Surfaces and Interfaces 15 (2019) 70–77

Contents lists available at ScienceDirect

Surfaces and Interfaces

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

Green fabrication of iron oxide nanoparticles using grey mangrove Avicennia T

marina for antibiofilm activity and in vitro toxicity
Vaikundamoorthy Ramalingama,b, Pandurangan Dhinesha, Subramaniam Sundaramahalingamc,
Rajendran Rajarama,
⁎

a
DNA Barcoding and Marine Genomics Lab, Department of Marine Science, Bharathidasan University, Tiruchirappalli 620 024, Tamil Nadu, India
b
Department of Animal Science, Chonbuk National University, Jeonju, Republic of Korea
c
Department of Electrical and Electronics Engineering, Mepco Schlenk Engineering College, Sivakasi, Tamil Nadu, India

ARTICLE INFO ABSTRACT

Keywords: The aim of the present study is a biogenic synthesis of iron oxide nanoparticles (FeO-NPs) using grey mangrove
Avicennia marina Avicennia marina to control marine and pathogenic biofilm forming bacteria. Initially, the synthesized nano-
FeO-NPs particles were characterized by UV-visible spectrometer, scanning electron microscope and transmission electron
Characterization microscope for size and shape, Edax analysis for elemental confirmation, zeta potential for stability, X-ray dif-
Antibiofilm activity
fraction for structure and Fourier transform infrared spectroscopy for functional group analysis. Further, the
Toxicity
antibiofilm activity of FeO-NPs were inhibited the initial attachment and biofilm development of primary biofilm
forming bacterial strains Escherichia coli, Pseudomonas aeruginosa and Staphylococcus aureus. Almost 72% of
quorum sensing factors of P. aeruginosa were impeded by 200 µg/ml of FeO-NPs, 63% and 46% for S. aureus and
E. coli respectively. The FeO-NPs also decrease the cell surface hydrophobicity of E. coli, P. aeruginosa and S.
aureus from 16% to 9%, 19% to 11% and 15% to 11% respectively. Moreover, the FeO-NPs inhibit the pro-
duction of exopolysaccharide in E. coli, P. aeruginosa and S. aureus from 90% to 69%, 92% to 65% and 86% to
60% respectively. Further, the FeO-NPs also showed less toxicity against human HBL100 cells using the MTT
assay. In conclusion, the biogenic FeO-NPs could be used as a potential antibiofilm agent against marine and
medical pathogenic biofilm forming bacteria.

1. Introduction
In the modern technology, nanoparticles are used for numerous
biomedical applications where they facilitate laboratory diagnostics
and therapeutics [1]. More specifically for drug delivery purposes, the
use of green synthesized nanoparticles is attracting increasing attention
due to their unique capabilities and their negligible side effects not only
in cancer therapy but also in the treatment of other ailments [2]. Re-
cently researchers focusing on magnetic nanoparticles especially iron
oxide nanoparticles (FeO-NPs), due to their various applications in-
cluding terabit magnetic storage devices, catalysis, sensors and high-
sensitivity biomolecular magnetic resonance imaging (MRI) for medical
diagnosis and therapeutics [3]. Apart from this, the antibiofilm activity
of FeO-NPs was not clearly studied.
The leading problem in medical, food and marine industries is the
attachment of bacteria on solid surfaces called “Biofilms”. These bio-
films are a serious concern, developed by bacterial communities [4,5]
and difficult to remove after provides shelter for bacteria [6]. Biofilms
can act as carrier for pathogenic organisms and cause diseases like otitis
media, otolaryngologic infections, osteomyelitis, bacterial endocarditis,
cystic fibrosis and non-healing wounds [7]. A conventional path to
inhibit the biofilm formation of bacteria is using the antimicrobial
agents. But these bacteria are remains in the presence of high con-
centration of antimicrobial agents due to the existence of poly-
saccharides, proteins, nucleic acids, phospholipids and humic sub-
stances in extracellular polymeric matrix (EPM) that actively involved
in stability of biofilm on surface [8]. The higher antimicrobial re-
sistance of biofilm bacteria can be a protective wall to EPM, conse-
quently the antimicrobial agents acquire to disturb the EPM and de-
struct the biofilm bacteria [9]. In recent times the scientists are focusing
on destruction of EPM of biofilm bacteria and to prevent the pre-ex-
isting biofilms completely [10].
In modern technology researchers are focusing on the compounds
with capability of decrease the gene expression that responsible for
biofilm formation and quorum sensing factor (QSF) [11]. The QSF are
determined by auto inducer molecules like acylated homoserine lactone

⁎
Corresponding author.
E-mail address: rajaramdms@bdu.ac.in (R. Rajaram).

