This article reports a novel approach for the evaluation of fetal lung maturity based on fluorescence polarization (FP). The technique determines the intrinsic fluorescence polarization ratio (IFPR) of the amniotic fluid (AF). In vitro measurements of the IFPR indicate a clear dichotomy: high values for young pregnancies and low values for mature pregnancies. The new method has the potential to be a noninvasive procedure because the excitation of the AF and the collection of its fluorescence emission can be performed through the intact cervical amniotic membranes.

This report concerns physiological function of mycosporinelike amino acids (MAA) as an active defense against the photooxidative effects of sunlight in marine organisms. Mycosporine glycine (MG) is a representative member of MAA family and was found to effectively suppress various detrimental effects of the Type-II photosensitization in biological systems, such as inactivation of mitochondrial electron transport, lipid peroxidation of microsomes, hemolysis of erythrocytes and growth inhibition of Escherichia coli. The presence of MG in solutions of eosin Y or methylene blue resulted in a marked decrease in the level of singlet oxygen (¹O₂) produced by the sensitizers under illumination. The rate constant of ¹O₂ quenching by MG was determined to be 5.6 × 10⁷ M⁻¹s⁻¹ by the time-resolved ¹O₂ luminescence decay method, which is higher than, or at least comparable to, the values for ¹O₂ reaction of well-known quenchers such as 1,4-diazabicyclo[2,2,2]octane and furfuryl alcohol. The results suggest that MG probably together with some other active MAA may play an important role in protecting marine organisms against sunlight damage by eliminating ¹O₂ generated from certain endogenous photosensitizers.

In protein–cofactor reaction center (RC) complexes of purple photosynthetic bacteria, the major role of the bound carotenoid (C) is to quench the triplet state formed on the primary electron donor (P) before its sensitization of the excited singlet state of molecular oxygen from its ground triplet state. This triplet energy is transferred from P to C via the bacteriochlorophyll monomer B_B. Using time-resolved electron paramagnetic resonance (TREPR), we have examined the temperature dependence of the rates of this triplet energy transfer reaction in the RC of three wild-type species of purple nonsulfur bacteria. Species-specific differences in the rate of transfer were observed. Wild-type Rhodobacter capsulatus RCs were less efficient at the triplet transfer reaction than Rhodobacter sphaeroides RCs, but were more efficient than Rhodospirillum rubrum RCs. In addition, RCs from three mutant strains of R. capsulatus carrying substitutions of amino acids near P and B_B were examined. Two of the mutant RCs showed decreased triplet transfer rates compared with wild-type RCs, whereas one of the mutant RCs demonstrated a slight increase in triplet transfer rate at low temperatures. The results show that site-specific changes within the RC of R. capsulatus can mimic interspecies differences in the rates of triplet energy transfer. This application of TREPR was instrumental in defining critical energetic and coupling factors that dictate the efficiency of this photoprotective process.

Photodynamic therapy (PDT) has been used for many years for both palliative and curative treatment of bronchial carcinomas. However, prolonged skin phototoxicity and reduced depth of penetration has limited the widespread use of PDT. We studied the endobronchial phototoxicity of a novel photosensitizer, WST 09 (Tookad®). Fourteen pairs of Large White-Landrace male piglets were given intravenous WST 09 followed by laser light illumination of the left mainstem bronchus. Different settings for light dose (fluence), fluence rate (FR), drug dose (D) and drug–light interval (DLI) were applied to each pair. Bronchial toxicity was assessed with repeat bronchoscopic photographic evaluation as well as by pathologic examination following autopsy. Animals developed no toxicity, moderate toxicity or severe toxicity. Increased toxicity was seen with increasing D and fluence and decreasing DLI, whereas no increased toxicity was seen with higher FR. PDT-related histological changes in the normal bronchus confirm the vascular effect of WST 09. Depending on the parameter settings for fluence, D and DLI, the lesions ranged from focal intramucosal ischemia to transmural infarction with subsequent acute inflammation and fibrosis. Clinically feasible parameters for drug and light dosimetry were documented. These data will be important in determining safe starting doses for human phase I/II studies.

