Results in Chemistry 4 (2022) 100285

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Results in Chemistry
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Synthesis and characterization of nickel oxide nanoparticles using Clitoria

ternatea flower extract: Photocatalytic dye degradation under sunlight and
antibacterial activity applications
S. Prabhu a, b, *, T. Daniel Thangadurai c, P. Vijai Bharathy d, Pon. Kalugasalam e
a
Department of Research and Development, Bharathiar University, Coimbatore, Tamil Nadu, India
b
Department of Physics, Hindusthan College of Engineering and Technology, Coimbatore, Tamil Nadu, India
c
Department of Chemistry, KPR Institute of Engineering and Technology, Coimbatore, Tamil Nadu, India
d
Department of Physics, C.B.M College, Coimbatore, Tamil Nadu, India
e
Department of Physics, Anna University Regional Campus, Coimbatore, Tamil Nadu, India

A R T I C L E I N F O A B S T R A C T

Keywords: The present study is aimed to synthesize nickel oxide nanoparticles (NiO NPs) by using medicinal flower extract
Green synthesis of Clitoria ternatea and examine its antibacterial and photocatalytic dye degradation activities. The synthesized
NiO NPs NPs were investigated using UV–visible spectroscopy wherein they exhibited a color change pattern and
Clitoria ternatea
confirmed their maximum absorption peak at 280 nm, which is distinct for NiO NPs. The FTIR analysis showed
Antibacterial
the presence of various functional groups in the flower extract as well as on the NiO NPs. XRD studies revealed
Dye degradation
the high crystalline nature of NiO NPs with 9.6 nm average size. The TEM micrograph shows hexagonal shape of
the NPs that are non-agglomerated with a particle size of 13 nm. The composition of nickel and oxygen elements
was confirmed through EDX studies. Using the well diffusion method, the antibacterial activity of NiO NPs was
tested against both gram positive and gram-negative bacterial strains of Staphylococcus aureus and Escherichia
coli. Additionally, photocatalytic degradation of Fast Green (FG) and Rose Bengal (RB) dye molecules were
performed by using NiO NPs. Based on the obtained results, we confirmed that NiO NPs degrade FG by 89 % and
RB by 77 % in the presence of sun light.

1. Introduction NPs, namely, co-precipitation [8], hydrothermal [9], sol–gel [10],

electrochemical [11], solvothermal [12], and microwave-assisted [13].
Recently, metal oxide (MO) based nanomaterials have gained a lot of Nevertheless, these methods are prone to various disadvantages such as
attention because their unique properties such as optical, chemical, usage of toxic chemicals, high cost, and time consumption. Therefore, in
mechanical, electronic, and magnetic properties [1]. These unique this study we took the advantage of the green synthesis method it is
properties make metal oxide (MO) nanomaterials suitable for diverse simple, eco-friendly, cost-effective, non-toxic, fast reaction times,
applications in the field of agriculture, biomedicine, electronics, food biodegradable in nature and high yields production. Recently, NiO NPs
processing, textile and material science [2–4]. Among MOs, NiO is an were synthesized using various plants extracts such as opuntia ficus
important transition MO with a wide bandgap from 3.6 to 4.0 eV, p-type indica [14], Leucas Aspera [15], Calotropis gigantea [16], Callistemon
semiconductor with cubic crystallinity which exhibits significant prop­ viminalis [17], Gymnema Sylvestre [8] and Rhamnus virgata [18] has
erties such as outstanding electrochemical stability, high reactivity, already been reported.
good durability, excellent biocompatibility and bacterial resistivity [5]. In this study, the Clitoria ternatea flower is selected for the synthesis
The positive charge of NiO NPs enables their use as catalyst. The of NiO NPs. The Clitoria ternatea is a traditional Ayurvedic medicinal
controllable size, surface volume, and morphological properties of NiO plant, commonly known as Butterfly pea or Blue pea, and it belongs to
NPs have promoted their usage in catalysis, sensor and biomedical field the family of Fabaceae [19]. The flower is comprised of various phyto­
[6–7]. There are various methods have been employed to synthesize NiO chemicals such as alkaloids, terpenoids, aldehydes, flavonoids,

* Corresponding author at: Department of Research and Development, Bharathiar University, Coimbatore, Tamil Nadu, India.
E-mail address: [email protected] (S. Prabhu).

