植物在低溫逆境下的耐受性是決定其光合作用產能的重要因子之一。目前對於低溫導致光合作用減緩的原因有著數種說法，主要為低溫可能造成（1）類囊膜的流動性降低；（2）光系統II（PSII）被抑制；（3）光系統I（PSI）被抑制；和（4）暗反應被抑制等。以往探討植物光合作用在低溫下的反應多為長期實驗（數小時、數天到數個月），只能看到低溫對植物所造成的中、長期效果，但無法了解其早期的影響。本研究選擇了耐冷植物豌豆（Pisum sativum L.）與不耐冷植物小黃瓜（Cucumis sativus L.），欲研究它們的光合作用對突然出現的低溫所做的早期反應。
本研究經由葉綠素螢光來觀察豌豆和小黃瓜在溫度逆境下整個光合作用（包括光反應和暗反應）的運作，也利用酵素連結系統來測量Rubisco在低溫下活化狀態的改變。我們發現豌豆和小黃瓜突然遭遇5℃後，豌豆的電子傳遞速率會下降但迅速地恢復（5分鐘內），而小黃瓜下降的幅度更大而恢復的情況則比較差；如果植物事先浸泡methyl viologen可以明顯地消除此低溫下的抑制現象。另外，二氧化碳會使得低溫下的抑制情況更嚴重，而降低氧氣含量可以促進光合作用的進行。我們也由酵素連結實驗發現Rubisco的活性變化和葉綠素螢光所觀察到的電子傳遞的變化有一致性。

從葉綠素螢光實驗可以觀察到豌豆的qN伴隨ETR恢復與qP的上升而逐漸升高，再加上從qN relaxation實驗觀察到大部份的qN是在十幾分鐘內衰減，我們推測qN主要的組成成分是QT（state transition）。也就是耐冷植物豌豆受到在低溫強光照射，PSII會將部份的光能傳導給PSI，因有這個機制的保護，所以光合作用可以在低溫下保持良好狀態。而小黃瓜在低溫下qN不會快速升高，光合作用也沒有獲得抒解。

綜合以上實驗，我們可以發現植物在低溫逆境下，其光合作用抑制的主要位置應為暗反應。而豌豆在5℃可以維持高光合作用產率可能和豌豆能利用QT來排解過多激發能量，還有它的Rubisco具調節性相關，5℃下小黃瓜的沒有qN的累積，Rubisco也不具有調節能力，故產率會下降。由於光合作用反應的複雜性，使得偵測到的螢光往往是多種因素混合而產生的結果，所以仍需要更多的實驗以了解光合作用的各個部分在面對低溫時的反應與扮演的角色。

The tolerance to chilling temperature is one of the most important factors in determining photosynthetic yield of plants. The chilling temperature may lead to (1) a decrease in the fluidity of thylokoid membrane; (2) an inhibition of photosystem II (PSII) and/or photosystem I (PSI); and (3) slowing down of the dark reaction. Former studies concerning with photosynthesis in chilling temperature were almost long-term experiments (several hours, several days and several months), thus we can only figure out the middle and long term effects, while the information about plants’ early responses are still sparse. In this study, we choose a cold-tolerant plant, pea (Pisum sativum L.), and a cold-sensitive plant, cucumber (Cucumis sativus L.), to investigate their early response of photosynthesis to chilling temperature.
The photosynthesis of pea and cucumber, including light and dark reactions were monitored by chlorophyll fluorescenc. Enzymatic assay was used to measure Rubisco activity. We found that when facing a sudden lowing of temperature (5℃), the electron transport rate (ETR) of pea droped but with a rapid recovery while that of cucumber was more seriously inhibited and with a recovery to much less extent. We also found that the inhibition under 5℃ could be alleviated if we pre-treated leaves sample with methyl viologen, which is an efficient electron acceptor of PSI. In addition, high CO2 concentration would make the inhibition more seriously, while reducing O2 concentration would ease it. In the enzyme-linked assay, we found the activation state of Rubisco paralleled with the change of electron transport rate.

From the chlorophyll fluorescence experiment, we found that qN of pea rose following the recovery of Yield and qP. We also found that most of its qN relaxed in ten minutes. Thus we infer that the major component of qN is QT (state transition). It means that PSII of pea may transfer excess radiation energy to PSI, and with the protection mechanism pea can keep well photosynthesis under chilling stress. On the contrary, qN of cucumber didn’t rise too much so that we could observe its photosynthetic yield still inhibited under chilling stress.

