Enhanced production of pinosylvin stilbene with aging of Pinus strobus callus and nematicidal activity of callus extracts against pinewood nematodes

Pinosylvin stilbenes are phenolic compounds mainly occurring in the Pinaceae family. We previously reported that the accumulation of two pinosylvin stilbene compounds, dihydropinosylvin methyl ether (DPME) and pinosylvin monomethyl ether (PME), in Pinus strobus trees was highly enhanced by infection with pine wood nematodes (PWNs: Bursaphelenchus xylophilus), and these two compounds showed strong nematicidal activity against PWNs. In this work, we established a system of pinosylvin stilbene (DPME and PME) production via the in vitro culture of P. strobus calli, and we examined the nematicidal activity of callus extracts. Calli were induced from the culture of mature zygotic embryos of P. strobus. Optimized growth of calli was obtained in 1/2 Litvay medium with 1.0 mg/L 2,4-D and 0.5 mg/L BA. DPME and PME accumulation did not occur in nonaged (one-month-old) calli but increased greatly with prolonged callus culture. The concentrations of DPME and PME in three-month-old dark-brown calli were 6.4 mg/g DW and 0.28 mg/g DW, respectively. The effect of methyl jasmonate treatment on the accumulation of DPME and PME was evaluated in cell suspension culture of P. strobus. However, the treatment appeared to show slight increase of DPME accumulation compared to callus browning. A test solution prepared from crude ethanol extracts from aged calli (three months old) containing 120 µg/ml DPME and 5.16 µg/ml PME treated with PWNs resulted in 100% immobilization of the adult PWNs and 66.7% immobilization of the juvenile PWNs within 24 h. However, nonaged callus extracts did not show any nematicidal activity against juvenile PWNs and showed less than 20% nematicidal activity against adult PWNs. These results indicate that pinosylvin stilbenes can be effectively produced by prolonged culture of P. strobus calli, can be isolated using simple ethanolic extraction, and are applicable as beneficial eco-friendly compounds with nematicidal activity against PWNs.

Trace analysis of emerging and regulated mycotoxins in infant stool by LC-MS/MS

Infants are sensitive to negative effects caused by food contaminants such as mycotoxins. To date, analytical methods assessing mycotoxin mixture exposure in infant stool are absent. Herein, we present a novel multi-mycotoxin LC-MS/MS assay capable of detecting 30+ analytes including the regulated mycotoxin classes (aflatoxins, trichothecenes, ochratoxins, zearalenone, citrinin), emerging Alternaria and Fusarium toxins, and several key metabolites. Sample preparation consisted of a ‘dilute, filter, and shoot’ approach. The method was in-house validated and demonstrated that 25 analytes fulfilled all required criteria despite the high diversity of chemical structures included. Extraction recoveries for most of the analytes were in the range of 65–114% with standard deviations below 30% and limits of detection between 0.03 and 11.3 ng/g dry weight. To prove the methods’ applicability, 22 human stool samples from premature Austrian infants (n = 12) and 12-month-old Nigerian infants (n = 10) were analyzed. The majority of the Nigerian samples were contaminated with alternariol monomethyl ether (8/10) and fumonisin B1 (8/10), while fumonisin B2 and citrinin were quantified in some samples. No mycotoxins were detected in any of the Austrian samples. The method can be used for sensitive human biomonitoring (HBM) purposes and to support exposure and, potentially, risk assessment of mycotoxins. Moreover, it allows for investigating potential associations between toxicant exposure and the infants’ developing gut microbiome. Graphical abstract

Assessment of the Optimum Linker Tethering Site of Alternariol Haptens for Antibody Generation and Immunoassay Development

Immunochemical methods for mycotoxin analysis require antigens with well-defined structures and antibodies with outstanding binding properties. Immunoreagents for the mycotoxins alternariol and/or alternariol monomethyl ether have typically been obtained with chemically uncharacterized haptens, and antigen conjugates have most likely been prepared with mixtures of functionalized molecules. For the first time, total synthesis was performed, in the present study, to obtain two haptens with opposite linker attachment locations. The functionalized synthetic haptens were purified and deeply characterized by different spectrometric methods, allowing the preparation of bioconjugates with unequivocal structures. Direct and indirect competitive enzyme-linked immunosorbent assays, using homologous and heterologous conjugates, were employed to extensively evaluate the generated immunoreagents. Antibodies with high affinity were raised from conjugates of both haptens, and a structure-activity relationship between the synthetic haptens and the specificity of the generated antibodies could be established. These results pave the way for the development of novel highly sensitive immunoassays selective of one or two of these Alternaria mycotoxins.

Preparation of Polyethylene Glycol Monomethyl Ether Chitosan-diethylenetriamine Pentaacetic Acid and Its Effect on 89SrCl2 Excretion and Radiation Protection in the Digestive Tract of Rats

In this study, hydrophilic polymer polyethylene glycol monomethyl ether chitosan-diethylenetriamine pentaacetic acid (mPEGCS-DTPA) was synthesized via chemical synthesis and cross-linking, and its cytotoxicity and radiation protective effect in vivo were studied. The results revealed that mPEGCS-DTPA exhibits good cytocompatibility and increases the excretion of radionuclides through the digestiveand urinary tracts. The pathological results of the small intestine revealed that mPEGCS-DTPA exerts a good radiation-protective effect and could reduce the α and β-emitting radiation-induced damage of the intestinal mucosa. MPEGCS-DTPA can increase the excretion of radionuclides in the digestive tract and exhibits effective protection against radiation.

