2019/4/17 Fabrication of silver nanoparticle sponge leather with durable antibacterial property - ScienceDirect

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Journal of Colloid and Interface ScienceVolume 514, 15 March 2018, Pages 338-348

Regular Article

Fabrication of silver nanoparticle sponge leather with durable antibacterialproperty

Gongyan Liu , Haiqi, Gao , Kaijun Li , Jun Xiang , Tianxiang Lan , Zongcai Zhang

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https://doi.org/10.1016/j.jcis.2017.09.049

AbstractLeather product with durable antibacterial property is of great interest both from industry and consumer’s pointof view. To fabricate such functional leather, gallic acid modified silver nanoparticles (GA@AgNPs) were first insitu synthesized with a core–shell structure and an average size of 15.3 nm. Due to its hydrophilic gallic acidsurface, the GA@AgNPs possessed excellent stability and dispersibility in wide pH range from 3 to 12 and alsoshowed effective antibacterial activity with a minimum inhibitory concentration (MIC) of around 10 μg mL .Then, such GA@AgNPs were used as retanning agent to be successfully filled into leather matrix during theleather manufacturing process. Moreover, taking the advantage of its high surface density of carboxyl groups,these GA@AgNPs could be further chemically cross-linked onto collagen fibers by chrome tanning agent. Afterretanning, the resultant leather was given a “AgNPs sponge” feature with high payload of silver nanoparticlesagainst laundry, exhibiting high and durable antibacterial activity.

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2019/4/17 Fabrication of silver nanoparticle sponge leather with durable antibacterial property - ScienceDirect

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Antibacterial; Silver nanoparticle; Gallic acid; Leather; Retanning

1. IntroductionLeather is a durable and flexible material produced by tanning the collagen fiber network of animal hides andskins [1]. In leather manufacture, chrome tanning is the most popular tanning method based on chemical cross-linking of the collagen fibers by forming complexation between Cr and carboxyl groups on the collagen fibers,giving the leather matrix known as wet-blue leather with enhanced hydrothermal stability and mechanicalstrength [2]. After chrome tanning, retanning process is usually needed by using acrylic resin e.g. poly(acrylicacid) (PAA) with multi carboxyl groups to fill into leather matrix and form complexation with Cr for improvingutilization of chrome tanning agent and leather fullness [3]. Then, the resultant leather are suitable for makingvarious goods. Because of its breathable, soft and comfortable nature, leather goods, such as shoes, clothing andbags have been widely used in daily life [4]. Especially, the excellent moisture permeability of leather goods canabsorb sweat from the sweat glands of skin, which bring comfort to users [5]. But in the meantime, the absorbedsweat containing proteins may provide nutrient sources for the growth of bacteria on the leather surface [6].Besides, the collagen fibrous network of leather can also offer other ideal conditions such as moisture,temperature and oxygen for bacterial growth and rapid colonization, resulting in the formation of a biofilm whichcan lead to unpleasant odor, discoloration, reduced mechanical strength and even risks of skin infection [7], [8].Therefore, the antibacterial property of leather product has been a major issue to be addressed, both at industriallevel and from the consumer’s point of view.

To solve this problem, considerable efforts have been directed toward constructing various antibacterial coatingson leather surface [9], [10], [11], [12], [13]. For example, chitosan has been applied as antimicrobial coatingmaterial for leather surface based on its contact-active disruption of microbe cell membrane [12], [14], [15], [16],[17], [18]. Recently, our group have prepared a PEGylated chitosan (PEG-g-CS) coating on leather, whichexhibited more efficiently antibacterial activity than that of single chitosan coating due to the synergistic effectbetween bacterial resistance of PEG and contact-killing of chitosan [13]. Despite the high antibacterial efficiencyof these polymer coating, one important defect of these coatings in application is that they can be easily damagedand even fell off from leather surface by abrasion during wearing and laundry, causing loss of antibacterialactivity [19]. Therefore, incorporating biocides into leather matrix with sustained release behavior will be an idealstrategy for leather products with durable antibacterial property. Though chemical biocides were commonly usedin leather industry, they were mainly play the role in preventing biodeterioration of hides rather than to endowantibacterial property to the final products [6], [20]. What’s more, some of these biocides have been restricteddue to human health and environmental issues [8]. Thus, the development of eco-friendly and effectivelyantimicrobial bactericide that can be applied in leather tanning process will be great interesting.