https://doi.org/10.1016/j.rechem.2022.100285
Received 15 November 2021; Accepted 10 January 2022
Available online 19 January 2022
2211-7156/© 2022 The Author(s). Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-
nc-nd/4.0/).
S. Prabhu et al. Results in Chemistry 4 (2022) 100285

glycosides, phenols, alkaloids, and anthocyanin which play a significant

role in the bioreduction of metal ions into metal oxide nanoparticles
[20]. It has also been reported that Clitoria ternatea flowers contain high
amounts of anthocyanin pigments and rich in antioxidant properties as
compared to other medicinal flower elements. The flower extract
composition has significant impact on nanoparticle synthesis. The
flower exhibits antimicrobial, antiviral, anti-inflammatory properties,
used to treat diabetes, neurological disorders, protects the cardiovas­
cular system [21].
The novelty of the present study is the use of Clitoria ternatea flower
extract as a strong reducing and capping agent for NiO NPs synthesis.
The variation in the morphology of NiO NPs is attributed to the presence
of varying concentrations of secondary metabolites dissolved in the
plant extract [22]. Through an active phytochemical reaction, metal
ions become their metal oxides. Finally, the formation of oxygen results
in the linking of metal ions, and they take on a defined shape [23].
Further, the functional group of hydroxyl forms hydrogen bonds in the
solvent, which in turn reacts with metal ions coordination complexes as Fig. 1. UV–visible spectrum of Clitoria ternatea flower extract and NiO NPs; (in
ligands leading to formation of nanoparticles of different morphologies set). (a) Nickel acetate tetra hydrate solution, (b) Clitoria ternatea flower extract,
[12]. The annealing temperature also affects the morphology of the and (c) NiO NPs.
nanoparticles. A study has reported that NiO NPs of different mor­
phologies such as spherical, elliptical, sheet/snowflake like, rod-like, at 60 ◦ C for 6 h. Later, the dried powder was calcinated in a muffle
etc. can be obtained using solvents of different polarities. The polarity furnace for 2 h at 400 ◦ C. The resultant powder was used for further
of the solvent affects the nucleation resulting in different morphologies studies.
[24–25].
The present study focuses on the synthesis of NiO nanostructures 2.4. Antibacterial studies
using Clitoria ternatea flower extract. Photocatalytic efficiency and
antibacterial activity of the synthesized NiO nanostructures have been Well diffusion method was used to investigate the antibacterial ac­
incorporated. The degradation of the textile dyes Fast Green (FG) and tivity of green synthesized NiO NPs against pathogenic bacteria namely,
Rose Bengal (RB) has been chosen to demonstrate the catalytic potential E. coli and S. aureus [26]. The test organism was swabbed with sterile
of the green synthesized NiO nanostructures under sunlight. Addition­ cotton swabs on Muller Hinton agar plates and dried for 10 min before
ally, we evaluated the antibacterial activity of NiO NPs against gram- adding the samples. The wells on the agar plates were prepared by
negative bacteria such as E. coli and gram-positive bacteria such as punching with sterile micropipette tips and filling the wells with 50 μL of
S. aureus. There is no research report of Clitoria ternatea flower extract NiO NPs at various concentrations. These plates were incubated at 35 ◦ C
mediated green synthesis of NiO NPs and application in antibacterial for 24 h. The diameter of the zone of inhibition was used to calculate the
and photocatalytic activity to date. obstruction area of NiO NPs on bacteria growth.

