Preparation of silver iodide nanoparticles using laser ablation in liquid for antibacterial applications

Abstract

In this study, the authors reported the first synthesis process of silver iodide (AgI) nanoparticles (NPs) by pulsed laser ablation of the AgI target in deionised distilled water. The optical and structural properties of AgI NPs were investigated by using UV–vis absorption, X‐ray diffraction, scanning electron microscope (SEM), energy dispersive X‐ray, Fourier transform infrared spectroscopy, and transmission electron microscope (TEM). The optical data showed the presence of plasmon peak at 434 nm and the optical bandgap was found to be 2.6 eV at room temperature. SEM results confirm the agglomeration and aggregation of synthesised AgI NPs. TEM investigation showed that AgI NPs have a spherical shape and the average particle size was around 20 nm. The particle size distribution was the Gaussian type. The results showed that the synthesised AgI NPs have antibacterial activities against both bacterial strains and the activities were more potent against gram‐negative bacteria.

1 Introduction

Silver iodide (AgI) is a well‐known ionic conductor whose high‐temperature α ‐phase shows a superionic conductivity >1 Ω⁻¹ cm⁻¹ [1]. Its ionic conductivity arises from the highly disorder phase structure. It was reported that below 146°C, AgI undergoes a phase transition into the α ‐AgI which is considered as superionic conductor [2]. It has a direct bandgap of 2.9 eV at room temperature. It is reported that it's ionic conductivity and phase transition temperature strongly depended on the particle size. AgI is widely used in photography, medicine, antibacterial, electronic, magnetic, and optical applications [3, 4, 5]. Many methods were adapted to synthesis AgI nanoparticles (AgI NPs) such as ultrasonic [6], mechanochemical reaction [7], hydrothermal [8], laser ablation of Ag in a solution containing sodium dodecylsulphate surfactants and iodine [9], reverse micelles [10], electrochemical [11], ultrasonic spray pyrolysis [12]. Synthesis of nanoparticles by laser ablation in liquid has many advantages over other methods; recently [13], AgI NPs have been successfully used to reduce bacterial numbers of the Gram‐negative Escherichia coli (E. coli) and Pseudomonas aeruginosa (P. aeruginosa) as well as to the Gram‐positive Staphylococcus aureus (S. aureus) within 24 h and the effects were more effective against both gram‐negative bacteria. Up to our knowledge, no available reports in the published literature evaluating the bactericidal effect of AgI NPs synthesised by laser ablation in liquid. The pulsed laser ablation of nanoparticles has many advantages over the conventional methods; it is a cost‐effective method, it can produce high purity nanoparticles, no catalyst is needed, it exhibits good control on the size (<10 nm) and morphology of the synthesised nanoparticles, synthesis of core–shell nanocomposite, preserving the stoichiometry, and it's offering short reaction time [14]. Currently, there is no available information in the published literature evaluating the bactericidal action of AgI NPs synthesised by using laser ablation technique. In comparison, little research has been conducted on metal–halide nanoparticles, which are composed of a heavy metal compound such as silver and iodine to form a salt. Halogens themselves are considered potent antimicrobial agents [15, 16]; thus, it is feasible that a combination of heavy metal compounds with halogens could be engineered to enhance their antimicrobial efficacy [16]. Also, the concept of using a low solubility material like AgI NPs as the shell is to prevent the diffusion of Ag into the water, thus preventing the toxicity of the material. This phenomenon of materials is well characterised and showed its potential as a good antibacterial agent for repeated use [17]. Thus, here we report on the preparation and characterisation of AgI NPs by laser ablation in water and to investigate the antibacterial efficacy of prepared nanoparticles against three common bacterial pathogens using agar‐well diffusion method.

