Contribution of hydroxycinnamates and flavonoids to epidermal shielding of UV‐A and UV‐B radiation in developing rye primary leaves as assessed by ultraviolet‐induced chlorophyll fluorescence measurements

Epidermally located ultraviolet (UV)‐absorbing phenolic compounds, flavonoids and hydroxycinnamic acid esters (HCAs), can shield the underlying tissues in plants against harmful UV‐radiation. The relative importance of the two different classes of phenolic compounds for UV‐screening was a matter of recent debate. Using a non‐invasive method based on chlorophyll fluorescence measurements to estimate epidermal UV transmittance, the relationship between epidermal UV shielding and the content of the two different groups of secondary phenolic compounds in the epidermal layers and the underlying photosynthetic mesophyll of developing rye primary leaves grown under supplementary UV‐B radiation was investigated. From the fourth to the tenth day after sowing, epidermally located flavonoids increased in an age‐ and irradiation‐dependent manner, whereas mesophyll flavonoids and epidermal HCAs, mainly ferulic acid and p‐coumaric acid esters, were constitutively present and did not vary in their contents over the observed time period. There was an excellent correlation between epidermal UV‐A and UV‐B absorbances as assessed by chlorophyll fluorescence measurements and contents of epidermal flavonoids. However, HCAs showed an additional contribution to UV‐B shielding. In contrast, mesophyll flavonoids did not seem to play a respective role. When absorbances of the abaxial and adaxial epidermal layers were compared, it became apparent that in fully expanded primary leaves epidermal tissues from both sides were equally effective in absorption of UV‐radiation. However, the earlier and more UV‐exposed abaxial epidermis of young unrolling leaves showed a significantly higher absorption. It is shown that in early stages of development the epidermal HCAs are the dominant UV‐B protective compounds of the primary leaf. This function is increasingly replaced by the epidermal flavonoids during leaf development and acclimation. The application of chlorophyll fluorescence measurements has been proven to be a useful tool for estimating relative contents of these compounds in epidermal tissue.     

Leaf expansion and development of PHOTOSYNTHETIC CAPACITY AND PIGMENTS IN Liquidambar styraciflua (Hamamelidaceae)—EFFECTS OF UV-B RADIATION

In order to perform their functions as photosynthetic organs, leaves must cope with excess heat and potentially damaging ultraviolet radiation. Possible increases in the UV-B portion of the solar spectrum may place an additional burden on leaves, and this could be particularly important for young expanding leaves with poorly developed UV-B defense mechanisms. We evaluated the effects of supplemental UV-B radiation on leaf expansion and the development of photosynthetic capacity and pigments in sweetgum (Liquidambar styraciflua L.) seedlings. Seedlings were grown in the field under either ambient or ambient plus 3 or 5.0 kJ of biologically effective supplemental UV-B radiation. Although final leaf size was unaffected, the rate of leaf elongation and accumulation of leaf area was slower in leaves exposed to the lower supplemental UV-B irradiance. In contrast, chlorophyll accumulation and the development of photosynthetic capacity was more rapid in plants exposed to the higher, compared to the lower supplemental UV-B irradiance. The accumulation of anthocyanins and other putative flavonoids or UV-absorbing compounds was scarcely affected by exposure to supplemental UV-B radiation. These results suggest that the UV-B portion of the solar spectrum may, in the absence of gross affects on biomass, exert subtle influences on leaf ontogeny and the development of photosynthetic pigments and capacity in sweetgum.     

Developmental changes in spatial distribution of in vivo fluorescence and epidermal UV absorbance over Quercus petraea leaves

Background and Aims

Epidermal phenolic compounds (mainly flavonoids) constitute a vital screen that protects the leaf from damage by natural ultraviolet (UV) radiation. The effectiveness of epidermal UV-screening depends on leaf anatomy, the content of UV-screening compounds and their spatial uniformity over the leaf area. To investigate in vivo the spatial pattern of the epidermal UV-screen during leaf development, a fluorescence imaging method was developed to map the epidermal UV-absorbance at a microscopic scale. This study was done on oak (Quercus petraea) leaves that were used as a model of woody dicotyledonous leaves.

