Psychoactive property of Tasmannia piperita leaf extract in zebrafish by behavioral approach
DOI:
https://doi.org/10.53992/njns.v11i3.342Keywords:
Tasmannia piperita, Psychoactive Properties, Zebrafish, Behavioral Assay, Phytochemical AnalysisAbstract
This study evaluates the psychoactive properties of Tasmannia piperita leaf extract using zebrafish (Danio rerio) as a behavioral model through behavioral assays. T. piperita crude leaf extract was processed via ethanolic extraction with characterization performed using spectrophotometry. Relatively, TAC, TPC, and TFC assays revealed that the extract contained high levels of phenolic compounds (3,380 µg GAE/g), moderate alkaloid content (1,010 µg AE/g), and low flavonoid concentration (270 µg QE/g). Adult zebrafish were exposed to 30, 60, and 120 mg/L of plant extract concentrations, alongside negative and diazepam-treated (0.16 mg/L) controls, then subjected to mirror-biting, novel tank, and shoaling tests. Principal Component Analysis (PCA) demonstrated behavioral variation associated with locomotion, exploratory activity, and social cohesion across treatment groups. In the mirror-biting test, the 60 mg/L treatment significantly reduced mirror-zone engagement, suggesting suppression of aggressive or socially motivated behavior. In the novel tank test, the 120 mg/L treatment significantly decreased upper-zone entries, indicating increased anxiety-like bottom-dwelling behavior without affecting general locomotion. Shoaling behavior showed no statistically significant differences among treatments; however, the 120 mg/L concentration caused complete mortality, suggesting group-related acute toxicity. The study highlights the dose-dependent psychoactive and behavioral effects of T. piperita in zebrafish, with moderate concentrations producing behavioral modulation and higher concentrations demonstrating potential toxicological effects which provide preliminary insights into the psychoactive influence of the plant extract.
References
Dapar, M. L. G., Alejandro, G. J. D., Meve, U., & Liede-Schumann, S. (2020). Quantitative ethnopharmacological documentation and molecular confirmation of medicinal plants used by the Manobo tribe of Agusan del Sur, Philippines. Journal of Ethnobiology and Ethnomedicine, 16(1), Article 34.
Banua, A. S. (2022). Medicinal plants utilized by the traditional healers and their patients in Bicol Region, Philippines. Bicol University (BU) R&D Journal, 25, 1–15.
Cordero, C. S., Meve, U., & Alejandro, G. J. D. (2022). Ethnobotanical documentation of medicinal plants used by the Indigenous Panay Bukidnon in Lambunao, Iloilo, Philippines. Frontiers in Pharmacology, 12, Article 790567.
Andalan, J. R., Mondejar, A. J. S., Sumaya, N. H. N., Guihawan, J. Q., Madamba, M. R. S. B., Tabelin, C. B., Guilingen, D., Paglinawan, F. C., Maulas, K. M., Arquisal, I., Beltran, A. B., Orbecido, A. H., Promentilla, M. A., Alonzo, D., Pisda, P. F., Ananayo, A., Suelto, M., Dalona, I. M., Resabal, V. J., & Villacorte-Tabelin, M. (2024). Ethnobotanical survey of medicinal and ritual plants utilized by the indigenous communities of Benguet province, Philippines. Tropical Medicine and Health, 52(1), Article 29.
Bersamin, A. T., Tayaben, J. L., Balangcod, K. D., Balangcod, A. K. D., Cendana, A. C., Dom-Ogen, E. T., Licnachan, L. O. C., Siadto, B., Wong, F. M., & Balangcod, T. D. (2020). Utilization of plant resources among the Kankanaeys in Kibungan, Benguet Province, Philippines. Biodiversitas Journal of Biological Diversity, 22(1), 384–398.
Letchuman, S., Madhuranga, H. D., Kaushalya, M., Premarathna, A. D., & Saravanan, M. (2024). Alkaloids unveiled: A comprehensive analysis of novel therapeutic properties, mechanisms, and plant-based innovations. Intelligent Pharmacy, 2(4), 512–530.