Sonodynamic Effects of a Novel Ether-Group Modified Porphyrin Derivative Combined With Pulsed Low-Intensity Ultrasound on PC-9 Cells

Sonodynamic therapy (SDT) is a developing modality for cancer treatment based on the synergistic effect of ultrasound and chemical compounds which are known as sonosensitizers. The development of more efficient sonosensitizers has become an urgent issue in this field. In this study, a novel porphyrin derivative (BBTPP) mediated SDT was evaluated on PC-9 cells. Pulsed low-intensity ultrasound (PLIU) was used for its little thermal and mechanical damage. The accumulation of drugs in cells was evaluated through porphyrin fluorescence, and the cytotoxicity of BBTPP was evaluated using a cell counting kit-8 assay. The sonodynamic effect was investigated by Hoechst 33342/PI and Annexin V-FITC/PI double staining, which showed an apoptotic rate of 18.87% in the BBTPP-SDT group, as compared with 1.71%, 1.4%, 1.57%, 3.61%, 11.18% in the control, BBTPP, hematoporphyrin monomethyl ether (HMME), ultrasound, and HMME-SDT groups, respectively. The sono-toxic effect of BBTPP was significantly superior to HMME. Our results showed that BBTPP-SDT resulted in much higher intracellular reactive oxygen species (ROS) and lipid peroxidation levels which were evaluated by 2′,7′-dichlorodihydrofluorescein diacetate (H2DCFDA) and Liperfluo assay, respectively. The expressions of Bax, Bcl-2, caspase-9, caspase-8, and cleaved caspase-3 proteins were evaluated to investigate the apoptotic mechanism of BBTPP-SDT. The results of this study showed that the combination of BBTPP and PLIU induced the generation of ROS, resulting in lipid peroxidation, and activated both the extrinsic and intrinsic apoptotic pathways of PC-9 cells. Our results also suggested that the ether group introduced in the side chain of porphyrin could enhance the sono-toxicity of porphyrin-based sensitizers under the sonication of PLIU. These results supported the possibility of BBTPP as a promising sonosensitizer, and an appropriate side chain could enhance the sono-sensitivity of porphyrins.

Biodegradable reduce expenditure bioreactor for augmented sonodynamic therapy via regulating tumor hypoxia and inducing pro-death autophagy

Backgrounds Sonodynamic therapy (SDT) as an emerging reactive oxygen species (ROS)-mediated antitumor strategy is challenged by the rapid depletion of oxygen, as well as the hypoxic tumor microenvironment. Instead of the presently available coping strategies that amplify the endogenous O2 level, we have proposed a biodegradable O2 economizer to reduce expenditure for augmenting SDT efficacy in the present study. Results We successfully fabricated the O2 economizer (HMME@HMONs-3BP-PEG, HHBP) via conjugation of respiration inhibitor 3-bromopyruvate (3BP) with hollow mesoporous organosilica nanoparticles (HMONs), followed by the loading of organic sonosensitizers (hematoporphyrin monomethyl ether; HMME) and further surface modification of poly(ethylene glycol) (PEG). The engineered HHBP features controllable pH/GSH/US-sensitive drug release. The exposed 3BP could effectively inhibit cell respiration for restraining the oxygen consumption, which could alleviate the tumor hypoxia conditions. More interestingly, it could exorbitantly elevate the autophagy level, which in turn induced excessive activation of autophagy for promoting the therapeutic efficacy. As a result, when accompanied with suppressing O2-consumption and triggering pro-death autophagy strategy, the HHBP could achieve the remarkable antitumor activity, which was systematically validated both in vivo and in vitro assays. Conclusions This work not only provides a reduce expenditure means for enduring SDT, but also represents an inquisitive strategy for tumor treatments by inducing pro-death autophagy. Graphical Abstract

Separation of Polar Compounds using Poly(Ethylene Oxide) Bonded Stationary Capillary Liquid Chromatography

Poly(ethylene oxide) (PEO) bonded stationary phase has been synthesized by a single and simple step reaction. Poly(ethylene glycol monomethyl ether p-toluene sulfonate) (tosylated-PEO, molecular weight 900, n ≈ 18) was chemically bonded to 3-aminopropyl silica (TSKgel NH2-60, 5 µm particle size, and 60 Å mean pore diameter). The prepared stationary phase was able to separate polar compounds such as phenolics and nucleobases in capillary liquid chromatography. The retention and separation of phenolics and nucleobases could be achieved under isocratic elution condition. Nucleobases such as thymine, adenine, uracil, uridine, cytidine and toluene and phenolics (phenol, pyrocatechol, pyrogallol) were baseline separated in less than 6 min using 98% acetonitrile and less than 7 minutes using 80% acetonitrile, respectively. We demonstrated that the retention of nucleobases as analyte decreased with decreasing eluent concentration. The retention of these polar compounds was believed to be based on dipole-dipole and/or hydrogen-bonding interactions.