Biocontrol activities of yeasts or lactic acid bacteria isolated from Robusta coffee against Aspergillus carbonarius growth and ochratoxin A production in vitro.

Biocontrol Agents (BCAs) can be an eco-friendly alternative to fungicides to reduce the contamination with mycotoxigenic fungi on coffee. In the present study, different strains of bacteria and yeasts were isolated from Ivorian Robusta coffee. Their ability to reduce fungal growth and Ochratoxin A (OTA) production during their confrontation against Aspergillus carbonarius was screened on solid media. Some strains were able to reduce growth and OTA production by 85 % and 90 % and were molecularly identified as two yeasts, Rhodosporidiobolus ruineniae and Meyerozyma caribbica. Subsequent tests on liquid media with A. carbonarius or solely with OTA revealed adhesion of R. ruineniae to the mycelium of A. carbonarius through Scanning Electron Microscopy, and an OTA adsorption efficiency of 50 %. For M. caribbica potential degradation of OTA after 24 h incubation was observed. Both yeasts could be potential BCAs good candidates for Ivorian Robusta coffee protection against A. carbonarius and OTA contamination.

A green-footprint approach for parallel multiclass analysis of contaminants in roasted coffee via LC-HRMS.

The development and validation of a simple, comprehensive, and environment-friendly procedure to determine pesticide residues, naturally occurring and processing contaminants in roasted coffee is presented. A solid-liquid extraction of pesticides and mycotoxins with ethyl acetate and the concurrent partition of acrylamide to an aqueous phase follows a parallel analytical strategy that requires a single analytical portion to determine contaminants that are typically analyzed by dedicated single residue methods. The partition rules the lipids out of the aqueous extract before an "in-tube" dispersive solid phase microextraction (dSPME) for acrylamide retention. This is followed by the elution with buffer prior to injection. This extract is independently introduced into the system front end followed by the injection of the compounds from the organic phase, yet all spotted in the same run. A novel liquid chromatography high-resolution mass spectrometry (LC-HRMS) method setup enables the quantification of 186 compounds at 10 µg/kg, 226 at 5 µg/kg, and the acrylamide at 200 µg/kg for a total of 414 molecules, with acceptable recoveries (70-120%) and precision (RSD < 20%) making this strategy significantly faster and cost-effective than the dedicated single residue methods. Even though the presence of chlorpyrifos, acrylamide, and ochratoxin A was confirmed on samples of different origins, the findings were below the limit of quantification. During the storage of raw coffee, no proof of masking of OTA was found; however, condensation with glucose was evidenced during thermal processing experiments with sucrose by using stable isotope labeling (SIL). No detected conjugates were found in roasted nor in commercial sugar-added torrefacto samples, an industrial processing usually carried out above the decomposition temperature of the disaccharide.

Ochratoxin A detoxification potentials of basil, chan, and chia seeds.

The most toxic of the ochratoxins is ochratoxin A (OTA), which is primarily produced by species of Aspergillus and Penicillium that can be found in maize, wheat, coffee, red wine, and various grains. OTA induces immunotoxicity, nephrotoxicity, hepatotoxicity, teratogenicity, and carcinogenicity in both animals and humans. Thus, there is a need to identify mycotoxin detoxification agents that can effectively decontaminate OTA. Seeds of basil (Ocimum basilicum L.), chan (Hyptis suaveolens L.), and chia (Salvia hispanica L.) are functional foods capable of eliminating harmful substances. Despite this potential, the impact of these seeds on OTA detoxification remains unclear. This study reveals that milled basil, chan, and chia seeds adsorb significant levels of OTA, with chia demonstrating the highest adsorption capacity, followed by chan and basil seeds showing the least efficiency. Furthermore, milled basil, chan, and chia seeds effectively reduced OTA residues in artificial gastric and intestinal fluids, where they achieved up to 93% OTA adsorption in the former. In addition, these milled seeds were able to remove OTAs from canned, drip, and instant coffee. This study is the first to report the OTA elimination potential of basil, chan, and chia seeds.

