A review on licorice

Abstract

Licorice (Glycyrrhizaglabra L) is an important herb used in almost all systems of medicine. The author tries to present in this article a comprehensive review on all aspects of Licorice.

Full text

Pages 57 - 88
Ancient Science of Life, Vol No. XIII Nos. 1 & 2, July-October 1993, Pages 57 - 88
A REVIEW ON LICORICE
K. VENKATA SUBBA RAO
Department of Genetics, P.G. Institute of Basic Medical Sciences, University of Madras,
Taramani, Madras 600 113, India.
Received: 1 November, 1991 Accepted: 20 December, 1991
ABSTRACT: Licorice (Glycyrrhizaglabra L) is an important herb used in almost all systems of
medicine. The author tries to present in this article a comprehensive review on all aspects of
Licorice.
INTRODUCTION
Licorice, Glycyrrhiza glabra L. belongs to
the Family – Fabaceae; Tribe – Astragaleae.
The name Glycyrrhiza is of Greek origin
which means “sweet wood”. Since ancient
times, licorice finds an important place in
Chinese medicine. It was considered to
rejuvenate those consuming it for longer
periods [Chopra et al 1958]. Licorice is
referred to by Theophrastus. Romans
cultivated it after thirteenth centuryand
called it as Radix dulis. In UK it is in
cultivation from sixteenth century. [Trease
and Evans 1983]. In Indian medicine;
licorice is one of the principal drugs of
Susruta. It is referred to as Mulethi,
Malahatti, Yastimadhu and in Malayalam
called as Erattimathuram. In Japan,
commonly known as Kanzo. Species other
than G.glabra were also found to yield
compounds of commercial value.
In virtue of its importance in food and
pharmaceutical industry, licorice was
extensively subjected to scientific
investigations. Attempts were made to
understand the biochemistry of licorice and
its derivatives. New compounds were
derived from this plant utilizing new and
improved analytical methods. Recent
advents in molecular biology and
biotechnology led to the efforts to
improve/increase the yield of compounds
which are of economic importance.
Occurrence
Different varieties of G. glabra are known to
yield commercial drugs [Trease and Evans
1983].
G. glabra var. typica Reg. et. Herd. The
plant is about 1.5 m high bearing publish –
blue flowers. The underground portion
consists of long root and thin rhizome or
stolons and penetrates the soil upto a depth
of 1m or more. It is the source of Spanish
licorice and is grown in Spain, Italy,
England, France, Germany and the USA.
G. glabra var. glandulifera Wald. et Kit. is
the source for Russian licorice and grows
wild in Central and Southern Russia. The
underground portion consists of large root
stock bearing numerous long roots but no
stolons.
G. glabra var B.violacea Bioss. is Persian
licorice which bears violet flowers. It is
available in Iran, Iraq and in the valleys of
Tigris and Euphrates. The other

Pages 57 - 88
macrosopic and microscopical characters
were detailed by Trease and Evans [1983].
G.lepidota was reported to grow in rich soil
of low or moist land in the valleys or on the
plains 20 to 4000 feet throughout California
[Jepson 1970].
Cultivation of Licorice
Licorice can be cultivated or obtained from
wild plants. The plants are known to grow
well in deep fertile sandy soils near streams
in the subtropics. Dry seasons are beneficial
to the crop and thrives well in warm regions
where the annual rainfall is not more than
50cm. Fertile sandy of sandy-loan soils
devoid of any stones are optimal for licorice.
Manuring is not required unless the soil is
not fertile [Singh et al 1984]. G.uralensis,
another species of Glycyrrhiiza known to
yield commercial products was reported by
Nadezhina et al [1981] to grow in dense
sands.
G.glabra was found to exist under wide
range of soil salinity [Mirkin et al 1971].
Drought resistant variety of G. glabra was
reported by Aprasidi [1978] from the flood
plains of river Amudarya. Khafizova [1978]
recorded the highest yield of roots and top
growth of Golodnaya steppe and Amudarya
populations of licorice in chloride sulphate
soils of Uzbek, USSR.
Mohammad and Rehman [1985] compared
the cultivation of licorice in irrigated and
rainfed sanddunes. Survival percentage was
more in irrigated sands. To achieve stable
yields of licorice, irrigation of G.glabra was
a must in oasis region sands [Durmeshev
1986]. It was reported by Osipov [1987]
that variations in the yield of root mass of
licorice was caused by different
hydrogeologic regimes of Amudarya flood
plain. Increase growth was recorded where
subsoil waters were at a depth from 177cm
to 195cm.
Reclamation of desert sands upon
cultivation of licorice was demonstrated in
sands adjacent to oases in the Russian desers
[Kel’dzhaev and Gladyshev 1982]. A study
carried out by Varganov and Gladyshev [
1981] revealed the utility of cultivation of
licroce on oasis sands in stabilization of
sand and quick improvement of soil.
Similar report was also on record
[Mohammad and Rehman 1985] which
shows the stabilization of sanddunes after
cultivation of licorice.
Though inter-plantation of carrot, potato or
cabbage crops along with licorice is feasible
for the first two years, it is discouraged in
the view of the increase in weed population
[Singh et al 1984].
Reports are available on the cultivation of
licorice in India. It was found to grow well
in Patiala, Hissar of Haryana State [Singh
1964], Uttar Pradesh [Uniyal et al 1978] and
in South India [Ahmad and Khaleefathullah
1986]. Abrus precatorius Linn. Is
commonly called as wild licorice, Indian
licorice or licorice bush. Though the roots
of this plant are found as a substitute for
genuine roots, it was not recommended by
Chopra et al [1958] due to the toxic
properties associated with it.
Propagation
Propagation is generally performed by
employing 15cm long stolons with two or
three buds obtained from the planting stock.
Old crowns were also used as planting
materials [Singh et al 1984]. Germination
capacity of G.glabra seeds at different
stages was studies by Gladyshev and
Kerbabaev [1967]. Seeds in waxy-ripe stage
were found to show highest germination

