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Sheath blight (ShB) is one of the most common diseases in rice that significantly affects yield production. However, the underlying mechanisms of rice defense remain largely unknown. Our previous transcriptome analysis identified that infection with Rhizoctonia solani significantly induced the expression level of SWEET2a, a member of the SWEET sugar transporter. The sweet2a genome-editing mutants were less susceptible to ShB. Further yeast-one hybrid, ChIP, and transient assays demonstrated that WRKY53 binds to the SWEET2a promoter to activate its expression. WRKY53 is a key brassinosteroid (BR) signaling transcription factor. Similar to the BR receptor gene BRI1 and biosynthetic gene D2 mutants, the WRKY53 mutant and overexpressor were less and more susceptible to ShB compared to wild-type, respectively. Inoculation with R. solani induced expression of BRI1, D2, and WRKY53, but inhibited MPK6 (a BR-signaling regulator) activity. Also, MPK6 is known to phosphorylate WRKY53 to enhance its transcription activation activity. Transient assay results indicated that co-expression of MPK6 and WRKY53 enhanced WRKY53 trans-activation activity to SWEET2a. Furthermore, expression of WRKY53 SD (the active phosphorylated forms of WRKY53) but not WRKY53 SA (the inactive phosphorylated forms of WRKY53), enhanced WRKY53-mediated activation of SWEET2a compared to expression of WRKY53 alone. Taken together, our analyses showed that R. solani infection may activate BR signaling to induce SWEET2a expression via WRKY53 through negative regulation of ShB resistance in rice.

Polynitrogen heterocycles are readily available and have recently arisen as versatile synthons for the formation of various C–C and C–X bonds, and medicinally active nitrogen-containing heterocycles. Several cascade reactions, including annulation, radical cascade, and borylation reactions, have been reported in which polynitrogen heterocycles are applied as arylation reagents. The success of these exceptional reactions illustrates the great synthetic potential of polynitrogen heterocycles, which provides a direct and useful approach to arylation reactions and the synthesis of nitrogen-containing heterocycles. The use of photocatalysts to effectively transfer energy from visible light to non-absorbing compounds has gained increasing attention as this method allows for the mild and efficient generation of radicals in a controlled manner. This approach has thus led to new methods that involve unique bond formation reactions. In addition, the use of free radical intermediates stabilized by transition metal catalysts is a powerful way to construct new chemical bonds. The aim of this review is to highlight the rapidly expanding area of radical-initiated denitrogenative cascade reactions of polynitrogen heterocycles and elaborate on their mechanisms from a new perspective by using photocatalysis and metal-based catalysis.

A novel and efficient strategy for trifluoromethylthiolation and dearomatization of activated alkynes with stable and readily available AgSCF3 has been developed. Reported herein is the unprecedented electrochemical generation of the SCF3 radical in the absence of persulfate for the synthesis of SCF3-containing spiro[5,5]trienones in good yields via a 6-exo-trig radical cyclization.

A metal-free and oxidant-free procedure for the sulfonylation of Baylis–Hillman acetates with sulfonyl hydrazides under the irradiation of visible light and open-air conditions at room temperature is described. This transformation has high yields (up to 92%), excellent stereoselectivity, broad substrate scope, good functional group tolerance, and mild reaction conditions. Moreover, mechanistic studies indicate that a sulfonyl radical pathway is involved in this reaction.

Cancer stem cells (CSCs) have an established role in cancer progression and therapeutic resistance. The p63 proteins are important transcription factors which belong to the p53 family, but their function and mechanism in CSCs remain elusive. Here, we investigated the role of TAp63α in colorectal CSCs and the effects of sulforaphane on TAp63α. We found that TAp63α was upregulated in spheres with stem cell properties compared to the parental cells. Overexpression of TAp63α promoted self-renewal capacity and enhanced CSC markers expression in colorectal sphere-forming cells. Furthermore, we showed that TAp63α directly bound to the promoter region of Lgr5 to enhance its expression and activate its downstream β-catenin pathway. Functional experiments revealed that sulforaphane suppressed the stemness of colorectal CSCs both in vitro and in vivo. Upregulation of TAp63α attenuated the inhibitory effect of sulforaphane on colorectal CSCs, indicating the role of TAp63α in sulforaphane suppression of the stemness in colorectal cancer. The present study elucidated for the first time that TAp63α promoted CSCs through targeting Lgr5/β-catenin axis and participated in sulforaphane inhibition of the stem cell properties in colorectal cancer.

The large catalytic subunit of acetohydroxyacid synthase (AHAS, EC 2.2.1.6) of Thermotoga maritima (TmcAHAS) was prepared in this study. It possesses high specific activity and excellent stability. The protein and a whole cell catalyst overexpressing the protein were applied to the preparation of α-hydroxyketones including acetoin (AC), 3-hydroxy-2-pentanone (HP), and (R)-phenylacetylcarbinol (R-PAC). The results show that AC and HP could be produced in high yields (84% and 62%, respectively), while R-PAC could be synthesized in a high yield (about 78%) with an R/S ratio of 9:1. Therefore, TmcAHAS and the whole cell catalyst overexpressing the protein could be practically useful bio-catalysts in the preparation of α-hydroxyketones including AC, HP, and R-PAC. To the best of our knowledge, this is the first time that bacterial AHAS was used as a catalyst to prepare HP with a good yield, and also the first time that TmcAHAS was employed to synthesize AC and R-PAC.

Background The purpose of this study was to investigate the safety, tolerability and pharmacokinetics of tetramethylpyrazine nitrone (TBN) in healthy Chinese volunteers. Methods A single-ascending-dose (SAD) study where 68 subjects were randomized to a single dose of placebo or TBN (50, 100, 200, 400, 700, 1,000, 1,400, or 1,800 mg) through IV infusion over 30 min. A multiple-ascending-dose (MAD) study where 24 subjects received TBN twice daily (with 12 hr interval) for total 6.5 days at doses of either 700 or 1,400 mg. Adverse events were recorded and pharmacokinetic samples were collected during the whole study period. Results No serious adverse events were found in the study. All of the observed adverse events, including increased white blood cell (4.4% subjects) and neutrophil counts (4.4% subjects), and decreased hemoglobin levels (4.2% subjects), were laboratory test abnormalities. All the adverse events were mild and tolerable, and returned to normal without any intervention. In the SAD study, linear Cmax values were observed in the dose interval of 50-1,800 mg. In the MAD study, the average steady-state concentrations (Cavg.ss ) of TBN in the 700 and 1,400 mg dose group were 2,407 and 5,837 ng/ml, respectively. No drug accumulation was observed in this study. Conclusions TBN is well tolerated in healthy volunteers. Linear Cmax values were observed in the interval of 50-1,800 mg, and target exposures of TBN were achieved without accumulation after twice daily administration to subjects. (This study has been registered at ChiCTR.org.cn. Identifier: ChiCTR1800016225 and ChiCTR1800019627.).

