The digital images of cellulose fabric, matrix and the CNCFs using 1-5 mmol/L aq. AgNO3 solutions were photographed and are presented in Fig. 1. From Fig. 1, it is evident that cellulose fabric was white in color while the Vitex leaf extract infused cotton fabric (matrix) appeared light brown. On the other hand, the color of the CNCFs varied from medium brown to dark brown with an increasing concentration of the source solution. The color change preliminarily indicates the in situ generation of the AgNPs in the matrix. Similar observation was made by Logeswari et al. (2013) and Banerjee et al. (2014) in the generation of metal nanoparticles by some leaf extracts. In order to visualize the formed AgNPs, the SEM images of the CNCFs were recorded. As an example, the SEM images of the CNCFs using 1 mmol/L (minimum) and 5 mmol/L (maximum) aq. AgNO3 solutions are presented in Fig. 2. The EDX spectra recorded simultaneously for the corresponding SEM images are also presented in the same figure. Using the micrographs, the particle size distribution in both these cases was determined and is presented in the same figure. From the SEM images (Fig. 2a and 2b), it is clearly evident that the formed AgNPs were spherical in shape. From Fig. 2c and 2d, the EDX spectra confirmed the presence of silver particles. It can be further seen that the intensity of the EDX peak corresponding to Ag element for CNCF using 5 mmol/L source solution was higher than that formed using 1 mmol/L source solution. It clearly indicates that larger number of AgNPs was generated when higher concentrated source solution was used. The particle size distribution of the prepared CNCFs using 1 mmol/L and 5 mmol/L source solution is presented in Fig. 2e and 2f, respectively. From the distribution curves, it is evident that in both cases, maximum numbers of generated AgNPs were in the particle range of 91-100 nm. However, the average size of the formed AgNPs in the CNCFs made using 1 mmol/L and 5 mmol/L source solutions was found to be 86 nm and 102 nm, respectively. This may be due to the agglomeration of some of the nanoparticles formed when higher concentrated source solutions were used. In the case of the CNCFs made using other concentrated source solutions (2, 3 and 4 mmol/L), the average particle size was found to be between these two values.
Figure 1. Digital photographs of cellulose fabric (a); matrix (b) and CNCFs with in situ generated AgNPs using 1 mmol/L (c); 2 mmol/L (d); 3 mmol/L (e); 4 mmol/L (f) and 5 mmol/L (g) source solutions.
Figure 2. Scanning electron microscope (SEM) micrographs (a, b); energy-dispersive X-ray (EDX) spectra (c, d) and particle size distribution (e, f) of cellulose nanocomposite fabrics (CNCFs) with in situ generated silver nanoparticles (AgNPs) using 1 mmol/L and 5 mmol/L aq. AgNO3 solutions respectively.
In order to examine the interaction between the cotton fabric and the Vitex leaf extract, their FT-IR spectra are presented in Fig. 3a. From Fig. 3a, it is evident that both the spectra had similar peaks indicating similar chemical groups in them. However, the intensity of the peaks at 3337 cm-1 (OH st. vibration) and 1022 cm-1 (C-O-C st. vibration) of the matrix was higher than that of the cotton fabric, indicating that additional OH and C-O-C groups were contributed by the leaf extract. In order to examine the influence of in situ generated AgNPs on the chemical structure of matrix, the FT-IR spectra of the matrix and the CNCFs using 1-5 mmol/L, source solutions are presented in Fig. 3b. For clarity, the spectra of matrix and the CNCF using 5 mmol/L source solution are presented separately in Fig. 3c. From Fig. 3b and 3c, it is evident that no major changes were observed except the lowering of intensity of the bands corresponding to O-H, C-O groups in the case of nano composites fabrics. This clearly indicates the role of O-H and C-O groups of the matrix in reducing the silver salts into AgNPs. The other common bands appearing at 2905 cm-1 (CH st.), 1640 cm-1 (crystallization of water) 1425 cm-1 (CH2 b.), 1359 and 1313 cm-1 (CH2 wagging) and 878 cm-1 (β-glucosidic linkage) indicate the cellulosic structure in both the matrix and the CNCFs. Similar observation was made in the case of synthesis of AgNPs using banana peel extract (Ibrahim et al., 2015) and cellulose nanofibers (Li et al., 2015). Though the OH and CO groups presented in cellulose can act as mild reducing agents (Vainio et al., 2007), their increased number by the leaf extract is expected to increase the reducing capacity of silver salts to AgNPs.
