Diketopyrrolopyrrole-based fluorescent probe for visualizing over-expressed carboxylesterase in fever via ratiometric imaging.

Fever is the result of inflammation and the innate self-defense response of organisms, can cause abnormal changes in the activity of many enzymes in organisms, including the important carboxylesterase (CE). Monitoring the activity changes of CE in vivo during a fever will help to understand heat-related pathological mechanisms. In this paper, we designed diketopyrrolopyrrole-based ratiometric fluorescent probes DPP-FBC-P and DPP-FBO-P containing alkyl chain and diethylene glycol monomethyl ether chain respective for detection of CE. Both probes could realized fast response to CE and displayed good selectivity and high sensitivity. Compared with DPP-FBO-P, DPP-FBC-P had better biocompatibility, larger signal to noise ratio (225-fold vs 125-fold) and lower detection limit (1.6 × 10-5 U/mL vs 4.2 × 10-5 U/mL). Moreover, the probe DPP-FBC-P had been successfully applied to image the endogenous CE in HepG2 cells and solid tumors, and also visualized the over expressed CE in fever cells. Most importantly, the changes of CE level in the liver of fever mice model induced by LPS were monitored with the assistance of DPP-FBC-Pvia dual channel ratio imaging for the first time. In addition, fluorescence color signal in solution was captured by smart phone, and the linear relationship between RGB ratio (G/R) and CE concentration was established. This work will provide a potential approach for investigating the physiological and pathological processes of heat related diseases.

Preparation and properties of films loaded with cellulose nanocrystals stabilized Thymus vulgaris essential oil Pickering emulsion based on modified tapioca starch/polyvinyl alcohol.

Pickering emulsions were prepared by stabilizing thymus vulgaris essential oil (TEVO) with cellulose nanocrystals (CNCs), which formed composite films by loading the emulsions into modified tapioca/polyvinyl alcohol (PVA)-based films. The results showed that the 1.0 % CNCs-15 % TEVO emulsion had optimal stability and smaller particle size. The emulsion increased the thickness of the composite film in the form of solid material additions (thickness, 0.062-0.099 mm), which opacity given the laminating film's superior UV-blocking ability compared to blank film. The emulsion plasticizing effect enhanced the film's elongation at break (EAB, 123-159 %). In addition, due to the hydrophobicity and influencing the diffusion path of water molecules in the emulsion, the denser microstructure composite film had a lower water vapor transmission coefficient (WVP, 6.22 × 10-11-5.35 × 10-11g∙cm/cm2∙s∙Pa) to impede moisture penetration. Meanwhile, the composite film can effectively maintain the color and inhibit the growth of microorganisms to extend the storage time of fish fillets.

DNA hypomethylation-associated transcriptional rewiring enables resistance to heavy metal mercury (Hg) stress in rice.

Mercury (Hg) is an important hazardous pollutant that can cause phytotoxicity and harm human health through the food chain. Recently, rice (Oryza sativa L.) has been confirmed as a potential Hg bioaccumulator. Although the genetic and molecular mechanisms involved in heavy metal absorption and translocation in rice have been investigated for several heavy metals, Hg is largely neglected. Here, we analyzed one Hg-resistant line in rice (RHg) derived from a DNA methyltransferase-coding gene, OsMET1-2 heterozygous mutant. Compared with its isogenic wild-type (WT), RHg exhibited a significantly higher survival rate after Hg treatment, ameliorated oxidative damage, and lower Hg uptake and translocation. RNAseq-based comparative transcriptomic analysis identified 34 potential Hg resistance-related genes involved in phytohormone signaling, abiotic stress response, and zinc (Zn) transport. Importantly, the elevated expression of Hg resistance-related genes in RHg was highly correlated with DNA hypomethylation in their putative promoter regions. An ionomic analysis unraveled a negative correlation between Zn and Hg in roots. Moreover, Hg concentration was effectively decreased by exogenous application of Zn in Hg-stressed rice plants. Our findings indicate an epigenetic basis of Hg resistance and reveal an antagonistic relationship between Hg and Zn, providing new hints towards Hg detoxification in plants. ENVIRONMENTAL IMPLICATION: Mercury (Hg) as an important hazardous pollutant adversely impacts the environment and jeopardizes human health, due to its chronicity, transferability, persistence, bioaccumulation and toxicity. In this paper, we identified 34 potential genes that may significantly contribute to Hg resistance in rice. We find the expression of Hg resistance-related genes was highly correlated with DNA hypomethylation in their putative promoter regions. Our results also revealed an antagonistic relationship between Hg and Zinc (Zn), providing new hints towards Hg detoxification in plants. Together, findings of this study extend our current understanding of Hg tolerance in rice and are informative to breed seed non-accumulating rice cultivars.

