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Trakya University Journal of Natural Sciences, 21(1): 21-32, 2020
ISSN 2147-0294, e-ISSN 2528-9691
DOI: 10.23902/trkjnat.641367
OPEN ACCESS
© Copyright 2020 Erdoğan
Research Article
TEXTILE FINISHING WITH CHITOSAN AND SILVER NANOPARTICLES
AGAINST Escherichia coli ATCC 8739
Sevil ERDOĞAN
Trakya University, Keşan Vocational College, Laborant and Veterinary Health Programme, 22800, Keşan, Edirne,
TURKEY
Cite this article as:
Erdoğan S. 2020. Textile Finishing with Chitosan and Silver Nanoparticles against Escherichia coli ATCC 8739. Trakya Univ J Nat Sci, 21(1): 21-32,
DOI: 10.23902/trkjnat.641367
Received: 01 November 2019, Accepted: 07 February 2020, Online First: 17 February 2020, Published: 15 April 2020
Edited by:
Hatice Korkmaz Güvenmez
Corresponding Author:
Sevil Erdoğan
sevilerdogan@trakya.edu.tr
ORCID ID:
orcid.org/0000-0001-9148-911X
Key words:
Textile finishing process
Biomaterial
Nano-Ag
Antibacterial
Shrimp
Crayfish
Abstract: The finishing process with the antibacterial agents that protect the environment and
human health is gaining importance. This study aims 1) to develop new generation antibacterial
finishes using chitosan as a binder for nano-Ag coatings, 2) to determine the applicability of
chitosan from shrimp and crayfish for textile production and 3) to contribute to environmentally
friendly textile production. Chitosan from shrimp and crayfish wastes were used as adhesive in the
binding of nanoparticles to fabric surfaces. The bonding properties of the nano-Ag particles on the
fabric surfaces were investigated by Fourier transform infrared spectroscopy (FTIR), Scanning
electron microscopy (SEM), and Energy dispersive x-ray spectroscopy (EDX) analysis. The
antibacterial effectiveness of fabrics against Escherichia coli ATCC 8739 were tested according to
JIS L 1902-2015 standard. The crayfish and shrimp chitosan formed a colorless film and coated the
nano-Ag particles homogeneously on the cotton fabric. Antibacterial activity values were
calculated as 3.10 and 5.74 for crayfish and shrimp chitosan coated cotton fabrics and as 5.37 and
5.10 for crayfish and shrimp chitosan+nano Ag coated cotton fabrics, respectively. Chitosan nano-
Ag coating which exhibited a good antibacterial activity (99.99% reduction) against E. coli ATCC
8739 can be used in the manufacture of garments such as medical textiles, baby clothes, and
underwear. The use of chitosan as a binder can reduce the use of chemicals in textile printing and
pigment dying in finishing materials, pollutant discharges and emissions from industrial sources.
Also, it presents innovative solutions for the protection of human and environmental health.
Özet: Çevreyi ve insan sağlığını koruyan antibakteriyel malzemelerle bitim işlemi son yıllarda
önem kazanmaktadır. Bu çalışmanın amacı da 1) kitosanı nano-gümüş (Ag) kaplamalar için
bağlayıcı olarak kullanarak yeni nesil antibakteriyel apreler geliştirmek, 2) karides ve kerevitlerden
üretilen kitosanın tekstil üretimi için uygulanabilirliğini belirlemek ve 3) çevre dostu tekstil
üretimine katkıda bulunmaktır. Çalışmada, karides ve kerevit atıklarından üretilen kitosan, Ag
nanopartiküllerinin kumaş yüzeylerine bağlanmasında yapışkan olarak kullanılmıştır. Nano-Ag
partiküllerinin kitosan aracılığıyla kumaş yüzeylerine bağlanma özellikleri Fourier dönüşümlü
kızılötesi spektroskopi (FTIR), Taramalı elektron mikroskobu (SEM) ve Enerji dağılımlı x ışını
(EDX) analizleri ile incelenmiştir. Kumaşların Escherichia coli ATCC 8739' ye karşı antibakteriyel
aktiviteleri JIS L 1902-2015 standardına göre test edilmiştir. Çalışmanın sonuçları kerevit ve
karides kitosanlarının renksiz bir film oluşturduğunu ve Ag nanoparçacıklarını pamuklu kumaş
üzerine homojen bir şekilde kapladığını göstermiştir. Kerevit kitosanı ve karides kitosanı ile kaplı
pamuklu kumaşların antibakteriyel aktivite değerleri sırasıyla, 3,10 ve 5,74 olarak hesaplanırken,
kerevit kitosanı+nano-Ag ve karides kitosanı+nano-Ag ile kaplanmış pamuklu kumaşların
antibakteriyel aktivite değerleri sırasıyla 5,37 ve 5,10 olarak bulundu. E. coli ATCC 8739' ye karşı
iyi bir antibakteriyel aktivite sergileyen (% 99,99 azalma) kitosan+nano-Ag kaplamalar, tıbbi
tekstiller, bebek kıyafetleri ve iç çamaşırları gibi giysilerin imalatında kullanılabilir. Binder olarak
kitosanın kullanılması, tekstil baskısında, pigment boyamada, terbiye maddelerinde, kirletici
deşarjlarında ve endüstriyel kaynaklı emisyonlarda kimyasalların kullanımını azaltabilir. Ayrıca,
insan ve çevre sağlığının korunmasına yönelik yenilikçi çözümler sunar.
