Changes in phytochemical content and pharmacological activities of three <Emphasis Type="Italic">Chlorella</Emphasis> strains grown in different nitrogen conditions

The phytochemical content and biological activity of three Chlorella strains cultured in low (35 mg L^?1) or high (700 mg L^?1) nitrogen (N) and harvested on days 5 and 10 were evaluated. High N resulted in a higher biomass in Chlorella MACC 438 and MACC 452 while MACC 555 produced a higher biomass in low N. MACC 555 (low N/day 5) had the highest phenolic content, and MACC 438 in low N/day 5 and high N/day 5 accumulated the highest flavonoids and condensed tannins, respectively. Iridoids were most abundant in MACC 452 on low N/day 10. Harvest time had the greatest effect on the phytochemical content with phenolics, flavonoids, and condensed tannins decreasing over time and iridoids increasing in low N and decreasing in high N conditions. Extracts were more active in β-carotene-linoleic acid model compared to 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging assay. Most extracts had good antimicrobial activity. Extracts became less potent over time in the antioxidant, acetylcholinesterase inhibitory (AChE), and antimicrobial assays when growing in low N and more potent in the antioxidant and AChE assays when grown in high N. Thus, phytochemical content and biological activities of the three Chlorella strains were affected by N levels, harvest time, and strain.     

<Emphasis Type="Italic">In vivo</Emphasis> and <Emphasis Type="Italic">in vitro</Emphasis> antioxidant effects of three Veronica species

The aim of the present study was to examine the antioxidant activity of three Veronica species (Plantaginaceae). The antioxidant potential of various extracts obtained from aerial flowering parts was evaluated by DPPH-free (1,1-diphenyl-2-picrylhydrazyl-free) radical scavenging activity and ferric-reducing antioxidant power assays. Considerable antioxidant activity was observed in the plant samples (FRAP values ranged from 0.97 to 4.85 mmol Fe²⁺/g, and DPPH IC₅₀ values from 12.58 to 66.34 μg/ml); however, these levels were lower than the activity of the control compound butylated hydroxytoluene (BHT) (FRAP: 10.58 mmol Fe²⁺/g; DPPH IC₅₀: 9.57 μg/ml). Also, the in vivo antioxidant effects were evaluated in several hepatic antioxidant systems in rats (activities of glutathione peroxidase, glutathione reductase, peroxidase, catalase, xanthine oxidase, glutathione content and level of thiobarbituric acid reactive substances) after treatment with different Veronica extracts, or in combination with carbon tetrachloride (CCl₄). Pretreatment with 100 mg/kg b.w. of Veronica extracts inhibited CCl₄-induced liver injury by decreasing TBA-RS level, increasing GSH content, and bringing the activities of CAT and Px to control levels. The present study suggests that the extracts analyzed could protect the liver cells from CCl₄-induced liver damage by their antioxidative effect on hepatocytes.     

<Emphasis Type="Italic">In vitro</Emphasis> culture of <Emphasis Type="Italic">Hypericum rumeliacum</Emphasis> Boiss. and production of phenolics and flavonoids

The Balkan endemic species, Hypericum rumeliacum, Guttiferae was introduced in vitro for the first time with the aim to study the type of morphogenetic response to plant growth regulators and ability to produce phenolics and flavonoid compounds. The morphoregulatory effect of 2,4-dichlorophenoxyacetic acid (2,4-D), 1-naphtaleneacetic acid (NAA), 6-benzyladenine (BA) and combination of BA with NAA in Murashige–Skoog's basal medium on leaf lamina, internode stem segment, stem node and root cuttings was studied. Histological analysis of the structures regenerated from the primary explants proved the presence of both, embryoids and meristemoids. The node explants cultivated on BA-supplemented medium were the most favourable for regeneration through meristemoids. Therefore a double-stage culture approach, allowing an effective multiplication of large quantities of plant shoots in vitro along with maintenance of the biosynthetic capacity of the culture was developed. It comprised one subculture of three-nodal stem explants derived from the stock shoot cultures on MS medium supplemented with 0.2 mg/l BA followed by subculture of the induced multiple shoots on cytokinin-free MS medium. Determination of the total phenolics and flavonoids showed that the decrease of the levels of these secondary metabolites is transitional, as the exclusionof BA from the medium resulted in an increase of their total content.     

