Photosystem II Excitation Pressure and Development of Resistance to Photoinhibition (I. Light-Harvesting Complex II Abundance and Zeaxanthin Content in Chlorella vulgaris)

The basis of the increased resistance to photoinhibition upon growth at low temperature was investigated. Photosystem II (PSII) excitation pressure was estimated in vivo as 1 - qp (photochemical quenching). We established that Chlorella vulgaris exposed to either 5[deg]C/150 [mu]mol m-2 s-1 or 27[deg]C/2200 [mu]mol m-2 s-1 experienced a high PSII excitation pressure of 0.70 to 0.75. In contrast, Chlorella exposed to either 27[deg]C/150 [mu]mol m-2 s-1 or 5[deg]C/20 [mu]mol m-2 s-1 experienced a low PSII excitation pressure of 0.10 to 0.20. Chlorella grown under either regime at high PSII excitation pressure exhibited: (a) 3-fold higher light-saturated rates of O2 evolution; (b) the complete conversion of PSII[alpha] centers to PSII[beta] centers; (c) a 3-fold lower epoxidation state of the xanthophyll cycle intermediates; (d) a 2.4-fold higher ratio of chlorophyll a/b; and (e) a lower abundance of light-harvesting polypeptides than Chlorella grown at either regime at low PSII excitation pressure. In addition, cells grown at 5[deg]C/150 [mu]mol m-2 s-1 exhibited resistance to photoinhibition comparable to that of cells grown at 27[deg]C/2200 [mu]mol m-2 s-1 and were 3- to 4-fold more resistant to photoinhibition than cells grown at either regime at low excitation pressure. We conclude that increased resistance to photoinhibition upon growth at low temperature reflects photosynthetic adjustment to high excitation pressure, which results in an increased capacity for nonradiative dissipation of excess light through zeaxanthin coupled with a lower probability of light absorption due to reduced chlorophyll per cell and decreased abundance of light-harvesting polypeptides.     

Photosynthetic vesicles with bound phycobilisomes from Anabaena variabilis.

Photosynthetically active vesicles with attached phycobilisomes from Anabaena variabilis, were isolated and shown to transfer excitation energy from phycobiliproteins to F696 chlorophyll (Photosystem II). The best results were obtained when cells were disrupted in a sucrose/phosphate/citrate mixture (0.3 : 0.5 : 0.3 M, respectively) containing 1.5% serum albumin. The vesicles showed a phycocyanin/chlorophyll ratio essentially identical to that of whole cells, and oxygen evolution rates of 250 mumol O2/h per mg chlorophyll (with 4 mM ferricyanide added as oxidant), whereas whole cells had rates of up to 450. Excitation of the vesicles by 600 nm light produced fluorescence peaks (-196 degrees C) at 644, 662, 685, 695, and 730 nm. On aging of the vesicles, or upon dilution, the fluorescence yield of the 695 nm emission peak gradually decreased with an accompanying increase and final predominant peak at 685 nm. This shift was accompanied by a decrease in the quantum efficiency of Photosystem II activity from an initial 0.05 to as low as 0.01 mol O2/einstein (605 nm), with a lesser change in the Vmax values. The decrease in the quantum efficiency is mainly attributed to excitation uncoupling between phycobilisomes and Photosystem II. It is concluded that the F685 nm emission peak, often exclusively attributed to Photosystem II chlorophyll, arises from more than one component with phycobilisome emission being a major contributor. Vesicles from which phycobilisomes had been removed, as verified by electron microscopy and spectroscopy, had an almost negligible emission at 685 nm.     

In vitro preparation and characterization of a 700 nm absorbing chlorophyll-water adduct according to the proposed primary molecular unit in photosynthesis.

