Urinary biomarkers for hepatocellular carcinoma: current knowledge for clinicians.

Hepatocellular carcinoma (HCC) is the most predominant primary liver cancer, causing many illnesses and deaths worldwide. The insidious clinical presentation, difficulty in early diagnosis, and the highly malignant nature make the prognosis of HCC extremely poor. The complex and heterogeneous pathogenesis of HCC poses significant challenges to developing therapies. Urine-based biomarkers for HCC, including diagnostic, prognostic, and monitoring markers, may be valuable supplements to current tools such as serum α-fetoprotein (AFP) and seem promising for progress in precision medicine. Herein, we reviewed the major urinary biomarkers for HCC and assessed their potential for clinical application. Molecular types, testing platforms, and methods for building multimolecule models in the included studies have shown great diversity, thus providing abundant novel tools for future clinical transformation and applications.

Identification of RASSF1A promoter hypermethylation as a biomarker for hepatocellular carcinoma.

BACKGROUND: RAS association domain family protein 1A (RASSF1A) promoter hypermethylation is suggested to be linked to hepatocellular carcinoma (HCC), but the results remained controversial. METHODS: We evaluated how RASSF1A promoter hypermethylation affects HCC risk and its clinicopathological characteristics through meta-analysis. Data on DNA methylation in HCC and relevant clinical data were also collected based on The Cancer Genome Atlas (TCGA) database to investigate the prognostic role of RASSF1A promoter hypermethylation in HCC. RESULTS: Forty-four articles involving 4777 individuals were enrolled in the pooled analyses. The RASSF1A promoter methylation rate was notably higher in the HCC cases than the non-tumor cases and healthy individuals, and was significantly related to hepatitis B virus (HBV) infection-positivity and large tumor size. Kaplan-Meier survival analysis revealed that HCC cases with RASSF1A promoter hypermethylation had worse outcomes. Receiver operating characteristic curves confirmed that RASSF1A promoter methylation may be a marker of HCC-related prognoses. CONCLUSIONS: RASSF1A promoter hypermethylation is a promising biomarker for the diagnosis of HCC from tissue and peripheral blood, and is an emerging therapeutic target against HCC.

Screening of potential probiotic lactic acid bacteria based on gastrointestinal properties and perfluorooctanoate toxicity.

The consumption of lactic acid bacteria capable of binding or degrading food-borne carcinogens may reduce human exposure to these deleterious compounds. In this study, 25 Lactobacillus strains isolated from human, plant, or dairy environments were investigated for their potential probiotic capacity against perfluorooctanoate (PFOA) toxicity. The PFOA binding, tolerance ability, and acid and bile salt tolerance were investigated and assessed by principal component analysis. Additionally, the effect of different pH levels and binding times was assessed. These strains exhibited different degrees of PFOA binding; the strain with the highest PFOA binding capability was Lactobacillus plantarum CCFM738, which bound to 49.40 ± 1.5 % of available PFOA. This strain also exhibited relatively good cellular antioxidative properties, acid and bile salt tolerance, and adhesion to Caco-2 cells. This study suggests that L. plantarum CCFM738 could be used as a potential probiotic in food applications against PFOA toxicity.

Comparison of the Wells score with the revised Geneva score for assessing suspected pulmonary embolism: a systematic review and meta-analysis.

The Wells score and the revised Geneva score are two most commonly used clinical rules for excluding pulmonary embolism (PE). In this study, we aimed to assess the diagnostic accuracy of these two rules; we also compared the diagnostic accuracy between them. We searched PubMed and Web of science up to April 2015. Studies assessed Wells score and revised Geneva score for diagnosis suspected PE were included. The summary area under the curve (AUC) and the 95 % confidence interval (CI) were calculated. Eleven studies were included in this meta-analysis. For Wells score, the sensitivity ranged from 63.8 to 79.3 %, and the specificity ranged from 48.8 to 90.0 %. The overall weighted AUC was 0.778 (95 % CI 0.740-0.818; Z = 9.88, P < 0.001). For revised Geneva score, the sensitivity ranged from 55.3 to 73.6 %. The overall weighted AUC was 0.693 (95 % CI 0.653-0.736; Z = 11.96, P < 0.001). 95 % CIs of two AUCs were not overlapped, which indicated Wells score was more accurate than revised Geneva score for predicting PE in suspected patients. Meta-regression showed diagnostic accuracy of these two rules was not related with PE prevalence. Sensitivity analysis by only included prospective studies showed the results were robust. Our results showed the Wells score was more effective than the revised Geneva score in discriminate PE in suspected patients.

