Integrated Metabolomics and Transcriptomics Analyses of the Biosynthesis of Arbutin and 6'-O-Caffeoylarbutin in Vaccinium dunalianum Cell Suspension Cultures Fed with Hydroquinone.

Arbutin and 6'-O-caffeoylarbutin (CA) from Vaccinium dunalianum Wight are known for their ability to inhibit melanin synthesis. To boost the production of arbutin and CA, precursor feeding with hydroquinone (HQ) was studied in V. dunalianum suspension cells. The effect of HQ on the biosynthesis of arbutin and CA in the suspension cells was investigated using high-performance liquid chromatography (HPLC), and possible molecular mechanisms were analyzed using metabolomics and transcriptomics analyses. HPLC analysis only showed that the addition of HQ significantly enhanced arbutin synthesis in cells, peaking at 15.52 ± 0.28 mg·g-1 after 0.5 mmol·L-1 HQ treatment for 12 h. Subsequently, metabolomics identified 78 differential expression metabolites (DEMs), of which arbutin and CA were significantly up-regulated metabolites. Moreover, transcriptomics found a total of 10,628 differential expression genes (DEGs). The integrated transcriptomics and metabolomics revealed that HQ significantly enhanced the expression of two arbutin synthase (AS) genes (Unigene0063512 and Unigene0063513), boosting arbutin synthesis. Additionally, it is speculated that CA was generated from arbutin and 3,4,5-tricaffeoylquinic acid catalyzed by caffeoyl transferase, with Unigene0044545, Unigene0043539, and Unigene0017356 as potentially associated genes with CA synthesis. These findings indicate that the precursor feeding strategy offers a promising approach for the mass production of arbutin and CA in V. dunalianum suspension cells and provides new insights for CA biosynthesis in V. dunalianum.

[Molecular modification of cyclodextrin glucosyltransferase and its application in the synthesis of α-arbutin].

α-arbutin has important applications in cosmetics and medicine. However, the extraction yield from plant tissues is relatively low, which restricts its application value. In this study, we investigated the synthesis of α-arbutin using maltodextrin as the donor and hydroquinone as the acceptor, using a cyclodextrin glucosyltransferase (CGTase) from Anaerobranca gottschalkii. We performed site-saturated and site-directed mutagenesis on AgCGTase. The activity of the variant AgCGTase-F235G-N166H was 3.48 times higher than that of the wild type. Moreover, we achieved a conversion rate of 63% by optimizing the reaction pH, temperature, and hydroquinone addition amount. Overall, this study successfully constructed a strain with improved conversion rate for the synthetic production of α-arbutin and hydroquinone. These findings have significant implications for reducing the industrial production cost of α-arbutin and enhancing the conversion rate of the product.

Tert-Butylhydroquinone retards longan fruit deterioration by regulating membrane lipid and energy metabolisms.

Longan fruit deteriorates rapidly after harvest, which limits its storability. This study aimed to investigate the effect of tert-butylhydroquinone (TBHQ) on quality maintenance, membrane lipid metabolism, and energy status of longan fruit during 25 °C storage. Compared with control fruit, TBHQ treatment maintained better marketable fruit rate and suppressed activities of phospholipase D (PLD), lipase, and lipoxygenase (LOX), and downregulated expressions of DlPLD, DlLOX, and Dllipase. TBHQ also increased the ratio of unsaturated fatty acids to saturated fatty acids (U/S) and the index of unsaturated fatty acids (IUFA). In addition, higher levels of ATP, ADP, energy charge, NADP+/ NADPH as well as higher activities of H+-ATPase, Ca2+-ATPase and NADK were also observed in TBHQ-treated fruit. These results suggested that TBHQ may maintain postharvest quality of longan fruit by regulating membrane lipid and energy metabolisms.

From Ground-State to Excited-State Activation Modes: Flavin-Dependent "Ene"-Reductases Catalyzed Non-natural Radical Reactions.

