Interactive Effect of Melatonin and UV-C on Phenylpropanoid Metabolite Production and Antioxidant Potential in Callus Cultures of Purple Basil (Ocimum basilicum L. var.s purpurascens).

The present study evaluated the interactive effect of melatonin and UV-C on phenylpropanoid metabolites profile and antioxidant potential of Ocimum basilicum L. Callus was treated with varying concentrations of melatonin and UV-C radiations for different time durations, either alone and/or in combination. Individual treatments of both UV-C and melatonin proved to be more effective than combine treatments. Results indicated that UV-C (10 min) exposure increased rosmarinic acid (134.5 mg/g dry weight (DW)), which was 2.3-fold greater than control. Chichoric acid (51.52 mg/g DW) and anthocyanin (cyanide 0.50 mg/g DW) were almost 4.1-fold, while peonidin was found 2.7-fold higher in UV-C (50 min) exposure. In the case of melatonin, 1.0 mg/L concentrations showed maximum rosmarinic acid (79.4 mg/g DW) accumulation; i.e., 1.4-fold more, as compared to the control. However, 2 mg/L melatonin accumulate chichoric acid (39.99 mg/g DW) and anthocyanin (cyanide: 0.45 mg/g DW and peonidin: 0.22 mg/g DW); i.e., 3.2, 3.7 and 2.0-fold increase, as compared to the control, respectively. On the other hand, melatonin-combined treatment (melatonin (Mel) (4 mg/L) + UV-C (20 min)) was proved to be effective in caffeic acid elicitation, which was 1.9-fold greater than the control. Furthermore, antioxidant potential was evaluated by both in vitro (DPPH, ABTS and FRAP assays) and in cellulo methods. Maximum in vitro antioxidant activity (DPPH: 90.6% and ABTS: 1909.5 µM) was observed for UV-C (50 min)-treated cultures. The highest in vitro antioxidant activity measured with the ABTS assay as compared to the FRAP assay, suggesting the main contribution of antioxidants from basil callus extracts acting through a hydrogen atom transfer (HAT) over an electron transfer (ET)-based mechanism. Cellular antioxidant assay was evaluated by production of ROS/RNS species using yeast cell cultures and further confirmed the protective action of the corresponding callus extracts against oxidative stress. Overall, both melatonin and UV-C are here proved to be effective elicitors since a positive correlation between the induced production of phenolic compounds, and in cellulo antioxidant action of basil callus extracts were observed.

De Novo Assembly and Comparative Transcriptome Analyses of Red and Green Morphs of Sweet Basil Grown in Full Sunlight.

Sweet basil (Ocimum basilicum), one of the most popular cultivated herbs worldwide, displays a number of varieties differing in several characteristics, such as the color of the leaves. The development of a reference transcriptome for sweet basil, and the analysis of differentially expressed genes in acyanic and cyanic cultivars exposed to natural sunlight irradiance, has interest from horticultural and biological point of views. There is still great uncertainty about the significance of anthocyanins in photoprotection, and how green and red morphs may perform when exposed to photo-inhibitory light, a condition plants face on daily and seasonal basis. We sequenced the leaf transcriptome of the green-leaved Tigullio (TIG) and the purple-leaved Red Rubin (RR) exposed to full sunlight over a four-week experimental period. We assembled and annotated 111,007 transcripts. A total of 5,468 and 5,969 potential SSRs were identified in TIG and RR, respectively, out of which 66 were polymorphic in silico. Comparative analysis of the two transcriptomes showed 2,372 differentially expressed genes (DEGs) clustered in 222 enriched Gene ontology terms. Green and red basil mostly differed for transcripts abundance of genes involved in secondary metabolism. While the biosynthesis of waxes was up-regulated in red basil, the biosynthesis of flavonols and carotenoids was up-regulated in green basil. Data from our study provides a comprehensive transcriptome survey, gene sequence resources and microsatellites that can be used for further investigations in sweet basil. The analysis of DEGs and their functional classification also offers new insights on the functional role of anthocyanins in photoprotection.

