Electrospun Cu–Co ferrite nanofibers: synthesis, structure, optical and magnetic properties, and anti-cancer activity

Abstract

In this study, we investigated Cu–Co ferrite nanofibers (NFs) that were synthesized for the first time employing the electrospinning technique. The structure, phase purity and crystallite size of all the prepared NFs were revealed by powder X-ray diffraction (PXRD) analysis. The NFs crystallized in the Fd3̄m (no. 227) space group and the cation distribution arrangement over distinct sites in their structure was analyzed. Scanning electron microscopy (SEM) together with energy-dispersive X-ray (EDX) spectroscopy analysis showed the microstructure of the NFs and verified their expected chemical compositions. High-resolution transmission electron microscopy (TEM) images confirmed the fibrous nature and the construction of the NFs. The band gap energies derived from the UV-vis reflectance spectra showed a blue shift with an increase in the amount of Cu in the sample from 1.42 eV to 1.86 eV. Magnetization (M) as a function of magnetic field (H) measurements performed at ambient and low temperatures showed the ferrimagnetic behavior of all the NFs. The magnetic parameters including coercivity (H_c), saturation magnetization (M_s), remanent magnetization (M_r), and squareness ratio were determined from the recorded magnetization curves. At 300 K, M_s was reduced from 78.8 to 42.4 emu g⁻¹, M_r reduced from 22.8 to 7.6 emu g⁻¹ and the Bohr magneton reduced from 3.3 to 1.8μ_B with an increase in the content of Cu in the samples. The same trend was observed at 10 K, where M_s was reduced from 93.7 to 50.9 emu g⁻¹, M_r reduced from 60.9 to 35.9 emu g⁻¹ and the Bohr magneton reduced from 3.94 to 2.16μ_B. Alternatively, H_c has the highest values for x = 0 (850 Oe at 300 K and 5220 Oe at 10 K) and x = 0.6 (800 Oe at 300 K and 5400 Oe at 10 K). The anti-cancer activity of the NFs was evaluated using the MTT cell viability assay, showing a reduction in the viability of both HCT-116 and HeLa cancer cells compared to non-cancerous HEK-293 cells after treatment with the NFs. Apoptotic activity was examined by DAPI staining, where treatment with the NFs induced chromatin condensation and nuclear disintegration in HCT-116 cells.

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1. Introduction

Nanotechnology plays a key role in the current renaissance of our life. The super and unusual properties of materials with a nano size compared to the bulk of the same materials make them optimum choices for state-of-the-art applications.^1–3 Among the materials applied in nanotechnology, nano-sized ferrites have attracted considerable interest owing to their low cost and high efficiency in a wide range of applications including industrial, medical, and environmental applications.¹ Ferrites are inorganic materials consisting of Fe³⁺ as the main cation, coupled with O²⁻ anions and other cations such as transition, rare earth and alkaline earth metals. They are grouped according to their structures and chemical formulas into spinel, garnet, ortho and hexaferrite.⁴ Spinel ferrites with the chemical formula AB₂O₄ (A is a divalent cation and B is Fe³⁺ and trivalent cations dopants if present) are the most well-known type of ferrites.⁵ Among the spinel ferrites, nano-sized Co ferrites are of scientific interest due to their eco-friendly nature, strong coercivity, acceptable magnetization and high resistivity, chemical stability and mechanical hardness.^2–6 Thus, they have been used in significant applications including microwave adsorption, magnetic resonance imaging, magnetic recording, organic dye adsorption, gas sensing, catalysis and biotechnology. However, Co ferrites suffer from low electrical conductivity, low magnetostriction coefficient and huge volume expansion.^7,8

One of the most significant features of ferrites is that their magnetic, optical, and electrical properties can be enhanced by varying their chemical composition and morphology utilizing appropriate synthesis techniques.^1,6

For instance, doping Co ferrite nanocrystals with other transition metals such as Mn, Ni and Zn can enhance their electrical and magnetic properties and expand their applications.^9–12 In the case of nanocrystalline Cu–Co ferrite, several studies have reported the synthesis of Cu-substituted Co ferrite nanoparticles (NPs) and their modified properties with respect to unsubstituted Co ferrite. N. Velinov et al. synthesized Cu–Co ferrite NPs utilizing the mechanochemical method and reported their catalytic activity.¹³ M. Hashim et al. synthesized Cu–Co ferrite NPs using the sol–gel method and studied their modified electrical properties.¹⁴ J. Balavijayalakshmi et al. synthesized Cu–Co ferrite NPs via a coprecipitation reaction and studied the influence of varying concentrations on their dielectric properties.¹⁵ B. Chandra Sekhar et al. synthesized Cu–Co ferrite via an auto-combustion reaction and reported the enhancement in their magnetostrictive properties and strain derivative values.¹⁶ However, to the best of our knowledge, there is no report to date on the synthesis and characterization of Cu–Co ferrite in nanofiber form.

