Colloids and Surfaces B: Biointerfaces 184 (2019) 110465

Contents lists available at ScienceDirect

Colloids and Surfaces B: Biointerfaces

journal homepage: www.elsevier.com/locate/colsurfb

Effect of Ficus carica L. leaves extract loaded gold nanoparticles against T

cisplatin-induced acute kidney injury
⁎
Samah M. El-Sayeda, Mehrez E. El-Naggarb, , Jihan Husseinc, Dalia Medhatc, Mona El-Bannac
a
Dairy Science Department, National Research Centre, Dokki, Giza, P.O. 12622 Egypt
b
Textile Research Division, National Research Centre, 33 El Bohouth st., P.O. 12622, Dokki, Giza, Egypt
c
Medical Biochemistry Department, Medical Research Division, National Research Centre, Dokki, Giza, Egypt

A R T I C LE I N FO A B S T R A C T

Keywords: Background: Cisplatin (CisPt) is one of the most widely used and highly effective drugs for the treatment of
Ficus carica L various solid tumors, unfortunately acute kidney injury (AKI) is considered one of its side effects through several
Gold nanoparticles mechanisms including production of reactive oxygen species (ROS), pro-inflammatory and pro-fibrotic cyto-
Cisplatin kines. Due to the poor effect of AKI therapy, the use of nanoparticles loaded with natural extracts for delivering
Homocysteine
to the kidney molecules are desirable.
Hydroxyproline
Aim: This study aims to investigate the effectiveness of different concentrations of gold nanoparticles (Au-NPs)
as a carrier for Ficus carica L. (Fig) leaves extract against CisPt induced AKI.
Methods: Seventy male albino rats were used and divided into seven groups. After the experimental period,
blood was withdrawn, serum was separated for determination of urea, creatinine, homocystein (Hcy) and folic
acid while reduced glutathione (GSH), nitric oxide (NO), malondialdehyde (MDA), total antioxidant capacity
(TAC) and hydroxyproline content (Hyp) were evaluated in kidney tissue homogenate.
Results: CisPt induced AKI in rats and results in a significant increase in the levels of serum urea, creatinine, Hcy
and kidney Hyp, lipid peroxidation along with a significant reduction of kidney GSH, NO and TAC compared to
the control rats. Treatment with Au-NPs and Fig extract particularly in a ratio of (3:2) respectively was shown to
improve renal functions with efficient capacity in scavenging ROS and reduced AKI severity.
Conclusion: Au-NPs enhanced the anti-oxidative properties of the Fig extract in targeting kidney damaged tissue
and reduced oxidative toxicity induced by CisPt.

1. Introduction
Cis-diamminedichloroplatinum II (CisPt) is a well-documented drug
in the treatment of several malignancy types; while a limitation of its
use is well observed because CisPt -induced nephrotoxicity (CIN) in
more than 50% of cases [1]. Several mechanisms were involved in
nephrotoxicity including endothelin-1, upregulation of transforming
growth factor-β, attenuation of oxidative stress, also, necrosis and
apoptosis in addition to the elevation of macrophage and monocyte
infiltration into the renal cortex [2]. Antioxidants have become a topic
of interest to health and food science researchers and medical experts.
Food that has antioxidants properties have an effective role in pro-
tecting against different diseases concerning oxidative stress such as
liver and kidney diseases in a long duration as chemo preventive agents
[3]. Nanotechnology can confer significant benefit to medicine, such as
the targeted delivery of drugs to specific tissues. Nanomedicines in the
clinic have increased drug solubility, reduced off-target side effects, and
provides novel diagnostic tools; there is an increasing cohort of nano-
materials which may have implications for kidney disease [4]. Of these
nanomedicines is gold nanoparticles (Au-NPs) which have attracted
intensive interest owing to some features such as being easily prepared,
high surface area (more active), low toxicity and can be readily at-
tached to molecules of biological interest.
Ficus carica L. (Fig) is a common tree in the Middle East and the
Mediterranean region, belonging to the botanical family. Fig is usually
consumed its dry or fresh grade [5]. It is an excellent source of minerals
and vitamins; it is fat and cholesterol-free and contain a high number of
amino acids [6] and a large number of phenolic compounds [7].
Therefore, the present work was designed to investigate the effec-
tiveness of both Au-NPs and Fig leaves extract against CisPt induced
AKI. Furthermore, the synergistic behavior of augmenting both agents
was thoroughly studied. Different concentrations of Au-NPs loaded with

⁎
Corresponding author.
E-mail addresses: Mehrez_chem@yahoo.com, mehrezeelnaggar@gmail.com (M.E. El-Naggar).

