UVB Dose Research Articles

UVB induces a genome-wide acting negative regulatory mechanism that operates at the level of transcription initiation in human cells.

Faithful transcription of DNA is constantly threatened by different endogenous and environmental genotoxic effects. Transcription coupled repair (TCR) has been described to stop transcription and quickly remove DNA lesions from the transcribed strand of active genes, permitting rapid resumption of blocked transcription. This repair mechanism has been well characterized in the past using individual target genes. Moreover, numerous efforts investigated the fate of blocked RNA polymerase II (Pol II) during DNA repair mechanisms and suggested that stopped Pol II complexes can either backtrack, be removed and degraded or bypass the lesions to allow TCR. We investigated the effect of a non-lethal dose of UVB on global DNA-bound Pol II distribution in human cells. We found that the used UVB dose did not induce Pol II degradation however surprisingly at about 93% of the promoters of all expressed genes Pol II occupancy was seriously reduced 2–4 hours following UVB irradiation. The presence of Pol II at these cleared promoters was restored 5–6 hours after irradiation, indicating that the negative regulation is very dynamic. We also identified a small set of genes (including several p53 regulated genes), where the UVB-induced Pol II clearing did not operate. Interestingly, at promoters, where Pol II promoter clearance occurs, TFIIH, but not TBP, follows the behavior of Pol II, suggesting that at these genes upon UVB treatment TFIIH is sequestered for DNA repair by the TCR machinery. In agreement, in cells where the TCR factor, the Cockayne Syndrome B protein, was depleted UVB did not induce Pol II and TFIIH clearance at promoters. Thus, our study reveals a UVB induced negative regulatory mechanism that targets Pol II transcription initiation on the large majority of transcribed gene promoters, and a small subset of genes, where Pol II escapes this negative regulation.

Low and high doses of UV-B differentially modulate chlorpyrifos-induced alterations in nitrogen metabolism of cyanobacteria

The present study assessed the comparative responses on the specific growth rate, nitrogen metabolism and enzymes associated with nitrogen metabolism in two nitrogen fixing cyanobacteria—Nostoc muscorum and Phormidium foveolarum exposed to two UV-B doses (low; UV-BL: 0.5472kJm−2 and high; UV-BH: 5.472kJm−2) and two doses of the insecticide chlorpyrifos (O,O-diethyl O-3,5,6-trichloro-2-pyridyl phosphorothioate; low i.e. CPL, 1µgml−1 and high i.e. CPH, 2µgml−1) singly and in combination. The specific growth rate, NO3− and NO2− uptake, nitrate assimilating enzymes – nitrate reductase and nitrite reductase and ammonium assimilating enzymes – glutamine synthetase and glutamate synthase were severely affected when treated either with CPH or/and UV-BH while glutamate dehydrogenase exhibited a stimulatory response. CPL also reduced all the measured parameters (except GDH activity) after 24h, however, a stimulatory effect was observed after 72h due to an increase in nitrogen metabolism (and other antioxidant) enzymes during this period. UV-BL did not cause significant alteration in the studied parameters while in combination with CP doses, it either alleviated the inhibitory effects or further enhanced the CPL induced activities of these enzymes (except GDH). Overall results indicate the resistant nature of P. foveolarum against the inhibitory doses of UV-B and chlorpyrifos in comparison to N. muscorum.

Photosensitized mefloquine induces ROS-mediated DNA damage and apoptosis in keratinocytes under ambient UVB and sunlight exposure.

