DOI QR코드

DOI QR Code

Evaluation of in vivo Genotoxicity of Plant Flavonoids, Quercetin and Isoquercetin

식물유래 플라보노이드 Quercetin과 Isoquercetin의 생체 내 유전독성평가

  • Pak, Bumsoo (Department of Biotechnology, Hoseo University) ;
  • Han, Sehee (Department of Biotechnology, Hoseo University) ;
  • Lee, Jiyeon (Department of Biotechnology, Hoseo University) ;
  • Chung, Young-Shin (Department of Biotechnology, Hoseo University)
  • Received : 2016.09.20
  • Accepted : 2016.09.30
  • Published : 2016.10.28

Abstract

In vivo genotoxic potential of isoquercetin, a plant common flavonoid, in comparison with quercetin was investigated for the DNA breakage and the clastogenicity endpoints. Male ICR mice were administered by oral gavage for 3 days with $3{\times}0.5%$ carboxymethyl cellulose (CMC), 3 ${\times}$ isoquercetin (250, 500 mg/kg/day), 3 ${\times}$ quercetin (250, 500 mg/kg/day) and 2 ${\times}$ ethyl methanesulfonate (EMS, 200 mg/kg/day). Tissues were collected 48 hours after the first treatment and within 3 hours after the last treatment. The DNA damages were evaluated using Comet assay in liver and stomach, while the clastogenicities were determined using micronucleus test in bone marrow of same animals. The treatment of isoquercetin as well as quercetin did not cause the DNA damages in liver and stomach, and not induce the frequencies of micronucleus polychromatic erythrocytes in bone marrow. In conclusion, isoquercetin as well as quercetin did not cause the DNA breakages and the chromosomal damages in vivo system in these study conditions.

식물에서 흔히 존재하는 isoquercetin의 유전독성을 평가하기 위하여 최종평가항목인 DNA 절단 및 염색체 손상을 quercetin과 비교 평가하였다. 7주령의 수컷 ICR 마우스를 사용하였고 3일 동안 시험물질을 경구로 투여하였다. 부형제로 0.5% carboxymethylcellulose를 사용하였고 isoquercetin과 quercetin은 각각 250, 500 mg/kg/day로 투여하였으며, 양성대조물질로 ethyl methanesulfonate 200 mg/kg/day를 사용하였다. 일차 투여 후 48시간에 그리고 마지막 투여 후 3시간 내에 부검하였고 조직을 적출하였다. DNA 손상은 Comet assay를 사용하여 위와 간세포에서 관찰하였고, 소핵시험은 골수세포에서 소핵 분석방법을 사용하여 평가하였다. 두 가지 유전독성 시험을 동일 마우스를 사용하여 수행하였다. 그 결과, isoquercetin과 quercetin의 경구 투여는 500 mg/kg/day에서도 위와 간에서 DNA 손상을 초래하지 않았으며 골수세포에서 소핵을 유발하지 않았다. 따라서 본 연구에서 사용한 플라보노이드는 본 실험 조건하에서 유전독성을 유발하지 않는 것으로 사료된다.

