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A new, high performance liquid chromatography method has been developed for the separation of monovinyl- and divinyl-protochlorophyllides, using commercially available, C30 reverse phase column and isocratic elution. This method can be used both for analytical applications and preparative scale purification of monovinyl- and divinyl-protochlorophyllides using the same column where submilimolar concentrations of the crude protochlorophyllide extract can be separated in one run. The purity of the obtained protochlorophyllides was demonstrated by spectroscopic methods, as well by the formation of aggregates in toluene. [ABSTRACT FROM AUTHOR]

The synthesis and assembly of functioning photosynthetic complexes in chloroplasts developing from etioplasts during the de-etiolation of angiosperm seedlings are imperative for the plant's autotrophic lifestyle. This study compared de-etiolation process under monochromatic red or blue light of equal photon flux density during a 24-hour illumination period of etiolated Arabidopsis seedlings. The aim was to elucidate the impact of these light wavelength on the etioplast-to-chloroplast transformation and the initiation of light-dependent photosynthetic reactions. Both treatments lead to the formation of functional young chloroplasts; however, the etioplast-to-chloroplast transition and the assembly of photosynthetic complexes occurred unevenly, with individual steps tuned by red or blue light. Ultrastructural analysis suggested faster prolamellar bodies disassembly under blue light, while low temperature fluorescence studies indicated a slower transformation of protochlorophyllide to chlorophyllide, and chlorophyll a, under these conditions. Red light further promoted the synthesis of chlorophyll b and LHCII antenna proteins. However, the efficiency of antennae in dissipating excess absorbed energy was higher for seedlings de-etiolated under blue light; the maximum quantum yield of the photosystem II reached 0.81 after 24-hour de-etiolation, equivalent to mature plants. Blue light seemed to enhance the development of well-functioning photosystems (I and II) and antennae. These findings are important for gaining a deeper understanding of photoreceptor regulation of de-etiolation and for utilizing selected light regimes to improve crop yield., (© The Author(s) 2024. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists.)

Being poikilohydric, lichens are inherently exposed to alternating desiccation and hydration cycles. They can exhibit extraordinary resistance to extreme temperatures in a dehydrated state but thermal thresholds for hydrated lichens are lower. The ability of the lichen Cetraria aculeata to recovery after high temperature treatment (40°C, 60°C) at different air humidity levels (relative humidity [RH]: <15%, 25%, 50%, 75%, ≅100%) was examined to find a linkage between passive dehydration of the lichen and its physiological resistance to heat stress. The response to heating was determined by measuring parameters related to photosynthesis and respiration after 2- and 24-h recovery. A higher RH level resulted in a slower decline in relative water content (RWC) in hydrated thalli. In turn, the stress resistance of active thalli depended on the ambient humidity and associated RWC reduction. Elevated temperature had a negative impact on bioenergetic processes, but only an unnatural state of permanent full hydration during heat stress resulted in a lethal effect. Hydrated lichen thalli heated at 40°C and 50% relative humidity (RH) tended to be least susceptible to stress-induced damage. Although atypical climatic conditions may lead lichens to lethal thresholds, the actual likelihood of deadly threat to lichens due to heat events per se is debatable., (© 2024 John Wiley & Sons Ltd.)

4-[5-(Naphthalen-1-ylmethyl)-1,3,4-thiadiazol-2-yl]benzene-1,3-diol (NTBD) was extensively studied through stationary UV-vis absorption and fluorescence measurements in various solvents and solvent mixtures and by first-principles quantum chemical calculations. It was observed that while in polar solvents (e.g., methanol) only a single emission band emerged; the analyzed 1,3,4-thiadiazole derivative was capable of producing dual fluorescence signals in low polarity solvents (e.g., n -hexane) and certain solvent mixtures (e.g., methanol/water). As clearly follows from the experimental spectroscopic studies and theoretical modeling, the specific emission characteristic of NTBD is triggered by the effect of enol → keto excited-state intramolecular proton transfer (ESIPT) that in the case of solvent mixture is reinforced by aggregation of thiadiazole molecules. Specifically, the restriction of intramolecular rotation (RIR) due to environmental hindrance suppresses the formation of non-emissive twisted intramolecular charge transfer (TICT) excited keto* states. As a result, this particular thiadiazole derivative is capable of simultaneously producing both ESIPT and aggregation-induced emission (AIE).