Enzymes are desired catalysts for chemical synthesis, because they can be engineered to provide unparalleled levels of efficiency and selectivity. Yet, despite the astonishing array of reactions catalyzed by natural enzymes, many reactivity patterns found in small molecule catalysts have no counterpart in the living world. With a detailed understanding of the mechanisms utilized by small molecule catalysts, we can identify existing enzymes with the potential to catalyze reactions that are currently unknown in nature. Over the past eight years, our group has demonstrated that flavin-dependent "ene"-reductases (EREDs) can catalyze various radical-mediated reactions with unparalleled levels of selectivity, solving long-standing challenges in asymmetric synthesis.This Account presents our development of EREDs as general catalysts for asymmetric radical reactions. While we have developed multiple mechanisms for generating radicals within protein active sites, this account will focus on examples where flavin mononucleotide hydroquinone (FMNhq) serves as an electron transfer radical initiator. While our initial mechanistic hypotheses were rooted in electron-transfer-based radical initiation mechanisms commonly used by synthetic organic chemists, we ultimately uncovered emergent mechanisms of radical initiation that are unique to the protein active site. We will begin by covering intramolecular reactions and discussing how the protein activates the substrate for reduction by altering the redox-potential of alkyl halides and templating the charge transfer complex between the substrate and flavin-cofactor. Protein engineering has been used to modify the fundamental photophysics of these reactions, highlighting the opportunity to tune these systems further by using directed evolution. This section highlights the range of coupling partners and radical termination mechanisms available to intramolecular reactions.The next section will focus on intermolecular reactions and the role of enzyme-templated ternary charge transfer complexes among the cofactor, alkyl halide, and coupling partner in gating electron transfer to ensure that it only occurs when both substrates are bound within the protein active site. We will highlight the synthetic applications available to this activation mode, including olefin hydroalkylation, carbohydroxylation, arene functionalization, and nitronate alkylation. This section also discusses how the protein can favor mechanistic steps that are elusive in solution for the asymmetric reductive coupling of alkyl halides and nitroalkanes. We are aware of several recent EREDs-catalyzed photoenzymatic transformations from other groups. We will discuss results from these papers in the context of understanding the nuances of radical initiation with various substrates.These biocatalytic asymmetric radical reactions often complement the state-of-the-art small-molecule-catalyzed reactions, making EREDs a valuable addition to a chemist's synthetic toolbox. Moreover, the underlying principles studied with these systems are potentially operative with other cofactor-dependent proteins, opening the door to different types of enzyme-catalyzed radical reactions. We anticipate that this Account will serve as a guide and inspire broad interest in repurposing existing enzymes to access new transformations.

Characterization of a novel sucrose phosphorylase from Paenibacillus elgii and its use in biosynthesis of α-arbutin.

α-Arbutin, a naturally occurring glycosylated derivative of hydroquinone (HQ), effectively inhibits melanin biosynthesis in epidermal cells. It is widely recognized as a fourth-generation whitening agent within the cosmetic industry. Currently, enzymatic catalysis is universally deemed the safest and most efficient method for α-arbutin synthesis. Sucrose phosphorylase (SPase), one of the most frequently employed glycosyltransferases, has been extensively reported for α-arbutin synthesis. In this study, a previously reported SPase known for its effectiveness in synthesizing α-arbutin, was used as a probe sequence to identify a novel SPase from Paenibacillus elgii (PeSP) in the protein database. The sequence similarity between PeSP and the probe was 39.71%, indicating a degree of novelty. Subsequently, the gene encoding PeSP was coexpressed with the molecular chaperone pG-Tf2 in Escherichia coli, significantly improving PeSP's solubility. Following this, PeSP was characterized and employed for α-arbutin biosynthesis. The specific activity of co-expressed PeSP reached 169.72 U/mg, exhibited optimal activity at 35℃ and pH 7.0, with a half-life of 3.6 h under the condition of 35℃. PeSP demonstrated excellent stability at pH 6.5-8.5 and sensitivity to high concentrations of metal ions. The kinetic parameters Km and kcat/Km were determined to be 14.50 mM and 9.79 min- 1·mM- 1, respectively.The reaction conditions for α-arbutin biosynthesis using recombinant PeSP were optimized, resulting in a maximum α-arbutin concentration of 52.60 g/L and a HQ conversion rate of 60.9%. The optimal conditions were achieved at 30℃ and pH 7.0 with 200 U/mL of PeSP, and by combining sucrose and hydroquinone at a molar ratio of 5:1 for a duration of 25 h.

