In vitro antioxidant assessment, screening of enzyme inhibitory activities of methanol and water extracts and gene expression in Hypericum lydium

  • Nuraniye EruygurEmail author
  • Esra Ucar
  • Hüseyin Aşkın Akpulat
  • Keyhan Shahsavari
  • Seyed Mehdi Safavi
  • Danial Kahrizi
Original Article


Hypericum lydium Boiss. is a perennial plant of the Hypericaceae family, which has been used in particular to treat depression. The aim of this study was to determine in vitro antioxidant, antimicrobial activities, anticholinesterase (acetylcholinesterase (AChE)/butyrylcholinesterase (BChE)), antidiabetic activities (α-glucosidase/α-amylase) and Tyrosinase inhibitor activity of methanol and water extracts of H. lydium. Also, gene expression has been evaluated in the shoot and root by microarray technology. So, in general, the purpose of this study is to study the active molecules such as antioxidant, antimicrobial, antidiabetic, enzymes and genes in the plant, which is the first to be reported. The experiments were conducted in a completely randomized design with three replications. In addition, gene expression was compared in the shoot and root parts. Expression profiling was carried out by microarrays. According to the results, the highest chemical components were determined in methanol extract rather than water extract. There was a difference between the obtained components. While the highest antioxidant activity was determined from the methanol extract of plant herbs for DPPH Free Radical Scavenging Activity, antioxidant activity was the same in both methanol and water extracts using the ABTS method. The methanol extract demonstrated stronger anticholinesterase (AChE and BChE) and α-amylase inhibition activity. This study was complemented by the detection of antioxidant activity and some enzyme inhibition activity in the methanol extract. Microarray showed 10,784 genes had significantly different expression in root and shoot. There was a positive effect of methanol extract in respect of different activities compared to the water extract. Gene expression showed that the number of expressed genes in the root was greater than the shoot.


Hypericum lydium Gene expression Anticholinesterase Antimicrobial and antidiabetic activity 







Diabetes mellitus


Food and drug administration


Gas chromatography mass spectrometry




Dimethyl sulfoxide


2,2′-Azino-bis-(3-ethylbenzothiazoline-6-sulphonic acid)




Phosphate-buffered saline


Dinitrosalicylic acid




Author contributions

NE: Execution research project, Data analysis, Manuscript preparation. EU: Experimental design, Data analysis, Manuscript preparation. HAA: Data analysis, proofreading of the article. KS: Experimental design, Data analysis, gene expression analysis. SMS: Data analysis, proofreading of the article, gene expression analysis. DK: Data analysis, gene expression analysis.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

This study was approved by the Ethics Committee of Razi University, Kermanshah, Iran.


