PHYTOTHERAPY RESEARCH

Phytother. Res. 31: 1635–1650 (2017)

Published online 18 August 2017 in Wiley Online Library
(wileyonlinelibrary.com) DOI: 10.1002/ptr.5893

REVIEW
Toxicological Effects of Glycyrrhiza glabra
(Licorice): A Review

Somayeh Nazari,1 Maryam Rameshrad2 and Hossein Hosseinzadeh2*

1
Department of Pharmacodynamics and Toxicology, School of Pharmacy, Mashhad University of Medical Sciences, Mashhad, Iran
2
Pharmaceutical Research Center, Department of Pharmacodynamics and Toxicology, School of Pharmacy, Mashhad University of
Medical Sciences, Mashhad, Iran

Licorice (Glycyrrhiza glabra) has been considered as an herbal drug since ancient time. Nowadays, it is a well-
known spice that possesses worth pharmacological effects. However, some relevant articles have revealed nega-
tive impacts of licorice in health. By considering the great wishes in using herbal medicine, it is important to show
adverse effects of herbal medicine in health. At present, there are misunderstandings toward the safety of herbal
medicines. Herein, we gathered scientific research projects on the toxicity effects of licorice and glycyrrhizin to
highlight their safety. In this regards, we categorized our findings about the toxicity effects of licorice and
glycyrrhizin in acute, sub-acute, sub-chronic, and chronic states. Besides, we discussed on the cytotoxicity,
genotoxicity, mutagenicity, and carcinogenicity of licorice and glycyrrhizin as well as their developmental toxicity.
This review disclosed that G. glabra and glycyrrhizin salts are moderately toxic. They need to be used with cau-
tion during pregnancy. G. glabra and glycyrrhizin possess selective cytotoxic effects on cancerous cells. The most
important side effects of licorice and glycyrrhizin are hypertension and hypokalemic-induced secondary disor-
ders. Licorice side effects are increased by hypokalemia, prolonged gastrointestinal transient time, decreased
type 2 11-beta-hydroxysteroid dehydrogenase activities, hypertension, anorexia nervosa, old age, and female
sex. Copyright © 2017 John Wiley & Sons, Ltd.
Keywords: Glycyrrhiza glabra; liquorice; glycyrrhizin; acute toxicity; developmental toxicity; cancer cell.

The application of licorice goes back to ancient Assyr-

INTRODUCTION
Liquorice or licorice is an important medicinal herb with
nutritional and therapeutic values. It is mainly obtained
from the root of Glycyrrhiza glabra (Fabaceae) that is
widely cultivated in numerous temperate or semitropi-
cal parts of Europe and Asia (Federation of American
Societies for Experimental and United, 1974).
There are several critical ingredients in licorice in-
cluding saponins, flavonoids (Isbrucker and Burdock,
2006), and coumarins (Kinoshita et al., 2005). Special
yellow color and sweet taste of this spice are resulted
from flavonoids and glycyrrhizin, respectively.
Glycyrrhizin (Fig. 1), a triterpenoid saponin, is the most
important constituent of licorice and is found in the
form of potassium and calcium salts of 18β-glycyrrhizic
acid (also known as glycyrrhizic or glycyrrhizinic acid
and a glycoside of glycyrrhetinic acid) in licorice root
and ammonium salt in the commercial preparations
(Isbrucker and Burdock, 2006). Licorice root
glycyrrhizin concentration is ranging up to 15% that
the presence of this saponin could be changed by the ex-
traction method and the root source (Fugh-Berman and
Ernst, 2001).
Licorice has a long history of use around the world,
which could be called as ‘the grandfather of plants’.
* Correspondence to: Hossein Hosseinzadeh, Pharmaceutical Research
Center, Department of Pharmacodynamics and Toxicology, School of
Pharmacy, Mashhad University of Medical Sciences, Mashhad, Iran.
E-mail: hosseinzadehh@mums.ac.ir
ian, Egyptian, Chinese, and Indian culture. Tradition-
ally, it has been used for treating asthma, hoarseness of
voice, cough, and lung diseases. It was a remedy for
mouth ulcerations, diseases of liver, and heart burning.
It has been prescribed for artery diseases, heart palpita-
tion, and angina. Besides, it has been suggested to treat
bladder and kidney pain, kidney stones, fever, neural-
gia, skin, and eye diseases (Fiore et al., 2005). Nowa-
days, it is a well-known spice for its taste and worth
pharmacological properties. Licorice extract and
glycyrrhizin derivatives are used for treating inflamma-
tion, allergy, and peptic ulcer and applied to cosmetic
preparation because of their antisensitizing and skin-
whitening effects (Hayashi and Sudo, 2009). The phar-
macology of licorice and its constituents has been
reviewed extensively by Nassiri Asl and Hosseinzadeh
(2007, 2008, 2015). G. glabra and its bioactive com-
pounds have antioxidant (Sil and Chakraborti, 2016),
antiviral (Nassiri Asl and Hosseinzadeh, 2007), antimi-
crobial, antiprotozoal, antiinflammatory, and immuno-
modulatory activities (Nassiri Asl and Hosseinzadeh,
2008). They could reverse drug resistance of
methicillin-resistant Staphylococcus aureus infections
(Gaur et al., 2016) and evoke hepatoprotective
(Chigurupati et al., 2016; Nassiri Asl and Hosseinzadeh,
2008) and antitumor effects (Nassiri Asl and
Hosseinzadeh, 2008). Besides, they could be used as a
remedy in cardiovascular diseases and diabetes owing
to their antiplatelet aggregation activity, thrombin inhib-
itory effects, vasorelaxant effects, antiangiogenic

Received 30 May 2017

Revised 22 July 2017
Copyright © 2017 John Wiley & Sons, Ltd. Accepted 24 July 2017
1636 S. NAZARI ET AL.

attributed to the inhibition of 11-beta-hydroxysteroid

Figure 1. Chemical structure of glycoside of glycyrrhetinic acid,
glycyrrhizin.
properties, and positive effects on peroxisome
proliferator-activated receptor ɤ (PPARɤ) (Nassiri Asl
and Hosseinzadeh, 2008; Hosseinzadeh and Nassiri
Asl, 2015). It has been reported that they have worth
sedative and muscle relaxant activities, antidepressant
and anticonvulsant effects, memory-enhancing activity
(Nassiri Asl and Hosseinzadeh, 2008), and protective ef-
fects against ischemia–reperfusion-induced cerebral and
skeletal muscle injury (Hosseinzadeh et al., 2005). Their
efficacy to treat recurrent aphthous stomatitis,
postextubation sore throat, gastrointestinal and skin dis-
orders, cancer, and hepatitis B and C have been evalu-
ated in clinical studies (Nassiri Asl and Hosseinzadeh,
2008; Hosseinzadeh and Nassiri Asl, 2015). Licorice
could be used in patients with postural hypotension
attributable to autonomic diabetic neuropathy (Basso
et al., 1994) and hirsutism and polycystic ovary syn-
drome (Armanini et al., 2004). The efficacy of licorice
in combination with Cinnamomum verum, Hypericum
perforatum, Paeonia lactiflora, Tribulus terrestris, and
lifestyle intervention has been proved in women with
polycystic ovary syndrome (Arentz et al., 2017). Neo-
Minophagen C® is a glycyrrhizin-based drug that its ef-
ficacy in treating viral hepatitis B is being approved
(Chen et al., 2014).
A number of mechanisms could cause the beneficial
effects of licorice. The basic of these effects could be
dehydrogenase (11βHSDH), key enzyme in converting
cortisol to cortisone (Luyckx, 2012), and accumulation
of glucocorticoids with antiinflammatory and mineralo-
corticoid properties. Besides, it possesses inhibitory
effects on both phospholipase A2 activity and platelet
aggregation (Okimasu et al., 1983). Structural similari-
ties between licorice constituents and steroid
hormones (Baker, 1995) could explain their mineralo-
corticoid and glucocorticoid activities (Armanini
et al., 1983). Furthermore, licorice-derived compounds
possess affinity to PPARɤ (Kuroda et al., 2010)
(Fig. 2).
Ploeger et al. (2001) reviewed the pharmacokinetic of
glycyrrhizic acid comprehensively (Ploeger et al., 2001).
Yamamura et al. (1992) showed that in normal human
after oral consumption of glycyrrhizin (100 mg), it was
partly absorbed in intact form from the intestinal, and
it was not detectable in plasma; however, a small
amount was detected in the urine. Hydrolysis of
glycyrrhizin by intestinal flora evoked glycyrrhetic acid
appearance in plasma but not in the urine. In intrave-
nous (IV) route, the plasma concentration of
glycyrrhizin decreased in a biphasic manner. Interest-
ingly, glycyrrhetic acid was not found in plasma. For
glycyrrhizin in three dosing experiments, they estimated
2.7–4.8 h for terminal half-life, 37–64 mL/kg for the
apparent volume of the central compartment,
59–98 mL/kg for the steady-state distribution volume,
and 16.25 mL/kg/h for total body clearance (Yamamura
et al., 1992).
However, it has been reported that glycyrrhetic acid is
detectable in plasma not only after oral consumption
but also after IV or IP administration of glycyrrhizin to
rat. IV or intraperitoneal (IP) bioavailability of
glycyrrhizin in rats is same and is more than its oral bio-
availability. However, the plasma glycyrrhetic acid level

