Structural Analysis
XRD pattern analyzed by the particle size and the crystal structure are shown in Figure.1 Here, the XRD pattern can be indexed well to the rhombohedral structure of Fe2O3 (JCPDS 89-2810) where the diffraction peaks are found at 24.22º, 33.23º, 35.53º, 40.77º, 49.25º, 54.04º and 62.71º with correspond to (220), (311), (110), (400), (202), (311) and (214) plane, respectively. The particle size is calculated by using Scherrer’s formula 13 given below,
![](data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAH4AAAA8CAYAAACgsw7vAAAFrElEQVR4Ae2bPU70MBCG9w7cgIIbcAJoaaGjpqbmCJwADoBooaagokO00NEg0SFxAH96on2X2ZHDfs7f7uKxFJzYjn/mmRlPkmWWIlUpgVmVq45FpwBfqRIE+ABfqQQqXXZYfICvVAKVLjssPsBXKoFKlx0WH+ArlUClyw6LD/CVSmCDlv309JRms1kiHzuFxXeU8OXlZQMJUPf39yt7OT09XbTn3ly6ublJLy8vqa0+d0/XsgDfQXKA3t3dTR8fH83B+W/wgc5B4h7ae7jv7+/p6+uraePrOkxx5S0BfqWIlht8f3+ng4ODJdBAp4w6n56fn9P+/n4DXHVWcVRm87u7O3s5ynmALxQrIPf29hK5Uq5MdTnItN/Z2VlSHrXH6tk+xk7jjzD2CibuPwdS7ps6nygDsleUNvDs8wHeS3EDrkvBSynsVpBTBpb2+fnZQL+4uGjOx1xuWHyhdHPgf3PddK96LFmHgkM7PMCvrq4Se/zr66utGvw8wBeKFIgle7zvXsGhj9x5jEMp2OMDvJfaBlwLHJavBETrylWey3NtgX10dJQeHh6aWwCvcwp41Bs6hcV3kCjQBVp7uFWEXJdSmJyLB/TZ2dniNqBj/QR6Y+33AX4h7rITLFf7tYUuwFIMu797986ICujsno4HQBHof6zXt73A29eQEgK53lKViTJaTymBXuCZKFps3ZdcnzR+ysXEWP8vgd7g7XtoDYvrw/KtC1Rd5JshgV7gZd0esMrD5W8G5NwseoEHuHXzGkABToCXRDYv7wUesLm9fB0Wb4PL3DmPTJF+JNAZvKw694iiR5hc3c/QcbZOCXQGn3t1qYXgCXJbgOo3Jc95hr9U9pucO4PHmnNuvi2ipz1CRSHe3t6ae/k0eXJysngRYj9fojwWQm4suzDbNncert5KK3X7N2nt4d6VC5aP8jUk7Tm4H49BYss4Pj5urqmjDxJtDg8PF79csXVNg/jTSwLFFg+AnEVRtsoqUQi+bNmfIqEA5+fnzSKATv8kylEIFIPkFaEpjD+dJVAMvvNILfC0NaA4gs4YnNvrqcDznlzfxBmfc8r+WpoMvFw6Fm+3Aqycr1By9whYbbUdUMY92gbGhMDHEftJlM+lwF9H4nMsYyOfoRVwMvC4c0BaS7ZW3FYu6DbwGwsCX8jwPDbxWZRj6sQPM1A6eRvAD6mAy6scYXVYqdw4oInquSZw0znQUQrg4hHIaaNjVeww1LSJ/D1k5oDFTZmAzbj2kyxzo2yoNFxPQ81ojf2ggPaxT//SJKubamqCbMfVjzOG+jVOgDc02d+BLcFjYbjcXCIOwRPRhsPGH77OeywUTPflXnQxDw6bNCd+uDFECvBGisCwv4RpE7bAKkglVwyiOuCSdC34/jH19vZ28a5CU2Ee7O+KL8ilKGrTNw/wcwniQhGuTSpDAWwCtEDacs6p81as+IU6nVsPYftoG1OKYNv2OV9eaZ+etvxeXDzC9Qll8MEd1twGnjoPXkGtvIDg07dXAD1Z2C1Gv8uzj5l+nqXXAX4uMaxaYCRE/dbdRtfU5axa91AHUHIlgbdl6se3FWQLXluODfa4H0WgHY95fo4auy0P8HPJsI9a8Ag5F2TRXCCttWLF19fXiz3dWj39ykPQ7vHxccGDPuy4VDAXPbPL9VtF0M0oDQnoaq+6VXmAn0sIN49LV0CFUBGmtzIJVPBpxyGw1Cugy9VZN0+9VR71zZgog97Y2YBTbShjfnrMK432A3xKDVwgYF3bkrBy7fnkPg5ZtY4An1KzTwJ+mxLWLvcf4DuSw1r8C5OOXU12m1VUtoVw9R1Ej4svFVyHYQa9BfAA11Ha+Xb5t9LV/dH2BH+5dw4lyw3wJdLakLYEdqXBnJ96gPcSqeQ6wFcC2i8zwHuJVHId4CsB7ZcZ4L1EKrkO8JWA9ssM8F4ilVz/AyRYZ528fcfvAAAAAElFTkSuQmCC)
where λ is the x-ray wavelength (Cu Kα = 1.5418Å), k is a constant (0.916), β is the full width at half maximum (FWHM) of the peak and θ is the Bragg angle peak position. The lattice constants for Fe2O3 nanoparticles are a = b = 5.040 Å and c = 13.74650 Å and the average crystal size has been calculated as 12 nm. From JCPDF chart 77-0777, the XRD confirmed that Er2O3 possesses the cubic structure. By applying the Scherrer’s formula, the FWHM for the strongest reflection peak of (222) with cell parameters a = 10.53 Å, the particle size was calculated to be 15 nm. Further, XRD pattern was also analyzed for the erbium doped iron composite material. It has been found that the peaks of iron oxide and erbium oxide are shifted to new position when they are used as composite; the particle size was calculated to be 17 nm