Characterization of the inner membrane cytochrome ImcH from Geobacter reveals its importance for extracellular electron transfer and energy conservation.

Electroactive bacteria combine the oxidation of carbon substrates with an extracellular electron transfer (EET) process that discharges electrons to an electron acceptor outside the cell. This process involves electron transfer through consecutive redox proteins that efficiently connect the inner membrane to the cell exterior. In this study, we isolated and characterized the quinone-interacting membrane cytochrome c ImcH from Geobacter sulfurreducens, which is involved in the EET process to high redox potential acceptors. Spectroscopic and electrochemical studies show that ImcH hemes have low midpoint redox potentials, ranging from -150 to -358 mV, and connect the oxidation of the quinol-pool to EET, transferring electrons to the highly abundant periplasmic cytochrome PpcA with higher affinity than to its homologues. Despite the larger number of hemes and transmembrane helices, the ImcH structural model has similarities with the NapC/NirT/NrfH superfamily, namely the presence of a quinone-binding site on the P-side of the membrane. In addition, the first heme, likely involved on the quinol oxidation, has apparently an unusual His/Gln coordination. Our work suggests that ImcH is electroneutral and transfers electrons and protons to the same side of the membrane, contributing to the maintenance of a proton motive force and playing a central role in recycling the menaquinone pool.

Boric acid group-functional ruthenium complex as a novel fluorescence probe for robust detection of propyl gallate and tert-butyl hydroquinone by tuning the pH.

This study reported a ruthenium complex-based fluorescence probe, achieving rapid and sequential detection of propyl gallate (PG) and tert-butyl hydroquinone (TBHQ) for the first time by tuning pH only. Under 480 nm excitation, probe exhibited intensive emission at 620 nm, which was selectively quenched by PG at pH 7.0 due to the covalent binding between the boric acid of probe and o-diphenol hydroxyl of PG. Then pH was tuned to 7.4, the emission was significantly quenched by TBHQ because of the π-π stacking between aromatic rings of probe and paraquinone of TBHQ. This probe realized specific and sensitive detection of PG and TBHQ with wide range and low detection limit (0.26 µM for PG and 0.66 µM for TBHQ). Furthermore, a portable visual test paper detection platform was built based on this probe for rapid and sensitive detection of antioxidants in food, which was of great significance for market regulation.

Exploring ND-011992, a quinazoline-type inhibitor targeting quinone reductases and quinol oxidases.

Bacterial energy metabolism has become a promising target for next-generation tuberculosis chemotherapy. One strategy to hamper ATP production is to inhibit the respiratory oxidases. The respiratory chain of Mycobacterium tuberculosis comprises a cytochrome bcc:aa3 and a cytochrome bd ubiquinol oxidase that require a combined approach to block their activity. A quinazoline-type compound called ND-011992 has previously been reported to ineffectively inhibit bd oxidases, but to act bactericidal in combination with inhibitors of cytochrome bcc:aa3 oxidase. Due to the structural similarity of ND-011992 to quinazoline-type inhibitors of respiratory complex I, we suspected that this compound is also capable of blocking other respiratory chain complexes. Here, we synthesized ND-011992 and a bromine derivative to study their effect on the respiratory chain complexes of Escherichia coli. And indeed, ND-011992 was found to inhibit respiratory complex I and bo3 oxidase in addition to bd-I and bd-II oxidases. The IC50 values are all in the low micromolar range, with inhibition of complex I providing the lowest value with an IC50 of 0.12 µM. Thus, ND-011992 acts on both, quinone reductases and quinol oxidases and could be very well suited to regulate the activity of the entire respiratory chain.