Pages 57 - 88
rate. Slow germination of scarified seeds of
licorice was revealed from the studies of
Yaskonis [1976].
The cuttings planted and irrigated in the
ridges of 45-60 cm in height facilated root
development. Poor quality of licorice yield
resulted when the plants were allowed to set
seeds. Removal of flowers immediately
after their emergence yielded good quality
of licorice [Singh et al 1984]. Seed
germination of G. uralensis, as reported by
Liu [1987] depends on the thickness and
texture of soil.
Licorice root production was successfully
enhanced by treating cuttings with 0.0025%
succinic acid followed by the application of
nitrogen and phosphorus. The root weight
was found two fold in three-year old plants
[Badalov 1978].
Different harvesting procedures were
detailed by Singh et al [1984]. Regeneration
occurred from underground root system
after harvesting and were ready for further
harvesting after two to five years.
The water product of licorice after extraction
was found useful as animal feed [Yumatni et
al 1980]. It was reported by Timofeyer
[1984] that the waste yields inorganic
substances and vitamins.
Diseases and Pests of Licorice
Only few reports are available on the disease
and pests of licorice. Gentrospora acerina
caused licorice rot [Bank 1963]. Viral
infection of licorice was documented by
Blattny et al [1950]. Heterodera
glycyrrhizae [Narbaev 1987] and
Acanthoscelides aureolus [Boe et al 1988]
were the pests of licorice reported from
USSR and USA respectively. According to
CIMAP [1982], G.glabra plants were
affected by weevil, Myllocerus
undecimpustulatus.
Developmental studies in Licorice
Galimova [1978a] conducted studies on
male and female gametophyte development
in G.glabra and G. uralensis.
Microsporogenesis and development of
gametophyte of Glycyrrhiza was
investigated by Ashurmetov et al [1979].
Embryological features in spontaneous
hybrids of Glycyrrhiza revealed normal
male and female gametophyte development
and seed production. Superior seed quality
was also reported from such hybrids
[Ashurmetov and Sakhibaeva 1985].
Factors affecting the seed productivity in
G.glabra and G.uralensis were assessed by
Galimova [1978b]. Crossess of G.glabra
and G.ralensis produced viable fruits and
seeds. Galimova and Murinova [1985]
further found that these hybrids were of
immense use from cultivation studies.
Mikailov and Mirzaliev [1978]
demonstrated distinctive morphological
characters such as color of flowers, their
size, weight, shape of pod as well as yield
and number of seeds to be helpful in
identification of useful forms of G. glabra.
Based on anatomical characters, Sukhova
and Kel’dzhaev [1987] differentiated
licorice grown in sands and natural
conditions in Amudarya alluvial plain
[USSR]. They showed an increase in
volume of stalk and root in G. glabra grown
in sands. To identify the crude drugs
present in licorice, a detailed key was
developed by Zeng et al [1988] by
evaluating the morphological and
microscopically similarities.
Cytogenetics of Licorice

Pages 57 - 88
The diploid chromosomes complement of
Glycyrrhiza glabra was established as 2n =
16 [Darlington and Wylie 1955].
Morphological and biological polymorphism
in different populations of G.glabra were
reported by Tashmukhamedov and Aprasidi
[1977]. Karyological investigations were
carried out in three species of Glycyrrhiza,
namely G. glabra, G. lepidota and G.
echinata by Barghi and Siljak-Yakovlev
[1990]. Inspite of their morphological
variations, these species revealed a stable
chromosomes number supporting the earlier
investigations [Taylor and Taylor 1977;
Mikailov and Mirzaliev 1978; Pauzner and
Tashmukhemdov 1978; Magulaev 1980;
Ashurmetov and Karshibaev 1982].
However, total and relative lengths of
chromosome pairs and centrometric indices
differed among species. Ashurmetov et al
[1979] recorded normal meiosis in six
different species of Glycyrrhiza.
Licorice Phytochemistry
The major constituents of licorice are
triterpenoids and flavonoids. Apart from
these, occurrence of other classes of
compounds in small quantities has also been
reported. The Sweetness of licorice is due
to glycyrrhizin [a triterpenoid compound].
Presence of flavonoids gives yellow colour
to licorice. Quantitative differences could
be observed in the compounds derived from
various sources of licorice. Analytical
methods such as TLC, GLC, HPLC were
employed for the separation of constituents
of licorice. Further evaluation and
determination were carried out using
spectrophotometric, mass
spectrophotometric, and mass
spectrophotometric and NMR procedures.
Flavonoids
Afcher et al [1980] isolated and identified
licuroside, liquiritigenin, isoliquiritigenin
and liquiritin from Iranian G. glabra var.
glandulifera. Epigeal portions of G. glabra
yielded new flavonoids galangin,
naringenin, tioxy-isoflavone and dioxy-
flavone apart from pinocembrin and
glabranins [Baturov et al 1986]. Structure
of licoricidin, a prenylated isoflavan isolated
from Si-pei licorice was characterized by
Fukai et al [1988]. They also studied the
structure of six isoprenoid substituted
flavonoids from Xibei licorice [Fukai et al
1989]. Licoflavanone isolated from the
leaves of G. glabra [Fukai et al 1988] was
found to possess antimicrobial property.
Neolicuroside, a new chalcone glycoside
was isolated by Miethin and Speicher-
Brinker [1989] from the roots of G. glabra.
Two flavanon glycosides have been isolated
by Yahara and Nishioka [1984] from G.
uralensis.
Nakanishi et al [1985] isolated flavonoid
glycosides from G. uralensis roots. On the
basis of irradiation experiments, Shiozawa et
al [1989] revised the structure of glycyrol
and isoglycyrol from G. Uralensis roots.
Gancaonins, the prenylated flavonoids, were
isolated from aerial parts of G.uralensis and
G.palladiflora and characterized by special
studies [Fukai et ali 1990a, b].
Studies of Ghisalberi et al [1981] showed
that aerial parts of G.acanthocarpa yielded
four isopreny-lated resocrcinol derivatives.
G.eurycarpa, new species was reported from
Gansu province in China. Liu and Liu
[1989] for the first time isolated glycyroside,
an isoflavone diglycoside, apart from
liquiritigenin, isoliquiritigenin and
schaftoside. Cultured cells of G.echinata,
also called as pseudoglycyrrhiza, yielded
two flavonoids viz. echinatin and licodione

