Effective Parameters in the Green Synthesis of Zero-valent Iron Nanoparticles as a Fenton-like Catalyst

Journal of Environmental Treatment Techniques

2020, Volume 8, Issue 1, Pages: 442-447

J. Environ. Treat. Tech.

ISSN: 2309-1185

Journal web link: http://www.jett.dormaj.com

Effective Parameters in the Green Synthesis of

Zero-valent Iron Nanoparticles as a Fenton-like

Catalyst

Seyedeh-Masoumeh Taghizadeh , Alireza Zare-Hoseinabadi , Aydin Berenjian , Younes

4,1,*

Ghasemi , and Alireza Ebrahiminezhad

Department of Pharmaceutical Biotechnology, School of Pharmacy and Pharmaceutical Sciences Research Center, Shiraz University of Medical

Sciences, Shiraz, Iran

Department of Medical Biotechnology, School of Medicine and Non-Communicable Diseases Research Centre, Fasa University of Medical

Sciences, Fasa, Iran

School of Engineering, Faculty of Science and Engineering, The University of Waikato, Hamilton, New Zealand

Department of Medical Nanotechnology, School of Advanced Medical Sciences and Technologies, Shiraz University of Medical Sciences,

Shiraz, Iran

Received: 03/09/2019

Accepted: 21/01/2020

Published: 20/02/2020

Abstract

Nowadays, utilization of new and sustainable techniques for the synthesis of INPs are increasing significantly. Common nettle

(

Urtica dioica) or stinging nettle is an herbaceous perennial flowering plant with a significant reduction potential. Leaf extract of

this plant can reduce iron ions to zero-valent INPs. Like all chemical and biochemical reactions, reaction conditions have an

immense effect on the process. In this experiment, impacts of different process parameters such as FeCl concentration, leaf extract

quantity, reaction temperature, and reaction time on the biosynthesis of INPs were evaluated. FeCl concentration, quantity of the

leaf extract, and reaction temperature were identified as effective factors. But, reaction time has not any significant impact. INPs

were characterized by transmission electron microscope (TEM) Fourier transform infrared (FTIR) spectroscopy, and X-ray

diffractometer. Particles were spherical and oval in shape and were measured to be 21–71 nm with mean particle size of 46 nm.

TEM illustrated that the zero-valent iron (ZVI) nanoparticles had mostly a spherical shape with 46 nm of mean diameter. The

particles tend to form irregular clusters with phytochemicals from leaf extract and form macrostructures in the range of 117–605

nm. Prepared nanostructures are a promising catalyst in Fenton reactions and can degrade Methyl orange dye with about 70 percent

efficiency in 5 h.

Keywords: Bioreduction; Biosynthesis; Common nettle; Fe nanoparticles; Plant mediated synthesis; Urtica dioica

Introduction¹

biology and biotechnology (6, 7), ferrofluids (8), and

medicine (9). Hence, production of INPs with varius

characteristics has attracted great attention (4, 10-12).

Chemical synthesis is one of the primary techniques for

synthesis of INPs. Since now chemical approaches has been

developed significantly and is able to produce pure INPs

with desirable characteristics. But, these techniques usually

employ organic solvents and noxious chemicals and also are

conducted at harsh conditions (13-17).

Biosynthesis has emerged as a potential sustainable

alternative for chemical procedures (18-21). Biological

compounds such as flavonoids, polyphenol, alkaloides,

carbohydrates, proteins and other biomolecules can act as

Nanoparticles retained so much attention in various

fields of sciences and technologies due to thire unique

physicochemical and biological properties (1, 2). Among all

nanoparticles, metal nanoparticles are employed in a variety

of fields such as environmental remediations, bioprocess

intensification, bioseparation, pharmaceotical sciences, and

nanomedicine (3). Among these nanoparticles, there has

been special focus on iron nanoparticles (INPs) due to their

unique properties such as ease of synthesis and

functionalization, magnetic behavior, catalytic activity, and

biocompatibility (4). These nanoparticles are now being

applied in various fields including bioremediation (5),

Corresponding author: Alireza Ebrahiminezhad, (a) Department of Pharmaceutical Biotechnology, School of Pharmacy and

Pharmaceutical Sciences Research Center, Shiraz University of Medical Sciences, Shiraz, Iran; (b) Department of Medical

Nanotechnology, School of Advanced Medical Sciences and Technologies, Shiraz University of Medical Sciences, Shiraz, Iran. E-

mail: a_ebrahimi@sums.ac.ir.