The photocycle in photoactive yellow protein (PYP) crystals was studied by single-crystal absorption spectroscopy with experimental setups for low-temperature and time-resolved measurements. Thin and flat PYP crystals, suitable for light absorption studies, were obtained using special crystallization conditions. Illumination of PYP crystals at 100 K led to the formation of a photostationary state, which includes at least one hypsochromic and one bathochromic photoproduct that resemble PYP_H and PYP_B, respectively. The effect of temperature, light color and light pulse duration on the occupancy of these low-temperature photoproducts was determined and appeared similar to that observed in solution. At room temperature a blueshifted photocycle intermediate was identified that corresponds to the blueshifted state of PYP (pB). Kinetic studies show that the decay of this blueshifted intermediate is biphasic at −12°C and 15-fold faster than that observed in solution at room temperature. These altered pB decay kinetics confirm a model that holds that the photocycle in crystals takes place in a shortcut version. In this version the key structural events of the photocycle, such as photoisomerization and reversible protonation of the chromophore, take place, but large conformational changes in the surrounding protein are limited by constraints imposed by the crystal lattice.

The macula of the human retina contains high amounts of the xanthophyll carotenoids lutein and zeaxanthin [a mixture of (3R,3′R)-zeaxanthin and (3R,3′S-meso)-zeaxanthin]. Recently, it was shown that the uptake and the stabilization of zeaxanthin and lutein into the retina are likely to be mediated by specific xanthophyll-binding proteins (XBP). Here, we have used femtosecond pump–probe spectroscopy to study the dynamics of the S₁ state of these xanthophylls in xanthophyll-enriched and native XBP. The results from the native XBP and the enriched XBP were then compared with those for carotenoids in organic solvents and in detergent micelles. Steady-state and transient absorption spectra show that the incorporation of xanthophylls into the protein causes a redshift of the spectra, which is stronger for lutein than for zeaxanthin. The transient absorption spectra further indicate that a part of the xanthophylls remains unbound in the xanthophyll-enriched XBP. The transient absorption spectra of the native XBP prove the presence of both xanthophylls in native XBP. Although the S₁ lifetime of lutein does not exhibit any changes when measured in solution, micelles or XBP, we have observed the influence of the environment on the S₁ lifetime of meso-zeaxanthin, which has a longer (12 ps) lifetime in XBP than in solution (9 ps). The most pronounced effect was found for vibrational relaxation in the S₁ state, which is significantly slower for xanthophylls in XBP compared with micelles and solution. This effect is more pronounced for meso-zeaxanthin, suggesting a specific site of binding of this carotenoid to XBP.

The quantum yield (QY) of the iodide–iodate chemical actinometer (0.6 M KI–0.1 M KIO₃) was determined for irradiation between 214 and 330 nm. The photoproduct, triiodide, was determined from the increase in absorbance at 352 nm, which together with a concomitant measurement of the UV fluence enabled the QY to be calculated. The QY at 254 nm was determined to be 0.73 ± 0.02 when calibration was carried out against a National Institute of Standards and Technology traceable radiometer or photometric device. At wavelengths below 254 nm the QY increased slightly, leveling off at ∼0.80 ± 0.05, whereas above 254 nm the QY decreases linearly with wavelength, reaching a value of 0.30 at 284 nm. In addition, the QY was measured at different iodide concentrations. There is a slight decrease in QY going from 0.6 to 0.15 M KI, whereas below 0.15 M KI the QY drops off sharply, decreasing to 0.23 by 0.006 M KI. Calibration of the QY was also done using potassium ferrioxalate actinometry to measure the irradiance. These results showed a 20% reduction in QY between 240 and 280 nm as compared with radiometry. This discrepancy suggests that the QY of the ferrioxalate actinometer in this region of the spectrum needs reexamination.

