Antonio Miceli

Characterization of two Arabidopsis thaliana glutathione S-transferases

Glutathione S-transferases (GST) are multifunctional proteins encoded by a large gene family, divided on the basis of sequence identity into phi, tau, theta, zeta and lambda classes. The phi and tau classes are present only in plants. GSTs appear to be ubiquitous in plants and are involved in herbicide detoxification and stress response, but little is known about the precise role of GSTs in normal plant physiology and during biotic and abiotic stress response. Two cDNAs representing the two plant classes tau and phi, AtGSTF9 and AtGSTU26, were expressed in vitro and the corresponding proteins were analysed. Both GSTs were able to catalyse a glutathione conjugation to 1-chloro-2,4-dinitrobenzene (CDNB), but they were inactive as transferases towards p-nitrobenzylchloride (pNBC). AtGSTF9 showed activity towards benzyl isothiocyanate (BITC) and an activity as glutathione peroxidase with cumene hydroperoxide (CumHPO). AtGSTU26 was not active as glutathione peroxidase and towards BITC. RT-PCR analysis was used to evaluate the expression of the two genes in response to treatment with herbicides and safeners, chemicals, low and high temperature. Our results reveal that AtGSTU26 is induced by the chloroacetanilide herbicides alachlor and metolachlor and the safener benoxacor, and after exposure to low temperatures. In contrast, AtGSTF9 seems not to be influenced by the treatments employed.

Betalains, Phenols and Antioxidant Capacity in Cactus Pear [Opuntia ficus-indica (L.) Mill.] Fruits from Apulia (South Italy) Genotypes

Betacyanin (betanin), total phenolics, vitamin C and antioxidant capacity (by Trolox-equivalent antioxidant capacity (TEAC) and oxygen radical absorbance capacity (ORAC) assays) were investigated in two differently colored cactus pear (Opuntia ficus-indica (L.) Mill.) genotypes, one with purple fruit and the other with orange fruit, from the Salento area, in Apulia (South Italy). In order to quantitate betanin in cactus pear fruit extracts (which is difficult by HPLC because of the presence of two isomers, betanin and isobetanin, and the lack of commercial standard with high purity), betanin was purified from Amaranthus retroflexus inflorescence, characterized by the presence of a single isomer. The purple cactus pear variety showed very high betanin content, with higher levels of phenolics, vitamin C, and antioxidant capacity (TEAC) than the orange variety. These findings confirm the potential for exploiting the autochthonous biodiversity of cactus pear fruits. In particular, the purple variety could be an interesting source of colored bioactive compounds which not only have coloring potential, but are also an excellent source of dietary antioxidant components which may have beneficial effects on consumers’ health.

Polyphenols, resveratrol, antioxidant activity and ochratoxin a contamination in red table wines, controlled denomination of origin (DOC) wines and wines obtained from organic farming

In this work, 15 red wines (five table wines, four Apulian Denominazione di origine controllata (DOC) wines and six wines obtained from organic farming) were assayed in relation to their content of polyphenolic compounds (total phenols, total flavonoids, total anthocyans, non-anthocyan flavonoids and orthodiphenols), resveratrol, antioxidant activity and ochratoxin A (OTA) contamination. The results showed that the quantity of the various classes of polyphenolic substances and the antioxidant activity was on average higher in wines obtained from organic farming and DOC wines; the contamination by OTA, present in all wines, proved to be lower in those obtained from organic farming, which averaged 0.14 μg l−1.

Biochemical characterization of a lipase from olive fruit (Olea europaea L.)

Lipase (triacylglycerol acylhydrolase; EC 3.1.1.3) is the first enzyme of the degradation path of stored triacylglycerols (TAGs). In olive fruits, lipase may determine the increase of free fatty acids (FFAs) which level is an important index of virgin olive oil quality. However, despite the importance of virgin olive oil for nutrition and human health, few studies have been realized on lipase activity in Olea europaea fruits. In order to characterize olive lipase, fruits of the cv. Ogliarola, widely diffused in Salento area (Puglia, Italy), were harvested at four stages of ripening according to their skin colour (green, spotted I, spotted II, purple). Lipase activity was detected in the fatty layer obtained after centrifugation of the olive mesocarp homogenate. The enzyme exhibited a maximum activity at pH 5.0. The addition of calcium in the lipase assay medium leads to an increment of activity, whereas in the presence of copper the activity was reduced by 75%. Furthermore, mesocarp lipase activity increases during olive development but declined at maturity (purple stage). The data represent the first contribution to the biochemical characterization of an olive fruit lipase associated to oil bodies.

Antimicrobial activity of essential oils from aromatic plants grown in the Mediterranean area.