Cardiolipin-mediated temporal response to hydroquinone toxicity in human retinal pigmented epithelial cell line.

Hydroquinone, a potent toxic agent of cigarette smoke, damages retinal pigmented epithelial cells by triggering oxidative stress and mitochondrial dysfunction, two events causally related to the development and progression of retinal diseases. The inner mitochondrial membrane is enriched in cardiolipin, a phospholipid susceptible of oxidative modifications which determine cell-fate decision. Using ARPE-19 cell line as a model of retinal pigmented epithelium, we analyzed the potential involvement of cardiolipin in hydroquinone toxicity. Hydroquinone exposure caused an early concentration-dependent increase in mitochondrial reactive oxygen species, decrease in mitochondrial membrane potential, and rise in the rate of oxygen consumption not accompanied by changes in ATP levels. Despite mitochondrial impairment, cell viability was preserved. Hydroquinone induced cardiolipin translocation to the outer mitochondrial membrane, and an increase in the colocalization of the autophagosome adapter protein LC3 with mitochondria, indicating the induction of protective mitophagy. A prolonged hydroquinone treatment induced pyroptotic cell death by cardiolipin-mediated caspase-1 and gasdermin-D activation. Cardiolipin-specific antioxidants counteracted hydroquinone effects pointing out that cardiolipin can act as a mitochondrial "eat-me signal" or as a pyroptotic cell death trigger. Our results indicate that cardiolipin may act as a timer for the mitophagy to pyroptosis switch and propose cardiolipin-targeting compounds as promising approaches for the treatment of oxidative stress-related retinal diseases.

Preparations of Terminal Oxidase Cytochrome bd-II Isolated from Escherichia coli Reveal Significant Hydrogen Peroxide Scavenging Activity.

Cytochrome bd-II is one of the three terminal quinol oxidases of the aerobic respiratory chain of Escherichia coli. Preparations of the detergent-solubilized untagged bd-II oxidase isolated from the bacterium were shown to scavenge hydrogen peroxide (H2O2) with high rate producing molecular oxygen (O2). Addition of H2O2 to the same buffer that does not contain enzyme or contains thermally denatured cytochrome bd-II does not lead to any O2 production. The latter observation rules out involvement of adventitious transition metals bound to the protein. The H2O2-induced O2 production is not susceptible to inhibition by N-ethylmaleimide (the sulfhydryl binding compound), antimycin A (the compound that binds specifically to a quinol binding site), and CO (diatomic gas that binds specifically to the reduced heme d). However, O2 formation is inhibited by cyanide (IC50 = 4.5 ± 0.5 µM) and azide. Addition of H2O2 in the presence of dithiothreitol and ubiquinone-1 does not inactivate cytochrome bd-II and apparently does not affect the O2 reductase activity of the enzyme. The ability of cytochrome bd-II to detoxify H2O2 could play a role in bacterial physiology by conferring resistance to the peroxide-mediated stress.

Tert-butylhydroquinone protects the retina from oxidative stress in STZ-induced diabetic rats via the PI3K/Akt/eNOS pathway.

