Saudi Pharmaceutical Journal 29 (2021) 820–832

Contents lists available at ScienceDirect

Saudi Pharmaceutical Journal

journal homepage: [Link]

Original article

Cinnamomum zeylanicum Blume (Ceylon cinnamon) bark extract

attenuates doxorubicin induced cardiotoxicity in Wistar rats
Jayasinghe Arachchige Nirosha Sandamali a, Ruwani Punyakanthi Hewawasam b,⇑,
Kamani Ayoma Perera Wijewardana Jayatilaka b, Lakmini Kumari Boralugoda Mudduwa c
a
Department of Medical Laboratory Science, Faculty of Allied Health Sciences, University of Ruhuna, 80000, Sri Lanka
b
Department of Biochemistry, Faculty of Medicine, University of Ruhuna, 80000, Sri Lanka
c
Department of Pathology, Faculty of Medicine, University of Ruhuna, 80000, Sri Lanka

a r t i c l e i n f o a b s t r a c t

Article history: Anti-tumour efficacy of doxorubicin is hindered by the cumulative dose-dependent cardiotoxicity
Received 23 February 2021 induced by reactive oxygen species during its metabolism. As Cinnamomum zeylanicum has proven
Accepted 13 June 2021 antioxidant potential, objective of this study was to investigate the cardioprotective activity of
Available online 20 June 2021
Cinnamomum bark extract against doxorubicin induced cardiotoxicity in Wistar rats. Physicochemical
and phytochemical analysis was carried out and dose response effect and the cardioprotective activity
Keywords: of Cinnamomum were determined in vivo. 180 mg/kg dexrazoxane was used as the positive control.
Doxorubicin
Plant extracts were free of heavy metals and toxic phytoconstituents. In vivo study carried out in
Cardiotoxicity
Oxidative-stress
Wistar rats revealed a significant increase (p < 0.05) in cardiac troponin I, NT-pro brain natriuretic pep-
Cinnamomum zeylanicum bark extract tide, AST and LDH concentrations in the doxorubicin control group (18 mg/kg) compared to the normal
Antioxidant effect control. Rats pre-treated with the optimum dosage of Cinnmamomum (2.0 g/kg) showed a significant
Myeloperoxidase reduction (p < 0.05) in all above parameters compared to the doxorubicin control. A significant reduction
was observed in the total antioxidant capacity, reduced glutathione, glutathione peroxidase, glutathione
reductase, superoxide dismutase and catalase activity while the lipid peroxidation and myeloperoxidase
activity were significantly increased in the doxorubicin control group compared to the normal control
(p < 0.05). Pre-treatment with Cinnamomum bark showed a significant decrease in lipid peroxidation,
myeloperoxidase activity and significant increase in rest of the parameters compared to the doxorubicin
control (p < 0.05). Histopathological analysis revealed a preserved appearance of the myocardium and
lesser degree of cellular changes of necrosis in rats pre-treated with Cinnamomum extract. In conclusion,
Cinnamomum bark extract has the potential to significantly reduce doxorubicin induced oxidative stress
and inflammation in Wistar rats.
Ó 2021 The Author(s). Published by Elsevier B.V. on behalf of King Saud University. This is an open access
article under the CC BY-NC-ND license ([Link]

Abbreviations: WHO, World Health Organization; DNA, Deoxyribonucleic acid; NADPH, Nicotinamide adenine dinucleotide phosphate hydrogen; SOD, Superoxide
dismutase; ROS, Reactive oxygen species; DPPH, 2,2-diphenyl-1-picrylhydrazyl; ABTS, 2,20 -azino-bis(3-ethylbenzothiazoline-6-sulfonic acid); ABEC, Aqueous bark extract of
Cinnamomum zeylanicum; FRAP, Ferric reducing antioxidant power; NO, Nitric oxide; IP, Intraperitoneal; ELISA, Enzyme-linked immunosorbent assay; cTnI, Cardiac troponin
I; NT-pro BNP, N terminal- pro brain natriuretic peptide; AST, Aspartate aminotransferase; LDH, Lactate dehydrogenase; PBS, Phosphate buffered saline; GSH, Reduced
glutathione; GPx, Glutathione peroxidase; GR, Glutathione reductase; MPO, Myeloperoxidase; USA, United States of America; H & E, Haematoxylin and eosin; MDA,
Malondialdehyde.
⇑ Corresponding author.
E-mail addresses: jansandamali@[Link] (J.A.N. Sandamali), rphewawasam@[Link] (R.P. Hewawasam), ayomaj@[Link] (Kamani Ayoma Perera
Wijewardana Jayatilaka).
Peer review under responsibility of King Saud University.

Production and hosting by Elsevier

[Link]
1319-0164/Ó 2021 The Author(s). Published by Elsevier B.V. on behalf of King Saud University.
This is an open access article under the CC BY-NC-ND license ([Link]
J.A.N. Sandamali, R.P. Hewawasam, K.A.P.W. Jayatilaka et al. Saudi Pharmaceutical Journal 29 (2021) 820–832