https://doi.org/10.1016/j.surfin.2019.01.008
Received 3 November 2018; Received in revised form 11 January 2019; Accepted 24 January 2019
Available online 28 January 2019
2468-0230/ © 2019 Elsevier B.V. All rights reserved.
V. Ramalingam et al.
(AHL) is the signal molecules produced in out of the cell and attached
with receptor protein that responsible for expression of genes [12]. N-
acetylcysteine (NAC) is a mucolytic agent that effectively decreases the
production of EPM and biofilm formation [13]. Cell Surface Hydro-
phobicity (CSH) of microorganism was also determining the biofilm
formation and influences the anti-infective potential of bacteria [14].
The characterization of CSH is depends upon the existence of hydro-
phobic proteins in EPM underneath an exterior fibrillar layer and can be
precisely determined using a hydrophobicity microsphere assay (HMA)
[15]. The characterization of CSH includes the techniques like sessile
liquid drop contact angle measurement, microbial adhesion to hydro-
carbon (MATH), infrared spectroscopy, X-ray photoelectron spectro-
scopy (XPS), electrophoretic mobility, electron microscopy, retention
on chromatographic resins and adhesion to inanimate materials [16]. In
this background, the present study focusing on to find out the efficiency
of green synthesized iron oxide nanoparticles (FeO-NPs) in disruption
of EPM of biofilm bacteria and its chemical characterization. Further,
the CSH of biofilm bacteria was characterized using MATH assay and
inhibition of QSF by FeO-NPs.
2. Materials and methods
2.1. Collection and preparation of plant extract
A grey mangrove Avicennia marina leaves were collected from
Muthupet Mangrove forest (Lat 10° 25′ N: Long 79° 39′ E), Southeast
Coast of India. Collected plant leaves were washed with sea water and
sterilized with distilled water, dried and powdered using mortar and
pestle. 1 g of leaf powder was mixed with 100 ml of water and kept on
boiling water at 60 °C for 30 min. After cooling, the extracts were fil-
tered with Whatman No. 1 filter paper and stored at 4 °C [17].
2.2. Biogenic synthesis and characterization of nanoparticles
For synthesis of FeO-NPs, 95 ml aqueous solution of 1 mM Ferric
chloride (Himedia) was added with 5 ml extract in 250 ml Erlenmeyer
flask and incubated in shaker [18]. The reaction mixture was analyzed
using UV-visible spectrometer and the reduction of ferric chloride was
examined by change in color. The synthesized NPs were characterized
using UV-visible spectrometer (Shimadzu model 1800), FESEM and
Edax (ΣIGMA model, CASL ZEISS, German), Zeta potential (Malvern
Zetasizer, Nano-ZS90), XRD pattern (Bruker AXS Microstar) and FTIR
(SAM-AV spectrum RXI).
2.3. Antibiofilm activity
2.3.1. Bacterial culture
Human clinical pathogens such as E. coli (MH701895) and P. aeru-
ginosa (MH703431) were collected from Government Hospital,
Tiruchirappalli, Tamil Nadu, India. The marine bacteria Staphylococcus
aureus (KF554244) was collected from contaminated area of Cuddalore
coast. The collected pathogens were cultured in Luria-Bertani broth
(HiMedia) was used and the 24 h old culture was stored as glycerol
stock at −70 °C. The cultures used for each experiment was grown
(OD600 = 1) under aerobic conditions at 37 °C and shaking condition at
150 rpm.
2.3.2. Screening of nanoparticles for biofilm inhibition
The biofilm formation of E. coli, P. aeruginosa and S. aureus
(KF554244) were determined by previously reported method [19]. The
anti-biofilm activity of FeO-NPs, the E. coli, P. aeruginosa and S. aureus
(density 0.05 OD at 600 nm) were grown on glass pieces (1 × 1 cm)
placed in 6 well cell culture plates. The 100 µg/ml concentration of
FeO-NPs were effectively inhibiting the biofilm formation in microtiter
plate were added to 6 well plates and incubated for 36 h at 30 °C. After
removal of non-adherence cells with phosphate buffered saline, the
71
Surfaces and Interfaces 15 (2019) 70–77
biofilm was stained using Live/Dead BacLight Viability Kit (BacLight;
Molecular Probes, Eugene, Oregon, USA), which employs STYO9 and
Propidium iodide (STYO9/PI) and subsequently analyzed with confocal
laser scanning microscope (CLSM - Carl Zeiss) [20]. Further, the biofilm
cultures were grown on glass surfaces and treated with FeO-NPs
(100 µg/ml) for 24 h. After treatment, the glass surfaces were washed
with PBS and stained with acridine orange (0.01%). The biofilm for-
mation was observed under CLSM with 488 nm Ar laser and a
500–640 nm band pass emission filters
2.3.3. Effect of FeO-NPs on QSF
Three biofilm forming bacteria were sub-cultured in M9 minimal
medium by incubating for 24 h at 37 °C and centrifuged at 8000x g for
10 min to separate the cell free supernatant. Equal volume of super-
natant and NPs (25–200 µg/ml) were incubated either in combination
