The MdMYB44-MdTPR1 repressive complex inhibits MdCCD4 and MdCYP97A3 expression through histone deacetylation to regulate carotenoid biosynthesis in apple.

Carotenoids are photosynthetic pigments and antioxidants that contribute to different plant colors. However, the involvement of TOPLESS (TPL/TPR)-mediated histone deacetylation in the modulation of carotenoid biosynthesis through ethylene-responsive element-binding factor-associated amphiphilic repression (EAR)-containing transcription factors (TFs) in apple (Malus domestica Borkh.) is poorly understood. MdMYB44 is a transcriptional repressor that contains an EAR repression motif. In the present study, we used functional analyses and molecular assays to elucidate the molecular mechanisms through which MdMYB44-MdTPR1-mediated histone deacetylation influences carotenoid biosynthesis in apples. We identified two carotenoid biosynthetic genes, MdCCD4 and MdCYP97A3, that were confirmed to be involved in MdMYB44-mediated carotenoid biosynthesis. MdMYB44 enhanced ß-branch carotenoid biosynthesis by repressing MdCCD4 expression, whereas MdMYB44 suppressed lutein level by repressing MdCYP97A3 expression. Moreover, MdMYB44 partially influences carotenoid biosynthesis by interacting with the co-repressor TPR1 through the EAR motif to inhibit MdCCD4 and MdCYP97A3 expression via histone deacetylation. Our findings indicate that the MdTPR1-MdMYB44 repressive cascade regulates carotenoid biosynthesis, providing profound insights into the molecular basis of histone deacetylation-mediated carotenoid biosynthesis in plants. These results also provide evidence that the EAR-harboring TF/TPL repressive complex plays a universal role in histone deacetylation-mediated inhibition of gene expression in various plants.

Abscisic acid activates transcription factor module MdABI5-MdMYBS1 during carotenoid-derived apple fruit coloration.

Carotenoids are major pigments contributing to fruit coloration. We previously reported that the apple (Malus domestica Borkh.) mutant fruits of "Beni Shogun" and "Yanfu 3" show a marked difference in fruit coloration. However, the regulatory mechanism underlying this phenomenon remains unclear. In this study, we determined that carotenoid is the main factor influencing fruit flesh color. We identified an R1-type MYB transcription factor (TF), MdMYBS1, which was found to be highly associated with carotenoids and abscisic acid (ABA) contents of apple fruits. Overexpression of MdMYBS1 promoted, and silencing of MdMYBS1 repressed, ß-branch carotenoids synthesis and ABA accumulation. MdMYBS1 regulates carotenoid biosynthesis by directly activating the major carotenoid biosynthetic genes encoding phytoene synthase (MdPSY2-1) and lycopene ß-cyclase (MdLCYb). 9-cis-epoxycarotenoid dioxygenase 1 (MdNCED1) contributes to ABA biosynthesis, and MdMYBS1 enhances endogenous ABA accumulation by activating the MdNCED1 promoter. In addition, the basic leucine zipper domain TF ABSCISIC ACID-INSENSITIVE5 (MdABI5) was identified as an upstream activator of MdMYBS1, which promotes carotenoid and ABA accumulation. Furthermore, ABA promotes carotenoid biosynthesis and enhances MdMYBS1 and MdABI5 promoter activities. Our findings demonstrate that the MdABI5-MdMYBS1 cascade activated by ABA regulates carotenoid-derived fruit coloration and ABA accumulation in apple, providing avenues in breeding and planting for improvement of fruit coloration and quality.

Method development, validation, and risk assessment of multiple pesticide residues of fruits in China.