https://doi.org/10.1016/j.colsurfb.2019.110465
Received 7 March 2019; Received in revised form 22 August 2019; Accepted 27 August 2019
Available online 12 September 2019
0927-7765/ © 2019 Elsevier B.V. All rights reserved.
S.M. El-Sayed, et al.
Fig leaves extract were examined in ameliorating CisPt induced ne-
phrotoxicity in albino rats.
2. Materials and methods
2.1. Materials
2.1.1. Chemicals
Tetrachloroauric Acid (HAuCL4) and Hcy standard {for high per-
formance liquid chromatography (HPLC)} were obtained from Sigma-
Aldrich Company USA. Tri-sodium Citrate was purchased from Fischer
Co. Germany. Cisplatin was purchased from Mylan, Saint Priest, France.
Chloramine T was purchased from Fisher, Fair Lawn, NJ, USA. P-di-
methylaminobenzaldehyde, and Hyp were obtained from Sigma
Chemical Company (U.S.A). Sodium acetate, sodium hydroxide, acetic
acid, citric acid, n-propanol and ethanol were of high grade and pur-
chased from Fisher Scientific (USA). All other chemicals were used as
received and Deionized water was used for experiments, characteriza-
tion and application.
2.1.2. Plant
Fresh Fig leaves were collected in May 2018 from a farm in
Menofyia governorate, Egypt. The plant was identified on the
Herbarium of the National Research Centre (NRC) of Egypt. The leaves
were washed and shade dried at room temperature for one week and
then cut into small pieces. Then ground into powder with special
electric mill and stored in polyethylene bags until extraction.
2.1.3. Experimental animals
Seventy male albino rats, weighing 180 ± 20 g from Animal House,
National Research Centre (NRC) Giza, Egypt was kept in clean cages of
polypropylene and maintained in controlled room temperature with
light and dark cycle, given a standard diet and water ad libitum along
the experimental period. The experiment was carried out in accordance
with guidelines and protocol approved by the Institutional Animal
Ethics Committee of National Research Centre (NRC), Giza, Egypt.
2.2. Methods
2.2.1. Cisplatin induction
Rats received a single dose of CisPt (20 mg/kg body weight) by
intraperitoneal injection; the dose was modified from a previous study
[8].
2.2.2. Preparation of tissue homogenate
One gram from kidney tissue was cut into tiny pieces and then
homogenized with phosphate buffer (pH was adjusted to 7.4). After
that, the homogenate was centrifuged using cooling centrifuge
(Laborzentrifugen, 2K15, Sigma, Germany) at 5000 rpm for 10 min at
4 °C; clear supernatant was then removed and used for estimation of
different parameters [9].
2.2.3. Preparation of Fig leaves extract
In order to prepare the extract, 50 g of dried leaves were extracted
with 250 ml of 95% ethanol (v/v) by cold extraction. The whole solu-
tion containing Fig leaves were kept under stirring at room temperature
for 70 h using an orbital shaker. At the end of stirring time, the mixture
was then filtered using filter paper (Whatman No.1). After that, the
solvent was evaporated at 40 °C from the extracted mixture using a
rotary evaporator under reduced pressure. Finally, the dried extract was
kept in dark sterile screw capped bottles and stored in 4 °C for further
characterization and application.
2.2.4. Preparation of gold nanoparticles (Au-NPs)
Commonly, the production of Au-NPs via the chemical reduction
method involves two main parts: the first part is the reduction of Au
2
Colloids and Surfaces B: Biointerfaces 184 (2019) 110465
ions to Au0. For this purpose, the reduction reaction between tetra-
chloroauric acid (HAuCl4) and tri-sodium citrate (Na3C6H5O7.2H2O)
was used in an aqueous solution. The second part is the stabilization
effect using cationic surfactant; cetyltrimethyl ammonium bromide
(CTAB) to avoid the aggregation of the formed nanoparticles. Finally,
the prepared Au-NPs with narrow size distribution were obtained.
Typically, 500 mg of tri-sodium citrate was dissolved in 50 ml of deio-
nized water, kept under magnetic stirring while the temperature was
raised to 70 °C. After complete dissolution, 500 mg of CTAB was added
under stirring. The pH of the reaction medium was adjusted to 11 by
using NaOH solution (0.01 M). At the end of mixing, 250 mg of HAuCl4/
5 ml of deionized water was added dropwise to the previous solution
and the whole reaction was kept for another 30 min under stirring. At
the end of the reaction, the color of the solution turned to red indicating
the successful preparation of Au-NPs. The solution was kept away from
light for further characterization and application. Techniques such as
UV–vis spectroscopy, TEM and XRD were used to characterize and
determine the size and size distribution of the synthesized Au-NPs.
2.2.5. Preparation of Fig leaves extract loaded Au-NPs
Different concentrations from both Au-NPs and Fig extract were
prepared as follows: Au-NPs and Fig extract in a ratio of (2:3) v/v, Au-
NPs and Fig extract in a ratio of (2:2) v/v and Au NPs and Fig extract in
a ratio of (3:2) v/v. Rats received different concentrations of Au-NPs/
Fig extract mixture (0.5 ml/Kg rat) by oral administration daily for 14
days.
2.2.6. Experimental design
Seventy rats were included in this study and equally divided into
seven groups as follow: Control group: animals received a vehicle, AKI
group: Animals received a single dose of Cispt and received a vehicle
daily, treated group I: Rats received a single dose of CisPt then received
Fig leaves extract (400 mg extract /Kg b.w. /day) orally for 14 days.
Treated group II: Rats received a single dose of CisPt then received Au-
NPs (1 mg/kg b.w. in aqueous solution/day) orally for 14 days. Treated
group III: Rats received a single dose of CisPt then received 0.5 ml of Au
NPs/Fig leaves extract mixture as (2:3) v/v respectively orally for 14
days, treated IV group: Rats received a single dose of CisPt then re-
ceived 0.5 ml of Au-NPs/Fig leaves extract mixture as (2:2) v/v re-
spectively orally for 14 days, treated group V: Rats received a single
dose of CisPt then received 0.5 ml of Au-NPs/Fig leaves extract mixture
as (3:2) v/v respectively orally for 14 days.
After finalizing the experimental period, all animals were kept
fasting for12 h for blood sampling that was collected in clean tubes and
centrifuged at 3000 rpm for separation of sera and then stored at