Novel Perspective on Mechanism in Muscle Growth Inhibited by Ochratoxin A Associated with Ferroptosis: Model of Juvenile Grass Carp (Ctenopharyngodon idella) In Vivo and In Vitro Trials.

Ochratoxin A (OTA) is a common mycotoxin in food and feed that seriously harms human and animal health. This study investigated the effect of OTA on the muscle growth of juvenile grass carp (Ctenopharyngodon idella) and its possible mechanism in vitro. Our results have the following innovative findings: (1) Dietary OTA increased the expression of increasing phase I metabolic enzymes and absorbing transporters while reducing the expression of efflux transporters, thereby increasing their residue in muscles; (2) OTA inhibited the expressions of cell cycle and myogenic regulatory factors (MyoD, MyoG, and MyHC) and induced ferroptosis by decreasing the mRNA and protein expressions of FTH, TFR1, GPX4, and Nrf2 both in vivo and in vitro; and (3) the addition of DFO improved OTA-induced ferroptosis of grass carp primary myoblasts and promoted cell proliferation, while the addition of AKT improved OTA-inhibited myoblast differentiation and fusion, thus inhibiting muscle growth. Overall, this study provides a potential research target to further mitigate the myotoxicity of OTA.

Barcode-style lateral flow immunochromatographic strip for the semi-quantitative detection of ochratoxin A in coffee samples.

Ochratoxin A (OTA) is a mycotoxin contaminating agricultural products produced by fungi, associated with important toxic effects. Thus, the development of fast, sensitive, and economical approaches for OTA detection is crucial. In this study, a barcode-style lateral flow assay for the semi-quantitative detection of OTA in coffee samples was developed. To achieve this goal, a BSA-OTA complex was immobilized in three test zones to compete with OTA molecules in the sample for binding with anti-OTA antibodies labeled with gold nanoparticles. Different concentrations of OTA in the sample produced distinct colour patterns, allowing semi-quantification of the analyte. The assay exhibited high sensitivity, with a limit of detection of 2.5 µg.L-1, and high reproducibility, with variation coefficient values between 2% and 13%. Moreover, the colour patterns obtained in the analysis with coffee samples were similar to the results obtained with standard OTA solutions, demonstrating a reliable applicability in real samples.

Mycotoxin detection in selected medicinal plants using chromatographic techniques.

Mycotoxins are toxic mycological products that when consumed, absorbed or inhaled cause sickness or even the death of humans. Therefore, the present study aimed to evaluate the contamination levels of mycotoxins (aflatoxins, AFB1 , AFB2 , AFG1 , AFG2 , and ochratoxin A, OTA) in selected medicinal herbs and shrubs using thin-layer chromatography (TLC) and high-performance liquid chromatography (HPLC). A total of 15 samples of medicinal herbs and shrubs were selected. Among them, four samples were aflatoxin contaminated while two samples were ochratoxin A contaminated. The highest level of aflatoxin was detected in Justicia adhathoda (4,704.94 ppb) through HPLC (153.4 ppb) and through TLC, while the lowest level of aflatoxin was detected in Pegnum harmala (205.1 ppb) through HPLC. Similarly, the highest level of OTA was detected in Dodonia viscosa (0.53 ppb) through HPLC (0.5 ppb) and through TLC, while the lowest level was detected in J. adhathoda (O.11 ppb) through HPLC (0.4 ppb) and through TLC. The OTA concentration was very low, being negligible and below permissible limits. The present study concludes that there is a potential risk for the consumption of herbal decoctions. Therefore, regular monitoring and proper management of mycotoxins, including aflatoxins and OTA, in herbal medicines are needed to ensure the safety of herbal drugs to protect consumers.

Plasmonic immunosensors based on spoon-shaped waveguides for fast and on-site ultra-low detection of ochratoxin A in coffee samples.

