Tailoring of Antibacterial and UV-protective Cotton Fabric by an in situ Synthesis of Silver Particles in the Presence of a Sol-gel Matrix and Sumac Leaf Extract Izdelava protibakterijske in UV zaščitne bombažne tkanine z in situ sintezo srebrovih delcev v prisotnosti sol-gel matrice in ekstrakta listov octovca

This research presents a new procedure for the chemical modifi cation of cotton fabric, which included a ‘’green’’ in situ synthesis of silver particles using an extract of sumac leaves as a reducing agent. To increase the adsorption ability of silver cations, a sol-gel matrix was previously created on cotton fabric using an organic–inorganic precursor sol-gel. The presence of silver particles on the cotton fabric was confi rmed by scanning electron microscopy and energy-dispersive X-ray spectroscopy. The results showed that silver particles were created on the cotton fabric in the presence of the sumac leaf extract, which colored the fi bers in brown. The presence of the sol-gel matrix increased the adsorption of silver cations and therefore the concentration of sliver particles, which resulted in a deeper color yield. Silver particles provided antibacterial protection, with a 99–100% reduction of E. coli in S. aureus bacteria, while the sumac leaf extract provided excellent protection against ultraviolet radiation, with an ultraviolet protective factor equaling 66.54. The coating was also highly durable in terms of its washing fastness.


Introduction
In textiles, silver particles (Ag Ps) have been recognized as eff ective antimicrobial agents, with broadspectrum activity against bacteria, fungi and viruses. Besides zinc oxide and titanium dioxide, Ag Ps are mostly used for the fabrication of medical and hygiene textiles. However, the antimicrobial mechanism of Ag Ps is not yet fully known, yet the activity has been attributed to silver cations (Ag + ), which are released from the surface of Ag Ps, and to Ag Ps, if their size is within a nanometer scale (Ag NPs) [1,2]. Both Ag + and Ag NPs can interact with the bacterial cell wall, where their accumulation causes membrane damage. Furthermore, Ag + and Ag NPs lower than 10 nm can penetrate the cell, where they hinder or deactivate its critical physiological functions and consequently destroy the cell. In the presence of oxygen, Ag + and Ag NPs may also catalytically accelerate the formation of reactive oxygen species (ROS), which are highly toxic to microorganism cells [1,2]. Th ere are several classical and contemporary approaches for the application of Ag Ps to textile substrates, which include the application of pre-synthesized Ag Ps using an appropriate fi nishing method or an in situ synthesis of Ag Ps in the presence of a textile substrate [3−5]. An important advantage of the in situ generation of Ag Ps is that it enables the growth of Ag Ps inside the textile fi bers, increases the homogeneity and uniformity of the particle distribution inside and on the fi ber surface, and reduces the agglomeration of particles. In this process, no additional methods or chemicals are needed to enhance dispersibility of the ex situ synthesized Ag Ps and to achieve their stability against agglomeration. However, in both approaches, Ag Ps are formed in the chemical reduction of silver salt, where diff erent environmentally harmful organic or inorganic reducing and stabilizing agents are usually used. To avoid toxic chemicals and perform the fabrication processes more sustainably, biological methods for the synthesis of Ag Ps, in which extracts of plants and microorganisms are used as reducing and stabilizing agents, have received increasing attention [6−8]. Th e in situ biosynthesis of Ag Ps represents a 'green' fabrication process of Ag-functionalized textile substrates. In this process, AgNO 3 , as a silver precursor, and plant extracts, as reducing and stabilizing agents, have mostly been used simultaneously.
Namely, natural biomolecules with carbonyl and phenolic hydroxyl functional groups, including alkaloids, tannins, fl avonoids, phenols, amino acids, and polysaccharides, have been extracted from leaves, seeds, peel, and fruits of diff erent plants and introduced in the reduction of silver precursors to Ag Ps [9−17]. Among plant extracts, extracts from sumac (Rhus spp.) could be introduced as a promising reducing agent because of the variety of biological active compounds present in sumac's bark, branches, roots, leaves, seeds and fruits [18,19]. Sumac is native to the temperate regions of North America, but it has also been spread worldwide and developed as a sustainable non-traditional economic plant. While diff erent parts of sumac have already been used in food and cosmetic industries, to the best of our knowledge, sumac has not yet been used for the production of the micro-and nanoparticles of metals or metal oxides. Th erefore, the aim of this research was to develop a novel process for the in situ synthesis of Ag Ps on cotton fi bers using sumac leaf extracts. To increase the adsorption ability of silver cations, an organicinorganic hybrid sol-gel precursor was applied to fi bers to create a sol-gel matrix, prior the immersion of the fi bers in AgNO 3 . Namely, we assumed that the presence of a sol-gel matrix on cotton fi bers will increase their concentration of Ag Ps, as well as enhance their coating durability in comparison to fi bers with no sol-gel matrix. Th e chemically modifi ed cotton fi bers were characterized by scanning electron microscopy (SEM) and energy-dispersive X-ray spectroscopy (EDS). Th eir antibacterial properties were investigated against the Gram-positive Staphylococcus aureus and the Gram-negative Escherichia coli bacteria. Th eir ultraviolet (UV) protection properties were determined in terms of the ultraviolet protection factor (UPF). An important goal of the research was also to determine the washing fastness of the coating.

