Estimated assessment of dietary exposure to artificial sweeteners from processed food in Nanjing, China.

The objective of this study was to measure the concentrations of three intensity sweeteners (Acesulfame-K, cyclamate and saccharin) in different categories of food available on the Nanjing market, and to investigate whether the Nanjing general population was at risk for exceeding the ADI of sweeteners. A set of 1885 foods was collected and analysed using the National Food Safety Standard procedure in order to establish the concentration levels of the sweeteners. Dietary exposure was estimated using probabilistic modelling software and compared directly with each sweetener's ADI. Consumption data from the China National Nutrition and Health Survey (conducted in 2010-2013) and the actual concentrations of sweeteners in the collected food products were used to perform the intake assessment. The results indicated that Acesulfame-K and cyclamate were commonly used in processed food, and processed nuts, preserved fruit, beverages, and bakery products are the main sources of sweeteners in Nanjing. The estimated exposure of sweeteners in Nanjing was well below the ADIs, as relative intakes at the 95th percentile were 29.7% for saccharin, 79.8% for cyclamate, and 35.9% for Acesulfame-K of the respective ADIs. It was concluded that adults were not at risk of exceeding ADIs for these sweeteners, but the intake of cyclamate at the higher percentiles by children may approach or slightly exceed ADI values.

Monitoring and exposure assessment of organophosphorus flame retardants in source and drinking water, Nanjing, China.

This study developed a new method to determine the residues of 13 organophosphorus flame retardants (OPFRs) in drinking water by gas chromatography-tandem mass spectrometry (GC-MS/MS) technique and investigated the chemical distribution in water samples from municipal plants along the Yangtze River in Nanjing. The linear calibration correlation coefficients R2 for all 13 OPFRs were at least 0.998 0. Three levels of spiked test were performed. Most of the recoveries were in the range of 80~120%, and the relative standard deviations (RSDs) for the 13 OPFRs were 2.1~17% (n = 6). Five OPFRs were 100% positively detected in the samples, and 3 OPFRs were positively detected in some samples. The concentrations of detected OPFR in the water ranged from 0.7 to 5780.0 ng L-1. The average concentrations of OPFRs in wet season were higher than those in dry season, and the contaminants mainly originated from the source water in the Yangtze River. The exposure assessments of individual and total OPFRs were investigated. The estimated daily intakes of total OPFRs via ingestion of drinking water reached up to 64.8 and 45.2 ng/kg bw/day in dry and wet season, respectively. This study demonstrates a profile of OPFR distribution in Nanjing municipal water and provides information on human exposure assessment via drinking water in the Nanjing District, China.

[Determination of three volatile N-nitrosamine in drinking water by solid phase extraction and gas chromatography-triple quadrupole mass spectrometry with programmable temperature vaporizer-based large volume injection].

A method had been developed for the determination of three N-nitrosamines, namely, N-nitrosodimethylamine (NDMA), N-nitrosomethylethylamine (NMEA), and N-nitrosodiethylamine (NDEA) in drinking water by solid phase extraction (SPE) and gas chromatography-triple quadrupole mass spectrometry (GC-QqQ-MS/MS) with programmable temperature vaporizer (PTV)-based large volume injection (LVI). The N-nitrosamine compounds were extracted from the water sample using a solid phase extraction (SPE) cartridge filled with coconut charcoal, and then eluted with 10 mL methylene chloride. The eluate was dried by anhydrous sodium sulfate and 10 µL was injected into the GC-MS/MS by PTV-LVI. The target compounds were detected by the multi-reaction monitoring (MRM) mode, and quantified with the external standard method. The results showed that the three compounds had good linearities in the range of 1-50 ng/L with correlation coefficients (r2) higher than 0.999. Drinking water samples were spiked with the target compounds at three concentration levels (10, 20, and 50 ng/L), and satisfactory recoveries (between 94.8% and 109%) and good reproductivities (relative standard deviation RSD<10%) were achieved. The limits of quantitation (LOQs) of the three N-nitrosamines were found to be in the range of 0.08-0.7 ng/L. The developed method was sensitive, accurate, convenient, and reliable for the determination of the three trace level N-nitrosamines in drinking water.

