Mitophagy and Mitochondrial DNA Dynamics During Ramadan Dry Fasting vs 16:8 Time-Restricted Feeding

This is a prospective, single-centre, parallel-group, longitudinal observational cohort study conducted across Ramadan 2027. The study aims to evaluate the distinct molecular and metabolic trajectories of intermittent dry fasting compared to standard fluid-permitted time-restricted feeding (TRF).

Participants will be allocated to one of two parallel groups based on their self-selected, established fasting practices:

• Group A (Ramadan Dry Fasting): Observing complete fluid and nutrient deprivation from dawn (Fajr) to sunset (Maghrib) for 30 consecutive days.
• Group B (16:8 TRF): Practicing a 16-hour daily fasting window with water/fluids permitted, observed concurrently during the same calendar period.

Participants in both groups will be pairwise-matched based on age and BMI at screening. Blood samples will be collected from all participants at five distinct milestones:

• T0: Pre-Ramadan baseline (Day -3)
• T1: Acute induction phase (Day 7)
• T2: Mitochondrial turnover phase (Day 14)
• T3: Saturation plateau / primary endpoint (Day 30)
• T4: Post-Ramadan recovery phase (Day 44)

Peripheral blood mononuclear cells (PBMCs) and serum will be isolated from the collected blood. Laboratory analyses will measure mitochondrial DNA (mtDNA) copy number changes via qPCR, alongside a quantitative gene expression panel evaluating key regulators of mitophagy (such as PINK1 and Parkin), mitochondrial biogenesis, and general autophagy. Secondary evaluations include routine serum metabolic indicators, inflammatory markers, and dietary monitoring to evaluate the physiological pathways predicted by computational systems pharmacology.

Inclusion criteria

• Common Criteria (Both Arms):
• Age 18-50 years
• BMI 18.5-30.0 kg/m²[cite: 1]
• Non-smoker for at least 12 months[cite: 1]
• No chronic disease requiring regular prescription medication[cite: 1]
• Stable sleep-wake schedule (self-reported sleep midpoint deviation less than 90 minutes)[cite: 1]
• Willing and able to attend all five study visits at the specified times[cite: 1]

Arm A Specific (Ramadan Dry Fasting):

• Documented religious commitment and intention to complete all 30 days of Ramadan 2027 fasting[cite: 1]

Arm B Specific (16:8 TRF Control):

• Confirmed practice of 16:8 TRF (fasting window 16 hours or more per day) throughout the study period[cite: 1]
• Not observing Ramadan dry fasting during the study period[cite: 1]

Exclusion criteria

• Established type 1 or type 2 diabetes (HbA1c 6.5% or higher or pharmacological antidiabetic therapy)[cite: 1]
• Severe gastrointestinal, cardiovascular, hepatic, or renal disease (eGFR less than 60 mL/min/1.73m²)[cite: 1]
• Active cancer or chemotherapy within five years[cite: 1]
• Use of metformin, statins, or exogenous antioxidant supplements (vitamins C, E, NAC, CoQ10) within 30 days of screening[cite: 1]
• Pregnancy, lactation, or planned conception during the study period[cite: 1]
• Participation in another clinical study within three months[cite: 1]
• Inability to provide written informed consent in Arabic or English[cite: 1]