Keywords

References

  1. Appleton, J.: Evaluating the Bioavailability of Isoquercetin. Nat. Med. J., 2, 1-6 (2010).
  2. Heim, K.E., agliaferro, A.R., Bobilya, D.J.: Flavonoid antioxidants: chemistry, metabolism and structure-activity relationships. J. Nutr. Biochem., 13, 572-584 (2002). https://doi.org/10.1016/S0955-2863(02)00208-5
  3. Murota, k., Hotta, A., Ido, H., Kawai, Y., Moon, J.H., Sekido, K., Hayashi, H., Inakuma, T., Terao, J.: Antioxidant capacity of albumin-bound quercetin metabolites after onion consumption in humans. Dep. Food Sci., 770-8503 (2007).
  4. Manach, C., Williamson, Gary., Morand, C., Scalbert, A., Remesy, C.: Bioavailability and bioefficacy of polyphenols in humans. I. Review of 97 bioavailability studies. Am. J. Clin. Nutr., 81, 230-42 (2005). https://doi.org/10.1093/ajcn/81.1.230S
  5. Ansari, M.A., Abdul, H.M., Joshi, G., Opii, W.O., Butterfield, D.A.: Protective effect of quercetin in primary neurons against $A{\beta}$(1-42): relevance to Alzheimer's disease. J. Nutr. Biochem., 20, 269-275 (2009). https://doi.org/10.1016/j.jnutbio.2008.03.002
  6. Dajas, F., Rivera-Megret F., Blasina, F., Arredondo, F., Abin-Carriquiry, J.A., Costa, G., Echeverry, C., Lafon, L., Heizen, H., Ferreira, M., Morquio, A.: Neuroprotection by flavonoids. Braz. J. Med . Biol. Res., 36, 1613-1620 (2003). https://doi.org/10.1590/S0100-879X2003001200002
  7. Hyang D.G., Lee, K.H., Kim, H.J., Lee, E.H., Lee, J.y., Song, Y.S., Lee, Y.H., Jin, C.b., Lee, Y.S., Choa, J.s.: N europrotective effects of antioxidative flavonoids, quercetin, (+)-dihydroquercetin and quercetin 3-methyl ether, isolated from Opuntia ficus-indica var. saboten. Brain Res., 965, 130-36 (2003). https://doi.org/10.1016/S0006-8993(02)04150-1
  8. Edwards, R.L., Lyon, T., Litwin, S.E., Rabovsky, A., Symons, J.D., Jalili, T.: Quercetin Reduces Blood Pressure in Hypertensive Subjects. J. Nutr., 137, 2405-2411 (2007). https://doi.org/10.1093/jn/137.11.2405
  9. Chopra, M., Fitzsimons, P.E.E., Strain, J.J., Thurnham, D.I., Howard, A.N.: Nonalcoholic Red Wine Extract and Quercetin Inhibit LDL Oxidation without Affecting Plasma Antioxidant Vitamin and Carotenoid Concentrations. Clin. Chem., 46, 1162-170 (2000).
  10. Hubbard, G.P., Wolffram, S., Lovegrove, J.A., Gibbins, J.M.: Ingestion of quercetin inhibits platelet aggregation and essential components of the collagen-stimulated platelet activation pathway in humans. J. Thromb. Haemost., 2, 2138-145 (2004). https://doi.org/10.1111/j.1538-7836.2004.01067.x
  11. Vitor, R.F., Mota-Filipe, H., Teixeira, G., Borges, C., Rodrigues, A.I., Teixeira, A., Paulo, A.: Flavonoids of an extract of Pterospartum tridentatum showing endothelial protection against oxidative injury. J. Ethnopharmacol., 93, 363-370 (2004). https://doi.org/10.1016/j.jep.2004.04.003
  12. Bouriche, H., Miles, E.A., Selloum, L., Calder, P.C.: Effect of Cleome arabica leaf extract, rutin and quercetin on soybean lipoxygenase activity and on generation of inflammatory eicosanoids by human neutrophils. Prostaglandins, Leukotrienes and Essential Fatty Acids, 72, 195-201 (2005). https://doi.org/10.1016/j.plefa.2004.10.018
  13. Comalada, M., Camuesco, D., Sierra, S., Ballester, I., Xaus, J., Glvez, J., A Zarzue, A.: In vivo quercitrin anti-inflammatory effect involves release of quercetin, which inhibits inflammation through down-regulation of the $NF-{\kappa}B$ pathway. Eur. J. Immunol., 35, 584-592 (2005). https://doi.org/10.1002/eji.200425778
  14. UEDA, H., YAMAZAKI, C., YAMAZAKI, M.:A Hydroxyl Group of Flavonoids Affects Oral Antiinflammatory Activity and Inhibition of Systemic Tumor Necrosis Factor-$\alpha$ Production. Biotechnol. Biochem., 68, 119-125 (2014).
