Optimization of a Liquid Culture System for Shoot Regeneration and Achieving an Enriched Level of Scopadulcic Acid B in the Leaf Organ Cultures of Scoparia dulcis L. by Response Surface Methodology
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Response surface methodology (RSM) approach was utilized in the present investigation to optimize the constituents of Murashige and Skoog’s (MS) liquid medium to enrich the scopadulcic acid B (SDB) content in Scoparia dulcis L. RSM approach using the central composite design (CCD) was employed to identify the precise concentration of growth regulators of medium and substantiated in shake-flask cultivation. CCD-RSM model revealed a strong agreement, with a predicted coefficient of determination (R2) values of 0.881 and 0.872 for the shoot regeneration percentage (Y1) and the average number of shoots per explant (Y2), respectively. RSM predicted augmented conditions of MS liquid medium considerably influence the shoot proliferation (Y1 and Y2) in the leaf explants of S. duclis. The medium was fortified with 3.59 μM kinetin (KN; X1), 6.00 μM 6-benzylaminopurine (BAP; X2), 3.93 μM indole-3-acetic acid (IAA; X3), and 25.84 g L−1 of sucrose (X4). The present investigation conquered a maximal response of Y1 and Y2 with 91.28 ± 3.85 and 82.26 ± 2.13, respectively. The experimentally detected values are in close agreement with the predicted values of 90.07 and 79.70, respectively. This proposed that the developed design using CCD had efficacy in the optimization of medium components. The enhanced level of SDB, 9.89 ± 0.98 mg g−1 FW (ca. 5-fold), in the plantlets grown on liquid medium was significantly greater than those accomplished in the other tested plant tissues, comprises of field-grown parent plant leaves and in vitro developed callus and micropropagated plants on MS solid medium. For the first time, the methodology was developed effectively using RSM to promote the shoot proliferation and enhance the SDB production in the leaf organ culture of S. dulcis on MS liquid medium.
KeywordsCCD-RSM model Growth regulators Murashige and Skoog’s medium Scoparia dulcis Shake-flask culture
The authors are grateful to Prof. Russell V. Lenth, Department of Statistics and Actuarial Science, The University of Iowa, USA, for assistance with R statistical programming. The authors also thank the anonymous reviewers for their constructive comments.
The financial support (grant reference No. MRP/3011/09) was from the University Grants Commission, New Delhi.
- Burkill HM (2000) The useful plants of west tropical Africa, 2nd Ed, 5, Royal Botanic Gardens, Kew, pp 648Google Scholar
- Hayashi T, Kawasaki M, Okamura K, Tamada Y, Morita N, Tezuka Y, Kikuchi T, Miwa Y, Taga T (1992) Scoparic acid A, a beta-glucuronidase inhibitor from Scoparia dulcis. J Nat Prod 55:1748–1755Google Scholar
- Hirata K, Yamanaka A, Kurano N, Miyamoto K, Miura Y (1987) Production of indole alkaloids in multiple shoot culture of Catharanthus roseus (L). G. Don. Agric Biol Chem 51:1311–1317Google Scholar
- Lee CL, Wang WL (1997) Biological statistics. Science Press, Beijing, ChinaGoogle Scholar
- Majumder S, Rahman MM, Bhadra SK (2011) Micropropagation of Scoparia dulcis Linn. through induction of indirect organogenesis. Asia Pac J Mol Biol Biotechnol 1:11–17Google Scholar
- Naveenchandra PM, Bhattacharya S, Ravishankar GA (2011) Culture media optimization through response surface methodology for in vitro shoot bud development of Solanum melongena L. for micropropagation. Int J Bioautomation 15:159–172Google Scholar
- Osei-Safo D, Chama MA, Addae-Mensah I, Waibel R (2009) Hispidulin and other constituents of Scoparia dulcis Linn. J Sci Technol 29:7–15Google Scholar
- R Core Team (2018) R: A language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, Austria. URL https://www.R-project.org/
- Taylor L (2006) Vassourinha monograph 4/Scoparia dulcis. In: The rainforest, pharmacy to the world. Raintree Nutrition, Inc., Carson CityGoogle Scholar