Silver nanoparticles (AgNPs) are well known for their strong and broad-spectrum of bacterial and fungal strainsthrough the sustained release of Ag ions, damaging the microbial cell membrane as well as disrupting thefunction of bacterial enzymes and nuclei acid groups in protein and DNA [21], [22], [23], [24], [25], [26], [27],[28]. In addition, AgNPs are less toxic to human as compared to other metals and have been widely selected assuitable antibacterial finishing agent for textile fibers [26], [29], [30], [31], [32], [33], [34]. Inspired by theseAgNPs-based antibacterial finishing, durable antibacterial leather is supposed to be fabricated via filling AgNPsinto leather matrix and subsequently immobilized on collagen fibers during leather tanning process. However,due to the complicated and versatile pH environments of the whole tanning process, the application of AgNPs inleather manufacture is still a challenge that requires high stability and dispersibility of AgNPs. In this work, gallicacid stabilized silver nanoparticles (GA@AgNPs) were in situ synthesized by chemical reduction of AgNO , withexcellent stability and dispersibility. Due to the high carboxyl group density on its surface, these GA@AgNPs wereused as retanning agent instead of traditional acrylic resin to filled into the leather matrix. Simultaneously, theseGA@AgNPs could be chemically immobilized onto collagen fibers through the cross-linking of carboxyl groupsbetween GA and collagen fiber by chrome tanning agent, resulting a “AgNPs sponge” leather with high payload ofsilver nanoparticles (Scheme 1). Based on the sustained release behavior of Ag , the resultant leather or its

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2019/4/17 Fabrication of silver nanoparticle sponge leather with durable antibacterial property - ScienceDirect

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products possessed durable and even long-term antibacterial activity against laundry and mechanical abrasion.To the best of our knowledge, this is the first report about durable antibacterial leather with “AgNPs sponge”feature.

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Scheme 1. Schematic application of GA@AgNPs as retanning agent for fabricating “AgNPs sponge leather”with durable antibacterial property.

2. Materials and methods

2.1. Materials

Gallic acid (GA, 99%) was purchased from AiBi Chemical Reagents Corporation (Shanghai, China). Silver Nitrate(AgNO , 99.8%) was purchased from Tianhua Technological Incorporated Company (Chengdu, China). BasicChromium Sulfate [Cr(OH)SO ] containing 25% Cr O was purchased from Tianjin Mingyang Chemical Co., Ltd.(Tianjin, China). Sodium borohydride (NaBH , >96%) and other Chemical materials are purchased from JinshanChemical Reagents Corporation (Chengdu, China). Silver standard solution with the silver concentration of1000 µg mL was purchased from Jinan Zhongbiao Technology Co. Ltd. (Jinan, China). Pickled pelt was madeby treating sheep skin follow the process of degreasing, unhairing, liming, deliming, bating and picking in ourown lab.

2.2. Synthesis of gallic acid modified silver nanoparticles (GA@AgNPs)

10 mL AgNO (5 mM) was first completely mixed with 10 mL gallic acid solution at the concentration of 0.5, 2.5,3 and 5 mM, respectively. Then the above mixture was added dropwise into 30 mL NaBH solution with aconcentration of 10 mM for reacting 2 h at dark. The obtained GA@AgNPs were further purified by centrifugationat 15,000 rpm for 20 min and re-dispersed into water with a concentration of 50 μg mL .

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2.3. The colloidal stability of GA@AgNPs

To determine the stability of GA@AgNPs with respect to pH, a series of GA@AgNPs solutions with the pH valuefrom 1 to 13 were prepared. Then, the UV–vis spectra, size distribution and zeta-potential of these nanoparticlesolutions were measured, respectively. To determine the stability of GA@AgNPs with respect to storage time, theUV–vis spectra, size distribution and zeta-potential of GA@AgNPs solutions storage at 1, 30 and 120 days underroom temperature were measured, respectively.