The co-production of xylitol and ethanol from agricultural straw has more economic advantages than the production of ethanol only. Saccharomyces cerevisiae, the most widely used ethanol-producing yeast, can be genetically engineered to ferment xylose to xylitol. In the present study, the effects of xylose-specificity, cofactor preference, and the gene copy number of xylose reductase (XR; encoding by XYL1 gene) on xylitol production of S. cerevisiae were investigated. The results showed that overexpression of XYL1 gene with a lower xylose-specificity and a higher NADPH preference favored the xylitol production. The copy number of XYL1 had a positive correlation with the XR activity but did not show a good correlation with the xylitol productivity. The overexpression of XYL1 from Candida tropicalis (CtXYL1) achieved a xylitol productivity of 0.83 g/L/h and a yield of 0.99 g/g-consumed xylose during batch fermentation with 43.5 g/L xylose and 17.0 g/L glucose. During simultaneous saccharification and fermentation (SSF) of pretreated corn stover, the strain overexpressing CtXYL1 produced 45.41 g/L xylitol and 50.19 g/L ethanol, suggesting its application potential for xylitol and ethanol co-production from straw feedstocks.

We previously demonstrated that circulating extracellular vesicles (EVs) from patients with valvular heart disease (VHD; vEVs) contain inflammatory components and inhibit endothelium-dependent vasodilation. Neutrophil chemotaxis plays a key role in renal dysfunction, and dexmedetomidine (DEX) can reduce renal dysfunction in cardiac surgery. However, the roles of vEVs in neutrophil chemotaxis and effects of DEX on vEVs are unknown. Here, we investigated the impact of vEVs on neutrophil chemotaxis in kidneys and the influence of DEX on vEVs. Circulating EVs were isolated from healthy subjects and patients with VHD. The effects of EVs on chemokine generation, forkhead box protein O3a (FOXO3a) pathway activation and neutrophil chemotaxis on cultured human umbilical vein endothelial cells (HUVECs) and kidneys in mice and the influence of DEX on EVs were detected. vEVs increased FOXO3a expression, decreased phosphorylation of Akt and FOXO3a, promoted FOXO3a nuclear translocation, and activated the FOXO3a signaling pathway in vitro. DEX pretreatment reduced vEV-induced CXCL4 and CCL5 expression and neutrophil chemotaxis in cultured HUVECs via the FOXO3a signaling pathway. vEVs were also found to suppress Akt phosphorylation and activate FOXO3a signaling to increase plasma levels of CXCL4 and CCL5 and neutrophil accumulation in kidney. The overall mechanism was inhibited in vivo with DEX pretreatment. Our data demonstrated that vEVs induced CXCL4-CCL5 to stimulate neutrophil infiltration in kidney, which can be inhibited by DEX via the FOXO3a signaling. Our findings reveal a unique mechanism involving vEVs in inducing neutrophils chemotaxis and may provide a novel basis for using DEX in reducing renal dysfunction in valvular heart surgery.

Ent-kaur-15-en-17-al-18-oic acid (LL-3) was demonstrated that it can inhibit LPS-induced nitric oxide (NO) production and macrophage migration, maintain homeostasis of oxidative stress, including increased mitochondrial membrane potential (MMP), decreased levels of reactive oxygen species (ROS) and malondialdehyde (MDA), and maintenance of superoxide dismutase (SOD) and glutathione (GSH) activities and inhibit oxidative stress-induced P38 and nuclear factor κB (NF-κB) pathways to decrease inducible nitric oxide synthase (iNOS), cyclooxygense-2 (COX-2), and tumour necrosis factor (TNF)-α mRNA expressions without marked cytotoxicity. These findings revealed that LL-3 could serve as a candidate lead compound for further studying anti-inflammatory therapies.[Formula: see text].

The aim of this work was to evaluate the general role of the vacuolar pyrophosphatase proton pump (V-PPase) in sucrose accumulation in citrus species. First, three citrus V-PPase genes, designated CsVPP-1, CsVPP-2, and CsVPP-4, were identified in the citrus genome. CsVPP-1 and CsVPP-2 belonging to citrus type I V-PPase genes are targeted to the tonoplast, and CsVPP-4 belonging to citrus type II V-PPase genes is located in the Golgi bodies. Moreover, there was a significantly positive correlation between transcript levels of type I V-PPase genes and sucrose, rather than hexose, content in fruits of seven citrus cultivars. Drought and abscisic acid treatments significantly induced the CsVPP-1 and CsVPP-2 transcript levels, as well as the sucrose content. The overexpression of type I V-PPase genes significantly increased PPase activity, decreased pyrophosphate contents, and increased sucrose contents, whereas V-PPase inhibition produced the opposite effect in both citrus fruits and leaves. Furthermore, altering the expression levels of type I V-PPase genes significantly influenced the transcript levels of sucrose transporter genes. Taken together, this study demonstrated that CsVPP-1 and CsVPP-2 play key roles in sucrose storage in the vacuole by regulating pyrophosphate homeostasis, ultimately the sucrose biosynthesis and transcript levels of sucrose transport genes, providing a novel lead for engineering or breeding modified taste in citrus and other fruits.

The growth and ethanol fermentation of Saccharomyces cerevisiae are vulnerable to weak acid that coexists with sugar in lignocellulosic hydrolysate. In this study, the molecular basis of S. cerevisiae in response to acetic and formic acids was investigated when fermented with glucose and xylose at pH 5.0. Transcriptional profiling revealed that 294 (657 in total) differentially expressed genes (DEGs) were exclusive to mixed acetic and formic acids (Aa_Fa) group, the DEGs in acetic acid (Aa) and formic acid (Fa) group were 308 and 326, respectively. The general responses to Aa, Fa, and Aa_Fa mainly involved carbohydrate metabolism, amino acid metabolism, transcriptional regulation, and stress response. In Aa_Fa group, the enrichment ratio of each pathway was much higher than being exposed to Aa or Fa stress individually, and some enrichments not existed in Aa or Fa group were observed, suggesting the synergistic anti-inhibition responses of S. cerevisiae was triggered. In yeast cells, bioconversion, stress response, and transmembrane transport could be activated to resist acetic and formic acids accompanying with energy consumption. Except the putative transcriptional factor (TF) - Haa1p, the other three TFs, Hap4p, Yox1p, and Mag1p, might as well be considered to participate in acetic and formic acids resistance responses.

