Studies of Mycotoxins in Medicinal Plants Conducted Worldwide over the Last Decade: A Systematic Review, Meta-Analysis, and Exposure Risk Assessment.

BACKGROUND: Mycotoxins have been reported to be present in medicinal plants. With the growing usage of medicinal plants, contamination of mycotoxins has emerged as one of the biggest threats to global food hygiene and ecological environment, posing a severe threat to human health. PURPOSE: This study aimed to determine the mycotoxin prevalence and levels in medicinal plants and conduct a risk assessment by conducting a systematic review and meta-analysis. METHODS: A thorough search on Web of Science and PubMed was conducted for the last decade, resulting in 54 studies (meeting the inclusion criteria) with 2829 data items that were included in the meta-analysis. RESULTS: The combined prevalence of mycotoxins in medicinal plants was 1.7% (95% confidence interval, CI = 1.1% - 2.4%), with a mean mycotoxin concentration in medicinal plants of 3.551 µg/kg (95% CI = 3.461 - 3.641 µg/kg). Risk assessment results indicated that aflatoxins and ochratoxin A found in several medicinal plants posed a health risk to humans; additionally, emerging enniatins exhibited possible health risks. CONCLUSION: Therefore, the study underlines the need for establishing stringent control measures to reduce the severity of mycotoxin contamination in medicinal plants.

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.

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.

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.

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.

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.

Molecular Docking and In Vitro Studies of Ochratoxin A (OTA) Biodetoxification Testing Three Endopeptidases.

Ochratoxin A (OTA) is considered one of the main mycotoxins responsible for health problems and considerable economic losses in the feed industry. The aim was to study OTA's detoxifying potential of commercial protease enzymes: (i) Ananas comosus bromelain cysteine-protease, (ii) bovine trypsin serine-protease and (iii) Bacillus subtilis neutral metalloendopeptidase. In silico studies were performed with reference ligands and T-2 toxin as control, and in vitro experiments. In silico study results showed that tested toxins interacted near the catalytic triad, similar to how the reference ligands behave in all tested proteases. Likewise, based on the proximity of the amino acids in the most stable poses, the chemical reaction mechanisms for the transformation of OTA were proposed. In vitro experiments showed that while bromelain reduced OTA's concentration in 7.64% at pH 4.6; trypsin at 10.69% and the neutral metalloendopeptidase in 8.2%, 14.44%, 45.26% at pH 4.6, 5 and 7, respectively (p < 0.05). The less harmful α-ochratoxin was confirmed with trypsin and the metalloendopeptidase. This study is the first attempt to demonstrate that: (i) bromelain and trypsin can hydrolyse OTA in acidic pH conditions with low efficiency and (ii) the metalloendopeptidase was an effective OTA bio-detoxifier. This study confirmed α-ochratoxin as a final product of the enzymatic reactions in real-time practical information on OTA degradation rate, since in vitro experiments simulated the time that food spends in poultry intestines, as well as their natural pH and temperature conditions.

The Mechanism of Ochratoxin Contamination of Artificially Inoculated Licorice Roots.

Ochratoxin (OT) contamination of medicinal herbs is a serious threat to human health. This study was performed to investigate the mechanism of OT contamination of licorice (Glycyrrhiza sp.) root. Licorice root samples were cut into eight parts, which were placed separately on sucrose-free Czapek Dox agar medium, inoculated with the spores of ochratoxigenic Aspergillus westerdijkiae. After incubation for 10 and 20 days, the OT contents of the samples were determined by high-performance liquid chromatography, and microtome sections prepared from the samples were analyzed by desorption electrospray ionization tandem mass spectrometry, to visualize OT localization. The same sections were further examined by light microscopy and scanning electron microscopy, to investigate the path of fungal mycelial penetration of the inner roots. OT concentrations tended to increase from the upper- to the middle-root parts. OTs were located in cut areas and areas of cork layer damage; they were not present in the undamaged cork layer, indicating that the structure of this layer prevents OT contamination of the licorice root.

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.

Detection of ochratoxin A using a "turn-on" fluorescence assay based on guanine quenching of the aptamer.

Ochratoxin A (OTA) is a common mycotoxin with high carcinogenicity; therefore, it is crucial to establish a simple, rapid, and sensitive method for its detection. In this study, we developed a "turn-on" fluorescence assay for detecting OTA based on guanine quenching of the aptamer. The method uses fluorescein (FAM) fluorophore to label the complementary strand of the OTA aptamer, Fc-DNA. In the absence of OTA, the Fc-DNA hybridizes with the aptamer to form a double strand. Due to the occurrence of photo-induced electron transfer (PET), the FAM fluorescence signal is quenched as the FAM on the Fc-DNA approaches the guanine of the aptamer at the 5' end. When OTA is present, the aptamer binds to it and thus, is unable to hybridize with Fc-DNA to form a double strand; the FAM fluorescence signal is restored as FAM moves away from the guanine of the aptamer. The assay achieved OTA detection at a detection limit of 28.4 nM. The application of the original guanine of the aptamer as the quenching agent helps avoid the complex designing and labeling of the aptamer, which ensures the high affinity of the aptamer for OTA. Meanwhile, this "turn-on" detection mode helps avoid potential false-positive results as in the "turn-off" mode and improves the assay's sensitivity. Additionally, the method has good selectivity and can be used to detect OTA in traditional Chinese medicine. This method provides a simple, low-cost, and rapid method for OTA detection.