  15. Middleton, E., Kandaswami, C., Theoharides, T.C.: The Effects of Plant Flavonoids on Mammalian Cells: Implications for Inflammation, Heart Disease, and Cancer. Pharmacol. Rev., 52, 673-51 (2000).
  16. Thornhill, S.M., Kelly, A.M.: Natural Treatment of Perennial Allergic Rhinitis. Altern. Med. Rev., 5, 448-454 (2000).
  17. Hollman, P.C.H., van Trijp, J.M.P., Mengelers, M.J.B., de Vries, J.H.M., Katan, M.B.: Bioavailability of the dietary antioxidant flavonol quercetin in man. Cancer. Lett., 114, 139-140 (1997). https://doi.org/10.1016/S0304-3835(97)04644-2
  18. Wolffram, S., Block, M., Ader, P.: Quercetin-3-Glucoside Is Transported by the Glucose Carrier SGLT1 across the Brush Border Membrane of Rat Small Intestine. J. Nutr., 132, 630-635 (2002). https://doi.org/10.1093/jn/132.4.630
  19. Ader P., Wessmann A.,Wolffram S.: Bioavailability and metabolism of the floavonol quercetin in the pig. Free Radical Bilo. Med., 28, 1056-1067 (2000). https://doi.org/10.1016/S0891-5849(00)00195-7
  20. Gee, J.M., DuPont, M.S., Day, A.J., Plumb, G.W., Williamson, G., Johnson, I.T.: Intestinal transport of quercetin glycosides in rats involves both deglycosylation and interaction with the hexose transport pathway. J. Nutr., 130, 2765-2771 (2000). https://doi.org/10.1093/jn/130.11.2765
  21. He, J., Feng, Y., Ouyang, H.-Z., Yu, B., Chang, Y.-X., Pan, G.-X., Dong, G.-Y., Wang, T., Gao, X.-M.: A sensitive LC-MS/ MS method for simultaneous determination of six flavonoids in rat plasma: application to a pharmacokinetic study of total flavonoids from mulberry leaves. J. Pharm. Biomed. Anal., 84, 189-195 (2013). https://doi.org/10.1016/j.jpba.2013.06.019
  22. Pang, Q., Tian, Y., Mi, J., Wang, J., Xu, Y.: Simultaneous determination and pharmacokinetic study of eight components in rat plasma by UHPLC-MS/MS after oral administration of Hypericum japonicum Thunb extract. J. Pharm. Biomed. Anal., 118, 228-234 (2016). https://doi.org/10.1016/j.jpba.2015.10.027
  23. Paulke, A., Eckert, G.P., Schubert-Zsilavecz, M., Wurglics, M.: Isoquercitrin provides better bioavailability than quercetin: comparison of quercetin metabolites in body tissue and brain sections after six days administration of isoquercitrin and quercetin. Pharm., 67, 991-996 (2012).
  24. Chaudhary, A., Willett, K.L.: Inhibition of human cytochrome CYP 1 enzymes by flavonoids of St. John's wort. Toxicology, 217, 194-205 (2006). https://doi.org/10.1016/j.tox.2005.09.010
  25. Bjeldanes, L.F., Chang, G.W.: Mutagenic activity of quercetin and related compounds. Sci., 197, 577-578 (1977). https://doi.org/10.1126/science.327550
  26. Schimmer, O., Hafele, F., Kruger, A.: The mutagenic of plant extracts containing quercetin in Salmonella typhimurium TA98 and TA100. Mutat. Res., 206, 201-208 (1988). https://doi.org/10.1016/0165-1218(88)90161-9
  27. Carver, J.H., Carrano, A.V., MacGregor, J.T.: Genetic effects of the flavonols quercetin, Kaempferol, and galangin on chineses hamster ovary cells in vitro. Mutat. Res., 113, 45-60 (1983). https://doi.org/10.1016/0165-1161(83)90240-6
  28. Engen, A., Maedaa, J., Wozniaka, D.E., Brentsa, C.A., Bell, J.J., Uesaka, M., Aizawa, Y., Katoa, T.A.:Induction of cytotoxic and genotoxic responses by natural and novel quercetin glycosides. Mutat. Res., 784-785, 15-22 (2015). https://doi.org/10.1016/j.mrgentox.2015.04.007
  29. Taj, S., Nagarajan, B.: Inhibition by quercetin and luteolin of chromosomal alterations induced by salted, deep-fried fish and mutton in rats. Mutat. Res., 369, 97-106 (1996). https://doi.org/10.1016/S0165-1218(96)90053-1
  30. da Silva J., Herrmann S. M., Heuser V., Peres W., Possa Marroni N., Gonzalez-Gallego J., Erdtmann B.: Evaluation of the genotoxic effect of rutin and quercetin by comet assay and micronucleus test. Food Chem. Toxicol., 40, 941-947 (2002). https://doi.org/10.1016/S0278-6915(02)00015-7