Anti-oxidant,hemolysis inhibition,and collagen-stimulating activities of a new hexapeptide derived from <Emphasis Type="Italic">Arthrospira</Emphasis> (<Emphasis Type="Italic">Spirulina</Emphasis>) <Emphasis Type="Italic">platensis</Emphasis>

In this study, the whole protein of Arthrospira (Spirulina) platensis was extracted and hydrolyzed with trypsin. Screening process for new peptides was driven by 2,2′-azinobis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS) radical scavenging activity assay, and a hexapeptide with sequence of GMCCSR was identified. Its anti-oxidant activity was measured based on ABTS, 1,1’-diphenyl-2-picrylhydrazyl (DPPH), and ferric reducing ability of plasma (FRAP) assays. Hemolysis inhibition on human erythrocytes and collagen-stimulating activities on human skin fibroblasts (HSFs) were investigated. Results showed that its radical scavenging effect could compete with ascorbic acid. Intracellular assays revealed the reduction of reactive oxygen species (ROS) and malondialdehyde (MDA) contents and the increase in activities of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px) in erythrocytes pretreated with hexapeptide, demonstrating that the peptide could protect erythrocytes from lipid peroxidation and protein oxidation. Moreover, results from HSFs showed that it could promote proliferation and collagen production of HSFs pro-damaged by UVB. These results suggest the potential of the hexapeptide in food and pharmaceutical industries.     

Phytochemicals accumulation and antioxidant activity in callus and suspension cultures of <Emphasis Type="Italic">Cynara scolymus</Emphasis> L.

The antioxidative phytochemicals in globe artichoke (Cynara scolymus L.) have received increasing attention for their health-promoting properties related to the high levels of caffeoylquinic acids and flavones in capitula and leaves. Since phytochemicals in plants vary in relation to both biotic and abiotic factors, we explored the possibility to use in vitro-derived materials as a source of antioxidant compounds. Two suspension cultures, an anthocyanin-producing and not-producing cultures, and the sourced callus were evaluated in terms of their total polyphenol (TP) content and qualitative profile, total anthocyanin (TA) content and antioxidant activity (AA). TP and TA content were quantified by spectrophotometric assays, while the polyphenol profile was estimated by HPLC analysis. AA was evaluated by 2,2-diphenyl-1-picrylhydrazyl (DPPH) and ferric reducing antioxidant power (FRAP) assays. Growth kinetics and polyphenol accumulation were investigated for 25 days in red suspension cultures. The latter accumulated a higher TP and TA content (25.7 and 2.61 g kg^?1 of DM, respectively) than calluses and green suspension cultures. During cell growth, the TA content in red suspension cultures ranged from 1.43 to 2.41 g kg^?1 of DM. Optimum production of polyphenols was achieved on day 25 of culture; a positive correlation existed between TP and both DPPH (r?=?0.84) and FRAP (r?=?0.85). The 1,5–O-dicaffeoylquinic acid and cyanidin malonylglucoside (21.18 and 1.24 g kg^?1 of DM, respectively) were the primary compounds. The results of this investigation indicate that cell suspension of globe artichoke could represent a potential source of bioactive compounds with high antioxidant properties for industrial applications.     

Antioxidant properties of <Emphasis Type="Italic">Galium verum</Emphasis> L. (Rubiaceae) extracts

The antioxidant properties of methanol extracts of Lady’s Bedstraw (Galium verum L., Rubiaceae) herb from two different localities in Serbia were evaluated. Antioxidant activity was assessed in four different model systems. Free radical scavenging capacity (RSC) was examined by measuring the scavenging activity of extracts on 2,2-diphenyl-1-pycrylhydrazil (DPPH) and hydroxyl radical (OH), as well as on hydrogen peroxide. In addition, the protective effects of lipid peroxidation (LP) in corn oil were evaluated by the TBA-assay using the Fe²⁺/ascorbate system of induction. The amount of dried extract, the content of total phenolics, flavonoids and chlorophylls was also determined. Extracts from both locations expressed very strong scavenger activity, reducing the DPPH^⊙ (IC₅₀=3.10 μg/mland 8.04 μg/ml) and OH radical formation (IC₅₀=0.05 μg/ml and 0.54 μg/ml) and neutralising H₂O₂ (IC₅₀=4.98 μg/ml and 3.80 μg/ml), in a dose dependant manner. Also, examined extracts showed notable inhibition of LP (IC₅₀=11.69 μg/ml and 19.47 μg/ml). The observed differences in antioxidant activity could be partially explained by the levels of phenolics (2.44–4.65 mg and 4.57–5.16 mg gallic acid equivalents/g dry extract), flavonoids (6.38–10.70 μg and 15.56–17.96 μg quercetin equivalents/g dry extract) and chlorophylls in the investigated Lady’s Bedstraw extracts.     

