Multiplex PCR system to track authorized and unauthorized genetically modified soybean events in food and feed

A diverse range of genetic elements has been used to develop genetically modified organisms (GMOs) over the last 18 years. Screening methods that target few elements, such as the Cauliflower Mosaic Virus 35S promoter (P-35S) and Agrobacterium tumefaciens nopaline terminator (T-nos), are not sufficient to screen GMOs. In the present study, a multiplex PCR system for all globally commercialized GM soybean events was developed to easily trace the events. For this purpose, screening elements of 24 GM soybean events were investigated and 9 screening targets were selected and divided into three individual triplex PCR systems: P-35S, ribulose-1,5-bisphosphate carboxylase small subunit promoter of Arabidopsis thaliana, T-nos, T-35S, pea E9 terminator, open reading frame 23 terminator of A. tumefaciens, proteinase inhibitor II terminator of potato, acetohydroxy acid synthase large subunit terminator of A. thaliana, and the revealed 3′ flanking sequences of DP-305423-1. The specificity of the assays was confirmed using thirteen GM soybean events as the respective positive/negative controls. The limit of detection of each multiplex set, as determined using certified reference materials of specific GM events, ranged from 0.03 to 0.5%, depending upon target. Furthermore, 26 food samples that contained soybean ingredients, which were purchased from the USA, China, Japan, and Korea, were analyzed, 17 of which contained one or more GM soybean events. These results suggest that the developed screening method can be used to efficiently track and identify 24 GM soybean events in food and feed.     

Antitumor Effects of Vitamin D Analogs on Hamster and Mouse Melanoma Cell Lines in Relation to Melanin Pigmentation

Deregulated melanogenesis is involved in melanomagenesis and melanoma progression and resistance to therapy. Vitamin D analogs have anti-melanoma activity. While the hypercalcaemic effect of the active form of Vitamin D (1,25(OH)₂D₃) limits its therapeutic use, novel Vitamin D analogs with a modified side chain demonstrate low calcaemic activity. We therefore examined the effect of secosteroidal analogs, both classic (1,25(OH)₂D₃ and 25(OH)D₃), and novel relatively non-calcemic ones (20(OH)D₃, calcipotriol, 21(OH)pD, pD and 20(OH)pL), on proliferation, colony formation in monolayer and soft-agar, and mRNA and protein expression by melanoma cells. Murine B16-F10 and hamster Bomirski Ab cell lines were shown to be effective models to study how melanogenesis affects anti-melanoma treatment. Novel Vitamin D analogs with a short side-chain and lumisterol-like 20(OH)pL efficiently inhibited rodent melanoma growth. Moderate pigmentation sensitized rodent melanoma cells towards Vitamin D analogs, and altered expression of key genes involved in Vitamin D signaling, which was opposite to the effect on heavily pigmented cells. Interestingly, melanogenesis inhibited ligand-induced Vitamin D receptor translocation and ligand-induced expression of VDR and CYP24A1 genes. These findings indicate that melanogenesis can affect the anti-melanoma activity of Vitamin D analogs in a complex manner.     

Simultaneous determination of mycotoxin in commercial coffee

Mycotoxins are secondary metabolites produced by filamentous fungi that usually contaminate food products. Coffee is a natural product susceptible to mycotoxin contamination. The present study evaluates the presence of nivalenol, deoxynivalenol, T-2 and HT-2 Toxin, diacetoxyscirpenol, aflatoxin B₁, aflatoxin B₂, aflatoxin G₁, aflatoxin G₂, fumonisin B₁, fumonisin B₂, ochratoxin A, zearalenone, enniatin A, enniatin A₁, enniatin B, enniatin B₁, and beauvericin in coffee samples, using liquid chromatography tandem mass spectrometry (LC-MS/MS). The results show that zearalenone was not present in any sample. In the positive samples the contents of fumonisins ranged from 58.62 to 537.45 μg/kg, emerging mycotoxins ranged from 0.10 to 3569.92 μg/kg, aflatoxins ranged from 0.25 to 13.12 μg/kg, and trichothecenes, excepting nivalenol, ranged from 5.70 to 325.68 μg/kg. Nivalenol presented the highest concentrations, from 0.40 to 25.86 mg/kg. Ochratoxin A ranged from 1.56 to 32.40 μg/kg, and five samples exceeded the maximum limit established by the European Commission.     

Biomineralization,strength and cytocompatibility improvement of bredigite scaffolds through doping/coating

Coprecipitation-derived, sacrificial polymeric (urethane) foam-fabricated bredigite (Ca₇MgSi₄O₁₆) scaffolds were processed by individual and combined treatments of fluoride doping and poly (lactic-co-glycolic acid) (PLGA) coating and then studied in terms of structure, mechanical strength, bioactivity and cell biocompatibility in vitro. According to scanning electron microscopy and Archimedes porosimetry, the geometrical characteristics of pores for all the scaffolds are in the appropriate range for hard tissue regeneration applications. The apatite-formation ability of the samples immersed in a simulated body fluid is improved by doping for both the bare and coated conditions, based on microscopic and energy-dispersive X-ray spectroscopic analyses. Both the treatments advantageously buffer physiological pH changes imposed due to the fast bioresorption of the ceramic. Also, the biodegradable PLGA coating typically enhances the compressive strength of the scaffolds, which is critical for bone tissue engineering. In accordance with the MTT assay on osteoblast-like cells (MG-63) cultures, both the processes individually enhance the cell viability, while the highest improvement is obtained for the combined application of them. It is finally concluded that fluoride doping and PLGA coating are impressive approaches to improve the bioperformance of bredigite-based scaffolds.     

