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Characterization and antimicrobial properties of cotton fabric loaded with green synthesized silver nanoparticles
Accepted Manuscript Title: Characterization and antimicrobial properties of cotton fabric loaded with green synthesized silver nanoparticles Author: Haytham M.M. Ibrahim Mahmoud S. Hassan PII: DOI: Reference:
S0144-8617(16)30563-X http://dx.doi.org/doi:10.1016/j.carbpol.2016.05.041 CARP 11110
To appear in: Received date: Revised date: Accepted date:
1-3-2016 28-4-2016 12-5-2016
Please cite this article as: Ibrahim, Haytham MM., & Hassan, Mahmoud S., Characterization and antimicrobial properties of cotton fabric loaded with green synthesized silver nanoparticles.Carbohydrate Polymers http://dx.doi.org/10.1016/j.carbpol.2016.05.041 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
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Characterization and antimicrobial properties of cotton fabric loaded with green synthesized silver nanoparticles
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Haytham M. M. Ibrahima* [email protected], Mahmoud S. Hassanb
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a
Department of Radiation Microbiology, National Center for Radiation Research and Technology (NCRRT), Atomic Energy Authority, Cairo, Egypt b
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Department of Radiation Chemistry, National Center for Radiation Research and Technology (NCRRT), Atomic Energy Authority, Cairo, Egypt
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* Corresponding author: Haytham M. M. Ibrahim, Ph.D. Radiation Microbiology Department, National Center for Radiation Research and Technology (NCRRT), Atomic Energy Authority, Egypt, Address: 3 Ahmed El Zomor St., Nasr City, P.O. Box 29, Cairo, Egypt, Tel.: +202 22748246 Fax: +202 22749298
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Highlight
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Silver nanoparticles (AgNPs) were prepared by eco-friendly, safe, and cost effective method using fungal filtrate.
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Antimicrobial formulation encompasses AgNPs and binder was applied to cotton fabric.
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Both AgNPs and treated cotton fabric were characterized.
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The antimicrobial activity of treated fabric was assessed qualitatively and quantitatively.
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Durability of antimicrobial properties was demonstrated after repeated washing cycles.
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1. Introduction
41 42
Cotton fabrics are prevalent with people due to their superior characteristics such as
43
biodegradation, regeneration, affinity to skin, softness, and hygroscopic property (Lim and
44
Hudson, 2004). Cotton fibers are in particular appropriate for industrialization of textiles for
45
sports, medical, and healthcare/hygiene products (Czajka, 2005). Clothes and other textile
46
materials, especially those made of natural fibers such as cotton and wool, considered as good
47
media for the growth of pathogenic or odor-generating microorganisms. They offer a perfect
48
environment for the growth of bacteria and fungi due to their large surface area and capability to
49
keep oxygen, humidity, heat, and nutrients from the body exudates (Dev et al., 2009). Therefore,
50
with the increasing concern for individual health and hygiene, textiles with antimicrobial
51
activities are becoming an increasingly desirable objective of textile manufacturers.
52
Antimicrobial finishes are applied to textiles for three essential goals: (1) to control the
53
propagation of disease and preclude the danger generated by injury infection, (2) to prevent the
2
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development of odor from aspiration, stains, and soil on textile materials and (3) to suppress the
55
damage of textiles caused by decomposition, especially textiles made of natural fibres (Gao and
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Cranston, 2008). Currently, there is an increasing attention to produce non-toxic, durable, cost-
57
effective, and efficient antimicrobial textiles with expanded applications in health care, hygienic
58
products, medical, and protective textile materials (Gao and Cranston, 2008). The antimicrobial
59
agents that have been used industrially included quaternary ammonium salts, metal salts
60
solutions, and antibiotics. Regrettably, some of these substances are toxic or poorly effective,
61
which renders them not appropriate for application in, filters, textiles, health foods and for the
62
exclusions of pollution. On the other hand, employment of nanoparticles and their
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nanocomposites would be good alternatives (Grace and Pandian, 2007). They can offer a new
64
opportunity for antimicrobial and multifunctional modification of textiles.
65
Despite the outstanding antimicrobial activity of silver nitrate, it’s not suitable for the
66
application to textile materials since it stains to black–brown when exposed to light and air,
67
because of uncontrolled reduction processes (Vigneshwaran et al., 2006). However, at defined
68
concentrations, a significant antimicrobial activity with an acceptable color change can be
69
achieved by using silver nanoparticles (AgNPs) deposited on the textile materials. The
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antimicrobial activity of AgNPs is affected by the size of the particles, i.e. the smaller the
71
particles the greater the antimicrobial effect (Morones et al., 2005). At the nanoscale, AgNPs
72
exhibit remarkably unusual, chemical, physical, and biological properties, as well as
73
antimicrobial activity (Chen and Schluesener, 2008). Chemical synthesis of nanoparticles has
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several hazards, like genotoxicity, cytotoxicity, carcinogenicity, and general toxicity (Mukherjee
75
et al., 2008). Therefore, there is a need to develop clean, non-toxic, and environment-friendly
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methods for synthesis of nanoparticles (Mukherjee et al., 2008).
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In the present study, AgNPs were prepared biologically employing the fungus Alternaria
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alternata. The synthesized nanoparticles were characterized by scanning electron microscopy
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(SEM), energy dispersive X-ray spectroscopy (EDX), transmission electron microscopy (TEM),
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and dynamic light scattering (DLS). The prepared nano-silver was applied to then cotton fabric.
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The treated cotton fabric was characterized by thermogravimetric analysis (TGA), SEM, and
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mechanical properties. The resistance of treated fabric to biodegradation, their antimicrobial
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properties, both qualitatively and quantitatively, against representative microorganisms, as well
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as durability of the antimicrobial effects, after repeated washing cycles, were demonstrated.
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2. Materials and methods
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2.1. Cotton fabric and chemicals
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Plain-weave cotton fabric (1mm thickness) were kindly provided by El-Nasr Company for
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spinning, weaving, and dyeing, El-Mahalla El-Kubra, Egypt. The fabric was scoured and not
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subjected to any finishing processes. The Impron MTP binder (butyl acrylate) as a crosslinking
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agent and Genapol BE 2410 as a non-ionic detergent were kindly supplied by Clariant, Germany.
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Silver nitrate (AgNO3) was purchased from Sigma, USA and used as received without further
95
purification, other chemicals used were of laboratory grade.
96 97
2.2. Microorganisms
98
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The fungus Alternaria alternata was used to prepare AgNPs. Four bacterial strains, Bacillus
100
subtilis local isolate and Staphylococcus aureus ATCC 6538 as Gram-positive bacteria and
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Pseudomonas aeruginosa ATCC 9027 and Escherichia coli ATCC 8739 as Gram-negative
102
bacteria, in addition to the fungus Aspergillus niger were employed to evaluate the antimicrobial
103
activity of the treated textile fabric. The standard bacterial strains were obtained from
104
Microbiologics, Inc. Minnesota, USA. Bacterial strains were maintained on nutrient agar slants
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and the fungus was maintained on potato dextrose agar (PDA) slants at 4 oC.
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2.3. Preparation of AgNPs
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The fungus A. alternata was inoculated into 500-ml Erlenmeyer conical flasks containing 200 ml
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of the mineral salts medium described by Kathiresan et al., (2009), with the following
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composition in (g/l): potassium dihydrogen phosphate (KH2PO4), 7.0; dipotassium hydrogen
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phosphate (K2HPO4), 2.0; magnesium sulfate (MgSO4), 0.1; ammonium sulfate [(NH4)2SO4],
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1.0; yeast extract, 0.6; and glucose, 10.0, pH was adjusted to 6.5-7 using 1N solutions of HCl and
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NaOH. Inoculated flasks were incubated at 28 oC and 150 rpm for 72 h in a rotary shaker
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incubator (Gallen Kamp, UK). The mycelial biomass was harvested by filtration through
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Whatman No.1 filter papers and extensively washed with distilled water to eliminate residual
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medium. The biomass was taken into a flask containing 100 ml distilled water and incubated for
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72 h at 28 oC. Thereafter, the aqueous solution (fungal filtrate) was separated by filtration and
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used for the synthesis of AgNPs. For reduction, AgNO3 was added to 100 ml of the biomass
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filtrate at a concentration of 1, 2, 3, 4, and 5 mM (Jaidev and Narasimha, 2010). The mixtures
123
were kept at room temperature for 48 h. Flasks with either fungal biomass filtrate or AgNO3
124
solution, served as positive and negative controls, were run simultaneously. The reduction of Ag+
125
into Ago (AgNPs) was monitored by using UV–Vis spectrophotometer (Helios Gamma, Thermo
126
Corporation, England).
127 128
2.4. Characterization of AgNPs
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The average size of obtained AgNPs was estimated by DLS using zeta potential/particle sizer
131
spectrophotometer (NICOMP 380-ZLS, USA). The shape was determined by using TEM
132
(JEOL-JEM-1200 EX, Japan), operating at accelerating voltage of 80 kV. TEM samples were
133
prepared by dropping 2 drops of the colloidal AgNPs solutions onto 200 mesh carbon-coated
134
copper grids. The samples were air dried overnight prior to imaging the particles. The elemental
135
profile of AgNPs was determined by using EDX spectroscopy.
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2.5. Silver nanoparticles loading on the textile fabric
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Before being used, cotton fabric was washed and dried. Antimicrobial formulations, consist of
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colloidal solutions of AgNPs with different concentrations (1, 2, 3, 4, and 5 mM), and the binder
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(10 %), were treated with ultrasound before being used in the finishing process, to ensure
142
homogeneous dispersion and deagglomeration of nanoparticles. Cotton fabric samples (5 x 8 cm)
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were immersed in the antimicrobial formulation with stirring and rotation and of the finishing
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bath for 5 min. The samples were then squeezed, air-dried at room temperature. For fixation of
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AgNPs on the fabric surface, the fabric samples were subjected to a curing process, utilizing
146
gamma-irradiation or a thermal method. Irradiation to the required doses (5, 10, 30, and 50 kGy)
147
was carried out in a 60Co gamma irradiation facility at NCRRT, the dose rate at the experiment
148
time was 0.276 kGy/h. In contrast, thermal curing was achieved by placing the samples in an
149
oven at 160 oC for 3 min.
150 151 152 153 154 155
2.6. Characterization of AgNPs loaded cotton fabric
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2.6.1. Thermogravimetric analysis (TGA)
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The thermogravimetric analysis (TGA) of control, binder coated, and AgNPs coated cotton
159
fabric was performed by using TGA-30 (Shimadzu, Japan) at a heating rate of 10oC/min in the
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air at different temperatures ranged from room temperature up to 600 oC.
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2.6.2. Scanning electron microscopy (SEM)
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The surface morphology of AgNPs loaded cotton fibers, as well as the untreated control fabric,
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was imaged at different magnifications by using SEM (JSM-5400, JEOL, Japan), working at an
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accelerating voltage of 30 kV. Before imaging, the dried sample was sputter-coated with gold
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using a microscope sputter coater.
