Magnetic aptamer copper nanoclusters fluorescent biosensor for the visual detection of zearalenone based on docking-aided rational tailoring.

To address the food safety issues caused by toxins, we established a fluorescent copper nanocluster biosensor based on magnetic aptamer for the visual and quantitative detection of ZEN. Specifically, we utilized the docking-aided rational tailoring (DART) strategy to analyze intermolecular force and interaction sites between zearalenone (ZEN) and the aptamer, and optimize the long-chain aptamer step by step to enhance the binding affinity by 3.4 times. The magnetic bead-modified aptamer underwent conformational changes when competing with complementary sequences to bind with ZEN. Then, the released complementary sequences will be amplified in template-free mode with the presence of the terminal deoxynucleotidyl transferase (TdT), and generating T-rich sequences as the core sequences for the luminescence of copper nanoclusters. The luminescence could be visualized and quantitatively detected through ultraviolet irradiation. The proposed label-free aptasensor exhibited high sensitivity and specificity, with a low limit of detection (LOD) of 0.1 ng/mL.

Zearalenone Decreases Food Intake by Disrupting the Gut-Liver-Hypothalamus Axis Signaling via Bile Acids.

Zearalenone (ZEN) is a mycotoxin that is harmful to humans and animals. In this study, female and male rats were exposed to ZEN, and the results showed that ZEN reduced the farnesoid X receptor (FXR) expression levels in the liver and disrupted the enterohepatic circulation of bile acids (BAs). A decrease in food intake induced by ZEN was negatively correlated with an increase in the level of total BAs. BA-targeted metabolomics revealed that ZEN increased glycochenodeoxycholic acid levels and decreased the ratio of conjugated BAs to unconjugated BAs, which further increased the hypothalamic FXR expression levels. Preventing the increase in total BA levels induced by ZEN via Lactobacillus rhamnosus GG intervention restored the appetite. In conclusion, ZEN disrupted the enterohepatic circulation of BAs to decrease the level of food intake. This study reveals a possible mechanism by which ZEN affects food intake and provides a new approach to decrease the toxic effects of ZEN.

Biotransformation of zearalenone to non-estrogenic compounds with two novel recombinant lactonases from Gliocladium.

BACKGROUND: The mycotoxin zearalenone (ZEA) produced by toxigenic fungi is widely present in cereals and its downstream products. The danger of ZEA linked to various human health issues has attracted increasing attention. Thus, powerful ZEA-degrading or detoxifying strategies are urgently needed. Biology-based detoxification methods are specific, efficient, and environmentally friendly and do not lead to negative effects during cereal decontamination. Among these, ZEA detoxification using degrading enzymes was documented to be a promising strategy in broad research. Here, two efficient ZEA-degrading lactonases from the genus Gliocladium, ZHDR52 and ZHDP83, were identified for the first time. This work studied the degradation capacity and properties of ZEA using purified recombinant ZHDR52 and ZHDP83. RESULTS: According to the ZEA degradation study, transformed Escherichia coli BL21(DE3) PLySs cells harboring the zhdr52 or zhdp83 gene could transform 20 µg/mL ZEA within 2 h and degrade > 90% of ZEA toxic derivatives, α/ß-zearalanol and α/ß-zearalenol, within 6 h. Biochemical analysis demonstrated that the optimal pH was 9.0 for ZHDR52 and ZHDP83, and the optimum temperature was 45 °C. The purified recombinant ZHDR52 and ZHDP83 retained > 90% activity over a wide range of pH values and temperatures (pH 7.0-10.0 and 35-50 °C). In addition, the specific activities of purified ZHDR52 and ZHDP83 against ZEA were 196.11 and 229.64 U/mg, respectively. The results of these two novel lactonases suggested that, compared with ZHD101, these two novel lactonases transformed ZEA into different products. The slight position variations in E126 and H242 in ZDHR52/ZEA and ZHDP83/ZEA obtained via structural modelling may explain the difference in degradation products. Moreover, the MCF-7 cell proliferation assay indicated that the products of ZEA degradation using ZHDR52 and ZHDP83 did not exhibit estrogenic activity. CONCLUSIONS: ZHDR52 and ZHDP83 are alkali ZEA-degrading enzymes that can efficiently and irreversibly degrade ZEA into non-estrogenic products, indicating that they are potential candidates for commercial application. This study identified two excellent lactonases for industrial ZEA detoxification.

