Gentamicin-assisted selenium nanoparticles synthesized under gamma irradiation for

controlling the spread of resistant urinary tract infection-causing pathogens

Abstract
• The purpose of this research is to compare and enhance the antimicrobial and antibiofilm potentials of
the biogenic Selenium nanoparticles (Se NPs) produced by cost-effective and eco-friendly green methods.
The synthesis of Se NPs is described in this manuscript by two different methods; a biogenic process
using Penicillium chrysogenum’s filtrate and by utilizing Gentamicin drug (CN) following the application
of gamma irradiation.
• Se NPs were characterized by UV-Vis., HRTM, FTIR, XRD, DLS, SEM and EDX mapping technique.
Antimicrobial and antibiofilm activities of the synthesized Se NPs were investigated against multidrug-
resistant (MDR) bacteria and yeast causing severe diseases such as Urinary Tract Infection (UTI).
• The biogenic Se NPs exhibited an absorption peak at 435.0 nm while Se NPs-CN showed an absorption
peak at 350.0 nm which is related to the Surface Plasmon Resonance (SPR).
• Data obtained from HRTEM, SEM/mapping and XRD analysis confirmed the mono-dispersion and
crystalline nature of the prepared samples with an average diameter of 33.84 nm and 22.37 nm for the
mycogenic Se NPs and Se NPs-CN, respectively.
• The synthesized Se NPs-CN possesses an encouraging antimicrobial potential with respect to the biogenic
Se NPs against all examined UTI-causing microbes. Remarkably, Se NPs-CN showed antimicrobial
potential toward Candida albicans with a Zone of Inhibition (ZOI) recorded at 26.0 mm, 23.0 mm ZOI
for Escherichia coli and 20.0 mm ZOI against Staphylococcus aureus.
• In addition, the incorporated Se NPs-CN displayed an enhanced percentage of biofilm inhibition of
88.67%, 87.93% and 85.20% against S. aureus, P. aeruginosa and E. coli, respectively. Accordingly, the
novelty of the present research involves the green synthesis of mono-dispersed Se NPs and combining
the synergistic potential of CN with Se NPs for potential biomedical, pharmaceutical and therapeutic
applications especially in the treatment of UTI
Key words: Selenium nanoparticles; Gentamicin; Gamma irradiation; E. coli; Penicillium chrysogenum
and Urinary Tract Pathogens
Introduction
Nanotechnology is supposed to be the subsequent industrial revolution and is considered to have a
tremendous influence on community, economics and the common world [1-3]. This field possesses a great
impact on various perspectives including industry, biomedicine, infection control, agriculture, wastewater
treatment, drug and gene control and nano-biotechnology [1, 2, 4-9].