2 Materials and methods

2.1 Preparation of AgI NPs

All the chemicals used in this route were reagent grade without further purification. AgI pellet was placed at the bottom of a glass vessel containing about 3 ml of deionised distilled water and irradiated by Nd: YAG laser pulses operated at 1064 nm with a pulse duration of 7 ns and repetition frequency of 1 Hz. The laser beam was focused on AgI target by using a converging lens to spot size about 1 mm. Fig. 1 shows the schematic diagram of experimental set‐up of PLAL system used in this work. The ablation process was performed at normal temperature with laser fluence 5.5 Jcm⁻² and the ablation time was 10 min. The laser fluence was measured using a laser joule meter taking into account the water transmittance percentage. Our investigations revealed that the values of laser fluence and ablation time were optimum. The liquid was continuously stirred during laser irradiation to prevent any probable particles agglomeration.

Fig. 1.

Schematic diagram of PLAL system used in this study

2.2 Characterisation of AgI NPs

The optical absorption of colloidal AgI NPs was measured using UV–vis spectrophotometer with double beam (Shimadzu UV‐1800). The structure and crystallinity of synthesised nanoparticles was investigated using X‐ray diffraction (XRD), X‐ray diffractometer (XRD‐8000, Shimadzu) using a CuKα source (λ  = 0.15419 nm). The surface morphology of AgI NPs was measured using scanning electron microscopy (SEM) (Jeol JSM‐6335F). The shape and size of AgI NPs were examined using transmission electron microscopy (TEM) (type CM10 pw6020, Philips‐Germany).

2.3 Antibacterial activity of AgI NPs

E. coli, P. aeruginosa (G −ve) and S. aureus (G +ve) were used to investigate the antibacterial activity of prepared AgI NPs. The synthesised AgI NPs with different concentrations (0.62, 1.25, 3.50 and 7.00 µg ml⁻¹) were tested for antibacterial activity via agar‐well diffusion method. The agar wells (8 mm, diameter) were prepared with the help of a sterilised stainless steel cork borer. Each strain was swabbed uniformly onto individual plates using sterile cotton swabs. The plates, containing the test organism and AgI NPs were incubated at 37°C for 24 h. After the incubation, the zones of growth inhibition of well diffusion were measured in millimetre and recorded as mean ±SD. The experiments were conducted in triplicate according to the standard protocol.

2.4 Colony‐forming unit (CFU) assay

CFUs were carried out according to the method of Maqbool et al. [18]. Briefly, three tubes with 2 ml of sterilised medium containing E. coli, P. aeruginosa and S. aureus were inoculated with 10⁷ CFU and then placed in a shaking incubator at 37°C. Thereafter, the bacterial culture tubes were loaded with AgI NPs (25 μg 5 μl⁻¹) solution. Non‐treated cultures were used as control. After an interval of 30 min or 120 min, 10 μl from each sample was directly cultured over the already prepared agar plate surface at 37°C for 24 h. Later on, the CFUs were counted.

3 Results and discussions

The optical absorption spectrum of AgI is shown in Fig. 2; it decreases sharply as wavelength increase up to 400 nm. A strong absorption peak is noticed at 428 nm due to surface plasmon resonance (SPR) which is in good agreement with reported data in [19]. Inset of Fig. 2 is the freshly prepared colloidal AgI NPs with bright brown colour. No colour changing was observed after several hours of preparation. The optical bandgap E _g of AgI can be estimated from the Tauc's relationship [20]

$${\left(\alpha h\upsilon \right)}^{1/2}=A\left(E_g-h\upsilon \right)$$

where α is the absorption coefficient, A is constant, and (hν) is the photon energy.

Fig. 2.

Optical absorbance of colloidal AgI NPs. Inset is the photograph of freshly prepared colloidal AgI NPs

Fig. 3 shows the (αhυ)² versus photon energy plot of AgI. This figure confirms that the optical transition is the direct type and the value of the energy gap of AgI was around 2.6 eV.

Fig. 3.

(αhν)² versus photon energy plot

Fig. 4 shows the room temperature XRD pattern of AgI NPs. It is clearly seen that AgI consists of a mixture of two phases; hexagonal wurtizte β ‐AgI and cubic zinc blend γ ‐AgI phases. The XRD peaks located at 22.1°, 24.36°, 34.34°, 39.52°, 42.6° and 45.14° corresponding to (100), (002), (101), (102), (110), (112) and (103) planes, respectively, are related to hexagonal wurtzite phase [7, 21]. The peak located at 39.52° and 45.14° corresponding to (220) and (311) planes, respectively, are indexed to the cubic structure according to JCPD card no. (44‐1482). No diffraction peaks related to Ag and AgO phases or impurities were noticed in XRD spectrum. The average crystallite size of AgI was calculated using Scherrer's equation [22] and found to be 18 nm.