Methods

The leaf development of 2-year-old trees, grown outdoors, was monitored, at a macroscopic scale, by in vivo measurements of chlorophyll content per unit area and epidermal UV-absorbance using two optical leaf-clip meters. The distribution of pigments within leaves was assessed in vivo spectroscopically. The microscopic images of UV-induced fluorescence and UV-absorbance acquired in vivo during leaf development were interpreted from spectral characteristics of leaves.

Key Results

At a macroscopic scale, epidermal UV-absorbance was high on the upper leaf side during leaf development, while it increased on the lower leaf side during leaf expansion and reached the adaxial value at maturity. At a microscopic scale, in immature leaves, for both leaf sides, the spatial distribution of epidermal UV-absorbance was heterogeneous, with a pattern depending on the flavonoid content of vacuoles in developing epidermal cells. At maturity, epidermal UV-absorbance was uniform.

Conclusions

The spatial pattern of epidermal UV-screen over the area of oak leaves is related to leaf anatomy during development. In vivo spectroscopy and fluorescence imaging of the leaf surface showed the distribution of pigments within the leaf and hence can provide a tool to monitor optically the leaf development in nature.Key words: Blue-green fluorescence, chlorophyll fluorescence, epidermis, flavonoids, leaf development, microscopic imaging, polyphenols, Quercus petraea     

Spectral reflectance indices and pigment functions during leaf ontogenesis in six subtropical landscape plants

Pigment combinations are regulated during leaf ontogenesis. To better understand pigment function, alterations in chlorophyll, carotenoid and anthocyanin concentrations were investigated during different leaf development stages in six subtropical landscape plants, namely Ixora chinensis Lam, Camellia japonica Linn, Eugenia oleina Wight, Mangifera indica L., Osmanthus fragrans Lowr and Saraca dives Pierre. High concentrations of anthocyanin were associated with reduced chlorophyll in juvenile leaves. As leaves developed, the photosynthetic pigments (chlorophyll and carotenoid) of all six species increased while anthocyanin concentration declined. Chlorophyll fluorescence imaging of Φ_PSII (effective quantum yield of PSII) and of NPQ (non-photochemical fluorescence quenching) and determination of electron transport rate-rapid light curve (RLC) showed that maximum ETR (leaf electron transport rate), Φ_PSII and the saturation point in RLC increased during leaf development but declined as they aged. Juvenile leaves displayed higher values of NPQ and Car/Chl ratios than leaves at other developmental stages. Leaf reflectance spectra (400–800 nm) were measured to provide an in vivo non-destructive assessment of pigments in leaves during ontogenesis. Four reflectance indices, related to pigment characters, were compared with data obtained quantitatively from biochemical analysis. The results showed that the ARI (anthocyanin reflectance index) was linearly correlated to anthocyanin concentration in juvenile leaves, while a positive correlation of Chl NDI (chlorophyll normalized difference vegetation index) to chlorophyll a concentration was species dependent. Photosynthetic reflectance index was not closely related to Car/Chl ratio, while a structural-independent pigment index was not greatly altered by leaf development or species. Accordingly, it is suggested that the high concentration of anthocyanin, higher NPQ and Car/Chl ratio in juvenile leaves are important functional responses to cope with high radiation when the photosynthetic apparatus is not fully developed. Another two leaf reflectance indices, ARI and Chl NDI, are valuable for in vivo pigment evaluation during leaf development.     