Multichromic polymers based on pyrene clicked thienylpyrrole

In this study, the effect of pendant pyrene on the optical and electronic properties of poly(2,5‐dithienylpyrrole)s was studied. For this purpose a new pyrene coupled 2,5‐dithienylpyrrole derivative (SNS‐pyrene) was synthesized through click reaction. SNS‐pyrene was electrochemically polymerized and its electrochemical and optical properties were investigated by electrochemical and optical techniques. The polymer had a band gap of 3.36 eV and displayed light green to blue color variation upon oxidation in less than 2.48 s. Additionally, electrochemical copolymerization of SNS‐pyrene with 3,4‐ethylenedioxythiophene was achieved whilst a detailed investigation was performed on the effect of electrochemical polymerization conditions on the optoelectronic properties of the copolymers. Studies revealed that the copolymers exhibit multichromic reversible redox behavior with lower band gaps and shorter switching times than their parent polymer, P(SNS‐pyrene) © 2014 Society of Chemical Industry.     

Single-Crystalline TiO2(B) Nanobelts with Unusual Large Exposed {100} Facets and Enhanced Li-Storage Capacity

The {100} facet of single-crystalline TiO₂(B) is an ideal platform for inserting Li ions, but it is hard to be obtained due to its high surface energy. Here, the single-crystalline TiO₂(B) nanobelts from H₂Ti₃O₇ with nearly 70% {100} facets exposed are synthesized, which significantly enhances Li-storage capacity. The first-principle calculations demonstrate an ab in-plane 2D diffusion through the exposed {100} facets. As a consequence, the nanobelts can significantly accommodate Li ions in LiTiO₂ formula with specific capacity up to 335 mAh g⁻¹, which is in good agreement with the electrochemical characterizations. Coating with conductive and protective poly(3,4-ethylenedioxythiophene)-poly(styrenesulfonate), the cut-off discharge voltage is as low as 0.5 V, leading to a capacity of 160.7 mAh g⁻¹ after 1500 cycles with a retention rate of 66% at 1C. This work provides a practical strategy to increase the Li-ion capacity and cycle stability by tailoring the crystal orientation and nanostructures.     

Discovery of Affinity-Based Probes for Btk Occupancy Assays

Bruton's tyrosine kinase (Btk) is an attractive target for the treatment of a wide array of B-cell malignancies and autoimmune diseases. Small-molecule covalent irreversible Btk inhibitors targeting Cys481 have been developed for the treatment of such diseases. In clinical trials, probe molecules are required in occupancy studies to measure the level of engagement of the protein by these covalent irreversible inhibitors. The result of this pharmacodynamic (PD) activity provides guidance for appropriate dosage selection to optimize inhibition of the drug target and correlation of target inhibition with disease treatment efficacy. This information is crucial for successful evaluation of drug candidates in clinical trials. Based on the pyridine carboxamide scaffold of a novel solvent-accessible pocket (SAP) series of covalent irreversible Btk inhibitors, we successfully developed a potent and selective affinity-based biotinylated probe 12 (2-[(4-{4-[5-(1-{5-[(3aS,4S,6aR)-2-oxo-hexahydro-1H-thieno[3,4-d]imidazol-4-yl]pentanamido}-3,6,9,12-tetraoxapentadecan-15-amido)pentanoyl]piperazine-1-carbonyl}phenyl)amino]-6-[1-(prop-2-enoyl)piperidin-4-yl]pyridine-3-carboxamide). Compound 12 has been used in Btk occupancy assays for preclinical studies to determine the therapeutic efficacy of Btk inhibition in two mouse lupus models driven by TLR7 activation and type I interferon.     

Application of Vis–NIR hyperspectral imaging in classification between fresh and frozen-thawed pork Longissimus Dorsi muscles

Fresh and frozen-thawed (F-T) pork meats were classified by Vis–NIR hyperspectral imaging. Eight optimal wavelengths (624, 673, 460, 588, 583, 448, 552 and 609 nm) were selected by successive projections algorithm (SPA). The first three principal components (PCs) obtained by principal component analysis (PCA) accounted for over 99.98% of variance. Gray-level-gradient co-occurrence matrix (GLGCM) was applied to extract 45 textural features from the PC images. The correct classification rate (CCR) was employed to evaluate the performance of the partial least squares-discriminate analysis (PLS-DA) models, by using (A) the reflected spectra at full wavelengths and (B) those at the optimal wavelengths, (C) the extracted textures based on the PC images, and (D) the fused variables combining spectra at the optimal wavelengths and textures. The results showed that the best CCR of 97.73% was achieved by applying (D), confirming the high potential of textures for fresh and F-T meat discrimination.