Dey, P., Kundu, A., Kumar, A., Gupta, M., Lee, B. M., Bhakta, T., Dash, S., & Kim, H. S. (2020). Analysis of alkaloids (indole alkaloids, isoquinoline alkaloids, tropane alkaloids). In Dietary Phytochemicals and Microbes (pp. 505–567). Elsevier.
Heinrich, M., Mah, J., & Amirkia, V. (2021). Alkaloids used as medicines: Structural phytochemistry meets biodiversity - An update and forward look. Molecules, 26(7), Article 1836.
Marileo, A. M., Gavilán, J., Martín, V. P. S., Lara, C. O., Sazo, A., Muñoz-Montesino, C., Castro, P. A., Burgos, C. F., Leiva-Salcedo, E., Aguayo, L. G., Moraga-Cid, G., Fuentealba, J., & Yévenes, G. E. (2023). Modulation of GABAA receptors and of GABAergic synapses by the natural alkaloid gelsemine. Frontiers in Molecular Neuroscience, 15, Article 1083189.
Vezikov, L. V., & Simpson, M. (2023). Plant alkaloids toxicity. In StatPearls. StatPearls Publishing.
Zandavar, H., & Babazad, M. A. (2023). Secondary metabolites: Alkaloids and flavonoids in medicinal plants. In Secondary Metabolites - Sources and Applications. IntechOpen.
Rajput, A., Sharma, R., & Bharti, R. (2021). Pharmacological activities and toxicities of alkaloids on human health. Materials Today: Proceedings, 48, 1407–1415.
Dohárszky, A., Vági, E. M., Könczöl, Á., Simon, A., Várnagy, E., Muratov, M., Steiger, K. I., Várnai, B., Béni, S., Riethmüller, E., & Fejős, I. (2024). Kratom alkaloids: A blood–brain barrier specific membrane permeability assay-guided isolation and cyclodextrin complexation study. Molecules, 29(22), Article 5302.
Velasquez-Jiménez, D., Corella-Salazar, D., Martinez, B. S. Z., & Dominguez-Avila, J. A. (2021). Phenolic compounds that cross the blood-brain barrier exert positive health effects as central nervous system antioxidants. Food & Function, 12(21), 10300–10318.
Kumar, N., & Goel, N. (2019). Phenolic acids: Natural versatile molecules with promising therapeutic applications. Biotechnology Reports, 24, e00370.
Cordeiro, M. L. S., Martins, V. G. Q. A., Silva, A. P., Rocha, H. A. O., Rachetti, V. P. S., & Scortecci, K. C. (2022). Phenolic acids as antidepressant agents. Nutrients, 14(20), Article 4309.
Fiore, M., Terracina, S., & Ferraguti, G. (2025). Brain neurotrophins and plant polyphenols: A powerful connection. Molecules, 30(1), Article 265.
Etukudo, E. M., Usman, I., Oviosun, A., Ojiakor, V. O., Makena, W., Owembabazi, E., Aja, P. M., Vivalya, B. M. N., Archibong, V. B., & Anyanwu, E. (2025). Exploring the neuroprotective potentials of flavonoid metabolites in Syzygium aromaticum: A review with in-silico insight to therapeutic potential. Journal of Experimental Pharmacology, 17, 587–611.
Nicolucci, C., Padovani, M., De Castro Rodrigues, F., Fritsch, L. N., Santos, A. C., Priolli, D. G., & Sciani, J. M. (2023). Flavonoids: The use in mental health and related diseases. Natural Product Research, 38(23), 4223–4233.
Nájera-Maldonado, J., Salazar, R., Alvarez-Fitz, P., Acevedo-Quiroz, M., Flores-Alfaro, E., Hernandez-Sotelo, E., & Espinoza-Rojo, M. (2024). Phenolic compounds of therapeutic interest in neuroprotection. Journal of Xenobiotics, 14(1), 180–205.
Gopalakrishna, R., Aguilar, J., Lee, E., & Mack, W. J. (2024). Neuroprotection by resveratrol-glucuronide and quercetin-glucuronide via binding to polyphenol- and glycosaminoglycan-binding sites in the laminin receptor. Neural Regeneration Research, 19(11), 2465–2472.