Journal of Environmental Treatment Techniques

2020, Volume 8, Issue 1, Pages: 442-447

green bioreductant for synthesis of metal nanoparticles (22-

weighed to calculate the concentration of prepared

nanoparticles.

7). These metabolites can also act as stabilizing agent to

prevent agglomeration and improve dispersity and reactivity

of nanoparticles (19, 21, 28, 29). Biosynthesis reactions have

been conducted by using bioactive compounds from various

organisms such as microbial cells, algae, and plants (19-21,

2.4 Characterization of INPs

Visual appearance and morphological characteristics of

INPs were investigated by Philips CM 10, TEM, operated at

high voltage (HT) 100 kV. The INPs was diluted in millipore

water and dried on a carbon-coated copper grid.

0). But, use of plant extracts for synthesis of INPs have

considerable advantages over other organisms such as

cheapness, availability, safety, less recovery steps and

elimination of elaborate cell culture process (18, 31).

Recent studies have shown that INPs can be synthesized

by using extracts of eucalyptus, Castanea sativa, some kinds

of tea, mulberry, pomegranate, peach, pear, vine, coffee,

Salvia officinalis, Caricaya papaya, tangerine, sorghum, and

so many other plants (4). But, like all chemical and

biochemical reactions, biosynthesis of INPs can be affected

by reaction conditions (4, 30, 32). So, there is an increasing

demand for investigations toward recovery of the effective

factors in the biosynthesis of INPs.

Urtica dioica, also known as common nettle or stinging

nettle, is an herbaceous perennial flowering plant in the

family Urticaceae. U. dioica leaf extract contains various

phytochemicals such as phenolic and flavonoid compounds,

and has a long history of application in medicine and

fooditarian (11, 33). In our previous study we have shown

that leaf extract of this plant is so bioactive which can reduce

ferric ion to zero-valent iron nanoparticles (11). In this study,

Figure 1: Effects of FeCl concentration on the formation of INPs

Particles size distribution was obtained using Image J

software version 1.47v, an image analysis software

developed by the NIH. Main functionality of the INPs

were evaluated by Fourier transform infrared (FTIR)

spectroscopy using a Bruker, Vertex 70, FTIR spectrometer,

and the standard KBr pellet method in the range between 400

the effect of different process parameters such as FeCl

concentration, leaf extract quantity, reaction temperature,

and reaction time on the biosynthesis of INPs by using

Urtica dioica leaf extract were evaluated.

-1

cm and 4000 cm . Phase composition and crystallinity of

INPs were obtained using Siemens D5000 x-ray powder

diffractometer instrument. X-ray diffraction (XRD) pattern

of INPs samples were scanned within the 2θ range from 20°

to 90° and results were evaluated by PANalytical X’Pert

HighScore software.

Materials and methods

.1 Materials

3 2

Ferric chloride (FeCl .6H O, analytical grade), was

purchased from Merck Chemicals (Darmstadt, Hessen,

Germany) and used without any further treatment. Dried

leaves of U. dioica were purchased from a local shop (Fasa,

Fars, Iran). Millipore water (Millipore Corp., Bedford, MA,

USA) was used throughout the whole experiment.

.5 Fenton catalytic activity of INPs

Potential of the prepared nanostructures as a Fenton

catalyst was evaluated in a dye degradation reaction. The

experiment was performed at room condition in single-use

0 mL reaction volume (12). In brief, zero-valent iron

.2 Leaf extract preparation

Dried leaves were washed to remove any impurities and

nanoparticles (1 mg/mL final concentration in reaction) were

mixed with Methyl orange dye solution (20 mg/L)

were dried at room temperature. In order to prepare aqueous

extract of nettle leaves, deionized water was added to the

crushed leaves at a ratio of 1:20 (w/v) and boiled under

reflux for 15 min by using a heater mantel (11). The prepared

mixture was cooled to room temperature and filtered through

a Whatman filter paper (Reeve angel, Grade 201). The

filtrate was centrifuged for 5 min at 2000 rpm to eliminate

plant microparticles. The obtained clear solution was stored

in the refrigerator and was used as leaf extract.

