The Antioxidant properties of <Emphasis Type="Italic">para</Emphasis>-Aminobenzoic acid and its sodium salt

The antioxidant properties of para-aminobenzoic acid, a substance from the group of vitamins, and its sodium salt has been found using the reaction of adrenaline autoxidation in an alkaline medium as a superoxide-generating model system. These compounds inhibited the accumulation of adrenochrome, which is a product of adrenaline oxidation, and the formation of superoxide anions, detected by their reaction with nitro blue tetrazolium. Approaches have been developed to produce a true solution of para-aminobenzoic acid and conditions were established to measure the antioxidant activity of para-aminobenzoic acid and its sodium salt. The antioxidant properties of these compounds indicate their possible participation in the redox reactions of the cell and can also be one reason that they are essential.     

Effect of artificial redox mediators on the photoinduced oxygen reduction by photosystem I complexes

The peculiarities of interaction of cyanobacterial photosystem I with redox mediators 2,6-dichlorophenolindophenol (DCPIP) and N,N,N′,N′-tetramethyl-p-phenylenediamine (TMPD) were investigated. The higher donor efficiency of the reduced DCPIP form was demonstrated. The oxidized form of DCPIP was shown to be an efficient electron acceptor for terminal iron–sulfur cluster of photosystem I. Likewise methyl viologen, after one-electron reduction, DCPIP transfers an electron to the molecular oxygen. These results were discussed in terms of influence of these interactions on photosystem I reactions with the molecular oxygen and natural electron acceptors.     

Glutathione reductase gene expression depends on chloroplast signals in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

Glutathione reductase (EC 1.6.4.2) is one of the main antioxidant enzymes of the plant cell. In Arabidopsis thaliana, glutathione reductase is encoded by two genes: the gr1 gene encodes the cytosolic-peroxisomal form, and the gr2 gene encodes the chloroplast-mitochondrial form. Little is known about the regulation of expression of plant glutathione reductase genes. In the present work, we have demonstrated that gr2 (but not gr1) gene expression in Arabidopsis leaves changes depending on changes in redox state of the photosynthetic electron transport chain. Expression of both the gr1 and gr2 genes was induced by reactive oxygen species. In heterotrophic suspension cell culture of Arabidopsis, expression of both studied genes did not depend on H₂O₂ level or on changes in the redox state of the mitochondrial electron transport chain. Our data indicate that chloroplasts are involved in the regulation of the glutathione reductase gene expression in Arabidopsis.     

Electronic fine structure calculation of metal complexes with three-open-shell <Emphasis Type="Italic">s</Emphasis>, <Emphasis Type="Italic">d,and p</Emphasis> configurations

The ligand field density functional theory (LFDFT) algorithm is extended to treat the electronic structure and properties of systems with three-open-shell electron configurations, exemplified in this work by the calculation of the core and semi-core 1s, 2s, and 3s one-electron excitations in compounds containing transition metal ions. The work presents a model to non-empirically resolve the multiplet energy levels arising from the three-open-shell systems of non-equivalent ns, 3d, and 4p electrons and to calculate the oscillator strengths corresponding to the electric-dipole 3d ^m  → ns ¹3d ^m 4p ¹ transitions, with n = 1, 2, 3 and m = 0, 1, 2, …, 10 involved in the s electron excitation process. Using the concept of ligand field, the Slater-Condon integrals, the spin-orbit coupling constants, and the parameters of the ligand field potential are determined from density functional theory (DFT). Therefore, a theoretical procedure using LFDFT is established illustrating the spectroscopic details at the atomic scale that can be valuable in the analysis and characterization of the electronic spectra obtained from X-ray absorption fine structure or electron energy loss spectroscopies.     

