In higher plants, natural radiation simultaneously activates more than one photoreceptor. Five phytochromes (phyA through phyD), two cryptochromes (cry1, cry2) and phototropin have been identified in the model species Arabidopsis thaliana. There is light-dependent epistasis among certain photoreceptor genes because the action of one pigment can be affected by the activity of others. Under red light, phyA and phyB are antagonistic, but under far-red light, followed by brief red light, phyA and phyB are synergistic in the control of seedling morphology and the expression of some genes during de-etiolation. Under short photoperiods of red and blue light, cry1 and phyB are synergistic, but under continuous exposure to the same light field the actions of phyB and cry1 become independent and additive. Phototropic bending of the shoot toward unilateral blue light is mediated by phototropin, but cry1, cry2, phyA and phyB positively regulate the response. Finally, cry2 and phyB are antagonistic in the induction of flowering. At least some of these interactions are likely to result from cross talk of the photoreceptor signaling pathways and uncover new avenues to approach signal transduction. Experiments under natural radiation are beginning to show that the interactions create a phototransduction network with emergent properties. This provides a more robust system for light perception in plants.

Determining whether the first step of photooxygenation is Type I or Type II is a necessary prerequisite in order to establish the mechanism of photodynamic action. But this distinction is not sufficient, because other processes, both consecutive and competitive, commonly participate in the overall mechanism. Thus, in both Type I and Type II reactions, the initial products are often peroxides that can break down and induce free radical reactions. These aspects of photosensitization are discussed and illustrated by sensitizer/substrate systems involving (1) only radical reactions (decatungstate/alkane) and (2) reactions of singlet oxygen occurring in competitive and consecutive processes and possibly followed by radical reactions (methylene blue/2′-deoxyguanosine). Two other previously investigated systems involving, respectively, a Type II interaction followed by radical processes (methylene blue/alkene) and Type II reactions, some of which being competitive or consecutive (rose bengal/alkene), are briefly reconsidered.

O-Methyl methylene violet (OMeMV), O-methyl bromomethylene violet (OMeBrMV) and O-methyl iodomethylene violet (OMeIMV) have been prepared in order to test their potential utility as anti-viral and anti-tumor phototoxic dyes. Rates of photosensitized toxicity of KB cells with 633 nm irradiation are (×10⁻¹⁹ photon⁻¹): 2.4, 2.2, 1.9 and 0.17 for OMeIMV, OMeBrMV, methylene violet (MV) and OMeMV, respectively. Rates of photosensitized inactivation of Sindbis virus in phosphate-buffered saline with 633 nm irradiation are (×10⁻¹⁸ photon⁻¹): 3.3, 1.8, 0.99, 0.15 for MV, OMeIMV, OMeBrMV and OMeMV, respectively. Quantum efficiencies for singlet oxygen formation were determined as OMeIMV, 0.64; OMeBrMV, 0.40; OMeMV, 0.054. Titration of the dyes with double-stranded (ds)DNA resulted in bathochromic shifts and hypochromic effects in the visible absorption spectra. Association constants for interaction of the methylated dyes with dsDNA of ∼1 × 10⁵ M⁻¹ were determined by Scatchard analysis of equilibrium dialysis and UV absorption titration data. Photolysis of the halogenated dyes with DNA under argon led to covalent bond formation with the nucleic acid; there was no evidence of covalent binding in the dark.

The irradiation of colchicone 5 led to the formation of lumicolchicone 7. The same reaction cannot be obtained by using thiocolchicone 6 as substrate. Transient absorption spectroscopy of colchicone and β-lumicolchicone showed that probably the photoisomerization occurred on colchicone in its first excited singlet state. The spectroscopic data are in agreement with the hypothesis that lumicolchicone was generated in the ground state from the S₁ state of colchicone without the presence of any intermediate. Semiempirical calculations on colchicone and thiocolchicone showed that the highest single occuped molecular orbital and the lowest unoccupied molecular orbital of the singlet excited colchicone can give a disrotatory ring closure to 7, while thiocolchicone cannot give the same type of process.