Materials
Alkaline-scoured, bleached, and mercerized 100% cotton plain-weave fabric (Tekstina d.o.o., Aj dovščina, Slovenia), with a mass per unit area of 119 g/m 2 , was used for chemical modifi cation. To create a solgel matrix on the cotton fabric, iSys MTX (CHT R.

Tailoring of Antibacterial and UV-protective Cotton Fabric by an in situ Synthesis of Silver Particles in the Presence of a Sol-gel Matrix and Sumac Leaf Extract
Beitlich GmbH, Tübingen, Germany), a reactive organic-inorganic sol, which is miscible with water at every ratio, was used in combination with Kollasol CDO, an anti-foaming agent (CHT R. Beitlich GmbH, Tübingen, Germany). Silver nitrate (AgNO 3 ; 99.98%, Sigma Aldrich) was used as a silver precursor. Fresh leaves of sumac were supplied by the public holding company, JP VOKA SNAGA d.o.o. All solutions were prepared in double-distilled water.

Preparation of the sumac leaf extract solution
First, 20 g of dried sumac leaves were crushed and poured with 1000 ml of water. Th e mixture was heated to 98 °C and let to boil for 20 min at a gentle boiling. Aft erwards, the extract was fi ltered and cooled at room temperature.

Chemical modifi cation of the cotton fabric
Th e modifi cation of the cotton fabric samples was performed in a two-step procedure ( Figure 1). Firstly, 15 g/l iSys MTX in the combination with 1 g/l Kollasol CDO were prepared in double-distilled water and applied to the cotton samples by a pad-dry-cure method, including full immersion at room temperature, a wet-pick-up of 80%, drying at 100 °C and curing for 3 min at 150 °C. Th e concentrations of the agents and the application conditions were those recommended by the producer. Aft er the treatment, the samples were left for seven days under standard atmospheric conditions (65% ± 2% relative humidity and 20 °C ± 1 °C) to allow for a complete sol-gel matrix formation. In the second step, the samples without and the samples with the sol-gel matrix were immersed in a 1.0 × 10 -3 M AgNO 3 solution, at a liquor ratio of 1:25, and treated for 10 min at 60 °C under constant stirring in a Girowash machine (James Heal, GB). Th en, the sumac leaf extract solution was added, until the liquor ratio was 1:50, and the samples were treated in the solution for 60 min under the same conditions. For comparison, the fabric samples were treated with the sumac leaf extract solution, without the previous application of AgNO 3 . Aft er the treatment, the samples were rinsed in cold distilled water, squeezed and dried at room temperature. Th e procedures of the chemical modifi cations of the fabric samples and the corresponding sample codes are summarized in Table 1.

Analyses and measurements 2.4.1 Scanning electron microscopy (SEM) and
energy-dispersive X-ray spectroscopy (EDS) Untreated and chemically modifi ed cotton fabric samples were analyzed using a fi eld emission scanning electron microscope, FEG-SEM Th ermo Scientifi c Quattro S (Th ermoFischer Scientifi c, USA). Th e sample analysis was performed using an Oxford Instruments Ultim Max 65 Energy-dispersive Detector (EDS) and AZtec soft ware. Th e samples were coated with a thin layer of carbon before observation to provide conductivity and hence the quality of the images.

