A theoretical study of the electron paramagnetic resonance spectra and local environment in copper(II) complexes with different imidazole and chlorido ligands.

Copper(II) chloride anionic coordination complexes with different imidazole-derived ligands due to the potential cytotoxic activity play the important role in protein. By investigating the experimental electron paramagnetic resonance (EPR) and ultraviolet-visible (UV-vis) spectra of [CuCl(C6H10N2)4]Cl, [CuCl(C6H10N2)4]Cl, [CuCl2(C4H6N2)4], and [Cu2Cl2(C5H8N2)6]Cl2·2H2O, the local structure of the corresponding Cu2+ centers and the role of different ligands are obtained. Based on the well-agreed EPR parameters and the d-d transitions (10Dq), the four Cu2+ centers show tetragonal and orthorhombic distortion, corresponding to the different anisotropies of EPR signals. In addition, the general rules of governing the impact of methanol in imidazolylalkyl derivatives are also discussed, especially the influence on the local environment (symmetry, distortion, covalency, and crystal field) of above four copper(II) chloride anionic coordination complexes. Therefore, the obtained results in this study will be beneficial to provide a theoretical basis for the experimental design of desired copper-containing imidazolyl alkyl derivatives.

First report of Rhizopus stolonifer causing flower rot of yellow passion fruit (Passiflora edulis f. flavicarpa Deg.) in China.

The genus Passiflora, commonly known as passion fruit, originated in South America, is an economically important horticulture crop and widely distributed in the tropics and subtropics. Yellow passion fruit (Passiflora edulis f. flavicarpa) and purple passion fruit (Passiflora edulis f. edulis) are the two most planted species (Santos-Jiménez et al., 2022), which have been largely cultivated in southern China. The average annual production reaches 600,000 tons, of which yellow fruit accounts for more than 70% (Zhou et al., 2022). In 2022 to 2023, a disease caused flower rot severely in passion fruit plantations. The incidence rate was generally 10% in purple passion fruit, with an incidence up to 60% in yellow passion fruit 'Qinmi No. 9'. Flower rot occurs mainly in the rainy season, especially during periods of prolonged rain. Infected flowers had black patches that were water-soaked on the interior of the flower bud. The patches covered the entire flower bud, and fluffy mycelium and sporangia developed, which caused the flower bud rotten and abscised easily. Five symptomatic flowers from Wuhua, Guangdong (23°23'N, 115°18'E) and 8 symptomatic flowers from Shangsi, Guangxi (21°15'N, 107°98'E) of 'Qinmi No. 9' were collected during flowering period in 2022 and 2023. Diseased flower pieces were surface-sterilized with 70% ethanol for 2 to 3 min, rinsed with sterile distilled water 3 times, and placed on PDA medium at 25℃ in darkness. Four and 6 fungal isolates with similar morphology were isolated from the infected samples of Wuhua and Shangsi, respectively. Two isolates, PRFJ01 from Wuhua and PRGX02 from Shangsi, were randomly selected for further study. Purified fungal colonies at the age of 3 days accompany with diffuse cottony mycelia, turned white to gray later. The mycelia were hyaline and aseptate. Sporangiophores with 0.56 (0.22~1.10) mm in length and 6.1 (3.18~10.87) µm in width (n=100) were erect, light brown, and had rhizoids and stolons at their bases. Sporangia with 48.0 (23.45~92.85) µm in diameter (n=100) were dark-colored, near spherical and having dark ovoid sporangiospores with 3.56 (2.34~6.39) µm × 2.82 (1.73~4.70) µm (n=100). The morphology of the fungus were identical to Rhizopus stolonifer (Ehrenb.) Vuill (Haque et al. 2023). The two isolates were molecularly identified using genomic regions of 28S large ribosomal subunit (LSU) with NL1 and LR3 primers (Cruz-Lachica et al., 2018). The phylogenetic trees revealed the sequences of PRFJ01 (OR801560.1) and PRGX02 (OR801561.1) were 100% and 99% identical to R. stolonifer (MK705761.1 and KC412868.1), respectively. Pathogenicity tests were conducted on healthy flowers and leaves of 5-month-old grafted 'Qinmi No. 9' plants. Mycelial plugs with 5-mm diameter were placed on the flowers and leaves. Three plants were performed for each of the isolates, and the test was repeated twice. The inoculated plants were moisturized with plastic bags. Healthy flowers and leaves inoculated with sterile PDA plugs were used as control. Typical symptoms were observed on inoculated plants after 2 days. The dark grey mycelia and sporangia covered the entire flower after 4 days inoculation. The flower bud became putrid and the flower stalk split off. Lesions on leaves expanded accompany with numerous aerial mycelium. However, the controls were symptomless. R. stolonifer was reisolated from inoculated tissues. Previously, flower rot on passion fruit caused by R. stolonifer has only been recorded in Brazil (Ploetz, 2003). To our knowledge, this is the first report of R. stolonifer causing flower rot on passion fruit in China.

