Production and characterization of GPC3-N protein and its nanobody.

Glypican-3 (GPC3) has a promise to be the diagnostic biomarker as well as therapeutic target for hepatocellular carcinoma (HCC). Nanobody have the great potential in clinical diagnosis and treatment for their characteristics of small size, high solubility, stability, manipulability, binding advantages, and ease of production. In this study, the recombinant glypican-3-N terminal (GPC3-N) protein was expressed as inclusion body in E. coli BL21(DE3)pLysS cells and then purified, which is then used as the immunogen to construct nanobody phage library. The positive clone (named MF15) was obtained by four rounds of bio-panning, and then transformed into the E. coil TOP10F' cells to express nanobody protein, with the molecular weight of 19 kDa. Both Western blot and immunofluorescence analysis revealed that bacterially expressed GPC3-N protein is biologically active, and MF15 could specifically recognized native GPC3 expressed in HepG2 cells. The results in this study would provide the technical support for the development of diagnostic kits and antibody drugs targeting GPC3.

A bifunctional immunosensor based on osmium nano-hydrangeas as a catalytic chromogenic and tinctorial signal output for folic acid detection.

As a neglected member of the platinum group elements, osmium, the metal with the highest density in the earth, is very suitable for the preparation of a peroxidase with high catalytic activity and stability, and can also be associated with the development of a sensor. In this study, we accessed Os nano-hydrangeas (OsNHs) with one-pot synthesis and utilized them in a bifunctional immunosensor that can present both catalytic chromogenic and tinctorial signal for nanozyme-linked immunosorbent assay (NLISA) and lateral flow immunoassay (LFIA) for use in folic acid (FA) detection. In the OsNHs-NLISA, the linear range is from 9.42 to 167.53 ng mL-1. The limit of detection (LOD) is 4.03 ng mL-1 and the IC50 value is 39.73 ng mL-1. In OsNHs-LFIA, the visual cut-off value and limit of detection (v-LOD) are 100 ng mL-1 and 0.01 ng mL-1, respectively. Additionally, the outcome from the specificity and spiked sample analysis offered recovery from the spiked milk powder sample ranging from 93.9 to 103.6% with a coefficient of variation under 4.9%, compared with UPLC-MS/MS for a correlation of R2 = 0.999 and admirable validation. The promising application of the OsNHs can also be used in other bioprobes, and this bifunctional immunosensor analysis mode is suitable for diversified analytes.

An ultrasensitive colorimetric assay based on a multi-amplification strategy employing Pt/IrO2@SA@HRP nanoflowers for the detection of progesterone in saliva samples.

Progesterone (P4) belongs to a factor that affects stress response and is a potential carcinogen, and saliva levels are expected to be a standard measurement for clinical diagnosis. In this study, a new type of nanoflower with both recognition functionality and catalytic substrate ability was prepared by copper phosphate, Pt/IrO2 nanocomposites (Pt/IrO2 NPs), streptavidin (SA) and horseradish peroxidase (HRP) via a one-pot co-precipitation strategy. Due to the enhanced catalytic activity and stability of Pt/IrO2@SA@HRP nanoflowers, we developed a powerful and sensitive multiple-catalysis ELISA to monitor progesterone in saliva. Multiple-catalysis ELISA based on a specific antibody and Pt/IrO2@SA@HRP nanoflowers exhibited a linear interval range from 0.217 ng mL-1 to 7.934 ng mL-1. The median inhibitory concentration (IC50) for progesterone is 1.311 ng mL-1 and the limit of detection (LOD = IC10) is 0.076 ng mL-1 in the proposed method. Satisfactory recoveries were in a range of 79.6-107% with an acceptable coefficient of variation (below 10.6%). Results of the multiple-catalysis ELISA and LC-MS/MS had a good coincidence. Our result unraveled that multiple-catalysis ELISA is a potentially serviceable tool for the detection of progesterone in saliva.

Au-Au/IrO2@Cu(PABA) Reactor with Tandem Enzyme-Mimicking Catalytic Activity for Organic Dye Degradation and Antibacterial Application.