2.4. Antibacterial assay for GA@AgNPs

The antibacterial activity of GA@AgNPs was conducted by the minimum inhibitory concentration (MIC) test,with Escherichia coli (E. coli, ATCC25922) and Staphylococcus aureus (S. aureus, ATCC6538) as model bacteria.GA@AgNPs in sterile water with an initial concentration of 50 μg mL were prepared and diluted to 30, 20, 10,and 5 μg mL by using sterile water, respectively. Then, 1 mL GA@AgNPs solution was added into tubes with5 mL LB medium at bacteria concentration of 5 × 10 cfu mL . For the blank samples, 1 mL sterile water wasadded into the control tube with 5 mL LB medium at bacteria concentration of 5 × 10 fu mL . The tubes wereincubated at 37 °C with shaking at 200 rpm for 24 h in a shaking bath. After incubation, 20 μL solution waswithdrawn from tubes and spread on Luria-Bertani agar in petri dish. The petri dishes were incubated for 24 h at37 °C, and counted for colony-forming units. Antibacterial activity was defined as the percentage of microbereduction. The percentage of colony number reduction was then determined according to the following equation:

where A is colony count of the petri dish containing the GA@AgNPs and B is the colony count of the petri dishcontaining the sterile water as blank control. According to the result, the MIC was defined as the lowestGA@AgNPs concentration that lead to the percentage of microbe reduction >90%.

2.5. The application of GA@AgNPs as retanning agent for chrome tanned leather

The detail of the tanning process could be found in Table S1. Briefly, 100 g wet pickled pelt with a pH of 3.0 wasplaced into a 500 mL beaker flask with 150 mL water and 1.0 g sodium formate, shaking in water bath with aspeed of 60 rpm for 1 h at 30 °C. For chrome tanning, 3.0 g basic chromium sulfate was added into the flask andreacted for 1.5 h at 30 °C. When the chrome tanning process was finished, the picked pelt was converted to wet-blue leather and 1.5 g sodium bicarbonate was added for increasing the pH of leather to 4.0. Then, the wet-blueleather was taken out, washed completely by 200 mL water to remove unutilized basic chromium sulfate. Thenthe leather was put into 150 mL water containing 1.0 g sodium bicarbonate for increasing the pH of leather to 5.5and accelerating the cross-linking between Cr and collagen fibers. For retanning, 200 mL GA@AgNPs solutionwith concentration of 500 μg mL was added into above solution and reacted with the wet-blue leather for 1.5 hat 30 °C. After retanning, the leather was taken out and washed by 200 mL water in a heating drum at 40 °C for1.0 h at a spinning rate of 40 rpm, removing the un-incorporated GA@AgNPs under mimicked laundry condition.Wet-blue leather after chrome tanning was used as control leather sample and defined as pristine leather in thefollowing investigation. Dry leather samples were prepared by vacuum drying for 24 h at 40 °C.

2.6. Antibacterial activity assays for GA@AgNPs retanned leather

2.6.1. Killing efficiency assay by contact or controlled release

Each rounded dry leather sample with a weight of 100 mg was fist washed by sterilized water for several timesand put into 5 mL sterilized water with bacterial concentration of 5 × 10 cfu mL in a tube. The tubes wereincubated at 37 °C with shaking at 200 rpm in a shaking bath. At pre-determined time point, 50 μL solution waswithdrawn from tubes for preparing a series of tenfold dilutions and spread on Luria-Bertani agar in petri dish.The petri dishes were incubated for 24 h at 37 °C, and counted for colony-forming units. The bacterial killingefficiency is defined as:

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2.7. Zone of inhibition test of GA@AgNPs retanned leather

50 μL sterilized water with bacterial concentration of 5 × 10 cfu mL was spread on LB agar in petri dish,incubating with rounded dry leather sample with a diameter of 13 mm for 24 h at 37 °C. The presence of clearzone around the samples was recorded as an inhibition against the bacteria.