Deciphering the tumour immune microenvironment of hepatocellular carcinoma.

Current treatments for hepatocellular carcinoma (HCC) are less effective and prone to recurrence after surgery, so it's needed to seek new ideas for its therapy. Tumour immune microenvironment (TME) is crucial for the pathogenesis, development and metastasis of HCC. Interactions between immune cells and tumour cells significantly impact responses to immunotherapies and patient prognosis. In recent years, immunotherapies for HCC have shown promising potential, but the response rate is still unsatisfactory. Understanding their cross-talks is helpful for selecting potential therapeutic targets, predicting immunotherapy responses, determining immunotherapy efficacy, identifying prognostic markers and selecting individualized treatment options. In this paper, we reviewed the research advances on the roles of immune cells and multi-omic research associated with HCC pathogenesis and therapy, and future perspectives on TME.

DEL-1, as an anti-neutrophil transepithelial migration molecule, inhibits airway neutrophilic inflammation in asthma.

BACKGROUND: Neutrophil migration into the airways is a key process in neutrophilic asthma. Developmental endothelial locus-1 (DEL-1), an extracellular matrix protein, is a neutrophil adhesion inhibitor that attenuates neutrophilic inflammation. METHODS: Levels of DEL-1 were measured in exhaled breath condensate (EBC) and serum in asthma patients by ELISA. DEL-1 modulation of neutrophil adhesion and transepithelial migration was examined in a co-culture model in vitro. The effects of DEL-1-adenoviral vector-mediated overexpression on ovalbumin/lipopolysaccharide (OVA/LPS)-induced neutrophilic asthma were studied in mice in vivo. RESULTS: DEL-1 was primarily expressed in human bronchial epithelial cells and was decreased in asthma patients. Serum DEL-1 concentrations were reduced in patients with severe asthma compared with normal subjects (567.1 ± 75.3 vs. 276.8 ± 29.36 pg/mL, p < .001) and were negatively correlated to blood neutrophils (r = -0.2881, p = .0384) and neutrophil-to-lymphocyte ratio (NLR) (r = -0.5469, p < .0001). DEL-1 concentrations in the EBC of severe asthmatic patients (113.2 ± 8.09 pg/mL) were also lower than normal subjects (193.0 ± 7.61 pg/mL, p < .001) and were positively correlated with the asthma control test (ACT) score (r = 0.3678, p = .0035) and negatively related to EBC IL-17 (r = -0.3756, p = .0131), myeloperoxidase (MPO) (r = -0.5967, p = .0055), and neutrophil elastase (NE) (r = -0.5488, p = .0009) expression in asthma patients. Neutrophil adhesion and transepithelial migration in asthma patients were associated with LFA-1 binding to ICAM-1 and inhibited by DEL-1. DEL-1 mRNA and protein expression in human bronchial epithelial cells were regulated by IL-17. Exogenous DEL-1 inhibited IL-17-enhanced neutrophil adhesion and migration. DEL-1 expression was decreased while neutrophil infiltration was increased in the airway of a murine model of neutrophilic asthma. This was prevented by DEL-1 overexpression. CONCLUSIONS: DEL-1 down-regulation leads to increased neutrophil migration across bronchial epithelial cells and is associated with neutrophilic airway inflammation in asthma.

The Molecular Biology of Prostate Cancer Stem Cells: From the Past to the Future.

