Branislav Šiler

Centauries as underestimated food additives: antioxidant and antimicrobial potential.

Methanol extracts of aerial parts and roots of five centaury species (Centaurium erythraea, C. tenuiflorum, C. littorale ssp. uliginosum, C. pulchellum, and Schenkia spicata) were analysed for their main secondary metabolites: secoiridoid glycosides, a group of monoterpenoid compounds, and phenolics (xanthones and flavonoids), and further investigated for antioxidant capacity and antimicrobial activity. The results of ABTS, DPPH, and FRAP assays showed that above ground parts generally displayed up to 13 times higher antioxidant activity compared to roots, which should be related to higher phenolics content, especially flavonoids, in green plant organs. Secoiridoid glycosides showed no antioxidant activity. All the tested extracts demonstrated appreciative antibacterial (0.05-0.5 mg ml(-1)) and strong antifungal activity (0.1-0.6 mg ml(-1)). Our results imply that above ground parts of all centaury species studied, could be recommended for human usage as a rich source of natural antioxidants and also in food industry as strong antimicrobial agents for food preservation.

Simultaneous UHPLC/DAD/(+/−)HESI–MS/MS Analysis of Phenolic Acids and Nepetalactones in Methanol Extracts of Nepeta Species: A Possible Application in Chemotaxonomic Studies

INTRODUCTION Nepeta species contain a variety of secondary metabolites, including iridoid monoterpenes - nepetalactones and phenolic acids - that are considered the main bioactive constituents. This work represents the first attempt to comparatively explore variations in these two major groups of secondary metabolites within the genus. OBJECTIVE To develop an efficient analytical methodology for simultaneous analysis of nepetalactones and phenolic acids in methanol extracts of selected Nepeta species, and to evaluate its potential application in chemotaxonomic studies. MATERIAL AND METHODS A UHPLC combined with linear-trap quadrupole (LTQ) orbitrap MS method was used to characterise chemical diversity and complexity of phenolics among 12 selected Nepeta species. A targeted metabolomic approach using UHPLC coupled to a diode array detector (DAD) and combined with (+/-) heated electrospray ionisation (HESI) MS/MS was developed and validated for quantitative analysis of six hydroxycinnamic acid derivatives and four nepetalactones. RESULTS Phenolic profiling provided a valuable database of bioactive compounds in the plant group studied, including phenolic acids (hydroxybenzoic and hydroxycinnamic acids) and flavonoids (flavones, flavonols and flavanones). Principal component analysis and cluster analysis suggested the applicability of 10 targeted compounds as chemomarkers for chemotaxonomic studies. Pearsons correlation analysis revealed significant positive correlations between metabolites involved in different biosynthetic pathways (phenylpropanoid or monoterpenoid). CONCLUSION The described targeted metabolomic approach proved to be highly beneficial in designing a phytochemical overview of the genus Nepeta, and might have applications in further clarification of phylogenetic relations. Furthermore, it has the potential to be implemented in a routine quality control of plant material and herbal preparations.

Nepetalactone content in shoot cultures of three endemic Nepeta species and the evaluation of their antimicrobial activity.

Rapid micropropagation of Nepeta rtanjensis, N. sibirica and N. nervosa was performed. Qualitative and quantitative nepetalactone content in methanol extracts of in vitro grown plants was analysed by reverse-phase HPLC coupled with UV and MS detection. Only trans,cis-nepetalactone was detected in shoots of N. rtanjensis, while cis,trans-nepetalactone stereoisomer was present in N. sibirica. No nepetalactone was observed in shoots of N. nervosa. The antimicrobial activity of methanol extracts, against eight bacterial and eight fungal species, was evaluated. All the tested extracts showed significant antibacterial and strong antifungal activity. However, N. rtanjensis extract exhibited the best antimicrobial potential.

Sugars and acid invertase mediate the physiological response of Schenkia spicata root cultures to salt stress

A heterotrophic model system was established in our studies in order to differentiate the effect of high salt concentrations in external medium on growth and sugar metabolism in roots from the effect of reduced sugar availability resulting from decreased photosynthesis under salinity. Soluble sugar content and the activity of acid invertase in root cultures of salt-tolerant (ST) and salt-sensitive (SS) Schenkia spicata (L.) Mansion genotypes were investigated during exposure to different NaCl concentrations (0-200 mM). Their response to severe salinity was characterized by a metabolic adjustment that led to the accumulation of sucrose (Suc) in root tissues. There was clear evidence that cell wall invertase (CW-Inv) is the major contributor to the Suc/hexose ratio in roots during exposure to elevated salinity. The results of CW-Inv activity and immunodetection assays in our study suggest that the regulation of CW-Inv expression is most likely achieved in a salt stress dependent manner. Also, NaCl modulated soluble acid invertase (SA-Inv) expression differentially in SS and ST genotypes of S. spicata. Regardless of the salt treatment, genotype, or the amount of enzyme, SA-Inv activity was generally low, indicating regulation at the posttranslational level. The results suggest no direct role of SA-Inv in the regulation of the root tissue carbohydrate pool and therefore in the control of the availability of glucose and fructose for the primary metabolism and/or osmotic adjustment in the present heterotrophic model system.

Centaurium erythraea methanol extract protects red blood cells from oxidative damage in streptozotocin-induced diabetic rats

ETHNOPHARMACOLOGICAL RELEVANCE Centaurium erythraea Rafn (CE) is a traditional medicinal herb in Serbia with antidiabetic, digestive, antipyretic and antiflatulent effects AIM OF THE STUDY: To investigate the potential protective effects of the methanol extract of the aerial parts of CE against glyco-oxidative stress in red blood cells (RBCs) in rats with experimentally induced diabetes. MATERIAL AND METHODS Diabetes was induced in Wistar rats by intraperitoneal (i.p.) injection of multiple low-dose streptozotocin (STZ) (40mg/kg, for five consecutive days), with the 1st day after the last STZ injection taken as the day of diabetes onset. The methanol extract of CE (100mg/kg) was administered orally and daily, two weeks before the first STZ injection, during the 5-day treatment with STZ, and for four weeks after the STZ injections (pre-treated group) or for four weeks after diabetes onset (post-treated group). The effect of CE extract administration on the redox status of RBCs was evaluated by assessing lipid peroxidation, the ratio of reduced/oxidized glutathione (GSH/GSSG), the level of S-glutathionylated proteins (GSSP) and the enzymatic activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and glutathione reductase (GR) in RBCs four weeks after diabetes onset. The major biochemical parameters of diabetes, protein glycation/glycosylation of erythrocytes and parameters which correlate with their aggregation and deformability were also evaluated. RESULTS Daily application of CE extract to STZ-induced diabetic rats provided important antidiabetic effects, observed in both pre-treated and post-treated groups of diabetic rats as elevated serum insulin concentration, reduction of blood glucose and glycated hemoglobin concentrations and an improved lipid profile. Antioxidant effects of CE extract were detected in RBCs of diabetic rats and observed as decreased lipid peroxidation and ameliorated oxidative damage as a result of increased SOD, CAT and GR activities, an improved GSH/GSSG ratio and reduced GSSP levels. Moreover, the CE extract protected RBC proteins from hyperglycemia-induced damage by reducing non-enzymatic glycation and enzymatic glycosylation processes. CE extract was more effective when applied before diabetes induction (pre-treated group). CONCLUSIONS The results of this study show that the Centaurium erythraea methanol extract protects RBCs in diabetic animals from oxidative damage. They provide additional support for the application of this traditionally used plant in diabetes management.