Hattem Mekky, Samah El Sohafy, Rasha Abu El-khair, Amr El Hawiet


Objective: The present study was designed to study the effect of supplementing Silybum marianum cultures with different combinations and concentrations of growth regulators on the total phenolic content and the antioxidant activity of the produced cultures.

Methods: The total polyphenolic content of 23 extracts of calli was calculated as µg of gallic acid/mg of dry weight extract (DWE) using Folin–Ciocalteu reagent. The antioxidant activity was calculated as trolox equivalent antioxidant capacity using 1,1-Diphenyl-2-picryl-hydrazyl (DPPH) assay for extracts containing more than 10 µg gallic acid/mg DWE.

Results: Most of the combinations showed an increase in the total polyphenolic concentration compared to the wild plant. However, cultures grown on media supplemented with combinations of benzyl aminopurine (BAP) and gebberellic acid (Gb), adenine (Ad) and kinetin (Kin), BAP and indole acetic acid (IAA) possessed the highest total polyphenolic contents 20.5, 13.09 and 12.15 µg gallic acid/mg DWE, respectively. However, only (BAP+Gb), (BAP+1-Naphthaleneacetic acid (NAA), IAA, kin and (BAP+Ad) showed a significant increase in the antioxidant activity calculated as trolox equivalent with 2.65, 1.56, 1.56, 1.35 and 1.22 folds increase over the wild plant, respectively.

Conclusion: The results obtained clearly indicated that changing the growth regulator system of Silybum marianum cultures serves as an easy and reliable method for increasing the total phenolic content and the antioxidant activity of cultures.


Silybum marianum, Growth regulators, Polyphenolic, Antioxidant, Trolox, DPPH, Folin–Ciocalteu


Ram VJ. Herbal preparations as a source of hepatoprotective agents. Drug News Perspect 2001;14:353-63.

Byass P. The global burden of liver disease: a challenge for methods and for public health. BMC Med 2014;12:159.

Mokdad AA, Lopez AD, Shahraz S, Lozano R, Mokdad AH, Stanaway J, et al. Liver cirrhosis mortality in 187 countries between 1980 and 2010:a systematic analysis. BMC Med 2014;12:145.

Awadalla HI, Ragab MH, Nassar NA, Osman M. Risk factors of hepatitis C infection among Egyptian blood donors. Cent Eur J Public Health 2011;19:217-21.

Darwish MA, Faris R, Darwish N, Shouman A, Gadallah M, El-Sharkawy M, et al. Hepatitis C and cirrhotic liver disease in the Nile Delta of Egypt: a community-based study. Am J Trop Med Hyg 2001;64:147-53.

Waked IA, Saleh SM, Moustafa MS, Raouf AA, Thomas DL, Strickland GT. High prevalence of hepatitis C in Egyptian patients with the chronic liver disease. Gut 1995;37:105-7.

Strickland GT. Liver disease in Egypt: hepatitis C superseded Schistosomiasis as a result of iatrogenic and biological factors. Hepatol 2006;43:915-22.

Lehman EM, Wilson ML. Epidemic hepatitis C virus infection in Egypt: estimates of past incidence and future morbidity and mortality. J Viral Hepat 2009;16:650-8.

Riley TR, Smith JP. Preventive care in chronic liver disease. J Gen Int Med 1999;14:699-704.

El Sohafy SM, Alqasoumi SI, Metwally AM, Omar AA, Amer MM, Abou-Shoer MI, et al. Evaluation of the hepatoprotective activity of some plants belonging to the tribe Cynareae growing in Egypt. J Med Plants Res 2013;7:324-8.

Witte I, Berlin J, Wray V, Schubert W, Kohl W, Hofle G, et al. Mono and diterpenes from cell culture of Thuja Occidenalis. Planta Med 1983;49:216-21.

Bahmani M, Shirzad H, Rafieian S, Rafieian-Kopaei M. Silybum marianum: Beyond hepatoprotection. J Evidence-Based Complementary Altern Med 2015;20:292-301.

Pradhan SC, Girish C. Hepatoprotective herbal drug, silymarin from experimental pharmacology to clinical medicine. Indian J Med Res 2006;124:491-504.

Morazzoni P, Bombardelli E. Silybum marianum (Carduus marianus). Fitoterapia 1995;66:3-42.

Vargas-Mendoza N, Madrigal-Santillán E, Morales-González A, Esquivel-Soto J, Esquivel-Chirino C. Hepatoprotective effect of silymarin. World J Hepatol 2014;6:144-9.

Polyak SJ, Ferenci P, Pawlotsky J. Hepatoprotective and antiviral functions of silymarin components in HCV infection. Hepatol 2013;57:1262-71.

Dixit N, Baboota S, Kohli K, Ahmad S, Ali J. Silymarin: a review of pharmacological aspects and bioavailability enhancement approaches. Indian J Pharmacol 2007;39:172-9.

Fraschini F, Demartini G, Esposti D. Pharmacology of silymarin. Clin Drug Invest 2002;22:51-65.

Luper S. A review of plants used in the treatment of liver disease: part 1. Altern Med Rev 1998;3:410-21.

Ligeret H, Brault A, Vallerand D, Haddad Y, Haddad PS. Antioxidant and mitochondrial protective effects of silibinin in cold preservation–warm reperfusion liver injury. J Ethnopharmacol 2008;115:507-14.

Dvorak Z, Kosina P, Walterova D, Simanek V, Bachleda P, Ulrichova J. Primary cultures of human hepatocytes as a tool in cytotoxicity studies: cell protection against model toxins by flavonolignans obtained from Silybum marianum. Toxicol Lett 2003;137:201-12.

