Sodium chloride, colchicine, and 6-Benzylaminopurine can change antioxidant property and phenol content in Hypericum perforatum L.: an in vitro study

Ara MANTEGHI TAFRESHI; Reza MOHAMMADHASSAN

doi:10.14720/aas.2025.121.3.18392

Authors

Ara MANTEGHI TAFRESHI Plant Sciences Department, Amino Techno Gene Private Virtual Laboratory (NGO), Tehran, Iran. https://orcid.org/0000-0002-2190-6130
Reza MOHAMMADHASSAN Plant Sciences Department, Amino Techno Gene Private Virtual Laboratory (NGO), Tehran, Iran. https://orcid.org/0000-0002-0509-4187

DOI:

https://doi.org/10.14720/aas.2025.121.3.18392

Keywords:

Hypericum perforatum, Antioxidant property, Phenol content, Colchicine, Sodium chloride, 6-Benzylaminopurine, Plant tissue culture

Abstract

Hypericum perforatum, a medicinal plant from the Hypericaceae family, is known for its wide range of bioactive properties, including antidepressant, antiviral, antibacterial, and anti-cancer activity. These effects are attributed mainly to its antioxidants and among them phenolic secondary metabolites. This in vitro study aimed to investigate the impact of various concentrations of colchicine (0.05, 0.1, 0.2 mg l^-1), sodium chloride (NaCl; 0.5, 1. 2 mg l^-1), and 6-benzylaminopurine (BAP; 0.25, 0.5, 1 mg l^-1) on antioxidant activity and total phenol content in H. perforatum over three weeks. A factorial experiment was conducted in a completely randomized design with three replications. The results revealed that all treatments significantly enhanced (p < 0.01) antioxidant capacity, phenol content, and morphological characteristics. Notably, the highest levels of antioxidant activity and total phenol content were observed in the third week following treatment with 0.25 mg l^-1 BAP, 2 mg l^-1 NaCl, and 2 mg l^-1 colchicine, separately. These findings suggest that the higher concentrations of NaCl and colchicine, along with a lower concentration of BAP, can effectively enhance the biosynthesis of phenolic compounds and antioxidant activity in H. perforatum.

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Author Biographies

Ara MANTEGHI TAFRESHI, Plant Sciences Department, Amino Techno Gene Private Virtual Laboratory (NGO), Tehran, Iran.
- Plant Sciences Department, Amino Techno Gene Private Virtual Laboratory (NGO), Tehran, Iran.
Reza MOHAMMADHASSAN, Plant Sciences Department, Amino Techno Gene Private Virtual Laboratory (NGO), Tehran, Iran.
- Plant Sciences Department, Amino Techno Gene Private Virtual Laboratory (NGO), Tehran, Iran.

References

Abd El-Latif, F. M., El-Gioushy, S. F., Islam, S. E., & Zakry, T. A. (2018). Impact of papaya seed soaking in different BA, colchicine and EMS solutions on germination, growth and chromosomal behaviour. Asian Journal of Biotechnology and Genetic Engineering, 1(1), 1-17. https://doi.org/10.9734/AJBGE/2018/40538.

Albuquerque, B. R., Heleno, S. A., Oliveira, M. B. P., Barros, L., & Ferreira, I. C. (2021). Phenolic compounds: Current industrial applications, limitations and future challenges. Food & function, 12(1), 14-29. https://doi.org/10.1039/D0FO02324H.

Alexandri, S., Tsaktsira, M., Hatzilazarou, S., Kostas, S., Nianiou-Obeidat, I., Economou, A., ... & Tsoulpha, P. (2023). Selection for sustainable preservation through in vitro propagation of mature Pyrus spinosa genotypes rich in total phenolics and antioxidants. Sustainability, 15(5), 4511. https://doi.org/10.3390/su15054511.

Aremu, A. O., Fawole, O. A., Makunga, N. P., Masondo, N. A., Moyo, M., Buthelezi, N. M., ... & Doležal, K. (2020). Applications of cytokinins in horticultural fruit crops: Trends and future prospects. Biomolecules, 10(9), 1222. https://doi.org/10.3390/biom10091222.

Arumugam, G., Sinniah, U. R., Swamy, M. K., & Lynch, P. T. (2020). Micropropagation and essential oil characterization of Plectranthus amboinicus (Lour.) Sprengel, an aromatic medicinal plant. In Vitro Cellular & Developmental Biology-Plant, 56, 491-503.https://doi.org/10.1007/s11627-020-10056-1.