Selective photocatalytic semihydrogenation of alkynols to alkenols on Pd-C3N4 nanosheets under ambient conditions.

Selective hydrogenation of alkynols to alkenols is an essential process for producing fine and intermediate chemicals. Currently, thermocatalytic alkynol hydrogenation faces several challenges, e.g., the safety of high-pressure hydrogen (H2) gas and the need for elevated temperature, and unavoidable side reactions, e.g., overhydrogenation. Here, a novel photocatalytic strategy is proposed for selectively reducing alkynols to alkenols with water as a hydrogen source under ambient temperature and pressure. Under the irradiation of simulated solar light, carbon nitride (C3N4) nanosheets with palladium (Pd) nanoparticles as cocatalysts (Pd-C3N4 NSs) exhibit a 2-methyl-3-butyn-2-ol (MBY) conversion of 98% and 2-methyl-3-buten-2-ol (MBE) selectivity of 95%, outperforming state-of-the-art thermocatalysts and electrocatalysts. After natural-sunlight irradiation (average light intensity of 25.13 mW cm-2) for 36 h, a MBY conversion of 98% and MBE selectivity of 92% was achieved in a large-scale photocatalytic system (2500 cm2). Experimental and theoretical investigations reveal that Pd cocatalysts on C3N4 facilitate the adsorption and hydrogenation of MBY as well as the formation of active hydrogen species, which promote the selective semihydrogenation of alkynols. Moreover, the proposed strategy is applicable to various water-soluble alkynols. This work paves the way for photocatalytic strategies to replace thermocatalytic hydrogenation processes using pressurized hydrogen.

Cooperative adsorption of interfacial Ga-N dual-site in GaOOH@N-doped carbon nanotubes for enhanced electrocatalytic reduction of carbon dioxide.

To reduce activation energy barrier and promote the kinetics of electrocatalytic CO2 reduction reaction (eCO2RR), the performance of CO2 adsorption and activation on electrocatalysts should be optimized. Here, GaOOH is successfully coupled with N-doped carbon nanotubes (NC) via a facile self-assembly-calcination process. The obtained GaOOH@N-doped carbon nanotubes (Ga-NC) display the best CO faradaic efficiency of 96.1 % at -0.6 V (vs. reversible hydrogen electrode). Control-experiment and characterization results suggest Ga-N dual-site in interface between GaOOH and NC shows cooperative adsorption of CO2. C atom in CO2 is adsorbed on N site while O atom in CO2 is adsorbed on Ga site. This cooperative adsorption efficiently promotes the CO2 adsorption and activation performance, as well as the breaking of CO bond due to opposite attraction from Ga-N dual-site. Moreover, in-situ Fourier transform infrared spectroscopy confirms decreased reaction barrier for formation of *CO2- and *COOH intermediates. This work inspires us to construct interfacial dual-site structure with cooperative adsorption property for promoting eCO2RR activity.

The environmental risk assessment of constructed wetlands filled with iron and manganese ores in typical antibiotic treatment.

