POTENTIAL OF CHEMICAL ELICITORS TO ENHANCE PRODUCTION SECONDARY METABOLITES IN INDUCED CALLUS FROM HYOSCYAMUS NIGER LEAVES
DOI:
https://doi.org/10.36103/h8r61b57Keywords:
Black henbane, glycosides, HPLC, tropane alkaloidsAbstract
Hyoscyamus niger have the main pharmaceutical tropane alkaloids and glycosides in their tissues. The study aimed to enhance the biosynthesis of these compounds by tissue culture techniques. Callus cultures were established from leaf explants on MS medium. Three chemical salts were added separately in different concentrations to the culture medium to evaluate their effects on callus growth and content of pharmaceutical compounds. The results demonstrate that copper (Cu) and cadmium (Cd) decreased callus weights, while the aluminum (Al) did not exhibit any significant effects, compared to the control treatment. However, the HPLC assay proved that the chemical stress caused beneficial effects in promoting the production of alkaloid and glycoside compounds in callus cultures. Succinctly, the highest concentration of Cu (36 mg/L) caused a significant increasing in the content of hyoscyamine, scopolamine, and hyoscyammoside-A. Whereas, Al and Cd at low concentration (5 mg/L) produced the varying significant increasing in the substances of tropane alkaloids and glycosides, compared to the other treatments and the control. These finding concludes that application of chemical elicitation in certain concentrations offer advantageous visions for developing the conditions that related with biosynthesis of medicinal compounds in H. niger.
Received date: 12/8/2023
Accepted date: 27/12/2023
Published date: 26/1/2026
References
Abed, A. S., Ismail, E. N., Majeed, D. M., and Al-Jibouri, A. M. J. (2023). Increasing amounts of secondary metabolites and medicinal compounds in callus culture of Moringa oleifera (Lam.) using abiotic elicitors. Iraqi Journal of Science, 64(8), 3902–3913. https://doi.org/https://doi.org/10.24996/ijs.2023.64.8.17
Akhtar, M. T., Lu, Z., Ren, S., Zou, H., Noor, I., and Jin, B. (2026). Copper homeostasis: Crosstalk with plant secondary metabolism and stress responses. Plant Science, 362(January). https://doi.org/https://doi.org/10.1016/j.plantsci.2025.112795
Al-Khayri, J. M., and Naik, P. M. (2020). Elicitor-induced production of biomass and pharmaceutical phenolic compounds in cell suspension culture of date palm (Phoenix dactylifera L.). Molecules, 25(4669). https://doi.org/10.3390/molecules25204669
Al-mayahi, A. M. W. (2014). Effect of copper sulphate and cobalt chloride on growth of the in vitro culture tissues for date palm (Phoenix dactylifera L.) cv. Ashgar. American Journal of Agricultural and Biological Sciences, 9(1), 6–18. https://doi.org/10.3844/ajabssp.2014.6.18
Alamholo, M., Katayounchah, R. N., and Soltani, J. (2019). Hyoscyamine and scopolamine production in induced callus and hairy roots of Hyoscyamus niger. Academia Journal of Biotechnology, 7(12), 207–215. https://doi.org/10.15413/ajb.2019.0114
Anjitha, K. S., Sameena, P. P., and Puthur, J. T. (2021). Plant stress functional aspects of plant secondary metabolites in metal stress tolerance and their importance in pharmacology. Plant Stress, 2(100038), 1–14. https://doi.org/https://doi.org/10.1016/j.stress.2021.100038
Aslam, M. M., Okal, E. J., and Waseem, M. (2023). Cadmium toxicity impacts plant growth and plant remediation strategies. Plant Growth Regul, 99, 397–41 2. https://doi.org/https://doi.org/1 0.1 007/s1 0725-022-0091 7-7
Azhar, M., and Mustehasan. (2020). Phytopharmacology of an important Unani drug bazr-ul-banj (Hyoscyamus niger Linn.) – Review. Asian J Pharm Clin Res, 13(9), 28–32. https://doi.org/https://doi.org/10.22159/ajpcr.2020.v13i9.38166
Bakhtiari, M. A., and Golkar, P. (2022). The effects of callus elicitation on lepidine, phenolic content, and antioxidant activity of Lepidium sativum L.: chitosan and gibberellic acid. Journal of Plant Growth Regulation, 41, 1 1 48–1 1 60. https://doi.org/https://doi.org/10.1007/s00344-021-10368-5
Chauhan, D. K., Yadav, V., Vaculík, M., Gassmann, W., Pike, S., and Arif, N. (2021). Aluminum toxicity and aluminum stress- induced physiological tolerance responses in higher plants. Critical Reviews in Biotechnology, 41(5). https://doi.org/https://doi.org/10.1080/07388551.2021.1874282
Duncan, D. B. (1955). Multiple range and multiple F tests. Biometrics, 11(1), 1–42.