Ass. Prof. Dr. Gharieb S. El-Sayyad, Egyptian Atomic Energy Authority 1

The physical methods used for the synthesis of Se NPs varied, including UV radiation, laser ablation,
gamma rays and the hydrothermal method [2, 8, 10-14].
Gamma irradiation was confirmed to be a simple yet effective method for the synthesis of different NPs
at room temperature (without any pressure) and in the aqueous solution (as a source of water) to produce
solvated electrons and free radicals (by the aid of water radiolysis) as a reducing agents for NPs synthesis
[2, 15-19].
On the other hand, the chemical methods used for the preparation of NPs included the precipitation
technique, the acid breakdown, and the catalytic reduction utilizing glucose, ascorbic acid, sodium
dodecyl sulfate and sulfur dioxide [2, 20-22].
Chemical methods still need higher temperature and pH adjustment to acidic conditions beside the
hazardous chemicals [10, 23, 24], which may lead to the synthesis dangerous NPs not applicable for the
biomedical treatments [2].
On the contrary, the green biosynthesis of Se NPs is harmless and cost-effective, utilizing eco-friendly
and non-toxic substances [5, 18, 25-30]. Moreover, the resulted Se NPs synthesized by the green
biosynthesis are constant and stable for a long time, because of the natural capping agent like the organic
molecules and amino acids which prevent its precipitation. In addition, it needs no external additives as
stabilizing agents like the methods of chemical synthesis [30-32].
A vast development in the microbial infection was noticeable particularly due to the increased microbial
resistance to the synthetic antibiotics like CN [1, 2, 33]. Staphylococcus aureus and Escherichia coli are
definite classes of bacteria observed in the intestinal area of all humans and many animals. E. coli is
creating an enteric disease and additional infection includes intestinal dysfunction and UTI [34].
The bacterial biofilm is composed of multi-cellular polysaccharides present at the surfaces [35, 36]. The
pathogenic microbes initiate biofilm formation in the wet location due to the adequate course of nutrients
and the obtained surface connections [36, 37].
Biofilms are used to protect the microbial population from the environmental pressure [36]. Therefore,
the maturation of the bacterial biofilm allows the resistance to different bactericidal, the host protection
responses, bacteriophages and certain drugs, by the creation of extracellular polysaccharide substances
acts as a shielding block [35, 36].
UTI is considered a severe and common health difficulty [36]. It is the second abundant general bacterial
disease, after the respiratory tract disease [38]. This kind of disease can lead to a high danger of morbidity,
death and raised healthcare payments. Although many pathogenic microbes can cause UTI, Escherichia
coli is responsible for the strong preponderance of UTI problems (about 80.0% of UTI records) [36, 39].

Ass. Prof. Dr. Gharieb S. El-Sayyad, Egyptian Atomic Energy Authority 2

There are some difficulties after UTI treatment due to the administrated drugs which lead to a
modification in the abdomen micro-biome (commensal microbiota) and eventually the generation of
multidrug-resistant (MDR) microbes [40].
There are some significant attempts to improve a distinct antimicrobial agents by choosing native
products, changing some structure of antibiotics by increasing its activity through the substitution and
finally by applied nanotechnology [2, 41, 42]. Some of the produced NPs may be serve as a protection
agents against MDR microbes causing UTI [36]. The antimicrobial efficiency is due to their higher surface
area allowing a fine synergy resulting from multivalent attractions [43].
A great strategy to improve the potency of different NPs toward microbial population is to incorporate
them among certain ineffective antibiotics. Incorporation of NPs with antimicrobials is considered a
revolutionary way to target MDR-bacteria causing UTI and increasing popular therapeutic approaches.
Our research proposed synthesis of Se NPs by two different methods; a biogenic synthesis by P.
chrysogenum filtrate and green synthesis by the incorporation with CN under the effect of gamma
irradiation. Full validation and characterization were performed to obtain confirmatory data about the size,
distribution, purity, crystallinity and the major appearance of the synthesized samples. Additionally,
comparisons of the antimicrobial and anti-biofilm activities of the synthesized Se NPs and Se NPs-CN
against pathogenic yeast and bacteria that induce UTI were estimated.
Materials and methods
Chemicals and Reagents
Media ingredients and components were purchased from Hi-Media and Difco. Chemicals such as sodium
selenite; Na2SeO4, Gentamycin (CN) and isopropyl alcohol and other reagents (used in the following
examinations) were obtained at the analytical standard grade from Sigma-Aldrich.
Preparation of cell-free filtrates and fungal strains
In the present screening assay ten fungal isolates were tested for their capabilities towards the biosynthesis
of Se NPs. They named Penicillium citrinum, Penicillium chrysogenum, Penicillium digitatum,
Penicillium expansum, Aspergillus fumigatus, Aspergillus niger, Aspergillus oryzea, Aspergillus flavus,
Apergillus terrus and Monascus purpureus. They were obtained from the culture collection of Drug
Microbiology Lab., NCRRT, Cairo, Egypt. Sterile potato dextrose broth (PDB) containing 24.0 g/l (Hi-
Media) was inoculated with the tested fungal strains and incubated with continuous shaking (LAB-Line
R Orbit Environ, USA; 200.0 rpm) at 30.0 ± 0.5 ˚C for 14.0 days and pH was adjusted at 5.50 ± 0.5.
After that, the fungal cultures were filtrated using Whatman filter paper No.1 and centrifuged through
cooling centrifuge (Hettich Universal 16 R, Germany) for 30.0 min. at 5.0 ± 1.0 ˚C. The cell-free