Herein, a Au-Au/IrO2 nanocomposite with tandem enzyme-mimicking activity was innovatively synthesized, which can show outstanding glucose oxidase (GOx)-like activity and peroxidase-like activity simultaneously under neutral conditions. Moreover, a Au-Au/IrO2@Cu(PABA) reactor was prepared via encapsulation of the Au-Au/IrO2 nanocomposite in a Cu(PABA) metal organic framework. The reactor not only exhibits excellent organic solvent stability, acid resistance, and reusability but also displays better cascade reaction catalytic efficiency (kcat/Km = 148.86 min-1 mM-1) than the natural free enzyme system (GOx/HRP) (kcat/Km = 98.20 min-1 mM-1) and Au-Au/IrO2 nanocomposite (kcat/Km = 135.24 min-1 mM-1). In addition, it is found that the reactor can catalyze glucose or dissolved oxygen to produce active oxygen species (ROS) including HO, 1O2, and O2-· through its enzyme-mimicking activity. Finally, the novel reactor was successfully used in organic dye degradation and antibacterial application. The results show that it can effectively degrade methyl orange, methylene blue, and rhodamine B, which all can reach a degradation rate of nearly 100% after interacting with Au-Au/IrO2@Cu (PABA) for 3.5 h. Furthermore, the reactor also exhibits excellent antibacterial activity, so as to achieve a complete bactericidal effect to Staphylococcus aureus and Escherichia coli at a concentration of 12.5 µg mL-1.

Photodegradation kinetics, mechanism and aquatic toxicity of deltamethrin, permethrin and dihaloacetylated heterocyclic pyrethroids.

Photochemical methods attracted much research interests for their high-efficiency and low secondary pollution. Decomposition of synthetic pyrethroids, the fourth major group of insecticides in use worldwide, was also of great significance due to their possible environmental risks. The photodegradation of deltamethrin, permethrin and dihaloacetylated heterocyclic pyrethroids in methanol/acetone = 9/1 (by volume) by a 400 W mercury lamp was examined. The t1/2 of tested pyrethroids was less than 25 min, except for cis-permethrin with a t1/2 of up to 50 min. The trans-isomer of permethrin and compound DCA-01 with a smaller t1/2 might be more susceptible to degradation than their cis-isomer. Besides, the photodegradation of pyrethroids was divided into twelve pathways including isomerization, ester hydrolysis, ester bond cleavage, CO bond cleavage, 3,3-dimethylacrylate formation, double bond break, C1-C3 bond cleavage in cyclopropyl, reductive dehalogenation, decarboxylation, nucleophilic reagents attack on lone pair electrons on oxygen atoms in the phenyl ether, cyano hydrolysis, and halogenated hydrocarbon hydrolysis. The ECOSAR program displayed that pyrethroids and most of their photodegradation products were toxic to fish, daphnid, green algae. Particularly, some photodegradation products were more harmful to aquatic organisms than their parents.

Acute and chronic toxicity of deltamethrin, permethrin, and dihaloacetylated heterocyclic pyrethroids in mice.

BACKGROUND: Pyrethroids, a class of insecticides, that act on the nervous system of insects. Frequent consumption of foods with pyrethroid residues increase the risk of developmental and neurological diseases in humans. Assessing the toxicity of novel synthetic pyrethroids to mammals is also critical to the development of agrochemicals. RESULTS: Using mice as models, the acute and chronic toxicity of deltamethrin, permethrin, dihaloacetylated pyrethroids to mammals was researched by gavage administration. Acute toxicity assessment displayed that the median lethal dose (LD50 ) of deltamethrin, permethrin and dihaloacetylated heterocyclic pyrethroids DCA-O, DCA-01, and DCA-11 tested were greater than 500 mg/kg of weight. Furthermore, chronic toxicity assessment demonstrated that deltamethrin, permethrin caused epidermal damage near the genitals, while dihaloacetylated heterocyclic pyrethroids DCA-O, DCA-01, and DCA-11 showed no relevant symptoms. However, both the acute and chronic toxicity assessment suggested that pyrethroids exposure induced mice loss weight. Additionally, the elevated plus maze (EPM) test showed that pyrethroids caused anxiety-like behaviors and no motor defects in Kunming mice. Beside, during the sucrose preference test (SPT), 60-day pyrethroids exposure increased excitatory behaviors in mice. However, the neurochemical studies displayed that pyrethroids exposure increased the total amount of glutamate (Gln), glutamine (Glu) and γ-aminobutyric acid (GABA) in the mice's blood. CONCLUSION: Pyrethroids exposure induced weight loss in mice, although the acute oral toxicity of deltamethrin, permethrin and dihaloacetylated heterocyclic pyrethroids DCA-O, DCA-01, and DCA-11 was low. However, regarding chronic toxicity, deltamethrin, permethrin and dihaloacetylated heterocyclic pyrethroids DCA-O, DCA-01, and DCA-11 induced anxiety-like behaviors, excitatory behaviors, Gln-Glu-GABA circulatory dysfunction in blood. Particularly, deltamethrin, though permethrin also had reproductive toxicity. © 2020 Society of Chemical Industry.