Giglou, R. H., Giglou, M. T., Sobhanizadeh, A., and Szumny, A. (2025). Enhancing the accumulation of scopolamine and atropine in Hyoscyamus niger L. callus by LED light and glycine-a cheap method. BMC Plant Biology, 25(1162), 1–14. https://doi.org/https://doi.org/10.1186/s12870-025-07062-z
Hemmati, N., Cheniany, M., and 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/https://doi.org/10.2298/BOTSERB2002163H
Kamada, H., Okamura, N., Satake, M., Harada, H., and Shimomura, K. (1986). Alkaloid production by hairy root cultures in Atropa belladonna. Plant Cell Reports, 5(4), 239–242. https://doi.org/10.1007/BF00269811
Karimi, F., and Khataee, E. (2011). Aluminum elicits tropane alkaloid production and antioxidant system activity in micro propagated Datura innoxia plantlets. Acta Physiologiae Plantarum, 34(3). https://doi.org/10.1007/s11738-011-0900-z
Khalaf, M. A., and Taha, A. J. (2023). Photoperiod and UV light influence secondary metabolites of Hyoscyamus niger callus induced from leaves. Journal of Biotechnology Research Center, 17(2), 16–26. https://doi.org/https://doi.org/10.24126/jobrc.2023.17.2.684
Khan, H., Khan, T., Ahmad, N., Zaman, G., Khan, T., Ahmad, W., Batool, S., Hussain, Z., Drouet, S., Hano, C., and Abbasi, B. H. (2021). Chemical elicitors-induced variation in cellular biomass, biosynthesis of secondary cell products, and antioxidant system in callus cultures of Fagonia indica. Molecules, 26(6340). https://doi.org/10.3390/molecules26216340
Liu, J., Al-namazi, A. A., and Wan, J. S. H. (2024). Plants generally suffer less enemy damage and are more defended in a copper mine than in a closely adjacent site. South African Journal of Botany, 171(August), 164–172. https://doi.org/10.1016/j.sajb.2024.06.010
Mohammed, Z. H. (2019). Determination of nicotine extracted from eggplant and green pepper by HPLC. Baghdad Science Journal, 16(1), 61–67. https://doi.org/10.21123/bsj.2019.16.1.0061
Murashige, T., and Skoog, F. (1962). A revised medium for rapid growth and bio assays with Tobacco tissue cultures. Physiologia Plantarum, 15, 473–497. https://doi.org/https://doi.org/10.1111/j.1399-3054.1962.tb08052.x
Naik, P. M., and Al-Khayri, J. M. (2016). Impact of abiotic elicitors on in vitro production of plant secondary metabolites: A review. Journal of Advanced Research in Biotechnology, 1(2), 1–7. https://doi.org/10.15226/2475-4714/1/2/00102
Ofoe, R., Thomas, R. H., Asiedu, S. K., Wang-Pruski, G., Fofana, B., and Abbey, Lord. (2023). Aluminum in plant: Benefits, toxicity and tolerance mechanisms. Frontiers in Plant Science, 13. https://doi.org/10.3389/fpls.2022.1085998
Panda, S. K., Gupta, D., Patel, M., Vyver, C. Van Der, and Koyama, H. (2024). Functionality of reactive oxygen species (ROS) in plants: Toxicity and control in Poaceae crops exposed to abiotic stress. Plants, 13(2071). https://doi.org/10.3390/plants13152071
Prażak, R., and Molas, J. (2015). Effect of copper concentration on micropropagation and accumulation of some metals in the Dendrobium kingianum Bidwill Orchid. Journal of Elementology, 20(3), 693–703. https://doi.org/10.5601/jelem.2014.19.4.748
Rai, A., and Han, S. (2022). Critical review on key approaches to enhance synthesis and production of Steviol glycosides: A blueprint for zero-calorie sweetener. Applied Sciences, 12(8640). https://doi.org/https://doi.org/10.3390/app12178640
Rajput, V., Rani, K. S., & Patan, P. (2024). A review on exploring the medicinal value of Zingiber officinale and Hyoscyamus niger to prevent or overcome seasickness. Journal of Population Therapeutics and Clinical Pharmacology, 31(11), 1140–1150. https://doi.org/https://doi.org/10.53555/1pszpb02