In this study, a muti-residue analysis method of 40 pesticides in five different categories of fruits in China was developed based on ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). Five hundred real samples were analyzed and assessed for the dietary exposure risk. The sample treatment method was optimized by comparing four clean-up methods. The matrix effects of different fruits were evaluated. The analytical method was validated in terms of linearity, limit of detection (LOD), limit of quantification (LOQ), accuracy, and precision. The results showed that the optimal method was the treatment by clean up with ODS (octadecylsilane) and MgSO4. The matrix effect was the strongest in orange and weakest in apple. The LOD and LOQ of pesticides were 0.04-5.9 µg kg-1 and 0.13-19.5 µg kg-1, respectively. The recoveries at three spiked levels were ranged from 71.2 to 115.2% with the RSDs from 0.1 to 19.6%. Twenty-two pesticides were detected in 500 fruit samples from the major production regions of China, with concentrations ranging from 0.1 to 1930 µg kg-1. A total of 13, 10, 9, 8, and 4 pesticides were detected in peach, orange, grape, apple, and strawberry. Both the acceptable daily intake (ADI) and acute reference dose (ARfD) for all the detected pesticides were lower than 100%, indicating that the dietary intake risks are acceptable and would not pose potential health risks.

The 14-3-3 protein GRF8 modulates salt stress tolerance in apple via the WRKY18-SOS pathway.

Salinity is a severe abiotic stress that limits plant survival, growth, and development. 14-3-3 proteins are phosphopeptide-binding proteins that are involved in numerous signaling pathways, such as metabolism, development, and stress responses. However, their roles in salt tolerance are unclear in woody plants. Here, we characterized an apple (Malus domestica) 14-3-3 gene, GENERAL REGULATORY FACTOR 8 (MdGRF8), the product of which promotes salinity tolerance. MdGRF8 overexpression improved salt tolerance in apple plants, whereas MdGRF8-RNA interference (RNAi) weakened it. Yeast 2-hybrid, bimolecular fluorescence complementation, pull-down, and coimmunoprecipitation assays revealed that MdGRF8 interacts with the transcription factor MdWRKY18. As with MdGRF8, overexpressing MdWRKY18 enhanced salt tolerance in apple plants, whereas silencing MdWRKY18 had the opposite effect. We also determined that MdWRKY18 binds to the promoters of the salt-related genes SALT OVERLY SENSITIVE 2 (MdSOS2) and MdSOS3. Moreover, we showed that the 14-3-3 protein MdGRF8 binds to the phosphorylated form of MdWRKY18, enhancing its stability and transcriptional activation activity. Our findings reveal a regulatory mechanism by the MdGRF8-MdWRKY18 module for promoting the salinity stress response in apple.

Deciphering the degradation characteristics of the fungicides imazalil and penflufen and their effects on soil bacterial community composition, assembly, and functional profiles.

The adsorption-desorption and degradation characteristics of two widely applied fungicides, imazalil and penflufen, and the responses of soil bacterial diversity, structure, function, and interaction after long-term exposure were systemically studied in eight different soils. The adsorption ability of imazalil in soil was significantly higher than that of penflufen. Both imazalil and penflufen degraded slowly in most soils following the order: imazalil > penflufen, with soil pH, silt, and clay content being the potential major influencing factors. Both imazalil and penflufen obviously inhibited the soil microbial functional diversity, altered the soil bacterial community and decreased its diversity. Although exposure to low and high concentrations of imazalil and penflufen strengthened the interactions among the soil bacterial communities, the functional diversity of the co-occurrence network tended to be simple at high concentrations, especially in penflufen treatment. Both imazalil and penflufen markedly disturbed soil nitrogen cycling, especially penflufen seriously inhibited most nitrogen cycling processes, such as nitrogen fixation and nitrification. Meanwhile, sixteen and ten potential degradative bacteria of imazalil and penflufen, respectively, were found in soils, including Kaistobacter and Lysobacter. Collectively, the long-term application of imazalil and penflufen could cause residual accumulation in soils and subsequently result in serious negative effects on soil ecology.

Combined transcriptomic and metabolomic analyses identifies CsERF003, a citrus ERF transcription factor, as flavonoid activator.

