Excess Peroxide Research Articles

Early modification of cytochrome c by hydrogen peroxide triggers its fast degradation

Cytochrome c (cyt c), in addition to its function as an electron shuttle in respiratory chain, is able to perform as a pseudo-peroxidase with a critical role during apoptosis. Incubation of cyt c with an excess of hydrogen peroxide leads to a suicide inactivation of the protein, which is accompanied by heme destruction and covalent modification of numerous amino acid residues. Although steady-state reactions of cyt c with an excess of hydrogen peroxide represent non-physiological conditions, they might be used for analysis of the first-modified amino acid in in vivo. Here, we observed oxidation of tyrosine residues 67 and 74 and heme as the first modifications found upon incubation with hydrogen peroxide. The positions of the oxidized tyrosines suggest a possible migration pathway of hydrogen peroxide-induced radicals from the site of heme localization to the protein surface. Analysis of a size of folded fraction of cyt c upon limited incubation with hydrogen peroxide indicates that the early oxidation of amino acids triggers an accelerated destruction of cyt c. Position of channels from molecular dynamics simulation structures of cyt c points to a location of amino acid residues exposed to reactive oxidants that are thus more prone to covalent modification.

Silicon nanowires decorated with platinum nanoparticles were applied for photothermal-enhanced sonodynamic therapy.

Sonodynamic therapy (SDT) triggered by ultrasound (US) can overcome pivotal limitations of photo-therapy owing to its high depth-penetration and low phototoxicity. However, there is still a need to develop more efficient sonosensitizes to enhance the therapy efficiency.Methods: In this study, Pt nanoparticles (Pt NPs) are reduced on silicon nanowires (SiNWs) by in situ reduction to prepare Si-Pt nanocomposites (Si-Pt NCs).Results: Si-Pt NCs can produce reactive oxygen radicals (ROS) under ultrasound (US) irradiation, which have sonodynamic therapy (SDT) effect. Meanwhile, Si-Pt NCs can convert excess hydrogen peroxide (H2O2) into ROS in the tumor microenvironment, which endow strong chemodynamic therapy (CDT) effect. Taking the advantages of the mesoporous structure of SiNWs, the SDT and CDT effects of Si-Pt NCs are stronger than those of the pure Pt NPs and SiNWs. Besides, the mild photothermal effect of Si-Pt NCs further improves the SDT&CDT activity and realizes the combined cancer therapy.Conclusion: The developed Si-Pt NCs with the ability of photothermal enhanced SDT/CDT combined therapy play a momentous role in the novel cancer treatment.

Exogenous melatonin improves salt stress adaptation of cotton seedlings by regulating active oxygen metabolism.

Melatonin is a small-molecule indole hormone that plays an important role in participating in biotic and abiotic stress resistance. Melatonin has been confirmed to promote the normal development of plants under adversity stress by mediating physiological regulation mechanisms. However, the mechanisms by which exogenous melatonin mediates salt tolerance via regulation of antioxidant activity and osmosis in cotton seedlings remain largely unknown. In this study, the regulatory effects of melatonin on reactive oxygen species (ROS), the antioxidant system, and osmotic modulators of cotton seedlings were determined under 0–500 µM melatonin treatments with salt stress induced by 150 mM NaCl treatment. Cotton seedlings under salt stress exhibited an inhibition of growth, excessive hydrogen peroxide (H2O2), superoxide anion (O2−), and malondialdehyde (MDA) accumulations in leaves, increased activity levels of superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and ascorbate peroxidase (APX), and elevated ascorbic acid (AsA) and glutathione (GSH) content in leaves. However, the content of osmotic regulators (i.e., soluble sugars and proteins) in leaves was reduced under salt stress. This indicates high levels of ROS were produced, and the cell membrane was damaged. Additionally, osmotic regulatory substance content was reduced, resulting in osmotic stress, which seriously affected cotton seedling growth under salt stress. However, exogenous melatonin at different concentrations reduced the contents of H2O2, O2−, and MDA in cotton leaves, increased the activity of antioxidant enzymes and the content of reductive substances (i.e., AsA and GSH), and promoted the accumulation of osmotic regulatory substances in leaves under salt stress. These results suggest that melatonin can inhibit ROS production in cotton seedlings, improve the activity of the antioxidant enzyme system, raise the content of osmotic regulation substances, reduce the level of membrane lipid peroxidation, and protect the integrity of the lipid membrane under salt stress, which reduces damage caused by salt stress to seedlings and effectively enhances inhibition of salt stress on cotton seedling growth. These results indicate that 200 µM melatonin treatment has the best effect on the growth and salt tolerance of cotton seedlings.