Barzin, R., Abbaspour, H., Hajkazemian, M., Mahmoudi, A., & Hassan, R. M. (2016). Study of genetic diversity in oat seeds by using SSR molecular markers. International Journal of Advanced Biotechnology and Research, 7(4), 1493-1497. http://www.bipublication.com/barzin2016/74-1493-97.

Budantsev, A. L., Prikhodko, V. A., Varganova, I. V., & Okovityi, S. V. (2021). Biological activity of Hypericum perforatum L.(Hypericaceae): a review. Pharmacy & Pharmacology, 9(1), 17-31. https://doi.org/10.19163/2307-9266-2021-9-1-17-31.

Chatoui, K., Harhar, H., El Kamli, T., & Tabyaoui, M. (2020). Chemical composition and antioxidant capacity of Lepidium sativum seeds from four regions of Morocco. Evidence-based Complementary and Alternative Medicine, 2020, 7302727. https://doi.org/10.1155/2020/7302727.

Çömlekçioğlu, N., & Özden, M. (2020). Effects of colchicine applications and ploidy level on fruit secondary metabolite profiles of goldenberry (Physalis peruviana L.). Applied Ecology & Environmental Research, 18(1), 289-302. http://dx.doi.org/10.15666/aeer/1801_289302.

Cortleven, A., Leuendorf, J. E., Frank, M., Pezzetta, D., Bolt, S., & Schmülling, T. (2019). Cytokinin action in response to abiotic and biotic stresses in plants. Plant, Cell & Environment, 42(3), 998-1018. https://doi.org/10.1111/pce.13494.

Cui, L., Liu, Z., Yin, Y., Zou, Y., Faizan, M., Alam, P., & Yu, F. (2023). Research progress of chromosome doubling and 2 n gametes of ornamental plants. Horticulturae, 9(7), 752. https://doi.org/10.3390/horticulturae9070752.

de Vasconcelos Dias, M., Rodrigues, F. A., de Souza Ribeiro, M., Dambroz, C., Dória, J., & Pasqual, M. (2025). Physiological and morphological responses of Selenicereus species to salt stress in vitro. Plant Cell, Tissue and Organ Culture (PCTOC), 162(2), 26. https://doi.org/10.1007/s11240-025-03082-7.

Delcheh, K. S., Kashefi, B., & Mohammadhassan, R. (2014). A review optimization of tissue culture medium medicinal plant: Thyme. International Journal of Farming and Allied Sciences, 3(9), 1015-1019. https://ijfas.com/wp-content/uploads/2014/10/1015-1019.pdf.

Eng, W. H., & Ho, W. S. (2019). Polyploidization using colchicine in horticultural plants: A review. Scientia Horticulturae, 246, 604-617. https://doi.org/10.3390/horticulturae9070752.

Eng, W. H., Ho, W. S., & Ling, K. H. (2021). Effects of colchicine treatment on morphological variations of Neolamarckia cadamba. International Journal of Agricultural Technology, 17(1), 47-66. https://www.cabidigitallibrary.org/doi/full/10.5555/20210228445.

Etesami, H., Fatemi, H., & Rizwan, M. (2021). Interactions of nanoparticles and salinity stress at physiological, biochemical and molecular levels in plants: A review. Ecotoxicology and Environmental Safety, 225, 112769. https://doi.org/10.1016/j.ecoenv.2021.112769.

Ghasemi-Omran, V. O., Ghorbani, A., & Sajjadi-Otaghsara, S. A. (2021). Melatonin alleviates NaCl-induced damage by regulating ionic homeostasis, antioxidant system, redox homeostasis, and expression of steviol glycosides-related biosynthetic genes in in vitro cultured Stevia rebaudiana Bertoni. In Vitro Cellular & Developmental Biology-Plant, 57(2), 319-331. https://doi.org/10.1007/s11627-021-10161-9.

Giordano, M., Petropoulos, S. A., & Rouphael, Y. (2021). Response and defence mechanisms of vegetable crops against drought, heat and salinity stress. Agriculture, 11(5), 463. https://doi.org/10.3390/agriculture11050463.

Gracheva, I. A., Shchegravina, E. S., Schmalz, H. G., Beletskaya, I. P., & Fedorov, A. Y. (2020). Colchicine alkaloids and synthetic analogues: current progress and perspectives. Journal of Medicinal Chemistry, 63(19), 10618-10651. https://doi.org/10.1021/acs.jmedchem.0c00222.

Gulcin, İ. (2020). Antioxidants and antioxidant methods: An updated overview. Archives of Toxicology, 94(3), 651-715. https://doi.org/10.1007/s00204-020-02689-3.