2. Experimental
2.5. Photocatalytic degradation of FG and RB dye
2.1. Material
The photocatalytic dye degradation activity of the synthesized NiO
Fresh flowers of Clitoria ternatea (F) were collected from Coimbatore, NPs were performed under sunlight as described in the previous reports
India. Nickel acetate tetra hydrate (C4H6NiO4) with AR grade used for with minor modifications [27]. The green synthesized NiO NPs were
the synthesis was purchased from Merck. Throughout the green syn­ used as a photocatalyst for solar light induced dye degradation of FG and
thesis process, deionized (DI) water was used as solvent. RB dyes. In 25 mL of aqueous dye solution, 25 mg of NiO NPs was added
and irradiated with solar light for 150 min. 2 mL of reaction solution was
2.2. Preparation of Clitoria ternatea flower extract taken at appropriate time intervals of 30 min. The absorption intensity
of the collected solution was studied using UV–Visible spectrophotom­
The flower extract was prepared using the methods described in eter. The color of the solution disappeared showing that the catalyst was
earlier reports [26]. The fresh Clitoria ternatea flowers were washed with performing well. The photocatalytic experiment was performed during
DI water two times and allowed to dry in the open air to remove exterior the cloudless clear sky time between 11:30 am and 2:00 pm. The average
dust particles. The flower extract was prepared by adding 10 g of flowers sun light intensity observed during the photocatalytic study was around
in 100 mL DI water and boiling at 60 ◦ C for 1 h. The residues were fully 0.95 × 105 lx.
filtered by No.1 Whatman filter paper and kept in an airtight glass
container and maintained at 4 ◦ C to be used in the synthesis for NiO NPs. 2.6. Characterization techniques

2.3. Synthesis of NiO-NPs The UV–visible spectra were recorded by using Perkin Elmer Lambda
25 spectrometer (JASCO V650). The FTIR spectrum was recorded using
Green synthesis of the NiO NPs was carried out using the methods a Shimadzu IR Affinity 1S series spectrometer. The XRD pattern was
demonstrated in previous reports with minor modifications. For the obtained using Rigaku X-ray Diffractometer at room temperature using
synthesis of NiO NPs, 40 mL of 0.1 M C4H6NiO4 solution was added to Cu-Kα as radiation source (λ = 1.5406 Å) and scanning angle 2θ over the
10 mL of 10% flower extract of Clitoria ternatea. The flower extract acts range of 20 to 80◦ at room temperature. The size distribution and zeta
as a reducing and capping agent. The reaction solution color changed potential values were obtained using Dynamic Light Scattering (DLS)
from violet to bluish green indicating the formation of NiO NPs. The measurements carried out in a Zeta sizer Nano series (Malvern) instru­
obtained precipitate was subjected to centrifugation at 5000 rpm for 10 ment. TEM with EDX measurement was carried out using JEOL 2100
min, followed by washing with DI water to remove impurities and dried HRTEM.

2
S. Prabhu et al. Results in Chemistry 4 (2022) 100285

3000
8000 (c) 100

(200)
2500
80
6000
(αhν)2 (109nm eV)2

(111)
2000

Intensity (%)
Intensity

(220)
60
4000 1500
40

(311)
(222)
2000 1000
20

500
0 2.2 eV 0
10 20 30 40 50 60 70 80
1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5 6.0 6.5 2theta (degrees)

hν (eV) Fig. 4. XRD pattern of NiO NPs.

Fig. 2. Optical band gap diagram of NiO NPs.

Table 1
XRD peaks at various diffraction angles for NiO NPs.
NiO
S. No 2θ FWHM Miller Indices Particle size (D)
Flower extract
(Ni-O)

1 37.4 0.017444 111 9.75

2338 (C=C)

1635 (C=O)
(C-H)

2 43.5 0.034889 200 9.89

(C-O)
(OH)

3 63.08 0.052333 220 9.22

Transmittance (%)

486
2970

4 75.6 0.069778 311 6.28

3386

1111

601
547

formation of NiO NPs [26]. The FTIR spectra of the Clitoria ternatea
(C=C)

(C=O)

1126 (C-O)

flower extract and NiO NPs recorded over the range of 4000–400 cm− 1
(C-H)

(Fig. 3. The broad peaks found at 3317 and 3386 cm− 1 corresponding to
478
(OH)

