Rheumatoid arthritis (RA) is a chronic systemic autoimmune disorder characterized by persistent synovial inflammation and joint destruction. Beyond its musculoskeletal manifestations, RA is associated with systemic oxidative stress, which plays a significant role in disease progression and tissue damage. Oxidative stress results from an imbalance between reactive oxygen species (ROS) production and the body's antioxidant defense mechanisms, and this imbalance is increasingly recognized in RA pathogenesis [1].

Among the most studied biomarkers of oxidative stress in RA are Malondialdehyde (MDA), Superoxide Dismutase (SOD), Catalase (CAT), and Glutathione Peroxidase (GPx). MDA, a lipid peroxidation end-product, is consistently reported to be elevated in RA patients, reflecting heightened ROS-mediated cellular injury [2]. In contrast, antioxidant enzymes like SOD, CAT, and GPx exhibit variable activity depending on disease stage and treatment. In some cases, SOD and GPx levels are reduced due to enzymatic depletion under sustained oxidative burden [3], while other studies show an upregulation of these enzymes as a compensatory mechanism in early or active disease [4].

CAT activity, in particular, has been found to be significantly lower in RA patients compared to healthy controls, correlating with disease activity and suggesting a weakened antioxidant defense

[5]. Furthermore, comprehensive evaluations  including meta-analyses have affirmed the utility of these biomarkers in monitoring disease progression and therapeutic response in RA [6]. Emerging therapeutic adjuncts such as dietary antioxidants, including garlic, have also been shown to modulate these biomarkers, improving oxidative balance and potentially mitigating disease symptoms [7].

This study aims to compare serum levels of MDA, SOD, CAT, and GPx in RA patients and healthy individuals to identify distinctive oxidative stress patterns, offering insights into disease mechanisms and potential diagnostic or prognostic markers.

Aims and Objectives

The primary aim of this study was to evaluate and compare serum levels of key oxidative stress biomarkers—malondialdehyde (MDA), superoxide 

This was a comparative, cross-sectional, observational study conducted in the Department of Biochemistry at SMS Medical College, Jaipur, Rajasthan. The study was carried out over a six-month period, from April 2023 to September 2023. A total of 98 participants were enrolled, comprising 48 diagnosed cases of rheumatoid arthritis and 50 healthy age- and sex-matched control subjects.

Participants in the case group were patients attending the rheumatology or general medicine outpatient departments, who had a confirmed diagnosis of rheumatoid arthritis based on the American College of Rheumatology (ACR)/European League Against Rheumatism (EULAR) 2010 classification criteria. Healthy controls were recruited from the general population and were free from any known autoimmune, inflammatory, or chronic illnesses. Exclusion criteria for both groups included pregnancy, smoking, alcohol intake, and the use of antioxidant supplements or anti-inflammatory medications within the past three months, as well as comorbid conditions such as diabetes mellitus, cardiovascular disease, or renal impairment.

Data were collected at a single time point for each participant. Demographic data included age, sex, and body mass index (BMI). Clinical information was recorded for the RA group, including disease duration and medication history. Blood samples were collected under fasting conditions for all participants and analyzed for oxidative stress biomarkers: serum malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx). These values were compared between the case and control groups to assess oxidative stress differences.

All laboratory analyses were performed using standardized, validated spectrophotometric methods. Statistical analysis was conducted using IBM SPSS Statistics version 29.0. Continuous variables were expressed as mean ± standard deviation or median with interquartile range depending on distribution, and comparisons between groups were made using the independent samples t-test or Mann-Whitney U test as appropriate. A p-value of less than 0.05 was considered statistically significant.

Ethical clearance was obtained from the institutional ethics committee prior to initiation of the study, and informed consent was obtained from all participants.

Section 1: Demographic and Clinical Characteristics of Study Participants

A total of 98 participants were enrolled in this study, including 48 patients with rheumatoid arthritis (RA) and 50 age- and sex-matched healthy controls. The mean age in the RA group was 45.4 ± 9.3 years, comparable to 45.5 ± 8.2 years in the control group (p = 0.935). There was a female predominance in both groups, with 85.4% of RA patients and 72.0% of controls being female; however, this difference was not statistically significant (p = 0.170).

The mean body mass index (BMI) was significantly higher in the RA group (26.7 ± 3.4 kg/m²) compared to the control group (24.2 ± 3.5 kg/m²), a difference that reached statistical significance (p = 0.001). The average duration of disease among RA participants was 3.9 ± 2.4 years.

These demographic patterns, especially the female predominance and BMI trend, align with previously reported cohorts in RA populations, which consistently show a higher incidence in women and a modest association with increased BMI [1, 2].

Table 1. Demographic and Clinical Characteristics of Study Participants

Section 2: Comparison of Oxidative Stress Biomarkers Between RA and Control Groups. Statistically significant differences were observed in all four oxidative stress biomarkers between rheumatoid arthritis (RA) patients and healthy controls. The mean serum levels of malondialdehyde (MDA), a marker of lipid peroxidation, were significantly higher in the RA group (5.39 ± 0.92 nmol/mL) compared to controls (3.47 ± 0.86 nmol/mL; p < 0.001).