The present study illustrates the photosensitizing behavior of mefloquine (MQ) in human skin keratinocytes under ambient doses of UVB and sunlight exposure. Photochemically, MQ generated reactive oxygen species superoxide radical, hydroxyl radical, and singlet oxygen through type I and type II photodynamic reactions, respectively, which caused photooxidative damage to DNA and formed localized DNA lesions cyclobutane pyrimidine dimers. Photosensitized MQ reduced the viability of keratinocytes to 25 %. Significant level of intracellular reactive oxygen species (ROS) generation was estimated through fluorescence probe DCF-H2. Increased apoptotic cells were evident through AO/EB staining and phosphatidyl serine translocation in cell membrane. Single-stranded DNA damage was marked through single-cell gel electrophoresis. Mitochondrial membrane depolarization and lysosomal destabilization were evident. Upregulation of Bax and p21 and downregulation of Bcl-2 genes and corresponding protein levels supported apoptotic cell death of keratinocyte cells. Cyclobutane pyrimidine dimers (CPDs) were confirmed through immunofluorescence. In addition, hallmarks of apoptosis and G2/M phase cell cycle arrest were confirmed through flow cytometry analysis. Our findings suggest that MQ may damage DNA and produce DNA lesions which may induce differential biological responses in the skin on brief exposure to UVB and sunlight.

Study of morphological deformity ofTigriopus japonicuss. l. by mid-ultraviolet radiation (UVB)

The damaging effect of mid-ultraviolet (UVB, 280–315 nm) radiation on a marine copepod, Tigriopus japonicus sensu lato was investigated. Copepods were collected from tidal pools on rocky shores, Yeosu, on the south coast of Korea, and were cultured under constant conditions (temperature: 20°C, salinity: 20 psu, L : D = 12 : 12 h) in the laboratory. Each stage of nauplius (N1–N6) and copepodite (CI–CVI) was harvested from the culture and exposed to one of eight irradiation doses (1, 5, 10, 15, 20, 25, 30, and 35 kJ m−2) of UVB. No deleterious effect was induced by UVA or PAR. Extensive morphological deformities were observed, i.e., shrunken body (in groups 5 kJ m−2 at N4; 1 and 15 kJ m−2 at CI), in the urosome and caudal rami (5 and 15 kJ m−2 at N6; 10 kJ m−2 at CI), in the swimming legs (5, 10, and 15 kJ m−2 at CI; 10 kJ m−2 CIV), and in the antennule (5 kJ m−2 at CIII; 1 kJ m−2 at CIV). The types of deformity observed depended on the developmental stages of copepods that were exposed to UVB radiation. UVB radiation had a damaging effect on the morphology of T. japonicus s. l. Exposure of earlier larval stages to mid-ultraviolet affected the shape of the entire body, and the prosome shape was abnormal. Exposure of later larval stages to mid-ultraviolet resulted in abnormalities at the appendage level, i.e., in the antennules and swimming legs.

Re-interpreting plant morphological responses to UV-B radiation.

There is a need to reappraise the effects of UV-B radiation on plant morphology in light of improved mechanistic understanding of UV-B effects, particularly elucidation of the UV RESISTANCE LOCUS 8 (UVR8) photoreceptor. We review responses at cell and organismal levels, and explore their underlying regulatory mechanisms, function in UV protection and consequences for plant fitness. UV-induced morphological changes include thicker leaves, shorter petioles, shorter stems, increased axillary branching and altered root:shoot ratios. At the cellular level, UV-B morphogenesis comprises changes in cell division, elongation and/or differentiation. However, notwithstanding substantial new knowledge of molecular, cellular and organismal UV-B responses, there remains a clear gap in our understanding of the interactions between these organizational levels, and how they control plant architecture. Furthermore, despite a broad consensus that UV-B induces relatively compact architecture, we note substantial diversity in reported phenotypes. This may relate to UV-induced morphological changes being underpinned by different mechanisms at high and low UV-B doses. It remains unproven whether UV-induced morphological changes have a protective function involving shading and decreased leaf penetration of UV-B, counterbalancing trade-offs such as decreased photosynthetic light capture and plant-competitive abilities. Future research will need to disentangle seemingly contradictory interactions occurring at the threshold UV dose where regulation and stress-induced morphogenesis overlap.