Sultanbawa, Y. (2016). Tasmanian pepper leaf (Tasmannia lanceolata) oils. In Essential Oils in Food Preservation, Flavor and Safety (pp. 819–823). Elsevier.
Shalom, J., & Cock, I. E. (2023). Tasmannia lanceolata (Poir.) A.C.Sm. berry and leaf extracts inhibit proliferation and induce apoptosis in selected human carcinoma cell lines. Pharmacognosy Communications, 13(3), 126–140.
Rayan, P., Matthews, B., Donnell, P. M., & Cock, I. E. (2016). Phytochemical analysis of Tasmannia lanceolata extracts and inhibition of Giardia duodenalis proliferation. Pharmacognosy Journal, 8(3), 291–299.
Bahekar, S., & Kale, R. (2013). Herbal plants used for the treatment of malaria: A literature review. Journal of Pharmacognosy and Phytochemistry, 1(6), 141–146.
Pelser, P. B., Barcelona, J. F., &
Nickrent, D. L. (Eds.). (2024). Winteraceae. Co's Digital Flora of the Philippines.
Padio, L. P., & Villaseñor, I. M. (2004). Anti-spasmodic constituents from Drimys piperita Hook. f. leaves. Philippine Journal of Science, 133(1), 17–21.
Zaragoza, E. G. C. (2019). Organic fungicide developed from Tasmannia piperita (Hook.f.) Miers. Philippine Council for Agriculture, Aquatic and Natural Resources Research and Development (DOST-PCAARRD).
Balangcod, T. D., & Balangcod, A. K. D. (2011). Ethnomedical knowledge of plants and healthcare practices among the Kalanguya tribe in Tinoc, Ifugao, Luzon, Philippines. Indian Journal of Traditional Knowledge, 10(2), 227–238.
Taguiling, M. L. G., & Villena, C. (2017). Phytochemical, antimicrobial and sensory qualities of herbal tea made from Sarcandra glabra and Drimys piperita. Proceedings of the 7th International Conference on Chemical, Agricultural, Biological and Environmental Sciences, 55–66.
Amoroso, V., Mendez, R., Junio, H., Emerson, R. J., Molino, J., Pescadero, I., & Villalobos, A. (2021). Characterization of a natural fungicide from an indigenous plant Tasmannia piperita (Hook.f.) Miers extract: Stability, metabolomics, and in silico studies. The Philippine Journal of Science, 150(2), 395–408.
Kolling, G. J., Stivanin, S. C. B., Gabbi, A. M., Machado, F. S., Ferreira, A. L., Campos, M. M., Tomich, T. R., Cunha, C. S., Dill, S. W., Pereira, L. G. R., & Fischer, V. (2022). Behavior of Holstein and Holstein-Gyr lactating cows supplemented with oregano and green tea extracts. Journal of Veterinary Behavior, 49, 75–79.
Clemen-Pascual, L. M., Macahig, R. A. S., & Rojas, N. R. L. (2022). Comparative toxicity, phytochemistry, and use of 53 Philippine medicinal plants. Toxicology Reports, 9, 22–35.
Neelkantan, N., Mikhaylova, A., Stewart, A. M., Arnold, R., Gjeloshi, V., Kondaveeti, D., Poudel, M. K., & Kalueff, A. V. (2013). Perspectives on zebrafish models of hallucinogenic drugs and related psychotropic compounds. ACS Chemical Neuroscience, 4(8), 1137–1150.
Cueto-Escobedo, J., German-Ponciano, L. J., Guillén-Ruiz, G., Soria-Fregozo, C., & Herrera-Huerta, E. V. (2022). Zebrafish as a useful tool in the research of natural products with potential anxiolytic effects. Frontiers in Behavioral Neuroscience, 15, Article 795285.
Syed, O. A., Tsang, B., & Gerlai, R. (2023). The zebrafish for preclinical psilocybin research. Neuroscience & Biobehavioral Reviews, 153, Article 105381.
Hipol, R. L., Wayas, H., Bacuyag, F. M., Daquigan, M., Hipol, R., & Cabanlong, J. (2024). Phytochemical, nutraceutical and pharmacological aspects of the Philippine native Acalypha angatensis Blanco. Indonesian Journal of Pharmacy, 35(3), 321–331.