1. Introduction diphenyl-1-picrylhydrazyl (DPPH) radicals and 2,20 -azino-bis(3-et

hylbenzothiazoline-6-sulfonic acid) (ABTS) radical cations
Anthracyclines are a group of antibiotics which play a key role (Ranasinghe et al., 2013). Although it has already been proven that
in the treatment of cancer in the modern era and they are among Cinnamomum zeylanicum Blume has a significant antioxidant activ-
the essential medicine listed by the World Health Organization ity, effect of its bark extract has never been investigated against
(WHO) (McGowan et al., 2017). These anti-neoplastic agents are doxorubicin induced cardiotoxicity in an animal model. Hence,
used in the treatment of a wide range of malignancies including the objective of this study was to investigate the ameliorative
haematologic malignancies such as leukemia, lymphoma and solid effects of Cinnamomum zeylanicum Blume bark against doxorubicin
organ tumors such as sarcoma, breast cancer, multiple myeloma induced cardiac injury by the attenuation of oxidative stress and
and lung cancer. Among the members of the anthracycline family, structural cardiomyocyte changes in rats.
doxorubicin is the most effective and commonly used antineoplas-
tic drug (Shaker et al., 2018).
2. Material and methods
The clinical usefulness of doxorubicin is hampered by its dose
related cardiotoxicity (Khattry et al., 2009; Minotti et al., 2004).
2.1. Collection of Cinnamomum zeylanicum Blume bark
Although the underlying mechanism of doxorubicin induced car-
diotoxicity is not fully understood, reactive oxygen species induced
The cultivated Cinnamomum zeylanicum Blume bark was col-
oxidative stress is considered as the most accepted phenomenon
lected and identified according to the descriptions given by
(Chatterjee et al., 2010; Mobaraki et al., 2017). Some of the other
Jayaweera (1982). The species identification was confirmed by
proposed mechanisms are alterations in protein synthesis, deoxyri-
the curator of the National Herbarium, Royal Botanical Gardens,
bonucleic acid (DNA) damage, lipid peroxidation, cell membrane
Peradeniya, Sri Lanka. A voucher specimen (2015/PG/VS/02) was
lesions, induction of immunogenic reactions and dysregulation of
deposited at the Department of Biochemistry, Faculty of Medicine,
calcium homeostasis. As a result of doxorubicin metabolism, a
University of Ruhuna, Sri Lanka.
semiquinone form is produced which is mediated by reduced
flavoenzymes such as nicotinamide adenine dinucleotide phos-
phate hydrogen (NADPH)-cytochrome P450 reductase (Mitry and 2.2. Standardization of plant material
Edwards, 2016; Mobaraki et al., 2017). This semiquinone is able
to complex with iron (Fe2+) and the free radical complex which 2.2.1. Physicochemical analysis
reduce molecular oxygen to superoxide. In addition to this, enzy- The bark parts (cut into small pieces) of Cinnamomum were
matic pathway produces free oxygen radicals by accepting elec- dried at 40 °C until a constant weight was reached and finely
trons from nicotinamide adenine dinucleotide (NADH) or NADPH grounded. The powdered plant material was taken for the physic-
when doxorubicin gets reduced at Complex I of the electron trans- ochemical analysis. Tests for moisture content, extractable matter
port chain. This reaction sequence is known as the redox cycling and heavy metal analysis were followed according to the WHO
and it may be very harmful as even a low amount of doxorubicin standards (1996). Microscopic analysis of the plant was carried
is capable of producing many superoxide radicals. As doxorubicin out according to the WHO (2011) guidelines on quality control
has high affinity for cardiac-specific phospholipid named cardi- and standardization of plant materials.
olipin found in the internal membrane of mitochondria, it enters
the mitochondria easily and suppresses the respiratory chain 2.2.2. Phytochemical analysis
(Goormaghtigh et al., 1990; Aryal and Rao, 2016). Doxorubicin also Phytochemical screening of Cinnamomum zeylanicum bark was
diminishes the activity of cardiac enzymes such as catalase, super- followed to identify medicinally active substances found in the
oxide dismutase (SOD) and glutathione S- plant. Plant material was dried at 40 °C for three days, ground coar-
transferase. Furthermore, the heart tissues are more susceptible sely, and extracted in distilled water or organic solvents according
to damage by doxorubicin as the antioxidant storage is low in heart to the method used. The relevant extracts were subjected to qual-
tissues compared to other organs in the body (Halestrap, 2006). itative phytochemical screening assays for the detection of anthra-
Therapeutic strategies which have the ability to supplement the cene glycosides, cyanogenic glycosides, cardenoloid glycosides,
cellular endogenous defence systems have been recognized as saponins, polyphenols, alkaloids, flavonoids, tannins, reducing sug-
hopeful tactics to combat the oxidative stress conditions and in ars and proteins (Trease and Evans, 2009; Mushtaq et al., 2014;
this respect, natural products which enhance endogenous antioxi- Yusuf et al., 2014).
dants, have been found to offer protection in preventing doxoru-
bicin induced cardiotoxicity (Mukherjee et al., 2003). 2.2.3. Total polyphenol content and in vitro antioxidant activity of
Cinnamomum zeylanicum Blume (Ceylon Cinnamon) is a commonly aqueous bark extract of Cinnamomum zeylanicum (ABEC)
found medicinal plant in Sri Lanka with proven antioxidant activ- Constant weight of the Cinnamomum zeylanicum bark was
ity. It belongs to the family Lauraceae and has been reported to ensured by drying the plant material at 40 °C and coarsely ground.
have many beneficial activities such as being an antioxidant, Then the plant material (2.50 g) was mixed with distilled water
antimicrobial, anticancer, anti-inflammatory, antidiabetic, anti- (60 mL) and extracted using a reflux system for the preparation
mutagenic and as an anti-tyrosinase agent (Rao and Gan, 2014). of the aqueous extract. After cooling, the mixture was filtered using
It was previously reported that the bark extract of Ceylon cinna- a cheese cloth and the final volume was concentrated to 50 mL.
mon has numerous antioxidant compounds, which can effectively Then final concentration of the refluxed ABEC was 0.05 g/mL. A
counteract with reactive oxygen species (ROS) such as hydroxyl concentration series (1–500 mg/mL) of plant extract was prepared
radicals, superoxide anions as well as other free radicals. Many for the in vitro antioxidant assays.
in vitro studies reported the antioxidant effect of Cinnamomum zey- Folin-Ciocalteau spectrophotometric method described by
lanicum Blume in the recent past (Rao and Gan, 2014; Ghosh et al., Singleton et al. (1999) was used to measure the total polyphenol
2015; Ranasinghe and Galappaththy, 2016; Premakumara and content in ABEC. Result was expressed as milligrams of gallic acid
Abeysekera, 2020). The essential oils obtained from the bark of Cin- equivalent per gram of extract dry weight (mgGAE/g dw). Ferric
namomum zeylanicum Blume and eugenol have shown very power- reducing antioxidant power (FRAP) was determined according to
ful antioxidant activities (Chericoni et al., 2005) and in vitro studies the method described by Galketiya et al. (2017). DPPH assay was
revealed that Cinnamomum bark extracts effectively scavenged 2,2- used to determine the radical scavenging ability of the ABEC
821
J.A.N. Sandamali, R.P. Hewawasam, K.A.P.W. Jayatilaka et al. Saudi Pharmaceutical Journal 29 (2021) 820–832