or alone in microtiter plate containing biofilm forming culture super-
natant (density 0.05 OD at 600 nm) and biofilm formation was esti-
mated [21].
2.3.4. Effect of FeO-NPs on cell surface hydrophobicity
Hydrophobicity of biofilm bacteria was estimated using microbial
adhesion to hydrocarbons (MATH) assay and adhesion of bacteria on
glass with hydrophobic hydrocarbons (toluene) was measured [22].
Briefly, 1 ml of biofilm bacterial culture (E. coli – 1.675; P. aeruginosa –
1.645; S. aureus – 1.623 at OD 600 nm) was taken in test tube and
100 µl of toluene with NP (25–200 µg/ml) was added and vortexed for
2 min. After incubation for 10 min. at room temperature, the mixture
was allowed for phase separation and the lower aqueous phase was
measured at 530 nm. The percentage of hydrophobicity was calculated
using the measurements of before and after vortex, the biofilm bacterial
cells with toluene alone in the tube were used as control.
2.3.5. Effect of FeO-NPs on EPS production
The production of EPS was quantified in control and FeO-NPs
treated biofilm forming bacteria by previously described method [23].
The overnight culture of E. coli, P. aeruginosa, and S. aureus was added
to 10 ml LB broth treated with and without FeO-NPs extract and in-
cubated for 24 h at 37 °C. After 24 h of incubation the equal volume of
Trichloroacetic acid (10%) and acetone was added and incubated at
4 °C for overnight. The contents were centrifuged again at 10,000 rpm
for 10 min and the pellet was weighed.
2.4. Toxicity assessment
Human normal HBL100 cells was obtained from NCCS (National
Centre for Cell Sciences, Pune, India). These cell lines were grown in
high glucose DMEM with 50 mM glutamine, supplemented with 10%
FBS, 100 U/ml penicillin and 100 mg/ml streptomycin. Cells were
maintained in a humidified 5% CO2 incubator at 37 °C. The cells were
seeded in 96-well plates at the density of 1 × 105 cells/well in DMEM
media supplemented with 10% FBS and incubated at 37 °C in 5% CO2
incubator. Cells were treated with different concentrations (0–200 μg/
ml) of FeO-NPs for 24 h at 37 °C. After 24 h of incubation, 20 μl of MTT
(5 mg/ml) was added and incubated for 4 h, MTT was aspirated and
200 μl of DMSO was added to dissolve the formazan crystals [24]. The
absorbance was measured at 570 nm using microplate reader (Bio-rad,
USA). The experiments were conducted in triplicates and the significant
difference was calculated.
3. Results and discussion
3.1. Biogenic synthesis and characterization of FeO-NPs
The bacterial biofilms are familiar in nature and its development
leads researchers to investigate many pathogenic diseases caused by
biofilms. These biofilm forming bacteria can damage the medical
V. Ramalingam et al.
Fig. 1. Represents the UV-visible spectrum of 1 mM ferric chloride and biogenic synthesized FeO-NPs (a), SEM image of FeO-NPs (b), TEM images of FeO-NPs (c,d),
the histogram of FeO-NPs size (e) and Edax analysis of FeO-NPs (f).
devices such as urinary catheters, hemodialysis equipment and medical
and dental implants that increases the risk of patient infection. The
present study involved in inhibiting the attachment of biofilm forming
bacteria on solid surface at initial stage of adherence using FeO-NPs.
The extracts of A. marina were used for reduction of FeCl3 into nano-
materials was confirmed by spectrometrically. The synthesis of NPs was
characterized by strong surface plasmon peak at 478 nm in UV-visible
spectrophotometer (Fig. 1a). The size of FeO-NPs was analyzed using
FESEM and the results showed that size ranges from 10–25 nm with
spherical shape (Fig. 1b). Additionally, the spherical shape of FeO-NPs
was analyzed with TEM (Fig. 1c & d) and the results were confirmed the
size of NPs (10–25 nm) with FESEM and the average size of the FeO-NPs
was found to be 16 nm (Fig. 1e). Previously, the size of the NPs was
72
Surfaces and Interfaces 15 (2019) 70–77
∼10 nm [25,26], 15 nm [27] and 64 nm [28] were reported. Moreover,
the Edax analysis showed the presence of Fe element (54%) and the
oxygen molecules (22%) in FeO-NPs (Fig. 1f) and confirmed the green
fabrication of FeO-NPs is highly pure and without any impurities.
The XRD pattern of FeO-NPs was showed the presence of FeO at five
peaks (Fig. 2a) 36.42, 42.24, 61.29, 73.24 and 77.07 were corresponds
to (1 1 1), (2 0 0), (2 2 0), (3 1 1) and (2 2 2) planes of FeO (PDF No.:
46–1312) and also the FeO-NP was cubic crystal in nature and con-
firmed with JCPDS (Joint Committee on Powder Diffraction Standards).
FTIR spectrum of A. marina and FeO-NPs (Fig. 2b) showed a broad peak
at 1383 cm−1, 1635 cm−1 and 3440 cm−1 that corresponds to alco-
holic OeH stretching band, hydroxyl bending mode and hydroxyl
functional group of alcohol. The presence of γ-Fe2O3 NPs was at five
V. Ramalingam et al. Surfaces and Interfaces 15 (2019) 70–77