Epiphytic lichens constitute an important component of biodiversity in both deforested and forest ecosystems. Widespread occurrence is the domain of generalist lichens or those that prefer open areas. While, many stenoecious lichens find shelter only in a shaded interior of forests. Light is one of the factors known to be responsible for lichen distribution. Nevertheless, the effect of light intensity on photosynthesis of lichen photobionts remain largely unknown. We investigated photosynthesis in lichens with different ecological properties in relation to light as the only parameter modified during the experiments. The aim was to find links between this parameter and habitat requirements of a given lichen. We applied the methods based on a saturating light pulse and modulated light to perform comprehensive analyses of fast and slow chlorophyll fluorescence transient (OJIP and PSMT) combined with quenching analysis. We also examined the rate of CO 2 assimilation. Common or generalist lichens, i.e. Hypogymnia physodes, Flavoparmelia caperata and Parmelia sulcata, are able to adapt to a wide range of light intensity. Moreover, the latter species, which prefers open areas, dissipates the excess energy most efficiently. Conversely, Cetrelia cetrarioides considered an old-growth forest indicator, demonstrates definitely lower range of energy dissipation than other species, although it assimilates CO 2 efficiently both at low and high light. We conclude that functional plasticity of the thylakoid membranes of photobionts largely determines the dispersal abilities of lichens and light intensity is one of the most important factors determining the specificity of a species to a given habitat., (© 2023. The Author(s).)

In the presented study, advanced experimental techniques, including electronic absorption and fluorescence spectroscopies [with Resonance Light Scattering (RLS)], measurements of fluorescence lifetimes in the frequency domain, calculations of dipole moment fluctuations, quantum yields, and radiative and non-radiative transfer constants, were used to characterize a selected analogue from the group of 1,3,4-thiadiazole, namely: 4-[5-(naphthalen-1-ylmethyl)-1,3,4-thiadiazol-2-yl]benzene-1,3-diol (NTBD), intrinsically capable to demonstrate enol → keto excited-states intramolecular proton transfer (ESIPT) effects. The results of spectroscopic analyses conducted in solvent media as well as selected mixtures were complemented by considering biological properties of the derivative in question, particularly in terms of its potential microbiological activity. The compound demonstrated a dual fluorescence effect in non-polar solvents, e.g. chloroform and DMSO/H 2 O mixtures, while in polar solvents only a single emission maximum was detected. In the studied systems, ESIPT effects were indeed observed, as was the associated phenomenon of dual fluorescence, and, as demonstrated for the DMSO: H 2 O mixtures, the same could be relatively easily induced by aggregation effects related to aggregation-induced emission (AIE). Subsequently conducted quantum-chemical (TD-)DFT calculations supported further possibility of ESIPT effects. The following article provides a comprehensive description of the spectroscopic and biological properties of the analyzed 1,3,4-thiadiazole derivatives, highlighting its potential applicability as a very good fluorescence probes as well as a compound capable of high microbiological activity., (© 2022. The Author(s).)

The aim of the study was to measure the tensile strength of 4-year-old alfalfa leaves cultivated from seeds exposed to pre-sowing stimulation with He-Ne laser light for 1 or 5 min (designated respectively as F1 and F5) and alternating magnetic field with the exposure time of 1 or 5 min (respectively, L1 and L5). The leaves were measured in terms of blade length and width as well as petiole thickness prior to the tensile test. Measurements were also conducted to determine the chlorophyll fluorescence lifetime and content of photosynthetic pigments (chlorophyll a, b, a + b and carotenoids). The observed tensile strength was between 1.59 and 2.45 N. In the test group, the observed strength was lower in leaves collected from the top and central section of the stem but higher in the bottom part of the stem as compared to the control. The maximum increase of the tearing tensile force relative to the control (C) was observed for the L1 and F1 stimulation samples in leaves collected from the top and bottom parts of the stem, while the maximum decrease for that force was recorded for L5 leaves from the middle and top part of the stem. Chlorophyll fluorescence lifetimes and the overall content of photosynthetic pigments (chlorophyll a and b and carotenoids) were noticeably decreased in the leaves subjected to the stressors/stimulants applied. The results obtained for F1, L5 and, L1 stimulation revealed a decrease in fluorescence lifetimes. The content of photosynthetic pigments was also decreased under the influence of laser light stimulation (L1). This was a clear indication of plant ageing., (© 2022. The Author(s).)