Pages 57 - 88
and their structure were evaluated by Ayabe
et al [1980].
Triterpenoids
The biosynthetic pathway of glycyrrhizic
acid was demonstrated by Fraz et al [1983].
Glabranin A and B were isolated from
G.glabra root. Varshney et al [1983]
elucidated their structure employing TLC
and HPLC. Shu et al [1987] determined the
structure of uralenolide, a new triterpenoid
lactone from G. uralensis. Employing a
chromatographic system, a new triterpene
namely glyuranolide was isolated from the
crude sapogenin of G.uralensis [Jia et al
1989]. Recently, Mirhom et al [1990]
isolated a new triterpenoid from the roots of
G. echinata. Licorice roots yielded 5-penta
cyclic triterpenoids [Elgamal et al 1990].
Polysaccharides
A neutral polysaccharide, glycyrrhizin, was
isolated by Shimizu et al [1990] from the
roots of G. uralensis. Similarly Tomoda et
al [1990] characterized two polysaccharides,
glycyrrhizans UA and UB from the same
source.
Phenolic compounds of Licorice
Hatano et al [1989] isolated
licopyranocoumarin, an anti HIV phenolic
compound from Si-pei licorice from the
north – western region of China. Four new
phenolic compounds were derived from the
roots of Glycyrrhiza species. Kiuchi et al
[1990] elucidated their structure on the basis
of spectroscopic and chemical studies.
Similarly, phenolic constituents of
G.uralensis were studied by Fukai et al
[1991].
Other constituents of Licorice
Phrrolo – pyrimidine alkaloid and
tetrahydroquinoline alkaloids were isolated
from the roots of G.uralensis [Han dna
Chung 1990; Han et al 1990].
Licocoumarone, a new benzofuran
derivative from licorice was isolated and
characterized by Demizu et al [1988].
Antimicrobial glepidotin, a bibenzyl from
American licorice, G. lepidota was reported
[Mitscher et al 1983; Sitaraghav et al 1989].
Volatile flavor components were isolated by
Miyazawa and Kameoka [1990] from G.
glabra var. glandulifera growing in north-
east China.
Isolation and Determination of Licorice
Constituents
Various methods employed to isolate the
constituents of licorice were modified from
time to time in accordance with
improvement in technology. The traditional
method of licorice extraction involves the
shredding of dried roots and stolons and
extraction with hot water. The aqueous
extract was then allowed to concentrate after
evaporation in vacuum pans. A paste with
18-25% moisture was obtained and moulded
to sticks or blocks of required size and shape
[Singh et al 1984].
Modern continuous extraction plants have
been developed in later years [Masters 1972;
Molyneux 1975]. An equipment was
devised by Muraviev et al [1985] for
gravitational multistep extraction of solid-
liquid system to extract licorice roots to
yield higher quantities of biologically active
agents. Stepanova [1985] demonstrated the
usefulness of fermentation of harvested
plant material. Fermentation led to an
increase in the yield of triterpene saponins
from G. glabra when compared to the non-
fermented sample.

Pages 57 - 88
An array of analytical methods are on record
for the determination of licorice derivatives.
High performance liquid chromatography
[HPLC] was applied to estimate the content
of Glycyrrhiza in traditional Chinese drug
‘Kampo” [Akada et al 1978]. Sulfostyrene
cationic exchanger was used by Manyak and
Muraviev [1984] for the spectrophotometric
determination of glycyrrhizic acid in
Glycyrrihiza roots and drug extracts. Sagara
et al [1985] developed a simple precise
method to determine glycyrrhizin in
Glycyrrhiza radix employing ionpair HPLC.
Using bilayer column chromatography,
determination of glycyrrhizhin in cosmetic
lotions and creams was demonstrated
[Mikami et al 1988]. Spectrophotometric
quantitation of hexoses and pentoses [free
and linked to polysaccharide chain] from
G.glabra extracts was reported by Riccio
and Riviera [1988]. A second derivative
[D2] spectrophotometric method was
developed [Song et al 1990] recently to
detect the total glycyrrhizic acid in G.
glabra using ion-pair extraction technique.
Zeng et al [1990a] devised a rapid HPLC
method for simultaneous separation and
determination of flavonoids and coumarins
from licorice.
Chemical Synthesis / Modification of
Licorice Derivatives
Attempts were made on chemical
modification, biotransformation of licorice
phytochemicals to obtain more derivatives
of pharmacological importance. A perusal
of literature, however, reveals no reports on
direct chemical synthesis of licorice
products.
Glycyrrhetinic acid was subjected to
chemical modification to derive
deoxyglycyrrhetol with a view to eliminated
pseudoaldosteronism, a side effect
accompanied with glycyrrhetinic acid
administration. Chemically derived
deoxyglycyrrhetol was found to lact
aldosteronic effect, while it maintained the
therapeutic activities [Shibata et al 1987].
Chemical synthesis of glycopeptides from
glycyrrhizhic acid was demonstrated from
glycyrrhizhic acid was demonstrated by
Tolstikov et al [1989]. The derivatives were
shown to possess anti-inflammatory activity
and stimulating effects on hormonal factors
of immunity. Chemical synthesis of
glycyrrhizin from glycyrrhetinic acid was
achieved by coupling of methyl
glycyrrhetate with per-O-acetylated glycosyl
bromides of mono and di-saccharides
[Hirooka et al 1989]. The synthesized
glycosides were found to be beta-type on the
basis of NMR studies.
Amano1 [1984] obtained patent for
biotransformation of glycyrrhetic acid to 3-
epiglycyrrhetic acid employing intestinal
bacteria. The compound was found to have
same effect as glycyrrhetinic acid excepting
pseudo-aldosteronic effect.
Biotransformation was attempted in virtue
of the difficulty associated with the
synthesis of compounds by chemical means.
Conversion of glycyrrhetinic acid to 18-
beta-glycyrrhetic acid by Streptomyces G 20
[isolated from soil] was demonstrated by
Sakano and Ohshima [1986] and their
structures were demonstrated. Daiichi-
Pharm3 [1986] employed Chainia
antibiotica to transform glycyrrhetinic acid
to its derivatives which have pharmaceutical
value. Recent investigations of Tanaka et al
[1990] suggested the use of soil bacterium,
Pseudomonas saccharophila strain 11, for
industrial use to convert glycyrrhizin to
glycyrrhizic acid. The maximum yield was
attributed to the higher beta – glucurudinase
activity possessed by this bacterial strain
when compared to its counterparts.

