International Journal of Agriculture and Biosciences 2025 xx(x): xx-xx.

[Link]
This is an open-access article under the CC BY-NC-ND license ([Link]
RESEARCH ARTICLE eISSN: 2306-3599; pISSN: 2305-6622

Investigation of Oxidative Stress and Antioxidative Enzymes in Erythrocytes and Bone

Marrow of Albino Rats Treated with Different Concentrations of Copper Ferrite
Nanoparticles

Roshan Riaz1, Muhammad Asif2, Gulnaz Afzal2, Urva-Til-Wusqa2, Mubeen Talib2, Moeen Afzal2, Shanzab Noor3,
Hafiz Muhammad Nouman1, Arooj Ali4, Konul Ahmadova5, Rashid Iqbal5,6 and Riaz Hussain7,*

1Department of Animal Nutrition and Nutritional Diseases, Faculty of Veterinary Medicine, Kafkas University, 36100 Kars, Türkiye
2Department of Zoology, The Islamia University of Bahawalpur 63100, Pakistan
3Department of Biomedical Engineering (Medicine), Shenzhen University, China
4Institute of Physics, Faculty of Physical & Mathematical Sciences, The Islamia University of Bahawalpur, Institute of Physics,

63100, Pakistan
5Department of Life Sciences, Western Caspian University, Baku, Azerbaijan

6Department of Agronomy, Faculty of Agriculture and Environment, The Islamia University of Bahawalpur 63100, Pakistan
7Department of Pathology, Faculty of Veterinary and Animal Sciences, The Islamia University of Bahawalpur 63100, Pakistan

*Corresponding author: [Link]@[Link]

ABSTRACT Article History

Copper ferrite nanoparticles are widely studied for their biomedical applications; however, their Article # 25-059
potential toxicity to hematopoietic and dental tissues remains unclear. This study evaluates Received: 10-Feb-25
oxidative stress, antioxidative enzymatic contents, and histopathological alterations in albino Revised: 16-Jun-25
rats exposed to copper ferrite nanoparticles. Male Wistar albino rats were divided into four Accepted: 04-Jul-25
groups: Group A (control, 0.0mg/kg), Group B (2.5mg/kg), Group C (5.0mg/kg), and Group D Online First: 11-Jul-25
(7.5mg/kg) received copper ferrite nanoparticles intravenously for 15 days. Oxidative stress was
assessed by measuring thiobarbituric acid-reactive substances (TBARS) and reactive oxygen
species (ROS). At the same time, antioxidant defense was evaluated in terms of the estimation
of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) contents. The results
showed a dose-dependent increase in TBARS and ROS levels, indicating elevated oxidative
stress, along with a significant decline in SOD, POD, and CAT contents, suggesting impaired
antioxidant defense mechanisms. Histopathological analysis of teeth revealed structural
alterations, including enamel erosion, dentin degeneration, and inflammatory changes,
particularly at higher doses. The most severe oxidative and histopathological effects were
observed in Group D (7.5mg/kg), indicating potential toxicity associated with exposure to
copper ferrite nanoparticles. These findings suggest the need for further research on the long-
term effects of copper ferrite nanoparticles on erythrocytes, hematopoietic, and dental tissues,
to ensure their safe biomedical use.

Keywords: Albino Rats, Copper ferrite, Erythrocytes, Bone marrow, Oxidative stress,
Antioxidative enzymes

INTRODUCTION oxide nanoparticles (NPs) belong to a class of nanomaterials

and are synthesized from silver, copper, magnesium, zinc,
Nanotechnology has transformed numerous scientific gold, titanium and alginate (Ghonimi et al., 2022). Among
domains by allowing for the manipulation of materials at the these, copper ferrite (CuFe2O4) NPs have attracted
atomic and molecular levels. Among the diverse Considerable attention due to their remarkable magnetic
nanoparticles, metal oxide nanoparticles have gained a lot catalytic, optical, and structural characteristics, making them
of attention for their distinct physicochemical features and promising candidates for applications in catalysis, magnetic
potential biological uses (Nikolova & Chavali, 2020). Metal storage, and biomedicine (Saikova et al., 2023).