Fig. 2. XRD pattern (a), the FTIR spectrum of A. marina and FeO-NPs (b) and the zeta potential of NPs (c).

peaks correspondence to (1 1 1), (2 2 0), (3 1 1), (2 2 2), (4 0 0), (4 2 2),

(5 1 1) and (4 4 0) [29]. The peak from 570–625 cm−1 represents the
presence of FeO stretching bend [30]. The average zeta potential of
FeO-NPs was measured as −17.2 mv (Fig. 2c) showed that the NPs are
stable due to the magnetic property of the FeO-NPs.

3.2. Antibiofilm activity of FeO-NPs using microtiter plate assay

Initially, the biofilm forming efficacy was estimated using 96 well

microtiter plate method and the results showed that E. coli and P. aer-
uginosa were strongly adherent and S. aureus was moderately adherent
on surface of plate. The antibiofilm activity of FeO-NPs was assessed
using concentration dependent method in microtiter plate assay against
E. coli, P. aeruginosa and S. aureus (Fig. 3). The activity of NPs was
commensurate and slightly higher for P. aeruginosa at the concentration
of 200 µg/ml, the biofilm disruption was 84%, 79% and 68% on P.
aeruginosa, E. coli and S. aureus respectively. The present results were
compared with the previous results [19, 31] revealed that nanomater-
ials were effectively involved in biofilm disturbance.
The competence of FeO-NPs in reduction of growth rate of biofilm
Fig. 3. Effect of different concentration of FeO-NPs on biofilm formation
bacteria was measured using growth curve analysis in the presence and
measured using 96 microtiter plate assays. Error bar represents the mean
absence of NPs (25–200 µg/ml). The outcome of growth curve analysis (n = 3).
showed NPs (200 µg/ml) were effectively inhibiting the growth of
biofilm bacteria P. aeruginosa (2.453 ± 0.05 to 1.624 ± 0.03), E. coli
(2.157 ± 0.02 to 1.483 ± 0.03) and S. aureus (2.236 ± 0.06 to 1.581) was dose dependent and the complete inhibition by E. coli and P. aer-
after 24 h of period. The antibiofilm effect of nanomaterials was species uginosa was achieved at 28 and 52 µg/ml of NPs [33]. More than 65% of
specific and it effectively disturbed the biofilm formation at low con- biofilm formation was inhibited at 2 ppm concentration of chitosan-
centration (5 µg/ml) [32]. The antibiofilm activity of nanomaterials stabilized NPs in P. aeruginosa and 5 ppm for S. aureus [34].