Identification of a second <Emphasis Type="Italic">PAD1</Emphasis> in <Emphasis Type="Italic">Brettanomyces bruxellensis</Emphasis> LAMAP2480

Volatile phenols are aromatic compounds produced by some yeasts of the genus Brettanomyces as defense against the toxicity of hydroxycinnamic acids (p-coumaric acid, ferulic acid and caffeic acid). The origin of these compounds in winemaking involves the sequential action of two enzymes: coumarate decarboxylase and vinylphenol reductase. The first one converts hydroxycinnamic acids into hydroxystyrenes, which are then reduced to ethyl derivatives by vinylphenol reductase. Volatile phenols derived from p-coumaric acid (4-vinylphenol and 4-ethylphenol) have been described as the major contributors to self-defeating aromas associated with stable, gouache, wet mouse, etc., which generates large economic losses in the wine industry. The gene responsible for the production of 4-vinylphenol from p-coumaric acid has been identified as PAD1, which encodes a phenylacrylic acid decarboxylase. PAD1 has been described for many species, among them Candida albicans, Candida dubliniensis, Debaryomyces hansenii and Pichia anomala. In Brettanomyces bruxellensis LAMAP2480, a 666 bp reading frame (DbPAD) encodes a coumarate decarboxylase. Recent studies have reported the existence of a new reading frame belonging to DbPAD called DbPAD2 of 531 bp, which could encode a protein with similar enzymatic activity to PAD1. The present study confirmed that the transformation of Saccharomyces cerevisiae strain BY4722 with reading frame DbPAD2 under the control of the B. bruxellensis ACT1 promoter, encodes an enzyme with coumarate decarboxylase activity. This work has provided deeper insight into the origin of aroma defects in wine due to contamination by Brettanomyces spp.     

Estimation of biophysical characteristics for <Emphasis Type="Italic">Chlamydomonas reinhardtii</Emphasis> pigment mutants with an M-PEA-2 fluorometer

Light green pigment mutants with a reduced chlorophyll b content were constructed in the microalga Chlamydomonas reinhardtii Dangeard. A simultaneous recording of the induction curves for prompt and delayed fluorescence and the redox state of P700 in the microsecond range with a M-PEA-2 fluorometer revealed decreases in the quantum yield of electron transport in PS2 (φE₀) and the performance index (PI_ABS) and increases in the quantum efficiency of energy dissipation (φD₀) and ΔpH-dependent nonphotochemical quenching (qE and NPQ). The light-dependence curves of the fluorescence parameters confirmed a decrease in the coefficient of maximum utilization of light energy (α) for the mutants. However, the mutants showed an adequate rate of electron transport at a medium light intensity under steady-state conditions. The mutations did not directly affect the oxidation reactions of the PS1 pigment (P700) and the decrease in delayed fluorescence. Experience in using the mutants to test polluted waters of Kazakhstan confirmed that the mutants are promising for use in biomonitoring for mutagens.     

Evaluation of <Emphasis Type="Italic">Galleria mellonella</Emphasis> larvae as an in vivo model for assessing the relative toxicity of food preservative agents

Larvae of Galleria mellonella are widely used for evaluating the virulence of microbial pathogens and for measuring the efficacy of anti-microbial agents and produce results comparable to those that can be obtained using mammals. In this work, the suitability of using G. mellonella larvae to measure the relative toxicity of a variety of food preservatives was evaluated. The response of larvae to eight commonly used food preservatives (potassium nitrate, potassium nitrite, potassium sorbate, sodium benzoate, sodium nitrate, sodium chloride, sodium nitrite and sodium acetate) administered by feeding or by intra-haemocoel injection was measured. A significant correlation between the LD₅₀ (R ²?=?0.8766, p?=?0.0006) and LD₈₀ (R ²?=?0.7629, p?=?0.0046) values obtained due to oral or intra-haemocoel administration of compounds was established. The response of HEp-2 cells to the food preservatives was determined, and a significant correlation (R ²?=?0.7217, p?=?0.0076) between the LD₅₀ values of the compounds administered by feeding in larvae with the IC₅₀ values of the compounds in HEp-2 cells was established. A strong correlation between the LD₅₀ values of the eight food preservatives in G. mellonella larvae and rats (R ²?=?0.6506, p?=?0.0156) was demonstrated. The results presented here indicate that G. mellonella larvae may be used as a model to evaluate the relative toxicity of food preservatives, and the results show a strong positive correlation to those obtained using established cell culture and mammalian models.     