2.8. Long-acting antibacterial test of GA@AgNPs retanned leather

10 mL liquid LB media with bacterial concentration at an OD of 0.05 was first prepared. The bacteria wereincubated with 100 mg leather samples in test tubes on an automated shaker at 37 °C. At pre-determined timepoints, 150 μL of growth media removed, and the OD was measured using UV–vis spectrophotometer.

2.9. The release behavior of Ag from GA@AgNPs retanned leather

The release profile of silver ions from GA@AgNPs retanned leather was investigated by using inductively coupledplasma atomic emission spectroscopy (ICP-AES). First, the silver content of the GA@AgNPs retanned leather wasmeasured by ICP-AES. Briefly, 1 g dry GA@AgNPs retanned leather was cut into pieces and dissolved in 10 mLmixture of concentrated nitric acid (HNO ) and H O (8:2, v:v) for nitrolysis. Then the acidic solution was dilutedwith water to 100 mL, and the silver concentration was measured by ICP-AES and determined by the silvercalibration curve. The silver calibration curve was prepared by diluting the Silver standard solution with the silverconcentration of 1000 μg mL to 20, 10, 5, 1, 0.5 μg mL , respectively.

For determining the release rate of silver ions, 1 g GA@AgNPs retanned leather was put into 10 mL water andincubated on an automated shaker at 37 °C. At predetermined time intervals, 1 mL solution was withdrawn fromthe release media and another 1 mL fresh water was added. Then the amount of released Ag was determined byusing ICP-AES.

2.10. Characterization

The formation of AgNPs using GA as stabilizer in the aqueous solution was monitored by UV–visspectrophotometer (F96Pro, China). The size distribution and zeta potential of GA@AgNPs were tested by NanoZS instrument (Malvern, England). The zeta potential of GA@AgNPs was measure in water. Before measuring thesize of GA@AgNPs, the GA@AgNPs solution was first filtered by using 0.22 μm membrane filter. The morphologyof GA@AgNPs was measured by transmission electron microscope (TEM, JEOL, Japan). TEM sample wasprepared by drying a drop of GA@AgNPs solution onto a carbon-coated copper grid. The surface morphology ofthe leather fibers immobilized GA@AgNPs was observed by scanning electron microscopy (SEM, Hitachi S-4800)coupled with energy-dispersive X-ray spectroscopy (EDX) at an accelerating voltage of 3.0 kV. X-rayphotoelectron spectra (XPS, PHI-5300 ESCA, USA) of the control leather sample and GA@AgNPs retannedleather were recorded with an Al Kα source (hv = 1486.6 eV). Water contact angle was used to measure thesurface hydrophilicity of leather via using OCA20 (Dataphysis, Germany). All the contact angles were determinedby averaging values measured at 5–6 different points on each surface. The shrinkage temperature (T ) of leathersample was tested by shrinkage temperature tester (DL07-PS-83, China).

3. Results and discussion

3.1. Preparation and characterization of gallic acids stabilized silver nanoparticles (GA@AgNPs)

As shown in Scheme 2, GA@AgNPs were in situ synthesized by chemical reduction of AgNO with gallic acids asstabilizers based on the chelation between two phenolic hydroxyl groups of GA and silver ion [35], [36], [37]. Dueto the hydrophilic carboxyl groups, gallic acids were supposed to prevent the aggregation of the nanoparticles andprovide good dispersibility in water. To investigate the optimum synthesis condition of the silver nanoparticles,four groups of GA@AgNPs were prepared with the molar ratio of GA, AgNO and NaBH at 0.1: 1.0: 6.0, 0.5: 1.0:6.0, 0.6: 1.0: 6.0 and 1.0: 1.0: 6.0, respectively. UV–visible spectrum was first used to evaluate these GA@AgNPsand the results were shown in Fig. 1A. It was clear that all spectra showed the characteristic surface plasmonresonance (SPR) absorption peak of AgNPs at around 405 nm, indicating the successful formation of spherical