The complexation dynamics of radical cations with cyclodextrins (CD) was studied using photophysical techniques. Radical cations of 4-vinylanisole and trans-anethole were formed within α- and β-CD cavities by two-photon photolysis of the respective styrene precursors. Exit of the radical cations from α-CD complexes with 1:1 and 1:2 (guest : CD) stoichiometries and β-CD complexes with 1:1 stoichiometries occurred with lifetimes shorter than 100 ns. Most of the radical cations formed escape from the CD cavities, but a small portion do react with α-CD when this host is present in high concentrations.

The spectral and kinetic properties of reduced radicals and the triplet state of neutral red (NR), a phenazine-based dye, have been investigated using pulse radiolysis technique. A mixed water–isopropanol–acetone solvent has been used to study the reduced radicals of NR for a wide pH range of about 1–13, due to limitation of solubility of the dye in aqueous solutions particularly above pH 8. From pH-dependent absorption studies it has been established that the reduced radicals of NR can exist in four different prototropic forms in solution. Three pK_a values for the corresponding prototropic equilibria have been estimated. The formation and decay rate constants of reduced radicals have also been measured. The triplet state characteristics of the dye have been investigated in neat benzene solutions, both in the presence and in the absence of triplet sensitizers. The T₁ → T_n absorption spectrum and decay kinetics of the triplet state have been measured. The triplet state energy (E_T) of NR in benzene have been estimated to be within 36–42 kcal mol⁻¹, using an energy transfer method.

We describe the synthesis and photophysical properties of a series of neutral and cationic 3,4-tetrapyridinoporphyrazines, potential lead photosensitizers for photodynamic inactivation of bacteria. Tetracationic TPyPzs exist essentially as monomers in aqueous systems, but the presence of trialkylated compounds due to incomplete quaternization of the outer nitrogen atoms induces severe aggregation. The absorption, fluorescence, triplet, and singlet oxygen quantum yields for both the neutral and cationic compounds are comparable to those of the related phthalocyanines.

Cumulative and intermittent sun exposure are risk factors for skin cancer, highlighting the need to monitor exposure during childhood. There is currently very little available information concerning the accuracy of self-reported levels of sun exposure, particularly for very young children. In this study, UV radiation (UVR) exposure measured by polysulfone dosimeters worn on the wrist was compared with a measure of estimated exposure using a diary based on recall at the end of the 4 day study period and ambient dosimetric measures. Results of the study indicate that the relative UVR exposures expressed as a fraction of daily total ambient received during the 4 day period by young children and mothers are similar. A high level of association was obtained for the estimated levels of exposure between young children and their mothers. Moderate correlations were evident for dosimeter readings of mother and child on weekends with no significant association on weekdays. The association between estimated exposure and dosimeter readings was poor and needs improvement. This may be achieved by greater consideration of structural and environmental factors that influence the levels of UVR exposure received by individuals and by increasing the level of specificity in the measurement instruments. Methodological issues such as recall of exposure after several days, quantity of sun exposure and more precise measurement of grades of exposure when outside may have more impact than previously expected, and further information is being sought with a larger sample for exposure during the summer months.

The effects of a single dose of ultraviolet B (UVB) radiation (0.4 J/cm²) on immunological functions by blood leukocytes and on hematological parameters was studied in roach (Rutilus rutilus), a teleostean fish. The respiratory burst of phorbol 12-myristate 13-acetate stimulated whole blood phagocytes increased significantly after UVB irradiation but spontaneous cytotoxicity of blood leukocytes toward ⁵¹chromium-labeled K562 target cells was not markedly altered. Differential cell counting revealed that UVB exposure significantly increased the proportion of granulocytes and significantly decreased the proportion of lymphocytes in the peripheral blood, whereas hematocrit and the total number of white and red blood cells were unchanged. Plasma cortisol concentration increased in UVB-exposed fish. Severe handling stress caused similar, although not as potent, effects on the measured parameters of fish blood as UVB irradiation. These observations suggest that in fish UVB brings about a stress response, which may account for the observed alterations in the immune parameters and leukocyte composition of blood. Exposure of fish to strong visible light induced no alterations in immunological or hematological parameters, making it unlikely that ultraviolet radiation mediates its effects through visual perception.