Thermostable phycocyanin from the red microalga <Emphasis Type="Italic">Cyanidioschyzon merolae</Emphasis>, a new natural blue food colorant

The demand for natural food colorants is growing as consumers question the use of artificial colorants more and more. The phycobiliprotein C-phycocyanin of Arthospira platensis is used as a natural blue colorant in certain food products. The thermoacidophilic red microalga Cyanidioschyzon merolae might provide an alternative source of phycocyanin. Cyanidioschyzon merolae belongs to the order Cyanidiophyceae of the phylum Rhodophyta. Its natural habitat are sulfuric hot springs and geysers found near volcanic areas in, e.g., Yellowstone National Park in the USA and in Java, Indonesia. It grows optimally at a pH between 0.5 and 3.0 and at temperatures up to 56 °C. The low pH at which C. merolae grows minimizes the risk of microbial contamination and could limit production loss. As C. merolae lacks a cell wall, phycocyanin with a high purity number of 9.9 could be extracted by an osmotic shock using a simple ultrapure water extraction followed by centrifugation. The denaturation midpoint at pH 5 was 83 °C, being considerably higher than the A. platensis phycocyanin (65 °C). The C. merolae phycocyanin was relatively stable at pH 4 and 5 up to 80 °C. The high thermostability at slightly acidic pH makes the C. merolae phycocyanin an interesting alternative to A. platensis phycocyanin as a natural blue food colorant.     

Association of genes of different functional classes with type 1 diabetes

The genetic structure of susceptibility to type 1 diabetes (T1D) in the population of Tomsk was studied. We had a group of T1D patients (N = 285) and a population sample (N = 300) and we studied 58 SNPs localized in the 47 genes which products are involved in various metabolic pathways and processes as fibrogenesis, endothelial dysfunction, and inflammation. Genotyping was performed by mass spectrometry using the Sequenom MassARRAY system (United States). We compared the group of T1D patients and the population sample and found an association with the predisposition to disease for seven markers: rs3765124 of the ADAMDEC1 gene, genotype AA (p = 0.004), allele A (p = 0.033); rs1007856 of the ITGB5 gene, genotype TT (p = 0.015), allele T (p = 0.036); rs20579 of the LIG1 gene, genotype CC (p = 0.004), allele C (p = 0.002); rs12980602 of the IFNL2 gene, allele C (p = 0.029); rs4986819 of the PARP4 gene, allele C (p = 0.044); rs1143674 of the ITGA4 gene genotype GG (p = 0.002); rs679620 of the MMP3 gene, genotype AA (p = 0.008). Thus, the products of genes associated with T1D belong to different molecular classes: metalloproteases (ADAMDEC1, MMP3), cytokines (IL28A), cell surface receptors (ITGA4), adhesion molecules (ITGB5), DNA ligases (LIG1), and ribosyltransferase enzymes (PARP4). The ADAMDEC1, ITGA4, and ITGB5 genes belong to two biological processes: cell communication and signal transduction. The LIG1 and PARP4 genes regulate the metabolism of nucleic acids, MMP3 is involved in the regulation of protein metabolism, and the IFNL2 is involved in the immune response.     

Association of <Emphasis Type="Italic">p</Emphasis>53 codon 72 polymorphism and survival of North Indian lung cancer patients treated with platinum-based chemotherapy

p53 helps in maintaining genomic stability by undergoing cellular arrest, DNA repair or cellular apoptosis during DNA damage. So, as to find the association of p53Arg ⁷² Pro towards lung carcinogenesis and overall survival of North Indian lung cancer patients, single nucleotide polymorphic variant (rs1042522) was analyzed. 840 subjects including 420 cases and 420 controls were recruited and genotyped using PCR-RFLP technique for p53Arg ⁷² Pro polymorphic site. Association was analyzed using adjusted odds ratio along with its confidence intervals (95?% CI) and p value predicted from logistic regression whereas overall survival for lung cancer patients was obtained using Kaplan–Meir and Cox regression model for different parameters to obtain hazard ratio and survival time with statistical significance (log-rank p value). None of the variant genotypes for p53Arg ⁷² Pro showed any association towards lung cancer risk or any specific histological subtype. Lung cancer subjects with Pro/Pro genotype had better median survival time as compared to Arg/Pro genotype (10 months; HR?=?0.65; 95?% CI?=?0.45–0.95; p?=?0.03). Furthermore, female lung cancer patients with Arg/Pro (HR?=?0.08; 95?% CI?=?0.02–0.34; p?=?0.0005) and Pro/Pro (HR?=?0.21; 95?% CI?=?0.06–0.67; p?=?0.008) genotypes showed a better overall survival and hence a better prognosis as compared to males. Our data also reveals that lung cancer patients with ECOG scores between 0 and 1 and carrying the Pro/Pro had better chances of survival. p53 codon 72 polymorphism could play a role as a prognostic marker in lung cancer patients.     

