Silene vulgaris subsp. macrocarpa leaves and roots from Morocco: assessment of the efficiency of different extraction techniques and solvents on their antioxidant capacity,brine shrimp toxicity and phenolic characterization

Abstract

This study was undertaken to investigate the effect of various solvents and techniques on the extractability of antioxidant compounds, particularly phenolics, from leaves and roots of Silene vulgaris subsp. macrocarpa grown wild in Morocco. Maceration and hot extraction with methanol or water and Soxhlet ethanol extraction were utilized. Aimed at establishing the potential safety of the extracts, Artemia salina lethality bioassay was performed. All the extracts were found to be non-toxic, except for the leaf Soxhlet ethanol. The antioxidant potential of the extracts was evaluated in vitro by DPPH, reducing power, and ferrous ions chelating activity assays. The leaf extracts displayed noticeable radical scavenging and chelating activities, and maceration with methanol (Mac-MeOH) resulted the most suitable extraction method for an effective recovery of antioxidants; further, the root Mac-MeOH extract demonstrated good chelating properties (IC₅₀ = 335.49?±?0.70?µg/mL). Thus, leaf and root Mac-MeOH extracts were subjected to phytochemical investigations. The total phenolic, flavonoid and condensed tannin content was determined spectrophotometrically. Thirteen polyphenolic compounds were positively identified, by HPLC-PDA-ESI-MS, in the leaf extract for the first time, with p-coumaric acid derivatives being the most abundant ones (81%), whereas only catechin and procyanidin B1 were found in the root extract.     

Characterization and analysis of the subclasses and molecular species of choline phosphoglycerides from porcine heart by successive chemical hydrolyses and reverse phase high-performance liquid chromatography

Choline phosphoglycerides (CPG) represent the major fraction of heart phospholipids. Since depletion of membrane phospholipids and accumulation of lyso-compounds, particularly lysophosphatidylcholines, have been implicated in arrhythmogenesis, it was of great interest to study the composition of this major phospholipid fraction of the heart at a molecular level in an established animal model. The data presented here describe the first report on the detailed chemical examination of CPG and resolution, characterization and quantitative analysis of the molecular species of this phospholipid fraction from porcine heart by high performance liquid chromatography (HPLC). This fraction constitutes 37.5 ± 0.7% (n = 21) of the total phospholipids and upon successive mild acid and alkaline hydrolyses revealed the presence of essentially three subclasses: diacyl-, alkenylacyl-, and alkylacyl glycerophosphorylcholines, in a relative abundance of 57.7 ± 2.2% (n = 8), 37.3 ± 1.3% (n = 8) and 4.6 ± 0.2% (n = 8), respectively. The fourth subclass, dialkyl CPG was found only in minute amounts (0.43 ± 0.05%, n = 8) and the presence of dialkenyl and alkenylalkyl analogues could not be detected. Alternatively, by converting the CPG fraction to benzoate derivatives after phospholipase C digestion, it was possible to isolate and quantitate subclass composition by TLC/spectroscopy or both subclass compositions and molecular species analysis by HPLC directly by a UV detector online with the column. By these techniques, subclass composition was found to be very similar to that obtained by the chemical hydrolysis technique. By HPLC, up to 25 species can be identified and quantitated in each subclass, their identity being confirmed by gas-liquid chromatography, after their isolation from the column. The analyses showed that up to 74% of the diacyl moiety consisted of 16:0–18:2 (34%), 16:0–18:1 (27%), and 18:0–18:2 (13%) species, while the major species of the alkenylacyl moiety were 16:0–18:2 (44%) 16:0–18:1 (13%), 16:0–20:4 (12%) and 18:1–18:2 (9%) making up more than 75% of the total mass of this subclass. The major molecular species of the alkylacyl moiety was 16:0–18:2, constituting up to 47% of this fraction, while others constituted about 10% (16:0–18:1), 9% (18:1–18:2), 8% (16:0–20:4) and 6% (18:0–18:2), making up 80% of the total mass.The ether chain composition of alkylacyl CPG whether determined after isolation of this fraction by the chemical hydrolysis technique or by HPLC was indistinguishable. Similarly, the aliphatic moieties of diradylglycerols, and their subclasses, whether analysed directly or reconstituted from the molecular species data, were very similar in composition, confirming the accuracy of the data and the reproducibility of the technique devised. This also suggests that this method is suitable to distinguish minor changes in the molecular species of CPG in the heart during the early phase of ischemia and in arrhythmias, and should facilitate further studies on the metabolism of the individual species in health and disease.     