Constructed wetlands (CWs) is considered as an efficient and environmentally friendly technology for advanced wastewater treatment to eliminate organic pollutants such as sulfamethoxazole (SMX) and trimethoprim (TMP). Iron (Fe) and manganese (Mn) ores have attracted more and more attention as CWs substrates in treating SMX and TMP, but the potentially negative environmental effects of wetland effluents, ore contaminants leached from the substrates and the risk of transmission of antibiotic resistance genes (ARGs) are still not clear. Three CW groups with different substrates (river sand (C-CW), Fe ore (Fe-CW), and Mn ore (Mn-CW)) were set up to evaluate the average removal rates and environmental risk in treating wastewater containing SMX and TMP. The results showed that the average removal rates of SMX and TMP by Fe-CW and Mn-CW were significantly higher than that of C-CW by 12.46%, 6.59% and 38.93%,15.39% respectively (p < 0.05), suggesting that both Fe and Mn ores facilitated the removal of antibiotics. However, the least abundance of ARGs was found in the layer of Fe ore at the middle layer (ML) in Fe-CW among all CWs, which suggested that Fe ore could reduce the risk of ARGs transmission. Although the environmental risk of Fe-CW and Mn-CW effluent was low, Fe-CW effluent inhibited the growth of Chlorella in both 48h and 72h experiments, while Mn-CW effluent showed an inhibitory effect in 48h and then promoted the growth in 72h. Meanwhile, these findings offer valuable insights for wetland health assessment and substrate selection for CWs.

Whole-body aging mediates the association between exposure to volatile organic compounds and osteoarthritis among U.S. middle-to-old-aged adults.

BACKGROUND: Humans are constantly exposed to various volatile organic compounds (VOCs) because of their widespread sources and characteristic of easy evaporation. Existing evidence regarding the association between VOC exposure and osteoarthritis (OA) risk is limited. PURPOSE: This study aimed to investigate the associations between individual urinary VOC metabolites (VOCMs) and the VOCM mixture, representing internal exposure levels of VOCs, with prevalent OA risk and to explore the mediating effect of aging and oxidative stress (OS) in these associations. METHODS: Data from the National Health and Nutrition Examination Surveys 2005-2020 were analyzed. Weighted generalized linear regression was employed to explore the associations between individual VOCMs and OA risk, as well as aging and OS biomarkers. A five-repeated ten-fold cross-validation elastic net model was used to identify critical VOCMs for the weight quantile sum (WQS) analysis, which was performed to explore the VOCM mixture and OA risk association. Parallel and serial mediation analyses were conducted to identify the potential mediators and mediation pathways. RESULTS: This study included 6578 American adults aged ≥40 years, among whom 1052 (16.0 %) individuals reported prevalent OA. Urinary levels of N-acetyl-S-(benzyl)-L-cysteine, mandelic acid and phenylglyoxylic acid were positively associated with OA risk. Eleven VOCMs with nonzero coefficients were identified and included in the WQS analysis, and results revealed an average increase of 24.4 % in OA risk (OR = 1.244, 95 % CI: 1.041, 1.486) per one-quantile increment in the VOCM mixture. Two aging biomarkers, phenotypic age and biological age, parallelly mediated the association between the VOCM mixture and OA risk, with mediation effect proportions of 9.0 % and 16.4 %, respectively. CONCLUSIONS: Exposure to VOCs is associated with an increased OA risk in middle-to-old aged American adults. The mediating effect of aging contributes to the association between co-exposure to VOCs and OA risk. Further prospective studies are required to substantiate these findings.

Imaging hydrological dynamics in karst unsaturated zones by time-lapse electrical resistivity tomography.