Inhibition of lipase activity by low-molecular-weight chitosan

Inhibition of enzymatic activity of lipase (EC 3.1.1.3) from the fungus Candida rugosa and wheat (Triticum aestivum L.) germ by low-molecular-weight chitosan with an average molecular weight of 5.7 kDa in reactions of p-nitrophenyl palmitate cleavage was studied. Preincubation of lipases with chitosan, prior to addition of the substrate to solution, showed that equilibrium during the lipase-inhibitor complex formation was reached within 30 min. The inhibition constants for C. rugosa lipase and wheat germ lipase were 1.4 and 0.9 mM, respectively. The contribution of electrostatic interactions to the complex formation between chitosan and lipases is insignificant.     

Photosynthetic activity and components of the electron transport chain in the aerobic bacteriochlorophyll <Emphasis Type="Italic">a</Emphasis>-containing bacterium <Emphasis Type="Italic">Roseinatronobacter thiooxidans</Emphasis>

Bioenergetics of the aerobic bacteriochlorophyll a-containing (BCl a) bacterium (ABC bacterium) Roseinatronobacter thiooxidans is a combination of photosynthesis, oxygen respiration, and oxidation of sulfur compounds under alkaliphilic conditions. The photosynthetic activity of Rna. thiooxidans cells was established by the photoinhibition of cell respiration and reversible photobleaching discoloration of the BCl a of reaction centers (RC), connected by the chain of electron transfer with cytochrome c ₅₅₁ oxidation. The species under study, like many purple bacteria and some of the known ABC bacteria, possesses a light-harvesting pigment-protein (LHI) complex with the average number of 30 molecules of antenna BCl a per one photosynthetic RC. Under microaerobic growth conditions, the cells contained bc ₁ complex and two terminal oxidases: cbb ₃-cytochrome oxidase and the alternative cytochrome oxidase of the a ₃ type. Besides, Rna. thiooxidans was shown to have several different soluble low- and high-potential cytochromes c, probably associated with the ability of utilizing sulfur compounds as additional electron donors.     

Inheritance of Traits Controlled by Odd Chromosomes Using Data on Transmission of Monosomic Addition S<Superscript>t</Superscript> Chromosome of the <Emphasis Type="Italic">Elymus Sibiricus</Emphasis> Genome

On the basis of the winter bread wheat cultivar Obryi, two independent disomic addition lines BC₁₂F_∞ with the chromosome of the E. sibiricus S^t genome are created. A practical algorithm for determining the probabilities of transmission of the odd chromosome separately through male and female gametes in selfpollination of hemizygous hybrids from the equation p²–(1 + f₁–f₄) × p + f₁ = 0 is proposed, where p is the probability of the formation of viable gametes with the considered chromosome and f₁ and f₄ are the empirical frequencies of the corresponding homozygotes with and without the trait. The probability of transmission of an alien univalent chromosome through pollen (p_♂) is associated with the frequency of its transmission through the egg cell (p_♀) in backcrosses and in self-pollination (1–f₄) by the equation p_♂ = 1–f₄/(1–p_♀). The calculated empirically dependent estimates of the probabilities of transmission of the added chromosome through the egg cell p_♀ = 18.7% and through pollen p_♂ = 4.3% correspond to the empirical frequencies obtained for backcrosses. The coefficients of the gamete selection V_♀ = 0.748 and V_♂ = 0.172 are calculated, and the expected segregation for the alien trait controlled by a dominant gene located in the added chromosome is determined—with the trait: without the trait is 0.222: 0.778 in F₂; 0.187: 0.813 in equational and 0.043: 0.957 in certational backcrosses.     