  31. Okada, E., Fujiishi, Y., Narumi, K., Yasutake, N., Ohyama, W.: A four-day oral treatment regimen for simultaneous micronucleus analyses in the glandular stomach, colon, and bone marrow of rats. Mutat. Res., 758, 87-94 (2013). https://doi.org/10.1016/j.mrgentox.2013.10.002
  32. Harwood, M., Danielewska-Nikiel, B., Borzelleca, J.F., Flamm, G.W., Williams, G.M., Lines, T.C.: A critical review of the data related to the safety of quercetin and lack of evidence of in vivo toxicity, including lack of genotoxic/carcinogenic properties. Food Chem. Toxicol., 45, 2179-2205 (2007). https://doi.org/10.1016/j.fct.2007.05.015
  33. Hasumura, M., Yasuhara, K., Tamura, T., Imai, T., Mitsumori, K., Hirose, M.: Evaluation of the toxicity of enzymatically decomposed rutin with 13-weeks dietary administration to Wistar rats. Food Chem. Toxicol., 42, 439-444 (2004). https://doi.org/10.1016/j.fct.2003.10.006
  34. Tamura, T., Mitsumori, K., Muto, S., Kasahara, H., Kobayashi, S., Okuhara, Y., Hayashi, M., Nagasawa, T., Onozato, T., Kuroda, J.: Fifty-two week chronic toxicity of enzymatically decomposed rutin in Wistar rats. Food Chem. Toxicol., 48, 2312-2318 (2010). https://doi.org/10.1016/j.fct.2010.05.065
  35. Zan, M.A., Ferraz, A.B.F., Richter, M.F., Picada, J.N., de Andrade, H.H.R., Lehmann, M., Dihl, Emilene Nunes, R.R., Semedo, J., Silva, J.D.: In vivo Genotoxicity Evaluation of an Artichoke (Cynara scolymus L.) Aqueous Extract. J. Food Sci., 78, T367-T371 (2013). https://doi.org/10.1111/1750-3841.12034
  36. Tice, R., Agurell, E., Anderson, D., Burlinson, B., Hartmann, A., Kobayashi, H., Miyamae, Y., Rojas, E., Ryu, J. C., and Sasaki, Y. F.: Single cell gel/comet assay: Guidelines for in vitro and in vivo genetic toxicology testing. Environ. Mol. Mutagen., 35, 206-221 (2000). https://doi.org/10.1002/(SICI)1098-2280(2000)35:3<206::AID-EM8>3.0.CO;2-J
  37. The comet assay International Validation Management Team. (2013). Report of the JaCVAM initiative international pre-validation studies of the in vivo rodent alkaline Comet assay for the detection of genotoxic carcinogens, version 1.4. Available at: http://www.oecd.org/env/ehs/testing/Come%20assay%20revised%20pre-validation%20report%202013.pdf.
  38. Schmid, W.: The micronucleus test. Mutat. Res., 31, 9-15 (1975). https://doi.org/10.1016/0165-1161(75)90058-8
  39. OECD Paris. (1997). OECD Guideline for the Testing of Chemicals: Mammalian Erythrocyte Micronucleus Test (No. 474).
  40. Utesch D., Feige, K., Dasenbrock, J., Broschard, T.H., Harwood, M., Danielewska-Nikiel, B., Lines, T.C.: Evaluation of the potential in vivo genotoxicity of quercetin. Mutat. Res., 654, 38-44 (2008) https://doi.org/10.1016/j.mrgentox.2008.04.008
  41. Valentova, K., Vrba, J., Bancirova, M., Ulrichova, J., Kren, V.: Isoquercitrin: Pharmacology, toxicology, and metabolism. Food Chem. Toxicol., 68, 267-282 (2014). https://doi.org/10.1016/j.fct.2014.03.018
  42. Snijman, P.W., Swanevelder, S., Joubert, E., Green, J.r., Gelderblom, W.C.: The antimutagenic activity of the major flavonoids of rooibos (Aspalathus linearis): some dose-response effects on mutagen activation-flavonoid interactions. Mutat. Res., 631, 111-123 (2007). https://doi.org/10.1016/j.mrgentox.2007.03.009
  43. Sekeroglu, V., Aydin B., Sekeroglu, Z.A.: Viscum album L. extract and quercetin reduce cyclophosphamide-induced cardiotoxicity, urotoxidity and genotoxicity in mice. Asian Pacific J Cancer Prev., 12, 2925-2931 (2011).
  44. Valentova, K., Sima, P., Rybkova, Z., Krizan, J., Malachova, K., Kren, V.: (Anti)mutagenic and immunomodulatory properties of quercetin glycosides. J Sci Food Agric., DOI: 10.1002/jsfa.7251 (2015).