Comparison of methods for determination of total antioxidant status: application to analysis of medicinal plant essential oils

The free radical scavenging capacity of a wide range of plant oil extracts, principally those used in traditional European herbal medicine (with novel therapeutic potential for patients with degenerative disorders of the CNS), has been compared in vitro. The antioxidant capacity of individual plant extracts was determined via three complementary assay procedures, based on: (i) attenuation of the generation of ABTS√⁺ radical (quantitated colorimetrically), by a metmyoglobin catalyst/hydrogen peroxide system; (ii) inhibition of iodophenol enhanced chemiluminescence by a horseradish peroxidase/perborate/luminol system; (iii) protection of a target enzyme (human brain alanyl aminopeptidase, activity quantitated via fluorimetric assay) against oxidative damage by √OH or O√⁻₂ generated by Co⁶⁰γ radiolysis. In assays (i) and (ii), only three plant extracts (cinnamon, pimento, bay) showed substantial antioxidant activity, although the two assays yielded quantitatively different values of antioxidant activity (Trolox equivalent values of 16–25 M (method ii) and 0.25–2.1 M (method (i)). None of the plant extracts investigated showed significant antioxidant protective activity against √OH or O√⁻₂ species in assay (iii). The data obtained thus demonstrate that the apparent antioxidant capacity of putative free radical scavenging agents depends entirely on the assay method utilized and particular free radical species generated. We therefore suggest that antioxidant capacity determined by a single assay method (particularly via competitive assay with ABTS√⁺) should be interpreted with some caution. This conclusion may be of particular potential importance in clinical chemistry, in view of the current interest in the assessment of the antioxidant status of tissues of patients with a variety of disorders.