Figure 2. Both cortisol and aldosterone bind with equal affinity to mineralocorticoid receptors that are activated predominantly with aldoste-
rone. 11-Beta-hydroxysteroid dehydrogenase type 2 (11-βHSD2) converts cortisol to its inactive form, cortisone, and evokes corticosteroid
specificity. By inhibitory effects of Glycyrrhiza glabra and glycyrrhizin on 11-βHSD2, the MR receptors are activated by cortisol, leading to
accumulation of glucocorticoids with antiinflammatory and mineralocorticoid properties, severe hypokalemia, and mineralocorticoid-related
hypertension (for more details, refer to the text). MR, mineralocorticoid receptor; GR, glucocorticoid receptor; blocking sign: , side

effects: , protective effects: . [Colour figure can be viewed at wileyonlinelibrary.com]

Copyright © 2017 John Wiley & Sons, Ltd. Phytother. Res. 31: 1635–1650 (2017)
 TOXICOLOGY OF LICORICE
following administration via different routes is identical
to each other (Egashira et al., 2003).
In comparison of pure glycyrrhizin bioavailability
with glycyrrhizin in licorice extract, it has been reported
that several components in licorice root extract interact
by intestinal absorption of glycyrrhizin and reduce its
oral bioavailability in comparison with pure glycyrrhizin
(Cantelli-Forti et al., 1994).
Pharmacokinetic parameters of glycyrrhizin (20, 50,
or 100 mg/kg) and glycyrrhetic acid (2.5 or 12.5 mg/kg)
in rat plasma after single treating in IV injection showed
that the decline in plasma is biexponential and concen-
tration dependent and has a saturable elimination rate.
Their pharmacokinetic obeys a two-compartment phar-
macokinetic model with Michaelis–Menten elimination
(Ishida et al., 1989; Tsai et al., 1992). The results showed
that pharmacokinetic parameters of glycyrrhizin is non-
linear owing to its saturable hepatic metabolism (Tsai
et al., 1992). Besides, it has been suggested that the
pharmacokinetic of glycyrrhizin is dose dependent be-
cause of its saturable biliary excretion rates that causes
its dose-dependent biliary clearance (Ishida et al.,
1992). Van Rossum et al. (1999) showed that in patients
with chronic hepatitis C infection, the pharmacokinetic
of glycyrrhizin is linear up to 200 mg (Van Rossum et al.,
1999).
Glycyrrhetic acid and its parent, glycyrrhizin, have a
great albumin-binding affinity which could be saturated
in higher concentration (Ploeger et al., 2001). Koga et al.
(2008) showed that the serum concentration of the
glycyrrhizin is mostly dependent to elimination of
glycyrrhizin from liver into bile, and decrease in serum
albumin level has no effects on the serum concentration
of glycyrrhizin (Koga et al., 2008). The pharmacokinetic
of glycyrrhizin and its total body clearance in hepatitis
and cirrhosis patients are directly dependent to the liver
function (Yamamura et al., 1995).
Based on a multi-center retrospective review of data
from European and Brazilian poison centers gathered
between 2006 and 2010, G. glabra had been reported
as 1 of the 10 most frequently plant foods that caused
adverse effects (Lüde et al., 2016).
METHODS
A comprehensive literature search was performed using
relevant key words including G. glabra, liquorice,
licorice, glycyrrhizin, acute toxicity, sub-acute toxicity,
sub-chronic toxicity, chronic toxicity, mutagenic, devel-
opmental, miscarriage, cancer, and clinical trial in the
following databases: PubMed, Scopus, Google Scholar,
and Web of Sciences. All kinds of relevant articles,
abstracts, or books were included. Furthermore, bibliog-
raphies of eligible articles were also examined for addi-
tional relevant studies. Both in vivo and in vitro studies
were included to this investigation. No time or language
limitation was considered in this review. Congress
abstracts, as well as studies on the other species of
Glycyrrhiza, were considered ineligible for inclusion,
but glycyrrhetinic acid as the metabolite of glycyrrhizin
was included.
The toxic effects of licorice and glycyrrhizin were cat-
egorized in the following main headings: toxicological
findings, mutagenicity, genotoxicity, carcinogenicity
Copyright © 2017 John Wiley & Sons, Ltd.
1637
and developmental toxicity, cytotoxicity, side effects,
and drug interactions.
TOXICOLOGICAL FINDINGS
Acute toxicity test
Acute toxicity test is the first toxicity analysis that is de-
scribed from single exposure during 14 days. The most
often used species are mice and rats. The outcomes are
used for evaluating the intrinsic toxicity of the chemical
[approximate lethal dose (LD50)], target organ/s, inter-
species sensitivity, and dose selection for longer term
studies. It is important to notice that LD50 value is not
a constant level, and many factors influence on it. It de-
pends on the animal age, weight, and strain. Also, type
of feed and animal caging can have an effect on it
(Eaton and Gilbert, 2013). According to the LD50
values, the chemical could be allocated to one of the five
category toxicity levels: dangerously toxic (<1 mg/kg),
extremely toxic (1–50 mg/kg), very toxic (50–500 mg/kg),
moderately toxic (500–5000 mg/kg), slightly toxic
(5000–15,000 mg/kg), and practically non-toxic
(>15,000 mg/kg) (Bone and Mills, 2013).
G. glabra root aqueous and ethanol extract did not
show any mortality up to 1000 mg/kg with a single oral
dose, in female albino rats. Behavioral assessment
showed that the animal’s alertness, spontaneous loco-
motor activity, and reactivity to touch behavior were de-
creased by all doses up to 3 h (Chowdhury et al., 2013).
Single oral dose of G. glabra root aqueous extract was
safe up to 1500 mg/kg in both sexes of Swiss albino mice.
No mortality was reported during the following 14 days.
However, some physical changes were observed at
higher doses up to 1500 mg/kg within the second hours
(Gupta et al., 2016).
The acute toxicity values of glycyrrhizin salts have
been reported from 412 mg/kg (potassium
glycyrrhizinate, IV) to 12700 mg/kg (crude ammonium
glycyrrhizate, orally), depending on the salt type tested.
Besides, according to gathered data, there is a signifi-
cant difference in LD50 values after IV or IP administra-
tion in comparison with oral exposure (Federation of
American Societies for Experimental and United,
1974), which might be affected from first-pass metabo-
lism and lower absorption in the oral treatment
(Hosseinzadeh et al., 2013) (Table 1).
Besides, it has been shown that the IP administration
of 2 g/kg of α-isomer of 18-glycyrrhetic acid was lethal to
adult female Sprague–Dawley rats. It induced atrioven-
tricular block between 130 and 140 min (Rossi et al.,
1999).
Skin irritation
For evaluating the effect/s of a chemical on the skin, it
acutely exposed on the rabbit skin (Eaton and Gilbert,
2013). Glycyrrhetinic acid (100 mg/mL, about 0.5 cc)
was introduced on the shaven skin of a rabbit’s back,
both normal and injured. After 24 and 72 h, there was
no sign of edema or erythema on the both treated
groups. It showed that glycyrrhetinic acid had no
primary skin irritant effects (Finney et al., 1958).
Phytother. Res. 31: 1635–1650 (2017)
1638 S. NAZARI ET AL.