The high toxicity and occurrence of ochratoxin A (OTA) in grains and foods has been a growing concern due to the impacts on health and the economy in many countries. In this sense, simplified devices with high sensitivity and specificity for local monitoring are enthusiastically pursued. In this work, we report for the first time the detection of ochratoxin A in coffee samples using a spoon-shaped waveguide immunosensor. The biosensor was built with the surface of the spoon-shaped waveguide covered by a 60 nm layer of gold to enable the SPR phenomenon. The measurements indicated a linear relationship between the change in the SPR phenomenon values and the OTA concentration in the range from 0.2 ppt to 5 ppt. When analyzed in coffee samples, the biosensor was highly selective and did not suffer matrix interference. The developed biosensor represents a promising analytical device for coffee quality analyses, as it is portable, simple, and suitable for onsite detection of target analytes.

Nucleic acid aptamer based thermally oxidized porous silicon/zinc oxide microarray chip for detection of ochratoxin A in cereals.

A nucleic acid aptamer based thermally oxidized porous silicon/zinc oxide microarray chip was constructed for the detection of ochratoxin A. The hybrid chains formed by aptamer and complementary chains labeled with fluorescent groups and fluorescent burst groups were used as recognition molecules, and the detection of toxins was accomplished on the chip by the principle of fluorescence signal burst and recovery. The modified QuEChERS method was used for sample pretreatment and the performance of the method was evaluated. The results showed that the linear range was 0.02 âˆ¼ 200 ng/kg with the detection limit of 0.0196 ng/kg under the optimal detection conditions. The method was applied to different cereals with the recoveries of 90.30 âˆ¼ 111.69 %. The developed microarray chip has the advantages of being cost-effective, easy to prepare, sensitive and specific, and can provide a new method for the detection of other toxins.

Pharmacokinetics, dynamics, toxicology and molecular docking of bioactive alkaloid vasicine from Adhatoda vasica: a promising toxin binder against aflatoxin B1 and ochratoxin A.

Vasicine from Adhatoda vasica was investigated in the management of aflatoxicosis and ochratoxicosis by in silico molecular docking approach. The computational analysis was carried out using Discovery Studio Autodock 4.5 tool. Absorption, distribution, metabolism, and excretion (ADME), pharmacodynamics and toxicity studies were also carried out using Swiss ADME and PASS online server, respectively. The standard drug compound used was silymarin and the structure were retrieved from the protein data bank for both the test compound vasicine and the standard drug. Vasicine interacted with aflatoxin B1 at 10 different poses and the maximum dock score was found to be 83.04 and the binding energy was -37.54 kcal/mol. Silymarin interacted with aflatoxin B1 at 10 different poses and the maximum dock score was found to be 143.578 and the binding energy was -67.32 kcal/mol. Vasicine interacted with ochratoxin A at 10 different poses and the maximum dock score was found to be 73.75 and the binding energy was -56.20 kcal/mol. Silymarin interacted with ochratoxin A at 10 different poses and the maximum dock score was found to be 89.23 and the binding energy was -98.86 kcal/mol. The compounds possess good gastro intestinal absorption with antioxidant property and exhibits minimum adverse effects. The obtained results support the toxin mitigating potential of the test compound with minimum adverse effects and hence vasicine can be regarded as a potential toxin binder of aflatoxin B1 and ochratoxin A, wherein it can be implemented for alleviating aflatoxicosis and ochratoxicosis.

Effects of nano-selenium on cecum microbial community and metabolomics in chickens challenged with Ochratoxin A.

