Introduction

In recent times, nanoparticles have emerged as a highly promising substitute for conventional materials, finding extensive applications in the domains of science and engineering. The advancement of nanotechnology necessitates the ongoing progress of alternative nanofabrication methods that align with the demands of being cost-effective, straightforward, environmentally friendly, and capable of high-throughput production. Extensive research has been conducted in the field of nanoscience on small particles (NPs) of colloidal form noble metals. Nanoparticles are commonly distinguished based on their dimensions, morphology, and heterogeneity. The initial assessment of the synthesized nanoparticles is conducted through the utilization of ultraviolet spectroscopy [1, 2]. Nanotechnology employs two primary methodologies: the “top-down” approach, in which nano-objects are fabricated from bigger objects without precise atomic-level manipulation, and the bottom-up method, also known as the reduction method, wherein nanostructures are generated by assembling atoms produced through ion reduction [3,4,5]. Nanoparticles can be generated through the short-pulse laser destruction of a solid target immersed in a liquid medium. Gold nanoparticles are created using a variety of techniques. A method frequently used for the creation of gold nanoparticles (Au NPs) is laser ablation. One often used method for the creation of metal nanoparticles is laser destruction in liquid. In this method, bulk metals are ablated in liquid media like water using pulsed lasers [6]. It is being used to create a wide variety of nanostructured materials with different sizes and forms using pulsed laser ablation in liquid. The intensity, wavelengths, and pulse duration of the used laser are just a few of the specific features that are directly related to them [7,8,9,10]. A portion of the laser energy is converted into a mechanical impulse by vaporization and plasma formation in the liquid. The plasma formation, when the laser beam interacts with the Au target, causes localized heating which causes the gold to ionize and vaporize, forming a plasma plume with high energy. This plasma contains extremely energetic species like ions and electrons. The vaporized atoms and ions condense to produce nanoparticles as the plasma rapidly cools upon expansion into a liquid medium [2, 6]. The rate of fabrication and the morphology of the nanoparticles prepared by pulsed laser ablation in liquid are dependent on several factors, including the laser wavelength, energy of the laser pulses, the absorption of laser energy by the liquid medium, and the duration of the ablation process [11,12,13]. The distinctive characteristics exhibited by Au nanoparticles (Au NPs) can be attributed to their elevated surface-to-volume ratio and the augmented atom count at their grain boundaries. Gold nanoparticles (Au NPs) exhibit distinct optical characteristics within the visible spectrum due to the occurrence of surface plasmon oscillation involving unbound electrons [14,15,16], which makes them suitable to apply in many application fields [17,18,19,20,21,22]. Therefore, in this study, pulsed laser ablation in liquid (PLAL) was utilized to synthesize Au nanoparticles and evaluate their antibacterial activity against several strains of microbes, such as Pseudomonas aeruginosa, Staphylococcus aureus, Streptococcus mutans, and Acinetobacter baumannii. The characterization of prepared Au NPs was investigated using XRD to determine the phase and crystal structure; SEM, to examine the morphology of nanoparticles, and the optical properties were carried out using Shimadzu spectrophotometers and photoluminescence.

Materials and Methods

Gold Nanoparticle Synthesis

A 99.99% pure gold plate was employed as the target material for laser ablation. The laser ablation of gold in deionized distilled water (D.D.W.) using a Q-switched Nd:YAG laser operating at the fundamental wavelength of 1064 nm, 9 ns, and 10 Hz, as shown in Fig. 1. In order to attain a significant level of laser flounce, the laser beam was focused by employing a positive lens with a focal length of 120 mm. The experiment was done using two procedures. In the first procedure, the laser fluence was varied from (2.2, 4.4, 6.64, 8.85, and 11.1) J/cm2 with a constant number of laser pulses (300 pulses). In the second procedure, the number of laser pulses varied from 100, 200, 300, and 400 pulses, while the laser fluence was fixed at 4.4 J/cm2. The laser light is directed and concentrated to form a laser spot with a diameter of approximately 2.4 mm. The target plate was affixed to the base of a baking vessel containing a volume of 3 mL of liquid.