1. This study characterized chlorophyll a-H2O adducts in vitro in order to establish their generic relationship to the recently proposed [15, 18-20, 31] primary molecular adduct in photosynthesis. The effects of water titration and temperature on the absorption, fluorescence, excitation, and redox properties of the various in vitro chlorophyll a aggregate species are investigated. 2. From fluorescence measurements, we conclude that the driest chlorophyll a sample contains an equimolar amount of water. This conclusion is consistent with earlier experimental work [2, 3, 14, 17, 31], and clarifies the origin of the controversial [15] Katz model [14] of chlorophyll a-H2O interactions. 3. With increasing water concentration or as the temperature is lowered below room temperature, the A-663 monohydrate chlorophyll a-H2O (species absorbing at 663 nm) is favored at the expense of the A-678 anhydrous aggregate according to the equilibrium 2H2O+chlorophyll a2in equilibrium2 chlorophyll a-H2O. Under excess water conditions, A-663 is converted to A-743 (chlorophyll a-2H2O)n. 4. On slow sample cooling to T less than or approximately 200 degrees K, we observe the growth of A-700 at the expense of A-663. There is a direct correspondence between the increasing (decreasing) absorption by A-700 (A-633) and increasing (decreasing) fluorescence at 720 nm (664 nm). 5. It is concluded that A-700 is most probably the dimer participating in the equilibrium 2 chlorophyll a-H2O in equilibrium (chlorophyll a-H2O)2. The A-700 band consists of two exciton components (separated by approximately 280 cm1) that are interpretable in terms of the dimeric origin of A-700. 6. The deconvoluted A-700 absorption spectrum and the excitation spectrum of the 720 nm fluorescence are compared with the light-minus-dark spectra of P-700. 7. It is found that A-700 is reversibly bleached by I2 (E0 equals 0.54 V). The significance of this observation is discussed in terms of the redox properties of monomeric chlorophyll a and P-700.     

Regulation of the distribution of excitation energy in Ochromonas danica,an organism containing a chlorophyll-A/C/carotenoid light harvesting antenna

A chlorophyll a, c-fucoxanthin pigment-protein complex8 functions as the major light harvesting antenna in the Chrysophyte Ochromonas danica. The regulated distribution of excitation energy between the two photosystems was investigated in these organisms and was shown to be strongly wavelength dependent. A light state transition was induced by pre-illumination of cells using light 2 (640 nm) and light 1 (700 nm) of equal absorbed intensity, and detected by reversible changes in the 77 K chlorophyll fluorescence emission spectra. Peaks at 690 nm and 720 nm in the low temperature spectra are most likely associated with PS2 and PS1 respectively. A room temperature fluorescence emission at 680 nm induced by modulated light 2 (500 nm) was strongly quenched in the presence of background light 1 (720 nm). Removal of light 1 led to an increase in fluorescence followed by a slow quenching. The room temperature fluorescence changes were directly correlated with changes in the 77 K emission spectra that indicated a change in the distribution of excitation energy between the two photosystems. It was established that DCMU (1 mol) prevented the state 2. The conversion to state 1 followed a simple photochemical dose dependence and had a half-time of 20 s-1.5 min at 6 W m^-2. In contrast, the conversion to state 2 was independent of light intensity. These data indicate that O. danica undergoes a light state transition in response to the preferential excitation of PS2 or PS1.Abbreviations PS2 photosystem 2 - PS1 photosystem 1 - LHC light harvesting chlorophyll a/b protein - fx fucoxanthin - PQ plastoquinone - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethyl urea     

Direct fluorescence measurement of diffusional water permeability in the vasopressin-sensitive kidney collecting tubule.