Pages 57 - 88
Licorice in vitro
Efforts were made to increase the yield in
licorice, employing plant tissue culture
techniques. Shah and Dalal [1982]
attempted for in vitro multiplication under
various cultural conditions employing
modifications of MS medium. Their trails
yielded successful establishment of plantlets
and found 15-20 fold increase in
multiplication rate when compared to
propagation through stolon cuttings.
Similarly, Syrtanova and Mukhitdinova
[1984] tried colonel propagation of G.
glabra and G. uralensis.
Investigations were also carried out to derive
commercially important phytochemicals
from licorice. Wu et al [1974] reported the
absence of glycyrrhizin in suspension
cultures of licorice. Hayashi et al [1988]
recorded similar observations in callus and
cell suspension cultures of G. glabra.
The cells failed to produce detectable
amounts of glycyrrhizin though the
intermediate compounds such as betulinic
acid, beta-amyrin were detected. They
speculated the absence of glycyrrhizin
production was to be due to interruption in
the biosynthetic pathway of glycyrrhizin.
Their later studies also revealed the failure
of suspension cultures to produce
glycyrrhizin after exogenous
supplementation of 18 β-glycyrrhizic acid
[Hayashi et al 1990a]. Triterpenoid
biosynthesis in the cultured tissues of
G.glabra var. glandulifera was ascertained
by Ayabe et al [1990]. Their studies
demonstrated the quantitative differences in
the metabolic alterations in the intermediate
products of glycyrrhizin biosynthesis in
stolon segments, roots and callus cultures.
However, in suspension cultures of
G.glabra, accumulation of soyasaponins
could be observed and their production was
found to be influenced by culture strains and
growth harmones [Hayashi et al 1990b].
Callus cultures of G. uralensis under
optimal conditions were found to yield
formononetin, isoliquiritigenin, echinatin,
liquiritigenin, p-hydroxy benzoic acid and
isobarachalcone [Kobayashi et al 1985]. It
was Ko et al [1989] to demonstrate the
production of glycyrrhizin in Agrobacterium
rhizogenes transformed hairy roots of
G.uralensis. Contrary to this, Saito et al
[1990] reported the absence of glycyrrhizin
in a similar investigation.
Though contradicting reports are available
on the production of glycyrrhizin through
plant tissue culture techniques, patents were
obtained by commercial establishments.
Adventitious roots of licorice produced in
vitro [PCC Technol10] were demonstrated to
contain glycyrrhizic acid. Babcock-
Hitachi2 developed a novel procedure to
regenerate plants from calli which yielded
glycyrrhizic acid. Production of
glycyrrhizic acid by A.rhizogenes
transformed G.uralensis tissue was claimed
by Mitsui-Toatsu-Chem9.
In vitro studies on G.echinata revealed the
production of flavonoids. Investigations of
Ayabe et al [1980] led to the isolation of
echinatin and licodione from G.echinata.
They studies the biosynthetic pathway apart
from characterization of the flavonoids.
G.echinata cells when transferred to fresh
medium or immobilized, exhibited a rapid
transient accumulation of retrochalcone,
echinatin in both cells and the medium
[Ayabe et al 1986a]. According to their
further studies, addition of yeast extract or
calcium alginate beads stimulated the
production of echinatin and retrochalcone
which was found to be under the influence

Pages 57 - 88
of O-methyl-transferases [Ayabe et al
1986b, 1987].
Suspension cultures of G.glabra [Dorisse I
1988] and G. echinata [Ushimaya et al
1989] were found to effectively
biotransform papaverine hydrochloride and
phenylcarboxylic acids respectively.
Licorice in Food industry
Licorice is widely used as a flavouring agent
especially in tobacco industry. In
combination with sugar, the sweetness
increased by 100 times. In pharmaceuticals
and medicinal tea it not only acts as a
flavouring agent but also reduces the
unpleasant taste of other constituents. It is
used as a sweetner and flavouring agent in
low caloric and non-cariogenic food. It
gives sparkle and aroma to confectionary
products and beer respectively. Licorice
serves as a preservative in food industry.
Excessive consumption, however, leads to
harmful consequences.
Pharmacological Activities of Licorice
Though the major use is as a flavouring
agent in food industry, licorice finds its
place in pharmaceuticals too owing to its
diverse pharmacological activities.
According to Chinese Materia Medica,
licorice increases the physical strength and
cures wounds. In India, licorice powder
mixed with fat and honey is applied to cuts
and wounds [Singh et al 1984].
It has been attempted to use licorice for the
treatment of common ailments, for eg.
Spasms, to dreaded diseases like AIDS,
which is the present days challenge to
scientists for its effective management.
Aleshinskaya et al [1964] reported the
usefulness of cabonoxolone, a derivative of
glycyrrhizic acid as an anti-inflammatory
agent. Glycopeptides synthesized from
glycyrrhizic acid [Baltina et al 1988],
glyderinine, a derivative of G. glabra
[Azimov et al 1988] and glycyrrhizin
[Ichikawa et al 1989] exhibited anti-
inflammatory effects in animal models.
Glycyrrhizin reduced hepatotoxicity in
experimental animals exposed to Paraquat
[Kim and Hong 1988]. According to
larkworthy [1977], deglycyrrhinized licorice
was useful for the treatment of both peptic
and duodenal ulcers, supporting the earlier
studies [Li et al 1960; Takagi et al 1963].
Antiulcerogenic activity in mice and rats by
glycyrrhitinic acid derivatives were reported
by Yano et al [1989]. Glycyrrhiza also
protected the rat liver damage caused by
Ischemia [Nagai et al 1991].
The antioxidative property of Glycyrrhizai
flavonoids and glycyrrhizic salts in liver
tissues were on record [Syrov et al 1987; Ju
et al 1989; Abdugafurova et al 1990]. It
was Berger and Holler [1957], Jo et al
[1986] who demonstrated the analgesic
activity of licorice in animal cells.
Hashiguchi et al [1990] suggested the use of
glycyrrhetinic acid as an alternative to local
anaesthetic.
Glycyrrhetinic acid inhibited the activity of
11 beta-dehydrogenase resulting in a
blockade in the conversion of cortisol to
cortisone in human [Mackenzie et al 1990].
Mineralocortecoid effect of licorice
derivatives was reported by Stewart et al
[1990] in healthy volunteers.
Isoliquiritigenin prevented the diabetes
related complications in rats [Aida et al
1990].
Sureshkumar and Prabhakar [1990] reported
the cario-active property of licorice. Zinc
deficiency in children was effectively
managed by licorice extracts [Qiao et al