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2.6.3. Mechanical properties
169
The mechanical properties [tensile strength and elongation to break (%)] of treated and untreated
170
cotton fabrics were evaluated before the contact of the different systems with the soil and after
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burring in moist soil for 14 days, were tested at room temperature, according to the standard test
172
method for breaking strength and elongation of textile fabrics (strip test); ASTM D5035-95,
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using Mecmesin tester (Mecmesin Limited, UK), employing a crosshead speed of 50 mm/min
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and equipped with software. In this system, the mechanical parameters were calculated directly.
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Every data point is the average of 5 measurements.
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2.6.4. Color-difference measurements
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A microcolor unit, attached to a data station manufactured by Dr. Lange, Germany, was used for
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color strength measurements. The L*, a* and b* interceptions used in this system are based on
181
the CIE-color triangle (Commission International de l'Eclairage units x, y and z). In this system,
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the L* value represents the dark-white axis, a* represents the green-red axis and b* represents
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the blue-yellow axis. The recorded values of the different color interceptions are the average of 5
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measurements in different positions along the investigated samples. The L*, a*, and b* values of
185
the untreated samples were measured and taken as a reference, the total color differences (∆E*)
186
of the AgNPs treated samples were determined according to the following equation:
187
∆Ε ∗ =
( ∆ L∗ )2 + ( ∆ a ∗ )2 + ( ∆ b ∗ )2
(1)
188
Where ∆ L* is the color lightness difference between the fabric loaded with AgNPs and the
189
control sample; ∆ a* is the red/green difference between the treated and the control samples; ∆ b*
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190
is the yellow/blue difference between the treated and the control samples (Ilić et al., 2009). The
191
recorded values of the different color interceptions are the average of ten measurements.
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2.7. Antimicrobial activity of AgNPs loaded textile
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The antimicrobial activities of AgNPs loaded cotton fabric were assessed qualitatively and
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quantitatively, the untreated cotton fabric was used as a control. In addition, the resistance of the
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AgNPs coated textile fabric together with uncoated fabric (control) to the action of soil
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microflora was demonstrated by the soil burial test.
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2.7.1. Qualitative method using agar diffusion assay
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The bacterial strains under study were grown overnight in nutrient broth medium (Oxoid), then,
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the cultures broth (containing ~108 CFU/ml) were swabbed on nutrient agar plates using sterile
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cotton swabs. AgNPs loaded textile fabric was cut into small pieces (1cm x 1cm, 1mm
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thickness), then, treated and untreated (control) samples were planted on the nutrient agar plates.
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After being incubated at 32 oC for 24 h, the plates were examined for possible clear zone
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formation (Ravindra et al., 2010). The presence of clear zone was recorded as an inhibition
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against the tested microorganism. The extent of inhibition zone diameter was taken as a relative
209
measure of the antimicrobial effectiveness. The means of three replicates were tabulated.
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2.7.2. Quantitative method using percentage reduction of bacterial count
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The antimicrobial activity of the treated fabric was tested against E. coli (Gram-negative) and S.
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aureus (Gram-positive) bacteria following the method described by Perelshtein et al., (2009).
214
The bacterial strains were grown in a nutrient broth medium at 32 °C overnight. Next day, they
215
were transferred into fresh medium at an initial optical density (O.D600) of 0.1. When the culture
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reached an O.D600 of 0.3, the cells were harvested by centrifugation and washed twice with
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sterile saline solution (NaCl 0.9 %, pH 6.5). Then, the cells were diluted to a final concentration
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of 0.65 (O.D600) with saline solution. A 500 µl of the cells suspensions was pipetted into test
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tubes, each containing 4.5 ml of sterile saline solution along with AgNPs treated or untreated
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fabric samples; each of 1 cm2. The initial bacterial concentration was approximately 107 colony-
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forming units (CFU)/ml. The test tubes were incubated for 24 h at 32 °C and 150 rpm in a rotary
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shaker incubator (Gallen Kamp, UK). A 1 ml was taken at 0 h and after 24 h and serially diluted
223
up to 108 fold. Then, 100 µl of the appropriate dilutions was spread onto nutrient agar plates. The
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plates were incubated at 32 °C for 24 h. The viable bacterial count (CFU/ml) was determined and
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the antimicrobial activity was expressed as the percentage reduction (R, %) using the following
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formula. R ( %) =
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Co – C ×100 Co
(2)
228
Where R is the percentage reduction, Co is the number of bacterial colonies from the untreated
229
fabric, and C is the number of bacterial colonies from the treated fabric (Ilić et al., 2009).
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2.8. Soil burial test
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The resistance of AgNPs loaded and unloaded textile fabric to the actions of soil microflora was
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evaluated by soil burial test according to the method of Mitchell et al., (2012). The samples were
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buried in a naturally fertile top soil, placed in a dry wooden box to a depth of 13 cm. The soil
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was moisturized by gradual addition of water accompanied by mixing. Uniform moisture content
238
was maintained by covering the soil container with a suitable lid. Before burying, the treated
239
samples were first moistened with an aqueous solution containing 0.05 % of a non-ionic wetting
240
agent. The samples were then buried horizontally. Precautions were taken to ensure uniform
241
covering with the soil along the length of the fabric samples. The soil moisture content was
242
maintained between 20 and 30% (based on the dry weight) and the temperature was kept at 28 oC
243
throughout the burying time (14 days). Later, the samples were removed from the soil, slightly
244
rinsed with tap water, and air-dried at room temperature. The mechanical properties [tensile
245
strength and elongation at break (%)] of the fabric samples were evaluated. Moreover, their
246
resistance to the action of soil microflora, in terms of biodegradation, was determined by using
247
SEM.
248 249
2.9. Durability of the antimicrobial finishing for washing
250 251
To evaluate the durability of AgNPs-treated cotton fabric toward repeated laundering, the
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samples treated with the antimicrobial formulation followed by thermal or γ-irradiation curing
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were submerged in a washing solution containing non-ionic detergent (2 g/l). The samples were
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stirred for 15 min at 50 oC. Then, the samples were rinsed with tap water and air dried. This
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procedure was repeated 1, 10, and 20 times. Later, the antimicrobial properties of the washed
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samples were determined.
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257 258
3. Results and discussions
259 260
3.1. Synthesis and characterization of AgNPs
261 262
In the present work, AgNPs were prepared by a green biological method using the biomass
263
filtrate of fungus A. alternata. This process has many advantages: (a) it is an environmentally
264
safe technique, (b) no need to use additional reducing agents or stabilizers, (c) it can be
265
conducted at room temperature, and (d) the developed AgNPs have excellent characteristics,
266
such as dispersion stability over a long time. As depicted in Fig. 1, upon the addition of AgNO3
267
aqueous solution (Fig. 1 a) to the pale yellow biomass filtrate solution (Fig. 1 b) the solution’s
268
color turned to reddish brown (Fig. 1 c). This color was mainly due to the surface plasmon
269
resonance of the AgNPs. The successful fabrication of AgNPs was confirmed by UV˗Vis
270
spectroscopy, where strong broad peaks located at λmax = 430 nm with an absorption tail in the
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longer wavelengths were detected (Fig. 1). It is well known that AgNPs display ruby red color in
272
aqueous solutions, having a strong absorbance band around 400 - 450 nm emanating as a result
273
of surface plasmon excitation vibrations in the metal nanoparticles (Duran et al., 2007). This
274
peak is well documented for various metal nanoparticles with size range (2˗100 nm) (Shervani
275
et al., 2008). As illustrated in Fig. 1, the peak intensity, which represents the AgNPs
276
concentration, was increased with increasing the AgNO3 concentration (1-5 mM). The TEM
277
micrograph revealed that the individual AgNPs were spherical in shape with a mean diameter of
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∼25–30 nm (Fig. 2a). The DLS technique showed relatively larger particle size, i.e. 35±20 nm
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(Fig. 2b). The EDX spectrum of AgNPs showed signal characteristic for Ag+, as seen in Fig. 2c,
280
which confirms the presence of silver metal. Generally, metallic silver nanocrystallites reveal
281
ideal optical absorption peak approximately at 3 keV due to their surface plasmon resonance
282
(Kalimuthu et al., 2008).
283 284
3.2. Characterization of AgNPs treated cotton fabric
285 286
3.2.1. Thermal stability
287
The thermal stability of untreated cotton fabric, binder coated fabric, and fabric treated with the
288
antimicrobial formulation, containing a binder (10%), based on the total volume of treatment
289
solution, and AgNPs (1mM), followed by thermal curing at 160 oC for 3 min, have been
290
investigated by thermogravimetric analysis. The TGA thermograms were shown in Fig. 3. It
291
could be noticed that, the thermal degradation of the coated fabric with binder only begins at a
292
lower initial temperature than the uncoated cotton fabric, in which the initial temperature of the
293
overall degradation was ~238 °C, i.e. lower than the uncoated cotton fabric by ~50 °C,
294
accompanied with wt. loss (%) of 5% and 19%, for the uncoated and coated fabrics with acrylate
295
binder, respectively. These results could be attributed to the presence of acrylate binder thin film
296
which degrades faster than the textile component, reducing the initial degradation temperature
297
(Alongi et al., 2012). The addition of AgNPs to the coating formulation showed slight increases
298
in the thermal stability more than that of the fabric coated with acrylate binder, but still less than
299
the untreated cotton fabrics, with a wt. loss of 13% which improves the binding of AgNPs to the
300
cotton fabric encapsulated thought the acrylate binder, causing the increasing of the total thermal
301
stability more than of the fabrics coated with binder only (Dhas et al., 2015).
13
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For all fabric samples, the thermal degradation undergoes three steps. The first step illustrates the
303
water elimination from cotton fabrics, besides the initial degradation of the acrylate binder of
304
treated fabrics, which begins at 325, 238, and 238 °C, with a maximum wt. loss of 33, 37, and
305
33% for cotton, cotton/binder, and cotton/binder/AgNPs, respectively. The second step
306
corresponds to the degradation of the cross-linked acrylate binder, which begins at 385, 365, and
307
385 °C with a maximum wt. loss of 32% for cotton, cotton/binder, and cotton/binder/AgNPs,
308
respectively. The third step is accompanied with the degradation of the textile component, which
309
begins at 446 °C with a maximum wt. loss of 97% for all samples. Table 1 illustrated the total
310
weight loss (%) at different temperatures, in which the temperatures of maximum rates of the
311
thermal decomposition (Tmax) were obtained at 399±2.8, 358±3.1, and 377±4.2 oC for cotton,
312
cotton/binder, and cotton/binder/AgNPs, respectively.