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silver nanoparticles. In Fig. 1B, the average sizes of these GA@AgNPs were determined by dynamic lightscattering (DLS) as 21.1 nm, 19.6 nm, 15.8 nm and 15.3 nm, corresponding to molar ratio of GA at 0.1, 0.5, 0.6and 1.0, respectively. It can be seen that the average sizes of GA@AgNPs decreased slightly as the increase of GAmolar ratio. For the GA@AgNPs with GA molar ratio at 0.1, the zeta-potential was only −5.6 mV which was muchlower than that of other three silver nanoparticles with similar zeta-potential value at around −25.0 mV (Fig. 1B).That was due to its lowest GA density on the nanoparticle surface. Actually, GA@AgNPs with GA molar ratio at0.1 and 0.5 were not stable and would aggregate to visible precipitation within several hours. The result indicatedthe GA density of these two silver nanoparticles was too low to provide enough hydrophilicity for keeping theirstability in water. On the contrary, once the molar ratio of GA was above 0.5, GA@AgNPs could remain stable inwater for long time. For example, the GA@AgNPs with GA molar ratio at 1.0 was able to keep stable in water forseveral months under room temperature. Therefore, the molar ratio of GA, AgNO and NaBH at 1.0: 1.0: 6.0 wasconsidered as the optimum synthetic condition, and GA@AgNPs prepared under this condition were used in thefollowing studies.

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Scheme 2. Schematic illustration of synthetic rout of gallic acid stabilized AgNPs.

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Fig. 1. (A) UV–vis spectra, (B) average size (■) and zeta-potential (▴) of four GA@AgNPs with differentmolar ratios of gallic acids; TEM image (C) and size distribution histograms (D) of GA@AgNPs preparedwith the molar ratio of GA, AgNO and NaBH at 1.0: 1.0: 6.0. The inset in TEM image showed thephotographs of GA@AgNPs solution.

The morphology of GA@AgNPs was then observed by TEM, which demonstrated the well-formed sphericalnanoparticles with good dispersion (Fig. 1C). The photograph in the inset of TEM image exhibited bright yellowin color of these silver nanoparticles. The size distribution histograms based on artificially analyzing 200 AgNPsfrom TEM images was shown in Fig. 1D. The average diameter of the silver nanoparticle was calculated as about8.5 nm which was smaller than 15.3 nm determined by DLS, implying a core–shell structure of such GA@AgNPswith the silver core of 8.5 nm and the GA shell thickness of about 3.4 nm. The hydrophilic GA layer could stabilizethe silver nanoparticles and provide the excellent dispersibility, consisting with the observed single silver particlein Fig. 1C.

3.2. The colloidal stability and antibacterial activity of GA@AgNPs

As mentioned above, GA@AgNPs prepared with the molar ratio of GA, AgNO and NaBH at 1.0:1.0:6.0 couldkeep their stability for several months. Fig. 2A and B showed that the UV–vis spectra, average size and zeta-potential of these nanoparticles almost didn’t change within 120 days, indicating excellent stability during storageat room temperature. The colloidal stability of GA@AgNPs was also investigated under different pH conditions.Fig. 2C revealed the UV adsorption curves of GA@AgNPs solutions with pH from 3 to 12 were nearly same,demonstrating the AgNPs were stable in a wide pH ranged from 3 to 12. Fig. 2D also showed the average sizes andzeta-potential of GA@AgNPs were stable from pH 3 to 12. It is worth noting that the characteristic absorptionpeak of GA@AgNPs at pH 1 and pH 2 disappeared in Fig. 2C. In fact, GA@AgNPs were not stable when pH waslower than 3 and would aggregate immediately, which was due to the protonation of the carboxyl groups of gallicacid in strong acid solution. After protonation, the hydrophilicity of gallic acid was reduced and could not provideenough stability for GA@AgNPs. While for GA@AgNPs at pH 13, its maximum absorbance at 405 nm was lowerthan that at pH 3–12 with a larger nanoparticle size. That might be ascribed to the weak chelation effect betweengallic acid and silver nanoparticle surface in strong basic solution, leading to slight aggregation.

