The screening method for 39 phytotoxins and mycotoxins in blood and urine with liquid chromatography-high resolution mass spectrometry.

BACKGROUND: Poisonings caused by plant toxins and mycotoxins occur frequently, which do great harm to human health and social public health safety. When a poisoning incident occurs, biological samples are commonly be used to conduct the detection of toxic substances and their metabolites for targeted clinical treatment and incident analysis. OBJECTIVE: To establish an efficient and accurate analysis method of 39 phytotoxins and mycotoxins in blood and urine by high performance liquid chromatography quadrupole tandem orbitrap mass spectrometry (HPLC-Orbitrap MS). METHOD: After 3 mL of methanol being added to 1 mL blood and urine respectively for extraction and protein precipitation, the supernatant was injected into HPLC-Orbitrap MS for analysis. The phytotoxins and mycotoxins were separated by Hypersil GOLD PFP column with gradient elution using methanol-5 mmol/L ammonium acetate as mobile phase. The data were collected in ESI positive ion mode using Full MS/dd-MS2 for mass spectrometry detection. RESULT: The mass database of 39 phytotoxins and mycotoxins was developed, and accurate qualitative analysis can be obtained by matching with the database using the proposed identification criteria. Limit of detections (LODs) were 1.34 × 10-4 âˆ¼ 1.92 ng/mL and 1.92 × 10-4 âˆ¼ 9.80 ng/mL for blood and urine samples, respectively. Limits of quantification (LOQ) of toxins in blood and urine ranged from 4.47 × 10-4 âˆ¼ 6.32 ng/mL and 6.39 × 10-4 âˆ¼ 32.67 ng/mL, respectively. Intra-day relative standard deviations (RSDs) were 0.79 % âˆ¼ 10.90 %, and inter-day RSDs were 1.08 % âˆ¼ 18.93 %. The recoveries can reach 90 % âˆ¼ 110 % with matrix matching calibration curves. CONCLUSION: The established method is simple and rapid to operate, which can complete the sample analysis within 30 min, providing technical support for clinical poisoning treatment and public health poisoning analysis.

Endogenous H2S-activated Ag nanoparticles embedded in programmed DNA-cubes for specific visualization of colorectal cancer cells.

To avoid the unexpected aggregation and reduce the cytotoxicity of nanomaterials as optical probes in cell imaging applications, we propose a programmed DNA-cube as a carrier for silver nanoparticles (Ag NPs) to construct a specific hydrogen sulfide (H2S) responsive platform (Ag NP@DNA-cube) for diagnosing colorectal cancer (CRC) in this study. The DNA-cube maintains good dispersion of Ag NPs while providing excellent biocompatibility. Based on the characteristic overexpression of endogenous H2S in CRC cells, the Ag NPs are etched by H2S within target cells into silver sulfide quantum dots, thereby selectively illuminating the target cells. The Ag NP@DNA-cube exhibits a specific fluorescence response to CRC cells and achieves satisfactory imaging.

The Efficacy and Safety of Shashen-Maidong Decoction Combined with Western Medicine in the Treatment of Omicron Infected Individuals Over 85 Years Old: A Retrospective Study.

Background: During the Omicron variant outbreak of COVID-19 (2022-2023), Chinese healthcare institutions combined traditional Chinese medicine (TCM) with Western medical practices to treat COVID-19 patients, especially the elderly. The efficacy and safety of this approach, especially for individuals aged over 85, need further investigation. Methods: In this retrospective study, a cohort of 62 patients aged over 85 years, diagnosed with COVID-19 infection, was examined. Among them, 34 patients were administered Shashen-Maidong decoction in conjunction with Western medicine (SMD+WM group), while the remaining 28 patients received only Western medicine (WM group). Comparative analysis was conducted between the two groups, encompassing parameters such as the duration for the nucleic acid test to turn negative, length of intensive care unit (ICU) stay, mortality rate, utilization of high-flow nasal cannula oxygen (HFNC), occurrence of endo-tracheal intubation, frequency of recurrent respiratory infections within three months, and various laboratory indicators. Results: There were no significant differences observed between the two groups in terms of the duration for the nucleic acid test to turn negative, the length of ICU stay, mortality rate, utilization of HFNC, performance of endo-tracheal intubation, or the frequency of recurrent respiratory infections within three months (P > 0.05). However, in comparison to the WM group, the SMD+WM group exhibited notably lower growth rates in white blood cell (WBC) and neutrophil (NEUT) values. Additionally, the SMD+WM group demonstrated superior improvement in cardiac troponin I (cTnI) and B-type natriuretic peptide (BNP) values. Conclusion: In contrast to the administration of Western medicine alone, the combined use of Shashen-Maidong decoction with Western medicine significantly suppresses the increase in WBC count, particularly in NEUT levels, in elderly patients diagnosed with COVID-19. Moreover, this combined treatment exhibits a protective effect on cardiac function and demonstrates a relatively safe profile.