The conversion of chlorophyll a (Chl a) monomers into large aggregates in six polar solvents upon addition of water has been studied by means of absorption, fluorescence spectroscopy and fluorescence lifetime measurements for the purpose of elucidating the various environmental factors promoting Chl a self-assembly and determining the type of its organization. Two empirical solvent parameter scales were used for quantitative characterization of the different solvation properties of the solvents and their mixtures with water. The mole fractions of water f_1/2 giving rise to the midpoint values of the relative fluorescence quantum yield were determined for each solvent, and then various solvent–water mixture parameters for the f_1/2 values were compared. On the basis of their comparison, it is concluded that the hydrogen-bonding ability and the dipole–dipole interactions (function of the dielectric constant) of the solvent–water mixtures are those that promote Chl a self-assembly. The influence of the different nature of the nonaqueous solvents on the Chl aggregation is manifested by both the different water contents required to induce Chl monomer → aggregate transition and the formation of two types of aggregates at the completion of the transition: species absorbing at 740–760 nm (in methanol, ethanol, acetonitrile, acetone) and at 667–670 nm (in pyridine and tetrahydrofuran). It is concluded that the type of Chl organization depends on the coordination ability and the polarizability (function of the index of refraction) of the organic solvent. The ordering of the solvents with respect to the f_1/2 values—methanol < ethanol < acetonitrile < acetone < pyridine < tetrahydrofuran—yielded a typical lyotropic (Hofmeister) series. On the basis of this solvent ordering and the disparate effects of the two groups of solvents on the Chl a aggregate organization, it is pointed out that the mechanism of Chl a self-assembly in aqueous media can be considered a manifestation of the Hofmeister effect, as displayed in the lipid-phase behavior (Koynova et al., Eur. J. Biophys. 25, 261–274, 1997). It relates to the solvent ability to modify the bulk structure and to distribute unevenly between the Chl–water interface and bulk liquid.

This paper reports that a new photobleaching compound, 2-(2-chloro-5-propoxycarbonylphenyl)aminomethylidene-5,5-dimethyl-cyclohexane-1,3-dione (RWH-21), stimulates accumulation of 13²-hydroxychlorophyll a in cultured tobacco cells. This was shown based on isolation of 13²-hydroxychlorophyll a from pigment extracts of cultured tobacco cells by diode-array HPLC and subsequent fast atom bombardment mass spectrometry analysis. 13²-Hydroxychlorophyll a rapidly accumulates in tobacco cells both in the light and dark in the presence of RWH-21 (50 μM). Analysis of 13²-hydroxychlorophyll a formation in tobacco cells indicates that 13²-hydroxychlorophyll a is rapidly accumulated within 20 h incubation time both in the dark and light. Although the amount of 13²-hydroxychlorophyll a is continuously increased in the dark, the amount of 13²-hydroxychlorophyll a decreased remarkably in the light after 20 h incubation. Analysis of 13²-hydroxychlorophyll a formation and lipid peroxidation by determination malondialdehyde in tobacco cells suggests that RWH-21-induced 13²-hydroxychlorophyll a has the potential to cause a photodynamic action in cultured tobacco cells.

Photoinactivation of vesicular stomatitis virus (VSV) in stroma-free hemoglobin (SFH) was carried out using methylene blue (MB) or 1,9-dimethylmethylene blue (DMMB). The VSV was more sensitive to inactivation by 660 nm light with 1 μM DMMB than with the same concentration of MB. Under conditions that inactivated 6 log₁₀ of VSV, the methemoglobin content (Met-Hb[%]) and P₅₀ of hemoglobin were changed by 1 μM MB phototreatment but were not changed by 1 μM DMMB phototreatment. The migration of hemoglobin during electophoresis and the activity of superoxide dismutase were not changed by MB or DMMB phototreatment. In contrast to the results obtained with DMMB at 660 nm, 580 nm irradiation of SFH with DMMB resulted in a significant increase of Met-Hb(%) under conditions that only inactivated 1.19 log₁₀ VSV. The 580 nm irradiation primarily activates the dimer and higher-order aggregates of the dyes, while 660 nm irradiation primarily activates the monomer. These results indicate that the monomer form of DMMB can effectively inactivate viruses without damage to SFH.