Prostate cancer (PCa) continues to rank as the second leading cause of cancer-related mortality in western countries, despite the golden treatment using androgen deprivation therapy (ADT) or anti-androgen therapy. With decades of research, scientists have gradually realized that the existence of prostate cancer stem cells (PCSCs) successfully explains tumor recurrence, metastasis and therapeutic failure of PCa. Theoretically, eradication of this small population may improve the efficacy of current therapeutic approaches and prolong PCa survival. However, several characteristics of PCSCs make their diminishment extremely challenging: inherent resistance to anti-androgen and chemotherapy treatment, over-activation of the survival pathway, adaptation to tumor micro-environments, escape from immune attack and being easier to metastasize. For this end, a better understanding of PCSC biology at the molecular level will definitely inspire us to develop PCSC targeted approaches. In this review, we comprehensively summarize signaling pathways responsible for homeostatic regulation of PCSCs and discuss how to eliminate these fractional cells in clinical practice. Overall, this study deeply pinpoints PCSC biology at the molecular level and provides us some research perspectives.

First report of leaf spot on passion fruit in China, caused by Alternaria alternata.

Passion fruit (Passiflora edulis Sims), which is native to South America, is an important fruit crop in tropical and subtropical countries. Passion fruit growing areas have increased rapidly in southern China. In 2018 to 2019, circular spots on passion fruit were observed in Shangsi, Guangxi, China (21°15'N, 107°98'E). The disease occurred from June to April of the following year. The disease incidence was generally between 10% to 30%, but could reach up to 50% in purple passion fruit 'Tainong No.1'. The initial lesions on the fruits were small, with a brown center and a greasy margin, and then became sunken and lighter brown with a diameter of about 1 cm in later stages. The spots on the leaves were often surrounded by a yellow halo and turned into larger lesions after coalescence.. Five typical symptomatic fruit and leaves were collected from Shangsi county for the presumed pathogen isolation. Section of the samples were surface sterilized to isolate the fungus on potato dextrose agar (PDA) at 28°C. Five fungal isolates with similar morphology on PDA were obtained by single spore isolation. Colonies at the age of 7 days accompany with flourishing aerial hyphae, showed surface color varying from white to grey. Conidia were ovate or elliptic, light brown to brown, with 2 to 5 diaphragms, 0 to 4 longitudinal-oblique diaphragms, and mostly 8.2 to 36.7 µm × 5.4 to 15.8 µm. The morphology of the fungus resembled Alternaria alternata (Fr.) Keissl (Simmons, 2007). Each of the five isolates (SF-001, SF-002, SF-003, SF-004 and SF-005) was molecularly identified using genomic regions of 18S nrDNA (SSU), 28S nrDNA (LSU), RNA polymerase second largest subunit (RPB2), internal transcribed spacer (ITS), glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and translation elongation factor 1-alpha (TEF1) (Jayawardena et al., 2019). Nucleotide sequences of SSU (MZ275254, ON055696, ON055697, ON055698 and ON055699), LSU (MZ275253, ON062947, ON062948, ON062949, ON062950), RPB2 (MZ275251, ON055377, ON055378, ON055379 and ON055380), ITS (MW866522, MW866523, ON053451, ON053452 and ON053453), GAPDH (MZ286628, ON055381, ON055382, ON055383 and ON055384) and TEF1 (MZ275255, ON055373, ON055374, ON055375 and ON055376) were deposited in GenBank database. The LSU, GAPDH and TEF1 sequences showed 100% identity with A. alternata in NCBI (KX609773, MK683852 and MK637432, respectively). The SSU, RPB2 and ITS sequences showed 99% identity to A. alternata (U05194, MK605898 and MN856409, respectively). In pathogenicity test (Zhang et al., 2020), 3-month-old grafted 'Tainong No.1' seedlings and mature fruit were used. Five-mm-diameter mycelial plugs taken from 7-day-old PDA colonies of each of 5 isolates were placed on the leaves and fruit that were wounded with a sterilized needle to form 3 pinpricks. Sterile PDA plugs were used as control. Three plants and three fruits were used in each treatment, and the test was repeated twice. The inoculated plants and fruit were kept in plastic bags and grown in a chamber at 28℃. Typical lesions were observed on inoculated plants and fruit after 3 days, but the controls remained healthy. A. alternata was consistently reisolated from these typical lesions. Previously, leaf spot on passion fruit caused by A. alternata has only been recorded in New Zealand (Rheinländer, 2010). To our knowledge, this is the first report of A. alternata (Fr.) Keissl. causing leaf spot on passion fruit in China. The identification of the pathogen may help to take effective management strategies of controlling this disease.