Isolation,characterization, and phylogenic analysis of three new severe fever with thrombocytopenia syndrome bunyavirus strains derived from Hubei Province,China

Hubei Province is a major epidemic area of severe fever with thrombocytopenia syndrome bunyavirus(SFTSV) in China. However, to date, a few SFTSV strains have been isolated from Hubei Province, preventing effective studies of epidemic outbreaks. Here, we report three confirmed patients(2015–2016) with typical symptoms of severe fever with thrombocytopenia syndrome disease(SFTS) who were farmers resident in different regions in Hubei Province. Three new SFTSV strains were isolated from the serum samples of each patient. Characterization of viral growth properties showed that there were no significant differences in virus production. All strains were completely sequenced, and phylogenetic analysis showed that unlike the other strains from Hubei province, which belonged to the SFTSV C3 genotype, one of the three strains belonged to the SFTSV C2 genotype. These results suggested that multiple SFTSV genotypes have been circulating in Hubei Province, providing insights into SFTSV evolution and improving our understanding of SFTSV prevalence in Hubei Province.     

Enantioselective synthesis and antioxidant activity of 3‐(3,4‐dihydroxyphenyl)‐glyceric acid—Basic monomeric moiety of a biologically active polyether from Symphytum asperum and S. caucasicum

The racemic and enantioselective synthesis of a novel glyceric acid derivative, namely, 2,3‐dihydroxy‐3‐(3,4‐dihydroxyphenyl)‐propionic acid as well as the antioxidant activities is described. The virtually pure enantiomers, (+)‐(2R,3S)‐2,3‐dihydroxy‐3‐(3,4‐dihydroxyphenyl)‐propionic acid and (?)‐(2S,3R)‐2,3‐dihydroxy‐3‐(3,4‐dihydroxyphenyl)‐propionic acid were synthesized for the first time via Sharpless asymmetric dihydroxylation of trans‐caffeic acid derivatives using the enantiocomplementary catalysts, (DHQD)₂‐PHAL and (DHQ)₂‐PHAL. The determination of enantiomeric purity of the novel chiral glyceric acid derivatives was performed by high‐performance liquid chromatographic techniques on the stage of their alkylated precursors. The novel glyceric acid derivatives show strong antioxidant activity against hypochlorite and N,N‐diphenyl‐N‐picryl‐hydrazyl free radical. Their antioxidant activity is about 40‐fold higher than that of the corresponding natural polyether and three‐fold higher of trans‐caffeic acid itself. Chirality, 2010. © 2010 Wiley‐Liss, Inc.     

Water soluble feruloyl arabinoxylans from rice and ragi: changes upon malting and their consequence on antioxidant activity

The objective of this study is to determine the changes brought about by germination on water soluble feruloyl arabinoxylans (feraxans), one of the major components of soluble fibre from rice and ragi and their consequence on antioxidant activity. Soluble feraxans, isolated from native and malted rice and ragi were fractionated on DEAE-cellulose. Ferulic acid content of the major [0.1 molar ammonium carbonate (AC) eluted] fraction was higher in malts (rice: 1045 microg/g; ragi: 1404 microg/g) than in native (rice: 119 microg/g; ragi: 147 microg/g) and this fraction was separated by Sephacryl S-300 chromatography into two peaks each in rice (native: 232 and 24.4 kDa; malt: 75.4 and 39.6 kDa) and ragi (native: 140 and 15.4 kDa; malt: 38.9 and 15.4 kDa). 0.1 molar AC eluted fractions showed very strong antioxidant activity in vitro as determined by beta-carotene-linoleate emulsion (IC50: 0.16-0.24 mg), DPPH* (IC50: 4.1-11.4 mg) and Ferric reducing/antioxidant power, FRAP (EC1: 0.76-3.1mg) assays. Antioxidant activity of feraxans was several (4.9-1400) folds higher than the expected activity due to their bound ferulic acid content. Apart from ferulic acid, presence of sugars with >C=O (uronyl/acetyl) groups and degree/nature of glycan-polymerization were observed to influence antioxidant activity of the polysaccharides. Malting resulted in many dynamic changes in the ferulic acid content in different feraxan types and affected their antioxidant activity.     