Taking glucose as intermediate bridge-signal-molecule for on-site and convenient detection of ochratoxin A in rice with portable glucose meter.

On-site screening of biotoxins is of great importance for food safety. A new electrochemical-biosensing strategy was constructed for ochratoxin A (OTA) detection by direct using ready-made commercial portable-glucose-meter (PGM). Aptamer against OTA was adopted as the recognition probe and pre-immobilized onto the sensing interface. The complementary biotin-modified probe was further decorated by hybridization. Biotinylated invertase was further introduced onto the sensing system with streptavidin, which also acted as the signal amplification unit. The invertase, which was depended on the amount of OTA, produced the glucose from sucrose in the sensing solution. The glucose could be directly and conveniently measured with PGM. Quantitative analysis of OTA was achieved with a linear range from 0.5 ng/mL to 10 ng/mL and detection limit of 0.45 ng/mL. Of significance, it has been successfully applied for OTA analysis in rice with satisfied recoveries. This unique PGM-based electrochemical platform reveals prospective potential in food safety monitoring.

Non-aqueous bonding of leuprorelin to ochratoxin A for peptide-based solid-phase extraction.

The anticancer therapeutic leuprorelin was found to have excellent affinity to the carcinogen ochratoxin A (OTA), with an equilibrium constant of 2.2 × 108 M-1 at 273 K (dissociation constant Kd = 4.5 nM) when functionalized into a mesoporous polymer. Binding between the surface-bound leuprorelin and mycotoxin was corroborated with DFT calculations, and it was extended to the extraction of OTA from the heavily fatty matrices of coffee, achieving 95% recovery with improved cyclability as compared with immunoaffinity. This work presents the potential of peptide-mycotoxin interactions for durable non-aqueous extraction.

Nanobody-Nanoluciferase Fusion Protein-Enabled Immunoassay for Ochratoxin A in Coffee with Enhanced Specificity and Sensitivity.

Ochratoxin A (OTA), one of the best-known mycotoxins, causes problems concerning food safety with potential toxic effects in humans and animals. So, it is crucial to develop simple and sensitive methods for the detection of OTA. Herein, a nanoluciferase-nanobody fusion protein (Nb28-Nluc)-retaining antibody recognition and enzymatic activity was first prepared, which was then applied as a bifunctional tracer to construct a one-step bioluminescent enzyme-linked immunosorbent assay (BLEIA) for OTA in coffee samples. On the basis of Nb28-Nluc, the BLEIA can be completed with a one-step incubation and detection, with only a substrate replacement from 3,3',5,5'-tetramethylbenzidine (TMB) to a Nluc assay reagent (Furimazine). Under the optimal experimental conditions, the proposed one-step BLEIA achieved a detection limit of 3.7 ng/mL (IC10) within 3 h. Moreover, the BLEIA method showed good repeatability and accuracy in the spike recovery experiments with recoveries of 83.88% to 120.23% and relative standard deviations (RSDs) of 5.2% to 24.7%, respectively. Particularly, the BLEIA displayed superior performances, such as fewer operations and more rapid and sensitive detection as compared with Nb28-based enzyme-linked immunosorbent assay. Therefore, the proposed one-step BLEIA has great potential for the sensitive and accurate screening of OTA in food samples.

Exposure assessment and risk-based limit levels evaluation of ochratoxin A in Astragali Radix in China.

Ochratoxin A (OTA) is a mycotoxin found in a variety of foods and herbal medicines, and several governmental bodies around the world have set maximum allowable levels of OTA in different foods and herbal medicines. This study aims to evaluate the health risk of OTA in Astragali Radix (AR) in China, and to evaluate the effects of different limit levels on the risk control of OTA in AR. The concentrations of OTA in 187 samples of AR were investigated, and 61 (32.6%) samples were positive. The mean, 50th and 95th percentile values of OTA in positive samples were 56.2, 5.1 and 304.5 µg/kg, respectively. A margin of exposure (MOE) approach was applied to assess the risk. Considering other food sources, long-term consumers have a relatively high risk of OTA exposure due to the ingestion of AR. Theoretical limit levels of OTA in AR were evaluated from two dimensions by weighing the costs and the benefits. The results indicated that the limit levels that might be applied to the management of OTA contamination in AR in China could be screened out through risk-based evaluation of limit levels.

Assessment of ochratoxin A exposure risk from the consumption of coffee beans in Phnom Penh, Cambodia.

This survey aimed to determine OTA contamination in roasted coffee samples commercialised in Phnom Penh, Cambodia and to assess the potential health risk from OTA exposure. Forty locally grown and imported coffee samples were collected and analysed. Analytical validation methods were fully performed. In 3 of 40 samples (7.5%), the results showed detectable levels of OTA, ranging from 0.19 to 1.12 µg kg-1, with an overall average of 0.26 µg kg-1 and an average over the LOQ (n = 3) at 0.81 µg kg-1. OTA estimated daily intake (EDI) of both values were 0.05 (overall average) and 0.17 ng/kg bw/day (the worst-case scenario) with the calculated risk of OTA exposure expressed as a Hazard Quotient at 0.003 and 0.01, respectively. This result could imply a low health risk to Cambodian coffee consumers.