according to modified method of Rahman et al. (2015). Nitric oxide Group I (normal control); distilled water administered orally for
(NO) assay was performed according to the modified Griess reac- 14 days, single IP injection of normal saline (10 mL/kg) on the
tion (Boora et al., 2014). Following formula was used to calculate day11 after 16hr fast
the radical scavenging activity in terms of percentage inhibition Group II (plant extract control); freeze dried ABEC (2.0 g/kg)
of free radicals by the sample. administered orally for 14 days, single IP injection of normal saline
Percentage inhibition = [(Abs control – Abs test)]/ (Abs (10 mL/kg) on the day 11 after 16hr fast
control)]  100 Group III (doxorubicin control); first distilled water adminis-
IC50 value (concentration of the plant extract or standard tered orally for 14 days, then, a single dose of doxorubicin
required to inhibit DPPH radical formation by 50%) was finally cal- (18 mg/kg) intraperitoneally on the day 11 after 16hr fast
culated to measure the antioxidant activity of the bark. Group IV (plant + doxorubicin); freeze dried ABEC at 2.0 g/kg
L- Ascorbic acid was used as the standard for DPPH, NO radical administered orally for 14 days, a single dose of doxorubicin at
scavenging assay and the FRAP assay. 8 mg/kg administered intraperitoneally on the day 11 after 16hr
fast
2.3. Preparation of plant extract for in vivo studies Group V (positive control group); Distilled water was adminis-
tered orally for 14 days, then a single injection of dexrazoxane
The bark (cut into small pieces) of Cinnamomum was dried at (180 mg/kg, IP) was administered 30 min before the single dose
40 °C until a constant weight was reached and coarsely ground. of doxorubicin (18 mg/kg) was administered intraperitoneally on
Ground plant material (24.00 g) was refluxed in distilled water the day 11
for 4hrs to be compatible with the extraction method used by On day15, all Wistar rats were sacrificed and blood was drawn
the traditional ayurvedic medical practitioners in Sri Lanka. The fil- by cardiac puncture for the estimation of AST activity, LDH activity,
tered mixture was freeze dried after adjusting the final volume to N terminal- pro brain natriuretic peptide (NT-pro BNP) cTnI con-
500.0 mL. centration, concentration and myeloperoxidase (MPO, EC
[Link]) activity. A portion of heart tissue was collected into phos-
phate buffered saline (PBS) to prepare the homogenate for the esti-
2.4. Experimental animals
mation of anti-oxidant parameters such as total antioxidant level,
reduced glutathione (GSH), glutathione peroxidase (GPx, EC
Healthy, Wistar albino rats in both sexes which are 6–8 weeks
[Link]), glutathione reductase (GR, EC [Link]), catalase (EC
old weighing 175 ± 25 g were purchased from the Medical
[Link]) activity, SOD (EC [Link]) activity, and the lipid peroxida-
Research Institute, Colombo, Sri Lanka. They were kept in a well-
tion. Remaining portion of heart tissues was stored in 10% formal
ventilated animal house located in the Faculty of Medicine, Univer-
saline for the histological assessment myocardial damage.
sity of Ruhuna, Sri Lanka. A standard laboratory diet of rat pellets
was used for feeding and water ad libitum. Rats were used in exper-
iments after they were allowed to acclimatize to the settings of the 2.7. Assessment of blood parameters
new animal house such as the temperature (23 ± 2 °C), relative
humidity (50 ± 5%), and 12hr light–dark cycle) for one week prior The separated serum was used for the estimation of cardiac
to the experiments. Approval was obtained from the Ethical biomarkers and MPO activity. NT-pro BNP and cTnI concentrations
Review Committee of the Faculty of Medicine, University of were estimated based on sandwich-Enzyme-linked immunosor-
Ruhuna, Sri Lanka (23.10.2014:3.10). bent assay (ELISA) method using the test kits purchased from
Elabscience Biotechnology Co., Ltd, China. AST activity and LDH
activity were measured using spectrophotometric enzyme assay
2.5. Dose response effect of ABEC for cardioprotective effect in
kit purchased from Biorex Diagnostic, United Kingdom. MPO
doxorubicin induced cardiotoxicity in vivo
activity was estimated using the ELISA kit purchased from DRG
International Inc., (USA).
Healthy Wistar albino rats (male and female) were divided into
seven groups as ten animals in each group (Beery, 2018). Group I
was the control group which was given distilled water orally for 2.8. Assessment of antioxidant parameters and lipid peroxidation in
14 days and on the 11th day a single intraperitoneal (IP) injection the homogenate of heart tissues
(10 mL/kg) of saline was injected after a 16hr fast. Group 2 was
considered as the doxorubicin control group and they were admin- Homogenate of the heart tissues was prepared by using ice-cold
istered distilled water orally for 14 days. On the 11th day, a single PBS buffer (tissue weight to homogenization buffer; 1:10). The
injection of 18 mg/kg of doxorubicin was administered intraperito- supernatant of the homogenate was collected to assess the total
nially after a 16hr fast. Group 3–7 received freeze dried ABEC antioxidant activity, GSH concentration, GR and GPx activities
(0.125, 0.25, 0.5, 1.0 & 2.0 g/kg) via oral administration for 14 days. using commercially available spectrophotometric assay kits pur-
Then, on the 11th day, a single dose of doxorubicin was injected chased from Biorex Diagnostic (UK). Activity of catalase was mea-
intraperitoneally after a 16hr fast. All animals were sacrificed on sured by an assay kit purchased from [Link] (USA).
the 15th day, blood was collected for the estimation of serum con- SOD activity and lipid peroxidation were assessed by the colouri-
centration of cardiac troponin I (cTnI), aspartate aminotransferase metric assay kits purchased from Sigma Aldrich (USA).
(AST, EC [Link]) and lactate dehydrogenase (LDH, EC [Link]) and
heart tissues were collected in to 10% formal saline to be processed 2.9. Histological assessment of myocardial damage
for the histological assessment of myocardial damage.
The left half of the heart tissues fixed in 10% formal saline was
2.6. Experimental procedure for screening of ABEC for cardioprotective processed and sections with 3 mm thickness were stained with rou-
effect against doxorubicin induced cardiotoxicity in vivo tine histological stain, haematoxylin and eosin (H & E). The sec-
tions cut from each group were examined under the light
Healthy male and female Wistar albino rats were randomly microscope and necrotic changes were scored. The scoring system
allocated to five groups of ten animals in each group. Following test mentioned below was developed by the authors by observing the
protocol was followed (Beery, 2018, Sandamali et al., 2020). myocardium of rats (tissue section with 5 mm diameter).
822
J.A.N. Sandamali, R.P. Hewawasam, K.A.P.W. Jayatilaka et al. Saudi Pharmaceutical Journal 29 (2021) 820–832

Cells without necrotic changes: 0; Up to 10 cells with necrotic substantial positive correlation (0.95 to 0.98) was detected
changes: 1; 10–50 cells with necrotic changes: 2; 50–100 cells between the polyphenolic content and antioxidant activities.
with necrotic changes: 3; >100 cells with necrotic changes: 4 Therefore, it can be assumed that there is a significant influence
Cardiomyocytes with early necrotic changes including hyper of phenolic substances to the recognized antioxidant activity of
eosinophilic cytoplasm with no striations and nuclear changes ABEC.
such as pyknosis, karrheorhexis or karyolysis were identified as
necrotic cells. Density of necrotic myocytes was assessed in the 3.3. Dose response effect of ABEC
peripheral and sub- endocardial regions of the myocardium
separately. Doxorubicin control group showed significant increase
(p < 0.001) in serum cTnI concentration (161.9 ± 25.7 pg/mL) com-
2.10. Statistical analysis pared to the control group (Fig. 1). When the dosage of ABEC was
increased gradually, a gradual decrease in serum cTnI concentra-
Results are expressed as mean ± SD. The significance of inter- tion was observed in rat groups treated with 0.25, 0.5, 1.0 and
group differences was evaluated by one-way analysis of variance 2.0 g/kg dosages of plant extract compared to the doxorubicin con-
using SPSS 22.0 software. Differences between groups were consid- trol group (P < 0.05). When consider the measurement of serum
ered statistically significant at P < 0.05. AST and LDH activity, rat groups treated with doxorubicin alone
showed a significant increase (p < 0.001) in enzyme activities com-
3. Results pared to the control (Figs. 2 & 3). However, all groups of rats trea-
ted with ABEC showed a significant decrease (p < 0.05) in AST and
3.1. Physicochemical and phytochemical analysis LDH activities compared to the doxorubicin control group.
A normal morphology was observed in the cross section of the
The physicochemical properties of Cinnamomum zeylanicum myocardial tissues of normal control group (Figs. 4 & 5). However,
bark are shown in Table 1 (Supplementary data). When consider group of rats that were treated with doxorubicin alone showed
the extractable mater in water and methanol, hot extraction extensive early changes of necrosis in both peripheral and suben-
resulted in a higher yield of the plant. None of the heavy metals docardium of the myocardial tissue giving the highest score of
including lead (Pb), cadmium (Cd), arsenic (As) and mercury (Hg) 7.9 (Supplementary data Table 4, Figs. 4 & 5). All rat groups treated
were detected in the plant extract. Microscopic observations are with the ABEC (Group 3–7) showed a gradual reduction of the
also shown in Table 1 (Supplementary data). score for the early changes of necrosis with increasing dosage of
In phytochemical analysis, Cinnamomum bark was positive for the plant extracts (Supplementary data Table 4). Reversible histo-
saponins, polyphenols, alkaloids, tannins, proteins and reducing logical changes such as inflammatory infiltrations, haemorrhages,
sugars as shown in Table 2 (Supplementary data). The Cinnamo- interstitial oedema, congestion of blood vessels, intracellular vac-
mum plant extract was negative for toxic phytochemicals including uoles and wavy myocardial fibers were observed in doxorubicin
anthracene, cyanogenic and cardenoloid glycosides. control group (Supplementary data Table 5, Fig. 6). Groups of rats
treated with the ABEC showed different degree of reversible histo-
3.2. Total polyphenol content and in vitro antioxidant activity of ABEC logical changes while some changes were completely absent in the
groups treated with higher doses of the ABEC.
Total polyphenol content and the in vitro antioxidant activity of
ABEC are shown in Table 3 (Supplementary data). The correlation 3.4. Screening of ABEC for cardioprotective effect
between the polyphenol content and the antioxidant activities of
ABEC was determined to evaluate the appropriateness and consis- 3.4.1. Effect on serum cardiac biomarkers
tency of the in vitro antioxidant assay methods. The linear regres- The concentration of cTnI as a specific marker of myocardial
sion analysis results are shown in Fig. 1 (Supplementary data). A damage was significantly increased (p < 0.001) in doxorubicin

Fig. 1. Serum cTnI concentration of rats treated with different concentrations of ABEC cTnI; cardiac troponinI, Dox; Doxorubicin, ABEC; Aqueous bark extract of Cinnamomum
zeylanicum.