2978

2360

the O–H stretching band from hydroxyl groups [8]. Absorption bands at
1635

2978 and 2970 cm− 1 are due to C–H stretching of the alkanes group
555

(b)
3317

[31]. The absorption bands at 2360 and 2368 cm− 1 belong to CO2 and
601

C–– C groups, respectively [5] and the strong absorption at 1635 cm− 1 is
4000 3500 3000 2500 2000 1500 1000 500 – O groups [29]. The peaks found at 1126 and 1111 cm− 1
assigned to C–
Wavenumber (cm-1) attributed to the C-O group indicate the presence of alcohols and car­
boxylic acid groups involved in the nanoparticle formation [28]. The
Fig. 3. FTIR spectra of NiO NPs and Clitoria ternatea flower extract.
peaks at 486 and 547 cm− 1 indicate the frequency of the Ni-O bond
bending vibrations. In general, the peaks between 400 and 600 cm− 1 are
3. Results and discussion characteristic of the metal–oxygen bond [4]. In addition to the similar
peak profiles and predominant shifting in peaks of the Clitoria ternatea
3.1. UV–visible study flower extract and the NiO NPs result justifies that the biomolecules are
responsible for the capping and reduction process [32–33]. Phenols,
The Clitoria ternatea flower extract was added to the aqueous solution Alcohol, carboxylic acids and alkanes were noticed in this study,
of C4H6NiO4 which changes color from green to bluish green showing whereas phenols and alcohols were found to bind with the NiO NPs more
the formation of NiO NPs (Fig. 1 inset). The presence of active secondary effectively [31]. We found that the FTIR spectrum analysis was consis­
metabolites in the flower extract reduces nickel ions in the solution to tent with the results obtained by XRD and that no other impurity phases
NiO. The flower extract acts as both capping and reducing agent. This were observed.
was confirmed by the UV–visible spectrum given in Fig. 1. The
maximum absorption spectra peak (λ) was found at 283 nm which is
specific for NiO NPs [5]. The Clitoria ternatea flower extract shows two 3.3. X-ray diffraction study
absorbance peaks at 575 and 620 nm confirming the presence of
anthocyanin [28]. The direct optical band gap (Eg) for NiO NPs was Fig. 4 shows the X-ray diffraction pattern with intense and sharp
calculated using Tauc’s plot approach and was found to be 2.24 eV peaks which demonstrate that the synthesized NiO NPs are pure and
(Fig. 2 and the obtained band gap values confirm that NiO NPs are semi- crystalline in nature. The XRD peaks observed at 2θ angles of 37.40,
conductors. The estimated Eg value of bare NiO NPs should be in the 43.50, 63.080, and 75.60 representing (1 1 1), (2 0 0), (2 2 0) and (3 1 1)
range of 3.6–4.0 eV but decreased bandgap values would be due to the planes, respectively, of the cubic crystal structure matched with JCPDS
chemical defects or vacancies present in the crystal structure and NO 47–1049 (Table 1 [34]. The crystallite size of the NiO NPs was
annealing temperature are resulting to reduce the band gap energy [29]. calculated using the Debye-Scherrer formula,
These results are in close agreement with earlier reports [30]. 0.89λ
d=
βcosθ
3.2. FTIR spectra study
where, λ, β and θ are the X-ray wavelength, full width at half maximum
FTIR spectroscopy was used to analyze the various functional groups (FWHM) of XRD diffraction peaks, and angle of Bragg diffraction,
present in the Clitoria ternatea flower extract and to confirm the respectively [26]. The average crystalline size of the NiO NPs was

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S. Prabhu et al. Results in Chemistry 4 (2022) 100285

Fig. 5. (a) Size distribution and (b) Zeta potential of NiO NPs.

Fig. 6. HRTEM images of NiO NPs at different magnifications; (a) 100 nm, (b) 10 nm, (c) 5 nm, and (d) SAED pattern.

calculated to be 9.62 nm. The calculated average crystallite size of the were quantified by Dynamic Light Scattering (DLS) analysis (Fig. 5a and
CuO NPs was in concordance with the previous findings [35]. The 5b. The NiO NPs (0.001 mg) were dispersed in 5 mL of DI water to find
various phytochemicals present in the aqueous flower extract of Clitoria the average particle size. The average particle size obtained from
ternatea acting as a reducing and capping agent are primarily responsible colloidal solution mixture was found to be 119 nm with a polydispersity
for smaller crystallite size of NiO NPs [34]. The synthesized NiO NPs index (PDI) of 1.00 and zeta deviation resulted in − 17.4 mV. Generally,
with high crystallinity corresponding with reduced particle size and PDI values > 0.7 were observed in samples that had increased size dis­
high surface area would give effective antibacterial and catalytic tributions [35]. The negative zeta potential and large particle size of NiO
activities. NPs confirms that the active biomolecules such as flavonoids, anthocy­
anin, and phenols present in the Clitoria ternatea flower extract cover the
surface of the NPs, which established electrostatic repulsive force be­
3.4. Particle size and zeta potential analysis tween the particles providing moderate stability for the green synthe­
sized NiO NPs. It has been reported that the particle size and negative
The nano size particle distribution and surface charge of NiO NPs