Antibacterial activity
Th e bacterial reduction on the functionalized samples was evaluated against the Gram-positive Staphylococcus aureus (ATCC 6538) and the Gram-negative

Tailoring of Antibacterial and UV-protective Cotton Fabric by an in situ Synthesis of Silver Particles in the Presence of a Sol-gel Matrix and Sumac Leaf Extract
Escherichia coli (ATCC 25922) bacteria, according to the standard method, ASTM E 2149-01. Th e reduction in the number of bacteria, R, was calculated as follows [20]: where R is the bacterial reduction, A is the number of bacteria colony forming units per ml (CFU/ml) in a fl ask containing a chemically modifi ed sample, aft er 1 hour of contact time, and B is the number of bacteria colony forming units per ml (CFU/ml) in a fl ask containing an unmodifi ed reference sample, aft er 1 hour of contact time. Two parallel assessments with eight CFU counts were carried out for each functionalized sample and the R value was reported as the mean value and the standard error.

UV protection properties
Th e UV protection properties of untreated and chemically modifi ed cotton fabric samples, before and aft er repetitive washings, were determined according to the AATCC TM 183 standard. Th e measurements were performed using a Varian CARY 1E UV/Vis spectrophotometer (Varian, Australia), containing a DRA-CA-301 integration sphere and Solar Screen soft ware. Th e transmission of the ultraviolet radiation through the samples were measured within the 280-400 nm spectral region, and the average transmittance (T) at the wavelengths between 315 nm and 400 nm (UV-A), 280 nm and 315 nm (UV-B) and 280 nm and 400 nm (UV-R) were determined from the measurements. Th e ultraviolet protection factor (UPF) was calculated as follows [21]: where E λ is the relative erythemal spectral eff ectiveness, S λ is the solar spectral irradiance, T λ is the spectral transmittance of the specimen, and Δλ is the measured wavelength interval in nm. Th e higher the UPF, the higher the protection. Th e UPF rating and UVR protection categories were determined from the calculated UPF values, according to the Australian/New Zealand Standard: Sun protective clothing -Evaluation and classifi cation [22]. Additionally, the transmission of the samples was measured within the 280-800 nm spectral region with the use of the UV/Vis spectrophotometer Lambda 800 (Perkin Elmer, UK) equipped by the integrating sphere PELA-1000.

Washing fastness
Th e fabric samples were washed once (1 W) and 5 times (5 W) in a Girowash machine (James Heal, GB), according to the ISO 105-C06 standard method. Th e washing cycles were performed in a SDC standard detergent solution, at a concentration of 4 g/l, at 40 °C for 45 min. Aft er washing, the samples were rinsed in distilled water at 40 °C for 1 min, subsequently rinsed in tap water, and then dried in air at room temperature.

Colour measurements
Th e CIELAB color coordinates of the untreated and chemically modifi ed cotton samples, before and after repetitive washings and illumination, were determined using a Datacolor Spectrafl ash 600 PLUS-CT spectrophotometer. Th e measurements were performed with a 30-mm aperture under D 65 illumination and an observation angle of 10°. Th e average of ten measurements was provided for each sample, and the color diff erence, ΔE*, was calculated using the following equation [23]: where ΔL*, Δa* and Δb* are diff erences between the color coordinates of the two samples.

Sample characterization
Th e SEM/BSE images, shown in Figure 2, revealed numerous bright spots on the surface of the CO/ Ag-S and CO-M/Ag-S samples, confi rming that the presence of phenols, such as gallic acid, myricetin and quercetin derivatives, myricetin 3-rhamnoside, quercetin 3-glucoside as well as penta to decagalloyl-glucosides in the sumac leaf extract [18,19,24], successfully converted Ag + to Ag o in the reduction reaction. Th e results also show that the application of a sol-gel matrix (CO-M sample) did not significantly change the fi ber surface morphology, but it importantly infl uenced the adsorption ability of Ag + . A comparison of CO/Ag-S and CO-M/Ag-S clearly showed that the amount of Ag was signifi cantly  Th e presence and distribution of the Ag element on the surface of the CO-M/Ag-S sample was confi rmed by the EDS spectrum and element mapping images of C, Ag and O (Figure 3). Th e EDS spectrum showed a strong characteristic peak, corresponding to Ag at 2.984 keV. Furthermore, the element mapping images suggested that Ag was rather homogeneously distributed on the fi ber surface, along with the intrinsic elements, C and O.