73
V. Ramalingam et al.
Fig. 4. CLSM shows the effect of FeO-NPs (100 µg/ml) on inhibition of biofilm formation on glass surface.
3.3. CLSM analysis
The confocal microscopic analysis also evaluated to determine the
efficacy of NPs against biofilm formation on solid surfaces. This tech-
nique proved the inhibition of biofilm formation was due to the in-
creased concentration of NPs, although the low concentration of NPs
showed significant inhibition. The complete inhibition of biofilm was
detected at 100 µg/ml concentration of NPs in P. aeruginosa and mod-
erate activity on biofilm formation was observed in E. coli and S. aureus
at 100 µg/ml of concentration (Fig. 4). 5 mg/ml concentration of FeO-
NPs effectively inhibits the formation of biofilm by E. faecalis, B. subtilis
C. krusei, P. aeruginosa and E. coli [35]. The chemically synthesized FeO-
NPs were decreased the number of bacteria at 2 mg/ml concentration
[36] (Taylor and Webster, 2009). Further, FeO-NPs also inhibit the
growth of biofilm forming bacteria. After 48h treatment of super-
paramagnetic FeO-NPs (10 µg/ml), 65% of dead S. epidermis bacteria
were quantified by fluorescence microscopic method [36] (Fig. 5).
3.4. Live and dead cells staining assay
Moreover, the live and dead cells were also visualized using SYTO9
and propidium iodide staining method. The results indicated that in-
crease the concentration of FeO-NPs increase the number of dead cells.
Among the three bacteria, S. aureus was highly killed followed by P.
aeruginosa and E. coli by FeO-NPs. Previously, 24 h treatment with FeO-
NPs (200 mg/ml) was decrease the P. aeruginosa attachment on glass
slide and the complete detachment was observed in NPs combined with
10 mM of fructose metabolites [37]. Nguyen et al. [38] reported that
treatments with FeO-NPs induces 19.7% decrease in biofilms which
may be due to the presence of iron in the nanoparticles. There is a
significant reduction (P < 0.05) in biofilm growth on all the three
74
Surfaces and Interfaces 15 (2019) 70–77
bacteria was observed in the presence of iron-oxide nanoparticles
compared to control [39].
3.5. Effect of FeO-NPs on QSF
The ability of NPs to inhibit the QSF was assessed by evaluating the
competence between NPs and culture supernatant. Roughly 72% of QSF
inhibiting activity was obtained in P. aeruginosa at 200 µg/ml of NPs
and 63% for S. aureus and 46% for E. coli (Fig. 6). The different con-
centration of iron and oxygen affect the quorum sensing regulatory
protein lasR. The increase of iron concentration effectively decreases
the β—galactosidase activity leads to the expression of lasR that de-
creases the virulence activity of bacteria [40]. The probability of po-
tential inhibition of biofilm formation suggested that the biofilm in-
hibiting NPs were interacted with membrane biofilm bacteria and
induces the oxidative stress [41].
3.6. Effect of FeO-NPs on hydrophobicity of biofilm bacteria
CSH plays an important role in initial adhesion of bacteria on solid
surfaces. In our result, the initial hydrophobicity was 16%, 19% and
15% for E. coli, P. aeruginosa and S. aureus respectively (Fig. 7). After
24 h treatment with 200 µg/ml of NPs, the CSH was decreased to 9%,
11% and 11% for E. coli, P. aeruginosa and S. aureus respectively. The
increase or decrease of absorbance at 530 nm depends upon the pre-
sence of FeO-NPs on cell wall of biofilm bacteria. FeO-NPs were at-
tached on the cell wall of biofilm bacteria and increase the interaction
with hydrocarbons that leads to hydrophobicity on cell wall. Recently,
the single dosage of nanomaterials with numerous corrosion rates could
be adequate to increase the antimicrobial resistance over the MIC level
and evade the biofilm formation [42]. The increase of absorbance was
V. Ramalingam et al.
Fig. 5. Effect of FeO-NPs on biofilm formation of E. coli, P. aeruginosa and S. aureus using live/dead cells staining assay.
due to the raise of boundary between oil and water during the phase
separation in MATH assay and vice versa for decrease.
3.7. Effect of FeO-NPs on EPS production of biofilm bacteria
EPS represents the house of biofilms and determines the life of
biofilm formation, the biofilm forming ability of microorganisms is lost
