Relationships between leaf chlorophyll content and spectral reflectance and algorithms for non-destructive chlorophyll assessment in higher plant leaves

Leaf chlorophyll content provides valuable information about physiological status of plants. Reflectance measurement makes it possible to quickly and non-destructively assess, in situ, the chlorophyll content in leaves. Our objective was to investigate the spectral behavior of the relationship between reflectance and chlorophyll content and to develop a technique for non-destructive chlorophyll estimation in leaves with a wide range of pigment content and composition using reflectance in a few broad spectral bands. Spectral reflectance of maple, chestnut, wild vine and beech leaves in a wide range of pigment content and composition was investigated. It was shown that reciprocal reflectance (R lambda)-1 in the spectral range lambda from 520 to 550 nm and 695 to 705 nm related closely to the total chlorophyll content in leaves of all species. Subtraction of near infra-red reciprocal reflectance, (RNIR)-1, from (R lambda)-1 made index [(R lambda)(-1)-(RNIR)-1] linearly proportional to the total chlorophyll content in spectral ranges lambda from 525 to 555 nm and from 695 to 725 nm with coefficient of determination r2 > 0.94. To adjust for differences in leaf structure, the product of the latter index and NIR reflectance [(R lambda)(-1)-(RNIR)-1]*(RNIR) was used; this further increased the accuracy of the chlorophyll estimation in the range lambda from 520 to 585 nm and from 695 to 740 nm. Two independent data sets were used to validate the developed algorithms. The root mean square error of the chlorophyll prediction did not exceed 50 mumol/m2 in leaves with total chlorophyll ranged from 1 to 830 mumol/m2.     

Extremely rapid effects of polyunsaturated fatty acids and N-acetylglucosamine on free-radical metabolism in cultured potato plant cells

The effects of arachidonic and linoleic acids, separately and in co-operation with N-acetylglucosamine oligomers, on Solanum tuberosum plant suspension cell cultures were investigated in terms of the fluorescent oxygen-activated-species-sensitive dye 2',7'-dichlorofluorescin diacetate. The inductors used triggered extremely rapid (within 2-10 min) generation of H(2)O(2) in the cells; the majority was expressed in cultures treated with combined polyunsaturated fatty acid and N-acetylglucosamine oligomers. The stimulation of free-radical generation may be related to defensive mechanisms modulating a plant-pathogenic-micro-organism interaction.     

Bright monomeric red fluorescent protein with an extended fluorescence lifetime

Fluorescent proteins have become extremely popular tools for in vivo imaging and especially for the study of localization, motility and interaction of proteins in living cells. Here we report TagRFP, a monomeric red fluorescent protein, which is characterized by high brightness, complete chromophore maturation, prolonged fluorescence lifetime and high pH-stability. These properties make TagRFP an excellent tag for protein localization studies and fluorescence resonance energy transfer (FRET) applications.     

Apple flavonols during fruit adaptation to solar radiation: spectral features and technique for non-destructive assessment

Spectral properties of flavonols of three varieties (Golden Delicious, Antonovka, and Renet Simirenko) of anthocyanin-free apple fruit were investigated with reflectance spectroscopy. The results of spectral and biochemical analyses suggested that fruit reflectance in a broad spectral range 365-430 nm is strongly dependent on and, in sunlit fruit surfaces, governed by flavonols. The build up of peel flavonols (mainly rutin and other quercetin glycosides) resulted in a dramatic decrease of fruit reflectance in this range, flattening of the spectrum, and extending the region with low reflectance (4-5%) to ca. 410 nm. The spectral features observed suggest that flavonols contribute significantly to screening of excessive radiation, not only UV-A, but in the short-wave bands of chlorophyll and carotenoid absorption in the visible part of the spectrum as well. To retrieve quantitatively flavonol content from reflectance spectra, we tested the applicability of an inversion technique developed for non-destructive leaf pigment assessment. The model for flavonol content assessment was suggested in the form (R(-1)410 - R(-1)460)R800, where Rlambda is reflectance at wavelength lambda. The model was linearly related to flavonol content between 8 and 220nmol/cm2 with the coefficient of determination r2=0.92 and root mean square error of flavonol estimation of 20 nmol/ cm2 regardless of cultivar, chlorophyll, and carotenoid content.     

The allomerization of chlorophylls a and b with superoxide anion

The interaction of chlorophylls a and b with electrochemically prepared superoxide anion was studied in aprotic solvent. It was found that O₂^?·causes the deprotonation at carbon C-10 of ring V and production of chlorophyll enolate ions. The intermediate anions undergo rapid oxidation into corresponding chlorins. Pyrochlorophyll a, which lacks the C-10 carboxymethyl group, did not show the transformation. It is suggested that more strong free radical oxidants (e.g., HO₂·, or RO₂·) are capable of abstracting the hydrogen atom at C-10. The possible significance of free radical deprotonation and oxidation in chlorophyll allomerization is discussed.     