Ass. Prof. Dr. Gharieb S. El-Sayyad, Egyptian Atomic Energy Authority 3

supernatants (CFS) of all the tested fungal strains were isolated from the solid cells and were used for the
synthesis of biogenic Se NPs.
Gamma irradiation
60
Gamma irradiation method was carried out at NCRRT, Cairo, Egypt. The irradiation source was Co-
Gamma chamber 4000-A-India. The dose rate was determined at 2.10kGy/h. Gamma irradiation exhibits
solvated electrons and a hydrated and free radical through the water radiolysis.
Green synthesis of Se NPs
Synthesis and optimization of Se NPs by CN and gamma rays
Se NPs were synthesized by CN as a stabilizing agent and gamma ray as reducing agent. Sodium selenite
solution (at different conc.) was mixed with different concentrations of CN solution through a ratio 1:1
(v/v) and 0.2% isopropyl alcohol. The resultant mixtures were stirred at room temperature and displayed
to various gamma ray’s doses (5.0, 10.0, 15.0, 20.0, 25.0, 30.0 kGy). The optimum gamma dose was
estimated after established the O.D. Additionally, the stability of CN was examined after the exposure to
the screened gamma ray’s doses.
An experimental investigation was conducted to estimate the impact sodium selenite and CN
concentrations regarding Se NPs synthesis after displayed to the most potent gamma ray. The considered
factorial study was included two variables (the concentration of sodium selenite and CN) in 12 levels
(Table 2). The main idea for factors chosen in the present research that, they provide multiple important
impact towards Se NPs biosynthesis [44].
Sodium selenite solution (at various conc.; Table 1) was mixed with different concentrations of CN
solution and 0.2% isopropyl alcohol. Mixtures were stirred at room temperature (24.0 ± 1.0 oC) and
exposed to the most effective gamma ray which conducted from the first screening experiment. UV-Vis.
spectroscopy was estimated the main influence of the parameters towards Se NPs biosynthesis.
Table 2: Experimental factorial design for the optimization of Se NPs production using
Gentamicin (CN) and sodium selenite after displayed to gamma rays at 10.0 kGy, wavelengths
(nm) and the corresponding optical density.

Runs Sodium selenite Gentamicin (CN) Absorption Wavelength

concentration concentration of Se NPs of Se NPs
(mM) (mM) (O.D.) (nm)
1 1.0 0.5 6.425 325.0
2 1.0 1.0 7.815 375.0
3 1.0 1.5 5.895 375.0
4 1.0 2.0 3.452 375.0
5 1.0 2.5 4.120 370.0
6 1.0 3.0 3.525 375.0