In vitro studies on the mechanisms involved in chemoprevention using Calluna vulgaris on vascular endothelial cells exposed to UVB

The study aims to investigate the mechanisms involved in the in vitro effect of UVB on endothelial vascular cells (HUVECs) pretreated with a photochemopreventive agent, the Calluna vulgaris (Cv) extract.Two concentrations of Cv, below the limit of cytotoxicity IC50 (2.5 and 7.5μg GAE/ml) and two doses of UVB (50 and 100mJ/cm2) were used. Oxidative stress parameters were quantified at 1h and 24h after irradiation and apoptosis, DNA damage and the induction/activation of NF-κB were evaluated at 24h.UVB exposure led to the formation of lipid peroxides in a dose dependent manner (p<0.001), induced apoptosis, increased the γ-H2AX levels and the activation of NF-κB. Pretreatment with 2.5μg GAE/ml Cv improved the antioxidant defense, protected against DNA lesions and was able to decrease cellular death at low dose of irradiation. 7.5μg GAE/ml Cv was prooxidant, favored the formation of DNA lesions, amplified the NF-κB activation UVB-induced (p<0.01) and led to high levels of cellular death. Both doses of Cv inhibited caspase-3 activation.The modulatory effect of Cv extract on endothelial cells exposed to UVB depend on the concentration of Cv used. This study provides insides into the mechanisms triggered by UVB and antioxidants on skin endothelial cells.

Cortisol release in response to UVB exposure in Xiphophorus fish

Xiphophorus fishes are comprised of 26 known species. Interspecies hybridization between select species has been utilized to produce experimental models to study melanoma development. Xiphophorus melanoma induction protocols utilize ultraviolet light (UVB) to induce DNA damage and associated downstream tumorigenesis. However, the impact of induced stress caused by the UVB treatment of the experimental animals undergoing tumor induction protocols has not been assessed. Stress is an adaptive physiological response to excessive or unpredictable environmental stimuli. The stress response in fishes may be measured by an assay of cortisol released into the water. Here, we present results from investigations of stress response during an experimental treatment and UVB exposure in Xiphophorus maculatus Jp 163 B, Xiphophorus couchianus, and F1 interspecies hybrids produced from the mating X. maculatus Jp 163 B×X. couchianus. Overall, cortisol release rates for males and females after UVB exposure showed no statistical differences. At lower UVB doses (8 and 16kJ/m2), X. couchianus exhibited 2 fold higher levels of DNA damage then either X. maculatus or the F1 hybrid. However, based on the cortisol release rates, none of the fish types tested induced a primary stress response at the UVB lower doses (8 and 16kJ/m2). In contrast, at a very high UVB dose (32kJ/m2) both X. maculatus and the F1 hybrid showed a 5 fold increase in the cortisol release rate. To determine the effect of pigmentation on UVB induced stress, wild type and albino Xiphophorus hellerii were exposed to UVB (32kJ/m2). Albino X. hellerii exhibited 3.7 fold increase in the cortisol release while wild type X. hellerii did not exhibit a significant cortisol response to UVB. Overall, the data suggest the rather low UVB doses often employed in tumor induction protocols do not induce a primary stress response in Xiphophorus fishes.

Detrimental effects of UV‐B radiation in a xeroderma pigmentosum‐variant cell line

DNA polymerase η (pol η), of the Y-family, is well known for its in vitro DNA lesion bypass ability. The most well-characterized lesion bypassed by this polymerase is the cyclobutane pyrimidine dimer (CPD) caused by ultraviolet (UV) light. Historically, cellular and whole-animal models for this area of research have been conducted using UV-C (λ=100-280 nm) owing to its ability to generate large quantities of CPDs and also the more structurally distorting 6-4 photoproduct. Although UV-C is useful as a laboratory tool, exposure to these wavelengths is generally very low owing to being filtered by stratospheric ozone. We are interested in the more environmentally relevant wavelength range of UV-B (λ=280-315 nm) for its role in causing cytotoxicity and mutagenesis. We evaluated these endpoints in both a normal human fibroblast control line and a Xeroderma pigmentosum variant cell line in which the POLH gene contains a truncating point mutation, leading to a nonfunctional polymerase. We demonstrate that UV-B has similar but less striking effects compared to UV-C in both its cytotoxic and its mutagenic effects. Analysis of the mutation spectra after a single dose of UV-B shows that a majority of mutations can be attributed to mutagenic bypass of dipyrimidine sequences. However, we do note additional types of mutations with UV-B that are not previously reported after UV-C exposure. We speculate that these differences are attributed to a change in the spectra of photoproduct lesions rather than other lesions caused by oxidative stress.

