INTERFERENCE BY NEAR ULTRAVIOLET AND GREEN LIGHT WITH GROWTH OF ANIMAL AND PLANT CELL CULTURES*

Abstract— Near ultraviolet (365 nm) and green-yellow (546-579nm) radiations repress the growth of liquid cell cultures of Ginkgo pollen and of monolayer cultures of HeLa. The deleterious effects of green-yellow wavelengths can be negated by red radiation; near-u.v.-induced growth repressions are insensitive to visible light photorestoration. These wavelengths do not interact synergistically and evoke different kinetics of response.     

Photoinactivation Sensitivity of Staphylococcus carnosus to Visible-light Irradiation as a Function of Wavelength

Inactivation properties of visible light are of increasing interest due to multiple possible fields of application concerning antibacterial treatment. For violet wavelengths, the generation of reactive oxygen species by porphyrins is accepted as underlying mechanism. However, there is still little knowledge about photosensitizers at blue wavelengths. While flavins were named as possible candidates, there is still no experimental evidence. This study investigates the photoinactivation sensitivity of Staphylococcus carnosus to selected wavelengths between 390 and 500 nm in 10- to 25-nm intervals. Absorption and fluorescence measurements in bacterial lysates confirmed inactivation findings. By means of a mathematical calculation in MATLAB^®, a fit of different photosensitizer absorption spectra to the measured action spectrum was determined to gain knowledge about the extent to which specific photosensitizers are involved. The most effective wavelength for S. carnosus at 415 nm could be explained by the involvement of zinc protoporphyrin IX. Between 450 and 470 nm, inactivation results indicated a broad plateau, statistically distinguishable from 440 and 480 nm. This observation points to flavins as responsible photosensitizers, which furthermore seem to be involved at violet wavelengths. A spectral scan of sensitivities might generally be an advantageous approach for examining irradiation impact.     

PHOTOBIOLOGY OF RNA BACTERIOPHAGES—I. ULTRAVIOLET INACTIVATION AND PHOTOREACTIVATION STUDIES*

Abstract— Biologically active f2-RNA, Obtained from bacteriophage f2, was inactivated by ultraviolet (u.v) light (2537 Å) with a quantum yield of 3.3 ± 0.3 times 10^-3 when assayed in the dark with protoplasts of an F- strain of E. coli k12. Assay under “black light” gave a quantum yield of 2.7 ± 0.5 times 10^-3 which was just enough lower to suggest that 17 per cent photorecovery of the u.v. lesions has taken place. Intact phage f2 was inactivated by u.v. radiation with a quantum yield of 0.7 ± 0.12 times 10^-3, Thus the whole phage is much less sensitive than the free RNA. No evidence of photorecovery was found in u.v.-irradiated RNA phage 7S assayed in its host Pseudomonas aeruginosa.     

Action spectra for oxygen-dependent and independent inactivation of Escherichia coli WP2s from 254 to 460 nm.

Abstract— Action spectra for lethality of E. coli WP2s under aerobic and anaerobic conditions. based on final slopes of the survival curves, reveal the absence of oxygen dependence at 313 nm and shorter wavelengths and a strong oxygen dependence (OER of 12 at 334 nm and 16 at 365 nm) at wavelengths longer than 313 nm. Shoulders or small peaks at 340, 365 , 410 and 500 nm suggest the participation of non-DNA chromophores in aerobic lethality at these wavelength ranges.     

High-Intensity 405 nm Light Inactivation of Listeria monocytogenes

The antimicrobial properties of light is an area of increasing interest. This study investigates the sensitivity of the significant foodborne pathogen Listeria monocytogenes to selected wavelengths of visible light. Results demonstrate that exposure to wavelength region 400–450 nm, at sufficiently high dose levels (750 J cm^?2), induced complete inactivation of a 5 log₁₀ population. Exposure to wavelengths longer than 450 nm did not cause significant inactivation. Analysis of 10 nm bandwidths between 400 and 450 nm confirmed 405(±5) nm light to be most effective for the inactivation of L. monocytogenes, with a lesser bactericidal effect also evident at other wavelengths between 400 and 440 nm. Identification of the optimum bactericidal wavelength enabled the comparison of inactivation using 405(±5) nm filtered light and a 405 nm light‐emitting diode (LED) array (14 nm FWHM). Results demonstrate similar inactivation kinetics, indicating that the applied dose of 405 nm light is the important factor. Use of the 405 nm LED array for the inactivation of L. monocytogenes and other Listeria species resulted in similar kinetics, with up to 5 log₁₀ reductions with a dose of 185 J cm^?2. Comparative data for the 405 nm light inactivation of L. monocytogenes and other important foodborne pathogens, Escherichia coli, Salmonella enteritidis and Shigella sonnei, are also presented, with L. monocytogenes showing higher susceptibility to inactivation through 405 nm light exposure.     

