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Abstract: The callus culture from shoots (from
aseptically germinated seeds) and roots (from cultivated
plant) of Podophyllum hexandrum were initiated and
maintained on Murashiage and Skoog (MS) basal medium
with various concentrations of Phytohormones. The study
showed that combined use of Naphthalene Acetic acid (NAA)
and 6-Benzyl amino purine (BAP) exhibits better results
(NAA at 0.5µM to 1.5µM and BAP at 1.5 µM to 2.5 µM ) for
shoot callus initiation and maintenance over that of
root callus. Growth phase study was measured with fresh
weight method for shoot callus and revealed its growth
rate correlation with sigmoid curve. Further, TLC
chromatogram was developed to identify the presence of
podophyllotoxin (active constituent) in shoot callus.
Introduction
Podophyllum hexandrum Royle (Family: Berberidaceae), is
a perennial herb. It is a source of highly valued
podophyllotoxin has been subjected to heavy collection
from the wild and cultivated land. It grows on the lower
slopes of the Himalayas in scrub and forest from
Afghanistan eastwards to central China. The ever
increasing demand of Podophyllum is mainly due to two
semi-synthetic derivatives obtained from rhizomes of
podophyllotoxin (lignan derivative) viz. etoposide and
teniposide which are used in the treatment of various
types of cancer. The anticancer lignan derivative
podophyllotoxin in Podophyllum hexandrum is
biosynthesized at very low quantities in intact plant,
so the biotechnological production of podophyllotoxin
has been considered essential.
Literature survey revealed somatic embryogenesis in
callus derived from zygotic embryos of P. hexandrum on
MS medium supplemented with Phytohormones (Arumugam et
al., 1990 and Ewald et al., 1995). Callus formation from
root segments, on White medium and B5 medium with
phytohormone combination were also reported (Heyenga et
al., 1990 and Fujii, 1991). Production of increased
levels of podophyllotoxin from cell and suspension
cultures has already been initiated from root cultures (Woerdenbag
et al., 1990 and Chattopadhyay et al., 2001). In
relation to growth phase study, it is required to
observe the effects of Phytohormones on callus growth
rates in the aseptic medium. Generally the studies are
used for quantitative purpose using several methods viz.
fresh weight method, cell count method, dry weight
method, density cell count, total cell count, mitotic
index etc. Few of these methods were used for growth
rate study of other species of Podophyllum and
Trigonella (Kadkade, 1982 and Hardman & Stevans, 1978).
The present investigation was carried out to compare,
establish and standardize condition for in vitro culture
and growth rate study of initiated callus of Podophyllum
hexandrum and to identify the presence of
podophyllotoxin by the method.
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