Fig. 4.

XRD pattern of AgI NPs

Fig. 5 a shows the SEM image of AgI NPs, agglomeration and aggregation of micro‐sized spherical particles are observed. This aggregation may interpret due to the van der Waals force between particles [23]. The magnified SEM image of AgI particles is given in the inset of Fig. 5 a. It is clearly seen that agglomerated and irregular shape particles with different sizes were found. The EDX analysis was performed to determine the chemical composition of synthesised AgI NPs. The EDX result illustrated in Fig. 5 b clearly shows the presence of silver and iodine elements, the [Ag]/[I] ratio was calculated and found to be about 0.95 which indicates the formation of fully stoichiometric AgI phase. A peak related to Cl element was observed in the EDX spectrum, which is probably originated from the glass substrate and from the SEM chamber.

Fig. 5.

SEM image and EDX spectrum of AgI NPs

(a) SEM image of AgI NPs, inset is the magnified SEM image, (b) EDX spectrum of AgI NPs. Inset is atomic weight percentage of elements

Fig. 6 illustrates the Fourier transform infrared (FT‐IR) spectrum (transmission mode) of AgI NPs. Five absorption bands were noticed at a wavenumber of 560, 1624, 2967, 3018 and 3402 cm⁻¹, the 1624 and 3402 bands are indexed to ν (OH) stretching and bending vibration modes which given an indication of the existence of physisorbed water attached to AgI NPs [24].

Fig. 6.

FT‐IR spectrum of AgI NPs

The FT‐IR band at 2967 cm⁻¹ can assign to CH₂ comes from organic contaminants. The peak at 3018 cm⁻¹ indexed to C = C stretching mode [25]. The absorption band of 560 cm⁻¹ corresponds to the vibration of AgI [26]. Moreover, no absorption bands were noticed in the range of 4000–900 cm⁻¹ related to the AgI indicating that no lattice vibration for AgI in the mid‐infrared region. The size and shape of AgI NPs were obtained from TEM image and analysis. The TEM of fresh synthesised AgI NPs shown in Fig. 7 a revealed that the some of the synthesised AgI particles are well dispersed and have a spherical shape and were largely homogeneous. The average particle size was 25 nm which is in good agreement with that obtained by XRD investigation. As we can see from the TEM image, some of AgI particles are agglomerated and tend to form bigger size which may form via the repulsive electrostatic forces between attracting particles [23]. Fig. 7 b shows the particle size distribution of AgI NPs which determined from the TEM investigation with the aid of using the software. It is clear from this figure that this distribution is Gaussian type fitting and confirms that the average particle size is around 20 nm.

Fig. 7.