  1. 1.
    Ames BN (1983) Dietary carcinogens and anticarcinogens: oxygen radicals and degenerative diseases. Science 221(4617):1256–1264Google Scholar
  2. 2.
    Bhattacharya S (2011) Natural antimutagens: a review. Res J Med Plant 5(2):116–126Google Scholar
  3. 3.
    Boran R, Ugur A (2017) The mutagenic, antimutagenic and antioxidant properties of Hypericum lydium. Pharm Biol 55(1):402–405Google Scholar
  4. 4.
    Halliwell B, Gutteridge JM (2015) Free radicals in biology and medicine. Oxford University Press, OxfordGoogle Scholar
  5. 5.
    Sathuvan M et al (2012) In vitro antioxidant and anticancer potential of bark of Costus pictus D. Don. Asian Pac J Trop Biomed 2(2):S741–S749Google Scholar
  6. 6.
    Ozsoy N et al (2008) Antioxidant activity of Smilax excelsa L. leaf extracts. Food Chem 110(3):571–583Google Scholar
  7. 7.
    Caro AA et al (2019) Antioxidant and pro-oxidant mechanisms of (+) catechin in microsomal CYP2E1-dependent oxidative stress. Toxicol In Vitro 54:1–9Google Scholar
  8. 8.
    Silva S et al (2006) Phenolic compounds and antioxidant activity of Olea europaea L. fruits and leaves. Food Sci Technol Int 12(5):385–395Google Scholar
  9. 9.
    Oreopoulou A, Tsimogiannis D, Oreopoulou V (2019) Extraction of polyphenols from aromatic and medicinal plants: an overview of the methods and the effect of extraction parameters. In: Polyphenols in plants. Elsevier, Amsterdam, pp 243–259Google Scholar
  10. 10.
    Do J-R et al (2004) Antimicrobial and antioxidant activities and phenolic contents in the water extract of medicinal plants. Food Sci Biotechnol 13(5):640–645Google Scholar
  11. 11.
    Albayrak S, Sağdiç O, Aksoy A (2010) Bitkisel ürünlerin ve gıdaların antioksidan kapasitelerinin belirlenmesinde kullanılan yöntemler. Erciyes Üniversitesi Fen Bilimleri Enstitüsü Fen Bilimleri Dergisi 26(4):401–409Google Scholar
  12. 12.
    Soory M (2009) Relevance of nutritional antioxidants in metabolic syndrome, ageing and cancer: potential for therapeutic targeting. Infect Disord Drug Targets 9(4):400–414Google Scholar
  13. 13.
    Abdel-Daim MM et al. (2018) Aging, metabolic, and degenerative disorders: biomedical value of antioxidants. Oxid Med Cell Longev 2018:2098123Google Scholar
  14. 14.
    Amar A et al (2018) Determination of oxidative stress levels and some antioxidant enzyme activities in prostate cancer. Aging Male 1–9Google Scholar
  15. 15.
    Amin F, Bano B (2018) Spectroscopic studies on free radical coalescing antioxidants and brain protein cystatin. J Biomol Struct Dyn 1–11Google Scholar
  16. 16.
    Liguori I et al (2018) Oxidative stress, aging, and diseases. Clin Interv Aging 13:757–772Google Scholar
  17. 17.
    Liu Z et al (2018) Role of ROS and nutritional antioxidants in human diseases. Front Physiol 9:477Google Scholar
  18. 18.
    Crockett SL, Robson NK (2011) Taxonomy and chemotaxonomy of the genus Hypericum. In: Odabaş MS, Çırak C (eds) Hypericum. Medicinal and aromatic plant science and biotechnology, vol 5(Special Issue 1), pp 1Google Scholar
  19. 19.
    Bingol U, Cosge B, Gurbuz B (2011) Hypericum species in flora of Turkey. In: Odabas, MS, Cirak C (eds) Hypericum. Medicinal and aromatic plant science and biotechnology vol 5, pp 86–90Google Scholar
  20. 20.
    Sezik E et al (2001) Traditional medicine in Turkey X. Folk medicine in central Anatolia. J Ethnopharmacol 75(2–3):95–115Google Scholar
  21. 21.
    Baytop T (1999) Therapy with medicinal plants in Turkey (past and present), 2 edn. Istanbul University: Nobel Medicine Publications, Istanbul, pp 312Google Scholar
  22. 22.
    Yeşilada E et al (1995) Traditional medicine in Turkey. V. Folk medicine in the inner Taurus Mountains. J Ethnopharmacol 46(3):133–152Google Scholar
  23. 23.
    Cakir A et al (2003) Isolation and characterization of antioxidant phenolic compounds from the aerial parts of Hypericum hyssopifolium L. by activity-guided fractionation. J Ethnopharmacol 87(1):73–83Google Scholar
  24. 24.
    Öztürk N, Tunçel M, Potoğlu-Erkara İ (2009) Phenolic compounds and antioxidant activities of some Hypericum species: a comparative study with H. perforatum. Pharm Biol 47(2):120–127Google Scholar