We thus suggest that (1) Dark reaction is the major inhibition site under chilling stress. (2) The photosynthesis rate of cold-tolerant plant like pea decreases in response to a sudden chilling stress, but it recovers within l minutes. On the other hand, cold-sensitive plant like cucumber doesn’t possess thus kind of capability. The phenomenon was not observed by former long-term experiments. (3) The ability of pea to keep normal photosynthetic yield in chilling temperature may be related to the Rubisco adjustment and the ability to mediate the excess radiation energy (QT), and which is absent in cucumber.

Owning to the complexity of photosynthesis, the fluorescence detected is always a result of effects of multi-factors. We still need more experimental data to find out the roles played by various parts of photosynthesis under chilling stress.

Allen D. J. and Ort D. R. (2001) Impacts of chilling temperatures on photosynthesis in warm-climate plants. Trends Plant Sci., 6, 36-42.
Allen J. F. (2003) Cyclic, pseudocyclic and noncyclic photophosphory-
lation: new links in the chain. Trends Plant Sci., 8, 15-19.
Asada K. (1999) The water-water cycle in chloroplasts: scavenging of active oxygens and dissipation of excess photons. Annu. Rev. Plant Physiol. Plant Mol. Biol., 50, 601-639.
Bahr J. T. and Jensen R. G. (1978) Activation of ribulose bisphosphate carboxylase in intact chloroplast by CO2 and light. Arch. Biochem. Biophys., 185, 39-48.
Bowes G. (1991) Growth at elevated CO2: photosynthetic responses mediated through Rubisco. Plant Cell Environ., 14, 795-806.
Byrd G. T., Ort D. R., and Ogren W. L. (1995) The effects of chilling in the light on Ribulose-1,5-bisphosphate carboxylase/oxygenase activation in tomato. Plant Physiol., 107, 585-591.
Choi S. M., Jeong S. W., and Jeong W. J. (2002) Chloroplast Cu/Zn-superoxide dismutase is a highly sensitive site in cucumber leaves chilled in the light. Planta, 216, 315-324.
Clarke J. E. and Johnson G. N. (2001) In vivo temperature dependence of cyclic and pseudocyclic electron transport in barley. Planta., 212, 808-816.
Fryer M. J., Andrews J. R., Oxborough K., Bloxers D. A., and Baker N. R. (1998) Relationship between CO2 assimilation, photosynthetic electron transport, and active O2 metabolism in leaves of maize in the field during periods of low temperature. Plant Physiol., 116, 571-580.
Hall D. O. and Rao K. K. (1999). Photosynthesis, 6th edn., pp.103-107. Cambridge University Press, Cambridge.
Holaday A. S., Martindale W., Alred R., Brooks A. L., and Leegood R. C. (1992) Changes in activities of enzymes of carbon metabolism in leaves during exposure of plants to low temperature. Plant Physiol., 98, 1105-1114.
Hymus G. J., Baker N. R., and Long S. P. (2001) Growth in elevated CO2 can both increase and decrease photochemistry and photoinhibition of photosynthesis in a predictable manner. Dactylis glomerata grown in two levels of nitrogen nutrition. Plant Physiol., 127, 1204-1211.
Iba K. (2002) Acclimative response to temperature stress in higher plants: approaches of gene engineering for temperature tolerance. Annu. Rev. Plant Biol., 53, 225-2`45.
Kim S. J., Lee C. H., Hope A. B. and Chow W. S. (2001) Inhibition of photosystem I and II and enhanced back flow of photosystem I electrons in cucumber leaf discs chilled in the light. Plant cell physiol., 42, 842-848.
Kingston-Smith A. H., Harbinson J., Williams J., and Foyer C. H. (1997) Effect of chilling on carbon assimilation, enzyme activation, and photosynthetic electron transport in the absence of photoinhibition in maize leaves. Plant Physiol., 114, 1039-1046.
Kudoh H. and Sonoike K. (2002) Irreversible damage to photosystem I by chilling in the light: cause of the degradation of chlorophyll after returning to normal growth temperature. Planta, 215, 541-548.
Labate C. A., Adcock M. D., and Leegood R. C. (1990) Effects of temperature on the regulation of photosynthetic carbon assimilation in leaves of maize and barley. Planta., 181, 547-554.
Lazár D. (1999) Chlorophyll a fluorescence induction. Biochem. Biophys. Acta, 1412, 1-28.