This study aims to investigate whether tert-butylhydroquinone protects the retina from oxidative stress in STZ-induced experimental diabetic rats through the activation of phosphinositide 3-kinase (PI3K)/Akt/endothelial nitric oxide synthase (eNOS) pathway.In vitro, NO, reactive oxygen species(ROS), eNOS, p-eNOS Ser1179, Akt, p-Akt Ser473 and L-NAME protein expression was analyzed within rMC-1 cells cultivated within normal control(NC), high glucose (HG) and HG-containing tert-butyl hydroquinone (tBHQ) (5 µM) medium. We confirmed tBHQ's protection through administering inhibitors of PI3K and Akt. In vivo, tBHQ was administered at a ratio of 1% (w/w) to diabetic rats was induced through an STZ injection (65 mg/kg) for a 3-month period, and the retinal expression of eNOS, p-eNOS Ser1179, Akt, and p-Akt Ser473 proteins was measured using Western blotting (WB) assay. We also utilized the TUNEL kit for detecting retinal cell apoptosis. The changes of retinal morphology and visual function were measured by performing hematoxylin-eosin staining (HE staining) and electroretinograms. In vitro, ROS levels were increased in the high glucose group, NO levels were decreased, and the relative expression of Akt/p-Akt Ser473 and eNOs/p-eNOS Ser1179 was reduced. tBHQ abolished these changes, and these effects were suppressed by specific inhibitors. In vivo, tBHQ upregulated retinal protein expression in STZ-induced diabetic rats, reduced retinal apoptotic cell numbers, and partially prevented abnormalities in retinal function and structure caused by diabetes. tBHQ alleviates oxidative stress during diabetic retinopathy by upregulating the PI3K/Akt/eNOS pathway and partially restoring the structure and function of the retina. It may play a role in delaying vision loss caused by diabetic retinopathy.

Efficient degradation of hydroquinone by a metabolically engineered Pseudarthrobacter sulfonivorans strain.

Pseudarthrobacter sulfonivorans strain Ar51 can degrade crude oil and multi-substituted benzene compounds efficiently at low temperatures. However, it cannot degrade hydroquinone, which is a key intermediate in the degradation of several other compounds of environmental importance, such as 4-nitrophenol, g-hexachlorocyclohexane, 4-hydroxyacetophenone and 4-aminophenol. Here we co-expressed the two subunits of hydroquinone dioxygenase from Sphingomonas sp. strain TTNP3 with different promoters in the strain Ar51. The strain with 2 hdnO promoters exhibited the strongest hydroquinone catabolic activity. However, in the absence of antibiotic selection this ability to degrade hydroquinone was lost due to plasmid instability. Consequently, we constructed a hisD knockout strain, which was unable to synthesise histidine. By introducing the hisD gene onto the plasmid, the ability to degrade hydroquinone in the absence of antibiotic selection was stabilised. In addition, to make the strain more stable for industrial applications, we knocked out the recA gene and integrated the hydroquinone dioxygenase genes at this chromosomal locus. This strain exhibited the strongest activity in catabolizing hydroquinone, up to 470 mg/L in 16 h without antibiotic selection. In addition, this activity was shown to be stable when the strain has cultured in medium without antibiotic selection after 20 passages.

Evidence for Quinol Oxidation Activity of ImoA, a Novel NapC/NirT Family Protein from the Neutrophilic Fe(II)-Oxidizing Bacterium Sideroxydans lithotrophicus ES-1.