One-dimensional nanofibers (NFs) possess many advantages compared to other forms of nanocrystals including light weight, high surface-to-volume ratio, accessible pore structure and easy-to-modify surface. NFs can be produced in different forms including, organic, metallic and organometallic compositions. Among them, metal oxide NFs exhibit high thermal and chemical stability, making them suitable for a variety of applications. One of the most employed methods to produce nanofibers is the electrospinning technique due to its low cost, easy preparation, probability of mixing materials and mass production capability.¹⁷

Besides the technical importance of Co ferrites in industry, they have been used for medical applications due to their bio-compatibility, activity and low toxicity.^18–20 Doping of Cu in Co ferrites is expected to enhance their bio-activity due to the role that Cu²⁺ plays in medical applications, especially in anti-cancer agents.^21–24 In this regard, the substitution of Co with Cu has been found to improve the antibacterial activity of Co ferrite nanoparticles.²⁵ Regarding anti-cancer activity, a Co–Cu ferrite/Na₂Ca₂Si₃O₉ composite (Bioglass) has been reported to have cytotoxicity against osteoblast cancer cells.²⁶ To the best of our knowledge, the anti-cancer activity of pure nanocrystalline Cu–Co ferrite has not been reported to date.

Herein, we introduce the use of the electrospinning technique to synthesize Cu–Co ferrite NFs for the first time. Furthermore, the structure, morphology, optical property, magnetism, and anti-cancer activity of the prepared NFs are discussed based on an increase in the content of Cu dopant in the sample.

2. Materials and methods

2.1. Instrumentation

Nanofibers (NFs) were synthesized utilizing an electrospinning system (NanoSpinner Unit INOVENSO, Standard Type, NS1, Co. Turkey). The phase purity of the obtained NFs was verified by PXRD (Rigaku Benchtop Miniflex, Tokyo, Japan X-ray powder diffractometer operating using a Cu Kα radiation generator). The NF morphology was imaged using scanning electron microscopy (SEM/TEM, FEI Titan 80–300 kV FEGS/TEM, Hillsboro, OR, USA). The presence and percentage of various merged elements were confirmed by energy dispersive X-ray spectroscopy (EDX) analysis. UV-vis diffuse reflectance spectra were measured using a JASCO V-700 spectrophotometer in the range of 200–900 nm at room temperature.

A Quantum Design SQUID-PPMS vibrating sample magnetometer (PPMS DynaCool, Quantum Design, San Diego, CA, USA) was used to measure the magnetization of the synthesized samples.

2.2. Synthesis

High-purity metal nitrates, polyvinylpyrrolidone (PVP, average M_w ∼ 1 300 000) and absolute ethanol were used as received (Sigma-Aldrich). To prepare the gel solution, a stoichiometric amount of Co(NO₃)₂·6H₂O, Fe(NO₃)₃·9H₂O, and Cu(NO₃)₂·3H₂O was mixed with PVP in a weight ratio of about 1 : 3 before adding 10 mL ethanol and 6.6 mL H₂O and kept stirring magnetically for 2 h until the formation of a clear gel. According to the systemic study, this metal salt to PVP ratio was found to give a clear gel that was viscous enough to spin without dropping. The electrospinning synthesis was done according to our previously published procedure.²⁷ The obtained electrospun NF samples were left to dry in air at 100 °C for 2 h before calcination at 600 °C for 5 h.

2.3. Anti-cancer assay

2.3.1. In vitro cell culture. Colorectal carcinoma HCT-116 and cervical HeLa cells were selected to examine the effect of the NFs on the number and proliferation of cells. Non-cancerous HEK-293 embryonic kidney cells were used as the control group. All cell lines were purchased from ATCC (American Type Culture Collection), Manassas, Virginia, United States. The cells were cultured in 96 well culture plates with DMEM and other nutrients according to the previously described method.²⁸ Cell growth was performed in 96-well culture plates placed in a 5% CO₂ incubator at a temperature of 37 °C. After 24 h, when cells were 75–80% confluent, they were ready for MTT examination.