Induction of stress volatiles and changes in essential oil content and composition upon microwave exposure in the aromatic plant Ocimum basilicum.

Exposure to sustained low intensity microwaves can constitute a stress for the plants, but its effects on plant secondary chemistry are poorly known. We studied the influence of GSM and WLAN-frequency microwaves on emissions of volatile organic compounds and content of essential oil in the aromatic plant Ocimum basilicum L. hypothesizing that microwave exposure leads to enhanced emissions of stress volatiles and overall greater investment in secondary compounds. Compared to the control plants, microwave irradiation led to decreased emissions of ß-pinene, α-phellandrene, bornyl acetate, ß-myrcene, α-caryophyllene and benzaldehyde, but increased emissions of eucalyptol, estragole, caryophyllene oxide, and α-bergamotene. The highest increase in emission, 21 times greater compared to control, was observed for caryophyllene oxide. The irradiation resulted in increases in the essential oil content, except for the content of phytol which decreased by 41% in the case of GSM-frequency, and 82% in the case of WLAN-frequency microwave irradiation. The strongest increase in response to WLAN irradiation, >17 times greater, was observed for hexadecane and octane contents. Comparisons of volatile compositions by multivariate analyses demonstrated a clear separation of different irradiance treatments, and according to the changes in the volatile emissions, the WLAN-frequency irradiation represented a more severe stress than the GSM-frequency irradiation. Overall, these results demonstrating important modifications in the emission rates, essential oil content and composition indicate that microwave irradiation influences the quality of herbage of this economically important spice plant.

Daytime Solar Heating Controls Downy Mildew Peronospora belbahrii in Sweet Basil.

The biotrophic oomycete Peronospora belbahrii causes a devastating downy mildew disease in sweet basil. Due to the lack of resistant cultivars current control measures rely heavily on fungicides. However, resistance to fungicides and strict regulation on their deployment greatly restrict their use. Here we report on a 'green' method to control this disease. Growth chamber studies showed that P. belbahrii could hardly withstand exposure to high temperatures; exposure of spores, infected leaves, or infected plants to 35-45 °C for 6-9 hours suppressed its survival. Therefore, daytime solar heating was employed in the field to control the downy mildew disease it causes in basil. Covering growth houses of sweet basil already infected with downy mildew with transparent infra-red-impermeable, transparent polyethylene sheets raised the daily maximal temperature during sunny hours by 11-22 °C reaching 40-58 °C (greenhouse effect). Such coverage, applied for a few hours during 1-3 consecutive days, had a detrimental effect on the survival of P. belbahrii: killing the pathogen and/or suppressing disease progress while enhancing growth of the host basil plants.

Photoprotection by foliar anthocyanins mitigates effects of boron toxicity in sweet basil (Ocimum basilicum).

Boron (B) toxicity is an important agricultural problem in arid environments. Excess edaphic B compromises photosynthetic efficiency, limits growth and reduces crop yield. However, some purple-leafed cultivars of sweet basil (Ocimum basilicum) exhibit greater tolerance to high B concentrations than do green-leafed cultivars. We hypothesised that foliar anthocyanins protect basil leaf mesophyll from photo-oxidative stress when chloroplast function is compromised by B toxicity. Purple-leafed 'Red Rubin' and green-leafed 'Tigullio' cultivars, grown with high or negligible edaphic B, were given a photoinhibitory light treatment. Possible effects of photoabatement by anthocyanins were simulated by superimposing a purple polycarbonate filter on the green leaves. An ameliorative effect of light filtering on photosynthetic quantum yield and on photo-oxidative load was observed in B-stressed plants. In addition, when green protoplasts from both cultivars were treated with B and illuminated through a screen of anthocyanic protoplasts or a polycarbonate film which approximated cyanidin-3-O-glucoside optical properties, the degree of photoinhibition, hydrogen peroxide production, and malondialdehyde content were reduced. The data provide evidence that anthocyanins exert a photoprotective role in purple-leafed basil mesophyll cells, thereby contributing to improved tolerance to high B concentrations.