Severe reactive astrocytes precipitate pathological hallmarks of Alzheimer's disease via H2O2- production.

Although the pathological contributions of reactive astrocytes have been implicated in Alzheimer's disease (AD), their in vivo functions remain elusive due to the lack of appropriate experimental models and precise molecular mechanisms. Here, we show the importance of astrocytic reactivity on the pathogenesis of AD using GiD, a newly developed animal model of reactive astrocytes, where the reactivity of astrocytes can be manipulated as mild (GiDm) or severe (GiDs). Mechanistically, excessive hydrogen peroxide (H2O2) originated from monoamine oxidase B in severe reactive astrocytes causes glial activation, tauopathy, neuronal death, brain atrophy, cognitive impairment and eventual death, which are significantly prevented by AAD-2004, a potent H2O2 scavenger. These H2O2--induced pathological features of AD in GiDs are consistently recapitulated in a three-dimensional culture AD model, virus-infected APP/PS1 mice and the brains of patients with AD. Our study identifies H2O2 from severe but not mild reactive astrocytes as a key determinant of neurodegeneration in AD.

Endophytic Bacillus altitudinis WR10 alleviates Cu toxicity in wheat by augmenting reactive oxygen species scavenging and phenylpropanoid biosynthesis

Soil copper (Cu) pollution severely stunts crops growth and limits sustainable agri-food production. Many microbes are widely used for remediation of polluted soil, including Cu pollution. In this study, the potential of an endophytic Bacillus altitudinis WR10 to protect wheat from Cu stress and the molecular mechanisms were investigated using hydroponic model. The Cu resistance assay showed B. altitudinis WR10 can resist up to 2 mM Cu and remove about 74% Cu in medium after 24 h of fermentation. Co-culture study demonstrated WR10 increased roots length and dry weight in wheat seedlings under 50 μM Cu. These results indicated that WR10 was a Cu-resistant strain and reduced Cu toxicity in wheat. Transcriptome data and biochemical tests of wheat roots indicated that WR10 alleviated Cu toxicity through enhancing peroxidases (PODs) gene expression and activity to remove excess hydrogen peroxide (H2O2) and down-regulating glutathione S-transferases (GSTs) to increase glutathione (GSH) level. Moreover, enrichment and pathway analysis indicated WR10 regulated the expression of genes involved in phenylpropanoid biosynthesis, which may improve phenolic acids accumulation for protecting plant cells from Cu toxicity. Overall, this study revealed that B. altitudinis WR10 alleviated Cu toxicity in wheat via augmenting reactive oxygen species scavenging and phenylpropanoid biosynthesis.

Different Oxidation Pathways of 2-Selenouracil and 2-Thiouracil, Natural Components of Transfer RNA.

Sulfur- and selenium-modified uridines present in the wobble position of transfer RNAs (tRNAs) play an important role in the precise reading of genetic information and tuning of protein biosynthesis in all three domains of life. Both sulfur and selenium chalcogens functionally operate as key elements of biological molecules involved in the protection of cells against oxidative damage. In this work, 2-thiouracil (S2Ura) and 2-selenouracil (Se2Ura) were treated with hydrogen peroxide at 1:0.5, 1:1, and 1:10 molar ratios and at selected pH values ranging from 5 to 8. It was found that Se2Ura was more prone to oxidation than its sulfur analog, and if reacted with H2O2 at a 1:1 or lower molar ratio, it predominantly produced diselenide Ura-Se-Se-Ura, which spontaneously transformed to a previously unknown Se-containing two-ring compound. Its deselenation furnished the major reaction product, a structure not related to any known biological species. Under the same conditions, only a small amount of S2Ura was oxidized to form Ura-SO2H and uracil (Ura). In contrast, 10-fold excess hydrogen peroxide converted Se2Ura and S2Ura into corresponding Ura-SeOnH and Ura-SOnH intermediates, which decomposed with the release of selenium and sulfur oxide(s) to yield Ura as either a predominant or exclusive product, respectively. Our results confirmed significantly different oxidation pathways of 2-selenouracil and 2-thiouracil.

