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When chilling sensitive plants are exposed to low temperature, photosynthetic activity is rapidly reduced. To investigate the biochemical mechanisms involved in chilling-induced reduction in photosynthetic activity of the chloroplast, it is necessary to isolate active chloroplasts in a coupled state. When chloroplasts were isolated from chilling-sensitive melon (Cucumis melo L.), bell pepper (Capsicum annuum L.), and eggplant (Solanum melongena L.) by the current, conventional method, active, coupled chloroplasts were not obtained from melon and bell pepper leaves. To improve the method for isolating active chloroplasts from these plants, sucrose as an osmoticant was replaced with NaCl and sorbitol in the grinding and suspension media, respectively; isoascorbate and EDTA were also added in the grinding medium. With these media, isolated, chilled chloroplasts had measurable electron transport rates, and sufficient coupling to investigate the site of inhibition of photochemical processes. Melon leaves exposed to 5 °C for 24 hr under 100 μmol•m-2•s-1 showed a much reduced photosynthetic rate at 25 °C, compared to unchilled leaves. Measurements of activity of chloroplasts isolated from chilled and unchilled leaves by the improved method revealed that the whole-chain electron transport rates were unaffected by chilling, whereas the extent of coupling was decreased. This result suggests that photophosphorylation would be involved in the inhibition of photosynthesis in chilled melon leaves. Thus, the improved method for isolating chloroplasts is applicable to study the inhibition site of photochemical reactions in chilling-sensitive melon, bell pepper, and eggplant.

Improved Methods for Isolating Active Chloroplast from Chilling-Sensitive Melon, Bell Pepper and Eggplant.