Transcription factor (TF) modulation is a promising strategy for plant flavonoid improvement. Here, we observed evident decreases in some major flavones and flavonols and the expression of some key related genes in a 'Newhall' navel orange mutant (MT) relative to the wild type (WT). A consistently downregulated ERF TF CsERF003 in MT could increase the contents of major flavonoids and the precursor phenylalanine when transiently overexpressed in citrus fruit. Overexpression of CsERF003 in 'Micro-Tom' tomato (OE) resulted in a darker and redder fruit color than wild type 'Micro-Tom' (WTm). Two major flavonoids, naringeninchalcone and kaempferolrutinoside, were averagely induced by 7.99- and 36.83-fold in OEs, respectively, while little change was observed in other polyphenols, such as caffeic acid, ferulic acid, and gallic acid. Key genes involved in the initiation of phenylpropanoid (PAL, 4CH, and 4CL) and flavonoid (CHS and CHI) biosynthesis were up-regulated, while most genes participating in the biosynthesis of other polyphenols, such as HCT and CCR, were down-regulated in OEs. Therefore, it could be concluded that carbon flux floods into the phenylpropanoid biosynthetic pathway and is then specifically directed for flavonoid biosynthesis. CsERF003 may be a potentially promising gene for fruit quality improvement and engineering of natural flavonoid components.

Comprehensive review on patulin and Alternaria toxins in fruit and derived products.

Mycotoxins are toxic secondary metabolites produced by certain fungi, which can contaminate various food commodities, including fruits and their derived products. Patulin and Alternaria toxins are among the most commonly encountered mycotoxins in fruit and their derived products. In this review, the sources, toxicity, and regulations related to these mycotoxins, as well as their detection and mitigation strategies are widely discussed. Patulin is a mycotoxin produced mainly by the fungal genera Penicillium, Aspergillus, and Byssochlamys. Alternaria toxins, produced by fungi in the Alternaria genus, are another common group of mycotoxins found in fruits and fruit products. The most prevalent Alternaria toxins are alternariol (AOH) and alternariol monomethyl ether (AME). These mycotoxins are of concern due to their potential negative effects on human health. Ingesting fruits contaminated with these mycotoxins can cause acute and chronic health problems. Detection of patulin and Alternaria toxins in fruit and their derived products can be challenging due to their low concentrations and the complexity of the food matrices. Common analytical methods, good agricultural practices, and contamination monitoring of these mycotoxins are important for safe consumption of fruits and derived products. And Future research will continue to explore new methods for detecting and managing these mycotoxins, with the ultimate goal of ensuring the safety and quality of fruits and derived product supply.

Heavy metal contamination and risk assessment in winter jujube (Ziziphus jujuba Mill. cv. Dongzao).

Winter jujube (Ziziphus jujuba Mill. cv. Dongzao) is a major fresh-eating jujube fruit with various important nutrients for humans. It can absorb heavy metals from polluted air, water and soils and applied pesticides, which may pose potential threats to consumers. Here, to evaluate the content of heavy metals in winter jujube and systematically evaluate the potential risks, we collected 212 winter jujube samples from four main producing areas in China and determined the contents of eight heavy metals (Cd, Cr, Pb, Ni, Cu, Zn, As, and Mn) using inductively coupled plasma mass spectrometer (ICP-MS). Based on the integrated pollution index (IPI) evaluation standard, more than 99.06% of samples were at safe levels. Moreover, clustering analysis divided the eight heavy metals into four groups, namely Cr/Ni, Cd/Pb, Cu/Mn/Zn, and As. Importantly, none of the analyzed heavy metals posed risks to adults as indicted by the average carcinogenic and non-carcinogenic risks. Notably, Cr and Cd could pose low carcinogenic risks to children (≤12 age group) when their concentration reached the 90th percentile. This study systematically assessed the health risks associated with heavy metal intake through winter jujube consumption and highlighted the necessity of constant heavy metal monitoring in winter jujube.

DNA methylation variation is crucial to restore adventitious rooting ability during in vitro shoot culture-induced rejuvenation in apple rootstock.