UVB-induced gene expression in the skin of Xiphophorus maculatus Jp 163 B

Xiphophorus fish and interspecies hybrids represent long-standing models to study the genetics underlying spontaneous and induced tumorigenesis. The recent release of the Xiphophorus maculatus genome sequence will allow global genetic regulation studies of genes involved in the inherited susceptibility to UVB-induced melanoma within select backcross hybrids. As a first step toward this goal, we report results of an RNA-Seq approach to identify genes and pathways showing modulated transcription within the skin of X. maculatus Jp 163 B upon UVB exposure. X. maculatus Jp 163 B were exposed to various doses of UVB followed by RNA-Seq analysis at each dose to investigate overall gene expression in each sample. A total of 357 genes with a minimum expression change of 4-fold (p-adj<0.05) were identified as responsive to UVB. The molecular genetic response of Xiphophorus skin to UVB exposure permitted assessment of; (1) the basal expression level of each transcript for each skin sample, (2) the changes in expression levels for each gene in the transcriptome upon exposure to increasing doses of UVB, and (3) clusters of genes that exhibit similar patterns of change in expression upon UVB exposure. These data provide a foundation for understanding the molecular genetic response of fish skin to UVB exposure.

Ultraviolet radiation dose calculation for algal suspensions using UVA and UVB extinction coefficients

Although the biological importance of ultraviolet light (UVR) attenuation has been recognised in marine and freshwater environments, it is not generally considered in in vitro ecotoxicological studies using algal cell suspensions. In this study, UVA and UVB extinction were determined for cultures of algae with varying cell densities, and the data were used to calculate the corresponding extinction coefficients for both UVA and UVB wavelength ranges. Integrating the Beer–Lambert equation to account for changes in the radiation intensity reaching each depth, from the surface until the bottom of the experimental vessel, we obtained the average UVA and UVB intensity to which the cultured algal cells were exposed. We found that UVR intensity measured at the surface of Chlamydomonas reinhardtii cultures lead to a overestimation of the UVR dose received by the algae by 2–40 times. The approach used in this study allowed for a more accurate estimation of UVA and UVB doses.

Acute exposure to ultraviolet‐B radiation modulates sex steroid hormones and receptor expression in the skin and may contribute to the sex bias of melanoma in a fish model

Using the Xiphophorus fish melanoma model, we show a strong male bias for sunlight-induced malignant melanoma, consistent with that seen in the human population. To examine underlying factors, we exposed adult X. couchianus fish to a single, sublethal dose of UVB and measured circulating sex steroid hormones and expression of associated hormone receptor genes over a 24-h period. We found that a single exposure had profound effects on circulating levels of steroid hormones with significant decreases for all free sex steroids at 6 and 24 h and increases in conjugated 2-estradiol and 11-ketotestosterone at 6 and 24 h, respectively. Whereas ARα expression increased in male and female skin, neither ARβ nor either of the ERs showed significant responses to UVB in either sex. The rapid response of male androgens and their receptors in the skin after UVB irradiation implicates hormones in the male bias of skin cancer and suggests that the photoendocrine response immediately after UV exposure may be relevant to melanomagenesis.