Non-destructive optical detection of pigment changes during leaf senescence and fruit ripening

Reflectance spectra in the visible and near infra-red range of the spectrum, acquired for maple ( Acer platanoides L.), chestnut ( Aesculus hippocastanum L.), potato ( Solanum tuberosum L.), coleus ( Coleus blumei Benth.), leaves and lemon (Citrus limon L.) and apple ( Malus domestica Borkh.) fruits were studied. An increase of reflectance between 550 and 740 nm accompanied senescence-induced degradation of chlorophyll (Chl), whereas in the range 400–500 nm it remained low, due to retention of carotenoids (Car). It was found that both leaf senescence and fruit ripening affect the difference between reflectance ( R ) near 670 and 500 nm ( R ₆₇₈− R ₅₀₀), depending on pigment composition. The plant senescing reflectance index in the form ( R ₆₇₈− R ₅₀₀)/ R ₇₅₀ was found to be sensitive to the Car/Chl ratio, and was used as a quantitative measure of leaf senescence and fruit ripening. The changes in the index were followed during leaf senescence, and natural and ethylene-induced fruit ripening. This novel index can be used for estimating the onset, the stage, relative rates and kinetics of senescence/ripening processes.     

Lipid peroxidation in and photo-damage to a light-sensitive chlorescence pea mutant

Leaves of 10- to 12-day-old chlorescence lethal Pisum sativum L. mutant are similar to control plants with respect to the content of chlorophylls, carotenoids, fatty acids and α-tocopherol. Subsequent development of the mutant under high irradiation resulted in th destruction of the photosynthetic pigments, polyunsaturated fatty acids, α-tocopherol, and also in the accumulation of liposoluble fluorescent products. No increase in the level of malondialdehyde was observed. In chloroplasts isolated from mutant plants the contents of chlorophyll a and β-carotene were decreased to a greater extent than the more oxidized pigments (xanthophylls and chlorophyll b ). The data obtained are discussed with special reference to the role of lipid peroxidation in the injury of plant cells under the action of visible light and to the antioxidative mechanisms stabilizing photosynthetic membranes.     

Effects of illumination and nitrogen starvation on accumulation of arachidonic acid by the microalga Parietochloris incisa

Parietochloris incisa is a unicellular freshwater green alga capable of accumulating high amounts of the valuable long-chain polyunsaturated arachidonic acid (AA) in triacylglycerols (TAG) of cytoplasmic oil bodies. To find the cultivation conditions providing maximum AA yield, the effects of illumination and N-availability on the dry weight (DW), chlorophyll, carotenoid, and AA content were studied. Under nitrogen starvation, TAG accounted for over 30% of dry weight (DW) and the AA content became as high as about 55% of total fatty acids. For biomass accumulation, light intensity of ca 400 μE m^?2 s^?1 was found to be optimal for growing P. incisa on a complete medium. Lower light intensities (or a higher cell density of inoculum) resulted in a higher AA yield when the alga was cultivated on nitrogen-free media. In the absence of nitrogen, algal cells were unable to cope with high illumination and suffered from photooxidative damage, whereas the nutrientsufficient culture survived under such illumination conditions, probably due to accumulation of carotenoids. Nitrogen-deprived P. incisa cells displayed elevated sensitivity to light.     

Nondestructive estimation of anthocyanins and chlorophylls in anthocyanic leaves

The anthocyanin and chlorophyll contents in leaves provide valuable information about the physiological status of plants. Thus, there is a need for accurate, efficient, and practical methodologies to estimate these biochemical parameters of vegetation. In this study, we tested the performance and accuracy of several nondestructive, reflectance-based techniques for estimating anthocyanin and chlorophyll contents in leaves of four unrelated species, European hazel (Corylus avellana), Siberian dogwood (Cornus alba =Swida alba), Norway maple (Acer platanoides), and Virginia creeper (Parthenocissus quinquefolia), with widely variable pigment content and composition. An anthocyanin reflectance index, which uses reflectances in the green and red edge spectral bands, and a modified anthocyanin reflectance index, employing, in addition, the near-infrared (NIR) band, were able to accurately estimate leaf anthocyanin for all species taken together with no reparameterization of algorithms. Total chlorophyll content was accurately estimated by a red edge chlorophyll index that uses spectral bands in the red edge and the NIR. These approaches can be used to estimate anthocyanin and chlorophyll nondestructively and allow the development of simple handheld field instrumentation.     

Plasmalemmal VDAC controversies and maxi-anion channel puzzle

The maxi-anion channel has been observed in many cell types from the very beginning of the patch-clamp era. The channel is highly conductive for chloride and thus can modulate the resting membrane potential and play a role in fluid secretion/absorption and cell volume regulation. A wide nanoscopic pore of the maxi-anion channel permits passage of excitatory amino acids and nucleotides. The channel-mediated release of these signaling molecules is associated with kidney tubuloglomerular feedback, cardiac ischemia/hypoxia, as well as brain ischemia/hypoxia and excitotoxic neurodegeneration. Despite the ubiquitous expression and physiological/pathophysiological significance, the molecular identity of the maxi-anion channel is still obscure. VDAC is primarily a mitochondrial protein; however several groups detected it on the cellular surface. VDAC in lipid bilayers reproduced the most important biophysical properties of the maxi-anion channel, such as a wide nano-sized pore, closure in response to moderately high voltages, ATP-block and ATP-permeability. However, these similarities turned out to be superficial, and the hypothesis of plasmalemmal VDAC as the maxi-anion channel did not withstand the test by genetic manipulations of VDAC protein expression. VDAC on the cellular surface could also function as a ferricyanide reductase or a receptor for plasminogen kringle 5 and for neuroactive steroids. These ideas, as well as the very presence of VDAC on plasmalemma, remain to be scrutinized by genetic manipulations of the VDAC protein expression. This article is part of a Special Issue entitled: VDAC structure, function, and regulation of mitochondrial metabolism.