Altitude-Associated Variation in Phenolic Composition and Oxidative Stress–Related Antioxidant Activity in Rhododendron arboreum Leaves

Rhododendron arboreum leaf samples from a high-altitude site (2709 m; Chopta) and mid-altitude site (1700 m; Barlowganj) in Uttarakhand, India, were compared using quantitative phytochemical analysis, targeted HPLC profiling, in vitro antioxidant assays, biomolecular protection assays, and enzymatic measurements. Chopta extracts showed significantly higher antioxidant activity (P < 0.05), with elevated total phenolics (327 mg GAE/g), flavonoids (32 mg QE/g), and proteins (21 mg/g). High-altitude leaves accumulated ~40% more total phenolics, ~50% more flavonoids, and 2–3-fold higher concentrations of gallic acid, ellagic acid, rutin, and quercetin, as determined by HPLC. These differences correlated with enhanced antioxidant activities, including lower IC50 values in DPPH, ABTS, and metal-chelation assays (Chopta: 25–27 µg/mL vs. Barlowganj: 29–31 µg/mL), indicating superior radical-scavenging and Fe2+-chelating efficacy. Functional assays confirmed superior Chopta inhibition of lipid peroxidation (~50% at 8 h) and protein oxidation (~80% at 1 mg/mL). Key antioxidant enzymes (SOD, CAT, APX) were also elevated in high-altitude samples, evidencing coordinated enzymatic upregulation. Unlike prior studies focused on total antioxidant capacity or floral tissues, this work integrates global biochemical parameters, targeted phenylpropanoid quantification, oxidative-damage assays, and enzymatic profiling of leaves. Overall, altitude-linked secondary metabolite enrichment and strengthened antioxidant enzymes underpin R. arboreum leaf adaptation to high-elevation oxidative stress.