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Polymers 2020, 12, 440 11 of 15 3.3. Analysis of Ag-NPs and Films’ Antibacterial Activity The Ag-NPs solution showed strong antibacterial properties. Based on the difference in OD600 values, the MIC values against E. coli and S. aureus were 32 μg/mL and 64 μg/mL, respectively, revealing that the solution has been able to inhibit bacterial growth and reproduction at lower concentrations. Studies have shown that the bactericidal mechanism of Ag-NPs is mainly caused by silver clustering and anchoring to the negatively charged sites of bacterial cell wall, and then destroying the cell wall to kill bacteria [44]. However, the resistance of Gram-positive S. aureus with thicker cell wall to antibacterial agents is higher than that of Gram-negative E. coli with thinner cell wall [45]. Therefore, the antibacterial ability of Ag-NPs against E. coli was slightly stronger than S. aureus, and the MIC values of S. aureus were higher than E. coli. Based on the MBC test result, the bactericidal ratio of E. coli was more than 95% when the Ag-NPs concentration increased to 64 μg/mL, indicating that 64 μg/mL was the MBC value of Ag-NPs against E. coli, and the MBC value of Ag-NPs against S. aureus was 128 μg/mL can be determined as well. Lower MIC and MBC values suggest that a small amount of silver Ag-NPs could make the film antibacterial. The results of the inhibition zone test (Figure 10) showed that only CANF0.02, CANF0.04, and CANF0.08 have obvious circles against both bacteria. The increase of Ag-NPs concentration enlarged the Inhibition Zone of films, suggesting that the antibacterial activity of films is determined by the concentration of Ag-NPs. The inhibitory zone diameter of CANF0.04 and CANF0.08 against E. coli was greater than 5 mm, and the diameter against S. aureus was greater than 3 mm. This indicated that the Ag-NPs in composite films owned antibacterial diffusivity when the concentration reached to a certain degree. Studies have shown that Ag-NPs has cytotoxicity in vitro experiments. When the concentration was exceeded 1 mg/kg, experimental mice exhibited liver toxicity via taken orally [46]. The ICP results showed the residual silver of CANF0.04 diffused in NA plate was 35.3 μg, which was lower than the dose that produced cytotoxicity. The results suggest that the diffusivity of Ag-NPs may be less harmful to the organism, and cytotoxicity need to be further characterized. Thus, the composite 12 of 15 films may have the potential to be applied in some biological fields. Polymers 2020, 12, 440 10 10 88 66 44 22 00 (a) (b) Figure 10. Inhibitory zone of composite films: (a) against E. coli and (b) against S. aureus. Figure 10. Inhibitory zone of composite films: (a) against E. coli and (b) against S. aureus. Two kinds of bacteria incubated on the blank NA plate and RCF NA pllatte grew welll ((Table 1),, whereasthebacteriaincubattedontthesurffaceoffttheCANF0.04 didnotmultiply.Itcanbeseenthatthe 0.04 contactsterilizationeffffecttofCANF0.04 agaiinstbothbacteriaexceed99.9%.Therremarkablebactericidal 0.04 effffect can be attributed to the large number of Ag-‐NPs uniformly distributed on the surface of the CANF0.04 ((Fiigure5b),whichmakessititeeaassieierrfoforrthteheAAg-gN‐NPsPasttaatctahcehdetdotohethbeacbtaecritaelricaelllcwellalwlwalhlewnhtehne 0.04 tfihlemfiwlmaswinascointcaocnt twacithwbitahctbearicat.eria. Sample Bacterial Growth 1 Blank control + Sterilization Rate (%) 0 Bacterial Growth 1 + Table 1. Test results of films’ contact sterilization ability. E. coli S. aureus Sterilization Rate (%) 0 Diameters of inhibition zone (mm) Diameters of inhibition zone (mm) CANF0.005 RCF CANF0.02 CANF0.01 CANF0.08 CANF0.04 CANF0.005 RCF CANF0.02 CANF0.01 CANF0.08 CANF0.04PDF Image | One-Pot Synthesis of Antibacterial Silver Nanoparticle
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