Sideroxydans species are important chemolithoautotrophic Fe(II)-oxidizing bacteria in freshwater environments and play a role in biogeochemical cycling of multiple elements. Due to difficulties in laboratory cultivation and genetic intractability, the electron transport proteins required for the growth and survival of this organism remain understudied. In Sideroxydans lithotrophicus ES-1, it is proposed that the Mto pathway transfers electrons from extracellular Fe(II) oxidation across the periplasm to an inner membrane NapC/NirT family protein encoded by Slit_2495 to reduce the quinone pool. Based on sequence similarity, Slit_2495 has been putatively called CymA, a NapC/NirT family protein which in Shewanella oneidensis MR-1 oxidizes the quinol pool during anaerobic respiration of a wide range of substrates. However, our phylogenetic analysis using the alignment of different NapC/NirT family proteins shows that Slit_2495 clusters closer to NirT sequences than to CymA. We propose the name ImoA (inner membrane oxidoreductase) for Slit_2495. Our data demonstrate that ImoA can oxidize quinol pools in the inner membrane and is able to functionally replace CymA in S. oneidensis. The ability of ImoA to oxidize quinol in vivo as opposed to its proposed function of reducing quinone raises questions about the directionality and/or reversibility of electron flow through the Mto pathway in S. lithotrophicus. IMPORTANCE Fe(II)-oxidizing bacteria play an important role in biogeochemical cycles. At circumneutral pH, these organisms perform extracellular electron transfer, taking up electrons from Fe(II) outside the cell, potentially through a porin-cytochrome complex in the outer membrane encoded by the Mto pathway. Electrons from Fe(II) oxidation would then be transported to the quinone pool in the inner membrane via periplasmic and inner membrane electron transfer proteins. Directly demonstrating the functionality of genes in neutrophilic iron oxidizers is challenging due to the absence of robust genetic methods. Here, we heterologously expressed a NapC/NirT family tetraheme cytochrome ImoA, encoded by Slit_2495, an inner membrane protein from the Gram-negative Fe(II)-oxidizing bacterium Sideroxydans lithotrophicus ES-1, proposed to be involved in extracellular electron transfer to reduce the quinone pool. ImoA functionally replaced the inner membrane c-type cytochrome CymA in the Fe(III)-reducing bacterium Shewanella oneidensis. We suggest that ImoA may function primarily to oxidize quinol in S. lithotrophicus.

Mechanism of enhanced production of triterpenoids in algal-fungal consortium.

Chlorella pyrenoidosa-Ganoderma lucidum symbiotic systems were constructed. The mechanism of enhanced production of triterpenoids in algal-fungal consortium by comparing the contents of triterpenoids in individual fungal systems and algal-fungal consortium systems was investigated. The production of triterpenoids in C. pyrenoidosa-G. lucidum consortium increased significantly (P < 0.05). The categories and relative abundances of metabolites in the individual systems and algal-fungal systems were measured and analyzed by metabonomic tests. There were 57 significant different metabolites (VIP > 1 and P < 0.05) including 12 downregulated metabolites and 45 upregulated metabolites were obtained. The significant enriched metabolic pathways (VIP > 1 and P < 0.05) of citrate cycle (TCA cycle), tyrosine metabolism, glycolysis, and terpenoid backbone biosynthesis in algal-fungal consortium were obtained. The relative abundances of important precursors of triterpenoids including mevalonic acid, lanosterol, and hydroquinone were 1.4 times, 1.7 times, and 2 times, respectively, in algal-fungal consortium than that in the individual fungal systems. The presence of C. pyrenoidosa in algal-fungal consortium promoted the biosynthesis of triterpenoids in G. lucidum.

A non-canonical vitamin K cycle is a potent ferroptosis suppressor.

Ferroptosis, a non-apoptotic form of cell death marked by iron-dependent lipid peroxidation1, has a key role in organ injury, degenerative disease and vulnerability of therapy-resistant cancers2. Although substantial progress has been made in understanding the molecular processes relevant to ferroptosis, additional cell-extrinsic and cell-intrinsic processes that determine cell sensitivity toward ferroptosis remain unknown. Here we show that the fully reduced forms of vitamin K-a group of naphthoquinones that includes menaquinone and phylloquinone3-confer a strong anti-ferroptotic function, in addition to the conventional function linked to blood clotting by acting as a cofactor for γ-glutamyl carboxylase. Ferroptosis suppressor protein 1 (FSP1), a NAD(P)H-ubiquinone reductase and the second mainstay of ferroptosis control after glutathione peroxidase-44,5, was found to efficiently reduce vitamin K to its hydroquinone, a potent radical-trapping antioxidant and inhibitor of (phospho)lipid peroxidation. The FSP1-mediated reduction of vitamin K was also responsible for the antidotal effect of vitamin K against warfarin poisoning. It follows that FSP1 is the enzyme mediating warfarin-resistant vitamin K reduction in the canonical vitamin K cycle6. The FSP1-dependent non-canonical vitamin K cycle can act to protect cells against detrimental lipid peroxidation and ferroptosis.