313 314 315
3.2.2. Surface morphology
316
To demonstrate that the AgNPs were deposited on the cotton fibers, the fabric before and after
317
antibacterial treatment, with a formulation containing 10 % binder and 1mM AgNPs followed by
318
thermal curing at 160 oC for 3 min, were scanned under the SEM (Fig. 4). The SEM
319
photomicrographs revealed that the fibers of control cotton fabric (Fig. 4a) and fabric treated
320
with the binder only (Fig. 4b) exhibiting uniform neat plain spun structures and the cotton fibers
321
manifested smooth surfaces. Whereas, AgNPs treated cotton fibers clearly became more coarse
322
and exhibited characteristic semi-granulated pattern (Fig. 4c,d), due to the formation of a thin
323
layer around the fibers, composed of the binder and AgNPs encapsulated in the treatment
324
formulation. The images demonstrated also that the AgNPs were spherical in nature. Clusters of
14
325
AgNPs are demonstrated on the fabric surface due to the agglomeration process. Agglomeration
326
of AgNPs on the cotton fibers could not be prohibited in spite of the utilization of ultrasound
327
processing before finishing and by the implementation of the finish by the exhaustion method,
328
which is guaranteed by stirring and rotation of the finishing bath. The elemental profile of
329
AgNPs treated fabric showed higher counts at 3 keV due to silver, which confirms the presence
330
of AgNPs as indicated in Fig. 1.
331 332
3.2.3. Color differences of AgNPs coated cotton fabric
333
The data summarized in Table 2 represent the effect of increasing AgNPs concentration on the
334
color changes of the treated cotton fabric, which was quantitatively evaluated by the CIE L*, a*,
335
and b* color parameters, and the total color differences (∆E*). As given in Table 2, the cotton
336
fabric treated with binder only was a white/cream color, as indicated by the high value of L*
337
coordinate (whiteness), this value decreased progressively with introducing increased
338
concentrations of AgNPs (1˗5 mM). In contrast, the values of a* (red) and b* (blue) coordinates
339
increased. Therefore, the value of the total color difference (∆E*) increased from 1.8±0.3 to
340
41.8±1. The AgNPs/binder/cotton fabric composites color changed from white/cream (untreated
341
fabric) to yellowish red to fade brownish red to dark brownish red to fade bluish red to dark
342
bluish red. This color is attributed to the surface plasmon resonance effects of AgNPs, the
343
successive change in color resulting from an increase in the size of the nanoparticles as more Ag0
344
is formed on the nanoparticle surface with increasing the AgNPs concentration (Kelly and
345
Johnston, 2011).
346 347
3.3. Antimicrobial properties of AgNPs coated fabric
15
348 349
As shown in Table 3 the AgNPs treated cotton fabric revealed high antimicrobial activity at all
350
the tested concentrations, whereas the untreated fabric did not show any activity. The lowest
351
AgNPs concentration (1mM) was found to be sufficient to inhibit the growth of tested
352
microorganisms as indicated by the inhibition zone diameter (13-17 mm). Further increase in
353
AgNPs concentration was insignificant, as revealed by the trivial increase in inhibition zone
354
diameter (20 mm at 4 and 5 mM). In general, the antibacterial activity was higher against E.coli
355
than S. aureus. The AgNPs containing cotton fibers developed in the present work have
356
exhibited inhibition zone > 1.5 mm in all cases. These results indicate that they are highly
357
important, from the technological point of view, for establishing antibacterial finishing (Ravindra
358
et al., 2010).
359
The quantitative antimicrobial activity of the loaded fabric was studied against E. coli (Gram-
360
negative bacterium) and S. aureus (Gram-positive bacterium). The viable bacteria were
361
monitored by counting the number of CFU/ml on nutrient agar plates. As shown in Table 4,
362
treatment for 24h with the loaded fabric achieved 99.9 % inhibition of E. coli and S. aureus
363
growth. Upon increasing the concentration of the applied nano-silver colloidal solutions from 1
364
to 5 mM, the growth of S. aureus and E. coli was completely inhibited, i.e. 100%. These results
365
agree with that obtained by the agar diffusion method. The control fabric did not show any
366
antibacterial activity. In agreement, Zhang et al., (2009) reported that the silver-treated cotton
367
fabric showed 99.01% and 99.26% bacterial reduction of S. aureus and E. coli, respectively,
368
while the silver content of cotton was about 88 mg/kg. The bacterial reduction rates increased
369
slightly from 99.01% to 100% with the increase of the silver content of the silver-treated cotton
370
fabric to 158.49 and 173.62, respectively.
16
371
Based on the results of Tables 2, 3, and 4, it apparent that the AgNPs concentration of 1 mM is
372
sufficient and efficient concentration, which gives qualitatively noticeable inhibition zone and
373
quantitatively an excellent microbial reduction (99.9%) against the tested bacterial strains,
374
accompanied with lower and acceptable color change of the treated cotton fabric.
375 376
3.4. Mechanism of antimicrobial activity of AgNPs
377 378
Silver nanoparticles have been reported to exhibit antimicrobial properties (Virender et al.,
379
2009). Using AgNPs leads to increasing the number of particles per unit area and, thus, the
380
antibacterial effects can be maximized (Yeo and Jeong, 2003). The antibacterial property of
381
AgNPs treated cotton fabric is related to their Ag contents. AgNPs are capable of generating
382
reactive oxygen species (ROS), such as superoxide (•O2−), hydrogen peroxide (H2O2), and
383
hydrogen radical (•OH) from its surface. Several main mechanisms underlie the biocidal
384
properties of AgNPs against microorganisms have been postulated. (a) Ag0 nanoparticles attach
385
to the negatively charged cell surface, alter the physical and chemical properties of the cell
386
membranes and the cell wall and disturb important functions such as permeability,
387
osmoregulation, electron transport, and respiration. Besides, they increase the cell permeability
388
and leak the intracellular contents by cell disruption (Kora and Arunachalam, 2011), (b) they can
389
cause further damage to bacterial cells by permeating the cell, where they interact with DNA,
390
proteins, and other phosphorus- and sulfur-containing cell constituents. It is believed that the
391
high affinity of silver towards sulfur or phosphorus is the key element of this effect. Due to the
392
abundance of sulfur containing proteins on the bacterial cell membrane, AgNPs can react with
393
sulfur-containing proteins inside or outside the cell membrane, which in turn affects bacterial cell
17
394
viability (Morones et al. 2005), (c) silver ions (particularly Ag+) released from (Ag0) can interact
395
with phosphorus moieties in DNA, resulting in inactivation of DNA replication, or can react with
396
sulfur-containing proteins, leading to the inhibition of enzyme functions (Mastsumure et al.,
397
2003; AshaRani et al., 2009, Nel et al., 2009, Marambio-Jones and Hoek, 2010).
398 399
3.5. Soil burial test
400 401
The efficiency of AgNPs finishes to protect the cotton fibers against the microbial degradation
402
process was evaluated by the soil burial test. As depicted in Fig. 5, after 14 days, the untreated
403
sample, as well as samples treatment with binder only, showed intensive brownish color, while,
404
the protection acquired by AgNPs (1˗5 mM) treatment was evident i.e., no color change was
405
observed. In addition, AgNPs prepared from 1mM AgNO3 solution was sufficient to achieve
406
maximum protection of the treated fabric against the adverse effects normally occur by soil
407
microflora. These results confirm those obtained in the previous section. The differences in
408
morphological changes and rotting intensities of the untreated and treated cotton fabric were
409
demonstrated by SEM (Fig. 6). The SEM micrographs showed that the morphological changes
410
obtained on the control untreated samples (Fig. 6 a, b) were much greater than those
411
demonstrated in the treated samples (Fig. 6 c, d), which were nearly unaffected by the soil
412
microflora. The SEM image of the control sample (Fig. 6b) revealed highly degraded parts of the
413
fibers, which again affirm the potential role exhibited by AgNPs in the protection of treated
414
samples against the damage arises by the actions of soil microflora. It is believed that excellent
415
antimicrobial properties of AgNPs were caused by the synergistic action of AgNPs and Ag+
416
present on the finished cellulose fibers. In addition, the large surface area of these particles
18
417
enabled a high rise of the released Ag+ concentration from the particles surface, which resulted in
418
the augmentation of the biocidal activity of the treatment (Klemenčič et al., 2010). In a similar
419
study, Klemenčič et al., (2010) reported that cotton woven fabric treated with AgNPs was almost
420
unaffected by the soil microflora as indicated by the SEM micrographs.
421
The influence of the Ag finishes on the cotton fabric biodegradation process was investigated by
422
measuring their mechanical properties in terms of tensile strength (MPa) and the elongation at
423
break (%); since these parameters are directly affected by the degree of sample degradation. As
424
expected, the tensile strength (MPa) and the elongation at break (%) of the control samples were
425
highly decreased after 14 days of soil burial (Fig. 7). The same trend was observed for the fabric
426
coated with binder only. In contrast, introducing AgNPs to the formulation ingredients inhibited
427
the biodegradation process of treated samples, which resulted in a sharp increase in both of the
428
tensile strength and elongation at break (%) (Fig. 7), this increase may be attributed to the
429
antibacterial effects of AgNPs. As illustrated in Fig. 7, with increasing of AgNPs concentration
430
in the treatment formulation, the tensile strength was slightly increased. This behavior could be
431
ascribed to the adsorption of AgNPs onto the cotton fibers, which in turn improved the
432
antimicrobial properties of the coated fabric, leading to enhanced tensile strength (MPa) of the
433
treated fabric. On the other hand, the elongation at break (%) was slightly decreased by
434
increasing AgNPs concentration. This trend may be attributed to the improved antimicrobial
435
properties of treated fabric, which in turn, decreased the degradation of the formed binder thin
436
layers, thereby, increases the mechanical strength and decreases the overall elongation of the
437
treated fabric. In a similar study, cotton fibers impregnated with AgNPs showed improved
438
mechanical properties due to binding of AgNPs onto the hydroxyl groups of the cellulose chains
439
of the cotton fibers (Ravindra et al., 2010).
19
440 441
3.6. Effect of curing method on the durability of antimicrobial treatment for washing
442 443
Table 5 illustrates the effect of γ-irradiation as a curing system, at different doses, on the color
444
differences (∆E*) of the treated fabric compared with the thermally cured (160 oC for 3 min)
445
treated fabric, and its durability against repeated washing cycles (1, 10, and 20) by using 2 g/l of
446
non-ionic detergent at temperature 50 oC. It must be mentioned that the smallest decreasing in
447
the color differences represents the higher durability for washing of the tested samples. It can be
448
seen that the durability of the thermally cured treated fabric was slightly decreased with
449
increasing the number of washing cycles, i.e., it decreased by 9.3% after 20 washing cycles,
450
compared with the unwashed samples. This could be ascribed to the good encapsulation of the
451
AgNPs upon the fabric’s surface through the used formulation. On the other hand, the color
452
differences (∆E*) was sharply decreased (38.5%) by using low dose of γ-irradiation (5 kGy) as a
453
curing system, and it increased gradually with increasing the irradiation dose up to (50 kGy), at
454
which the color differences decreased by 11.9% compared to the unwashed sample after 20
455
washing cycles, which gave an acceptable durability for washing nearly equal to that achieved by
456
thermally cured samples. The maintenance of color even after rinsing in water and drying
457
confirm the binding of the AgNPs on the surface of the cotton fibers (Falletta et al., 2008).
