"Topical photodynamic therapy with endogenous porphyrins after application of 5-aminolevulinic acid: An alternative treatment modality for solar keratoses, superficial squamous cell carcinomas, and basal cell carcinomas","volume":"28","author":"Wolf","year":"1993","journal-title":"J. Am. Acad. Dermatol."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"275","DOI":"10.1016\/1011-1344(92)85108-7","article-title":"New trends in photobiology: Endogenous protoporphyrin IX, a clinically useful photosensitizer for photodynamic therapy","volume":"14","author":"Kennedy","year":"1992","journal-title":"J. Photochem. Photobiol. B"},{"key":"ref_13","doi-asserted-by":"crossref","unstructured":"De Annunzio, S.R., Costa, N.C.S., Mezzina, R.D., Graminha, M.A.S., and Fontana, C.R. (2019). Chlorin, Phthalocyanine, and Porphyrin Types Derivatives in Phototreatment of Cutaneous Manifestations: A Review. Int. J. Mol. Sci., 20.","DOI":"10.3390\/ijms20163861"},{"key":"ref_14","first-page":"266","article-title":"The syntheses of phthalocyanine hybrid derivatives and their properties","volume":"24","author":"Kim","year":"2013","journal-title":"Appl. Chem. Eng."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"693","DOI":"10.1038\/89068","article-title":"Heme oxygenase-1 protects against vascular constriction and proliferation","volume":"7","author":"Duckers","year":"2001","journal-title":"Nat. Med."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"993S","DOI":"10.1590\/0001-3765201820170811","article-title":"Cancer, Photodynamic Therapy and Porphyrin-Type Derivatives","volume":"90","author":"Gomes","year":"2018","journal-title":"An. Acad. Bras. Cienc."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"445","DOI":"10.1055\/s-2002-32073","article-title":"Interference of plant extracts, phytoestrogens and antioxidants with the MTT tetrazolium assay","volume":"68","author":"Bruggisser","year":"2002","journal-title":"Planta Med."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"189","DOI":"10.1007\/s10616-007-9057-4","article-title":"Influence of medium type and serum on MTT reduction by flavonoids in the absence of cells","volume":"52","author":"Talorete","year":"2006","journal-title":"Cytotechnology"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"754","DOI":"10.9721\/KJFST.2011.43.6.754","article-title":"Changes in color response of MTT formazan by zinc protoporphyrin","volume":"43","author":"Park","year":"2011","journal-title":"Korean J. Food Sci. Technol."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"42","DOI":"10.1016\/j.ab.2017.09.011","article-title":"Menadione-mediated WST1 reduction assay for the determination of metabolic activity of cultured neural cells","volume":"538","author":"Stapelfeldt","year":"2017","journal-title":"Anal. Biochem."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"15","DOI":"10.1007\/BF01404408","article-title":"A rapid and simple MTT-based spectrophotometric assay for determining drug sensitivity in monolayer cultures","volume":"11","author":"Edmondson","year":"1998","journal-title":"J. Tissue Cult. Method."},{"key":"ref_22","first-page":"165","article-title":"Cell sensitivity assays: The MTT assay","volume":"Volume 88","author":"Langdon","year":"2004","journal-title":"Cancer Cell Culture: Methods and Protocols"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"e1900132","DOI":"10.1002\/adhm.201900132","article-title":"Photosensitizers for Photodynamic Therapy","volume":"8","author":"Lan","year":"2019","journal-title":"Adv. Healthc. Mater."},{"key":"ref_24","doi-asserted-by":"crossref","unstructured":"Klausen, M., Ucuncu, M., and Bradley, M. (2020). Design of Photosensitizing Agents for Targeted Antimicrobial Photodynamic Therapy. Molecules, 25.","DOI":"10.3390\/molecules25225239"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"13454","DOI":"10.1021\/acs.chemrev.1c00381","article-title":"Recent Strategies to Develop Innovative Photosensitizers for Enhanced Photodynamic Therapy","volume":"121","author":"Pham","year":"2021","journal-title":"Chem. Rev."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"227","DOI":"10.1021\/tx00026a012","article-title":"Evaluation of the probe 2\u2032, 7\u2032-dichlorofluorescin as an indicator of reactive oxygen species formation and oxidative stress","volume":"5","author":"LeBel","year":"1992","journal-title":"Chem. Res. Toxicol."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"612","DOI":"10.1016\/S0891-5849(99)00107-0","article-title":"Quantifying cellular oxidative stress by dichlorofluorescein assay using microplate reader","volume":"27","author":"Wang","year":"1999","journal-title":"Free Radic. Biol. Med."},{"key":"ref_28","doi-asserted-by":"crossref","unstructured":"Chiang, S.K., Chen, S.E., and Chang, L.C. (2018). A Dual Role of Heme Oxygenase-1 in Cancer Cells. Int. J. Mol. Sci., 20.","DOI":"10.3390\/ijms20010039"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"837","DOI":"10.1007\/s00217-011-1451-7","article-title":"Antioxidant properties of BHT estimated by ABTS assay in systems differing in pH or metal ion or water concentration","volume":"232","author":"Dawidowicz","year":"2011","journal-title":"Eur. Food Res. Technol."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"25","DOI":"10.1016\/S0023-6438(95)80008-5","article-title":"Use of a free radical method to evaluate antioxidant activity","volume":"28","author":"Cuvelier","year":"1995","journal-title":"LWT Food Sci. Technol."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"3597","DOI":"10.1021\/jf000220w","article-title":"Assessing antioxidant and prooxidant activities of phenolic compounds","volume":"48","author":"Fukumoto","year":"2000","journal-title":"J. Agric. Food Chem."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"2071","DOI":"10.1007\/s00216-006-0998-9","article-title":"New Noncellular Fluorescence Microplate Screening Assay for Scavenging Activity Against Singlet Oxygen","volume":"387","author":"Costa","year":"2007","journal-title":"Anal. Bioanal. Chem."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"1315S","DOI":"10.1093\/ajcn\/62.6.1315S","article-title":"Vitamins E and C, \u03b2-Carotene, and Other Carotenoids as Antioxidants","volume":"62","author":"Sies","year":"1995","journal-title":"Am. J. Clin. Nutr."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"149","DOI":"10.1007\/BF02537155","article-title":"Singlet oxygen quenching ability of naturally occurring carotenoids","volume":"29","author":"Hirayama","year":"1994","journal-title":"Lipids"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"29","DOI":"10.1615\/JEnvironPatholToxicolOncol.v25.i1-2.30","article-title":"Singlet oxygen in photosensitization","volume":"25","author":"Moan","year":"2006","journal-title":"J. Environ. Pathol. Toxicol. 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