by inhibiting the EPS production. Previously many reports stated that
EPS plays a crucial role in the biofilm formation and by destroying the
EPS leads to inhibition of biofilm formation [43,44]. Among the tested
concentration of FeO-NPs, almost 69%, 65% and 60% of EPS produc-
tion was inhibited in E. coli, P. aeruginosa and S. aureus respectively by
200 µg/ml of NPs after 24 h treatment. Initially, 90%, 92% and 86% of
EPS was produced by biofilm forming bacteria E. coli, P. aeruginosa and
75
Surfaces and Interfaces 15 (2019) 70–77
S. aureus respectively (Fig. 8). These findings suggested that the bio-
genic FeO-NPs can act as a potential antibiofilm agent. Recently, Le-
wisoscar et al. [45] reported that the nanoparticles effectively inhibited
the EPS production and thus resulted into reduce the EPS efficiency that
prevent the penetration of antimicrobial compounds and provide re-
sistance to antimicrobial agents.
3.8. Toxicity assessment of FeO-NPs
Further, the excellent antibiofilm activity of FeO-NPs was assessed
for toxicity against human normal cell lines using MTT assay. The re-
sults showed that the increasing the concentration of FeO-NPs from 0 to
200 µg/ml are not cause the viability of cells. Instead, the number of
cells at higher concentration after 24 h treatment with FeO-NPs was
V. Ramalingam et al.
Fig. 6. Competence between QSF and different concentrations of FeO-NPs.
Error bar represents the mean (n = 3).
Fig. 7. Inhibitory effect of FeO-NPs on the activity of cell surface hydro-
phobicity using MATH assay. Error bar represents the mean (n = 3).
82% reveals the biogenic fabricated FeO-NPs has less toxicity in human
cell lines (Fig. 9). A comparative study between iron oxide nano-
particles and human normal cell has demonstrated that FeO-NPs not
toxic or they contain the lowest cytotoxicity effect has been reported
previously [46].
4. Conclusion
FeO-NPs synthesized by grey mangrove A. marina have many ap-
plications that afford a gadget for enhanced the study of innovative
anti-adhesive materials. In this report, the FeO-NP effectively inhibited
the biofilm formation and also the growth of biofilm bacteria. FeO-NPs
also compete with QSF and CSH of biofilm bacteria and diffuse the
activity. FeO-NPs enhance the inhibition of EPS production in biofilm
forming bacteria. Moreover, the toxicity assessment of FeO-NPs showed
less toxicity against HBL100 cells at higher concentration revealed that
the NPs could be used as a potential antibiofilm agent.
76
Surfaces and Interfaces 15 (2019) 70–77
Fig. 8. Inhibition of EPS production of biofilm forming bacteria using biogenic
FeO-NPs. Error bar represents the mean (n = 3).
Fig. 9. Toxicity assessment of FeO-NPs against HBL100 cells using MTT assay.
Acknowledgment
The authors gratefully acknowledge the authorities of
Bharathidasan University for providing CLSM facility under the DST-
PURSE scheme (SR/FT/LS-113/2009).
Conflict of interest
Authors declares that the there is no conflict of interest to disclose.
References
[1] V. Ramalingam, S. Dhanasundari, P. Nithiya, R. Rajaram, Catalytic degradation of
methyl orange using biogenic nanosilver and its phytotoxicity evaluation, Ind. J.
Chem. Tech. 24 (3) (2017) 336–343.
[2] V. Ramalingam, S. Revathidevi, T.S. Shanmuganayagam, L. Muthulakshmi,
R. Rajaram, Gold nanoparticle induces mitochondria-mediated apoptosis and cell
cycle arrest in nonsmall cell lung cancer cells, Gold Bull. 50 (2) (2017) 177–189.
[3] W. Wu, Z. Wu, T. Yu, C. Jiang, W.S. Kim, Recent progress on magnetic iron oxide
nanoparticles: synthesis, surface functional strategies and biomedical applications,
Sci. Technol. Adv. Mater. 16 (2) (2015) 023501.
[4] G. Applerot, J. Lellouche, N. Perkas, N. Yeshayahu, A. Gedanken, E. Banin, ZnO
nanoparticle-coated surfaces inhibit bacterial biofilm formation and increase
V. Ramalingam et al.
antibiotic susceptibility, RSC Adv. 2 (2012) 2314–2321.
[5] A. Hequet, V. Humblot, J.M. Berjeaud, C.M. Pradier, Optimized grafting of anti-
microbial peptides on stainless steel surface and biofilm resistance tests, Colloids
Surf. B 84 (2011) 301–309.
[6] D. Pavithra, M. Doble, Biofilm formation bacterial adhesion and host response on
polymeric implants–issues and prevention, Biomed. Mater. 3 (2008) 34003–34016.