823
J.A.N. Sandamali, R.P. Hewawasam, K.A.P.W. Jayatilaka et al. Saudi Pharmaceutical Journal 29 (2021) 820–832

Fig. 2. Serum AST activity of rats treated with different concentrations of ABEC AST; Aspartate amino transferase, Dox; Doxorubicin, ABEC; Aqueous bark extract of
Cinnamomum zeylanicum.

Fig. 3. Serum LDH activity of rats treated with different concentrations of ABEC LDH; Lactate dehydrogenase, Dox; Doxorubicin, ABEC; Aqueous bark extract of Cinnamomum
zeylanicum.

control group exhibiting the mean value of 145.15 pg/mL com- The doxorubicin treated rats showed a significant increase
pared to the normal control group (0.00 ng/mL). The treatment (p < 0.001) in AST and LDH activity in the serum compared to
with ABEC alone (plant extract control) didn’t show any significant the normal control group. Pre-treatment with ABEC significantly
change in cTnI concentration compared to the normal control. Pre- decreased (p < 0.001) the elevated level of AST and LDH activity
treatment with ABEC showed a significant reduction (p < 0.001) in in rats compared to the doxorubicin control group. Plant extract
cTnI concentration with the mean value of 21.85 pg/mL compared control group didn’t show any significant changes in AST and
to the doxorubicin control group (Table 1). LDH activity compared to the normal control group (Table 1).
The serum concentration of NT-pro BNP was significantly
higher (371.14 ± 9.69 pg/mL) in animals treated with doxorubicin 3.4.2. Effect on antioxidant parameters
compared to the normal control group (41.57 ± 7.29 pg/mL) and The mean values of GSH level, GPx and GR enzyme activities
there was no significant change in NT-pro BNP concentration in were 2.3 nmol/mL, 277.60 U/L and 30.84 U/L respectively in heart
plant extract control (35.86 ± 3.10 pg/mL). Serum NT-pro BNP con- tissues of rats treated with doxorubicin and a significant decrease
centration was significantly decreased (p < 0.001) in animals trea- (p < 0.001) was observed in these antioxidant parameters com-
ted with ABEC before the administration of doxorubicin showing pared to the normal control group (4.7 ± 0.49 nmol/mL, 378.25 ± 3
the mean value, 198.57 pg/mL compared to the doxorubicin con- .81 U/L and 76.04 ± 3.09 U/L respectively) (Table 2). Pre-treatment
trol group (Table 1). with ABEC showed significant increase (p < 0.001) in GSH level,
824
J.A.N. Sandamali, R.P. Hewawasam, K.A.P.W. Jayatilaka et al. Saudi Pharmaceutical Journal 29 (2021) 820–832

Fig. 4. Cells with early necrotic changes in subendocardial region of rat heart treated with different doses of ABEC (Light micrograph, H & E, 400). a – Control, b -
Doxorubicin control, c - Dox + 0.125 g/kg of ABEC, d – Dox + 0.25 g/kg of ABEC, e – Dox + 0.5 g/kg of ABEC, f – Dox + 1.0 g/kg of ABEC, g – Dox + 2.0 g/kg of ABEC. ABEC; Aqueous
bark extract of Cinnamomum zeylanicum, Dox; doxorubicin.

GPx and GR enzyme activities with the respective mean values, doxorubicin showed significant reduction (p < 0.001) in MPO
4.01 nmol/mL, 342.79 U/L and 57.90 U/L compared to the doxoru- enzyme activity (210.46 ± 4.60 AAU/mL) compared to the doxoru-
bicin control group. bicin control group.
Doxorubicin treatment in Wistar rats caused significant Positive control group was administered with dexrazoxane
decrease (p < 0.001) in SOD activity and catalase activity in the before the injection of doxorubicin. Dexrazoxane treatment in rats
myocardium as compared to the normal control group (Table 2). showed a significant change (p < 0.001) in biochemical parameters
Pre-treatment with freeze dried ABEC significantly increased compared to the doxorubicin control exhibiting the considerable
(p < 0.001) the SOD and catalase activity compared to the doxoru- cardio-protection. Although all biochemical parameters except
bicin treated animals. No significant changes in SOD and catalase catalase showed a significant difference between the positive con-
activities were noticed in plant extract control compared to the trol and the ABEC pre-treated group, pre-treatment with ABEC
normal control group. showed a significant difference (p < 0.001) in all parameters
against the doxorubicin control group and showed a considerable
protection against doxorubicin induced cardiotoxicity in Wistar
3.4.3. Effect on lipid peroxidation in the cardiac tissues
rats.
There was a significant increase (p < 0.001) in the myocardial
lipid peroxidation evident with the increased malondialdehyde
levels (MDA) in the doxorubicin treated rat group (2.05 ± 0.023 n
3.4.5. Effect on histology of the myocardial tissues
mol/mL) compared to the normal control (1.19 ± 0.009 nmol/mL)
Control group of rats showed normal morphology in both
(Table 2). Pre-treatment with ABEC prevented the elevation of this
subendocardial as well as the peripheral region of the myocardium
oxidative stress marker giving the mean value of 1.51 nmol/mL.
as shown in Figs. 7 and 8. Necrotic changes were extensively seen
Plant extract control, however, exhibited no significant effect on
in doxorubicin control group and they showed the highest score
the MDA level (1.21 ± 0.012 nmol/mL) when compared to the nor-
among the study groups (7.8) (Table 3). However, necrotic changes
mal control group.
were more visible in the subendocardial region when compared to
the peripheral region (Figs. 7 and 8). Doxorubicin treated rats also
3.4.4. Effect on MPO enzyme activity showed many other histological changes of cell injury showed in
Any changes in the activity of MPO enzyme was not observed in Fig. 6 including intracellular vacuoles, wavy myocardial fibers,
the rats treated with ABEC alone. Doxorubicin control group exhib- inflammatory infiltration, haemorrhages, interstitial oedema and
ited significant (p < 0.001) elevation in the MPO activity (285.32 ± congestion of blood vessel (Supplementary data Table 6). Pre-
1.64 AAU/mL) compared to the control group (157.74 ± 1.76 AAU/ treatment with ABEC was capable to lessen the degree of damage
mL) (Table 2). Pre-treatment with ABEC in animals injected with in the myocardium showing significant reduction (p < 0.001) in
825
J.A.N. Sandamali, R.P. Hewawasam, K.A.P.W. Jayatilaka et al. Saudi Pharmaceutical Journal 29 (2021) 820–832

Fig. 5. Cells with early necrotic changes in peripheral region of rat heart treated with different doses of ABEC (Light micrograph, H & E, 400). a – Control, b - Doxorubicin
control, c - Dox + 0.125 g/kg of ABEC, d – Dox + 0.25 g/kg of ABEC, e – Dox + 0.5 g/kg of ABEC, f – Dox + 1.0 g/kg of ABEC, g – Dox + 2.0 g/kg of ABEC. ABEC; Aqueous bark extract
of Cinnamomum zeylanicum, Dox; doxorubicin.