2.3.2. MTT assay. During the MTT assay, NFs (5.0 μg to 50 μg mL⁻¹) were added to HCT-116, HeLa and HEK-293 cells and kept in a CO₂ incubator. After 48 h, the cells were treated with MTT (5 mg mL⁻¹) for 4 h in the incubator. Then, a plate reader was used to examine the plate at 570 nm. Finally, the cell viability was determined utilizing the GraphPad Prism Software 6.0 (GraphPad Software, USA).^29,30

2.3.3. Apoptotic DAPI staining. The apoptotic nuclear structure of CHT-116 cells was studied using DAPI staining 48 h post-treatment with NFs. In the experimental group, NFs were added at 25 μg mL⁻¹, whereas in the control group, no NFs were added. Then, 48 h post-treatment, the cells were fixed in paraformaldehyde, stained with DAPI and examined by confocal microscopy (Zeiss, Germany).^31,32

3. Results and discussion

3.1. Microstructure and morphology

The structure of Cu_xCo_1−xFe₂O₄ (x ≤ 0.6) NFs was studied via PXRD analysis. Fig. 1 displays the PXRD patterns of all the samples fitted by the Rietveld method, confirming the presence of a pure phase in each sample corresponding to the spinel ferrite structure (Fd3̄m space group). The as-measured PXRD patterns are presented in Fig. 1S (ESI).† The refinement of the experimental PXRD data was performed using the Rietveld method and FullProf software. Rietveld refinement is a powerful technique used to extract the crystal structure characteristics of a material from its PXRD reflections by estimating the peak intensity, position, height and width.³³ The crystallite size values were calculated using the Scherrer formula (Table 1). As shown in Table 1, the lattice constant a increased with an increase in the content of Cu²⁺ in the sample, suggesting that the dopant cation has a bigger ionic radius than the substituted one. In the present NFs, Cu²⁺ with an ionic radius of 0.70 Å substituted Fe³⁺, which has an ionic radius of 0.67 Å ionic radius, as confirmed by the cation distribution analysis shown in Table 2. Thus, the lattice to expands with an increase in the concentration of Cu²⁺, which is in agreement with the reported results.¹⁰ Fig. S2† shows a plot of the relation between the lattice constant a and Cu²⁺ concentration in the sample, as represented by the x value, which shows a direct proportional relation. The crystallite size (represented by D_XRD in Table 1) decreases with an increase in Cu²⁺ content in the sample in response to the elongation of the lattice constant, which has the same trend as the reported Cu-substituted Co ferrite nanoparticles.^15,25,34

Fig. 1 PXRD patterns of Cu_xCo_1−xFe₂O₄ (0 ≤ x ≤ 0.6) NFs fitted by the Rietveld method.

Table 1 The structural parameter refinement calculation for Cu_xCo_1−xFe₂O₄ (0 ≤ x ≤ 0.6) NFs with the Fd3̄m (no. 227) space group

x	a (Å)	V (Å)^3	DXRD (nm) ±0.12	χ^2 (chi^2)	RBragg
0.0	8.381(4)	588.78	35.89	1.63	2.10
0.2	8.383(0)	589.10	31.73	1.21	1.06
0.4	8.390(8)	590.76	27.14	1.59	1.05
0.6	8.400(6)	592.83	26.31	1.72	1.23

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Table 2 The cation distribution calculation for Cu_xCo_1−xFe₂O₄ (0 ≤ x ≤ 0.6) NFs

x	Tetrahedral A-site	Octahedral B-site
0.0	Co0.2Fe0.8	Co0.8Fe1.2
0.2	Co0.1Fe0.9	Co0.7Cu0.2Fe1.1
0.4	Co0.05Fe0.95	Co0.55Cu0.4Fe1.05
0.6	Cu0.1Fe0.9	Co0.4Cu0.5Fe1.1