Table 1. The median lethal dose (LD50) values of Glycyrrhiza glabra and glycyrrhizin salts

Route of
Compound Animals administration LD50 value (mg/kg) Ref.

G. glabra Leaf ethanol extract Mice IP 3030 Yazdi et al. (2011)

Root ethanol and aqueous Rat Oral No mortality was Chowdhury et al. (2013)
extract observed at >1000
Root aqueous extract Mice Oral No mortality was Gupta et al. (2016)
observed at >1500
Glycyrrhizin — Oral >2000 Sasaki et al. (2002)
Glycyrrhetinic acid Mice IP 308 Finney et al. (1958)
oral 610
Ammonium glycyrrhizate Mice IP (crude) 1050 Federation of American
Oral (crude) 12,700 Societies for Experimental
IP 1070 and United (1974)
Diammonium glycyrrhizate Oral 9600
IP 1250
Potassium glycyrrhizate Oral 1220
SC 697
IV 412
IM 695
Dipotassium glycyrrhizate Oral 8100
IP 1400
Potassium glycyrrhizate (crude) Oral 12,400
IP 1260

IP, intraperitoneal injection; SC, subcutaneous injection; IV, intravenous injection; IM, intramuscular injection.

Sub-acute toxicity
Sub-acute toxicity is described from three to four differ-
ent dosages of the chemical in repeated administration
during 14–28 days of exposure. Rats and dogs in both
sexes are used. The outcomes give a platform about
the toxicity of the chemical after repeated administra-
tion. The results are used for dose selection in sub-
chronic studies (Eaton and Gilbert, 2013).
It has been shown that G. glabra root aqueous extract
(100, 250, and 500 mg/kg, gavage, for 15 days, in male
Wistar rats) has a strong inhibitory effects on the
adrenal–pituitary axis in a dose-dependent manner. It
induced a hypermineralocorticoidism state with
decrease in the level of adrenocorticotropic hormone,
cortisol, aldosterone, and potassium and increase in
the rennin concentration and natrium (Al-Qarawi
et al., 2002).
Furthermore, the inhibitory effect of glycyrrhizin on
the pituitary–adrenal stress response has been proved
in sub-acute exposure. The resistance of mice to
prolonged cold stress was evaluated by treating the ani-
mals with 0.4% ammoniated glycyrrhizin in drinking
water for 4 days before inducing cold stress. The next
animals were exposed to cold stress (for 8 h with 5°C).
In comparison with control, ammoniated glycyrrhizin
decreased survival time. Besides, it has been shown that
treated rats with 0.4% ammoniated glycyrrhizin in
drinking water for 1 week were more sensitive to 48-h
fasting and showed severe hypoglycemia versus control
(Kraus, 1958). However, Finney et al. (1958) showed
the safety of glycyrrhetinic acid (10, 20 mg/kg, IM, three
times a week for 4 weeks) in rats. According to their
study, it did not have clinical effects (Finney et al., 1958).
Former study found the adverse influence of
glycyrrhizin on the iron liver state in a short-term treat-
ment. It found that feeding male Sprague–Dawley
weanling rats with 2% ammoniated glycyrrhizin in diet,
Copyright © 2017 John Wiley & Sons, Ltd.
for 14 days, decreased liver iron content and increased
fecal iron exertion significantly. Besides, treated animals
had a significant weight gain. However, this diet had no
effect on the level of the other minerals (zinc and mag-
nesium) (West et al., 1979) (Table 2).
Sub-chronic toxicity
Sub-chronic tests are performed on both sex of two spe-
cies, usually rodents (rat and mouse) and dog, in at least
three doses for about 30 to 90 days. Gathered data are
used for evaluating affected organs, ‘no observed ad-
verse effect level’ (NOAEL), ‘lowest observed adverse
effect level’, and a worth prediction of appropriate doses
for chronic studies. Biochemical and hematological
parameters, body weight, and food consumption are
the other evaluated factors (Eaton and Gilbert, 2013).
Long-term exposure to water extract of licorice (500,
1000, and 2000 mg/kg, orally, for 9 weeks) showed that
NOAEL is higher than 2000 mg/kg. Besides, it had no
significant toxicity effects in reproductive organs (Shin
et al., 2008).
Sub-chronic glycyrrhizin consumption (0.1–1 mg/mL,
in drinking water, for 12 weeks) had cardiovascular side
effects in male Sprague–Dawley rats. It induced miner-
alocorticoid excess syndrome accompanied with sys-
temic hypertension, water retention, hypernatremia,
and hypokalemia. It proved the positive effects of
glycyrrhizic acid on increasing right atrial pressure and
pulmonary arterial pressure. Histological study proved
pulmonary hypertension (Ruszymah et al., 1995).
In comparison with female B6C3F1 mice, male
B6C3F1 mice were more sensitive to chronic exposure
with disodium glycyrrhizinate in drinking water, where
maximum tolerable dose in male mice was 0.15% and
in female mice was 0.3% after 10-week exposure
(Kobuke et al., 1985).
Phytother. Res. 31: 1635–1650 (2017)
 TOXICOLOGY OF LICORICE 1639

Table 2. Sub-acute toxicity findings of Glycyrrhiza glabra and glycyrrhizin salts

Compound Animals Route of administration Findings Ref.

G. glabra root Rat 100, 250, and 500 mg/kg, Decrease in the plasma concentration of Al-Qarawi et al. (2002)
+
aqueous extract orally for 15 days cortisol, ACTH, aldosterone, and K dose
dependently
Increase in the plasma concentration of renin
+
and Na in a dose-dependent manner
Suppression of the adrenal–pituitary axis dose
dependently
Ammoniated Mice 0.4% in drinking water for Increase in the mortality rate in comparison Kraus (1958)
glycyrrhizin 4 days before inducing cold with the untreated animals
stress
Rat 0.4% in drinking water for Fall in the blood glucose in comparison with the
7 days before inducing 48-h untreated animals
fasting
Ammoniated Rat 2% mixed in rat chow, for Increase in body weight and average fecal West et al. (1979)
glycyrrhizin 14 days weight
Increase in the fecal excretion of iron and a
decrease in liver iron level
Glycyrrhetinic Rat 10 or 20 mg/kg, IM, three The animals were alive with no clinical effects Finney et al. (1958)
acid times a week for 4 weeks At necropsy, all tissues were normal except for
a slight thinning of the lipid in the zona
glomerulosa of the adrenal glands.

ACTH, adrenocorticotropic hormone; IM, intramuscular injection.