Light-induced accumulation of ortho-dihydroxylated flavonoids as non-destructively monitored by chlorophyll fluorescence excitation techniques

Chlorophyll fluorescence excitation techniques have been used in plant science for more than one decade to non-destructively estimate phenolic compounds in the epidermal cell layer. These techniques have been used here to evaluate the effect of different light intensities and spectral quality on the accumulation of ortho-dihydroxylated flavonoids in Phyllirea latifolia L., Myrtus communis L. and Ligustrum vulgare L. In a first experiment, chlorophyll fluorescence excitation spectra were measured (with a double arm optical fiber bundle connected to a spectrofluorimeter) on the adaxial and abaxial leaf surfaces of container-grown P. latifolia and M. communis exposed to 20% or 100% full sunlight. Differences in epidermal absorption spectra (referred to as epidermal absorption spectra throughout the paper) were then calculated from the relative chlorophyll fluorescence excitation spectra. This allowed comparing the content of UV-absorbing compounds between differentially irradiated leaves as well as between adaxial and abaxial epidermal layers. The absorption spectra were characterized by a band centered at 360–380 nm, which was greater in sun than in shade leaves and in the adaxial than in the abaxial surfaces, irrespective of species. Based upon HPLC-DAD and HPLC–MS analyses of leaf extracts and UV-spectral features of individual flavonoids, we conclude that quercetin, luteolin and myricetin derivatives were responsible for the observed light-induced changes in the spectral features of examined tissues.In a second experiment, we grew L. vulgare potted plants at 30% or 85% full sunlight in the presence or in the absence of UV radiation. We measured the absorbance characteristics at 370 nm of three leaf-pairs located in the apical portion of each shoot, using a portable fluorimetric sensor, the Multiplex^®. This allowed estimating, non-destructively, an index closely related to the concentration of epidermal flavonols and flavones having an ortho-dihydroxyl substitution in the B-ring. This index was greater in leaves growing at 85% than at 30% sunlight, irrespective of UV irradiance. When plants acclimated for 3 weeks to 30% sunlight, in the presence or in the absence of UV irradiance, were transferred to 85% sunlight, the flavonoid index exponentially increased, reaching a maximum within 10 days.On the whole, our experiments conclusively show that light-responsive flavonoids are mostly the dihydroxy B-ring-substituted structures. These flavonoids are the most effective, among the wide array of flavonoid structures, in preventing the generation and scavenging reactive oxygen species. As a consequence, we suggest that flavonoids do not merely serve as UV-screening agents, but behave as ROS-detoxifying agents in the mechanisms of photoprotection.     

Uptake of diuron and concomitant loss of photosynthetic activity in leaves as visualized by imaging the red chlorophyll fluorescence

The principles of the chlorophyll (Chl) fluorescence induction kinetics (known as Kautsky effect) and their change by the photosystem II herbicide diuron are presented together with the Chl fluorescence emission spectra of a normal and diuron-inhibited leaf. By imaging the Chl fluorescence emission of green leaves the successive uptake of diuron and the concomitant loss of photosynthetic quantum conversion from the leaf base to the leaf tip are documented.     

Measurement of leaf epidermal transmittance of UV radiation by chlorophyll fluorescence

In higher plants one of the important functions of the leaf epidermis is the effective screening of ultraviolet-B (280–320 nm, UV-B) radiation, due mostly to phenolic compounds. The assessment of the contribution of this function is necessary for an evaluation of the impact of increasing UV-B radiation. A method is proposed to estimate epidermal transmittance on the basis of chlorophyll fluorescence measurements. Fluorescence of chlorophyll induced by UV-A (320–400 nm, measuring beam centered at 366 nm, half band width 32 nm) or UV-B (measuring beam centered at 314 nm, half band width 18 nm) is compared to that induced by a blue-green measuring light (475 nm, half band width 140 nm). It is shown that the ratios of UV-and blue-green (BG)-induced fluorescence, F(UV-A)/F(BG) and F(UV-B)/F(BG), are relatively constant among leaf samples of various species ( Vicia faba, Spinacia oleracea, Rumex scutatus ) from which the epidermis was removed. In epidermis-free leaves no significant differences were found between adaxial and abaxial leaf sides, suggesting that leaf structure has negligible influence on the F(UV)/F(BG) ratios. On the other hand, fluorescence excitation ratios varied over a vast range when intact leaves from different species and habitats were investigated. Ratios were low in sun leaves and relatively high in shade- and greenhouse-grown leaves. By relating these results to those obtained with epidermis-free leaves, epidermal transmittances for UV-B radiation could be estimated, with values ranging between 1 and 45%. The data demonstrate a large adaptability of epidermal UV-A and UV-B transmittance in higher plants. The proposed method may prove a versatile and relatively simple tool for investigating epidermal UV transmittance complementing established methods.     