Structural and Biochemical Study of the Mono-ADP-Ribosyltransferase Domain of SdeA,a Ubiquitylating/Deubiquitylating Enzyme from Legionella pneumophila

Conventional ubiquitylation occurs through an ATP-dependent three-enzyme cascade (E1, E2, and E3) that mediates the covalent conjugation of the C-terminus of ubiquitin to a lysine on the substrate. SdeA, which belongs to the SidE effector family of Legionella pneumophila, can transfer ubiquitin to endoplasmic reticulum-associated Rab-family GTPases in a manner independent of E1 and E2 enzymes. The novel ubiquitin-modifying enzyme SdeA utilizes NAD⁺ as a cofactor to attach ubiquitin to a serine residue of the substrate. Here, to elucidate the coupled enzymatic reaction of NAD + hydrolysis and ADP-ribosylation of ubiquitin in SdeA, we characterized the mono-ADP-ribosyltransferase domain of SdeA and show that it consists of two sub-domains termed mART-N and mART-C. The crystal structure of the mART-C domain of SdeA was also determined in free form and in complex with NAD⁺ at high resolution. Furthermore, the spatial orientations of the N-terminal deubiquitylase, phosphodiesterase, mono-ADP-ribosyltransferase, and C-terminal coiled-coil domains within the 180-kDa full-length SdeA were determined. These results provide insight into the unusual ubiquitylation mechanism of SdeA and expand our knowledge on the structure–function of mono-ADP-ribosyltransferases.     

Molecular analysis of SMN1, SMN2, NAIP, GTF2H2, and H4F5 genes in 157 Chinese patients with spinal muscular atrophy

Spinal muscular atrophy (SMA) is a common and lethal autosomal recessive neurodegenerative disorder, which is caused by mutations of the survival motor neuron 1 (SMN1) gene. Additionally, the phenotype is modified by several genes nearby SMN1 in the 5q13 region. In this study, we analyzed mutations in SMN1 and quantified the modifying genes, including SMN2, NAIP, GTF2H2, and H4F5 by polymerase chain reaction–restriction fragment length polymorphism (PCR-RFLP), multiplex ligation-dependent probe amplification (MLPA), TA cloning, allele-specific long-range PCR, and Sanger sequencing in 157 SMA patients. Most SMA patients (94.90%) possessed a homozygous SMN1 deletion, while 10 patients demonstrated only the absence of exon 7, but the presence of exon 8. Two missense mutations (c.689 C > T and c.844 C > T) were identified in 2 patients who both carried a single copy of SMN1. We found inverse correlations between SMN2, the NAIP copy number, and the clinical severity of the disease. Furthermore, 7 severe type I patients possessed large-scale deletions, including SMN1, NAIP, and GTF2H2. We conclude that SMN1 gene conversion, SMN1 subtle mutations, SMN2 copy number, and the extent of deletion in the 5q13 region should all be considered in the genotype–phenotype analysis of SMA.     

Intra-specific hybridization: generator of genetic diversification and heterosis in Andrographis paniculata Nees. A bridge from extinction to survival

Andrographis paniculata (AP) has been stated as a low-diverse, endangered and red-listed plant species. Self-pollinated mating system, being an introduced species and experiencing a bottleneck as well as over exploitation cause such a consequence. Inter and intra-specific hybridizations have been suggested as essential techniques for generating genetic diversity. To test the effect of intra-specific hybridization on diversification and heterosis of AP, seven accessions were outcrossed manually in all 21 possible combinations. Three types of markers including morphological, phytochemical and RAPD markers were employed to evaluate the mentioned hypothesis. The results revealed that hybridization acted as a powerful engine for diversification of AP as it caused heterotic expression of the studied traits, simultaneously. Initially, it seems that additive and non-additive gene effects both can be considered as the genetic basis of heterosis in AP for the investigated traits. Agronomic and morphological traits were differentiated from each other, while positive heterosis was recorded mainly for agronomic traits but not for the morphological traits. Intra-specific hybridization increased the genetic diversity in AP population. Nevertheless, a part of this variation could also be attributed to the negative heterosis. The current exploration demonstrated the first ever conducted manual intra-specific hybridization among AP accessions in a mass scale. However, the 17 RAPD primers produced a monomorph pattern, but perhaps increasing the number of markers can feature a new genetic profile in this plant.