Impact of predicted climate change environmental conditions on the growth of Fusarium asiaticum strains and mycotoxins production on a wheat-based matrix.

Fusarium asiaticum is a predominant fungal pathogen causing Fusarium Head Blight (FHB) in wheat and barley in China and is associated with approximately £201 million in annual losses due to grains contaminated with mycotoxins. F. asiaticum produces deoxynivalenol and zearalenone whose maximum limits in cereals and cereals-derived products have been established in different countries including the EU. Few studies are available on the ecophysiological behaviour of this fungal pathogen, but nothing is known about the impact of projected climate change scenarios on its growth and mycotoxin production. Therefore, this study aimed to examine the interacting effect of i) current and increased temperature (25 vs 30 °C), ii) drought stress variation (0.98 vs 0.95 water activity; aw) and iii) existing and predicted CO2 concentrations (400 vs 1000 ppm) on fungal growth and mycotoxin production (type B trichothecenes and zearalenone) by three F. asiaticum strains (CH024b, 82, 0982) on a wheat-based matrix after 10 days of incubation. The results showed that, when exposed to increased CO2 concentration (1000 ppm) there was a significant reduction of fungal growth compared to current concentration (400 ppm) both at 25 and 30 °C, especially at 0.95 aw. The multi-mycotoxin analysis performed by LC-MS/MS qTRAP showed a significant increase of deoxynivalenol and 15-acetyldeoxynivalenol production when the CH024b strain was exposed to elevated CO2 compared to current CO2 levels. Zearalenone production by the strain 0982 was significantly stimulated by mild water stress (0.95 aw) and increased CO2 concentration (1000 ppm) regardless of the temperature. Such results highlight that intraspecies variability exist among F. asiaticum strains with some mycotoxins likely to exceed current EU legislative limits under prospected climate change conditions.

Effect of DON and ZEN and their metabolites DOM-1 and HZEN on B cell proliferation and antibody production.

Introduction: The mycotoxins deoxynivalenol (DON) and zearalenone (ZEN), produced by Fusarium fungi, are frequently found in the cereal-rich diet of pigs and can modulate the immune system. Some enzymes or bacteria present in the digestive tract can de-epoxydize DON to deepoxy-deoxynivalenol (DOM-1) and biotransform ZEN into hydrolyzed ZEN (HZEN). The effects of these metabolites on immune cells, particularly with respect to the vaccine responses, are poorly documented. The aim of this study was to address the impact of DON and ZEN and their respective derivatives, on proliferation, and antibody production of porcine B cells in vitro. Methods: Peripheral blood mononuclear cells (PBMCs), isolated from healthy pigs, were stimulated with the Toll-like receptor (TLR) 7/8-agonist Resiquimod (R848) or the TLR/1/2-agonist Pam3Cys-SKKKK in combination with DON [0.1-1.6 µM] or DOM-1 [1.6 µM and 16 µM] and ZEN [2.5-40 µM] or HZEN [40 µM]. Results: A strong decrease in B-cell proliferation was observed at DON concentrations equal to or exceeding 0.8 µM and at ZEN concentrations equal to or exceeding 20 µM. Treatment with 1.6 µM DON or 40 µM ZEN led to almost a complete loss of live CD79α+ B cells. Moreover, CD21 expression of proliferating IgG+ and IgM+ B-cell subsets was decreased at DON concentrations equal to and exceeding 0.4 µM and at ZEN concentrations equal to or exceeding 10 µM. ELISpot assays revealed a decrease of IgG-secreting B cells at concentrations of and exceeding 0.4 µM and at ZEN concentrations equal to and exceeding 10 µM. ELISA assays showed a decrease of IgM, IgG, and IgA secretion at concentrations equal to or exceeding 0.4 µM DON. ZEN reduced IgM secretion at 20-40 µM (both R848 and Pam3Cys-SKKKK), IgG secretion at 40 µM (both R848 and Pam3Cys-SKKKK) and IgA secretion at 20-40 µM. Discussion: Our in vitro experiments show that while DON and ZEN impair immunoglobulin production and B-cell proliferation, this effect is abrogated by HZEN and DOM-1.