Cite this Article as: Riaz R, Asif M, Afzal G, Ur-Til-Wusqa, Talib M, Afzal M, Noor S, Nouman HM, Ali
A, Ahmadova K, Iqbal R and Hussain R, 2025. Investigation of oxidative stress and antioxidative enzymes
in erythrocytes and bone marrow of albino rats treated with different concentrations of copper ferrite
nanoparticles. International Journal of Agriculture and Biosciences xx(x): xx-xx.
[Link] A Publication of Unique
Scientific Publishers

1
 2 Int J Agri Biosci, 2025, xx(x): xxx-xxx.

Despite the advantageous properties of copper ferrite, teeth of albino rats, aiming to gain a deeper understanding
concerns have emerged regarding its toxicological effects of the potential risks associated with their widespread
on biological systems. Nanoparticles may influence stress applications and exposure.
response mechanisms and interact with proteins in living
organisms. Nanoparticle exposure can cause oxidative MATERIALS & METHODS
stress and disrupt proteins involved in stress response
pathways (CAT, SODs, GPXs, GR, PRDXs, PDI and CAT). It is This research was approved by the Board of Studies,
now widely accepted that manufactured NPs cause Department of Zoology and Advanced Studies, and the
oxidative stress by producing damaging reactive oxygen Advanced Studies and Research Board, the Islamia
species. Nanoparticles' propensity to create ROS is University of Bahawalpur (IUB), Pakistan.
influenced by their physicochemical features, which include
size, shape, structure, and metal contents (Cameron et al., Study Animal
2022). Increased oxidative stress is one of the most Research was conducted in the Animal Rooms at
important processes in nanoparticle-induced toxicity, Baghdad ul Jadeed Campus, Department of Agriculture, IUB.
described as “a shift in the prooxidant or antioxidant Twenty male adult albino rats free of any clinical ailments
balance in favor of the former” (Horie & Tabei, 2021). were used in this study. The rats were obtained from IUB,
Enhanced levels of ROS lead to pathophysiological Baghdad ul Jadeed campus (Pakistan). Each rat was housed
outcomes, including inflammation, DNA damage, and in a metal wire cage with a regular dark/light interval and
fibrosis, which can contribute to the development of various constant temperature. Water and food were accessible
diseases, such as cancer, atherosclerosis, neurological every day. The CuFe2O4 NPs were obtained from the
disorders, autoimmune diseases, and type 2 diabetes (Xuan Institute of Physics, IUB.
et al., 2023). Due to their small size, nanoparticles can pass
through and traverse physiological barriers, traveling via the Experimental Design and Treatment
circulatory system to affect multiple tissues and thereby The rats were divided into four treatment groups: A, B,
impact animal and human health (Ghonimi et al., 2022). C, and D. Each group consisted of five rats. Group A was
Copper ferrite nanoparticles are frequently used in named the control group. All rats were fed commercial
biomedicine, magnetic resonance imaging, magnetic cell chicken feed. The weekly body weight of every male in each
separation, as well as energy storage devices, magnetic group was determined using a digital weight computerized
storage medium and spintronic and electromagnetic weight balance. For 15 days, rats in categories B, C and D
