A ratiometric fluorescence sensor for detection of organophosphorus pesticides based on enzyme-regulated multifunctional Fe-based metal-organic framework.

The residues of organophosphorus pesticides (OPs) are increasing environmental pollution and public health concerns. Thus, the development of simple, convenient and sensitive method for detection of OPs is crucial. Herein, a multifunctional Fe-based MOF with fluorescence, catalytic and adsorption, is synthesized by a simple one-pot hydrothermal method. The ratiometric fluorescence sensor for detection of OPs is constructed by using only one multifunctional sensing material. The NH2-MIL-101(Fe) is able catalyze the o-phenylenediamine (OPD) into 2,3-diaminophenazine (DAP) in the presence of H2O2. The generated DAP can significantly quench the intrinsic fluorescence of NH2-MIL-101(Fe) by the fluorescence resonance energy transfer (FRET) and internal filtration effect (IFE), while producing a new measurable fluorescence. Without immobilization or molecular imprinting, pyrophosphate ion (PPi) can inhibit the peroxidase-like activity of the NH2-MIL-101(Fe) by chelating with Fe3+/Fe2+ redox couple. Moreover, PPi can also be hydrolyzed by alkaline phosphatase (ALP), the presence of OPs inhibits the activity of ALP, resulting in the increase of extra PPi preservation and signal changes of ratiometric fluorescence, the interactions of ALP with different OPs are explored by molecular docking, the OPs (e.g., glyphosate) interact with crucial amino acid residues (Asp, Ser, Ala, Lys and Arg) are indicated. The proposed sensor exhibits excellent detection performance for OPs with the detection limit of 18.7 nM, which provides a promising strategy for detection of OPs.

Optimizing the Spatial Density of Single Co Sites via Molecular Spacing for Facilitating Sustainable Water Oxidation.

Advances in single-atom (-site) catalysts (SACs) provide a new solution of atomic economy and accuracy for designing efficient electrocatalysts. In addition to a precise local coordination environment, controllable spatial active structure and tolerance under harsh operating conditions remain great challenges in the development of SACs. Here, we show a series of molecule-spaced SACs (msSACs) using different acid anhydrides to regulate the spatial density of discrete metal phthalocyanines with single Co sites, which significantly improve the effective active-site numbers and mass transfer, enabling one of the msSACs connected by pyromellitic dianhydride to exhibit an outstanding mass activity of (1.63 ± 0.01) × 105 A·g-1 and TOFbulk of 27.66 ± 1.59 s-1 at 1.58 V (vs RHE) and long-term durability at an ultrahigh current density of 2.0 A·cm-2 under industrial conditions for oxygen evolution reaction. This study demonstrates that the accessible spatial density of single atom sites can be another important parameter to enhance the overall performance of catalysts.

Chromosome-Scale Assembly of the Dendrobium nobile Genome Provides Insights Into the Molecular Mechanism of the Biosynthesis of the Medicinal Active Ingredient of Dendrobium.

Orchids constitute approximately 10% of flowering plant species. However, only about 10 orchid genomes have been published. Metabolites are the main way through which orchids respond to their environment. Dendrobium nobile, belonging to Dendrobium, the second largest genus in Orchidaceae, has high ornamental, medicinal, and ecological value. D. nobile is the source of many popular horticultural varieties. Among the Dendrobium species, D. nobile has the highest amount of dendrobine, which is regarded as one of the criteria for evaluating medicinal quality. Due to lack of data and analysis at the genomic level, the biosynthesis pathways of dendrobine and other related medicinal ingredients in D. nobile are unknown. In this paper, we report a chromosome-scale reference genome of D. nobile to facilitate the investigation of its genomic characteristics for comparison with other Dendrobium species. The assembled genome size of D. nobile was 1.19 Gb. Of the sequences, 99.45% were anchored to 19 chromosomes. Furthermore, we identified differences in gene number and gene expression patterns compared with two other Dendrobium species by integrating whole-genome sequencing and transcriptomic analysis [e.g., genes in the polysaccharide biosynthesis pathway and upstream of the alkaloid (dendrobine) biosynthesis pathway]. Differences in the TPS and CYP450 gene families were also found among orchid species. All the above differences might contribute to the species-specific medicinal ingredient biosynthesis pathways. The metabolic pathway-related analysis will provide further insight into orchid responses to the environment. Additionally, the reference genome will provide important insights for further molecular elucidation of the medicinal active ingredients of Dendrobium and enhance the understanding of orchid evolution.