The accumulation of soluble sugars in fleshy fruits largely determines their sweetness or taste. A spontaneous sweet orange mutant 'Hong Anliu' (HAL, Citrus sinensis) accumulates low soluble sugar content in fruit juice sacs than its wild type, 'Anliu' (AL) orange; however, the cause of reduced sugar content in 'HAL' fruit remains unclear. In this study, sugar content and expression profiles of genes involved in sugar metabolism and transport were compared between 'HAL' and 'AL' fruit juice sacs. In both cultivars, fructose and glucose displayed the increasing trends with significantly lower contents in 'HAL' than 'AL' after 160 DAF; moreover, sucrose had a declining trend in 'HAL' and increasing trend in 'AL' with fruit development. On the other hand, transcript levels of VINV, CWINV1, CWINV2, SUS4, SUS5, SPS1, SPS2, VPP-1, VPP-2, and some sugar transporter genes were significantly decreased in 'HAL' compared with 'AL' after 100 DAF or 160 DAF. Interestingly, the transcript levels of SPS2 and SUT2 exhibited a similar trend as it was found for sucrose content in both cultivars. These results suggested that the low sugar accumulation in 'HAL' fruit JS is accompanied by the reduced sink strength, sucrose-synthesis ability, and vacuolar storage ability compared with 'AL'; reduction of CWINVs, VINV, SPS2, SUT2, VPP-1, and VPP-2 transcript levels possibly plays a key role in the low storage of soluble sugars in the vacuoles of mutant juice sacs.

A novel and facile approach for one-pot synthesis of spinel cobalt ferrites (CoFe₂O₄) nanoparticle (NPs) and is photocatalytic activity for degradation of pharmaceuticals waste and organic dye. The synthesis involves homogeneous chemical precipitation followed by hydrothermal heating, using of hexamine as a hydroxylating agent. As-synthesized CoFe₂O₄ photocatalyst was characterised by XRD, SEM, SEM Mapping, TEM, XPS, and BET-surface area analysis. The TEM image reveals cubic shapes with an average size of 10-20 nm. The surface area of the CoFe₂O₄ NPs was found to be 16.42 m² g-1. The photocatalytic activity of CoFe₂O₄ has proved to be an excellent photocatalyt for degradation of metronidazole and organic dyes such as methylene blue (MB) and rhodamine B (RhB). The rate of the reaction was found to be 0.102, 0.198 and 0.213 min-1 for metronidazole, MB and RhB respectively. The catalyst also proved to be noteworthy as it does not loss in its catalytic activity even after five cycles of reuse.

Heparinase I (Hep I) specifically degrades heparin to oligosaccharide or unsaturated disaccharide and has been widely used in preparation of low molecular weight heparin (LMWH). In this wor...

Excessive release of some hazardous chemicals, such as Fe3+, Cu2+, Cr2O72- and nitrobenzene, may endanger public health and the environment; therefore, targeted effective sensing strategies are important. In this report, a series of lanthanide-based metal-organic frameworks (Ln-MOFs), namely {[Ln(dpc)(2H2O)]·(Hbibp)0.5}n (H4dpc = 2-(3',4'-dicarboxylphenoxy) isophthalic acid, bibp = 4,4'-bis(imidazolyl) biphenyl, for I-VI, Ln = La, Ce, Pr, Nd, Sm, and Eu) were hydrothermally synthesized and characterized. Single-crystal X-ray diffraction indicates that I-VI are isostructural and the lanthanide center is nine-coordinated with a distorted tetrakaidecahedral configuration. The as-synthesized Ln-MOFs are assembled into three-dimensional frameworks through the connections of dpc4- ligands and hydrogen bonding interactions. Notably, Eu-MOF (VI) behaves as a multi-responsive luminescent sensor toward Fe3+, Cu2+, Cr2O72- and nitrobenzene with high sensitivity, selectivity, stability and anti-interference ability against the coexistence of other ions or molecules based on high luminescence quenching efficiency. Additionally, Eu-MOF (VI) shows excellent luminescence stability and retains its structural integrity within the pH range of 2-12 in an aqueous solution and its solid sample maintains high thermodynamic stability up to 320 °C. Furthermore, the possible luminescence sensing mechanisms have been discussed in detail, and are supported by PXRD analysis, UV-vis spectroscopy, X-ray photoelectron spectroscopy (XPS) or density functional theory (DFT).

A series of d10 based metal-organic frameworks (MOFs), namely, [Cd2(dpc)(bipy)(H2O)3]n (Ⅰ), {[Cd2(dpc)(bib)(H2O)]·H2O}n (Ⅱ), {[Cd2(dpc)(bitb)1.5(H2O)2]·H2O}n (Ⅲ) (H4dpc = 2-(3′,4′-dicarboxylphenoxy)isophthalic acid, bipy = 4,4′-dipyridyl, bib = 1,4-bis (1-imidazolyl)benzene, bitb = 1,4-bis (imidazol-1-ylmethyl)benzene) have been designed and successfully synthesized using a flexible tetrabasic carboxylic acid along with three ancillary ligands under hydrothermal conditions, which exhibit structural diversities of targeted Cd-MOFs. Single-crystal X-ray diffraction analysis reveals that they contain building units of {CdN1O6} and {CdN1O5} (for I), {CdN1O5} (for II), as well as {CdN2O4} and {CdN5O1} (for Ⅲ), based on which to assemble microporous three-dimensional (3D) frameworks via covalent bonds and/or hydrogen bonds using multiple coordination modes of H4dpc ligand and auxiliary linkers. The solid-state luminescence performances of these Cd-MOFs were investigated at ambient temperature. Furthermore, luminescent sensing responses of Ⅰ-Ⅲ to amino acids and metal cations were systematically explored. Notably, Ⅰ and II may act as potential dual-functional luminescent sensors for Aspartic acid and Cu2+ cations as well as Aspartic acid and Fe3+ cations with high selectivity and sensitivity in aqueous solutions, respectively. The mechanisms of the quenching effect and sensing properties of the as-synthesized Cd-MOFs have also been discussed in this report.