Both steady-state (SS) and time-resolved (TR) studies show that the fluorescence of the dye Nile red (NR) is quenched by various aromatic amines (ArA). Bimolecular quenching constants (k_q) from both SS and TR measurements are observed to match well, indicating that the interaction is dynamic in nature. The quenching interaction in the present systems has been attributed to electron transfer (ET) from ArA to excited NR, based on the variations in the k_q values with the oxidation potentials of the amines. The k_q values calculated within the framework of Marcus' outer-sphere ET theory at different free-energy changes (ΔG⁰) of the ET reactions match well with the experimental ones, supporting the ET mechanism in the systems studied. The reorganization energy (λ) estimated from the correlation of the experimental and the calculated k_q values is quite similar to the solvent reorganization energy (λ_s), calculated on the basis of the solvent dielectric continuum model along with the assumption that the reactants are the effective spheres. Although a modest error is involved in this λ_s calculation, the similarity in λ and λ_s values suggests that the solvent reorganization plays a dominant role in governing the ET dynamics in the present systems.

The reaction pathways for thermal and photochemical formation of cyclobutane pyrimidine dimers in DNA are explored using density functional theory techniques. Although it is found that the thermal [2 2] cycloadditions of thymine thymine (T T → T⋄T), cytosine cytosine (C C → C⋄C) and cytosine thymine (C T → C⋄T) all are similarly unfavorable in terms of energy barriers and reaction energies, the excited-state energy curves associated with the corresponding photochemical cycloadditions display differences that—in line with experimental findings—unanimously point to the predominance of T⋄T in UV-irradiated DNA. It is shown that the photocycloaddition of thymines is facilitated by the fact that the S₁ state of the corresponding reactant complex lies comparatively high in energy. Moreover, at a nuclear configuration coinciding with the ground-state transition structure, the excited-state energy curve displays an absolute minimum only for the T T system. Finally, the T T system is also associated with the most favorable excited-state energy barriers and has the smallest S₂–S₀ energy gap at the ground-state transition structure.

UVA radiation penetrates deeply into the skin reaching both the epidermis and the dermis. We thus investigated the effects of naturally occurring doses of UVA radiation on mitogen-activated protein kinase (MAPK) activities in human dermal fibroblasts. We demonstrated that UVA selectively activates p38 MAPK with no effect on extracellular-regulated kinases (ERK1–ERK2) or JNK–SAPK (cJun NH₂-terminal kinase–stress-activated protein kinase) activities. We then investigated the signaling pathway used by UVA to activate p38 MAPK. l-Histidine and sodium azide had an inhibitory effect on UVA activation of p38 MAPK, pointing to a role of singlet oxygen in transduction of the UVA effect. Afterward, using prolonged cell treatments with growth factors to desensitize their signaling pathways or suramin to block growth factor receptors, we demonstrated that UVA signaling pathways shared elements with growth factor signaling pathways. In addition, using emetine (a translation inhibitor altering ribosome functioning) we detected the involvement of ribotoxic stress in p38 MAPK activation by UVA. Our observations suggest that p38 activation by UVA in dermal fibroblasts involves singlet oxygen–dependent activation of ligand–receptor signaling pathways or ribotoxic stress mechanism (or both). Despite the activation of these two distinct signaling mechanisms, the selective activation of p38 MAPK suggests a critical role of this kinase in the effects of UVA radiation.

A hairpin polyamide–chlorobenzenesulfonyl conjugate was synthesized in solution by a haloform reaction and the dicyclohexylcarbodiimide/1-hydroxybenzotriazole (DCC/HOBT) coupling reaction. Its ability to cleave DNA was investigated with supercoiled DNA (pBluescript SK), and the DNA cleaving efficiency of chlorobenzenesulfonyl group was enhanced by the hairpin polyamide under UV irradiation (365 nm).

Broadband field measurements were conducted beneath three different-sized public shade structures, small, medium and large, during winter in the Southern Hemisphere. These measurements were compared with the diffuse UV to quantify the relationship of the UV under and around the shade structures to the diffuse UV. For the shade structures, a relationship between the diffuse UV and the UV in the shade has been provided for clear skies and solar zenith angles (SZA) of 49–76°. This allows the prediction of the UV in the shade of these structures if the diffuse UV is known. The ultraviolet protection factors for the three shade structures ranged from 1.5 to 5.4 for decreasing SZA. For the greater SZA of 70–76°, the erythemal UV in the shade was 65%, 59% and 51% of that in full sun for the small, medium and large structures, respectively. For the smaller SZA of 50–53° the erythemal UV in the shade was 35%, 41% and 18% for the small, medium and large shade structures, respectively. From this research it can be concluded that the UV radiation levels in the shade in winter could cause erythema and other sun-related disorders.

