Alexei Solovchenko

Reflectance spectral features and non-destructive estimation of chlorophyll, carotenoid and anthocyanin content in apple fruit

Fruit reflectance spectra of five apple (Malus domestica Borkh.) cultivars (Zhigulevskoe, Antonovka, Granny Smith, Golden Delicious and Renet Simirenko) with a wide range of peel pigment (chlorophylls a and b , carotenoids and anthocyanins) content have been studied to develop non-destructive techniques for pigment assessment. In addition to chlorophylls, positions of in vivo absorption maxima were established for carotenoids (480, 455 and a shoulder at 425 nm) and for anthocyanins (near 550 nm). In anthocyanin-free fruit, a close relationship between reflectance at 550 nm (R550) and 700 nm (R700) has been found (r 2 � /0.95). In fruit with chlorophyll content more than 5 nmol/cm 2 , the reflectance near 678 nm was insensitiv et o avariation in chlorophylls, whereas, reflectance in the bands 550 � /650 nm and 690 � /705 nm remained sensitive to chlorophyll content in a wide range of its variation. The reflectance ratios, R800/ R700 and R800/R640, were directly proportional to total chlorophyll content ranging from 0.4 to 11 nmol/cm 2 (r 2 � /0.93). The reflectance in the band 520 � /530 nm was found to be dependent mostly on carotenoids absorption. The index R800(1/R520� /1/R700) was suggested for estimation of carotenoid content in the range 0.6 � /4.5 nmol/cm 2 . The index for assessment of a carotenoid/chlorophyll ratio was proposed in the form, (R480� /R678)/R800. Reflectance in the green region of the spectrum proved to be sensitive to anthocyanin content. The index R800(1/R550� /1/R700) was developed to estimate anthocyanin content in peel ranging from 2.5 to 50 nmol/cm 2 ; the determination coefficient of the index with anthocyanin content was higher than 0.93. # 2002 Elsevier Science B.V. All rights reserved.

Screening of Visible and UV Radiation as a Photoprotective Mechanism in Plants

Prolonged exposure of plants to high fluxes of solar radiation as well as to other environmental stressors disturbs the balance between absorbed light energy and capacity of its photochemical utilization resulting in photoinhibition of and eventually in damage to plants. Under such circumstances, the limiting of the light absorption by the photosynthetic apparatus efficiently augments the general photoprotective mechanisms of the plant cell, such as reparation of macromolecules, elimination of reactive oxygen species, and thermal dissipation of the excessive light energy absorbed. Under stressful conditions, plants accumulate, in different cell compartments and tissue structures, pigments capable of attenuation of the radiation in the UV and visible parts of the spectrum. To the date, four principle key groups of photoprotective pigments are known: mycosporine-like amino acids, phenolic compounds (including phenolic acids, flavonols, and anthocyanins), alkaloids (betalains), and carotenoids. The accumulation of UV-absorbing compounds (mycosporine-like amino acids and phenolics in lower and higher plants, respectively) is a ubiquitous mechanism of adaptation to and protection from the damage by high fluxes of solar radiation developed by photoautotrophic organisms at the early stages of their evolution. Extrathylakoid carotenoids, betalains, and anthocyanins play an important role in long-term adaptation to the illumination conditions and in protection of plants against photodamage. A prominent feature of certain plant taxa lacking some classes of photoprotective pigments (such as anthocyanins) is their substitution by other compounds (e.g. keto-carotenoids or betalains) disparate in terms of chemical structure and subcellular localization but possessing close spectral properties.

Light absorption by anthocyanins in juvenile, stressed, and senescing leaves

The optical properties of leaves from five species, Norway maple (Acer platanoides L.), cotoneaster (Cotoneaster alaunica Golite), hazel (Corylus avellana L.), Siberian dogwood (Cornus alba L.), and Virginia creeper (Parthenocissus quinquefolia (L.) Planch.), differing in pigment composition and at different stages of ontogenesis, were studied. Anthocyanin absorption maxima in vivo, as estimated with spectrophotometry of intact anthocyanic versus acyanic leaves and microspectrophotometry of vacuoles in the leaf cross-sections, were found between 537 nm and 542 nm, showing a red shift of 5–20 nm compared with the corresponding maxima in acidic water–methanol extracts. In non-senescent leaves, strong anthocyanin absorption was found between 500 nm and 600 nm (with a 70–80 nm apparent bandwidth). By and large, absorption by anthocyanin in leaves followed a modified form of the Lambert–Beer law, showing a linear trend up to a content of nearly 50 nmol cm−2, and permitting thereby a non-invasive determination of anthocyanin content. The apparent specific absorption coefficients of anthocyanins at 550 nm showed no substantial dependence on the species. Anthocyanin contribution to total light absorption at 550 nm was followed in maple leaves in the course of autumn senescence. Photoprotection by vacuolar anthocyanins is discussed with special regard to their distribution within a leaf; radiation screening by anthocyanins predominantly localized in the epidermal cells in A. platanoides and C. avellana leaves was also evaluated.

