Clinical use of chemiluminescence assays for the determination of systemic oxidative stress.

REPETTO, MARISA

Handbook of chemiluminescent methods in oxidative stress assessment.

Transworld Research Network

Lugar: Kerala; Año: 2008; p. 163 - 194

The high sensitivity of chemiluminescence was utilized to develop two assays for the determination of systemic oxidative stress in humans: (a) the tert-butyl hydroperoxide initiated chemiluminescence (BOOH-CL) of erythrocytes, and (b) the plasma total reactive antioxidant potential (TRAP). The two complementary assays determine the blood level of antioxidants and report on the situation of systemic oxidative stress that may reflect a whole body situation or a localized dysfunction (i.e., the brain in neurodegenerative diseases). Plasma and erythrocyte antioxidants decrease in their concentration during systemic oxidative damage. In the BOOH-CL assay, washed erythrocytes are supplemented with tert-butyl hydroperoxide (BOOH) and the following burst of chemilumnescence is measured. The assay can be applied to erythrocytes, blood samples, biopsies and tissue homogenates. Added BOOH reacts with the heme-Fe2+ of oxyhemoglobin or hemoproteins producing peroxyl and alcoxyl free radicals, which enter into the propagation phase of the lipid peroxidation of erythrocyte unsaturated phospholipids. The termination steps of the free-radical chain reaction generate compounds in an excited state: singlet oxygen and carbonyl groups. The photoemission rate and level limiting step is erythrocyte a-tocopherol and the assay is useful to evaluate the integral level of systemic antioxidants. In the TRAP assay the plasma antioxidant capacity is determined by the inhibition of the burst of luminol chemiluminescence elicited by the thermal decomposition of the azo-compound ABAP that produces peroxyl radicals. The inhibition or lag time before the burst of luminol chemiluminescence is directly proportional to the radical trapping capacity of the plasma sample, including antioxidant concentration and the stoichiometric relationship of peroxyl trapping. Both tests are able to determine situations of systemic oxidative stress in neurodegenerative diseases, such as Parkinson?s and Alzheimer?s diseases and vascular dementia; in immunological diseases, such as HIV infection and AIDS, and in endocrine pathological situations, such as hyperthyroidism and hypothyroidism. The assays were also used to evaluate oxidative damage and lipid peroxidation in experimental rat models. Oxidative damage and antioxidant protection in tissues and plasma were observed in rat gastric ulcers produced by acute ethanol intake, and in the necrotic damage to liver, kidney, heart, and brain induced by a choline deficient diet.