−20 °C. After that, the biochemical analysis was carried for the sepa-
rated sera while kidney tissues were removed quickly from each rat,
washed with ice-cold saline and kept at -80 °C until used for determi-
nation of biochemical parameters.
2.3. Characterization
2.3.1. Characterization of fig leaves extract
Total phenol content of the Fig leaves plant (TPC) was assessed
calorimetrically by means of the Folin–Ciocalteau reagent at 625 nm
using the process designated by Rashidinejad et al., [10]. The solution
of Fig leaves extract (0.5 ml) mixed with 0.625 mL of the Folin–Cio-
calteau reagent were added to 20 mL of deionized water in a 25 mL
volumetric flask. After 3 min, 2.5 mL of sodium carbonate (35%) was
added to the previous solution. The content was mixed and diluted to
volume with deionized water. After 60 min, the absorbance of the
sample was measured at 625 nm against a blank using a double-beam
ultraviolet–visible spectrophotometer (UV–vis) Hitachi U-3210 (Hi-
tachi, Ltd., Tokyo, Japan). Gallic acid served as a standard compound
for preparing the calibration curve, and ranged from 2.5 to 20 μg/25 μL
of assay solution.
S.M. El-Sayed, et al.
Additionally, the antioxidant activity of the phenol extracts was
evaluated according to the method described by Bandoneon et al., [11].
The method was carried by mixing 3.9 mL methanolic solution of 2, 2-
diphenyl-1-picryl-hydrazyl radical (DPPH) (0.0025 g/100 mL CH3OH)
and 0.1 mL of methanolic solutions of phenol extracts in a cuvette and
the absorbance at 515 nm was measured against methanol using a
double-beam Uv–vis spectroscopy. On the other hand, the absorbance
at this definite wavelength was measured for the blank sample which
consists of a mixture containing 0.1 mL methanol and 3.9 mL metha-
nolic solution of DPPH). The inhibition percentage of DPPH for the
phenol extract and blank samples were calculated as follow:
DPPH (%) = 100 X (A-A0)/A0
A0 is coded to the absorbance for the blank sample at 515 nm, time
t = 0 min while, A is for the final absorbance of the tested sample at
515 nm.
2.3.2. Characterization of the prepared Au-NPs
The prepared colloidal solution of gold nanoparticles (Au-NPs) was
characterized by making use of ultraviolet–visible spectrophotometer
(UV–vis), transmission electron microscopy (TEM) and X-ray diffraction
pattern (XRD). The absorbance and wavelength (nm) of Au-NPs was
determined using UV–vis (UV 1800 Shimadzu, Germany). While the
particle shape and particle distribution of Au-NPs were evaluated using
TEM (a JEOL JEM 3010; Japan) at 300 kV. The colloidal solution of Au-
NPs was precipitated using centrifuge at 12,000 rpm and the resultant
powder was taken for examination by using an X-ray diff ;ractometer
(RINT2000, Rigaku, Japan).
2.3.3. Biochemical analysis
2.3.3.1. Kidney functions. Blood urea was estimated by colorimetric
method as described previously [12] while serum creatinine was
determined by kinetic method according to Laresn et al. [13].
2.3.3.2. Determination of kidney oxidants/antioxidants
parameters. Kidney reduced glutathione (GSH), MDANO TAC were
determined according to Watanabe et al. [14], Uchiyamara et al.
[15], Nakano et al. [16], and Mahmoud et al. [17], respectively.
2.3.3.3. Determination of kidney hyp content. A colorimetric method was
carried out according to the method described previously [18] and
modified by El-Khayat et al. [19]. Briefly, 0.5 g from kidney tissue was
hydrolyzed in hydrochloric acid (HCl; 6 mol/L) at 100 °C for 20 h; the
solution was then centrifuged at 3000 rpm for 10 min; the clear
supernatant was then kept in chloramine-T (0.05 mol/L) for 10 min at
room temperature, then in Ehrlich' sperchloric acid solution for 15 min
at 65 °C; sample was measured spectrophotometrically at wavelength
561 nm and the resulting absorbance of each sample was obtained from
a drawn standard curve.
2.3.3.4. Estimation of hcy. Hcy was estimated by the chromatographic
method using HPLC system (Agilent technologies, 1100 series) with a
quaternary pump according to Hussein et al., [20]. The sample was
prepared as follow: 16 μL from trichloroacetic acid (TCA, 1.2 mol/L)
were added to 200 μL serum (from each rat) then mixed well for 20 s
and incubated in −20 °C for 30 min to precipitate protein; the mixture
was then centrifuged for 20 min at 4000 rpm at 4 °C; the supernatant
was filtered through sample filter (PVDF, 0.45μ m).
2.3.3.5. HPLC condition. 30 μL from each sample were injected onto
HPLC column; separation was completed on reversed phase (RF)
column (C18, 25, 0.46 cm i.d. 5 μm) with isocratic method in which
the mobile phase (M.P) consisted of equal volumes of 1- sodium
phosphate (40 mmol/L). 2- heptanesulfonic acid (8 mmol/L) 3-
methanol (18%) v/v; M.P was adjusted to pH 3.1 by addition of few
3
Colloids and Surfaces B: Biointerfaces 184 (2019) 110465
drops from phosphoric acid ; then filtered twice through a membrane
filter (Whatman, 47 mm Dia.,0.45-μm). The M.P was delivered through
the column at a flow rate of 1 ml/min a controlled temperature (40 °C)
using UV detector that was set at 260 nm. The peak areas of samples
were determined and the concentration of each sample was calculated
from the standard curve.
2.3.3.6. Determination of serum folic acid. Serum folic acid was
measured using ELISA Kit (NOVA, Bioneovan Co., Ltd) according to
the manufacturer’s instructions.
2.3.4. Statistical analysis
In the present study, we used statistical package for social science
(SPSS software version 12, Chicago, Illinois). All numeric variables
were expressed as mean ± standard error (SE) and analyzed by one-
way analysis of variance (ANOVA) followed by Duncan’s multiple range
test for post-hoc comparison of group means. For all tests a probability
(P < 0.05) was considered significant.
3. Results and discussion
Before the formation of the Fig leaves extract, total phenol content
(mg/g) and antioxidant activity (%) are determined for the Fig leaves
plant and the obtained data are recorded in Table 1. It is clearly seen
that the total phenol content is 5.13 mg/g while the average value of
the antioxidant activity (%) determined by DPPH for the Fig leaves is
about 47.34% confirming that the Fig leaves plant has large amount of
phenol compounds and has antioxidant activity. These valuable prop-