Introduction
Microorganisms can live in all kinds of environments
including air, water and soil, and they can even survive in
extreme environments such as deep-sea bottoms and
volcano mouths. They are most commonly found in foods,
living organisms, and clothing. In particular, cotton
garments create favorable environments for bacteria with
22 S. Erdoğan
their natural fibrous, porous and hydrophilic structure and
by providing moisture, temperature and nutrient in areas
that come into contact with the skin (Akaydın & Kalkancı
2014, Zhou & Kan 2014). The damage caused by
microorganisms affects cotton fabrics more than other
types of fabrics. Microorganisms not only cause odor
formation and deterioration in the fabric, but also they
cause dermal infections and allergic reactions (Tania et al.
2019). Colclasure et al. (2015) reported that clinically
important coliform bacteria, such as Escherichia coli, can
be present in significant amounts, especially on moisture-
retaining fabrics at room temperatures and in dark
conditions and can survive for long periods. The
proliferation on the fabric surface of this pathogen
bacteria, which often causes gastrointestinal infections
and outbreaks of food poisoning, needs to be controlled
(Pevzner 2018). Therefore, this study focused on E. coli
ATCC 8739 and developing textile finishing agents which
would give antimicrobial activity to fabrics against this
pathogenic bacteria.
Polymers are materials used in various applications
and are of great importance in human life. Although
industrial polymers have high potential properties, they
create pollution as a result of not being destroyed by
natural processes (Sher et al. 2013). This led researchers
to renewable natural resources. Chitosan is a renewable
biopolymer and it is produced by deacetylation of chitin,
which is a protective and supportive structure in a wide
variety of living groups such as crustaceans, mollusks,
insects and fungi (Abdou et al. 2008, Erdogan & Kaya
2016, Erdogan et al. 2017, Arrouze et al. 2017, Zhang et
al. 2017, Song et al. 2018). Chitosan, which is a natural,
biocompatible, biodegradable and non-toxic biopolymer,
can be easily modified and functionalized thanks to its
reactive hydroxyl and amino groups and it is used in
diverse application areas (Anitha et al. 2014).
Furthermore, thanks to these reagent groups, chitosan can
be used in finishing and in textile dyeing (Islam & Butola
2019). The cationic structure of chitosan plays an active
role by exhibiting antimicrobial, antioxidant, antitumor
and anticancer properties which enables it to be used as a
therapeutic and antibacterial agent in many medical
applications (Anitha et al. 2014).
Nanoparticles are used for odor removal on textiles or
controlled release of antifungals and biocides (Rivero et
al. 2015). Nanoparticles with antimicrobial or
antibacterial properties can be added to the fibers with
nanotubes for long-term protection or can be added to
polymers in fabric coatings. Nanosilver (nano-Ag)
attracts attention with its antibacterial properties that
provide an easy and lasting effect on pathogenic
microorganisms like bacteria and viruses that threaten
human health (Morones et al. 2005, Panacek et al. 2006).
Besides, silver (Ag) nanoparticles are commercially used
as a broad-spectrum antibiotic agent and exhibit high
performance even at low concentrations (Govindan et al.
2012, Chen et al. 2014). Since they have a strong
antibacterial effect and do not create toxic effects, Ag and
Ag compounds can be used on many surfaces and areas
where harmful microorganisms are abundant during
production or after production (Becenen & Altun 2016).
Therefore, in this study, we coated cotton fabric samples
with nano-Ag particles using chitosan as a crosslinker and
investigated the antibacterial activity of nano-Ag and
chitosan together against E. coli.
Recently, a large amount of studies have been carried
out to give antimicrobial properties to textile samples
(Wang et al. 2016, Souza et al. 2017, Scacchetti et al.
2018, Fan et al. 2018, Rehan et al. 2018, Xu et al. 2019).
In these studies, chitosan was coated on fabric surfaces
via various crosslinkers or using different techniques as
composite prepared with various antibacterial agents.
Furthermore, in most antibacterial textile coatings, fabric
samples are pretreated and modified using various
chemicals before or during coating. In this study, nano-
Ag particles were coated via chitosan on fabric made of
100% cellulose without pretreating and using additional
chemicals. Thus, we strove to achieve in a single step both
to bond nano-Ag on the fabric surface and to give
antibacterial properties to the fabric, by evaluating
environmental waste materials and reducing the use of
chemicals. Previous studies pointed out that chitosan
source affects both the physicochemical and biological
properties of chitosan (Rinaudo 2006). However, there is
no comparative study on how chitosan obtained from
different sources affect the antibacterial performance of
fabric samples. In this study, we tested the antibacterial
activity of fabric samples coated with chitosan produced
from two different sources (saltwater shrimp Parapenaus
longirostris (Lucas, 1846) and freshwater crayfish
Astacus leptodactylus (Eschscholtz, 1823)) by using the
same method under controlled and standard conditions.
Also, unlike other studies, we used JIS L 1902: 2015 test
method, instead of the commonly used ATCC 147 and
AATCC 100 tests for determining antibacterial activity of
chitosan+nano-Ag coatings. Thus, we expect to obtain
more realistic and accurate results with this method which
reflects the real-life conditions by keeping limited the
amount of nutrients in the medium.
This study aims 1) to develop new generation
antibacterial finishes using chitosan as a binder for nano-
Ag coatings, 2) to determine the applicability of chitosan
produced from shrimp and crayfish for textile production
and 3) to contribute to environmentally friendly textile
production.
Materials and Methods
Materials used in the experiment
Waste shells forming the exoskeleton of shrimp and
crayfish were utilized as sources of chitin and chitosan.