Rao, K., Chodisett, B., Gandi, S., Giri, A., and Kishor, P. B. K. (2021). Cadmium chloride elicitation of Abutilon indicum cell suspension cultures for enhanced stigmasterol production. Plant Biosystems - An International Journal Dealing with All Aspects of Plant Biology. 156(3), 613–618. https://doi.org/https://doi.org/10.1080/11263504.2021.1891151
Salih, M. I., and Al Dabagh, F. M. K. (2021). Comparative analysis of some phenolic acids of in vitro and in vivo grown plant leaves of Salvia hispanica. Iraqi Journal of Agricultural Sciences, 52(1), 189–195. https://doi.org/10.36103/IJAS.V52I1.1250
Senekoviˇ, J. (2025). Copper sulfate elicitation effect on biomass production, phenolic compounds accumulation, and antioxidant activity of Morus nigra L. stem node culture. Plants, 14(5) (766). https://doi.org/https://doi.org/10.3390/plants14050766
Sidik, N. J., Agha, H. M., Alkamil, A. A., Alsayadi, M. M. S., and Mohammed, A. A. (2024). A mini review of plant tissue culture: The role of media optimization, growth regulators in modern agriculture, callus induction and the applications. AUIQ Complementary Biological System, 1(2024), 96–109. https://doi.org/10.70176/3007-973X.1019
Sivanandhan, G., Selvaraj, N., Ganapathi, A., and Manickavasagam, M. (2014). Enhanced biosynthesis of withanolides by elicitation and precursor feeding in cell suspension culture of Withania somnifera (L.) Dunal in shake-flask culture and bioreactor. PLoS ONE, 9(8). https://doi.org/10.1371/journal.pone.0104005
Soni, S., Chawra, H. S., Sharma, R. K., and Garg, R. (2018). Development of drying protocol for Withania somnifera roots. Asian Journal of Pharmaceutical and Clinical Research, 11(10), 86–92. https://doi.org/10.22159/ajpcr.2018.v11i10.28402
Surchy, S. M., Alzebari, I. N., Saeed, S. M., and Essa, I. M. (2025). Callus culture as a tool for producing phytochemicals in Glycyrrhiza glabra: A medicinal plant study. Qalaai Zanist Scientific Journal, 10(3), 1272–1293. https://doi.org/10.25212/lfu.qzj.10.3.49
Trettel, J. R., Gazim, Z. C., Gonçalves, J. E., Stracieri, J., and Magalhães, H. M. (2018). Effects of copper sulphate (CuSO4) elicitation on the chemical constitution of volatile compounds and the in vitro development of Basil. Scientia Horticulturae, 234(19–26). https://doi.org/10.1016/j.scienta.2018.01.062
Vats, S. (2018). Biotic and abiotic stress tolerance in plants. Springer Nature Singapore Pte Ltd. https://doi.org/10.1007/978-981-10-9029-5
Zafar, N., Mujib, A., Ali, M., Tonk, D., and Gulzar, B. (2017). Aluminum chloride elicitation (amendment) improves callus biomass growth and reserpine yield in Rauvolfia serpentina leaf callus. Plant Cell, Tissue and Organ Culture (PCTOC), 130, 357–368. https://doi.org/10.1007/s11240-017-1230-7
Zafar, N., Mujib, A., Ali, M., Tonk, D., Gulzar, B., Malik, M. Q., Mamgain, J., and Sayeed, R. (2020). Cadmium chloride (CdCl2) elicitation improves reserpine and ajmalicine yield in Rauvolfia serpentina as revealed by high‑performance thin‑layer chromatography (HPTLC). 3 Biotech, 10(344). https://doi.org/https://doi.org/10.1007/s13205-020-02339-6
Zandalinas, S. I., Fritschi, F. B., and Mittler, R. (2020). Signal transduction networks during stress combination. Journal of Experimental Botany, 71(5), 1734–1741. https://doi.org/10.1093/jxb/erz486
Zarad, M. M., Toaima, N. M., Refaey, K. A., Atta, R. F., and Elateeq, A. A. (2021). Copper sulfate and Cobalt chloride effect on total phenolics accumulation and antioxidant activity of Artemisia annua L. callus cultures. Al-Azhar Journal of Agricultural Research, 46(2), 26–40. https://doi.org/10.21608/ajar.2021.245610
Zheng, C., Chen, J., Wang, X., and Li, P. (2025). Reactive oxygen species in plants: Metabolism, signaling, and oxidative modifications. Antioxidants, 14(617), 1–17. https://doi.org/10.3390/antiox14060617
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