Although ultraviolet-B (UV-B) radiation is potentially harmful, it is an important component of terrestrial radiation to which plants have been exposed since invading land. Since then, plants have evolved mechanisms to avoid and repair UV radiation damage; therefore, it is not surprising that photomorphogenic responses to UV-B are often assumed to be adaptations to harmful radiation. This presupposes that the function of the observed responses is to prevent UV damage. It has been hypothesized that, as with blue light, UV-B provides a signal important for normal plant development and might be perceived within developing plants through nondestructive processes, perhaps through UV-specific signal perception mechanisms. UV signal perception can lead to photomorphogenic responses that may confer adaptive advantages under conditions associated with high-light environments, such as water stress. Plant responses to UV radiation in this regard include changes in leaf area, leaf thickness, stomatal density, photosynthetic pigment production and altered stem elongation and branching patterns. Such responses may lead to altered transpiration rates and water-use efficiencies. For example, we found that the cumulative effect of ambient UV-B radiation upon stomatal density and conductance can lead to altered water-use efficiencies. In field settings, UV might more properly be viewed as a photomorphogenic signal than as a stressor. Hence, it might be insufficient to attempt to fully evaluate the adaptive roles of plant responses to UV-B cues upon stress tolerance by the simultaneous application of UV and drought stress during development. We propose that rather than examining a plant's response to combinations of stressors one might also examine how a plant's response to UV induces tolerance to subsequently applied stresses.

There is growing interest regarding the joint effects of elevated levels of surface ultraviolet B (UV-B) radiation, carbon dioxide (CO₂) and ozone (O₃) on plants. Our current knowledge of this subject is too limited to draw any specific conclusions, although one might state that such effects are likely to be highly species dependent and may be more than additive, additive or less than additive. There are a number of uncertainties associated with the experimental protocols used and the conclusions reached in many studies. Nevertheless, in North America, there appear to be genotypes of three monocot crop species (Avena sativa L., Oryza sativa L. and Sorghum vulgare L.); six dicot crops (Cucumis sativus L., Lactuca sativa L., Lycopersicon esculentum Mill., Phaseolus vulgaris L., Pisum sativum L. and Solanum tuberosum L.) and two conifer species (Pinus ponderosa Dougl. and Pinus taeda L.) that may be considered sensitive to the joint effects of elevated levels of UV-B, CO₂ and O₃. However, to provide a more reliable assessment or validation of the predictions, future research must consider the concept of plant response surfaces and describe them more fully in numerical terms. Achieving that objective will require close cooperation among a number of scientists representing geographic locations with known spatial and temporal differences in UV-B, CO₂ and O₃ to conduct experiments under their site-specific conditions, using common plant materials and experimental protocols.

Protochlorophyllide (Pchlide) reductases are key enzymes in the process of chlorophyll biosynthesis. In this review, current knowledge on the molecular organization, substrate specificity and assembly of the light-dependent reduced nicotinamide adenine dinucleotide phosphate:Pchlide oxidoreductases are discussed. Characteristics of light-independent enzymes are also described briefly, and the possible reasons for the selection of light-dependent enzymes during the course of evolution are discussed.

A photophysical and photochemical study of a biphotochromic compound where two naphthopyran units are linked by an acetylene–thiophene–acetylene bridge has been carried out in toluene. Both fluorescence and intersystem crossing to the triplet manifold were found to compete with the photocoloration process. Two photoproducts (transoid-trans and transoid-cis stereoisomers), absorbing at ∼480 nm and corresponding to the opening of a single photochromic unit, were detected by spectrophotometric analysis after short irradiation time in diluted solution and identified by ¹H–nuclear magnetic resonance (NMR) spectroscopy. After prolonged irradiation at 228 K of highly concentrated solutions (up to 3 × 10⁻³ mol dm⁻³), two additional isomers, absorbing at ∼550 nm, were formed. Their NMR spectra indicate the opening of both photochromic units. An interesting effect of selective vibronic excitation was found, showing that the photoreaction is favored at excited vibronic levels to the detriment of the radiative relaxation.