458
After being subjected to repeated washing cycles, it is evident from the data in Table 6 that
459
the reduction of bacterial colonies was always higher than 99% of S. aureus and E. coli, for
460
AgNPs treated samples. Subjecting the same fabric to increased number of washing cycles (20
461
cycles) did not affect the antibacterial properties; for thermally cured samples it was 99.1%,
20
462
98.7% and for γ-irradiation cured samples it was 99%, 98.6% for E. coil and S. aureus,
463
respectively.
464
The results in Table 6 indicate that cotton fabric treated with an antimicrobial formulation (1mM
465
of AgNPs and 10% binder) and cured thermally or by using γ-irradiation exhibited excellent
466
antibacterial properties, which could be ascribed to deposition of AgNPs onto the molecular
467
structure of cellulose of the cotton fabric and their fixation therein via chemical and physical
468
bonding. In agreement, Hebeish et al. (2011) reported that bacterial reduction displayed with
469
silver nanoparticles treated cotton fabric amounts to 96.4% and 95% for S. aureus and E. coli,
470
respectively. After 20 washing cycles, fabric samples exhibited 94.8% and 92%, which reflect
471
the significance of binder and curing system in the fixation of AgNPs deposits within the
472
molecular structure of cotton. El-Rafie et al. (2010) mentioned that the efficiency of the
473
antibacterial finish (AgNPs + binder) on the cotton fabric, expressed as the bacterial reduction
474
(%), reached 94% and 85% for S. aureus and E. coli, respectively, after 20 washing cycles. On
475
the other hand, Ilić et al. (2009) found that the desirable antimicrobial efficiency of the cotton
476
fabric loaded with silver nanoparticles from 50 ppm colloid was preserved after 5 washing
477
cycles.
478 479
4. Conclusions
480 481
Silver nanoparticles functionalized cotton fabric was developed by using a safe, cost-effective,
482
and eco-friendly process. The deposition of AgNPs onto cotton fibers improved their thermal
483
stability and elongation properties. The fabric coated with AgNPs, prepared from 1 mM AgNO3
484
solution, exhibited excellent quantitative and qualitative antimicrobial activity, against
21
485
representative pathogens namely, E. coli and S. aureus bacteria, without considerable change in
486
their color. Curing systems have a significant role in fixation of AgNPs deposited on the fabric
487
surface. This treatment can be employed in the manufacture of antimicrobial finishing and
488
textiles.
489 490 491 492
Acknowledgements The authors would like to thank the National Center for Radiation Research and Technology
493
(NCRRT), Egyptian Atomic Energy Authority for providing the facilities and financial support
494
throughout this work.
495 496 497 498 499 500 501
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627 628
Fig. 1. UV–vis spectra of synthesized AgNPs, using different AgNO3 concentrations (1-5 mM),
629
after 24 h of incubation. Inset showing: (a) negative control (AgNO3 aqueous solution), (b)
630
positive control (biomass filtrate), (c) biomass filtrate treated with AgNO3. 26
631 632
Fig. 2. TEM micrograph of synthesized AgNPs, the scale bar corresponds to 100 nm (a), inset
633
showing selected area electron diffraction pattern recorded from one of the particle, particle size
634
distribution histogram (b), and EDX pattern showing strong signal from the silver atoms in the
635
nanoparticles (c).
636 637
Fig. 3. TGA thermograms of untreated cotton samples, treated with binder only, and treated with
638
the antimicrobial formulation.
639 640
Fig. 4. Scanning electron microscope images of untreated cotton fabric (a), fabric treated with
641
binder only (b), and fabric treated with the antimicrobial formulation (c, d).
642 643
Fig. 5. Images of untreated and treated cotton fabric before (0 days) and after burring in moist
644
soil for 14 days.
645 646
Fig. 6. SEM micrographs of untreated cotton fabrics (a, b) and cotton fabrics treated with AgNPs
647
(1mM) (c, d), after burring in moist soil for 14 days.
648 649
Fig. 7. Mechanical properties of treated and untreated cotton fabrics, after burring in moist soil
650
for 14 days.
651 652 653
Table 1 Weight loss (%) of untreated cotton and cotton fabric treated with the antibacterial
654
formulation at different characteristic temperatures.
27
655
Weight loss (%) Sample
o
200 C
o
o
250 C
o
300 C
400 oC
350 C
450 oC
500 oC
550 oC
T(max) oC
Cotton
0.7±0.1 0.82±0.1 1.5±0.2 12.6±0.5 66.2±3.2 84.9±4.1 89.9±4.4 95.1±3.9
399±2.8
Cotton/binder
2.6±0.3
4.6±0.4
8.0±0.2 39.8±1.2 72.2±2.1 84.4±3.0 91.9±3.9 99.4±5.1
358±3.1
Cotton/binder/AgNPs 1.6±0.2
2.9±0.1
7.9±0.6 24.2±0.9 70.8±3.6 82.3±2.6 95.9±5.0 96.8±4.5
377±4.2
656 657 658 659
Table 2 Effect of AgNPs concentration on the color properties of the treated cotton fabrics.
660
Color parameters
Sample
Description
L*
a*
b*
Total color difference (∆E*)
Cotton+ binder
88±3
4±0.5
-3±0.3
1.8±0.3
white/cream
Cotton+ binder+ AgNPs (1 mM)
61±4
13±1
-21±1
13.4±1
yellowish red
Cotton+ binder+ AgNPs (2 mM)
52±4
26±0.5
-28±1
25.9±2
fade brownish red
Cotton+ binder+ AgNPs (3 mM)
49±2
34±2
-35±0.5
33.3±1
dark brownish red
Cotton+ binder+ AgNPs (4 mM)
38±2
42±2
-46±2
37.0±2
fade bluish red
Cotton+ binder+ AgNPs (5 mM)
36±1
49±1
-52±2
41.8±1
dark bluish red
661 662 663
Table 3 Qualitative antimicrobial activity of textile fabric coated with different concentrations of
664
AgNPs.
665
Inhibition zone diameter (mm) Microorganism
AgNPs concentration (mM) Control
1 mM
2 mM
3 mM
4 mM
5 mM
S. aureus
0
17 ± 0.5
17 ± 2.2
18 ± 1.3
18 ± 0.5
18 ± 2.1
B. subtilis
0
13 ± 1.1
14 ± 1.5
14 ± 1.1
15 ± 2.2
15 ± 1.8
P. aeruginosa
0
15 ± 0.3
15 ± 0.9
16 ± 0.7
16 ± 1.4
16 ± 0.9
28
E. coli
0
17 ± 2.1
17 ± 1.5
18 ± 0.5
20 ± 0.8
20 ± 1.4
A. niger
0
14 ± 1.6
15 ± 2.0
17 ± 0.2
20 ± 0.7
20 ± 2.1
666 667 668 669
Table 4 Quantitative antimicrobial activity of AgNPs treated textile fabric. AgNPs
Antimicrobial activities
colloidal Sample
S. aureus
E. coli
solution
Initial count
Surviving
Reduction
Count
Surviving
Reduction
(mM)
(CFU/ml)
cells
(%)
(CFU/ml)
cells
(%)
(CFU/ml) Untreated
0
fabric
3.91 x108
3.9 x 108
±0.71
±0.35 2.4 x 104
1
(CFU/ml) 0
3.6 x 108
3.5 x 108
±0.33
±0.54 3.0 x103
99.9
±0.28 AgNPs
2
treated fabric
99.9
±0.84
3.91 x 108
3.1x103
±0.92
±0.63
99.9
3.6 x 108
2.2 x103
±0.39
±0.11
99.9
3
0
100
0
100
4
0
100
0
100
5
0
100
0
100
670 671 672 673
0
Table 5 Durability of the antimicrobial finishing for washing using different curing systems.
674
Color difference (∆E*) Number of washing cycles
Curing system None
1
10
20
Thermally (160 oC for 3 min)
33.5±0.9
32.8±1.2
32.0±1.1
30.4±0.8
γ-irradiation (5 kGy)
33.5±0.9
29.5±0.7
25.4±1.2
20.6±0.9
29
γ-irradiation (10 kGy)
33.5±0.9
28.9±1.1
27.1±1.4
22.1±1.0
γ-irradiation (30 kGy)
33.5±0.9
31.2±0.9
27.9±0.7
25.7±1.3
γ-irradiation (50 kGy)
33.5±0.9
32.1±1.0
31.8±0.9
29.5±0.9
675 676 677
Table 6 Effect of repeated washing on the antibacterial properties of cotton fabrics treated with
678
AgNPs and cured thermally at 160 oC for 3 min or by γ-irradiation at 50 kGy.
679 Antibacterial activities γ-irradiation cured
Thermally cured Fabric
Number of
sample
washing
Surviving
Bacterial
Surviving
Bacterial
Surviving
Bacterial
Surviving
Bacterial
cycles
cells
reduction
cells
reduction
cells
reduction
cells
reduction
(CFU/ml)
(%)
(CFU/ml)
(%)
(CFU/ml)
(%)
(CFU/ml)
(%)
0
3.5 x 108
0
2.3 x 107
0
3.5 x 108
0
2.3 x 107
0
1
2.6 x 10
4
2.8 x 10
4
3.1 x 10
6
Untreated
AgNPstreated
10 20
E. coli
S. aureus
99.9 99.9 99.1
2.0 x 10
3
2.4 x 10
3
2.9 x 10
5
680 681 682
30
E. coli
99.9 99.9 98.7
2.4 x 10
4
2.5 x 10
4
3.2 x 10
6
S. aureus
99.9 99.9 99.0
2.1 x 10
3
99.9
2.3 x 10
3
99.9
3.2 x 10
5
98.6
a
b
c
683 684
Fig. 1.
685
(a)
(b)
31
(c) 686 687 688
Fig. 2.
689
32
120 Cotton Cotton/ binder Cotton/ binder/ AgNPs
Weight loss (%)
100 80 60 40 20 0 -20 0
100
200
300
400
500
600
o
Temperature ( C)
690 691
Fig. 3.
692 693 694
(a)
(b)
33
700
(c)
(d)
695 696
Fig. 4.
697
14 days
0 days
698 699
Untreated
With binder
AgNPs (1 mM)
700 701
Fig. 5.
702
(a)
(b)
34
AgNPs (3 mM)
AgNPs (5 mM)
(c)
(d)
703 704
Fig. 6.
705 706
35
.
Tensile strength (MPa) ( )
40
Buried samples 60
30 50
20
10 40 0 30 ly ied ted r on bur re a e t n d n U u bin
M 2m
M 3m
AgNPs concentration (mM)
707 708
M 1m
Fig. 7.