Study on potential antigenicity and functional properties of whey protein treated by high hydrostatic pressure based on structural analysis.

High hydrostatic pressure (HHP) is extensively utilized in the field of food processing due to its remarkable ability to preserve the freshness of food. The potential antigenicity of ß-lactoglobulin (ß-LG) in whey protein isolate (WPI, 3%) treated by HHP was detected by enzyme linked immunosorbent assay (ELISA) using monoclonal antibodies. Furthermore, the impact of pressure-induced structural alterations on the emulsification properties and antioxidant activity of WPI was investigated. The findings revealed that pressures exceeding 300 MPa resulted in molecular aggregation, the formation of inter-molecular disulfide bonds, and an increase in surface hydrophobicity (H0). The percentage of ß-sheet decreased along with the pressure. The results showed the increment of α-helix and ß-turn with pressure. ELISA demonstrated a significant reduction in the antigenicity of ß-LG following HHP treatment (100-600 MPa), with a slight recovery observed at 300 MPa. These spatial structural modifications led to the unfolding of the ß-LG molecule, thereby enhancing its digestibility. Moreover, HHP treatment substantially improved the antioxidant properties, with the exposure to hydrophobic amino acids contributing to increased antioxidant properties and emulsion stability.

Fe-loaded alginate hydrogel beads activating peroxymonosulfate for enhancing anaerobic fermentation of waste activated sludge: Performance and potential mechanism.

The recovery of volatile fatty acids (VFAs) through anaerobic fermentation (AF) is usually restricted by the poor biodegradability of waste activated sludge (WAS). This study proposed a novel strategy, i.e. peroxymonosulfate (PMS) activated by Fe-loaded sodium alginate hydrogel beads (Fe-SA), to enhance AF performance. Experimental results demonstrated that the as-synthesized Fe-SA and PMS co-pretreatment synergistically enhanced WAS solubilization and VFAs production. The maximal VFAs yield of 2013 mg COD/L was achieved at the Fe-SA dosage of 4.0 mM/g TSS, which was 93.7% higher than that with sole PMS addition and 8.82 times higher than that of the control. Mechanistic studies elucidated that the generation of reactive radicals such as SO4•- and •OH from PMS was greatly induced by Fe-SA, which contributed to WAS disintegration and degradation of refractory compounds. Additionally, analysis of the key enzyme activities indicated that the Fe-SA could strengthen biological hydrolysis and acidogenesis of sludge during AF. Microbial analysis illustrated that Fe-SA evidently improved the abundances of fermentative microorganisms as well as functional gene expression via creating a favorable environment for microbial growth. This study demonstrated the applicable potential of Fe-SA hydrogel beads activating PMS for VFAs production and provides an important reference for developing advanced oxidation processes-based application in AF.

Purification and identification of xanthine oxidase inhibitory peptides from enzymatic hydrolysate of α-lactalbumin and bovine colostrum casein.

The development of food-derived Xanthine Oxidase (XO) inhibitors is critical to the treatment of hyperuricemia and oxidative stress-related disease. Few studies report on milk protein hydrolysates' XO inhibitory activity, with the mechanism of their interaction remaining elusive. Here, different commercial enzymes were used to hydrolyze α-lactalbumin and bovine colostrum casein. The two proteins hydrolyzed by alkaline protease exhibited the most potent XO inhibitory activity (bovine casein: IC50 = 0.13 mg mL-1; α-lactalbumin: IC50 = 0.28 mg mL-1). Eight potential XO inhibitory peptides including VYPFPGPI, GPVRGPFPIIV, VYPFPGPIPN, VYPFPGPIHN, QLKRFSFRSFIWR, LVYPFPGPIHN, AVFPSIVGR, and GFININSLR (IC50 of 4.67-8.02 mM) were purified and identified from alkaline protease hydrolysates by using gel filtration, LC-MS/MS and PeptideRanker. The most important role of inhibiting activity of peptides is linked to hydrophobic interactions and hydrogen bonding based on the results of molecular docking and molecular dynamics simulation. The enzymatic hydrolysate of α-lactalbumin and bovine colostrum casein could be a competitive candidates for hyperuricemia-resisting functional food.