DNA Damage and Oxidative Stress of Tobacco Smoke Condensate in Human Bladder Epithelial Cells.

Smoking is a major risk factor for bladder cancer (BC), with up to 50% of BC cases being attributed to smoking. There are 70 known carcinogens in tobacco smoke; however, the principal chemicals responsible for BC remain uncertain. The aromatic amines 4-aminobiphenyl (4-ABP) and 2-naphthylamine (2-NA) are implicated in BC pathogenesis of smokers on the basis of the elevated BC risk in factory workers exposed to these chemicals. However, 4-ABP and 2-NA only occur at several nanograms per cigarette and may be insufficient to induce BC. In contrast, other genotoxicants, including acrolein, occur at 1000-fold or higher levels in tobacco smoke. There is limited data on the toxicological effects of tobacco smoke in human bladder cells. We have assessed the cytotoxicity, oxidative stress, and DNA damage of tobacco smoke condensate (TSC) in human RT4 bladder cells. TSC was fractionated by liquid-liquid extraction into an acid-neutral fraction (NF), containing polycyclic aromatic hydrocarbons (PAHs), nitro-PAHs, phenols, and aldehydes, and a basic fraction (BF) containing aromatic amines, heterocyclic aromatic amines, and N-nitroso compounds. The TSC and NF induced a time- and concentration-dependent cytotoxicity associated with oxidative stress, lipid peroxide formation, glutathione (GSH) depletion, and apurinic/apyrimidinic (AP) site formation, while the BF showed weak effects. LC/MS-based metabolomic approaches showed that TSC and NF altered GSH biosynthesis pathways and induced more than 40 GSH conjugates. GSH conjugates of several hydroquinones were among the most abundant conjugates. RT4 cell treatment with synthetic hydroquinones and cresol mixtures at levels present in tobacco smoke accounted for most of the TSC-induced cytotoxicity and the AP sites formed. GSH conjugates of acrolein, methyl vinyl ketone, and crotonaldehyde levels also increased owing to TSC-induced oxidative stress. Thus, TSC is a potent toxicant and DNA-damaging agent, inducing deleterious effects in human bladder cells at concentrations of <1% of a cigarette in cell culture media.

Hydroquinone predisposes for retinal pigment epithelial (RPE) cell degeneration in inflammatory conditions.

In addition to hypoxia, inflammation is capable of inducing vascular endothelial growth factor (VEGF) expression in human retinal pigment epithelial (RPE) cells. Excessive levels of VEGF promote choroidal neovascularization and thereby contribute to the pathogenesis of wet age-related macular degeneration (AMD). Intravitreal anti-VEGF injections ameliorate pathological vessel neoformation in wet AMD but excessive dampening of VEGF can result in a degeneration of the RPE. In the present study, we induced VEGF production by exposing human ARPE-19 cells to the pro-inflammatory IL-1α and subsequently to hydroquinone, a component of tobacco smoke that is a major environmental risk factor for AMD. Effects were monitored by measuring the levels of VEGF and anti-angiogenic pigment epithelium-derived factor (PEDF) using an enzyme-linked immunosorbent assay (ELISA) technique. In addition, we measured the production of reactive oxygen species (ROS) using the 2',7'-dichlorofluorescin diacetate (H2DCFDA) probe and studied the effects of two anti-oxidants, ammonium pyrrolidinedithiocarbamate (APDC) and N-acetyl-cysteine (NAC), on VEGF production. Cellular and secreted VEGF as well as secreted PEDF levels were reduced at all tested hydroquinone concentrations (10, 50, or 200 µM); these effects were evident prior to any reduction of cell viability evoked by hydroquinone. Cell viability was carefully explored in our previous study and verified by microscoping in the present study. APDC further reduced the VEGF levels, whereas NAC increased them. The 50 µM concentration of hydroquinone increased ROS production in ARPE-19 cells primed with IL-1α. Hydroquinone disturbs the regulatory balance of VEGF and PEDF in inflammatory conditions. These data support the idea that hydroquinone mediates RPE degeneration by reducing VEGF levels and may predispose to dry AMD since VEGF is as well important for retinal integrity.