Transcriptional defects and reprogramming barriers in somatic cell nuclear reprogramming as revealed by single-embryo RNA sequencing.

BACKGROUND: Nuclear reprogramming reinstates totipotency or pluripotency in somatic cells by changing their gene transcription profile. This technology is widely used in medicine, animal husbandry and other industries. However, certain deficiencies severely restrict the applications of this technology. RESULTS: Using single-embryo RNA-seq, our study provides complete transcriptome blueprints of embryos generated by cumulus cell (CC) donor nuclear transfer (NT), embryos generated by mouse embryonic fibroblast (MEF) donor NT and in vivo embryos at each stage (zygote, 2-cell, 4-cell, 8-cell, morula, and blastocyst). According to the results from further analyses, NT embryos exhibit RNA processing and translation initiation defects during the zygotic genome activation (ZGA) period, and protein kinase activity and protein phosphorylation are defective during blastocyst formation. Two thousand three constant genes are not able to be reprogrammed in CCs and MEFs. Among these constant genes, 136 genes are continuously mis-transcribed throughout all developmental stages. These 136 differential genes may be reprogramming barrier genes (RBGs) and more studies are needed to identify. CONCLUSIONS: These embryonic transcriptome blueprints provide new data for further mechanistic studies of somatic nuclear reprogramming. These findings may improve the efficiency of somatic cell nuclear transfer.

Long non-coding RNAs potentially function synergistically in the cellular reprogramming of SCNT embryos.

BACKGROUND: Long non-coding RNAs (lncRNAs), a type of epigenetic regulator, are thought to play important roles in embryonic development in mice, and several developmental defects are associated with epigenetic modification disorders. The most dramatic epigenetic reprogramming event occurs during somatic cell nuclear transfer (SCNT) when the expression profile of a differentiated cell is abolished, and a newly embryo-specific expression profile is established. However, the molecular mechanism underlying somatic reprogramming remains unclear, and the dynamics and functions of lncRNAs in this process have not yet been illustrated, resulting in inefficient reprogramming. RESULTS: In this study, 63 single-cell RNA-seq libraries were first generated and sequenced. A total of 7009 mouse polyadenylation lncRNAs (including 5204 novel lncRNAs) were obtained, and a comprehensive analysis of in vivo and SCNT mouse pre-implantation embryo lncRNAs was further performed based on our single-cell RNA sequencing data. Expression profile analysis revealed that lncRNAs were expressed in a developmental stage-specific manner during mouse early-stage embryonic development, whereas a more temporal and spatially specific expression pattern was identified in mouse SCNT embryos with changes in the state of chromatin during somatic cell reprogramming, leading to incomplete zygotic genome activation, oocyte to embryo transition and 2-cell to 4-cell transition. No obvious differences between other stages and mouse NTC or NTM embryos at the same stage were observed. Gene oncology (GO) enrichment analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis and weighted gene co-expression network analysis (WGCNA) of lncRNAs and their association with known protein-coding genes suggested that several lncRNAs and their associated with known protein-coding genes might be involved in mouse embryonic development and cell reprogramming. CONCLUSIONS: This is a novel report on the expression landscapes of lncRNAs of mouse NT embryos by scRNA-seq analysis. This study will provide insight into the molecular mechanism underlying the involvement of lncRNAs in mouse pre-implantation embryonic development and epigenetic reprogramming in mammalian species after SCNT-based cloning.

Oct4 regulates DNA methyltransferase 1 transcription by direct binding of the regulatory element.

BACKGROUND: The transcription factor Oct4 plays a pivotal role in the pre-implantation development of the mouse embryo. DNA methyltransferase 1 (Dnmt1) maintains the changes in DNA methylation during mammalian early embryonic development. Little is known of the role of Oct4 in DNA methylation in mice. In this study, Kunming white mice were used as an animal model to reveal any correlation between DNA methylation and Oct4 during mammalian embryonic development. RESULTS: The expressions of Dnmt1 and Oct4 were initially studied using real-time PCR. They exhibited different patterns during the pre-implantation stage. Moreover, by using a promoter assay and ChIP analysis, we found that the transcriptional activities of Dnmt1 in mouse NIH/3 T3 cells and CCE cells were regulated by Oct4 through direct binding to the - 554 to - 294 fragment of the upstream regulation element of Dnmt1. The downregulation of Dnmt1 expression and enzyme activity by mouse Oct4 were further confirmed by transfecting Oct4 siRNA into mouse CCE cells. CONCLUSION: Our results indicate that Oct4 is involved in DNA methylation through the regulation of Dnmt1 transcription, especially during the early stages of mouse pre-implantation embryo development.