Wavelength sensitivity of the photoinduced reaction in polycarbonate

Bisphenol A polycarbonate (PC) was irradiated with monochromatic light of wavelengths 260, 280, 300, 320, 340, 400, and 500 nm by use of the Okazaki large spectrograph (OLS). The quantum yield of main-chain scission (?_cs), efficiency of photo-Fries rearrangement (E_r), and effects of wavelength on ?_cs and E_r were investigated. It was found that photodegradation and photo-Fries rearrangement of PC took place by the irradiation of 260–300 nm light, but did not by the irradiation at λ ≧ 320 nm. The ?_cs has a maximum value in the case of the irradiation with 260 nm light, while E_r was found to have a maximum value by the irradiation of 280 nm light. © 1993 John Wiley & Sons, Inc.     

Ultraviolet action spectra in perspective: with special reference to mutation

Abstract— Problems of determining action spectra are considered as well as various types of action spectra for U.V. action upon cell activities. U.V. is an effective mutagenic agent producing point mutations and chromosomal changes. U.V. is readily absorbed by superficial layers of cells in tissues; therefore, special experimental procedures are necessary for induction of mutations in animals or plants. U.V. is, however, suitable for mutagenesis in microorganisms because their cells are small, permitting the radiation to reach the nuclei. Action spectrum studies reveal that u.v. mutagenesis results from absorption of the radiation by nucleic acid. The most prominent alteration in DNA following absorption of u.v. is dimerization of pyrimidines, chiefly thymine. Such a change not only retards DNA replication but results in errors (mutations). U.V. mutagenesis therefore depends upon the conditions before, during and after irradiation. Thus immediate post-treatment with visible and long u.v. light splits pyrimidine dimers, thereby reversing impending u.v. mutagenesis. For cells kept in the dark, conditions which prevent DNA replication by interfering with the metabolism of the cell provide time for dark repair of the DNA lesion and so for reversal of the impending mutation.     

PHOTOACTIVATION OF UROCANASE IN PSEUDOMONAS PUTIDA: POSSIBLE CONFORMATIONAL CHANGE DURING ACTIVATION

Earlier studies of urocanase indicated that it is converted to an inactive form in resting cells of Pseudomonas putida . When cell extracts are irradiated by ultraviolet (u.v.) light, the enzyme is activated[1]. The chromophore is closely associated with the enzyme [ 1 ,2]. The action spectrum for photoactivation showed that, although near-u.v. light was effective, the most efficient wavelengths were at 275–280 nm[3]. The study we present here suggests that urocanase undergoes a conformational change upon photoactivation.     

Evidence from action and fluorescence spectra that UV-induced violet-blue-green fluorescence enhances leaf photosynthesis

We assessed the contribution of UV-induced violet-blue-green leaf fluorescence to photosynthesis in Poa annua, Sorghum halepense and Nerium oleander by measuring UV-induced fluorescence spectra (280-380 nm excitation, 400-550 nm emission) from leaf surfaces and determining the monochromatic UV action spectra for leaf photosynthetic O2-evolution. Peak fluorescence emission wavelengths from leaf surfaces ranged from violet (408 nm) to blue (448 nm), while excitation peaks for these maxima ranged from 333 to 344 nm. Action spectra were developed by supplementing monochromatic radiation from 280 to 440 nm, in 20 nm increments, to a visible nonsaturating background of 500 mumol m-2 s-1 photosynthetically active radiation and measuring photosynthetic O2-evolution rates. Photosynthetic rates tended to be higher with the 340 nm supplement than with higher or lower wavelength UV supplements. Comparing photosynthetic rates with the 340 nm supplement to those with the 400 nm supplement, the percentage enhancement in photosynthetic rates at 340 nm ranged from 7.8 to 9.8%. We suspect that 340 nm UV improves photosynthetic rates via fluorescence that provides violet-blue-green photons for photosynthetic energy conversion because (1) the peak excitation wavelength (340 nm) for violet-blue-green fluorescence from leaves was also the most effective UV wavelength at enhancing photosynthetic rates, and (2) the magnitude of photosynthetic enhancements attributable to supplemental 340 nm UV was well correlated (R2 = 0.90) with the apparent intensity of 340 nm UV-induced violet-blue-green fluorescence emission from leaves.     