The below article presents the results of spectroscopic research, theoretical (time-dependent density functional theory (TD-DFT)), microbiological, and antioxidative calculations for three compounds from the group of 1,3,4-thiadiazoles: 2-amino-5-phenyl-1,3,4-thiadiazole (TB), 2-amino-5-(2-hydroxyphenyl)-1,3,4-thiadiazole (TS), 2-amino-5-(2-hydroxy-5-sulfobenzoyl)-1,3,4-thiadiazole (TSF). In the fluorescence emission spectra (TS) of solutions with varying concentrations of hydrogen ions, a particularly interesting effect of dual fluorescence was observed. The aforementioned effect was observed even more clearly in the environment of butan-1-ol, relative to the compound's concentration. Depending on the modification of the resorcylic substituent (TS and TSF), we observed the emergence of two separate, partially overlapping, fluorescence emission spectra or a single emission spectrum. Interpretation of the obtained spectra using stationary and time-resolved spectroscopy allowed the correlation of the effect's emergence with the phenomenon of molecular aggregation (of a particular type) as well as, above all, the structure of the substituent system. The overlap of said effects most likely induces the processes related to the phenomenon of charge transfer (in TS) and is responsible for the observed fluorescence effects. Also, the position of the -OH group (in the resorcylic ring) is significant and can facilitate the charge transfer (CT). The determinations of the changes in the dipole moment and TD-DFT calculations further corroborate the above assumption. The following paper presents the analysis (the first for this particular group of analogues) of the fluorescence effects relative to the changes in the structure of the resorcylic group combined with pH effects. The results of biological studies also indicate the highest pharmacological potential of the analogue in the case where the effects of dual fluorescence emission are observed, which predisposes this particular group of fluorophores as effective fluorescence probes or potential pharmaceuticals with antimycotic properties., Competing Interests: The authors declare no conflicts of interest.

Heavy metals are known for their negative impact on the physiological processes of lichen photobiont. In spite of this, certain lichens are known to be effective pioneers of polluted sites. Cladonia cariosa, C. rei, and Diploschistes muscorum are prominent examples of lichens that spontaneously colonise post-industrial wastes. We examined the effect of total and intracellular Zn, Pb, Cd, As, Cu, and Ni accumulation in the thalli of these species on the physiological parameters of photobiont. Increased accumulation of Zn, Cd, Cu, and Ni in D. muscorum and of Zn and Ni in C. rei negatively affected contents of photosynthetic pigments, whereas concentrations of Pb had a positive effect in all lichen species. Moreover, pigment contents were positively associated with the concentrations of most examined elements in C. cariosa. The results indicate that even if chlorophyll contents reduced, its degradation does not progress. This suggests that metal stress may exert a negative effect on the synthesis rather than on the integrity of chlorophyll. Most importantly, lichen samples of each of the species from polluted sites proved to possess significantly higher F V /F M ratios than those from a reference site; moreover, the contents of elements of lichen thalli positively influenced this parameter. The efficient functioning of the algal component under heavy-metal stress conditions indicates that the examined lichens are well adapted to extremely contaminated substrates., (Copyright © 2019 Elsevier B.V. All rights reserved.)