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Table 2 displays the cation distribution of Cu_xCo_1−xFe₂O₄ (0 ≤ x ≤ 0.6) NFs over the tetrahedral and octahedral sites obtained from the PXRD patterns following the Bertaut method.^35,36 The XRD intensity ratios of the I₂₂₀/I₄₄₀ and I₄₂₂/I₄₀₀ planes were considered for the identification of the cation distribution.³⁶ The results show that the Fe³⁺ and Co²⁺ ions occupy both the tetrahedral sites (A-site) and octahedral sites (B-site); however, most of them are distributed over the B-site. The ratio of Co²⁺ decreases as the substitution ratio of Cu²⁺ increases. The Cu²⁺ ions only occupy the B-site at a substitution amount lower than x = 0.6; however, a small fraction of Cu²⁺ ions occupy the A-site for the x = 0.6 sample. The occupancy of the Co²⁺, Cu²⁺ and Fe³⁺ ions depicted here is consistent with other reports.³⁷

Scanning electron microscopy (SEM) and energy-dispersive X-ray spectroscopy (EDX) were used to analyze the NF samples. Fig. 2 shows the SEM/EDX images and the elemental mapping results for the lowest and highest x values of the Cu-containing samples, i.e., Cu_xCo_1−xFe₂O₄ (x = 0.2, 0.6). The results show that the aggregation of nanofibers containing all the involved elements with atomic ratios agrees well with the proposed chemical compositions.

Fig. 2 SEM/EDX and elemental mapping for Cu_xCo_1−xFe₂O₄ (x = 0.2 (top panel) and 0.6 (bottom panel)) NFs. In the elemental analysis graph, the X-axis represents energy (eV) and the Y-axis represents intensity (k).

Fig. 3 displays the high-resolution TEM images for Cu_xCo_1−xFe₂O₄ (x = 0.2, 0.6) at different resolutions. The images captured at around 200 nm confirm the fibrous nature of the prepared material with a nano size. The images captured at 50 nm and 20 nm illustrate the multifaced grains constructing the nanofibers for both samples. The average grain size for the lowest x value sample Cu_0.2Co_0.8Fe₂O₄ is obviously bigger than the average size of that for the highest x value sample Cu_0.6Co_0.4Fe₂O₄. Thus, the grain size was reduced by increasing the amount of Cu in the sample, which agrees with the PXRD data calculation in Table 1. This result suggests that introducing Cu in the Co ferrite lattice can address one of the issues associated with Co ferrites, which is the huge volume expansion presented by their large grain size.⁸ Given that the fibers are formed from the connection between the grain boundaries, a smaller grain size provides a stronger connection between the grains due to the larger contact area.³⁸ Moreover, the thickness of the NFs is associated with the grain size. Fig. 3a shows a thinner NF for the x = 0.2 sample constructed by bigger grains overlapping each other, appearing as a chain-like array. Fig. 3b shows a thicker NF for the x = 0.6 sample constructed by multi-grain attachments along the NF due to the large contact area of the smaller grains.

Fig. 3 High-resolution TEM images for Cu_xCo_1−xFe₂O₄ (x = 0.2 (a) and 0.6 (b)) NFs.

3.2. UV-vis optical analysis

The optical property of the prepared NFs was studied by measuring their UV-vis diffuse reflectance spectra (Fig. S3†). The absorption data was derived from the diffuse reflectance spectra using the Kubelka–Munk function F(R).³⁹ To determine the band gap energies (E_g) of the prepared NFs, the Tauc method was employed using the following expression:

(F(R)hν)^1/γ = B(hν − E_g) (1)

where h is Planck's constant, ν is the frequency and B is a constant. The γ factor depends on the nature of the material band gap, which has a value of ½ for a direct electron transition or 2 for an indirect electron transition.^40,41 Fig. 4 displays the Tauc plots for all the prepared NFs, which obey the direct band gap transition equation. All the prepared NFs are semiconductors with estimated band gap edges varying from 1.42 to 1.86 eV, which increase with an increase in the amount of Cu in the sample. The blue shift in the band gap throughout the samples is attributed to the decrease in crystallite size with an increase in Cu concentration from x = 0 to x = 0.6, which leads to the Brus effective mass model.^42,43 Moreover, this finding agrees well with the above-mentioned TEM results and PXRD calculation. The same behaviour has been reported for Co–Cu ferrite nanoparticles (NPs), where the band gap energies decrease with an increase in Cu concentration in the sample.⁴³ However, the present NF samples (1.4 < E_g < 1.9) have narrower band gap edges than their NP counterpart (1.7 < E_g < 2.5).

Fig. 4 Tauc plots for Cu_xCo_1−xFe₂O₄ (0 ≤ x ≤ 0.6) NFs. The dashed red lines are the Tauc plot leaner fits to determine the band gap energies.