erties tend for the utilization of Fig in medical domains as shown in this
current work.
The synthesis of Au-NPs with very small diameter size is obtained
via using CTAB as a cationic stabilizing agent in presence of tri-sodium
citrate which act as reductant (reducing agent). Firstly, seeds of Au
acting as nucleation centers which permit to Au ions to be reduced.
Secondly, the starting oxidation state of Au0 in the synthesis is Au3+
and the final oxidation state was Au0. As a consequence, tri-sodium
citrate (reducing agent) is needed in order to reduce Au3+ to Au0. The
formed particles have the ability to aggregate because of the huge ac-
tive surface area and attributed to the absence of strong stabilizing
agent [21–24]. CTAB as stabilizing agent is necessary to protect these
formed nanoparticles from aggregation. The formation of Au-NPs is
confirmed by the change of color during reaction due to the conversion
of Au+ to Au-NPs. The colorless of solution is changed to red color with
the presence of tri-sodium citrate and CTAB. After the formation of Au-
NPs, different concentrations of Au-NPs are blended with Fig leaves
extract to form different ratios of Au-NPs/Fig leaves extract mixtures.
As mentioned above in experimental part, Fig leaves extract is obtained
by the cold extraction of the dried Fig leaves using ethanol followed by
filtration. Scheme 1 clarifies the extraction and the formation of Au-NPs
and the color change due to the formation of Au-NPs and Au-NPs/Fig
leaves extract as well.
After the formation of Au-NPs, there are many tools used to char-
acterize the originated nanoparticles. From these tools; UV–vis spec-
troscopy, TEM and XRD. Fig. 1 represents the UV–vis, TEM and XRD of
Au-NPs and Au-NPs blended with Fig leaves extract. As shown in
UV–vis spectroscopy (Fig. 1A), a definite optical feature generally
Table 1
Antioxidant activity % by DPPH and total phenol content mg/g dry weight
samples.
plant Total phenol content (mg/ Antioxidant activity % by
g) D. W DPPH
Fig (Ficus carica L.) 5.13 ± 0.18 47.34 ± 1.15
Mean ± standard deviation, D.W: Dry Weight.
S.M. El-Sayed, et al.
Scheme 1. Formation of Fig leaves extract, Au-NPs and Fig leaves extract blended with Au-NPs.
exhibited for the identified surface plasmon resonance of Au-NPs. This
identified surface plasmon resonance of Au-NPs results in a strong ab-
sorbance band in the visible region; 500 nm–560 nm. As displayed from
Fig. 1A that the formed Au-NPs colloidal solution exhibits a peak at
520 nm confirming the successful conversion of Au ions to Au-NPs. The
full conversion of ions to nanoparticles is confirmed by the addition of
NaCl to the formed colloidal solution of Au-NPs. There is no white
precipitate is formed upon the addition of NaCl to Au-NPs solution that
confirms that there is no noticeable existence of ions at the end of the
reaction.
TEM was characteristically used to establish the physical size and
morphological structure of Au-NPs. Fig. 1B shows a TEM image of Au-
NPs reduced via tri-sodium citrate as reductant and CTAB as a capping
agent. As clearly seen from the onset image of TEM (Fig. 1B), the
particles are formed with a very small diameter size. The high-resolu-
tion image (Fig. 1B) is taken for better understanding the shape of the
particles which reveal that the hexagonal shapes of Au-NPs are formed.
For further characterization, XRD was used to study the structural
properties of the as-prepared Au-NPs. Fig. 1C shows XRD of Au-NPs in
its powder form after precipitation of the colloidal solution using cen-
trifuge instrument with 15,000 rpm for 2 h. As presented from XRD, the
formed Au-NPs exhibit characteristic peaks at 38°, 43.8° and 65° as-
cribed to the crystallographic planes (1 1 1), (2 0 0) and (2 2 0), cor-
respondingly. The peak width of Au-NPs from crystalline plane (1 1 1),
sizes of the Au0 crystallite were found to be approximately 20 nm.
On the other hand, after mixing Fig leaves extract with Au-NPs in
equal amounts and subjected to 30 min of sonication, it is essential to
assess the particle shape of the formed hybrids. Fig. 1D displays TE
image of Fig leaves extract blended with Au-NPs. It is evidently seen
4
Colloids and Surfaces B: Biointerfaces 184 (2019) 110465
that the particle shape turned to spherical shape with a marginal in-
crease in the particle size due to the coating effect of Fig leaves extract
on the surface of Au-NPs but still in the nanoform state. Thus, TEM was
effectively used to analyze the shape and diameter of the as synthesized
Au-NPs with and without mixing the Fig leaves extract.
Moving to the second part of the current work to evaluate the hy-
brids of Fig leaves extract blended with different concentrations of Au-
NPs against cisplatin induced acute kidney injury. CisPt, the anti-neo-
plastic drug, is used in the treatment of several types of malignancies
include head, bladder, breast, ovarian and lung cancer. In spite of its
role in improvement of patient’s life expectancy [25]. It is considered as
a strong toxin and the nephrotoxicity is one of its important compli-
cations in clinical and also in experimental models that can be pro-
gressive in a lot of cases [26]. Thus, acute renal toxicity remains the
main problem of CisPt' use [25].
In the present study, CisPt injection significantly increased blood
urea and serum creatinine in CisPt group in compared with control
animals (Table 2). Elevation of kidney functions in this work is related
to the deleterious side effects of CisPt that generates ROS as was ob-
served in Table 3, thus it significantly increased lipid peroxidation and
significantly decreased GSH, NO and TAC in CisPt group compared to
control.
Liberation of reactive oxygen species (ROS) and renal micro-
vasculature vasoconstriction are well dependable on CIN. Moreover,
cisplatin selectively injured S3 segment of proximal tubules that is
found in the outer stripe of the medulla that is found outside, as evi-
denced by necrosis & apoptosis. It was reported that generation of NOS,
the reactive nitrogen species, have been also involved in CIN; CisPt
exalts the liberation of NO and peroxynitrite in renal tissues; the later
S.M. El-Sayed, et al. Colloids and Surfaces B: Biointerfaces 184 (2019) 110465