Epithelial expression and role of secreted STC1 on asthma airway hyperresponsiveness through calcium channel modulation.

BACKGROUND: Asthma is characterized by airway hyperresponsiveness (AHR), inflammation, and airway remodeling. Airway hyperresponsiveness results from enhanced airway smooth muscle (ASM) contraction potentially under the control of an epithelium-derived relaxing factor (EpDRF). However, relatively rare is known about EpDRF. We aimed to elucidate the role of epithelium-derived stanniocalcin-1 (STC1) on AHR and ASM contraction. METHODS: Stanniocalcin-1 levels in the serum of asthmatic patients and healthy volunteers and in bronchoalveolar lavage fluid (BALF) from ovalbumin (OVA)-challenged mice were measured by ELISA. The effects of exogenous STC1 on AHR and on inflammation were examined in mice. IL-13 modulation of STC1 mRNA and protein levels was studied in human bronchial epithelial cell lines (16HBE). The function of STC1 on Ca2+ influx and ASM contraction was examined ex vivo. RESULTS: Serum STC1 was decreased in asthma (n = 93) compared with healthy volunteers (1071 ± 30.4 vs 1414 ± 75.1 pg/ml, p < 0.0001, n = 23) and correlated with asthma control (p = 0.0270), lung function (FEV1, p = 0.0130), and serum IL-13 levels (p = 0.0009). Treatment of ten asthmatic subjects with inhaled corticosteroids/long-acting beta2-agonists (ICS/LABA) for 1 year enhanced STC1 expression which correlated with improved asthma control (p = 0.022). STC1 was mainly expressed in bronchial epithelium and intranasal administration of recombinant human STC1 (rhSTC1) reduced AHR and inflammation in mice. IL-13 suppressed STC1 release from 16HBE, whereas rhSTC1 blocked store-operated Ca2+ entry (SOCE) by suppressing stromal interaction molecule 1 (STIM1) and further inhibited ASM cell contractility by suppressing Ca2+ -dependent myosin light chain (MLC) phosphorylation. CONCLUSION: Our data indicate that STC1 deficiency in asthmatic airways promotes STIM1 hyperactivity, enhanced ASM contraction, and AHR. STC1 may be a candidate EpDRF.

Identification and functional study of immortalized mouse thymic epithelial cells.

As the key cells in a three-dimensional scaffold within the thymus, Thymic epithelial cells (TECs) play critical roles in the homing, migration and differentiation of T cell precursors through adhesive interactions and the release of various cytokines. In this study, primary cultures of mouse TECs were isolated and identified with TEC-specific antibodies CK5 and CK8. These TECs were immortalized by retroviral transduction of simian virus (SV) 40 large T antigen. We then compared the functions of TECs and immortalized TECs (iTECs). Cell morphology and the proliferative capacity of TECs and iTECs were observed by inverted microscope photography and crystal violet assay after passage. A soft agar assay was then performed to observe their clone formation ability. The expression levels of epithelial cell related factors, such as IL-7, Lptin, Pax-9, Sema3A and et al., were detected by IF and qPCR. TECs were co-cultured with human acute monocytic leukemia cells (THP-1), and the effect of TECs on promoting THP-1 proliferation was observed with flow cytometry and CFSE labeling. Senescence-associated ß-galactosidase assay was measured to detect the anti-aging capabilities of the cells. Cell cycle distribution was analyzed by propidium iodide (PI) staining, and paclitaxel (PTX)-induced apoptosis was detected by Annexin V-PI staining to evaluate the anti-apoptotic ability of the cells. Throughout, we found that the immortalized TECs still retain the characteristics of primary TECs, such as the morphology, function and epithelial characteristics; however, iTECs have stronger capabilities in proliferation and anti-aging. Our research suggests that the iTECs were successfully immortalized by SV40 large T antigen, and that the biological characteristics and functions of iTECs were similar to the original TECs. This immortalized cell can be used as an efficient cell model in functional research of the thymus substituting primary TECs with iTECs.

Ezrin, a Membrane Cytoskeleton Cross-Linker Protein, as a Marker of Epithelial Damage in Asthma.