Aggregation-induced emission nanoparticles facilitating multicolor lateral flow immunoassay for rapid and simultaneous detection of aflatoxin B1 and zearalenone.

Herein we developed a multicolor lateral flow immunoassay (LFIA) test strip for rapid and simultaneous quantitative detection of aflatoxin B1 (AFB1) and zearalenone (ZEN). Three differently colored aggregation-induced emission nanoparticles (AIENPs) were designed as LFIA signal tags, with red and green AIENPs for targeting AFB1 and ZEN at the test line, and yellow AIENPs for indicating the validity of the test strip at the control (C) line. After surface functionalization with antibodies, the developed AIENP-based multicolor LFIA allows simultaneous and accurate quantification of AFB1 and ZEN using an independent C-line assisted ratiometric signal output strategy. The detection limits of AFB1 and ZEN were 6.12 and 26 pg/mL, respectively. The potential of this method for real-world applications was well demonstrated in corn and wheat. Overall, this multicolor LFIA shows great potential for field screening of multiple mycotoxins and can be extended to rapid and simultaneous monitoring of other small molecule targets.

Development of a flow injection chemiluminescence immunoassay based on DES-mediated CuCo2O4 nanoenzyme for ultrasensitive detection of zearalenone in foods.

Nanoenzymes have been widely used to construct biosensors because of their cost-effectiveness, high stability, and easy modification. At the same time, the discovery of deep eutectic solvents (DES) was a great breakthrough in green chemistry, and their combination with different materials can improve the sensing performance of biosensors. In this work, we report an immunosensor using CuCo2O4 nanoenzyme combined with flow injection chemiluminescence immunoassay for the automated detection of zearalenone (ZEN). The immunosensor exhibited excellent sensing performance. Under the optimal conditions, the detection range of ZEN was 0.0001-100 ng mL-1, and the limit of detection (LOD) was 0.076 pg mL-1 (S/N = 3). In addition, the immunosensor showed excellent stability with a relative standard deviation (RSD) of 2.65% for  15 repetitive  injections. The method has been successfully applied to the analysis of real samples with satisfactory recovery results, and can hence provide a reference for the detection of small molecules in food and feed.

Duration of Zearalenone Exposure Has Implications on Health Parameters of Lactating Cows.

There is a limited research focus on evaluating the detrimental effects of prolonged zearalenone (ZEN) intake on dairy cows' health under controlled conditions. This experiment was conducted to evaluate whether the length of exposure to a ZEN-contaminated total mixed ration (TMR) at a level of 9.45 mg per day can negatively influence animal health parameters, such as milk composition, rumen and fecal fermentation, and the chewing activity of lactating dairy cows. For this experiment, we used 18 lactating Simmental cows that were fed a diet of 60% forage and 40% concentrate (on dry matter basis) for 26 consecutive days. The first 4 days were for adaptation prior to the first sampling day (day 0). The sampling events took place on day 0 (baseline) without ZEN, followed by day 1, day 7, day 14, and day 21 (with toxin). Dry matter intake (DMI) and ruminating chews per minute increased on the third week of ZEN inclusion; meanwhile, ruminating, eating, and drinking times were not affected. Most milk composition variables were also unaffected. Rumen fluid osmolality increased on day 21 and total short-chain fatty acids (SCFA) of ruminal fluid decreased on day 7. Fecal SCFA increased on day 21 and the acetate-to-propionate ratio increased from day 1 onwards, showing the influence of toxin intake. Animal health parameters, like heart rate, respiratory rate, and body temperature, were negatively influenced by ZEN intake, all increasing consistently on days 4 and 6, 9 and 12, and 16 and 18, respectively. The liver enzyme glutamate dehydrogenase decreased in response to ZEN intake on day 7. A total daily ZEN intake at the level of 9.45 mg did not show detrimental effects on DMI. Nevertheless, certain health parameters were negatively affected, including body temperature, respiratory rate, and heart rate, starting from the 7th day of ZEN intake, with additional signs of possible loss of water balance on the last sampling day.