4
S. Prabhu et al. Results in Chemistry 4 (2022) 100285

of 10 nm, it can be observed that the NPs are hexagonal in shape and
non-agglomerated. The d-spacing between the planes was found to be
8 0.218 nm corresponding to (2 0 0) plane of NiO (Fig. 5c. This result
confirmed the synthesized NPs were pure crystalline in nature. The
Selected Area Electron Diffraction (SAED) pattern of the synthesized
6 NiO NPs in Fig. 5d shows diffraction rings with concentric circles. These
Frequency

rings correspond to the different diffraction planes namely (1 1 1),

(2 0 0), (2 2 0), (3 1 1), and (2 2 2) planes, which can be indexed to the
4
cubic crystal structure that matches with the XRD data [37]. The average
particle size calculated from the size distribution histogram (Fig. 7 was
2
found to be 13 nm, which is in concordance with XRD results. The EDAX
analysis that confirms the presence of nickel and oxygen elements
(Fig. 8. The as-synthesized NiO NPs are composed of 80.69% of Ni and
0 1.34% of oxygen. In addition, the atomic % of Ni was found to be 78.93
6 8 10 12 14 16 18 20
and oxygen 4.84. There is no affirmation for the appearance of impu­
Size (nm) rities. The decrease in particle size (13 nm) leads to an increase in the
surface area and this enhances the photocatalytic and antibacterial ap­
Fig. 7. Particle size distribution of NiO NPs.
plications [16].

charge distribution of the synthesized NPs enhanced the photocatalytic

activity and antibacterial activity of the NiO NPs [36]. 3.6. Antibacterial activity

3.5. HRTEM study Using the well diffusion method, the antibacterial activity of NiO NPs
was investigated upon E. coli and S. aureus bacteria [26]. NiO NPs
The HRTEM micrographs of the NiO NPs at two different magnifi­ concentration was varied as 50, 100 and 200 μg/mL and the corre­
cations (100 and 10 nm) are shown in Fig. 6a and 6b. At a magnification sponding inhibition zones are shown in Fig. 9a and 9b. The antibacterial

Fig. 8. EDAX spectrum of NiO NPs.

Fig. 9. Antibacterial activity of NiO NPs against (a) S. aureus, (b) E. coli, and (c) zone of inhibition.

5
S. Prabhu et al. Results in Chemistry 4 (2022) 100285

Table 2 adsorption of phytochemical compounds of the Clitoria ternatea flower

Zone of inhibition of S. aureus and E. coli. extract on the surface of NiO NPs, which also functioned together with
Microorganism Average zone of inhibition in (mm) nanoparticles to prevent the bacteria’s growth. Hence, it is believed that
the synthesized NiO NPs with reduced particle size (13 nm) and high
50 μg/mL 100 μg/mL 200 μg/mL
surface area will support enhanced antibacterial activity [8].
S. aureus 13 ± 1.0 16 ± 1.0 19 ± 1.0
E. coli 15 ± 1.0 17 ± 1.0 22 ± 1.0

