BACKGROUND: Silymarin, an active polyphenolic fraction of Silybum marianum, and high flavonoid content of Fagopyrum possess various interesting biological activities. The substituted pyrazine-2-carboxamides were previously used as effective elicitors of studied secondary metabolites. OBJECTIVE: To study the effect of new synthetic pyrazine carboxamide derivatives, N-(4-chlorobenzyl)-5-tert-butylpyrazine-2-carboxamide (1) and 3-(3-((trifluoromethyl) benzyl) amino) pyrazine-2-carboxamide (2), on flavonolignan and flavonoid production in S. marianum and Fagopyrumes culentum in vitro cultures. MATERIALS AND METHODS: Callus and suspension cultures were cultured on MS medium containing α-naphtaleneacetic acid or 2,4-D. Three elicitor concentrations for different exposure times were tested. Dried and powdered samples of callus and suspension cultures were extracted with methanol and analyzed by DAD-HPLC. RESULTS: Compound 1 showed as a good elicitor of taxifolin production. The effect on silymarin complex was less visible with a maximum between 24 and 48 h after 3.292 ×10(-4) mol/L concentration. The detailed analysis showed that silychristin was the most abundant. Compound 2 was effective in rutin production only in callus culture with maximum 24 h and 168 h after application of 3.3756 ×10(-3) mol/L concentration and 48 and 72 h after 3.3756 ×10(-4) mol/L concentration. CONCLUSION: From the results of the performed experiments, it can be concluded that compound 1 shows to be suitable elicitor for enhanced production of taxifolin and silychristin in S. marianum, mainly when 3.292 ×10(-4) mol/L concentration was used, and compound 2 is suitable for increase rutin production in callus cultures and less appropriate for suspension cultures of F. esculentum. SUMMARY: The influence of two new synthetic pyrazine-2-carboxamidesderivatives on secondary metabolite content of Silybum marianum and Fagopyrum esculentum in vitro cultures was tested.In S. marianum, the derivate N-(4-chlorobenzyl)-5-tert-butylpyrazine-2-carboxamide showed as a good elicitor of taxifolin production and less effective for silymarin complex production with silychristin as the most abundant.The derivate 3-(3-((trifluoromethyl) benzyl) amino) pyrazine-2-carboxamide is suitable for increase rutin production in callus cultures and less appropriate for suspension cultures of F. esculentum.

Department of Biology Faculty of Science University of Hradec Kralove Rokitanskeho 62 500 03 Hradec Kralove Czech Republic

Department of Pharmaceutical Chemistry and Pharmaceutical Analysis Faculty of Pharmacy in Hradec Kralove Charles University Prague Heyrovskeho 1203 500 05 Hradec Kralove Czech Republic

Department of Pharmacognosy Faculty of Pharmacy in Hradec Kralove Charles University Prague Heyrovskeho 1203 500 05 Hradec Kralove Czech Republic

Zobrazit více v PubMed

Křen V, Walterova D. Silybin and silymarin – New effects and applications. Biomed Pap. 2005;149:29–41. PubMed

Shaker E, Mahmoud H, Mnaa S. Silymarin, the antioxidant component and Silybum marianum extracts prevent liver damage. Food Chem Toxicol. 2010;48:803–6. PubMed

Vaid M, Singh T, Prasad R, Katiyar SK. Silymarin inhibits melanoma cell growth both in vitro and in vivo by targeting cell cycle regulators, angiogenic biomarkers and induction of apoptosis. Mol Carcinog. 2014 doi: 10.1002/mc.22208. [Epub ahead of print] PubMed

Pérez HJ, Carrillo SC, García E, Ruiz-Mar G, Pérez-Tamayo R, Chavarría A. Neuroprotective effect of silymarin in a MPTP mouse model of Parkinson's disease. Toxicology. 2014;319:38–43. PubMed

Raza SS, Khan MM, Ashafaq M, Ahmad A, Khuwaja G, Khan A, et al. Silymarin protects neurons from oxidative stress associated damages in focal cerebral ischemia: a behavioral, biochemical and immunohistological study in Wistar rats. J Neurol Sci. 2011;309:45–54. PubMed

Singhal NK, Srivastava G, Patel DK, Jain SK, Singh MP. Melatonin or silymarin reduces maneb- and paraquat-induced Parkinson's disease phenotype in the mouse. J Pineal Res. 2011;50:97–109. PubMed

Inglett GE, Chen D, Berhow M, Lee S. Anti-oxidant activity of commercial buckwheat flours and their free and bound phenolic compositions. Food Chem. 2011;125:923–9.