Ass. Prof. Dr. Gharieb S. El-Sayyad, Egyptian Atomic Energy Authority 4

 7 0.5 1.0 0.892 355.0
8 1.0 1.0 6.058 380.0
9 1.5 1.0 7.927 350.0
10 2.0 1.0 9.715 360.0
11 2.5 1.0 9.875 350.0
12 3.0 1.0 8.071 325.0
Other methods of conjugation
Sodium selenite solution (at different conc.) was synthesized by the plant extract like (green tea, or Gum
Arabic) and after the color change and confirmed the Se NPs synthesis they were mixed with different
concentrations of CN solution through a ratio 1:1 (v/v). To achieve the most effective method for preparing
the combination, the reaction parameters were established at 30 °C for the incubation time and 24 hours
for the reaction period over 200 rpm stirring in an incubator with shaking [45].
Characterization of the synthesized Se NPs
The optical property of Se NPs by was characterized by UV-Vis. spectrophotometer (JASCO V-560. UV-
Vis. spectrophotometer) which were compared with a negative control which including the irradiated
sample without selenium salt. The crystallinity and the crystallite size and/or lattice of the synthesized Se
NPs were estimated by the XRD-6000 lists, Shimadzu apparatus, SSI, Japan. The intensity of the
diffracted X- rays was tested as diffracted angle 2θ.
The most predominate Se NPs size and their distribution was defined by Dynamic Light Scattering (DLS-
PSS-NICOMP 380-USA). Additionally, the microstructure, mean particle size and the shape of the
synthesized Se NPs were evaluated by using High-Resolution Transmission Electron Microscope
(HRTEM, JEM2100, Jeol, Japan).
The surface morphology and the grain size of Se NPs were investigated by SEM, ZEISS, EVO-MA10,
Germany. Additionally, EDX analysis (BRUKER, Nano GmbH, D-12489, 410-M, Germany) was used to
estimate the elemental structure, purity and the percentage of each metal presented in our samples. On the
other hand, the mapping technique after conducting SEM/EDX analysis was performed to obtain a
complete knowledge regarding the purity, distribution and the placement of the metals which founded in
the synthesized Se NPs.
Finally, FT-IR spectral analysis was a vital target that gives information regarding the chemical functional
groups presented in both fungal filtrates and CN drug. The experiments were carried out by using a JASCO
FT-IR 3600 Infra-Red spectrometer after conducted KBr pellet technique. It was determined at a wave
number range from 400 to 4000cm-1.

Ass. Prof. Dr. Gharieb S. El-Sayyad, Egyptian Atomic Energy Authority 5

Antimicrobial activity of the synthesized Se NPs
The antimicrobial potential of the synthesized Se NPs (the biogenic and/or incorporated with CN at
25.0µg/ml), fungal filtrate, CN and selenium ions were tested against some selected UTI-causing microbes
by the agar disc diffusion assay.
The tested microbes were obtained kindly from the culture collections in Drug Microbiology Lab., Drug
Radiation Research Dep., NCRRT, Cairo, Egypt. They include Gram-positive (Staphylococcus aureus
and Bacillus subtilis) and Gram-negative bacteria (Pseudomonas aeruginosa, Escherichia coli, and
Klebsiella pneumoniae) in addition to the pathogenic yeast Candida albicans.
The bacterial inoculums were adjusted at 0.5 McFarland (1–4) ×108 CFU/ml, and the tested C. albicans
was set at 0.5 McFarland (2–5 ×107 CFU/ml) (at 600 nm) [46].
Nystatin (NS 100; 100 μg/ml) and Amoxicillin/Clavulanic acid (AMC; 20/10 μg/ml) were used as a
standard antibiotic. The growth inhibition of the pathogenic bacteria and yeast was estimated through the
zone of inhibition (ZOI) after 24 hrs. of incubation [2, 47].
The minimum inhibitory concentrations (MIC) was investigated after Luria-Bertani (LB) broth
inoculation by tested organism and applied the serial dilutions assay [48, 49]. In these purposes, a negative
control (the nutrient broth only), positive one (the tested microbes and the nutrient broth) and the tested
Se NPs-CN (starting with 250 µg/ml conc.) were used and was determined after 24 h. at 37.0 ± 0.5 °C
[47].
The bacterial inoculums were set at 4–5×107 CFU/ml while the tested yeast inoculums was adjusted at 1–
3×107 CFU/ml [46]. MIC was tested by conducted the ELISA plate assay at 600 nm [2, 47]. The MIC was
defined as the lowest Se NPs-CN concentration that effectively represses 99.0% in the growth of the
examined pathogens.
Antibiofilm potential of the synthesized Se NPs-CN
A qualitative estimation regarding the biofilm inhibition was defined as reported by Christensen et al.
[50]. The biofilm test was tested in the absence and present of the synthesized Se NPs-CN and was
observed at the tube wall. The antibiofilm of the synthesized Se NPs-CN at 25.0µg/ml was examined
toward the sensitive UTI-causing microbes (bacteria and yeast that were selected from the antimicrobial
results) and was determined and compared with the control non-treated one.
Briefly, 5.0 ml of the nutrient broth medium was added inside all tubes and the examined bacteria were
inoculated after adjusted 0.5 McFarland to be from 1.0 to 2.5 ×108 CFU/ml. After that, they were incubated
at 37.0 ± 0.5 oC for 24.0 hrs. Additionally, the contents presented in the control and treated tubes were
discarded, mixed with Phosphate Buffer Saline (PBS; pH 7.0) and finally desiccated [36, 51, 52].