appliances (Saikova et al., 2023). Humans are at high risk of were given daily injections of CuFe2O4 NPs. Rats in group B
exposure to nanoparticles, which can enter the body were administered CuFe2O4 NPs at a dose of 2.5mg/kg BW.
through various pathways, including the digestive, Rats in group C received 5.0mg/kg while rats in group D
respiratory, and skin systems, during the production, use, were administered CuFe2O4 NPs @7.5mg/kg BW. The
and disposal of nanoparticles. Once deposited in specific nanoparticles were administered intraperitoneally daily for
areas of the body, nanoparticles are transported to different 15 days. The rats were keenly observed daily. Sampling was
tissues through the lymphatic or circulatory system. Due to conducted on days 5, 10, and 15 of the experiment.
their stability, nanomaterials persist in the body and
environment for an extended period, with potential health Sample Processing and Biochemical Analysis
repercussions from accelerated exposure to nanoparticles Erythrocytes and bone marrow cells were collected
that are not yet fully defined. However, there is scarce from all experimental rats at days 5, 10, and 15th of the
evidence indicating possible toxic effects (Dobrzyńska et al., research trial. The cells were then washed with phosphate
2014). buffer saline and lysed with distilled water to release their
Earlier studies have demonstrated the harmful effects of contents. The lysates were centrifuged at 3000rpm for ten
copper ferrite nanoparticles in human cell culture, minutes and the resulting supernatants were stored at -
highlighting significant alterations in cellular morphology 20˚C. TBARS and ROS were measured at absorbance of 532
and survival (Ahmad et al., 2016). Additionally, a study on and 505nm according to previous protocol (Akram et al.,
Wistar rats demonstrated that intraperitoneal injections of 2021). Other parameters such as CAT, POD and SOD were
CuFe₂O₄ nanoparticles can disrupt antioxidant activities and measured at absorbance of 240, 470, and 560nm
blood plasma parameters in a gender-specific manner, respectively, using ultraviolet spectrophotometer in
indicating overall toxicity (Riaz et al., 2020). Exposure to erythrocytes and bone marrow of rats following the
metal oxide NPs, including silver and copper nanomaterials, previous protocol (Ghazanfar et al., 2018).
has been shown to adversely affect survival, body weight,
size and structure in aquatic species, signifying potential Histopathology Analysis
environmental and health hazards (Ostaszewska et al., For histopathological analysis, at day 15, the different
2016). Erythrocytes are highly susceptible to nanoparticles, teeth were carefully extracted from each rat. The teeth were
which can induce oxidative stress, thereby impairing their fixed in 10% formalin. Following decalcification, the teeth
function and lifespan. This study investigates the oxidative were then embedded in paraffin and thin blocks and
stress induced by copper ferrite nanoparticles, as well as the sections were cut using a microtome and stained using
antioxidant mechanism, particularly in erythrocytes and Hematoxylin and Eosin. These stained sections were
bone marrow, and the histopathological changes in the observed for microscopic changes (Nursanti et al., 2017).
 3 Int J Agri Biosci, 2025, xx(x): xxx-xxx.