ACTION SPECTRA FOR LETHALITY IN RECOMBINATION-LESS STRAINS OF SALMONELLA TYPHIMURIUM AND ESCHERICHIA COLI*

Abstract— Action spectra for lethality of both stationary and exponentially growing cells of recombinationless (recA) mutants of Salmonella typhimurium and Escherichia coli were obtained. Maximum sensitivity was observed at 260nm which corresponds to the maximum absorbance of DNA. However, a shoulder occurred in the 280–300 nm range that departed significantly from the absorption spectrum of DNA. At wavelengths longer than 320nm, the shapes of inactivation curves departed significantly from those at wavelengths shorter than 320nm and survival curves at wavelengths longer than 320nm had a large shoulder. A small peak or shoulder occurred in the 330–340nm region of the action spectra. The special sensitivity of recA mutants to broad spectrum near-UV radiation may be due to synergistic effects of different wavelengths. Parallels between the inactivation of recA mutants and the induction of a photoproduct of l -tryptophan toxic for recA mutants (now known to be H₂O₂) suggest that H₂O₂ photoproduct from endogenous tryptophan may be involved in the high sensitivity of these strains to broad spectrum near-UV radiation.     

Light induced synchronization of algal species that divide preferentially in darkness

Abstract— Three algal species (Protosiphon botryoides, Chlorella pyrenoidosa and Chlamy-domonas gymnogama) which divide preferentially at night during natural or simulated day-night conditions are shown to have cell division inhibited by light in the blue region of the visible spectrum (400–500 nm) and stimulated by the remainder of this spectrum (500–730 nm). Synchronous division has been established in cultures of these organisms on a circadian or longer period by alternating fluorescent cool-white light and cool-white with the blue component removed. This procedure is suggested as potentially superior to other methods for synchronizing cultures of algae that prefer, in nature, to undergo cell division at night. Where this procedure does not work it is recommended that an action spectrum for cell division be obtained, since this may provide information for achieving optimal synchrony through the use of other specific wavelength regimes.     

Photoreactivation of killing in Streptomyces. 3. Action spectra for photolysis of pyrimidine dimers and adducts in S. griseus and S. griseus PHR-1

Abstract— Photolysis of tritium-labelled thymine-derived photoproducts by 254-nm ultraviolet radiation (u.v.) in conidia of Streptomyces griseus was measured by chromatography of cell hydrolysates. The relative photolysis cross-sections of uracilthymine dimer (UT○) at various wavelengths are the same as those of thymine-thymine dimer (TT○), and their ratios at 313, 365, 405 and 436 nm are 2:1:2:3. Except at 436 nm, these relative values agree very well with cross-sections previously reported for photoreactivation of u.v. killing in this organism, leading to the conclusion that photoreactivation in the wild type is due to repair of cyclobutane-type pyrimidine dimers. In a mutant showing restricted photoreactivation (S. griseus PHR-1), post-u.v. treatments at the above wavelengths did not affect UT○ and TT○ in the conidia, supporting the earlier suggestion that this organism does not contain active PR enzyme. Another u.v. photoproduct, the precursor of a pyrimidine adduct (PO-T) that appears in cell hydrolysates, was removed from both wild-type and mutant cells very efficiently at 313 nm. This is presumably a direct photochemical reaction. In addition, in wild-type cells, the precursor of PO-T appeared to be inefficiently removed photoenzymatically at all wavelengths. Removal of the precursor of PO-T appears to be biologically significant, however, only in the mutant.     