Pages 57 - 88
1987]. Licorice served as a brain tonic for
mental disorders [Upadhyaya 1986].
Glycyrrhizin was found useful in the
treatment of skin diseases like eczema and
dermatitis as reported by Hayakawa et al
[1987]. Glycyrrhizin reduced the morphine
induced harmful effects in vitro systems
[Huh et al 1988].
Non-mutagenic property of glycyrrhitinic
acid in Chinese Hamster V-79 H3 cells was
reported by Tsuda and Okamoto [1986].
Antimutagenic potential of Glycyrrhiza
extracts and glycyrrhizin against methyl
cholanthrene, imidazole and diethyl
nitrosamine-induced mutagenicity was
demonstrated in mammalian cells [Tanaka et
al 1987]. Investigations of Minematsu et al
[990] on Shosaiko-to-go-Keishikash-
hakuyaku-to [TJ-960] revealed the
protective effects against valpronic acid-
induced anomalies in rats.
Protective role of licorice crude extracts
against radiation-induced lethal damage in
mice was recorded by Ohta et al [1987].
Glycyrrhiza flavonoids are reported to
possess radical scavenging capacity [Hatano
et al 1988; Ju et al 1989] in various test
systems.
Anticarcinogenic activity of licorice in
mammalian cells was on record. Abe et al
[1987] reported that glycyrrhizin and
glycyrrhitinic acid inhibited melanogenesis
in cultured B 16 melanoma cells.
Benzanthracene and tetradecanoyl phorbol-
13 acetate [TPA] activated carci nogenesis
was found inhibited by glycyrrhizin in
mouse skin tumors [Yasukava et al 1988;
O’Brain et al 1990]. It was also shown that
inhibition of protein kinase was the cause
for the suppression of carcinogenicity in
these cell lines. Observations of Mashiba
and Matsunaga [1990] revealed that
glycyrrhizin in combination with diethyl
dithio carbamate inhibited the in vitro
proliferation of mammalian tumor cells.
Licorice exhibited antibacterial activity.
Isoflavonoids and related susbstances from
G. glabra var. typical and prenylated
falvonoids from G.lepidota demonstrated
their antimicrobial activity [Mitscher et al
1980]. Investigations carried out by Hattori
et al [1986] revealed the inhibitory property
of G. uralensis derivatives on cariogenic
bacterium, Streptococcus mutans. On the
other hand, glycyrrhizin stimulated the
growth of Eubacterium strain GLH, a
human intestinal bacterium [Akao et al
1988].
Derivatives of licorice exhibited inhibitory
action against different classes of virus. A
triterpenoid component of G.glabra [Pompei
et al 1980] and Glycyrrhiza polysaccharides
[Chang et al 1989] were shown to possess
antiviral properties against DNA and RNA
viruses. It was reported by Ohtsuki and
Iahida [1988] that direct binding of
glycyrrhizin to virus causes a dose-
dependent direct inactivation of virus
associated kinase and hence the reduction of
viral infectivity. Segal and Pisanty [1987]
suggested the use of glycyrrhizin gel along
with iododeoxyuridine to reduce the healing
time in patients with herpes of lips and nose.
Glycyrrhizin was also found useful as an
additive agent, in chemotherapy of herpes
zoster patients [Aikawa et al 1990].
Glycyrrhizin was shown to have antiviral
activity against Hepatitis A virus replication
in vitro. A dose-dependent inhibition of
HAV antigen and HAV infectivity was
reported [Crance et al 1989, 1990]. They
suggested the utility of Glycyrrhiza for
chemotherapy of acute Hepatitis A.
According to Hayashi et al [1989], treatment
of chronic Hepatitis patients with

Pages 57 - 88
glycyrrhizin was effective without any side
effects.
Recent investigations further revealed a
significant antiviral property of licorice
derivatives against Human Immuno
deficiency Virus [HIV]. Glycyrrhizin
showed a dose-dependent inhibition of the
replication of HIV-1 in MOLT-4 cells. It
was suggested by Ito et al [1988] that
suppression of HIV replication was due to
the inhibitory action of glycyrrhizin on
protein kinase C. Administration of
glycyrrhizin i.v. for a period of more than
30d in individuals with AIDS resulted in
inhibition of HIV replication in vitro
[Hattori et al 1989c]. According to Mori et
al [1990], glycyrrhizin treatment to
individuals with asymptomatic carriers of
AIDS and AIDS related complexes
prevented them to develop AIDS.
Licorice extracts were found to exert their
effect on plant virus also. A dose dependent
inhibitory effect of licorice extract was
found in spinach mosaic virus [Zaidi et al
1988].
Insecticidal property of licorice was on
record. Erion and Mahrous [1983] showed
the toxic effects of G.glabra root extracts on
Spodoptera littoralis.
Pharmacological Studies on Drugs
Containing Licorice
Glycyrrhiza was found to be a constituent in
various Chinese and Japanese herbal drug
formulations. These drugs were in use for a
wide range of ailments.
“TCM-WM”, a Chinese traditional drug,
reduced biliary gastritis in patients
[Gouzeng 1987]. Reduction of
hepatotoxicity was reported by Kim et al
[1986] upon administration of ‘Sosiho
Tang”. Brunner’s glands of duodenal
ulcerated rats exhibited reduced β-
glucuronidase activity. Nadar and Pillai
[1989] observed an increase in
betaglycyronidase after administration of
“Shanka Bhasma” and thus showed a
protection against duodenal ulcers. “TJ-
8014”, a Japanese herbal drug found to have
antinephritic action in rats [Hattori et al
[1989a,b].
Antidiabetic effect of “Ganshao Jiantong”
tablets was reported by Wang [1986] in
diabetes mellitus patients. Plasma
adrenaline levels reduced when Amagaya
and Ogihara [1989] administered rats with
“Shosaiko-to”.
Parikh et al [1984] studies revealed 50%
improvement in Schizophrenia patients after
treatment with an indigenous drug [G K
022]. The herbal drug ‘Sipmidojuskam”
showed sedative, antipyretic, analgesic,
anticonvulsive, antiedemic properties in
mice. It reduced blood pressure and caused
dialation of blood vessels. It also caused
relaxing effect in smooth muscles of
digestive organs in rats [Hong et al 1989].
Sedative effect of “Sanayangsohap-Won”
and “Woohwang-poryong-hwan” was
reported by Lee and Han [1986] in
experimental animals.
The herbal drug “Lactare” improved the
sizes of mammary gland and teat in guinea
pigs and goats [Narendranath et al 1986].
Their clinical trials also showed that lactare
benefited the lactation in mothers and
improved appetite in mothers. Further
evaluation revealed ed no toxic effects.
Capsules of lactare were also found to exert
similar effect [Sholapurkar 1986].
“Tiao Wei Cheng Qi Tang” decoction along
with liquid diet was effective for bowel
cleansing and Zhang et al [1986] found this