Alleviative Effect of Rutin on Zearalenone-Induced Reproductive Toxicity in Male Mice by Preventing Spermatogenic Cell Apoptosis and Modulating Gene Expression in the Hypothalamic-Pituitary-Gonadal Axis.

Zearalenone (ZEN) is a non-steroidal estrogenic mycotoxin found in many agricultural products and can cause reproductive disorders, mainly affecting spermatogenesis in male animals. Rutin (RUT) is a natural flavonoid compound recognized for its significant antioxidant, anti-inflammatory and estrogenic properties. The present study aimed to determine the protective role of RUT against ZEN-induced reproductive toxicity in male mice. Twenty-four adult Kunming male mice were divided into four groups: control, RUT (500 mg/kg RUT), ZEN (10 mg/kg ZEN), ZEN + RUT (500 mg/kg RUT + 10 mg/kg ZEN), with six replicates per treatment. The results indicated that RUT mitigated ZEN-induced disruption in spermatogenic cell arrangement, decreased spermatozoa count, and increased sperm mortality in the testes. RUT significantly restored ZEN-induced reduction in T, FSH, LH, and E2 serum levels. Moreover, RUT mitigated ZEN-induced apoptosis by increasing the mRNA expression level of bcl-2, decreasing the mRNA expression level of kiss1-r, and decreasing the protein expression level of caspase 8 in reproductive tissues. These findings indicate the protective role of RUT against ZEN-induced reproductive toxicity in male mice by regulating gonadotropin and testosterone secretions to maintain normal spermatogenesis via the HPG axis, which may provide a new application direction for RUT as a therapeutic agent to mitigate ZEN-induced reproductive toxicity.

Association of Urinary Mycotoxins with Sperm Quality: A Case-Control Study in Southern Nigeria.

The objective of this study was to determine the association between mycotoxins and the quality of spermatozoa in Nigeria. We designed a prospective case-control study involving 136 men diagnosed with reduced sperm count and quality in five infertility clinics in southwest Nigeria and 154 normal fertile controls. Sperm analysis was conducted in accordance with the recommendations of the World Health OrganizationWHO, while Liquid Chromatography-Mass Spectrometry was used to assay three metabolites of mycotoxins (zearalenone, ochratoxin A, and deoxyvinelol) in the urine samples of cases and controls. The data were analysed with descriptive statistics and non-parametric linear regression. The results showed no overall significant difference in levels of these metabolites between the cases and control groups. In contrast, higher levels of zearalenone and ochratoxin A significantly decreased sperm motility in the cases. Similarly, an increase in the level of ochratoxin A decreased sperm morphology in the unadjusted model in the cases. We conclude that exposure to mycotoxins reduces the quality of spermatozoa (motility and morphology) in Nigerian men but may have no effect on sperm count. Efforts to reduce the exposure of men to mycotoxins are important interventions to improve sperm quality and reduce the prevalence of male infertility in the country.

Construction of a self-reporting molecularly-imprinted electrochemical sensor based on CuHCF modified by rGNR-rGO for the detection of zearalenone.

A self-reporting molecularly-imprinted electrochemical sensor is prepared for the detection of Zearalenone (ZEA). Firstly, the reduced graphene nanoribbons and reduced graphene oxide (rGNR-rGO) were simultaneously modified onto a glassy carbon electrode (GCE) to improve the sensor's sensitivity. After electrodepositing copper nanoparticles onto the rGNR-rGO/GCE, cyclic voltammetry scanning was performed in potassium ferrocyanide solution, and copper hexacyanoferrate (CuHCF) was deposited onto rGNR-rGO/GCE to further improve the sensor's sensitivity while giving it self-reporting capability. Then, molecularly-imprinted polymer films were prepared on the CuHCF/rGNR-rGO/GCE to ensure the selectivity of the sensor. It is found that the linear range of ZEA detection by the constructed sensor is 0.25-500 ng·mL -1, with a detection limit of 0.09 ng·mL -1. This sensor shows the merits of good selectivity, high sensitivity and accurate detection, providing a great possibility for the precise detection of low concentration ZEA in food.