Ass. Prof. Dr. Gharieb S. El-Sayyad, Egyptian Atomic Energy Authority 6

The bacterial and yeast cells which adhered to the tube walls were fixed with 5.0 ml sodium acetate (3.0
%) for about 15.0 minutes and finally, they were rinsed with de-ionized water. Biofilms which adhered
inside tubes were stained with 15.0 ml Crystal Violet (CV; 0.1%) and washed with de-ionized water to
remove the rest of the non-reacted CV.
It must be noted that, for the semi-quantitative antibiofilm estimation, 5.0 ml of the absolute ethanol was
added to dissolve the stained bacterial and yeast biofilms [36, 52]. The O.D. of the stained bacterial and
yeast biofilms was examined by UV-Vis. spectrophotometer at 570.0 nm. The bacterial and yeast biofilms
inhibition % was calculated by the following equation [2, 36, 50, 52].
𝐏𝐞𝐫𝐜𝐞𝐧𝐭𝐚𝐠𝐞 𝐨𝐟 𝐛𝐚𝐜𝐭𝐞𝐫𝐢𝐚𝐥 𝐚𝐧𝐝 𝐲𝐞𝐚𝐬𝐭 𝐛𝐢𝐨𝐟𝐢𝐥𝐦 𝐢𝐧𝐡𝐢𝐛𝐢𝐭𝐢𝐨𝐧 (%)
𝐎. 𝐃. 𝐨𝐟 𝐭𝐡𝐞 𝐜𝐨𝐧𝐭𝐫𝐨𝐥 𝐬𝐚𝐦𝐩𝐥𝐞 − 𝐎. 𝐃. 𝐨𝐟 𝐭𝐡𝐞 𝐭𝐫𝐞𝐚𝐭𝐞𝐝 𝐬𝐚𝐦𝐩𝐥𝐞
= 𝐗 𝟏𝟎𝟎
𝐎. 𝐃. 𝐨𝐟 𝐭𝐡𝐞 𝐜𝐨𝐧𝐭𝐫𝐨𝐥 𝐬𝐚𝐦𝐩𝐥𝐞

Scanning electron microscopy and elemental analysis of the control and treated microbial cell
The sensitive bacterial cells which estimated from the antibiofilm result were treated with PBS and
preserved with 5.0 ml glutaraldehyde (3.5 %). The maintained bacterial cells were rinsed repeatedly with
PBS and dehydrated by washing in a series of ethanol solutions with concentrations of 30.0, 50.0, 70.0,
80.0, 95.0, and 100.0 for 10.0 minutes at 24.0 ± 1.0 oC which served as significant method for the drying
process [36, 52].
The bacterial cells were attached and placed above the aluminum stubs for the imaging assay [36, 51, 52].
The surface morphology of the non-treated and treated bacterial cells with the synthesized Se NPs-CN
was examined using scanning electron microscopy (SEM-ZEISS, EVO-MA10). Additionally, the
complete elemental analysis of the sensitive bacterial cell was tested by using Energy-Dispersive X-ray
spectra (EDX-BRUKER, Nano GmbH, D-12489, 410-M, Germany).
Statistical analysis
The statistical examination of the results were calculated and checked by applied the ONE WAY ANOVA
(at P < 0.05), Duncan's multiple effects and the least significant difference income (LSD) [53]. The results
and data were analyzed and calculated by SPSS software version 15.
Results and discussion
Synthesis and optimization of Se NPs by CN and gamma rays
The biosynthesized Se NPs solutions displayed an intense red appearance due to the SPR [54, 55]. Fig.
2A represented the effective gamma rays’ dose that chooses for Se NPs synthesis. It was determined by
UV-Vis. spectroscopy and was estimated to be at 10.0 kGy with the elevated O.D. (1.734) at the
appropriate wavelength (415.0 nm).