the necrotic changes which was evident with decreased score (3.8) 2016; Ojha et al., 2016; Afsar et al., 2017; Ibrahim et al., 2017;
(Table 3). This group also showed much necrotic changes in suben- Sergazy et al., 2020).
docardial region than the peripheral region (Figs. 7 and 8). Only the Previous studies have shown that proinflammatory cytokine
intracellular vacuoles and congestion of blood vessels were expression, inflammatory cell infiltration and necrosis are com-
observed as reversible histological changes (Supplementary data monly found in doxorubicin induced oxidative stress which ulti-
Table 6). Treatment with ABEC alone did not reveal histological mately lead to increased release of cardiac markers and
changes in the myocardium and showed normal morphology as natriuretic peptides to blood (Ikegami et al., 2007; Riad et al.,
in the normal control group. 2009). A study done by Baniahmad et al. (2020) showed that dox-
orubicin treatment increases cardiac biomarkers including cTnI,
AST and LDH levels in rat serum while treatment with vanillic acid,
4. Discussion a pharmaceutical compound which belongs to phenolic acid family
significantly reduce the release of cardiac biomarkers indicating its
Among the strategies to attenuate doxorubicin induced car- cardioprotective activity based on antioxidant effect. Similar to
diotoxicity, optimization of dosage, nanoencapsulation, usage of these results ABEC which has high polyphenolic content and
different analogues that lessen the oxidative stress have been iden- in vitro antioxidant effect also showed cardio protection against
tified as effective approaches. Although dexrazoxane is the only doxorubicin treatment by significantly reducing the release of car-
Food and Drug Administration (FDA) approved drug to treat diac biomarkers. Several other studies also reported results consis-
anthracycline induced cardiotoxicity, it has several limitations tent with the present study (Afsar et al., 2017; Oyagbemi et al.,
(Bansal et al., 2019). Therefore, the present study sheds light on 2018; Li et al., 2020). In the present study, high concentrations of
the ameliorative effect of Cinnamomum zeylanicum against doxoru- NT-proBNP was observed in doxorubicin treated rats indicating
bicin induced cardiotoxicity and its potential to be developed fur- an increased ventricular dysfunction while the pre-treatment with
ther as a therapeutic agent. ABEC lead to a significant reduction in NT-proBNP concentration.
According to the results obtained for the qualitative and quan- Previous studies conducted on experimental rats also showed sim-
titative analysis of antioxidants in the present study, important ilar results to the present study (Koh et al., 2004; Argun et al.,
phytochemicals such as polyphenols, alkaloids and tannins were 2016).
present in ABEC in significant quantities while toxic phytochemi- Doxorubicin induced cardiotoxicity was biochemically con-
cals such as cyanogenic glycosides and cardenoloid glycosides firmed by the increase in oxidative stress as shown by the reduced
were absent. These results corroborated with many previous find- antioxidant markers such as GSH, GPx, GR, SOD and catalase and
ings where the presence of antioxidants such as polyphenols con- total antioxidant capacity (Baniahmad et al., 2020). Several previ-
tributed significantly to the protective effect against doxorubicin ous investigations showed that doxorubicin treatment increases
induced cardiotoxicity (Kaiserová et al., 2007; Hamza et al., oxidative stress in rats and plant extracts or some other com-
826
J.A.N. Sandamali, R.P. Hewawasam, K.A.P.W. Jayatilaka et al. Saudi Pharmaceutical Journal 29 (2021) 820–832

Fig. 6. Photomicrographs of reversible cellular changes in myocardium of rats treated with doxorubicin (H & E, 400) a- Inflammatory infiltrations, b- Interstitial oedema, c-
Haemorrhages, d- Congestion of blood vessel, e- Wavy myocardial fibers, f- Intracellular vacuoles. Arrows indicate reversible cellular changes.

Table 1
Effect of ABEC on serum cardiac biomarkers.

Cardiac biomarkers Group I Group II Group III Group IV Group V

(Control) (Plant control) (Dox Control) (ABEC + Dox) (Positive control)
cTnI concentration (pg/mL) 0.00 0.00 145.15 ± 10.77 21.85 ± 3.84*** 11.46 ± 2.59***
NT-pro BNP concentration (pg/mL) 41.57 ± 7.29 35.86 ± 3.10 371.14 ± 9.69 198.57 ± 7.07*** 159.43 ± 12.39***
AST activity (U/L) 25.71 ± 1.41 24.18 ± 1.60 66.10 ± 2.07 28.79 ± 1.98*** 26.90 ± 1.26***
LDH activity (U/L) 1057.21 ± 38.6 1076.64 ± 49.8 1584.19 ± 83.4 1190.77 ± 77.2*** 1104.97 ± 58.7***

ABEC; Aqueous bark extract of Cinnamomum zeylanicum, Dox; Doxorubicin, cTnI; cardiac Troponin I, NT-pro BNP; N-terminal pro brain natriuretic peptide, AST; Aspartate
amino transferase, LDH; Lactate dehydrogenase. All values are expressed as mean ± SD (n = 10). p values: * < 0.05,**< 0.01, *** <0.001 compared to the doxorubicin control
group (Group III).

pounds with significant antioxidant activity have the ability to Harthi et al., 2014; Hamza et al., 2016; Afsar et al., 2017; Alam
attenuate free radical induced oxidative stress indicating an et al., 2018; Shaker et al., 2018; Li et al., 2020). In the present study,
increased activity of antioxidant markers (Singh et al., 2008; Al- pre-treatment with ABEC also exhibited a significant increase in
827
J.A.N. Sandamali, R.P. Hewawasam, K.A.P.W. Jayatilaka et al. Saudi Pharmaceutical Journal 29 (2021) 820–832

Table 2
Effect of ABEC on antioxidant parameters, lipid peroxidation and MPO activity.

Biochemical parameters Group I Group II Group III Group IV Group V

(Normal control) (Plant control) (Dox Control) (ABEC + Dox) (Positive control)
GSH (nmol/mL) 4.70 ± 0.49 4.83 ± 0.45 2.30 ± 0.37 4.01 ± 0.35*** 4.42 ± 0.36***
GPx (U/L) 378.25 ± 3.81 377.78 ± 5.48 277.60 ± 6.03 342.79 ± 6.95*** 362.94 ± 5.24***
GR (U/L) 76.04 ± 3.09 75.84 ± 2.33 30.84 ± 4.15 57.90 ± 3.48*** 67.57 ± 3.88***
SOD activity (%) 87.60 ± 2.09 86.90 ± 2.11 49.42 ± 2.28 70.12 ± 1.59*** 81.03 ± 3.44***
Catalase activity (mmol/L) 193.31 ± 3.46 190.23 ± 4.43 150.74 ± 6.68 173.46 ± 6.6*** 178.61 ± 4.34***
Total antioxidant capacity (mmol/L) 5.52 ± 0.33 5.65 ± 0.25 2.92 ± 0.32 4.03 ± 0.30*** 4.85 ± 0.35***
Lipid peroxidation (nmol/mL) 1.19 ± 0.009 1.21 ± 0.012 2.05 ± 0.023 1.51 ± 0.020*** 1.28 ± 0.015***
MPO activity (AAU/mL) 157.74 ± 1.76 155.29 ± 2.27 285.32 ± 1.64 210.46 ± 4.6*** 197.66 ± 1.97***

ABEC; Aqueous bark extract of Cinnamomum zeylanicum, Dox; Doxorubicin, GSH; Reduced glutathione, GPx; Glutathione peroxidase, GR; Glutathione reductase, SOD;
Superoxide dismutase, MPO; Myeloperoxidase. All values are expressed as mean ± SD (n = 10). P values: * < 0.05, **< 0.01 *** <0.001 compared to the doxorubicin control
group (Group III).

Fig. 7. Effect of optimum dose of ABEC: Photomicrographs of subendocardial region of rat heart (H & E, 400). a- Group I (normal control); no any histological changes, b-
Group II (Plant control); shows normal architecture, c- Group III (Doxorubicin control); shows a higher degree of necrotic changes (hyper-eosinophilic cytoplasm and nuclear
changes in cell death; pyknosis/ karrheorhexis/ karyolysis), d– Group IV (Dox + 2.0 g/kg of ABEC); shows lesser degree of necrosis, e– Group V (Positive control); shows
occasional cells with early changes of necrosis. Arrows indicate areas of sub-endocardium with necrotic changes. ABEC; Aqueous bark extract of Cinnamomum zeylanicum,
Dox; doxorubicin.

