An Assessment of the Effect of Green Synthesized Silver Nanoparticles Using Sage Leaves (Salvia officinalis L.) on Germinated Plants of Maize (Zea mays L.)

Abstract

AgNPs have attracted considerable attention in many applications including industrial use, and their antibacterial properties have been widely investigated. Due to the green synthesis process employed, the nanoparticle surface can be coated with molecules with biologically important characteristics. It has been reported that increased use of nanoparticles elevates the risk of their release into the environment. However, little is known about the behaviour of AgNPs in the eco-environment. In this study, the effect of green synthesized AgNPs on germinated plants of maize was examined. The effects on germination, basic growth and physiological parameters of the plants were monitored. Moreover, the effect of AgNPs was compared with that of Ag(I) ions in the form of AgNO₃ solution. It was found that the growth inhibition of the above-ground parts of plants was about 40%, and AgNPs exhibited a significant effect on photosynthetic pigments. Significant differences in the following parameters were observed: weights of the caryopses and fresh weight (FW) of primary roots after 96 h of exposure to Ag(I) ions and AgNPs compared to the control and between Ag compounds. In addition, the coefficient of velocity of germination (CVG) between the control and the AgNPs varied and that between the Ag(I) ions and AgNPs was also different. Phytotoxicity was proved in the following sequence: control < AgNPs < Ag(I) ions.

Keywords: green synthesis; phyto-nanotechnology; phytotoxicity; plant physiology; thiol compounds.

Figure 1

A simplified scheme of the presence and movement of nanoparticles in the environment. Nanoparticles (NPs) get into the environment in a natural way (natural nanoparticles) in the inorganic (volcanic dust, etc.) or possibly organic form (cellular debris), as well as due to human activity (engineered nanoparticles). Such nanoparticles are purposefully synthesized or generated by unwanted processes or as a part of industrial waste. These particles can bind to the soil. From there they are mobilized into the environment as a water fraction, in which they can be solubilized. Thus, the particles can enter the plants through the root system [14].

Figure 2

Biologically important secondary metabolites found in Salvia officinalis at high concentrations. Antibacterially active molecules such as salviol and thujone have been identified. In inset: A proposed scheme for reduction of silver ions in the presence of phenolic compounds is also shown. The resulting reduced ions subsequently form aggregates in the form of nanoparticles.

Figure 3

Preparation and characterization of silver nanoparticles (AgNPs)—a simplified scheme for preparing AgNPs. (A,B) The plant was washed in distilled water, (C) dried at 60 °C, (D) homogenized to 1-mm particle size, (E) and mixed with water and extracted for 1 h at different temperatures (20, 40, 60 and 80 °C). (F) The extract was filtered, (G) 0.1 M AgNO₃ (1:1) was added and (H) the mixture was stirred for 24 hours. AgNPs have been prepared, (I) purified with methanol (J) dried and characterized.

Figure 4

Preparation and characterization of silver nanoparticles (AgNPs)—characteristics of plant extract. (A) A typical appearance of AgNPs obtained using a plant extract prepared at 20, 40, 60 and 80 °C. The color intensity was evaluated by ColorTest, (B) a typical dependence of total phenolic compounds on the plant extract temperature, (C) changes in integrals of thiol compounds of AgNPs prepared at 20 and 80 °C, (D) typical Vis spectrum of the prepared AgNPs (scan 0.2 nm), (E) hydrodynamic size, (F) typical voltammogram of thiol compounds bound to the surface of AgNPs, (G) XRD analysis of AgNPs, (H) the high-resolution transmission electron microscopic image of AgNPs. (I) Polydispersed AgNPs ranged between 2 and 50 nm.

Figure 5

A typical experimental arrangement of Ag(I) ions and AgNPs toxicity tests on germinated plants of maize (10 × 5). (A) The germination was evaluated at 96 h. The experiment was run at 25 °C in a cultivation box. The typical appearance of the germinated seeds was evaluated after 96 h. (B) Total weight of caryopses after 96 h of exposure. (C) Germination index determined after 96 h, (D) Germination energy after 96 h, (E) Fresh weight (FW) of root after 96 h of exposure, (F) Coefficient of velocity of germination. All results presented were determined as the mean of all concentrations (1, 50 and 150 mg/L) tested.

Figure 6

Chemical analysis of the primary roots of germinating maize plants after 96 h exposure to Ag(I) ions and AgNPs. In the study, the following parameters were analysed for all tested concentrations of Ag(I) ions and AgNPs. (A) Total protein concentration determined by the biuret method. (B) The average total protein ‒ biuret method. (C) Total protein concentration determined by the pyrogallol red-molybdate method. (D) The average total protein-pyrogallol red-molybdate method. (E) Total flavonoid concentration. (F) The average value of the total flavonoid concentration.

Figure 7

A basic study of the phytotoxic effect of AgNPs on plants of Zea mays in hydroponics. (A) A typical photo of germinated plants of Zea mays—(a) control group, (b) AgNO₃, (c) AgNPs—after 96 h of exposure to the hydroponic system with tap water. (B) A typical course of plant height change at different applied concentrations and exposure time. (C) Summarized average plant heights in individual studied variants. (D) A typical photo of germinated plants of Zea mays—(a) control group, (b) Ag(I) ions, (c) AgNPs—after 96 h of exposure to the hydroponic system with tap water; Ag concentration of 150 mg/L. Plants were collected in triplicate every day and then processed according to the procedure described in the Material and Methods section.

Figure 8

Effect of Ag(I) ions and AgNPs on the root system of germinal plants. (A) Root length after 96 h exposure expressed as summary data for each test concentration. (B) Summary data for each test group. (C) The root dry weight (DW) for each test concentration. (D) Changes in the anatomical structure of the stem and root of germinated plants exposed to Ag(I) ions and AgNPs. Plants were collected after 96 h of exposure to hydroponics. Plants were collected in triplicate every day and then processed according to the procedure described in the Material and Methods section.

Figure 9

Influence of AgNPs and Ag(I) ions on the above-ground parts of germinated plants. The amount of (A) silver ions, (B) phenolic compounds, (C) glucose, (D) total proteins, and (E) total flavonoids. (F) ABTS activity. Plants were collected after 96 h of exposure to hydroponics. Plants were collected in triplicate every day and then processed according to the procedure described in the Material and Methods section. Data are presented as the average of all experimental data.

Figure 10

Influence of Ag(I) ions and AgNPs on the root system of seedlings. (A) A typical root system appearance after exposure for 96 h. The amount of (B) silver ions, (C) phenolic compounds, (D) glucose, (E) total proteins, and (F) total flavonoids. (G) ABTS activity. Plants were collected after 96 h of exposure to hydroponics. Plants were collected in triplicate every day and then processed according to the procedure described in the Material and Methods section. Data are presented as the average of all experimental data.

Figure 11

Changes in selected plant parameters exposed to Ag(I) and AgNPs. (A) Weight of the above-ground parts of plants as summary data for individual tested variants. (B) Changes in photosynthetic pigment contents as average values in individual variants. (C) Changes in the total weight of above-ground parts of plants in the Ag(I) ion and AgNPs groups. (D) Total thiols in roots. (E) Total thiols in the above-ground parts. Plants were collected after 96 h of exposure to hydroponics. Plants were collected in triplicate every day and then processed according to the procedure described in the Material and Methods section. Data are presented as the average of all experimental data.