Salmon calcitonin phospholipid complex loaded-solid lipid nanoparticles (sCT-PC-SLNs) were prepared from Salmon calcitonin phospholipid complex (sCT-PC) by a film dispersion method. The formulation was optimized by the Box-Behnken response surface design using quality properties such as particle size, polydispersity index (PDI), zeta potential value, entrapment efficiency (EE), and drug loading capacity (DL). The relative bioavailability of different dosage forms of salmon calcitonin (sCT) were calculated separately for plasma calcium percentage of SD female rats after subcutaneous injection and colonic administration. The results show that each dosage form offers reduced plasma calcium levels after colonic administration, except for blank loaded-solid lipid nanoparticles (SLN). The relative bioavailability (0.32 ± 0.17%) for sCT-PC-SLN is twice as high as that of aqueous sCT solution (0.16 ± 0.03%), as well as values for Salmon calcitonin loaded-solid lipid nanoparticles (sCT-SLN) (0.15 ± 0.01%); these results are statistically significant. Compared to SLN, uptake of sCT in a rat colon can be greatly improved by phospholipid complex loaded-solid lipid nanoparticles (PC-SLN). The new dosage form developed here overcomes the low EE of the water-soluble drug, enhances the EE of SLN, and reduces the degradation of gastrointestinal enzymes due to its encapsulation in lipids.

Cysteine, a critical residue for catalytic process but also vulnerable to oxidative damage, was conventionally expressed as a buried catalytic site in most redox enzymes. In the present work, specific surface-exposed sites of a NADH oxidase from Lactobacillus rhamnosus (LrNox) were selected and mutated to cysteine to investigate its effects on catalytic function because LrNox has a buried catalytic cysteine but no surface-exposed one. The results showed that exception of the sites on dimer interface, the activities of LrNox mutants were improved to vary degrees when the polar uncharged and alanine residues were mutated to cysteine. But the cysteine mutations of polar charged and nonpolar residues except alanine showed obvious decline in catalytic activity. Substituting of Ala85 and Thr96 with other residues suggested that the cysteine mutation showed the highest activity. Structural analysis suggested that even a single cysteine mutation on the specific non-conserved surface area of LrNox could induce changes on the conformation of catalytic cysteine and lower the activation free energy to improve the catalytic activity.

A heterologous xylose utilization pathway, either xylose reductase-xylitol dehydrogenase (XR-XDH) or xylose isomerase (XI), is usually introduced into Saccharomyces cerevisiae to construct a xylose-fermenting strain for lignocellulosic ethanol production. To investigate the molecular basis underlying the effect of different xylose utilization pathways on the xylose metabolism and ethanol fermentation, transcriptomes of flocculating industrial strains with the same genetic background harboring different xylose utilization pathways were studied. A different source of xylA did not obviously affect the change of the strains transcriptome, but compared with the XR-XDH strain, several key genes in the central carbon pathway were downregulated in the XI strains, suggesting a lower carbon flow to ethanol. The carbon starvation caused by lower xylose metabolism in XI strains further influenced the stress response and cell metabolism of amino acid, nucleobase, and vitamin. Besides, the downregulated genes mostly included those involved in mitotic cell cycle and the cell division-related process. Moreover, the transcriptomes analysis indicated that the after integrate xylA in the δ region, the DNA and chromosome stability and cell wall integrity of the strains were affected to some extent. The aim of this was to provide some reference for constructing efficient xylose-fermenting strains.

Withaminimas A-F (1-6), six new withaphysalin-type withanolides were isolated from the aerial parts of Physalis minima L.. The structures of these compounds were elucidated through a variety of spectroscopic techniques including HR-MS, NMR, and ECD. Compound 1 belongs to rare 18-norwithanolides, and 2-3 were 13/14-secowithanolides. According to the traditional usage of P. minima, inhibitory effects on nitric oxide (NO) production in lipopolysaccaride-activated RAW264.7 macrophages were evaluated, and compounds 1-4 exhibited significant inhibitory effects with IC50 values among 3.91-18.46 μmol·L-1.

A new approach for discrimination of polycyclic aromatic hydrocarbons (PAHs) in environment was proposed based on fluorescence coupled with CS-SVM. Two groups of experiments were carried out on PAHs with similar spectra. The first one was PAHs any two among benzo[k]fluoranthene (BkF), benzo[b]fluoranthene (BbF) and benzo[a]pyrene (BaP). The second one was naphthalene (NAP) and fluorene (FLU) with similar spectra. The BaP-BkF mixture, BaP-BbF mixture and BbF-BkF mixture with similar fluorescence properties, cuckoo search algorithm (CS) optimizing support vector machine (SVM) was used to discriminate these three mixtures. By comparison with the basic grid search algorithm (GS), genetic algorithm (GA) and particle swarm optimization algorithm (PSO) optimizing SVM, it was found that fitting degree of CS was the best and the convergence speed was also the fastest. The test sample classification accuracy of CS-SVM can reach 100%, which was higher than that of GS-SVM, GA-SVM and PSO-SVM. In order to verify the validity of the proposed approach, all the above methods were applied to discriminate NAP and FLU with extremely similar spectra. The test sample classification accuracy can reach 100%. The satisfying results indicated that the proposed approach had potential to be an alternative approach for discriminating PAHs in environment.

Graphene oxide/attapulgite (GO/APT) composite membranes were successfully fabricated by the vacuum-assisted filtration for efficient dyes wastewater treatment. By characterization of FTIR, XPS, Raman, XRD and FESEM, APT nanorods were confirmed to be incorporated into GO laminar layers via grafting modification, which would influence GO interlayer distance (d-spacing), membrane surface microstructure (laminate morphology, structure, and hydrophilicity) and even water separation performance. Comparison of those of pristine GO membrane, the calculated d-spacing of GO/APT membranes gradually increased from 0.90 nm to 1.07 nm, while water contact angles decreased from 71.0° to 43.3° with the increasing APT/GO ratios. Moreover, GO/APT membranes exhibited rough hierarchical microstructure and higher surface hydrophilicity, which was in conjunction with larger interlayer spacing to synergistically improve separation performance. The water permeated flux increased from 3.4 of pristine GO membrane to 13.3 L m−2 h−1 of GO/APT membrane with preserving high rejection nearly to 100% for 7.5 mg L−1 Rh B wastewater under optimized conditions. Similarly, membrane thickness, dye concentrations and separating species in feed solutions were also found to affect membrane separation performance. Dye molecules were efficiently rejected through GO/APT nanofiltration membranes by the synergistic separation mechanism: size exclusion effect because of the unimpeded water channels formed into 3D network laminate structure, and electrostatic interactions between the oxygen-containing functional groups on membrane surface and charged molecules. Such these GO/APT membranes demonstrated efficiently separation properties and thus provided new insight into the potential applications in water purification and dyes wastewater treatment.