828
J.A.N. Sandamali, R.P. Hewawasam, K.A.P.W. Jayatilaka et al. Saudi Pharmaceutical Journal 29 (2021) 820–832

Fig. 8. Effect of optimum dose of ABEC: Photomicrographs of peripheral region of rat heart (H & E, 400). a- Group I (normal control); no any histological changes, b- Group II
(Plant control); shows normal architecture, c- Group III (Doxorubicin control); shows a higher degree of necrotic changes (hyper-eosinophilic cytoplasm and nuclear changes
in cell death; pyknosis/ karrheorhexis/ karyolysis), d– Group IV (Dox + 2.0 g/kg of ABEC); shows lesser degree of necrosis, e– Group V (Positive control); shows occasional cells
with early changes of necrosis. Arrows indicate areas of peripheral region of myocardium with necrotic changes. ABEC; Aqueous bark extract of Cinnamomum zeylanicum,
Dox; doxorubicin.

Table 3
Average grading of cells with necrotic changes in myocardial tissues.

Group I Group II Group III Group IV Group V

(Normal control) (Plant control) (Dox Control) (ABEC + Dox) (Positive control)
Sub-endocardial region of heart tissues (score out of 4) 0 0 4 2.6 1.9
Peripheral region of heart tissues (score out of 4) 0 0 3.8 1.2 0.8
Total score (out of 8) 0 0 7.8 3.8 2.7

ABEC; Aqueous bark extract of Cinnamomum zeylanicum, Dox; Doxorubicin. Grading scale; no cells with necrotic changes: 0; up to 10 cells with necrotic changes: 1; 10–50
cells with necrotic changes: 2; 50–100 cells with necrotic changes: 3; >100 cells with necrotic changes: 4.

GSH and other antioxidant enzymes and total antioxidant capacity GSH is an important antioxidant compound in cellular defence
suggesting that pre-treatment with the plant extract may replenish system and depletion of GSH may contribute to enhanced lipid
the cardiomyocytes with antioxidant enzymes which exert cardio- peroxidation in the cell membrane (Afsar et al., 2017). Doxoru-
protective effect against doxorubicin induced cardiotoxicity. bicin treatment that increases the free radical formation in
829
J.A.N. Sandamali, R.P. Hewawasam, K.A.P.W. Jayatilaka et al. Saudi Pharmaceutical Journal 29 (2021) 820–832

cardiomyocytes consequently enhance the lipid peroxidation results, ABEC exerted its protective effect against doxorubicin
indicated by an increase in MDA concentration, which is a stable induced cardiotoxicity via antioxidant and anti-inflammatory
end product of lipid peroxidation (Xiao, 2015). Previous investi- activities.
gations demonstrated that some herbal plants or related com-
pounds with high antioxidant activity are capable of reducing Author contributions
MDA concentration in myocardial tissues of rats treated with
doxorubicin (Hamza et al., 2008; Singh et al., 2008; Hamza JANS performed all experiments, analysed data and wrote the
et al., 2016; Kwatra et al., 2016; Afsar et al., 2017). In the present original draft of the manuscript. RPH conceptualized the study,
study also ABEC with high antioxidant effect was capable of sig- critically reviewed and edited the manuscript, supervised &
nificantly reducing the MDA concentration in homogenates of administered the project and secured funding. KAPWJ reviewed
heart tissues suggesting that reduction of lipid peroxidation and revised the manuscript and supervised the project. LKB super-
may be due to the radical scavenging ability of Cinnamomum vised experiments on histopathological evaluation of cardiac dam-
bark. age and data interpretation. All authors approved the final
Free radical induced oxidative stress in doxorubicin treatment manuscript submitted and agreed to be accountable for all aspects
may up regulate the inflammation through activation of NF-jB of the manuscript.
which stimulate the pro-inflammatory cytokine production
(Hamza et al., 2016). Therefore, several investigations have shown Declaration of Competing Interest
that doxorubicin treatment increases MPO activity in rat serum
which is considered as an inflammatory marker (Hamza et al., The authors declare that they have no known competing finan-
2008; Hamza et al., 2016; Ibrahim et al., 2017; Oyagbemi et al., cial interests or personal relationships that could have appeared
2017; Bin Jardan et al., 2020). All these studies have shown that to influence the work reported in this paper.
pre-treatment with compounds which have antioxidant effect are
effective to reduce the MPO activity. Pre-treatment with ABEC also Acknowledgements
showed a significant reduction in MPO activity exhibiting its cardio
protective activity through antioxidant effect. Further, Cinnamo- Mr. G.H.J.M. Priyashantha and Mr. E.G. Rukman Asiri of the
mum zeylanicum contains high amount of cinnamaldehyde which Department of Biochemistry, Faculty of Medicine, University of
has anti-inflammatory effect which may contribute to down regu- Ruhuna are acknowledged for the assistance provided in conduct-
late the inflammatory pathway induced by the doxorubicin treat- ing animal experiments.
ment (Han and Parker, 2017).
Histopathological assessment of myocardial damage is consid- Funding
ered as the gold standard to diagnose acute doxorubicin induced
cardiotoxicity (Octavia et al., 2012). In the present study, biochem- This work was supported by the National Research Council, Sri
ical changes in doxorubicin induced cardiotoxicity were confirmed Lanka (Grant No: 18-050) and University Grants Commission, Sri
by histological changes in myocardial tissues including early Lanka (UGC/VC/DRIC/PG2015 (III)/RUH/01).
changes of necrosis, inflammatory infiltrations, haemorrhages,
interstitial oedema and wavy myocardial fibers. A previous study
Appendix A. Supplementary data
conducted by Zhang et al. (2017) also reported that doxorubicin
produces massive changes in rat myocardium, consisting of necro-
Supplementary data to this article can be found online at
sis, intracellular oedema, swollen and damaged mitochondria, and
[Link]
wavy degeneration of cardiac muscle fibers. In addition, El-Agamy
et al. (2016) also demonstrated focal necrosis, signs of necrosis
with inflammatory infiltrations and loss of muscle striations in
rat heart treated with single dose of 20 mg/kg doxorubicin. Koti References
et al. (2013) and Erboga et al. (2016) also showed similar histolog-
ical changes in rats after the doxorubicin treatment. However, the Afsar, T., Razak, S., Batoo, K.M., Khan, M.R., 2017. Acacia hydaspica R. Parker prevents
doxorubicin-induced cardiac injury by attenuation of oxidative stress and
pre-treatment with ABEC in doxorubicin treated rats showed bet-
structural cardiomyocyte alterations in rats. BMC Complement. Altern. Med. 17.
ter preservation of myocardium by reducing the reversible histo- [Link]
logical changes and mean score of early changes of necrosis Alam, M.F., Khan, G., Safhi, M.M., Alshahrani, S., Siddiqui, R., Moni, S.S., Anwer, T.,
indicating that Cinnamomum zeylanicum has a potential cardiopro- 2018. Thymoquinone ameliorates doxorubicin-induced cardiotoxicity in Swiss
Albino mice by modulating oxidative damage and cellular inflammation.
tective activity. Cardiol. Res. Pract. 2018. [Link]
Collectively, biochemical and histopathological findings con- Al-Harthi, S.E., Alarabi, O.M., Ramadan, W.S., Alaama, M.N., Al-Kreathy, H.M.,
firmed a potential cardioprotective effect of ABEC against doxoru- Damanhouri, Z.A., Khan, L.M., Osman, A.M., 2014. Amelioration of doxorubicin-
induced cardiotoxicity by resveratrol. Mol. Med. Rep. 10, 1455–1460. https://
bicin induced cardiotoxicity. Therefore, the antioxidant and anti- [Link]/10.3892/mmr.2014.2384.
inflammatory effect of Cinnamomum zeylanicum may be a strong Argun, M., Üzüm, K., Sönmez, M.F., Özyurt, A., Derya, K., Çilenk, K.T., Unalmısß, S.,
contributing factor which protects the cells from degenerative Pamukcu, Ö., Baykan, A., Narin, F., Elmalı, F., Narin, N., 2016. Cardioprotective
effect of metformin against doxorubicin cardiotoxicity in rats. Anatol. J. Cardiol.
changes. Thus, in this study, ABEC effectively prevented the tissue 16, 234–241. [Link]
damage by decreasing the oxidative-stress and restoring the Aryal, B., Rao, V.A., 2016. Deficiency in cardiolipin reduces doxorubicin-induced
antioxidant status. oxidative stress and mitochondrial damage in human B lymphocytes. PLoS ONE.
[Link]
Baniahmad, B., Safaeian, L., Vaseghi, G., Rabbani, M., Mohammadi, B., 2020.
Cardioprotective effect of vanillic acid against doxorubicin-induced
5. Conclusion cardiotoxicity in rat. Res. Pharm. Sci. 15, 87–96. [Link]
5362.278718.
In conclusion, the present study revealed an ameliorative Bansal, N., Adams, M.J., Ganatra, S., Colan, S.D., Aggarwal, S., Steiner, R., Amdani, S.,
Lipshultz, E.R., Lipshultz, S.E., 2019. Strategies to prevent anthracycline-induced
effect of Cinnamomum zeylanicum to counteract with doxorubicin cardiotoxicity in cancer survivors. Cardio-oncol. 5. [Link]
induced cardiac injury for the first time. According to our s40959-019-0054-5.