Leaf development in Ricinus communis during drought stress: dynamics of growth processes, of cellular structure and of sink-source transition

Dicot leaf growth is characterized by partly transient tip-to-base gradients of growth processes, structure and function. These gradients develop dynamically and interact with dynamically developing stress conditions like drought. In Ricinus communis plants growing under well-watered and drought conditions growth rates peaked during the late night and minimal values occurred in the late afternoon. During this diurnal course the leaf base always showed much higher rates than the leaf tip. The amplitude of this diurnal course decreased when leaves approached maturity and during drought stress without any significant alteration of the diurnal pattern and it increased during the first days after rewatering. Unique relationships between leaf size and cytological structure were observed. This provided the framework for the analysis of changes in assimilation, transpiration and dark respiration, chlorophyll, protein, carbohydrate, and amino acid concentrations, and of activities of sink-source-related enzymes at the leaf tip and base during leaf development in well-watered and drought-stressed plants. Gas exchange was dominated by physiological rather than by anatomical properties (stomatal density). Tip-to-base gradients in carbohydrate concentrations per dry weight and sink-source-related enzymes were absent, whereas significant gradients were found in amino acid concentrations per dry weight. During drought stress, growing leaves developed source function at smaller leaf size, before specific physiological adaptations to drought occurred. The relevance of the developmental status of individual leaves for the drought-stress response and of the structural changes for the biochemical composition changes is discussed.     

叶角、光呼吸和热耗散协同作用减轻大豆幼叶光抑制

研究了大豆叶片逐步展开过程中的色素组成、气体交换、荧光动力学以及叶片角度等特性。随着叶片展开程度的增加 ,叶绿素含量和叶绿素 a/ b比值增加 ;光合速率 (Pn)也增加 ,揭示叶片展开过程中光合机构是逐步完善的。自然状态下 ,不同展开程度的叶片均未发生明显的光抑制 ;但将叶片平展并暴露在 12 0 0μmol/ (m2 · s)光下时幼叶发生严重的光抑制 ,伴随叶面积的增加光抑制程度减轻。强光下 ,尽管幼叶光呼吸 (Pr)的测定值较低 ,但幼叶光呼吸与总光合之比 (Pr/ Pm)较高。将叶片平展置于强光下时 ,幼叶的实际光化学效率 (ΦPSII)明显下调 ,非光化学猝灭 (NPQ)大幅增加 ;幼叶叶黄素库较大 ,光下积累较多的脱环氧化组分 ,揭示幼叶依赖叶黄素循环的热耗散增强。自然条件下测量叶片角度 ,观察到在叶片展开过程中叶柄夹角逐渐增加 ;日动态过程中幼叶的悬挂角随光强增加而明显减小 ,完全展开叶的悬挂角变化幅度很小。叶片角度的变化使实际照射到幼叶叶表的光强减少。推测较强的光呼吸、依赖叶黄素循环的热耗散以及较大的叶角变化可能是自然状态下幼叶未发生严重光抑制的原因     