This work presents spectroscopic studies of the keto-enol equilibrium induced by solvent polarizability in 4-[5-(naphthalen-1-ylmethyl)-1,3,4-thiadiazol-2-yl]benzene-1,3-diol a strong antiproliferative and anticancer thiadiazol derivative. Electronic absorption, steady state and time resolved fluorescence, and infrared spectroscopies were applied to investigate the keto and enol forms of this compound in a series of polar and non-polar solvents. The enol form dominates in polar solvents while, surprisingly, the keto form dominates in non-polar solvents with high average electric dipole polarizability e.g. n-alkenes. The electronic absorption spectrum of this derivative is more dependent on spatially averaged electric dipole polarizability of the solvent than on Kirkwood's correlation or on Lorenz-Lorenz electric polarizability. By analogy of n-alkanes to the alkyl parts of lipids, one can expect that the transformation of 1,3,4-thiadiazoles to the keto form may be facilitated in the hydrophobic core of the lipid membrane. Such a transition may be of great practical importance for the design of biologically active pharmaceutics, which are able to interact with the hydrophobic regions of cell membranes in a specific manner.

Ongoing research on DNA binding of cationic porphyrin derivatives and their conjugates is a subject of growing interest because of their possible DNA binding and demonstrated biological properties. In this study nucleoprotein binding of tri-cationic meso-tri(4-N-methylpyridyl)-mono-(4-carboxyphenyl)porphyrin (TMPCP) and tetrapeptides conjugated TMPCP (TMPCP-4P) and bi-cationic meso-5,10-bis(4-N-methylpyridyl)-15,20-di-(4-carboxyphenyl)porphyrin (BMPCP-4P2) was investigated with comprehensive spectroscopic methods. The key observation is that tetrapeptide-conjugates of cationic porphyrins with two or three positive charges bind to encapsidated DNA in T7 phage nucleoprotein complex. The binding modes were analyzed by fluorescent energy transfer, fluorescent life time and CD measurements. Intercalative binding is most feasible when tricationic ligands complex with DNA, especially when it is in close connection with protein capsid. It was found that larger ligand BMPCP-4P2 binds externally to encapsidated T7 DNA, and complex externally as well as by intercalation when the DNA accommodate to relaxed B-conformation. In the case of TMPCP and TMPCP-4P the intercalation is the predominant binding form both in nucleoprotein (NP) and preheated complexes. Further, melting experiments revealed that bound porphyrins do not influence the capsid stability or protein-DNA interactions, but efficiently stabilize the double helical structure of DNA without respect to binding form. A good correlation was found between porphyrin/base pair ration and DNA strand separation temperature., (Copyright © 2013 Elsevier B.V. All rights reserved.)

This study deals with the influence of cadmium on the structure and function of ferredoxin:NADP(+) oxidoreductase (FNR), one of the key photosynthetic enzymes. We describe changes in the secondary and tertiary structure of the enzyme upon the action of metal ions using circular dichroism measurements, Fourier transform infrared spectroscopy and fluorometry, both steady-state and time resolved. The decrease in FNR activity corresponds to a gentle unfolding of the protein, caused mostly by a nonspecific binding of metal ions to multiple sites all over the enzyme molecule. The final inhibition event is most probably related to a bond created between cadmium and cysteine in close proximity to the FNR active center. As a result, the flavin cofactor is released. The cadmium effect is compared to changes related to ionic strength and other ions known to interact with cysteine. The complete molecular mechanism of FNR inhibition by heavy metals is discussed.Electronic supplementary material The online version of this article (doi:10.1007/s10867-012-9262-z) contains supplementary material, which is available to authorized users.

Protochlorophyllide (Pchlide) accumulation and xantophyll composition were studied in 5-day old etiolated seedlings of three ecotypes of Arabidopsis thaliana: Columbia (Col-0), Landsberg erecta (Ler) and Wassiliewska (Ws). The total Pchlide level as measured by fluorescence spectroscopy varied significantly between ecotypes. A rapid HPLC method revealed quantitative differences in carotenoid composition. It was found that in the Ler ecotype any enhanced accumulation of Pchlide correlates with an increased level of lutein, suggesting the role of enzymes involved in lutein synthesis in cross-regulation between chlorophyll and carotenoid biosynthetic pathways. The function of the dark-accumulated carotenoid pool in seedling de-etiolation is discussed.