Dereplication of natural products from complex extracts by regression analysis and molecular networking: case study of redox-active compounds from <Emphasis Type="Italic">Viola alba</Emphasis> subsp. <Emphasis Type="Italic">dehnhardtii</Emphasis>

Introduction

In natural product research, bioassay-guided fractionation was previously widely employed but is now judged to be inadequate in terms of time and cost, particularly if only known compounds are ultimately isolated. The development of metabolomics, along with improvements in analytical tools, allows comprehensive metabolite profiling. This enables dereplication to target unknown active compounds early in the purification workflow.

Objectives

Starting from an ethanolic extract of violet leaves, this study aims to predict redox active compounds within a complex matrix through an untargeted metabolomics approach and correlation analysis.

Methods

Rapid fractionation of crude extracts was carried out followed by multivariate data analysis (MVA) of liquid chromatography–high resolution mass spectrometry (LC–HRMS) profiles. In parallel, redox active properties were evaluated by the capacity of the molecules to reduce 2,2-diphenyl-1-picrylhydrazyl (DPPH^·) and superoxide (O₂ ^·?) radicals using UV–Vis and electron spin resonance spectroscopies (ESR), respectively. A spectral similarity network (molecular networking) was used to highlight clusters involved in the observed redox activities.

Results

Dereplication on Viola alba subsp. dehnhardtii highlighted a reproducible pool of redox active molecules. Polyphenols, particularly O-glycosylated coumarins and C-glycosylated flavonoids, were identified and de novo dereplicated through molecular networking. Confirmatory analyses were undertaken by thin layer chromatography (TLC)–DPPH–MS assays and nuclear magnetic resonance (NMR) spectra of the most active compounds.

Conclusion

Our dereplication strategy allowed the screening of leaf extracts to highlight new biologically active metabolites in few steps with a limited amount of crude material and reduced time-consuming manipulations. This approach could be applied to any kind of natural extract for the study of various biological activities.

     

Differences in specificity and compensatory functions among three major starch synthases determine the structure of amylopectin in rice endosperm

Key message

This work adds a new player, HER2, downstream of the perception of E-2-hexenal, a green leaf volatile, and shows that E-2-hexenal specifically changes the redox status of the mitochondria.

Abstract

It is widely accepted that plants produce and respond to green leaf volatiles (GLVs), but the molecular components involved in transducing their perception are largely unknown. The GLV E-2-hexenal inhibits root elongation in seedlings and, using this phenotype, we isolated E-2-hexenal response (her) Arabidopsis thaliana mutants. Using map-based cloning we positioned the her2 mutation to the At5g63620 locus, resulting in a phenylalanine instead of serine on position 223. Knockdown and overexpression lines of HER2 confirmed the role of HER2, which encodes an oxidoreductase, in the responsiveness to E-2-hexenal. Since E-2-hexenal is a reactive electrophile species, which are known to influence the redox status of cells, we utilized redox sensitive GFP2 (roGFP2) to determine the redox status of E-2-hexenal-treated root cells. Since the signal peptide of HER2 directed mCherry to the mitochondria, we targeted the expression of roGFP2 to this organelle besides the cytosol. E-2-hexenal specifically induced a change in the redox status in the mitochondria. We did not see a difference in the redox status in her2 compared to wild-type Arabidopsis. Still, the mitochondrial redox status did not change with Z-3-hexenol, another abundant GLV. These results indicate that HER2 is involved in transducing the perception of E-2-hexenal, which changes the redox status of the mitochondria.

     

Inhibition of lipase activity by low-molecular-weight chitosan

Inhibition of enzymatic activity of lipase (EC 3.1.1.3) from the fungus Candida rugosa and wheat (Triticum aestivum L.) germ by low-molecular-weight chitosan with an average molecular weight of 5.7 kDa in reactions of p-nitrophenyl palmitate cleavage was studied. Preincubation of lipases with chitosan, prior to addition of the substrate to solution, showed that equilibrium during the lipase-inhibitor complex formation was reached within 30 min. The inhibition constants for C. rugosa lipase and wheat germ lipase were 1.4 and 0.9 mM, respectively. The contribution of electrostatic interactions to the complex formation between chitosan and lipases is insignificant.     