The acetoxymethyl ester of chlorin e6 (CAME) was initially designed to be a hydrophobic photosensitizing agent that would be recognized by an endocytic pathway and initially accumulated in lysosomes. This was expected to lead to hydrolysis of the ester groups, followed by redistribution of the free chlorin to other subcellular sites. In this study, we examined the patterns of localization of CAME and of subsequent photodamage in murine leukemia L1210 cells. The drug was initially localized at intracellular sites, yielding a pattern similar to that obtained with a fluorescent probe for acidic intracellular vesicles and endosomes. A brief (30 min) incubation with 10 μM CAME followed by irradiation led to mitochondrial photodamage and apoptotic cell death. At a higher drug level, or with a longer incubation time, we observed additional photodamage to the plasma membrane and to lysosomes. The higher photodynamic therapy dose led to inhibition of apoptosis, with cell death likely occurring via a necrotic process. Distribution of CAME among the components of human plasma was to albumin > high-density lipoprotein > low-density lipoprotein. These results have implications concerning the likely mechanism of CAME accumulation and subcellular distribution.

Oxidized LDL (ox-LDL) have been involved in the pathogenesis of several human diseases including dermatological pathologies. Oxidative modification of low-density lipoproteins (LDL) is accompanied by both extensive degradation of its polyunsaturated fatty acids and production of lipoperoxides. These highly reactive products induce an intracellular oxidative stress with a variety of cytotoxic effects. In order to evaluate cellular damage induced by oxidative stress in epidermal cells, a human epidermoid carcinoma cell line in culture (A 431) was used as experimental model. Cell treatment with UV-oxidized LDL resulted in cytostatic and cytotoxic effects characterized by morphological and functional alterations: inhibition of cell proliferation, modifications of cytoskeleton network, microtubular derangement, loss of cell–cell and cell–substrate contacts, cell detachment and cell death by apoptosis. The ox-LDL-induced alterations were almost completely prevented by pre-incubating cells with α-tocopherol. The results presented here could be of relevance for a better comprehension of the pathogenic mechanisms of several human diseases, including dermatological pathologies, and could indicate that antioxidants such as α-tocopherol could represent an important therapeutic challenge in the maintenance of cell and tissue homeostasis in the long run.

Possible association of photodynamic sensitization by cytochrome b₆/f complex (cyt b₆/f) via singlet oxygen (¹O₂) mechanism with photoinhibition damage to photosystem II (PS II) was studied using such subthylakoid preparations as photosystem I (PS I) particles, PS II core complex and cyt b₆/f from spinach leaves. Upon exposure to bright light, PS II core complex lost photosynthetic electron transport activity to a certain extent, whose spectral dependence implied that pheophytin a is likely involved in photoinactivation of PS II core complex in itself. The presence of PS I particles exerted virtually no effect on PS II core photoinactivation. However, the inclusion of cyt b₆/f in samples resulted in a marked exacerbation of the photoinactivation, particularly in UV-A and blue light. Such effect of cyt b₆/f was suppressed by azide and enhanced by the medium deuteration. Photogeneration of ¹O₂ from cyt b₆/f was confirmed by ESR and spectrophotometry, chemically trapping ¹O₂. Action spectra for both ¹O₂ photoproduction and PS II core photoinactivation by cyt b₆/f bore a close resemblance to each other, seemingly carrying the absorption characteristics of the Rieske Fe-S protein. A complex deficient in the Rieske protein prepared from intact cyt b₆/f showed virtually no generation of ¹O₂ in light, whereas an efficient photoformation of ¹O₂ was seen in the Rieske protein preparation. The results suggest that cyt b₆/f, rather specifically the Rieske center, may play a prominent role in photoinhibition processes through type II photosensitization in thylakoids.