We have extended our study of the decomposition of urocanic acid (UCA) with ultraviolet A radiation (UV-A) by the self-sensitized generation of singlet oxygen (see Photochem. Photobiol. 75, 565 [2002]). The chemistry has been found to be partially dependent on the presence of trace metal, most likely iron. Rigorous removal of metal impurities from the reaction mixture, using Chelex, retarded (but did not eliminate) the UV-A–initiated UCA degradation. The addition of small amounts of ferric chloride to the Chelex-treated solutions restored reactivity. Chelex treatment had a modest effect on the previously reported ability of UCA photoproducts to photonick supercoiled plasmid DNA. Also, photoinactivation of Sindbis virus on irradiation with the UCA photoproducts is now reported. Inactivation of the virus by a photoproduct mixture derived from a UCA solution that had been pretreated with Chelex was less rapid and gave better behaved time-course plots than was observed for photoproducts from non–Chelex treated solutions. These results are particularly noteworthy in light of the ubiquitous presence of both UCA and iron in the skin.

Excited state lifetime and amplitude measurements were made on thiazole orange dimer (TOTO), a dimeric DNA-intercalating fluorophore, at single-molecule concentrations. As expected from previous study, the excited state lifetime of TOTO intercalated in DNA is dependent on the sequence of the double-stranded DNA, having values of 2.2 ns in poly-GC DNA and 1.8 ns in poly-AT DNA. The distribution of excited state lifetimes of single molecules of TOTO intercalated into oligonucleotides having varying proportions of poly-GC sequences relative to poly-AT sequences were analyzed as a function of the fraction of poly-GC. By using excited state lifetime distributions from the purely GC and purely AT oligonucleotides as a basis set, it was possible to estimate the GC content of oligonucleotides with intermediate GC composition to within a few percent error. This serves as a model for the analysis of equilibrium binding distributions in DNA using single-molecule methods.

The sequence dependence of the double-stranded DNA (dsDNA)–binding affinity of TOTO, a thiazole orange dimer that functions as a DNA-intercalating fluorophore, was measured using single-molecule methods. An analysis was performed of the distribution of excited-state lifetimes of single molecules of TOTO intercalated into dsDNA fragments containing four–base pair sequences shown previously to have high affinity for TOTO under conditions used in nuclear magnetic resonance (NMR) spectroscopy. For the current studies, the putative binding sites were located centrally in 30–base pair–long dsDNA fragments in which the remaining sequence was either all poly-AT or poly-GC. The lifetime of TOTO fluorescence when bound to these fragments was entirely determined by the background sequence, i.e. DNA fragments with a poly-AT background predominantly gave a fluorescence lifetime of 1.7 ns, whereas DNA fragments with a poly-GC background gave a lifetime of 2.0 ns, independent of the presence or absence of the putative binding sequence. By performing competitive binding experiments in which these DNA fragments competed for TOTO binding with pure poly-AT fragments and using single-molecule fluorescence methods to determine the number of each type of DNA fragment having a TOTO bound in an equilibrium mixture, the relative binding affinity of each putative binding site was determined. The results of these experiments showed clearly that TOTO has no preference for binding to the putative binding sites over binding poly-AT or poly-GC under the conditions of these measurements. This suggests that there is very little sequence dependence of TOTO binding under conditions that would likely predominate in most biological applications of this intercalating dye.

This study relates to nanoparticle (NP) platforms that attach to tumor cells externally and only deliver singlet oxygen for photodynamic therapy (PDT) while conserving the embedded photosensitizers (PS). As a model, we demonstrate the successful embedding of the PS meta-tetra(hydroxyphenyl)-chlorin (m-THPC) in NP that are based on a sol–gel silica matrix and also show its positive effect on the singlet oxygen production. The embedding of m-THPC inside silica NP is accomplished by a modified Stöber sol–gel process, in which (3-aminopropyl)-triethoxysilane is introduced during the reaction. Singlet oxygen delivery by the targetable photodynamic NP exceeds that from free PS molecules. In the physiological pH range, there is no significant pH-induced decrease in the fluorescence of m-THPC embedded in silica NP, which might otherwise affect the efficiency of PDT.