Structural Features, Physicochemical Properties, and In Vitro Digestibility of the Starch-Lipid Complexes Formed between High Amylose Starch and Stearic Acid or Potassium Stearate.

Starch-lipid complexes were prepared from high amylose starch (HAS) with stearic acid (SA) or potassium stearate (PS) at different molar concentrations. The complexes (HAS-PS) formed between HAS and PS showed polyelectrolyte characteristics with ζ-potential ranging from -22.2 to -32.8 mV, and the electrostatic repulsion between anionic charges restricted the starch chain reassociation and facilitated the formation of V-type crystalline structures upon cooling. The hydrophobic effects enabled recrystallization of the SA, and the HAS-SA complexes exhibited weaker V-type crystalline structures than the HAS-PS complexes; both HAS-SA/PS complexes were of a similar "mass fractal" type, with a dimension varied from 2.15 to 2.96. The HAS-SA complexes had a considerable content of resistant starch (RS, 16.1~29.2%), whereas negligible RS was found in the HAS-PS complexes. The findings from the present study imply that the molecular order of starch chains and the macro-structures of starch particles are more important to regulate the digestibility of starch-lipid complexes than the crystalline structures.

A novel and sensitive method for simultaneous determination of 6 low-calorie bulk sweeteners by HPLC-ELSD.

A novel and sensitive method for the simultaneous analysis of six low-calorie bulk sweeteners (D-allulose, D-tagatose, D-mannitol, mycose, palatinose, and erythritol) without derivatisation was developed using high-performance liquid chromatography-evaporative light scattering detector (HPLC-ELSD). Chromatographic separations were carried out on a Zorbax Original NH2 (5 µm particle size, 250 mm×4.60 mm id, 70 Å) column with flow rate gradient elution with acetonitrile: water (80:20, v/v). Drift tube temperature was set at 50 ℃, the nebuliser carrier gas flow rate was 1.0 mL·min-1, and nitrogen pressure was regulated to 276 kPa with gain:3. The regression equation showed good linearity (R2 = 0.9985-0.9998) for all six low-calorie bulk sweeteners in the tested range (0.060-0.60 mg·mL-1). The limits of detection (LOD) for the six low-calorie bulk sweeteners ranged from 0.02 to 0.06 mg·mL-1. The proposed HPLC-ELSD method was validated for the quantification of the low-calorie bulk sweeteners in 14 types of foods, and the results were satisfactory. In addition, the results showed that the number of sweeteners in each food product varied. The presence of multiple low-calorie bulk sweeteners in certain foods is interesting. This method is successful in monitoring low-calorie bulk sweeteners in food.

Exploration of prognostic and treatment markers in hepatocellular carcinoma via GPCR-related genes analysis.

Background: G protein-coupled receptors (GPCRs), the biggest family of signaling receptors, account for 34 % of all the drug targets approved by the Food and Drug Administration (FDA). It has been gradually recognized that GPCRs are of significance for tumorigenesis, but in-depth studies are still required to explore specific mechanisms. In this study, the role of GPCRs in hepatocellular carcinoma (HCC) was elucidated, and GPCR-related genes were employed for building a risk-score model for the prognosis and treatment efficacy prediction of HCC patients. Methods: Patients' data on HCC were sourced from the Liver Hepatocellular Carcinoma-Japan (LIRI-JP) and The Cancer Genome Atlas (TCGA) databases, while GPCR-related genes were obtained from the Molecular Signatures Database (MSigDB). Univariant and multivariant Cox regression analyses, as well as least absolute shrinkage and selection operator (LASSO) were performed with the aim of identifying differentially expressed GPCR-related genes and grouping patients. Differential expression and functional enrichment analyses were performed; protein-protein interaction (PPI) mechanisms were explored; hub genes and micro ribonucleic acid (miRNA)-target gene regulatory networks were constructed. The tumor immune dysfunction and exclusion (TIDE) algorithm was utilized to evaluate immune infiltration levels and genetic variations. Sensitivity to immunotherapy and common antitumor drugs was predicted via the database Genomics of Drug Sensitivity in Cancer (GDSC). Results: A GPCR-related risk score containing eight GPCR-related genes (atypical chemokine receptor 3 (ACKR3), C-C chemokine receptor type 3 (CCR3), CCR7, frizzled homolog 5 (FZD5), metabotropic glutamate receptor 8 (GRM8), hydroxycarboxylic acid receptor 1 (HCAR1), 5-hydroxytryptamine receptor 5A (HTR5A) and nucleotide-binding oligomerization domain-like receptor family pyrin domain containing 6 (NLRP6)) was set up. In addition, patients were classified into groups with high and low risks. Patients in the high-risk group exhibited a worse prognosis but demonstrated a more favorable immunotherapy response rate compared with those in the low-risk group. Distinct sensitivity to chemotherapeutic drugs was observed. A clinical prediction model on the basis of GPCR-related risk scores was constructed. Areas under the curves (AUC) corresponding to one-, three- and five-year survival were 0.731, 0.765 and 0.731, respectively. Conclusions: In this study, an efficient HCC prognostic prediction model was constructed by only GPCR-related genes, which are all potential targets for HCC treatment.