Targeting myomiRs by tocotrienol-rich fraction to promote myoblast differentiation

Background

Several muscle-specific microRNAs (myomiRs) are differentially expressed during cellular senescence. However, the role of dietary compounds on myomiRs remains elusive. This study aimed to elucidate the modulatory role of tocotrienol-rich fraction (TRF) on myomiRs and myogenic genes during differentiation of human myoblasts. Young and senescent human skeletal muscle myoblasts (HSMM) were treated with 50 μg/mL TRF for 24 h before and after inducing differentiation.

Results

The fusion index and myotube surface area were higher (p?<?0.05) on days 3 and 5 than that on day 1 of differentiation. Ageing reduced the differentiation rate, as observed by a decrease in both fusion index and myotube surface area in senescent cells (p?<?0.05). Treatment with TRF significantly increased differentiation at days 1, 3 and 5 of young and senescent myoblasts. In senescent myoblasts, TRF increased the expression of miR-206 and miR-486 and decreased PTEN and PAX7 expression. However, the expression of IGF1R was upregulated during early differentiation and decreased at late differentiation when treated with TRF. In young myoblasts, TRF promoted differentiation by modulating the expression of miR-206, which resulted in the reduction of PAX7 expression and upregulation of IGF1R.

Conclusion

TRF can potentially promote myoblast differentiation by modulating the expression of myomiRs, which regulate the expression of myogenic genes.

     

An Investigation into Membrane Bound Redox Carriers Involved in Energy Transduction Mechanism in <Emphasis Type="Italic">Brevibacterium linens</Emphasis> DSM 20158 with Unsequenced Genome

Brevibacterium linens (B. linens) DSM 20158 with an unsequenced genome can be used as a non-pathogenic model to study features it has in common with other unsequenced pathogens of the same genus on the basis of comparative proteome analysis. The most efficient way to kill a pathogen is to target its energy transduction mechanism. In the present study, we have identified the redox protein complexes involved in the electron transport chain of B. linens DSM 20158 from their clear homology with the shot-gun genome sequenced strain BL2 of B. linens by using the SDS–Polyacrylamide gel electrophoresis coupled with nano LC–MS/MS mass spectrometry. B. linens is found to have a branched electron transport chain (Respiratory chain), in which electrons can enter the respiratory chain either at NADH (Complex I) or at Complex II level or at the cytochrome level. Moreover, we are able to isolate, purify, and characterize the membrane bound Complex II (succinate dehydrogenase), Complex III (menaquinone cytochrome c reductase cytochrome c subunit, Complex IV (cytochrome c oxidase), and Complex V (ATP synthase) of B. linens strain DSM 20158.     

Bioaccessibility of phenolic compounds,lutein, and bioelements of preparations containing <Emphasis Type="Italic">Chlorella vulgaris</Emphasis> in artificial digestive juices

Chlorella vulgaris Beijerinck is a spherical, green alga belonging to the genus Chlorella and family Chlorellaceae. It has high nutritional value and shows multiple biological effects. Dietary supplements that contain extracts of C. vulgaris are sold in the form of tablets, capsules, powders, and aqueous solutions. To the best of our knowledge, this is the first study to determine the content of bioelements (zinc, iron, and magnesium), phenolic compounds, and lutein before and after incubation with artificial digestive juices from preparations containing C. vulgaris. In this study, we used commercial preparations in the form of powder and tablets. The samples were incubated in artificial gastric juice and then in artificial intestinal juice for 30 and 90 min. The contents of bioelements were determined by using the flame atomic absorption spectrometric method. Lutein and phenolic compounds were analyzed by high-pressure liquid chromatography. We also aimed to evaluate the quality of chlorella-containing formulations by using the methods described in the European Pharmacopoeia 8th edition. According to the results, the preparations containing C. vulgaris demonstrated the presence of phenolic compounds and lutein. Therefore, daily supplementation of preparations containing C. vulgaris substantiates its usefulness for humans. The qualitative composition of the examined organic substances and bioelements was found to be in accordance with the manufacturer’s declarations on the packaging containing C. vulgaris compared with the control samples; however, the contents of bioelements were found to be negligible after incubation with artificial digestive juices. This shows that the examined preparations containing C. vulgaris are not good sources of bioelements such as zinc, iron, or magnesium.