Ass. Prof. Dr. Gharieb S. El-Sayyad, Egyptian Atomic Energy Authority 7

Table (1) confirmed the production of Se NPs during the reduction of different sodium selenite
concentration (0.5, 1.0, 1.5, 2.0, 2.5 and 3.0 mM) in the presence of different CN concentrations (0.5, 1.0,
1.5, 2.0, 2.5 and 3.0 mM) after mixing with 0.2 % isopropyl alcohol solution and displayed to 10.0 kGy
gamma rays.
The result listed in Table (1) and displayed in Fig. 1 B & C confirmed that; the run 11 was the superior
concentrations (2.5 mM sodium selenite and 1.0 mM CN) which applied for the Se NPs biosynthesis
(9.875) at 350.0 nm (shorter wavelength which corresponding to smaller size). Also, the same ideas for
the highest selenite concentration were associated with the presence of extra functional groups in CN
which applied for Se NPs biosynthesis after displayed to 10.0 kGy.
Additionally, Fig. 1B displayed the continuous increase in the O.D. regarding the first runs (runs 1 to 6)
where sodium selenite concentration was adjusted at 1.0 mM and CN concentration was changed from
0.5 to 3.0 mM. Our results indicated that, run 2 contains the best concentrations used for the highest Se
NPs production where the CN concentration was at 1.0 mM.
Additionally, Fig. 1C shows that there is a continuous increase in the O.D regarding the second runs (runs
7 to 12) where CN concentration was adjusted at 1.0 mM and sodium selenite concentration was varied
from 0.5 to 3.0 mM.
The presented results concluded that, Se NPs production actually depended on sodium selenite
concentration. It must be noted that, the results of the O.D with a wavelength of the second runs (7 to 12
runs; Table 2) indicated that, the synthesized Se NPs were small in size (blue shift) with the highest yield
as noted in the run 11 and Fig. 2C which discussed that, the production of small size and highest yield of
Se NPs were depends on sodium selenite concentration [56].
Finally, Fig. 1D estimates the stability of CN solution at various concentrations (the same concentrations
utilized in Se NPs biosynthesis) after displayed to 10.0 kGy. The results presented in Fig. 2D showed that,
CN was actually stable after gamma irradiation exposure which supported its availability and stability in
Se NPs incorporation and subsequent its activity. The mean peak of CN was noted at 290.0 nm and the
O.D. was decreased as the concentration of CN reduced.
Proposal reaction mechanism for Se NPs synthesis
Using P. chrysogenum filtrate and gamma rays
Fermentation processes were described as one of the greatest and the most suitable methods which
exhausted low energy for storing food and improving the nutritional properties [57]. P. chrysogenum
metabolites were noted to be an essential function agent in the bio-reduction of selenium ions, stabilization
and the flexibility of Se NPs. The characteristics of P. chrysogenum amino acids and protein are fitted for