In vitro shoot culture has been widely used for restoring adventitious rooting ability in rooting recalcitrant woody perennial species for the past few decades, but its molecular mechanism is largely uncovered. DNA methylation is an essential epigenetic mark that participates in many biological processes. Recent reports suggested a role of DNA methylation in vitro culture in plants. In this study, we characterized the single-base resolution DNA methylome and transcriptome of adult and in vitro shoot culture-induced rejuvenation cuttings of apple rootstock M9T337. We found a global decrease in DNA methylation during rejuvenation, which may be correlated with increased expression of DNA demethylase genes and decreased expression of DNA methyltransferase genes. We additionally documented DNA hypomethylation in 'T337'_R in gene protomer associated with higher transcript levels of several adventitious rooting-related genes. The application of a DNA methylation inhibitor (5-azacytidine) enhanced the adventitious rooting ability and the expression level of adventitious rooting-related genes, such as, MdANT, MdMPK3, MdABCB21, MdCDC48, MdKIN8B, pri-MdMIR156a5 and pri-MdMIR156a12. Together, the DNA hypomethylation is critical for the rejuvenation-dependent adventitious rooting ability in apple rootstock. In addition, increased DNA methylation was also found in thousands of genes in 'T337'_R. We additionally documented that DNA hypermethylation is required for inhibition of adventitious rooting-repressed genes, such as MdGAD5a, encoding glutamate decarboxylase, which can catalyze glutamate decarboxylated to form γ-aminobutyric acid (GABA). Our results revealed that in vitro shoot culture-dependent DNA methylation variation plays important roles in adventitious rooting in apple rootstock.

Deciphering the diversity, composition, function, and network complexity of the soil microbial community after repeated exposure to a fungicide boscalid.

Boscalid is a novel, highly effective carboximide fungicide that has been substantially and irrationally applied in greenhouses. However, little is known about the residual characteristics of boscalid and its ecological effects in long-term polluted greenhouse soils. Therefore, actual boscalid pollution status in greenhouse soils was simulated by repeatedly introducing boscalid into the soil under laboratory conditions. The degradation characteristics of boscalid, and its effects on the diversity, composition, function, and co-occurrence patterns of the soil microbial community were systematically investigated. Boscalid degraded slowly, with its degradation half-lives ranging from 31.5 days to 180.1 days in the soil. Boscalid degradation was further delayed by repeated treatment and increasing its initial concentration. Boscalid significantly decreased soil microbial diversity, particularly at the recommended dosage. Amplicon sequencing analysis showed that boscalid altered the soil microbial community and further stimulated the phylum Proteobacteria and four potential boscalid-degrading bacterial genera, Sphingomonas, Starkeya, Citrobacter, and Castellaniella. Although the network analysis revealed that boscalid significantly reduced the microbial network complexity, it enhanced the vital roles of Proteobacteria by increasing its proportion and strengthening the relationships among the internal bacteria in the network. The soil microbial function in the boscalid treatment were simulated at the recommended dosage and two-fold recommended dosage but showed an inhibition-recovery-stimulation trend at the five-fold recommended dosage with an increase in treatment frequency. Moreover, the expression of nitrogen cycling functional genes, nifH, AOA amoA, AOB amoA, nirK, and nirS in all boscalid treatments displayed an inhibition-recovery-stimulation trend during the entire experimental period, and the effects were more pronounced at the five-fold recommended dosage. In conclusion, repeated boscalid treatments delayed degradation, reduced soil microbial diversity and network complexity, disturbed soil microbial community, and interfered with soil microbial function.

Indigenous functional microbial degradation of the chiral fungicide mandipropamid in repeatedly treated soils: Preferential changes in the R-enantiomer.

This study investigated the indigenous functional microbial communities associated with the degradation of chiral fungicide mandipropamid enantiomers in soils repeatedly treated with a single enantiomer. The R-enantiomer degraded faster than the S-enantiomer, with degradation half-lives ranging from 10.2 d to 79.2 d for the R-enantiomer and 10.4 d to 130.5 d for the S-enantiomer. Six bacterial genera, (Burkholderia, Paraburkholderia, Hyphomicrobium, Methylobacterium, Caballeronia, and Ralstonia) with R-enantiomer substrate preference and three bacterial genera (Haliangium, Sorangium, and Sandaracinus) with S-enantiomer substate preference were responsible for the preferential degradation of the R-enantiomer and S-enantiomer, respectively. KEGG analysis indicated that Burkholderia, Paraburkholderia, Hyphomicrobium, and Methylobacterium were the dominant contributors to soil microbial metabolic functions. Notably, six microbial metabolic pathways and twelve functional enzyme genes were associated with the preferential degradation of the R-enantiomer, whose relative abundances in the R-enantiomer treatment were higher than those in the S-enantiomer treatment. A constructed biodegradation gene (BDG) protein database analysis further confirmed that Burkholderia, Paraburkholderia, Hyphomicrobium, Methylobacterium, and Ralstonia were the potential hosts of five dominant BDGs, bphA1, benA, bph, p450, and ppah. We concluded that bacterial genera Burkholderia, Paraburkholderia, Hyphomicrobium, and Methylobacterium may play pivotal roles in the preferential degradation of mandipropamid R-enantiomer in repeatedly treated soils.