A fluorescence method has been developed for accurate and instantaneous measurement of transepithelial diffusional water permeability (Pd) in perfused kidney tubules based on the sensitivity of the fluorophore aminonapthelane trisulfonic acid (ANTS) to solution H2O/D2O content. The fluorescence of ANTS was 3.2-fold lower in an H2O buffer than in a D2O buffer. The response of ANTS fluorescence to a change in solution H2O/D2O content occurred in less than 1 ms and was due to a collisional quenching mechanism. Isolated cortical (CCT) and outer medullary (OMCT) collecting tubules from rabbit were perfused with an isosmotic D2O buffer at specified lumen flow rates (2-100 nl/min); tubules were bathed in isosmotic H2O or D2O buffers in which vasopressin (VP) could be added rapidly. Lumen fluorescence was monitored by quantitative epifluorescence microscopy at 380 +/- 5 nm excitation and greater than 530 emission wavelengths. Pd was determined from tubule geometry, lumen flow, ANTS fluorescence, and ANTS fluorescence vs. H2O/D2O calibration relation. The instrument response time for a change in bath H2O/D2O content was less than 4 s. At 37 degrees C, Pd values (mean +/- SE in cm/s x 10(4] were 6.4 +/- 1.0 (-VP, n = 9) and 14.3 +/- 1.1 (+250 microU/ml bath VP, n = 9) in the CCT, and 5.8 +/- 1.0 (-VP, n = 6) and 15.3 +/- 2.0 (+VP, n = 6) in the OMCT; at 23 degrees C, Pd was 5.1 +/- 0.6 (-VP, n = 4) and 7.8 +/- 0.6 (+VP, n = 4) in the CCT.(ABSTRACT TRUNCATED AT 250 WORDS)     

Execution of programmed cell death by singlet oxygen generated inside the chloroplasts of Arabidopsis thaliana

Protoplasma - Absorption of excess excitation energy induces overproduction of singlet oxygen (1O2) in plants. The major sources of singlet oxygen production are chlorophyll and its intermediates...     

Studies on organomercury(II) complexes of isoniazid

A number of organomercury(II) complexes involving isoniazid (I), of the type RHgCl(L)(II) [R = phenyl(C6H5), o-hydroxyphenyl (o-HOC6H4), p-hydroxyphenyl (p-HOC6H4), p-acetoxyphenyl (p-AcOC6H4), 2-furyl (2-C4H3O); L = isoniazid] have been synthesized and characterized. Conductance measurements indicate that the complexes are nonelectrolytes. From IR and UV studies, it is concluded that isoniazid acts as a bidentate ligand, coordinating through hydrazinic nitrogen and carbonyl oxygen. 1H and 13C NMR support the stoichiometry of the complexes. From fluoroscence studies a number of photochemical parameters have been elucidated. For the C6H5HgCl(L), p-HOC6H4HgCl(L), and p-AcOC6H4HgCl(L) complexes, thermogravimetric studies have been carried out and relevant kinetic and thermodynamic parameters for thermal degradation have been enumerated. In addition, the fragmentation pattern of the complexes has been analyzed on the basis of mass spectra. The C6H5HgCl(L) and p-HOC6H4HgCl(L) complexes have been screened for tuberculosis activity.     

Photosystem II Excitation Pressure and Photosynthetic Carbon Metabolism in Chlorella vulgaris

Chlorella vulgaris grown at 5[deg]C/150 [mu]mol m-2 s-1 mimics cells grown under high irradiance (27[deg]C/2200 [mu]mol m-2 s-1). This has been rationalized through the suggestion that both populations of cells were exposed to comparable photosystem II (PSII) excitation pressures measured as the chlorophyll a fluorescence quenching parameter, 1 - qP (D.P. Maxwell, S. Falk, N.P.A. Huner [1995] Plant Physiol 107: 687-694). To assess the possible role(s) of feed-back mechanisms on PSII excitation pressure, stromal and cytosolic carbon metabolism were examined. Sucrose phosphate synthase and fructose-1,6-bisphosphatase activities as well as the ratios of fructose-1,6-bisphosphate/fructose-6-phosphate and sucrose/starch indicated that cells grown at 27[deg]C/2200 [mu]mol m-2 s-1 appeared to exhibit a restriction in starch metabolism. In contrast, cells grown at 5[deg]C/150 [mu]mol m-2 s-1 appeared to exhibit a restriction in the sucrose metabolism based on decreased cytosolic fructose-1,6- bisphosphatase and sucrose phosphate synthase activities as well as a low sucrose/starch ratio. These metabolic restrictions may feed-back on photosynthetic electron transport and, thus, contribute to the observed PSII excitation pressure. We conclude that, although PSII excitation pressure may reflect redox regulation of photosynthetic acclimation to light and temperature in C. vulgaris, it cannot be considered the primary redox signal. Alternative metabolic sensing/signaling mechanisms are discussed.     

