{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,7]],"date-time":"2026-07-07T19:45:15Z","timestamp":1783453515518,"version":"3.55.0"},"reference-count":46,"publisher":"MDPI AG","issue":"7","license":[{"start":{"date-parts":[[2020,7,19]],"date-time":"2020-07-19T00:00:00Z","timestamp":1595116800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Marine Drugs"],"abstract":"<jats:p>Polyhydroxylated naphthoquinones (PHNQs), known as spinochromes that can be extracted from sea urchins, are bioactive compounds reported to have medicinal properties and antioxidant activity. The MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) cell viability assay showed that pure echinochrome A exhibited a cytotoxic effect on Saos-2 cells in a dose-dependent manner within the test concentration range (15.625\u201365.5 \u00b5g\/mL). The PHNQ extract from New Zealand sea urchin Evechinus chloroticus did not induce any cytotoxicity within the same concentration range after 21 days of incubation. Adding calcium chloride (CaCl2) with echinochrome A increased the number of viable cells, but when CaCl2 was added with the PHNQs, cell viability decreased. The effect of PHNQs extracted on mineralized nodule formation in Saos-2 cells was investigated using xylenol orange and von Kossa staining methods. Echinochrome A decreased the mineralized nodule formation significantly (p &lt; 0.05), while nodule formation was not affected in the PHNQ treatment group. A significant (p &lt; 0.05) increase in mineralization was observed in the presence of PHNQs (62.5 \u00b5g\/mL) supplemented with 1.5 mM CaCl2. In conclusion, the results indicate that PHNQs have the potential to improve the formation of bone mineral phase in vitro, and future research in an animal model is warranted.<\/jats:p>","DOI":"10.3390\/md18070373","type":"journal-article","created":{"date-parts":[[2020,7,20]],"date-time":"2020-07-20T06:08:17Z","timestamp":1595225297000},"page":"373","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":5,"title":["PHNQ from Evechinus chloroticus Sea Urchin Supplemented with Calcium Promotes Mineralization in Saos-2 Human Bone Cell Line"],"prefix":"10.3390","volume":"18","author":[{"given":"Yakun","family":"Hou","sequence":"first","affiliation":[{"name":"Department of Food Science, University of Otago, Dunedin 9016, New Zealand"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alan","family":"Carne","sequence":"additional","affiliation":[{"name":"Department of Biochemistry, University of Otago, Dunedin 9016, New Zealand"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Michelle","family":"McConnell","sequence":"additional","affiliation":[{"name":"Department of Microbiology and Immunology, University of Otago, Dunedin 9016, New Zealand"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sonya","family":"Mros","sequence":"additional","affiliation":[{"name":"Department of Microbiology and Immunology, University of Otago, Dunedin 9016, New Zealand"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7526-026X","authenticated-orcid":false,"given":"Elena A.","family":"Vasileva","sequence":"additional","affiliation":[{"name":"G.B. Elyakov Pacific Institute of Bioorganic Chemistry, Far Eastern Branch of Russian Academy of Sciences, 690041 Vladivostok, Russia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7616-574X","authenticated-orcid":false,"given":"Natalia P.","family":"Mishchenko","sequence":"additional","affiliation":[{"name":"G.B. Elyakov Pacific Institute of Bioorganic Chemistry, Far Eastern Branch of Russian Academy of Sciences, 690041 Vladivostok, Russia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5411-7832","authenticated-orcid":false,"given":"Keegan","family":"Burrow","sequence":"additional","affiliation":[{"name":"Department of Food Science, University of Otago, Dunedin 9016, New Zealand"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ke","family":"Wang","sequence":"additional","affiliation":[{"name":"Department of Food Science, University of Otago, Dunedin 9016, New Zealand"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Adnan A.","family":"Bekhit","sequence":"additional","affiliation":[{"name":"Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Alexandria University, Alexandria 21521, Egypt"},{"name":"Pharmacy Program, Allied Health Department, College of Health Sciences, University of Bahrain, Sakheer P.O. Box 32 038, Bahrain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2569-8620","authenticated-orcid":false,"given":"Alaa El-Din A.","family":"Bekhit","sequence":"additional","affiliation":[{"name":"Department of Food Science, University of Otago, Dunedin 9016, New Zealand"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2020,7,19]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"32637","DOI":"10.1039\/C8RA04777D","article-title":"Naphthoquinones of the spinochrome class: Occurrence, isolation, biosynthesis and biomedical applications","volume":"8","author":"Hou","year":"2018","journal-title":"RSC Adv."