Functional properties of the chemically modifi ed samples
Th e photo images of the untreated and chemically modifi ed cotton fabric samples, shown in Figure 4, revealed that the application of the sol-gel matrix did not cause a visible color change in the cotton fi bers, which generally remained white. In contrast, the treatment of cellulose fi bers with sumac leaf extract colored the CO/S and CO-M/S samples in yellow (an increase in the positive value of the coordinate b*), which was slightly more yellow, if the sol-gel matrix was present (CO-M/S sample). Th e in situ synthesis of Ag Ps in the presence of the sumac leaf extract converted the yellow color of the cotton fi bers into a brown color, caused by a decrease in the values of the L* and b* coordinates, as well as a change of the a* coordinate sign from negative to positive. Th e color change was more intense for the CO-M/Ag-S sample than for CO/Ag-S sample. Th ese results clearly indicated that the presence of the sol-gel matrix increased the adsorption ability of the cotton fi bers, for both the sumac leaf extract and AgNO 3 , and that the reduction of Ag + to Ag° was accompanied by an intense color change. Th e latter was in accordance with the reports in the literature, in which the color change of the solution or of the textile substrates was chosen as the criteria for the formation of Ag Ps [25]. Th e antibacterial properties, presented in Figure 5, showed that not only Ag Ps (the CO-M/Ag-S sample), but also the sumac leaf extract (the CO-M/S sample) exhibited antibacterial activity. While the concentration of Ag on the cotton fi bers was high enough to cause a 99-100% reduction of both E. coli and S. aureus bacteria, the phenolic compounds present in the water extract of the sumac leaves caused an excellent 99% growth reduction of S. aureus. On the other hand, the sumac leaf extract did not inhibit the growth of E. coli, but in contrast, it even promoted the bacterial growth which resulted in negative values of R. Th ese fi ndings were reasonable, since the substances in the sumac water and alcohol extracts are, in general, recognized as strong antibacterial agents against Gram-positive bacteria. Th e results also showed that the antibacterial activity of the CO-M/Ag-S sample was highly wash resistant, since a 100% bacterial reduction was obtained aft er fi ve washings. Th is phenomenon was not observed for the CO-M/S sample, where the antibacterial substances were partially desorbed from the cotton fi bers during the washing of the sample. A desorption of the sumac extract during repetitive washing resulted in a lightening of the color, which is expressed by the values of ΔE* ab in Figure 6. Th e lowest color change was determined for the CO-M/Ag-S sample, suggesting the durability of the chemical modifi cation.
Th e results in Figure 7 revealed that the presence of the sumac leaf extract (CO/S and CO-M/S samples) signifi cantly decreased the transmission in the 280-400 nm spectral region in comparison to the CO and CO-M samples. Th is was attributed to the UV absorbing action of the aromatic phenolic compounds included in the sumac leaf extract. In the visible light spectrum (400-800 nm), the transmission of CO/S and CO-M/S samples gradually increased with increasing wavelength and almost reached the values of 37-40 % in the 700-800 nm spectral region, which were characteristic for the CO and CO-M samples, respectively, in the whole visible spectrum. Th ese results confi rmed that lower wavelengths of visible light were absorbed by the yellow pigments of the sumac leaf extract. In contrast, the transmission of the CO/Ag-S and CO-M/ Ag-S samples was very low in both UV and visible spectral region and it did not exceed 6 % even at 800 nm. Th is phenomenon implies that the brown colored Ag Ps successfully prevented the transmission in the whole measured spectral region. Th e calculated UPF values summarized in Table 2 are in accordance with the results presented in Figure 7. Accordingly, the presence of the sumac leaf extract drastically increased the UPF, from 3.

Conclusion
In this research, we successfully created a highly durable antimicrobial and UV-protective coating on cellulose fi bers by an in situ synthesis of Ag Ps in the presence of an extract of sumac leaves, which was used as a reducing agent. Th e results showed that: the sumac leaf extract colored the cellulose fi bers in yellow, and the conversion of Ag cations to Ag Ps in the presence of the sumac leaf extract caused the fi ber to be colored in brown; the concentration of Ag on the cotton fi bers was high enough to cause a 99-100% reduction of both E. coli and S. aureus bacteria; the sumac leaf extract exhibited excellent anti-bacterial activity against S. aureus but did not inhibit the growth of E. coli; the sumac leaf extract provided high UV-protec-tion, with a UPF value equal to 44.44, which was increased to 66.7, if Ag Ps were present on the cellulose fi bers; the presence of the sol-gel matrix increased the adhesion ability of the cellulose fi bers for the sumac leaf extract and Ag cations, resulting in an increased antibacterial activity and UV-protection properties of the chemically modifi ed fi bers, as well as their washing fastness.