温室栽培对软枣猕猴桃叶片形态及叶绿素荧光特性的影响

【目的】深入研究温室栽培条件下软枣猕猴桃叶片的形态与叶绿素荧光特性,探明温室与露地栽培环境下叶片光形态建成的差异。【方法】以软枣猕猴桃品种‘佳绿’和‘魁绿’的5年生植株为试材,测定温室及露地栽培条件下不同叶龄叶片的叶绿素相对含量及叶绿素荧光参数,分析这些参数在温室与露地栽培条件下的差异。【结果】温室与露地栽培软枣猕猴桃的光形态建成均处于1~40 d叶龄间,不同栽培条件下的同一品种荧光特性趋于一致;花期后,不同栽培条件下的同一品种荧光特性差异较大。温室栽培的软枣猕猴桃叶片叶面积较大,叶绿素b含量较高,趋于阴生叶特性,以吸收光能为基础的性能指数等显著低于露地栽培,对光能的吸收捕获能力较强,但热耗散较高,用于电子传递的能量低于露地栽培,叶绿素荧光参数表现出对环境的适应性变化。【结论】软枣猕猴桃叶片的光形态建成时间为展叶后1~40 d,即在花期之前完成,不同栽培环境无明显差异;温室栽培软枣猕猴桃叶片的光形态建成与露地栽培具显著差异,叶面积明显增大;温室栽培在一定程度上改变了叶片的叶绿素荧光特性,降低了光合性能。     

Intra-leaf gradients of photoinhibition induced by different color lights: implications for the dual mechanisms of photoinhibition and for the application of conventional chlorophyll fluorometers

? We studied how different color lights cause gradients of photoinhibition within a leaf, to attempt to resolve the controversy of whether photon absorption by chlorophyll or by manganese (Mn) is the primary cause of photoinhibition, as suggested by the excess-energy hypothesis or the two-step hypothesis, respectively. ? Lincomycin-treated leaf discs were photoinhibited by white, blue, green or red light. Combining a microfiber fluorometer, a fiber-thinning technique and a micro-manipulator enabled us to measure the chlorophyll fluorescence signals within a leaf. Photoinhibition gradients were also compared with results from various conventional fluorometers to estimate their depth of signal detection. ? The severity of photoinhibition was in the descending order of blue, red and green light near the adaxial surface, and in the descending order of blue, green and red light in the deeper tissue, which correlated with the chlorophyll and the Mn absorption spectrums, respectively. These results cannot be explained by either hypothesis alone. ? These data strongly suggest that both the excess-energy and the two-step mechanisms occur in photoinhibition, and fluorometers with red or blue measuring light give overestimated or underestimated F(v)/F(m) values of photoinhibited leaves compared with the whole tissue average, respectively; that is, they measured deeper or shallower leaf tissue, respectively.     

Photosynthesis and Chlorophyll Fluorescence Characteristics in Relationship to Changes in Pigment and Element Composition of Leaves of Platanus occidentalis L. during Autumnal Leaf Senescence

The loss of chlorophyll and total leaf nitrogen during autumnal senescence of leaves from the deciduous tree Platanus occidentalis L. was accompanied by a marked decline in the photosynthetic capacity of O₂ evolution on a leaf area basis. When expressed on a chlorophyll basis, however, the capacity for light-and CO₂-saturated O₂ evolution did not decline, but rather increased as leaf chlorophyll content decreased. The photon yield of O₂ evolution in white light (400-700 nanometers) declined markedly with decreases in leaf chlorophyll content below 150 milligrams of chlorophyll per square meter on both an incident and an absorbed basis, due largely to the absorption of light by nonphotosynthetic pigments which were not degraded as rapidly as the chlorophylls. Photon yields measured in, and corrected for the absorptance of, red light (630-700 nanometers) exhibited little change with the loss of chlorophyll. Furthermore, PSII photochemical efficiency, as determined from chlorophyll fluorescence, remained high, and the chlorophyll a/b ratio exhibited no decline except in leaves with extremely low chlorophyll contents. These data indicate that the efficiency for photochemical energy conversion of the remaining functional components was maintained at a high level during the natural course of autumnal senescence, and are consistent with previous studies which have characterized leaf senescence as being a controlled process. The loss of chlorophyll during senescence was also accompanied by a decline in fluorescence emanating from PSI, whereas there was little change in PSII fluorescence (measured at 77 Kelvin), presumably due to decreased reabsorption of PSII fluorescence by chlorophyll. Nitrogen was the only element examined to exhibit a decline with senescence on a dry weight basis. However, on a leaf area basis, all elements (C, Ca, K, Mg, N, P, S) declined in senescent leaves, although the contents of sulfur and calcium, which are not easily retranslocated, decreased to the smallest extent.     