Fixation of DNA and proteins in the isolated rat hepatocyte nuclei stained with ethidium bromide and irradiated with visible light was analyzed in this study. It was shown that irradiation results in the following modifications of higher-level nucleoprotein complexes of interphase chromatin: (1) the complexes acquire resistance to decondensing treatments, which may be indicative of the formation of links between proteins or proteins and DNA in the chromatin; (2) the linking rate for both DNA and proteins is dose dependent; (3) the irradiation induces intermolecular link formation between DNA molecules, which brings about an increase in the average molecular weight of DNA fragments; (4) some modifications (dimerization, etc.) of histones and nonhistone proteins occur; and (5) histone proteins are not effectively cross-linked to DNA. The structural stabilization of interphase chromatin is possibly mediated by free radical–based mechanisms, whereas disulfide bonds seem to play no significant role in the cross-linking.

We describe a fluorescent method that allows to differentiate the worms Eisenia fetida and Eisenia andrei. In fact, the coelomic fluid of E. andrei displays specific fluorescence absent in that of E. fetida. The two species do not metabolize the same types of molecules and thus can be differentiated at the molecular level. Each species has specific fluorescence fingerprints.

Aspects of different calibration procedures for erythemally weighing broadband radiometers are presented in this study. These instruments are common in projects dealing with ultraviolet radiation effects on humans. Many erythemally weighing broadband radiometers are still operated using a single calibration factor (cf) that is provided with the instrument. The individual characteristics of every instrument are strongly dependent on the total ozone amount and the solar elevation. Therefore, a calibration procedure also has to take into account the ozone concentrations and the solar elevation to compensate for the effects of the individual characteristics and to provide comparable measurements. Given the variation of the ozone concentrations and the solar elevation, an individual cf has to be calculated for every measurement. Using a simplified version of the calibration procedure, which is presented in this study, can lessen this effort. Taking into account the relevant meteorological conditions for a measuring site, a single cf is calculated to compensate the individual characteristics of the instruments and therefore deliver comparable measurements with less effort.

The aim of this study was to investigate the in vitro cellular accumulation, distribution and photocytotoxic effect of hypericin in two-dimensional (2-D) and three-dimensional (3-D) cultured RT-112 transitional cell carcinoma cells of the bladder. In addition, two iodinated derivatives of hypericin were incorporated to investigate whether these analogs, with their increased lipophilicity and heavy–atom effect, display a different biological behavior and optimized photodynamic effect. The results indicate that hypericin and mono–iodohypericin behave similarly in terms of cellular accumulation, spheroidal distribution and photocytotoxic effect. In contrast, di-iodohypericin concentrated to a higher extent in monolayers and spheroids, but the accumulation was restricted to the outermost part of the spheroid. An inverse correlation therefore seems to exist between the extent of cellular uptake under 2-D conditions and the penetration of the compounds in multicellular systems. Moreover, a less pronounced photocytotoxic effect was observed for di-iodohypericin in both 2-D and 3-D cell culture systems. It can be concluded that iodinated derivatives of hypericin do not show an increased cytotoxic effect upon irradiation in either monolayers or spheroids. Moreover, this study shows that when new photosensitizers are preclinically developed, the use of 3-D cell aggregates is critical for a correct evaluation of their efficacy.

Cellular responses to photodynamic therapy (PDT) include induction of heat shock proteins (HSP). We examined meso-tetrahydroxyphenyl chlorin (mTHPC) PDT–mediated HSP activation in EMT6 cells stably transfected with a plasmid containing the gene for green fluorescent protein (GFP) driven by an hsp70 promoter. mTHPC incubation induced concentration-dependent GFP expression. Irradiation of cells exposed to a sensitizer concentration that induced a slight increase in GFP and no loss of cell viability resulted in fluence-dependent GFP accumulation. In response to drug only and to PDT, GFP levels increased to a maximum of four- to five-fold above control levels with increasing drug or fluence and then decreased at higher doses. A trypan blue–exclusion assay confirmed that decreased GFP levels in both cases were due to a loss of cell viability. For initial evaluation in vivo, HSP70/GFP–transfected EMT6 tumors were grown in BALB/c mice and subjected to mTHPC-PDT with a fluence of 1 J/cm². Six hours after PDT, GFP fluorescence was imaged in these tumors through the intact skin in vivo. These results indicate that sublethal doses of mTHPC-PDT stimulate GFP expression under the control of an hsp70 promoter and illustrate the potential of noninvasively monitoring reporter protein fluorescence as a measure of molecular response to PDT.