Ultrasensitive detection of zearalenone based on electrochemiluminescent immunoassay with Zr-MOF nanoplates and Au@MoS2 nanoflowers.

In this work, an effective competitive-type electrochemiluminescence (ECL) immunosensor was constructed for zearalenone determination by using Zr-MOF nanoplates as the ECL luminophore and Au@MoS2 nanoflowers as the substrate material. Zr-MOF have an ultra-thin sheet-like structure that accelerates the transfer of electrons, ions and co-reactant intermediates, which exhibited strong and stable anodic luminescence. The three-dimensional Au@MoS2 nanoflowers would form a thin film modification layer on the glassy carbon electrode (GCE). And its good electrical conductivity and higher specific surface area utilization further improving the sensitivity of the ECL immunosensor. Under the optimized conditions, the proposed immunosensor exhibited satisfactory stability, sensitivity and accuracy, and its ECL signal was proportional to the logarithm of ZEN concentration (0.0001-100 ng/mL) and the limit of detection (LOD) was 0.034 pg/mL. In addition, the results of recovery experiment acquired for wheat flour and pig urine samples further proved the feasibility of the immunosensor for the detection of real samples, indicating its potential for ultrasensitive detection of ZEN.

Heme Oxygenase-1 Regulates Zearalenone-Induced Oxidative Stress and Apoptosis in Sheep Follicular Granulosa Cells.

Zearalenone (ZEA) is a common non-steroidal estrogenic mycotoxin found in a range of animal feeds and poses a serious threat to the reproductive health of farm animals and humans. However, the mechanism underlying ZEA-induced reproductive toxicity in sheep remains unknown. Granulosa cells are crucial for egg maturation and the fertility of female sheep. In this study, we aimed to examine the impact of different ZEA concentrations on sheep follicular granulosa cells and to elucidate the potential molecular mechanism underlying ZEA-induced toxicity using transcriptome sequencing and molecular biological approaches. Treating primary sheep follicular granulosa cells with different concentrations of ZEA promoted the overproduction of reactive oxygen species (ROS), increased lipid peroxidation products, led to cellular oxidative stress, decreased antioxidant enzyme activities, and induced cell apoptosis. Using transcriptome approaches, 1395 differentially expressed genes were obtained from sheep follicular granulosa cells cultured in vitro after ZEA treatment. Among them, heme oxygenase-1 (HMOX1) was involved in 11 biological processes. The protein interaction network indicated interactions between HMOX1 and oxidative and apoptotic proteins. In addition, N-acetylcysteine pretreatment effectively reduced the ZEA-induced increase in the expression of HMOX1 and Caspase3 by eliminating ROS. Hence, we suggest that HMOX1 is a key differential gene involved in the regulation of ZEA-induced oxidative stress and apoptosis in follicular granulosa cells. These findings provide novel insights into the prevention and control of mycotoxins in livestock.

Improve the accuracy of FT-NIR for determination of zearalenone content in wheat by using the characteristic wavelength optimization algorithm.

The research contributes a unique method to achieve high-precision quantification of zearalenone (ZEN) in wheat, significantly improving accuracy in the analysis. Fourier transform near infrared spectroscopy (FT-NIR) was employed to capture the spectral information of wheat with different mildew degrees. Three feature selection models, competitive adaptive reweighted sampling (CARS), support vector machine-recursive feature elimination (SVM-RFE), and multiple feature-spaces ensemble-least absolute shrinkage and selection operator (MFE-LASSO) were introduced to processed pre-processed near-infrared spectral data and established partial least squares (PLS) regression according to the selected features. The outcomes indicated that the optimal generalization performance was achieved by the PLS model optimized through the MFE-LASSO model. The root mean square error of prediction (RMSEP) was 18.6442 µg·kg-1, coefficient of predictive determination (RP2) was 0.9545, and relative percent deviation (RPD) was 4.3198. According to the results, it is feasible to construct a stoichiometric model for the quantitative determination of ZEN in wheat by using FT-NIR combined with feature selection algorithm, and this method can also be extended to the detection of various molds in other cereals in the future.