Recently cationic porphyrin-peptide conjugates were synthesized to enhance the cellular uptake of porphyrins or deliver the peptide moiety to the close vicinity of nucleic acids. DNA binding of such compounds was not systematically studied yet. We synthesized two new porphyrin-tetrapeptide conjugates which can be considered as a typical monomer unit corresponding to the branches of porphyrin-polymeric branched chain polypeptide conjugates. Tetra-peptides were linked to the tri-cationic meso-tri(4-N-methylpyridyl)-mono-(4-carboxyphenyl)porphyrin and bi-cationic meso-5,10-bis(4-N-methylpyridyl)-15,20-di-(4-carboxyphenyl)porphyrin. DNA binding of porphyrin derivatives, and their peptide conjugates was investigated with comprehensive spectroscopic methods. Titration of porphyrin conjugates with DNA showed changes in Soret bands with bathocromic shifts and hypochromicities. Decomposition of absorption spectra suggested the formation of two populations of bound porphyrins. Evidence provided by the decomposition of absorption spectra, fluorescence decay components, fluorescence energy transfer and induced CD signals reveals that peptide conjugates of di- and tricationic porphyrins bind to DNA by two distinct binding modes which can be identified as intercalation and external binding. Tri-cationic structure and elimination of negative charges in the peptide conjugates are preferable for the binding. Our findings provide essential information for the design of DNA-targeted porphyrin-peptide conjugates., (Copyright © 2011 Elsevier B.V. All rights reserved.)

One of the final reactions of chlorophyll (Chl) biosynthesis, e.g: photoreduction of protochlorophyllide (Pchlid) to chlorophyllide (Chlid) is a light-induced process in Angiosperm plants and it is catalyzed by light-dependent NADPH-Pchlid oxidoreductase (1.3.1.33; LPOR). In darkness, Chl biosynthesis is stopped at the stage of Pchlid formation. Seedlings and plastids develop according to a different pattern than that observed in the light. Moreover, synthesis of some proteins of the photosynthetic apparatus is inhibited. Light triggers the Pchlid photoreduction to Chlid, which induces the cascade of biochemical reactions and structural changes leading to the assembly of thylakoid membranes. In the present paper, the current knowledge on LPOR protein, mechanism of Pchlid to Chlid photoreduction, the role of lipid structure in etioplasts as well as spectral properties of Pchlid in etiolated seedlings and model systems is summarized.

Amphotericin B (AmB) is a life-saving polyene antibiotic used to treat deep-seated mycotic infections. Both the mode of therapeutic action as well as toxic side effects are directly dependent on molecular organization of the drug. Binding of AmB to lipid monolayers formed with dipalmitoylphosphatidylcholine, pure and containing 40 mol% cholesterol or ergosterol, the sterols of human and fungi respectively, has been examined by means of Fluorescence Lifetime Imaging Microscopy. AmB emits fluorescence with the characteristic lifetimes dependent on actual molecular organization: tau(M2) < or = 10 ps and tau(M1) = 0.35 ns in the monomeric state, the emission from the S(2) and the S(1) states respectively and tau(D) = 14 ns and tau(A) = 3.5 ns in the form of a dimer and associated dimers respectively. Analysis of the Langmuir-Blodgett films reveals that AmB binds to the lipid membranes and to the cholesterol-containing lipid membranes preferentially in the form of associated dimers. The same form of AmB appears in the membranes containing ergosterol but additionally the monomers and dimers of the drug can be observed, which can severely affect molecular organization of the lipid membrane. The results are discussed in terms of selectivity of AmB towards the ergosterol-containing biomembranes of fungi.