The effectiveness of the combination of retinoids with 8-methoxypsoralen (8-MOP) and ultraviolet-A (UV-A) light in the treatment of some cutaneous proliferative diseases has motivated the synthesis of new “chimera-type” molecules built from psoralen derivatives and retinoic amides and related molecules. The chimeras result from the combination of 8-(3-bromopropyloxy)-psoralen with amides prepared by reacting 4-amino-pyridine with 13E- and 13Z-retinoic acids or a “retinoid-like” derivative with an alkene chain of only three double bonds. The synthesis of chimeras built with the 8-(3-bromopropyloxy)-psoralen and the amide of cinnamic acid or its 4-methoxy derivative has also been carried out. In contrast to 8-MOP, all the chimeras exhibit strong molar absorptivities in the range 20 000–40 000 M⁻¹ cm⁻¹ in the 340–390 nm UV-A region. The “retinoid-like”– and retinoid–psoralen chimeras are characterized by a marked dark toxicity toward proliferating NCTC 2544 keratinocytes (with a lethal dose corresponding to 50% cell survival [LD₅₀] of 1–5 μM) as compared with that of the cinnamic acid derivative–psoralen chimeras (LD₅₀ ≥ 50 μM). This toxicity leads to alteration of the mitochondrial membrane potential. At nontoxic concentrations, the chimeras demonstrate effective psoralens UV-A–induced photocytotoxicity. They are moderate photosensitizers of membrane lipid peroxidation. Cell apoptosis is a major photocytotoxic process as suggested by the fluorescence-activated cell-sorting technique using annexin–fluorescein isothiocyanate and propidium iodide as apoptotic markers.

In a Danish population, basal cell carcinoma (BCC) patients have a higher dermal mast cell prevalence in buttock skin than controls. This finding was supported by a functional link in mice between histamine-staining dermal mast cells and the extent of susceptibility to UV-B–induced systemic immunomodulation. It was important to confirm that this association was maintained in an Australian population with very different ancestry and sun exposure patterns. Australian BCC patients (n = 26) had significantly higher densities of mast cells in the dermis of buttock skin than control subjects (n = 25) (P = 0.0003, Mann–Whitney U-test). However, this correlation was lost at the sun-exposed site of the hand (P = 0.547, Mann–Whitney U-test). To further evaluate whether a relationship exists between dermal mast cell prevalence in sun-exposed skin and incidence of BCC in a larger study, biopsies of dorsal hand skin were obtained from an age-stratified random sample of 166 Queensland subjects, together with the 51 South Australian subjects, and dermal mast cell prevalence was quantified. Older subjects (over the median age of 42 years) had a greater incidence of BCC development (P = 0.0001, chi-square test) and significantly higher mast cell densities in hand skin (P = 0.0001, chi-square test) than younger subjects. However, mast cell density in sun-exposed hand skin was not significantly associated with BCC incidence. Finally, cellular expression of c-kit correlated with mast cell prevalence in non–sun-exposed skin, thereby implicating the stem cell factor–c-kit axis in the intrinsic mechanisms that regulate prevalence. These results show that high prevalence of dermal mast cells in buttock skin but not hand is associated with BCC development in an Australian population.

The nonlinear optical membrane used for image processing was prepared using tetraethyl silicate by sol–gel method under mild conditions. Ru(bpy)₃Cl₂ (bpy, bipyridine) was immobilized in the sol–gel matrix. This light-excitable membrane containing Ru(bpy)₃Cl₂ as photocatalyst could catalyze the Belousov–Zhabotinsky reaction by illumination. Our results indicated that a fine-definition image processing could be autoperformed when projecting a picture on this medium. The imaging process was an image evolution process, which was similar to that of the human vision process.