Shamsa, F., Monsef, H., Ghamooshi, R., & Verdian-Rizi, M. (2008). Spectrophotometric determination of total alkaloids in some Iranian medicinal plants. Thai Journal of Pharmaceutical Sciences, 32(1), 17–20.
Alnajar, Z. A., Abdulla, M. A., Ali, H. M., Alshawsh, M. A., & Hadi, A. H. (2012). Acute toxicity evaluation, antibacterial, antioxidant and immunomodulatory effects of Melastoma malabathricum. Molecules, 17(3), 3547–3559.
Al-Dhabi, N. A., Ponmurugan, K., & Maran Jeganathan, P. (2017). Development and validation of ultrasound-assisted solid-liquid extraction of phenolic compounds from waste spent coffee grounds. Ultrasonics Sonochemistry, 34, 206–213.
Parker, M. O., Millington, M. E., Combe, F. J., & Brennan, C. H. (2012). Housing conditions differentially affect physiological and behavioural stress responses of zebrafish, as well as the response to anxiolytics. PLoS ONE, 7(4), e34992.
Siregar, P., Audira, G., Castillo, A. L., Roldan, M. J. M., Suryanto, M. E., Liu, R., Lin, Y., Lai, Y., & Hsiao, C. (2022). Comparison of the psychoactive activity of four primary Areca nut alkaloids in zebrafish by behavioral approach and molecular docking. Biomedicine & Pharmacotherapy, 155, Article 113809.
Dametto, F. S., Fior, D., Idalencio, R., Rosa, J. G. S., Fagundes, M., Marqueze, A., Barreto, R. E., Piato, A., & Barcellos, L. J. (2018). Feeding regimen modulates zebrafish behavior. PeerJ, 6, e5343.
Shishis, S., Tsang, B., Ren, G. J., & Gerlai, R. (2023). Effects of different handling methods on the behavior of adult zebrafish. Physiology & Behavior, 262, Article 114106.
Gebauer, D. L., Pagnussat, N., Piato, Â. L., Schaefer, I. C., Bonan, C. D., & Lara, D. R. (2011). Effects of anxiolytics in zebrafish: Similarities and differences between benzodiazepines, buspirone and ethanol. Pharmacology Biochemistry and Behavior, 99(3), 480–486.
Audira, G., Sampurna, B. P., Juniardi, S., Liang, S., Lai, Y., & Hsiao, C. (2018). A versatile setup for measuring multiple behavior endpoints in zebrafish. Inventions, 3(4), Article 75.
Pham, M., Raymond, J., Hester, J., Kyzar, E., Gaikwad, S., Bruce, I., Fryar, C., Chanin, S., Enriquez, J., Bagawandoss, S., Zapolsky, I., Green, J., Stewart, A. M., Robison, B. D., & Kalueff, A. V. (2012). Assessing social behavior phenotypes in adult zebrafish: Shoaling, social preference, and mirror biting tests. In Neuromethods (pp. 231–246). Humana Press.
Lim, C. J., Platt, B., Janhunen, S. K., & Riedel, G. (2023). Comparison of automated video tracking systems in the open field test: ANY-maze versus EthoVision XT. Journal of Neuroscience Methods, 397, Article 109940.
Haghani, S., Karia, M., Cheng, R., & Mathuru, A. S. (2019). An automated assay system to study novel tank induced anxiety. Frontiers in Behavioral Neuroscience, 13, Article 247.
Shinkai, T., Nakai, M., Takeshita, U., Morita, K., & Shimada, Y. (2025). Refinement of the novel tank diving test: Toward standardized and robust analysis of anxiety-like behavior in zebrafish. Frontiers in Behavioral Neuroscience, 19, Article 1624277.
Johnson, A., Loh, E., Verbitsky, R., Slessor, J., Franczak, B. C., Schalomon, M., & Hamilton, T. J. (2023). Examining behavioural test sensitivity and locomotor proxies of anxiety-like behaviour in zebrafish. Scientific Reports, 13(1), Article 3958.