830
J.A.N. Sandamali, R.P. Hewawasam, K.A.P.W. Jayatilaka et al. Saudi Pharmaceutical Journal 29 (2021) 820–832

Beery, A.K., 2018. Inclusion of females does not increase variability in rodent McGowan, J.V., Chung, R., Maulik, A., Piotrowska, I., Walker, J.M., Yellon, D.M., 2017.
research studies. Curr. Opin. Behav. Sci. 23, 143–149. [Link] Anthracycline chemotherapy and cardiotoxicity. Cardiovasc. Drugs Ther. 31,
[Link].2018.06.016. 63–75. [Link]
Bin Jardan, Y.A., Ansari, M.A., Raish, M., Alkharfy, K.M., Ahad, A., Al-Jenoobi, F.I., Haq, Minotti, G., Menna, P., Salvatorelli, E., Cairo, G., Gianni, L., 2004. Anthracyclines:
N., Khan, M.R., Ahmad, A., 2020. Sinapic acid ameliorates oxidative Stress, molecular advances and pharmacologic developments in antitumor activity and
Inflammation, and apoptosis in acute doxorubicin-induced cardiotoxicity via cardiotoxicity. Pharmacol. Rev. 56, 185–229. [Link]
the NF-jB-mediated pathway. Biomed Res. Int. 2020. [Link] Mitry, M.A., Edwards, J.G., 2016. Doxorubicin induced heart failure: Phenotype and
2020/3921796. molecular mechanisms. IJC Heart Vasc. 10, 17–24. [Link]
Boora, F., Chirisa, E., Mukanganyama, S., 2014. Evaluation of nitrite radical ijcha.2015.11.004.
scavenging properties of selected zimbabwean plant extracts and their Mobaraki, M., Faraji, A., Zare1, M., Dolati, P., Ataei, M., Manshadi, H.R.D., 2017.
phytoconstituents. J. Food Process. 2014. [Link] Molecular mechanisms of cardiotoxicity: A review on major side-effect of
Chatterjee, K., Zhang, J., Honbo, N., Karliner, J.S., 2010. Doxorubicin cardiomyopathy. doxorubicin. Indian J. Pharm. Sci. 79, 335-344. [Link]
Cardiology. 115, 155–162. [Link] pharmaceutical-sciences.1000235.
Chericoni, S., Prieto, J.M., Iacopini, P., Cioni, P., Morelli, I., 2005. In vitro activity of the Mukherjee, S., Banerjee, S.K., Maulik, M., Dinda, A.K., Talwar, K.K., Maulik, S.K., 2003.
essential oil of Cinnamomum zeylanicum and eugenol in peroxynitrite-induced Protection against acute adriamycin-induced cardiotoxicity by garlic: role of
oxidative processes. J. Agric. Food Chem. 53, 4762–4765. [Link] endogenous antioxidants and inhibition of TNF-alpha expression. BMC
10.1021/jf050183e. Pharmacol. 3. [Link]
El-Agamy, D.S., Abo-Haded, H.M., Elkablawy, M.A., 2016. Cardioprotective effects of Mushtaq, A., Akbar, S., Zargar, M.A., Wali, A.F., Malik, A.H., Dar, M.Y., Hamid, R.,
sitagliptin against doxorubicin-induced cardiotoxicity in rats. Exp. Biol. Med. Ganai, B.A., 2014. Phytochemical screening, physicochemical properties, acute
(Maywood, N.J.). 241, 1577–1587. [Link] toxicity testing and screening of hypoglycaemic activity of extracts of Eremurus
Erboga, M., Donmez, Y.B., Sener, U., Erboga, Z.F., Aktas, C., Kanter, M., 2016. Effect of himalaicus baker in normoglycaemic Wistar strain albino rats. Biomed Res. Int.
Urtica Dioica against doxorubicin-induced cardiotoxicity in rats through 2014. [Link]
suppression of histological damage, oxidative stress and lipid peroxidation. Octavia, Y., Tocchetti, C.G., Gabrielson, K.L., Janssens, S., Crijns, H.J., Moens, A.L.,
Eur. J. Gen. Med. 13, 139-144. [Link] 2012. Doxorubicin-induced cardiomyopathy: From molecular mechanisms to
Galketiya, C., Weerarathna, T.S., Punchihewa, J.C., Wickramaratne, M.N., therapeutic strategies. J. Mol. Cell. Cardiol. 52, 1213–1225. [Link]
Wickramaratne, D.B.M., 2017. Screening of edible plants in Sri Lanka for 10.1016/[Link].2012.03.006.
antioxidant activity. J. Med. Plants Stud. 5, 91–95. Ojha, S., Taee, H.A., Goyal, S., Mahajan, U.B., Patil, C.R., Arya, D.S., Rajesh, M., 2016.
Ghosh, T., Basu, A., Adhikari, D., Roy, D., Pal, A.K., 2015. Antioxidant activity and Cardioprotective potentials of plant-derived small molecules against
structural features of Cinnamomum zeylanicum. 3. Biotech. 5, 939–947. https:// doxorubicin associated cardiotoxicity. Oxid. Med. Cell. Longev. 2016. https://
[Link]/10.1007/s13205-015-0296-3. [Link]/10.1155/2016/5724973.
Goormaghtigh, E., Huart, P., Praet, M., Brasseur, R., Ruysschaert, J.M., 1990. Structure Oyagbemi, A.A., Omobowale, T.O., Olopade, J.O., Farombi, E.O., 2018. Kolaviron and
of the adriamycin-cardiolipin complex. Role in mitochondrial toxicity. Biophys. Garcinia kola attenuate doxorubicin-induced cardiotoxicity in Wistar rats. J.
Chem. 35, 247–257. [Link] Complement. Integr. Med. 15. [Link]
Halestrap, A.P., 2006. Calcium, mitochondria and reperfusion injury: a pore way to Premakumara G.A.S., Abeysekera W.P.K.M., 2020. Pharmacological properties of
die. Biochem. Soc. Trans. 34, 232–237. [Link] Ceylon cinnamon. In: Senaratne R., Pathirana R. (eds) Cinnamon. Springer,
Hamza, A.A., Ahmed, M.M., Elwey, H.M., Amin, A., 2016. Melissa officinalis protects Cham. [Link]
against doxorubicin-induced cardiotoxicity in rats and potentiates its Rahman, M.M., Islam, M.B., Biswas, M., Alam, A.H.M.K., 2015. In vitro antioxidant
anticancer activity on MCF-7 cells. PLoS ONE 11. [Link] and free radical scavenging activity of different parts of Tabebuia pallida
[Link].0167049. growing in Bangladesh. BMC Res. Notes. 8. [Link]
Hamza, A.A., Amin, A., Daoud, S., 2008. The protective effect of a purified extract of 1618-6.
Withania somnifera against doxorubicin-induced cardiac toxicity in rats. Cell Ranasinghe, P., Galappaththy, P., 2016. Health benefits of Ceylon cinnamon
Biol. Toxicol. 24, 63–73. [Link] (Cinnamomum zeylanicum): a summary of the current evidence. Ceylon Med.
