An Amalgam of Mg-Doped TiO2 Nanoparticles Prepared by Sol-Gel Method for Effective Antimicrobial and Photocatalytic Activity


 In this study, undoped and Magnesium doped TiO2 nanoparticles (Mg-TiO2 NPs) were successfully synthesized via a simple sol-gel method cost-effectively. The prepared Mg- TiO2 NPs were characterized by UV-Vis, FTIR, PL, XRD, FESEM, TEM, and EDAX. UV – Visible Spectroscopy showed that an increase in the optical bandgap concerning the concentration of dopant Mg increases. The bandgap values were found to be 3.57-3.54 eV. FTIR spectra showed that the presence of the characteristic stretching and bending vibrational band of Ti – O bonding at 468 cm-1 and shifts in vibrational bands were observed for Mg-TiO2 NPs. PL spectra of Mg- TiO2 NPs at different concentrations exhibit a strong UV emission band. X-ray diffraction confirmed the formation of the tetragonal anatase phase. The average crystallite size of prepared samples was found to be 22-19 nm. The average crystallite size of Mg- TiO2 NPs decreases with increasing the concentration of dopant Mg. The FESEM and TEM analysis confirmed that the spherical morphology for both TiO2 and Mg-TiO2 NPs. SAED pattern confirms the crystalline nature of prepared samples. EDAX spectra confirm the presence of Ti, O, and Mg and confirm that Mg2+ ions are present in the TiO2 lattices. The prepared samples were investigated against gram-positive (B. subtilis and S. aureus) and gram-negative (P. eugenia and K. Pneumonia) bacteria. The prepared samples showed potent antibacterial activity against gram-negative bacteria than the gram-positive bacteria. The prepared samples show the excellent photocatalytic degradation for Methylene blue.


Introduction
Nanotechnology has a wide range of applications such as electronics, catalysis, agriculture, optical communications, food packaging, etc. [1][2][3]. In the present era, nanomaterials have a greater interest in many fields because of change their optical and physical properties when particle size reduces to the nanoscale. The recent studies on semiconductor nanoparticles also suggested that optical bandgap becomes increased that as particle size decreased thereby change in its optical and electrical properties and thus making the nanomaterials suitable for several applications [4][5][6]. The performance of nanomaterials depending on the size and shape that are affected by the high surface to volume ratio. Different types of nanomaterials are used to enhance the optical, electrical, thermal, photocatalytic, antibacterial, and gas sensing properties [7,8]. Fe2O3, WO3, Bi2O3, MgO, ZnO, and TiO2 are the most semiconductor nanomaterials that are used for photocatalyst, antibacterial applications, and safe for human beings, animals, and plants [9]. Among all the metal oxide nanomaterials, TiO2 is an n-type semiconductor with a wide bandgap of 3.2 eV, UV light absorption, high chemical and thermal stability, and tetragonal structure [10]. TiO2 has many applications in the field of biomedical, photocatalytic activity, antibacterial activity, gas sensors, solar cells, agriculture, water purification, textiles, food packaging, etc. [11].
TiO2 occurs in three crystalline forms such as anatase, rutile, and brookite phase. These three forms have high refractive index values, which are 2.488, 2.609, and 2.583 respectively.
Among these three forms, anatase is metastable, rutile is highly stable and brookite is unstable [12].
The anatase form is considered the most physically and chemically active phase of TiO2 [13]. Shape and size-controlled synthesis of TiO2 nanoparticles enhance its properties have been extensively studied in recent years [14]. There are several methods for the synthesis of TiO2 nanoparticles such as sol-gel [15], wet chemical [16] co-precipitation [17] hydrothermal [18] ball milling [19] combustion [20], and biological method [21]. Among these methods, sol-gel is the most feasible method for the synthesis of TiO2 nanoparticles because of its ability to control size and surface morphology. The sol-gel method has greater advantages, which include high purity, the low temperature required for synthesis, and excellent homogeneity of nanoparticles [22,23]. TiO2 nanoparticles also exhibit potent antibacterial properties that are useful in many biological applications.
Recently, there are increasing research in alkali metal ion (Al, Ca, Ce, Co, Cu, Fe, Ga, In, Mn, Mg, Nb, Sn, and Sr) doping method, which leads to change in physical and chemical properties of TiO2 nanoparticles and enhances the antibacterial applications [24]. Among these metals, Mg-TiO2 NPs exhibit potent antibacterial activity, because Mg 2+ can be substituted into the Ti 4+ ion owing to their smaller ionic radii. The smaller ionic radii of Mg 2+ ion helpful to enhance the antibacterial efficiency [25]. The efficiency of antibacterial activity also depends on the structure of microbes.
Matsunaga et al [26] reported that TiO2 nanoparticles showed good antimicrobial activity against Escherichia coli under UV irradiation. To overcome the drawbacks transition metals are doped to enhance the antibacterial efficiency under visible light. Karunakaran et al [27] demonstrated that Cu doped TiO2 nanoparticles effective antibacterial activity towards E. coli and S. aureus under visible light. Meanwhile, they also studied Ni-TiO2 NPs against grampositive and gram-negative bacteria. Hamal et al [28] reported that Ag-doped TiO2 nanoparticles enhance antibacterial activity against E. coli and B. subtilis and suggesting that the Ag is responsible for the enhancement of antibacterial efficiency. According to earlier reports, few works have been carried out on the effect of TiO2 nanoparticles on antimicrobial activity [29].
For the preparation of Mg-TiO2 NPs, 0.2g of Magnesium nitrate was prepared in 100 ml of deionized water. Subsequently, 5ml of Titanium (IV) Isopropoxide was prepared in 100 ml of Isopropyl Alcohol. Then the aqueous solution of Magnesium nitrate was added drop wise to form a homogenous mixture. After that, 0.8 M of aqueous NaOH solution was added dropwise to this homogenous mixture to form white precipitation. Then the homogenous mixture was stirred at room temperature for 5 hrs. Further, a homogenous mixture could age for 24 hrs. and then the white precipitate was washed with ethanol and distilled water to removed unwanted impurities present in the solution. Then the solution was centrifuged at 10000 rpm for 30 min. Finally, the precipitate was dried at 120 o C for 2 hrs and annealed at 450 o C for 5 hrs. to obtain Mg-TiO2 NPs. The same procedure was followed for different concentrations of dopant Mg (0.4g, 0.6g and 0.8g). The obtained samples were ground with pestle and mortar and stored in an airtight container. The annealed samples were used for further studies. The same method was followed for TiO2 nanoparticles without the addition of Magnesium nitrate.