RATIONALE: Bronchial epithelial cell damage occurs in patients with bronchial asthma. Ezrin, a membrane-cytoskeleton protein, maintains cellular morphology and intercellular adhesion and protects the barrier function of epithelial cells. OBJECTIVES: To study the role of ezrin in bronchial epithelial cells injury and correlate its expression with asthma severity. METHODS: Levels of ezrin were measured in exhaled breath condensate (EBC) and serum in patients with asthma and BAL fluid (BALF) from a mouse model of asthma by ELISA. The regulation of IL-13 on ezrin protein levels was studied in primary bronchial epithelial cells. Ezrin knockdown using shRNA was studied in human bronchial epithelial 16HBE cells. MEASUREMENTS AND MAIN RESULTS: Ezrin levels were decreased in asthmatic EBC (92.7 ± 34.99 vs. 150.5 ± 10.22 pg/ml, P < 0.0001) and serum (700.7 ± 55.59 vs. 279.2 ± 25.83 pg/ml, P < 0.0001) compared with normal subjects. Levels were much lower in uncontrolled (P < 0.001) and partly controlled patients (P < 0.01) compared with well-controlled subjects. EBC and serum ezrin levels correlated with lung function in patients with asthma and serum ezrin levels were negatively correlated with serum IL-13 and periostin. IL-13-induced downregulation of ezrin expression in primary bronchial epithelial cells was significantly attenuated by the Janus tyrosine kinase 2 inhibitor, TG101348. Ezrin knockdown changed 16HBE cell morphology, enlarged intercellular spaces, and increased their permeability. Ezrin expression was decreased in the lung tissue and BALF of "asthmatic" mice and negatively correlated with BALF IL-13 level. CONCLUSIONS: Ezrin downregulation is associated with IL-13-induced epithelial damage and might be a potential biomarker of asthma control.

Determination of the levels of tryptophan and 12 metabolites in milk by liquid chromatography-tandem mass spectrometry with the QuEChERS method.

Tryptophan and metabolites have important biological functions in humans. Milk is an important source of tryptophan intake. In this study, we developed a method to detect levels of tryptophan and 12 metabolites in milk. The analytes were extracted by using the QuEChERS (quick, easy, cheap, effective, rugged, and safe) procedure and analyzed by liquid chromatography-tandem mass spectrometry with electrospray ionization. The proposed method resulted in suitable accuracy (standard deviation ≤10.31%) and high sensitivity (the limits of quantification were between 0.05 and 5 ng/mL). Recoveries were in the range of 44 to 126%. Finally, the developed method was successfully applied to compare the content of tryptophan and metabolites in 4 milk products produced by different processes: pasteurized milk, UHT milk, milk powder, and yogurt. The results of partial least squares-discriminant analysis (PLS-DA) showed that different types of processed milk could be distinguished clearly according to the method used here. The determined tryptophan and metabolites levels in milk can provide a new reference for evaluation of milk.

Cost-effective imprinting to minimize consumption of template in room-temperature ionic liquid for fast purification of chlorogenic acid from the extract of E. ulmoides leaves.

One of the main challenges in large-scale applications of molecularly imprinted polymers (MIPs) is the significant amount of template needed in polymer preparation. A new strategy based on room-temperature ionic liquids (RTILs) was suggested to solve this problem by reducing the amount of template in the polymerization recipe. The MIP was synthesized with a mixture of dimethyl sulfoxide and RTIL (1-butyl-3-methylimidazolium tetrafluoroborate) as porogen, in which chlorogenic acid (CGA) was used as template, 4-vinylpyridine (4-VP) as functional monomer, and ethylene glycol dimethacrylate (EDMA) as cross-linker. The influence of polymerization variables, including CGA concentrations, and the ratio of 4-VP to EDMA on imprinting effect were investigated comprehensively. Moreover, the properties involving the column permeability, the number of binding sites, and the polymer morphology of the CGA-MIP monoliths were studied thoroughly. The MIP monolith had an excellent column permeability (1.53 × 10-13 m2) and allowed an ultra-fast on-line SPE, which dramatically shortens the separation time (< 10 min) and improves the separation efficiency. At high flow velocity (5.0 mL min-1), 50 µL of the extract from Eucommia ulmoides leaves can be loaded directly on the CGA-MIP monoliths and CGA with high purity can be obtained with a recovery of 89.01 ± 0.05%. As a conclusion, the resulting RTIL-induced approach of preparing MIP may be an effective tool in fabricating MIP in a low-cost way. Graphical abstract ᅟ.