Introduction: Ochratoxin A (OTA) is a widely distributed mycotoxin. Nano-selenium (Nano-Se) is an emerging form of selenium known for its superior bioavailability, remarkable catalytic efficiency, and robust adsorbing capacity. Despite these characteristics, its impact on the microbial community and metabolomics in the cecum of chickens exposed to OTA has been infrequently investigated. This research examined the microbiota and metabolomic alterations linked to OTA in chickens, with or without Nano-Se present. Methods: A cohort of 80 healthy chickens at the age of 1 day was randomly distributed into four groups of equal numbers, namely the Se cohort (1 mg/kg Nano-Se), the OTA cohort (50 µg/kg OTA), the OTA-Se cohort (50 µg/kg OTA + 1 mg/kg Nano-Se), and the control group. Each chicken group's caecal microbiome and metabolome were characterized using 16S rRNA sequencing and Liquid chromatography coupled with mass spectrometry (LC-MS) analyses. Results and discussion: Our results showed that the on day 21, the final body weight was significantly reduced in response to OTA treatments (p < 0.05), the average daily gain in the OTA group was found to be inferior to the other groups (p < 0.01). In addition, Nano-Se supplementation could reduce the jejunum and liver pathological injuries caused by OTA exposure. The 16S rRNA sequencing suggest that Nano-Se supplementation in OTA-exposed chickens mitigated gut microbiota imbalances by promoting beneficial microbiota and suppressing detrimental bacteria. Moreover, untargeted metabolomics revealed a significant difference in caecal metabolites by Nano-Se pretreatment. Collectively, the dataset outcomes highlighted that Nano-Se augmentation regulates intestinal microbiota and associated metabolite profiles, thus influencing critical metabolic pathways, and points to a possible food-additive product.

Quantification and health risk assessment of ochratoxin A in dried fruit, spices, and coffee.

Ochratoxin A (OTA) is a stable toxin produced by fungal strains of Aspergillus and Penicillium. It is commonly found in a variety of food products, including dried fruit, coffee, and spices, raising concerns about their safety. This study was aimed to quantify OTA levels in different food products using HPLC with fluorescence detection. The pre-treatment process was optimised by employing immunoaffinity columns with Tween 20 to effectively remove interfering substances. An analytical method was developed, validated, and applied for OTA analysis in dried fruit, spices, and coffee samples. The validation procedure included determining detection and quantification limits, linearity, precision, and accuracy, as per the criteria specified by AOAC International. The validated method was successfully applied for OTA analysis in the selected food samples. Furthermore, health risk assessment was conducted based on the average intake and body weight of the Korean population. From the results, concentrations of OTA in the samples were found to be very low and therefore concluded not to pose significant threats to consumer health.

Effects of Combinations of Toxin Binders with or without Natural Components on Broiler Breeders Exposed to Ochratoxin A.

The objective of this study was to investigate the effects of toxin binders on broiler breeders fed an ochratoxin A (OTA)-contaminated diet. A total of 60 45-week-old female Arbor Acres broiler breeder birds with an initial body weight of 3.65 ± 0.35 kg were randomly divided into 6 treatment groups, with 10 replicates per group and 1 bird per replicate. The trial was conducted for 9 weeks (including 1 week of adaptation). Feed additive 1 (FA1) was composed of clay minerals (85% bentonite and 12% clinoptilolite) with 3% charcoal. FA2 was composed of clay minerals (66.1% aluminosilicates) with natural components (0.8% artichoke and rosemary plant extracts), 7% yeast extract, 0.5% beta-glucans, and 25.6% carriers. The dietary treatment groups were as follows: (1) birds fed an OTA-free basal diet (Negative Control; NC); (2) lipopolysaccharide (LPS)-challenged birds fed a diet including OTA (4 mg/kg) (Positive Control, PC); (3) the PC with 0.05% FA1 (Treatment 1, T1); (4) the PC with 0.10% FA1 (Treatment 2, T2); (5) the PC with 0.10% FA2 (Treatment 3, T3); and (6) the PC with 0.20% FA2 (Treatment 4, T4). The LPS challenge (an intramuscular injection of 1 mg E. coli O55:B5 LPS per kg of body weight) was performed on the first day of the experiment. The results of this experiment show that the PC treatment negatively affected (p < 0.05) egg production, hatchability, Haugh unit, bone mineralization, relative organ weight (abdominal fat, liver), the levels of glutamic oxaloacetic transaminase (GOT), high-density lipoprotein (HDL), and total cholesterol in the blood, and OTA accumulation in the liver compared with the NC. However, supplementation with toxin binders mitigated (p < 0.05) the negative effects of the OTA. Specifically, supplementation with 0.10% FA1 and 0.10% FA2 increased (p < 0.05) eggshell strength by week 4, and the Haugh unit and bone mineralization (phosphorous) by week 8, while decreasing (p < 0.05) the relative weight of the liver and the levels of GOT and HDL in the blood. Supplementation with 0.10% FA2 led to greater improvements in various parameters, including laying performance and bone mineralization, than the other treatments. In conclusion, toxin binders with or without natural components can be effective tools in the mitigation of OTA-induced problems due to their synergistic effects.