The hydrodynamics of karst terrain are highly complex due to the diverse fractures and reservoirs within limestone formations. The time delay between rainfall events and subsequent flow into reservoirs exhibits significant variability. However, these hydrological processes are not easily visualized in karst topography. Subsurface geophysics, specifically 2D time-lapse electrical resistivity tomography (ERT), provides an effective method for studying the relationships between hydrological and geophysical features. In our research, we adopted ERT in the Karst Critical Zone (KCZ) to visualize specific karstic zones, including cave galleries, water storage reservoirs, wetting fronts, soil layers, and potential preferential flow paths down to a depth of 20 m. To capture spatial and seasonal variations in resistivity, we presented a comprehensive approach by combining sixteen inversion models obtained between February 2020 and September 2022 above the Villars Cave in SW-France-a well-known prehistoric cave. We used a multi-dimensional statistical technique called Hierarchical Agglomerative Clustering (HAC) to create a composite model that divided the synthetic ERT image into eight clusters representing different karst critical zones. The ERT image clearly visualized the cave gallery with high resistivity values that remained consistent throughout the seasons. Our analysis revealed a close seasonal relationship between water excess and resistivity variations in most infiltration zones, with time delays increasing with depth. The karst reservoirs, located at significant depths compared to other clusters, displayed sensitivity to changes in water excess but were primarily affected by fluctuations in water conductivity, particularly during summer or dry periods. These findings have significant implications for predicting rainwater infiltration pathways into caves, thereby assisting in the conservation and preservation of prehistoric caves and their cultural heritage.

Highly efficient nitrate removal in sulfur-based constructed wetlands: Microbial mechanisms and environmental risks.

Nitrate is widely distributed in groundwater, posing an increasing threat to both water resources and human health. In this study, the treatment performance, removal mechanisms and environmental risks of sulfur-based constructed wetlands (CWs) for purifying nitrate-contaminated groundwater were investigated. Results showed that sulfur-based CWs could achieve the highest nitrate removal (95%). However, sulfate was largely produced as a by-product in sulfur-based CWs, which declined the nitrogen and phosphorus assimilation by plants. Metagenomic analysis indicated that autotrophs denitrifiers (e.g., Thiobacillus) were enriched, and the abundance of nitrate removal genes was enhanced in sulfur-based CWs. Additionally, sulfur cycle was formed in sulfur-based CWs, which explained the highest nitrate removal reasonably. This study provides comprehensive insights into the nitrate removal mechanisms in sulfur-based CWs and the associated environmental risks in purifying the polluted groundwater.

Contribution of zeolite to nitrogen retention in chicken manure and straw compost: Reduction of NH3 and N2O emissions and increase of nitrate.

Co-composting of chicken manure, straw and zeolite was investigated in a water bath heating system to estimate the effect of zeolite on physicochemical properties and metabolic functions related to nitrogen conversion. The results indicated that NH3 catches by zeolite was concentrated in the early stage and zeolite with 10 % addition reduced 28 % NH3 and 55 % N2O emissions as compost ended. The nitrate content in 10 % zeolite group was 17 % higher than that in control group. There was no significant increase of NO2- in zeolite group. More NO2- formed NH3, rather than being converted to NOx through denitrification. The abundance of nitrification genes amoA and hao increased except nxrA in zeolite groups. Denitrification was the most obvious at 20 d and zeolite decreased the abundance of denitrification genes narG, nirK and nosZ at this time.

FADH2-mediated radical polymerization amplification for microRNA-21 detection.

For early diagnosis of disease, ultrasensitive mircoRNA-21 detection has considerable potential. In this paper, an ultra-sensitive fluorescence detection method for microRNA was developed by atom transfer radical polymerization (ATRP). This ATRP reaction was first initiated by using flavin mononucleotide (FADH2). The DNA probe 1 modified with amino group was fixed on the magnetic nanoparticle Fe3O4, and microRNA-21 was added to form the probe 1-microRNA-21. Another carboxy-modified DNA 2 forms a sandwich structure with the bound microRNA-21. Two terminally modified DNA types are used as microRNA probes, using complementary base pairing to form a stable super-sandwich structure between the DNA probe and the microRNA. Under optimal conditions, microRNA was detected in PBS buffer with a detection limit of 0.19 fM. And even in 10% of human serum, microRNA-21 can be detected with a detection limit of 47.8 fM. Results show that this method has high selectivity, efficiency and stability, which broad application prospect in microRNA ultra-sensitive detection.