Vulnerability of Patagonian planktonic copepods to fluctuations in temperature and UV radiation

The aim of this investigation is to address the impact of fluctuations in temperature and ultraviolet radiation (UVR) on three species of South American copepods, Boeckella antiqua, B. gracilis and B. brevicaudata. These copepods are cold stenotherm and occur in high latitude lakes of South America and in mountain lakes in the Andes. The forecast scenarios for climate change in southern South America anticipate raising temperature and UVR levels. These changes may have the potential to impact high altitude and latitude ecosystems, including lakes and their cold adapted biota, such as those in Patagonia. Laboratory experiments, consisting of 10 day and 2 day incubations, were set up to analyse copepod mortality in relation with: (i) temperature, and (ii) the combined effect of temperature (5, 8, 12, 16, 20°C) and UV-B dose (61, 194 and 324 J m−2). The results obtained showed up that temperature is a limiting factor for B. brevicaudata that did not survive above 12°C. B. antiqua and B. gracilis withstood the temperature range although their mortality was higher at 12-16°C. The survivorship of these copepod species to radiation was found to depend on the UV-B dose, resulting in higher mortality at the highest UV-B dose. Overall, at least one Boeckella species showed an acute sensitivity to increasing temperature, and the three species studied proved tolerant to the UV-B experimental exposure. The survivorship patterns observed in Boeckella species reflect clearly their adaptation to high solar radiation exposure and to temperate to cold environmental conditions.

EFFECTS OF UV-B ON PHOTOSYNTHETIC PARAMETERS, LIPID PEROXIDATION, FLAVONOIDS AND GROWTH TRAITS OF &lt;i&gt;CONOCARPUS LANCIFOLIUS&lt;/i&gt; (ENGL.)

Conocarpus lancifolius is a perennial ornamental plant adapted to the semi-arid conditions of Kuwait. It accumulates most of its biomass during the stressful summer conditions which includes UV-B irradiation. The study evaluated equivalent UV-B irradiation intensities that plants would normally experience sometime during the course of a year in Kuwait. Leaf area, plant height, photosynthetic parameters, Lipid Peroxidation (LP) and Phenyl Alanine Lyase (PAL) were measured in C. lancifolius plants exposed to different UV-B treatments. UV-B treatments decreased leaf area and plant height but induced the formation of lateral branches. Chlorophyll content, photosynthesis, chlorophyll fluorescence measured as maximum quantum efficiency of photosystem II (Fv/Fm) and Electron Transport Rate (ETR) decreased in the short term or the first 10-20 days after treatment. Lipid Peroxidation (LP) measured in terms of Malondiadehyde (MDA) content decreased in the long term after a short term increase but Phenyl Alanine Lyase (PAL) activity increased linearly by 41.2, 66.8 and 78.6% after 30 days. There was also Positive Linear Correlation between UV-B dosage and total flavonoid accumulation which was localized in epidermal cell walls, trichomes and cuticular layer. The response of C. lancifolius to ameliorate and/or tolerate UV-B stress involved multiple mechanisms that included increased concentrations of PAL activity, increased flavonoids production; and reduced lipid peroxidation. The net effect of flavonoid accumulation and PAL enzyme activity probably contributed significantly to the protection or tolerance of the species to the effects of UV-B.

Evaluation of protective effect of Thymol on UVBinduced damage in an ex-vivo human skin tissue model: morphological analysis and genotoxic evaluations

insult, among which is the most important solar ultraviolet (UV) radiation (Bernerd et al., 2001). For this reason, the use of human skin tissue obtained from plastic aesthetic surgery represents a simple but efficient experimental approach to reproduce a physiological condition to test the early effects of an exogenous stimulus as UV radiation and the possibility of preventing or reducing the early epidermal effects. Normal human skin explants were obtained from healthy young non smoking women 20-40 years old (n=5) after informed consent and cultured epidermal side up at the air-liquid interface overnight in a Transwell system before treatment (Donetti et al., 2005; Bedoni et al., 2007). They were further divided in two groups: the first was exposed to UVB doses ranging from 0.24 J/cm2 to 0.72 J/cm2 and the other one pretreated for 1 h with Thymol (natural monoterpene phenol, 6.6 μM), before the UVB irradiation. In each experiment a cultured sample was not UVB exposed and represented the internal control. Samples were harvested 24 hours after of UVB exposure. Lactate Dehydrogenase (LDH) assay and alkaline comet and micronucleus tests were used to assess cytotoxicity and genotoxicity, respectively. Bioptic fragments were processed both for transmission electron (TEM) and light (LM) microscopy. Epidermal proliferation was investigated by indirect immunofluorescence after incorporation of 5-Bromo- 2’-deoxyuridine (BrdU). UVB induced evident ultrastructural alterations in nucleus and cytoskeleton, while the pretreatment with Thymol showed a reduction of damage in UVB exposed samples both from the morphological point of view that genotoxic aspects. Cell proliferation was strongly inhibited by UVB exposure, while in Thylom pretreated samples was comparable to control. Furthermore these results strongly support the use of ex vivo human skin as a relevant method for safety evaluation of UV skin exposure.