The first report of <Emphasis Type="Italic">Spondias</Emphasis> native to Madagascar: <Emphasis Type="Italic">Spondias tefyi</Emphasis>, sp. nov. (Anacardiaceae)

Spondias represents a genus new to Madagascar’s native flora. Like Campnosperma, it is now known from both American and Asian tropics and Madagascar but not from continental Africa. The new species Spondias tefyi is easily distinguished from all of its Asian congeners by having the stamens shorter than the pistil and fruits brown and lenticellate at maturity (vs. greenish, yellow, orange or red, and relatively smooth). The new species is one of several Anacardiaceae whose fruits are eaten by lemurs in the Analavelona Forest, highlighting the importance of conserving this threatened subhumid forest remnant in southern Madagascar.     

Effects of nitrogen on the lipid and carotenoid accumulation of oleaginous yeast <Emphasis Type="Italic">Sporidiobolus pararoseus</Emphasis>

Nitrogen limited but carbon excess condition was used to obtain high cellular lipid content and production. The maximum lipid production was 51 g/L, the lipid content in the dry cell was 60 %, and the lipid productivity was 0.53 g/L/h. In the fermentation, the content of lipid was raised from 20 % of dry cell weight to 60 %, and the proportion of oleic acid was raised from 66.8 to 72.5 %. Meanwhile, the metabolism of carotenoids switched to torulene, and its proportion was raised from 30 to 58 %. This was according to torulene had the better antioxidant ability than β-carotene to protect the strain from oxidative damage proved by their ABTS* radical scavenging activity and lipid peroxidation inhibition ability. Sporidiobolus pararoseus lipid was a good source of lipid not only because of its high oleic acid composition, but also the antioxidant ability of carotenoids in the lipid.     

Phytochemicals and biological activities of <Emphasis Type="Italic">Artemisia sieversiana</Emphasis>

This review discusses a whole plant’s chemical nature and biological effects of Artemisia sieversiana Ehrhart ex willd (ASS). Several types of chemical compositions have been isolated from A. sieversiana, including 26 terpenoids (21 guaiane-type sesquiterpenes, 3 germacrane-type sesquiterpenes, 1 muurolane-type sesquiterpene, and 1 diterpenoid), 16 sesamin-type lignans, 9 flavonoids, 3 steroids, and 3 alkaloids. Some of them have shown promising bioactivities, such as anti-tumour, anti-inflammatory, antioxidant functions and so on. Herein, we have summarized the phytochemical and pharmacological progress of ASS.     

Lycopene accumulation and cyclic carotenoid deficiency in heterotrophic <Emphasis Type="Italic">Chlorella</Emphasis> treated with nicotine

We studied the effects of nicotine on Chlorella regularis Y-21 grown under heterotrophic conditions. Nicotine repressed growth, doubled cell size, and changed the culture coloration from dark-green to orange. These effects are likely due to the change of the chloroplast into a red irregular vesicle. This morphological change was associated with alteration of the carotenoid composition. Lycopene accounted for more than 80% of total carotenoids in nicotine-treated cells. The red irregular vesicles had a high electron density; we supposed them to be immature chloroplasts accumulating lycopene.     

Callus and cell suspension culture of <Emphasis Type="Italic">Viola odorata</Emphasis> as in vitro production platforms of known and novel cyclotides