  25. 25.
    Russo D et al (2015) Evaluation of antioxidant, antidiabetic and anticholinesterase activities of Smallanthus sonchifolius landraces and correlation with their phytochemical profiles. Int J Mol Sci 16(8):17696–17718Google Scholar
  26. 26.
    López S et al (2002) Acetylcholinesterase inhibitory activity of some Amaryllidaceae alkaloids and Narcissus extracts. Life Sci 71(21):2521–2529Google Scholar
  27. 27.
    Ertaş A et al (2014) Essential oil compositions and anticholinesterase activities of two edible plants Tragopogon latifolius var. angustifolius and Lycopsis orientalis. Nat Prod Res 28(17):1405–1408Google Scholar
  28. 28.
    Aktumsek A et al (2013) Antioxidant potentials and anticholinesterase activities of methanolic and aqueous extracts of three endemic Centaurea L. species. Food Chem Toxicol 55:290–296Google Scholar
  29. 29.
    Ertaş A et al (2014) Antioxidant, anticholinesterase, and antimicrobial activities and fatty acid constituents of Achillea cappadocica Hausskn. et Bornm. Turk J Chem 38(4):592–599Google Scholar
  30. 30.
    Hacıbekiroğlu I, Kolak U (2015) Screening antioxidant and anticholinesterase potential of Iris albicans extracts. Arab J Chem 8(2):264–268Google Scholar
  31. 31.
    Orhan I et al (2007) Antioxidant and anticholinesterase evaluation of selected Turkish Salvia species. Food Chem 103(4):1247–1254Google Scholar
  32. 32.
    Öztürk M et al (2011) In vitro antioxidant, anticholinesterase and antimicrobial activity studies on three Agaricus species with fatty acid compositions and iron contents: a comparative study on the three most edible mushrooms. Food Chem Toxicol 49(6):1353–1360Google Scholar
  33. 33.
    Bhandari MR et al (2008) α-Glucosidase and α-amylase inhibitory activities of Nepalese medicinal herb Pakhanbhed (Bergenia ciliata, Haw.). Food Chem 106(1):247–252Google Scholar
  34. 34.
    El Essawy B, Kandeel F (2019) Pre, peri and posttransplant diabetes mellitus. Curr Opin Nephrol Hypertens 28(1):47–57Google Scholar
  35. 35.
    Lordan S et al (2013) The α-amylase and α-glucosidase inhibitory effects of Irish seaweed extracts. Food Chem 141(3):2170–2176Google Scholar
  36. 36.
    Kim Y-M et al (2005) Inhibitory effect of pine extract on α-glucosidase activity and postprandial hyperglycemia. Nutrition 21(6):756–761Google Scholar
  37. 37.
    Biswas R et al (2017) Tyrosinase inhibitory mechanism of betulinic acid from Dillenia indica. Food Chem 232:689–696Google Scholar
  38. 38.
    Kim JH et al (2017) Tyrosinase inhibitory components from Aloe vera and their antiviral activity. J Enzyme Inhib Med Chem 32(1):78–83Google Scholar
  39. 39.
    Sanni DM, Omotoyinbo OV (2016) Phytochemical screening, tyrosinase inhibitory effects and kinetics of cam wood dye extracts. Adv Biochem 4(2):16–20Google Scholar
  40. 40.
    Muddathir A et al (2017) Anti-tyrosinase, total phenolic content and antioxidant activity of selected Sudanese medicinal plants. South Afr J Bot 109:9–15Google Scholar
  41. 41.
    Neagu E et al (2016) Antioxidant activity, acetylcholinesterase and tyrosinase inhibitory potential of Pulmonaria officinalis and Centarium umbellatum extracts. Saudi J Biol Sci 25(3):578–585Google Scholar
  42. 42.
    Esmaeili F et al (2018) Effects of various glutamine concentrations on gene expression and steviol glycosides accumulation in Stevia rebaudiana Bertoni. Cell Mol Biol 64(2):1–5Google Scholar
  43. 43.
    Fallah F et al (2017) Effect of salinity on gene expression, morphological and biochemical characteristics of Stevia rebaudiana Bertoni under in vitro conditions. Cell Mol Biol 63(7):102–106Google Scholar
  44. 44.
    Ghaheri M et al (2018) Study of gene expression and steviol glycosides accumulation in Stevia rebaudiana Bertoni under various mannitol concentrations. Mol Biol Rep. Google Scholar
  45. 45.
    Tian C et al (2019) A gene expression map of shoot domains reveals regulatory mechanisms. Nat Commun 10(1):141Google Scholar
  46. 46.
