Abstract
Piper longum L. is a well known spice plant belonging to the family Piperaceae with high pharmacognosy potential, but it is becoming threatened due to overexploitation. Thus, this investigation aims to standardize a cost effective protocol for in vitro propagation of this economically important plant. Internodal segments were used as explant for callogenesis in Murashige and Skoog medium with 3% sucrose and 0.8% agar, with NAA or 2,4 D. Optimum callus induction was observed in MS medium with 5.0 mg/L NAA. Calli were subcultured on shoot regeneration media containing different concentrations of cytokinin (KIN/BAP) along with 0.1 mg/L NAA. Best shoot regeneration was obtained on MS media supplemented with 2.0 mg/L KIN and 0.1 mg/L NAA. Induced shoots were rooted in either NAA or IBA and highest rooting was induced in MS medium enriched with 0.5 mg/L NAA. Rooted plantlets were acclimatized and 88% of hardened plants survived. Field emission scanning electron microscopic showed that regeneration from callus had occurred by somatic embryogenesis. A comparative study on identification and quantification of piperine (the chief alkaloid of the cultivar) were done from root and fruit of both in vitro and in vivo grown plants through Reverse Phase-High Performance Liquid Chromatography method. In vitro grown fruit was found to have the maximum amount of piperine.
Key message
For the first time, a comparative study in identification and quantification of piperine was done from both in vitro and in vivo grown Piper longum through RP-HPLC method.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
Data availability
Authors certify that the work contained in this paper is original, and has not been submitted to any other journal for publication.
Abbreviations
- MS:
-
Murashige and Skoog
- NAA:
-
1-Napthaleneacetic acid
- 2,4-D:
-
2,4-Dichlorophenoxyacetic acid
- KIN:
-
Kinetin
- BAP:
-
N6-Benzylaminopurine
- IBA:
-
Indole-3-butyric acid
- mg/L:
-
Milligram/Litre
- FE-SEM:
-
Field emission scanning electron microscope
- RP-HPLC:
-
Reverse Phase-High Performance Liquid Chromatography
- Rt :
-
Retention time
- ANOVA:
-
Analysis of variance
References
Bhat SR, Kackar A, Chandel KPS (1992) Plant regeneration from callus cultures of Piper longum L. by organogenesis. Plant Cell Rep 11:525–528
Cabral GB, Carneiro VTC, Lacerda AL, Do valle CB, Martinelli AP, Dusi DMA (2011) Somatic embryogenesis and organogenesis in apomictic and sexual Brachiaria brizantha. Plant Cell Tissue Organ Cult 107:271–282
Chaudhury A, Qu R (2000) Somatic embryogenesis and plant regeneration of turf-type bermudagrass: effect of 6-benzyladenine in callus induction medium. Plant Cell Tissue Organ Cult 60:113–120
Gowthami R, Sharma N, Pandey R, Agarwal A (2021) Status and consolidated list of threatened medicinal plants of India. Genet Resour Crop Evol 68:2235–2263. https://doi.org/10.1007/s10722-021-01199-0
http://www.agritech.tnau.ac.in/horticulture/horti_medicine%20crops_tippili.html. Accessed on 8th Oct 2021
http://www.ayushveda.com/herbs/piper-longum.html. Accessed August 2021
http://www.gosumitup.com/about-indian-long-pepper-know-the-spice-piper-longum. Accessed on 8th Oct 2021
http://www.indiabiodiversity.org/species/show/230721. Accessed on 8th Oct 2021
India Biodiversity Portal. Available at http://indiabiodiversity.Org/species/show/230721. Accessed on 8th Oct 2021
Isah T, Umar S, Mujib A, Sharma MP (2018) Secondary metabolism of pharmaceuticals in the plant in vitro cultures, strategies, approaches and limitations to achieving higher yield. Plant Cell Tissue Organ Cult 132:239–265. https://doi.org/10.1007/s11240-017-1332-2
Kanaki N, Dave M, Padh H, Rajani M (2008) A rapid method for isolation of piperine from the fruits of Piper nigrum Linn. J Nat Med 62:281–283
Khound A, Barua P, Saud BK, Saikia A, Kumar S (2017) Piperine content variation in different Piper longum germplasms of North East India determined through RP-HPLC method. J Appl Nat Sci 9(2):960–965. https://doi.org/10.31018/jans.v9i2.1304
Kumar S, Kamboj J, Suman, Sharma S (2011) Overview for various aspects of the health benefits of Piper longum Linn. fruit. J Acupunct Meridian Stud 4(2):134–140
Kumar V, Markovic T, Emerald M, Dey A (2016) Herbs: composition and dietary importance. Encycl Food Health. https://doi.org/10.1016/B978-0-12-384947-2.00376-7
Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue culture. Physiol Plant 15:473–497
Na H, Kim KW, Kwack Y, Sung KK, Chun C (2007) Comparative anatomy of embryogenic and non embryogenic calli from Pimpinella brachycarpa. J Plant Biol 50:344–350. https://doi.org/10.1007/BF03030665