Tailored core‒shell dual metal–organic frameworks as a versatile nanomotor for effective synergistic antitumor therapy

Malignant tumor has become an urgent threat to global public healthcare. Because of the heterogeneity of tumor, single therapy presents great limitations while synergistic therapy is arousing much attention, which shows desperate need of intelligent carrier for co-delivery. A core‒shell dual metal–organic frameworks (MOFs) system was delicately designed in this study, which not only possessed the unique properties of both materials, but also provided two individual specific functional zones for co-drug delivery. Photosensitizer indocyanine green (ICG) and chemotherapeutic agent doxorubicin (DOX) were stepwisely encapsulated into the nanopores of MIL-88 core and ZIF-8 shell to construct a synergistic photothermal/photodynamic/chemotherapy nanoplatform. Except for efficient drug delivery, the MIL-88 could be functioned as a nanomotor to convert the excessive hydrogen peroxide at tumor microenvironment into adequate oxygen for photodynamic therapy. The DOX release from MIL-88-ICG@ZIF-8-DOX nanoparticles was triggered at tumor acidic microenvironment and further accelerated by near-infrared (NIR) light irradiation. The in vivo antitumor study showed superior synergistic antitumor effect by concentrating the nanoparticles into dissolving microneedles as compared to intravenous and intratumoral injection of nanoparticles, with a significantly higher inhibition rate. It is anticipated that the multi-model synergistic system based on dual-MOFs was promising for further biomedical application.

Excess hydrogen peroxide inhibits head and foot regeneration in hydra by affecting DNA repair and expression of essential genes.

Reactive oxygen species (ROS) are necessary for various cellular processes. However, excess ROS cause damage to many biological molecules and therefore must be tightly regulated in time and space. Hydrogen peroxide (H2 O2 ) is the most commonly used ROS as second messenger in the cell. It is a relatively long-lived freely diffusible signaling molecule during early events of injury. In the Cnidarian hydra, injury-induced ROS production is essential for regeneration to proceed. In the present study, we have examined influence of varying exposure to H2 O2 on head and foot regeneration in the middlepieces of trisected hydra. We find that longer (4 hours) exposure to 1 mM H2 O2 inhibits both head and foot regeneration while shorter exposure (2 hours) does not. Longer exposure to H2 O2 resulted in extensive damage to DNA that could not be repaired, probably due to suboptimal induction of APE1, an enzyme necessary for base excision repair (BER). Concomitantly, genes involved in activation of Wnt pathway, necessary for head regeneration, were significantly downregulated. This appeared to be due to failure of both stabilization and transient nuclear localization of β-catenin. Similarly, genes involved in foot regeneration were also downregulated on longer exposure to H2 O2 . Thus, exposure to excess ROS inhibits regenerative processes in hydra through reduced expression of genes involved in regeneration and diminished DNA repair.

Nuclear factor erythroid-2 related factor 2 inhibits human disc nucleus pulpous cells apoptosis induced by excessive hydrogen peroxide.