Interaction of mycotoxins zearalenone, α-zearalenol, and ß-zearalenol with cytochrome P450 (CYP1A2, 2C9, 2C19, 2D6, and 3A4) enzymes and organic anion transporting polypeptides (OATP1A2, OATP1B1, OATP1B3, and OATP2B1).

Zearalenone (ZEN) is a mycoestrogen produced by Fusarium fungi. ZEN is a frequent contaminant in cereal-based products, representing significant health threat. The major reduced metabolites of ZEN are α-zearalenol (α-ZEL) and ß-zearalenol (ß-ZEL). Since the toxicokinetic interactions of ZEN/ZELs with cytochrome P450 enzymes (CYPs) and organic anion transporting polypeptides (OATPs) have been barely characterized, we examined these interactions applying in vitro models. ZEN and ZELs were relatively strong inhibitors of CYP3A4 and moderate inhibitors of CYP1A2 and CYP2C9. Both CYP1A2 and CYP3A4 decreased ZEN and ß-ZEL concentrations in depletion assays, while only CYP1A2 reduced α-ZEL levels. OATPs tested were strongly or moderately inhibited by ZEN and ZELs; however, these mycotoxins did not show higher cytotoxicity in OATP-overexpressing cells. Our results help the deeper understanding of the toxicokinetic/pharmacokinetic interactions of ZEN, α-ZEL, and ß-ZEL.

pH-induced interaction mechanism of zearalenone with zein: Binding characteristics, conformational structure and intermolecular forces.

Zein-bound zearalenone (ZEN) complexes are naturally existed in maize by their spontaneous interaction, which significantly impacts the risk assessment of ZEN. Additionally, the pH levels in processing could affect the binding or release of zein-bound ZEN. In this study, pH-induced interaction mechanism of ZEN with zein were studied. Results showed that the acid conditions increased the binding constant (Ka) from 3.46 to 10.0 × 104 L/mol, binding energy from -17.38 to -43.49 kJ mol-1. By increasing hydrophobic interaction and hydrogen bond of ZEN with zein, the binding of ZEN with zein was promoted, forming zein-bound ZEN. Whereas, alkaline conditions decreased the Ka to 1.45 × 104 L/mol and binding energy to 148.48 kJ mol-1, weakened ZEN-zein interaction and stretched zein molecules, resulting the release of ZEN from zein. This study could provide important theoretical basis for perfecting risk assessment and controlling zein-bound ZEN during processing.

Programmable DNA tweezers-SDA for ultra-sensitive signal amplification fluorescence sensing strategy.

BACKGROUND: DNA tweezers, classified as DNA nanomachines, have gained prominence as multifunctional biosensors due to their advantages, including a straightforward structure, response mechanism, and high programmability. While the DNA tweezers demonstrate simultaneous, rapid, and stable responses to different targets, their detection sensitivity requires enhancement. Some small molecules, such as mycotoxins, often require more sensitive detection due to their extremely high toxicity. Therefore, more effective signal amplification strategies are needed to further enhance the sensitivity of DNA tweezers in biosensing. RESULTS: We designed programmable DNA tweezers that detect small-molecule mycotoxins and miRNAs through simple sequence substitution. While the DNA tweezers demonstrate simultaneous, rapid, and stable responses to different targets, their detection sensitivity requires enhancement. We introduced the Strand Displacement Amplification (SDA) technique to address this limitation, proposing a strategy of novel programmable DNA tweezers-SDA ultrasensitive signal amplification fluorescence sensing. We specifically investigate the effectiveness of this approach concerning signal amplification for two critical mycotoxins: aflatoxin B1 (AFB1) and zearalenone (ZEN). Results indicate that the detection ranges of AFB1 and ZEN via this strategy were 1-10,000 pg mL -1 and 10-100,000 pg mL -1, respectively, with corresponding detection limits of 0.933 pg mL -1 and 1.07 pg mL -1. Compared with the DNA tweezers direct detection method for mycotoxins, the newly constructed programmable DNA tweezers-SDA fluorescence sensing strategy achieved a remarkable 104-fold increase in the detection sensitivity for AFB1 and ZEN. SIGNIFICANCE: The constructed programmable DNA tweezers-SDA ultrasensitive signal-amplified fluorescence sensing strategy exhibits excellent detection performance for mycotoxins. The superb versatility of this strategy allows the developed method to be easily used for detecting other analytes by simply replacing the aptamer and cDNA, which has incredible potential in various fields such as food safety screening, clinical diagnostics, and environmental analysis.