Cellular Antioxidant Activity and Whitening Effects of Dendropanax morbifera Leaf Extracts

Su Ah Park , Jun Park , Chan Il Park , Young Jong Jie , Yun Chan Hwang , Yong Hyun Kim , So Ha Jeon , Hye Mi Lee , Ji Hoon Ha , Kyeong Jin Kim and Soo Nam Park . Microbiol. Biotechnol. Lett. 2013;41:407-15. https://doi.org/10.4014/kjmb.1311.11001

Short term UV-B radiation-mediated transcriptional responses and altered secondary metabolism of in vitro propagated plantlets of Artemisia annua L.

In vitro propagated Artemisia annua plantlets were exposed to low dose (2.8 W m−2) of UV-B (280–315 nm) radiation for different short-term (1, 2, 3 and 4 h) durations that resulted in 103 and 100 % enhanced artemisinin and flavonoid yield respectively after 3 h of UV-B irradiation. UV-B irradiation resulted in reduced chlorophyll a, b and carotenoids while the concentrations of UV-B-absorbing anthocyanins and phenolics were induced. Under UV-B radiation, superoxide radical formation increased in a time dependent manner and was maximum at 4 h. This lead to differential responses of enzymatic antioxidants (superoxide dismutase, catalase, peroxidases and glutathione reductase) to withstand the UV-B radiation-induced reactive oxygen species burst. The transcript analysis through RT-PCR revealed the significant up-regulation of HMGR, DXR, IPPi, FPS, ADS, CYP71AV1 and RED1 gene transcripts leading to higher artemisinin accumulation. Differential expression of stress regulated genes (AOX1a, NDB2, UPOX, PAL, LOX) helped the plant towards better adaptability at least up to 3 h UV-B irradiation period. Simultaneous clustering of genes and UV-B treatments using complete linkage and a euclidean distance matrix revealed that 3 h irradiation period is optimum for high artemisinin yield with better tolerance. Histological studies revealed the larger trichome size in UV-B treated plants as compared to untreated plants. The present study conclude that short term UV-B treatment to in vitro raised A. annua may be a safe approach for continuous supply of high artemisinin producing plantlets with simultaneous action of stress- regulated genes to keep the plant healthy.

Deciduous and evergreen tree responses to enhanced UV-B treatment during three years

This paper reports a study of the strategies in Norway spruce (Picea abies (L.) Karst.) and European beech (Fagus sylvatica L.) for coping with enhanced UV-B radiation. Trees, as plants in general, possess diverse systems which respond to UV-B radiation. Changes in physiology, biochemistry and morphology have been observed in trees under enhanced UV-B radiation. The efficiency of trees’ UV-B protective systems depends on plant characteristics and state of development as well as can be correlated with the UV-B dose and the environmental conditions. The two tree species were exposed outdoors to enhanced UV-B simulating 17% ozone depletion for three years during which time, selected parameters were monitored. Selected physiological parameters were monitored three times a year on beech leaves and three needle age classes of spruce. Spruce and beech exhibited great variability in the amounts of chlorophyll, methanol-soluble UV-B and UV-A absorbing compounds, and optimum quantum yield of photosystem II. The effects of UV-B radiation also varied with needle and leaf development stage and interaction with environmental conditions. Enhanced UV-B radiation triggered responses in both trees and a reduced negative effect of UV-B radiation on spruce photochemical efficiency was observed during prolonged drought. The results show high UV-B tolerance of both tree species and indicate the complexity of plant response to UV-B, involving multilevel interactions with environmental factors and thus emphasizes the necessity of long-term investigations on trees in a natural ecosystem.