Nair KKN (2000) Manual of non-wood forest produce plants of Kerala. Kerala forest Research Institute, Kerala
Parida R, Dhal Y (2011) A study on the micro-propagation and antioxidant activity of Piper longum (an important medicinal plant). J Med Plants Res 5:6991–6994
Pradhan B (2015) Regeneration of plantlets of Piper longum L. through in vitro culture from nodal segments. J Appl Biol Biotechnol 3(05):035–039. https://doi.org/10.7324/JABB.2015.35076
Prajapati V, Patel MM, Jha SK, Makwana K (2019) Direct adventitious shoot regeneration in Piper longum L. from spike explants. Int J Chem Stud 7(2):1418–1420
Rajopadhye AA, Namjoshi TP, Upadhye AS (2012) Rapid validated HPTLC method for estimation of piperine and piperlongumine in root of Piper longum extract and its commercial formulation. Braz J Pharmacogn 22(6):1355–1361. https://doi.org/10.1590/S0102-695X2012005000113
Rani D, Dantu PK (2012) Direct shoot regeneration from nodal, intermodal and petiolar segments of Piper longum L. and in vitro conservation of indexed plantlets. Plant Cell Tissue Organ Cult 109:9–17
Ravindran CP, Manokari M, Shekhawat MS (2016) In vitro propagation through ex vitro rooting of a medicinal spice Piper longum Linn. World Sci News 37:12–24
Santosh MK, Shaila D, Rajyalakshmi I, Rao S (2005) RP-HPLC method for determination of Piperine from Piper longum Linn. and Piper nigrum Linn. J Chem 2(2):131–135. https://doi.org/10.1155/2005/627029
Sarasan V, Thomas E, Lawrence B, Nair GM (1993) Plant regeneration in Piper longum L. (Piperaceae) through direct and indirect shoot development. J Spices Arom Crops 2:34–40
Saravanan S (2019) Efficient method of regeneration from nodal explants of Piper Longum L. (Piperaceae). RJLBPCS 5(4):184–192. https://doi.org/10.26479/2019.0504.16
Sathelly K, Podha S, Pandey S, Mangamuri U, Kaul T (2016) Establishment of efficient regeneration system from leaf discs in long pepper an important medicinal plant (Piper longum L). Med Aromat Plants 5:3. https://doi.org/10.4172/2167-0412.1000248
Satyavati G, Gupta K, Ashok, Tandon N (1987) Medicinal plants of India. Indian Council of Medical Research, New Delhi
Siva G, Sivakumar S, Prem KG (2015) Optimization of elicitation condition with Jasmonic acid, characterization and antimicrobial activity of Psoralen from direct regenerated plants of Psoralea corylifolia L. Biocatal Agric Biotechnol 4:624–631. https://doi.org/10.1016/j.bcab.2015.10.012
Sivanesan I, Kyoung KE, Kyoung KM, Young KE, Park SW (2015) Somatic embryogenesis and plant regeneration from zygotic embryo explants of onion. Hortic Bras 33:441–447. https://doi.org/10.1590/S0102-053620150000400006
Soniya EV, Das MR (2002) In vitro micropropagation of Piper longum—an important medicinal plant. Plant Cell Tissue Organ Cult 70:325–327
Vikaspedia. Available at http://vikaspedia.in/agriculture/crop-production/package-of-practices/medicinal-and-aromatic-plants/piper-longum. Accessed on 8 th Oct 2021
Yadav V, Krishnan A, Vohora D (2020) A systematic review on Piper longum L.: bridging traditional knowledge and pharmacological evidence for future translational research. J Ethnopharmacol 247:112255. https://doi.org/10.1016/j.jep.2019.112255
Acknowledgements
Authors are thankful to the University of Burdwan, Department of Biotechnology for providing us the infrastructural facilities and opportunity to done this research work. Authors also want to thank and convey their sincere gratitutude to Mr. Kaushik Sarkar (Technical assistant of Dept. of Biotechnology, The university of Burdwan, WB, India) for helping in HPLC analysis.
Funding
This is a self-funded research work.
Author information
Authors and Affiliations
Contributions
The experiment was designed by MC and Dr. IC Collection of plant, sample preparation and whole research work was executed and statistically analyzed by MC. Dr. SC monitored the research significantly. This experimental work is a part of Ph.D. thesis of MC. All authors read the manuscript, revised critically and approved the final version.
Corresponding author
Ethics declarations
Conflict of interest
All authors declare that they have no conflict of interest.
Additional information
Communicated by Patricia Marconi.
Publisher's Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary Information
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Chatterjee, M., Chatterjee, S. & Chandra, I. In vitro regeneration of Piper longum L. and comparative RP-HPLC analysis of piperine production of in vitro and in vivo grown plants. Plant Cell Tiss Organ Cult 149, 205–212 (2022). https://doi.org/10.1007/s11240-022-02237-0
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s11240-022-02237-0