Pages 57 - 88
method to be cheap and convenient with no
side effects. Eye drops and capsules
consisting of G. glanra, Berberis aristata
and Curcuma longa were reported to be
highly effective in curing all kinds of
allergic conjunctivitis without any
recurrence of clinical symptoms [Athneria et
al 1987]. Licorice extracts enhanced the
absorption of iron in intestinal segments of
rats as revealed from the studies of El-
chobaki et al [1990]. They recommended it
as a preventive agent to iron deficiency
anemia in both children and adults and as a
bioavailability source of medicinal iron.
Safety Evaluation of Drugs Containing
Licorice
Investigations were carried out to ascertain
the toxic effects, if any, of drug formulations
containing Licorice.
“Sairei-to” and “Saiboku-to”, the Chinese
herbal medicines, were found non-toxic to
rats after 90 days of administration. No
changes occurred in body weight, food
intake, urological, haematological,
ophthalmological and pathoanatomical
features. [Kiwaki et al 1989a,c]. Similarly,
Minematsu et al [1989] conducted a study
on the effect of “Junjentaiho-to”. The drug
was shown to have no toxic effects in mice
and rats as assessed from death rate,
abnormal symptoms, difference in body
weight, food intake, urological,
haematological, ophthalmological and
histopathological data.
Teratological evaluation of “Sairei-to” was
carried out by Kiwaki et al [1989b] on rats.
Their studies revealed no harmful effects of
the drugs as evidenced by the absence of
increase in fetal mortality and inhibition of
fetal growth. No developmental and
reproductive deformities of offsprings
recorded, thus, exhibiting non-teratogenic
effect of the drug.
Patents of Licorice
Owing to the economic importance of
licorice, attempts were made for commercial
production of licorice derivatives. Recent
advents in biotechnological procedures led
to either improvement or enhancement of
licorice derivatives.
Irkutsk State Medical Institute5,6 obtained
patent on extraction of licorice from ground
roots. Stable crude drugs were prepared by
Gelia-Shinyaku4 from Glycyrrhiza radix
employing two surfactants viz, polyoxy
ethylene curing, castor oil and polyoxy
ethylene polyoxy propylene condensate.
Biotransformation studies on licorice
constituents were conducted to derive
pharmaceutical compounds. Amano1
patented for biotransformation of
glycyrrhitinic acid to 3-epiglycyrrhetic acid
employing intestinal bacteria. The
compound was found to have same effect as
that of glycyrrhitinic acid. Further it has no
pseudo-aldosteronic effect which
glycyrrhitinic acid generally possesses.
Biotransformation was attempted since it
was difficult to derive the compound by
chemical means. Similarly, Daiichi Pharm3
employed chainia antibiotica to transform
glycyrrhitinic acid to its derivatives which
are antiallergic, anti-inflammatory and
antitumerogenic.
Tsumura-Juntendo13 demonstrated the
antitumor effect of Glycyrrhiza extracts in
combination with oyster shell powder.
Benzopyranone glycosides possessing
antianemic activity were isolated from G.
glabra [Juntendo Inc7]. Eye drops produced
by Tsumura-Juntendo14, where licorice
extract was one of its constituent was found

Pages 57 - 88
effective for the treatment of conjunctivitis.
Licorice extract in combination with other
herbal drugs prevented tooth decay by
inhibiting the growth of Streptococcus
mutans. Takasge Perfum12 suggested the
use of this drug formulation in chewing
gum, tooth paste and mouth washes.
Compounds derived from licorice were also
of importance in food industry. Aqueous
extracts of starchy material of licorice
yielded edible sweetner upon incubaction
with cyclodextrin [Kabushiki8]. The
product, a sweetner, found suitable for
incorporation into low calorie or non-
cariogenic food and beverages. It has a
milder flavor than glycyrrhizin without
unwanted flavors or medicinal odors.
Ueno15 prepared phytoncides from licorice,
which were of use as food preservatives.
Antioxidant and antimicrobial substances
from licorice [Ueno16] were also useful as
food preservatives.
Extracts of licorice find their place in
cosmetic preparations too. Rohto11
developed a hair tonic preparation from
licorice which was claimed to promote hair
growth and also effective in preventing hair
loss.
Patents on Licorice Cell Cultures
In vitro production of adventitious roots by
Glycyrrhiza cells was demonstrated by
PCC-Technol10. These roots were found to
contain glycyrrhizic acid. Babcock-Hitachi2
developed a new method to regenerate
plants through in vitro manipulation from
calli of licorice rhizome. The regenerated
plants were reported to possess glycyrrhizic
acid. Production of glycyrrhizic acid from
Agrobacterium rhizogenes transformed
Glycyrrhiz uralensis tissues in vitro was
demonstrated by Mitsui-Toatsu-Chem9.
Market Potential of Licorice*
Spain supplied the world’s requirement of
licorice until 1970. The other countries
which are major commercial producers of
licorice are : Spain, France, Italy,
Afghanistan, Iran, Iraq, Turkey, Syria,
Lebanon, Israel, USSR and China.
The data presented in the following table depicts the import of Licorice in India.
Year
Quantity
[in tones]
Value
[in lakhs of Rs.]
1976-77
1977-78
1978-79
1979-80
1980-81
1981-82
1982-83
364.42
752.16
578.98
642.77
286.88
515.63
574.41
9.85
28.97
21.86
26.20
13.37
30.04
29.38

Pages 57 - 88
1983-84
1984-85
1985-86
1986-87
1165.00
372.23
1273.57
401.84
68.80
22.15
78.53
19.79
The data show the increasing requirements
of licorice and sudden spurt in its
requirement was seen during the period
1983-84 and 1985 – 86. However, India too
entered the international market by
exporting licorice, though it is to a lesser
extent. The export statistics show that
during the years 1984 – 85 and 1985- 86,
our country exported one tonne of licorice in
each year which is equivalent to Rs.0.16 and
Rs. 0.18 lakhs respectively. The cost of
powdered and raw licorice/kg in 1989 was
US$2.2 and US$0.9 respectively.
Source: Current Research on Medicinal and
Aromatic Plants [Quarterly Journal
Published by Central Institute of Medicinal
and Aromatic Plants, Lucknow].
Future Prospects of Research on Licorice
From the preceding part of this text, the
economic potential of licorice is
unequivocally evident from its highly
diverse intrinsic pharmacological properties
apart from its use in food industry. With
further advancement in knowledge on its
pharmacological activities and use in food
industry, the commercial requirement of
licorice will certainly increase in future.
The statistical data on licorice imports
shows the increasing demand. To conserve
foreign exchange and to meet the increasing
demand, it is highly necessary to focus our
attention on licorice research. Attempts on
various aspects can be made to indigenize
the commercial production of licorice. The
goal can be achieved, in a broad manner, by
the application of plant breeding techniques,
micropropagation and metabolite production
in vitro. Many investigations manifest that a
large number of food products / medically
important compounds of plant origin have
been obtained through plant tissue culture,
direct enzyme application and utilization of
whole cells in biocatalysts in fermentation
technology.
1. Cultivation and Plant Breeding
Techniques
Investigations on licorice cultivation reveal
that the plant requires sandy or sandy loam
soils with rainfall less that 50cm per year.
Manuring was not required when soils were
fertile [Singh et al 1984]. Irrigated sands
were found beneficial for the cultivation of
licorice [Mohammad and Rehman 1985].
Its existence under wide salinity and drought
conditions was on record [Mirkin et al 1971;
Aprasidi 1978]. Soils containing chloride
sulphate were reported to be advantageous
for licorice cultivation [Khafizova 1978].
Licorice cultivation helps the
reclamation/improvement of soil condition
[Varganov and Gladyshev 1981; Kel’dznaev
and Gladyshev 1982; Mohammad and
Rehman 1985].
Though earlier reports on its cultivation in
India are available [Chopra and Kapoor
1952; Kapoor et al 1955; Singh 1964,
Verma 1969; Uniyal et al 1978; Chandra
1970], subsequent reports for the past