Gupta, G., Memon, A. G., Pandey, B., Khan, M. S., Iqbal, M. S., & Srivastava, J. K. (2021). Colchicine induced mutation in plant for the assessment of morpho-physiological and biochemical parameter anti-inflammatory activity. The Open Biotechnology Journal, 15(1), 173-182. http://dx.doi.org/10.2174/1874070702115010173.

Hallmark, H. T., & Rashotte, A. M. (2019). Review–cytokinin response factors: responding to more than cytokinin. Plant Science, 289, 110251. https://doi.org/10.1016/j.tplants.2018.10.012.

Hameed, A., Ahmed, M. Z., Hussain, T., Aziz, I., Ahmad, N., Gul, B., & Nielsen, B. L. (2021). Effects of salinity stress on chloroplast structure and function. Cells, 10(8), 2023. https://doi.org/10.3390/cells10082023.

Hao, S., Wang, Y., Yan, Y., Liu, Y., Wang, J., & Chen, S. (2021). A review on plant responses to salt stress and their mechanisms of salt resistance. Horticulturae, 7(6), 132. https://doi.org/10.3390/horticulturae7060132.

Hasanuzzaman, M., Raihan, M. R. H., Masud, A. A. C., Rahman, K., Nowroz, F., Rahman, M., ... & Fujita, M. (2021). Regulation of reactive oxygen species and antioxidant defense in plants under salinity. International Journal of Molecular Sciences, 22(17), 9326. https://doi.org/10.3390/ijms22179326.

Hemmati, N., Cheniany, M., & Ganjeali, A. (2020). Effect of plant growth regulators and explants on callus induction and study of antioxidant potentials and phenolic metabolites in Salvia tebesana Bunge. Botanica Serbica, 44(2), 163-173. https://doi.org/10.2298/BOTSERB2002163H.

Imreova, P., Feruszova, J., Kyzek, S., Bodnarova, K., Zduriencikova, M., Kozics, K., ... & Chalupa, I. (2017). Hyperforin exhibits antigenotoxic activity on human and bacterial cells. Molecules, 22(1), 167. https://doi.org/10.3390/molecules22010167.

Kara, E., Taşkın, H., Shimira, F., Karaköy, T., & Baktemur, G. (2025). Phenotypic responses of Capsicum annuum L. to salinity stress under in vitro conditions. Vegetos, 1-8. https://doi.org/10.1007/s42535-025-01412-w.

Karakas, F. P. (2020). Efficient plant regeneration and callus induction from nodal and hypocotyl explants of goji berry (Lycium barbarum L.) and comparison of phenolic profiles in calli formed under different combinations of plant growth regulators. Plant Physiology and Biochemistry, 146, 384-391. https://doi.org/10.1016/j.plaphy.2019.11.009.

Khan, W. U. D., Tanveer, M., Shaukat, R., Ali, M., & Pirdad, F. (2020). An overview of salinity tolerance mechanism in plants. In M. Hassanuzzaman & M. Tanveer (Eds), Salt and drought stress tolerance in plants: Signaling networks and adaptive mechanisms (1-16). Cham: Springer. https://doi.org/10.1007/978-3-030-40277-8_1.

Kharel, P., Creech, M. R., Nguyen, C. D., Vendrame, W. A., Munoz, P. R., & Huo, H. (2022). Effect of explant type, culture medium, and BAP concentration on in vitro shoot development in highbush blueberry (Vaccinium corymbosum L.) cultivars. In Vitro Cellular & Developmental Biology-Plant, 58(6), 1057-1065.https://doi.org/10.1007/s11627-022-10299-0.

Kozak, D., Parzymies, M., Swistowska, A., Marcinek, B., & Pogroszewska, E. (2021). The influence of growth regulators and explant position on the growth and development of Mandevilla sanderi (Hemsl.) woodson in vitro. Acta Scientiarum Polonorum - Hortorum Cultus, 20(5), 127-138. https://www.cabidigitallibrary.org/doi/full/10.5555/20210502599.

Kulus, D. (2020). Influence of growth regulators on the development, quality, and physiological state of in vitro-propagated Lamprocapnos spectabilis (L.) Fukuhara. In Vitro Cellular & Developmental Biology-Plant, 56(4), 447-457. https://doi.org/10.1007/s11627-020-10064-1.

Kumar, K., Debnath, P., Singh, S., & Kumar, N. (2023). An overview of plant phenolics and their involvement in abiotic stress tolerance. Stresses, 3(3), 570-585. https://doi.org/10.3390/stresses3030040.

Leandro, V. Ã., de Oliveira Guerra, M., & Peters, V. M. (2017). Effect of the extract of Hypericum perforatum on neurodevelopment of regions related to pain control and convulsion. Journal of Medicinal Plants Research, 11(6), 107-117. https://doi.org/10.5897/JMPR2016.6305.