Electrochemically Driven Photosynthetic Electron Transport in Cyanobacteria Lacking Photosystem II.

Light-activated photosystem II (PSII) carries out the critical step of splitting water in photosynthesis. However, PSII is susceptible to light-induced damage. Here, results are presented from a novel microbial electro-photosynthetic system (MEPS) that uses redox mediators in conjunction with an electrode to drive electron transport in live Synechocystis (ΔpsbB) cells lacking PSII. MEPS-generated, light-dependent current increased with light intensity up to 2050 µmol photons m-2 s-1, which yielded a delivery rate of 113 µmol electrons h-1 mg-chl-1 and an average current density of 150 A m-2 s-1 mg-chl-1. P700+ re-reduction kinetics demonstrated that initial rates exceeded wildtype PSII-driven electron delivery. The electron delivery occurs ahead of the cytochrome b6f complex to enable both NADPH and ATP production. This work demonstrates an electrochemical system that can drive photosynthetic electron transport, provides a platform for photosynthetic foundational studies, and has the potential for improving photosynthetic performance at high light intensities.

Three-Dimensional Hierarchical HRP-MIL-100(Fe)@TiO2@Fe3O4 Janus Magnetic Micromotor as a Smart Active Platform for Detection and Degradation of Hydroquinone.

A novel multifunctional Janus magnetic micromotor was designed and constructed by using MIL-100(Fe)@TiO2@Fe3O4 multicore-shells modified with horseradish peroxidase (HRP) as a smart active platform to realize detection and degradation of hydroquinone (HQ). The obtained micromotor showed a unique three-dimensional (3D) hierarchical architecture with highly exposed active sites and could autonomously move at a speed of 140 ± 7.0 µm·s-1 by O2 bubbles generated from the catalytic decomposition of H2O2 fuel. Benefiting from the combination of active self-propulsive motion, high peroxidase-like activity, tuned heterojunctions with matching band structures, and a 3D hierarchical structure, an effective platform involving dynamically sensitive detection and quick removal of HQ from water was established by using the multifunctional HRP-integrated MIL-100(Fe)@TiO2@Fe3O4 Janus micromotor. The proposed multifunctional Janus magnetic micromotor had advantages of simple and feasible fabrication, sensitive detection and effective photo-Fenton degradation of HQ in a wide pH range of 4-7, and magnetic recycling, revealing potential for environmental remediation applications.

Enhancement of laccase production from a newly isolated Trichoderma harzianum S7113 using submerged fermentation: Optimization of production medium via central composite design and its application for hydroquinone degradation.

Trichoderma harzianum S7113 as an efficient fungal isolate for laccase production was identified using the 18S rRNA sequencing. T. harzianum S7113 attained its maximal laccase production level on the 14th day of static incubation at 28 °C and pH 5.0 using the inoculum size of 5 discs (14 mm), according to the one factor per time (OFT) method. The most appropriate carbon, organic and inorganic nitrogen sources to promote maximal laccase synthesis were glucose (15 g/L), beef extract (5 g/L), and ammonium chloride (4 g/L), respectively. Results of Response Surface Methodology (RSM) revealed that glucose, meat extract, and ammonium chloride concentrations of 17.54, 7.17, and 4.36 g/L respectively, at a pH value of 6.74 are the favorite conditions for high titer production. The ANOVA analysis highlighted an excellent match between the actual experimental results and the model predicted laccase production levels. The biodegradation of hydroquinone (HQ) by T. harzianum S7113 laccase was most efficient in the pH range of 5.0 to 6.5. The increase in laccase concentration led to a significant increase in the HQ conversion to get a biodegradation rate of 92 ± 2.6% with a laccase concentration of 0.75 U/mL after 3 h of reaction.