Evaluation of Cadmium Oxide Nanoparticles’ Antibacterial and Antifungal Potential

INTRODUCTION
According to [1], pathogenic microorganisms are a serious health problem since they result in a significant number of hospital admissions and fatalities each year. Antibiotic-resistant bacteria are developing alarmingly [2].
Antimicrobial modification is a desirable goal to stop the growth of resistant microorganisms. Microbial invasion can result in serious infection [3]. Therefore, a high-priority topic for study is the creation of new antimicrobial chemicals or modifying existing ones [4]. The creation of antimicrobial materials and surfaces for use in healthcare, food, biomedicine, and individual care applications is therefore of tremendous interest [3,5,6].
A wide range of fungi, bacteria, and viruses are sensitive to the biocidal activity of metal ion-based nanomaterials [7]. Organic antimicrobial substances have been utilized as bactericides and insecticides for a very long time. Due to their antibacterial capabilities, manufacturing process heat was lowered. On the other hand, inorganic compounds demonstrated good thermal stability and resistance to bacteria [8]. Since the simplest structures are nanoparticles (sizes: 1-100 nm), any bound atom with a structural radius smaller than 100 nm is regarded as a nanoparticle. Nanoparticles have characteristics that set them apart from bulk materials, including low melting temperatures, greater surface areas, distinctive optical qualities, mechanical strength, and distinctive magnetizations [9].
Such “nanoparticles,” which are essentially smaller versions of contemporary particles and are formed of various metals, can be produced via nanotechnology [10,6]. Although nanoparticles are still in the research and development stage, progress indicates that they have a promising future, especially in computer and sensor applications [11]. 
Metal nanoparticles can perform a variety of tasks that are not possible in the bulk phase [12,13]. Due to their distinctive physicochemical characteristics, such as optical, catalytic, electrical, antibacterial activity, and magnetic capabilities, metal nanoparticles have received much research. Since bacteria are the lowest-level organisms in ecosystems and so enter the food chain, the impact of nanoparticles on them is crucial [14].
An essential property of cadmium oxide (CdO), an n-type II-VI semiconductor, is its low electrical resistivity, broadband gap, and excellent visual transmittance. Burning Cd in the air produces the brownish CdO that is seen. It can be converted to conductive oxides because it absorbs CO2 from the air and is insoluble in water. CdO has a direct band gap of 2.5 eV and an indirect band gap of 1.98 eV. It is possible to sterilize and get rid of industrial and environmental microorganisms by manufacturing efficient nanomaterials with high activity against specific bacteria. Therefore, a study of its antibacterial activity is required. We evaluated the antibacterial and antifungal properties of cadmium oxide nanoparticles in this study.

MATERIALS AND METHODS 
Microbial culture 
The bacterial pathogens used in the study were obtained in 2015 from AL-Sadder University Hospital’s Bank Culture Collection in Maysan, Iraq. They consist of seven gram-negative bacteria, such as P. aeruginosa and S. typhi, M. morganii, K. pneumoniae, Serratia sp, P. vulgaris, and the reference isolate Escherichia coli (ATCC 25922), as well as seven gram-positive bacteria, such as S. epidermidis, S. pneumonia. Nutrient Agar (NA) was utilized to cultivate bacteria, while Sabouraud Dextrose Agar (SDA) slants were employed to cultivate yeast.

CdO nanoparticle preparation
Pure CdO was obtained by sol–gel spin coating. All chemicals were from Sigma Aldrich. Cadmium acetate dihydrate (Cd(CH3COO)2), methanol (CH3OH), triethylamine (N(CH2CH3)3) and glycerol (C3H5(OH)3) were used as starting precursors with analytical purity. The CdO precursor consists of two solutions. The first was made by dissolving 4.0 g dehydrated cadmium acetate in 7 ml methanol. Then slowly add 0.25 ml of glycerin while stirring constantly. The second solution was prepared separately by dissolving 1 mL of triethylamine in 7 mL of methanol. This solution was slowly added to the first solution.The coating solution was applied to the slide and centrifuged at 1000 rpm for 2 minutes. The film was then dried in an oven at 350°C for 8 minutes to evaporate the solvent. This process was repeated three times before finally placing the sheet in a 450°C oven for 30 minutes and immersing in air, followed by slow oven cooling.