高、低温胁迫对牡丹叶片PSⅡ功能和生理特性的影响

以牡丹‘肉芙蓉’离体叶片为试材,以25 ℃为对照,研究了强光(1400 μmol·m-2·s-1)下高温(40℃)和低温(15℃)处理对牡丹叶片PSⅡ光化学活性和生理特性的影响.结果表明:随处理时间的延长,各处理叶片的PSⅡ最大光化学效率(Fv/Fm)、PSⅡ实际光量子效率(φPsⅡ)和光下开放的PSⅡ反应中心激发能捕获效率(Fv’/Fm’)均持续降低.暗恢复4h后,对照和15℃处理叶片的Fv/Fm基本上完全恢复,而40℃处理叶片仅恢复到处理前的75.5％,即使15 h后也不能完全恢复;强光下40℃处理使PSⅠ和PSⅡ间的激发能分配严重偏离平衡状态.强光下40 ℃处理抑制了超氧化物歧化酶活性,加剧了O2、H2O2、丙二醛的产生,导致叶绿素和可溶性蛋白含量不断下降.说明强光下40℃高温胁迫对牡丹叶片光合机构造成了不可逆的破坏,而15℃低温处理对其光合机构的影响相对较弱.     

FTIR study of the primary electron donor of photosystem I (P700) revealing delocalization of the charge in P700(+) and localization of the triplet character in (3)P700.

The effect of global (15)N or (2)H labeling on the light-induced P700(+)/P700 FTIR difference spectra has been investigated in photosystem I samples from Synechocystis at 90 K. The small isotope-induced frequency shifts of the carbonyl modes observed in the P700(+)/P700 spectra are compared to those of isolated chlorophyll a. This comparison shows that bands at 1749 and 1733 cm(-)(1) and at 1697 and 1637 cm(-)(1), which upshift upon formation of P700(+), are candidates for the 10a-ester and 9-keto C=O groups of P700, respectively. A broad and relatively weak band peaking at 3300 cm(-)(1), which does not shift upon global labeling or (1)H-(2)H exchange, is ascribed to an electronic transition of P700(+), indicating that at least two chlorophyll a molecules (denoted P(1) and P(2)) participate in P700(+). Comparisons of the (3)P700/P700 FTIR difference spectrum at 90 K with spectra of triplet formation in isolated chlorophyll a or in RCs from photosystem II or purple bacteria identify the bands at 1733 and 1637 cm(-)(1), which downshift upon formation of (3)P700, as the 10a-ester and 9-keto C=O modes, respectively, of the half of P700 that bears the triplet (P(1)). Thus, while the P(2) carbonyls are free from interaction, both the 10a-ester and the 9-keto C=O of P(1) are hydrogen bonded and the latter group is drastically perturbed compared to chlorophyll a in solution. The Mg atoms of P(1) and P(2) appear to be five-coordinated. No localization of the triplet on the P(2) half of P700 is observed in the temperature range of 90-200 K. Upon P700 photooxidation, the 9-keto C=O bands of P(1) and P(2) upshift by almost the same amount, giving rise to the 1656(+)/1637(-) and 1717(+)/1697(-) cm(-)(1) differential signals, respectively. The relative amplitudes of these differential signals, as well as of those of the 10a-ester C=O modes, appear to be slightly dependent on sample orientation and temperature and on the organism used to generate the P700(+)/P700 spectrum. If it is assumed that the charge density on ring V of chlorophyll a, as measured by the perturbation of the 10a-ester or 9-keto C=O IR vibrations, mainly reflects the spin density on the two halves of the oxidized P700 special pair, a charge distribution ranging from 1:1 to 2:1 (in favor of P(2)) is deduced from the measurements presented here. The extreme downshift of the 9-keto C=O group of P(1), indicative of an unusually strong hydrogen bond, is discussed in relation with the models previously proposed for the PSI special pair.     