Three metal-organic frameworks (MOFs), namely, [Mn(CPP)2(H2O)]n (I), [Cd(CPP)2(H2O)]n (II) and {[Zn2(CPP)2(CO3)]·(ClO4)·(H2O)2}n (III) (HCPP = 4'-(4-(4-carboxylphenoxy)-2,2':6′,2′-terpyridine), were successfully achieved under hydrothermal conditions. The as-synthesized coordination polymers have been characterized by IR spectroscopy, elemental analyses and single crystal X-ray diffraction. Single-crystal X-ray diffraction analyses indicate that compounds Ⅰ and Ⅱ display isomorphous and isostructural, in which the central metal ions present distorted octahedral configurations. Ⅰ and Ⅱ feature fascinating 3D frameworks based on π⋯π and C H⋯π interactions between the pyridine rings appertaining CPP− ligands, while compound Ⅲ possesses a 3D net interlinked via C-H⋯π interactions between the phenyl rings belonging to the CPP− ligands. In their 3D frameworks, the {MnN3O3}, {CdN3O3} and {ZnO2N3}–{ZnO2N3} building units are observed, respectively. Luminescent properties of coordination polymer Ⅱ and Ⅲ demonstrate that they may be acted as fluorescent sensors toward Cu2+ cation and Tryptophan, respectively. Furthermore, investigations on magnetic property reveal that coordination polymer Ⅰ presents antiferromagnetic coupling.

A new strategy to exfoliate inorganic layered double hydroxide (LDH) solids to micrometer-sized and positively-charged nanosheets in aqueous solution is presented. The procedure involves intercalation of zwitterionic surfactant into the decarbonated LDH by ion exchange, followed by simply stirring the surfactant-intercalated LDH in an aqueous solution of pH = 2. Since the charge of zwitterionic surfactant can be varied from anionic to cationic by adjusting the pH, the zwitterionic ions present in the interlayer were converted to cationic at pH = 2, resulting in the exfoliation of LDH by electrostatic repulsion. The delaminated LDHs nanosheets were confirmed by XRD, TEM, and AFM analyses. This simple method did not need reflux at high temperature, overcame the drawback of using organic solvents, and even resulted in a stably colloidal dispersion of nanosheets.

The first Cu-catalyzed asymmetric borylative cyclization of cyclohexadienone-containing 1,6-enynes is achieved through a tandem process: selective β-borylation of propargylic ether and subsequent conjugate addition to cyclohexadienone. The reaction proceeds with excellent regioselectivity and enantioselectivity to afford an optically pure cis-hydrobenzofuran framework bearing alkenylboronate and enone substructures. Furthermore, the resulting bicyclic products could be converted to bridged and tricyclic ring structures. This method extends the realm of Cu-catalyzed asymmetric tandem reactions using bis(pinacolato)diboron (B2pin2).

Hydroxycamptothecin (SN38) is a natural plant extract isolated from Camptotheca acuminate. It has a broad spectrum of anticancer activity through inhibition of DNA topoisomerase I, which could affect DNA synthesis and lead to DNA damage. Thus, the action of SN38 against cancers could inevitably affect endogenous levels of ribonucleotide (RNs) and deoxyribonucleotide (dRNs) that play critical roles in many biological processes, especially in DNA synthesis and repair. However, the exact impact of SN38 on RNs and dRNs is yet to be fully elucidated. In this study, we evaluated the anticancer effect and associated mechanism of SN38 in human colorectal carcinoma HCT 116 cells. As a result, SN38 could decrease the cell viability and induce DNA damage in a concentration-dependent manner. Furthermore, cell cycle arrest and intracellular nucleotide metabolism were perturbed due to DNA damage response, of which ATP, UTP, dATP, and TTP may be the critical metabolites during the whole process. Combined with the expression of deoxyribonucleoside triphosphates synthesis enzymes, our results demonstrated that the alteration and imbalance of deoxyribonucleoside triphosphates caused by SN38 was mainly due to the de novo nucleotide synthesis at 24 h, and subsequently the salvage pathways at 48 h. The unique features of SN38 suggested that it might be recommended as an effective supplementary drug with an anticancer effect.

In maize, the ear shank is a short branch that connects the ear to the stalk. The length of the ear shank mainly affects the transportation of photosynthetic products to the ear, and also influences the dehydration of the grain by adjusting the tightness of the husks. However, the molecular mechanisms of maize shank elongation have rarely been described. It has been reported that the maize ear shank length is a quantitative trait, but its genetic basis is still unclear. In this study, RNA-seq was performed to explore the transcriptional dynamics and determine the key genes involved in maize shank elongation at four different developmental stages. A total of 8145 differentially expressed genes (DEGs) were identified, including 729 transcription factors (TFs). Some important genes which participate in shank elongation were detected via function annotation and temporal expression pattern analyses, including genes related to signal transduction hormones (auxin, brassinosteroids, gibberellin, etc.), xyloglucan and xyloglucan xyloglucosyl transferase, and transcription factor families. The results provide insights into the genetic architecture of maize ear shanks and developing new varieties with ideal ear shank lengths, enabling adjustments for mechanized harvesting in the future.

Remdesivir (RDV) has generated much anticipation for its moderate effect in treating severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. However, the unsatisfactory survival rates of hospitalized patients limit its application to the treatment of coronavirus disease 2019 (COVID-19). Therefore, improvement of antiviral efficacy of RDV is urgently needed. As a typical nucleotide analog, the activation of RDV to bioactive triphosphate will affect the biosynthesis of endogenous ribonucleotides (RNs) and deoxyribonucleotides (dRNs), which are essential to RNA and DNA replication in host cells. The imbalance of RN pools will inhibit virus replication as well. In order to investigate the effects of RDV on cellular nucleotide pools and on RNA transcription and DNA replication, cellular RNs and dRNs concentrations were measured by the liquid chromatography-mass spectrometry method, and the synthesis of RNA and DNA was monitored using click chemistry. The results showed that the IC50 values for BEAS-2B cells at exposure durations of 48 and 72 h were 25.3 ± 2.6 and 9.6 ± 0.7 μM, respectively. Ten (10) μM RDV caused BEAS-2B arrest at S-phase and significant suppression of RNA and DNA synthesis after treatment for 24 h. In addition, a general increase in the abundance of nucleotides and an increase of specific nucleotides more than 2 folds were observed. However, the variation of pyrimidine ribonucleotides was relatively slight or even absent, resulting in an obvious imbalance between purine and pyrimidine ribonucleotides. Interestingly, the very marked disequilibrium between cytidine triphosphate (CTP) and cytidine monophosphate might result from the inhibition of CTP synthase. Due to nucleotides which are also precursors for the synthesis of viral nucleic acids, the perturbation of nucleotide pools would block viral RNA replication. Considering the metabolic vulnerability of endogenous nucleotides, exacerbating the imbalance of nucleotide pools imparts great promise to enhance the efficacy of RDV, which possibly has special implications for treatment of COVID-19.