Pages 57 - 88
decade are scandy. Singh et al [1984]
proclaimed in their view that inspite of the
attempts made on its cultivation, commercial
production was not achieved in our country.
From the agronomical point of view, it is
necessary to explore the suitable locations in
our country for its cultivation on a large
scale basis for the commercial propagation
of licorice. Morphological features which
were found useful to identify the useful
forms of licorice as described by Mikailov
and Mirzaliev [1978] and Zeng et al [1988]
may be utilized and cultivation can be
attempted with such plants by adopting
appropriate propagation methods.
Not much importance was given to breeding
experiments on licorice, as evidenced by
very few reports on this theme. Perhaps the
longer duration to attain flowering stage
would have restricted such studies. To
overcome this problem, prtoplast fusion
techniques may be of immense use.
2. How can Biotechnology Help Us to
Improve Cultivation Methods and
Commercial Production of Useful
Compounds of Licorice?
The impact of plant tissue culture on
agriculture and pharmaceutical industries is
tremendous. The plant tissue culture
techniques have become powerful tools for
studying basic and applied aspects of plant
sciences. These methods have found wide
rage of applications from propagation of
plants to the use of bioreactors and
immobilized cell technology.
Micropropagation
A recent review by Balaj et al [1988]
enunciated the usefulness of in vitro
techniques of higher plants for industrial
production of medicinally important
compounds. In vitro micropropagation
techniques were found advantageous to
supersede the problems encountered with
propagation by conventional methods. The
benefits associated with this technology are,
increase in the rate of propagation, rapid
multiplication of plants which in a particular
climate do not promote germination of
seeds, availability of plants throughout the
year, uniformity of selected genotypes,
production of uniform clones and
conservation of genetic resources.
Additionally, this type of multiplication
procedure of medicinal plants avoid the
problems connected with the loss of
biosynthesis pathways in dedifferentiated
tissues in vitro.
Different types of clonal propagation
methods are in use for the micropropagation
of various crops. The most routinely used
practice is direct propagation from existing
meristems, through which identical plants
with desired characters can be obtained
[Bajaj 1988].
Attempts have already been made on the
micropropagation of Glycyrrhiza in this
direction [Shah and Dalal 1982; Syrtanova
and Mukhitdinova 1984]. Attention can
now be focused on the utilization of somatic
embryoids derived from callus and
production of “artificial seeds” for the
multiplication of licorice. Cryopreservation
of germplasm and cultured tissues would
also meet the industrial requirement of
licorice.
Metabolite Production in vitro
Apart from the use of micropropagation
techniques, other plant tissue culture
procedure may also be considered. From a
biotechnological point of view, exploitation
of cells suspension cultures provide an
appropriate system for the production of
secondary metabolites on an economical

Pages 57 - 88
scale in bioreactors. The metabolic
pathways involved in secondary metabolism
are often complex in nature and living cells
have become the only source as it is also
turn with licorice, since the approaches on
chemical synthesis of licorice derivatives
were unsuccessful. The following are the
few familiar procedures to increase the yield
of medicinally important compounds.
Selection of High Yielding Cells for
Culture
The variability in any population for any
trait has long been recognized. The
production of a secondary metabolite by a
plant is no exception. Hence, the selection
of a variety with a high yield of a desired
natural product is a prerequisite for the
development of cell culture system [Misawa,
1985; Hoekstra et al 1988; Rhodes et al
1988; Roberts 1988]. An admirable
example has been the selection of
Catharanthus roseus plants with high yields
of vindoline, ajmalicine and serpentine
[Zenk et al 1977]. Continuous selection was
also found necessary to maintain high
yielding cell cultures [Deus and Zenk 1982].
Hayashi et al [1990b] reported that the
accumulation of soyasaponins in
Glycyrrhiza glabra suspension cultures was
under the influence of culture strains
employed. This clearly shows the
importance of selection of high yielding
cells. Similar types of studies can be
performed to derive other important
constituents of licorice. Investigations,
however, reveal contradictory reports on the
production of glycyrrhizin under culture
conditions [Wu et al 1974; Hayashi et al
1988; PCC Technol10]. These reports
perhaps demonstrate the varietal difference
to play an important role. Hence,
meticulous screening procedures to identify
high yielding strains and their utilization in
vitro might help to derive increased yield of
licorice products.
Optimization of Environmental
Conditions
The yield of secondary metabolities can be
improved by the optimization of culture
conditions [Mantell and Smith 1983].
Changes in hormonal concentration in
culture medium enhanced the alkaloid
production from Catharanthus roseus [Deus
and Zenk 1982], Thalictrum minus
[Nakagawa et al 1986] and Rauvolfia
serpentine [Yamamoto and Yamada 1987].
Other environmental factors are also critical.
Dissolved oxygen content of the medium
influenced the production of secondary
metabolites [Breuling et al 1986]. Light and
temperature also play a significant role
[Courtois and Guern 1980; Ohlssen et al
1983; Misawa 1985; Hobbs and Yeoman
1988]. Metal ions were also found to
influence secondary metabolism [Threlfall
and Whitehead 1988].
In many instances, synthesis of secondary
metabolites was in response to abiotic and
biotic stress [Timmermann et al 1984].
Fungal elictors of secondary metabolism
have been effective to increase the
phytochemicals [ Funk et al 1987; Di
Cosmo et al 1987; Holden et al 1988; van
der Heijden et al 1988]. Special attention
can be paid to find optimal environmental
conditions for the production of licorice
derivatives in vitro by supplementing the
medium with hormones, elicitors and
altering other environmental conditions.
Supplementation with Precursors and
Biotransformation
Supplementation of the media with
appropriate precursors or related
compounds, in few cases, stimulates the

