Breathing and ROS. Breathing oxygen inevitably leads to formation of reactive oxygen species (ROS) in the body which play important roles in cellular signaling processes. The generation of small amounts of ROS and free radicals is a normal side effect of aerobic metabolism and necessary for normal functioning of the human body.1 Why do ROS then typically carry such a negative connotation? When the body’s intricate antioxidant defense mechanisms malfunction, or break down, the delicate balance between oxidation and antioxidation is lost. The circulating levels of ROS spiral out of control causing disruption in redox signaling and control, and progressive damage to macromolecules, cells and tissues.
Does an illness set in first, or do the free radicals trigger it? Scientist largely agree that free radicals are the by-products of disease processes, although, in certain pathological conditions, a causative link between free radicals and disease has also been suggested.2 Irrespective of what comes first, it is now widely recognized that oxidative stress is an important factor associated with many acute and chronic diseases, including: cancer, diabetes, cardiovascular, and neurodegenerative diseases. Several decades of research focused on studying the connection between oxidative stress and disease have resulted in identification of important oxidative stress biomarkers – the products of oxidation of biological molecules: lipids, proteins and DNA.
Lipid peroxidation. Peroxidation of membrane lipids leads to loss of membrane fluidity and elasticity, impaired cellular functioning, and even cell rupture. The various direct products of lipid peroxidation, such as malondialdehyde (MDA), isoprostanes, and 4-hydroxynonenal are considered among the most important biomarkers of oxidative stress in tissues. Malondialdehyde is a reactive carbonyl compound and is both mutagenic and carcinogenic. It reacts with DNA to form DNA adducts that are believed to contribute significantly to cancers linked to lifestyle and dietary factors.3 Isoprostanes are prostaglandin-like substances produced by free radical-induced peroxidation of arachidonic acid, which exert potent biological effects. Increased levels of isoprostanes in biological fluids and tissue specimens have important clinical implications. The increase in F2-isoprostane levels has been identified as an early event in asthma,4 Alzheimer’s disease,5 scleroderma, 6 and hepatic cirrhosis.7 The non-specific Thiobarbituric Acid-Reactive Substances (TBARS) measurement is a well-established assay used to screen and monitor lipid peroxidation in the presence of malondialdehyde. Over the years, this assay has been adapted for use on a variety of samples including human and animal tissues, natural extracts and foods.
Protein damage. Protein oxidation can cause fragmentation at amino acid residues, formation of protein-protein cross-linkages, and oxidation of the protein backbone which ultimately leads to loss of function. Damaged proteins affect intracellular pathways and are contributing factors to different disorders and diseases. If the proteolytic mechanisms responsible for protein degradation do not function properly, altered proteins accumulate in the cell and may contribute to the development of pathological conditions. Several in vitro assays have been developed to specifically detect biomarkers of oxidative protein damage. Nitrotyrosine is a product of ROS-mediated tyrosine nitration and a biomarker of inflammation and NO production which can be detected using LCMSMS and ELISA methods. Protein carbonyl (CO) groups are produced on protein side chains during oxidation. High levels of protein CO groups have been observed in rheumatoid arthritis, Alzheimer’s disease, diabetes, sepsis and chronic renal failure.8
DNA damage. Oxidative damage to DNA causes alterations in DNA bases. If left unrepaired, the modifications of DNA bases in turn lead to genetic defects. Since guanine is especially susceptible to oxidation, 8-hydroxy-deoxyguanosine has been traditionally utilized as a biomarker of oxidative DNA damage.9
Clinical implications. Oxidation products of biological macromolecules may be quantified in biological fluids (plasma, serum, urine) using non-invasive, high-end analytical and bioanalytical techniques such as LC-MS/MS, GC-MS, and ELISA. In the scope of a clinical study, a comprehensive assessment of the levels of oxidative stress products in biological fluids and tissues is useful in gaining a better understanding of the patient’s overall health status. Specific biomarkers of lipid peroxidation have also been used as indicators of disease progression. Furthermore, clinical studies evaluating the effects of nutritional interventions in healthy patients, as well as in disease models, can also benefit from including oxidative stress biomarker analyses. Science-backed functional food and dietary supplements have a greater chance of succeeding in a competitive market and finding long-term customers, while at the same time giving credibility to the producer.
- 1. Wachtel-Galor S, Benzie IFF. Series Preface. In: Benzie IFF, Wachtel-Galor S, eds. Herbal Medicine Biomolecular and Clinical Aspects. 2nd ed. Boca Raton, FL: CRC Press; 2011:1-10.
- 2. Montuschi P, Barnes JP, Roberts JL. Isoprostanes: markers and mediators of oxidative stress. The FASEB Journal. 2004; 18(15): 1791-1800.
- 3. Marnett LJ. Lipid peroxidation – DNA damage by malondialdehyde. Mutat Res-Fund Mol M. 1999; 424(1-2):83-95.
- 4. Montuschi P, Corradi M, Ciabattoni G, et al. Increased 8-isoprostane, a marker of oxidative stress, in exhaled condensate of asthma patients. Am J Respir Crit Care Med. 1999; 160:216–220.
- 5. Montine TJ, Neely MD, Quinn JF, et al. Lipid peroxidation in aging brain and Alzheimer’s disease. Free Radic Biol Med. 2002; 33:620–626.
- 6. Cracowski JL, Marpeau C, Carpentier PH, et al. Enhanced in vivo lipid peroxidation in scleroderma spectrum disorders. Arthritis Rheum. 2001; 44:1143–1148.
- 7. Pratico D, Iuliano L, Basili S, et al. Enhanced lipid peroxidation in hepatic cirrhosis. J Invest Med. 1998; 46:51–57.
- 8. Dalle-Donne I, Rossi R, Giustarini D, et al. Protein carbonyl groups as biomarkers of oxidative stress. Clin Chim Acta. 2003; 329:23-38.
- 9. Tsuboi H, Kouda K, Takeuchi H, et al. 8-Hydroxydeoxyguanosine in Urine As an Index of Oxidative Damage to DNA in the Evaluation of Atopic Dermatitis. Br J Dermatol. 1998; 138:1033-1035.
Jasenka Piljac Zegarac is a scientist and freelance writer. She holds a PhD in biology and a BS degree in biochemistry, and contributes on a regular basis to several health and science blogs. She may be contacted for assistance with a variety of science and medical writing projects. Find Jasenka on LinkedIn.