Chitin was isolated by the chemical synthesis method
from waste shells of shrimp (Parapenaus longirostris
(Lucas, 1846)) purchased from a local market and
crayfish (Astacus leptodactylus (Eschscholtz, 1823))
caught from Altınyazı dam lake (in Turkey). Isolated
chitin were then converted to chitosan by deacetylation.
Textile Finishing with Chitosan and Silver Nanoparticles against E. coli ATCC 8739 23
Trakya Univ J Nat Sci, 21(1): 21-32, 2020
Pretreated optic white cotton calico fabric made of 100%
cotton yarn was used to apply chitosan and nano-Ag
particles. Nano-Ag particles (≤100 nm particle size,
purity: 99.5%) were purchased from Sigma Aldrich. The
nanoparticles were vortexed for 10 seconds before use to
ensure homogenization of the particles.
Chitin isolation and chitosan production
Shrimp and crayfish waste shells were cleaned with
water to remove impurities and then rinsed with distilled
water and dried in the oven at 50 °C for 2-3 days. Chitin
isolation and chitosan production from both organisms
were performed following the same procedure (Fig. 1).
Firstly, crushed shrimp and crayfish shells (40 g) were
refluxed with 1.8 M 400 mL HCl at 60-70°C for 6 hours
at 700 rpm. The acid-treated shells were then filtered
through common filter paper and rinced with pure water
until free of acid. The demineralized shells were then
treated with 1.8 M 400 mL of NaOH at 70°C for 20 hours
by stirring at 700 rpm. The mixture was then filtered again
and rinsed with distilled water until the pH was neutral.
Employing these two processes, minerals and proteins in
waste shells were removed and chitin was obtained. Since
the shells of both organisms contained a high percentage
of astaxanthin pigment, the chitin obtained from these
organisms were subjected to decolorization process.
Decolorization was achieved by treating chitin samples
with a mixture of 200 mL including chloroform, methanol
and distilled water in a 1: 2: 4 ratio, respectively. This was
accomplished by stirring the mixture containing chitin
samples at 800 rpm for 2 hours at room temperature.
Finally, the mixture was filtered and the collected chitin
samples were washed with pure water and dried in the
oven at 50 °C.
The dried chitin samples were refluxed with 70%
NaOH solution for 3 hours at 150 °C, 800 rpm to obtain
chitosan (Fig. 1). The deacetylated chitin samples were
then washed continuously with pure water and the process
continued until the samples were cleared of base and
reached pH: 7.0. The obtained chitosan sample was placed
in a petri dish and dried at 50 °C in an oven.
Fig. 1. Scheme illustrating the production of chitosan from shrimp and crayfish.
Fig. 2. Application of nano-Ag to fabric through chitosan. a) Dissolution of chitosan b) Coating machine c) Coated fabric.
24 S. Erdoğan
Coating of cotton fabric with chitosan and nano-Ag
mixture
A mixture including 10% by weight of chitosan and
1% acetic acid (CH3COOH) was prepared using chitosan.
To dissolve the chitosan, the mixture was heated at 40 °C
by stirring at 400 rpm for 5 hours (Fig. 2a). Nano-Ag
(0.5% by weight) was added to the prepared chitosan
solutions and stirring was continued for a further 1 hour.
The solutions containing chitosan and nano-Ag were
impregnated with the pad-dry method on fabrics in the
Ataç Brand foular machine with 100% squeezing pressure
(Fig. 2b). After impregnation, the fabrics were dried at
105-110 °C and then fixed in the same machine at 120 °C
(Fig. 2c).
Fourier Transform Infrared Spectroscopy (FTIR)
Chemical structure characterization was performed to
confirm that the material isolated from shrimp and
crayfish was chitin. Furthermore, to determine whether
the chitosan+nano-Ag mixture was coated on cotton
fabrics, changes in chemical bonds were determined by
FTIR peaks. FTIR spectra were obtained with Perkin–
Elmer ATR FTIR device in the range of 4000 to 400 cm−1
wavelength. Surface analyses of chitosan nano-Ag coated
fabric samples were performed in Trakya University
Technology Research Development Application and
Research Center (TUTAGEM).
Scanning electron microscopy (SEM) and Energy-
dispersive X-ray analysis (EDX)
The success of chitosan obtained from shrimp and
crayfish wastes in coating nano-Ag particles on fabric
samples, bonding properties between fabric and coating
and the change in fabric topography was determined by
SEM and EDX analyses. Whether shrimp and crayfish
chitosan was successfully coated on the cotton fabric was
investigated by taking surface images at different
magnifications. SEM images were taken with ZEISS
LEVO LS 10 and FEİ QUANTA FEG 250 SEM
microscopes at 500X, 1000X and 2000X magnifications.
Dispersions of nano-Ag particles on the cotton fabric
surface were examined by energy dispersive x-ray
analysis.
Determination of the antibacterial activity of chitosan
+ nano-Ag treated cotton fabrics
The antibacterial efficacy of chitosan+nano-Ag coated
fabric samples against Escherichia coli ATCC 8739 was
tested by the JIS L 1902-2015 standard test method (JIS
L 1902). It is a quantitative method used to test the ability
of antibacterial finished fabrics to inhibit microbial
growth and kill microorganisms.
Two experimental groups as shrimp chitosan and
crayfish chitosan-coated samples were formed according
to chitosan source. These samples were then subdivided
into 2 groups with and without nano-Ag. The
antimicrobial activity of the fabric samples in each group
against E. coli was analyzed according to the following
method.