Phytol, a C20 alcohol esterifying the C-17(3) propionate, and Mg2+ ion chelated in the central cavity, are conservative structural constituents of chlorophylls. To evaluate their intramolecular structural effects we prepared a series of metal- and phytyl-free derivatives of bacteriochlorophyll a and applied them as model chlorophylls. A detailed spectroscopic study on the model pigments reveals meaningful differences in the spectral characteristics of the phytylated and non-phytylated pigments. Their analysis in terms of solvatochromism and axial coordination shows how the central Mg and phytyl residue shape the properties of the pigment. Surprisingly, the presence/absence of the central Mg has no effect on the solvatochromism of (bacterio)chlorophyll pi-electron system and the hydrophobicity of phytyl does not interfere with the first solvation shell of the chromophore. However, both residues significantly influence the conformation of the pigment macrocycle and the removal of either residue increases the macrocycle flexibility. The chelation of Mg has a flattening effect on the macrocycle whereas bulky phytyl residue seems to control the conformation of the chromophore via steric interactions with ring V and its substituents. The analysis of spectroscopic properties of bacteriochlorophyllide (free acid) shows that esterification of the C-17(3) propionate is necessary in chlorophylls because the carboxyl group may act as a strong chelator of the central Mg. These observations imply that the truncated chlorophylls used in theoretical studies are not adequate as models of native chromophores, especially when fine effects are to be modeled.

Fluorescence spectra and fluorescence lifetimes of protochlorophyll (Pchl) were measured in organic solvents having different physical and chemical properties and were analyzed taking into account the nonspecific (dependent on bulk solvent parameters), and specific (e.g. H bonds, Mg coordination) solvent-solute interactions. The energy of the fluorescence emission band decreased, while the Stokes shift increased for increasing solvent orientation polarizability, which is a function of both the dielectric constant (epsilon) and the refractive index (n). The extent of the dependence of the Stokes shift on solvent orientation polarizability was higher in protic (i.e. those able to form hydrogen-binding) than in aprotic solvents. High value of the Stokes shift was also observed in pyridine and methanol, i.e. in solvents hexacoordinating the central Mg atom. The fluorescence decay of Pchl was monoexponential in all of the investigated solvents. The fluorescence lifetime decreased for increasing solvent orientation polarizability from 5.5 +/- 0.1 ns in 1,4-dioxane to 3.3 +/- 0.1 ns in methanol. Longer lifetime values were observed in the case of aprotic solvents than in protic solvents. The hexacoordination of Mg had no effect on the fluorescence lifetime. The present data are discussed with respect to results found for protochlorophyllide (Pchlide) (Myśliwa-Kurdziel et al. in Photochem Photobiol 79:62-67, 2004), and they indicate that the presence of phytol chain in the porphyrin ring influences the spectral properties of the whole chromophore. This is the first complex analysis comparing the fluorescence emission and fluorescence lifetimes of purified Pchl and Pchlide.

The steady-state and time-resolved fluorescence characteristics of protochlorophyll (Pchl) dissolved in neat Triton X-100 and in Triton X-100 micelles were investigated, and the fluorescence lifetimes of different Pchl spectral forms were studied. Varying the concentration of Pchl or diluting the micellar solutions either with a buffer or with a micellar solution, 631-634, 645-655, 680-692 and above 700 nm emitting Pchl complexes were prepared, the ratios of which varied from one another. The fluorescence decay of the 631-634 nm emitting (monomeric) form had a mono-exponential character with a 5.4-ns fluorescence lifetime. The long-wavelength Pchl complexes (aggregates) had two fluorescence lifetime values within a range of 1.4-3.9 ns and 0.15-0.84 ns, which showed high variability in different environments. Depending on the conditions, either mono- or double-exponential fluorescence decay was found for a fluorescence band at 680-685 nm. These data show that despite their very similar steady-state fluorescence properties, Pchl complexes can differ in fluorescence lifetimes, which may reflect different molecular structures, intrinsic geometries or different molecular interactions. This underlines the importance of complex spectroscopic analysis for a precise description of native and artificial chlorophyllous pigment forms.

We have found that for biological prenyllipids, such as plastoquinol-9, alpha-tocopherol quinol, and alpha-tocopherol, the shortest fluorescence lifetimes were found in aprotic solvents (hexane, ethyl acetate) whereas the longest lifetimes were those of ubiquinonol-10 in these solvents. For all the investigated prenyllipids, fluorescence lifetime in alcohols increased along with an increase in solvent viscosity. In a concentrated hexane solution, the lifetimes of prenylquinols considerably decreased. This contrasts with methanol solutions, which is probably due to the self-association of these compounds in aprotic solvents. We have also found a correlation of the Stokes shift of prenyllipids fluorescence with the orientation polarizability of the solvents. Based on data obtained in organic solvents, measurements of the fluorescence lifetimes of prenyllipids in liposomes allowed an estimation of the relative distance of their fluorescent rings from the liposome membrane surface, and was found to be the shortest for alpha-tocopherol quinol in egg yolk phosphatidylcholine liposomes, and increased in the following order: alpha-tocopherol in dipalmitoyl phosphatidylcholine liposomes < alpha-tocopherol < plastoquinol-9 < ubiquinol-10 in egg-yolk phosphatidylcholine liposomes.