Abstract The chemical composition and antimicrobial activity of the essential oils isolated from four Lamiaceae [Thymbra capitata (L.) Cav., Origanum vulgare L. subsp. hirtum (Link) Ietsw., Rosmarinus officinalis L. and Satureja cuneifolia Ten.] harvested at three different vegetative stages, spontaneously growing in the South of Puglia, towards Gram-(+)ve and Gram-(-)ve bacteria and pathogenic yeasts were evaluated. The four oils were analyzed by GC-FID and GC/MS, and exhibited similarities and differences with respect to data reported in the literature. Specifically, T. capitata and O. vulgare subsp. hirtum oils were found to be rich in thymol (60.9–67.5% and 29.8–56.8%, respectively) and R. officinalis oil had large amounts of 1,8-cineole (8.0–51.3%) and α-pinene (3.9–31.4%). The composition of the oil of S. cuneifolia varied most of all, its main constituents being linalool (9.6–32.7%), borneol (12.9–24.0%) and α-pinene (9.5–11.7%). Each of the oils was found to possess some antimicrobial properties using the agar diffusion method. The effectiveness and behavioral specificity varied from one species to another, and within each species, in relation to the vegetative period and the microorganism tested. The most effective oils were those of O. vulgare subsp. hirtum and T. capitata. The latter was also biologically active towards P. aeruginosa which was not sensitive to either R. officinalis or S. cuneifolia oils.

Influence of Environmental Factors on Essential Oil Variability in Thymbra capitata (L.) Cav. Growing Wild in Southern Puglia (Italy)

Abstract The composition of officinal plants in terms of active principles exhibits high intraspecific variability which is determined not only genetically but also by environmental conditions, growth area and harvesting time. This study analyzed the essential oil variability in Thymbra capitata (L.) Cav. in specimens growing wild in 23 places in Southern Puglia (Italy). The results show high chemical polymorphism in the oil content of T. capitata. Application of the spatial autocorrelation index to the principal components highlighted a relationship between thermal parameters and the composition in terms of active principles. The carvacrol chemotype was only present under the hottest and driest conditions. These results show that not only is the biosynthetic pathway of phenolic monoterpenes in T. capitata favored in high-temperature environments, as reported in other Lamiaceae, but also that carvacrol is present only in markedly “Mediterranean-like” environments.

Pigment production from in vitro cultures of Alkanna tinctoria Tausch.

SummaryAn in vitro culture of Alkanna tinctoria Tausch cells was set up in order to investigate the possibility of producing alkannin, a red naphthoquinone naturally present in the root bark of this plant. Furthermore, an in vitro culture of callusderived roots was established and the production of alkannin evaluated. In the different experimental conditions investigated, differences in the production of alkannin derivatives as well as in the type of pigments produced, were observed. The potential use of this technology is discussed.

Intraspecific variability of the essential oil of Calamintha nepeta subsp. nepeta from Southern Italy (Apulia)

The essential oil of 46 spontaneous plants of Calamintha nepeta (L.) Savi subsp. nepeta growing wild in Sud, Italy (Salento, Apulia), were investigated by GC/MS. Fifty-seven components were identified in the oil representing over the 98% of the total oil composition. Four chemotypes were identified: piperitone oxide, piperitenone oxide, piperitone-menthone and pulegone.

Antioxidant activity of Buglossoides purpureocaerulea (L.) I.M. Johnst. extracts

Buglossoides purpureocaerulea is a little-known plant used in the folk tradition for the preparation of a decoction in Sud, Italy, where it is appreciated for its beneficial effects on liver diseases. These properties may be due to the presence of antioxidant compounds. This study presents the phenolic characterisation and the antioxidant activity (AA) of B. purpureocaerulea extracts obtained by decoction, ethanol infusion and ethanol and methanol macerations. Total phenols ranged between 69 and 100 mg g−1 dry weight (DW). The main compounds were: rosmarinic acid, caffeic acid, lithospermic acid and salvianolic acid C. The extracts showed a good AA, particularly high for the decoction (142–283 µmol TE g−1 DW; TE, Trolox equivalent) and ethanol maceration extract (214–364 µmol TE g−1 DW) when determined by DPPH and ferric reducing antioxidant power test, respectively.

Essential Oil Variability of Satureja cuneifolia Ten. Growing Wild in Southern Puglia (Italy)

Abstract Satureja cuneifolia Ten. spontaneously growing in 12 sites in Southern Puglia (Italy), was characterized on the basis of its essential oil composition. Qualitative and quantitative analyses of steam-distilled oil were performed by GC and GC/MS. Cluster analysis made it possible to identify four different chemotypes: linalool, borneol, α-pinene, and borneol/α-pinene. Principal Component Analysis (PCA) highlighted the relations existing between the compounds that constitute the essential oils. Moreover, spatial autocorrelation, evaluated in the space described by the principal components, was used to analyse the relationships between chemical polymorphism and certain large-scale environmental conditions proper to sampling sites. Specifically, the arrangement of values in relation to Yearly Rainfall (YR) was found not to be random: the findings showed that increased water availability in the soil causes an arise in the presence of the borneol chemotype. In contrast, reduced water availability is directly correlated with the presence of the linalool and α-pinene chemotypes.