Samples (0.4 g in weight) were cut from the cotton
fabric coated with chitosan and nano-Ag and placed in test
tubes. Before the experiment, the fabric samples were
sterilized at 121 °C for 15 minutes. As the test method, the
absorption method was applied in which the bacterial
suspension was directly grafted onto the samples. The
fabric samples were then inoculated with the test
organism of 0.2 mL at a concentration of 2.9x105
CFU/mL grown in a liquid culture medium. Initial
microbial concentrations were determined at time zero.
Bacteria were counted according to Plate Count Method.
The control and test fabric samples inoculated with E. coli
were then incubated in closed containers at 37 ± 2 °C for
18-24 hours. After that, final bacterial concentrations
were determined. The reduction of bacteria in initial
concentrations and the control samples were calculated.
The bacterial growth rate for the control sample was
calculated according to the formula F = Ct-C0 and for the
test sample it was calculated according to the formula G
= Tt-T0, where
A is antibacterial activity,
C0 is the average logarithmic value of the untreated
sample immediately after inoculation (0 h contact time),
Ct is the average logarithmic value of the untreated
sample after 24 hours,
Tt is the average logarithmic value of the treated test
sample after 24 hours and
T0 is the average logarithmic value of the treated test
specimen immediately after inoculation (0 hours).
Antibacterial activity value was calculated according
to the formula A = F-G.
According to JIS L 1902 method, if 2≤A <3 then the
tested sample has an effect and if A≥3, then it has a strong
effect.
Results and Discussion
Chitosan as a binder
The crayfish and shrimp chitosan, used as binding
agents in the coating of cotton fabrics, were treated with
1% acetic acid to form water-soluble solutions. Although
there was no significant difference between the solutions,
chitosan produced from shrimp wastes was easier to
prepare. The shrimp chitosan produced a more
homogeneous solution than crayfish chitosan and it was
more easily applied to the fabric. However, the
distribution of nano-Ag particles on the fabric surface was
more homogeneous in the coating with crayfish chitosan
compared to coatings with shrimp chitosan. Analysis
results of crayfish chitosan coating were comparable to
the finding of Xu et al. (2019) who used carboxymethyl
chitosan to fix nano-Ag particles to the cotton fabric
surface. Crayfish chitosan was found to be a more suitable
to deposite nano-Ag particles on the fabric surface than
shrimp chitosan in terms of homogeneous distribution of
nanoparticles.
Textile Finishing with Chitosan and Silver Nanoparticles against E. coli ATCC 8739 25
Trakya Univ J Nat Sci, 21(1): 21-32, 2020
In coating with chitosan, the film layer on the fabric
surface was colorless and did not change the color of the
fabric. The film layer did not show adhesive properties
after drying. Furthermore, after treatment, the chitosan
solution was easily removed from the machine rollers.
The chitosan used as the binding agent had no adverse
effects on the binding quality of the nano-Ag particles to
the fabric. These are the expected properties of fiber-
bonding materials which provide the adhesion of the
finishing agent to the fiber in textile finishing processes.
For this reason, chitosan produced from shrimp and
crayfish waste shells by the chemical method were found
to be suitable for use as binder polymer in nano-Ag
coatings. The use of chitosan, which is a natural and
biodegradable polymer, as a binder in textile will enable
to minimize the negative effects of chemicals used in
finishing enterprises to the environment and contribute to
environmentally friendly textile production.
FTIR analysis results
FTIR analysis was performed to characterize the chitin
samples isolated from shrimp and crayfish waste shells
and to confirm whether the shrimp and crayfish chitosans
formed a bond between nano-Ag and cotton fabric. Chitin
is available in 3 different forms as alpha, beta and gamma
chitin in nature (Lavall et al. 2007). In the FTIR spectrum
of alpha chitin, the two absorption bands at 1650 and 1620
cm-1 refer to the stretching of amide I, and the absorption
band at 1550 cm-1 is attributed to amide II (N-H bending)
(Focher et al. 1992, Lavall et al. 2007). The band at 650
cm-1 was referred to the C = O group hydrogen-bonded to
N-H of the neighboring chain (Dahmane et al. 2016). In
our study, characteristic absorption bands showing
stretching of amide I of chitin were observed at 1653 and
1620 cm−1 for shrimp chitin and at 1655 and 1619 cm-1 for
crayfish chitin. Another characteristic band showing
amide II stretching was recorded at 1552 cm-1 for both
shrimp and crayfish chitin (Fig. 3). This also shows that
the isolated chitin is in alpha form.
Fig. 3. FTIR spectra of isolated chitin samples. a) from crayfish,
b) from shrimp.
Fig. 4. FTIR images of cotton fabric coated with nano-Ag
via chitosan. a) Shrimp chitosan+nano-Ag coated fabric, b)
Crayfish chitosan+nano-Ag coated fabric, c) Crayfish chitosan-
coated fabric and d) Shrimp chitosan-coated fabric.
The FTIR spectra of coatings using shrimp and crayfish
chitosan as binders are presented in Fig. 4. Previous studies
showed that the characteristic absorption bands of cellulose
in the FTIR spectra of cotton fabric were observed at 1430
cm-1 (C-H wagging), 1364 cm-1 (C-H bending), 1105 cm-1
(C-O-C, asymmetric bridge stretching), and 1160, 1060 and
1028 cm−1 (C-O stretching) (Xu et al. 2019). These peaks
were observed in our chitosan and nano-Ag coated fabric
samples at 1428-1428, 1361-1368, 1105-1109, 1158-1162,
1035-1055 and 1026-1030 cm-1, respectively. Mujtaba et
al. (2016) observed a sharp peak for cellulose at 3336 and
3330 cm-1 attributed to the hydroxyl group and hydrogen
bond, respectively. They also observed a peak at around
896 cm-1 correspondings to the structure of the glucose ring.