Sub-chronic toxicity findings of G. glabra and Chronic administration of disodium glycyrrhizinate to

glycyrrhizin salts are summarized in Table 3.
Chronic toxicity
Chronic toxicity test evaluates the effect of chemical on
the development of toxicity in longer than 3-month ex-
posure. In rodents, its longitude is about 6 months to
2 years, and in non-rodents, it is about 1 year or longer.
The evaluation of maximum tolerable dose and the uri-
nary metabolite indexes, carcinogenicity, and physiolog-
ical and pharmacokinetic aspects of the chemicals are
the main goal of the chronic toxicity studies (Eaton
and Gilbert, 2013).
B6C3F1 mice up to maximum tolerated dose for
96 weeks (male up to 0.15% and female up to 0.3% in
drinking water) had no tumorigenicity effects (Kobuke
et al., 1985). Male black molly fishes were exposed to
G. glabra root extract (1, 2, and 4 mg/L, for 45 days).
In this model, NOAEL and first observed effect concen-
tration were 1 and 4 mg/L, respectively. Liver toxicity
was reported in the treated fishes (Radhakrishnan
et al., 2005). Furthermore, NOAEL for subcutaneous in-
jection of monoammonium glycyrrhizinate in rat has
been introduced as 25 mg/kg for 26-week exposure
(Akasaka et al., 2008). NOAEL has different severity
between sexes. Respectively, it has been estimated as
800 and 400 mg/kg/day for oral consumption of licorice

Table 3. Sub-chronic toxicity findings of Glycyrrhiza glabra and glycyrrhizin salts

Compound Animals Route of administration Findings Ref.

Water extract Rat 500, 1000, and Slight decrease in the prostate Shin et al. (2008)
of licorice 2000 mg/kg, weight and daily sperm production
orally for 9 weeks without any changes in the other
sexual organs
Glycyrrhizic Rat 0.1 and 1 mg/mL in Increase in tail blood pressure as Ruszymah et al. (1995)
acid drinking water for well as right atrial pressure
+
12 weeks Increase in serum Na level and
+
decrease in K content
Thickening of the pulmonary
vessels
Disodium Male and female 0.04, 0.08, 0.15 The maximum tolerated dose Kobuke et al. (1985)
glycyrrhizinate B6C3F1 mice and 0.3, 0.6, or was 0.3 and 0.15% for female
1.25% in drinking and male, respectively
water for 10 weeks Decrease in body weight with MTD
Death of all animals with 0.6
and 1.25% in 10th week which
was accompanied with starvation

Copyright © 2017 John Wiley & Sons, Ltd. Phytother. Res. 31: 1635–1650 (2017)
1640 S. NAZARI ET AL.

flavonoid oil in female and male rats after 90-day treat-
ment (Nakagawa et al., 2008b).
Chronic toxicity findings of G. glabra and glycyrrhizin
salts are gathered in Table 4.
MUTAGENICITY, GENOTOXICITY,
CARCINOGENICITY, AND DEVELOPMENTAL
TOXICITY
A carcinogen, capable of causing cancer, could be
genotoxic and mutagenic. In toxicology studies, poten-
tial carcinogenicity of a substance is evaluated by both
in vitro and in vivo studies. Based on the duration re-
quired to conduct the test, these studies are categorized
to (1) short-term tests that last days to a few weeks that
evaluate mutagenesis and transformation in cell culture;
(2) intermediate-term tests with longitude about weeks
up to a year that include qualitative and quantitative
analysis of preneoplasia; and (3) chronic bioassay in
animals that persists 6 months to 2 years (Eaton and
Gilbert, 2013).
Developmental toxicology is the other aspect of toxi-
cological studies. It evaluates the adverse effects of a
substance on the development of the organism resulting
from exposure to the agent/s prenatal or postnatal until
the time of puberty. It encompasses structural
malformations, growth retardation, functional or meta-
bolic impairment, and/or death of the organism. Repro-
ductive toxicity and teratogenicity were also estimated.
Reproductive toxicity is a part of developmental toxic-
ity, in which the associations between specific exposures
of the father or mother and pregnancy are assessed. The
substances that cause structural birth defects are called
teratogen (Eaton and Gilbert, 2013).
Antimutagenic effects of G. glabra (Tanaka et al.,
1987; Shankel and Clarke, 1990; Hrelia et al., 1996;
Inami et al., 2017; Wang et al., 1991) against

Table 4. Chronic toxicity findings of Glycyrrhiza glabra and glycyrrhizin salts

Compound Animals Route of administration Findings Ref.

Disodium Male and female 0.04, 0.08, and 0.15% Dose related decrease in water Kobuke et al.
glycyrrhizinate B6C3F1 mice (both sex) and 0.3% (female), consumption (1985)
in drinking water for 96 weeks No effects on tumor incidence in
which followed 14 weeks comparison with control group
with normal drinking water
(110 weeks)
G. glabra propylene Male black Feeding with 1, 2, and The mortality rate for 4 mg/L was Radhakrishnan
glycol root extract molly fish 4 mg/L for 45 days 100% on 7th day, for 2 mg/L was et al. (2005)
34% on 15th day and for 1 mg/L
was 17% on 25th day. The latest
group were alive even after 45th day
With 4 mg/L, fish’s appetite was
decreased after 3 days
In a dose-dependent manner, body
weight was decreased, and damages
to the liver were increased
Monoammonium Rat 25, 75, and 225 mg/kg, Mortality was happened in the Akasaka et al.
glycyrrhizinate SC, 26 weeks highest dose (2008)
Increase of serum bilirubin and
decrease of serum chloride and
potassium with 75 or 225 mg/kg
Decrease of erythrocytes and
increase of reticulocytes, neutrophils,
and monocytes in 225 mg/kg
Increase of kidney weights dose
dependently with swollen and
degenerated tubuli contorti with
brownish pigment
Increase of liver weights with
hypertrophy of the hepatocytes in
75 or 225 mg/kg
Licorice Rat 400, 600, 800, and 1600 mg/kg, Decrease in erythrocytes, hemoglobin, Nakagawa et al.
flavonoid oil (both sex) orally for 90 days and hematocrit (2008b)
Increase in MCH, MCHC, and WBC
Prolongation of APTT and PT
Hemorrhage
Increase in urine volume and urine
+
Na level

MCH, mean corpuscular hemoglobin; MCHC, mean corpuscular hemoglobin concentration; WBC, white blood cell count; APPT, activated
partial thromboplastin time; PT, prothrombin time.