Li, S. M., Zheng, H. X., Zhang, X. S., & Sui, N. (2021). Cytokinins as central regulators during plant growth and stress response. Plant Cell Reports, 40, 271-282.https://doi.org/10.1007/s00299-020-02612-1.

Lima, E. M. F., Winans, S. C., & Pinto, U. M. (2023). Quorum sensing interference by phenolic compounds–A matter of bacterial misunderstanding. Heliyon, 9(7), e17657. https://doi.org/10.1016/j.heliyon.2023.e17657.

Liu, Y., Zhang, M., Meng, Z., Wang, B., & Chen, M. (2020). Research progress on the roles of cytokinin in plant response to stress. International Journal of Molecular Sciences, 21(18), 6574. https://doi.org/10.3390/ijms21186574.

Mangena, P. (2020). Research article in vivo and in vitro application of colchicine on germination and shoot proliferation in soybean [Glycine max (L.) Merr.]. Asian Journal Crop Science, 12, 34-42. https://doi.org/10.3923/ajcs.2020.34.42.

Mangena, P., & Mushadu, P. N. (2023). Colchicine-induced polyploidy in leguminous crops enhances morpho-physiological characteristics for drought stress tolerance. Life, 13(10), 1966. https://doi.org/10.3390/life13101966.

Manjusa, A., & Pradeep, K. (2022). Herbal anthelmintic agents: a narrative review. Journal of Traditional Chinese Medicine, 42(4), 641. https://doi.org/10.19852%2Fj.cnki.jtcm.2022.04.007.

Manzoor, A., Ahmad, T., Bashir, M. A., Hafiz, I. A., & Silvestri, C. (2019). Studies on colchicine induced chromosome doubling for enhancement of quality traits in ornamental plants. Plants, 8(7), 194. https://doi.org/10.3390/plants8070194.

Marawne, H., Mohammadhassan, R., Mohammadalipour, Z., & Ahmadpour, S. (2022). Valerian (Valeriana officinalis) extract inhibits TNF-α and iNOS gene expression in mouse LPS-activated microglial cells. Traditional Medicine Research, 7(5), 47. https://doi.org/10.53388/TMR20220320003.

Mohammadhassan, R., Ferdosi, A., Seifalian, A. M., Seifalian, M., & Malmir, S. (2021). Nanoelicitors application promote antioxidant capacity of Asparagus officinalis (in vitro). Journal of Tropical Life Science, 11(3), 259–265. http://dx.doi.org/10.11594/jtls.11.03.01.

Mok, M. C. (2019). Cytokinins and plant development—an overview. Cytokinins, 155-166. http://dx.doi.org/10.1201/9781351071284-12.

Mujib, A., Aslam, J., & Bansal, Y. (2023). Low colchicine doses improved callus induction, biomass growth, and shoot regeneration in in vitro culture of Dracaena sanderiana Sander ex Mast. Propagation of Ornamental Plants, 23, 81-87. https://www.journal-pop.org/2023_23_3_81-87.html.

Murashige, T., & Skoog, F. (1962). A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiologia Plantarum, 15(3), 473. https://doi.org/10.1111/j.1399-3054.1962.tb08052.x

Nazir, U., Gul, Z., Shah, G. M., & Khan, N. I. (2022). Interaction effect of auxin and cytokinin on in vitro shoot regeneration and rooting of endangered medicinal plant Valeriana jatamansi jones through tissue culture. American Journal of Plant Sciences, 13(2), 223-240. https://doi.org/10.4236/ajps.2022.132014.

Nett, R. S., & Sattely, E. S. (2021). Total biosynthesis of the tubulin-binding alkaloid colchicine. Journal of the American Chemical Society, 143(46), 19454-19465. https://doi.org/10.1021/jacs.1c08659.

Nouri Dashlibroon, H., Khorasaninejad, S., Mousavizadeh, S. J., & Mirjalili, M. H. (2020). Effects of colchicine treatment and polyploidy induction on yield components and some morphological and biochemical characteristics of Lavandula stricta Delile. Iranian Journal of Medicinal and Aromatic Plants Research, 36(4), 572-589. https://doi.org/10.22092/ijmapr.2020.341489.2705.

Osman, N. A. E., Shatnawi, M., Shibli, R., Majdalawi, M., Al Tawaha, A. R. M., & Qudah, T. (2021). Salts induced salinity and in vitro multiplication of Paronychia argentea. Ecological Engineering & Environmental Technology, 22(5), 55-64. http://dx.doi.org/10.12912/27197050/139408.