OBJECTIVE Nuclear factor erythroid-2 related factor 2 (Nrf2)/ antioxidant response element (ARE) is a novel defensive pathway involved in the oxidative and chemical stress of cells. The aim of the study was to explore the role of Nrf2 on the apoptosis of human disc nucleus pulpous cells induced by hydrogen peroxide (H2O2). METHODS The degeneration model of human intervertebral disc nucleus pulpous cells was established. The expression of Nrf2 was interfered with using sulforaphane (SFN); for that end, three groups were established: a blank group (H2O2-/SFN-), control group (H2O2+/SFN-), and an experimental group (H2O2+/SFN+). CCK8, Hoechst 33258 living cell staining was used to detect reactive oxygen species (ROS) content. RESULTS The apoptotic rates of the three groups were [(0.40±0.46)%], [(25.98±11.28)%], and [(3.83±2.06)%, respectively. The difference was statistically significant (p<0.05). The relative content of ROS in the three groups was [(100±7)%], [(1538±91)%], and [(818±63)%]; the difference was statistically significant (p<0.05). In Western blotting, Nrf2 content in the experimental group was higher than that in the control group. CONCLUSION Nrf2 exists in the nucleus pulpous cells of human intervertebral discs, which is related to the degeneration of the intervertebral disc. It has negative feedback regulation and can prevent the degeneration of the intervertebral disc by inhibiting the apoptosis of nucleus pulpous cells of human intervertebral discs caused by excessive ROS, which provides a new intervention strategy for the prevention and treatment of the degeneration of intervertebral discs.

A novel fluorogenic probe for visualizing the hydrogen peroxide in Parkinson’s disease models

Parkinson’s disease (PD) is closely related to the oxidative stress induced by excess hydrogen peroxide (H2O2) in organisms. Developing an efficient method for noninvasive and real-time H2O2 detection is beneficial to investigate the role played by H2O2 in PD. In this work, a novel fluorogenic probe (CBH) for living organisms H2O2 detection has been designed, synthesized and characterized. The emission of CBH in PBS solution is very weak. However, when H2O2 was added, the fluorescence of CBH solution was sharply increased for 12-fold, accompanied by the emission peak blue-shifted from 600 to 530 nm. Moreover, the response of CBH to H2O2 is highly sensitive and selective and is not affected by various ROS/RNS, anions, cations, and amino acids. Based on the good performance of CBH for H2O2 detection, it has been successfully applied to visualizing the H2O2 concentration in living cells, Zebrafish and [Formula: see text]. elegans PD models.

Melatonin ameliorates murine fetal oocyte meiotic dysfunction in F1 and F2 offspring caused by nicotine exposure during pregnancy

Although there is abundant evidence to demonstrate that maternal smoking during pregnancy will harm the health of future generations, the impact of nicotine use by pregnant woman upon the oogenesis and folliculogenesis of female offspring has not been as widely scrutinized. Here we focus on the effects of nicotine on the meiotic progression of fetal oocytes. The data indicated that in pregnant mice treated with nicotine, intracellular ROS increased in follicles within the fetal ovary. Excessive intracellular hydrogen peroxide (H2O2) and superoxide anion (O2-) decreased mitochondrial membrane potential, inducing mitochondrial dysfunction, triggering an autophagic cascade and inhibiting anti-autophagic proteins. Fetal oocytes in F1 offspring of pregnant mice treated with nicotine exhibited a delay in meiotic prophase I, especially from the stage of pachytene to diplotene. In pubertal F1 offspring we observed a reduced number of follicles; the same reduction was also observed in F2 offspring. Of note, we found that melatonin ameliorated nicotine-induced oocyte damage and increased the expression of MnSOD, which decreased the production of nicotine-induced intracellular ROS. In addition, melatonin also maintained normal H3K4 and H3K9 di- and tri-methylation in F1 and F2 ovaries. Taken together, the current evidence suggests that, in the mouse, melatonin could prevent nicotine-impaired fetal oogenesis and folliculogenesis in offspring.

Process Optimization for the Production of Ferric Sulfate Coagulant by the Oxidation of Ferrous sulfate with Hydrogen Peroxide

AbstractMotivated by the oxidative power of hydrogen peroxide and its environmentally attractive properties, the present study aimed to determine the optimum conditions for the production of ferric sulfate coagulant from ferrous sulfate The independent variables studied were the temperature (7.5–27.5 °C), amount of hydrogen peroxide (100–300 %) of the stoichiometric amount for the oxidation reaction, and dilution of the reaction medium using water (100–300 %) of the stoichiometric amount for the oxidation reaction. For the optimum conditions achieved, it was possible to obtain a conversion of 96.17 % of Fe+2to Fe+3, using a small hydrogen peroxide excess of 50 %, resulting in a product suitable for use as a coagulant in water treatment. It was found that an oxidation potential (Eh) greater than 0.7 volts corresponded to the conversion of Fe+2to Fe+3greater than 90 %, indicating Eh to be useful for reaction control in an industrial process.

