ERA’S JOURNAL OF MEDICAL RESEARCHVOL.2 NO.1
IN–VITRO PROPAGATION OF CHAMOMILLA RECUTITA FROM CAPITULUM
INFLORESCENCE: A MEDICINAL PLANT WITH MULTIPLE THERAPEUTIC
Department of Pathology
Era's Lucknow Medical College and Hospital, Lucknow, Uttar Pradesh India– 226003.
College of Natural and Applied Sciences (CONAS),
Crescent University, Abeokuta, Nigeria
A protocol has been developed for induction of somatic embryogenesis
Address for correspondence
from whole inﬂorescence explants of Chamomilla recutita L.
(chamomile). Chamomile is a well–known medicinal plant from the
Dr. Rumana Ahmad
Asteraceae family often referred to as the “star among medicinal species.”
Department of Pathology
Nowadays, it is a highly favoured medicinal plant in folk and traditional
Era's Lucknow Medical College &
medicine. Its multitherapeutic, cosmetic and nutritional values have been
established through the years of traditional and scientiﬁc use and
Sarfarazganj, Hardoi Road,
research. Chamomile has an established domestic (Indian) and
international market, which is increasing day by day. Among the various
major constituents, α–bisabolol and chamazulene have been reported to
be more useful than others. Chamazulene occurs in the capitula of the
ﬂowers in minute quantities and has been demonstrated to exert anti–
inﬂammatory activity in–vivo. Moreover, chamomile is a seasonal 4–5 months winter crop in India but is extensively
required in various medicinal applications. Therefore, to increase the overall yield of this plant, its in–vitro
propagation is needed. In the present study, somatic embryos were developed from capitulum explants after 2–4 weeks
of culture on MS medium supplemented with 26.8 µM NAA and 11.5 µM Kin. The somatic embryos were further
subcultured in–vitro, where new plantlets regenerated from embryos. It is concluded that in–vitro propagation is
possible in case of chamomile and can be used to increase the overall yield of chamazulene present in the capitula of
ﬂowers as well as augment the overall yield of this important plant, which is conventionally propagated by seeds.
Key words: Capitulum, Chamomilla recutita, Inﬂorescence, Plant regeneration , Somatic embryogenesis,
Abbreviations: NAA α–napthalene Acetic Acid, Kin Kinetin, MS medium Murashige and Skoog (1962) basal
of oil (1).
Chamomilla recutita (synonyms: Matricaria recutita,
It was introduced to India during the Mughal period,
Matricaria chamomilla), commonly known as
now it is grown in Punjab, Uttar Pradesh, Maharashtra
chamomile (also spelled camomile), German
and Jammu & Kashmir. It was introduced in Jammu in
chamomile, Hungarian chamomile (kamilla), wild
1957 by Handa et al (2). The plant was ﬁrst introduced
chamomile or scented mayweed, is an annual plant
in alkaline soils of Lucknow in 1964–65 by Chandra et
belonging to Asteraceae family. Chamomilla recutita is
al (3,4). There is a great demand for ﬂowers of
the most popular source of the herbal product
chamomile (Fig.1). Presently, two ﬁrms, namely, M/s
chamomile, although other species are also used as
Ranbaxy Labs Limited, New Delhi and M/s German
chamomile. Chamomile is one of the important
Remedies are the main growers of chamomile for its
medicinal herbs native to southern and eastern Europe.
Hungary is the main producer of the plant biomass. In
Hungary, it also grows abundantly in poor soils and it is a
Chamomile has been used in herbal
source of income to poor inhabitants of these areas.
remedies for thousands of years and has been
Flowers are exported to Germany in bulk for distillation
included in the pharmacopoeia of 26 countries (5).
1 ERA’S JOURNAL OF MEDICAL RESEARCHJan.–June.2015VOL.2 NO.1
inﬂammatory and antiseptic, also as an antispasmodic
and mildly sudoriﬁc (13). The other pharmacological
properties include carminative, healing, sedative and
spasmolytic activity (14). M. chamomilla has been
shown to exhibit both positive and negative
bactericidal activity with Mycobacterium tuberculosis,
Salmonella typhimurium and Staphylococcus aureus.