EFFECT OF NITROGEN STARVATION ON OPTICAL PROPERTIES, PIGMENTS, AND ARACHIDONIC ACID CONTENT OF THE UNICELLULAR GREEN ALGA PARIETOCHLORIS INCISA (TREBOUXIOPHYCEAE, CHLOROPHYTA) 1

Spectral properties of cell suspensions, individual cells, and extracts of the unicellular green alga Parietochloris incisa (Reisigl) Shin Watan. grown under low light were studied. Long‐term nitrogen (N) deprivation resulted in a decrease of chloroplast volume, appearance of numerous large cytoplasmic oil bodies, and the deposition of triacylglycerols with a high proportion of arachidonic acid. Chlorophylls a and b underwent a synchronous decline, whereas carotenoids (Car) showed a relative increase. Simultaneously, significant qualitative changes in the spectral properties of P. incisa individual cells, cell extracts, and cell suspensions were observed. To a large extent, the spectral changes observed in cell suspension could be attributed to a decrease in overall pigment content, leading to a gradual weakening of the so‐called package effect and accumulation of additional amounts of Car over chl, most probably, in oil bodies. Several optical characteristics of cell suspensions could serve as sensitive indicators of N‐deficiency in P. incisa. Furthermore, the absorption ratios, A476/A676 and A650/A676, showed close correlations with the Car‐to‐chl ratio and relative arachidonic acid (AA) content, respectively. The latter makes it possible to suggest that the increase in AA percentage in P. incisa proceeds in parallel with a decrease in cell chl content, accounting for the weakening of the package effect. N‐replenishment resulted in complete recovery of cell optical properties. The possible significance of the changes in cell ultrastructure, pigments, lipids, and optical properties is discussed with special reference to the ability of algae to adapt to and survive under conditions of long‐term nutrient deficiency.

Physiological role of neutral lipid accumulation in eukaryotic microalgae under stresses

In the studies of lipid metabolism of unicellular photoautotrophic eukaryotes (microalgae), the main attention is commonly paid to polar membrane lipids and their fatty acid (FA) composition, whereas neutral lipids, represented predominantly by triacylglycerols (TAG), are insufficiently studied. As was reported recently, the role of these compounds in microalgae is not limited to their storage function. It was found that TAG are frequently involved in adaptation to environmental conditions. This review summarizes experimental data obtained so far allowing to distinguish at least three aspects of TAG adaptive function in microalgae. First, these compounds are the source of long-chain FA, the building blocks for membranes necessary for rearrangements of the photosynthetic apparatus. Second, TAG biosynthesis consumes excessive photoassimilates preventing photooxidative injuries under stresses which reduce cell capacity of photosynthesis product utilization. Third, TAG deposited as cytoplasmic oil bodies form a depot for secondary carotenoids in carotenogenic microalgae producing an optical screen protecting the cell against photodamage by excessive PAR.

Photostability of pigments in ripening apple fruit: a possible photoprotective role of carotenoids during plant senescence

Light-induced pigment destruction was studied in ripening apple (Malus domestica Borkh., cv. Antonovka) fruits with reflectance spectroscopy. The reflectance spectral changes attributable to light-induced transformation of xanthophyll cycle carotenoids (Car) preceded the pigment degradation. In green fruits, the destruction of chlorophyll and Car proceeded synchronously up to complete disappearance of both pigments. In ripening fruits with a molar chlorophyll/carotenoid ratio B/2.5 � /3, significant amounts of peel Car were retained at the deep stages of chlorophyll degradation. Car were especially resistant to irradiation in yellow fruits (a chlorophyll/carotenoid ratio B/0.3); the extent of their bleaching after prolonged irradiation did not exceed 20%. Irradiation of pigment solutions showed that apple fruit Car alone exhibit much higher light stability than in the presence of chlorophyll. The extent, kinetics and stoichiometry of light-induced pigment destruction in apples are consistent with the existence of two carotenoid pools, (i) closely associated with chlorophyll in chloroplast thylakoid membranes and (ii) exhibiting higher light stability localised in plastoglobuli appearing in chloroplasts undergoing transformation to gerontoplasts � /chromoplast. It is suggested that the induction of carotenoid synthesis during senescence provides the protection of plastoglobuli and light-sensitive constituents of plant tissues from irradiation in the blue part of the visible spectrum. # 2002 Elsevier Science Ireland Ltd. All rights reserved.