Leegood R. C. (1993) Carbon metabolism. In: Photosynthesis and production in a changing environment: A field and laborantory manual. Hall D. O., Scurlock J. M. O., Bolhàr-nordenkampf H. R., Leegood R. C., and Long S. P. (eds) Chapman & Hall, London. pp. 248-267.
Lormer G. H., Badger M. R., and Andrews T. J. (1976) The activation of ribulose-1,5-bisphosphate carboxylase by carbon dioxide and magnesium ions. Equilibria, kinetics, a suggested mechanism, and physiological implications. Biochemistry, 15, 529-536.
Mathai J. C., Sauua Z. E., John O., and Sitaramam V. (1993) Rate-limiting steps in electron transport. Osmotically sensitive diffusion of quinines through voids in the bilayer. J. Biol. Chem., 268, 15442-15454.
Moon B. Y., Higashi S. I., Gombos Z. Gombos Z., and Murata N. (1995) Unsaturation of the membrane lipids of chloroplasts stabilizes the photosynthetic machinery against low-temperature photoinhibition in transgenic tobacco plants. Proc. Natl. Acad. Sci. USA, 92, 6219-6223.
Müller P., Li X. P., and Niyogi K. K. (2001) Non-photochemical quenching. A response to excess light energy. Plant Physiol., 125, 1558-1566.
Perchorwicz J. T., Raynes D. A., and Jensen R. G. (1981) Light limitation of photosynthesis and activation of ribulose bisphosphate carboxylase in wheat seedlings. Proc. Natl. Acad. Sci. USA, 78, 2985-2989.
Portis A. R., Jr. (1990) Rubisco activase. Biochem. Biophys. Acta, 1015, 15-28.
Raison J. K. and Orr G. R. (1986) Phase transitions in thylakoid polar lipids of chilling-sensitive plants. Plant Physiol., 80, 638-645.
Rohácek, K. and Barták, M. (1999) Technique of the modulated chlorophyll fluorescence: basic concepts, useful parameters, and some applications. Photosynthetica, 37, 339-363.
Sage R. F., Sharkey T. D., and Seemann J. R. (1989) Acclimation of photosynthesis to elevated CO2 in five C3 species. Plant Physiol., 89, 590-596.
Sage R. F., Sharkey T. D., and Seemann J. R. (1990) Regulation of Ribulose-1,5-bisphosphate carboxylase activity in response to light and CO2 in the C3 annuals Chenopodium album L. and Phaseolus vulgaris L. Plant Physiol., 94, 1735-1742.
Salvucci M. E., Osteryoung K. W., Crafts-Brandner S. J., and Vierling E. (2001) Exceptional sensitivity of Rubisco activase to thermal denaturation in vitro and in vivo. Plant Physiol., 127, 1053-1064.
Sharkey T. D., Savitch L. V., and Butz N. D. (1991) Photometric for routine determination of kcat and carbamylation of rubisco. Photosynthesis Research, 28, 41-48.
Spreitzer R. J. and Salvucci M. E. (2002) Rubisco: structure, regulatory interactions, and possibilities for a better enzyme. Annu. Rev. Plant Biol., 53, 449-475.
Stitt M (1991) Rising CO2 level and their potential significance for carbon flow in plant cells. Plant Cell Environ., 14, 741-762.
Stitt M. (2003) Metabolic Regulation of photosynthesis. In: Baker N. R. ed., Photosynthesis and environment. Kluwer Academic Publishers, pp151-190.
Tjus S. E., Møller B. L., and Scheller H. V. (1998) Photosystem I is an early target of photoinhibition in barley illuminated at chilling temperatures. Plant physiol., 116, 755-764.
Tjus S. E., Scheller H. V., Andersson B., and Møller B. L. (2001) Active oxygen produced during selective excitation of photosystem I is damaging not only to photosystem I, but also to photosystem II. Plant Physiol., 125, 2007-2015.
Van Mieghem, F., Brettel K., Hillmann B., Kamlowski A., Rutherford A. W., and Schlodder E. (1995) Charge recombination reactions in photosystem II. 1. Yields, recombination pathways, and kinetics of the primary pair. Biochemistry, 34, 4798-4813.
Vu C. V., Allen L. H., Jr., and Bowes George. (1983) Effects of light and elevated atmospheric CO2 on the Ribulose bisphosphate carboxylase activity and Riulose bisphosphate level of soybean leaves. Plant Physiol., 73, 729-734,
Ward D. A. and Keys A. J. (1989) A comparison between the spectrometric and uncoupled radiometric assays for RuBP carboxylase. Photosynthesis Research, 22, 167-171.
Wolfe D. W., Gifford R. M., Hilbert D., and Luo Y. (1998) Integration of photosynthetic acclimation to CO2 at the whole-plant level. Global change biology, 4, 879-893.
Xu D. Q., Gifford R. M., and Chow W. S. (1994) Photosynthetic acclimation in pea and soybean to high atmospheric CO2 partial pressure. Plant Physiol., 106, 661-671.