Fourier transforms infrared spectroscopy (FTIR) Studies
The FTIR spectra of Fe2O3, Er2O3 and Er2O3/Fe2O3 NPs were shown in Fig. 2 where the three spectra were combined and the functional groups were also analyzed. From the functional groups present in the three were compared and found the peaks ranges from 406 cm-1 to 555.49 cm-1 have the halogen compounds, the peaks of 769.97 cm-1 and 759.95 cm-1 of iron oxide and iron oxide composited with Erbium oxide have the halogen compound. Peaks of 530.42 – 528.48 cm-1 and 555.49 cm-1 show the flattened peak of Fe-O and Er-O bonds for metal oxides (M-O) 14. The absorption band of 109.27 cm-1 is due to the vibration of crystalline Fe-O modes 15. The broadband at 3203.76 - 3069.10 cm-1 may be assigned to OH stretching vibration of hydroxyl group. The band at 1643.36 and 1693.60 cm-1correspond to C-O stretching vibration and C = O symmetry, respectively. FTIR functional information gives the justification of good chemical property and consistent with the XRD pattern16.
Optical Absorption Spectral Studies
An optical absorption spectrum of the Fe2O3, Er2O3and Er2O3/Fe2O3 NPs was carried out between from 300 to 900 nm using UV visible spectrometer and the results are shown in Fig. 3(a). The optical studies reveal that the absorption peaks were found to be 377 nm for Fe2O3 NPs. For Er2O3 NPs, the absorption peaks were found to be 204, 379, 519, 655, 802,976 and 1520 nm. Similarly, for Er2O3/Fe2O3 NPs, the band peaks were present in 348 nm17 (Stjepko et al.2019).
Owing to the direct band gap, Fe2O3, Er2O3and Er2O3/Fe2O3 NPs under study have an absorption coefficient (α) obeying the following relation for high photon energies (hν);
![](data:image/png;base64,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)
Here, Eg is optical band gap, h is Planck’s constant and ν is the frequency of the incident light. The variation of (αhν)2 against of hν is shown in Figure. 3(b). The Eg is evaluated by the extrapolation of the linear part and the band gap for Fe2O3, Er2O3and for Er2O3/Fe2O3 NPs was found to be 2.07, 4.85 and 2.33 eV, respectively18,19
Photoluminescence Spectra
Photoluminescence (PL) spectroscopy is a suitable technique to determine the crystalline quality and the exciton fine structure. Figure 4 represents the PL spectrum of Fe2O3, Er2O3 and Fe2O3/Er2O3 as a composite nanoparticle at room-temperature. The sample exhibits two strong emission bands, namely, blue and blue green emissions at 368 and 438 nm, respectively. These peaks are due to the structural defect existence including oxygen vacancies and the results are consistent with the earlier reports20. The recorded spectra show sharp PL peaks for green and red centered about 654, 656 668 and 733 nm due to phonon assisted population mechanism of transition of Fe3+ and Er3+21, 22.