Epimedin C Protects H2O2-Induced Peroxidation Injury by Enhancing the Function of Endothelial Progenitor HUVEC Populations.

Endothelial cell injury and apoptosis induced by oxidative stress serve important roles in many vascular diseases. The repair of endothelial cell vascular injury relies on the function of local endothelial progenitor cells (EPCs). Our previous study indicated that epimedin C, a major flavonoid derived from Herba epimedii (yin yang huo), could promote vascularization by inducing endothelial-like differentiation of mesenchymal stem cells C3H/10T1/2 both in vivo and in vitro. In view of the significant cardiovascular protective effects of Herba epimedii, we detected a protective effect of epimedin C on hydrogen peroxide (H2O2)-induced peroxidation injury in human umbilical vein endothelial cells (HUVECs) and the role of EPC in this process. The results show that epimedin C increased the expression of the stem cell marker, CD34 and PROM1, and subsequently enhanced the expression and function of vascular endothelial growth factor and matrix metalloproteinase (MMP)-2 in local vascular endothelial cells. In conclusion, epimedin C protects H2O2-induced peroxidation injury by enhancing the function of endothelial progenitor HUVEC populations.

The combined use of 1 H and 2D NMR-based metabolomics and chemometrics for non-targeted screening of biomarkers and identification of reconstituted milk.

BACKGROUND: The illegal undeclared addition of reconstituted milk powder to ultra-heat treated (UHT) milk to lower production costs is an example of economically motivated adulteration. This activity not only defrauds consumers but also places honest traders at a disadvantage, which could damage the reputation of milk producers and reduce the integrity of the markets. In this research, a non-targeted analytical strategy that combines proton (1 H) nuclear magnetic resonance (NMR) spectroscopy with a chemometrics data mining tool was developed for the authentication of bovine UHT milk. RESULTS: Unsupervised principal component analysis was used to distinguish UHT and tap-water-reconstituted powdered milk. Partial least squares-discriminant analysis (PLS-DA) with R2 (Y) and Q2 equal to 0.859 and 0.748, respectively, was used to differentiate UHT and reconstituted milk samples. Three compounds were selected as biomarkers to distinguish UHT and reconstituted milk and identified according to the standard NMR-spectra database. Finally, a PLS-DA model was established, according to the characteristic spectral bands, to identify UHT milk and reconstituted milk. CONCLUSION: This procedure demonstrated the feasibility of using non-targeted NMR profiling combined with chemometric analysis to combat mislabeling and fraudulent practices in milk production. © 2019 Society of Chemical Industry.

Gracilibacillus eburneus sp.nov., a moderately halophilic bacterium isolated from Xinjiang province, China.

A novel Gram-staining positive, moderately halophilic, endospore-forming, motile, rod-shaped and strictly aerobic strain, designated YIM 93565T, was isolated from a salt lake in Xinjiang province of China and subjected to a polyphasic taxonomic study. Strain YIM 93565T grew in the range of pH 6.0-9.0 (optimum pH 7.0), 10-45 °C (optimum 35-40 °C) and at salinities of 2-24% (w/v) NaCl (optimum 7-10%). Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain YIM 93565T clustered with members of the genera Gracilibacillus and form a clade with Gracilibacillus bigeumensis KCTC 13130T (95.6% similarity) and Gracilibacillus halophilus DSM 17856T (94.9%), which was well separated from others. The DNA G + C content of this novel strain was 36.8 mol%. The major fatty acids were anteiso-C15:0, iso-C15:0, C16:0 and anteiso-C17:0 and its polar lipids consisted of diphosphatidylglycerol, phosphatidylglycerol, one unidentified glycolipid and two unidentified phospholipids. The predominant menaquinone was MK-7. The cell-wall peptidoglycan was based on meso-diaminopimelic acid. Based on the results of phylogenetic, physiological and chemotaxonomic comparative analyses, the isolate is assigned to a novel species of the genus Gracilibacillus, for which the name Gracilibacillus eburneus sp. nov. is proposed, with the type strain YIM 93565T (= DSM 23710T = CCTCC AB 2013249T).

Determination of four main components of gentamicin in animal tissues after solid-phase extraction by high-performance liquid chromatography/tandem mass spectrometry.