Characterization, Structural Analysis, and Thermal Stability Mutation of a New Zearalenone-Degrading Enzyme Mined from Bacillus subtilis.

Zearalenone (ZEN) is a widespread mycotoxin that causes serious damage to animal husbandry and poses a threat to human health. A screen of ZEN-degrading soil bacteria yielded Bacillus subtilis YT-4, which yielded 80% ZEN degradation after 6 h and 95% after 36 h. The gene sequence encoding the degradative enzyme ZENY was mined from the genome of YT-4 and expressed in yeast. ZENY is an α/ß-hydrolase with an optimal enzyme activity at 37 °C and pH 8. By breaking the lactone ring of ZEN, it produces ZENY-C18H24O5 with a molecular weight of 320.16 g/mol. Sequence comparison and molecular docking analyses identified the catalytic ZENY triad 99S-245H-123E and the primary ZEN-binding mode within the hydrophobic pocket of the enzyme. To improve the thermal stability of the enzyme for industrial applications, we introduced a mutation at the N-terminus, specifically replacing the fifth residue N with V, and achieved a 25% improvement in stability at 45 °C. These findings aim to achieve ZEN biodegradation and provide insight into the structure and function of ZEN hydrolases.

Dual function of magnetic nanocomposites-based SERS lateral flow strip for simultaneous detection of aflatoxin B1 and zearalenone.

Aflatoxin B1 (AFB1) and zearalenone (ZEN) are two mycotoxins that often co-occur in corn. A surface-enhanced Raman scattering-based lateral flow immunoassay (SERS-LFIA) that can simultaneously detect AFB1 and ZEN in corn samples was developed employing the core-interlayer-satellite magnetic nanocomposites (Fe3O4@PEI/AuMBA@AgMBA) as dual-functional SERS tags. Under the optimal conditions, the detection ranges of AFB1 and ZEN in corn samples were 0.1-10 µg/kg and 4-400 µg/kg, respectively. Moreover, the test results for two mycotoxins in contaminated corn samples employing the suggested SERS-LFIA was in line with those of the HPLC technique. In view of its satisfactory sensitivity, accuracy, precision and short testing time (20 min), the developed system has a promising application prospect in the on-site simultaneous detection of AFB1 and ZEN.

Site-directed display of zearalenone lactonase on spilt-intein functionalized nanocarrier for green and efficient detoxification of zearalenone.

In this study, we prepared a functional organic-inorganic hybrid nanoflower (InHNF) via split intein moiety in a biomineralization process without using organic solvents. InHNF could specifically bind the target enzymes from crude cell lysates within seconds and site-directedly display them on the surface by forming a peptide bond with enzyme's terminal amino acid residue. This unique feature enabled InHNF to increase the specific activity of zearalenone detoxifying enzyme ZHD518 by 40 âˆ¼ 60% at all tested temperatures and prevented enzyme denaturation even under extreme pH conditions (pH 3-11). Furthermore, it exhibited excellent operational stability, with a residual activity of over 70% after eight reaction cycles. Strikingly, InHNF-ZHD518 achieved above 50% ZEN degradation despite the near inactivation of free ZHD518 in beer sample. Overall, InHNF nanocarriers can achieve environmentally friendly, purification-free, and site-directed immobilization of food enzymes and enhance their catalytic properties, making them suitable for a wide range of industrial applications.