709
36
M 4m
M 5m
Elongation at break (%) (∆)
70
50
S0144-8617(16)30563-X http://dx.doi.org/doi:10.1016/j.carbpol.2016.05.041 CARP 11110
To appear in: Received date: Revised date: Accepted date:
1-3-2016 28-4-2016 12-5-2016
Please cite this article as: Ibrahim, Haytham MM., & Hassan, Mahmoud S., Characterization and antimicrobial properties of cotton fabric loaded with green synthesized silver nanoparticles.Carbohydrate Polymers http://dx.doi.org/10.1016/j.carbpol.2016.05.041 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
1 2 3
Characterization and antimicrobial properties of cotton fabric loaded with green synthesized silver nanoparticles
4 5 6
Haytham M. M. Ibrahima* [email protected], Mahmoud S. Hassanb
7 8 9
a
Department of Radiation Microbiology, National Center for Radiation Research and Technology (NCRRT), Atomic Energy Authority, Cairo, Egypt b
10 11
Department of Radiation Chemistry, National Center for Radiation Research and Technology (NCRRT), Atomic Energy Authority, Cairo, Egypt
12 13 14 15 16
* Corresponding author: Haytham M. M. Ibrahim, Ph.D. Radiation Microbiology Department, National Center for Radiation Research and Technology (NCRRT), Atomic Energy Authority, Egypt, Address: 3 Ahmed El Zomor St., Nasr City, P.O. Box 29, Cairo, Egypt, Tel.: +202 22748246 Fax: +202 22749298
17 18 19 20 21 22 23 24 25 26 27 28 29
1
30 31
Highlight
32 33
•
Silver nanoparticles (AgNPs) were prepared by eco-friendly, safe, and cost effective method using fungal filtrate.
34
•
Antimicrobial formulation encompasses AgNPs and binder was applied to cotton fabric.
35
•
Both AgNPs and treated cotton fabric were characterized.
36
•
The antimicrobial activity of treated fabric was assessed qualitatively and quantitatively.
37
•
Durability of antimicrobial properties was demonstrated after repeated washing cycles.
38 39 40
1. Introduction
41 42
Cotton fabrics are prevalent with people due to their superior characteristics such as
43
biodegradation, regeneration, affinity to skin, softness, and hygroscopic property (Lim and
44
Hudson, 2004). Cotton fibers are in particular appropriate for industrialization of textiles for
45
sports, medical, and healthcare/hygiene products (Czajka, 2005). Clothes and other textile
46
materials, especially those made of natural fibers such as cotton and wool, considered as good
47
media for the growth of pathogenic or odor-generating microorganisms. They offer a perfect
48
environment for the growth of bacteria and fungi due to their large surface area and capability to
49
keep oxygen, humidity, heat, and nutrients from the body exudates (Dev et al., 2009). Therefore,
50
with the increasing concern for individual health and hygiene, textiles with antimicrobial
51
activities are becoming an increasingly desirable objective of textile manufacturers.
52
Antimicrobial finishes are applied to textiles for three essential goals: (1) to control the
53
propagation of disease and preclude the danger generated by injury infection, (2) to prevent the
2
54
development of odor from aspiration, stains, and soil on textile materials and (3) to suppress the
55
damage of textiles caused by decomposition, especially textiles made of natural fibres (Gao and
56
Cranston, 2008). Currently, there is an increasing attention to produce non-toxic, durable, cost-
57
effective, and efficient antimicrobial textiles with expanded applications in health care, hygienic
58
products, medical, and protective textile materials (Gao and Cranston, 2008). The antimicrobial
59
agents that have been used industrially included quaternary ammonium salts, metal salts
60
solutions, and antibiotics. Regrettably, some of these substances are toxic or poorly effective,
61
which renders them not appropriate for application in, filters, textiles, health foods and for the
62
exclusions of pollution. On the other hand, employment of nanoparticles and their
63
nanocomposites would be good alternatives (Grace and Pandian, 2007). They can offer a new
64
opportunity for antimicrobial and multifunctional modification of textiles.
65
Despite the outstanding antimicrobial activity of silver nitrate, it’s not suitable for the
66
application to textile materials since it stains to black–brown when exposed to light and air,
67
because of uncontrolled reduction processes (Vigneshwaran et al., 2006). However, at defined
68
concentrations, a significant antimicrobial activity with an acceptable color change can be
69
achieved by using silver nanoparticles (AgNPs) deposited on the textile materials. The
70
antimicrobial activity of AgNPs is affected by the size of the particles, i.e. the smaller the
71
particles the greater the antimicrobial effect (Morones et al., 2005). At the nanoscale, AgNPs
72
exhibit remarkably unusual, chemical, physical, and biological properties, as well as
73
antimicrobial activity (Chen and Schluesener, 2008). Chemical synthesis of nanoparticles has
74
several hazards, like genotoxicity, cytotoxicity, carcinogenicity, and general toxicity (Mukherjee
75
et al., 2008). Therefore, there is a need to develop clean, non-toxic, and environment-friendly
76
methods for synthesis of nanoparticles (Mukherjee et al., 2008).
3
77
In the present study, AgNPs were prepared biologically employing the fungus Alternaria
78
alternata. The synthesized nanoparticles were characterized by scanning electron microscopy
79
(SEM), energy dispersive X-ray spectroscopy (EDX), transmission electron microscopy (TEM),
80
and dynamic light scattering (DLS). The prepared nano-silver was applied to then cotton fabric.
81
The treated cotton fabric was characterized by thermogravimetric analysis (TGA), SEM, and
82
mechanical properties. The resistance of treated fabric to biodegradation, their antimicrobial
83
properties, both qualitatively and quantitatively, against representative microorganisms, as well
84
as durability of the antimicrobial effects, after repeated washing cycles, were demonstrated.
85 86
2. Materials and methods
87 88
2.1. Cotton fabric and chemicals
89 90
Plain-weave cotton fabric (1mm thickness) were kindly provided by El-Nasr Company for
91
spinning, weaving, and dyeing, El-Mahalla El-Kubra, Egypt. The fabric was scoured and not
92
subjected to any finishing processes. The Impron MTP binder (butyl acrylate) as a crosslinking
93
agent and Genapol BE 2410 as a non-ionic detergent were kindly supplied by Clariant, Germany.
94
Silver nitrate (AgNO3) was purchased from Sigma, USA and used as received without further
95
purification, other chemicals used were of laboratory grade.
96 97
2.2. Microorganisms
98
4
99
The fungus Alternaria alternata was used to prepare AgNPs. Four bacterial strains, Bacillus
100
subtilis local isolate and Staphylococcus aureus ATCC 6538 as Gram-positive bacteria and
101
Pseudomonas aeruginosa ATCC 9027 and Escherichia coli ATCC 8739 as Gram-negative
102
bacteria, in addition to the fungus Aspergillus niger were employed to evaluate the antimicrobial
103
activity of the treated textile fabric. The standard bacterial strains were obtained from
104
Microbiologics, Inc. Minnesota, USA. Bacterial strains were maintained on nutrient agar slants
105
and the fungus was maintained on potato dextrose agar (PDA) slants at 4 oC.
106 107 108 109
2.3. Preparation of AgNPs
110 111
The fungus A. alternata was inoculated into 500-ml Erlenmeyer conical flasks containing 200 ml
112
of the mineral salts medium described by Kathiresan et al., (2009), with the following
113
composition in (g/l): potassium dihydrogen phosphate (KH2PO4), 7.0; dipotassium hydrogen
114
phosphate (K2HPO4), 2.0; magnesium sulfate (MgSO4), 0.1; ammonium sulfate [(NH4)2SO4],
115
1.0; yeast extract, 0.6; and glucose, 10.0, pH was adjusted to 6.5-7 using 1N solutions of HCl and
116
NaOH. Inoculated flasks were incubated at 28 oC and 150 rpm for 72 h in a rotary shaker
117
incubator (Gallen Kamp, UK). The mycelial biomass was harvested by filtration through
118
Whatman No.1 filter papers and extensively washed with distilled water to eliminate residual
119
medium. The biomass was taken into a flask containing 100 ml distilled water and incubated for
120
72 h at 28 oC. Thereafter, the aqueous solution (fungal filtrate) was separated by filtration and
121
used for the synthesis of AgNPs. For reduction, AgNO3 was added to 100 ml of the biomass
5
122
filtrate at a concentration of 1, 2, 3, 4, and 5 mM (Jaidev and Narasimha, 2010). The mixtures
123
were kept at room temperature for 48 h. Flasks with either fungal biomass filtrate or AgNO3
124
solution, served as positive and negative controls, were run simultaneously. The reduction of Ag+
125
into Ago (AgNPs) was monitored by using UV–Vis spectrophotometer (Helios Gamma, Thermo
126
Corporation, England).
127 128
2.4. Characterization of AgNPs
129 130
The average size of obtained AgNPs was estimated by DLS using zeta potential/particle sizer
131
spectrophotometer (NICOMP 380-ZLS, USA). The shape was determined by using TEM
132
(JEOL-JEM-1200 EX, Japan), operating at accelerating voltage of 80 kV. TEM samples were
133
prepared by dropping 2 drops of the colloidal AgNPs solutions onto 200 mesh carbon-coated
134
copper grids. The samples were air dried overnight prior to imaging the particles. The elemental
135
profile of AgNPs was determined by using EDX spectroscopy.
136 137
2.5. Silver nanoparticles loading on the textile fabric
138 139
Before being used, cotton fabric was washed and dried. Antimicrobial formulations, consist of
140
colloidal solutions of AgNPs with different concentrations (1, 2, 3, 4, and 5 mM), and the binder
141
(10 %), were treated with ultrasound before being used in the finishing process, to ensure
142
homogeneous dispersion and deagglomeration of nanoparticles. Cotton fabric samples (5 x 8 cm)
143
were immersed in the antimicrobial formulation with stirring and rotation and of the finishing
144
bath for 5 min. The samples were then squeezed, air-dried at room temperature. For fixation of
6
145
AgNPs on the fabric surface, the fabric samples were subjected to a curing process, utilizing
146
gamma-irradiation or a thermal method. Irradiation to the required doses (5, 10, 30, and 50 kGy)
147
was carried out in a 60Co gamma irradiation facility at NCRRT, the dose rate at the experiment
148
time was 0.276 kGy/h. In contrast, thermal curing was achieved by placing the samples in an
149
oven at 160 oC for 3 min.
150 151 152 153 154 155
2.6. Characterization of AgNPs loaded cotton fabric
156 157
2.6.1. Thermogravimetric analysis (TGA)
158
The thermogravimetric analysis (TGA) of control, binder coated, and AgNPs coated cotton
159
fabric was performed by using TGA-30 (Shimadzu, Japan) at a heating rate of 10oC/min in the
160
air at different temperatures ranged from room temperature up to 600 oC.
161 162
2.6.2. Scanning electron microscopy (SEM)
163
The surface morphology of AgNPs loaded cotton fibers, as well as the untreated control fabric,
164
was imaged at different magnifications by using SEM (JSM-5400, JEOL, Japan), working at an
165
accelerating voltage of 30 kV. Before imaging, the dried sample was sputter-coated with gold
166
using a microscope sputter coater.