[Effect of Biomass Burning on Carbonaceous Aerosol Composition and Light Absorption in Guangxi Regional Background Site].

Carbonaceous aerosols are an important component of fine particulate matter (PM2.5) in the atmosphere, having great impacts on air quality, human health, and the climate. In this study, PM2.5 samples were collected from November 2017 to October 2018 in a background site of Guangxi Province to investigate the potential impacts of biomass burning, an essential source of carbonaceous aerosols, on carbonaceous aerosols. Further, the composition of carbonaceous aerosols, sugar compounds, and the light absorption coefficient (babs) of water-soluble brown carbon (BrC) were also conducted. Considering the effect of the degradation of atmospheric levoglucosan (LG), the concentration of the corrected LG was quantified using the aging of air masses (AAM) index. Then, the contribution of biomass burning (BB) to organic carbon (OC) [BB-OC] was quantified using the corrected LG-derived molecular tracer method combined with the Bayesian mixing model. Here, we further explored the potential sources of water-soluble BrC using correlation analysis. In this research, the mean AAM index was 0.40±0.28 during the study period, indicating that the atmospheric LG had undergone a photochemical degradation process. The characteristic ratio combined with the Bayesian mixing model indicated that the crop straw (i.e., corn, rice, and sugarcane straw) was the dominant biomass fuel type in the Guangxi Region, contributing 22%, 23%, and 18% of OC without the correction of LG and 16%, 21%, and 17% with the corrected LG concentration, respectively. The neglection of LG degradation led to the underestimation of BB-OC, in which the BB-OC values with and without correction were 49.0% and 21.1%, respectively. Here, the annual mean babs of water-soluble BrC was (8.7±10.7) Mm-1, and its main sources were BB, fossil fuel combustion, and vegetation emission.

Pickering emulsion stabilized by parasin I and chitosan nanoparticles enhances protection against intestinal microbiota homeostasis by reducing inflammation in peritonitis mice.

Although various researches evaluated the stability and drug loading efficiency of chitosan Pickering emulsion, few studies assessed the role and mechanism of emulsions in gut flora homeostasis. Thus, in the basics of our previously published natural and antimicrobial Pickering emulsions, the function of emulsion on the intestinal microbiota and inflammation response was explored in Kunming mice with peritonitis. The results showed that lipid/peptide nanoparticles emulsion (LPNE) and the chitosan peptide-embedded nanoparticles emulsion (CPENE) presented less collagen fiber than parasin I in peritoneal tissue, and CPENE could reduce peritoneal inflammation by decreasing the expression of NOD-, LRR- and pyrin domain-containing protein 3 (NLRP3). The CPENE showed better histological morphology with a smaller fibrosis area in the spleen. Moreover, CPENE, LPNE, and parasin I-conjugated chitosan nanoparticle emulsion (PCNE) groups can increase the abundance of ABC transporters, DNA repair, and recombination proteins, and improve gut microbial. Furthermore, the Pickering emulsion showed a better protection effect on the composition and function of intestinal microbiota by decreasing interleukin-1ß secretion and assembly of the inflammasome of NLRP3. These results could provide evidence for intestinal microbiota homeostasis of chitosan Pickering emulsion in inflammation-related diseases.

Application and Progress of Cultured Models of Gallbladder Carcinoma.