Yeast culture plus enzymatically hydrolyzed yeast cell wall (YC-EHY) contains crude protein, mannan-oligosaccharide, β-glucan and yeast culture. This study was carried out to explore the effects of dietary YC-EHY at different levels on growth performance, antioxidant capacity, and immune function of broiler chickens. A total of 320 one-day-age male broiler chicks were allocated into 4 groups and were fed with a basal diet supplemented with 0 mg/kg (the control group), 50 mg/kg, 100 mg/kg, 150 mg/kg YC-EHY for 42 d. Dietary YC-EHY improved average daily gain and feed efficiency during the starter, grower, and overall periods as well as average body weight of broiler chickens on 42 d (linear and quadratic, P < 0.05). Broiler chickens fed with YC-EHY quadratically increased jejunal sucrase activity on 21 d (quadratic, P < 0.05), and linearly and quadratically enhanced maltase activity on 21 and 42 d (linear and quadratic, P < 0.05). Supplementing YC-EHY linearly and quadratically enhanced jejunal superoxide dismutase (SOD) activity on 21 and 42 d and glutathione peroxidase (GPX) activity on 42 d whereas decreased malonaldehyde (MDA) concentration on 42 d (linear and quadratic, P < 0.05). Consistently, the jejunal genes expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and SOD1 on 21 and 42 d, heme oxygenase-1 (HO-1) and GPX1 on 42 d were enhanced by YC-EHY supplementation (linear and quadratic, P < 0.05). The concentrations of jejunal immunoglobulin G (IgG) on 21 and 42 d and secreted immunoglobulin A (SIgA) on 42 d were linearly and quadratically elevated by supplementing YC-EHY (linear and quadratic, P < 0.05). Dietary YC-EHY quadratically increased jejunal IgG and IgM genes expression on 21 d (quadratic, P < 0.05), and linearly and quadratically enhanced the genes expression of IgG and IgM on 42 d (linear and quadratic, P < 0.05). Overall, this study indicated that supplementing YC-EHY could exert beneficial effects on growth performance, intestinal antioxidant capacity and immune function in broiler chickens.

Background Renewable chemicals have attracted attention due to increasing interest in environmental concerns and resource utilization. Biobased production of industrial compounds from nonfood biomass has become increasingly important as a sustainable replacement for traditional petroleum-based production processes depending on fossil resources. Therefore, we engineered an Enterobacter cloacae budC and ldhA double-deletion strain (namely, EC∆budC∆ldhA) to redirect carbon fluxes and optimized the culture conditions to co-produce succinic acid and acetoin. Results In this work, E. cloacae was metabolically engineered to enhance its combined succinic acid and acetoin production during fermentation. Strain EC∆budC∆ldhA was constructed by deleting 2,3-butanediol dehydrogenase (budC), which is involved in 2,3-butanediol production, and lactate dehydrogenase (ldhA), which is involved in lactic acid production, from the E. cloacae genome. After redirecting and fine-tuning the E. cloacae metabolic flux, succinic acid and acetoin production was enhanced, and the combined production titers of acetoin and succinic acid from glucose were 17.75 and 2.75 g L−1, respectively. Moreover, to further improve acetoin and succinic acid production, glucose and NaHCO3 modes and times of feeding were optimized during fermentation of the EC∆budC∆ldhA strain. The maximum titers of acetoin and succinic acid were 39.5 and 20.3 g L−1 at 72 h, respectively. Conclusions The engineered strain EC∆budC∆ldhA is useful for the co-production of acetoin and succinic acid and for reducing microbial fermentation costs by combining processes into a single step.

Citrate is an important intermediate product for the biosynthesis of several metabolites in plants. As two important organs of the citrus plant, fruits and leaves have their own metabolites characteristics; among them, citrate is normally high in fruit juice sacs (JS) and low in leaves. In this study, citrate content and transcript levels of citrate synthesis, transport, storage, and utilization related genes were compared between leaves and fruit JS of Citrus reticulata cv. 'Huagan No. 2', C. grandis cv. 'Hirado Buntan', and C. sinensis cv. 'Anliu'. Results indicated that the citrate content in fruit JS was significantly higher than in leaves of each cultivar. Only the relative mRNA levels of a P-type proton pump gene, CsPH8, was significantly lower in leaves than in fruit JS of three citrus cultivars, while other genes related to citrate biosynthesis, transport, storage, and utilization were highly expressed in leaves as compared to fruit JS. Furthermore, CsPH8 transient and stable transformation in leaves indicated that the change in citrate content is highly consistent with the change of CsPH8 transcript levels. Taken together, our results strongly suggest that the low accumulation of citrate in citrus leaves is mainly due to the low expression level of CsPH8; additionally, the high level of expression of citrate-utilizing genes would prevent citrate accumulation in the leaf organ.

l-Valine, l-isoleucine, and l-leucine are three key proteinogenic amino acids, and they are also the essential amino acids required for mammalian growth, possessing important and to some extent, special physiological and biological functions. Because of the branched structures in their carbon chains, they are also named as branched-chain amino acids (BCAAs). This review will highlight the advance in studies of the enzymes involved in the biosynthetic pathway of BCAAs, concentrating on their chemical mechanisms and applications in screening herbicides and antibacterial agents. The uses of some of these enzymes in lab scale organic synthesis are also discussed.