Fig. 1
figure 1

Experimental setup of the PLAL

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Characterization Techniques

The chemical composition of gold nanoparticles was examined using the Fourier-transform infrared spectroscopy FTIR (SPECTRUM TWO N spectrometer, PerkinElmer, Yokohama, Japan) by the KBr pellet technique in the range of 400–4000 cm−1. The crystalline structure of Au NPs was characterized using an X-ray diffractometer (PODWE XRD 2700AB, Haoyuan Instrument, Dandong, China) with a Cu K target and a tube operating at 45 kV, while the size and shape of Au NPs were examined using field emission scanning electron microscopy (FE-SEM, Czech Republic, Tescan, Mira III). Before proceeding with their examination, the samples were made by drop–casting a nanoparticle suspension on a silicon wafer. The optical properties of Au nanoparticle suspensions were carried out using a Shimadzu spectrophotometer (1900i, Shimadzu, Kyoto, Japan) in the wavelength range 300–900 nm with quartz cuvettes of 1 cm path lengths and distilled water as a blank and photoluminescence (PL) spectroscopy (G9800A, Xe lamp). These measurements were achieved using a quartzite cell with a 1-cm optical path at room temperature.

Bacterial Isolates

Various pathogenic bacteria were used: two-gram negative bacteria, Acinetobacter baumannii (A. baumannii) and Pseudomonas aeruginosa (P. aeruginosa), and two-gram positive bacteria, Streptococcus mutans (S. mutans) and Staphylococcus aureus (S. aureus). The pathogenic isolates were kindly provided from the microbiology laboratory of the Medical City Hospital, Baghdad, Iraq. Isolates were identified using the VITEK system (VITEK, Biomérieux, Marcy-l’Etoile, France) kindly supplied by the Division of Biotechnology, Department of Applied Science, University of Technology, Baghdad, Iraq.

Preparation of Mueller-Hinton Agar

Mueller-Hinton agar was prepared to reveal the antibacterial activity of Au NPs. According to the guidelines of the manufacturer, the agar was prepared by dissolving 28 g of the powder in 1000 mL of deionized distilled water with the assistance of heating and continuous shaking. After autoclaving at 121 °C for 15 min, the agar medium was cooled at 45 °C with cold water and solidified in Petri dishes which were left for about 15 min till they were cooled at room temperature.

Preparation of McFarland Solution

McFarland standard solution was prepared by adding Ba Cl2.2H2O (0.05 mL, 1.175%) to H2SO4 (9.95 mL, 1%). The standard solution that symbolized 5 × 107 bacterial cells per milliliter was compared with turbidity bacterial suspension, which combined and made sure from its suspending. In order to mitigate the effects of evaporation, the McFarland solution was densely packed and hermetically sealed with aluminum foil to restrict the amount of light exposure.

Anti-Bacterial Efficiency Tests

The activity of Au nanoparticles against A. baumannii, P. aeruginosa, S. mutans, and S. aureus was investigated using an agar well diffusion assay. The bacteria were subcultured on brain heart infusion agar. The agar was streaked with the organisms’ respective broth cultures 0.5 McFarland solution, which corresponded to the microbial concentration of 5 × 107 CFU/mL, and left for 15 min for the absorption to occur. Afterward, the end of a sterile Pasteur pipette (5 mm in diameter) was used to make wells in agar, and 70 µL of Au NPs at various concentrations (500, 750, 1000, and 1250 µg/mL) was poured into each hole. Deionized distilled water was used as a negative control. After 24 h of incubation at 37 °C, the diameters of the inhibition zones were measured in millimeters by measuring the diameter of circular inhibition zones around the well using a physical ruler.