Gallbladder carcinoma (GBC) is a malignant tumor of the biliary system that is aggressive, difficult to detect early, and has a low surgical resection rate and poor prognosis. Appropriate in vitro growth models are expected to focus on the study of the biological behavior and assess treatment effects. Nonetheless, cancer initiation, progression, and invasion include spatiotemporal changes and changes in the cell microenvironment intracellular communication, and intracellular molecules, making the development of in vitro growth models very challenging. Recent advances in biomaterial methods and tissue engineering, particularly advances in bioprinting procedures, have paved the way for advances in the creative phase of in vitro cancer research. To date, an increasing number of cultured models of gallbladder disease have emerged, such as two-dimensional (2D) GBC growth cell cultures, three-dimensional (3D) GBC growth cell cultures, xenograft models, and 3D bioprinting methods. These models can serve as stronger platforms, focusing on tumor growth initiation, the association with the microenvironment, angiogenesis, motility, aggression, and infiltration. Bioprinted growth models can also be used for high-throughput drug screening and validation, as well as translational opportunities for individual cancer therapy. This study focused on the exploration, progress, and significance of the development of GBC cultural models. We present our views on the shortcomings of existing models, investigate new innovations, and plan future improvements and application possibilities for cancer models.

Simultaneous Rapid Determination of Seven Alternaria Toxins in Tuberous Crops during Storage Using QuEChERS Coupled with Ultrahigh-Performance Liquid Chromatography-Tandem Mass Spectrometry.

Robust and sensitive ultrahigh-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) combined with the quick, easy, cheap, effective, rugged, and safe (QuEChERS) method was applied for the detection of seven Alternaria toxins (ATs) in tuberous crops. The influence of tuber conditions (fresh, germinated, and moldy) during storage on the concentration of the seven ATs is also investigated. ATs were extracted with acetonitrile under acidic conditions and purified with a C18 adsorbent. ATs were scanned with electrospray ionization (positive/negative ion) dynamic switching and detected in MRM mode. Calibration curve analysis results reveal good linear relationships in all toxin concentration ranges (R2 > 0.99). The limit of detection and limit of quantification were 0.25-0.70 and 0.83-2.31 µg/kg, respectively. The average recoveries of the seven ATs were 83.2-104% with intra-/inter-day precision at 3.52-6.55% and 4.02-7.26%, respectively. The developed method provided adequate selectivity, sensitivity, and precision in detecting the seven ATs at trace levels, and dispensed with standard addition or matrix-matched calibration to compensate for matrix effects. ATs in the fresh, germinated, and moldy samples of tuberous crops in storage (taro, potato, sweet potato, yam, cassava) were analyzed with this method, and the concentrations were 2.01-14.51 µg/kg and significantly increased with storage duration. ALS was detected in most samples, whereas no quantities of ALT and ATX-I were detected. AME was often detected in combination with AOH in sweet potatoes. TeA and Ten were mostly detected in taro, potato, and yam. The established method could be used for the simultaneous detection and quantification of multicomponent toxins in elaborate matrices.

Long-Term Retrogradation Properties and In Vitro Digestibility of Waxy Rice Starch Modified with Pectin.

In recent years, the blending of hydrocolloids and natural starch to improve the properties of natural starch has become a research hotspot. In this study, the effects of pectin (PEC) on the retrogradation properties and in vitro digestibility of waxy rice starch (WRS) were investigated. The results showed that PEC could significantly (p < 0.05) reduce the retrogradation enthalpy and reduce the hardness of WRS gel. X-ray diffraction results indicated that PEC could reduce the relative crystallinity of the composite system, and the higher the PEC content, the lower the relative crystallinity. When the PEC content was 10%, the relative crystallinity of the composite system was only 10.6% after 21 d of cold storage. Fourier transform infrared spectroscopy results proved that the interaction between PEC and WRS was mainly a hydrogen bond interaction. Furthermore, after 21 d of cold storage, the T23 free water signal appeared in the natural WRS paste, while only a small free water signal appeared in the compound system with 2% PEC addition. Moreover, addition of PEC could reduce the starch digestion rate and digestibility. When the content of PEC increased from 0% to 10%, the digestibility decreased from 82.31% to 71.84%. This study provides a theoretical basis for the further application of hydrocolloids in starch-based foods.

[Simultaneous determination of six rare sugars in solid foods by high performance liquid chromatography-evaporative light-scattering detection].