Characterization:
The prepared undoped and Mg-TiO2 NPs were examined using the following characterization techniques. UV-Visible absorption spectroscopy was obtained in the wavelength range 200 -800 nm using a UV visible spectrophotometer (JASCO-V-770 Spectroscopy).
Fourier transform infrared spectroscopy (FTIR) was carried out by using Bruker Alpha FTIR spectrometer at a wavenumber range of 400 cm-1 -4000 cm-1. Photoluminescence spectroscopy of the prepared samples was analyzed using an FP-3800 spectrofluorometer. XRD diffraction pattern was analyzed using Bruker D8 Advance X-ray diffractometer with Cukα1 (l = 1.

Antibacterial experiment
Bacillus subtilis, Staphylococcus aureus, Pseudomonas aureginosa, and Klebsiella pneumonia were chosen as microbes for antibacterial assays. The antibacterial activity of undoped and Mg-TiO2 NPs was tested using the disc diffusion method.
In brief, the microbes were cultivated in Müller-Hinton broth at 35°C ± 2°C on detour shuddering incubator (Remi, India) at 160 rpm. A pasture of microbial culture was arranged by dispersion of 10 mL culture broth of all test microbes on dense nutrient agar plates. The dishes were permitted to stand for 10-15 minutes for culture absorption. The 5 mm size discs/wells were perforated into the agar with the dome of sterilized micropipette tips. Using a spatula, 100 μg of undoped and Mg-TiO2 NPs were kept into each of the discs on all plates. The microbes were inoculated to the culture media by inoculation in the petri dishes and incubated at 35±2°C for 24 hours for culturing bacteria. After incubation, the diameter of zone of inhibition were examined.

Photocatalytic degradation study
Where A0 is the initial absorption, A is the absorption after a time t and k is the first-order rate constant. The Scherrer's formula was used to calculate the crystallite size of undoped and Mg-TiO2 NPs as follows [32], Where K is the Scherrer's constant,  is the full wave half maximum (FWHM) of the X-ray diffraction (radians),  is the wavelength of the X-ray (nm) and  is the diffraction angle. The assessed crystallite size of as-prepared nanoparticles was found to be 22 nm, 21 nm, 20.4 nm, 20 nm, and 19.6 nm respectively. The crystallite size is found to decrease with Mg-TiO2 NPs increases which are due to Mg 2+ ion is incorporated into the TiO2 lattice. The doping with Mg with TiO2 also increases the oxygen species and these oxygen species are responsible to enhance the antibacterial and photocatalytic activity.
The lattice constant of the tetragonal anatase phase of undoped and Mg-TiO2 NPs was calculated using the formula, Where d is the interplanar spacing, a and c are lattice constants, h k and l are the miller indices.
Positional parameter (u), bond length (l), and volume of the unit cell (V) of as-prepared samples were obtained using the following relation.
The obtained value of crystallite size, lattice parameter, positional parameter, bond length, and unit cell volume of undoped and Mg-TiO2 NPs are summarized in table 1. As presented in table   1, the crystallite size of as-prepared nanoparticles decreases when increases the dopant concentration Mg and also a slight variation in the positional parameter, bond length, and volume of the unit cell values, this might be due to the incorporation of Mg 2+ ion into the TiO2 lattice.