Fig. 1. (A) Uv–vis, (B) TEM, (C) XRD of Au-NPs and (D) TEM of Au-NPs loaded with Fig leaves extract.

Table 2 tubular cells necrosis that leads to the reduction of glomerular filtration
Blood urea and serum creatinine levels in different studied groups. rate (GFR) and kidney dysfunction [2].
Parameters Groups Urea (mg/dl) Creatinine (mg/dl) Elevation of oxidative stress parameters in this study may be related
to accumulation of CisPt in the kidney by peritubular uptake. Whereas,
Control 25 ± 5.7 0.83 ± 0.14 such this concentration of the drug in the renal cortex is greater than
Cisplatin 113a ± 6.6 2.2a ± 0.20
other organs.
Treated I 70a,b ± 5.0 1.4a,b ± 0.14
Treated II 38.3b ± 7.2 0.7b ± 0.05
The pathological mechanisms of renal toxicity involve elevation of
Treated III 40b,c ± 12.5 0.96b,c ± 0.18 endothelin-1 and transforming growth factor β, oxidative stress, ne-
Treated VI 40b,c ± 2.8 0.66b,c ± 0.21 crosis, apoptosis and increase in macrophage/monocyte infiltration into
Treated V 33.3b,c ± 4.4 0.40b,c ± 0.05 the renal cortex and medulla [2].
Renal dysfunction e.g. chronic kidney disease (CKD) preexisting
P: a significant difference compared to Pa) the control group.
when CisPt chemotherapy is started may induce tubulo-interstitial le-
Pb) CisPt group, Pc) treated I group and Pd)treated II group.
sions, which may further evolve towards interstitial inflammation and
fibrosis. In this study, hydroxyproline content in the kidney was sig-
effectively stimulates the changes of lipid functions and structure, DNA,
nificantly increased by CisPt injection compared with control subjects
proteins and also the reduction of cellular defenses system by oxidation
(Table 4) that appeared the role of CisPt in inducing AKI. In the same
of thiol pools [1].
line, El-Khayat et al., [19] indicated the accumulation of hydroxypro-
In addition, renal toxicity involves kidney free radical production
line content in cirrhotic rat's liver.
and increasing antioxidant defense mechanisms, and also acute renal
Additionally, this study showed a significant increase in serum Hcy