Synthesis, insecticidal activity, resistance, photodegradation and toxicity of pyrethroids (A review).

Pyrethroids are a class of highly effective, broad-spectrum, less toxic, biodegradable synthetic pesticides. However, despite the extremely wide application of pyrethroids, there are many problems, such as insecticide resistance, lethal/sub-lethal toxicity to mammals, aquatic organisms or other beneficial organisms. The objectives of this review were to cover the main structures, synthesis, steroisomers, mechanisms of action, anti-mosquito activities, resistance, photodegradation and toxicities of pyrethroids. That was to provide a reference for synthesizing or screening novel pyrethroids with low insecticide resistance and low toxicity to beneficial organisms, evaluating the environmental pollution of pyrethroids and its metabolites. Besides, pyrethroids are mainly used for the control of vectors such as insects, and the non-target organisms are mammals, aquatic organisms etc. While maintaining the insecticidal activity is important, its toxic effects on non-target organisms should be also considered. Pyrethroid resistance is present not only in insect mosquitoes but also in environmental microorganisms, which results in anti-pyrethroids resistance (APR) strains. Besides, photodegradation product dibenzofurans is harmful to mammals and environment. Additionally, pyrethroid metabolites may have higher hormonal interference than the parents. Particularly, delivery of pyrethroids in nanoform can reduce the discharge of more toxic substances (such as organic solvents, etc.) to the environment.

Synthesis, insecticidal activities and resistance in Aedes albopictus and cytotoxicity of novel dihaloacetylated heterocyclic pyrethroids.

BACKGROUND: The use of safe and effective insecticides against mosquito-borne stocks is still one of the most effective ways to rapidly interrupt the spread of diseases such as malaria, dengue etc., but many insects are resistant to most insecticides and most of the traditional pyrethroids (like permethrin, deltamethrin etc.) contain phenoxy groups so it is easy to form dibenzofuran, a pollutant, after photodegradation. It is therefore necessary to synthesize novel pyrethroids which have better mosquito-killing activity, poor resistance to mosquitoes, and no dibenzofuran formation during photodegradation. RESULTS: A series of novel dihaloacetylated heterocyclic pyrethroids were synthesized to enhance antimosquito activity. Bioassays based on the guidelines of the Chinese Center for Disease Control and Prevention (CDC) indicate that the synthesized compounds DCA-O (LC50  = 2.95 ± 0.05 µg L-1 ), DCA-01 (LC50  = 5.83 ± 0.36 µg L-1 ) and DCA-11 (LC50  = 5.79 ± 1.51 µg L-1 ) exhibit high insecticidal activities and sensitivity against Aedes albopictus while deltamethrin and permethrin produced moderate levels of resistance. Moreover, the analysis of the photodegradation implies that the novel compounds synthesized by substituting a phenoxy group with a heterocyclic ring do not form dibenzofuran, a pollutant. Lastly, the cytotoxicity data of pyrethroids on human dopaminergic neuroblastoma SH-SY5Y cells confirm that the synthesized compounds have low toxicity for the cells. CONCLUSION: Novel dihaloacetylated heterocyclic pyrethroids, made by simple synthesis steps, have better antimosquito activity, lower drug resistance, are less polluting to the environment and show low toxicity to human nerve cells. This also provides an effective idea for the synthesis of pyrethroids. Novel dihaloacetylated heterocyclic pyrethroids have better antimosquito activity, lower drug resistance, are less polluting to the environment, ahow low toxicity to human nerve cells and simple synthesis steps. © 2019 Society of Chemical Industry.