In vitro preparation and characterization of a 700 nm absorbing chlorophyll-water adduct according to the proposed primary molecular unit in photosynthesis

1. This study characterizes chlorophyll a-H₂O adducts in vitro in order to establish their generic relationship to the recently proposed [15, 18–20, 31] primary molecular adduct in photosynthesis. The effects of water titration and temperature on the absorption, fluorescence, excitation, and redox properties of the various in vitro chlorophyll a aggregate species are investigated.2. From fluorescence measurements, we conclude that the driest chlorophyll a sample contains an equimolar amount of water. This conclusion is consistent with earlier experimental work [2, 3, 14, 17, 31], and clarifies the origin of the controversial [15] Katz model [14] of chlorophyll a-H₂O interactions.3. With increasing water concentration or as the temperature is lowered below room temperature, the A-663 monohydrate chlorophyll a · H₂O (species absorbing at 663 nm) is favored at the expense of the A-678 anhydrous aggregate according to the equilibrium 2H₂O + chlorophyll a₂ai 2 chlorophyll a · H₂O. Under excess water conditions, A-663 is converted to A-743 (chlorophyll a · 2H₂O)_n.4. On slow sample cooling to T ? 200 °K, we observe the growth of A-700 at the expense of A-663. There is a direct correspondence between the increasing (decreasing) absorption by A-700 (A-663) and increasing (decreasing) fluorescence at 720 nm (664 nm).5. It is concluded that A-700 is most probably the dimer participating in the equilibrium 2 chlorophyll a · H₂O ai (chlorophyll a · H₂O)₂. The A-700 band consists of two exciton components (separated by ≈ 280 cm^?1) that are interpretable in terms of the dimeric origin of A-700.6. The deconvoluted A-700 absorption spectrum and the excitation spectrum of the 720 nm fluorescence are compared with the light-minus-dark spectra of P-700.7. It is found that A-700 is reversibly bleached by I₂ (E₀ = 0.54 V). The significance of this observation is discussed in terms of the redox properties of monomeric chlorophyll a and P-700.     

Growth at Low Temperature Mimics High-Light Acclimation in Chlorella vulgaris

Structural and functional alterations to the photosynthetic apparatus after growth at low temperature (5[deg]C) were investigated in the green alga Chlorella vulgaris Beijer. Cells grown at 5[deg]C had a 2-fold higher ratio of chlorophyll a/b, 5-fold lower chlorophyll content, and an increased xanthophyll content compared to cells grown at 27[deg]C even though growth irradiance was kept constant at 150 [mu]mol m-2 s-1. Concomitant with the increase in the chlorophyll a/b ratio was a lower abundance of light-harvesting polypeptides in 5[deg]C-grown cells as observed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and confirmed by western blotting.The differences in pigment composition were found to be alleviated within 12 h of transferring 5[deg]C-grown cells to 27[deg]C. Furthermore, exposure of 5[deg]C-grown cells to a 30-fold lower growth irradiance (5 [mu]mol m-2 s-1) resulted in pigment content and composition similar to that in cells grown at 27[deg]C and 150 [mu]mol m-2 s-1. Although both cell types exhibited similar measuring-temperature effects on CO2-saturated O2 evolution, 5[deg]C-grown cells exhibited light-saturated rates of O2 evolution that were 2.8-and 3.9-fold higher than 27[deg]C-grown cells measured at 27[deg]C and 5[deg]C, respectively. Steady-state chlorophyll a fluorescence indicated that the yield of photosystem II electron transport of 5[deg]C-grown cells was less temperature sensitive than that of 27[deg]C-grown cells. This appears to be due to an increased capacity to keep the primary, stable quinone electron acceptor of photosystem II (QA) oxidized at low temperature in 5[deg]C- compared with 27[deg]C-grown cells regardless of irradiance. We conclude that Chlorella acclimated to low temperature adjusts its photosynthetic apparatus in response to the excitation pressure on photosystem II and not to the absolute external irradiance. We suggest that the redox state of QA may act as a signal for this photosynthetic acclimation to low temperature in Chlorella.     

Excitation spectra for photosystem I and photosystem II in chloroplasts and the spectral characteristics of the distributions of quanta between the two photosystems.

