European Journal of Cardiovascular Medicine

Sleep is a fundamental physiological process essential for various restorative functions in the human body, including immune regulation, hormonal balance, cognitive processing, and metabolic stability (1). Disruption in sleep duration or quality—commonly referred to as irregular sleep patterns—has been increasingly recognized as a public health concern due to its adverse impact on multiple biological systems (2). One such system significantly influenced by sleep is the oxidative stress defense mechanism, which relies on the activity of key antioxidant enzymes to neutralize reactive oxygen species (ROS) generated during normal cellular metabolism (3).

Antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) serve as the first line of defense against oxidative damage. SOD catalyzes the dismutation of superoxide radicals into hydrogen peroxide, which is subsequently broken down by CAT and GPx into water and oxygen, thereby minimizing cellular injury (4). An imbalance between ROS production and antioxidant defenses leads to oxidative stress, which has been implicated in the pathogenesis of various chronic disorders, including cardiovascular disease, diabetes mellitus, and neurodegenerative conditions (5,6).

Recent studies have reported that sleep deprivation or irregular sleep schedules are associated with elevated oxidative stress markers and decreased antioxidant activity (7,8). This reduction in enzymatic antioxidant capacity may result from altered circadian regulation, systemic inflammation, or metabolic disturbances commonly observed in individuals with poor sleep hygiene (9). Despite growing evidence, limited research has specifically examined the comparative status of antioxidant enzyme levels in individuals with regular versus irregular sleep patterns in the general population.

Hence, this study aims to estimate and compare the levels of key antioxidant enzymes—SOD, CAT, and GPx—in individuals with regular and irregular sleep habits. Understanding the relationship between sleep patterns and oxidative stress markers may contribute to strategies aimed at preventing oxidative damage and its associated health risks.

Study Design and Participants
A cross-sectional analytical study was conducted over a period of three months among healthy adult volunteers aged between 18 and 40 years. A total of 60 participants were recruited and equally divided into two groups based on their self-reported sleep patterns:

• Group A (Regular Sleep Group): Individuals reporting consistent sleep duration of 7–8 hours per night for at least 5 days a week over the past 3 months.
• Group B (Irregular Sleep Group): Individuals with inconsistent sleep patterns, including sleep duration less than 6 hours, variable bedtimes, or frequent night-time awakenings for at least 5 days a week over the same period.

Participants with chronic illnesses, those on antioxidant supplements or medications affecting oxidative stress, smokers, alcohol users, and shift workers were excluded to eliminate confounding variables.

Data Collection
Each participant completed a validated questionnaire to assess sleep habits, lifestyle factors, and general health status. Anthropometric data including age, gender, BMI, and vital signs were recorded.

Sample Collection and Biochemical Analysis
Fasting venous blood samples (5 mL) were collected in plain vacutainers between 7:00 AM and 9:00 AM to minimize circadian influence. Serum was separated by centrifugation at 3000 rpm for 10 minutes and stored at -20°C until analysis.

The following antioxidant enzymes were quantified using spectrophotometric methods:

• Superoxide Dismutase (SOD): Measured based on the inhibition of pyrogallol autoxidation.
• Catalase (CAT): Estimated by the breakdown rate of hydrogen peroxide (H₂O₂).
• Glutathione Peroxidase (GPx): Determined using the oxidation of glutathione in the presence of cumene hydroperoxide.

All enzyme activity was expressed in standard units (U/mL or kU/L), and each sample was analyzed in duplicate for accuracy.

Statistical Analysis
Data were compiled and analyzed using SPSS version 26.0. Mean and standard deviation (SD) were calculated for each parameter. Independent sample t-tests were applied to compare antioxidant enzyme levels between the two groups. A p-value of less than 0.05 was considered statistically significant.

A total of 60 participants were included in the study, with 30 individuals in each group. Group A (regular sleep pattern) had a mean age of 26.4 ± 3.9 years, while Group B (irregular sleep pattern) had a mean age of 27.1 ± 4.2 years. The gender distribution and BMI were comparable between the groups, with no statistically significant differences (p > 0.05).