Copyright © 2017 John Wiley & Sons, Ltd. Phytother. Res. 31: 1635–1650 (2017)
 TOXICOLOGY OF LICORICE
ethyl methanesulfonate, N-methyl-N0 -nitro-N-
nitrosoguanidine, and ribose-lysine Maillard model
using Ames test have been proved (Zani et al., 1993).
Licorice root extract evokes antagonistic effects against
arecoline and fluoride-induced genotoxicity (Lee Ching
et al., 1996). However, there are controversial reports
about mutagenesis effects of G. glabra. Licorice flavo-
noid oil showed negative results in the reverse mutation
assay test, chromosomal aberration test, and liver and
peripheral blood micronucleus test. Nevertheless, at
higher concentrations (>0.6 mg/mL) in the short-period
test with metabolic activation, chromosomal aberration
test was positive. Bone marrow micronucleus test was
negative, but the ratio of polychromatic erythrocytes
to total erythrocytes was decreased at 5000 mg/kg/day
for 2 days (Nakagawa et al., 2008a). In line with this
study, Chandrasekaran et al. (2011) proved the non-
genotoxic effects of G. glabra extract. The Ames II™
tests in two bacterial Salmonella typhimurium strains
(TA98 and TAMix) with or without metabolic activa-
tion, for standardized deglycyrrhizinated extract of G.
glabra root (1.6–501 μg/mL), were negative. Besides,
in vitro chromosome aberration test and micronucleus
formation in CHO-K1 cells were negative at 4–
40 μg/mL. However, mitotic index was decreased by
the highest concentration of the extract (40 μg/mL)
(Chandrasekaran et al., 2011).
Ishidate et al. (1984) have shown non-mutagenic ef-
fects of disodium glycyrrhizinate (up to 4 mg/mL) in
Ames test with TA92, TA1535, TA100, TA1537, TA94,
and TA98 strains, with or without metabolic activation
(Ishidate et al., 1984).
Furthermore, the result of Ames test for
monoammonium glycyrrhizinate (100 to 5000 μg/plate)
with five S. typhimurium strains (TA98, TA100,
TA102, TA1535, TA1537) without and with metabolic
activation by S9 was negative. In vitro chromosomal ab-
breviation test in human lymphocyte culture
with/without S9 showed hemolysis at 1500 μg/mL; how-
ever, no chromosomal or chromatid damage was de-
tected. In vivo mouse bone marrow micronucleus test
was used for evaluating chromosomal or mitotic dam-
ages. There were no evidences in its mutagenic effects
up to 240 mg/kg, IV, after 24 and 48 h. However, a slight
ataxia has been recorded with 240 mg/kg, IV, during
5 min after injection. It has no effects on the polychro-
matic erythrocytes to normochromatic erthrocytes ra-
tios and the numbers of micronucleated polychromatic
erythrocytes (Akasaka et al., 2009).
According to comet assay test, administration of sin-
gle dose of glycyrrhizin (2000 mg/kg, orally) did not in-
duce any DNA damages in mice bone marrow,
glandular stomach, colon, liver, kidney, urinary bladder,
lung, and brain tissues after 3- or 8-h treatment (Sasaki
et al., 2002).
Conversely, in the other studies, the mutagenic activ-
ity of aqueous extract of licorice has been reported in
Ames test for TA100 strain, but not in TA98 (Martínez
et al., 1999), only at the highest concentration (25 mg/mL)
(Abudayyak et al., 2015).
Sheu et al. (1986) showed that glycyrrhizate has no
heritable genetic toxicity in mice. Besides, they evalu-
ated ammonium glycyrrhizate dominant lethal effects
in mice and rats. Female mice were mated with male
mice that had been treated with ammonium
glycyrrhizate (2 or 2.5%, in the diet chew, for 8 weeks).
Copyright © 2017 John Wiley & Sons, Ltd.
1641
The fetuses were analyzed at 12–15th gestation days.
Ammonium glycyrrhizate had no infertility or dominant
lethal effects in mice. However, it increased dead im-
plants in mated female rats with male rats that had been
treated with ammonium glycyrrhizate (4%, in the diet
chew, for 10 weeks) (Sheu et al., 1986).
It has been shown that IV injection of
monoammonium glycyrrhizinate with 25 and 75 mg/kg
in rats has no-observed effects in the maternal genera-
tion and for the fetuses, respectively. Rat prenatal and
postnatal development study showed that no-observed
effect level (NOEL) was 25 mg/kg for parental F0 gener-
ation. It was higher than 150 mg/kg for parental F1 gen-
eration and on the development of the offspring (F2
generation). According to rat and rabbit embryo–fetal
developmental studies, NOEL for the dams was
75 mg/kg. NOEL for rat and rabbit fetuses was 25 and
75 mg/kg, respectively. Toxicokinetic study of pregnant
and lactating rats showed that monoammonium
glycyrrhizinate and its derivatives have very low concen-
tration in the milk of lactating rats (Yoshida et al., 2011).
The other study revealed the reproductive and devel-
opmental safety of aqueous licorice extract. In this
study, male and female rats were exposed to the extract
(500, 1000, and 2000 mg/kg in drinking water) from
9 weeks before mating until copulation day and from
2 weeks before mating to gestation day 19, respectively
(Shin et al., 2005). According to the former study,
treating pregnant rats with ammonium glycyrrhizinate
(21, 239, and 680 mg/kg/day, from days 7 to 17 of gesta-
tion) evoked polydipsia in the dams. Specially in higher
doses, it induced a slight embryo lethality, skeletal
anomalies, and renal ectopy (Mantovani et al., 1988).
The conversion of cortisone (11-
dehydrocorticosterone in rodent) into cortisol (cortico-
sterone) has a pivotal role in the fetal lung maturation.
In rats, glycyrrhetinic acid consumption (1000 mg/kg/
day, in chow diet) during pregnancy (on gestational
day 13 until delivery) by inhibition of 11βHSDH im-
paired fetal lung development. It was accompanied with
decrease in the level of surfactant protein-A, amniotic
fluid lecithin/sphingomyelin ratio, and lung surfactant
(Hundertmark et al., 2002).
Investigations of Itami et al. (1985) on disodium
glycyrrhizinate effects on pregnant rats and their off-
spring proved its non-teratogenic effects. Pregnant rats
were fed with disodium glycyrrhizinate (2, 0.4, and
0.08% in diet) from 0 to 20th day of gestation. Mother
food intake and gain weight during pregnancy were nor-
mal. The numbers of corpora lutea and implants of
dams and the number of live fetuses and intrauterine
dead fetuses per litter were not significant in compari-
son with control. The sex ratios and their body weight,
the placental weight, the degrees of ossification in the
sternebrae and caudal vertebrae of the fetuses, or the
live birth index, survival rate, or body weight gain of
the offspring within 8 weeks after birth had no differ-
ence with control group. However, in treated pregnant
rats with highest doses, body weight gain after delivery
was significantly lower. In disodium glycyrrhizinate-
treated rats, fetus skeletal variation and one fetus with
dilatation of the renal pelvis were reported in non-
significant differences versus control (Itami et al.,
1985). It has been suggested that licorice is not a major
teratogen substance, and its effects on the risk of still-
births should be evaluated (Choi et al., 2013).
Phytother. Res. 31: 1635–1650 (2017)
1642 S. NAZARI ET AL.
In humans, licorice intake during pregnancy signifi-
cantly reduces gestational age (Strandberg et al., 2001;
Cuzzolin et al., 2010) and induces preterm delivery
(Strandberg et al., 2002). It could be due to the effect
of licorice constituents on the metabolism of cortisol
and prostaglandins (Lockwood, 2002). Licorice con-
sumption during pregnancy is associated with cognitive
dysfunction (Raikkonen et al., 2009) and some change
in hypothalamic–pituitary–adrenocortical axis function
(Räikkönen et al., 2010) in children in a dose-dependent
manner. Furthermore, it may increase the rate of pre-
eclampsia in mothers who have familial or genetic his-
tory of this disease (Hauksdottir et al., 2015).
Mutagenicity, genotoxicity, carcinogenicity, and de-
velopmental toxicity findings of G. glabra and
glycyrrhizin salts are summarized in Table 5.
CYTOTOXICITY: SAFETY COMPARISON IN
NORMAL VERSUS CANCEROUS CELLS
Licorice and its main constituents decrease cell viability
in different types of cancer cells such as gastrointestinal
(Khazraei-Moradian et al., 2017; Nourazarian et al.,
2016; Wang et al., 2015; Shen et al., 2015;
Mehdinejadiani et al., 2015; Khazraei-Moradian et al.,
2014; Hibasami et al., 2006; Hibasami et al., 2005), leuke-
mia (Hibasami et al., 2006; Hibasami et al., 2005; Chueh
et al., 2012), breast (Lee et al., 2013; Dong et al., 2007; Jo
et al., 2005), prostate (Yo et al., 2009; Thiugnanam et al.,
2008; Hawthorne and Gallagher, 2008; Thirugnanam
et al., 2008), cervix, uterus (Lee et al., 2008), and mela-
noma (Abe et al., 1987) cancer cell lines. Besides, their
tumor-suppressive effects have been evaluated in vivo
(Yang et al., 2015; Lee et al., 2007; Agarwal and
Mukhtar, 1991; Shibata et al., 1991; Shiota et al., 1999).
Wang and Nixon (2001) reviewed the worth effects of
licorice in cancer therapy (Wang and Nixon, 2001).
Herein, we focus to licorice cytotoxic and tumor speci-
ficity with a brief view to its cytotoxic mechanisms
(Table 6).
According to MTT assay data, licorice ethanol extract
showed cytotoxic activity (63% at 0.24 mg/mL) in Vero
cells (Martínez et al., 1999). Treating CT25, HT29, and
Hek293 cells with full extract of licorice (0, 20, 50, 100,
and 200 μg/mL) for 24 h decreases the viability of cancer
cell line significantly more than non-cancerous cell line
where IC50 value in CT26 and HT29 reported about
50 μg/mL and in HEK293 about 200 μg/mL. It was
showed that licorice extract increases apoptosis in
cancerous cells but not in the non-cancerous cells. Cell
viability was decreased by increasing dose (Khazraei-
Moradian et al., 2017). Besides, ethanol extract of
roasted licorice (2.5, 5, 7.5, and 10 μg/mL) reduced the
viability of MDA-MB-231 cells, dose and time depen-
dently, without any cytotoxic effects in hFOB1.19 hu-
man osteoblast cells and murine bone marrow-derived
macrophages, both normal cells. In vivo, administration
of this extract (0.5, 1, and 2 mg/kg, gavage, daily for the
first 5 days followed by weekly administration for
44 days) suppressed tumor growth and bone destruction
in the model of MDA-MB-231 metastatic human breast
cancer cell injection, subcutaneously, into mice tibia
(Lee et al., 2013). The other study showed that G. glabra
Copyright © 2017 John Wiley & Sons, Ltd.
(200 μg/mL) inhibited proliferation of HT-29 cell line in
a dose-dependent and time-dependent manner. Cell
proliferation was decreased to 52, 62, and 68% after
24, 48, and 72 h, respectively. The cytotoxicity of G.
glabra was attributed to downregulation of HSP90 gene
expression and apoptosis (Nourazarian et al., 2016). A
Chinese group attributed glycyrrhizin cytotoxicity in
BGC-823 cells to regulation of cell cycle via Wnt/β-
catenin signaling pathways. They showed that
glycyrrhizin decreases cell viability and arrests the cells
in G1/S phase. Besides, it (40 μM) inhibits cell adhesion
and migration and the level of β-catenin, Bcl-2, cyclin
D1, and survivin (Wang et al., 2015).
Trypan blue dye exclusion method showed that
treating KATO III and HL-60 cells with glycyrrhizin
(3 mg/mL) for 3 days reduces cell viability to 50%
(Hibasami et al., 2005). In addition, glycyrrhizin IC50
values in androgen-dependent LNCaP and androgen-
independent DU-145 cells after 48-h exposure have
been reported as 8.5 and 12.5 μM, respectively
(Thirugnanam et al., 2008). It has been shown that
glycyrrhetinic acid is more toxic than glycyrrhizin in
B16 melanoma cells, where cell growth is inhibited
completely with glycyrrhetinic acid (10 μg/mL) but
inhibited up to 40% with glycyrrhizin (200 μg/mL) after
3-day incubation (Abe et al., 1987). The other study
mentioned that cytotoxic effects of glycyrrhizin are less
than glycyrrhetinic acid. Treating SiHa cells with
glycyrrhetinic acid (100 μM, 24 h) decreased cell
viability about 66%, whereas glycyrrhizin with that
same concentration evoked no cell death (Lee et al.,
2008). The incubation of WEHI-3 cells with glycyrrhizin
(300 μM, for 48 h) inhibited cell viability up to 50%
(Chueh et al., 2012).
Some physicochemical descriptors influence on the
cytotoxicity and tumor specificity of licorice flavonoids.
The molecular volume and number of phenolic OH
groups correlate with cytotoxic effects. With arise in
the number of sugar units in the molecule, a reduction
in cytotoxicity was shown. Solvation energy was associ-
ated with the tumor specificity (Ohno et al., 2013).
The other studies have mentioned on the selective cy-
totoxicity of glycyrrhetinic acid. The IC50 values of the
normal SFME cells and cancerous r/m HM-SFME-1
cells treated with glycyrrhetinic acid for 24 h have been
reported 18 and 7.3 μM, respectively (Yu et al., 2010).
Glycyrrhetinic acid cytotoxicity in HepG2 cells have
been attributed to inhibition of cell growth at G1 phase
and induction of apoptosis (Y S, H N, and S S., 2005).
LICORICE AND GLYCYRRHIZIN-RELATED
SIDE EFFECTS
According to animal and human studies, some compli-
cation may be caused by licorice and glycyrrhizin con-
sumption. The main side effects could be categorized
to hypertension and hypokalemic-induced secondary
complications (Table 7).
Aldosterone and cortisol bind with equal affinity to
mineralocorticoid receptors that are activated predomi-
nantly with aldosterone. Cortisol but not aldosterone is
metabolized to its inactive form by 11βHSDH type 2.
Licorice by inhibiting 11βHSDH activity causes the
Phytother. Res. 31: 1635–1650 (2017)
 TOXICOLOGY OF LICORICE 1643