RATIONALE: An analytical method for gentamicin in animal tissues was developed and validated. An alkaline mobile phase with an HPH C8 column was selected so that all the four gentamicin components were retained and eluted without using fluorinated ion-pairing reagents. METHODS: The method is sufficiently sensitive and highly selective, using a strong cation-exchange solid-phase extraction cartridge (PCX) to clean up the samples. Different types of solid-phase extraction columns and membranes were considered to obtain a high recovery. The method was validated on spiking samples, recovery, inter- and intra-assay variation, to ensure its accuracy and precision. RESULTS: The LOQ (S/N ≥ 10) for gentamicin in goat meat, liver, kidney and adipose tissue was 25, 50, 30 and 30 ng/g, respectively; the LOD (S/N ≥ 3) was 5, 10, 10 and 10 ng/g, respectively. The recoveries were between 88% and 106%. The method in all animal tissues was calibrated from 10 to 1000 µg/L in the matrix-assisted standard solution. CONCLUSIONS: The novelty of this method is that the commonly used fluorinated ion-pairing reagent was not used in the mobile phase in our analysis, greatly reducing the contamination of the ESI source in negative mode. Moreover, the four gentamicin components were clearly separated via chromatographic separation.

A strategy of utilizing Zn(II) as metallic pivot in room temperature ionic liquid to prepare molecularly imprinted polymers for compound with intramolecular hydrogen bonds.

A method of preparing molecularly imprinted polymers (MIPs) with Zn(II) as a metallic pivot was adopted to solve the problem of imprinting compound with intramolecular hydrogen bonds by forming stronger coordination binding interaction among the template-functional monomer-Zn2+ complex. A ternary porogenic system including dimethyl sulfoxide, dimethylformamide, and room temperature ionic liquid 1-butyl-3-methylimidazolium tetrafluoroborate was employed to fabricate imprinted monolith with high porosity and good flow-through properties, in which chicoric acid (CA), zinc acetate, 4-vinylpyridine (4-VP), and ethylene glycol dimethacrylate (EDMA) was the template, metallic ion, functional monomer, as well as crosslinker, respectively. The influence of polymerization factors including the 4-VP-CA ratio, monomer-crosslinker ratio, template-Zn2+ ratio on imprinting factors was systematically investigated. When the ratio of 4-VP to CA was 24:1, the greatest IF value (24.81) was achieved on the CA-MIP prepared with zinc acetate. In addition, off-line SPE with the optimal MIPs monolith led to high purity of CA (98.0% ± 0.5%) from extraction of Cichorium intybus L. roots with the recovery of 77.5% ± 2.5% (n = 6). As a conclusion, the strategy of introducing metal ions as metal pivot to prepare MIPs was a powerful method for the MIPs synthesis to the template molecules with intramolecular hydrogen bonds.

Improvement of imprinting effect of ionic liquid molecularly imprinted polymers by use of a molecular crowding agent.

We aimed to improve the imprinting effect of ionic liquid molecularly imprinted polymers (MIPs) by use of a molecular crowding agent. The ionic liquid 1-vinyl-3-ethylimidazolium tetrafluoroborate ([VEIm][BF4]) was used as the functional monomer and aesculetin was used as the template molecule in a crowding environment, which was made up of a tetrahydrofuran solution of polystyrene. The ionic liquid MIPs that were prepared in the crowding environment displayed an enhanced imprinting effect. NMR peak shifts of active hydrogen of aesculetin suggested that interaction between the functional monomer and the template could be increased by the use of a crowding agent in the self-assembly process. The retention and selectivity of aesculetin were affected greatly by high molecular crowding, the amount of high molecular weight crowding agent, and the ratio of [VEIm][BF4] to aesculetin. The optimal MIPs were used as solid-phase extraction sorbents to extract aesculetin from Cichorium glandulosum. A calibration curve was obtained with aesculetin concentrations from 0.0005 to 0.05 mg mL-1 (correlation coefficient R 2 of 0.9999, y = 1519x + 0.0923). The limit of quantification was 0.12 µg mL-1, and the limit of detection was 0.05 µg mL-1. The absolute recovery of aesculetin was (80 ± 2)% (n = 3), and the purity of aesculetin was (92 ± 0.5)% (n = 5). As a conclusion, molecular crowding is an effective approach to obtain ionic liquid MIPs with high selectivity even in a polar solvent environment.

Increased neutrophil gelatinase-associated lipocalin (NGAL) promotes airway remodelling in chronic obstructive pulmonary disease.