The international demand for chamomile oil has been
steadily growing. As a result, the plant is widely
cultivated in Europe and has been introduced in some
Asian countries for the production of its essential oil.
The oil is used as a mild sedative(15,16) and also for
digestion(17–19) besides being antibacterial and
fungicidal in action (20,21). In addition to
pharmaceutical uses, the oil is extensively used in
perfumery, cosmetics and aromatherapy,(22–24) and in
food industry (25). Gowda et al (26)found that the
essential oil present in the ﬂower heads contains
azulene and is used in perfumery, cosmetic creams, hair
preparations, skin lotions, tooth pastes, and also in ﬁne
liquors (26). The dry ﬂowers of chamomile are also in
great demand for use in herbal tea, baby massage oil,
for promoting the gastric ﬂow of secretion and for the
treatment of cough and cold (27).
With an ever–increasing global inclination towards
herbal medicine, there is not only an obligatory
demand for a huge raw material of medicinal plants, but
also of the right stage when the active principles are
available in optimum quantities at the requisite time for
standardization of herbal preparations. Commensurate
with this, the intervention of biotechnology or to be
precise, plant tissue culture for accelerating clonal
multiplication of desired clones and strains (high–
yielding) of medicinal plants through micro
propagation and their conservation through
establishing Tissue Banks or Gene Banks are warranted
in the right earnest. Ideally, the herbal plants should be
Fig: 1 (a) Chamomile recutita Habit (b) Flowers
grown under uniform environmental conditions and the
(c) Flower and Leaf Morphology
planting material must have the same genetic make–up
It is an ingredient of several traditional, Unani and
as of the selected high–yielding clones, which is
homeopathy medicinal preparations (6–9). As a drug, it
possible when they are cloned through an in vitro
ﬁnds use in ﬂatulence, colic, hysteria and intermittent
strategy, that is, micro propagation, at least in cases
fever (10). The ﬂowers of M. chamomilla contain the
where conventional vegetative propagation methods
blue essential oil from 0.2 to 1.9% (11, 12) which ﬁnds a
are insufﬁcient or wanting to achieve the goal. In the
variety of uses. Chamomile is used mainly as an anti–
2IN VITRO PROPAGATION OF CHAMOMILLA RECUTITA FROM CAPITULUM INFLORESCENCE: A MEDICINAL PLANT WITH MUTIPLE
Sub–Culturing of Roots, Shoots and Embryos
present study, a protocol has been developed for rapid in–
The jars containing developing embryos were opened
vitro propagation of Chamomilla recutita.
under aseptic conditions and the embryos taken out
very carefully using sterilized forceps. Roots and
MATERIALS AND METHODS
shoots were separated very carefully from the embryos.
Plant material and disinfection
All three were sub–cultured in separate jars containing
This research work was carried out in the School of Life
Sciences, ITM University, Gwalior, MP, India. In this
study, young inﬂorescences from Chamomile recutita
were used as explants. All explants were washed under
After 7–10 days of culturing, some of the cultured
running tap water for 10 min then surface sterilized in a
explants (Fig. 2a) showed adaptation to the
70% ethanol for 60 seconds followed by sodium
environment while others showed necrosis (browning
hypochlorite solution (NaCl with 1% active chlorine) for
of explants) (Fig. 2b). In 15–18 days, early stages of
10 min. This was followed by 2–3 rinses in distilled water.
callus formation were observed (Fig. 2c, d). Complete
callus formation occurred within 4 weeks of culture
initiation (Figs.2e,f). The combination of an auxin with
Immediately after surface sterilization, the explants were
a cytokinin was more favorable for callus induction
transferred to laminar airﬂow cabinet and further
than addition of only one plant growth regulator.