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Fig. 2. UV–vis spectra of GA@AgNPs at different storage time (A) and different pH (C); The average size(■) and zeta-potential (●) of GA@AgNPs at different storage time (B) and different pH (D).

The antibacterial activity of the GA@AgNPs was evaluated by testing its minimum inhibitory concentration(MIC) against E. coli and S. aureus as Gram-negative and Gram-positive bacteria models, respectively. Becausethe MIC value was defined as the lowest material concentration that led to the percentage of microbe reduction≥90%, which can be quantitatively evaluated using viable cell count method. GA@AgNPs with an initialnanoparticle concentration of 50 μg mL were diluted to 30, 20, 10, and 5 μg mL by using LB medium,respectively, and LB medium with equal volume was used as control. The MIC value will be the dilutedconcentration which can give the antibacterial percentage ≥90%. Fig. 3B and E showed the antibacterial rates ofGA@AgNPs at concentration of 5 μg mL were lower than 90% against E. coli and S. aureus. However, it wasclear that the antibacterial activities of GA@AgNPs at concentration of 10 μg mL against E. coli and S. aureuswere higher than 99.9% (Fig. 3C and F). Therefore, the MIC value of GA@AgNPs was demonstrated as about10 μg mL .

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Fig. 3. The antibacterial activities of GA@AgNPs at concentration of 5 μg mL (B, E) and 10 μg mL (C,F) against E. coli and S. aureus after 24 h incubation. (A) and (D) are blank controls.

3.3. The application of GA@AgNPs as retanning agent for chrome tanned leather

Tanning is the process of treating skins and hides of animals to produce leather which is more durable and lesssusceptible to decomposition. As shown in Scheme 1, chrome tanning usually started from pickled pelt with a pHof 3. The acidic pH was able to ensure that chrome tanning agent [Cr(OH)SO ] could uniformly permeate into theinterval of the collagen fibers. Once the desired level of penetration of chrome was achieved, the pH was raised to5.5 for facilitating the chemical complexation between chrome ions (Cr ) and carboxyl groups of collagen fibers,resulting in cross-linking of collagen fibers. The cross-linking of collagen fibers by Cr converted pickled peltinto wet-blue leather with higher hydrothermal stability and mechanical strength. For example, the shrinkagetemperature (T ) of pickled pelt would be increased from 53 °C to 85 °C after chrome tanning.

For traditional leather tanning process, acrylic resin such as poly(acrylic acid) (PAA) with multi carboxyl groupswas commonly used as retanning agent to fill into leather matrix and form complexation with Cr , leading tohigher utilization of chrome tanning agent and improved leather fullness [3]. Therefore, GA@AgNPs with highdensity of carboxyl groups on their particle surface were supposed to be used as retanning agent instead of PAA.Besides, the interval between collagen fibers is several hundred nanometers (100–1000 nm) which is alsosuitable for the filling of GA@AgNPs with average size of ∼15.3 nm [38]. More than that, it can be alsohypothesized that GA@AgNPs will be further chemically immobilized onto collagen fibers by Cr cross-linkingduring the retanning process (Scheme 1). To verify above hypothesis, the UV absorbance (ABS) of GA@AgNPssolutions was tested before and after retanning to investigate the filling of sliver nanoparticles to leather. Asshown in Fig. 4, the maximum ABS of GA@AgNPs solutions at 405 nm was 1.82 before retanning and thesolution color was black due to the high concentration of the silver nanoparticles. However, after retanning, itsABS was dramatically decreased to 0.34 after retanning and the color of GA@AgNPs solution also became clear.These result revealed that almost all GA@AgNPs had been filled into the leather matrix during retanning process.In addition, the color of leather sample was also changed from blue to black after retanning, due to the filling oflarge amount of GA@AgNPs inside leather. After retanning, the leather was taken out and washed by water in aheating drum at 40 °C for 1.0 h at a spinning rate of 40 rpm for mimicking laundry condition, and the washingsolution was collected for UV absorbance testing. The ABS at 405 nm of the washing solution was determined asonly 0.03, indicating nearly no GA@AgNPs was washed out. The above results implied that GA@AgNPs not onlyhad been filled into the leather matrix but also chemically immobilized onto collagen fibers, resulting in a “AgNPssponge” leather with high payload of silver nanoparticles and laundry resistant property.