Complete genome sequencing and comparative genome analysis of the extremely halophilic archaea, Haloterrigena daqingensis.

In the present study, we report the complete genome sequencing of Haloterrigena daqingensis species. The genome of H. daqingensis JX313T consisted of a circular chromosome with three plasmids. The genome size and G+C content were estimated to be 3835796 bp and 61.7%, respectively. A total of 4158 genes were predicted with six rRNAs and 45 tRNAs. Metabolic pathway analysis suggests that H. daqingensis JX313T codes for all the necessary genes responsible to sustain its life at saline environment. The pan-genome analysis suggests that the number of singleton-gene between H. daqingensis and other Haloterrigena species varied. The study not only helps us understand H. daqingensis strategy for dealing with high stress, but it also provides an overview of its genomic makeup.

A competitive fluorescence assay based on free-complementary DNA for ochratoxin A detection.

An ultrasensitive, rapid, and specific method for Ochratoxin A (OTA) detection was designed using complementary sequence to aptamer as a target of molecular beacon (MB). The designed loop structure of the MB has the same sequence as the aptamer with a complementary DNA (cDNA) which translates the level of the target into a measurable response. The presence of the target holds aptamer at the corresponding amount and the additional cDNAs are consumed by unbound aptamers which avails free cDNAs that resulting in fluorescence rising due to unfolding of MBs. Under the optimized conditions, the fluorescence intensity increased linearly with OTA concentration over the range of 10 pg mL-1-1 µg mL-1 with the detection limit of 0.247 pg mL-1. The application of this assay in wheat sample in comparison with HPLC-MS/MS method, demonstrated that the new assay could be a potential sensing platform for OTA detection.

Design, synthesis and evaluation of novel tacrine-multialkoxybenzene hybrids as multi-targeted compounds against Alzheimer's disease.

A series of benzoates (or phenylacetates or cinnamates) - tacrine hybrids (7a-o) were designed, synthesized and evaluated as multi-potent anti-Alzheimer drug candidates. The screening results showed that most of them exhibited a significant ability to inhibit ChEs, certain selectivity for AChE over BuChE and strong potency inhibitory of self-induced ß-amyloid (Aß) aggregation. All IC50 values of biological activity were at the nanomolar range. Especially, compound 7c displayed the greatest ability to inhibit AChE with an IC50 value of 5.63 nM and the highest selectivity with ratio of BuChE/AChE value of 64.6. Moreover, it also exhibited a potent inhibitory of Aß aggregation with an IC50 value of 51.81 nM. A Lineweaver-Burk plot and molecular modeling study showed that compound 7c targeted both the CAS and PAS of ChEs. A structure-activity relationship analysis suggested that the electron density of aromatic ring which was linked with tacrine through acetyl group played a significant role in determining the inhibitory activity.

[Recurrence of Cerebral Infarction Associated Aspirin Resistance or Chinese Medical Constitutions: a Correlation Study].