The parameters listed in the title were determined within the context of a model for the photochemical apparatus of photosynthesis. The fluorescence of variable yield at 750 nm at -196 degrees C is due to energy transfer from Photosystem II to Photosystem I. Fluorescence excitation spectra were measured at -196 degrees C at the minimum, FO, level and the maximum, FM, level of the emission at 750 nm. The difference spectrum, FM-FO, which represents the excitation spectrum for FV is presented as a pure Photosystem II excitation spectrum. This spectrum shows a maximum at 677 nm, attributable to the antenna chlorophyll a of Photosystem II units, with a shoulder at 670 nm and a smaller maximum at 650 nm, presumably due to chlorophyll a and chlorophyll b of the light-harvesting chlorophyll complex. Fluoresence at the FO level at 750 nm can be considered in two parts; one part due to the fraction of absorbed quanta, alpha, which excites Photosystem I more-or-less directly and another part due to energy transfer from Photosystem II to Photosystem I. The latter contribution can be estimated from the ratio of FO/FV measured at 692 nm and the extent of FV at 750 nm. According to this procedure the excitation spectrum of Photosystem I at -196 degrees C was determined by subtracting 1/3 of the excitation spectrum of FV at 750 nm from the excitation spectrum of FO at 750 nm. The spectrum shows a relatively sharp maximum at 681 nm due to the antenna chlorophyll a of Photosystem I units with probably some energy transfer from the light-harvesting chlorophyll complex. The wavelength dependence of alpha was determined from fluorescence measurements at 692 and 750 nm at -196 degrees C. Alpha is constant to within a few percent from 400 to 680 nm, the maximum deviation being at 515 nm where alpha shows a broad maximum increasing from 0.30 to 0.34. At wavelengths between 680 and 700 nm, alpha increases to unity as Photosystem I becomes the dominant absorber in the photochemical apparatus.     

外源H2O2对小麦幼苗耐盐性的调节作用

以高原448小麦品种为材料,分别测定了4个处理(Hoagland营养液、Hoagland营养液 150 mmol/LNaCl、Hoagland营养液 150 mmol/L NaCl 10μmol/L H2O2和Hoagland营养液 10μmol/L H2O2)的小麦幼苗在第2、4、6、8天叶片叶绿素、丙二醛、可溶性糖和还原性谷胱甘肽含量.结果显示:外源H2O2提高了NaCl胁迫下4个时段小麦幼苗的叶绿素含量(8.27%、32.57%、10.19%、4.86%)及还原性谷胱甘肽含量(3.09%、23.97%、5.85%、2.11%),显著提高了可溶性糖含量(14.58%、8.43%、16.68%、5.8%,P<0.05),而显著降低了其丙二醛含量(17.53%、14.04%、4.75%、8.47%,P<0.05).外源H2O2(10μmol/L)使NaCl胁迫下叶绿素含量和还原性谷胱甘肽含量峰值提前,同时推迟了丙二醛峰值出现的时间.研究表明,外源H2O2通过提高叶片叶绿素、可溶性糖和还原性谷胱甘肽含量以有效地增强小麦幼苗的耐盐性.     

Quantum Yields and Rate Constants of Photochemical and Nonphotochemical Excitation Quenching (Experiment and Model)

Sunflower (Helianthus annuus L.), cotton (Gossypium hirsutum L.), tobacco (Nicotiana tabacum L.), sorghum (Sorghum bicolor Moench.), amaranth (Amaranthus cruentus L.), and cytochrome b6f complex-deficient transgenic tobacco leaves were used to test the response of plants exposed to differnt light intensities and CO2 concentrations before and after photoinhibition at 4000 [mu]mol photons m-2 s-1 and to thermoinhibition up to 45[deg]C. Quantum yields of photochemical and nonphotochemical excitation quenching (YP and YN) and the corresponding relative rate constants for excitation capture from the antenna-primary radical pair equilibrium system (k[prime]P and k[prime]N) were calculated from measured fluorescence parameters. The above treatments resulted in decreases in YP and K[prime]P and in approximately complementary increases in YN and K[prime]N under normal and inhibitory conditions. The results were reproduced by a mathematical model of electron/proton transport and O2 evolution/CO2 assimilation in photosynthesis based on budget equations for the intermediates of photosynthesis. Quantitative differences between model predictions and experiments are explainable, assuming that electron transport is organized into domains that contain relatively complete electron and proton transport chains (e.g. thylakoids). With the complementation that occurs between the photochemical and nonphotochemical excitation quenching, the regulatory system can constantly maintain the shortest lifetime of excitation necessary to avoid the formation of chlorophyll triplet states and singlet oxygen.     