Inhibition of Aspergillus carbonarius growth and Ochratoxin A production using lactic acid bacteria cultivated in an optimized medium.

The Gram-positive bacteria lactic acid bacteria (LAB) are used in the food industry but are also known for inhibiting certain food spoilage microorganisms, especially fungi. Sources of nitrogen (N) for culture media are generally organic and expensive. Many attempts have been made to formulate economical culture media with alternative N sources obtained from agricultural and industrial byproducts. This study describes the design and optimization of an inexpensive culture medium for Lactiplantibacillus plantarum (formerly Lactobacillus plantarum) MZ809351 strain B31. The culture medium was optimized using statistical experimental designs to identify the factors with the most significant effects on biomass concentration to reduce the overall cost, aiming to obtain a biomass concentration similar to that obtained with the reference LAB culture medium (de Man, Rogosa and Sharpe; MRS). Sodium acetate and magnesium sulfate were the most significant factors (p < 0.005), and their contents were reduced by 22 % and 40 %, respectively, without affecting biomass concentration. Malt germ extract (MGE) was used as an alternative nitrogen source to replace meat extract (ME) and proteose peptone (PP). Through these experiments, the composition of a culture medium that is less expensive than MRS broth was defined, which produced a biomass concentration (3.8 g/L) similar to that obtained with MRS medium. The inhibitory effects of two LAB strains isolated from the Ivory Coast and Mexico on the growth and production of ochratoxin A (OTA) in an ochratoxigenic fungus was tested. The minimum cellular concentration of the LAB to prevent the development of Aspergillus carbonarius Ac 089 and the production of OTA was determined in a model assay in Petri dishes. The conditions to inhibit the germination of A. carbonarius Ac 089 and the production of OTA were found. Using the optimized medium and a ratio of 2 × 104 LAB/spore (1 × 108 CFU/mL) strain B7 (L. plantarum MZ809351) and 2 × 103 LAB/spore (1 × 107 CFU/mL) strain B31 (L. plantarum MN922335) completely inhibited the growth of the fungus. A ratio of 2 × 105 LAB/spore (1 × 109 CFU/mL) was required to inhibit OTA production with strains B7 and B31. This study indicates the potential of cultivating LAB in an optimized and inexpensive culture medium for use as a biological control agent against ochratoxigenic fungi in food.

Effects of Aspergillus niger Infection on the Quality of Jujube and Ochratoxin A Cumulative Effect.

The jujube is one of the most popular fruits in China because of its delicious taste and high nutritional value. It has a long history of usage as an important food or traditional medicine. However, the jujube is easily infected by fungi, which causes economic losses and threatens human health. When the jujube was infected by Aspergillus niger (H1), the changes in nutritional qualities were determined, such as the content of total acid, vitamin C, reducing sugar, etc. In addition, the ability of A. niger (H1) to produce ochratoxin A (OTA) in different inoculation times and culture media was evaluated, and the content of OTA in jujubes was also analyzed. After jujubes were infected by A. niger (H1), the total acid, and vitamin C contents increased, while the total phenol content decreased, and the reducing sugar content increased after an initial decrease. Although A. niger (H1) infection caused the jujubes to rot and affected its quality, OTA had not been detected. This research provides a theoretical foundation for maximizing edible safety and evaluating the losses caused by fungal disease in jujubes.

Passion fruit-inspired dendritic mesoporous silica nanospheres-enriched quantum dots coupled with magnetism-controllable aptasensor enable sensitive detection of ochratoxin A in food products.