Facile Synthesis of High Performance Iron Oxide/Carbon Nanocatalysts Derived from the Calcination of Ferrocenium for the Decomposition of Methylene Blue

Iron oxide/carbon nanocatalysts were successfully synthesized by the calcination of ferrocenium at high temperatures ranging from 500 to 900 °C. Then the synthesized nanocomposites were characterized by XRD (X-Ray Diffraction), TEM (Transmission Electron Microscopy), VSM (Vibrating-Sample Magnetometry), BET (Brunauer-Emmett-Teller surface area measurements), TGA (Thermogravimetric Analysis), XPS (X-Ray Photoelectron Spectroscopy), EPR (Electron Paramagnetic Resonance), and CHN elemental analysis. The prepared nanocatalysts were applied for the decomposition of methylene blue as a model in wastewater treatment. It was unexpected to discover that the prepared nanocatalysts were highly active for the reaction with methylene blue in the dark even though no excess of hydrogen peroxide was added. The nanocatalyst calcined at 800 °C exhibited the rod shape with the best catalytic activity. The nanocatalysts could be reused for 12 times without the significant loss of the catalytic activity.

Isolation, purification and characterization of an ascorbate peroxidase from celery and overexpression of the AgAPX1 gene enhanced ascorbate content and drought tolerance in Arabidopsis

BackgroundCelery is a widely cultivated vegetable abundant in ascorbate (AsA), a natural plant antioxidant capable of scavenging free radicals generated by abiotic stress in plants. Ascorbate peroxidase (APX) is a plant antioxidant enzyme that is important in the synthesis of AsA and scavenging of excess hydrogen peroxide. However, the characteristics and functions of APX in celery remain unclear to date.ResultsIn this study, a gene encoding APX was cloned from celery and named AgAPX1. The transcription level of the AgAPX1 gene was significantly upregulated under drought stress. AgAPX1 was expressed in Escherichia coli BL21 (DE3) and purified. The predicted molecular mass of rAgAPX1 was 33.16 kDa, which was verified by SDS-PAGE assay. The optimum pH and temperature for rAgAPX1 were 7.0 and 55 °C, respectively. Transgenic Arabidopsis hosting the AgAPX1 gene showed elevated AsA content, antioxidant capacity and drought resistance. Less decrease in net photosynthetic rate, chlorophyll content, and relative water content contributed to the high survival rate of transgenic Arabidopsis lines after drought.ConclusionsThe characteristics of APX in celery were different from that in other species. The enhanced drought resistance of overexpressing AgAPX1 in Arabidopsis may be achieved by increasing the accumulation of AsA, enhancing the activities of various antioxidant enzymes, and promoting stomatal closure. Our work provides new evidence to understand APX and its response mechanisms to drought stress in celery.

The Comparison of Parameters of Oxidative Stress in Native Rat Livers Between Different Immunosuppressive Regimens.

BackgroundIt is thought that immunosuppressive treatment, besides anti-rejection properties, leads to pathological changes within the organ due to activation of mechanisms associated with oxidative stress. The aim of this study was to examine the parameters of oxidative stress in the livers of rats treated with the most commonly used transplant recipient drug regimens.Material/MethodsThe rat livers were obtained from archival material obtained from the previously performed experiment. Malondialdehyde (MDA), reduced glutathione (GSH) concentrations, superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) activities were analyzed.ResultsOnly the group treated with tacrolimus (T), mycophenolate mofetil (M), and prednisone (P), the TMP group, showed a slight increase in lipid peroxide concentration compared to the control group, though the difference was not statistically significant. Comparison of lipid peroxide concentration between the other treatment combinations and the control group showed a significant decrease. Additionally, a difference in lipid peroxide concentrations in the livers was observed between the cyclosporine A (C) group and tacrolimus (T) group. Alterations of other oxidative stress parameters were also observed in different regimens.ConclusionsLong-lasting immunosuppressive treatment does indeed affect redox status; however, the antioxidant defenses of the liver against the effects of excess hydrogen peroxide are efficient, so the superoxide dismutase/glutathione peroxidase (SOD/GPx) and superoxide dismutase/catalase (SOD/CAT) ratios were not significant.