Photosystem II Excitation Pressure and Development of Resistance to Photoinhibition (II. Adjustment of Photosynthetic Capacity in Winter Wheat and Winter Rye)

Winter wheat (Triticum aestivum L. cv Monopol), spring wheat (Triticum aestivum L. cv Katepwa), and winter rye (Secale cereale L. cv Musketeer) grown at 5[deg]C and moderate irradiance (250 [mu]mol m-2 s-1) (5/250) exhibit an increased tolerance to photoinhibition at low temperature in comparison to plants grown at 20[deg]C and 250 [mu]mol m-2 s-1 (20/250). However, 5/250 plants exhibited a higher photosystem II (PSII) excitation pressure (0.32-0.63) than 20/250 plants (0.18-0.21), measured as 1 - qP, the coefficient of photochemical quenching. Plants grown at 20[deg]C and a high irradiance (800 [mu]mol m-2 s-1) (20/800) also exhibited a high PSII excitation pressure (0.32-0.48). Similarly, plants grown at 20/800 exhibited a comparable tolerance to photoinhibition relative to plants grown at 5/250. In contrast to a recent report for Chlorella vulgaris (D.P. Maxwell, S. Falk, N.P.A. Huner [1995] Plant Physiol 107: 687-694), this tolerance to photoinhibition occurs in winter rye with minimal adjustment to polypeptides of the PSII light-harvesting complex, chlorophyll a/b ratios, or xanthophyll cycle carotenoids. However, Monopol winter wheat exhibited a 2.5-fold stimulation of sucrosephosphate synthase activity upon growth at 5/250, in comparison to Katepwa spring wheat. We demonstrate that low-temperature-induced tolerance to photoinhibition is not a low-temperature-growth effect per se but, instead, reflects increased photosynthetic capacity in response to elevated PSII excitation pressure, which may be modulated by either temperature or irradiance.     

Phosphorescence of protochlorophyll(ide) and chlorophyll(ide) in etiolated and greening bean leaves

The assignment is presented for the principal phosphorescence bands of protochlorophyll(ide), chlorophyllide and chlorophyll in etiolated and greening bean leaves measured at -196°C using a mechanical phosphoroscope. Protochlorophyll(ide) phosophorescence spectra in etiolated leaves consist of three bands with maxima at 870, 920 and 970 nm. Excitation spectra show that the 870 nm band belongs to the short wavelength protochlorophyll(ide), P627. The latter two bands correspond to the protochlorophyll(ide) forms, P637 and P650. The overall quantum yield for P650 phosphorescence in etiolated leaves is near to that in solutions of monomeric protochlorophyll, indicating a rather high efficiency of the protochlorophyll(ide) triplet state formation in frozen plant material. Short-term (2–20 min) illumination of etiolated leaves at the temperature range from -30 to 20°C leads to the appearance of new phosphorescence bands at about 990–1000 and 940 nm. Judging from excitation and emission spectra, the former band belongs to aggregated chlorophyllide, the latter one, to monomeric chlorophyll or chlorophyllide. This indicates that both monomeric and aggregated pigments are formed at this stage of leaf greening. After preillumination for 1 h at room temperature, chlorophyll phosphorescence predominates. The spectral maximum of this phosphorescence is at 955–960 nm, the lifetime is about 2 ms, and the maximum of the excitation spectrum lies at 668 nm. Further greening leads to a sharp drop of the chlorophyll phosphorescence intensity and to a shift of the phosphorescence maximum to 980 nm, while the phosphorescence lifetime and a maximum of the phosphorescence excitation spectrum remains unaltered. The data suggest that chlorophyll phosphorescence belongs to the short wavelength, newly synthesized chlorophyll, not bound to chloroplast carotenoids. Thus, the phosphorescence measurement can be efficiently used to study newly formed chlorophyll and its precursors in etiolated and greening leaves and to address various problems arising in the analysis of chlorophyll biosynthesis.Abbreviations Pchl protochlorophyll and protochlorophyllide - Chld chlorophyllide - Chl chlorophyll     