Epidermal coumaroyl anthocyanins protect sweet basil against excess light stress: multiple consequences of light attenuation.

The putative photoprotective role of foliar anthocyanins continues to attract heated debate. Strikingly different experimental set-ups coupled with a poor knowledge of anthocyanin identity have likely contributed to such disparate opinions. Here, the photosynthetic responses to 30 or 100% solar irradiance were compared in two cultivars of basil, the green-leafed Tigullio (TG) and the purple-leafed Red Rubin (RR). Coumaroyl anthocyanins in RR leaf epidermis significantly mitigated the effects of high light stress. In full sunlight, RR leaves displayed several shade-plant traits; they transferred less energy than did TG to photosystem II (PSII), and non-photochemical quenching was lower. The higher xanthophyll cycle activity in TG was insufficient to prevent inactivation of PSII in full sunlight. However, TG was the more efficient in the shade; RR was far less able to accommodate a large change in irradiance. Investment of carbon to phenylpropanoid biosynthesis was more in RR than in TG in the shade, and was either greatly enhanced in TG or varied little in RR because of high sunlight. The metabolic cost of photoprotection was lower whereas light-induced increase in biomass production was higher in RR than in TG, thus making purple basil the more light tolerant. Purple basil appears indeed to display the conservative resource-use strategy usually observed in highly stress tolerant species. We conclude that the presence of epidermal coumaroyl anthocyanins confers protective benefits under high light, but it is associated with a reduced plasticity to accommodate changing light fluxes as compared with green leaves.

ZnO nanofertilizer and He Ne laser irradiation for promoting growth and yield of sweet basil plant.

This study was conducted to evaluate the effectiveness of zinc nanofertilizer strategy on sweet basil yield, through alone application or combined with pre-sowing laser irradiation. Furthermore, evaluate the growth of plant and the level of active essential oil constituents. Zinc oxide (ZnO) nanoparticles (NPs) were synthesized, and transmission electron microscope revealed particle size of approximately 10.5-15.5 nm. ZnO NPs were applied to sweet basil plants by foliar spray at varying concentrations (10, 20 and 30 mg/L); He Ne laser of power 3mW was used for red light irradiation of sweet basil seeds for 2 min. exposure time. Total chlorophyll, total carbohydrate, essential oil levels, zinc content, plant height, branches/plant and fresh weight were measured. In general, the combined foliar spray application of ZnO nanofertilizer with pre-sowing He Ne laser irradiation showed more effectiveness than ZnO nanofertilizer alone and 20mg/L concentration gave the highest results of all measured traits. Statistical analysis (t-test) showed significant differences among the effects of the various concentrations of zinc oxide NPs on these attributes. The results showed an inverse relationship between the total carbohydrate content and the percentage of essential oil in the leaves. Together these findings support the usefulness and effectiveness of zinc oxide nanofertilizer and laser irradiation treatment to enhance the growth and yield of sweet basil plants. The article presents some promising patents on ZnO nanofertilizer and He Ne laser irradiation.

Aroma characterisation and UV elicitation of purple basil from different plant tissue cultures.

Exposure to stressful environmental conditions can induce severe metabolic variations in basil (Ocimum basilicum) aroma. The aromatic profiles of Dark Opal and Red Rubim varieties (in vivo plants, in vitro shoots, callus, and suspension cultures) were investigated for the first time. The established calli represented the most interesting miniaturised aromatic plant systems, as they were able to emit many typical basil volatiles with very low amounts of phenylpropanoids (1-2%). The hydrocarbon monoterpenes and oxygenated volatiles emitted from calli of both varieties were greatly and conversely affected by UV-C and UV-B, in comparison with the non-irradiated samples. As calli of both varieties still maintained very low levels of phenylpropanoids even after UV elicitation, they might be regarded not only as efficient in vitro plant models to study volatile compounds under UV stress conditions, but also as safe aromatic biomass in comparison with in vivo basil plants.

Changes in nitrate and nitrite concentrations over 24h for sweet basil and scallions.