2 2

containing one per cent H O . The reaction was followed for

h and rate of dye degradation was calculated against methyl

orange standard curve at 465 nm (Hitachi U-0080D UV–vis

spectrophotometer, Tokyo Japan). A solution of Methyl

orange and hydrogen peroxide without any nanostructure

was set as the control.

3 Results and discussion

.1 Effective factors in the biosynthesis reaction

.3 Effective factors in the biosynthesis reaction

The effects of iron precursor concentration on the

The effects of several parameters (i.e. FeCl

biosynthesis of INPs were evaluated from 2.5 mM to 320

mM ferric chloride and results were shown in Figure 1.

Increase in the concentration of iron precursor up to 20 mM

resulted to increase in the amount of prepared INPs. But,

interestingly, more increase in the iron precursor has shown

negative effect on the formation of nanoparticles. Similar

effects were also reported for the green synthesis of silver

concentration, leaf extract quantity, reaction temperature,

and reaction time) on the biosynthesis of INPs were

evaluated by one factor at a time approach. After completion

of each reaction, the prepared INPs were harvested by

centrifugation and washed with deionized water. The

resulting black pellets were oven-dried at 50 °C for 48 h and

Journal of Environmental Treatment Techniques

2020, Volume 8, Issue 1, Pages: 442-447

nanoparticles (AgNPs). It is shown that high concentrations

of silver precursor (AgNO ) have an immense negative

effect on the biosynthesis of AgNPs (25, 26).

process and industrial production. Similar results were also

reported for biosynthesis of INPs by using Amaranthus

dubius leaf extract (32).

Figure 2: Effects of leaf extract quantity on the formation of INPs

Figure 3: Effects of reaction temperature on the formation of INPs

Figure 5: TEM image of the biosynthesized INPs and

corresponding particle size distribution pattern

It has been shown that at increased temperatures up to

7°C leaf extract has a higher reduction potential for INPs

biosynthesis. But, increase in the reaction temperature above

7°C have a negative effect on the formation of INPs (32).

Negative effects of increased temperatures on the

bioreduction of metallic ions were also reported for Zataria

multiflora leaf extract, Alcea rosea flower extract and

Ephedra intermedia stem extract (25, 26, 30). These results

indicate that increase in the reaction temperature can degrade

plants antioxidant compounds and decreases amount of the

reduced nanoparticles (34-37).

Effect of reaction time on the production of INPs was

evaluated from 12 h to 48 h. As depicted in Figure 4, increase

in the reaction time has no effect on the amount of

synthesized nanoparticles. To provide enough time for

biological interactions time periods below 12 h are not

commonly set for biosynthesis reactions (11, 12, 19, 25, 26,

29, 30). But, there is a report that 90 min is sufficient to

achieve highest efficiency in production of INPs by using

Amaranthus dubius leaf extract (32). Reaction times more

than 48 h usually associated with the waste of time,

aggregation of nanoparticles, and production of multi-

Figure 4: Effects of reaction times on the formation of INPs

The effect of U. dioica leaf extract quantity on the

production of INPs is shown in Figure 2. The pattern

revealed that amount of U. dioica leaf extract plays a

significant role in the production of INPs and increase in the

leaf extract quantity resulted in more nanoparticle

production. Increase in the amount of leaf extract is equal to

increase in the concentrations of bioactive compounds in the

reaction and therefor more reduction of iron ions. The effect

of reaction temperature was evaluated from 25°C to 75°C.

Fortunately, room temperature (25°C) was identified as the

best temperature for the bioreduction of iron ions (Figure 3).

This means that bioreduction of iron ions by using U. dioica

leaf extract is an economic reaction from energy

consumption point of view which is so critical for scaling up

Journal of Environmental Treatment Techniques

2020, Volume 8, Issue 1, Pages: 442-447

shaped nanoparticles (38-41).

extracts (47), Sorghum bran extracts (59), Terminalia

chebula (60), and Syzygium jambos L. (61).

