Isolated chloroplasts of Euglena gracilis Klebs were kept for 10 d in complete darkness at 4° C in a maintenance buffer (pH 7.5) without shaking. During incubation, the qualitative and quantitative changes in the pattern of photosynthetic pigments were evaluated by the combined use of spectrophotometry in the visible range of whole chloroplasts and their acetone extracts, of in vivo spectrofluorimetry and of reversed-phase HPLC. Microscopic and submicroscopic modifications were also followed by UV and transmission electron microscopy. The main findings were as follows: (1) a fast decay of all photosynthetic pigments, chiefly chlorophylls, not accompanied by evident signs of alteration of the thylakoid system during the first 5 d; (2) a higher stability of PSII compared to PSI and of antenna complexes compared to the relative reaction centres during the first 24-48 h; (3) a low accumulation of phaeoderivative compounds in spite of the marked decrease of chlorophyll content; (4) a lack of dephytylated compounds; (5) a quicker decay of the intensity of fluorescence emission with respect to the decreasing chlorophyll a content; and (6) a fast degradation of xanthophylls and ß-carotene with the consequent lack of defence from the ageing oxidative stresses. This accounts for the rapid loss of pigments, although the lack of other antioxidant defence mechanisms is not excluded. The characterization of some of the steps involved in plastid degradation may render this experimental model viable for further studies on plastid senescence, a multifactorial process still awaiting definite answers.
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