TEM image and particle size distribution of AgI NPs

(a) TEM image of AgI NPs, (b) Particle size distribution of AgI NPs

In this study, the application of AgI NPs as an antibacterial agent was evaluated and demonstrated that the zone of inhibition increased according to the concentration of AgI NPs in all pathogenic bacteria (see Fig. 8 and Table 1). For all organisms, the antibacterial effect of AgI NPs appeared to be concentration dependent, though the AgI NPs was more effective at higher concentrations. The tested gram‐negative bacteria show a higher sensitivity against AgI NPs than tested gram‐positive bacteria. In general, all four concentrations of the AgI NPs produced significantly (P  ≤ 0.05) more rapid and greater reductions in P. aeruginosa and E. coli than they did in S. aureus. The lower sensitivity of gram‐positive bacteria against silver nanoparticles is primarily due to the thickness of peptidoglycan layer, which may prevent transport of those nanoparticles through the bacterial cell wall [27, 28]. For the further antibacterial activity of synthesised AgI NPs, the bactericidal efficiency of nanoparticles was further confirmed by performing a CFU assay against P. aeruginosa and E. coli and S. aureus (Fig. 9). The results of the time kinetic histograms revealed that AgI NPs could significantly kill E. coli and P. aeruginosa within the first 30 min of exposure to AgI NPs. In contrast, the bactericidal activity of AgI NPs against S. aureus was observed after 2 h of treatment. This bactericidal activity of AgI NPs may be attributed to the strong interactions between AgI NPs and the macromolecules such as proteins and the DNA of bacteria [29]. However, researchers suggested different mechanism(s) of the nanoparticles action onto bacteria. However, is still not very clear in many of the cases whether the killing is due to any one individual mechanism or due to the combination of more than one mechanism [30]. One of them may be due to the combination of the interaction of AgI NPs with the bacterial cell wall and simultaneous penetration of Ag⁺ ions inside the bacterial cells. Li et al. [31] illustrated the Ag NPs bacterial cell membrane permeability and respiratory function, which lead to cell death. The smaller size of Ag NPs provides a large surface area that is available for interaction, which would provide a more considerable activity than the larger Ag⁺ [32, 33]. In addition, Ag NPs not only interact with the membrane surface but may also penetrate inside the structure of bacteria, resulting in a disruption of adenosine triphosphate (ATP) production, DNA replication and activation of reactive oxygen species (ROS), which lead to cell damage and death [34]. Maqbool et al. observed that the antibacterial activity of CeO₂ NPs primarily depends upon the electrostatic attraction between positively charged NPs and negatively charged bacterial cell surface and is crucial for the activity of NPs as a bactericidal agent [18]. Furthermore, the antibacterial potential of AgNPs might be related to the inactivation of phosphomannose isomerase enzyme, which catalyses the conversion of mannose‐6‐phosphate to fructose‐6‐phosphate. The latter compound is an important intermediate of glycolysis, which is the most common pathway associated with sugar catabolism in bacteria [35]. According to the third concept the antibacterial activity of AgNPs is determined by the oxidative stress of hydrogen peroxide, hydroxyl radical and superoxide radical. ROS production is closely related to the efficiency of a photocatalyst, depending on the generation rate, rate of migration, and energy levels of the photoexcited electron–hole pairs [36]. The formed free radicals can attack membrane lipids and lead to denaturing of cell membrane permeability with leakage of potassium ions, ultimately causing bacterial cell death [37].

Fig. 8.

Images of inhibition zone showing the antibacterial activity of AgI NPs against three pathogens

(a) 0.62 µg ml⁻¹, (b) 1.25 µg ml⁻¹, (c) 3.50 µg ml⁻¹, (d) 7.00 µg ml⁻¹

Table 1.

Inhibition zone for AgI NPs against the tested bacterial strains

Concentration, µg ml−1	Zone of inhibition, mma
	P. aeruginosa	E. coli	S. aureus
0.62	7.4 ± 1.4	7.0 ± 0.8	6.9 ± 1.4
1.25	10.4 ± 1.6	10.2 ± 1.6	9.9 ± 1.5
3.50	12.0 ± 1.4	12.3 ± 1.2	8.2 ± 1.3
7.00	17.6 ± 1.6	16.2 ± 1.8	11.8 ± 1.5

^a Zone of inhibitions is the mean values of three experiments.

Fig. 9.

Time‐dependent effect of AgI NPs on the bacterial survival using CFU assay at 30 and 120 min of treatment

(a) P. aeruginosa, (b) E. coli, (c) S. aureus

4 Conclusion

In this study, we have successfully synthesised AgI NPs by using a simple laser ablation technique without using any catalyst. XRD data confirm of that the structure of AgI NPs is a mixture of two phases; β ‐AgI and γ ‐AgI. EDX analysis proved of preparation of stoichiometric AgI. The synthesised AgI NPs have almost spherical shape with a size of 20 nm and the particle size distribution obeys to Gaussian profile. AgI NPs exhibited a strong antimicrobial activity against several pathogens that were tested in this study. The tested AgI NPs are more effective against gram‐negative bacteria than gram‐positive bacteria. Further work could involve implementing the molecular level interaction of AgI NPs in bacterial growth inhibition.