Chiral fungicide penconazole: Absolute configuration, bioactivity, toxicity, and stereoselective degradation in apples.

Traditional evaluation of chiral pesticides can lead to inaccurate results, as their enantiomers may show different properties. Penconazole, a chiral triazole fungicide with two enantiomers, is widely applied to protect against phytopathogens. In this study, its absolute configuration, bioactivity, ecotoxicity, and stereoselective degradation were investigated at the enantiomeric level in detail. The absolute configuration of the two enantiomers (R-(+)-penconazole and S-(-)-penconazole) was first confirmed by electronic circular dichroism (ECD), and their enantioseparation method was developed and optimized using UPLC-MS/MS. S-(-)-penconazole showed high bioactivity, as its fungicidal activity against four target phytopathogens (Alternaria alternate f. sp. mali, Botryosphaeria berengeriana f. sp. piricola, Colletotrichum gloeosporioides, and Fusarium oxysporum) was 1.8-4.4 times higher than that of R-(+)-penconazole. The results of an acute toxicity test showed that the LC50 values of S-(-)-penconazole against Daphnia magna were 32.5 times higher than those of R-(+)-penconazole at 24 h during the test period. Stereoselective degradation behaviors were found in nonbagging and bagging Fuji apples collected from three major apple-producing regions in China, with half-lives of 23.5-51.6 d (nonbagging treatment) and 23.0-57.5 d (bagging treatment) for R-(+)-penconazole and 41.1-60.9 d (nonbagging treatment) and 52.5-91.2 d (bagging treatment) for S-(+)-penconazole, respectively. This study provided new insights into the bioactivity, ecotoxicity, and stereoselective degradation of penconazole enantiomers. The above results also emphasized the importance of risk assessments of chiral pesticides at the enantiomeric level.

Multielement authentication of apples from the cold highlands in southwest China.

BACKGROUND: Half of all apple production worldwide comes from China. However, the geographic authentication of Chinese apples has not been well studied. We highlight the multi-element-based geographical discrimination of apples from the southwest cold highlands (SCH) of China. 565 samples from the SCH (138) and others (427) were obtained, and the content of fifteen elements were applied to construct models for discrimination. RESULTS: The SCH apples from 2017 to 2019 had higher concentrations of Mn, Zn, Cr, Cd, Se, Pb, and Fe, but lower concentrations of Na, B, Ni, and P. With sufficient training, linear discriminant analysis (LDA) discriminated the SCH, and the testing accuracy averaged 92.5% and 92.2%. Nonlinear discrimination models were more suitable than the linear models. Optimized random forest analysis was the model with the best fit, and with averaged training and testing it obtained a level of accuracy of 98.2% and 98.5%. CONCLUSION: The multielement-based discrimination of SCH apples could aid further studies of geographical origins. © 2021 Society of Chemical Industry.

Application, advancement and green aspects of magnetic molecularly imprinted polymers in pesticide residue detection.

Molecularly imprinted polymers (MIPs) have added a vital contribution to food quality and safety with the effective extraction of pesticide residues due to their unique properties. Magnetic molecularly imprinted polymers (MMIPs) are a superior approach to overcome stereotypical limitations due to their unique core-shell and novel composite structure, including high chemothermal stability, rapid extraction, and high selectivity. Over the past two decades, different MMIPs have been developed for pesticide extraction in actual food samples with a complex matrix. Nevertheless, such developments are desirable, yet the synthesis and mode of application of MMIP have great potential as a green chemistry approach that can significantly reduce environmental pollution and minimize resource utilization. In this review, the MMIP application for single or multipesticide detection has been summarized by critiquing each method's uniqueness and efficiency in real sample analysis and providing a possible green chemistry exploration procedure for MMIP synthesis and application for escalated food and environmental safety.