In this work, absorption and fluorescence spectra of protochlorophyllide (Pchlide), as well as its fluorescence lifetime, were investigated in organic solvents having different physical properties. The obtained Pchlide spectral features are discussed in relation to the parameters describing solvent properties (refractive index and dielectric constant) and taking into account the specific solvent-Pchlide interaction. The correlation of Pchlide Qy and Soret absorption bands with solvent polarizability function ((n2 - 1)/(n2 + 2)) has been found; however, the dispersion of the observed points was rather high. A small Stokes shift of a magnitude between 50 and 300 cm(-1) was found, which indicates low sensitivity of Pchlide to nonspecific solvation. The fluorescence decay of Pchlide was single exponential in all the investigated solvents, with the lifetime value ranging from 5.2 ns for dioxane to 3.5 ns for methanol. Dependence of the obtained fluorescence lifetimes on the solvent orientation polarizability, a parameter being the function of both refractive index and dielectric constant, was discussed. In water-methanol mixtures, a further decrease of the fluorescence lifetime was observed, giving values of 2.9 ns for 25% methanol. Double-exponential decay of Pchlide fluorescence was found for Pchlide in a solution of 15% methanol with the lifetimes of 4.5 +/- 0.5 ns and 1.2 +/- 0.3 ns and in pure water with the lifetimes of 2.5 +/- 0.5 ns and 0.4 +/- 0.1 ns. The obtained results are discussed in relation to spectroscopic properties of Pchlide in vivo.

Dark-grown leaves of maize (Zea mays), wheat (Triticum aestivum), wild-type pea (Pisum sativum) and its light-independent photomorphogenesis mutant (lip1) have different proportions of protochlorophyllide (Pchlide) forms as revealed by low-temperature fluorescence emission spectra. Four discrete spectral forms of Pchlide, with emission peaks around 633, 640, 656 and 670 nm, could be distinguished after Gaussian deconvolution. In maize and wheat the 656 nm component was the most prominent, whereas for wild-type pea and its lip1 mutant, the 633 and 640 nm components contributed mostly to the fluorescence emission spectra. For the fluorescence lifetimes measured at 77 K a double exponential model was the most adequate to describe the Pchlide fluorescence decay not only for the Pchlide(650-656) form but also for the short-wavelength Pchlide forms. A fast component in the range 0.3-0.8 ns and a slow component in the range 5.1-7.1 ns were present in all samples, but the values varied, depending on species. The long-wavelength Pchlide(650-656) form had a slow component with a lifetime between 5.1 and 6.7 ns, probably reflecting the fluorescence from aggregated Pchlide. The short-wavelength Pchlide(628-633) form had values of the slow component varying between 6.2 and 7.1 ns. This represents a monomeric but probably protein-bound Pchlide form because the free Pchlide in solution has a much longer lifetime around 10 ns at 77 K. The contribution of different Pchlide forms to the measured lifetime values is discussed.

The origin of heat-induced chlorophyll fluorescence rise that appears at about 55-60 degrees C during linear heating of leaves, chloroplasts or thylakoids (especially with a reduced content of grana thylakoids) was studied. This fluorescence rise was earlier attributed to photosystem I (PSI) emission. Our data show that the fluorescence rise originates from chlorophyll a (Chl a) molecules released from chlorophyll-containing protein complexes denaturing at 55-60 degrees C. This conclusion results mainly from Chl a fluorescence lifetime measurements with barley leaves of different Chl a content and absorption and emission spectra measurements with barley leaves preheated to selected temperatures. These data, supported by measurements of liposomes with different Chl a/lipid ratios, suggest that the released Chl a is dissolved in lipids of thylakoid membranes and that with increasing Chl a content in the lipid phase, the released Chl a tends to form low-fluorescing aggregates. This is probably the reason for the suppressed fluorescence rise at 55-60 degrees C and the decreasing fluorescence course at 60-75 degrees C, which are observable during linear heating of plant material with a high Chl a/lipid ratio (e.g. green leaves, grana thylakoids, isolated PSII particles).