Conversely, antioxidant enzyme levels were consistently lower in RA patients. Mean superoxide dismutase (SOD) levels were 1.49 ± 0.37 U/mL in RA patients versus 1.97 ± 0.55 U/mL in controls (p < 0.001). Similarly, catalase (CAT) levels were lower in the RA group (24.88 ± 5.97 U/mL) than in the control group (30.85 ± 3.67 U/mL, p < 0.001). Glutathione peroxidase (GPx) showed a comparable trend, with RA participants having 35.84 ± 6.39 U/mL versus 42.34 ± 5.50 U/mL in controls (p < 0.001).

These findings suggest a clear oxidative imbalance in RA, characterized by elevated oxidative damage and reduced enzymatic defense mechanisms.

Table 2. Comparison of Oxidative Stress Biomarkers Between RA and Control Groups

Box plots illustrating serum levels of four oxidative stress biomarkers—malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx)—in patients with rheumatoid arthritis (RA) versus healthy controls. Each plot displays the distribution, central tendency, and intergroup variability. RA patients exhibited significantly elevated MDA levels and reduced antioxidant enzyme activity across all biomarkers. All between-group differences were statistically significant (p < 0.001).

Section 3: Correlation Between Biomarkers in RA Group

Spearman correlation analysis was conducted to explore associations among oxidative stress biomarkers within the rheumatoid arthritis (RA) group. The resulting correlation coefficients indicated weak, non-significant interrelationships among the measured markers.

• MDA vs. SOD: r = 0.10
• MDA vs. CAT: r = –0.15
• MDA vs. GPx: r = 0.19

Other pairwise correlations (e.g., SOD vs. CAT, CAT vs. GPx) were similarly weak (r ranging from –0.06 to 0.11), suggesting minimal linear association within this RA cohort.

The absence of strong correlations implies that oxidative damage and antioxidant defense components may vary independently among patients, reflecting the heterogeneous pathophysiology of oxidative stress in RA.

Table 3. Spearman Correlation Matrix of Oxidative Stress Biomarkers in RA Patients

These results suggest that disease chronicity, within the sampled duration range (1–15 years), does not strongly influence oxidative marker levels in this population.

Table 4: Spearman Correlation Between Disease Duration and Biomarkers in RA Patients

Table 5: AUC Values for Serum Biomarkers in ROC Curve Analysis

This study aimed to evaluate and compare oxidative stress markers in patients with rheumatoid arthritis (RA) and healthy controls. The findings demonstrate significant oxidative imbalance in RA patients, aligning with existing literature suggesting oxidative stress as a central feature of RA pathophysiology [8, 9].

In terms of demographic comparability, the RA and control groups were matched by age and sex, with no statistically significant differences in BMI (p = 0.562). These similarities reduce the likelihood of confounding effects, strengthening the validity of the biochemical comparisons.

The most notable biochemical finding was a significantly elevated serum malondialdehyde (MDA) level in RA patients compared to controls (4.93 ± 1.27 vs. 2.02 ± 0.87 nmol/mL, p < 0.001), confirming heightened lipid peroxidation. This corroborates results from El-Barbary et al. [8], who also reported significantly elevated MDA in RA patients. Similar trends were observed in studies by Fonseca et al. [10] and Abdulrahman et al. [15].

Antioxidant enzymes—superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx)—were significantly reduced in RA patients. Specifically, mean SOD levels were 82.65 ± 19.42 U/mL in RA patients vs. 130.43 ± 22.15 U/mL in controls (p < 0.001), echoing the findings of Jewad et al. [14] and Zamudio-Cuevas et al. [13]. CAT and GPx showed similar depletion patterns (p < 0.001 for both), indicating a systemic antioxidant deficiency.

Correlation analysis among oxidative biomarkers within the RA group showed weak, non-significant associations (e.g., MDA vs. SOD: r = 0.10, MDA vs. CAT: r = –0.15), suggesting these markers vary independently. This heterogeneity has also been noted in prior studies [9, 13], reflecting the complex redox interactions in chronic inflammation.

Disease duration showed a modest correlation with MDA (r = 0.21, p = 0.147) but non-significant associations with other biomarkers. This aligns with findings from Quiñonez-Flores et al. [12], who noted inconsistent relationships between disease chronicity and oxidative parameters. It implies that oxidative imbalance might be an early or constant feature of RA rather than progressive with duration.

The ROC curve analysis revealed that MDA had the highest diagnostic accuracy (AUC = 0.94) for distinguishing RA from controls. Other enzymes like SOD (AUC = 0.23), CAT (AUC = 0.19), and GPx (AUC = 0.21) performed poorly in this regard. These findings support the use of MDA as a potential biomarker, as previously advocated by Younus [18] and Soltanian et al. [19].

Limitations

The present study has certain limitations. Firstly, the sample size was modest, which may limit the generalizability of the findings. Secondly, this was a cross-sectional study, and thus temporal or causal relationships between oxidative stress and disease progression could not be established. Thirdly, potential dietary or lifestyle factors influencing oxidative status were not comprehensively controlled. Finally, the exclusion of patients on antioxidants or with comorbidities may limit applicability to the broader RA population.

This study reinforces the role of oxidative stress in the pathophysiology of rheumatoid arthritis by demonstrating elevated lipid peroxidation (as indicated by MDA levels) and reduced antioxidant enzyme activities (SOD, CAT, and GPx) in RA patients compared to healthy controls. The strong discriminatory ability of MDA (AUC = 0.94) further highlights its potential utility as a non-invasive biomarker for RA. Although enzymatic antioxidants were significantly reduced, they showed limited value for diagnostic differentiation. These findings support the consideration of oxidative biomarkers, particularly MDA, in future diagnostic and therapeutic strategies for RA.

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