These peaks were observed in our samples at 3335 and
3336 cm-1, and between 894 and 896 cm-1, respectively.
The characteristic absorptive peaks for chitosan are I.
carbonyl (C = O) band around 1650 cm−1 and II. amide (NH
2) band around 1590 cm−1 (Chen et al. 2014). When
considering the FTIR spectra of the cotton fabrics coated
with nano-Ag via chitosan, it was seen that the peak at 1650
cm-1 was shifted to 1633, 1641 and 1647 cm-1 and the peak
at 1590 cm-1 was shifted to 1538, 1562 and 1550 cm-1.
Murugan et al. (2017) reported that the FTIR spectra of
Ag/chitosan composite differed from those of chitosan. The
characteristic peaks of chitosan at 1658 and 1600 cm-1
assigned to the stretching vibrations of amide C–O bonds
sharply reduced and shifted to 1628 cm-1. Another study
stated that the intensity of the peak at 3434 cm-1 referred to
the hydroxyl and primary amino groups decreased because
26 S. Erdoğan
the amino and hydroxyl groups of chitosan chelated the
nano-Ag particles (Chen et al. 2014).
When compared to the FTIR bands of crayfish and
shrimp chitosan-coated cotton fabric (Figs 4c, 4d),
changes were observed in the FTIR spectra of
chitosan+nano Ag coated fabric samples (Figs 4a, 4b). As
mentioned in previous studies, these shifting in the FTIR
peaks indicate that the formation of coordination bonds
between the amine groups of the chitosan and the nano-
Ag particles (Xu et al. 2019), or that Ag is chelated by
both amino and hydroxyl groups of the chitosan (Chen et
al. 2014). This shows that nano-Ag is chemically bonded
to cotton fabric via shrimp and crayfish chitosan.
SEM and EDX analysis results
Scanning electron microscopy was used to evaluate
the presence and bonding properties of chitosan and nano-
Ag particles on cotton fabric. SEM and EDX images of
cotton fabrics before and after the coating showed that
these fabrics were successfully coated with shrimp and
crayfish chitosan and nano-Ag (Figs 5, 6). Zhou & Kan
(2014) observed that while the SEM images of the fibers
in the pure cotton fabric were smooth, the SEM images of
the chitosan-coated cotton fibers were not smooth. Our
study indicates the same results. Also, in this study, the
surface of crayfish chitosan-coated fabric appeared to be
rougher (Fig. 5b) than the surface of shrimp chitosan-
coated fabric (Fig. 5c).
The shrimp and crayfish chitosans adhered to the
cellulose fibers and formed a film by surrounding the
nano-Ag particles (Figs 6a, 6d). Other studies have also
confirmed that chitosan forms a film on a cotton fabric
surface (Chattopadhyay & Inamdar 2013). In the coating
with crayfish chitosan, nano-Ag particles exhibited a
relatively homogeneous distribution (Fig. 6f), while nano-
Ag particles coated on the fabric surface with shrimp
chitosan appeared to aggregate (Fig. 6c). Govindan et al.
(2012) stated that the FESEM image of chitosan–Ag
nanocomposite showed that nano-Ag particles were
wrapped with chitosan and agglomeration was observed.
The authors also reported that agglomeration can be
prevented by slowly dissolving the chitosan a good while
and dispersing Ag in the chitosan with stirring for a longer
time and thereby, nano-Ag particles will be embedded in
a chitosan matrix.
Fig. 5. SEM images of cotton fabric surfaces. a) Uncoated fabric b) Crayfish chitosan-coated fabric c) Shrimp chitosan-coated fabric.
Fig. 6. SEM and EDX images of chitosan and nano-Ag coated cotton fabrics. a) SEM image of shrimp chitosan+Nano-Ag coated
fabric, b and c) EDX images of shrimp chitosan+Nano-Ag coated fabric (C, O, Ag elements and Ag element only), d) SEM image of
crayfish chitosan+Nano-Ag coated fabric, e) and f) EDX images of the crayfish chitosan+Nano-Ag coated fabric (C, O, Ag elements
and Ag element only).
Textile Finishing with Chitosan and Silver Nanoparticles against E. coli ATCC 8739 27
Trakya Univ J Nat Sci, 21(1): 21-32, 2020
Table 1. Elemental composition of chitosan-nano Ag coatings.
Element
Wt %
At %
K-Ratio
Z
A
F
Shrimp chitosan + Nano-Ag coating
C
46.22
53.40
0.2373
1.0076
0.5094
1.0004
O
53.73
46.60
0.1052
0.9935
0.1970
1.0000
Ag
0.046
0.006
0.0009
0.7667
2.7016
1.0000
Total
100.00
100.00
Crayfish chitosan + Nano-Ag coating
C
46.26
53.44
0.2377
1.0076
0.5096
1.0004
O
53.67
46.54
0.1049
0.9935
0.1968
1.0000
Ag
0.06
0.01
0.0013
0.7667
2.6772
1.0000
Total
100.00
100.00
EDX analysis was performed to observe the elemental
distributions on the surface of cotton fabrics coated with
chitosan and nano-Ag particles and to detect the presence
of nano-Ag particles on the fabric surface. Previous
studies reported that 3 keV peaks attributed to the Ag
formation signal were observed in the EDX analysis graph
of the nano-Ag deposited fabric (Gharibshahi et al. 2017,
Tania et al. 2019). In this study, the presence of the peak
of the Ag element in the EDX spectrum of cotton fabrics
confirmed the deposition of silver nanoparticles on the
fabric. Table 1 shows the distribution by weight of C, O
and nano-Ag particles on the fabric surface. In previous
studies, the % by weight of nano-Ag particles deposited
on the fabric surface was recorded as 0.006%
(Gharibshahi et al. 2017) and 0.10% (Arif et al. 2015). In
our study, the weight of nano-Ag on the surface of the
fabric was 0.046% in the coating with shrimp chitosan and
0.06% in the coating with crayfish chitosan. Although the
amount of nano-Ag particles used in coatings is same, it
appears that crayfish chitosan binds a higher amount of
nano-Ag particles to the fabric in comparison to shrimp
chitosan. This may be due to the acquisition of EDX
images from selected regions and the aggregation of
shrimp chitosan in some regions.