Changes in Prooxidant-Antioxidant System Indices in the Blood and Brain of Rats with Modelled Acute Hypoxia which Consumed a Deuterium-Depleted Drinking Diet

Using the rat hypoxia model, the imbalanced work of antiradical defense enzymes with insufficient dismutase activity in blood leading to the development of oxidative stress, which was more pronounced in animals receiving a natural drinking diet, has been confirmed on the basis of the comparative analysis of the antiradical defense enzyme functioning index. It has been observed that acute hypoxia in the brain tissues is characterized by the development of catalase deficiency and the risk of excessive hydrogen peroxide production. It has been demonstrated that a decrease in the D/H (deuterium/protium) ratio in the blood and brain reduces the severity of the impairment of antioxidant enzyme work in hypoxia.

Lipid Composition and State of Oxidation of Fortified Infant Flours in Low‐Income Countries Are Not Optimal and Strongly Affected by the Time of Storage

AbstractFood fortification is widely used to address the public health problem of nutrient deficiencies. This study's purpose is to assess the lipid profile and nutritional quality of 13 fortified infant flours (FI) collected “in the field” in Africa and Asia after different periods of storage. The lipid content, fatty acid profiles, lipophilic vitamin content, and lipid oxidation state (peroxide values, secondary oxidation products) are determined. Mycotoxins and packaging materials are also characterized. The lipid content averages 9.1 ± 3.5 g/100 g. Fatty acid profiles are dominated by linoleic (43.3 ± 8.8%), oleic (29.5 ± 7.4%), and palmitic acid (17.8 ± 6.7%) and result in an average ω6/ω3 ratio of 12.2 ± 5.9 but with high values for some FI. Very high overages in vitamins A, D, and E are observed in products stored for short periods (1–6 months), whereas FI stored for more than 12 months has insufficient vitamin content. Lipid oxidation is acceptable but for six products presenting excessive peroxide values. Most products are contaminated by low amounts of mycotoxins but only two FI do not abide by the regulation. A strong correlation between peroxide values, hexanal content, and time of storage is observed.Practical Applications: The expiration dates for FI commercialized in low‐income countries should be shortened from 36 to 12 months so as to guarantee their nutritional quality of these functional foods and to abide by the fortified infant flour legislation. Indeed, FI quality significantly decreases over time of storage. The use of high barrier packaging materials must be generalized, although it is a necessary criterion but not sufficient to ensure the long‐term stability of FI. Special attention should be given to reduce lipophilic vitamin overages and to improve their lipid profile, especially the ω6/ω3 ratio, which has to be lower than 15.

FePt@MnO-Based Nanotheranostic Platform with Acidity-Triggered Dual-Ions Release for Enhanced MR Imaging-Guided Ferroptosis Chemodynamic Therapy.

Reactive oxygen species (ROS)-based anticancer therapy methods were heavily dependent on specific tumor microenvironments such as acidity and excess hydrogen peroxide (H2O2). In this work, an acidity-sensitive nanotheranostic agent (FePt@MnO)@DSPE-PEG5000-FA (FMDF NPs) was successfully constructed for MR imaging guided ferroptosis chemodynamic therapy (FCDT)of cancer. The FMDF NPs could specifically target folic acid (FA) receptor-positive tumor cells (HeLa etc.) and induce ferroptosis efficiently by rapidly releasing active Fe2+ to catalyze intracellular H2O2 into ROS based on Fentonreaction.On the other hand, the Mn2+ could also be released dueto acidity and further coordinate with GSH to enhance the longitudinal-transverse relaxivity (T1/T2-weighted MR imaging), which could obviously strengthen the contrast distinction between solid tumors and the surrounding tissue to accurately real-time monitor the tumor location. Furthermore, the in vivo anticancer study revealed that the growth of solid tumor models could be suppressed remarkably after treating with FMDF NPs and no obvious damage to other major organs. Therefore, the FMDF NPs were competent simultaneously as an enhanced imaging diagnosis contrast agent and efficient therapy agent for promoting more precise and effective treatment in the bionanomedicine field.

