Phytosynthesis of Silver Nanoparticles Using Perilla frutescens Leaf Extract: Characterization and Evaluation of Antibacterial, Antioxidant, and Anticancer Activities

Abstract

Purpose: The present study investigates the phytosynthesis of silver nanoparticles (AgNPs) using Perilla frutescens leaf extract, which acts as a reducing agent for the conversion of silver ions (Ag+) into AgNPs. P. frutescens leaf synthesized AgNPs (PF@AgNPs) were evaluated for biomedical properties including antibacterial, antioxidant and anticancer activities.

Materials and methods: PF@AgNPs were synthesized using P. frutescens leaf extract and silver nitrate solution. The morphology and physical properties of PF@AgNPs were studied by spectroscopic techniques including, UV-Vis, FTIR, TEM, XRD, DLS, and TGA. Antibacterial activity of PF@AgNPs was evaluated by disk diffusion assay. Antioxidant activity of PF@AgNPs was checked by 2.2-diphenyl-1-picrylhydrazyl (DPPH), and 2.2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) free radical scavenging assays. Anticancer activity of PF@AgNPs was checked by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay. Cytotoxic effects of PF@AgNPs on most susceptible cancer cell lines were observed by phase contrast microscopy.

Results: PF@AgNPs showed surface plasmon resonance peak at 461 nm. XRD pattern showed that the PF@AgNPs were face-centered cubic crystals with a mean size of 25.71 nm. TEM analysis revealed the different shapes (spherical, rhombic, triangle, and rod) of PF@AgNPs. Zeta potential value (-25.83 mV) indicated that PF@AgNPs were long-term stable and not agglomerated. A low polydispersity index value (0.389) indicated the monodispersity of PF@AgNPs. TGA revealed the high thermal stability of PF@AgNPs. PF@AgNPs exhibited maximum inhibition against Escherichia coli, followed by Bacillus subtilis and Staphylococcus aureus. PF@AgNPs showed maximum inhibition of 68.02 and 62.93% against DPPH and ABTS-free radicals, respectively. PF@AgNPs showed significant anticancer activity against human colon cancer (COLO205) and prostate adenocarcinoma (LNCaP). PF@AgNPs exhibited apoptotic effects on LNCaP cells including cell shrinkage, membrane blebbing, chromatin condensation, fragmentation of nuclei, and formation of apoptotic bodies.

Conclusion: The present study reports the successful synthesis of PF@AgNPs using P. frutescens leaf extract. The synthesized PF@AgNPs are FCC crystals, monodispersed, long-term stable, and non-agglomerated. The observed antibacterial, antioxidant, and anticancer activities demonstrate the potential biomedical applications of PF@AgNPs.

Keywords: LNCaP; Perilla frutescens; antibacterial activity; anticancer activity; antioxidant activity; silver nanoparticles.

Figure 1

Color change of the reaction solution from light yellow to dark brown after 2 h indicates the formation of PF@AgNPs.

Figure 2

UV-Vis analysis of P. frutescens leaf extract and PF@AgNPs. UV-Vis spectrum of PF@AgNPs showed surface plasmon resonance peak at 461 nm.

Figure 3

Schematic diagram represents the possible mechanism of plant-mediated synthesis (phytosynthesis) of AgNPs.

Figure 4

FTIR analysis of (A) P. frutescens leaf extract and (B) PF@AgNPs.

Figure 5

XRD pattern of PF@AgNPs.

Figure 6

TEM analysis PF@AgNPs (A) TEM micrograph at 100 nm shows monodispersed AgNPs with different shapes including spherical, rod, triangle, and rhombic; (B) rod-shaped, rhombic shaped, and spiral-shaped AgNPs observed at 50 nm scale; (C) triangle, rhombic and spherical shaped AgNPs at 50 nm; (D) rod-shaped and triangle-shaped AgNPs at 20 nm; (E) spherical shaped AgNPs at 20 nm; (F) SAED pattern of AgNPs.

Figure 7

Zeta potential measurement of PF@AgNPs.

Figure 8

TG and DTG analysis PF@AgNPs.

Figure 9

Antibacterial activity of PF@AgNPs against (A) S. aureus, (B) B. subtilis, and (C) E. coli; PTLE indicates pristine leaf extract; Ab indicates antibiotic; DMSO indicates dimethyl sulfoxide.

Figure 10

Comparison of antibacterial activity of pristine leaf extract (PTLE), PF@AgNPs and standard antibiotic streptomycin. Results were expressed as mean ± SD (n=3). Different lowercase letters above the bars indicate significant differences (P < 0.05) by SPSS test.

Figure 11

Scheme represents the possible mechanism of antibacterial activity of AgNPs.

Figure 12

Antioxidant activities of PTLE, PF@AgNPs and ascorbic acid (A) DPPH radical scavenging activity, and (B) ABTS radical scavenging activity. All the data were represented as mean ± SD (n =3) in the form of bar graph. Data were analyzed by one-way analysis of variance (ANOVA, P < 0.05). Different lowercase letters above the bars indicate significant differences (P < 0.05).

Figure 13

Dose-dependent cytotoxic effects of PF@AgNPs against human colon carcinoma (COLO205) and prostate carcinoma (LNCaP) All the data were represented as mean ± SD (n =3) in the form of bar graph. Bars with different superscripts indicate significant differences (P < 0.05).

Figure 14

Anticancer activity of PF@AgNPs against LNCaP cell lines. The effects of PF@AgNPs on morphological changes of LNCaP cells were studied after 24 h exposure at different concentrations. PF@AgNPs affected the cell viability by inducing apoptotic symptoms, including warping of cells, rounding of cells, membrane blebbing, and cell shrinkage.