OBJECTIVE: To explore the correlation between the recurrence of cerebral infarction and aspirin resistance (AR)/Chinese medical (CM) constitutions. METHODS: Totally 413 cerebral infarction patients took Aspirin Enteric-coated Tablet (100 mg per day) while receiving routine therapy, 5 days at least in a week. They were followed-up for 12 months. Aspirin sensitivity (AS) was determined using turbidimetry. CM constitutions among patients with different AS were compared. Ratios of AR patients and AS patients of different CM constitutions in cerebral infarction recurrent patients were compared. Platelet membrane glycoproteins (GP) II b HPA-3 gene polymorphism was detected by polymerase chain reaction (PCR) method. Correlation between recurrence of cerebral infarction and AR, bb genotypes, CM constitutions times AS were analyzed by Logistic regression. RESULTS: Totally 11 patients dropped out, 101 (25.12%)with recurrent cerebral infarction and 301 (74.88%) without recurrent cerebral infarction. There were 152 (37.81%) AR patients and 250 (62.19%) AS patients. AR accounted for 26.6% (80/ 301) and AS accounted for 73.4% (221/301) in non-recurrent cerebral infarction patients. AR accounted for 71.3% (72/101) and AS accounted for 28.7% (29/101) in recurrent cerebral infarction patients. There was statistical difference in AR and AS ratios (χ2 = 64.287, P = 0.000). The proportion of yin deficiency constitution (YDC) was the largest [28.3% (43/152)] in AR patients. The proportion of blood stasis constitution (BSC) was the largest [23.6% (59/250)] in AS patients. There was statistical difference in CM constitutions between AR patients and AS patients (χ2 = 21.574, P < 0.01). The former 4 recurrent rates occurred in AR patients of YDC, BSC, damp-phlegm constitution (DPC), qi deficiency constitution (QDC). YDC occupied the first place [22.4% (34/152)]. The former 4 recurrent rates occurred in AS patients of BSC, QDC, DPC, damp-heat constitution (DHC). BSC occupied the first place [3.2% (2/250)]. Compared with non-recurrent cerebral infarction patients and AS patients, bb gene occurred most often, but aa gene and ab gene occurred obviously lesser in non-recurrent cerebral infarction patients and AR patients (χ2 = 20.171, χ2 = 55.139, P < 0.01). AR and bb gene were positively correlated with recurrent cerebral infarction (OR = 18.423, P = 0.000; OR = 1.304, P = 0.028). Body constitutions interacted with AS (OR = 0.707, P = 0.000). CONCLUSIONS: Recurrent cerebral infarction was closely related to AR and constitutional types. The recurrence rate was higher in AR patients of YDC. GP I b HPA-3 bb genotype might be a risk factor for AR and recurrent cerebral infarction.

Antioxidant and tyrosinase inhibitory effects of neolignan glycosides from Crataegus pinnatifida seeds.

In our efforts to find an inhibitor of melanin formation and develop potential depigmenting agents for skin-protecting cosmetics and medicinal products from natural resources, we focused on the seeds of Crataegus pinnatifida which showed antioxidant and tyrosinase-inhibiting activities. By activity-guided fractionation of an extract of C. pinnatifida seeds, four new neolignan glycosides, pinnatifidaninsides A-D (1-4), along with two known compounds (5-6), were isolated. Their structures were elucidated by spectroscopic analyses, especially 1D, 2D NMR and CD spectra. The antioxidant and tyrosinase-inhibiting activities of all isolates were assayed. Compound 6 showed good activity against 2,2-diphenyl-1-pikrylhydrazyl, while compounds 1, 2, 5, and 6 exhibited strong 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) free radical scavenging activity, being as effective as, or even more effective than the positive control Trolox. Moreover, compounds 5 and 6 displayed a moderate mushroom tyrosinase inhibitory activity.

Tomensides A-D, new antiproliferative phenylpropanoid sucrose esters from Prunus tomentosa leaves.

To search for novel cytotoxic constituents against cancer cells as lead structures for drug development, four new 3-phenylpropanoid-triacetyl sucrose esters, named tomensides A-D (1-4), and three known analogs (5-7) were isolated from the leaves of Prunus tomentosa. Their structures were elucidated by spectroscopic analyses (1D, 2D NMR, CD and HRESIMS). The cytotoxic activities of all isolates against four human cancer cell lines (MCF-7, A549, HeLa and HT-29) were assayed, and the results showed that these isolates displayed stronger inhibitory activities compared with positive control 5-fluorouracil. Tomenside A (1) was the most active compound with IC50 values of 0.11-0.62 µM against the four tested cell lines. The structure-activity relationship (SAR) of the isolates was also discussed. The primary screening results indicated that these 3-phenylpropanoid-triacetyl sucrose esters might be valuable source for new potent anticancer drug candidates.

Mechanisms of antibacterial activity of MgO: non-ROS mediated toxicity of MgO nanoparticles towards Escherichia coli.

The toxicity of metal oxide nanomaterials and their antimicrobial activity is attracting increasing attention. Among these materials, MgO is particularly interesting as a low cost, environmentally-friendly material. The toxicity of MgO, similar to other metal oxide nanomaterials, is commonly attributed to the production of reactive oxygen species (ROS). We investigated the toxicity of three different MgO nanoparticle samples, and clearly demonstrated robust toxicity towards Escherichia coli bacterial cells in the absence of ROS production for two MgO nanoparticle samples. Proteomics data also clearly demonstrate the absence of oxidative stress and indicate that the primary mechanism of cell death is related to the cell membrane damage, which does not appear to be due to lipid peroxidation.