Anti-bacterial Studies
As prepared samples of Fe2O3, Er2O3 and Er2O3/Fe2O3 NPs against E. coli (Gram –ve) and Bacillus sp (Gram +ve) show good antibacterial activity 23. The mechanism of transition of Fe3+ and Er3+ results in the generation of more reactive and potent oxygen species 24,25.
Sample with Fe2O3nanoparticles of 100 μl demonstrated higher activity against Bacillus sp and the inhibition zone was 12 mm, followed by E. coli whose inhibition zone was found to be 19 mm, respectively. The iron oxide nanoparticles of 200 μl and 300 μl demonstrated increasing tendency of the inhibition zone and the details are presented Table1. Similarly, the antibacterial activity of Fe2O3 nanoparticles against the bacterial such as Bacillus sp, and E. coli is shown in Figs. 5(a) and 5(d).
From Figs.5(b) and 5(e) and Table1(a), it can be observed that the sample with Er2O3 nanoparticles of 100 μl were active against Bacillus sp, and the inhibition zone was 19 mm, followed by E. coli whose inhibition zone is 13 mm. When the concentration of sample increases with the 200 μl, the sample was active against bacterial but the inhibition zone increases to 22 mm against Bacillus sp and 15 mm against E. coli. Finally, the inhibition zone is 23 mm in 300 μl extract for Bacillus sp which is higher to the standard value. In the case of E. coli, the inhibition rises to 15 mm was observed.
From Figs. 5(c) and 5(f) and Table1(a), it has been found that the sample (Fe2O3/Er2O3 nanoparticles) of 100 μl was active against Bacillus sp, and the inhibition zone was 21 mm, followed by E. coli whose inhibition zone was 21 mm. When the concentration of sample is increased to 200 μl, the sample was active against bacterial but the inhibition zone increases to 23 mm against Bacillus sp and 22 mm against E. coli. However, for the sample with 300 μl, the sample was active and inhibition zone is 27 mm for Bacillus sp which is higher than the standard value. In the case of E. coli, the enhanced inhibition of 23 mm was observed.
It is envisaged that there exists an electrostatic interaction between positive charged nanoparticles and negative charged bacteria. Owing to this, the small-sized nanoparticles penetrate inside the cell wall and eventually they cause the cell damage. During the generation of reactive oxygen species (ROS), small-sized particles of constituent ions are released through reflux mechanism 26. Therefore, the antibacterial activity of the Fe2O3, Er2O3 and Fe2O3/Er2O3 was found to be enhanced rate.
Anti-Fungal Studies
The antifungal activity of Fe2O3, Er2O3and Er2O3/Fe2O3 NPs is carried out. The nanoparticles were also subjected to the evaluation of their antifungal activities against the different strains such as Aspergillus and Mucor with different concentrations.
Sample with Fe2O3 nanoparticles of 100 μl demonstrated higher activity against Aspergillus and the inhibition zone was 11 mm, followed by Mucor whose inhibition zone was found to be 18 mm. The iron oxide nanoparticles of 200 μl and 300 μl demonstrated increasing tendency of the inhibition zone and the detailed results are presented in Table.1(b). Similarly, the antifungal activity of iron oxide nanoparticles against the fungi such as Aspergillus, and Mucor is shown in Figs. 6 (a) and 6(d).
From Figs. 6 (b) and 6 (e) and Table1 (b), it has been observed that the Er203 nanoparticles with 100 μl were active against Aspergillus and the inhibition zone was 18 mm and the Mucor’s inhibition zone was 13 mm. When the concentration of sample is increased to 200 μl, the sample was also active against fungi but the inhibition zone is increased to 21 mm against Aspergillus but 14 mm against Mucor. However, for 300 μl extract, the inhibition zone was found to be 22 mm for Aspergillus and this is higher than the standard value. In the case of Mucor, the inhibition has been raised to 17 mm.
