AGING OF CUCUMBER AND ONION SEEDS - PHOSPHOLIPASE-D, LIPOXYGENASE ACTIVITY AND CHANGES IN PHOSPHOLIPID CONTENT

Cucumber (Cucumis sativus L. cv. Mesa) and onion (Allium cepa L. cv. R ijnsburger Heldis) seeds were rapidly aged at 40-degrees-C and 74% rel ative humidity. Onion seeds were also slowly aged at 40-degrees-C with 15% relative humidity for 11 months and one more month at 28% relativ e humidity. Significant loss of total and individual phospholipids was an early event during both storage treatments. With slow ageing of on ion, loss of phosphatidylcholine occurred several months before loss o f viability and vigour was detected. Phosphatidic acid, the lipid prod uct of phospholipase D action, increased during rapid ageing of both c ucumber and onion. Phosphatidic acid was present in onion seeds before the ageing treatments and its content remained unchanged in the slowl y aged seeds. There was 1600 (cucumber) and 2000 (onion) times more ph ospholipase D activity (6 x 10(5) and 2-9 x 10(5) nmol g-1 d-1 in cucu mber and onion, respectively) in crude extracts from non-aged seeds th an was required to account for the fastest fall in phospholipids (72, 372 and 144 nmol g-1 d-1 for cotyledons and radicles of cucumber and o nion, respectively, over the first 9 d [cucumber] or 1 d [onion] of ag eing) and fastest increase in phosphatidic acid (7, 162 and 37 nmol g- 1 d-1). How accurate a guide the in vitro activity of phospholipase D was to the in vivo activity was unclear. However, the considerable exc ess activity seen with the formation of phosphatidic acid supports the proposal that hydrolysis of phospholipids by phospholipase D is a fir st step in deterioration during ageing. Substantial lipoxygenase activ ity was also detected (58 x 10(3) and 54 x 10(3) nmol g-1 d-1, respect ively, for non-aged cucumber and onion seeds). However, the increase i n conjugated dienes (an early product of peroxidation) in ageing cucum ber seeds was comparatively small (90 nmol g-1 d-1 over 21 d ageing), and increase in malondialdehyde could not be detected, indicating that peroxidation may not have been a major factor in cucumber. The increa se in conjugated dienes during rapid ageing of onion seeds was larger (1.5 x 10(3) nmol g-1 d-1 over days 0-2 of rapid ageing), much greater than the decrease in phospholipid acyl groups (260 nmol g-1 d-1 over days 0-2 of rapid ageing) indicating the occurrence of peroxidation of fatty acids released from reserve as well as from membrane lipids. Th is higher level of conjugated dienes during onion ageing was the main difference between cucumber and onion, indicating that the level of pe roxidation could be an important difference between fast and slow agei ng seeds. However, peroxidation is not the only possible deleterious p rocess since hydrolysis of the normal membrane phospholipids to phosph atidic acid increased the content of non-bilayer-forming lipids and th is too could be a direct membrane-destabilizing consequence of phospho lipase D action during ageing.