Steroidal saponins from Anemarrhena asphodeloides.

Two new steroidal saponins, named anemarnoside A and anemarnoside B, along with three known compounds, timosaponin J, timosaponin B II, and timosaponin B, have been isolated from Anemarrhena asphodeloides. Their structures were established by spectroscopic techniques (IR, MS, 1D NMR, and 2D NMR) and by comparison with published data.

[Genetic analysis of the UGT1A1 gene mutation sites in a Chinese family suffered from Gilbert's syndrome].

To learn the variation in the gene for UGT1A1 enzyme, the genetic mechanism in a Chinese Han nationality family suffered from Gilbert's syndrome was studied. At first, genomic DNA from peripheral blood of the sufferer in this family was used for amplifying all of the five exons of the UGT1A1 gene by PCR, and then direct sequencing of the PCR product was applied to analyze gene mutation. The results showed that there existed a G-->A homozygous transition at nucleotide 211 leading the substitution of arginine for glycine at position 71 of corresponding protein product (G71R) and a T-->G homozygous transition at nucleotide 1456 leading the substitution of aspartic acid for tyrosine at position 486 of corresponding protein product (Y486D). No mutation was detected in promoter region and the splicing junction sites. The relevant mutation sites of the other family members were sequenced and identified to be heterozygous in the two above-mentioned mutation sites and in the TA repeat mutation in the promoter region. Furthermore, fresh blood samples were collected from all of the members to detect the serum bilirubin levels to determine the sufferer. The result was consistent with the mutation analysis. It could thus be inferred that this family was caused by mutation in the open reading frame of the gene UGT1A1.

[The epitope analysis of beta Netrin and preparation and characterization of its antibodies].

AIM: To prepare and characterize anti-human beta-Netrin antibodies. METHODS: B cell dominant epitopes of human beta-Netrin C-terminal 114 amino acid sequences were predicated by the GoldKey software. One of the epitopes was synthesized and coupled with bovine serum album (BSA) by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC). The BALB/c mice were immunized with the coupled protein. The splenocytes of immunized mice were fused with Sp2/0 cells by routine method and the hybridomas were selected in HAT medium. The hybridoma cells secreting specific antibody were detected by ELISA and cloned by limiting dilution. The titer specificity, and Ig subclass of anti-beta-Netrin mAbs were characterized by ELISA, Western blot and immunocytochemical staining. In addition, New Zealand rabbits were immunized with the coupled protein to prepare polyclonal antibody against beta-Netrin. The specificity of the antiserum was verified by Western blot. RESULTS: A 16-mer peptide NH2-FRGKRTLYPES-WTDRG-COOH was the dominant epitope of the B cells. Synthesized peptide coupled with BSA was used as the immunogen to immunize BALB/c mice. Three hybridoma cell lines that stably secrete specific mAbs were obtained. The result of immunocytochemical staining showed that prepared mAb specifically recognize the antigen in the neuronal cells. The polyclonal antibody against beta-Netrin had high specificity. Western blot analysis showed that the antiserum bind with the prokaryotically expressed beta-Netrin specifically. CONCLUSION: Using the synthesized peptides as hapten, we have prepared epitope-specific mAbs and pAb against beta-Netrin successfully.

[Identification of interaction and interaction domains between neuroglobin and Na(+), K(+)-ATPase beta2 subunit].

The pre-transformed human fetal brain cDNA library was used to screen the protein interacting with neuroglobin by using yeast two hybrid system III from ClonTech Inc. The protein encoded by one of the clones interacting with neuroglobin (NGB) was confirmed to be the C terminus of the Na(+), K(+)-ATPase beta2 subunit (NKA1b2) based on amino acid sequences. Then the full-length coding region cDNA sequence of NKA1b2 was obtained from human fetal brain cDNA library by PCR. A set of experiments were designed to test the interaction between NGB and NKA1b2. Interaction between NGB and NKA1b2 was confirmed by binding assay in vitro. Furthermore, the interaction was also proved by co-immunoprecipitation test in vivo. Moreover, the structure integrity of neuroglobin was found to be essential for the interaction between NGB and NKA1b2 by yeast two hybrid method with a series of neuroglobin truncated mutants.