Excessive sugar consumption is associated with metabolic health problems. Rare sugars are gradually being used as substitutes for sugar, and their consumption is increasing daily, raising food-safety issues such as false advertising, adulteration, and overdosing. The determination of rare-sugar compounds has attracted considerable attention in recent years. However, no standard method for the simultaneous determination of six rare sugars (allulose, tagatose, trehalose, isomaltulose, erythritol, and mannitol) in solid foods is available. Therefore, establishing a suitable analytical method for these sugars is necessary. In this study, high performance liquid chromatography coupled with evaporative light-scattering detection was used to determine rare sugars in solid foods. The optimum chromatographic and detector conditions were determined by evaluating the instrument parameters. Analysis was carried out on a Zorbax Original NH2 column (250 mm×4.6 mm, 5 µm) via flow-rate gradient elution (0-15 min, 1.0 mL/min; 15-18 min, 1.0-2.0 mL/min; 18-25 min, 2.0 mL/min) with acetonitrile-water (80∶20, v/v) as the mobile phase. Sharp and symmetric chromatographic peaks were obtained under these conditions. The resolutions for all the six rare sugars were greater than 1.5. Optimization of the evaporative light-scattering detector was extremely important to the responses of the rare-sugar compounds. The two most significant parameters were the nebulizer carrier gas flow rate and drift tube temperature. The detection system was operated under the following conditions: the drift tube temperature was set to 50 ℃, the nebulizer carrier gas was high-purity nitrogen, the carrier gas flow rate was 1.0 mL/min, the nitrogen pressure was regulated to 275.79 kPa, and the gain factor was set to 3. The sample was extracted with 25 mL of water, shaken and vortexed for 10 min, purified with 200 µL of zinc acetate solution and 200 µL of potassium ferricyanide solution, and centrifuged at 4500 r/min for 10 min. Next, 1 mL of the supernatant was passed through a 0.22 µm aqueous-phase filter membrane, and the filtrate obtained was analyzed using the evaporative light-scattering detector. The six rare sugars were quantitatively analyzed using the external standard method and showed good linearity with coefficients of determination (R2) greater than 0.9985. The limits of detection and quantification were 0.020-0.60 and 0.60-1.8 g/100 g, respectively. In addition, when blank solid food samples were spiked with the analytes at three levels, the average recoveries of the six rare sugars were 92.6%-103.2%, with relative standard deviations (RSDs) of 0.7%-4.4%. An RSD of <5% indicated that the method had good precision. Interference experiments were performed to determine whether the sugars and artificial sweeteners commonly found in solid foods affected the targets. The method established in this study was used to analyze the contents of the six rare sugars in actual solid food samples. The experimental results showed various levels of rare glycoconjugates in different solid foods. Moreover, the actual compositions and labeled of rare glycoconjugates in the solid foods were generally consistent. The proposed method features simple operation, rapid results, high sensitivity, and good reproducibility; thus, it meets the requirements for the detection of the six rare sugars in solid foods. It also provides technical support for the development of methodological standards and detection limits for rare sugars in Chinese foods. The results of this study are of great relevance for the daily monitoring of the levels of the six rare sugars in solid foods.

[Organic Aerosols and Source Analysis of Fine Particles in the Background of Shiwanda Mountain, Guangxi].

The non-polar compounds in atmospheric fine particles (PM2.5) mainly include polycyclic aromatic hydrocarbons (PAHs) and normal alkanes (n-alkanes), etc., which are usually used to identify the source of the air pollution and have a great important impact on human health and the environment. To study the pollution characteristics and sources of non-polar organic aerosols in the background point PM2.5 in Guangxi, from November 2017 to October 2018, 17 types of PAHs and 20 types of n-alkanes were analyzed on PM2.5 samples collected in the field. It was found that the annual average concentrations of PAHs and n-alkanes were 4.28±4.25 ng·m-3 and 13.7±14.72 ng·m-3, respectively. The seasonal change was as follows:winter[(7.86±5.19) ng·m-3, (27.51±16.90) ng·m-3]>spring[(2.73±1.76) ng·m-3, (7.64±4.71) ng·m-3]>autumn[(2.34±145) ng·m-3, (7.01±4.55) ng·m-3]>summer[(1.91±1.67) ng·m-3, (3.98±3.12) ng·m-3]. In PAHs, 5-ring and 6-ring molecules accounted for more than 60%, followed by low- and medium-ring molecules (4-ring and 3-ring). The high molecular weight of n-alkanes was relatively high (C29>C31>C27), and the odd and even carbon numbers were significantly different. In addition, combined with the feature ratio method, principal component analysis method, and backward trajectory joint verification, it was found that 41.5% of non-organic aerosols in winter were affected by maritime traffic emissions and ocean source transportation, and 36.7% of the pollution was explained by the coal burning and local biomass burning; 25.2% of the pollution in spring came from biomass combustion and transportation, and 45.0% was attributed to marine transportation in the southern part of the study area and higher plant wax emissions pollution; 53.4% of pollution in summer came from polluted ship emissions, and 10.6% of pollution came from transportation in Southwest Thailand Source transportation. The organic aerosols at the background sites in Guangxi were affected by the combined effects of local emissions and transmission sources.