The levels of antioxidant enzymes—superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx)—were significantly higher in participants with regular sleep compared to those with irregular sleep patterns. Mean SOD activity was 4.82 ± 0.62 U/mL in Group A versus 3.09 ± 0.54 U/mL in Group B (p < 0.01). Catalase levels were 32.47 ± 4.28 kU/L in Group A and 21.63 ± 3.84 kU/L in Group B (p < 0.01). GPx activity showed a similar trend, with values of 56.91 ± 6.51 U/L in Group A and 39.23 ± 5.74 U/L in Group B (p < 0.01), as shown in Table 1.

Table 1: Comparison of Antioxidant Enzyme Levels Between Group A (Regular Sleep) and Group B (Irregular Sleep)

As indicated in Table 1, the differences in antioxidant enzyme activities between the two groups were statistically significant across all measured parameters. Participants maintaining regular sleep schedules demonstrated a more robust antioxidant profile, suggesting a beneficial effect of consistent sleep on oxidative stress regulation.

The present study assessed and compared the levels of key antioxidant enzymes—superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx)—in individuals with regular and irregular sleep patterns. The results clearly demonstrate that participants with regular sleep exhibited significantly higher antioxidant enzyme levels compared to those with irregular sleep schedules. These findings suggest a strong link between sleep regularity and the maintenance of oxidative balance.

Oxidative stress arises when there is an imbalance between the production of reactive oxygen species (ROS) and the body's antioxidant defense mechanisms (1). Sleep is known to play a vital role in modulating the redox status of the body. Regular and adequate sleep supports metabolic homeostasis and enhances the activity of endogenous antioxidant systems (2). Our results are consistent with earlier studies that demonstrated increased oxidative stress markers and decreased antioxidant enzyme levels in individuals subjected to sleep deprivation or irregular sleep cycles (3,4).

SOD is the primary antioxidant enzyme that catalyzes the dismutation of superoxide radicals into hydrogen peroxide, which is then broken down by CAT and GPx (5). In this study, significantly higher SOD levels in the regular sleep group align with findings by Everson et al., who reported suppressed antioxidant activity following chronic sleep restriction in rodents (6). Similarly, decreased CAT and GPx levels observed in the irregular sleep group correspond with reports indicating the impairment of these enzymes in individuals experiencing circadian rhythm disturbances and sleep loss (7,8).

Chronic sleep disturbances have also been associated with systemic inflammation and metabolic dysregulation, both of which can negatively impact antioxidant defenses (9). Proinflammatory cytokines such as interleukin-6 and tumor necrosis factor-alpha, which are elevated during sleep deprivation, may contribute to increased oxidative damage by reducing antioxidant capacity (10). This systemic inflammatory response may partially explain the diminished antioxidant enzyme levels observed in our irregular sleep group.

Furthermore, circadian rhythms, which regulate the timing of sleep, also influence the expression of genes responsible for antioxidant enzyme synthesis (11). Disruption of the circadian system, as seen in irregular sleepers, may lead to downregulation of these genes and consequently lower enzyme activity (12). Experimental studies have demonstrated that clock gene mutations result in reduced SOD and GPx activity, further supporting the role of circadian regulation in oxidative defense (13).

Additionally, poor sleep hygiene has been linked to increased sympathetic nervous system activity, which elevates metabolic rate and ROS production, thereby overwhelming the antioxidant system (14). Behavioral factors such as increased caffeine intake, screen exposure at night, and erratic lifestyle habits among irregular sleepers may further exacerbate oxidative imbalance (15).

The findings of this study highlight the critical importance of maintaining regular sleep patterns in promoting antioxidant enzyme activity and protecting against oxidative stress. Encouraging consistent sleep hygiene could be a simple yet effective strategy to improve overall health and reduce the risk of oxidative-stress-related diseases.

However, this study has some limitations. The cross-sectional design limits the establishment of a causal relationship between sleep patterns and antioxidant enzyme levels. Self-reported sleep data may also be subject to recall bias. Future studies incorporating actigraphy-based sleep tracking and longitudinal follow-ups are recommended for better validation.

This study demonstrates a significant association between regular sleep patterns and enhanced antioxidant enzyme activity. Individuals with consistent sleep exhibited higher levels of superoxide dismutase, catalase, and glutathione peroxidase compared to those with irregular sleep habits. These findings emphasize the potential role of sleep regularity in mitigating oxidative stress and maintaining cellular homeostasis. Promoting healthy sleep hygiene may serve as a preventive strategy against oxidative-stress-related health disorders.

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