Statistical Analysis significantly from the control group. Groups B, C and D

The collected data were statistically evaluated by using had higher ROS and TBARS levels than the untreated
the ANOVA test with the IBM statistical software package group. Groups B, C and D showed significantly reduced
(SPSS). Tukey’s test was employed to compare the means of CAT level in rats administered higher doses of
oxidative and antioxidant factors between nanoparticles. nanoparticles as compared to the control group. POD
P<0.05 was used as the level of significance. levels were significantly reduced in groups C and D relative
to the control group. The lowest POD level was recorded
RESULTS in group D, which received the highest dose of CuFe₂O₄
NPs. The findings indicate a statistically substantial
Behavioral and Physiological Effects difference in SOD level compared to the control group.
Rats given copper ferrite nanoparticles (CuFe₂O₄ NPs) SOD level was notably lower in groups C and D relative to
during the experiment showed physiological changes the control group (Table 3).
including watery feces and signs of depression as well as
Table 3: Oxidative and antioxidant profile in bone marrow of albino rats
behavioral changes like lethargy. On the other hand, the
treated with different doses of copper ferrite nanoparticles
untreated control rats remained active throughout the trial. Parameters/ Days Treatments
The administration of copper ferrite nanoparticles A (0.0) B (2.5mg/kg) C (5.0mg/kg) D (7.5mg/kg)
indicated no significant changes in body mass of each Bone Marrow
Reactive oxygen species (ROS) contents (optical density)
group. Rats in groups B, C and D did not exhibit progressive
5 0.34±0.02 0.37±0.07 0.38±0.05 0.41±0.09
increase/decrease in body weight corresponding to the 10 0.34±0.04 0.38±0.09 0.39±0.04 0.51±0.05
escalating dosages of copper ferrite nanoparticles (Table 1). 15 0.33±0.03 0.41±0.06 0.48±0.02 0.55±0.05
Thiobarbituric acid reactive substances (TBARS) content (nmol/TBARS
Table 1: Body weight (g) of albino rats treated with different doses of copper formed/mg protein/min)
ferrite nanoparticles 5 0.33±0.07 0.36±0.07 0.37±0.07 0.42±0.05
10 0.31±0.07 0.37±0.08 0.41±0.09 0.48±.0.04*
Experimental Days A (0.0) B (2.5mg/kg) C (5.0mg/kg) D (7.5mg/kg)
15 0.32±0.05 0.38±0.10 0.52±0.03* 0.62±0.02*
5 132.3±1.86 132.4±1.81 133.2±2.38 129.9±1.52
Antioxidant enzymes
10 133.2±2.61 131.3±3.60 131.1±1.24 128.4±1.40
Superoxide dismutase SOD (units/mg protein)
15 134.5±2.22 130.2±2.91 129.2±1.75 127.5±1.47
5 0.31±0.02 0.28±0.03 0.27±0.03 0.26±0.02
10 0.33±0.03 0.27±0.02 0.23±0.01* 0.21±0.02*
Oxidative Stress and Antioxidant Response 15 0.31±0.03 0.26±0.02 0.21±0.01* 0.19±0.03*
The levels of TBARS and ROS in erythrocytes were Catalase CAT (units/min)
5 0.43±0.04 0.42±0.09 0.39±0.08 0.38±0.07
markedly different compared to the control group.
10 0.44±0.02 0.41±0.08 0.38±0.07 0.34±0.06*
Specifically, groups B, C and D exhibited notably higher 15 0.45±0.02 0.39±0.07 0.33±0.06* 0.27±0.05*