[7] F. Martinez-Gutierrez, L. Boegli, A. Agostinho, E.M. Sanchez, H. Bach, F. Ruiz,
G. James, Anti-biofilm activity of silver nanoparticles against different micro-
organisms, Biofouling 29 (6) (2013) 651–660.
[8] D.E. Waturangi, Y.A. Bunardi, S. Magdalena, Antibiofilm activity of bacteria iso-
lated from marine environment in Indonesia against Vibrio cholerae, Res. J. Microb.
6 (12) (2011) 926–930.
[9] A. Shrestha, Z. Shi, K.G. Neoh, A. Kishen, Nanoparticulates for antibiofilm treat-
ment and effect of aging on its antibacterial activity, J. Endod. 36 (2010)
1030–1035.
[10] P. Jiang, J. Li, F. Han, G. Duan, X. Lu, Y. Gu, et al., Antibiofilm activity of an
Exopolysaccharide from Marine Bacterium Vibrio sp. QY101, PLoS One 6 (4) (2011)
e18514.
[11] I.A. Packiavathy, S. Priya, S.K. Pandian, A.V. Ravi, Inhibition of biofilm develop-
ment of uropathogens by curcumin - an anti-quorum sensing agent from Curcuma
longa, Food Chem. 148 (2014) 453–460.
[12] C.M. Waters, B.L. Bassler, Quorum sensing: cell-to-cell communication in bacteria,
Annu. Rev. Cell Dev. Biol. 21 (2005) 319–346.
[13] T. Zhao, Y. Liu, N-acetylcysteine inhibit biofilms produced by Pseudomonas aeru-
ginosa, BMC Microbiol. 10 (2010) 140–147.
[14] S. Chusri, P.N. Phatthalung, S.P. Voravuthikunchai, Anti-biofilm activity of Quercus
infectoria G. Olivier against methicillin-resistant Staphylococcus aureus, Lett. Appl.
Microbiol. 54 (2011) 511–517.
[15] M.A. Jabra-Rizk, W.A. Falkler, W.G. Merz, T.F. Meiller, New assay for measuring
cell surface hydrophobicities of Candida dubliniensis and Candida albicans, Clin.
Diagn. Lab. Immunol. 8 (3) (2001) 585–587.
[16] J. Masuoka, K.C. Hazen, Cell wall mannan and cell surface hydrophobicity in
Candida albicans Serotype A and B strains, Infect. Immun. 72 (11) (2004)
6230–6236.
[17] S. Gnanasekar, J. Murugaraj, D. Balakrishnan, V. Krishnamoorthy, P.K. Jha,
P. Seetharaman, R. Vilwanathan, S. Sivaperumal, Antibacterial and cytotoxicity
effects of biogenic palladium nanoparticles synthesized using fruit extract of
Couroupita guianensis Aubl, J. Appl. Biomed. 16 (2018) 59–65.
[18] V.V. Makarov, S.S. Makarova, A.J. Love, O.V. Sinitsyna, A.O. Dudnik,
I.V. Yaminsky, M.E. Taliansky, N.O. Kalinina, Biosynthesis of stable iron oxide
nanoparticles in aqueous extracts of Hordeum vulgare and Rumex acetosa plants,
Langmuir 30 (2014) 5982–5988.
[19] V. Ramalingam, S. Raja, S. Sundaramahalingam, R. Rajaram, Chemical fabrication
of graphene oxide nanosheets attenuates biofilm formation of human clinical pa-
thogens, Bioorg. Chem. 83 (2018) 326–335.
[20] R. Vaikundamoorthy, R. Rajendran, A. Selvaraju, K. Moorthy, S. Perumal,
Development of thermostable amylase enzyme from Bacillus cereus for potential
antibiofilm activity, Bioorg. Chem. 77 (2018) 494–506.
[21] S.M.A. Sayem, E. Manzo, L. Ciavatta, A. Tramice, A. Cordone, A. Zanfardino, et al.,
Anti-biofilm activity of an exopolysaccharide from a sponge-associated strain of
Bacillus licheniformis, Microb. Cell Fact. 10 (2011) 74.
[22] R.S. Pembrey, K.C. Marshall, R.P. Schneider, Cell surface analysis techniques: what
do cell preparation protocols do to cell surface properties? Appl. Environ.
Microbiol. 65 (7) (1999) 2877–2894.
[23] V. Vicente-Garcia, E. Rios-Leal, G. Calderon-Dominguez, R.O. Canizares-Villanueva,
R. Olvera-Ramirez, Detection, isolation, and characterization of exopolysaccharide
produced by a strain of Phormidium 94a isolated from an arid zone of Mexico,
Biotechnol. Bioeng. 85 (2004) 306–310.
[24] V. Ramalingam, R. Rajaram, Enhanced antimicrobial, antioxidant and anticancer
activity of Rhizophora apiculata: an experimental report, 3 Biotech. 8 (4) (2018)
200.
[25] A. Uheida, M. Iglesias, C. Fontas, M. Hidalgo, V. Salvado, Y. Zhang, M. Muhammed,
Sorption of palladium(II) rhodium(III) and platinum(IV) on Fe3O4 nanoparticle, J.
Colloid Interface Sci. 301 (2006) 402–408.
[26] A. Uheida, G. Salazar-Alvarez, E. Bjorkman, Z. Yu, M. Muhammed, Fe3O4 and γ-
77
Surfaces and Interfaces 15 (2019) 70–77
Fe2O3 nanoparticles for the adsorption of Co2+ from aqueous solution, J. Colloid
Interface Sci. 298 (2006) 501–507.
[27] K. Simeonidis, S. Mourdikoudis, M. Moulla, I. Tsiaoussis, C. Martinez-Boubetaa,
M. Angelakerisa, et al., Controlled synthesis and phase characterization of Fe-based