Black carp RNF5 inhibits STING/IFN signaling through promoting K48-linked ubiquitination and degradation of STING.

Ubiquitination is one of the important post-translational modifications (PTMs) of proteins that plays a vital role in regulating substrate degradation to ensure cellular homeostasis. Ring finger protein 5 (RNF5) is an essential E3 ubiquitin ligase for inhibiting STING-mediated interferon (IFN) signaling in mammals. Nevertheless, the function of RNF5 in STING/IFN pathway remains obscure in teleost. Here, we reported that over-expression of black carp RNF5 (bcRNF5) inhibited STING-mediated transcription activity of bcIFNa, DrIFNφ1, NF-κB and ISRE promoters and antiviral activity against SVCV. Moreover, knockdown of bcRNF5 increased the expression of host genes, including bcIFNa, bcIFNb, bcILß, bcMX1 and bcViperin, and also enhanced the antiviral capability of host cells. Immunofluorescence (IF) and Co-immunoprecipitation (Co-IP) assay confirmed that bcRNF5 was mainly localized in the cytoplasm and interacted with bcSTING. The expression level of bcSTING protein was attenuated by co-expressed bcRNF5 and MG132 treatment rescued this attenuating effect, suggesting that bcRNF5-mediated bcSTING degradation was dependent on the proteasome pathway. Subsequent, Co-IP and immunoblot (IB) experiments identified that bcRNF5 triggered the K48-linked but not K63-linked ubiquitination of bcSTING. Altogether, above results conclude that RNF5 suppresses STING/IFN signaling by enhancing K48-linked ubiquitination and protease degradation of STING in black carp.

Emerging strategies of engineering retinal organoids and organoid-on-a-chip in modeling intraocular drug delivery: Current progress and future perspectives.

Retinal diseases are a rising concern as major causes of blindness in an aging society; therapeutic options are limited, and the precise pathogenesis of these diseases remains largely unknown. Intraocular drug delivery and nanomedicines offering targeted, sustained, and controllable delivery are the most challenging and popular topics in ocular drug development and toxicological evaluation. Retinal organoids (ROs) and organoid-on-a-chip (ROoC) are both emerging as promising in-vitro models to faithfully recapitulate human eyes for retinal research in the replacement of experimental animals and primary cells. In this study, we review the generation and application of ROs resembling the human retina in cell subtypes and laminated structures and introduce the emerging engineered ROoC as a technological opportunity to address critical issues. On-chip vascularization, perfusion, and close inter-tissue interactions recreate physiological environments in vitro, whilst integrating with biosensors facilitates real-time analysis and monitoring during organogenesis of the retina representing engineering efforts in ROoC models. We also emphasize that ROs and ROoCs hold the potential for applications in modeling intraocular drug delivery in vitro and developing next-generation retinal drug delivery strategies.

Nanoemulsion: A novel delivery approach for thermosensitive IgG on inhibiting milk fat oxidation.

IgG, a biologically active substance in bovine colostrum, is easily inactivated during heat treatment and edible process to lose its biological activity. Nanoemulsion can effectively protect IgG to maintain its biological activity from injurious treatment. In this study, a food-grade nanoemulsion system was developed to protect IgG from heat and acid damage. It can be found that the residual rate of nanoemulsion-protected IgG reaches 87.1 % after 10 min at 72 °C. After 5 min at 82 °C, the residual rate of IgG in nanoemulsion was 18.7 % higher than that in PBS. In the simulated gastric fluid at pH 2.0, the residual rate of IgG in the nanoemulsion reacted for 4 h was 21.5 % higher than that in PBS. It indicated that nanoemulsion system can improve the heat and acid resistance of IgG compared with others, which is attributed to the lowest water activity of nanoemulsion. The contents of hydroperoxide and malondialdehyde in the milk after storage for 72 h with nanoemulsion-protected IgG were 0.12 meq/kg and 0.04 mg/kg, respectively, less than that of PBS-protected IgG. IgG is protected by nanoemulsion can effectively protect its activity during processing, which provides a theoretical basis for its direct application in liquid milk.