Antibacterial activity results of cotton fabric coated
with chitosan and nano-Ag
Antibacterial activity of cotton fabric coated with
chitosan and nano-Ag against E. coli ATCC 8739 was
investigated by JIS L 1902-2015 Quantitative standard
test method.
Various standard tests have been developed and used so
far to determine the antibacterial efficacy of textile
products. The most commonly used are qualitative tests
such as ISO 20645: 2004, AATCC 147: 2004, and halo
method of JIS L 1902: 2008, and quantitative tests such as
AATCC 100: 2004, absorption method of ISO 20743:
2007, and absorption method of JIS L 1902: 2008
(Palamutçu et al. 2008, Pinho et al. 2011). Qualitative tests
are inadequate to accurately determine the antibacterial
activity of fabric samples. In quantitative methods, the
reduction of bacterial growth is calculated by comparing it
to a control sample and the value of the antibacterial
activity can be accurately determined (Torlak 2008, Pinho
et al. 2011). Pinho et al. (2011) reported that the JIS L
1902-The adsorption method gives very sensitive and
accurate results. Palamutçu et al. (2008) stated that JIS L
1902-2002 standard is applied considering the moisture
content and nutrient amount on textiles under normal
clothing conditions. The authors also stated that tests are
performed in a situation similar to real-life conditions and
that the amount of nutrients in the medium is kept limited.
Thus, the antibacterial activity of textile samples with
antibacterial finishing is tested in an environment similar to
real-life conditions. Table 2 presents comparative
antibacterial activity values of cotton fabrics coated with
nano-Ag via chitosan according to JIS L 1902-2015.
The antibacterial activity value (A) of the fabric
samples covered with crayfish and shrimp chitosan were
calculated as 3.10 and 5.74, respectively, considering
bacterial growth rates. According to JIS L 1902-2015, if
A>3, the tested sample is considered to exhibit a very
good antibacterial activity (99.9% reduction). In this case,
the fabrics coated with crayfish and shrimp chitosan
showed high antibacterial activity against E. coli ATCC
8739. Besides, shrimp chitosan exhibited higher
antibacterial performance than crayfish chitosan. The
antibacterial activity values of cotton fabric samples
coated with nano-Ag via shrimp and crayfish chitosan
were calculated as 5.10 and 5.37, respectively. The
reduction ratio of 99.99% in bacterial viability indicated
that chitosan and nano-Ag composite coatings gave a
strong antibacterial activity to cotton fabrics. The
antibacterial activity values of fabrics coated with shrimp
chitosan+nano-Ag and crayfish chitosan+nano-Ag were
very close to each other. A significant improvement in the
antibacterial activity of coatings containing
chitosan+nano-Ag was observed in comparison to
crayfish chitosan-coated fabric sample. Interestingly,
shrimp chitosan coated fabric sample had the highest
bacterial reducing rate among other fabrics.
The biocompatibility, biodegradability, non-toxicity,
and antimicrobial and hypoallergenic properties of
chitosan make it a suitable antibacterial agent for fabric
surfaces (Zhou & Kan 2014). It has been reported that the
mechanism of the antibacterial action of chitosan is to
bind to the cell wall of the bacteria through its positively
charged amino groups and then to adhere to DNA and
prevent the proliferation of bacteria (Govindan et al.
2012, Zhou & Kan 2014). Results of various studies have
been published on determination of antibacterial activity
28 S. Erdoğan
Table 2. Antibacterial activities of chitosan and nano-Ag coated cotton fabrics.
Samples
Number of reproducing
microorganisms (CFU/sample)
Logaritmic reduction
Growth
value
Antibacterial
activity value
Contact time
(0 hours)
After incubation
(24 hours)
Log (C0)
and
Log (T0)
Log (Ct)
and
Log (Tt)
*F= Ct-C0
and
*G= Tt-T0
A=F-G
Uncoated cotton fabric
7x104
6.8x107
4.85
7.83
2.99
-
Shrimp chitosan and nano-
Ag coated fabric
5.14x104
4x102
4.71
2.60
-2.11
A=5.10
A>3
%99.99
Crayfish chitosan and
nano-Ag coated fabric
4.84x104
2x102
4.68
2.30
-2.38
A=5.37
A>3
%99.99
Shrimp Chitosan coated
fabric
4.1x104
2x102
4.61
2.30
-2.31
A=5.74
A>3
%99.99
Crayfish Chitosan coated
fabric
7.8x102
6x104
4.89
4.78
-0.11
A=3.10
A>3
%99.9
*F= Ct-C0: Growth value of uncoated fabric, *G= Tt-T0: Growth value of coated fabric
of chitosan against various bacterial species (Jiang et al.