Ochratoxin A (OTA) is a powerful mycotoxin present in a variety of food products, and its detection is important for human health. Here, a fluorescent aptasensor is reported for sensitive OTA determination. Specifically, the surface of bio-inspired passion fruit-like dendritic mesoporous silica nanospheres-enriched quantum dots (MSNQs-apt) was first modified with the OTA aptamer as the recognition unit and fluorescence emitter, while the aptamer-complementary DNA (MNPs-cDNA) was linked with the magnetic nanoparticles (MNPs) as the separation element. In the range of 2.56 pg/mL to 8 ng/mL, the proposed aptasensor exhibited satisfactory linearity and a detection limit of 1.402 pg/mL. The developed aptasensor achieved recoveries of 90.98-103.20% and 94.33-107.57 % in red wine and wheat flour samples, respectively. By simply replacing the aptamer, this aptasensor can be easily extended to detection of other analytes, suggesting its potential as a universal detection platform for mycotoxins in food products.

Polyurethane Foam as a Novel Material for Ochratoxin A Removal in Tea and Herbal Infusions-A Quantitative Approach.

A novel solid-phase extraction methodology followed by UHPLC-MS/MS has been developed for Ochratoxin A (OTA) analysis in herbal infusions. For this purpose, a commercial polyurethane foam (PUF) was used as sorbent, and the experimental conditions were fully optimized. The strategy was satisfactory for reducing the matrix effect and allowed for OTA quantification in black tea and herbal infusions, with suitable recoveries and quantitation limits in agreement with those required by the maximum levels allowed by current regulations. The achieved results demonstrated the unprecedented use of polyurethane foam as an effective alternative for OTA retention and quantification in herbal infusions with the advantages of simple preparation, time saving, sustainability, and low cost for routine analysis.

Label-free electrochemical immunosensor for Ochratoxin a detection in coffee samples.

Ochratoxin A (OTA) is a nephrotoxic and carcinogenic mycotoxin frequently found in coffee, which directly impacts human health and the economy of many countries. For this reason, there has been a growing need for simple and sensitive tools for the on-site detection of this mycotoxin. In this study, we developed a label-free impedimetric immunosensor to detect OTA. The biosensor was built on a thin-film gold electrode evaporated on glass substrtes, modified with a self-assembled cysteamine monolayer and anti-OTA antibodies. Atomic force microscopy and Microspectroscopy RAMAN confirmed the successful functionalization of the electrodes. The biosensor performance was evaluated by electrochemical impedance spectroscopy and the measurements indicated a linear relationship between the change in the impedance values and the OTA concentration in the range from 0.5 to 100 ng mL-1 with a limit of detection of 0.15 ng mL-1. The biosensor was highly selective and did not suffer matrix interference when analyzed in coffee samples. Furthermore, considering the small sample volumes, the short time required for analysis, and the possibility of miniaturization, the developed biosensor represents a promising analytical device for on-site coffee quality analyses.

Ultra-sensitive determination of Ochratoxin A in coffee and tea samples using a novel semi-automated in-syringe based coagulant-assisted fast mycotoxin extraction (FaMEx) technique coupled with UHPLC-MS/MS.

In this study, we demonstrated a novel semi-automated in-syringe-based coagulant-assisted liquid-liquid microextraction (IS-CGA-LLME) as fast mycotoxin extraction (FaMEx) technique coupled with ultra-high-performance liquid chromatography connected with a tandem-mass spectrometer (UHPLC-MS/MS) for the quantification of mycotoxin (Ochratoxin A, OT-A) in coffee and tea samples. IS-CGA-LLME is a three-step extraction process that includes extraction of OT-A from sample matrix using low-volume solvent extraction, then the extractant was cleaned-up using a coagulation process, and finally, the decolorized/matrix removed sample solution was processed for LLME for target analyte's pre-concentration. The final extractant was analyzed using UHPLC-MS/MS for OT-A quantification. Under the optimized experimental conditions, highly sensitive detection and quantification limits were obtained at 0.001 and 0.003 ng g-1 for OT-A with excellent extraction recovery (93-111%) and precision <10%. These results proved that the developed method is a simple, highly sensitive, semi-automated, low-matrix effect and efficient procedure for the determination of mycotoxins in food samples.