Callus from Opuntia streptacantha (cv. Tuna loca), Opuntia megacantha (cv. Rubí reina), and Opuntia ficus-indica (cv. Rojo vigor) were exposed to jasmonic acid (JA) and abiotic stress (drought and UV light) to improve the metabolite production. The callus growth curves, phenolic acids and flavonoids content, antioxidant activity and phenylalanine ammonia lyase (PAL) activity were analyzed under normal and stress conditions. In O. streptacantha callus, the phenolics concentration increased 1.6 to 3 times times in presence of 5% PEG or after irradiation with UV light for 240 min, respectively, while flavonoids triplicate with UV light. A significant increase in antioxidant activity was observed in calli from the three Opuntia species in media with 50 µM JA. The relationships between metabolites/PAL activity, and metabolites/antioxidant activity were analyzed using a surface response methodology. Results showed that PAL activity, induced with PEG and UV, correlated with flavonoids content in O. megacantha and O. ficus-indica calli; PAL activity was related to both flavonoids and phenolics compounds in O. ficus-indica and O. megacantha calli exposed to JA, but only to flavonoids in O. streptacantha callus. In general, the JA stimulated simultaneously the metabolic pathways for phenolics and flavonoids synthesis, while abiotic stress induced mainly flavonoids route. As the stressed Opuntia calli exhibited as high antioxidant activity as cladodes, they are a promising system for research on antioxidant biosynthesis and/or to identify new compounds with antioxidant properties.     

Carotenoids from <Emphasis Type="Italic">Rhodotorula</Emphasis> and <Emphasis Type="Italic">Phaffia</Emphasis>: yeasts of biotechnological importance

Carotenoids represent a group of valuable molecules for the pharmaceutical, chemical, food and feed industries, not only because they can act as vitamin A precursors, but also for their coloring, antioxidant and possible tumor-inhibiting activity. Animals cannot synthesize carotenoids, and these pigments must therefore be added to the feeds of farmed species. The synthesis of different natural commercially important carotenoids (β-carotene, torulene, torularhodin and astaxanthin) by several yeast species belonging to the genera Rhodotorula and Phaffia has led to consider these microorganisms as a potential pigment sources. In this review, we discuss the biosynthesis, factors affecting carotenogenesis in Rhodotorula and Phaffia strains, strategies for improving the production properties of the strains and directions for potential utility of carotenoid-synthesizing yeast as a alternative source of natural carotenoid pigments.     

Mapping and validation of a major QTL for yellow pigment content on 7AL in durum wheat (<Emphasis Type="Italic">Triticum turgidum</Emphasis> L. ssp. <Emphasis Type="Italic">durum</Emphasis>)

Yellow pigment content in durum wheat (Triticum turgidum L. ssp. durum) is an important criterion for both pasta bright yellow color and human health because of antioxidant properties of carotenoids involved in this pigmentation. In the present study, QTLs for yellow pigment content in durum wheat were mapped in a population of 140 RILs developed from a intraspecific cross between a released variety (PDW 233) and a landrace (Bhalegaon 4). This trait was evaluated in one location for 3 years and in two more locations for one additional year (five different year × location combinations further called “environments”). Yellow pigment content was highly heritable across the five different environments. Analysis of variance showed the significant effect of genotype, environment and genotype × environment interaction on the trait. Five different QTLs linked to yellow pigment content were identified on chromosome 1A, 3B, 5B, 7A and 7B across five different environments. The strongest one located on the distal part of the long arm of chromosome 7A, QYp.macs-7A, explained 55.22% of the variation in the trait, while, remaining four QTLs explained 5–8.75% of phenotypic variation in yellow pigment content. Marker analysis revealed significant association of one ISSR and one AFLP fragment with the trait. These two markers were linked to the major QTL QYp.macs-7A and were converted into SCAR markers. These SCAR markers were further validated on another population as well as 38 diverse genotypes so as to prove their potential in marker assisted selection. These markers will be very useful for the marker assisted breeding of durum wheat for higher yellow pigment content.     

In vitro evaluation of phenolic and osmolite compounds,ionic content,and antioxidant activity in safflower (<Emphasis Type="Italic">Carthamus tinctorius</Emphasis> L.) under salinity stress