Fluorescent conjugated microporous polymer based on perylene tetraanhydride bisimide for sensing o-nitrophenol.

A novel conjugated microporous polymer based on perylene tetraanhydride bisimide (DP2A2) has been synthesized through Sonogashira-Hagihara cross-coupling polymerization of tetrabromo-substituted perylene tetraanhydride bisimide derivative (DPBr2ABr2) with 1,4-diethynylbenzene, whose Brunauer-Emmett-Teller (BET) specific surface area is about 378 m2 g-1. The fluorescence quenching behaviors of the DP2A2 were investigated. It is found that the DP2A2 shows high sensitivity and selectivity to tracing o-nitrophenol (o-NP) in THF with KsV constant of 2.00 × 104 L mol-1. The detection limit (LOD) is 1.50 × 10-9 mol L-1. The possible sensing mechanism for the luminescent quenching of DP2A2 towards o-NP exciting at 365 nm was considered the donor-acceptor electron transfer mechanism, which is a combined result from both dynamic (collisional) and static quenching. Moreover, the static quenching process is dominant for DP2A2.

Supplement of Betaine into Embryo Culture Medium Can Rescue Injury Effect of Ethanol on Mouse Embryo Development.

Mammal embryos can be impaired by mother's excessive ethanol uptake, which induces a higher level of reactive oxygen species (ROS) and interferes in one carbon unit metabolism. Here, our analysis by in vitro culture system reveals immediate effect of ethanol in medium on mouse embryo development presents concentration dependent. A preimplantation embryo culture using medium contained 1% ethanol could impact greatly early embryos development, and harmful effect of ethanol on preimplantation embryos would last during the whole development period including of reducing ratio of blastocyst formation and implantation, and deteriorating postimplantation development. Supplement of 50 µg/ml betaine into culture medium can effectively reduce the level of ROS caused by ethanol in embryo cells and rescue embryo development at each stage damaged by ethanol, but supplement of glycine can't rescue embryo development as does betaine. Results of 5-methylcytosine immunodetection indicate that supplement of betaine into medium can reduce the rising global level of genome DNA methylation in blastocyst cells caused by 1% ethanol, but glycine can't play the same impact. The current findings demonstrate that betaine can effectively rescue development of embryos harmed by ethanol, and possibly by restoring global level of genome DNA methylation in blastocysts.

Organophosphorus pesticide and ester analysis by using comprehensive two-dimensional gas chromatography with flame photometric detection.

Thirty-seven phosphorus (P)-containing compounds comprising organophosphorus pesticides and organophosphate esters were analyzed by using comprehensive two-dimensional gas chromatography with flame photometric detection in P mode (GC × GC-FPD(P)), with a non-polar/moderately polar column set. A suitable modulation temperature and period was chosen based on experimental observation. A number of co-eluting peak pairs on the (1)D column were well separated in 2D space. Excellent FPD(P) detection selectivity, responding to compounds containing the P atom, produces clear 2D GC × GC plots with little interference from complex hydrocarbon matrices. Limits of detection (LOD) were within the range of 0.0021-0.048 µmol L(-1), and linear calibration correlation coefficients (R(2)) for all 37 P-compounds were at least 0.998. The P-compounds were spiked in 2% diesel and good reproducibility for their response areas and retention times was obtained. Spiked recoveries were 88%-157% for 5 µg L(-1) and 80%-138% for 10 µg L(-1) spiked levels. Both (1)tR and (2)tR shifts were noted when the content of diesel was in excess of 5% in the matrix. Soil samples were analyzed by using the developed method; some P-compounds were positively detected. In general, this study shows that GC × GC-FPD(P) is an accurate, sensitive and simple method for P-compound analysis in complicated environmental samples.