Characterization
The structure of the films was examined using X-ray diffraction (XRD) using the Philips PW-1710 apparatus with Cu-K radiation (0.15406 nm). The morphological profile of CdOnps was examined using scanning electron microscopy (SEM) with a Philips XL40. In order to investigate the composition, energy dispersive X-ray analysis (EDX) was employed. Using a collection of typical peaks for each element and their amounts, one can deduce their atomic composition.

CdO nanoparticles’ antimicrobial activity 
According to Rezaei-Zarchi et al. (2010), the agar disc diffusion test was used to determine the antibacterial activity of CdO-NPS. In order to evaluate the antimicrobial activity, Muller Hinton Agar (MHA) plates were prepared. An inoculum of bacteria, consisting of 100 µl, was spread out on plates. A 30-microliter volume of Cdo -nps, with a concentration of 50 mg/ml, was added to each well once it was prepared. The zone of inhibition was calculated using the Hi antibiotic zone scale during a 24-hour incubation of the MHA plates at 37°C. Following the completion of the experiment in three separate runs, the standard deviation was determined.

Minimum Inhibitory Concentrations 
Each the MICs (Minimum Inhibitory Concentration, Minimum Bactericidal Concentration, and Minimum Fungicidal Concentration) were determined using standard assay dilution [15]. This test was conducted on CdO-NPS as the lowest concentration did not show any turbidity after 24 hours of incubation compared to the control tubes . The bacterial inhibition test made use of both Müller-Hinton agar and Sabouraud’s yeast with dextrose. 

RESULTS AND DISCUSSION 
The XRD spectrum of the cadmium oxide nanoparticles that were generated is shown in Fig. 1. The cubic surfaces-focused CdO data (JCPDS.Nr: 00-005-0640) perfectly matches all of the diffraction peaks.According to Cullity and Stock (2001), the crystallite size of the prepared nanoparticles was determined using the Scherrer relation D=(0.89 λ)/(β cosƓ). Here, λ is the wavelength of the X-ray source, and β is the angular full width at half-maximum (FWHM) of the XRD peak at the diffraction angle. Nanoparticles had an average size of 36 nm, according to XRD. 
The cadmium oxide nanoparticles’ crystallites are spherical, and no peculiar or other impurity peaks were detected, according to scanning electron microscopy (SEM) pictures (Fig. 2) and an energy dispersive X-ray analysis (Fig. 3). 
Cadmium oxide nanoparticles exhibited considerable antibacterial activity in seven Gram-positive zones of inhibition (11.5 mm-23 mm) and seven Gram-negative zones of inhibition (10 mm-18.5 mm), our result agree with [16] who have found antibacterial behavior of green synthesized nanoparticles against four separate bacterial species, “Pseudomonas aeruginosa, Klebsiella pneumoniae, Staphylococcus aureus, and Escherichia coli”, using agar well diffusion.  The findings revealed that the green synthesized CdOx nanoparticles demonstrated superior antibacterial activity in the field of inhibition (ZOI) against bacterial strains.
For two yeasts, the antifungal efficacy and zones of inhibition range from 10 to 10.5 mm, as shown in (Table 1). The size of the inhibitory zone varies across different types of bacteria. Evidence suggests that CdO may explain why Gram-negative bacteria are more likely to display the zone of inhibition compared to Gram-positive bacteria. 
Gram-negative bacteria die as a result of nanoparticle damage to their cell membrane structure and a decrease in the function of certain membrane enzymes [17,18].
Carefully crafted nanoparticles exhibit strong antibacterial action, as demonstrated by recent studies [19]. Table 1 shows that CdOnps exhibited low MIC values (25–200 g/mL), MBC values (50–200 g/mL), and MFC values (100–200 g/mL) against several bacterial species. Conventional wisdom held that the minimum inhibitory concentration (MIC) was the concentration at which microorganisms could not grow. Given that microorganisms have negative charges and metal nanoparticles have positive charges, an electromagnetic attraction between the two is likely to be the mechanism of action [20]. The thiol groups (-SH) on the surface of bacterial cells can be activated by ions produced by nanomaterials, leading to cell lysis (Table 1) [21,22]. 

CONCLUSION
The developed nanoparticles were characterized by XRD and SEM methods with EDAX measurements demonstrated potent antibacterial and antifungal activity against the microbes tested, demonstrating that the generated nanoparticles effectively inhibited their growth.

CONFLICT OF INTEREST
The authors declare that there is no conflict of interests regarding the publication of this manuscript.