Rapid Identification and Analysis of Ochratoxin-A in Food and Agricultural Soil Samples Using a Novel Semi-Automated In-Syringe Based Fast Mycotoxin Extraction (FaMEx) Technique Coupled with UHPLC-MS/MS.

In this work, a fast mycotoxin extraction (FaMEx) technique was developed for the rapid identification and quantification of carcinogenic ochratoxin-A (OTA) in food (coffee and tea) and agricultural soil samples using ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) detection. The FaMEx technique advancement is based on two plastic syringes integrated setup for rapid extraction and its subsequent controlled clean-up process. In the extraction process, a 0.25-g sample and extraction solvent were added to the first syringe barrel for the vortex-based extraction. Then, the extraction syringe was connected to a clean-up syringe (pre-packed with C18, activated carbon, and MgSO4) with a syringe filter. Afterward, the whole set-up was placed in an automated programmable mechanical set-up for controlled elution. To enhance FaMEx technology performance, the various influencing sample pretreatment parameters were optimized. Furthermore, the developed FaMEx method indicated excellent linearity (0.9998 and 0.9996 for coffee/tea and soil) with highly sensitive detection (0.30 and 0.29 ng/mL for coffee/tea and soil) and quantification limits (1.0 and 0.96 for coffee/tea and soil), which is lower than the toxicity limit compliant with the European Union regulation for OTA (5 ng/g). The method showed acceptable relative recovery (84.48 to 100.59%) with <7.34% of relative standard deviation for evaluated real samples, and the matrix effects were calculated as <-13.77% for coffee/tea and -9.7 for soil samples. The obtained results revealed that the developed semi-automated FaMEx/UHPLC-MS/MS technique is easy, fast, low-cost, sensitive, and precise for mycotoxin detection in food and environmental samples.

Dual-signal output fluorescent aptasensor based on DNA programmability and gold nanoflowers for multiple mycotoxins detection.

Herein, a dual-signal output fluorescent aptamer sensor was constructed for the simultaneous detection of aflatoxin B1 (AFB1) and ochratoxin A (OTA) using the specific recognition ability of aptamers and the programmability of DNA. A functional capture probe (cDNA) was designed with the black hole quenching motif BHQ1 labeled at the 5' end and biotin (bio) labeled at the 3' end. The fluorescent dye Cy3-labeled aflatoxin B1 aptamer (AFB1-Apt) and the carboxyfluorescein FAM-labeled ochratoxin A aptamer (OTA-Apt) were used as two fluorescent probes. The cDNA is anchored to the quenching material gold nanoflowers (AuNFs) by the action of streptavidin (SA) and biotin. Its ends can be complementarily paired with two fluorescent probe bases to form a double-stranded structure. The fluorescence of Cy3 was quenched by AuNFs, and the fluorescence of FAM was quenched by BHQ1 through the fluorescence energy resonance transfer (FRET) effect, forming a fluorescence quenching system. Due to the high affinity of the target and the aptamer, the structure of the aptamer probe changes and detaches from the sensor when AFB1 and OTA are present, resulting in enhanced fluorescence. Under optimal conditions, the linear range of AFB1 was 0.1-100 ng/mL (R2 = 0.996), the limit of detection (LOD) was as low as 0.014 ng/mL, and the limit of quantification (LOQ) was 0.046 ng/mL. The linear range of OTA was 0.1-100 ng/mL (R2 = 0.995), the limit of detection (LOD) was as low as 0.027 ng/mL, and the limit of quantification (LOQ) was 0.089 ng/mL. The sensor had high accuracy in detecting both AFB1 and OTA in real sample analysis. The results of the t test show that there is no significant difference between the results of this study and the high-performance liquid phase (HPLC) method, indicating that the prepared sensor can be used as a potential platform for multiple mycotoxins detection.