不同叶色三叶海棠叶片成色色素分析

结合民族植物学和药理学的研究方法,对西双版纳地区傣族、哈尼族和基诺族等3个少数民族民间利用番石榴（Psidium guajava）、余甘子（Phyllanthus emblica）和水柳（Homonoia riparia）的传统知识进行调查研究及体外抗菌活性实验。结果表明：番石榴和余甘子在村寨中较为常见,当地少数民族将其种植于庭院中,常作为果蔬食用,食用番石榴嫩叶可缓解拉肚子的症状,治疗腹痛、腹泻。水柳生长在水边,傣族会将其叶作为腌酸鱼的配料之一。根据文献记载,番石榴、余甘子和水柳的叶部位作为药使用时,常煎水外洗,治疗皮肤瘙痒。对这3种药用植物叶部位采用80%乙醇浸泡制备的提取物进行体外抗菌实验,结果显示番石榴、余甘子和水柳3种药用植物对金黄色葡萄球菌和大肠埃希菌均有较好的抑菌和杀菌活性,其最小抑菌浓度MIC在98~390 μg·mL 1之间,最小杀菌浓度MBC在98~781 μg·mL 1之间。番石榴和水柳叶对铜绿假单胞菌有一定抑菌和杀菌活性,其MIC和MBC范围均为6 250~12 500 μg·mL 1。由此可见,这3种药用植物的民间利用具有一定的合理性和药用开发价值。     

Plastid differentiation and chlorophyll biosynthesis in different leaf layers of white cabbage (Brassica oleracea cv. capitata)

The contents of protochlorophyllide, protochlorophyll and chlorophyll together with the native arrangements of the pigments and the plastid ultrastructure were studied in different leaf layers of white cabbage (Brassica oleracea cv. capitata) using absorption, 77 K fluorescence spectroscopy and transmission electron microscopy. The developmental stage of the leaves was determined using the differentiation of the stoma complexes as seen by scanning electron microscopy and light microscopy. The pigment content showed a gradual decrease from the outer leaf layer towards the central leaves. The innermost leaves were in a primordial stage in many aspects; they were large but had typical proplastids with few simple inner membranes, and contained protochlorophyllide and its esters in a 2 : 1 ratio and no chlorophyll. Short‐wavelength, not flash‐photoactive protochlorophyllide and/or protochlorophyll forms emitting at 629 and 636 nm were dominant in the innermost leaves. These leaves also had small amounts of the 644 and 654 nm emitting, flash‐photoactive protochlorophyllide forms. Rarely prolamellar bodies were observed in this layer. The outermost leaves had the usual characteristics of fully developed green leaves. The intermediary layers contained chlorophyll a and chlorophyll b besides the protochlorophyll(ide) pigments and had various intermediary developmental stages. Spectroscopically two types of intermediary leaves could be distinguished: one with only a 680 nm emitting chlorophyll a form and a second with bands at 685, 695 and 730 nm, corresponding to chlorophyll–protein complexes of green leaves. In these leaves, a large variety of chloroplasts were found. The data of this work show that etioplasts, etio‐chloroplasts or chloro‐etioplasts as well as etiolated leaves do exist in the nature and not only under laboratory conditions. The specificity of cabbage leaves compared with those of dark‐grown seedlings is the retained primordial or intermediary developmental stage of leaves in the inner layers for very long (even for a few month) period. This opens new developmental routes leading to formation of specially developed plastids in the various cabbage leaf layers. The study of these plastids provided new information for a better understanding of the plastid differentiation and the greening process .     