Development and validation of the enzymatic kinetic photometric procedure for determination of residual quantities of dequalinium chloride on the surface of the pharmaceutical equipment

Aim. To develop and validate the new enzymatic kinetic photometric procedure for analysis of dequalinium chloride based on application of enzymatic acetylcholine hydrolysis to determine the residual quantities of dequalinium chloride while monitoring the completeness of cleaning of the pharmaceutical equipment.Results and discussion. The optimal conditions for the enzymatic reaction course were determined – the order of mixing and the concentration of acetylcholine (0.05 mg/mL), cholinesterase (0.4 mg/mL), hydrogen peroxide (10 %) and p-phenetedine (1 %), the time of the reaction mixture maintaining (20 min), pH (8.35), the effect of the nature of the buffer solution. Validation of the procedure developed was carried out – the application range of the procedure was proposed (40–160 % in the normalized coordinates, the maximum acceptable concentration of dequalinium chloride in washes from the pharmaceutical equipment хcrit = 0.5 μg/mL), the number of concentration levels (n = 7); the order of their location within the application range, taking these data into account the maximum acceptable uncertainty of the procedure (maxΔx = 3.05 %) and the acceptability criteria for linear dependence parameters, systematic and random errors were calculated. The validation characteristics of the procedure developed were shown to correspond to the acceptability criteria calculated. The degree of extraction of dequalinium chloride from swabs with flushing from the pharmaceutical equipment was determined.Experimental part. The residual quantities of dequalinium chloride on the surface of the pharmaceutical equipment were determined by the degree of inhibition of the enzymatic reaction assessed by the unreacted residue of the substrate of the biochemical reaction, such as acetylcholine. The residual quantities of acetylcholine in the reaction mixture was determined by the kinetic photometric method by the indicator reaction of p-phenetidine oxidation with peracetic acid (formed during the auxiliary perhydrolysis reaction when adding an excess of hydrogen peroxide to the reaction mixture) in the way of registering the light absorbance of the resulting reaction product – azoxifenetole (λmax = 358 nm) for a definite period of time.Conclusions. The enzymatic kinetic photometric procedure has been developed to determine the residual quantities of dequalinium chloride on the surface of the pharmaceutical equipment after its cleaning. The procedure developed has been validated by such parameters as linearity, accuracy, precision and the limit of quantification

Phosphoethanolamine N-methyltransferase 1 contributes to maintenance of root apical meristem by affecting ROS and auxin-regulated cell differentiation in Arabidopsis.

The continuous growth of roots requires the balance between cell division and differentiation. Reactive oxygen species (ROS) and auxin are important regulators of root development by affecting cell division and differentiation. The mechanism controlling the coordination of cell division and differentiation is not well understood. Using a forward genetic screen, we isolated a mutant, defective primary root 2 (dpr2), defective in root apical meristem (RAM) maintenance. The DPR2 gene encodes phosphoethanolamine N-methyltransferase 1 (PEAMT1) that catalyzes phosphocholine biosynthesis in Arabidopsis. We characterized the primary root phenotypes of dpr2 using various marker lines, using histochemical and pharmacological analysis to probe early root development. Loss-of-function of DPR2/PEAMT1 resulted in RAM consumption by affecting root stem cell niche, division zone, elongation and differentiation zone (EDZ). PIN-FORMED (PIN) protein abundance, PIN2 polar distribution and general endocytosis were impaired in the root tip of dpr2. Excess hydrogen peroxide and auxin accumulate in the EDZ of dpr2, leading to RAM consumption by accelerating cell differentiation. Suppression of ROS over-accumulation or inhibition of auxin signalling partially prevent RAM differentiation in dpr2 after choline starvation. Taken together, we conclude that the EDZ of the root tip is most sensitive to choline shortage, leading to RAM consumption through an ROS-auxin regulation module.