Modulation of keratin 1, 10 and involucrin expression as part of the complex response of the human keratinocyte cell line HaCaT to ultraviolet radiation

Skin exposure to ultraviolet (UV) light evokes a complex stress response in keratinocytes. Keratin filament organization provides structural stability and mechanical integrity of keratinocytes. Involucrin is a transglutaminase substrate protein contributing to the formation of insoluble cornified envelopes. However, a more complex role for keratins and involucrin has been proposed, including the regulation of cell stress response. The aim was to evaluate modulations of keratin 1, 10 and involucrin expression in HaCaT in the light of the complex response of these cells to UV-B radiation, including effects on c-Jun and matrix metalloproteinase 1 (MMP-1) gene expression and production of interleukin (IL) 6 and 8. A UV-B (300±5 nm) dose of 10 mJ/cm2 was selected since this dose resulted in a partial decrease in cell viability in contrast to higher UV-B doses, which induced complete cell death 48 h after treatment. The UV-B radiation induced significant expression of keratin 1 and 10 and decreased expression of involucrin. This was accompanied by increased expression of c-Jun and MMP-1 and IL-6 and IL-8 production. The data suggest that the expression of keratin 1, 10 and involucrin is modulated in HaCaT keratinocytes as a part of the complex stress response to UV radiation.

Acrylic nail curing UV lamps: High-intensity exposure warrants further research of skin cancer risk

Acrylic nail curing UV lamps: High-intensity exposure warrants further research of skin cancer risk

EFFECT OF OXIDASE STRESSES ON GLUCOSINOLATES PROFILE IN BROCCOLI FLORETS DURING POSTHARVEST STORAGE

Stresses cause production of reactive oxygen species in plant bodies and high levels of stress can be harmful to plant tissues. When sub-acute doses of stresses are applied to postharvest crops, beneficial effects can occur through activation of protection mechanisms. In broccoli (Brassica oleracea), such mechanisms can be induction of glucosinolates and phenolic compounds, which are also beneficial to human health. The objective of this work was to examine sub-acute or hormetic doses as well as extreme doses of UV-B, ozone and hydrogen peroxide on the content of glucosinolates during postharvest storage. Broccoli florets were exposed to hormetic and high doses of UV-B (0, 1.5 and 7.5 kJ m(-2)); O(3) (5 ppm for 0, 60 and 720 min) and H(2)O(2) (0, 1.25 and 5 mM for 3 h). The profile of glucosinolates and hydroxy-cinnamic acids was determined for up to 14 days in broccoli florets stored at 4°C by LC-MS. The expression of glucosinolate pathway genes (p450 of the CYP79 family-A2, B3 and F1) and phenylpropanoids pathway genes (PAL, CS and F3H1) were also monitored. Titer of neoglucobrassin in UV-B treated broccoli florets at 1.5 kJ m(-2) was the highest at the end of the 14 days of storage. However, up regulated expression of genes CYP79B3 and CoL were observed with the high UV-B dose of 7.5 kJ m-(2) throughout and at the beginning of the storage, respectively. Both O(3) and H(2)O(2) increased the titers of 4-methoxy-glucobrassicin and 4-hydroxy-glucobrassicin; and the florets treated with high ozone dose (5 ppm for 720 min) exhibited an over expression of genes CYP79F1 and CYP79B3, at the beginning of the storage. Results showed a good correlation between gene expression of CYP79B3, and the titers of indole glucosinolates in treated broccoli florets, suggesting that the target of the applied stresses is likely to be the branch pathway of indole glucosinolates.