Motor cortex repetitive transcranial magnetic stimulation in major depressive disorder - A preliminary randomized controlled clinical trial.

BACKGROUND: Repetitive transcranial magnetic stimulation (rTMS) at left dorsolateral prefrontal cortex (lDLPFC) is commonly used in major depressive disorder (MDD), even though its therapeutic efficacy is limited. Given that many MDD patients show psychomotor retardation, we aim to examine whether the left motor cortex (lMC) as a novel rTMS target would provide effective and well-tolerated treatment as being comparable to lDLPFC-rTMS. METHODS: In this prospective double-blind randomized single-center study, 131 MDD patients were randomly assigned to the lDLPFC or lMC group and were treated with 10 Hz rTMS (90 % motor threshold) applied twice daily for 4000 pulses continuously over five days. The primary endpoint was the Hamilton Depression Scale (HAMD) total score change after treatment. RESULTS: After the five-day rTMS treatment, there was no significant difference in both HAMD reduction rate (lDLPFC 59.3 % ± 20.4 %, lMC 51.3 % ± 26.3 %, P = 0.10) and adverse effects (P = 0.79) between 48 (73.8 %) lMC subjects and 51 (77.3 %) lDLPFC subjects. Furthermore, the lMC study group showed stable HAMD scores at follow-up compared to their endpoint scores (P = 0.08). LIMITATIONS: Sham-control group was not included and the sample size was small. Therefore, our results should be seen as exploratory and preliminary. CONCLUSIONS: The preliminary good therapeutic response, comparability, and tolerability of lMC-rTMS suggest lMC a potential and more easily accessible rTMS target. Together, our findings raise the possibility of symptom-specific rTMS in motor cortex (psychomotor retardation) or lDLPFC (cognitive deficits). This warrants larger clinical trials of rTMS in MDD with symptom-specific stimulation targets.

Highly efficient determination of trace ascorbic acid in vitamin C tablets by boronate affinity-modified magnetic metal-organic frameworks.

Ascorbic acid (AA) plays an important role in many life processes. The chronic nutritional deficiency of AA will lead to the symptoms of scurvy. Therefore, the sensitive quantitative detection of AA is most important in the pharmaceutical analysis, food industry and diagnostic application. In this study, a dual-functional magnetic metal-organic frameworks (Fe3O4@SiO2@UiO-PBA) nanoparticles was synthesized by modifying phenylboronic acid to the surface of magnetic UiO-66-NH2 via postsynthetic modification for selectively and sensitively florescent detection of AA. Due to the abundant amino groups and grafted phenylboronic acid, the proposed nanoparticles have the dual properties of hydrophilicity and boronate affinity. Under optimum conditions, the obtained Fe3O4@SiO2@UiO-PBA nanoparticles can detect AA within 30 s, and has a good linear relationship with the concentration of AA in the range of 5.0-60 µM with a detection limit of 2.5 µM (S/N = 3). In addition, the prepared Fe3O4@SiO2@UiO-PBA nanoparticles showed excellent selectivity and great potential application in the highly efficient determination of trace AA in vitamin C tablets. These results indicated that a convenient method was proposed to develop fluorescent probes for rapid and sensitive detection of trace AA in real samples.

Accurate construction of NIR probe for visualizing HClO fluctuations in type I, type II diabetes and diabetic liver disease assisted by theoretical calculation.