Appearance of a State 1 – State 2 transition during chloroplast development in the wheat leaf: Energetic and structural considerations

The ability of developing chloroplasts to dynamically regulate the distribution of excitation energy between photosystem 1 and photosystem 2, and thus perform a State 1 – State 2 transition, was examined from analyses of chlorophyll fluorescence kinetics in 4- and 8-day-old Triticum aestivum L. cv. Maris Dove leaves grown under a diurnal light regime. Chloroplasts at all stages of development in the two leaf systems could undergo a State 1 – State 2 transition, except those found in the basal 0.5 cm of the 4-day-old leaf. The ability to physiologically modify the excitation energy distribution between the chlorophyll matrices of the two photosystems developed after the development of mature, fully photochemically competent photosystem 2 units and the appearance of excitation energy transfer between photosystem 2 and photosystem 1. Also, changes in the degree of energetic interaction between the two photosystems, in vivo rates of electron transport and the chlorophyll a/b ratio could not be correlated with the appearance of a State 1 – State 2 transition. Ultrastructural studies demonstrated a 32% increase in the degree of thylakoid appression in chloroplasts at the base of the 8-day-old leaf compared to the situation in the basal 0.5 cm of the 4-day-old leaf. This difference in thylakoid stacking can account for the differing abilities of these two tissues to perform a State 1 – State 2 transition when considered in the context of the distribution of the two photosystems within appressed and non-appressed regions of thylakoid membranes.     

Sinapate esters provide greater UV-B attenuation than flavonoids in Arabidopsis thaliana (Brassicaceae)

Mutants affected in flavonoid (tt4) or sinapate ester (fah1) biosynthesis were used to assess the relative importance of these phenolic UV photoprotectants in Arabidopsis. Flavonoid and sinapate ester absorption was more specific for UV-B than major nonphenolic chromophores in crude extracts. A new method of evaluating phenolic UV-B attenuation was developed using fluorescence analysis. When excited by UV-B, sinapate ester containing leaves and cotyledons had enhanced sinapate ester fluorescence and reduced chlorophyll fluorescence relative to those without sinapate esters. Although fluorescence analysis gave no evidence of UV-B attenuation by flavonoids, enhanced chlorophyll and protein loss were observed upon UV-B exposure in flavonoid-deficient leaves, suggesting they have another mechanism of UV-B protection. The hydroxycinnamates have been largely ignored as UV-B attenuating pigments, and the results indicate that greater attention should be paid to their role in attenuating UV-B.     

THE CHLOROPHYLL-PROTEIN COMPOUND OF THE GREEN LEAF

1. Aqueous extracts of spinach and Aspidistra leaves yield highly opalescent preparations which are not in true solution. Such extracts differ markedly from colloidal chlorophyll in their spectrum and fluorescence. The differences between the green leaf pigment and chlorophyll in organic solvents are shown to be due to combination of chlorophyll with protein in the leaf. 2. The effect of some agents on extracts of the chlorophyll-protein compound has been investigated. Both strong acid and alkali modify the absorption spectrum, acid converting the compound to the phaeophytin derivative and alkali saponifying the esterified groups of chlorophyll. Even weakly acid solutions (pH 4.5) denature the protein. Heating denatures the protein and modifies the absorption spectrum and fluorescence as earlier described for the intact leaf. The protein is denatured by drying. Low concentrations of alcohol or acetone precipitate and denature the protein; higher concentrations cause dissociation liberating the pigments. 3. Detergents such as digitonin, bile salts, and sodium desoxycholate clarify the leaf extracts but denature the protein changing the spectrum and other properties. 4. Inhibiting agents of photosynthesis are without effect on the absorption spectrum of the chlorophyll-protein compound. 5. The red absorption band of chlorophyll possesses the same extinction value in organic solvents such as ether or petroleum ether, and in aqueous leaf extracts clarified by digitonin although the band positions are different. Using previously determined values of the extinction coefficients of purified chlorophylls a and b, the chlorophyll content of the leaf extracts may be estimated spectrophotometrically. 6. It was found that the average chlorophyll content of the purified chloroplasts was 7.86 per cent. The protein content was 46.5 per cent yielding an average value of 16.1 parts per 100 parts of protein. This corresponds to a chlorophyll content of three molecules of chlorophyll a and one of chlorophyll bfor the Svedberg unit of 17,500. It is suggested that this may represent a definite combining ratio of a and b in the protein molecule.     