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1007\/s11101-018-9547-3","article-title":"Naphthoquinone pigments from sea urchins: Chemistry and pharmacology","volume":"17","author":"Shikov","year":"2018","journal-title":"Phytochem. Rev."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"379","DOI":"10.1016\/j.foodchem.2018.08.046","article-title":"Extraction, structural characterization and stability of polyhydroxylated naphthoquinones from shell and spine of New Zealand sea urchin (Evechinus chloroticus)","volume":"272","author":"Hou","year":"2019","journal-title":"Food Chem."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1185","DOI":"10.1016\/j.lwt.2010.03.005","article-title":"Quantitative separation of antioxidant pigments in purple sea urchin shells using a reversed-phase high performance liquid chromatography","volume":"43","author":"Kuwahara","year":"2010","journal-title":"LWT Food Sci. Technol."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"331","DOI":"10.1007\/s00217-013-1996-8","article-title":"Extraction, structural characterization and antioxidant activity of polyhydroxylated 1,4-naphthoquinone pigments from spines of sea urchin Glyptocidaris crenularis and Strongylocentrotus intermedius","volume":"237","author":"Li","year":"2013","journal-title":"Eur. Food Res. Technol."},{"key":"ref_6","first-page":"455","article-title":"Extraction and identification of antioxidant polyhydroxynaphthoquinone pigments from the sea urchin, Psammechinus miliaris","volume":"59","author":"Powell","year":"2014","journal-title":"Eur. Food Res. Technol."},{"key":"ref_7","doi-asserted-by":"crossref","unstructured":"Brasseur, L., Hennebert, E., Fi\u00e9vez, L., Caulier, G., Bureau, F., Tafforeau, L., Flammang, P., Gerbaux, P., and Eeckhaut, I. (2017). The roles of spinochromes in four shallow water tropical sea urchins and their potential as bioactive pharmacological agents. Mar. Drugs., 15.","DOI":"10.3390\/md15060179"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"126339","DOI":"10.1016\/j.foodchem.2020.126339","article-title":"In vitro antioxidant and antimicrobial activities, and in vivo anti-inflammatory activity of crude and fractionated PHNQs from sea urchin (Evechinus chloroticus)","volume":"316","author":"Hou","year":"2020","journal-title":"Food Chem."},{"key":"ref_9","first-page":"34","article-title":"Histochrome, a new antioxidant, in the treatment of ocular diseases","volume":"115","author":"Egorov","year":"1998","journal-title":"Vestnik Oftalmol."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"48","DOI":"10.1023\/A:1023659331010","article-title":"Histochrome: A new original domestic drug","volume":"37","author":"Mishchenko","year":"2003","journal-title":"Pharm. Chem. J."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"223","DOI":"10.1007\/s10517-015-3134-z","article-title":"Assessment of nephroprotective potential of histochrome during induced arterial hypertension","volume":"160","author":"Agafonova","year":"2015","journal-title":"Bull. Exp. Biol. Med."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"739","DOI":"10.1007\/s10517-011-1238-7","article-title":"Evaluation of effects of Histochrome and Mexidol on structural and functional characteristics of the brain in senescence-accelerated OXYS Rats by magnetic resonance imaging","volume":"150","author":"Agafonova","year":"2011","journal-title":"Bull. Exp. Biol. Med."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"2922","DOI":"10.3390\/md12052922","article-title":"Echinochrome a protects mitochondrial function in cardiomyocytes against cardiotoxic drugs","volume":"12","author":"Jeong","year":"2014","journal-title":"Mar. Drugs."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"149","DOI":"10.1007\/s11094-014-1067-x","article-title":"Effects of histochrome on P53 expression in mouse red bone marrow cells in a model of chronic Stress","volume":"48","author":"Kareva","year":"2014","journal-title":"Pharm. Chem. J."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"41","DOI":"10.1016\/j.lfs.2016.03.007","article-title":"Mechanisms of echinochrome potency in modulating diabetic complications in liver","volume":"151","author":"Mohamed","year":"2016","journal-title":"Life Sci."