Hypochlorous acid (HClO) is a key signaling molecule which involved in various pathological and physiological processes and the immune system. It had been proved that excess HClO in the organisms was closely associated with diabetes. In this paper, we constructed a series of BODIPY-based fluorophores modified with olefinic bond. With the assistance of theoretical calculations, the optimized near-infrared (NIR) dye BDP-ENE-S-Me, which possessed the longest wavelength (690 nm) and the best stability, was screened and synthesized. Based on BDP-ENE-S-Me, we further introduced N, N-dimethylcarbamate group to construct a NIR fluorescent probe BDP-ENE-ClO. BDP-ENE-ClO displayed excellent selectivity and sensitivity with a low detection limit (49 nM) towards HClO. Besides, the probe was successfully applied in monitoring concentration fluctuations of HClO in vitro and in vivo caused by various stimuli. Most importantly, the over-production of HClO in the type I, type II diabetes and diabetic liver disease mice models could be visualized and assessed precisely with the assistance of BDP-ENE-ClO. By comparing fluorescent intensity of diabetic mice models with that of diabetic liver disease mice models, the probe was competent to assess the progression of diabetes.

Vitamin B12-catalyzed electro-polymerization for ultrasensitive RNA detection.

Vitamin B12 being a natural catalyst in atom transfer radical polymerization (ATRP), has the advantages of mild reaction conditions, good biocompatibility and high catalytic efficiency. In this report, an electrochemical biosensor of the lung cancer biomarker microRNA-21 (miRNA-21) is designed for early screening of lung cancer with high sensitivity at the femtomolar level. In this approach, hairpin DNA with N3 end group was first attached to the electrode surface. When miRNA-21 was present and paired with hairpin DNA, the N3 group released and attached to the ATRP initiator through "click reaction". Through eATRP, a large number of FerrocenylMethyl Methacrylate (FcMMA) monomers polymerized into long chains for signal amplification. These long chains had a distinct electrical signal in the square wave voltammetry (SWV), which can detect RNA with high sensitivity. The limit of detection (LOD) goes down to 1.010 fM after ATRP polymerization, which is lower than that of the majority of other ultra-sensitive RNA electrochemical assays. Results also show that the vitamin B12-based electrochemical biosensor is highly selective and suitable for RNA detection in complex biological samples.

Effective sequential combined therapy with carboplatin and a CDC7 inhibitor in ovarian cancer.

BACKGROUND: The enhancement of DNA damage repair is one of the important mechanisms of platinum resistance. Protein cell division cycle 7 (CDC7) is a conserved serine/threonine kinase that plays important roles in the initiation of DNA replication and is associated with chemotherapy resistance in ovarian cancer. However, whether the CDC7 inhibitor XL413 has antitumor activity against ovarian cancer and its relationship with chemosensitivity remain poorly elucidated. METHODS: We evaluated the antitumor effects of carboplatin combined with XL413 for ovarian cancer in vitro and in vivo. Cell viability inhibition, colony formation and apoptosis were assessed. The molecules related to DNA repair and damage were investigated. The antitumor effects of carboplatin combined with XL413 were also evaluated in SKOV-3 and OVCAR-3 xenografts in subcutaneous and intraperitoneal tumor models. RESULTS: Sequential administration of XL413 after carboplatin (CBP) prevented cellular proliferation and promoted apoptosis in ovarian cancer (OC) cells. Compared with the CBP group, the expression level of RAD51 was significantly decreased and the expression level of γH2AX was significantly increased in the sequential combination treatment group. The equential combination treatment could significantly inhibit tumor growth in the subcutaneous and intraperitoneal tumor models, with the expression of RAD51 and Ki67 significantly decreased and the expression of γH2AX increased. CONCLUSIONS: Sequential administration of CDC7 inhibitor XL413 after carboplatin can enhance the chemotherapeutic effect of carboplatin on ovarian cancer cells. The mechanism may be that CDC7 inhibitor XL413 increases the accumulation of chemotherapy-induced DNA damage by inhibiting homologous recombination repair activity.

Field migration of veterinary antibiotics via surface runoff from chicken-raising orchard in responding to natural rainfalls.