Somatic embryogenesis in callus tissue was initiated
processes were carried out under aseptic conditions. The
within 30 days of explant culture (Fig. 2g). Embryoids
explants were carefully transferred to another Petridish
neither formed nor developed on media not containing
containing sterilized blotting paper. All petals and sepals
growth regulators or supplemented only with auxin or
from the explants were excised with the help of sterilized
cytokinin. The plumules and radicles were excised
blades and forceps and the explants were carefully
from the embryos and sub–cultured separately (Fig.
inoculated on basal MS medium (Murashige & Skoog,
2h). Leaves could be seen arising from the plumule,
1962) supplemented with 26.8 µM NAA, 11.5 µM Kin,
roots developing from the radicle. whereas callus
3% (w/v) sucrose and solidiﬁed with 0.7% (w/v) agar.
forming on embryos within 2 weeks of subculturing
Sucrose was used as the carbon source and agar as
solidifying agent. The pH of the medium was adjusted to
5.8 before autoclaving at 121°C for 15 min and poured
into 800x800mm glass jars (50 ml medium per jar and
four explants per jar). Inoculated jars were sealed with
Paraﬁlmand incubated at 25°C on tissue culture racks
under controlled light regime (16:8 h light/dark
photoperiod) supplied by cool–white ﬂuorescent lamps
and the changes were observed.
Somatic embryo development
After 2–4 weeks in culture all callus tissues (with or
without somatic embryos at globular stage) were
transferred to solid media having the same composition
as the induction medium from which they were removed.
Culture in jars was maintained for 4–8 weeks at 25°C
under controlled light regime (16:8 h light/dark
photoperiod) supplied by cool–white ﬂuorescent lamps.
Somatic embryo maturation and germination
Cotyledonary stage embryos were dissected from callus
tissues and transferred onto solid induction medium
where they were maintained for 4–5 weeks at 25°C under
controlled light regime (16:8 h light/dark photoperiod)
supplied by cool–white ﬂuorescent lamps.
3 ERA’S JOURNAL OF MEDICAL RESEARCHJan.–June.2015VOL.2 NO.1
Fig.2 (a) Sterilized capitula from Chamomile as
explants. (b) Inoculation of explants on MS
medium. (c,d) early. and (e,f) late stages of callus
formation from cultured explants. (g) somatic
embryogenesis from transferred callus tissue.
(h,i) Sub–culturing of excised radicle, plumule
4IN VITRO PROPAGATION OF CHAMOMILLA RECUTITA FROM CAPITULUM INFLORESCENCE: A MEDICINAL PLANT WITH MUTIPLE
In the present study, an efﬁcient protocol was developed
Tissue culture is the culture and maintenance in vitro of
for rapid micro propagation of chamomile, a winter
plant cells or organs in sterile, nutritionally and
crop, from whole capitulum inﬂorescence. It can be
environmentally supportive conditions. It has
applications in research and commerce. In commercial
concluded from the study that micro propagation can be
settings, tissue culture is often referred to as micro
used in chamomile as an alternative to conventional
propagation, which is really only one form of a set of
propagation by seeds to increase the yield not only of
techniques. Micro propagation refers to the production
chamazulene but the entire herb for various medicinal
of whole plants from cell cultures derived from
explants, the initial piece of tissue put into culture of
meristem cells. In literature, there are reports on tissue
culture and micro propagation of chamomile using
The authors are thankful to the Chancellor and Vice
various plant parts as explant sources. Stem and leaf
Chancellor, ITM University, Gwalior, MP, India, for
explants have been used for the induction of callus
their support and co–operation in carrying out this study.
cultures; this approach has been taken by Reichling and
Baker (28) for the establishment of callus cultures from
CONFLICT OF INTEREST
two chamomile varieties (E40 and BK2) on a modiﬁed
The authors declare that they have no competing
MS medium supplemented with 2.4 µM NAA (28).
Reichling et al. (29), isolated a callus culture from
surface–sterilized shoot segments of the chamomile
variety BK2 on a modiﬁed MS medium supplemented
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