Figure 3. Fourier transform infrared (FT-IR) spectra of cotton fabric and matrix (a); matrix and CNCFs with in situ generated AgNPs using 1-5 mmol/L aq. AgNO3 source solutions (b) and matrix and cotton nanocomposite fabric with in situ generated AgNPs using 5 mmol/L source solution (c).
In order to examine the effect of AgNPs on the crystallinity of the nanocomposite fabrics, the X-ray diffractograms of the cotton fabric, matrix and the nanocomposite cotton fabrics with in situ generated AgNPs using 1-5 mmol/L aq. AgNO3 solutions were recorded and are presented in Fig. 4a. As the overlapped diffractograms did not yield much information, the diffractograms corresponding to the matrix and the CNCF using 5 mmol/L aq. AgNO3 solution are shown separately in Fig. 4b. From Fig. 4b, it is clearly evident that the intensity of the CNCF was slightly higher than that of the matrix indicating a slight increase in the crystallinity of the CNCFs due to the presence of AgNPs. As the presence of AgNPs is hard to observe in the overlapped diffractograms (due to the higher intensity of the peak corresponding to the matrix), the diffractogram of the CNCF using 5 mmol/L aq. AgNO3 was expanded in 2θ = 30°-67° and is presented in Fig. 4c. From Fig. 4c, it is evident that there are several peaks in the diffractograms of which the peaks at 2θ = 38.1°, 45.1° and 63.6° corresponding to the reflections from (111), (200) and (220) planes of AgNPs (Mahdieh et al., 2012). The other prominent peaks observed at 2θ = 32.0°, 38.1°, 54.8° and 65.3° correspond to the reflections from (111), (200), (220) and (311) planes of Ag-O nanoparticles (Kumar Trivedi, 2015), respectively. Thus in the developed CNCFs, both AgNPs and AgO nanoparticles were generated. These results indicate the in situ generation of AgNPs and AgO nanoparticles in the matrix.
Figure 4. The X-ray diffractograms (XRD) of cotton cloth, matrix and the CNCFs made using 1-5 mmol/L aq. AgNO3 as source solutions (a); matrix and CNCF using 5 mmol/L source solution (b) and CNCF using 5 mmol/L solution expanded in the 2θ = 30°-67° (c).
In order to examine the antibacterial activities of the prepared CNCFs using different concentrated source solutions, the disc method was used against two gram negative (Pseudomonas and E. coli) and two gram positive (Bassilus subtils and Stephylococus areus) bacteria. For comparison, the test for cotton fabric and the matrix was also carried. The photographs of the petri dishes in which the test was conducted are presented in Fig. 5. The cotton cloth and the matrix were designated as A and B whereas the CNCFs using 1, 2, 3, 4 and 5 mmol/L were designated as H, I, J, K and L respectively in the images. Using the Image J program, the diameter of the clear zones was calculated and is presented in Table 1.
Figure 5. Antibacterial activity of cotton fabric (A); matrix (B); CNCFs with in situ generated AgNPs using 1 mmol/L (H), 2 mmol/L (I), 3 mmol/L (J), 4 mmol/L (K) and 5 mmol/L (L) aq. AgNO3 source solutions and Vitex leaf extract as a reducing agent against Pseudomonas (a); Escherichia coli (b); Bassilus subtils (c) and Stephylococus areus (d) bacteria.
Concentration of aq. AgNO3 solution used Diameter (mm) Pseudomonas E. coli Bassilus subtils Stephylococus areus Cotton Fabric (A) - - - - Matrix (B) - - - - 1 mmol/L (H) 12.9 11.0 12.9 11.1 2 mmol/L (I) 12.6 11.1 14.1 11.4 3 mmol/L (J) 13.7 13.1 15.1 13.8 4 mmol/L (K) 13.7 14.4 14.3 14.7 5 mmol/L (L) 14.4 14.0 13.4 16.3
Table 1. Diameter of the clear zones observed for cotton fabric, matrix and CNCFs using 1-5 mmol/L aq. AgNO3 solutions against gram negative (Pseudomonas and Escherichia coli) and gram positive (Bassilus subtils and Stephylococus areus).
From Fig. 5 and Table 1, it is evident that both cotton fabric and matrix did not show any antibacterial activity. However, the CNCFs with in situ generated AgNPs using different concentrated source solutions and Vitex leaf extract as a reducing agent exhibite good antibacterial activities against gram negative and gram positive bacteria. In all the cases, the clear zone diameters varied between 11.0 mm and 16.3 mm and in most of the cases, incresed with increasing concentration of the source solutions. These observations indicate that the CNCFs can be used in medical applications as antibacterial materials.