GENETIC AND PHYSIOLOGICAL STUDIES OF THE EFFECT OF LIGHT ON THE DEVELOPMENT OF THE MOSS, PHYSCOMITRELLA PATENS

The germination of Physcomitrella patens spores only occurs when wet spores are exposed to light. Depending on their ripeness, spores require from 44 to 64 h illumination to bring about maximum germination. There is a lag period of about 15 h between the reception of sufficient light to elicit germination before germination can be observed. Wavelengths in the range 640–64080 nm are much more effective in inducing germination than longer or shorter wavelengths, but far-red reversal of red light induction of germination has not been demonstrated. Light also has very marked effects on protonemal and gametophore development. In darkness, only caulonemata are produced, and these grow negatively geotropically. No new gametophores develop but existing gametophores grow negatively geotropically, etiolate and bear only scale leaves. In light, chloronemata, as well as caulonemata are produced, the former grow positively phototropically, while the latter grow at right angles to the direction of light, and neither cell type is sensitive to gravity. In the light, gametophores grow positively phototropically, are indifferent to gravity, produce large leaves and do not etiolate. All these responses to light by protonemata and gametophores are shown by cultures growing in a 23 h dark/l h red light cycle, but if this red light treatment is followed by 15min far-red light, the effect of the red light is reversed, indicating an involvement of phytochrome in the mediation of these responses. Mutants showing abnormal growth in the dark have been isolated, as well as mutants having abnormal phototropic responses. The latter type has lost the phototropic response of both the protonemal cell types, as well as of gametophores, indicating that these different responses may share a common component.     

Intramolecular NH/pi complexes of 2-allylaniline derivatives in the ground and excited states

The photochemical and photophysical properties of three 8-allyl-1,2,3,4-tetrahydroquinolines (1a-c) have been studied. These compounds exhibit a 2-allylaniline-like photochemical behavior, undergoing photocyclization to lilolidines (3a-c). The absorption, emission, and excitation spectra of 1a-c, employing convenient model compounds for comparison, demonstrate the formation of a NH/pi intramolecular ground-state complex (AB). This species can absorb light at long wavelengths (330-340 nm), giving rise to the corresponding excited complex AB*. Emission from AB* is red-shifted (420 nm) with respect to that observed when the monomer A is excited (lambda(exc) = 300 nm). These experimental results have been rationalized by means of density-functional theory calculations.     

Near-U.V. photolysis of pyrimidine heteroadducts in E. coli DNA

Abstract— The cytosine-thymine precursor of the U–T adduct is not subject to enzymatic photoreactivation, but can be eliminated directly from u.v.-irradiated E. coli DNA by exposure to wavelengths around 313 nm.     

LIGHT-INDUCED SYNTHESIS OF ANTHOCYANIN IN CARROT CELLS IN SUSPENSION—IV. THE ACTION SPECTRUM

Abstract— Using carrot cell suspension in 2,4-dichlorophenoxyacetic acid (2,4-D)-depleted culture medium, fluence-response curves for the formation of anthocyanin were determined at various wavelengths from 250 to 800 nm. In the fluence-response curves at wavelengths between 260 and 330 nm, the response showed a sharp fluence-dependent increase after the fluence exceeded threshold level at the respective wavelength. Such a sharp increase in response was not observed by light at 450 nm or longer wavelengths, although the response obtained by higher fluence of such light was always higher than that in the dark control. Action spectra determined at the sharp increasing phase of the response showed the single peak at 280 nm which equals the absorption maximum of UV-B photoreceptor.
Although red (R)-light alone had a minor effect on anthocyanin accumulation, it modulated the action of UV-B light. That is, when carrot cells were irradiated with R-light either before or after UV-B irradiation, anthocyanin formation was greatly enhanced above the level enhanced by UV-B light alone. The most effective wavelength for this enhancement was 660 nm. The effect of R-light on the anthocyanin formation of the UV-B irradiated cells was reversed by immediately following it with far-red light, suggesting the involvement of phytochrome in the R-effect.     

In situ, rapid, and temporally resolved measurements of cellulase adsorption onto lignocellulosic substrates by UV-vis spectrophotometry

This study demonstrated two in situ UV-vis spectrophotometric methods for rapid and temporally resolved measurements of cellulase adsorption onto cellulosic and lignocellulosic substrates during enzymatic hydrolysis. The cellulase protein absorption peak at 280 nm was used for quantification. The spectral interferences from light scattering by small fibers (fines) and particulates and from absorptions by lignin leached from lignocelluloses were corrected using a dual-wavelength technique. Wavelengths of 500 and 255 nm were used as secondary wavelengths for correcting spectral interferences from light scattering and absorption of leached lignin. Spectral interferences can also be eliminated by taking the second derivative of the measured spectra of enzymatic hydrolysate of cellulose or lignocelluloses. The in situ measured cellulase adsorptions in cellulose and lignocellulose suspensions by these two spectrophotometric methods showed general agreement with batch sampling assayed by the Bradford method. The in situ methods not only eliminated tedious batch sampling but also can resolve the kinetics of the initial adsorption process. The measured time-dependent cellulase adsorptions were found to follow pseudo-second-order kinetics.     