3.7. Dye degradation studies

activity was observed to increase as the concentration of NiO NPs was
increased, (Fig. 9c, Table 2. For E. coli, the highest inhibition zone was Degradation of environmental pollutants like organic dye molecules
found to be 22 mm while for S. aureus was 19 mm at 200 μg/mL. The using active catalyst under solar light is one of the main objectives of this
surface of NiO NPs carries a positive charge while the bacterial cell wall study. Fig. 10a and 10b shows the photocatalytic degradation process of
has a negative charge. Consequently, it stimulates an electromagnetic FG and RB dyes by NiO NPs. When irradiating the FG and RB dye so­
interaction resulting in the destruction of the bacterial wall as well as lutions, the concentration was visibly reduced and it was confirmed
cytoplasm [8]. Further, NiO NPs produce reactive oxygen species (ROS) from reduction in the UV absorption intensity. The efficiency of degra­
that leads to protein oxidation and DNA destruction which kills the dation is determined by the Beer-Lamberts equation,
bacteria [34]. NiO NPs antibacterial activity is attributed to a combined Co− C
effect of the size of the particle, surface area, ROS and the concentration Degradation efficiency(%) =
C
× 100
of NPs. Interestingly, the NiO NPs exhibit significant inhibition activity
against both bacterial strains but best against gram-negative E. coli where the variables C0 and C denote as the initial and final concentra­
bacteria. A similar more antibacterial activities of NiO NPs against tions, respectively [27]. The degradation percentage of FG and RB has
Gram-positive than Gram-negative bacteria was also reported by earlier been valued to be around 89% and 77%, respectively.
studies [34,37]. The Clitoria ternatea flower extract treated NiO NPs Comparable results of the degradation efficiency against RB and FG
exhibited a significant antibacterial activity compared to previously dyes with different metal oxide nanoparticles are given in Table 3. The
reported against S. aureus, and E. coli [34,37]. It could be due to the photocatalytic degradation mechanism of NiO NPs against RB and FG
dyes under sunlight irradiation is shown in Fig. 11. The dye degradation

Fig. 10. Photodegradation (a) FG, (b) RB dye, (c) C/C0 vs Time interval graph of degradation, (d) –In(C/C0) vs Time interval graph of degradation.

Table 3
Comparison of photocatalytic activity for various photocatalyst and its reported values against FG and RB dyes.
S. No Photocatalyst Synthesis method Average Crystallite Size (nm) Dye % Of degradation References

1 ZnO NPs Green synthesis 15–25 RB 85 [1]

2 CuO NPs Green synthesis 34 RB 94.5 [2]
3 Ƴ Fe2O3 NPs Thermal decomposition 35–40 RB 96 [3]
4 NiO NPs Green synthesis 9.6 RB 88.8 Present Work
5 NiO NPs Green synthesis 9.6 FG 76.64 Present Work

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S. Prabhu et al. Results in Chemistry 4 (2022) 100285

degradation rates were found to be 0.032 and 0.022 min− 1 for NiO NPs
against FG and RB dye solutions, respectively. The values of k and R2 are
given in Table 4. The value of k corresponds to the slope of the line and it
is high and relates with rate of reaction along with the concentration of
the reaction components. The concentration of the dye, amount of the
catalyst, irradiation time along with the intensity of incident light affect
the process of dye degradation. The reaction rate slow down as the
irradiation time increases along with intermediate products, and re­
actants play an important role during the degradation [38]. Moreover,
the various phytochemicals present in plant extract and even in the
nanoparticles surface acts like catalyst to increase the photocatalytic
activity and the following improved degradation of dye molecules [34].

4. Conclusion

Herein, the NiO NPs were successfully synthesized by using Clitoria

ternatea flower extract is reported for the first time. The optical band gap
was found to be 2.2 eV and thus NiO is a p-type semiconductor which
can be utilized as a significant photocatalyst and strong antibacterial
agent. The FTIR results proved that the flower extract has an excellent
potential as stabilizing and capping agent in the formation of NiO NPs.
The morphology studies confirmed the cubic crystal structure and the
DLS studies revealed the size and charge distribution of the NiO NPs. The
HRTEM analysis reveal the hexagonal shaped particles with average size
of 13 nm. The antibacterial activity results divulge that the NiO NPs
have good potential to inhibit the growth of S. aureus and E. coli bacteria.
Fig. 11. Photocatalytic degradation mechanism of dye molecule by NiO NPs In addition, NiO NPs was found to be a promising photocatalyst for the
under sunlight. degradation of FG and RB dyes under sunlight without using other en­
ergy sources. Based on the obtained results, we conclude that the green
synthesized NiO NPs could have potential applications in cost-effective
Table 4 industrial waste water treatment and bacteria-based biological
Degradation efficiency, linear correlation coefficients (R2), and degradation rate applications.
constant (k) of FG and RB dyes.
S. Name of the Band gap Degradation efficiency k R2 Declaration of Competing Interest
No dye Solution (eV) (%) (150 min) (min− 1)

1 Fast green (20 2.24 88.8 0.032 0. The authors declare that they have no known competing financial
ppm) 89 interests or personal relationships that could have appeared to influence
2 Rose Bengal 2.24 76.64 0.022 0.93 the work reported in this paper.
(20 ppm)

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