Carthamus tinctorius L., rich in antioxidant compounds, is a herbal medicine. Biochemical mechanisms of adaptation to salinity stress in safflower are still poorly understood at the cellular level. For this purpose, callus cultures of four different genotypes of safflower were used in this study to evaluate changes in their biochemical (ionic content, proline, and glycine betaine), total phenolics content (TPC), total flavonoids content (TFD), antioxidant responses (2,2-diphenyl-1-picrylhydrazyl: DPPH assay and carotenoid content), and lipid peroxidation (malon dialdehyde content: MDA) under salinity stress. The calluses derived from hypocotyls were exposed to in vitro salt stress at different concentrations of sodium chloride (0, 100, 200, and 300 mM). A reducing trend was observed in K⁺ and carotenoid reserves of the calluses with increasing NaCl concentration while an increasing trend was observed in Na⁺ content, proline, MDA, TPC, TFD, and DPPH activity under the same conditions. Callus glycine betaine content was found to decrease in the medium containing 100 mM NaCl but increased beyond this concentration up to 300 mM NaCl. Positive and significant correlations were recognized between DPPH and total phenolics as well as DPPH and total flavonoid contents, demonstrating that phenolics are the main contributors to the potential antioxidant activity of safflower at the cellular level. Overall, the salt-tolerant genotypes of Mex.2-137 and Mex.2-138 were found capable of being processed for the production of secondary metabolites via NaCl elicitation.     

Endophytic fungi from <Emphasis Type="Italic">Nerium oleander</Emphasis> L (Apocynaceae): main constituents and antioxidant activity

Diverse endophytic fungi exist within plant aerial tissues, with a global estimate of up to a million undescribed species. These endophytes constitute a rich bio-resource for exploration to discover new natural products. Here we investigate fungal endophytes associated with a medicinal plant, Nerium oleander L. (Apocynaceae). A total of 42 endophytic fungal strains were isolated from the host plant. Total antioxidant capacity, xanthine oxidase inhibitory activity, antimicrobial activity, and total phenolic content (TPC) were evaluated for 16 representative fungal cultures grown in improved Czapek’s broth and for the host plant. The total antioxidant capacities and phenolic contents of the fungal cultures ranged from 9.59 to 150.79 μmol trolox/100 mL culture, and from 0.52 to 13.95 mg gallic acid/100 mL culture, respectively. The fungal culture of an endophytic strain Chaetomium sp. showed the strongest antioxidant capacity, contained the highest level of phenolics, and to some extent inhibited xanthine oxidase activity with an IC₅₀ value of 109.8 μg/mL. A significant positive correlation was found between antioxidant capacity and TPC in the tested samples. Most of the endophytic fungal cultures tested have a wide range of antimicrobial activities, which were not very strong, but much better than those of the host plant. The major bioactive constituents of the fungal cultures were investigated using LC-ESI-MS and GC-MS, and preliminary identification detected phenolics (e.g. phenolic acids and their derivatives, flavonoids) and volatile and aliphatic compounds. This study shows that the endophytic fungi isolated from N. oleander can be a potential antioxidant resource.     

Tentative Identification of Phytochemicals and Antioxidant Activities during Fruit-Ripening on <i>Chamaedorea radicalis</i> Mart.

This work aims to determine the phytochemical characterization of the pericarp of Chamaedorea radicalis Mart. fruit as a non-timber product with potential to obtain phytochemicals with potential applications in the industry. Fruit from C. radicalis were grouped in four ripening stages named as S1, S2, S3 and S4, according to maturity; S1 the most unripe stage and S4 the completely ripe stage. Determinations of total phenolic compounds, free radical scavenging activities and total flavonoid contents using spectrophotometric methods were done. Also, the tentative identification of phytochemicals during fruit ripening was done using UPLC-MS-MS. Total phenolic compound (TPC) content ranged from 7.24 to 12.53 mg gallic acid equivalents per gram of fresh weight (mg GAE/ g FW). Total flavonoids (TF) contents ranged from 0.163 to 0.23 mg of quercetin equivalents per g FW (mg QE/g FW). Free radical scavenging activity against DPPH and ABTS radicals varied from 40.80 to 53.68 and from 22.29 to 37.76 mmol Trolox equivalents g FW (mmol TE/g FW), respectively. Antioxidant assay in vitro by FRAP (ferric reducing antioxidant power) method showed that S3 was the highest level with antioxidant power while S4 was the lowest with Red ripeness stage showed the lowest contents for all determinations. Mass spectrometry allowed detection of 26 compounds, including phenolics, alkaloids and saponins. Afzelin, Kaempferol 3-neohesperidoside and the four saponins identified were present in all ripeness stages. Preliminary phytochemical identi- fication and the spectrophotometric determinations showed that the pericarp of C. radicalis presented antioxidants and compounds related to alkaloids, phenolics and saponins. The presence and abundance of each phytochemical regarding each ripeness stage should be considered.