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Fig. 4. The UV absorbance of GA@AgNPs solutions before retanning, after retanning and after washing.The inset is the photograph of GA@AgNPs solutions before retanning, after retanning and after washing.

The distribution of GA@AgNPs inside of such “AgNPs sponge” leather was further observed by scanning theparallel section using SEM. Compared to pristine leather without GA@AgNPs retanning (Fig. 5A), it was clearlyto see that the collagen fibers of GA@AgNPs retanned leather (Fig. 5B) were more tight and the inter spacebetween collagen fibers was completely filled by GA@AgNPs. What is more, GA@AgNPs observed as white dotswere uniformly distributed along the collagen fibers (as showed by red arrows). The EDX spectra in Fig. 5C and Dalso provided the obvious signal of silver element of the GA@AgNPs retanned leather. These resultsdemonstrated that GA@AgNPs have been successfully filled into the leather matrix during the retanning processand further chemically immobilized onto the collagen fibers through cross-linking by Cr . The chemical state ofsilver in leather was also ascertained by XPS analysis. Compared to Fig. 6A, the survey scan spectrum ofGA@AgNPs retanned leather exhibited the presence of Ag 3d signal in Fig. 6B. The Ag 3d core-level XPS spectrawas further shown in Fig. 6C, in which two peaks at 368.3 and 374.3 eV were corresponded to Ag 3d and Ag3d binding energies, respectively. The splitting of the 3d doublet of Ag is 6.0 eV, implying the formation ofmetallic AgNPs on the collagen fiber surface [39].

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Fig. 5. SEM images of parallel section of (A) pristine leather without GA@AgNPs retanning and (B) leatherwith GA@AgNPs retanning, with corresponding EDX spectra (C, D). The scale bar in the SEM image is100 nm.

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Fig. 6. XPS spectra of pristine leather (A), GA@AgNPs retanned leather (B) and Ag 3d core-level ofGA@AgNPs retanned leather (C).

Since silver nanoparticles were immobilized onto the micro-scaled collagen fibers (as shown in Fig. 5B), theroughness of collagen fiber surface was enhanced due to such micro- and nanoscaled dual structure, which mightchange the surface wettability of leather [26]. Therefore, water contact angle was used to investigated thewettability of leather surface. As shown in Fig. 7, the water contact angle of GA@AgNPs retanned leather at 10 swas 130° which was much larger than that of 86° of pristine leather, exhibiting a hydrophobic surface. Within120 s, GA@AgNPs retanned leather remained its hydrophobicity with the water contact angle slightly decreasingto 113°. While for pristine leather, the water contact angle rapidly decreased to 0° at 60 s. These results revealedthat the water repellency of leather was improved after GA@AgNPs retanning. In fact, water repellency was veryuseful for leather or its product. Because the mechanical strength of leather can be easily damaged by water

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penetration, causing leather product e.g. shoes out of its shape. Therefore, the GA@AgNPs immobilized oncollagen fiber not only endowed “AgNPs sponge” feature to resultant leather but also improved water repellencyof leather.

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Fig. 7. The change of water contact angle of GA@AgNPs retanned leather and pristine leather with time.