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"1035","DOI":"10.1002\/jcb.23016","article-title":"Effects of Cissus quadrangularis on the proliferation, differentiation and matrix mineralization of human osteoblast like SaOS-2 cells","volume":"112","author":"Muthusami","year":"2011","journal-title":"J. Cell. Biochem."},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"Marcus, R., and Majumder, S. (2001). Chapter 35, the Nature of Osteoporosis. Osteoporosis, Academic Press. [2nd ed.].","DOI":"10.1016\/B978-012470862-4\/50036-2"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"3318","DOI":"10.1172\/JCI27071","article-title":"Pathogenesis of osteoporosis: Concepts, conflicts, and prospects","volume":"115","author":"Raisz","year":"2005","journal-title":"J. Clin. Investig."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"46","DOI":"10.1016\/j.lfs.2016.01.024","article-title":"Natural products for treatment of osteoporosis: The effects and mechanisms on promoting osteoblast-mediated bone formation","volume":"147","author":"An","year":"2016","journal-title":"Life Sci."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"473","DOI":"10.1517\/14656566.2016.1124856","article-title":"Management of severe osteoporosis","volume":"17","author":"Miller","year":"2016","journal-title":"Expert Opin. Pharmacother."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"981","DOI":"10.3389\/fphar.2018.00981","article-title":"Bone health and natural products-An insight","volume":"9","author":"Suvarna","year":"2018","journal-title":"Front Pharmacol."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"897","DOI":"10.1111\/j.2042-7158.2012.01483.x","article-title":"Cistanche deserticola extract increases bone formation in osteoblasts","volume":"64","author":"Li","year":"2012","journal-title":"J. Pharm. Pharmacol."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"382","DOI":"10.1016\/j.jsbmb.2014.08.002","article-title":"Vanillic acid exerts oestrogen-like activities in osteoblast-like UMR 106 cells through MAP kinase (MEK\/ERK)-mediated ER signaling pathway","volume":"144","author":"Xiao","year":"2014","journal-title":"J. Steroid Biochem."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"683","DOI":"10.1016\/j.phymed.2013.03.001","article-title":"Assessment of the role of flavonoids for inducing osteoblast differentiation in isolated mouse bone marrow derived mesenchymal stem cells","volume":"20","author":"Srivastava","year":"2013","journal-title":"Phytomedicine"},{"key":"ref_25","first-page":"38","article-title":"Sea urchin chemical and pharmacological research","volume":"2","author":"Lu","year":"1994","journal-title":"Chin. J. Mar. Drugs."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"537","DOI":"10.2174\/1389557514666140529224147","article-title":"Traditional Chinese Medicine\u2014\u2014Sea urchin","volume":"14","author":"Shang","year":"2014","journal-title":"Mini Rev. Med. Chem."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"171","DOI":"10.1016\/j.toxlet.2005.07.001","article-title":"In vitro cytotoxicity assays: Comparison of LDH, neutral red, MTT and protein assay in hepatoma cell lines following exposure to cadmium chloride","volume":"160","author":"Fotakis","year":"2006","journal-title":"Toxicol. Lett."},{"key":"ref_28","doi-asserted-by":"crossref","unstructured":"Van Meerloo, J., Kaspers, G.J., and Cloos, J. (2011). Cell sensitivity assays: The MTT assay. Cancer Cell Culture, Humana Press.","DOI":"10.1007\/978-1-61779-080-5_20"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"e1700182","DOI":"10.1002\/cbdv.201700182","article-title":"Diversity of polyhydroxynaphthoquinone pigments in North Pacific sea urchins","volume":"14","author":"Vasileva","year":"2017","journal-title":"Chem. Biodivers."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"4602","DOI":"10.3390\/md12084602","article-title":"Echinochrome a increases mitochondrial mass and function by modulating mitochondrial biogenesis regulatory genes","volume":"12","author":"Jeong","year":"2014","journal-title":"Mar. Drug."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"270","DOI":"10.1002\/1097-4636(200011)52:2<270::AID-JBM5>3.0.CO;2-1","article-title":"Calcium and phosphate supplementation promotes bone cell mineralization: Implications for hydroxyapatite (HA)-enhanced bone formation","volume":"52","author":"Chang","year":"2000","journal-title":"J. Biomed. Mater. Res."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"1697","DOI":"10.1002\/bp060274b","article-title":"Examination of mineralized nodule formation in living osteoblastic cultures using fluorescent dyes","volume":"22","author":"Wang","year":"2006","journal-title":"Biotechnol. Prog."