Kvasnicka F, Bıba B, Sevcık R, Voldrich M, Kratka J. Analysis of the active components of silymarin. J Chromatogr 2003;990:239-45.

Lee DYW, Liu Y. Molecular structure and stereochemistry of silybin A, silybin B, isosilybin A and isosilybin B isolated from Silybum marianum (milk thistle). J Nat Prod 2003;66:1171-4.

Muriel P, Mourelle M. Prevention by silymarin of membrane alterations in acute CCl4 liver damage. J Appl Toxicol 1990; 10:275-9.

Ainsworth EA, Gillespie KM. Estimation of total phenolic content and other oxidation substrates in plant tissues using Folin–Ciocalteu reagent. Nat Protoc 2007;2:875-7.

Dal Toso R, Melandri F. Echinacea angustifolia cell culture extract, added value for sport and fitness. Natura Foods 2011;10:19-25.

Miguez M, Anundi I, Sainz-Pardo LA, Lindros KO. Hepatoprotective mechanism of silymarin: No evidence for the involvement of cytochrome P450 2E1. Chem Biol Interact 1994;91:51-63.

Nencini C, Giorgi G, Micheli L. Protective effect of silymarin on oxidative stress in rat brain. Phytomed 2007;14:129-35.

Bosisio E, Benelli C, Pirola O. Effect of the flavanolignans of Silybum marianum L. on lipid peroxidation in rat liver microsomes and freshly isolated hepatocytes. Pharmacol Res 1992;25:147-54.

Campos R, Garido A, Guerra R, Valenzuela A. Silybin dihemisuccinate protects against glutathione depletion and lipid peroxidation induced by acetaminophen on rat liver. Planta Med 1989;55:417-9.

Polyak SJ, Morishima C, Shuhart MC, Wang CC, Liu Y, Lee DY. Inhibition of T-cell inflammatory cytokines, hepatocyte NF-kappa B signaling, and HCV Infection by Standardized Silymarin. Gastroenterology 2007;132:1925-36.

De La Puerta R, Martinez E, Bravo L, Ahumada MC. Effect of silymarin on different acute inflammation models and on leukocyte migration. J Pharm Pharmacol 1996;48:968-70.

Sonnenbichler J, Zetl I. Biochemical effects of the flavanolignane silibinin on RNA, protein and DNA synthesis in rat livers. In: Cody V, Middleton E, Harbourne JB. Plant Flavonoids in Biology and Medicine: Biochemical, Pharmacological, and Structure-Activity Relationships. New York: Alan R. Liss, Inc; 1986. p. 319-31.

Tsai JH, Liu JY, Wu TT, Ho PC, Huang CY, Shyu JC, et al. Effects of silymarin on the resolution of liver fibrosis induced by carbon tetrachloride in rats. J Viral Hepat 2008;15:508-14.

Jia J, Bauer M, Cho J, Ruehl M, Milani S, Boigk G, et al. Antifibrotic effect of silymarin in rat secondary biliary fibrosis is mediated by downregulation of procollagen α1(I) and TIMP-1. J Hepatol 2001;35:392-8.

Gharagozloo M, Karimi M, Amirghofran Z. Immunomodulatory effects of silymarin in patients with β-thalassemia major. Int Immunopharmacol 2013;16:243-7.

Amirghofran Z, Azadbakht M, Karimi MH. Evaluation of the immunomodulatory effects of five herbal plants. J Ethnopharmacol 2000;72:167-72.

Fidranny I, Sukowati A, Sukrasno S. In vitro antioxidant activities of various leaves extracts from five varities of Rambutan (Nephelium lappaceum) and it’s correlation with total flavonoid, phenolic, carotenoid content. Asian J Pharm Clin Res 2015;8:139-43.

Fidranny I, Puspitasari N, Singgih M. Antioxidant activities, total flavonoid, phenolic, a carotenoid of various shells extracts from four species of legumes. Asian J Pharm Clin Res 2014;7:42-6.

Taviera M, Pereira DM, Sousa C, Ferreres F, Andrade PB, Martins A, et al. In vitro cultures of Brassica oleraceae L. var costata DC: Potential plant Bioreactor forantioxidant phenolic compounds. J Agric Food Chem 2009;57:1247-52.

Sayd SS, Taie HAA, Taha LS. Micropropagation, antioxidant activity, total phenolics and flavonoids content of Gardenia jasminoides Ellis as affected by growth regulators. Int J Acad Res 2010;2:184-91.

Anastasia E, Giannakoula F, Jelena J, Maksimovic M, Maksimovic DZ. The effects of plant growth regulators on growth, yield and phenolic profile of lentil plants. J Food Comp Anal 2012;28:46-53.

Jeong G, Woo J, Park D. Effect of plant growth regulators on growth and biosynthesis of phenolic compounds in genetically transformed hairy roots of Panax ginseng C. A. Meyer. Biotechnol Bioprocess Eng 2007;12:86-91.

About this article




Silybum marianum, Growth regulators, Polyphenolic, Antioxidant, Trolox, DPPH, Folin–Ciocalteu





Additional Links

Manuscript Submission


International Journal of Pharmacy and Pharmaceutical Sciences
Vol 9, Issue 4, 2017 Page: 44-47

Online ISSN



42 Views | 83 Downloads

Authors & Affiliations

Hattem Mekky
Department of Pharmacognosy, Faculty of Pharmacy, University of Alexandria, Egypt

Samah El Sohafy
Department of Pharmacognosy, Faculty of Pharmacy, University of Alexandria, Egypt

Rasha Abu El-khair
Department of Pharmacognosy, Faculty of Pharmacy, University of Alexandria, Egypt

Amr El Hawiet
Department of Pharmacognosy, Faculty of Pharmacy, University of Alexandria, Egypt


Article Tools



  • There are currently no refbacks.