Table 3
Oxidant and antioxidant levels in different studied groups.
Parameters Groups GSH (mg/g tissue) MDA (nmol/g tissue) NO (U/g tissue) TAC (mM/g tissue)

Control 111 ± 6.8 28.7 ± 0.8 11.7 ± 0.83 18.9 ± 1.1

Cisplatin 58.7a ± 2.2 81.5a ± 1.4 3.7a ± 0.34 7.3a ± 0.77
Treated I 83a,b ± 3.0 43.9a,b ± 1.1 10.6b ± 1.0 15.7a,b ± 0.30
Treated II 83.9a,b ± 6.6 43.2a,b ± 1.5 9.1b ± 0.17 16.7b ± 0.78
Treated III 83.1a,b ± 5.0 48.6a,b ± 4.4 7.5a,b,c ± 0.21 14.2a,b ± 0.60
Treated VI 84.9b ± 3.2 34.8b,c,d ± 2.5 8.4a,b,c ± 0.38 14.8a,b ± 1.4
Treated V 100.5b,c,d ± 2.0 35.1b,c,d ± 0.8 10.4b ± 0.5 17.8b ± 0.59

P: a significant difference compared to Pa) the control group, Pb) CisPt group, Pc) treated I group and Pd) treated II group.

5
S.M. El-Sayed, et al. Colloids and Surfaces B: Biointerfaces 184 (2019) 110465

Table 4 oxidative-stress associated diseases such as cancer [31], cardiovascular

Kidney hydroxyproline content in different studied groups. [32] or neurodegenerative diseases [33] and liver [3].
Parameters Groups Hyp content (mg/g tissue) Fig leaves is one of several plants that contain different anti-
oxidative fractions which are able to inhibit lipid oxidation effectively,
Control 51.6 ± 4.1 by different mechanisms of action [34]. It was observed that, admin-
Cisplatin 160a ± 2.8
istration of Fig leaves extract was significantly decreased kidney func-
Treated I 73.6a,b ± 3.3
Treated II 48b ± 1.2
tions, MDA, Hyp and Hcy in compared to CisPt group; however, these
Treated III 55.6b,c ± 0.88 parameters were still significantly increased compared to control.
Treated VI 52.2b,c ± 3.3 Contrarily, antioxidant parameters and folic acid were significantly
Treated V 49b,c ± 1.4 decreased by Fig extract administration compared to CisPt group al-
though these parameters were still significantly decreased compared to
P: a significant difference compared to Pa) the control group.
control (Tables 1–5). These findings were consistent with Kore et al.
Pb) CisPt group, Pc) treated I group and Pd) treated II group.
[35] who reported that Fig extract caused marked reduction in serum
urea and creatinine levels in gentamicin induced nephrotoxicity; these
Table 5
Serum homocystein and folic acid levels in different studied groups. findings are indicative of antioxidant properties of Fig which are mainly
due to its high contents of flavonoids and phenolic compounds.
Parameters Groups Hcy (μmol/L) Folic acid (Pg/ml) One of the most common uses of nanoparticles (NPs) is in the de-
Control 21.7 ± 3.1 11.6 ± 0.55 livery and controlled release of therapeutic molecules. Nanomaterials
Cisplatin 76.2a ± 3.4 3.9a ± 0.58 have been engineered to incorporate small molecule drugs as well as
Treated I 32.8a,b ± 4.2 8.3a,b ± 0.50 macromolecules such as nucleic acids, proteins, and peptides [4], in
Treated II 38.3a,b ± 1.2 5a ± 0.86 addition it was suggested that delivery of antioxidants to mitochondria
Treated III 27.6b,d ± 3.3 8.5a,b,d ± 0.76
Treated VI 26.6b,d ± 4.3 8.1a,b,d ± 0.37
is one of the most important mechanisms for prevention of CIN with the
Treated V 25.3b,d ± 0.98 9a,b,d ± 0.50 purpose of decreasing toxic oxidative stress injuries [36].
In this study, Fig leaves extracts are mixed with Au-NPs in different
P:a significant difference compared to Pa) the control group, Pb) CisPt group, Pc) concentrations to compare between groups in order to find the best
treated I group and P)treated II group. effect of using extract, besides using Au-NPs alone in treatment as well.
The combination between Au-NPs and Fig extract particularly in the
accompanied with a significant decrease in serum folic acid in CisPt ratio of 3/2 (Au-NPs/Fig leaves extract) respectively (treated V) sig-
group compared to the control group (Table 5). nificantly decreased MDA, hydroxyproline content, blood urea, serum
Hyperhomocysteinemia (HHcy) is a condition developed by genetic creatinine and homocystein levels compared to the CisPt group, these
enzymatic deficiencies and/or nutritional defects that interfere with the results were getting close to those of the control group. On the other
proper metabolism of methionine [27]. Genetic variation of the en- hand, it significantly increased antioxidant parameters and folic acid
zymes, or deficiency of nutritional factors, including folic acid, vitamin compared to CisPt group. Interestingly, all these parameters were sig-
B6 and B12, impairs Hcy metabolism and cause moderate HHcy. Ele- nificantly changed compared to other treated groups indicating the
vated plasma Hcy levels are prevalent in CKD patients and the lower beneficial effect of this ratio (3/2) of Au-NPs/Fig leaves extract; that
glomerular filtration rate (GFR) is associated with a higher plasma total indicated the effective role of Au-NPs in the delivery of therapeutic
homocysteine (tHcy) concentration [28]. Thus, there are considerable molecules (Fig leaves extract). Thus, Au-NPs in this study effectively
evidences showing that HHcy is implicated in the progression of CKD. improved the efficacy of using extract and gave the maximum effect
CisPt molecules can pass freely in the glomerulus and penetrate the more than using it alone or with other ratio with small amounts of Au-
tubular cells due to their low molecular weight, once these small mo- NPs.
lecules enter the tubular cells, they became activate and formed a Au-NPs improve the drugs specificity to target the injured organ
glutathione-conjugate which is metabolized to a cysteinyl glycine with minimized side effects, and also to circumvent the potential drug
conjugate and then to cysteine conjugate, and finally to a reactive thiol. resistance mechanisms that may develop by the diseased cells [37–39].
Interestingly, it was reported that pre-incubation of CisPt with cy- Thus, the current study offers a promising compound with a specific
steinyl-glycine, or N-acetyl-cysteine increased the toxicity of cisplatin ratio (3/2) of Au-NPs/fig leaves extract to verify the hypothesis of
due to the formation of cysteine conjugate. The oxidized Hcy in human improving the Fig extract properties by gold nanoparticles and to give
and animal plasma exists as a disulfide form, including homocystine the best effect for attenuation of renal dysfunction.
(Hcys-S-S-Hcys) and cysteinylhomocysteine (Cys-S-S-Hcys) [29].
Therefore, increased plasma levels of Hcy in hyperhomocysteinemic
(HHcy) mice might attenuate the amount of cysteine that promotes 4. Conclusion
cysteinyl-glycine-conjugate formation to enhance the toxicity of cis-
platin. Gold nanoparticles (Au-NPs) was successfully prepared via chemical
The highest concentration of CisPt is observed in mitochondria, reduction method using tri-sodium citrate as reductant and cetyl-
nuclei, cytosol and microsomes and is interlaced to glutathione and trimethyl ammonium bromide (CTAB) as stabilizing agent for pro-
next metabolized through gamma glutamyltranspeptidase and a cy- tecting the formed nanoparticles from agglomeration. It was concluded
steine S-conjugate β-lyase-dependent pathways to a reactive thiol as an that the Fig extract leaves blended with Au-NPs have an important role
active nephrotoxin [30]. in attenuating kidney toxicity induced by CisPt, through its high con-
Cisplatin-induced oxidative stress and inflammation in the kidney tents of polyphenolic compounds that have the ability in protecting
may be partially prohibited by several natural products as well as against oxidative stress induced by CisPt. In addition, Au-NPs as a
chemical compounds that have antioxidant properties. Plant-derived carrier enhanced effectively the antioxidative properties of Fig extract
antioxidant compounds are molecules that donate electrons or hy- in targeting kidney damaged tissue and in reducing oxidative toxicity
drogen atoms. These compounds have the ability to form less reactive induced by CisPt. These findings offer prospective approaches in uti-
species, which are professionally quenched by other electron or hy- lizing nanoparticles as drug delivery molecules in the treatment of
drogen sources to protect cellular damage. Therefore, they help in de- kidney diseases.
laying and inhibiting lipid oxidation and so protecting human and an-
imal cells against oxidative stress and decreasing the risk of several