Clinical Characteristics and Risk Factors for Pulmonary Infection in Emergency ICU Patients.

Background: Pulmonary infection in the emergency ICUs increases patient morbidity, hospital stay, treatment costs, and the risk of related adverse events. Methods: This study included 695 patients admitted to our emergency ICU between December 2019 and March 2021. Medical records of emergency ICU patients were reviewed to collect their clinical data, including antibiotic use, history of tracheostomy, history of mechanical ventilation, presence or absence of underlying disease, history of smoking, alcohol consumption, age, gender, and history of shock. Bacterial cultures were performed. The incidence, main clinical features, main pathogens, and risk factors of pulmonary infection in emergency ICU were analyzed. Results: In this study, 69 of the 695 emergency ICU patients (9.93%) developed pulmonary infection. The main clinical features of patients with pulmonary infection included cough and expectoration (97.10%), shortness of breath and chest tightness (95.65%), leukocyte elevation (69.57%), confusion (31.88%), drowsiness (28.99%), persistent fever (27.54%), and nausea and vomiting (10.14%). The main pathogenic bacteria in those with pulmonary infection included Klebsiella pneumoniae (62.32%), Pseudomonas aeruginosa (49.28%), Streptococcus pneumoniae (21.74%), Staphylococcus aureus (39.13%), Candida albicans (7.25%), Pneumococcus pneumoniae (15.95%), Pseudomonas aeruginosa (24.64%), and lung diplococcus inflammatory (13.04%). Univariate analysis showed that there were no significant differences in the occurrence of pulmonary infection with regard to sex, smoking, and alcohol consumption, but there were significant differences with regard to age, basic disease, invasive surgery, and shock. Logistic regression analysis confirmed that age ≥ 80 years, invasive surgery, shock, and basic diseases ≥ 2 were important risk factors for pulmonary infection in emergency ICU patients. Conclusion: Considering the clinical features and risk factors for pulmonary infection in the emergency ICU, preventive and control measures are required to minimize its occurrence and ensure good outcomes.

[Determination of 19 chlorophenols in fish by QuEChERS-gas chromatography-mass spectrometry].

With the increasing use of chlorine-containing pesticides, hypochlorous acid disinfection water as well as aquatic product insecticides and fungicides are widely used in the cultivation of fish. This has led to the contamination of fish by chlorophenol compounds. However, currently, there is no standard method for the simultaneous determination of 19 chlorophenol compounds in fish. In this study, the optimum chromatography and mass spectrometry conditions were determined by investigating the instrument parameters. The 19 chlorophenol compounds were well separated using the DB-5MS capillary chromatographic column (30 m×0.25 mm×0.25 µm) with a carrier gas flow rate of 1 mL/min. Under this condition, the chromatographic peak was sharp and symmetric. An analytical method was developed for the simultaneous determination of the 19 chlorophenol compounds in fish using gas chromatography-mass spectrometry coupled with QuEChERS pretreatment. The improved QuEChERS method was used in sample preparation. The 19 chlorophenol compounds were extracted with organic solvents and purified with purifying agents. During the experiment, the effect of the kinds and volumes of the extraction solvent, as well as the types and dosages of the purifying agent, on the recoveries of the 19 chlorophenol compounds were investigated. Moreover, the temperature and time of derivatization, as well as the dosage of the derivatization agent, were optimized. All aforementioned analyses were conducted with the aim of determining the optimal pretreatment method. Finally, the optimized gas chromatography-mass spectrometry conditions were employed for the quantitative determination of 19 chlorophenol compounds in fish samples. Based on the experimental results, the best extraction method was determined to be the one where the extraction agent (10 mL ethyl acetate) was added to 3 g sodium chloride and 5 g anhydrous magnesium sulfate in the test tube, followed by ultrasonication for 15 min. The sample was centrifuged at 4500 r/min for 5 min, and 500 mg C18 was selected as the purifying agent to purify the supernatant. The purified supernatant was blown with nitrogen to less than 1 mL at 45 ℃, and then redissolved with ethyl acetate to 1 mL. Subsequently, the sample solution was passed through a 0.22 µm organic filter membrane, following which 50 µL bis(trimethylsilyl)trifluoroacetamide was added for derivatization at 45 ℃ for 30 min. Lastly, the 19 chlorophenol compounds were determined by gas chromatography-mass spectrometry with an electrospray ionization source and selecting ion monitoring mode. The 19 chlorophenol compounds were then quantitatively analyzed by the external standard method. The compounds showed good linearity in the concentration range of 0.4-10 µg/L, with correlation coefficients (R2) greater than 0.998. The limits of detection and limits of quantification were 0.01-0.05 µg/kg and 0.04-0.16 µg/kg, respectively. Moreover, the average recoveries of the 19 chlorophenol compounds were in the range of 70.6%-115.0% at three spiked levels, and the relative standard deviations were in the range of 2.6%-10.5%. The established method in this study was applied to detect and analyze chlorophenol compounds in actual samples. The experimental results showed that various levels of chlorophenol compounds could be detected in different fishes. Among them, the total amount of chlorophenol compounds detected in the Corvina was 8.74 µg/kg, followed by the Crucian carp at 7.59 µg/kg, and the minimum detected amount in rice fish (1.59 µg/kg). With its simple operation, high sensitivity, and good repeatability, the established method simplifies the pre-treatment of fish samples. It can also meet the requirements for the high-throughput detection of 19 chlorophenol compounds in fish, thereby significantly improving the detection efficiency of chlorophenols. Moreover, the method provides crucial technical support and a theoretical basis for the establishment of feasible detection standards for chlorophenols in China, as well as for the control of residue levels of chlorophenol compounds in fish. The findings have important practical significance to implement management measures during fish breeding and transportation.