Airway remodelling is an important component of chronic obstructive pulmonary disease (COPD). Neutrophil gelatinase-associated lipocalin (NGAL) from neutrophils may drive COPD epithelial-mesenchymal transition (EMT). NGAL expression was quantified in the lungs of COPD patients and bronchoalveolar lavage fluid (BALF) of ozone-treated mice. Reticular basement membrane (RBM) thickness and E-cadherin and α-smooth muscle actin (α-SMA) expression were determined in mice airways. Effects of cigarette smoke extract (CSE) and inflammatory factors on NGAL expression in human neutrophils as well as the effects of NGAL on airway structural cells was assessed. NGAL was mainly distributed in neutrophils and enhanced in lung tissues of both COPD patients and BALF of ozone-treated mice. We showed decreased E-cadherin and increased α-SMA expression in bronchial epithelium and increased RBM thickness in ozone-treated animals. In vitro, CSE, IL-1ß and IL-17 enhanced NGAL mRNA expression in human neutrophils. NGAL, in turn, down-regulated the expression of E-cadherin and up-regulated α-SMA expression in 16HBE cells via the WNT/glycogensynthase kinase-3ß (GSK-3ß) pathway. Furthermore, NGAL promoted the proliferation and migration of human bronchial smooth muscle cells (HASMCs). The present study suggests that elevated NGAL promotes COPD airway remodelling possibly through altered EMT. NGAL may be a potential target for reversing airway obstruction and remodelling in COPD.

One-Pot Hydrothermal Synthesis of Carbon Dots with Efficient Up- and Down-Converted Photoluminescence for the Sensitive Detection of Morin in a Dual-Readout Assay.

Blue luminescent carbon dots (CDs) with a high photoluminescence (PL) quantum yield (48.3 ± 5.3%) were prepared by the one-pot hydrothermal reaction of citric acid with poly(ethylenimine) (PEI). The CDs display bright PL, narrow emission spectra, pH-dependent PL intensity, high photostability, and up-converted luminescence. The CDs exhibit a quenching of both down- and up-conversion PL in the presence of morin and thus serve as useful probes for morin detection. Both down- and up-conversion measurements allow the quantification of concentrations from 0 to 300 µmol/L with a detection limit of 0.6 µmol/L, and this dual-mode detection increases the reliability of the measurement. The proposed method of determination is simple, sensitive, and cost-effective, with potential applications in clinical and biochemical assays.

High Sensitive and Selective Detection of PFOS with Resonance Light Scattering Technology Based on the Interaction with Victoria Blue B.

This report presents a simple, sensitive and selective victoria blue B (VBB)-based resonance light scattering (RLS) assay of perfluorooctane sulfonate (PFOS). In pH 6.0 KH2PO4-NaOH buffer solution, VBB can be protonated and reacts with PFOS through electrostatic interactions to produce ionic-association complexes. Simultaneously, the interaction leads to enhanced resonance light scattering intensities greatly, which are characterized with a peak at 277 nm. It is found that the enhanced RLS intensity is proportional to the concentration of PFOS in a range of 0.05 to 4.0 µmol·L-1. The limit of detection is 5.0 nmol·L-1. While, under the optimal experimental conditions, almost no change of the resonance light scattering intensity was observed between VBB and perfluorooctane acid (PFOA) which is one kind of representative perfluorinated compounds (PFCs). The excellent selectivity of PFOS could be due to the hydrophobicity of PFOS higher than PFOA. It is worth noting that the proposed method is capable of differentiating PFOS and some other PFCs. UV/Vis absorption spectrum and scanning electron microscope (SEM) image both were investigated to further validate the reaction mechanism. The interference of coexisting foreign substances and the optimum tests of reaction conditions, including pH value, reaction time, experimental temperature and ionic strength, were also investigated. This method has been successfully applied to the determination of PFOS in environmental water samples with RSD ≤1.74%. The experimental process as followed: In 2 mL colorimetric tube, 200 µL pH 6.0 KH2PO4-NaOH buffer solution, followed by adding 600 µL 20 µmol·L-1 VBB solution, swirl evenly, then add the right amount of PFOS solution. After vortex mixing with ultrapure water volume to 2 mL, swirl to mix, stand for 10 min at room temperature. Then the mixture was transferred for RLS measurements and absorption measurement.