Results and Discussion

The absorbance spectra of gold nanoparticle Au NPs synthesized by PLAL as a function of laser fluence (2.2, 4.4, 6.64, 8.85, and 11.1) J/cm2 with 300 laser pulses are shown in Fig. 2. The peak of absorption intensity was observed at 530 nm, which increased as laser fluence increased. This indicates an increase in the concentration of Au nanoparticles. Also, the position and height of the peak in the absorption spectrum depend on the shape, size, and yield of the obtained particles. This phenomenon can be attributed to surface plasmon resonance’s impacts, which are contingent upon the size and shape of the nanoparticles [23, 24]. The absorbance shows that the height of the plasmon peak increases with increasing laser fluence indicating an increase in the concentration of gold nanoparticles. This finding was substantiated by an investigation by Kuriakose et al. and Nasiri et al. [25, 26]. It was also observed that the highest degree of light absorption occurs at wavelengths of 522 and 526 nm, respectively. According to a study conducted by Prakash et al. [27], the surface of the resonance plasmon characteristic is influenced by the dimensions and morphology of the gold nanoparticles that are produced [28,29,30]. Figure 3 shows an absorbance spectrum of Au NPs prepared at different numbers of laser pulses (100, 200, 300, 400, and 500) and laser fluence of about 4.4 J/cm2. The plasmon absorbance peak was shown at 433 nm and increased with an increasing number of laser pulses, indicating an increase in the concentration of gold nanoparticles.

Fig. 2
figure 2

UV-VIS absorbance spectra of Au NPs prepared with different laser fluence at 300 pulses

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Fig. 3
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UV-VIS absorbance spectra of Au NPs prepared at laser fluence 4.4 J/cm2 with different pulses

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Figures 4 displays the FTIR spectrum of gold NPs prepared with different laser fluences (2.2, 4.4, 6.64, 8.85, and 11.1) J/cm2 and 300 pls, while Fig. 5 shows the FTIR spectrum of gold NPs prepared with different numbers of laser pulses (100, 200, 300, 400, and 500) and 4.4 J/cm2. From all FTIR spectra, the broad peaks within the range of 3200–3600 cm−1 are ascribed to the O–H stretching vibration, as reported in reference [31]. The peak at ~2100 cm−1 is identical to the stretching vibration of C=C, which is ascribed to carbon dioxide present in the atmosphere. The peak between 1630 and1660 cm−1 is attributed to the O–H stretching vibration mode and the bending of water. The peaks at ~710 cm−1 demonstrate the vibration properties of NPs. These specified functional assemblies could have been involved in reducing the metal ions, donating electrons to the stability of NPs, and helping to avoid their agglomeration. Also, a peak about ~500 cm−1 is the fingerprint of Au-NPs. The confirmation of the creation of NPs of Au was established based on the results obtained from FTIR analysis. The spectral peaks exhibit a shift toward higher wavenumbers upon an increase in laser fluence, commonly referred to as a blue shift. These results are in full agreement with previously published data by Gurunathan et al. [32].

Fig. 4
figure 4

FTIR spectra of Au NPs prepared with different laser fluence: a 2.2, b 4.4, c 6.64, d 8.85, and e 11.1 J/cm2 and 300 pulses

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Fig. 5
figure 5

FTIR spectra of Au NPs prepared at laser fluence 4.4 J/cm2 and different number of pulses: a 100, b 200, c 300, d 400, and e 500 pulses

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Figure 6 shows the XRD pattern in the 2θ range 10°–80° for Au NPs prepared by PLAL in D.D.W. with different laser fluences (2.2, 4.4, 6.64, 8.85, and 11.1) J/cm2 and 300 pulses. Patterns exhibit peaks due to diffraction from (2θ) of ≈38.5°, 43.7°, 64.6°, and 77.5°, which return to the 111, 200, 220, and 311 reflection phases of the sample under consideration, which comprises metallic nanoparticles of gold arranged in distinct planes, as per JCPDS no. 04-0784. Based on the X-ray diffraction (XRD) pattern analysis, it can be inferred that the synthesized nanoparticles are crystalline in nature and exhibit a face-centered cubic structure, which is indicative of the presence of gold (Au) nanoparticles. The evident broadening of X-ray diffraction peaks suggests that specimens possess a nanocrystalline characteristic (Table 1) [33, 34].