Optical absorption and fluorescence parameters of chlorophyll a and the phytol-free chlorophyllide a, as well as of their Mg-depleted derivatives, were compared in a series of organic solvents. In contrast to prevailing opinion, the spectral properties of chlorophyll are not indifferent to the removal of phytol. The electronic absorption spectra of chlorophyll a and chlorophyllide a differ and display a different dependence on the nature of the solvent, which cannot be explained solely by the location of a charged carboxylic group in the proximity of the pi- electron system. In fact, measurements in media of varying basicity show that deprotonation of the free carboxylic group in chlorophyllide, i.e., the presence of a negative point charge near the macrocycle, has no effect on pigment absorption spectra. Analysis of the solvent effect on the Q(Y) energies in terms of solvent polarity reveals that the phytyl moiety perturbs the spectral features of chlorophyll, mainly due to its interactions with the pigment solvation shell. The phytyl residue might also be thus partly involved in controlling the central metal ligation in chlorophylls. This influence of phytol on the spectral features of chlorophyll should be taken into account when comparing the spectra in solution with various spectral forms of chlorophyll in vivo.

Our study was aimed to investigate the significance of the isoprenoid side chain size as well as redox state of the quinone ring for interaction of two main classes of prenylquinones: plastoquinones (PQ) and ubiquinones (UQ) with lipid bilayers. By use of differential scanning calorimetry (DSC) we have followed the thermotropic behaviour of multilamellar vesicles prepared from dipalmitoylphosphatidylcholine (DPPC) upon incorporation of increasing amount (1.3-12 mol%) of quinone (quinol) molecules. Our studies reveal that as the side chain is shorter (from 9 to 2 isoprenoid units) the height of the calorimetric profiles is reduced and the temperature of the main transition of DPPC (T(m)) decreases (T(m)=39.4 degrees C for a sample with 12 mol% of PQ-2), and then increases up to 39.8 degrees C for PQ-1. For the samples containing quinols the effect is more pronounced even at lower concentration. The greater influence of the added prenylquinones on the pretransition demonstrates a stronger distortion of the DPPC packing in the gel state. It seems that this is the isoprenoid side chain length rather than the redox state of prenylquinones that determines their effectiveness in perturbation of thermotropic properties of lipid bilayer.

An antiserum to alpha-tocoquinone was prepared by immunization of rabbits. Immunization was obtained by injection of a conjugate consisting of the hapten alpha-tocoquinone attached to methylated ovalbumin into the rabbit. The antiserum recognizes the 3,4-dimethyl-p-benzoquinone group of the molecule as well as part of the immediate vicinity to the side chain. This is concluded from the fact that the antibody has some affinity also to plastoquinone. No reaction of the antibody is observed with alpha-tocopherol hydroquinone or alpha-tocopherol. Reaction of the antiserum to alpha-tocoquinone with photosystem II-particle preparations from tobacco affects the functionality of the preparation. Chlorophyll(a)-fluorescence emission is quenched without an alteration of the emission spectrum. Concomitant with this fluorescence quenching, the lifetime of two fluorescence components namely that of a fast and a slower component are shortened. By analogy with the literature the fast component is associated with chlorophyll(a) of the reaction center core and that of the slow component with the antenna system in which the lifetime parameter is shortened by the antibody from 3.42 ns to 1.795 ns. The action on the fast component is less and leads to a shortening of the lifetime parameter from 0.373 ns to only 0.249 ns. The effect is interpreted in terms of an enhancement of linear photosynthetic electron transport possibly due to an inhibition of the cyclic electron transport around PS II. discovered by Gruszecki et al. (1995), Z. Naturforsch. 50c, 61-68.