TBARS and ROS levels than group A. The results also Peroxidase POD (units/min)
demonstrated a statistically significant variation in contents 5 0.37±0.02 0.36±0.03 0.34±0.01 0.32±0.09
10 0.36±0.03 0.34±0.02 0.32±0.04 0.27±0.08*
of POD, CAT and SOD in erythrocytes compared to control 15 0.36±0.04 0.33±0.08 0.26±0.07* 0.19±0.07*
group. Groups B, C and D displayed significantly lower CAT, Values (mean+SD) bearing asterisk within a row differ significantly (P<0.05).
POD and SOD contents than the control group (Table 2).
Histopathology Evaluation
Table 2: Oxidative and antioxidant profile in erythrocytes of albino rats Microscopic analysis of dental structures revealed
treated with different doses of copper ferrite nanoparticles
Parameters/ Groups
normal histological structures of the teeth of rats of group
Days A (0.0) B (2.5mg/kg) C (5.0mg/kg) D (7.5mg/kg) A. However, rats of groups B and C treated with 2.5 and
Erythrocytes 5.0mg/kg BW nanoparticles displayed mild and moderate
Reactive oxygen species (ROS) contents microscopic changes in their teeth at days 5, 10, and 15 th
5 0.58± 0.03 0.59±0.02 0.60±0.03 0.61±0.04
10 0.57±0.05 0.62±0.07 0.63±0.08 0.74±0.05*
of trial in groups B and C. These changes were pulp
15 0.58±0.04 0.63±0.04 0.69±0.06* 0.83±0.03* calcifications, inflammatory responses, changes in dentin
Thiobarbituric acid reactive substances (TBARS) content (nmol/TBARS bridge thickness, resorption of dentin, reduced
formed/mg protein/min) vascularization in the pulp tissue, thickness of inner
5 0.69±0.05 0.71±.0.04 0.73±0.03 0.74±1.42
10 0.67±0.04 0.72±0.02 0.75±0.03 0.86±0.08* enamel epithelial cells, increased cellularity of fibrocytes,
15 0.68±.0.07 0.73±0.06 0.82±0.08* 0.94±0.07* thickened periodontal tissue, periodontal tissue, necrosis
Antioxidant enzymes of odontoblasts, elevated percentage of osteoclasts and
Superoxide dismutase SOD (units/mg protein)
delayed growth of the periodontal tissues. Conversely,
5 0.34±0.06 0.32±0.08 0.31±0.04 0.29±0.07
10 0.35±0.05 0.31±0.03 0.28±0.04 0.27±0.02* mild to moderate effects were observed in the teeth of
15 0.35±0.02 0.29±0.05 0.23±0.05* 0.21±0.02* group B rats treated with 2.5mg/kg CuFe2O4 NPs. These
Catalase CAT (units/min) alterations were severe in rats of group D at day 15 of trial
5 0.20±0.07 0.21±0.06 0.19±0.04 0.17±0.06
10 0.21±0.03 0.20±0.07 0.19±0.02 0.15±0.01*
(Table 4 and Fig. 1).
15 0.21±0.05 0.19±0.05 0.16±0.03* 0.14±0.04*
Peroxidase POD (units/min) DISCUSSION
5 0.57±0.03 0.55±0.05 0.53±0.04 0.52±0.04
10 0.54±0.05 0.52±0.07 0.51±0.03 0.49±0.02
15 0.56±0.04 0.51±0.03 0.41±0.06* 0.36±0.07* With the advancement of science and technology, as
Values (mean+SD) bearing asterisk within a row differ significantly (P<0.05). well as the rapid growth of nanotechnology, NPs are
becoming increasingly prevalent in various fields such as
TBARS and ROS levels in bone marrow differed medical, agricultural environment, energy production and
 4 Int J Agri Biosci, 2025, xx(x): xxx-xxx.