Ass. Prof. Dr. Gharieb S. El-Sayyad, Egyptian Atomic Energy Authority 8

the chelating of selenium ions among their amine groups, carbonyl and/or the available electrons [7, 19,
48, 58].
This may surely demonstrate the efficacy of these composites to be adsorbed upon the surface of the
synthesized Se NPs. This apparently indicates that they are required in the first steps of Se NPs structure
in interest to the bio-reduction step [7, 58].
The mechanism of Se NPs biosynthesis by P. chrysogenum filtrate was involved in three principal stages.
The first one was the activation form; throughout the selenium ions reduction and the nucleation of the
degraded metal atoms were occurs.
The second phase was the growth stage; when the small nearby Se NPs quickly combine into an extended
size nano-scale particle, which is followed by a growth of the thermodynamically balanced Se NPs.
Finally, the third stage was the end phase; which defines the definitive shape and the configuration of the
synthesized Se NPs [7, 30, 59].
In expanding to the potency of P. chrysogenum filtrate to reduce and capped Se NPs, 10.0 kGy gamma
rays perform a necessary functional role in Se NPs biosynthesis as describes in the next section.
Using CN and gamma rays
When CN solution (1.0 mM) was mixed with (2.5 mM) sodium selenite solution (the most optimized
condition which produced a high Se NPs yield) and displayed to 10.0 kGy gamma rays, it was produced
the free radical species and solvated electrons (such as: OH•, H•, e-aq, H2O2 and H3O+) [16] (Equ. 1).
γ-radiolysis
H2O OH• + H• + e-aq + H2O2 + H3O+ (1)
The free solvated electrons (e-aq) and proton (H•) are powerful reducing agents which reduced selenium
ions (Se2+) to zero valent selenium atoms (Seo) [60] (Equ. 2&3).
Se2+ + 2 e-aq Seo (2)
Se2+ + H• Seo + H+ (3)
The zero valent Seo firstly dimerized during their incorporation with the remainder Seo and/or Se2+ (Equ.
4&5).
Seo + Seo Seo2 (4)
Seo + Se2+ Se+2 (5)
The present free radicals like OH• and H• are sufficient to remove hydrogen from CN drug and subsequent
production of CN free radicals (secondary radicals= C21H42N5O7•; Equ. 6). Furthermore, CN radicals were
reacted with Se2+ to produce Se NPs and regular stable CN drug (Equ. 7).
Finally, the permanent CN can integrate Se NPs by the incorporation for more stabilization of the
produced Se NPs as presented in Equ. 8.
C21H43N5O7 + OH• C21H42N5O7• + H2O (6)

Ass. Prof. Dr. Gharieb S. El-Sayyad, Egyptian Atomic Energy Authority 9

 2C21H42N5O7• + Se2+ +H2O Reduction
Seo + [C21H43N5O7]2 + H3O+ (7)
Stabilization and capping
C21H43N5O7 + Seo [C21H43N5O7] Seo (8)
In our results, UV-Vis. spectrum of Se NPs-CN at 10.0 kGy presents a peak at 350.0 nm, with extremely
sharp and the greatest intensity was recorded at 9.875 (Table 1).
It was noted that, the highest yield of Se NPs with small size distribution were conducted after the
application CN as stabilizing agents rather than the mycogenic synthesis (P. chrysogenum filtrate).
The speculation can be assumed to the production of the highest amount of the free electron and radical
kinds in the presence of a distinctive stabilizing factors (CN) that increasing the reduction of Se2+ to Seo
NPs [60]. Additionally, P. chrysogenum metabolites were qualified for the reducing Se NPs aggregation,
agglomeration and the precipitation [61].
It must be noted that, by increasing gamma rays’ exposure more than 10.0 kGy, the size distribution and
the average particle size of the synthesized Se NPs were increasing. This may be due to the aggregation
of the synthesized Se NPs by the aid of more electrons and free radicals produced during gamma radiolysis
of water in a time of reaction as explains in Eq. 1 [7, 8, 36, 47].

A B

Ass. Prof. Dr. Gharieb S. El-Sayyad, Egyptian Atomic Energy Authority 10

 C D

Figure 1: UV-Vis. spectroscopy of Se NPs synthesized by Gentamicin (CN) and Sodium selenite at [A] different
gamma irradiation doses, [B] different CN concentration (as mentioned in Table 1), [C] different Sodium selenite
concentration (as mentioned in Table 1) and [D] stability of CN (at different concentration) after displayed to 10.0
kGy.
Sodium Selenite Silver nitrate Gentamicin Amoxicillin-Clavulanic acid (Augmentin)
Fungal filtrate Green tea extract https://link.springer.com/article/10.1007/S12011-019-01842-Z

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