Table 5. Mutagenicity, genotoxicity, carcinogenicity, and developmental toxicity of Glycyrrhiza glabra and glycyrrhizin salts

Results

Test Compound Animal or cell culture Positive Negative Ref.

Ames test Escherichia coli Licorice flavonoid oil WP2uvrA/pKM101a ± S9b + Nakagawa et al. (2008a)
Salmonella TA98, TA100, TA1535, +
typhimurium and TA1537c ± S9
Standardized TA98 and TAMixd ± S9 + Chandrasekaran et al.
de-glycyrrhizinated (2011)
extract of G. glabra
root
Disodium TA92, TA1535, TA100, + Ishidate et al. (1984)
glycyrrhizinate TA1537, TA94, and
TA98 ± S9
Monoammonium TA98, TA100, TA102, + Akasaka et al. (2009)
glycyrrhizinate TA1535, TA1537 ± S9
e
Aqueous extract of TA100 and TA98 ± S9 + Martínez et al. (1999),
licorice Abudayyak et al. (2015)
Comet assay Glycyrrhizin Not explained + Sasaki et al. (2002)
Chromosomal aberration Standardized CHO-K1 cells ± S9 +f Chandrasekaran et al.
de-glycyrrhizinated (Chinese hamster ovary) (2011)
extract of G. glabra
root
Monoammonium Human lymphocyte + Akasaka et al. (2009)
glycyrrhizinate culture ± S9
Licorice flavonoid oil Chinese hamster lung +g Nakagawa et al. (2008a)
(CHL/IU)
cells, for 6, 24,
and 48 h ± S9
Micronucleus test In vivo: rat bone marrow +h
In vivo: rat peripheral +
blood and liver
Standardized CHO-K1 ± S9 + Chandrasekaran et al.
de-glycyrrhizinated (2011)
extract of G. glabra
root
Monoammonium In vivo: rat bone marrow +i Akasaka et al. (2009)
glycyrrhizinate
Dominant lethal effect Ammonium Mice + Sheu et al. (1986)
glycyrrhizate Rat +
Heritable translocation test Mice +
Reproductive toxicity Mice +
Monoammonium Rat + Yoshida et al. (2011)
glycyrrhizinate
Aqueous licorice extract Rat + Shin et al. (2005)
Developmental toxicity Ammonium glycyrrhizinate Rat + Mantovani et al. (1988)
Glycyrrhetinic acid Rat + Hundertmark et al. (2002)
Disodium glycyrrhizinate Rat + Itami et al. (1985)
a
Escherichia coli strain.
b
Prepared from the livers of rats pretreated with phenobarbital and 5,6-benzoflavone.
c
Salmonella typhimurium strains.
d
A mixture of the S. typhimurium strains TA7001, TA7002, TA7003, TA7004, TA7005, and TA700 is used for the detection of base pair
substitutions.
e
Positive results just for TA100.
f
But, mitotic index was decreased by the highest concentration, 40 μg/mL.
g
Positive results just for 6 h with S9 mix.
h
Positive results just with high dose (5 g/kg/day).
i
Slight ataxi.

mineralocorticoid receptors to be activated by both cor- visual problem secondary to hypertension, severe hypo-
tisol and aldosterone, a state leading to a mineralocorti- kalemia and metabolic alkalosis, rhabdomyolysis, acute
coid excess, mineralocorticoid-related hypertension, tubular necrosis, and uremic and alkalosis-induced
Copyright © 2017 John Wiley & Sons, Ltd. Phytother. Res. 31: 1635–1650 (2017)
1644 S. NAZARI ET AL.

Table 6. The effect of Glycyrrhiza glabra and its main constituents on normal and cancer cell line under in vitro conditions

Type Dose Toxic responses Ref.