Knowledge on runoff transport of manure-sourced antibiotics from farmland soil to aquatic environment is limited due to complexity of hydrological regime and pathways. This study monitored natural rainfalls in sloping orchard plots with free-range chickens, with an attempt to investigate the migration characteristics of typical antibiotics via surface runoff as well as the impact of manure presence. Results showed that rainstorms continuously carried away antibiotics in surface runoff and all target antibiotics including those with high affinities to soil were detected at the beginning of runoff production. Concentration of antibiotics was found to respond strongly to the instantaneous rainfall intensity, showing consistent fluctuations during rainfalls. Concentrations of sulfonamides and florfenicol were two orders of magnitude higher than that of tetracyclines and fluoroquinolones. Compared to the control without raising chickens, antibiotics migration was considerably increased with the increased runoff production due to soil surface changes caused by chicken activities. Additionally, dynamics of antibiotic concentration significantly correlated with variations of fluorescent DOM components. Chicken manure-derived DOM mainly contained tryptophan moiety, and laboratory fluorescence quenching test with 2D-COS analysis indicated that all antibiotics interacted more strongly and preferentially with tryptophan than humic-like species. Antibiotics bonded to manure DOM with an affinity corresponding to the significance level of their correlations. In this light, potential use of fluorescence indices based on the established correlations may provide a convenient tool for tracing runoff migration of antibiotics during rainfalls.

Solubility Enhancement and Antioxidation Maintenance of CBD Encapsulated in the P407-RUB Nano-micelle System.

AIM: The aim of this study is to improve the water solubility and stability of cannabidiol (CBD) utilizing micelle technology. MATERIALS AND METHODS: Mixing of rubusoside (RUB) with Poloxamer 407 (P407) was explored as the wall material for the preparation of CBD micelles. In this study, CBD-loaded mixed micelles (CBD-M) composed of P407 and RUB was successfully formed by self-assembly then solid were prepared by solvent evaporation. The saturated solubility of CBD-loaded micelles in water increased to 15.60 mg/mL, 1560-fold compared with its intrinsic solubility (0.01 mg/mL). The average size of CBD-M was 103 ± 2.66 nm and the encapsulation efficiency for CBD was 92.8 + 4.7 %, and 18.6 + 0.94% for drug-loading. RESULTS: The morphology and encapsulation of CBD-M were characterized by TEM, FI-IR, DSC and TG. The CBD-M solution was stable and CBD-M did not precipitate or leak after being diluted and centrifugated. Besides, CBD-M solution was stable for 6 months at 4°C and room temperature. In vitro antioxidant studies found that the antioxidant activity of CBD remained at the same level after micellization. CONCLUSION: These results suggest that CBD-M may be a promising and competitive formulation for the delivery of CBD, laying a foundation for improving the bioavailability in the future.