Spatial Confinement of Single-Drop System to Enhance Aggregation-Induced Emission for Detection of MicroRNAs.

Due to high incidence, poor prognosis, and easy transformation into pancreatic cancer (PC) with high mortality, early diagnosis and prevention of acute pancreatitis (AP) have become significant research focuses. In this work, we proposed a magnetic single-drop microextraction (SDME) system with spatial confinement to enhance the aggregation-induced emission (AIE) effect for simultaneous fluorescence detection of miRNA-155 (associated with AP) and miRNA-196a (associated with PC). The target miRNAs were selectively recognized by the hairpin probe and triggered the DNA amplification reaction; then, the DNA strands with two independent probes of G-quadruplex/TAIN and Cy5 were constructed on the surfaces of the magnetic beads. The SDME process, in which a drop containing the fluorescence probes was formed at the tip of the magnetic microextraction rod rapidly within 10 s, was performed by magnetic extraction. In this way, G-quadruplex/TAIN was enriched owing to the spatial confinement of the single-drop system, and the fluorescence signal given off (by G-quadruplex/TAIN) was highly enhanced (AIE effect). This was detected directly by fluorescence spectrophotometry. The approach achieved low limits of detection of 2.1 aM for miRNA-196a and 8.1 aM for miRNA-155 and wide linear ranges from 10 aM to 10 nM for miRNA-196a and from 25 aM to 10 nM for miRNA-155. This novel method was applied to the fluorescence detection of miRNAs in human serum samples. High relative recoveries from 95.6% to 104.8% were obtained.

Effect of ferulic acid covalent conjugation on the functional properties and antigenicity of ß-lactoglobulin.

Binding to phenolics can improve the functional properties of proteins. Changes in structure, functional properties, and antigenicity of ß-lactoglobulin (ß-LG) after covalent conjugation with ferulic acid (FA) at different mass ratios were reported here. The results of SDS-PAGE and gel exclusion chromatography confirmed that covalent complexes were formed. When the mass ratio of ß-LG and FA was 10:6, the binding content of FA was the highest. Fluorescence spectroscopy, UV-visible absorption spectrometry, and FTIR analysis showed that the structure of the complexes was more stretched compared to native ß-LG. The addition of FA significantly improved the emulsifying property of ß-LG. When the mass ratio was 10:6, the radical scavenging activities of DPPH and ABTS reached 65.06% and 88.22%, respectively, and the antigenicity of ß-LG reduced by about 35%. This study provides novel ß-LG-FA complexes in food systems to reduce the antigenicity of ß-LG and improve functional properties.

Vanillic Acid as a Promising Xanthine Oxidase Inhibitor: Extraction from Amomum villosum Lour and Biocompatibility Improvement via Extract Nanoemulsion.

Gout is an oxidative stress-related disease. Food-derived vanillic acid, a promising xanthine oxidase inhibitor, could potentially be used as a safe, supportive, and therapeutic product for gout. The extraction of vanillic acid from a classic Chinese herbal plant Amomum villosum with ethanol was investigated in the study. The optimum conditions were determined as extraction time of 74 min, extraction temperature of 48.36 °C, and a solid-to-liquid ratio of 1:35 g·mL-1 using the Box-Behnken design (BBD) of response surface methodology (RSM). The experimental extraction yield of 9.276 mg·g-1 matched with the theoretical value of 9.272 ± 0.011 mg·g-1 predicted by the model. The vanillic acid in Amomum villosum was determined to be 0.5450 mg·g-1 by high-performance liquid chromatography-diode array detection (HPLC-DAD) under the optimum extraction conditions and exhibited xanthine oxidase (XO) inhibitory activity, with the half-maximal inhibitory concentration (IC50) of 1.762 mg·mL-1. The nanoemulsion of Amomum villosum extract consists of 49.97% distilled water, 35.09% Smix (mixture of tween 80 and 95% ethanol with 2:1 ratio), and 14.94% n-octanol, with a particle size of 110.3 ± 1.9 nm. The nanoemulsion of Amomum villosum extract exhibited markable XO inhibitory activity, with an inhibition rate of 58.71%. The result demonstrated the potential benefit of Amomum villosum as an important dietary source of xanthine oxidase inhibitors for gout.