167
7
168
2.6.3. Mechanical properties
169
The mechanical properties [tensile strength and elongation to break (%)] of treated and untreated
170
cotton fabrics were evaluated before the contact of the different systems with the soil and after
171
burring in moist soil for 14 days, were tested at room temperature, according to the standard test
172
method for breaking strength and elongation of textile fabrics (strip test); ASTM D5035-95,
173
using Mecmesin tester (Mecmesin Limited, UK), employing a crosshead speed of 50 mm/min
174
and equipped with software. In this system, the mechanical parameters were calculated directly.
175
Every data point is the average of 5 measurements.
176 177 178
2.6.4. Color-difference measurements
179
A microcolor unit, attached to a data station manufactured by Dr. Lange, Germany, was used for
180
color strength measurements. The L*, a* and b* interceptions used in this system are based on
181
the CIE-color triangle (Commission International de l'Eclairage units x, y and z). In this system,
182
the L* value represents the dark-white axis, a* represents the green-red axis and b* represents
183
the blue-yellow axis. The recorded values of the different color interceptions are the average of 5
184
measurements in different positions along the investigated samples. The L*, a*, and b* values of
185
the untreated samples were measured and taken as a reference, the total color differences (∆E*)
186
of the AgNPs treated samples were determined according to the following equation:
187
∆Ε ∗ =
( ∆ L∗ )2 + ( ∆ a ∗ )2 + ( ∆ b ∗ )2
(1)
188
Where ∆ L* is the color lightness difference between the fabric loaded with AgNPs and the
189
control sample; ∆ a* is the red/green difference between the treated and the control samples; ∆ b*
8
190
is the yellow/blue difference between the treated and the control samples (Ilić et al., 2009). The
191
recorded values of the different color interceptions are the average of ten measurements.
192 193
2.7. Antimicrobial activity of AgNPs loaded textile
194 195
The antimicrobial activities of AgNPs loaded cotton fabric were assessed qualitatively and
196
quantitatively, the untreated cotton fabric was used as a control. In addition, the resistance of the
197
AgNPs coated textile fabric together with uncoated fabric (control) to the action of soil
198
microflora was demonstrated by the soil burial test.
199 200 201
2.7.1. Qualitative method using agar diffusion assay
202
The bacterial strains under study were grown overnight in nutrient broth medium (Oxoid), then,
203
the cultures broth (containing ~108 CFU/ml) were swabbed on nutrient agar plates using sterile
204
cotton swabs. AgNPs loaded textile fabric was cut into small pieces (1cm x 1cm, 1mm
205
thickness), then, treated and untreated (control) samples were planted on the nutrient agar plates.
206
After being incubated at 32 oC for 24 h, the plates were examined for possible clear zone
207
formation (Ravindra et al., 2010). The presence of clear zone was recorded as an inhibition
208
against the tested microorganism. The extent of inhibition zone diameter was taken as a relative
209
measure of the antimicrobial effectiveness. The means of three replicates were tabulated.
210 211
2.7.2. Quantitative method using percentage reduction of bacterial count
9
212
The antimicrobial activity of the treated fabric was tested against E. coli (Gram-negative) and S.
213
aureus (Gram-positive) bacteria following the method described by Perelshtein et al., (2009).
214
The bacterial strains were grown in a nutrient broth medium at 32 °C overnight. Next day, they
215
were transferred into fresh medium at an initial optical density (O.D600) of 0.1. When the culture
216
reached an O.D600 of 0.3, the cells were harvested by centrifugation and washed twice with
217
sterile saline solution (NaCl 0.9 %, pH 6.5). Then, the cells were diluted to a final concentration
218
of 0.65 (O.D600) with saline solution. A 500 µl of the cells suspensions was pipetted into test
219
tubes, each containing 4.5 ml of sterile saline solution along with AgNPs treated or untreated
220
fabric samples; each of 1 cm2. The initial bacterial concentration was approximately 107 colony-
221
forming units (CFU)/ml. The test tubes were incubated for 24 h at 32 °C and 150 rpm in a rotary
222
shaker incubator (Gallen Kamp, UK). A 1 ml was taken at 0 h and after 24 h and serially diluted
223
up to 108 fold. Then, 100 µl of the appropriate dilutions was spread onto nutrient agar plates. The
224
plates were incubated at 32 °C for 24 h. The viable bacterial count (CFU/ml) was determined and
225
the antimicrobial activity was expressed as the percentage reduction (R, %) using the following
226
formula. R ( %) =
227
Co – C ×100 Co
(2)
228
Where R is the percentage reduction, Co is the number of bacterial colonies from the untreated
229
fabric, and C is the number of bacterial colonies from the treated fabric (Ilić et al., 2009).
230 231 232
2.8. Soil burial test
233
10
234
The resistance of AgNPs loaded and unloaded textile fabric to the actions of soil microflora was
235
evaluated by soil burial test according to the method of Mitchell et al., (2012). The samples were
236
buried in a naturally fertile top soil, placed in a dry wooden box to a depth of 13 cm. The soil
237
was moisturized by gradual addition of water accompanied by mixing. Uniform moisture content
238
was maintained by covering the soil container with a suitable lid. Before burying, the treated
239
samples were first moistened with an aqueous solution containing 0.05 % of a non-ionic wetting
240
agent. The samples were then buried horizontally. Precautions were taken to ensure uniform
241
covering with the soil along the length of the fabric samples. The soil moisture content was
242
maintained between 20 and 30% (based on the dry weight) and the temperature was kept at 28 oC
243
throughout the burying time (14 days). Later, the samples were removed from the soil, slightly
244
rinsed with tap water, and air-dried at room temperature. The mechanical properties [tensile
245
strength and elongation at break (%)] of the fabric samples were evaluated. Moreover, their
246
resistance to the action of soil microflora, in terms of biodegradation, was determined by using
247
SEM.
248 249
2.9. Durability of the antimicrobial finishing for washing
250 251
To evaluate the durability of AgNPs-treated cotton fabric toward repeated laundering, the
252
samples treated with the antimicrobial formulation followed by thermal or γ-irradiation curing
253
were submerged in a washing solution containing non-ionic detergent (2 g/l). The samples were
254
stirred for 15 min at 50 oC. Then, the samples were rinsed with tap water and air dried. This
255
procedure was repeated 1, 10, and 20 times. Later, the antimicrobial properties of the washed
256
samples were determined.
11
257 258
3. Results and discussions
259 260
3.1. Synthesis and characterization of AgNPs
261 262
In the present work, AgNPs were prepared by a green biological method using the biomass
263
filtrate of fungus A. alternata. This process has many advantages: (a) it is an environmentally
264
safe technique, (b) no need to use additional reducing agents or stabilizers, (c) it can be
265
conducted at room temperature, and (d) the developed AgNPs have excellent characteristics,
266
such as dispersion stability over a long time. As depicted in Fig. 1, upon the addition of AgNO3
267
aqueous solution (Fig. 1 a) to the pale yellow biomass filtrate solution (Fig. 1 b) the solution’s
268
color turned to reddish brown (Fig. 1 c). This color was mainly due to the surface plasmon
269
resonance of the AgNPs. The successful fabrication of AgNPs was confirmed by UV˗Vis
270
spectroscopy, where strong broad peaks located at λmax = 430 nm with an absorption tail in the
271
longer wavelengths were detected (Fig. 1). It is well known that AgNPs display ruby red color in
272
aqueous solutions, having a strong absorbance band around 400 - 450 nm emanating as a result
273
of surface plasmon excitation vibrations in the metal nanoparticles (Duran et al., 2007). This
274
peak is well documented for various metal nanoparticles with size range (2˗100 nm) (Shervani
275
et al., 2008). As illustrated in Fig. 1, the peak intensity, which represents the AgNPs
276
concentration, was increased with increasing the AgNO3 concentration (1-5 mM). The TEM
277
micrograph revealed that the individual AgNPs were spherical in shape with a mean diameter of
278
∼25–30 nm (Fig. 2a). The DLS technique showed relatively larger particle size, i.e. 35±20 nm
12
279
(Fig. 2b). The EDX spectrum of AgNPs showed signal characteristic for Ag+, as seen in Fig. 2c,
280
which confirms the presence of silver metal. Generally, metallic silver nanocrystallites reveal
281
ideal optical absorption peak approximately at 3 keV due to their surface plasmon resonance
282
(Kalimuthu et al., 2008).
283 284
3.2. Characterization of AgNPs treated cotton fabric
285 286
3.2.1. Thermal stability
287
The thermal stability of untreated cotton fabric, binder coated fabric, and fabric treated with the
288
antimicrobial formulation, containing a binder (10%), based on the total volume of treatment
289
solution, and AgNPs (1mM), followed by thermal curing at 160 oC for 3 min, have been
290
investigated by thermogravimetric analysis. The TGA thermograms were shown in Fig. 3. It
291
could be noticed that, the thermal degradation of the coated fabric with binder only begins at a
292
lower initial temperature than the uncoated cotton fabric, in which the initial temperature of the
293
overall degradation was ~238 °C, i.e. lower than the uncoated cotton fabric by ~50 °C,
294
accompanied with wt. loss (%) of 5% and 19%, for the uncoated and coated fabrics with acrylate
295
binder, respectively. These results could be attributed to the presence of acrylate binder thin film
296
which degrades faster than the textile component, reducing the initial degradation temperature
297
(Alongi et al., 2012). The addition of AgNPs to the coating formulation showed slight increases
298
in the thermal stability more than that of the fabric coated with acrylate binder, but still less than
299
the untreated cotton fabrics, with a wt. loss of 13% which improves the binding of AgNPs to the
300
cotton fabric encapsulated thought the acrylate binder, causing the increasing of the total thermal
301
stability more than of the fabrics coated with binder only (Dhas et al., 2015).
13
302
For all fabric samples, the thermal degradation undergoes three steps. The first step illustrates the
303
water elimination from cotton fabrics, besides the initial degradation of the acrylate binder of
304
treated fabrics, which begins at 325, 238, and 238 °C, with a maximum wt. loss of 33, 37, and
305
33% for cotton, cotton/binder, and cotton/binder/AgNPs, respectively. The second step
306
corresponds to the degradation of the cross-linked acrylate binder, which begins at 385, 365, and
307
385 °C with a maximum wt. loss of 32% for cotton, cotton/binder, and cotton/binder/AgNPs,
308
respectively. The third step is accompanied with the degradation of the textile component, which
309
begins at 446 °C with a maximum wt. loss of 97% for all samples. Table 1 illustrated the total
310
weight loss (%) at different temperatures, in which the temperatures of maximum rates of the
311
thermal decomposition (Tmax) were obtained at 399±2.8, 358±3.1, and 377±4.2 oC for cotton,
312
cotton/binder, and cotton/binder/AgNPs, respectively.