Application of Reflectance Spectroscopy for Analysis of Higher Plant Pigments

Nondestructive techniques developed by the authors for assessment of chlorophylls, carotenoids, and anthocyanins in higher plant leaves and fruits are presented. The spectral features of leaf reflectance in the visible and near infrared regions are briefly considered. For pigment analysis only reflectance values at several specific wavelengths are required. The chlorophyll (Chl) content over a wide range of its changes can be assessed during leaf ontogeny using reflectance near 700 nm and, in the absence of anthocyanins, at 550 nm. The approaches used for elimination of Chl interference in the analysis of carotenoids (reflectance at 520 nm) and anthocyanins (at 550 nm) are described. The suitability of reflectance spectroscopy for estimates of carotenoid/chlorophyll ratios during leaf senescence and fruit ripening is demonstrated. The algorithms developed for pigment analysis are presented, and the conditions of their applicability are considered. Further perspectives for the application of reflectance spectroscopy including remote sensing for estimation of plant pigment content and physiological states are discussed.

Optical properties and contribution of cuticle to UV protection in plants: experiments with apple fruit

To assess the UV-screening capacity of plant surface structures, the optical properties of isolated cuticle and detached peel of apple fruit (Malus domestica Borkh., cv. Antonovka) have been studied. It was found that the cuticle exhibits considerable scattering of UV radiation, negligible absorption between 500-800 nm and attenuates UV radiation: on average, cuticular transmittance of non-reflected light amounts to 35.7 +/- 20.2 and 14.2 +/- 7.1% at 375 and 300 nm, respectively. The principal UV-A absorbers in the cuticle were identified as quercetin glycosides with an in vivo absorption maximum near 375 nm and content ranging from 10 to 70 nmol cm(-2). On the shaded side of apple fruit, both UV-A and UV-B absorption by the peel is, to a large extent, governed by cuticular phenolics, whereas on the sunlit surface, the absorption of the peel in the UV-A range is determined mainly by vacuolar peel flavonoids. It is concluded that a massive build-up of flavonoids in the peel cells located just below the cuticle, resulting in trapping of radiation in a broad spectral range, plays a dominant role in the long-term adaptation of apple fruit to elevated levels of solar radiation.

Effects of Light and Nitrogen Starvation on the Content and Composition of Carotenoids of the Green Microalga Parietochloris incisa

The changes in pigment content and composition of the unicellular alga Parietochloris incisa comb. nov (Trebouxiophyceae, Chlorophyta) were studied. This alga is unique in its ability to accumulate high amounts of arachidonic acid in the cell during cultivation under different irradiances and nitrogen availability in the medium. Under low irradiance of 35 μE/(m2 s) photosynthetically active radiation the P. incisa cultures possessed slow growth and a relatively low carotenoid-to-chlorophyll ratio. At higher irradiances (200 and 400 μE/(m2 s)) on complete medium, the alga displayed higher growth rate and an increase in the carotenoid content, especially that of β-carotene and lutein. Both on nitrogen-free (regardless of illumination intensity) and nitrogen-replete medium (under high light), a considerable increase in the ratio of carotenoid and chlorophyll contents was recorded. Predominant accumulation of xanthophylls took place in thylakoid membranes, whereas β-carotene deposition occurred mainly in the cytoplasmic lipid globules (oil bodies); lower amounts of carotenoids were accumulated in the absence of nitrogen. Under high light and nitrogen-deficiency conditions, an increase in violaxanthin de-epoxidation and nonphotochemical quenching was recorded together with a decline in variable chlorophyll fluorescence (Fv/Fm) level. A possible photoprotective role of carotenoids in adaptation of P. incisa to high light under nitrogen starvation conditions is discussed.

Optical properties of rhodoxanthin accumulated in Aloe arborescens Mill. leaves under high-light stress with special reference to its photoprotective function

In Aloe arborescens Mill. leaves, strong sunlight or its combination with drought induces the accumulation of the red keto-carotenoid, rhodoxanthin. Simultaneously, the transformation of chloroplasts into chromoplasts accompanied by degradation of thylakoid membranes and formation of plastoglobuli, large in size and number, takes place. Depending on stress conditions the build up of rhodoxantin occurred along with the loss of chlorophyll or on the background of relatively high content of the pigment in the leaves. Microspectrophotometrical measurements showed the presence of chlorophyll-free plastids and retention of carotenoids during leaf adaptation to strong sunlight. The plastid spectra contained absorption bands of common for higher plants carotenoids together with those of rhodoxantin, with absorption maxima situated in the blue (440-480 nm) and the green ranges of the spectrum, respectively. The studies of whole-leaf optical properties revealed a broad band of rhodoxanthin absorption in the blue-green range peaking near 540-550 nm. Within this spectral band the accumulation of rhodoxanthin occurring, probably, in plastoglobuli considerably increased light absorption by stressed Aloe leaves. A possible photoprotective function of rhodoxanthin and other carotenoids as an internal light trap analogous to that accomplished by anthocyanins in other plant species is discussed.