An apple (Malus domestica) AP2/ERF transcription factor modulates carotenoid accumulation.

Color is an important trait for horticultural crops. Carotenoids are one of the main pigments for coloration and have important implications for photosynthesis in plants and benefits for human health. Here, we identified an APETALA2 (AP2)/ETHYLENE RESPONSE FACTOR (ERF) transcription factor named MdAP2-34 in apple (Malus domestica Borkh.). MdAP2-34 expression exhibited a close correlation with carotenoid content in 'Benin Shogun' and 'Yanfu 3' fruit flesh. MdAP2-34 promotes carotenoid accumulation in MdAP2-34-OVX transgenic apple calli and fruits by participating in the carotenoid biosynthesis pathway. The major carotenoid contents of phytoene and ß-carotene were much higher in overexpressing MdAP2-34 transgenic calli and fruit skin, yet the predominant compound of lutein showed no obvious difference, indicating that MdAP2-34 regulates phytoene and ß-carotene accumulation but not lutein. MdPSY2-1 (phytoene synthase 2) is a major gene in the carotenoid biosynthesis pathway in apple fruit, and the MdPSY2-1 gene is directly bound and transcriptionally activated by MdAP2-34. In addition, overexpressing MdPSY2-1 in apple calli mainly increases phytoene and total carotenoid contents. Our findings will advance and extend our understanding of the complex molecular mechanisms of carotenoid biosynthesis in apple, and this research is valuable for accelerating the apple breeding process.

Repeated exposure to fungicide tebuconazole alters the degradation characteristics, soil microbial community and functional profiles.

Tebuconazole is a broad-spectrum triazole fungicide that has been extensively applied in agriculture, but its toxicity on soil ecology remains unknown after repeated introduction to soil. This study investigated the degradation of tebuconazole and the changes in soil microbial community composition and functional diversity as well as network complexity in soil repeatedly treated with tebuconazole. Tebuconazole degraded slowly as the degradation half-life initially increased and then decreased during the four repeated treatments. High concentration of tebuconazole treatment significantly delayed the degradation of tebuconazole. The soil microbial functional diversity in tebuconazole-treated soils showed an inhibition-recovery-stimulation trend with increasing treatment frequency, which was related to the increased degradation rates of tebuconazole. Tebuconazole significantly decreased soil microbial biomass and bacterial community diversity, and this decreasing trend became more pronounced with increasing treatment frequency and concentration. Moreover, tebuconazole significantly decreased soil bacterial community network complexity, particularly at high concentration of tebuconazole treatment. Notably, four bacterial genera, Methylobacterium, Burkholderia, Hyphomicrobium, and Dermacoccus, were identified as the potential tebuconazole-degrading bacteria, with the relative abundances in the tebuconazole treatment significantly increasing by 42.1-34687.1% compared to the control. High concentration of tebuconazole treatment delayed increases in the relative abundances of Methylobacterium but promoted those of Burkholderia, Hyphomicrobium and Dermacoccus. Additionally, repeated tebuconazole treatments improved only four metabolic pathways, cell motility, membrane transport, environmental information processing, and xenobiotics biodegradation and metabolism, which were associated with the degradation of tebuconazole. The above results indicated that repeated tebuconazole treatments resulted in the significant accumulation of residues and long-term negative effects on soil ecology, and also emphasized the potential roles of dominant indigenous microbial bacteria in the degradation of tebuconazole.

DNA sequencing, genomes and genetic markers of microbes on fruits and vegetables.

The development of DNA sequencing technology has provided an effective method for studying foodborne and phytopathogenic microorganisms on fruits and vegetables (F & V). DNA sequencing has successfully proceeded through three generations, including the tens of operating platforms. These advances have significantly promoted microbial whole-genome sequencing (WGS) and DNA polymorphism research. Based on genomic and regional polymorphisms, genetic markers have been widely obtained. These molecular markers are used as targets for PCR or chip analyses to detect microbes at the genetic level. Furthermore, metagenomic analyses conducted by sequencing the hypervariable regions of ribosomal DNA (rDNA) have revealed comprehensive microbial communities in various studies on F & V. This review highlights the basic principles of three generations of DNA sequencing, and summarizes the WGS studies of and available DNA markers for major bacterial foodborne pathogens and phytopathogenic fungi found on F & V. In addition, rDNA sequencing-based bacterial and fungal metagenomics are summarized under three topics. These findings deepen the understanding of DNA sequencing and its application in studies of foodborne and phytopathogenic microbes and shed light on strategies for the monitoring of F & V microbes and quality control.