Assessing the food safety risk of ochratoxin A in coffee: A toxicology-based approach to food safety planning.

Under the Food Safety Modernization Act (FSMA) and preventive controls (PCs) regulations, food manufacturers must consider whether PCs are needed for potential hazards present in food. The mycotoxin ochratoxin A (OTA) is considered a chemical hazard under FSMA. It is produced by several fungal species and can be present in various agricultural commodities, including coffee. OTA presents a unique scenario in food safety, because it is known to be a potential risk; because heating may destroy it, but not completely; and because the hazard profile suggests it is not acutely toxic at the occurrence levels in coffee, although at high exposure levels, it is potentially nephrotoxic and carcinogenic in animal models. In the absence of US compliance levels, it is important for the risk assessor and risk manager to determine whether PCs are warranted. To address this complex situation in the coffee industry, we combined food safety and toxicology risk assessment principles to examine the available information on OTA hazard and risk in coffee. Exposure and health-based benchmarks for OTA in coffee, established by reviewing peer-reviewed literature, food recall databases, and authoritative reviews, resulted in large margins-of-exposure for both single and repeated exposure scenarios. Furthermore, no evidence was identified from historical data to suggest OTA is acutely toxic in humans from coffee consumption or other exposure sources. Therefore, findings from this assessment indicate that no PC is warranted for US coffee manufactures, based on the low severity and likelihood of risk according to margin-of-exposure estimates and historical data.

An Immunoenzymatic Method for the Determination of Ochratoxin A in Biological Liquids (Colostrum and Cow's Milk).

Ochratoxins are mycotoxins that have been extensively studied lately due to the multiple toxic effects such as nephrotoxicity, hepatotoxicity, and carcinogenicity. These toxins contaminate plant and animal foods and after ingestion they reach into body fluids. The method of competitive direct enzyme immunoassay, in the solid phase, was validated through the determination of specific parameters (performance, linearity, recovery percentage, limit of detection, limit of quantification). The validated method was used to determine ochratoxin A in colostrum and cow's milk. The method applied for the determination of ochratoxin A was linear for the concentration range of 0.0-0.5 ng/mL, the value for the regression coefficient (r) was 0.9838. Ochratoxin A was present in 91.67% of the colostrum and in 93.33% of cow's milk samples. The linearity of the method, demonstrated for very low concentrations of analyte, the detection limit as well as the limit of quantification recommend the method for the determinations of micro-pollutants from foods, including biological fluids.

Evaluation of fungal contamination and ochratoxin A detection in different types of coffee by HPLC-based method.

BACKGROUND: Mycotoxins are secondary fungal metabolites that are produced by some toxigenic fungi on foodstuffs which are poisoning and potentiate for human's health hazards. In coffee samples, ochratoxin A and fungal contamination were examined. METHODS: Immunoaffinity columns were used for treating of all 50 samples from four types of coffee, after that high-performance liquid chromatography was used for determining the amount of ochratoxin. For the identification of fungi, all coffee samples were cultured in appropriated media. RESULTS: The results showed that all samples were contaminated by ochratoxin A but only up to 50% of them had toxins higher than acceptable level as detected in black beans (47%), green beans (33.3%), torch (33.3%), and espresso (25%). Black coffee had a higher mean concentration of ochratoxin A than green coffee. CONCLUSION: Predominant fungi isolated from coffee samples were Aspergillus species. Finally, careful monitoring of mycotoxins in coffee samples is essential to improve the quality of this favorable beverage in future.

Adsorption of aflatoxins and ochratoxins in edible vegetable oils with dopamine-coated magnetic multi-walled carbon nanotubes.

A new, green, and cost-effective magnetic solid-phase extraction of aflatoxins and ochratoxins from edible vegetable oils samples was developed using polydopamine-coated magnetic multi-walled carbon nanotubes (PDA@Fe3O4-MWCNTs) as the absorbent. PDA@Fe3O4-MWCNTs nanomaterials were prepared by chemical co-precipitation and in situ oxidation and self-polymerization of dopamine and was characterized. Factors affecting MSPE and the adsorption behavior of the adsorbent to mycotoxins were studied, and the optimal extraction conditions of MSPE and the complexity of the adsorption process were determined. Based on this, the magnetic solid-phase extraction-high-performance liquid chromatography-fluorescence detection method (MSPE-HPLC-FLD) was established for determining six mycotoxins [aflatoxin B1 (AFB1), AFB2, AFG1, and AFG2, and ochratoxin A (OTA) and OTB)] in vegetable oils. The recovery was 70.15%~89.25%, and RSD was ≤6.4%. PDA@Fe3O4-MWCNTs showed a high affinity toward aflatoxins and ochratoxins, allowing selective extraction and quantification of aflatoxins and ochratoxins from complex sample matrices.