Neurotoxicity study of ethyl acetate extract of Zanthoxylum armatum DC. on SH-SY5Y based on ROS mediated mitochondrial apoptosis pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Zanthoxylum armatum DC. (ZADC) is a traditional medicinal plant with various pharmacological activities and is widely used in China, Japan, India, and other regions. Previous studies have revealed that the methanol extract of ZADC can cause neurotoxicity symptoms in rats, such as drooling, decreased appetite, decreased movement, and increased respiratory rate. However, the basis of these toxic substances and the mechanism of neurotoxicity remain unclear. AIM OF THE STUDY: To evaluate the effects of ZADC on nerve cells and their damage mechanisms and discuss the possible toxic substance basis. MATERIALS AND METHODS: The ethyl acetate extract of ZADC is obtained by extracting the methanol extract of ZADC with ethyl acetate. The Q-Orbitrap LC-MS/MS method was employed to analyze the chemical composition of the EA extract of ZADC. SH-SY5Y cells were incubated with different concentrations of the ethyl acetate extract of ZADC. The cytotoxicity of the extract was evaluated using CCK-8, LDH, and ROS assays, and the oxidative stress status of cells was assessed using MDA, GSH, and SOD. Cell apoptosis was detected using flow cytometry. Damage to mitochondrial function was evaluated by labeling mitochondria, ATP, and MMP with fluorescence. Cyto-C, Caspase-3, Caspase-9, Apaf-1, Bax, and reduced Bcl2 expression were measured to evaluate the activation of the mitochondrial apoptosis pathway. Finally, NAC intervention was used to detect changes in the relevant indicators. The activation of mitochondrial apoptosis pathway was evaluated by measuring Cyto-C, Caspase-3, Caspase-9, Apaf-1, and Bax and Bcl2 expression. Finally, NAC intervention was utilized to detect changes in the relevant indicators. RESULTS: After treating SY-SY5Y cells with EA extract from ZADC, cell viability decreased significantly, and the intracellular ROS level increased in a dose-dependent manner. Meanwhile, ZADC can cause cellular oxidative stress and increase MDA and SOD concentrations while decreasing GSH concentrations. It can also shorten the mitochondrial cristae and decrease the number of mitochondria. In contrast, it can reduce ATP synthesis in the mitochondria and mitochondrial membrane potential (MMP). Furthermore, it increased the apoptosis rate and the expression of Cyto-C, Caspase-3, Caspase-9, Apaf-1, and Bax and reduced Bcl2 expression. NAC intervention alleviated the reduction in SH-SY5Y cell survival and the accumulation of reactive oxygen species induced by the EA extract in ZADC. It also inhibits signaling pathways dominated by proteins, such as Cyto-C, reducing cell apoptosis and cytotoxicity. A total of 46 compounds were identified in the extracts. CONCLUSIONS: The results suggest that EA extract of ZADC can induce the mitochondrial apoptotic pathway by accumulating ROS in cells, leading to apoptosis. Antioxidants had a good inhibitory and protective effect against cell damage caused by the EA extract of ZADC. The neurotoxic components of ZADC may be organic acids and compounds containing amino groups.

Zanthoxylum armatum DC fruit ethyl acetate extract site induced hepatotoxicity by activating endoplasmic reticulum stress and inhibiting autophagy in BRL-3A models.

ETHNOPHARMACOLOGICAL RELEVANCE: Zanthoxylum armatum DC (Z. armatum) is renowned not only as a culinary spice but also as a staple in traditional ethnic medicine, predominantly in Southeast Asia and various other regions. Recent research has unveiled its multifaceted pharmacological properties, including anti-inflammatory, antibacterial, and toothache relief effects. Nonetheless, some studies have reported the potential toxicity of Z. armatum, emphasizing the need to further explore its toxicity mechanisms for safer application. AIM OF THE STUDY: This study investigated the effect and mechanism of hepatotoxicity in BRL-3A cells induced by Z. armatum. MATERIALS AND METHODS: The compounds of the ethyl acetate extract of Z. armatum (ZADC-EA) were identified by ultrahigh performance liquid chromatography coupled with quadrupole-orbitrap high resolution mass spectrometry (UPLC-Q-Orbitrap HRMS). The hepatotoxicity of the extract was evaluated by detecting cell viability, aspartate aminotransferase (AST), alanine aminotransferase (ALT), and lactate dehydrogenase (LDH) activity, and apoptosis. Endoplasmic reticulum stress, autophagy, and apoptosis were detected by Ad-mCherry-GFP-LC3B, flow cytometry, and Western blot to explore the mechanism of hepatotoxicity induced by ZADC-EA. RESULTS: UPLC-Q-Orbitrap HRMS analysis revealed the presence of compounds belonging to flavonoids, terpenoids, and alkaloids. The IC50 value of ZADC-EA was 62.43 µg/mL, the cell viability of BRL-3A decreased in a time-dose dependent manner, and the levels of AST, ALT, and LDH were upregulated. In addition, ZADC-EA-induced increased expression of eIF2α-ATF4-CHOP pathway proteins, inhibited autophagy, and promoted apoptosis. CONCLUSIONS: This study provides insights into the hepatotoxicity mechanisms of ZADC-EA on BRL-3A cells. It was found that ZADC-EA could induce endoplasmic reticulum stress and inhibit autophagy, then intensify apoptosis, and endoplasmic reticulum stress could exacerbate autophagy inhibition.