Synthesis and characterization of magnetic molecularly imprinted polymers for effective extraction and determination of kaempferol from apple samples.

The specific of magnetic molecularly imprinted polymers (Fe3O4@SiO2-MIPs) for kaempferol were fabricated by using acrylamide (AM) as the functional monomer, azobisisobutyronitrile (AIBN) as the initiator and ethylene glycol dimethacrylate (EGDMA) as the cross-linker. The Fe3O4@SiO2-MIPs showed high adsorption capacity (3.84 mg/g) for kaempferol, and the adsorption equilibrium was achieved within 50 min. The specific recognition capacity of Fe3O4@SiO2-MIPs was 3.02 times as high as that of Fe3O4@SiO2-NIPs. The Fe3O4@SiO2-MIPs showed high selectivity towards kaempferol over structural analogues. The recoveries of proposed method at three spiked levels analysis were ranged from 90.5% to 95.4% with the relative standard deviations (RSD) less than 5%. The obtained Fe3O4@SiO2-MIPs were successfully applied for the extraction and determination of kaempferol from apple samples. The established method was simple and feasible, which showed high selectivity, fast separation and satisfactory recoveries for real sample analysis.

Enantioselective fate of mandipropamid in grape and during processing of grape wine.

Enantioselective monitoring of chiral fungicide mandipropamid enantiomers were carried out in grapes and wine-making process. The enantiomers of mandipropamid were separated on a Lux Cellulose-2 column and determined by ultra-high-performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS). The processing procedure included washing, fermentation, and clarification. Significant enantioselectivity was observed in grape under field conditions and during wine-making processing. The half-lives of R-mandipropamid and S-mandipropamid were 5.63 days and 7.79 days under field conditions 43.3 h and 69.3 h during wine-making processing, respectively. The EF values ranged from 0.498 to 0.283 in grape under field conditions, and the EF values were from 0.458 (0 h) to 0.362 (312 h) during the whole fermentation process. The results indicated that R-mandipropamid degraded faster than S-mandipropamid in grape under field conditions and during the fermentation process. The processing factors (PFs) were less than 1 for each procedure, and the PF ranged from 0.005 to 0.025 in the overall process, which indicated that the wine-making process can reduce mandipropamid residue in grape wine. The results of this study could help facilitate more accurate risk assessments of mandipropamid in table grapes and during wine-making process.

Etoxazole stereoselective determination, bioaccumulation, and resulting oxidative stress in Danio rerio (zebrafish).

An environmentally-friendly and fast analytical method for the stereoselective determination of etoxazole was developed and then applied to estimate stereoselective bioaccumulation and elimination in zebrafish using SFC-MS/MS. Optimal enantioseparation conditions were determined using a Chiralpak IG-3 column and CO2/MeOH mobile phase (80/20, v/v), at 3.0 mL/min within 1 min, 30°Me and 18 MPa. A modified QuEChERS method was developed for zebrafish sample pretreatment, and mean recoveries were 88.43-110.12% with relative standard deviations ranging from 0.32 to 5.34%. The enantioselectives of etoxazole enantiomers in zebrafish during uptake and depuration phases were evaluated. Significant enantioselective bioaccumulation was observed, with preferential accumulation of (-)-R-etoxazole compared to its antipode, during uptake at both low and high exposure concentrations. The toxic effects of etoxazole on zebrafish were further explored, and activities of antioxidant enzymes were determined in liver of zebrafish. Significant changes were observed in the SOD and GST activities and in the MDA levels, which indicated the occurrence of oxidative stress in liver of zebrafish. The toxic effects exhibited time- and dose-dependent properties. These results can facilitate the accurate risk evaluation of etoxazole and provide basic knowledge for further study of biotoxicity mechanisms.