SENSITIVITY OF STRAINS OF ESCHERICHIA COLI DIFFERING IN REPAIR CAPABILITY TO FAR UV,NEAR UV AND VISIBLE RADIATIONS*

Abstract— In stationary phase, strains of Escherichia coli deficient in excision (B/r Her) or recombination repair (K.12 AB2463) were more sensitive than a repair proficient strain (B/r) to monochromatic near-ultraviolet (365 nm) and visible (460 nm) radiations. The relative increase in sensitivity of mutants deficient in excision or recombination repair, in comparision to the wildtype, was less at 365 nm than at 254 nm. However, a strain deficient in both excision and recombination repair (K12 AB2480) showed a large, almost equal, increase in sensitivity over mutants deficient in either excision or recombination repair at 365 nm and 254 nm. All strains tested were highly resistant to 650 nm radiation. Action spectra for lethality of strains B/r and B/r Her in stationary phase reveal small peaks or shoulders in the 330–340, 400–410 and 490–510 nm wavelength ranges. The presence of 5μg/ml acriflavine (an inhibitor of repair) in the plating medium greatly increased the sensitivity of strain B/r to radiation at 254, 365 and 460 nm, while strains E. coli B/r Her and K12 AB2463 were sensitized by small amounts. At each of the wavelengths tested, acriflavine in the plating medium had at most a small effect on E. coli K.12 AB2480. Acriflavine failed to sensitize any strain tested at 650 nm. Evidence supports the interpretation that lesions induced in DNA by 365 nm and 460 nm radiations play the major role in the inactivation of E. coli by these wavelengths. Single-strand breaks (or alkali-labile bonds), but not pyrimidine dimers are candidates for the lethal DNA lesions in uvrA and repair proficient strains. At high fluences lethality may be enhanced by damage to the excision and recombination repair systems.     

Phototoxic effect of visible light on Porphyromonas gingivalis and Fusobacterium nucleatum: an in vitro study

The antibacterial effect of visible light irradiation combined with photosensitizers has been reported. The objective of this was to test the effect of visible light irradiation without photosensitizers on the viability of oral microorganisms. Strains of Porphyromonas gingivalis, Fusobacterium nucleatum, Streptococcus mutans and Streptococcus faecalis in suspension or grown on agar were exposed to visible light at wavelengths of 400-500 nm. These wavelengths are used to photopolymerize composite resins widely used for dental restoration. Three photocuring light sources, quartz-tungsten-halogen lamp, light-emitting diode and plasma-arc, at power densities between 260 and 1300 mW/cm2 were used for up to 3 min. Bacterial samples were also exposed to a near-infrared diode laser (wavelength, 830 nm), using identical irradiation parameters for comparison. The results show that blue light sources exert a phototoxic effect on P. gingivalis and F. nucleatum. The minimal inhibitory dose for P. gingivalis and F. nucleatum was 16-62 J/cm2, a value significantly lower than that for S. mutans and S. faecalis (159-212 J/cm2). Near-infrared diode laser irradiation did not affect any of the bacteria tested. Our results suggest that visible light sources without exogenous photosensitizers have a phototoxic effect mainly on Gram-negative periodontal pathogens.     

Protochlorophyllide a: A Comprehensive Photophysical Picture

The photochemistry of protochlorophyllide a, a precursor in the biosynthesis of chlorophyll and substrate of the light regulated enzyme protochlorophyllide oxidoreductase, is investigated by pump‐probe spectroscopy. Upon excitation into the lowest lying Q‐band the light induced changes are recorded over a wide range of probe wavelengths in the visible and near‐IR region between 500 and 1000 nm. Following excitation, an initial ultrafast 450 fs process is observed related to the motion out of the Franck‐Condon region on the excited state surface; thus directly unraveling previous suggestions based on time‐resolved fluorescence measurements (ChemPhysChem 2006 , 7, 1727–1733). Furthermore, the data reveals a previously concealed photointermediate, whose formation on a nanosecond timescale matches the overall fluorescence decay and is assigned to a triplet state. The implications of this finding with respect to the photochemistry of NADPH:protochlorophyllide oxidoreductase (POR) are discussed.