3.4. The antibacterial activity of the“AgNPs sponge” leather

After retanning, the resultant leather can be seemed as “AgNPs sponge” which will continuously release Ag andprovide durable antibacterial activity by killing the attached bacteria on leather surface. The release behavior ofAg from GA@AgNPs retanned leather was investigated by inductively coupled plasma atomic emissionspectroscopy (ICP-AES). Fig. 8 shows that the silver content was about 6.4 mg per gram dry leather and theaccumulative release rate of Ag was only 0.08% when 1 g dry GA@AgNPs retanned leather was dipped into10 mL water for 72 h at 37 °C, with the released Ag ions reaching a final concentration of 0.51 μg mL . Theseresults suggested that the release of Ag exhibited an extremely sustained behavior and such GA@AgNPsretanned leather could provide long-acting antibacterial activity. Therefore, the killing efficiency of GA@AgNPsretanned leather was first investigated by challenging the leather with E. coli and S. aureus at a concentration ofabout 5×10 cfu mL , respectively. As shown in Fig. 9A, the killing efficacy was increased with contact time andreached over 99% after 120 min, demonstrating excellent antibacterial activity against E. coli and S. aureus.Consistent with the contact killing result, distinct inhibition zones around GA@AgNPs retanned leather wereobserved on both E. coli and S. aureus agar plates (Fig. 9B). However, it was worth to not that the inhibition zoneof GA@AgNPs retanned leather was not such very remarkable in comparison with that in the contact killing test.That was because the release the release of Ag from dry leather would be very slow and only bacteria closed toleather surface could be killed. While for contact killing test, leather sample was immersed into bacterial solutionand the release of Ag from leather into bacterial solution would be faster and exhibited significant higher killingefficiency. On the contrary, the pristine leather that only tanned by Cr did not show any antibacterial activitywith growth of bacteria even on the leather sample. This also implied that the chrome tanning agent would notaffect the antibacterial property of GA@AgNPs and the antibacterial activity of the GA@AgNPs retanned leatherwas given by GA@AgNPs. The above results also demonstrated that such excellent killing efficacy of GA@AgNPsretanned leather was based on the synergistic effect of Ag release and its contact killing.

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2019/4/17 Fabrication of silver nanoparticle sponge leather with durable antibacterial property - ScienceDirect

https://www.sciencedirect.com/science/article/pii/S0021979717310809?via%3Dihub 13/19

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Fig. 8. The release of Ag from GA@AgNPs retanned leather.

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Fig. 9. The killing efficiency (A) and inhibition zones (B, C) of GA@AgNPs retanned leather against E. coliand S. aureus.

To demonstrate the long-acting antibacterial activity of such “AgNPs sponge”, 100 mg leather sample was addedinto 10 mL culture medium precultured E. coli and S. aureus with an optical density at 600 nm (OD ) ofaround 0.05. Sterilized water and pristine leather was used as control. The growth of OD values for both E. coliand S. aureus over time was recorded. As shown in Fig. 10, the OD values for pristine leather increased fastwith culture time, which was similar with blank control and demonstrated pristine leather do not have anyantibacterial property. In contrast, OD values for both E. coli and S. aureus incubated with GA@AgNPsretanned leather nearly did not increase within 24 h, suggesting the sustained release of Ag from the leathercould offer long-acting inhibition of bacterial growth on leather surface.

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2019/4/17 Fabrication of silver nanoparticle sponge leather with durable antibacterial property - ScienceDirect

https://www.sciencedirect.com/science/article/pii/S0021979717310809?via%3Dihub 14/19

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Fig. 10. Growth curves of E. coli (A) and S. aureus (B) as a function of culture time in the presence ofGA@AgNPs retanned leather. Sterilized water and pristine leather was used as control.

4. ConclusionIn this work, gallic acid modified silver nanoparticles with an average size of 15.3 nm were facilely prepared,possessing excellent colloidal stability and antibacterial activity. Due to its nanoscale size and high surfacedensity of carboxyl groups, these GA@AgNPs were selected as retanning agent instead of traditional acrylic resinfor leather tanning. During the retanning process, GA@AgNPs were filled into the leather matrix and furtherchemically immobilized onto the collagen fibers by chromium cross-linking, with laundering durability. Afterretanning, the resultant leather was like “AgNPs sponge” which exhibited excellent and long-term antibacterialability, taking advantage of its high payload of AgNPs and sustained release of Ag . Such GA@AgNPs retannedleather will be popular both by industry and consumer.

AcknowledgmentThis work was supported by Natural Science Foundation of China (NSFC 51403131) and China PostdoctoralScience Foundation Funded Project (2015M570783).

Appendix A. Supplementary material

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Supplementary data 1.

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