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Background
Wine has gained a high popularity among the world. According to statistics from the International Vine and Wine Organization (OIV), in 2022, wine occupies a high production volume of 25.8 billion and a large consumption of 23.2 billion litres. However, the European Food Safety Authority emphasizes that wine ranks second among exposures to Ochratoxin A, which is one of the most toxic mycotoxins in food by the World Health Organization (WHO).
Ochratoxin is another deadly fungal mycotoxin that has attracted widespread attention around the world after Aflatoxin. Ochratoxins can be mainly categorized into many types. Among them, OTA is considered the most toxic, abundant, virulence, toxigenic, pollutive to agricultural products, and closest relationship with human health. OTA shows extremely strong hepatorenal toxicity, teratogenic and carcinogenic effects, which can accumulate in human blood and viscera through food chain transmission effects with a huge threat to human health. Additionally, Ochratoxin A is classfied as a class 2B carcinogen by the International Agency for Research on Cancer (IARC) and one of the most toxic mycotoxins commonly found in food by the World Health Organization (WHO).
Thus, the accurate detection and degradation of OTA in wine is crucial for food safety.
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The limited index of OTA
Risk to public health related to the presence of OTA in wines was assessed on the basis of 113 dry wines from vintages spanning 2011-2016, analyzed by immunoaffinity and liquid chromatography with fluorescence detection. More than half of sampled wines (52.2%) contained OTA in quantifiable amounts (above 0.02 μg/L). Ochratoxin A has been detected in all types of wine. For example, red wine exhibited OTA levels ranging from 0.3 to 2.1 μg/L, and white wine showed levels in the range of 0.6 to 8.6 μg/L.
Table 1. Prevalence and measured concentrations (µg/L) of OTA among U.S. Wines.
*LOD: limit-of-detection (LOD = 0.1 µg/L)
However, both Europe and China have established a maximum allowable limit of 2 μg/L for OTAs.
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Detect and degrade OTA
Current solution
Currently, methods for detecting OTA consist of instrumental analysis and immunoassay techniques. Instrumental analysis includes gas chromatography-mass spectrometry (GC-MS), liquid chromatography-mass spectrometry (LC-MS), and high-performance liquid chromatography (HPLC). Immunoassay methods encompass Enzyme-Linked Immunosorbent Assay (ELISA), Immunochromatographic Assay (ICA), and so on. However, these methods are expensive, time-consuming, inconvenient, and susceptible to environmental influences.
When brewing wine, manufacturers will try to remove OTA during the fermentation stage. However, if the OTA levels detected remain above the permitted level before the wine is sold as a final product, this batch of wine can only be discarded, resulting in significant wastage.
Our answer
We designed two modules to detect and degrade OTA. The detecting module is based on OTA and OTA aptamer binding, with high affinity and specificity, which aflicates signals through a structure-switching locked hairpin triggered RCA with the assistance of a dumbbell template so that we can detect it conveniently.
The degradation module could convert OTA into phenylalanine and OTα that can be regarded as non-toxic to humans, which uses functional semi-interpenetrating polymer networks automatically assembled by engineering bacteria to fuse OTA degrading enzyme to form a living functional material.
Our method overcome the low pH and alcohol content of wine leading to the problem of activity reducing of degrading enzymes, rechieving safe and efficient degrading of OTA in wine compared to other approaches. Overall, we attempt to establish a rapid and efficient strategy to detect and degrade OTA, which supplements current methods efficiently. Our project provides a new idea for food safety testing, which has implications for the food safety of wine.
Fig. 1 Entire design of the project
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Reference
DE JESUS C L, BARTLEY A, WELCH A Z, et al. High Incidence and Levels of Ochratoxin A in Wines Sourced from the United States[J]. Toxins, 2017, 10(1).
WANG L, HUA X, SHI J, et al. Ochratoxin A: Occurrence and recent advances in detoxification[J]. Toxicon: Official Journal of the International Society on Toxinology, 2022, 210: 11-8.
https://www.who.int/news-room/fact-sheets/detail/mycotoxins
ORTIZ-VILLEDA B, LOBOS O, AGUILAR-ZUNIGA K, et al. Ochratoxins in Wines: A Review of Their Occurrence in the Last Decade, Toxicity, and Exposure Risk in Humans[J]. Toxins, 2021, 13(7).
DE JESUS C L, BARTLEY A, WELCH A Z, et al. High Incidence and Levels of Ochratoxin A in Wines Sourced from the United States[J]. Toxins, 2017, 10(1).
MEULENBERG E P. Immunochemical methods for ochratoxin A detection: a review[J]. Toxins, 2012, 4(4): 244-66.
XU G, ZHAO J, YU H, et al. Structural Insights into the Mechanism of High-Affinity Binding of Ochratoxin A by a DNA Aptamer[J]. Journal of the American Chemical Society, 2022, 144(17): 7731-40.
ZHANG J, LU Y, GAO W, et al. Structure-switching locked hairpin triggered rolling circle amplification for ochratoxin A (OTA) detection by ICP-MS[J]. Microchemical Journal, 2023, 186: 108365.
DAI Z, YANG X, WU F, et al. Living fabrication of functional semi-interpenetrating polymeric materials[J]. Nature Communications, 2021, 12(1): 3422.
XIONG L, PENG M, ZHAO M, et al. Truncated Expression of a Carboxypeptidase A from Bovine Improves Its Enzymatic Properties and Detoxification Efficiency of Ochratoxin A[J]. Toxins, 2020, 12(11).