From Figs. 6 (c) and 6 (f) and Table1 (b), it is clearly seen that the Er2O3/ Fe2O3 nanoparticles of 100 μl were active against Aspergillus and the inhibition zone was 13 mm, followed by Mucor whose inhibition zone was 15 mm. When the concentration of sample is increased to 200 μl, still the sample was active against fungi and the inhibition zone was extended to 19 mm against Aspergillus but 16 mm against Mucor. Lastly, for 300 μl, the sample was still active for Aspergillus and the inhibition zone was 21 mm which is higher than the standard value. In the case of Mucor, the inhibition has been extended to 16 mm.
The size of the bacteria/microbes is typically in the order of micrometer27, 28. Nearly, thousands of nanopores were found in their outer cellular membranes. At this juncture, it should be emphasized that the size of the particle of the iron oxide and erbium oxide and Fe2O3/Er2O3 is less than 100 nm, and they are unlikely to enter the cell wall and damage the fungi from the interior. In addition, the liberated metal ions from the doped nanoparticle may also impact the antifungal activity in disrupting the cell membrane and gaining entry 29.
Cytotoxicity and Cell viability Analysis of MDA MB-231 Cell Line
In the last two decades, there has been immense interest in the pharmacological effects of bioactive compounds on cancer treatments and prevention. The research investigations reveal that there are numerous anti-cancer activities in various cancer cells through different forms of cytotoxic effects without exhibiting considerable damage to normal cells 30-32. In vitro cytotoxic potential of Fe2O3, Er2O3and Er2O3/Fe2O3 NPs and human breast cancer MDA MB-231 cell line and viability of tumor cells were confirmed using MTT assay. When the cells were treated with various concentrations (0, 5, 10, 25, 50, 75, and 100 µg/mL) of iron oxide, erbium oxide and Fe2O3/Er2O3 NPs for 24 hrs, a significant decrease in cell viability was observed when compared to untreated cells whose viability is assumed to be 1 (i.e.,100 %). In this study, various percentages of cell viability were examined in cultured cells. The results showed that Fe2O3, Er2O3 and Er2O3/Fe2O3 NPs induced significant potential cytotoxic response which is clearly depicted in Figs.7 (a-d). Further, all these details are also clearly presented in Table 2. Upon treatment of the MDA MB-231 cells with Fe2O3NPs for 24 hrs at 37 °C with a concentration of 5 µg/mL, a maximum cell viability of 20% has been noticed. In the case of MDA MB-231 cells with Er2O3NPs for 24 hrs at 37 °C, a maximum cell viability of 17% has been observed for a concentration of 50 µg/mL. Finally, when MDA MB-231 cells are treated with a composite of Fe2O3 and Er2O3NPs, a maximum cell viability of 21% has been noted for a concentration of 75 µg/mL. Thus, based on the above discussions, a noticeable dose-dependent reduction in cell viability has been observed. Consequently, the results of MTT assay demonstrated that iron oxide, erbium oxide and Fe2O3/Er2O3 NPs have appreciable effects on a human breast cancer MDA MB-231 cells. Thus, these results revealed the non-toxic nature and best biocompatibility of our synthesized Fe2O3, Er2O3 and Er2O3/Fe2O3 NPs in vitro experiments against normal cell MDA MB-231 and toxic nature against cancerous cells. The outcomes of our study are in conformity with the modern fact that the mainly employed biocompatible material for the preparation of nanoparticles. Interestingly, Fe2O3, Er2O3 and Er2O3/Fe2O3 NPs have not exerted significant toxic effect as >80 % viability of normal cells MCF-10A at the highest concentration, whereas human breast cancer MDA MB-231 cancerous cells show the profound cytotoxic effect. Therefore, it is suggested that the severe cytotoxicity mostly is initiated from the cellular internalization of Fe2O3, Er2O3 and Er2O3/Fe2O3 NPs instead of physical injure on the cell membrane.
From the result of anti-cancer, the cytotoxic effect for the samples clearly demonstrated that metal oxide nanoparticles show cytoxicity by inducing apoptosis in MDA-MB231 breast cancer cells. However, further studies are needed to increase the therapeutic potential of metal oxide nanoparticles to much higher extent by functionalizing them with targeting by encapsulation with other drugs. It would be interesting to study the mechanism of metal oxide nanoparticles to formulate more potent and effective anticancer drug.