Han, X., Parker, T.L., 2017. Antiinflammatory activity of Cinnamon (Cinnamomum J. 61, 1–5. [Link]
zeylanicum) bark essential oil in a human skin disease model. Phytother. Res. Ranasinghe, P., Pigera, S., Premakumara, G.A., Galappaththy, P., Constantine, G.R.,
31, 1034–1038. [Link] Katulanda, P., 2013. Medicinal properties of ’true’ cinnamon (Cinnamomum
Ibrahim, D.M., Radwan, R.R., Fattah, S.M.A., 2017. Antioxidant and antiapoptotic zeylanicum): a systematic review. BMC Complement. Altern. Med. 22. https://
effects of sea cucumber and valsartan against doxorubicin-induced [Link]/10.1186/1472-6882-13-275.
cardiotoxicity in rats: The role of low dose gamma irradiation. J. Photochem. Rao, P.V., Gan, S.H., 2014. Cinnamon: A multifaceted medicinal plant. Evid. Based
Photobiol., B 170, 70–78. [Link] Complement. Alternat. Med. 2014. [Link]
Ikegami, E., Fukazawa, R., Kanbe, M., Watanabe, M., Abe, M., Watanabe, M., Riad, A., Bien, S., Westermann, D., Becher, P.M., Loya, K., Landmesser, U., Kroemer, H.
Kamisago, M., Hajikano, M., Katsube, Y., Ogawa, S., 2007. Edaravone, a potent K., Schultheiss, H.P., Tschöpe, C., 2009. Pretreatment with statin attenuates the
free radical scavenger, prevents anthracycline-induced myocardial cell death. cardiotoxicity of doxorubicin in mice. Cancer Res. 69, 695–699. [Link]
Circ. J. 71, 1815–1820. [Link] 10.1158/[Link]-08-3076.
Jayaweera, D.M.A., 1982. Medicinal plants (Indigenous and Exotic) used in Ceylon. Sandamali, J.A.N., Hewawasam, R.P., Jayatilaka, K.A.P.W., Mudduwa, L.K.B., 2020.
National Science Foundation in Sri Lanka, Sri Lanka. Cardioprotective potential of Murraya koenigii (L.) Spreng. leaf extract against
Kaiserová, H., Šimůnek, T., van der Vijgh, W.J.F., Bast, A., Kvasničková, E., 2007. doxorubicin-induced cardiotoxicity in rats. Evid. Based Complement. Alternat.
Flavonoids as protectors against doxorubicin cardiotoxicity: Role of iron Med. 2020. [Link]
chelation, antioxidant activity and inhibition of carbonyl reductase. Biochim. Sergazy, S., Shulgau, Z., Fedotovskikh, G., Chulenbayeva, L., Nurgozhina, A.,
Biophys. Acta, Mol. Basis Dis. 1772, 1065–1074. [Link] Nurgaziyev, M., Krivyh, E., Kamyshanskiy, Y., Kushugulova, A., Gulyayev, A.,
bbadis.2007.05.002. Aljofan, M., 2020. Cardioprotective effect of grape polyphenol extract against
Khattry, N., Malhotra, P., Grover, A., Sharma, S.C., Varma, S., 2009. Doxorubicin- doxorubicin induced cardiotoxicity. Sci. Rep. 10. [Link]
induced cardiotoxicity in adult Indian patients on chemotherapy. Indian J. Med. s41598-020-71827-9.
Paediatr. Oncol. 30, 9–13. [Link] Shaker, R.A., Abboud, S.H., Assad, H.C., Hadi, N., 2018. Enoxaparin attenuates
Koh, E., Nakamura, T., Takahashi, H., 2004. Troponin-T and brain natriuretic peptide doxorubicin induced cardiotoxicity in rats via interfering with oxidative stress,
as predictors for adriamycin-induced cardiomyopathy in rats. Circ. J. 68, 163– inflammation and apoptosis. BMC Pharmacol. Toxicol. 19. [Link]
167. [Link] 10.1186/s40360-017-0184-z.
Koti, B.C., Nagathan, S., Vishwanathswamy, A., Gadad, P.C., Thippeswamy, A., 2013. Singh, G., Singh, A.T., Abrahama, A., Bhat, B., Mukherjee, A., Verma, R., Agarwal, S.K.,
Cardioprotective effect of Vedic Guard against doxorubicin-induced Jha, S., Mukherjee, R., Burman, A.C., 2008. Protective effects of Terminalia arjuna
cardiotoxicity in rats: A biochemical, electrocardiographic, and against doxorubicin-induced cardiotoxicity. J. Ethnopharmacol. 117, 123–129.
histopathological study. Pharmacogn. Mag. 9, 176–181. [Link] [Link]
10.4103/0973-1296.111287. Singleton, V.L., Ortofer, R., Lamuda-raventos, R.M., 1999. Analysis of total phenols
Kwatra, M., Kumar, V., Jangra, A., Mishra, M., Ahmed, S., Ghosh, P., Vohora, D., and other oxidation substrates and antioxidants by means of Folin-Ciocalteu
Khanam, R., 2016. Ameliorative effect of naringin against doxorubicin-induced reagent. Meth. Enzymol. 299, 152–178. [Link]
acute cardiac toxicity in rats. Pharm. Biol. 54, 637–647. [Link] (99)99017-1.
13880209.2015.1070879. Trease, G.E., Evans, W.C., 2009. Pharmacognosy. Bailliere Tindall Ltd, London.
Li, Z., Chinnathambi, A., Alharbi, S.A., Yin, F., 2020. Plumbagin protects the World Health Organization, 1996. Guidelines for the assessment of herbal
myocardial damage by modulating the cardiac biomarkers, antioxidants, and medicines. WHO Technical Report Series, World Health Organization, Geneva.
apoptosis signaling in the doxorubicin-induced cardiotoxicity in rats. Environ. Available at: [Link]
Toxicol. 35, 1374–1385. [Link] medicine/publications/trs1010_annex2.pdf?ua=1.

831
J.A.N. Sandamali, R.P. Hewawasam, K.A.P.W. Jayatilaka et al. Saudi Pharmaceutical Journal 29 (2021) 820–832

World Health Organization, 2011. Quality control methods for herbal materials. Yusuf, A.Z., Zakir, A., Shemau, Z., Abdullahi, M., Halima, S.A., 2014. Phytochemical
WHO Press, World Health Organization, Geneva, Switzerland. Available at: analysis of the methanol leaves extract of Paullinia pinnata linn. J. Pharmacog.
[Link] Phytother. 6, 10–16. [Link]
pdf;jsessionid=698837D8E9627AA38A9A831738FC6F95?sequence=1. Zhang, Y.Y., Yi, M., Huang, Y.P., 2017. Oxymatrine ameliorates doxorubicin-induced
Xiao, N.N., 2015. Effects of resveratrol supplementation on oxidative damage and cardiotoxicity in rats. Cell. Physiol. Biochem. 43, 626–635. [Link]
lipid peroxidation induced by strenuous exercise in rats. Biomol. Ther. 23, 374– 10.1159/000480471.
378. [Link]

832