Morphology and elemental Analysis of as-prepared nanoparticles
The surface morphology of the as-prepared undoped and Mg-TiO2 NPs was examined using FESEM analysis and results are shown in fig 2. The morphology of as-prepared nanoparticles shows a spherical shape. The particle size of undoped and Mg-TiO2 NPs is around 25 nm. From the XRD results, it can be inferred that the crystallite size is less than the particle size and it proves that the prepared nanoparticle is in crystalline nature. In addition, the aggregation and agglomeration occur in the prepared nanoparticles and it is shown in the FESEM images. The decrease in agglomeration can be attributed to the increasing the dopant concentration Mg and particle size also decreases. The crystalline is defined as the lowest even crystallographic unit based on the disorientation of the adjacent atoms and the nanoparticle consists of more than one crystalline with dissimilar direction. Here, particle size obtained from  Mg-TiO2 NPs. d -spacing values are calculated using the relation [33].
Where L is the camera length (120 nm), l is the wavelength of the electron beam and R is the radius diffraction ring respectively. d -spacing values of undoped and Mg-TiO2 NPs were found to be 0.239 nm and 0.268 nm respectively which corresponds to the (1 0 1) tetragonal anatase phase of TiO2 nanoparticles. Lattice spacing values were found to be slight increases when Mg-TiO2 NPs which are ascribed to the imperfections in TiO2 lattice due to metal ion doping [34].
The intensity of the crystalline phase of TiO2 nanoparticles was decreased which are wellmatched with intensity peaks of XRD results. The crystallinity of as-prepared samples was assessed using the selected area diffraction pattern (SAED) and portrayed in fig 4 (E-F). The ring pattern confirms the anatase crystalline nature of as-prepared nanoparticles and a bright spot indicates the formation of high crystallinity nature of undoped and Mg-TiO2 NPs (1 0 1) anatase phase. The mean particle size of undoped and Mg-TiO2 NPs was obtained to be 24.6 nm and 21.9 nm respectively. The assessed particle size is in good accord with the crystalline size of XRD results. From the TEM results, particle size decreases the increasing the dopant concentration which is due to Mg 2+ ion is incorporated into the TiO2 lattice. The smaller particle size improves photocatalytic activity.

Optical properties and bandgap assessment of as-prepared nanoparticles.
The optical properties of as-prepared nanoparticles were examined using UV-visible absorption spectroscopy carried out at room temperature and depicted in fig 5. Generally, TiO2 nanoparticles tend to absorb UV light of bandgap 3.2 eV. The absorption peak exhibit UV cutoff wavelength which is attributed to photoexcitation of electron from the valence band (formed from 2p orbital of the oxide anion) to conduction band (formed from the 3d orbitals of the Ti 4+ cation [35]. The shift in the absorption edge was observed for Mg-TiO2 NPs which are ascribed to the acceptor tendency of Mg in the TiO2 and creation of additional state within the TiO2 lattice which leads to reducing the bandgap.
To estimate the bandgap of as-prepared samples, Tauc's formula is used from UV visible spectra [36]. nanoparticles. This peak plays a significant role in enhancing photocatalytic activity.

Antibacterial activity
Generally, the antibacterial activity of nanoparticles depends on various factors such as phase formation, particle size, surface morphology, specific surface area, chemical composition, and surface hydroxyl groups [42,43]. The several killing mechanisms of TiO2 nanoparticles explained in literature such as ROS generation, superficial tension leads to cell damage, Ti 2+ ion penetrates cell membrane leads to damage of cell wall, hole creation, and leakage of intracellular electrolytes [44,45]. Amongst, ROS creation was mostly used to describe the antibacterial activities of TiO2 nanoparticles.
According to the ROS creation of TiO2 nanoparticles, additional electron-hole pairs might be formed on the surface of nanoparticles. ROS creation mainly consists of hydroxyl radicals, hydrogen peroxide (H2O2), and superoxide anion radicals [46]. Furthermore, TiO2 nanoparticles bind with the external microbial membrane and enter the cell wall. This damage the cell wall, DNA, lipids, and protein synthesis and leads to bacteria viability [47]. The killing mechanism of TiO2 nanoparticles is given below [48].

TiO2
TiO2 * + e -+ h +  (9) H2O + h+ OH* + H +  In this study, Mg-TiO2 NPs (0.8g) exhibited higher antibacterial activity because of the smaller crystallite size with a larger surface area. In addition, Mg also increases the oxygen vacancies in ROS generation to enhance the antibacterial activity. The doping of Mg with TiO2 nanoparticles leads to the variation in particle size, morphology, and solubility of Ti 2+ . The results reveal that Mg-TiO2 NPs will be a promising candidate for a potential drug delivery system to cure some significant infections in the future.

Photocatalytic activities
The photo corrosion is caused by the reaction of oxygen species and holes present on the surface of TiO2 nanoparticles. According to XRD results, Mg-TiO2 NPs leads to increases in the oxygen species and also increases the chemical stability of photocatalytic reactions.   Photocatalytic degradation of A) undoped and B) Mg-TiO2 NPs (0.8g) Figure 12 Photocatalytic mechanism of undoped and Mg-TiO2 NPs Degradation e ciency of A) undoped and B) Mg-TiO2 NPs (0.8g)