Fig. 6
figure 6

XRD analysis of Au NPs prepared by PLAL in D.D.W at 300 pulses with different laser fluences: a 2.2, b 4.4, c 6.64, d 8.85, and e 11.1 J/cm2

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Table 1 Displays the findings of an XRD analysis of Au NPs
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Figure 7 shows the FE-SEM images of Au NPs prepared by PLAL in D.D.W. at 300 pulses with different laser fluences (2.2, 4.4, 6.64, 8.85, and 11.1) J/cm2, and Fig. 8 shows the FE-SEM images of Au NPs prepared by PLAL in D.D.W. at laser fluence 4.4 J/cm2 with different numbers of laser pulses (100, 200, 300, 400, and 500). This showed that the surface morphology of Au nanoparticles has an agglomerated form. The sample is made up of uniform nanoparticles of nearly spherical morphology with a narrow size distribution. This agglomeration is owing to electrostatic force among Au nanoparticles [35, 36].

Fig. 7
figure 7

FE-SEM images of Au NPs prepared by PLAL in D.D.W. with 300 pulses and different laser fluences a 2.2, b 4.4, c 6.64, d 8.85, and e 11.1 J/cm2

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Fig. 8
figure 8

FE-SEM images of Au NPs prepared by PLAL in D.D.W. with 4.4 J/cm2 and different numbers of laser pulses (a 100, b 200, c 300, d 400, and e 500)

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The PL spectra of the nanoparticles of gold are depicted in Fig. 9. Prepared by the PLAL method, with constant pulses (300) and different laser fluences (2.2, 4.4, 6.64, 8.85, and 11.1) J/cm2, the maximum intensity of fluorescent emission for Au NPs was obtained at a higher peak of ~380 nm. These results revealed that the increased laser fluence of AuNPs has a higher wavelength intensity. While Fig. 10 represents the PL spectra of the gold nanoparticles prepared by the PLAL method, with a 4.4 J/cm2 laser effect and different pulses (100, 200, 300, 400, and 500) revealing the maximum intensity of fluorescent emission for Au NPs and a higher peak at ~370 nm [37,38,39].

Fig. 9
figure 9

Photoluminescence spectrum of Au NPs prepared by PLAL with 300 pulses and different laser fluences

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Fig. 10
figure 10

Photoluminescence spectrum of Au NPs prepared by PLAL with a 4.4 J/cm2 laser fluence and different pulses

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Figure 11 shows the antibacterial activity of Au NPs against the four types of bacteria which included A. baumannii, P. aeruginosa, S. mutans, and S. aureus in different concentrations of Au NPs; the results revealed that the 1250 µg/mL has high activity against Pseudomonas aeruginosa and S. aureus, while Streptococcus has been affected by all concentrations of Au NPs; and A. baumannii has been highly affected by 1000 µg/mL and 1250 µg/mL [40,41,42]. The proposed mechanisms of the activity of the Au NPs might occur in two successive stages: firstly, by causing damage in the outer membrane of the bacterial cell wall through electrostatic or direct interaction between the cell wall and the nanoparticles. Secondly, by production and accumulation of oxygen reactive species (ROS) due to the presence of metal oxides [43]. Generation of ROS can cause changes in macromolecules such as proteins, nucleic acids, and lipids by effects of oxidative stress. These oxygen species produce free radicals that are short-lived and unstable, which influences nuclear viability and healthiness and finally leads to cell death [44, 45].

Fig. 11
figure 11

Antibacterial activity of Au NPs prepared with 4.4 J/cm2 and 500 pulses at various concentrations against P. aeruginosa., S. aureus., S. mutans, and A. baumannii. A control, B 500 µg/mL, C 750, D 1000 µg/mL, and E 1250 µg/mL

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Conclusion

In this study, we investigated the successful, rapid, and easy production of Au NPs via pulsed laser ablation of gold targets in water. Fourier-transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), field emission scanning electron microscopy (FE-SEM), UV-visible spectroscopy, and PL spectra were used to examine the Au nanoparticles’ features. The produced nanoparticles have an almost spherical shape and high aggregation, with a face-centered cubic structure depending on the laser fluence. Eco-friendly Au NPs demonstrated potent antibacterial activity in all tested strains; this nanomaterial could be used as an alternative ingredient in antimicrobial drugs.