materials research. Different features including synthesis, switching under various situations. Research on the
characterization, size, nanocomposites, nanomaterials and potential hazards of spinel ferrite nanoparticles has grown
route of exposure play important role in the pathogenesis considerably in recent years. Given their wide range of
of induction of toxic effects in target and non-target animals applications, it is crucial to evaluate their toxicity on
different tissues. The extensive utilization of CuFe₂O₄
Table 4: Severity of histopathological ailments in teeth of albino rats treated nanoparticles in biological fields has gained significant
with different doses of copper ferrite nanoparticles
scientific interest; however, their associated toxicity and
Histopathological Lesions TREATMENTS
A(0.0 B(2.5 C(5.0 D(7.5 environmental risks must also be carefully considered
mg/kg) mg/kg) mg/kg) mg/kg) (Srikanth & Nutalapati, 2022).
Inflammatory reactions - ++ ++ ++++ Prolonged contact to nanoparticles presents potential
Pulp calcifications - ++ ++ ++++
Increase in cellularity of fibrocytes - ++ +++ +++
risks to animal life and human health, requiring though
Increased resorption of dentin - ++ ++ +++ assessment and examination of environmental pollutants
Hyaline necrosis - ++ +++ ++++ to minimize their adverse impacts (Ali et al., 2024).
Thickness of periodontal tissue - ++ +++ ++++
Numerous studies have highlighted concerns regarding
Necrosis of odontoblasts - ++ +++ ++++
Disrupted fibers - ++ ++++ +++ the deleterious effects of manufactured nanomaterials in
Decrease vascularization of pulp tissue - ++ +++ ++++ living species primarily focusing on exposure levels (Anwar
Decrease of the mineral contents - ++ +++ ++++ et al., 2023). A key issue is the induction of oxidative stress
Increased dentin bridge thickness - + +++ ++++
Elevated percentage of osteoclasts - ++ +++ ++++
(Cao et al., 2020) in different tissues when exposed to NPs
which are utilized in (Pandey & Saha, 2023), surface
modification (Oliva et al., 2023), lubrication, stabilization,
cellular delivery (Dang et al., 2014; Hasan et al., 2013) and
energy harvesting (Samy et al., 2022). Due to their nanoscale
dimensions and elevated surface area relative to volume,
nanoparticles can interact with biological molecules
(Hussain et al., 2023). They also readily penetrate nuclear
and cell membranes, causing indirect damage such as
oxidative stress and inflammatory responses (Hasan et al.,
2021; Javed et al., 2023; Zafar et al., 2020).
Previously, potential concerns linked with the
applications of nanomaterials, including genotoxicity
mechanisms and health risks, have gained significant
attention (Ghouri et al., 2023). These nanoparticles can
generate reactive oxygen species either through direct and
indirect pathways, leading to genotoxicity, cellular damage,
and cell death (Huang et al., 2022; Huang et al., 2023).
Numerous studies examining various nanomaterials such as
copper iron oxide, calcium nanoparticles, copper
nanoparticles, nickel-iron oxide, and zinc-iron oxide have
reported harmful effects on multiple species, including
human cells, primarily through the induction of oxidative
stress. With nanotechnology rapidly advancing across
medicine, industry and nutrition, understanding and
mitigating these effects has become increasingly important.
A previous study also reported an increase in ROS
production as well as DNA damage on exposure to the ZnO
nanomaterials (Iqbal et al., 2024). ROS levels in cells range
from low to high, leading to a variety of effects, including
Fig. 1: Photomicrograph of teeth of albino rats of group C (a) and D (b)
treated with higher doses of nanoparticles at day 15 indicating various apoptosis, autophagy, and necrosis (Younas et al., 2024).
histopathological lesions. Dentin (D), increased osteoclast and resorption Different types of nanoparticles, insecticides and herbicides
lacunae (red arrows), increased fibrocytes (*) disrupted fibers (DF), are known to trigger oxidative stress by generating
inflammatory material (black arrow), mononuclear cell infiltration (red and
black arrow heads) increased thickness of peridentin (PD), necrosis of
intracellular reactive oxygen species (Horie & Tabei, 2021;
odontoblasts (blue arrow) and hyaline necrosis (HN). H & E stain; 400X. Kanwal et al., 2024; Shafqat et al., 2024)
The considerable rise in oxidative damage indicators
from the macroscopic to the microscopic level. Because and diminished levels of several antioxidant proteins in
of this shift in properties, NPs and nanotechnology are bone marrow and erythrocytes of CuFe2O4 nanoparticles
incredibly significant for a variety of applications including exposed rats in this study might be attributed to the
health management (Xuan et al., 2023). Copper ferrite activation of an immune response (Zhuo et al., 2024). Earlier,
nanoparticles (CuFe2O4NPs) are the most significant and different investigations (Srisuvetha et al., 2020; Zhuo et al.,
important ferrites that exhibit point transitions, changes in 2024) demonstrated that interaction with nanoparticles
semiconductor nature, tetragonality variation and electrical stimulates NLRP3 complexes resulting in cellular
 5 Int J Agri Biosci, 2025, xx(x): xxx-xxx.