Canzian, J., Fontana, B. D., Quadros, V. A., & Rosemberg, D. B. (2017). Conspecific alarm substance differently alters group behavior of zebrafish populations: Putative involvement of cholinergic and purinergic signaling in anxiety- and fear-like responses. Behavioural Brain Research, 320, 255–263.
Green, J., Collins, C., Kyzar, E. J., Pham, M., Roth, A., Gaikwad, S., Cachat, J., Stewart, A. M., Landsman, S., Grieco, F., Tegelenbosch, R., Noldus, L. P., & Kalueff, A. V. (2012). Automated high-throughput neurophenotyping of zebrafish social behavior. Journal of Neuroscience Methods, 210(2), 266–271.
Schmidel, A. J., Assmann, K. L., Werlang, C. C., Bertoncello, K. T., Francescon, F., Rambo, C. L., Beltrame, G. M., Calegari, D., Batista, C. B., Blaser, R. E., Júnior, W. A. R., Conterato, G. M., Piato, A. L., Zanatta, L., Magro, J. D., & Rosemberg, D. B. (2014). Subchronic atrazine exposure changes defensive behaviour profile and disrupts brain acetylcholinesterase activity of zebrafish. Neurotoxicology and Teratology, 44, 62–69.
Pérez-Escudero, A., Vicente-Page, J., Hinz, R. C., Arganda, S., & De Polavieja, G. G. (2014). idTracker: Tracking individuals in a group by automatic identification of unmarked animals. Nature Methods, 11(7), 743–748.
Lieschke, G. J., & Currie, P. D. (2007). Animal models of human disease: Zebrafish swim into view. Nature Reviews Genetics, 8(5), 353–367.
Li, H., Zhang, Q., Jiang, Q., Sa, K., Liang, J., Sun, D., & Chen, L. (2023). Research progress of plant-derived natural alkaloids in central nervous system diseases. Phytotherapy Research, 37(11), 4885–4907.
Matić, P., Sabljić, M., & Jakobek, L. (2017). Validation of spectrophotometric methods for the determination of total polyphenol and total flavonoid content. Journal of AOAC International, 100(6), 1795–1803.
Pękal, A., & Pyrzynska, K. (2014). Evaluation of aluminium complexation reaction for flavonoid content assay. Food Analytical Methods, 7(9), 1776–1782.
Sakulnarmrat, K., & Konczak, I. (2012). Composition of native Australian herbs polyphenolic-rich fractions and in vitro inhibitory activities against key enzymes relevant to metabolic syndrome. Food Chemistry, 134(2), 1011–1019.
Norton, W., & Bally-Cuif, L. (2010). Adult zebrafish as a model organism for behavioural genetics. BMC Neuroscience, 11(1), Article 90.
Kalueff, A. V., Stewart, A. M., &
Gerlai, R. (2014). Zebrafish as an emerging model for studying complex brain disorders. Trends in Pharmacological Sciences, 35(2), 63–75.
Stewart, A., Wu, N., Cachat, J., Hart, P., Gaikwad, S., Wong, K., Utterback, E., Gilder, T., Kyzar, E., Newman, A., Carlos, D., Chang, K., Hook, M., Rhymes, C., Caffery, M., Greenberg, M., Zadina, J., & Kalueff, A. V. (2011). Pharmacological modulation of anxiety-like phenotypes in adult zebrafish behavioral models. Progress in Neuro-Psychopharmacology and Biological Psychiatry, 35(6), 1421–1431.
Rowson, Z., Setzer, R. W., Padilla, S., Friedman, K. P., & Carstens, K. E. (2025). A data analysis approach for locomotor behavioral responses in developmentally exposed zebrafish. NAM Journal, 1, Article 100033.
Cachat, J., Stewart, A., Grossman, L., Gaikwad, S., Kadri, F., Chung, K. M., Wu, N., Wong, K., Roy, S., Suciu, C., Goodspeed, J., Elegante, M., Bartels, B., Elkhayat, S., Tien, D., Tan, J., Denmark, A., Gilder, T., Kyzar, E., ... Kalueff, A. V. (2010). Measuring behavioral and endocrine responses to novelty stress in adult zebrafish. Nature Protocols, 5(11), 1786–1799.