Nitrate and nitrite concentrations were determined for sweet basil and scallions over 24h to determine if time of sampling or harvest impacts concentrations in raw vegetables. Also, nitrate and nitrite concentrations were determined separately for various edible parts of these plants. Basil had significant changes in nitrate and nitrite concentrations over a 24h period. Nitrate was correlated to changes in light intensity with a 3h lag time. The highest nitrate concentrations in basil (2777 ppm) occurred around 3h after the light intensity peaked and had low values (165-574 ppm) during the dark period. The scallion nitrate and nitrite concentrations were always low but nitrate showed a peak a few hours before sunrise. Nitrate and nitrite concentrations in some raw vegetables may be reduced by harvesting at the best time of day for each type of plant. Nitrate concentrations were different in the edible plant parts tested.

The effects of different UV-B radiation intensities on morphological and biochemical characteristics in Ocimum basilicum L.

BACKGROUND: The effects of short-term ultraviolet B (UV-B) irradiation on sweet basil (Ocimum basilicum L. cv. Cinnamon) plants at the 3-4 leaf pair and flowering stages were examined in controlled environment growth chambers. Plants were exposed to 0 (reference), 2 and 4 kJ UV-B m(-2) day(-1) over 7 days. RESULTS: Exposure of basil plants to supplementary UV-B light resulted in increased assimilating leaf area, fresh biomass and dry biomass. Stimulation of physiological functions in young basil plants under either applied UV-B dose resulted in increased total chlorophyll content but no marked variation in carotenoid content. At the flowering stage the chlorophyll and carotenoid contents of basil were affected by supplementary UV-B radiation, decreasing with enhanced UV-B exposure. Both total antioxidant activity (2,2-diphenyl-1-picrylhydrazyl free radical assay) and total phenolic compound content were increased by UV-B light supplementation. Young and mature basil plants differed in their ascorbic acid content, which was dependent on UV-B dose and plant age. UV-B radiation resulted in decreased nitrate content in young basil plants (3-4 leaf pair stage). CONCLUSION: These results indicate that the application of short-exposure UV-B radiation beneficially influenced both growth parameters and biochemical constituents in young and mature basil plants.

Effects of gamma-irradiation on the free radical and antioxidant contents in nine aromatic herbs and spices.

Nine spice and aromatic herb samples (i.e., basil, bird pepper, black pepper, cinnamon, nutmeg, oregano, parsley, rosemary, and sage) were gamma-irradiated at a dose of 10 kGy according to commercial practices. The effects of the disinfection treatment on the content of organic radicals and some nutrients (namely, vitamin C and carotenoids) in the samples were investigated by chromatographic and spectroscopic techniques. Irradiation resulted in a general increase of quinone radical content in all of the investigated samples, as revealed by electron paramagnetic resonance spectroscopy. The fate of these radicals after storage for 3 months was also investigated. The cellulose radical was clearly observed in a few samples. Significant losses of total ascorbate were found for black pepper, cinnamon, nutmeg, oregano, and sage, whereas a significant decrease of carotenoids content was observed for cinnamon, oregano, parsley, rosemary, bird pepper, and sage.

UV-B is required for normal development of oil glands in Ocimum basilicum L. (sweet basil).

Plants of Ocimum basilicum L. grown under glass were exposed to short treatments with supplementary UV-B. The effect of UV-B on volatile essential oil content was analysed and compared with morphological effects on the peltate and capitate glandular trichomes. In the absence of UV-B, both peltate and capitate glands were incompletely developed in both mature and developing leaves, the oil sacs being wrinkled and only partially filled. UV-B was found to have two main effects on the glandular trichomes. During the first 4 d of treatment, both peltate and capitate glands filled and their morphology reflected their 'normal' mature development as reported in the literature. During the following days there was a large increase in the number of broken oil sacs among the peltate glands as the mature glands broke open, releasing volatiles. Neither the number of glands nor the qualitative or quantitative composition of the volatiles was affected by UV-B. There seems to be a requirement for UV-B for the filling of the glandular trichomes of basil.