6
S.M. El-Sayed, et al.
References
[1] F. Hayati, M. Hossainzadeh, S. Shayanpour, Z. Abedi-Gheshlaghi, S.S.B. Mousavi,
Prevention of cisplatin nephrotoxicity, J. Nephropharmacol. 5 (2016) 57.
[2] M. Reck, J. Von Pawel, P. Zatloukal, R. Ramlau, V. Gorbounova, V. Hirsh, N. Leighl,
J. Mezger, V. Archer, N. Moore, Overall survival with cisplatin–gemcitabine and
bevacizumab or placebo as first-line therapy for nonsquamous non-small-cell lung
cancer: results from a randomised phase III trial (AVAiL), Ann. Oncol. 21 (2010)
1804–1809.
[3] Z. El-Khayat, W. Rasheed, T. Ramzy, J. Hussein, M. Agaiby, S. Morsy, F. Morsy,
N. Shaffie, Protective effect of garlic oil against liver injury in experimental animals,
J. Med. Plants Res. 4 (2010) 2359–2369.
[4] R.M. Williams, E.A. Jaimes, D.A. Heller, Nanomedicines for kidney diseases, Kidney
Int. 90 (2016) 740–745.
[5] A.P. Oliveira, P. Valentão, J.A. Pereira, B.M. Silva, F. Tavares, P.B. Andrade, Ficus
carica L.: Metabolic and biological screening, Food Chem. Toxicol. 47 (2009)
2841–2846.
[6] M. Viuda-Martos, X. Barber, J.A. Perez-Alvarez, J. Fernandez-Lopez, Assessment of
chemical, physico-chemical, techno-functional and antioxidant properties of fig
(Ficuscarica L.) powder co-products, Ind. Crops Prod. 69 (2015) 472–479.
[7] R. Veberic, M. Colaric, F. Stampar, Phenolic acids and flavonoids of fig fruit (Ficus
carica L.) in the northern Mediterranean region, Food Chem. 106 (2008) 153–157.
[8] M. Nematbakhsh, F. Ashrafi, H. Nasri, A. Talebi, Z. Pezeshki, F. Eshraghi,
M. Haghighi, A model for prediction of cisplatin induced nephrotoxicity by kidney
weight in experimental rats, J. Res. Med. Sci. 18 (2013) 370.
[9] D. Medhat, M.A. El-Bana, M.N. Ashour, E. Badawy, Y. Diab, J. Hussein, New ap-
proaches in protecting against atherosclerosis in experimental model of post-
menopause, J. Appl. Pharm. Sci. 7 (2017) 090–096.
[10] A. Rashidinejad, E.J. Birch, D. Sun‐Waterhouse, D.W. Everett, Effects of catechin on
the phenolic content and antioxidant properties of low‐fat cheese, Int. J. Food Sci.
Technol. 48 (2013) 2448–2455.
[11] D. Bandonienė, M. Murkovic, W. Pfannhauser, P. Venskutonis, D. Gruzdienė,
Detection and activity evaluation of radical scavenging compounds by using DPPH
free radical and on-line HPLC-DPPH methods, Eur. Food Res. Technol. 214 (2002)
143–147.
[12] W.H. Marsh, B. Fingerhut, H. Miller, Automated and manual direct methods for the
determination of blood urea, Clin. Chem. 11 (1965) 624–627.
[13] K. Larsen, Creatinine assay by a reaction-kinetic principle, Clin. Chim. Acta 41
(1972) 209–217.
[14] M. Watanabe, K. Yamaguchi, K. Chijiiwa, M. Tanaka, FR167653 improves survival
and pulmonary injury after partial hepatectomy under ischemia/reperfusion in rats,
J. Surg. Res. 101 (2001) 146–151.
[15] M. Uchiyama, M. Mihara, Determination of malonaldehyde precursor in tissues by
thiobarbituric acid test, Anal. Biochem. 86 (1978) 271–278.
[16] H. Nakano, M. Yamaguchi, Y. Kaneshiro, K. Yoshida, G. Kigawa, H. Nagasaki,
Y. Fujiwara, F. Matsumoto, N. Kitamura, J. Sasaki, S-adenosyl-L-methionine at-
tenuates ischemia-reperfusion injury of steatotic livers, Transplantation
Proceedings (1998) 3735–3736.
[17] F. Khaled, A. Ramadan, I. Ashoush, Nanoencapsulation and nanoemulsion of
bioactive compounds to enhance their antioxidant activity in food, Int. J. Food Sci.
Technol. 4 (2014) 1–22.
[18] S. Patiyal, S. Katoch, Tissue specific and variable collagen proliferation in Swiss
albino mice treated with clenbuterol, Physiol. Res. 55 (2006).
[19] Z. El-Khayat, E. Mostafa, J. Hussein, M. El-Waseef, L. Rashed, A.R. Farrag,
D. Medhat, Mesenchymal stem cells therapy for thioacetamide induced liver cir-
rhosis, Int. J. Pharm. Pharm. Sci. 5 (2013) 196–203.
[20] J. Hussein, Z. El-Khayat, A.R. Farrag, D. Medhat, Y. Abdel, Inhibition of hyperho-
mocysteinemia in Indomethacin induced peptic ulcer: Impact of pomegranate juice
supplementation, Inhibition of hyperhomocysteinemia in Indomethacin induced
peptic ulcer: Impact of pomegranate juice supplementation, J. Chem. Pharm. Res. 6
Colloids and Surfaces B: Biointerfaces 184 (2019) 110465
(2014) 131–138.
[21] A.M. Abdelgawad, M.E. El-Naggar, W.H. Eisa, O.J. Rojas, Clean and high-