Tudorache, C., Schaaf, M. J. M., & Slabbekoorn, H. (2013). Covariation between behaviour and physiology indicators of coping style in zebrafish (Danio rerio). Journal of Endocrinology, 219(3), 251–258.
Tran, S., & Gerlai, R. (2013). Individual differences in activity levels in zebrafish (Danio rerio). Behavioural Brain Research, 257, 224–229.
Egan, R. J., Bergner, C. L., Hart, P. C., Cachat, J. M., Canavello, P. R., Elegante, M. F., Elkhayat, S. I., Bartels, B. K., Tien, A. K., Tien, D. H., Mohnot, S., Beeson, E., Glasgow, E., Amri, H., Zukowska, Z., & Kalueff, A. V. (2009). Understanding behavioral and physiological phenotypes of stress and anxiety in zebrafish. Behavioural Brain Research, 205(1), 38–44.
Kalueff, A. V., Gebhardt, M., Stewart, A. M., Cachat, J. M., Brimmer, M., Chawla, J. S., Craddock, C., Kyzar, E. J., Roth, A., Landsman, S., Gaikwad, S., Robinson, K., Baatrup, E., Tierney, K., Shamchuk, A., Norton, W., Miller, N., Nicolson, T., Braubach, O., ... Bally-Cuif, L. (2013). Towards a comprehensive catalog of zebrafish behavior 1.0 and beyond. Zebrafish, 10(1), 70–86.
Facciol, A., & Gerlai, R. (2020). Zebrafish shoaling, its behavioral and neurobiological mechanisms, and its alteration by embryonic alcohol exposure: A review. Frontiers in Behavioral Neuroscience, 14, Article 572175.
Sivaraman, A., Nandakumar, R., & Ramachandran, B. (2022). Conspecific identity determines interactive space area in zebrafish shoal. ACS Omega, 7(42), 37351–37358.
Way, G. P., Southwell, M., & McRobert, S. P. (2016). Boldness, aggression, and shoaling assays for zebrafish behavioral syndromes. Journal of Visualized Experiments, (114), e54049.
Hamilton, T. J., Krook, J., Szaszkiewicz, J., & Burggren, W. (2021). Shoaling, boldness, anxiety-like behavior and locomotion in zebrafish (Danio rerio) are altered by acute benzo[a]pyrene exposure. Science of the Total Environment, 774, Article 145702.
Rosa, L. V., Costa, F. V., Canzian, J., Borba, J. V., Quadros, V. A., & Rosemberg, D. B. (2020). Three- and bi-dimensional analyses of the shoaling behavior in zebrafish: Influence of modulators of anxiety-like responses. Progress in Neuro-Psycho-pharmacology and Biological Psychiatry, 102, Article 109957.
Xavier, J., & Kripasana, K. (2020).
Acute toxicity of leaf extracts of Enydra fluctuans Lour in zebrafish (Danio rerio Hamilton). Scientifica, 2020, Article 3965376.
Kyzar, E. J., Collins, C., Gaikwad, S., Green, J., Roth, A., Monnig, L., El-Ounsi, m., Davis, A., Freeman, A., Capezio, N., Stewart, A. M., & Kalueff, A. V. (2012). Effects of hallucinogenic agents mescaline and phencyclidine on zebrafish behavior and physiology. Progress in Neuro-Psycho-pharmacology and Biological Psychiatry, 37(1), 194–202.
Miller, A., Panneerselvam, J., & Liu, L. (2022). A review of regression and classification techniques for analysis of common and rare variants and gene-environmental factors. Neuro-computing, 489, 466–485.
Román, A., Vicente-Page, J., Pérez-Escudero, A., Carvajal-González, J. M., Fernández-Salguero, P. M., & De Polavieja, G. G. (2018). Histone H4 acetylation regulates behavioral inter-individual variability in zebrafish. Genome Biology, 19(1), Article 55.
Pagnussat, N., Piato, A. L., Schaefer, I. C., Blank, M., Tamborski, A. R., Guerim, L. D., Bonan, C. D., Vianna, M. R., & Lara, D. R. (2013). One for all and all for one: The importance of shoaling on behavioral and stress responses in zebrafish. Zebrafish, 10(3), 338–342.