[Determination of seven Alternaria toxins in infant milk powder by solid phase extraction coupled with ultra-performance liquid chromatography-tandem mass spectrometry].

Alternaria toxin is a general term for a class of toxic metabolites produced by Alternaria, which widely exists in soil, grain, vegetables, and fruits. This mycotoxin is extremely harmful to human health. It is well known that infant milk powder containing vegetable oil is easily contaminated by Alternaria alternata. Alternaria toxins have thus become an increasingly important focus in food. Rapid and accurate detection of Alternaria toxin residues in food is of great significance for food safety. This requires pretreatment to purify the target toxins and maximize the accuracy and precision of the analysis. In this study, a rapid method based on online solid phase extraction/purification and ultra-performance liquid chromatography-tandem mass spectrometry (online SPE UPLC MS/MS) was established to detect seven Alternaria toxins (alternariol monomethyl ether, altenuene, tenuazonic acid, alternariol, tentoxin, altenusin, and altertoxin Ⅰ) in infant milk powder. First, the mass spectrometry and chromatographic conditions were optimized. A BEH-C18 column (50 mm×2.1 mm, 1.7 µm) was selected, with 0.1% formic acid aqueous solution-acetonitrile as the mobile phase. Then, the extraction conditions (extraction agent ratio and extraction method) and the solid phase extraction process (extraction column, type and volume of the eluent, and pH of the sample loading solution) were optimized. One gram of milk powder (accurate to 0.01 g) was weighed into a 50 mL tip and bottom plug centrifuge tube. Acetonitrile-water (84∶16, v/v) was set as the extraction agent for the first two cycles, and acetonitrile-methanol-water (45∶10∶45, v/v/v) was set as the third extraction agent. Horizontal shaking for 30 min was the best extraction method. The sample was centrifuged at 9500 r/min for 10 min, and the supernatant extracted many times was mixed and blown with nitrogen at 40 ℃. The sample was redissolved in first-order water (pH 5.5), purified on an HLB column, and successively activated with 6 mL methanol and 6 mL first-order water (pH 5.5). The solution was then loaded onto the column, and the SPE was adjusted to ensure that the water sample flowed through the column at the rate of 1 mL/min so that the column did not dry up during the analysis process before the end of sample loading. The column was rinsed with 12 mL of first-order water. After leaching, the negative pressure filtration was continued for approximately 5 min, followed by elution with 10 mL methanol, and the eluted solution was directly tested after passing through a 0.22 µm filter membrane, without concentration. The analytes were determined by electrospray ionization (ESI) with alternating positive and negative ions. Under the optimal analysis conditions, the linear relationships of the seven Alternaria toxins were good in the mass concentration range of 0.5-200 µg/L, with coefficients of determination (R2)>0.9903. The limits of detection and limits of quantification were 0.15-0.64 µg/kg and 0.54-2.24 µg/kg, respectively. The recoveries of the seven Alternaria toxins were 79.1%-114.3%, and the relative standard deviations were less than 8.87% at different concentrations. The method was applied to the determination and analysis of 60 samples of infant milk powder, and the results showed that no toxin was found in stage one or stage two of the milk powder. Only one sample of the stage three of milk powder was detected, which was tentoxin, and the content was 4.97 µg/kg. The developed method is accurate, rapid, simple, sensitive, repeatable, and stable. It can be used for the practical determination of seven Alternaria toxins in infant milk powder.