The characteristics of the NFs, as a conjugated material, are mainly ascribed to the superstructure of connected particles, not only the properties of individual particles. The overlap between some conduction orbitals of the conjugated particles results in a delocalization of the charge carriers along the conjugation length. Thus, the high mobility along the NF backbone results in a reduction in the bandgap energies.⁴⁴

3.3. Magnetic analysis

The magnetic property of the Cu_xCo_1−xFe₂O₄ (0 ≤ x ≤ 0.6) NFs was analyzed by measuring the M–H hysteresis loops in the field range of ±7 T at a low temperature (10 K) and ambient temperature (300 K), as shown in Fig. 5. All the measured hysteresis loops show ferrimagnetic ordering behavior at both temperatures. Different magnetic parameters including saturation magnetization (M_s), coercivity (H_c), remanent magnetization (M_r) and squareness ratio were calculated using the measured curves, as listed in Table 3. The magnetization loops recorded at 300 K were saturated at high fields, whereas that recorded at 10 K were not. Therefore, the law of approach to saturation (LAS) method was used to determine the M_s values by extrapolating the linear fits of the M vs. 1/H² plots.⁴⁵

Fig. 5 M–H hysteresis loops of Cu_xCo_1−xFe₂O₄ (0 ≤ x ≤ 0.6) NFs at 10 K and 300 K.

Table 3 Magnetic parameters of Cu_xCo_1−xFe₂O₄ (0 ≤ x ≤ 0.6) NFs at 10 K and 300 K

x	Ms (emu g^−1)	Mr (emu g^−1)	Hc (Oe)	SQR	Bohr magneton, μB	Keff (erg g^−1)	Hci (Oe)
@300 K
0.0	78.8	22.8	850	0.29	3.31	10.5 × 10^4	2656
0.2	73.5	22.8	741	0.31	3.10	8.51 × 10^4	2316
0.4	62.0	13.6	297	0.22	2.63	2.88 × 10^4	928
0.6	42.4	7.6	800	0.18	1.80	5.30 × 10^4	2500
@10 K
0.0	93.7	60.9	5220	0.65	3.94	7.64 × 10^5	16 313
0.2	88.3	58.2	4160	0.66	3.72	5.74 × 10^5	13 000
0.4	74.8	45.1	3420	0.60	3.17	4.00 × 10^5	10 688
0.6	50.9	35.9	5400	0.71	2.16	4.29 × 10^5	16 875

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The saturation magnetization (M_s) values of the Cu–Co ferrite NFs decreased with an increase in the Cu content at both temperatures. The reduction in M_s for all the samples can be related to the site preference of the substituted cations in the structure of each sample, as described below.

The magnetic moments of the prepared NFs were estimated using the Néel theory in units of Bohr magneton (μ_B) according to the following equation:

where M_s is the saturation magnetization and M_w is the molecular weight of the Cu_xCo_1−xFe₂O₄ NFs.⁴⁹ As shown in Table 3, the magnetization of the Cu–Co ferrite NFs also decreased with an increase in Cu content in the unit of μ_B.

In simple Co spinel ferrites, where the Co²⁺ ions occupy the octahedral sites and Fe³⁺ ions are distributed equally over the tetrahedral and octahedral sites, the magnetic moment is ∼5μ_B.^46–48 In this case, tuning the cation distribution in the structure affects the magnetic properties of the nanoparticles.⁴³ The calculated magnetic moment of the unsubstituted CoFe₂O₄ NFs in the present series was reduced to ∼3μ_B. Unlike simple Co ferrites, the cations distribution analysis in Table 2 reveals that in Cu_xCo_1−xFe₂O₄ with x values from 0 to 0.4, most of the Co²⁺ ions are in the octahedral B-sites but a small amount is in the tetrahedral A-sites. When non-magnetic ions (Cu²⁺) replace the magnetic ions (Co²⁺ and Fe³⁺) in the octahedral site, the magnetization of the B-sublattice decreases, and thus the net magnetization, which is expressed by M = M_B − M_A, also decreased.^43,50 In the case of the highest Cu concentration sample (x = 0.6), a remarkable reduction in the magnetic moment occurred due to the site occupation rearrangement. In this case, Co²⁺ occupied only the B-sites, while a small amount of Cu²⁺ migrated to the A-sites, which is believed to be responsible for the further decrease in the magnetic moment of this sample. The magnetic ordering in spinel ferrites is subject to the superexchange mechanism between the metal ions in the octahedral and tetrahedral sites. The exchange interactions (A–A, A–B, and B–B) between the electrons of the ions in these two sublattices determine the magnetization of the ferrite structure, where the A–B interaction is the strongest one.^50,51