Xanthophyll cycle pool size and composition in relation to the nitrogen content of apple leaves

The objective of this study was to determine xanthophyll cycle pool size and composition in response to N status and their relationships to non-photochemical quenching in apple leaves. Bench-grafted Fuji/M.26 trees were fertilized with different N concentrations (0-20 mM) in a modified Hoagland's solution for 6 weeks to create a wide range of leaf N status (1-4.4 g m(-2)). Chlorophyll content, xanthophyll cycle pool size, lutein, total carotene, and neoxanthin on a leaf area basis all increased linearly with increasing leaf N. However, only the ratios of the xanthophyll cycle pool and of lutein to chlorophyll were higher in low N leaves than in high N leaves. Under high light at midday, both zeaxanthin (Z), expressed on a chlorophyll basis, and the percentage of the xanthophyll cycle pool present as Z, increased as leaf N decreased. Thermal dissipation of excitation energy, measured as non-photochemical quenching of chlorophyll fluorescence, was positively related to, whereas efficiency of excitation transfer and photosystem II quantum efficiency were negatively related to, Z, expressed on a chlorophyll basis or on a xanthophyll cycle pool basis. It is concluded that both xanthophyll cycle pool size (on a chlorophyll basis) and conversion of violaxanthin to zeaxanthin are enhanced in response to N limitation to dissipate excessive absorbed light under high irradiance.     

A resonance Raman study on a reaction intermediate of Pseudomonas L-phenylalanine oxidase (deaminating and decarboxylating).

Resonance Raman (RR) spectra of purple intermediates of L-phenylalanine oxidase (PAO) with non-labeled and isotopically labeled phenylalanines as substrates, i.e., [1-13C], [2-13C], [ring-U-13C6], and [15N]phenylalanines, were measured with excitation at 632.8 nm within the broad absorption band around 540 nm. The spectra obtained resemble those of purple intermediates of D-amino acid oxidase (DAO). The isotope effects on the 1,665 cm-1 band with [15N] or [2-13C]phenylalanine indicate that the band is due to the C = N stretching mode of an imino acid derived from phenylalanine, i.e., alpha-imino-beta-phenylpropionate. The intense band at 1,389 cm-1 is contributed to by the CO2- symmetric stretching and C-CO2- stretching modes of alpha-imino-beta-phenylpropionate. The 1,602 cm-1 band, which does not shift upon isotopic substitution of phenylalanine, corresponds to the 1,605 cm-1 band of DAO purple intermediates and was assigned to a vibrational mode associated with the C(10a) = C(4a) - C(4) = O moiety of reduced flavin. These results confirm that PAO purple intermediates consist of the reduced enzyme and an imino acid derived from a substrate, and suggest that the plane defined by C(10a) = C(4a) - C(4) = O of reduced flavin and the plane containing H2+N = C - CO2- of an imino acid are arranged in close contact to each other, generating a charge-transfer interaction.     

Effect of the spectral composition of light on the mechanism of reaction of chlorophyll oxidation]

Photochemical oxidation of chlorophyll "a" on excitation of the pigment in different spectrum region (400-800 nm) was studied by flash photolysis. The absorption spectrum of chlorophyll cation-radical Chl+ was obtained and the values of extinction coefficient found. An attempt was made of photochemical generation of dication form of chlorophyll. Thermodynamical calculation supporting the possibility of the following reaction is presented: (Chl+)*+Ae -- Chl2"Ae. The absence of Chl2+ is explained by a short life time of the excited cation-radical of chlorophyll (Chl+)*. The effect of the wave length of excited light on the kinetics of the decay of chlorophyll cation-radical is studied. It is shown that on excitation of chlorophyll "a" with white light the life time of Chl+ decreases and its death is described by an equation of first order, which is explained by the formation of ion-radical of the electron acceptor resulting from the direct excitation of Ae.