BackgroundXylitol accumulation is a major barrier for efficient ethanol production through heterologous xylose reductase-xylitol dehydrogenase (XR-XDH) pathway in recombinantSaccharomyces cerevisiae. Mutated NADH-preferring XR is usually employed to alleviate xylitol accumulation. However, it remains unclear how mutated XR affects the metabolic network for xylose metabolism. In this study, haploid and diploid strains were employed to investigate the transcriptional responses to changes in cofactor preference of XR through RNA-seq analysis during xylose fermentation.ResultsFor the haploid strains, genes involved in xylose-assimilation (XYL1,XYL2,XKS1), glycolysis, and alcohol fermentation had higher transcript levels in response to mutated XR, which was consistent with the improved xylose consumption rate and ethanol yield. For the diploid strains, genes related to protein biosynthesis were upregulated while genes involved in glyoxylate shunt were downregulated in response to mutated XR, which might contribute to the improved yields of biomass and ethanol. When comparing the diploids with the haploids, genes involved in glycolysis and MAPK signaling pathway were significantly downregulated, while oxidative stress related transcription factors (TFs) were significantly upregulated, irrespective of the cofactor preference of XR.ConclusionsOur results not only revealed the differences in transcriptional responses of the diploid and haploid strains to mutated XR, but also provided underlying basis for better understanding the differences in xylose metabolism between the diploid and haploid strains.

A novel glucose-Zn-based porous coordination polymer (PCP) was selected as a carrier of cell membranes (CMs) to fabricate CM-coated PCP (CMPCP) for rapid screening of potentially active compounds from natural products. The cell disruption and the amount of maximum CMs adsorbed on PCP were optimized according to the amount of immobilized protein. This new kind of matrix exhibited good reproducibility and stability, and was applied for fishing potentially active compounds from the extracts of Vaccinium corymbosum L. leaves (VCL). Using LC-MS/MS, chlorogenic acid and quercetin were identified as the potentially active compounds through comparison of normal and non-alcoholic fatty liver disease (NAFLD)-modeled CMPCP. Our results suggested that the proposed approach based on CMPCP was environmentally friendly, cost-effective, and convenient in terms of green porous material, stable protein loading capacity, and accessible operation process. The developed method could provide a promising platform for efficient drug discovery from natural product resources.Graphical abstract.

Engineered Saccharomyces cerevisiae can reduce xylose to xylitol. However, in S.cerevisiae, there are several endogenous enzymes including xylitol dehydrogenase encoded by XYL2, sorbitol dehydrogenases encoded by SOR1/SOR2 and xylulokinase encoded by XKS1 may lead to the assimilation of xylitol. In this study, to increase xylitol accumulation, these genes were separately deleted through CRISPR/Cas9 system. Their effects on xylitol yield of an industrial S. cerevisiae CK17 overexpressing Candida tropicalis XYL1 (encoding xylose reductase) were investigated. Deletion of SOR1/SOR2 or XKS1 increased the xylitol yield in both batch and fed-batch fermentation with different concentrations of glucose and xylose. The analysis of the transcription level of key genes in the mutants during fed-batch fermentation suggests that SOR1/SOR2 are more crucially responsible for xylitol oxidation than XYL2 under the genetic background of S.cerevisiae CK17. The deletion of XKS1 gene could also weaken SOR1/SOR2 expression, thereby increasing the xylitol accumulation. The XKS1-deleted strain CK17ΔXKS1 produced 46.17 g/L of xylitol and reached a xylitol yield of 0.92 g/g during simultaneous saccharification and fermentation (SSF) of pretreated corn stover slurry. Therefore, the deletion of XKS1 gene provides a promising strategy to meet the industrial demands for xylitol production from lignocellulosic biomass.

Sorbitol is the primary substrate translocated from source to sink in pear species. Among the many sorbitol transporters (SOTs), some are known to be involved in sorbitol accumulation in fruit; however, their particular roles are unclear. In this study, we examined the transcriptome and metabolome of a variety of pear samples from six time points to identify those SOTs. Similar to previous studies, sorbitol and sucrose differed significantly between the leaf and fruit, and sorbitol was consistently observed at higher concentrations at all time points. Interestingly, we found that sorbitol accumulation in pear fruit was cooperatively mediated by SOT3, SOT6/20, SOT19/21, and SOT22. In particular, the up-regulated SOT6/20 and SOT19/21 in fruit under 1 mg L-1 abscisic acid and 10 mg L-1 indole acetic acid treatments, respectively, resulted in an increased sorbitol concentration. In addition, sorbitol concentration showed positive correlations to fructose and glucose concentrations, indicating a role for sorbitol in the determination of fruit sweetness. Together with the deduced process of sugar biosynthesis, transport, conversion, and accumulation in pear, our study provides a foundation for further research into sugar accumulation processes in pear fruit, contributing to the improvement of fruit quality.

Endogenous ribonucleotides (RNs) and deoxyribonucleotides (dRNs) are important metabolites related with the pathogenesis of many diseases. In light of their physiological and pathological significances, a novel and sensitive pre-column derivatization method with N-(t-butyldimethylsilyl)-N-methyltrifluoroacetamide (MTBSTFA) was developed to determine RNs and dRNs in human cells using high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS). A one-step extraction of cells with 85 % methanol followed by a simple derivatization reaction within 5 min at room temperature contributed to shortened analysis time. The derivatives of 22 nucleoside mono-, di- and triphosphates were retained on the typical C18 column and eluted by ammonium acetate and acetonitrile in 9 min. Under these optimal conditions, good linearity was achieved in the tested calibration ranges. The lower limit of quantitation (LLOQ) was determined to be 0.1-0.4 μM for the tested RNs and 0.001-0.1 μM for dRNs. In addition, the precision (CV) was

A novel, rapid and sensitive chemiluminescence (CL) method for the determination of oxytetracycline hydrochloride (OTCH) is described in this paper. The presented method was based on the fact that OTCH could immensely enhance the CL of the reaction of cerium sulfate and tris(2,2-bipyridyl) ruthenium (II) in acidic medium. Under optimal experimental conditions, CL intensity was favorably linear for OTCH in the range 5.0 × 10-7 to 5.0 × 10-5 g/ml, with a detection limit of 1.5 × 10-7 g/ml (S/N = 3). The relative standard detection was 4.76% for 5.0 × 10-6 g/ml (n = 11). This method was successfully applied to the analysis of OTCH in milk and egg white samples. According to the results of the kinetic curves for OTCH in the Ru(bipy)32+ -Ce(SO4 )2 CL system, together with CL and ultraviolet (UV)-visible spectra, the possible mechanism of the CL reaction is discussed briefly.

Six new prenylated xanthones (1–6) and seventeen known xanthones were isolated from extracts of Garcinia bracteata leaves. Their structures were determined by extensive NMR and MS spectroscopic data analysis. The inhibitory activities of the isolates were assayed on HeLa, A549, PC-3, HT-29, and WPMY-1 cell lines. Compounds 1 and 15–17 showed moderate inhibitory effects on tumor cell growth, with IC50s ranging from 3.7 to 14.7 µM.