The squareness ratio (SQR) is another key parameter described in the Stoner–Wohlfarth model, where a value equal or close to 0.5 means that the particles have uniaxial crystal anisotropy and they are non-interacting, resulting in a single domain structure.⁵² The SQR values of the presented Cu–Co ferrite NFs are either below or above this threshold, and thus we conclude that they have a multi-domain structure, which agrees with the nature of the NFs as connected particles.

In the case of coercivity (H_c), the highest values were determined at x = 0 and 0.6 (Table 3). The H_c decreased with an increase in Cu content until x = 0.4, and then increased strongly with higher Cu concentration (x = 0.6). This observation can be explained based on the above-mentioned cation distribution rearrangement, which may result in a change in the Cu²⁺ spin arrangement due to its occupation of both the A and B sites in this sample only.⁵³ Generally, the H_c of magnetic nanoparticles is controlled by several factors such as shape anisotropy, grain size, particle distribution, morphology and magnetocrystalline anisotropy.^54,55

The effective anisotropy constant, K_eff, was calculated using the M_s and H_c values for cubic anisotropy according to the following the expression:

The calculated K_eff values are 10⁴ erg g⁻¹ and 10⁵ erg g⁻¹ orders at 300 K and 10 K, respectively.⁵⁶

The intrinsic coercivities (H_ci) for all NF samples are significantly higher compared to H_c and have the same tendency with an increase in the Cu-substitution ratio (Table 3).

The magnetic parameters including M_s, H_c, and M_r extracted from the M–H loops measured at 10 K are substantially higher than the same parameters calculated from the M–H curve measured at 300 K. As the temperature decreases, the surface spin disorder and thermal fluctuation of the nanoparticles diminish, which increases the various magnetic parameters.⁵⁷

3.4. Cell viability post treatment

The prepared NFs produced an inhibitory action on HCT-116 and HeLa cells, as revealed by the cell viability assay. The cell viability decreased in both groups after 48 h of treatment with the Cu_xCo_1−xFe₂O₄ (x ≤ 0.6) NFs (Fig. 6). The treated HCT-116 cells showed a significant decrease in viability, while that of the treated HeLa cells did not decrease significantly. Also, to examine whether the prepared NFs produced similar effects on normal and healthy cells, HEK-293 cells were treated with the same concentration. The viability of the HEK-293 cells decreased after the treatment, but to a lesser degree compared to HCT-116 and HeLa cells (Fig. 6).

Fig. 6 MTT assay showing the effect of 25 μg per mL Cu_xCo_1−xFe₂O₄ (x ≤ 0.60) NFs on HCT-116 cells, HeLa cells and HEK-293 cells after 48 h of treatment. *p < 0.05.

This finding suggests that these NFs produced better cytotoxic effects on cancerous cells than the healthy cells. The inhibitory concentrations (IC₅₀) of the NFs on HCT-116 cells were 29.50 μg mL⁻¹ (x = 0), 31.72 μg mL⁻¹ (x = 0.2), 33.25 μg mL⁻¹ (x = 0.4), and 36.25 μg mL⁻¹ (x = 0.6). In the case of HeLa cells, the IC₅₀ values were 29.20 μg mL⁻¹ (x = 0.0), x = 0.2 (39.52 μg mL⁻¹), x = 0.4 (41.15 μg mL⁻¹), and x = 0.6 (44.50 μg mL⁻¹). This is the first study to examine the cytotoxic effects of pure nano-sized Cu–Co ferrites on human cancer cells. The cell viability and cytotoxicity of a Co–Cu ferrite/Na₂Ca₂Si₃O₉ (Bioglass) composite was assessed using osteoblast (MG63) cell lines for hyperthermia application.²⁶ Another study showed that Cu and Co MOF NPs showed anticancer activity against the SKOV3, U87MG, and LN229 cell lines.⁵⁸