Pages 57 - 88
production of secondary metabolites.
Addition of amino acids enhanced the
production of tropane alkaloids, indole
alkaloids nad ephedrine [Reinhard and
Alfermann 1990; Misawa 1985; Roberts
1988]. However, to achieve maximum
benefits of medium supplementation, it is
necessary to have a thorough comprehension
on the biosynthetic pathway of
phytoproducts which are generally
complicated.
Biotransformation has been extensively
applied in the fermentation industry
[Misawa 1985] and a classical example of
this approach was the conversion of beta-
methyldigitoxin to beta-methyl digoxin.
Fungal metabolism of licorice derivatives
was reported by Tahara et al [1985]. A
thorough screening of various
microorganism led to the isolation of an
enzyme preparation [Muro et al 1986] from
a strain of Aspergillus niger strain GRM3
which effectively catalyzed the hydrolysis of
glycyrrhizic acid.
The influence of active principals of fungal
derivative in enhancing the licorice
secondary metabolites in vitro could be
assessed. Biotransformation studies carried
out by Hayashi et al [1990a] revealed the
failure of glycyrrhizic acid production in
vitro. Further strengthening of the
investigations in this direction might help in
increasing the in vitro production of
Glycyrrhiza secondary metabolities.
Enzyme Isolation and Localization
This is an important aspect for the complete
exploitation of plant biotechnology for the
production of secondary metabolites.
However, a sound knowledge on the
characteristic of enzymes involved in the
biosynthesis of these products is highly
essential. Though this area is still infancy,
few reports are available [Robers 1988].
Similar type of studies have not been carried
out so far in licorice. Though these
investigations are of academic importance,
they unravel the metabolic pathways
involved in secondary metabolism which
ultimately have great impact on
biotechnological approaches of licorice.
Induction of Mutants
In biotechnology, induction of genetic
mutant strains of micro organisms is
ubiquitous and auxotrophic and regulatory
mutants are used extensively to produce
amino acids, nucleotides, antibiotics etc.
The impediments for such as approach with
the plant system is due to the lacuna in the
knowledge on the regulatory mechanisms of
biosynthetic pathways of secondary
metabolism. However, attempts were made
by employing chemical and physical
mutagents [Misawa 1985] and extension of
such studies may be beneficial to derive
secondary metabolites of licorice.
Plant Cell Immobilization
The development of techniques for plant cell
and enzyme immobilization has increase the
flexibility of plant cell biotechnology for the
production of pharmaceuticals. Brodelius
and Nilsson [1980] described the two major
methods of plant cell immobilization –
adhesion and entrapment. In the former
type, plant cells from suspension culture will
spontaneously bind to a suitable matrix in
bioreactor. Polyurethane foam particles
have been found to support dense cell
masses and other matrices have also been
successfully used [Mavituna and Park 1985;
Rhodes et al 1985, 1987]. In the
entrapment techniques, plant cell
immobilization is an inert gel or bead matrix
[alginate, acrylamide, carageenan, chitin or
chitosan] is one of the most successful

Pages 57 - 88
cultivation methods [Nabajima et al 1986;
Rosevear and Lambe 1986]. Increased of
secondary metabolic production and
recovery are through possible. [Morris et al
1985; Yeoman 1986].
The advantages of this technology are -
reduced rate results in increased levels of
secondary metabolite production, greater
cell to cell contact increases levels of
secondary metabolites, chemical
composition of the medium can be
optimized to maximize product and less
production costs which is an important
criteria for commercial utilization.
Immobilized plant cells can also be
employed for biotransformation, eg. [-]
codeinone to [-] codeine and digitoxin to
digoxin. Precursors and elicitors can be
employed [Roberts 1988].
The exciting developments in immobilized
technology encouraged investigators to
employ similar methods for licorice.
Favourable observations were found by
Ayabe et al [1986a]. When G.echinata cells
immobilized, the accumulation of
flavonoids, retrochalcone and echinatin
transiently increased in both cells and
media. Similar attempts with other species
of Glycyrrhiza may be of immense use to
accumulate other constituents of licorice.
3. How can Recombinant DNA
Technology Help in Genetic
Manipulation of Licorice?
The recent past has seen the development of
techniques by which goes may be
transferred into plants. In addition to the
established methods, new and refined
methods for manipulating DNA in vitro will
offer many exciting and novel opportunities.
Fundamental questions regarding the control
of plant secondary metabolism can be
tackled and ultimately, the ability to
influence secondary product accumulation,
both qualitatively and quantitatively, in
plants and in tissues grown in vitro should
be possible.
Hairy Root Cultures
Transformed root cultures (also known as
‘hairy’ root cultures) are derived from
tissues after injection with Agrobacterium
rhizogenes and have a number of attractive
features regarding the synthesis of plant
secondary products. These include ease of
culture in vitro using simple media lacking
phytohormones, reproducible and
predictable levels of product synthesis and
genetic stability over prolonged periods of
growth in vitro features which are not
usually associated with cell suspension or
callus cultures. The advantages of
transformed root tissues have been reviewed
by many investigators [Flores et al 1987;
Hamill et al 1987; Weising et al 1988].
The state-of-the art of transformation of
roots by A.rhizogenes, the exciting
developments in other phytoproducts and
the benefits associated with this technology
opened new vistas for similar type of studies
on licorice. Efforts were taken to gain more
yield of Glycyrrhiza products which are of
commercial importance. Ko et al [1989]
was successful in demonstrating the
production of glycyrrhizin in A.rhizogenes
transformed hairy roots of G.uralensis.
Similar was the report from Mitsui-Toatsu-
Chem9. The production of glycyrrhizic acid
from transgenic roots of G.uralensis was
patented by them. Investigations carried
out by Saito et al [1990], however, manifest
the absence of production of glycyrrhizin in
transformed roots of G.uralensis.
As this area is in its infancy, with regard to
licorice detailed investigations will certainly
help the efficient utilization of the A.

Pages 57 - 88
rhizogenes transformed roots. Such studies
will throw more light on the production of
licorice derivatives in vitro and ultimately
benefit the industry.
Utilization of Vectors
Apart from the Agrobacterium another
vector of potential utility in plant genetic
engineering is the cauliflower mosaic runs.
Recent evidence indicates that the
cauliflower mosaic virus is capable of
vectoring in a certain amount of DNA in
recombinant form. The limitation to use this
virus as plant vector is its limited host range.
Microinjection and Encapsulation
Micromanipulation of isolated protoplasts
for the purpose of micro injection of
recombinant DNA fragments appears to be
gaining acceptance as a potent tools to
engineer the plant cells. Injection of
liposome encapsulated DNA into protoplasts
was also evaluated by investigators. These
approaches would avoid the complexities of
constructing a recombinant vector system.
Protoplast and Organelle Fusion
Protoplasts isolated were allowed to fuse
under in vitro conditions and the resultant
hybrid protoplasts were shown to regenerate
into hybrid plants (Vasil and Vasi 1980).
Advantages to such fusion systems include
the ability to recognize and micromanipulate
the hybrid cells, use of selective markets to
distinguish true hybrids. Mitochondria and
chloroplasts when incubated with
protoplasts under appropriate conditions can
be taken up by the stable manner for several
cell generations. As a considerable number
of secondary metabolites are formed in these
organelles, the potential for organelle
transplantation appears most promising.
It is evident from the literature that genetic
engineering approaches for the improvement
of licorice is meager. Success can be
achieved with licorice too by employing
these methods. However, a clear
establishment of the molecular mechanisms
involved in the metabolic pathways of its
derivative is a prerequisite for an effort
usage of the most appropriate recombinant
[DNA] methods.
ACKNOWLEDGMENT
I grateful thank Prof. P.M. Gopinath and Dr.
G. Jayaraman for their encouragement. My
thanks are also to K.S. Usha and S.
Rajeswari for their help during the course of
writing this article.
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