A monocationic ZnII tetrahedral cluster, [Zn4L3(μ3-OH)]·NO3·1.25H2O (Zn4L3) based on a new salicylamide imine ligand H2L, H2L = 1-(2-hydroxy-3-methoxy-benzamido)-3-(2-hydroxy-3-methoxy-benzylideneamino)-propane, has been prepared. Single crystal X-ray analysis reveals that three deprotonated ligands L2– chelate three Zn(II) centres with their salicylamide moiety in such a way that the three salicylimine groups attached to the other side of the three chelates are converged to bind another Zn(II) centre to provide a tetrahedral cluster with two phenyl groups of the same ligand L2– being close enough to present a strong intramolecular π⋯π stacking effect. Fluorescence studies indicate that Zn4L3 is stable in water and exhibits highly sensitive and selective recognition of phosphates against other common anions including CO32–, HCO3–, NO3–, F−, Cl−, Br−, I−, HSO4–, SO42–, OAc−, BF4–, ClO4– and CF3SO3– in HEPES buffer solution (pH = 7.4) + DMSO (V : V = 1 : 9). The excellent sensing capability of Zn4L3 for phosphate against other common anions with a low detection limit of 0.15 μM renders it a candidate probe for phosphate detection. Furthermore, the observed fluorescence quenching responses of Zn4L3 towards phosphates were highly reversible. The possible sensing mechanisms for phosphate detection by Zn4L3 was investigated by means of 1H NMR, UV-Vis spectra and high-resolution ESI-MS spectra and the results indicate that phosphates could exclusively decompose Zn4L3 to release H2L in HEPES buffer solution (pH = 7.4) + DMSO (V : V = 1 : 9).

A set of two cyclic [2]catenane dimers were self-assembled in aqueous media via selective hydrazone condensation, by means of either one-pot or post-functionalization procedures. Considering that each catenane is composed of two thirty-six member rings, each of these two cyclic [2]catenane dimers can be considered as a ultra-large macrocycle that contains more than fifty atoms in the ring framework. This cyclization methodology relying on catenation thus represents a novel approach for accomplishing ultramacrocyclization in pure water.

Sorbitol is a primary substrate that is translocated from its source to its sink in pear and apple by sorbitol transporters (SOTs). However, little is known about the expression profiles of SOTs in tree tissues, and their association with sorbitol accumulation. In this study, 64 SOT genes were isolated from the sequenced genomes of six rosaceae trees, which clustered phylogenetically into four groups. Groups I and III contained genes from pome trees, group II those from drupe trees, and group IV those from pome, drupe, and berry trees. A gene duplication analyses showed that the 24 SOT genes of pear arose from tandem, proximal, and dispersed duplications, which differs from their evolution in the other rosaceae fruit trees. A qRT-PCR analysis showed that the PbSOT6/20 gene in group IV is most strongly expressed in the leaf, branch, flower, fruit, and seed of pear. The expression pattern of PbSOT6/20 correlates more strongly with the pattern of sorbitol accumulation in the pear fruits than do those of other SOT genes. A low concentration (100 mg/g) of exogenous sorbitol induced the expression of PbSOT6/20 in the pear fruits, but not in the leaves. These results suggest that PbSOT6/20 is associated with sorbitol accumulation in pear fruits and plays important role in sorbitol translocation from its source to its sink. This study provides new insights into sorbitol translocation mediated by the SOT genes in pear.

BACKGROUND Coronavirus disease 2019 (COVID-19) caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) outbreak in China, constitutes a Public Health Emergency of International Concern. It is well known that COVID-19 patients may have increased serum lactate dehydrogenase (LDH) levels in the early stage. The clinical changes in LDH may have predictive value in disease evolution and prognosis in critically ill COVID-19 patients. AIM To examine serum LDH and clinical characteristics in patients with COVID-19 and their predictive value for prognosis. METHODS This retrospective study analyzed the clinical data of forty-seven critical COVID-19 patients in the intensive care unit of the Third People's Hospital of Yichang City from January 27 to March 25, 2020 and divided them into survivors and non-survivors. The patients were diagnosed according to the World Health Organization interim guidance and critical cases met any one of the following criteria: Respiratory failure and required mechanical ventilation, the occurrence of shock, and the combined failure of other organs that required intensive care unit monitoring and treatments, according to the diagnostic criteria of critical COVID-19. Clinical data including symptoms, detection of SARS-CoV-2, chest computed tomography (CT) images, changes in serum LDH in different clinical phases, and prognosis were collected. Statistical analysis of the data was performed. Continuous variables were expressed as median (interquartile range) and compared with the Mann-Whitney U test. Categorical variables were compared with the Chi-square test. Survival data were analyzed using Kaplan-Meier survival curves and log-rank tests. RESULTS According to chest CT images, we observed the alveolitis and fibrosis stages in all critical patients in this study. Most non-survivors died in the fibrosis stage. Non-survivors had fewer days of hospitalization, shorter disease duration, shorter duration of alveolitis and fibrosis, and had dyspnea symptoms at disease onset (P = 0.05). Both first and lowest LDH values in the alveolitis stage were more pronounced in non-survivors than in survivors (449.0 U/L vs 288.0 U/L, P = 0.0243; 445.0 U/L vs 288.0 U/L, P = 0.0199, respectively), while the first, lowest and highest values of serum LDH in non-survivors were all significantly increased compared to survivors in the fibrosis phase (449.0 U/L vs 225.5 U/L, P = 0.0028; 432.0 U/L vs 191.0 U/L, P = 0.0007; 1303.0 U/L vs 263.5 U/L, P = 0.0001, respectively). The cut-off points of first LDH values in the alveolitis and fibrosis phase for distinction of non-survivors from survivors were 397.0 U/L and 263.0 U/L, respectively. In the fibrosis stage, non-survivors had more days with high LDH than survivors (7.0 d vs 0.0 d, P = 0.0002). Importantly, patients with high LDH had a significantly shorter median survival time than patients with low LDH in the alveolitis phase (22.0 d vs 36.5 d, P = 0.0002), while patients with high LDH also had a significantly shorter median survival time than patients with low LDH in the fibrosis phase (27.5 d vs 40.0 d, P = 0.0008). The proportion of non-survivors with detectable SARS-CoV-2 until death in the alveolitis stage was significantly increased compared with that in the fibrosis stage (100% vs 35.7%, P = 0.0220). CONCLUSION High LDH and dyspnea symptoms were positive predictors of an adverse outcome in critical COVID-19. The rapid progressive fibrosis stage was more perilous than the alveolitis stage, even if SARS-CoV-2 is undetectable.