313 314 315
3.2.2. Surface morphology
316
To demonstrate that the AgNPs were deposited on the cotton fibers, the fabric before and after
317
antibacterial treatment, with a formulation containing 10 % binder and 1mM AgNPs followed by
318
thermal curing at 160 oC for 3 min, were scanned under the SEM (Fig. 4). The SEM
319
photomicrographs revealed that the fibers of control cotton fabric (Fig. 4a) and fabric treated
320
with the binder only (Fig. 4b) exhibiting uniform neat plain spun structures and the cotton fibers
321
manifested smooth surfaces. Whereas, AgNPs treated cotton fibers clearly became more coarse
322
and exhibited characteristic semi-granulated pattern (Fig. 4c,d), due to the formation of a thin
323
layer around the fibers, composed of the binder and AgNPs encapsulated in the treatment
324
formulation. The images demonstrated also that the AgNPs were spherical in nature. Clusters of
14
325
AgNPs are demonstrated on the fabric surface due to the agglomeration process. Agglomeration
326
of AgNPs on the cotton fibers could not be prohibited in spite of the utilization of ultrasound
327
processing before finishing and by the implementation of the finish by the exhaustion method,
328
which is guaranteed by stirring and rotation of the finishing bath. The elemental profile of
329
AgNPs treated fabric showed higher counts at 3 keV due to silver, which confirms the presence
330
of AgNPs as indicated in Fig. 1.
331 332
3.2.3. Color differences of AgNPs coated cotton fabric
333
The data summarized in Table 2 represent the effect of increasing AgNPs concentration on the
334
color changes of the treated cotton fabric, which was quantitatively evaluated by the CIE L*, a*,
335
and b* color parameters, and the total color differences (∆E*). As given in Table 2, the cotton
336
fabric treated with binder only was a white/cream color, as indicated by the high value of L*
337
coordinate (whiteness), this value decreased progressively with introducing increased
338
concentrations of AgNPs (1˗5 mM). In contrast, the values of a* (red) and b* (blue) coordinates
339
increased. Therefore, the value of the total color difference (∆E*) increased from 1.8±0.3 to
340
41.8±1. The AgNPs/binder/cotton fabric composites color changed from white/cream (untreated
341
fabric) to yellowish red to fade brownish red to dark brownish red to fade bluish red to dark
342
bluish red. This color is attributed to the surface plasmon resonance effects of AgNPs, the
343
successive change in color resulting from an increase in the size of the nanoparticles as more Ag0
344
is formed on the nanoparticle surface with increasing the AgNPs concentration (Kelly and
345
Johnston, 2011).
346 347
3.3. Antimicrobial properties of AgNPs coated fabric
15
348 349
As shown in Table 3 the AgNPs treated cotton fabric revealed high antimicrobial activity at all
350
the tested concentrations, whereas the untreated fabric did not show any activity. The lowest
351
AgNPs concentration (1mM) was found to be sufficient to inhibit the growth of tested
352
microorganisms as indicated by the inhibition zone diameter (13-17 mm). Further increase in
353
AgNPs concentration was insignificant, as revealed by the trivial increase in inhibition zone
354
diameter (20 mm at 4 and 5 mM). In general, the antibacterial activity was higher against E.coli
355
than S. aureus. The AgNPs containing cotton fibers developed in the present work have
356
exhibited inhibition zone > 1.5 mm in all cases. These results indicate that they are highly
357
important, from the technological point of view, for establishing antibacterial finishing (Ravindra
358
et al., 2010).
359
The quantitative antimicrobial activity of the loaded fabric was studied against E. coli (Gram-
360
negative bacterium) and S. aureus (Gram-positive bacterium). The viable bacteria were
361
monitored by counting the number of CFU/ml on nutrient agar plates. As shown in Table 4,
362
treatment for 24h with the loaded fabric achieved 99.9 % inhibition of E. coli and S. aureus
363
growth. Upon increasing the concentration of the applied nano-silver colloidal solutions from 1
364
to 5 mM, the growth of S. aureus and E. coli was completely inhibited, i.e. 100%. These results
365
agree with that obtained by the agar diffusion method. The control fabric did not show any
366
antibacterial activity. In agreement, Zhang et al., (2009) reported that the silver-treated cotton
367
fabric showed 99.01% and 99.26% bacterial reduction of S. aureus and E. coli, respectively,
368
while the silver content of cotton was about 88 mg/kg. The bacterial reduction rates increased
369
slightly from 99.01% to 100% with the increase of the silver content of the silver-treated cotton
370
fabric to 158.49 and 173.62, respectively.
16
371
Based on the results of Tables 2, 3, and 4, it apparent that the AgNPs concentration of 1 mM is
372
sufficient and efficient concentration, which gives qualitatively noticeable inhibition zone and
373
quantitatively an excellent microbial reduction (99.9%) against the tested bacterial strains,
374
accompanied with lower and acceptable color change of the treated cotton fabric.
375 376
3.4. Mechanism of antimicrobial activity of AgNPs
377 378
Silver nanoparticles have been reported to exhibit antimicrobial properties (Virender et al.,
379
2009). Using AgNPs leads to increasing the number of particles per unit area and, thus, the
380
antibacterial effects can be maximized (Yeo and Jeong, 2003). The antibacterial property of
381
AgNPs treated cotton fabric is related to their Ag contents. AgNPs are capable of generating
382
reactive oxygen species (ROS), such as superoxide (•O2−), hydrogen peroxide (H2O2), and
383
hydrogen radical (•OH) from its surface. Several main mechanisms underlie the biocidal
384
properties of AgNPs against microorganisms have been postulated. (a) Ag0 nanoparticles attach
385
to the negatively charged cell surface, alter the physical and chemical properties of the cell
386
membranes and the cell wall and disturb important functions such as permeability,
387
osmoregulation, electron transport, and respiration. Besides, they increase the cell permeability
388
and leak the intracellular contents by cell disruption (Kora and Arunachalam, 2011), (b) they can
389
cause further damage to bacterial cells by permeating the cell, where they interact with DNA,
390
proteins, and other phosphorus- and sulfur-containing cell constituents. It is believed that the
391
high affinity of silver towards sulfur or phosphorus is the key element of this effect. Due to the
392
abundance of sulfur containing proteins on the bacterial cell membrane, AgNPs can react with
393
sulfur-containing proteins inside or outside the cell membrane, which in turn affects bacterial cell
17
394
viability (Morones et al. 2005), (c) silver ions (particularly Ag+) released from (Ag0) can interact
395
with phosphorus moieties in DNA, resulting in inactivation of DNA replication, or can react with
396
sulfur-containing proteins, leading to the inhibition of enzyme functions (Mastsumure et al.,
397
2003; AshaRani et al., 2009, Nel et al., 2009, Marambio-Jones and Hoek, 2010).
398 399
3.5. Soil burial test
400 401
The efficiency of AgNPs finishes to protect the cotton fibers against the microbial degradation
402
process was evaluated by the soil burial test. As depicted in Fig. 5, after 14 days, the untreated
403
sample, as well as samples treatment with binder only, showed intensive brownish color, while,
404
the protection acquired by AgNPs (1˗5 mM) treatment was evident i.e., no color change was
405
observed. In addition, AgNPs prepared from 1mM AgNO3 solution was sufficient to achieve
406
maximum protection of the treated fabric against the adverse effects normally occur by soil
407
microflora. These results confirm those obtained in the previous section. The differences in
408
morphological changes and rotting intensities of the untreated and treated cotton fabric were
409
demonstrated by SEM (Fig. 6). The SEM micrographs showed that the morphological changes
410
obtained on the control untreated samples (Fig. 6 a, b) were much greater than those
411
demonstrated in the treated samples (Fig. 6 c, d), which were nearly unaffected by the soil
412
microflora. The SEM image of the control sample (Fig. 6b) revealed highly degraded parts of the
413
fibers, which again affirm the potential role exhibited by AgNPs in the protection of treated
414
samples against the damage arises by the actions of soil microflora. It is believed that excellent
415
antimicrobial properties of AgNPs were caused by the synergistic action of AgNPs and Ag+
416
present on the finished cellulose fibers. In addition, the large surface area of these particles
18
417
enabled a high rise of the released Ag+ concentration from the particles surface, which resulted in
418
the augmentation of the biocidal activity of the treatment (Klemenčič et al., 2010). In a similar
419
study, Klemenčič et al., (2010) reported that cotton woven fabric treated with AgNPs was almost
420
unaffected by the soil microflora as indicated by the SEM micrographs.
421
The influence of the Ag finishes on the cotton fabric biodegradation process was investigated by
422
measuring their mechanical properties in terms of tensile strength (MPa) and the elongation at
423
break (%); since these parameters are directly affected by the degree of sample degradation. As
424
expected, the tensile strength (MPa) and the elongation at break (%) of the control samples were
425
highly decreased after 14 days of soil burial (Fig. 7). The same trend was observed for the fabric
426
coated with binder only. In contrast, introducing AgNPs to the formulation ingredients inhibited
427
the biodegradation process of treated samples, which resulted in a sharp increase in both of the
428
tensile strength and elongation at break (%) (Fig. 7), this increase may be attributed to the
429
antibacterial effects of AgNPs. As illustrated in Fig. 7, with increasing of AgNPs concentration
430
in the treatment formulation, the tensile strength was slightly increased. This behavior could be
431
ascribed to the adsorption of AgNPs onto the cotton fibers, which in turn improved the
432
antimicrobial properties of the coated fabric, leading to enhanced tensile strength (MPa) of the
433
treated fabric. On the other hand, the elongation at break (%) was slightly decreased by
434
increasing AgNPs concentration. This trend may be attributed to the improved antimicrobial
435
properties of treated fabric, which in turn, decreased the degradation of the formed binder thin
436
layers, thereby, increases the mechanical strength and decreases the overall elongation of the
437
treated fabric. In a similar study, cotton fibers impregnated with AgNPs showed improved
438
mechanical properties due to binding of AgNPs onto the hydroxyl groups of the cellulose chains
439
of the cotton fibers (Ravindra et al., 2010).
19
440 441
3.6. Effect of curing method on the durability of antimicrobial treatment for washing
442 443
Table 5 illustrates the effect of γ-irradiation as a curing system, at different doses, on the color
444
differences (∆E*) of the treated fabric compared with the thermally cured (160 oC for 3 min)
445
treated fabric, and its durability against repeated washing cycles (1, 10, and 20) by using 2 g/l of
446
non-ionic detergent at temperature 50 oC. It must be mentioned that the smallest decreasing in
447
the color differences represents the higher durability for washing of the tested samples. It can be
448
seen that the durability of the thermally cured treated fabric was slightly decreased with
449
increasing the number of washing cycles, i.e., it decreased by 9.3% after 20 washing cycles,
450
compared with the unwashed samples. This could be ascribed to the good encapsulation of the
451
AgNPs upon the fabric’s surface through the used formulation. On the other hand, the color
452
differences (∆E*) was sharply decreased (38.5%) by using low dose of γ-irradiation (5 kGy) as a
453
curing system, and it increased gradually with increasing the irradiation dose up to (50 kGy), at
454
which the color differences decreased by 11.9% compared to the unwashed sample after 20
455
washing cycles, which gave an acceptable durability for washing nearly equal to that achieved by
456
thermally cured samples. The maintenance of color even after rinsing in water and drying
457
confirm the binding of the AgNPs on the surface of the cotton fibers (Falletta et al., 2008).
