Superoxide radical production by microsomal membranes from senescing carnation flowers: an effect on membrane fluidity

The lipid fluidity of carnation microsomal membranes decreases during in vitro aging in a manner that correlates with enzymatic production of superoxide anion (O 2 − dot ) by the membranes. Levels of (O 2 − dot ) were determined from ESR spectra of the semiquinone formed when (O 2 − dot ) reacts with Tiron (1,2-dihydroxybenzene-3,5-disulfonic acid). Heat denaturation of the microsomes or addition of n-propyl gallate, a free radical scavenger, eliminated the Tiron radical signal and prevented the change in membrane fluidity. By contrast, the addition of xanthine/xanthine oxidase, a reputed (O 2 − dot )-generating system, accentuated the decrease in membrane fluidity and greatly increased the Tiron radical signal. Superoxide dismutase, an enzyme that catalyses the dismutation of (O 2 − dot ), reduced the amplitude of the Tiron radical signal. When Tiron was used as a scavenger of (O 2 − dot ), there was no change in membrane fluidity. The data collectively suggest that (O 2 − dot ) or highly reactive derivatives of (O 2 − dot ) induce membrane rigidification. In addition, microsomes from senescent flowers produce more (O 2 − dot ) than those from freshly cut flowers, suggesting that this reactive species of oxygen also contributes to membrane rigidification during natural senescence.