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"345","DOI":"10.1016\/j.jep.2005.09.041","article-title":"Puerariae radix promotes differentiation and mineralization in human osteoblast-like SaOS-2 cells","volume":"104","author":"Huh","year":"2006","journal-title":"J. Ethnopharmacol."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"59","DOI":"10.1016\/j.jnutbio.2017.09.014","article-title":"Select polyphenolic fractions from dried plum enhance osteoblast activity through BMP-2 signaling","volume":"55","author":"Graef","year":"2018","journal-title":"J. Nutr. Biochem."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"483","DOI":"10.1097\/00075197-200111000-00003","article-title":"Effects of vitamin K on calcium and bone metabolism","volume":"4","author":"Zittermann","year":"2001","journal-title":"Curr. Opin. Clin. Nutr."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"1268","DOI":"10.1210\/jcem-60-6-1268","article-title":"Electrochemical detection of pepressed circulating levels of Vitamin K1 in osteoporosis","volume":"60","author":"Hart","year":"1985","journal-title":"J. Clin. Endocrinol. Metab."},{"key":"ref_37","unstructured":"Lancaster, C.E. (2015). Effects of Vitamin D, K1, K2 and Calcium on Bone Formation by Osteoblasts in vitro. [Master\u2019 Thesis, University of Toronto]."},{"key":"ref_38","first-page":"1364","article-title":"Vitamin K2 analog menaquinone-7 shows osteoblastic bone formation activity in vitro","volume":"28","author":"Zhu","year":"2017","journal-title":"Biomed. Res."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"254","DOI":"10.1016\/j.phytol.2015.01.009","article-title":"Naphthoquinone and flavonoid constituents from the carnivorous plant Nepenthes mirabilis and their anti-osteoporotic and antioxidant activities","volume":"11","author":"Thao","year":"2015","journal-title":"Phytochem. Lett."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"E616","DOI":"10.1152\/ajpendo.00229.2004","article-title":"Involvement of calcium-sensing receptor in osteoblastic differentiation of mouse MC3T3-E1 cells","volume":"288","author":"Yamauchi","year":"2005","journal-title":"Am. J. Physiol. Endocrinol. Metab."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"1452","DOI":"10.1002\/jbmr.5650081206","article-title":"Effects of high calcium concentration on the functions and interactions of osteoblastic cells and monocytes and on the formation of osteoclast-like cells","volume":"8","author":"Sugimoto","year":"1993","journal-title":"J. Bone Miner. Res."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"771","DOI":"10.1016\/j.bbrc.2016.09.137","article-title":"Inorganic polyphosphate triggers upregulation of interleukin 11 in human osteoblast-like SaOS-2 cells","volume":"479","author":"Lui","year":"2016","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"ref_43","unstructured":"Burrow, K.W. (2019). The Effects of New Zealand Sheep Milk on Mineral Absorption and Bone Health. [Ph.D. Thesis, University of Otago]."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"2643","DOI":"10.1002\/jbm.a.34937","article-title":"A phenotypic comparison of osteoblast cell lines versus human primary osteoblasts for biomaterials testing","volume":"102","author":"Czekanska","year":"2014","journal-title":"J. Biomed. Mater. Res. Part A."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"191","DOI":"10.1016\/S0003-9861(03)00111-5","article-title":"How do calcium ions induce free radical oxidation of hydroxy-1,4-naphthoquinone? Ca2+ stabilizes the naphthosemiquinone anion-radical of echinochrome A","volume":"413","author":"Lebedev","year":"2003","journal-title":"Arch. Biochem."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"885","DOI":"10.1023\/A:1011904819563","article-title":"Antioxidant properties, autooxidation, and mutagenic activity of echinochrome a compared with its etherified derivative","volume":"66","author":"Lebedev","year":"2001","journal-title":"Biochemistry (Moscow)"}],"container-title":["Marine Drugs"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1660-3397\/18\/7\/373\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T09:49:44Z","timestamp":1760176184000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1660-3397\/18\/7\/373"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2020,7,19]]},"references-count":46,"journal-issue":{"issue":"7","published-online":{"date-parts":[[2020,7]]}},"alternative-id":["md18070373"],"URL":"https:\/\/doi.org\/10.3390\/md18070373","relation":{},"ISSN":["1660-3397"],"issn-type":[{"value":"1660-3397","type":"electronic"}],"subject":[],"published":{"date-parts":[[2020,7,19]]}}}