3.5. Apoptotic cancer cell death

To examine whether cancer cell death is due to apoptosis or programmed cell death, the NF samples were stained with DAPI, which is a nuclear DNA stain.⁵⁹ DAPI staining is used as a marker for apoptosis, where chromatin condensation, chromatin augmentation, and cell shrinkage are the hallmarks of apoptosis. It was found that the NF treatment induced chromatin condensation and nuclear disintegration of the colon cancer cells, as revealed by the DAPI staining (Fig. 7B–E). In contrast, when colon cancer cells were not treated with NFs, no inhibitory action was observed in the cancer cells (Fig. 7A). A decrease in cancer cells may occur due to programmed cell death resulting from oxidative stress (apoptosis) as a reaction to nano-sized Co ferrites.^19,27 Moreover, the inhibitory action of the present NFs on cancer cells was found to increase with an increase in Cu concentration in the sample. The role of Cu is critical in cancer treatment, where in the case of nanomaterials containing Cu, several studies have examined their inhibitory action against cancer cells. Copper oxide nanowires decorated with folic acid (CuO-Nw–FA) showed enhanced cellular uptake in triple negative breast cancer (TNBC) cells.⁶⁰ CuFe₂O₄ hybrid nanoradiosensitizers (CuFe₂O₄@BSA–FA–CUR) produced a cytotoxic effect on tumor cells through an in vitro study, and also showed high cancer suppression efficiency an in vivo experiment, where CuFe₂O₄@BSA–FA–CUR nanoplatforms led to complete tumor ablation in almost all the treated mice.⁶¹ The cytotoxicity of a CuO NP and chitosan composite was examined against human breast cancer MDA-MB-231 and lung cancer A549 cell lines.⁶² The combination of disulfiram and Cu nanoparticles showed significant anti-cancer activity in CT26 colorectal cancer cells in both in vitro and in vivo studies, where significant apoptosis was induced, which caused the elimination of tumors in animals.⁶³ In addition, Cu nanoparticles showed a cytotoxic effect on the SW480 human colorectal cancer cell line.⁶⁴ Thus, the prepared NFs, which contain Co and Cu, were expected to have anti-cancer activity.

Fig. 7 (A–E) Impact of NFs on HeLa cells stained with DAPI, 48 h post-treatment. (A) (Control), (B) (x = 0.0), (C) (x = 0.2), (D) (x = 0.4), and (E) (x = 0.6). NF dose was 25 μg mL⁻¹. Arrows show nuclear condensation, fragmentation, and cell membrane disruption.

4. Conclusion

The electrospinning synthesis of Cu_xCo_1−xFe₂O₄ (0 ≤ x ≤ 0.6) NFs was successfully achieved. The structure information, phase purity, grain size and cation distribution for all prepared NFs were estimated from the PXRD data. The microstructure and chemical composition of the NF samples were investigated using SEM/EDX analysis. The high-resolution TEM images illustrated the nanofiber nature of the studied samples and the agglomeration of the grains within the NFs. The prepared NFs are semiconductors, with their band gap edges narrowing with an increase in Cu content in the sample. The possibility for the optical property engineering of these NFs makes them potential candidates for photocatalysis applications. Furthermore, the magnetic properties such as M_s, H_c and H_ci of these samples were determined by M–H magnetization measurements at low and room temperature. The tendency of these parameters to increase or decrease was explained based on the increase in the concentration of Cu²⁺ throughout the samples and the cation distribution over the A and B sites. At 10 K, both the M_s and H_c values for each sample are higher than that of the same sample at ambient temperature due to the weakening of the thermal fluctuations of magnetic moments.

The bioactivity of the prepared NFs was examined using a cell viability assay, which showed a substantial decline in the cell viability of cancerous cells (HCT-116) compared to normal cells (HEK-293) after treatment with NFs. Although the cancerous cells (HeLa) treated with NFs showed less inhibition action compared to the latter, it was still higher than that in the normal cells (HEK-293). Apoptotic activity was examined by DAPI staining after treatment with NFs, which caused chromatin condensation and nuclear disintegration in colon cancer cells. The results showed that an increase in the Cu dopant ratio enhanced the bioactivity of the Co ferrites, making them a potential candidate for biomedical applications.

Conflicts of interest

There are no conflicts to declare.

Acknowledgements

The study was supported by the Deanship of Scientific Research, Imam Abdulrahman Bin Faisal University, Dammam, Saudi Arabia, and project number IRMC-015-2019.

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Footnote

† Electronic supplementary information (ESI) available. See DOI: https://doi.org/10.1039/d3ra08087k

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