The thorough understanding of photosynthetic membrane assembly requires a deeper knowledge of the coordination of chlorophyll (Chl) and thylakoid apoprotein biosynthesis. As a working model for future investigations, we have proposed three Chl–thylakoid apoprotein biosynthesis models, namely, a single-branched Chl biosynthetic pathway (SBP) single-location model, an SBP multilocation model and a multibranched Chl biosynthetic pathway (MBP) sublocation model. Rejection or validation of these models can be probed by determination of resonance excitation energy transfer between various tetrapyrrole intermediates of the Chl biosynthetic pathway and various thylakoid Chl–protein complexes. In this study we describe the detection of resonance energy transfer between protoporphyrin IX (Proto), Mg-Proto and its monomethyl ester (Mp(e)) and divinyl and monovinyl protochlorophyllide a (Pchlide a) and several Chl–protein complexes. Induction of various amounts of tetrapyrrole accumulation in green photoperiodically grown cucumber cotyledons and barley leaves was achieved by dark incubation of excised tissues with delta-aminolevulinic acid (ALA) and various concentrations of 2,2′-dipyridyl for various periods of time. Controls were incubated in distilled water. After plastid isolation, treated and control plastids were diluted in buffered glycerol to the same Chl concentration. Excitation spectra were then recorded at 77 K at emission maxima of about 686, 694 and 738 nm. Resonance excitation energy transfer from Proto, Mp(e) and Pchlide a to Chl–protein complexes emitting at 686, 694 and 738 nm was observed by calculation of treated minus control difference excitation spectra. The occurrence of resonance excitation energy transfer between anabolic tetrapyrroles and Chl–protein complexes appeared as well-defined excitation bands with excitation maxima corresponding to those of Proto, Mp(e) and Pchlide a. Furthermore, it appeared that resonance excitation energy transfer from multiple short-wavelength, medium-wavelength and long-wavelength Proto, Mp(e) and Chlide a sites to various Chl–protein complexes took place. Because resonance excitation transfer from donors to acceptors cannot take place at distances larger than 100 Å, it is proposed that the observed resonance excitation energy transfers are not compatible with the SBP single-location Chl biosynthesis thylakoid membrane biogenesis model. The latter assumes that a single-branched Chl biosynthetic pathway located in the center of a 450 × 130 Å photosynthetic unit generates all of the Chl needed for the assembly of all Chl–protein complexes.

Native fluorescence characteristics of blood plasma were studied in the visible spectral region, at two different excitation wavelengths, 405 and 420 nm, to discriminate patients with different stages of oral malignancy from healthy subjects. The fluorescence spectra of blood plasma of oral malignant subjects exhibit characteristic spectral differences with respect to normal subjects. Different ratios were calculated using the fluorescence intensity values at those emission wavelengths that give characteristic spectral features of each group of experimental subjects studied. These fluorescence intensity ratios were used as input variables for a multiple linear discriminant analysis across different groups. Leave-one out cross-validation was used to check the reliability of each discriminant analysis performed. The discriminant analysis performed across normal and oral cancerous subjects classified 94.7% of the original grouped cases and 93.7% of the cross-validated grouped cases. A classification algorithm was developed on the basis of the score of the discriminant functions (discriminant score) resulted in the analyses. The diagnostic potentiality of the present technique was also estimated in the discrimination of malignant subjects from normal and nonmalignant diseased subjects such as liver diseases. In the discriminant analysis performed across the three groups, normal, oral malignancy (including early and advanced stages) and liver diseases, 99% of the original grouped cases and 95.9% of the cross-validated grouped cases were correctly classified. Similar analysis performed across normal, early stage of oral malignancy, advanced oral malignancy and liver diseases correctly classified 94.9% of the original grouped cases and 91.8% of the cross-validated grouped cases.

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.