增强UV-B辐射和氮对谷子叶光合色素及非酶促保护物质的影响

以谷子(Setaria italica(L)Beauv.)为对象,从拔节期开始持续给予低氮(1.875 mmol/L)和高氮(15 mmol/L)两种氮供应条件并从抽穗期开始进行26 d两种强度(4.29、7.12 kJ·m-2·d-1)的增强UV-B辐射处理,研究了谷子叶中光合色素含量、类黄酮含量和苯丙氨酸解氨酶(PAL)活性的变化.结果表明:与高氮供应条件相比,低氮供应条件明显降低了谷子叶中光合色素含量但提高了类胡萝卜素/叶绿素含量比值;在开花期中段和灌浆期中段,高氮供应条件下谷子叶中光合色素含量对增强UV-B辐射比低氮供应条件下的谷子更敏感.从灌浆期开始到处理结束,两种影响因子对谷子叶中类黄酮含量均有显著影响,增强UV-B辐射导致谷子叶中类黄酮含量逐渐升高,且相同增强UV-B辐射强度下低氮供应条件下的谷子叶中类黄酮含量明显高于高氮供应条件下的谷子.谷子叶中PAL活性对两种影响因子的响应较类黄酮含量更加敏感,低氮供应条件使谷子叶中PAL活性明显提高.结合上述指标的相关性分析结果可知,低氮供应条件加强了处于繁殖期主要阶段的谷子叶中类黄酮的积累,并使谷子叶中的类胡萝卜素/叶绿素含量比值明显提高,进而有助于维持谷子叶中光合色素含量在增强UV-B辐射条件下的相对稳定性,对植株抵抗UV-B辐射伤害有利.     

Effect of transient accumulation of anthocyanin on leaf development and photoprotection of Fagopyrum dibotrys mutant

Changes in pigments contents, leaf area, leaf dry mass per unit area (LMA), photosynthetic rate and chlorophyll a fluorescence were investigated in developing leaves of Fagopyrum dibotrys Hara. mutant. Anthocyanins transiently accumulate below the upper epidermis during leaf ontogeny of this mutant. Red leaves possessed lower Chl content, LMA, photosynthetic rate and apparent carboxylation efficiency than green leaves. However, content of anthocyanins declined and above mentioned parameters increased during further leaf development. In both red and green leaves, chronic photoinhibition did not take place according to variable to maximum chlorophyll fluorescence ratio (F_v/F_m). Red leaves had higher non-photochemical quenching (NPQ) and higher PS 2 efficiency.     

Effects of UV-B radiation on photosynthetic reactions in Rhizophora apiculata

Influences of UV-B radiation on Rhizophora apiculata were studied in terms of chlorophylls, their presence in protein complexes of the chloroplast, PS I and PS II photochemical activities, in vitro absorption spectrum of the chloroplast, in vivo leaf fluorescence and UV absorbing compounds. The seedlings were exposed to the various levels of UV-B radiations, equivalent to 0 (control), 10, 20, 30 and 40% stratospheric ozone depletion of the study area. The low doses of UV-B (10 and 20%) increased the reaction centre chlorophyll (10 and 8%) and activities of PS-I (98 and 39%) and PS-II (77 and 38%) respectively; whereas, 30 and 40% UV-B treatments decreased the reaction centre chlorophylls by 11 and 33% and PS II activity by 0 and 20%; while PS I activity did not show any inhibitory effect. Chloroplasts isolated from control and 10% UV-B treated plants exhibited the same level of absorption at 676 nm. In vivo leaf fluorescence was found to be diminished with UV-B radiation and at the 10% UV-B, variable fluorescence was promoted significantly by 10%. The content of UV-absorbing compounds was progressively enhanced with doses of UV-B radiation along with higher absorption at 276 and 330 nm.