Normal cells Vero cells Licorice ethanol extract
(240 μg/mL)
HEK293 Full extract of licorice
(0, 20, 50, 100,
and 200 μg/mL) for 24 h
hFOB1.19 Ethanol extract of roasted
human osteoblast licorice (2.5, 5, 7.5, and
cells 10 μg/mL) for 24 and 48 h
Bone marrow-derived Ethanol extract of roasted
macrophages licorice (2.5, 5, 7.5,
and 10 μg/mL) for 5 days
SFME Glycyrrhetinic acid for 24 h
Cancerous cells CT-26 Full extract of licorice (0,
HT-29 20, 50, 100, and
200 μg/mL) for 24 h
MDA-MB-231 Ethanol extract of roasted
cells licorice (2.5, 5, 7.5, and
10 μg/mL) for 24 and 72 h
r/m HM-SFME-1 Glycyrrhetinic acid for 24 h
HT-29 G. glabra (200 μg/mL) for
24, 48, and 72 h
BGC-823 Glycyrrhizin (40 μM)
KATO III Glycyrrhizin (3 mg/mL)
HL-60 for 3 days
LNCaP Glycyrrhizin for 48 h
DU-145
B16 melanoma Glycyrrhetinic acid
cells (10 μg/mL) for 72 h
Glycyrrhizin (200 μg/mL)
for 72 h
SiHa Glycyrrhetinic acid
(100 μM) for 24 h
Glycyrrhizin (100 μM)
for 24 h
WEHI-3 Glycyrrhizin (300 μM)
for 48 h
HepG2 Glycyrrhizin for 24 h
Glycyrrhetinic acid for 24 h
HepG2 Glycyrrhizin for 48 h
MHCC97-H
4T1 G. glabra methanol extract
(50, 100, 200, 400,
800 μg/mL), for 24,
48, and 72 h
Cytotoxic effects: 63% Martínez et al. (1999)
IC50 value: 200 μg/mL Khazraei-Moradian
et al. (2017)
Increased the viability of cells Lee et al. (2013)
Had no effect on the cell viability
IC50 value: 18 μM Yu et al. (2010)
IC50 value: 50 μg/mL Khazraei-Moradian
et al. (2017)
IC50 value: 8.9 μg/mL for Lee et al. (2013)
24 h and
6.9 μg/mL for 72 h
IC50 value: 7.3 μM Yu et al. (2010)
Cell proliferation was Nourazarian et al.
decreased to 52, 62, (2016)
and 68%, respectively
Decreased cell viability Wang et al. (2015)
and arrested the cells in
G1/S phase
Inhibited cell adhesion and
migration and the level of
β-catenin, Bcl-2, cyclin D1,
and survivin
Reduced cell viability to 50% Hibasami et al. (2005)
IC50 value: 8.5 μM Thirugnanam et al.
IC50 value: 12.5 μM (2008)
Growth is inhibited completely Abe et al. (1987)
Growth is inhibited up to 40%
Induced cell death about 66% Lee et al. (2008)
No cell death
Inhibited cell viability up to 50% Chueh et al. (2012)
IC50 value: >1200 μM Y S, H N, and S S.
IC50 value: 80 μM (2005)
IC50 value: 1000 μM Zhang et al. (2017)
Induced cell death up to Hamta et al. (2014)
62% in a dose-dependent
and time-dependent manner

neurological syndrome. Besides, licorice-induced hypo-
kalemia evokes QT prolongation and torsade de
pointes. This state totally is called pseudo-
hyperaldosteronism with low concentration of aldoste-
rone and renin in serum (Luyckx, 2012; Schröder et al.,
2015; Bisogni et al., 2014; Iida et al., 2006; Panduranga
and Al-Rawahi, 2013; Eid et al., 2011; Sontia et al.,
2008; Farese et al., 1991; Ishiguchi et al., 2004; Chubachi
et al., 1992; Velickovic-Radovanovic et al., 2011; Daniş
Copyright © 2017 John Wiley & Sons, Ltd.
et al., 2015; Heidemann and Kreuzfelder, 1983; Saito
et al., 1994). In one case, carpal tunnel syndrome was
reported that has been attributed to the salt and water
retention and edema accompanied with licorice con-
sumption (Tacconi et al., 2009). Asthenia and muscle
cramps (De Putter and Donck, 2014), occupational
asthma (Cartier et al., 2002), and contact allergy
(O’Connell et al., 2008) are the other recorded licorice-
induced complications (Fig. 2).
Phytother. Res. 31: 1635–1650 (2017)
 TOXICOLOGY OF LICORICE 1645

Table 7. Glycyrrhiza glabra and glycyrrhizin salt-related side effects

Direct toxicity Model Compound Maine complication Ref.

Cardiovascular Male Wistar rat Glycyrrhizin 375 mg/day, Increase in mineralocorticoid activity: Kageyama et al. (1994)
toxicity 10 days, BID, gavage increase in mean blood pressure,
hypokalemia, and hypernatremia
suppression of both plasma rennin
activity and plasma aldosterone
Male Sprague– Glycyrrhizin, 0.1 and Increase in tail blood pressure Ruszymah et al. (1995)
Dawley rats 1 mg/mL in drinking Increase in right atrial pressure
water for 12 weeks Thickening of the pulmonary vessels
Hypokalemia and hypernatremia
57-year-old man Licorice, 900 g/week Hypertension Schröder et al. (2015)
for 3–4 months Acute visual impairment
Severe hypokalemia
72-year-old man Liquorice, 2 oz/week Hypertension Bisogni et al. (2014)
for 1 month Metabolic alkalosis and severe
hypokalemia
Increase in rhabdomyolysis indexes
Myoglobinuria
Electrocardiography showed U-wave,
wide T-wave, and apparent prolongation
of QTc
77-year-old man Glycyrrhizin + ACEI Pseudo-aldosteronism Iida et al. (2006)
more than 10 years
38-year-old woman Licorice root tea, TID Polymorphic ventricular tachycardia Panduranga and
for 2 months Al-Rawahi (2013)
46-year-old woman Herbal tea containing Hypertension Eid et al. (2011)
licorice, one to two Hypokalemia
cups per day for 7 years Reduced plasma aldosterone and rennin
55-year-old woman Large amount of licorice, Hypokalemia and hypertension Sontia et al. (2008)
daily for 4 years
70-year-old man Licorice candies, Hypertension Farese et al. (1991)
60–100 g/day, 4–5 years Hypokalemia
Rhabdomyolysis
90-year-old woman Antacid that contained Hypokalemia, hypertension, myoclonus, Ishiguchi et al. (2004)
licorice and metabolic alkalosis
Nephrotoxicity 72-year-old man Glycyrrhizin Convulsion, delirium, muscle weakness, Chubachi et al. (1992)
90–240 mg/day, hypertension, edema, hypokalemia,
1–2 years acute renal failure
39-year-old female Herbal medicine Hypertension, hypokalemia, muscle Velickovic-Radovanovic
containing licorice/daily weakness, acute renal failure et al. (2011)
for 8 weeks
49-year-old man Herbal medicine Hypokalemia, confusion, somnolence, Daniş et al. (2015)
containing licorice, rhabdomyolysis, acute renal failure
for 1.5 years
Myopathy 54-year-old man Licorice 20–25 g/day, Hypokalemia, rhabdomyolysis, Heidemann and
for 2 years myoglobinuria Kreuzfelder (1983)
78-year-old man Glycyrrhizin 280 mg/day Muscular weakness, acute renal failure, Saito et al. (1994)
for 7 years hypokalemic rhabdomyolysis
Other 44-year-old woman Chewing pure licorice Ankle edema, bilateral nocturnal hand Tacconi et al. (2009)
sticks, 3 days pain, paresthesias in median-innervated
fingers, bilateral carpal tunnel syndrome
52-year-old man licorice1.5 g/day for Severe asthenia and muscle cramps, De Putter and Donck (2014)
2 months hypertension
33-year-old woman Handling several Occupational asthma Cartier et al. (2002)
products at work
including licorice
35-year-old woman Recovery cream Contact allergy O’Connell et al. (2008)
that contains licorice

BID, twice a day; ACEI, angiotensin converting enzyme inhibitors; TID, three times a day.