Oulahal, N., & Degraeve, P. (2022). Phenolic-rich plant extracts with antimicrobial activity: an alternative to food preservatives and biocides? Frontiers in Microbiology, 12, 753518. https://doi.org/10.3389/fmicb.2021.753518.

Parcheta, M., Świsłocka, R., Orzechowska, S., Akimowicz, M., Choińska, R., & Lewandowski, W. (2021). Recent developments in effective antioxidants: The structure and antioxidant properties. Materials, 14(8), 1984. https://doi.org/10.3390/ma14081984.

Ramedani, B., Akhavan, S., Mohammadhassan, R., Tutunchi, S., & Khazaei, A. (2015). Evaluation of DISC1 gene rs3738401 polymorphism in iranian parkinson patients affected by type 2 diabetes. Bulletin of Environment, Pharmacology and Life Sciences, 4, 20-23. https://bepls.com/beplssept2015/5.pdf.

Saleh, B. (2023). GC/MS Analysis of Hypericum perforatum L.(Hypericaceae) Species. Journal of Stress Physiology & Biochemistry, 19(2), 25-33. https://cyberleninka.ru/article/Saleh2023/192-2533.

Salem, J., Hassanein, A., El-Wakil, D. A., & Loutfy, N. (2022). Interaction between growth regulators controls in vitro shoot multiplication in Paulownia and selection of NaCl-tolerant variants. Plants, 11(4), 498. https://doi.org/10.3390/plants11040498.

Shamilov, E. N., Abdullaev, A. S., Shamilli, V. E., & Azizov, I. V. (2019). Antiradiation properties of extracts from Hypericum perforatum L.. Faktori Eksperimental’noi Evolucii Organizmiv, (24), 313-316. https://doi.org/10.7124/FEEO.v24.1121.

Su, Y., Wei, M., Guo, Q., Huang, J., Zhao, K., & Huang, J. (2023). Investigating the relationships between callus browning in Isatis indigotica Fortune, total phenol content, and PPO and POD activities. Plant Cell, Tissue and Organ Culture (PCTOC), 155(1), 175-182.https://doi.org/10.1007/s11240-023-02567-7.

Suryawanshi, M. V., Gujarathi, P. P., Mulla, T., & Bagban, I. (2024). Hypericum perforatum: a comprehensive review on pharmacognosy, preclinical studies, putative molecular mechanism, and clinical studies in neurodegenerative diseases. Naunyn-Schmiedeberg’s Archives of Pharmacology, 1-16. https://doi.org/10.1007/s00210-023-02915-6.

Tariq, S., Wani, S., Rasool, W., Shafi, K., Bhat, M. A., Prabhakar, A., ... & Rather, M. A. (2019). A comprehensive review of the antibacterial, antifungal and antiviral potential of essential oils and their chemical constituents against drug-resistant microbial pathogens. Microbial Pathogenesis, 134, 103580. https://doi.org/10.1016/j.micpath.2019.103580.

Velingkar, V. S., Gupta, G. L., & Hegde, N. B. (2017). A current update on phytochemistry, pharmacology and herb–drug interactions of Hypericum perforatum. Phytochemistry Reviews, 16, 725-744. https://doi.org/10.1007/s11101-017-9503-7.

Verma, S. K., Gantait, S., Mukherjee, E., & Gurel, E. (2022). Enhanced somatic embryogenesis, plant regeneration and total phenolic content estimation in Lycium barbarum L.: a highly nutritive and medicinal plant. Journal of Crop Science and Biotechnology, 25(5), 547-555.https://doi.org/10.1007/s12892-022-00150-8.

Wu, W., Du, K., Kang, X., & Wei, H. (2021). The diverse roles of cytokinins in regulating leaf development. Horticulture Research, 8, 118. https://doi.org/10.1038/s41438-021-00558-3.

Wybouw, B., & De Rybel, B. (2019). Cytokinin–a developing story. Trends in Plant Science, 24(2), 177-185. https://doi.org/10.1016/j.plantsci.2019.110251.

Yuling, L. I., Shaobo, Y. A. N., Xiuhong, M. A. O., Cuiyan, W. A. N. G., Jinna, W. A. N. G., Cuilan, L. I. U., ... & Yanhui, Q. I. A. O. (2022). Polyploidy induction by colchicine in forest trees: Research Progress. Journal of Agriculture, 12(8), 55. https://doi.org/10.11923/j.issn.2095-4050.cjas2021-0077.

Zeb, A. (2020). Concept, mechanism, and applications of phenolic antioxidants in foods. Journal of Food Biochemistry, 44(9), e13394. https://doi.org/10.1111/jfbc.13394.