Simultaneous detection of multiple phenolic compounds in fish by gas chromatography-mass spectrometry following a modified QuEChERS cleanup.

Phenolic compounds can cause health problems in humans through the food chain. Considering that fish play an important role in human diets, we established a rapid, simple and high-throughput method for the determination of 18 phenolic compounds in fish based on a modified QuEChERS sample preparation method combined with GC-MS. The average recovery of the 18 phenolic compounds was 81.3-116% at 3 spiked levels, and the relative standard deviations, RSDr and RSDwR, were in the range of 1.1-11.3% and 1.5-12.2%, respectively. The limit of detection was 2.0-10.1 µg/kg. Satisfactory linear relationships (R2 > 0.998) were observed for the phenolic compounds in their corresponding concentration ranges. Moreover, the established method exhibited a high sensitivity, good stability, and reliability. The development of this method has an important theoretical and practical significance for establishing standards and to control the residue levels of phenolic compounds in fish.

[Changes in Carbonaceous Aerosol in the Northern Suburbs of Nanjing from 2015 to 2019].

Carbonaceous aerosol is an important component of atmospheric fine particles that has an important impact on air quality, human health, and climate change. In order to explore the long-term changes in carbonaceous aerosol under the background of emission reduction, this study measured the mass concentrations of organic carbon (OC) and elemental carbon (EC) of PM2.5, which collected in the northern suburbs of Nanjing for five years (December 17, 2014 to January 5, 2020). The results showed that the five-year average ρ(OC) and ρ(EC) were (10.2±5.3) µg·m-3 and (1.6±1.1) µg·m-3, accounting for 31.1% and 5.2% of PM2.5, respectively. OC and EC concentrations were both high in winter and low in summer. According to the nonparametric Mann-Kendall test and Sen's slope, the mass concentrations of OC and PM2.5 decreased significantly[OC:P<0.0001, -0.79 µg·(m3·a)-1, -0.29%·a-1; PM2.5:P<0.0001, -4.59 µg·(m3·a)-1, -1.58%·a-1]. Although EC had an upward trend, the significance and range of change were not obvious[P=0.02, 0.05 µg·(m3·a)-1, 0.02%·a-1]. OC and EC decreased significantly during winter from 2014 to 2019[OC:P<0.0001, -2.05 µg·(m3·a)-1, -0.74%·a-1; EC:P=0.001, -0.15 µg·(m3·a)-1, -0.05%·a-1], and the decline was more obvious than the whole. The correlation between OC and EC showed that the sources in winter and summer were more complex than those in spring and autumn. According to the characteristic ratio of OC and EC, the contribution of coal combustion and biomass burning decreased from 2015 to 2019, whereas the impact of industrial sources and vehicle emissions became more significant. Corresponding to this was the obvious decline in OC and the slight recovery of EC. The OC/EC ratio was over 2.0, indicating that there was secondary pollution in the study area. Further calculation revealed that the variation in SOC was consistent with that in OC, showing a significant decrease[P<0.0001, -0.47 µg·(m3·a)-1, -0.17%·a-1]. The average mass concentration of SOC was (5.0±3.5) µg·m-3, accounting for 49.2% of OC. These changes indicate clear effects of the prevention and control of air pollution in Nanjing in recent years. Furthermore, future control can focus on the emissions of VOCs to reduce secondary pollution.

Characterization of Microbial Community in Cold-Chain Hairtail Fish by High-Throughput Sequencing Technology.

ABSTRACT: High-throughput DNA sequencing with the Illumina MiSeq platform was used to analyze the microbial communities in hairtail (Trichiurus haumela) muscle samples to study the diversity and dynamic changes in these communities during cold-chain circulation of these fish. The richness and diversity of the microbial community in hairtail muscle had a transient decline from 0 to 24 h and decreased after the first rise from 24 to 216 h. The diversity and richness of bacteria in cold-chain hairtail reached maximum at 168 h. The Shannon and Simpson diversity indices of the bacteria were 2.96 and 0.16, respectively, and their ACE and Chao1 richness indices were 254.84 and 155.10, respectively. The dominant bacteria belonged to phylum Proteobacteria, class Gammaproteobacteria, order Pseudomonadales, family Pseudomonadaceae, and genus Pseudomonas, and their relative abundances were 80.52, 72.11, 76.68, 23.25, and 53.50%, respectively. These results provide a basis for exploring how to maintain the freshness and predict the shelf life of hairtail.