throughput production of silver nanoparticles mediated by soy protein via solid
state synthesis, J. Clean. Prod. 144 (2017) 501–510.
[22] M.E. El-Naggar, T.I. Shaheen, M. Fouda, A. Hebeish, Eco-friendly microwave-as-
sisted green and rapid synthesis of well stabilized gold and core-shell silver-gold
nanoparticles, Carbohydr. Polym. 136 (2016) 1128–1136.
[23] A. Hebeish, T.I. Shaheen, M.E. El-Naggar, Solid state synthesis of starch-capped
silver nanoparticles, Int. J. Biol. Macromol. 87 (2016) 70–76.
[24] A.A. Yan Jinhua, A.M. Abdelgawad, M.E. El-Naggar, O. Rojas, Antibacterial activity
of silver nanoparticles synthesized in-situ by solution spraying onto cellulose,
Carbohydr. Polym. 147 (2016) 500–508.
[25] V. Bunel, Y. Tournay, T. Baudoux, E. De Prez, M. Marchand, Z. Mekinda,
R. Maréchal, T. Roumeguère, M.-H. Antoine, J.L. Nortier, Early detection of acute
cisplatin nephrotoxicity: interest of urinary monitoring of proximal tubular bio-
markers, Clin. Kidney J. 10 (2017) 639–647.
[26] K.K. Filipski, W.J. Loos, J. Verweij, A. Sparreboom, Interaction of Cisplatin with the
human organic cation transporter 2, Clin. Cancer Res. 14 (2008) 3875–3880.
[27] J. Suszyńska-Zajczyk, O. Utyro, H. Jakubowski, Methionine-induced hyperhomo-
cysteinemia and bleomycin hydrolase deficiency alter the expression of mouse
kidney proteins involved in renal disease, Mol. Genet. Metab. 112 (2014) 339–346.
[28] C. Sabanayagam, A. Shankar, Association between plasma homocysteine and mi-
croalbuminuria in persons without hypertension, diabetes mellitus, and cardio-
vascular disease, Clin. Exp. Nephrol. 15 (2011) 92–99.
[29] F. Yi, P.-L. Li, Mechanisms of homocysteine-induced glomerular injury and
sclerosis, Am. J. Nephrol. 28 (2008) 254–264.
[30] X. Yao, K. Panichpisal, N. Kurtzman, K. Nugent, Cisplatin nephrotoxicity: a review,
Am. J. Med. Sci. 334 (2007) 115–124.
[31] Z. El-khayat, O.A. Abas, D. Medhat, M. Elghreeb, A. Farrag, N. Mostafa, Biochemical
studies on the effect of flaxseed and corn oils on cell membrane phospholipids in
Ehrlich ascites carcinoma and solid tumor in mice, Pharm. Lett. 8 (2016) 90–101.
[32] S. Mubarak, S.A. Hamid, A.R. Farrag, N. Samir, J.S. Hussein, Atherogenic coefficient
and atherogenic index in Doxorubicin–induced cardiotoxicity: impact of date palm
extract, Comp. Clin. Path. 27 (2018) 1515–1522.
[33] A. Scalbert, C. Manach, C. Morand, C. Rémésy, L. Jiménez, Dietary polyphenols and
the prevention of diseases, Crit. Rev. Food Sci. Nutr. 45 (2005) 287–306.
[34] E. Maghsoudlou, R. Esmaeilzadeh Kenari, Z. Raftani Amiri, Evaluation of
Antioxidant Activity of Fig (Ficus carica) Pulp and Skin Extract and Its Application
in Enhancing Oxidative Stability of Canola Oil, J. Food Process. Preserv. 41 (2017)
e13077.
[35] K.J. Kore, R.V. Shete, B.N. Kale, A.S. Borade, Protective role of hydroalcoholic
extract of Ficus carica in gentamicin induced nephrotoxicity in rats, Int. J. Pharm.
Life Sci. 2 (2011).
[36] N. Santos, C.C. Bezerra, N. Martins, C. Curti, M.P. Bianchi, A.C. Santos, Hydroxyl
radical scavenger ameliorates cisplatin-induced nephrotoxicity by preventing oxi-
dative stress, redox state unbalance, impairment of energetic metabolism and
apoptosis in rat kidney mitochondria, Cancer Chemother. Pharmacol. 61 (2008)
145.
[37] T.I. Shaheen, M.E. El-Naggar, J.S. Hussein, M. El-Bana, E. Emara, Z. El-Khayat,
M.M. Fouda, H. Ebaid, A. Hebeish, Antidiabetic assessment; in vivo study of gold
and core-shell silver-gold nanoparticles on streptozotocin-induced diabetic rats,
Biomed. Pharmacother. 83 (2016) 865–875.
[38] D. Medhat, J. Hussein, M.E. El-Naggar, M.F. Attia, M. Anwar, Y.A. Latif, H.F. Booles,
S. Morsy, A.R. Farrag, W.K. Khalil, Effect of Au-dextran NPs as anti-tumor agent
against EAC and solid tumor in mice by biochemical evaluations and histopatho-
logical investigations, Biomed. Pharmacother. 91 (2017) 1006–1016.
[39] J. Hussein, M. El-Banna, K.F. Mahmoud, S. Morsy, Y.A. Latif, D. Medhat, E. Refaat,
A.R. Farrag, S.M. El-Daly, The therapeutic effect of nano-encapsulated and nano-
emulsion forms of carvacrol on experimental liver fibrosis, Biomed. Pharmacother.
90 (2017) 880–887.