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Piper nigrum: Micropropagation, Antioxidative enzyme activities, and Chromatographic Fingerprint Analysis for Quality Control

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Abstract

A reliable in vitro regeneration system for the economical and medicinally important Piper nigrum L. has been established. Callus and shoot regeneration was encouraged from leaf portions on Murashige and Skoog (MS) medium augmented with varied concentrations of plant growth regulators. A higher callus production (90 %) was observed in explants incubated on MS medium incorporated with 1.0 mg L−1 6-benzyladenine (BA) along with 0.5 mg L−1 gibberellic acid after 4 weeks of culture. Moreover, a callogenic response of 85 % was also recorded for 1.0 mg L−1 BA in combination with 0.25 mg L−1 α-naphthalene acetic acid (NAA) and 0.25 mg L−1 2,4-dichlorophenoxyacetic acid or 0.5 mg L−1 indole butyric acid (IBA) along with 0.25 mg L−1 NAA and indole acetic acid. Subsequent sub-culturing of callus after 4 weeks of culture onto MS medium supplemented with 1.5 mg L−1 thiodiazoran or 1.5 mg L−1 IBA induced 100 % shoot response. Rooted plantlets were achieved on medium containing varied concentrations of auxins. The antioxidative enzyme activities [superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and ascorbate peroxidase (APX)] revealed that significantly higher SOD was observed in regenerated plantlets than in other tissues. However, POD, CAT, and APX were higher in callus than in other tissues. A high-performance liquid chromatography (HPLC) fingerprint analysis protocol was established for quality control in different in vitro-regenerated tissues of P. nigrum L. During analysis, most of the common peaks represent the active principle “piperine.” The chemical contents, especially piperine, showed variation from callus culture to whole plantlet regeneration. Based on the deviation in chromatographic peaks, the in vitro-regenerated plantlets exhibit a nearly similar piperine profile to acclimated plantlets. The in vitro regeneration system and HPLC fingerprint analysis established here brought a novel approach to the quality control of in vitro plantlets, producing metabolites of interest with substantial applications for the conservation of germplasm.

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Abbreviations

TDZ:

Thiodiazoran

BA:

6-Benzyladenine

GA3 :

Gibberellic acid

NAA:

α-Naphthalene acetic acid

2,4-D:

2,4-Dichlorophenoxyacetic acid

IBA:

Indole butyric acid

IAA:

Indole acetic acid

MS0:

MS medium without plant growth regulators

PGRs:

Plant growth regulators

SOD:

Superoxide dismutase

POD:

Peroxidase

CAT:

Catalase

APX:

Ascorbate peroxidase

WHO:

World Health Organization

RSD:

Relative standard deviation

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Acknowledgments

Financial support of Higher Education Commission of Pakistan is acknowledged.

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Authors and Affiliations

  1. Department of Biotechnology, Faculty of Biological Sciences, Quaid-i-Azam University, Islamabad, 45320, Pakistan

    Nisar Ahmad & Bilal Haider Abbasi

  2. Pakistan Council of Scientific and Industrial Research (PCSIR) Laboratories Complex, Peshawar, 25100, Pakistan

    Inayat ur Rahman & Hina Fazal

Authors
  1. Nisar Ahmad
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  2. Bilal Haider Abbasi
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  3. Inayat ur Rahman
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  4. Hina Fazal
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Correspondence to Bilal Haider Abbasi.

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Ahmad, N., Abbasi, B.H., Rahman, I.u. et al. Piper nigrum: Micropropagation, Antioxidative enzyme activities, and Chromatographic Fingerprint Analysis for Quality Control. Appl Biochem Biotechnol 169, 2004–2015 (2013). https://doi.org/10.1007/s12010-013-0104-7

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  • DOI: https://doi.org/10.1007/s12010-013-0104-7

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