    Clarke G et al (2013) High correlation of 2, 2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging, ferric reducing activity potential and total phenolics content indicates redundancy in use of all three assays to screen for antioxidant activity of extracts of plants from the Malaysian rainforest. Antioxidants 2(1):1–10Google Scholar
  47. 47.
    Re R et al (1999) Antioxidant activity applying an improved ABTS radical cation decolorization assay. Free Radic Biol Med 26(9–10):1231–1237Google Scholar
  48. 48.
    Yang H et al (2011) Antioxidant compounds from propolis collected in Anhui, China. Mol 16:3444–3455Google Scholar
  49. 49.
    Ellman GL et al (1961) A new and rapid colorimetric determination of acetylcholinesterase activity. Biochem Pharmacol 7(2):88–95Google Scholar
  50. 50.
    Kumar D et al (2012) Bio-assay guided isolation of α-glucosidase inhibitory constituents from Hibiscus mutabilis leaves. Phytochem Anal 23(5):421–425Google Scholar
  51. 51.
    Kumar D et al (2013) α-Glucosidase and α-amylase inhibitory constituent of Carex baccans: Bio-assay guided isolation and quantification by validated RP-HPLC-DAD. J Funct Foods 5:211–218Google Scholar
  52. 52.
    Jeong SH et al (2009) Tyrosinase inhibitory polyphenols from roots of Morus lhou. J Agric Food Chem 57(4):1195–1203Google Scholar
  53. 53.
    Benjamini Y, Yekutieli D (2001) The control of the false discovery rate in multiple testing under dependency. Ann Stat 29:1165–1188Google Scholar
  54. 54.
    Çlrak C et al (2007) Chemical constituents of some Hypericum species growing in Turkey. J Plant Biol 50(6):632Google Scholar
  55. 55.
    Şerbetçi T et al (2012) Chemical composition of the essential oil and antioxidant activity of methanolic extracts from fruits and flowers of Hypericum lydium Boiss. Ind Crops Prod 36(1):599–606Google Scholar
  56. 56.
    Saddiqe Z, Naeem I, Maimoona A (2010) A review of the antibacterial activity of Hypericum perforatum L. J Ethnopharmacol 131(3):511–521Google Scholar
  57. 57.
    Akbarabadi A et al. (2018) Validation of expression stability of reference genes in response to herbicide stress in wild oat (Avena ludoviciana). Cell Mol Biol 64(4):113–118Google Scholar
  58. 58.
    Akbari F et al (2018) Effect of nitrogen sources on gene expression of Stevia rebaudiana (Bertoni) under in vitro conditions. Cell Mol Biol 64(2):11–16Google Scholar
  59. 59.
    Ghaheri M et al. (2018) Effects of life cycle and leaves location on gene expression and glycoside biosynthesis pathway in Stevia rebaudiana Bertoni. Cell Mol Biol 64(2):17–22Google Scholar
  60. 60.
    Ghaheri M et al (2018) Effect of Stevia rebaudiana Bertoni extract on sexual dysfunction in Streptozotocin-induced diabetic male rats. Cell Mol Biol 64(2):6–10Google Scholar
  61. 61.
    Ghorbani T et al (2017) Effect of sucrose concentrations on Stevia rebaudiana Bertoni tissue culture and gene expression. Cell Mol Biol 63(8):33–37Google Scholar
  62. 62.
    Hashempoor S et al (2018) Effects of different concentrations of mannitol on gene expression in Stevia rebaudiana Bertoni. Cell Mol Biol 64(2):28–31Google Scholar
  63. 63.
    Kahrizi D et al (2018) Investigation of different concentrations of MS media effects on gene expression and steviol glycosides accumulation in Stevia rebaudiana Bertoni. Cell Mol Biol 64(2):23–27Google Scholar
  64. 64.
    Kahrizi D et al (2017) Effect of KH2PO4 on gene expression, morphological and biochemical characteristics of Stevia rebaudiana Bertoni under in vitro conditions. Cell Mol Biol 63(7):107–111Google Scholar

Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  1. 1.Department of Pharmacognosy, Faculty of PharmacyUniversity of SelcukKonyaTurkey
  2. 2.Department of Crop and Animal Production, Sivas Vocational SchoolCumhuriyet UniversitySivasTurkey
  3. 3.Department of Biology, Faculty of EducationCumhuriyet UniversitySivasTurkey
  4. 4.Molecular Genetics LaboratoryResearcher of Zagros Bioidea CompanyKermanshahIran
  5. 5.Department of Agronomy and Plant Breeding, Kermanshah BranchIslamic Azad UniversityKermanshahIran
  6. 6.Agronomy and Plant Breeding DepartmentRazi UniversityKermanshahIran
  7. 7.Medical Biology Research CenterKermanshah University of Medical SciencesKermanshahIran

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