2010, Velmurugan et al. 2014, Tania et al. 2019). A study
reported that chitosan-coated cotton fabric samples
effectively inhibited the growth of Staphylococcus aureus
and Klebsiella pneumoniae (Zhou & Kan 2014). Şahan &
Demir (2016) reported that both chitosan and nano
chitosan showed good antibacterial activity against S.
aureus, E. coli and, K. pneumoniae. The results of this
study indicated that crayfish chitosan showed high
antibacterial effeciency against E.coli. Chitosan source
affects the physico-chemical and biological properties of
chitosan (Kumirska et al. 2011). The results of this study
confirmed that the source of chitosan affects the
biological properties of chitosan. Shrimp chitosan showed
higher antibacterial performance than crayfish chitosan.
Nano-Ag particles have been widely used in the
development of antibacterial textiles as effective
antibacterial agents in recent years. Previous studies
emphasized that Ag showed high biocompatibility and
low toxicity with human cells in both ionic and colloidal
forms (Marambio-Jones & Hoek 2010). Since nano-Ag
particles have a large surface area compared to their
volume, they provide better contact with microorganisms
and thus show good antibacterial properties (Velmurugan
et al. 2014). A study reported that Nano-Ag particle
coated cotton fabric has an excellent bacterial reduction
for S. aureus (95%) and E. coli (92%) (Tania et al. 2019).
Another study reported that the bacterial reduction rates
of nano-Ag particle coated polyester fabrics were 99.7%
for E. coli and 99.8% for S. aureus (Jiang et al. 2010).
Both studies agree that nano-Ag particles exhibit
excellent bacterial reduction and this effect related to the
amount of nano-Ag particles deposited on the fabric
surface. Microbial growth reduction values of nano-Ag -
containing composites for S. aureus and Pseudomonas
aeruginosa were measured between 3.3-7.0 log CFU by
JIS L 1902 method (Wiegand et al. 2015).
Chattopadhyay & Inamdar (2013) reported that nano
chitosan exhibited better antibacterial activity when
treated with nano-Ag. Chen et al. (2014) reported that
Ag/chitosan composites exhibit higher antibacterial
activity against both the gram-positive bacteria S. aureus
and Bacillus subtilis and the gram-negative bacteria E.
coli and Salmonella choleraesuis than chitosan. The
authors concluded that as well as the homogenously and
well distribution of nano-Ag particles in the chitosan
matrix, also synergistic effects of chitosan and nano-Ag
particles caused Ag/chitosan composites to exhibit higher
antibacterial activity. Arif et al. (2015) measured the
antibacterial activity of fabric samples coated with
chitosan+nano-Ag particles against S. aureus and E. coli
and they found the bacterial reduction rates to be around
98-99%. The authors stated that there was only a slight
reduction in antibacterial activity even after 20 washes.
Xu et al. (2019) coated carboxymethyl chitosan/Ag
nanoparticle colloidal solution on a cotton fiber surface by
the pad-dry-cure method using carboxymethyl chitosan as
a binder. The authors stated that this coating was
successfully coated on the fabric surface and imparted to
the fabric significant antibacterial activity against S.
aureus and E. coli. They discovered that even after 50
washes, the fabric retains its antibacterial activity. The
results of relevant studies have shown that the application
of chitosan together with Ag particles increases the
antibacterial effect, and some results of our study are
consistent with these studies. Crayfish chitosan+nano-Ag
composite (A= 5.37) showed a higher antibacterial effect
than crayfish chitosan (3.10). On the other hand, shrimp
chitosan (A= 5.74) exhibited higher antibacterial activity
then shrimp chitosan+nano-Ag composite (A= 5.10).
Evaluation of the chitosan and nano-Ag coatings in
terms of human and environmental health
It is expected that antibacterial textile coatings are safe
for human health and environmentally friendly. Chitosan
is a biocompatible, biodegradable and non-toxic
Textile Finishing with Chitosan and Silver Nanoparticles against E. coli ATCC 8739 29
Trakya Univ J Nat Sci, 21(1): 21-32, 2020
glycopolymer of biological origin. These properties of
chitosan, which are used for treatment in many medical
applications, have been emphasized in many studies (Li et
al. 2016, Liang et al. 2018, Zhao et al. 2018). Chitosan is
degradable within the body and it is safe and non-toxic
(Dutta et al. 2004). In vertebrates, it is known that
chitosan is mainly degraded by lysozyme and certain
bacterial enzymes in the large intestine (Dash et al. 2011).
It can also be degraded by many microorganisms in
nature. Nano-Ag particles are also not prohibited and are
one of the most suitable and commercially distributed
nanomaterials in the world (Korani et al. 2015). Nano-Ag
particles are widely used in medical and functional
textiles, wound dressings, medical devices implanted for
a long time, dental materials, water disinfectants, room
sprays, laundry powders and deodorants due to their
antibacterial and deodorizing properties (SCENIHR 2014,
Korani et al. 2015, Burdusel et al. 2018). However,
researches on the effects of nano-Ag particles on human
health are limited (Korani et al. 2015). In vitro studies
revealed that nano-Ag particles may show cytotoxic and
genotoxic effects (SCENIHR 2014). Hartemann et al.
(2015) reported that in vitro studies showed that
nanosilver induced cytokine production in macrophages.
Since there are few studies on the genotoxicity of nano-
Ag particles in vivo, it has been reported that the results
of the present studies can not confirm the positive or
negative effects of nano-Ag particles (SCENIHR 2014).