Vogrincic M, Kreft I, Filipic M, Zegura B. Antigenotoxic effect of Tartary (Fagopyrum tataricum) and common (Fagopyrum esculentum) buckwheat flour. J Med Food. 2013;16:944–52. PubMed

Orcic D, Svirčev E, Mimica-Dukic N, Beara I, Balog K, Francišković M, et al. Phenolic profile and anti-oxidant activity of buckwheat (Fagopyrum esculentum) herb and root extracts. Planta Med. 2012;78:11.

Namdeo AG. Plant cell elicitation for production of secondary metabolites: a review. Pharmacogn Rev. 2007;1:69–79.

Zhao J, Davis LC, Verpoorte R. Elicitor signal transduction leading to production of plant secondary metabolites. Biotechnol Adv. 2005;23:283–333. PubMed

Sánchez-Sampedro MA, Fernández-Tárrago J, Corchete P. Yeast extract and methyl jasmonate-induced silymarin production in cell cultures of Silybum marianum (L.) Gaertn. J Biotechnol. 2005;119:60–9. PubMed

Zhao G, Zhao J, Peng L, Zou L, Wang J, Zhong L, et al. Effects of yeast polysaccharide on growth and flavonoid accumulation in Fagopyrum tataricum sprout cultures. Molecules. 2012;17:11335–45. PubMed PMC

Elwekeel A, Elfishway A, AbouZid S. Enhanced accumulation of flavonolignans in Silybum marianum cultured roots by methyl jasmonate. Phytochem Lett. 2010;5:393–6.

Kim HJ, Park KJ, Lim JH. Metabolomic analysis of phenolic compounds in buckwheat (Fagopyrum esculentum M.) sprouts treated with methyl jasmonate. J Agric Food Chem. 2011;59:5707–13. PubMed

Khalili M, Hasanloo T, Kazemi Tabar SK, Rahnama H. Influence of exogenous salicylic acid on flavonolignans and lipoxygenase activity in the hairy root cultures of Silybum marianum. Cell Biol Int. 2009;33:988–94. PubMed

Ashtiani SR, Hasanloo T, Bihamta MR. Enhanced production of silymarin by Ag+elicitor in cell suspension cultures of Silybum marianum. Pharm Biol. 2010;48:708–15. PubMed

Chung WJ, Kornilov A, Brodsky BH, Higgins M, Sanchez T, Heifets LB, et al. Inhibition of M. tuberculosis in vitro in monocytes and in mice by aminomethylene pyrazinamide analogs. Tuberculosis (Edinb) 2008;88:410–9. PubMed

Servusová B, Eibinová D, Doležal M, Kubícek V, Paterová P, Peško M, et al. Substituted N-benzylpyrazine-2-carboxamides: Synthesis and biological evaluation. Molecules. 2012;17:13183–98. PubMed PMC

Doležal M, Kráľová K. Herbicides, Theory and Applications: Synthesis and Evaluation of Pyrazine Derivatives with Herbicidal Activity. InTech. 2011. Available from: http://www.intechopen.com/books/herbicides-theory-and-applications/synthesis-andevaluation-of-pyrazine-derivatives-with-herbicidal-activity .

Tumová L, Tuma J, Megusar K, Dolezal M. Substituted pyrazinecarboxamides as abiotic elicitors of flavolignan production in Silybum marianum (L.) gaertn cultures in vitro. Molecules. 2010;15:331–40. PubMed PMC

Tumova L, Tuma J, Dolezal M. Pyrazinecarboxamides as potential elicitors of flavonolignan and flavonoid production in Silybum marianum and Ononis arvensis cultures in vitro. Molecules. 2011;16:9142–52. PubMed PMC

Doležal M, Zitko J, Carillo C, Eibinová D, Tauchman M, Kuneš J, et al. Substituted N-Benzylpyrazine-2-Carboxamides, their Synthesis, Hydro-Lipophilic Properties and Evaluation of their Anti-Mycobacterial, and Photosynthesis-Inhibiting Activity. 15th International Electronic Conference on Synthetic Organic Chemistry (ECSOC-15). November 1-30, 2011. p. a64/1-a64/6

Murashige T, Skoog F. A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant. 1962;15:437–97.

Praha: Grada; 2009. The Ministry of Health of the Czech Republic. Fagopyriherba. In: Czech Pharmacopoeia; pp. 2069–70.

Dolezal M, Tumová L, Kesetovicová D, Tuma J, Králová K. Substituted N-phenylpyrazine-2-carboxamides, their synthesis and evaluation as herbicides and abiotic elicitors. Molecules. 2007;12:2589–98. PubMed PMC

Kašparová M, Siatka T, Klimešová V, Dušek J. New synthetic pyridine derivate as potential elicitor in production of isoflavonoids and flavonoids in Trifolium pratense L. suspension culture. ScientificWorldJournal 2012. 2012 746412. PubMed PMC