.2 Characterization of INPs

TEM micrograph of the INPs is provided in Figure 5.

.3 Fenton catalytic activity of INPs

Catalytic activity of the prepared nanostructures over 6

h is presented in Figure 8. More than 60 percent reduction in

the dye concentration was achieved after 4 h reaction. After

h reaction about 70 percent of Methyl orange was degraded

and no more significant dye degradation was recorded after

this time. Similar time dependent degradation pattern was

also reported for other iron nanostructures which synthesized

by using green tea leaves extract and leafy branches extract

of Mediterranean cypress (Cupressus sempervirens) (12,

2).

Figure 6: FTIR spectrum of the biosynthesized INPs

Figure 7: XRD pattern of the biosynthesized INPs

Figure 8: Methyl orange degradation pattern using INPs as Fenton

catalyst

Produced nanoparticles were spherical and oval in shape

and were measured to be 21–71 nm with mean particle size

of 46 nm. INPs with similar particles size distribution were

also obtained by using Eucalyptus tereticornis, Melaleuca

nesophila (42), Zataria multiflora (26), pistachio (43), green

tea (44-46), and eucalyptus (47). TEM investigations

revealed that prepared particles tend to form irregular

clusters with phytochemicals from leaf extract and form

macrostructures in the range of 117–605 nm. This finding

shows the role of plant organic compounds as reducing and

stabilizing agent in the synthesis of INPs (43, 48).

Conclusions

U. dioica is a potential plant for biosynthesis of metallic

nanoparticles. Nettle leaf extract has a significant reduction

potential which can reduce ferric ions to zero-valent INPs in

a sustainable, economic, and facile manner. Also, it has

phytochemicals which can act as biological stabilizer.

Reaction conditions (i.e. ferric ion concentration, leaf extract

quantity, and reaction temperature) have a significant impact

on the bioreduction of INPs. Bioactive compounds in the

plants are sensitive to increased temperatures and usually

bioreduction reactions are well done at room temperatures.

Also, high concentrations of metal precursor can prevent

formation of nanoparticles. Increase in the plant extract

quantity resulted in the increase in the formation of

nanoparticles which is due to increase of bioactive

compounds in the reaction. About 12 h of reaction time can

be sufficient for the formation of INPs and biological

interactions. The particles were efficient as a Fenton-like

catalyst and can be developed for technical applications in

future.

Fourier transformed infrared (FTIR) spectroscopy was

used as non-destructive way to identify and study the

interactions of biological compound with INPs (Figure 6)

(

49). The broad peak at 3467 cm is corresponding to the O–

H stretching vibrations (50, 51). The characteristic peak of

C–O bond and carbonyl group can be seen at 1070 cm and

-1

636 cm , respectively. These functionalities can be

corresponding to heterocyclic compounds from proteins and

unsaturated hydrocarbons (52-54). Absorption peaks from

aliphatic C–H groups are recorded at about 2800 cm (55,

6). Fe–O stretching vibration resulted in two characteristic

-1

peaks at about 640 cm and 450 cm . Absence of these

peaks in the recorded pattern is an indicative feature for zero-

valent INPs (57, 58).

Aknowledgment

This experiment was financially supported by Fasa

University of Medical Sciences under the grant No. 95095.

XRD pattern of the synthesized INPs is shown in Figure

. Lack of distinct diffraction peak shows amorphous nature

Ethical issue

of prepared nanoparticles. The broad hump peak in the

region 10-20 of 2θ degrees is corresponding to organic

compounds on the surface of INPs. Similar results were also

reported for INPs which synthetized by using eucalyptus leaf

Authors are aware of, and comply with, best practice in

publication ethics specifically with regard to authorship

(avoidance of guest authorship), dual submission,

manipulation of figures, competing interests and compliance

Journal of Environmental Treatment Techniques

2020, Volume 8, Issue 1, Pages: 442-447

with policies on research ethics. Authors adhere to

publication requirements that submitted work is original and

has not been published elsewhere in any language.

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Competing interests

The authors declare that there is no conflict of interest

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