458
After being subjected to repeated washing cycles, it is evident from the data in Table 6 that
459
the reduction of bacterial colonies was always higher than 99% of S. aureus and E. coli, for
460
AgNPs treated samples. Subjecting the same fabric to increased number of washing cycles (20
461
cycles) did not affect the antibacterial properties; for thermally cured samples it was 99.1%,
20
462
98.7% and for γ-irradiation cured samples it was 99%, 98.6% for E. coil and S. aureus,
463
respectively.
464
The results in Table 6 indicate that cotton fabric treated with an antimicrobial formulation (1mM
465
of AgNPs and 10% binder) and cured thermally or by using γ-irradiation exhibited excellent
466
antibacterial properties, which could be ascribed to deposition of AgNPs onto the molecular
467
structure of cellulose of the cotton fabric and their fixation therein via chemical and physical
468
bonding. In agreement, Hebeish et al. (2011) reported that bacterial reduction displayed with
469
silver nanoparticles treated cotton fabric amounts to 96.4% and 95% for S. aureus and E. coli,
470
respectively. After 20 washing cycles, fabric samples exhibited 94.8% and 92%, which reflect
471
the significance of binder and curing system in the fixation of AgNPs deposits within the
472
molecular structure of cotton. El-Rafie et al. (2010) mentioned that the efficiency of the
473
antibacterial finish (AgNPs + binder) on the cotton fabric, expressed as the bacterial reduction
474
(%), reached 94% and 85% for S. aureus and E. coli, respectively, after 20 washing cycles. On
475
the other hand, Ilić et al. (2009) found that the desirable antimicrobial efficiency of the cotton
476
fabric loaded with silver nanoparticles from 50 ppm colloid was preserved after 5 washing
477
cycles.
478 479
4. Conclusions
480 481
Silver nanoparticles functionalized cotton fabric was developed by using a safe, cost-effective,
482
and eco-friendly process. The deposition of AgNPs onto cotton fibers improved their thermal
483
stability and elongation properties. The fabric coated with AgNPs, prepared from 1 mM AgNO3
484
solution, exhibited excellent quantitative and qualitative antimicrobial activity, against
21
485
representative pathogens namely, E. coli and S. aureus bacteria, without considerable change in
486
their color. Curing systems have a significant role in fixation of AgNPs deposited on the fabric
487
surface. This treatment can be employed in the manufacture of antimicrobial finishing and
488
textiles.
489 490 491 492
Acknowledgements The authors would like to thank the National Center for Radiation Research and Technology
493
(NCRRT), Egyptian Atomic Energy Authority for providing the facilities and financial support
494
throughout this work.
495 496 497 498 499 500 501
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616
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617
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621 622 623
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624 625 626
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627 628
Fig. 1. UV–vis spectra of synthesized AgNPs, using different AgNO3 concentrations (1-5 mM),
629
after 24 h of incubation. Inset showing: (a) negative control (AgNO3 aqueous solution), (b)
630
positive control (biomass filtrate), (c) biomass filtrate treated with AgNO3. 26
631 632
Fig. 2. TEM micrograph of synthesized AgNPs, the scale bar corresponds to 100 nm (a), inset
633
showing selected area electron diffraction pattern recorded from one of the particle, particle size
634
distribution histogram (b), and EDX pattern showing strong signal from the silver atoms in the
635
nanoparticles (c).
636 637
Fig. 3. TGA thermograms of untreated cotton samples, treated with binder only, and treated with
638
the antimicrobial formulation.
639 640
Fig. 4. Scanning electron microscope images of untreated cotton fabric (a), fabric treated with
641
binder only (b), and fabric treated with the antimicrobial formulation (c, d).
642 643
Fig. 5. Images of untreated and treated cotton fabric before (0 days) and after burring in moist
644
soil for 14 days.
645 646
Fig. 6. SEM micrographs of untreated cotton fabrics (a, b) and cotton fabrics treated with AgNPs
647
(1mM) (c, d), after burring in moist soil for 14 days.
648 649
Fig. 7. Mechanical properties of treated and untreated cotton fabrics, after burring in moist soil
650
for 14 days.
651 652 653
Table 1 Weight loss (%) of untreated cotton and cotton fabric treated with the antibacterial
654
formulation at different characteristic temperatures.
27
655
Weight loss (%) Sample
o
200 C
o
o
250 C
o
300 C
400 oC
350 C
450 oC
500 oC
550 oC
T(max) oC
Cotton
0.7±0.1 0.82±0.1 1.5±0.2 12.6±0.5 66.2±3.2 84.9±4.1 89.9±4.4 95.1±3.9
399±2.8
Cotton/binder
2.6±0.3
4.6±0.4
8.0±0.2 39.8±1.2 72.2±2.1 84.4±3.0 91.9±3.9 99.4±5.1
358±3.1
Cotton/binder/AgNPs 1.6±0.2
2.9±0.1
7.9±0.6 24.2±0.9 70.8±3.6 82.3±2.6 95.9±5.0 96.8±4.5
377±4.2
656 657 658 659
Table 2 Effect of AgNPs concentration on the color properties of the treated cotton fabrics.
660
Color parameters
Sample
Description
L*
a*
b*
Total color difference (∆E*)
Cotton+ binder
88±3
4±0.5
-3±0.3
1.8±0.3
white/cream
Cotton+ binder+ AgNPs (1 mM)
61±4
13±1
-21±1
13.4±1
yellowish red
Cotton+ binder+ AgNPs (2 mM)
52±4
26±0.5
-28±1
25.9±2
fade brownish red
Cotton+ binder+ AgNPs (3 mM)
49±2
34±2
-35±0.5
33.3±1
dark brownish red
Cotton+ binder+ AgNPs (4 mM)
38±2
42±2
-46±2
37.0±2
fade bluish red
Cotton+ binder+ AgNPs (5 mM)
36±1
49±1
-52±2
41.8±1
dark bluish red
661 662 663
Table 3 Qualitative antimicrobial activity of textile fabric coated with different concentrations of
664
AgNPs.
665
Inhibition zone diameter (mm) Microorganism
AgNPs concentration (mM) Control
1 mM
2 mM
3 mM
4 mM
5 mM
S. aureus
0
17 ± 0.5
17 ± 2.2
18 ± 1.3
18 ± 0.5
18 ± 2.1
B. subtilis
0
13 ± 1.1
14 ± 1.5
14 ± 1.1
15 ± 2.2
15 ± 1.8
P. aeruginosa
0
15 ± 0.3
15 ± 0.9
16 ± 0.7
16 ± 1.4
16 ± 0.9
28
E. coli
0
17 ± 2.1
17 ± 1.5
18 ± 0.5
20 ± 0.8
20 ± 1.4
A. niger
0
14 ± 1.6
15 ± 2.0
17 ± 0.2
20 ± 0.7
20 ± 2.1
666 667 668 669
Table 4 Quantitative antimicrobial activity of AgNPs treated textile fabric. AgNPs
Antimicrobial activities
colloidal Sample
S. aureus
E. coli
solution
Initial count
Surviving
Reduction
Count
Surviving
Reduction
(mM)
(CFU/ml)
cells
(%)
(CFU/ml)
cells
(%)
(CFU/ml) Untreated
0
fabric
3.91 x108
3.9 x 108
±0.71
±0.35 2.4 x 104
1
(CFU/ml) 0
3.6 x 108
3.5 x 108
±0.33
±0.54 3.0 x103
99.9
±0.28 AgNPs
2
treated fabric
99.9
±0.84
3.91 x 108
3.1x103
±0.92
±0.63
99.9
3.6 x 108
2.2 x103
±0.39
±0.11
99.9
3
0
100
0
100
4
0
100
0
100
5
0
100
0
100
670 671 672 673
0
Table 5 Durability of the antimicrobial finishing for washing using different curing systems.
674
Color difference (∆E*) Number of washing cycles
Curing system None
1
10
20
Thermally (160 oC for 3 min)
33.5±0.9
32.8±1.2
32.0±1.1
30.4±0.8
γ-irradiation (5 kGy)
33.5±0.9
29.5±0.7
25.4±1.2
20.6±0.9
29
γ-irradiation (10 kGy)
33.5±0.9
28.9±1.1
27.1±1.4
22.1±1.0
γ-irradiation (30 kGy)
33.5±0.9
31.2±0.9
27.9±0.7
25.7±1.3
γ-irradiation (50 kGy)
33.5±0.9
32.1±1.0
31.8±0.9
29.5±0.9
675 676 677
Table 6 Effect of repeated washing on the antibacterial properties of cotton fabrics treated with
678
AgNPs and cured thermally at 160 oC for 3 min or by γ-irradiation at 50 kGy.
679 Antibacterial activities γ-irradiation cured
Thermally cured Fabric
Number of
sample
washing
Surviving
Bacterial
Surviving
Bacterial
Surviving
Bacterial
Surviving
Bacterial
cycles
cells
reduction
cells
reduction
cells
reduction
cells
reduction
(CFU/ml)
(%)
(CFU/ml)
(%)
(CFU/ml)
(%)
(CFU/ml)
(%)
0
3.5 x 108
0
2.3 x 107
0
3.5 x 108
0
2.3 x 107
0
1
2.6 x 10
4
2.8 x 10
4
3.1 x 10
6
Untreated
AgNPstreated
10 20
E. coli
S. aureus
99.9 99.9 99.1
2.0 x 10
3
2.4 x 10
3
2.9 x 10
5
680 681 682
30
E. coli
99.9 99.9 98.7
2.4 x 10
4
2.5 x 10
4
3.2 x 10
6
S. aureus
99.9 99.9 99.0
2.1 x 10
3
99.9
2.3 x 10
3
99.9
3.2 x 10
5
98.6
a
b
c
683 684
Fig. 1.
685
(a)
(b)
31
(c) 686 687 688
Fig. 2.
689
32
120 Cotton Cotton/ binder Cotton/ binder/ AgNPs
Weight loss (%)
100 80 60 40 20 0 -20 0
100
200
300
400
500
600
o
Temperature ( C)
690 691
Fig. 3.
692 693 694
(a)
(b)
33
700
(c)
(d)
695 696
Fig. 4.
697
14 days
0 days
698 699
Untreated
With binder
AgNPs (1 mM)
700 701
Fig. 5.
702
(a)
(b)
34
AgNPs (3 mM)
AgNPs (5 mM)
(c)
(d)
703 704
Fig. 6.
705 706
35
.
Tensile strength (MPa) ( )
40
Buried samples 60
30 50
20
10 40 0 30 ly ied ted r on bur re a e t n d n U u bin
M 2m
M 3m
AgNPs concentration (mM)
707 708
M 1m
Fig. 7.
709
36
M 4m
M 5m
Elongation at break (%) (∆)
70
50