impairment and the activation of inflammatory processes in nanoparticles induce oxidative stress and alter the
tissues producing oxidative stress. antioxidant defense system in the erythrocytes and bone
Therefore, oxidative stress, which elevates the marrow of albino rats in a dose-dependent manner. The
production of reactive species and causes the death of significant increase in TBARS and ROS levels, coupled with
numerous cellular organelles, may be the cause of the the depletion of antioxidative enzymes (SOD, POD and
reduced levels of diverse antioxidant biomarkers in rats' CAT), suggests that copper ferrite nanoparticles disrupt
teeth. According to several research in previously published redox homeostasis, leading to oxidative damage. The
literature, oxidative stress causes damage to various cells highest dose (7.5mg/kg) exhibited the most pronounced
and can interfere with regular physiological processes, toxic effects, indicating potential risks associated with
resulting in tissue necrosis through programmed cell death prolonged or high-dose exposure. These findings
(Ali et al., 2024; Samim et al., 2023). highlight the importance of evaluating the
There have been several histological abnormalities biocompatibility and toxicity of copper ferrite
found in rats' teeth as a consequence of oxidative stress nanoparticles before their biomedical applications. Further
induction (Yaman et al., 2018). CuFe2O4 NPs may cause in-depth studies, including long-term exposure
histological and oxidative illnesses in the current assessments and molecular mechanisms of toxicity, are
investigation due to excessive free radical release, essential to establish safe dosage limits and explore
antioxidant depletion and activation of signaling cascades. possible protective strategies against nanoparticle-
Free radicals have been shown to engage immediately with induced oxidative stress.
many micro and macromolecular structures, such as lipids,
proteins and DNA present in different types of cells, DECLARATIONS
resulting in oxidative stress-associated damages. The
significantly elevated levels of oxidative stress indices Funding: The authors express their sincere gratitude to the
(TBARS and ROS) in rat erythrocytes and bone marrow in Islamia University of Bahawalpur, Pakistan, for providing
this study show that CuFe2O4 NPs caused pathological financial and infrastructural support.
alterations in cell membrane integrity, resulting in reduced
antioxidant levels. Conflict of Interest: All authors of the manuscript declare
Furthermore, it has been reported that oxidative that they have no financial or personal interests.
damage caused by nanoparticles lowers the levels of
enzymatic antioxidants, a widely recognized secondary Data Availability: All the data is included in this manuscript
defensive mechanism (Ghaffar et al., 2021; Sanati et al., and can be obtained from the corresponding author on
2022; Wang et al., 2022). CAT and SOD neutralize reasonable request.
superoxide free radicals and detoxify H₂O₂, whereas POD
scavenges lipid hydroperoxides (Hussain et al., 2018; Kumar Ethics Statement: This study was approved by the Board of
et al., 2023). Therefore, the reduction of these proteins Studies of the relevant department and conducted
produces oxidative impairments in various organs. following the guidelines established by the Bioethics
Reactive oxygen species (oxygen-based free radicals) Committee of Islamia University of Bahawalpur, Pakistan.
are highly reactive and can cause damage to various
biological structures, including DNA. Bone marrow and Author’s Contribution: Muhammad Asif: Execution,
erythrocytes are vital parts of the blood-forming system, Investigation, Methodology, manuscript preparation.
and they are particularly vulnerable to reactive damage. Gulnaz Afzal and Riaz Hussain: Supervision and data
Investigating the harmful effects of CuFe₂O₄ nanoparticles analysis. Roshan Riaz and Hafiz Muhammad Nouman:
(NPs) is crucial to identifying possible hazards associated Conceptualization, involved in manuscript writing and
with their applications (Samy et al., 2022). Extensive formal analysis: Moeen Afzal, Ur-Til-Wusqa, Mubeen Talib
research has explored the cytotoxic effects of these NPs and Shanzab Noor: Writing of initial manuscript, data
using various animal models (Chong et al., 2021; Han et al., collection and Investigation: Arooj Ali: Data Curation,
2016). While some investigations have demonstrated that Methodology, Validation. Rashid Iqbal and Konul
these NPs exhibit dose-dependent cytotoxicity, others Ahmadova: Formal Analysis and Resources. Riaz Hussain:
have reported minimal to no adverse effects on cells (Chen Conceptualization, Writing – R.
et al., 2019; Hanley et al., 2009; Namvar et al., 2015). Our
results align with earlier research that has reported Generative AI Statement: The authors declare that no Gen
increased levels of oxidative stress indicators (ROS and AI/DeepSeek was used in the writing/creation of this
TBARS) and reduced antioxidant enzymatic antioxidants manuscript.
activity in tissues such as the bone marrow and teeth.
Histopathological examination of the treated rats' muscle Publisher’s Note: All claims stated in this article are
(gum) tissues revealed atrophied cell, muscular fiber exclusively those of the authors and do not necessarily
degeneration and the presence of inflammatory represent those of their affiliated organizations or those of
components (Mahmood et al., 2024). the publisher, the editors, and the reviewers. Any product
that may be evaluated/assessed in this article or claimed by
Conclusion its manufacturer is not guaranteed or endorsed by the
This study demonstrates that copper ferrite publisher/editors.
 6 Int J Agri Biosci, 2025, xx(x): xxx-xxx.

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