The SCENIHR report states that the results of the studies
are contradictory and some of them said that nano-Ag
showed high cytotoxicity at doses between 2-5 μg/mL,
while the others showed almost no cytotoxicity at doses
up to 100 μg/mL (SCENIHR 2014). Korani et al. (2013)
showed that nano-Ag particles cause histopathological
abnormalities in spleen, liver, and skin in animal
experiments. In another study, colloidal nano-Ag particles
were reported to be capable of producing a dose-
dependent toxic response in several organs (Korani et al.
2015). However, many studies point out that there is a
lack of consistent and reliable data on the toxicity and
biological behavior of nano-Ag particles in both in vitro
and in vivo toxicity studies and further studies are needed
to produce meaningful results (Korani et al. 2015,
Hartemann et al. 2015, Burdusel et al. 2018). Researchers
concluded that many parameters including deposition
rates, particle size, surface area, dose taken, interaction
with biological macromolecules, dispersion ratio,
concentration, surface charge, morphology, surface
oxidation, and conversion under biological conditions
affect the toxicity of nano-Ag particles (SCENIHR 2014,
Hartemann et al. 2015, Korani et al. 2015, Burdusel et al.
2018). Researchers emphasize the necessity of long-term
studies with wide-range doses and different particle sizes
to accurately determine the effects of nano-Ag particles
on human health, (SCENIHR 2014, Korani et al. 2015).
When the published data on the subject is examined, it
is seen that the findings related to the genotoxic and
cytotoxic effects of nano-Ag particles are generally
related to the intake of these nanomaterials as food
supplements or drugs. However, in antibacterial textile
products, nano-Ag is used on the fabric surface and it is
only in contact with the skin. As in this study, it is
generally used in low doses. In this study, SEM analyzes
of chitosan and nano-Ag coated fabrics revealed that
chitosan forms a film that encapsulates nano-Ag particles
and fixes them to the fabric. The nanoparticles are
embedded in the chitosan matrix. This is also noted in
previous studies (Govindan et al. 2012, Chattopadhyay &
Inamdar 2013). Chitosan is already a natural biomaterial
and it prevents direct contact of nano-Ag on the fabric
surface with the skin. Thus, it prevents possible toxic
effects by preventing or limiting the penetration of nano-
Ag particles into the body. Edward-Jones (2009) stated
that although, Ag element has been used intensively in the
treatment of burns for 50 years, there are limited reported
cases of Ag toxicity. The author states that Ag poisoning
is dependent on Ag levels absorbed into the body in time
and the amount absorbed by intact skin are lower than that
absorbed by open wounds. Furthermore, SCENIHR
(2014) reports that the uptake of nanomaterials via the
skin is generally very low. Considering all these data, it
can be concluded that the chitosan/nano-Ag coatings are
safe for human health and environmentally friendly
because they minimize the use of chemicals.
Little is known about the environmental impacts and
hazards of nano-Ag particles in aquatic systems and it is
not possible to draw general conclusions (SCENIHR
2014, Korani et al. 2015). Hartemann et al. (2015) state
that nano-Ag particles released into the environment have
a transformation such as aggregation, agglomeration,
dissolution or silver chloride and silver sulfide formation.
The author says that the chemical species of the
transformed nano-Ag determines the bioavailability and
toxicity of Ag in nature. He also stated that the effects of
nano-Ag particles on soil can be vary depending on both
nanoparticle and soil properties. As for water sources, Ag
can be used to control the bacteriological quality of
drinking water. The WHO (2017) report states that
although there is insufficient data to obtain a health-based
guide for Ag in drinking water, Ag levels up to 0.1 mg/L
can be tolerated.
Conclusion
Chitosan formed a colorless film and a matrix
enabling nano-Ag particles to deposit homogeneously on
the fabric surface. Chitosan source was effective in the
quality of chitosan coatings. Crayfish chitosan may be
more suitable for more homogeneous and thinner
coatings. These properties are sought after in textiles and
indicate that chitosan is appropriate to be used as a
finishing agent in textiles. Also, the method used to coat
fabrics is applicable to the long length and it is a useful
method. It can be applied to the industry due to its ease of
use and low cost.
Since the JIS L 1902 method allows comparative
evaluation of the antibacterial activity of treated and
untreated fabrics, it increases the reliability of the results
and is considered to be a suitable method for testing
30 S. Erdoğan
antibacterial activity. A very strong antibacterial effect
(99.99% reduction) was obtained against E. coli bacteria
on pre-treated cotton fabric surfaces with the effect of
both nano-Ag particles and chitosan. Chitosan nano-Ag
coating which exhibited a good antibacterial activity can
be used in the manufacture of garments such as medical
textiles, baby clothes, and underwear because of its
sterility, due to its biocompatible, biodegradable, non-
toxic and sterilizable properties.
Chitosan nano-Ag coating used as finishing
contributes to the protection of human health by reducing
the use of antibiotics. The evaluation of shrimp and
crayfish wastes as chitosan also contributes to the
protection of the environment. Besides, the chemical ratio
can be reduced by the use of chitosan instead of binder,
which is commonly used in textile printing, pigment
dyeing in finishing materials. Purification processes can
be reduced and thus cost is reduced. By structuring a clean
production approach in textile enterprises and sustainable
use of natural resources, it is possible to prevent or reduce
pollutant discharges and emissions from industrial
sources. Thus, this approach offers innovative solutions
for the protection of human and environmental health.
Acknowledgement
This study was supported financially by Trakya
University Reseach Fund with the project TUBAP
2018/198. The analyses were performed in the Trakya
University Technology Research Development
Application and Research Center (TUTAGEM)
laboratories.
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