nanoparticles obtained by thermal decomposition, J. Magn. Magn. Mater. 316
(2007) e1–e4.
[28] Z. Cheng, A.L.K. Tan, Y. Tao, D. Shan, K.E. Ting, X.J. Yin, Synthesis and char-
acterization of iron oxide nanoparticles and applications in the removal of heavy
metals from industrial wastewater, Int. J. Photoenergy (2007) 608298.
[29] V. Sreeja, P.A. Joy, Microwave–hydrothermal synthesis of γ-Fe2O3 nanoparticles
and their magnetic properties, Mater. Res. Bull. 42 (2007) 1570–1576.
[30] G. Shahnaz, C. Kremser, A. Reinisch, A. Vetter, F. Laffleur, D. Rahmat, et al.,
Efficient MRI labeling of endothelial progenitor cells: design of thiolated surface
stabilized superparamagnetic iron oxide nanoparticles, Eur. J. Pharm. Biopharm. 85
(2013) 346–355.
[31] G.D. Christensen, W.A. Simpson, J.A. Younger, L.M. Baddour, F.F. Barrett,
D.M. Melton, et al., Adherence of coagulase-negative Staphylococci to plastic tissue
culture plates: a quantitative model for the adherence of Staphylococci to medical
devices, J. Clin. Microbiol. 22 (1985) 996–1006.
[32] D. Inbakandan, C. Kumar, L.S. Abraham, R. Kirubagaran, R. Venkatesan, S.A. Khan,
Silver nanoparticles with anti microfouling effect: a study against marine biofilm
forming bacteria, Colloids Surf. B 111 (2013) 636–643.
[33] P. Kanmani, S.T. Lim, Synthesis and characterization of pullulan-mediated silver
nanoparticles and its antimicrobial activities, Carbohydr. Polym. 97 (2013)
421–428.
[34] P. Jena, S. Mohanty, R. Mallick, B. Jacob, A. Sonawane, Toxicity and antibacterial
assessment of chitosan coated silver nanoparticles on human pathogens and mac-
rophage cells, Int. J. Nanomed. 7 (2012) 1805–1818.
[35] A.M. Prodan, S.L. Iconaru, C.M. Chifiriuc, C. Bleotu, C.S. Ciobanu, M. Motelica-
Heino, et al., Magnetic properties and biological activity evaluation of iron oxide
nanoparticles, J. Nanomater. (2013) 893970.
[36] E.N. Taylor, T.J. Webster, The use of superparamagnetic nanoparticles for pros-
thetic biofilm prevention, Int. J. Nanomed. 4 (2009) 145–152.
[37] N.G. Durmus, E.N. Taylor, K.M. Kummer, T.J. Webster, Enhanced efficacy of
Superparamagnetic Iron Oxide nanoparticles against antibiotic-resistant biofilms in
the presence of metabolites, Adv. Mater. 25 (40) (2013) 5706–5713.
[38] T.K. Nguyen, H.T. Duong, R. Selvanayagam, C. Boyer, N. Barraud, Iron oxide na-
noparticle-mediated hyperthermia stimulates dispersal in bacterial biofilms and
enhances antibiotic efficacy, Sci. Rep. 5 (2015) 18385.
[39] T. Monica, S. Soundarya, K. Sathish kumar, S. Guruprakash, Antibacterial Efficacy
of iron-oxide nanoparticles against biofilms on different biomaterial surfaces, Int. J.
Biomat. 716080 (2014) 6.
[40] E.J. Kim, W. Wang, W.D. Deckwer, A.P. Zeng, Expression of the quorum-sensing
regulatory protein LasR is strongly affected by iron and oxygen concentrations in
cultures of Pseudomonas aeruginosa irrespective of cell density, Microbiology 151
(2005) 1127–1138.
[41] C. Kaweeteerawat, A. Ivask, R. Liu, H. Zhang, C.H. Chang, C. Low-Kam, et al.,
Toxicity of metal oxide nanoparticles in Escherichia coli correlates with conduction
band and hydration energies, Environ. Sci. Technol. 49 (2) (2015) 1105–1112.
[42] J.K. Miller, R. Neubig, C.B. Clemons, K.L. Kreider, J.P. Wilber, G.W. Young, et al.,
Nanoparticle deposition onto biofilms, Ann. Biomed. Eng. 41 (1) (2013) 53–67.
[43] P.C. Bogino, L. Oliva Mde, F.G. Sorroche, W. Giordano, The role of bacterial bio-
films and surface components in plant-bacterial associations, Int. J. Mol. Sci. 14 (8)
(2013) 15838–15859.
[44] M. Zubair, M. Ashraf, M. Arshad, A. Raza, B. Mustafa, A. Ahsan, Formation and
significance of bacterial biofilms, Int. J. Curr. Microbiol. App. Sci. 3 (12) (2014)
917–923.
[45] F. Lewisoscar, D. MubarakAli, C. Nithya, R. Priyanka, V. Gopinath, N.S. Alharbi,
N. Thajuddin, One pot synthesis and anti-biofilm potential of copper nanoparticles
(CuNPs) against clinical strains of Pseudomonas aeruginosa, Biofouling 31 (4) (2015)
379–391.
[46] L. Gholami, R.K. Oskuee, M. Tafaghodi, A.R. Farkhani, M. Darroudi, Green facile
synthesis of low-toxic superparamagnetic iron oxide nanoparticles (SPIONs) and
their cytotoxicity effects toward Neuro2A and HUVEC cell lines, Ceram. Int. 44
(2018) 9263–9268.