Besides, hypertension, increase in right atrial pres- (Kageyama et al., 1994; Ruszymah et al., 1995). In
sure, pulmonary hypertension, and thickening of the rats, aqueous extract of G. glabra (50, 100, and
pulmonary vessels have been recorded in animal studies 200 mg/kg/day for 30 days, IP) induced harmful effects
Copyright © 2017 John Wiley & Sons, Ltd. Phytother. Res. 31: 1635–1650 (2017)
1646 S. NAZARI ET AL.
on hepatocyte integrity and increased concentrations of
alkaline phosphatase, aspartate aminotransferase, and
alanine aminotransferase in serum (Hussein, 2013).
LICORICE AND GLYCYRRHIZIN-RELATED
DRUG INTERACTION
Phase I metabolism mediated by CYP 450 isoenzymes,
phase II metabolism mediated by mainly UDP
glucuronosyltransferases, and phase III metabolism me-
diated by transporter proteins play a pivotal role in drug
metabolism (Dietrich et al., 2003). Every change in these
pathways is important to notice in possible drug interac-
tions in combination therapy.
Licorice possesses some effects on the CYP enzymes
that are important in combination with narrow thera-
peutic index drugs. The licorice root-derived isoflavan
(glabridin) showed inhibitory effects on CYP3A4, 2B6,
and 2C9 (Kent et al., 2002). In vitro and in vivo,
glycyrrhetinic acid shows inhibitory effects on CYP3A
(Li et al., 2010; Liu et al., 2011; Lv et al., 2016), 2C9,
and 2C19 enzymes (Liu et al., 2011; Lv et al., 2016).
However, former studies showed that prolong intake
of licorice and glycyrrhizin induces CYP isoenzyme ac-
tivity on rodent liver (Paolini et al., 1998; Paolini et al.,
1999) that are proved using Vivid CYP450 Screening
Kits (Hou et al., 2012) and human study (Tu et al., 2010).
It has been shown that G. uralensis increases cyclo-
phosphamide teratogenicity potential due to induction
of CYP 2B activity (Park et al., 2011). This pharmacoki-
netic interaction should be noticed for G. glabra owing
to its inducing effects on CYP isoenzymes. Besides,
(3R)-vestitol, 4-hydroxyguaiacol apioglucoside, and
liquiritigenin 7,40 -diglucoside compounds isolated from
G. uralensis, a Glycyrrhiza species, possess CYP3A in-
hibitory effect. However, its glycyrrhizin content was in-
active for CYP3A4 inhibition (Tsukamoto et al., 2005).
It is important to notice that glycyrrhetinic acid in-
duces UDP glucuronosyltransferases in rat liver in vivo
(Lee and Ho, 2013).
In vitro, glycyrrhetinic acid possesses inhibitory ef-
fects on P-glycoprotein-mediated transport (Yoshida
et al., 2006; Nabekura et al., 2008). On one hand, it could
increase the efficacy of cancer chemotherapy (Nabekura
et al., 2008), but on the other hand may increase normal
cytotoxicity, decrease oral bioavailability of some drugs,
enhance blood–brain barrier permeability and neuro-
toxicity, and decrease renal elimination. This interaction
is important for P-glycoprotein substrate drugs such as
digoxin and loperamide (Lin and Yamazaki, 2003).
However, Hou et al. (2012) reported activator effects
for both glycyrrhizin and glycyrrhetinic acid on P-
glycoprotein function in vitro (Hou et al., 2012).
In this regard, there is a probable pharmacokinetic in-
teraction between ombitasvir/paritaprevir/ritonavir and
glycyrrhizin due to its inhibitory effects on P-
glycoprotein and its CYP3A activating potential. How-
ever, Zha et al. (2015) reported that no dose adjustment
is needed in this cotreatment therapy except of clinical
monitoring that this combination therapy may increase
glycyrrhizin AUC up to 50% (Zha et al., 2015).
The other study showed that glycyrrhizin decreases
Cmax and AUC0-t of ribavirin and its metabolite. It has
Copyright © 2017 John Wiley & Sons, Ltd.
been attributed to reduction in ribavirin transportation
in GI track due to inhibitory effect of glycyrrhizin and
glycyrrhetinic acid on nucleoside transporters (Liao
et al., 2016).
Besides, licorice influences on the metabolism of ste-
roid compounds. It enhances the activity of 5α reductase
(Fugh-Berman and Ernst, 2001). Takeuchi et al. (1991)
proposed activator effects for glycyrrhizin and
glycyrrhetinic acid on aromatase enzyme in homoge-
nized ovary tissue culture (Takeuchi et al., 1991). How-
ever, recently, licorice flavonoid isoliquiritigenin shows
inhibitory effects on this enzyme in MCF7 cell culture
(Ye et al., 2009).
Hypokalemia induced by licorice could increase the
risk of digoxin toxicity (Harada et al., 2002). Licorice
has interaction in coadministration with cilostazol,
potassium-depleting drugs, thiazide, and loop diuretics
that may cause hypokalemia, with immunosuppressive
agents that may decrease drug clearance, with predniso-
lone that may potentiate the action or increase levels of
drug, with midazolam and omeprazole that may de-
crease drug level, and with antihypertensive medica-
tions that decrease effectiveness of drugs (Bone and
Mills, 2013).
It has been shown that glycyrrhizin increases the skin
absorption of diclofenac sodium (Nokhodchi et al.,
2002), and glycyrrhetinic acid potentiates hydrocorti-
sone activity in skin (Teelucksingh et al., 1990).
CONCLUSION
Nowadays, there is a great scope in utilizing herbal med-
icine worldwide in direct or indirect way, and so many
research articles proved the worth traditional use of
them. However, in contrast to the general opinion,
herbal drugs are composed of many known and un-
known substances that may possess harmful impacts on
human health.
Herein, we have summarized the possible toxicity of
G. glabra and glycyrrhizin, its main secondary metabo-
lite. By considering their LD50 values and Bone and
Mills classification, G. glabra is moderately toxic. Be-
sides, it should be noticed that their toxicity depends
on the rout of administration and type of salts. It is clear
that the risk of toxicity after oral administration is low-
est than IP or IV administration due to first-pass metab-
olism and lower absorption after the oral dispensation.
Sub-acute exposure with G. glabra and glycyrrhizin
salts induces inhibitory effects on the adrenal–pituitary
axis and depletes the liver iron contents. The most im-
portant side effects by administration of licorice and
glycyrrhizin are hypertension and hypokalemic-induced
secondary disorders. Some factors may increase the risk
of toxicity with licorice and glycyrrhizin. The susceptibil-
ity to glycyrrhizin is increased by hypokalemia,
prolonged gastrointestinal transient time, decreased
11βHSDH type 2 activity, hypertension, anorexia
nervosa, old age, and female sex.
According to the literature reviews, G. glabra and
glycyrrhizin salts are not major teratogens and possess
weak mutagenicity, genotoxicity, carcinogenicity, and
developmental toxicity effects. However, the use of G.
glabra and its components should be cautioned in preg-
nancy and neonates. Clinical studies showed that the use
Phytother. Res. 31: 1635–1650 (2017)
 TOXICOLOGY OF LICORICE
of licorice during pregnancy is accompanied with reduc-
tion of gestational age, preterm delivery, and some
change in hypothalamic–pituitary–adrenocortical axis
function and cognitive dysfunction in delivered children.
In addition, it needs to be used with warning in woman
with familial history of pre-eclampsia.
Normal cells differ from cancer cell in response to cy-
totoxic drugs. Gathered data demonstrate the selective
cytotoxic effects of licorice and its main constituents on
cancerous cells that could engage them as a valuable ad-
juvant treatment in cancer therapy.
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