Abstract :
[en] Mycotoxins contaminants, one of the most serious global challenges, have been attracted more and more attention from International Agency for Research on Cancer (IARC) of the World Health Organization (WHO) and scientists in food safety sciences. Many countries and organizations have established the maximum contamination level of the main mycotoxins at very low values. Traditional analytic strategies are mainly based on instrumental quantitative method and immunoassays approaches. Aptamer, a novel molecules recognition element like or even superior to antibodies, has attracted more and more attentions for scientists. Therefore, specific aptamers could be employed to construct biosensors for the detection of mycotoxins. The objective of this thesis is to develop novel aptamer-based biosensors for sensitive determination of trace levels of mycotoxins and to provide a promising application of these aptasensors for more hazard factors in food safety sciences.
The main contents and results are as follows:
(1) Aptamer-based biosensor for detection of mycotoxins
Mycotoxins are a large types of secondary metabolites appeared by fungi, they pose a great hazard and toxic reactions to human and animals. A majority of countries and regulators, such as European Union, have established series of requirements and set the maximum tolerated levels. The development of high sensitive and specific analytical platform for mycotoxins is much in demand to address new challenges for food safety in worldwide. Due to the superiority of simple, rapid, and low-cost characteristics, aptamer-based biosensors are successfully developed for the detection of various mycotoxins with high sensitivity and selectivity compared with traditional instrumental methods and immunological approaches. In this article, we discuss and analyze the development of aptasensors for mycotoxins determination in food and agricultural products during the last eleven years and cover the literatures from the first report in 2008 until today. In addition, challenges and future trends for the selection of aptamers towards various mycotoxins and aptasensors for multi-mycotoxins analysis are summarized. Given the promising development and potential application of aptasensors, the future researches will witness the great practicability of aptamer-based biosensor for food safety field.
(2) A qPCR aptasensor for sensitive detection of aflatoxin M1
Aflatoxin M1 (AFM1), one of the most toxic mycotoxins, imposes serious health hazards. AFM1 had previously been classified as a group 2B carcinogen (IARC, 1993) and has been classified as a group 1 carcinogen by the International Agency for Research on Cancer (IARC) of the World Health Organization (WHO) (IARC, 2002). Determination of AFM1 thus plays an important role for quality control of food safety. In this work, a sensitive and reliable aptasensor was developed for the detection of AFM1. The immobilization of aptamer through a strong interaction with biotin–streptavidin was used as a molecular recognition element, and its complementary ssDNA was employed as the template for a real-time quantitative polymerase chain reaction (RT-qPCR) amplification. Under optimized assay conditions, a linear relationship (ranging from 1.0×10-4 to 1.0 µg L-1) was achieved with a limit of detection (LOD) down to 0.03 ng L-1. In addition, the aptasensor developed here exhibits high selectivity for AFM1 over other mycotoxins and small effects from cross-reaction with structural analogs. The method proposed here has been successfully applied to quantitative determination of AFM1 in infant rice cereal and infant milk powder samples. Results demonstrated that the current approach is potentially useful for food safety analysis, and it could be extended to a large number of targets.
(3) A novel graphene oxide-based aptasensor for amplified fluorescent detection of aflatoxin M1 in milk powder
In this paper, a rapid and sensitive fluorescent aptasensor for the detection of aflatoxin M1 (AFM1) in milk powder has been developed. Graphene oxide (GO) was employed to quench the fluorescence of carboxyfluorescein-labelled aptamer and protect the aptamer from nuclease cleavage. Upon the addition of AFM1, a formation of AFM1/aptamer complex resulted in the aptamer detached from the surface of GO, then the aptamer was cleaved by DNase I and the target AFM1 was released for a new cycle, which led to a great signal amplification and high sensitivity. Under optimized conditions, the GO-based detection of the aptasensor exhibited a linear response to AFM1 in a dynamic range from 0.2 to 10 μg/kg, with a limit of detection (LOD) of 0.05 μg/kg. Moreover, the developed aptasensor showed a high specificity towards AFM1 without interference from other mycotoxins. In addition, the technique has been successfully applied for detection of AFM1 in infant milk powder samples. This aptasensor proposed here offers a promising technology for food safety and can be extended to various targets.
(4) Graphene oxide driven fluorescent aptasensor for the detection of fumonisin B1
Fumonisin B1 (FB1), one of the most toxic mycotoxins, has been designated as possible 2B group carcinogen by the International Agency for Research on Cancer (IARC) in 2002. Therefore, simple, sensitive and specific approaches for the detection of FB1 are much in demand. In this study, a novel aptasensor was introduced for FB1 analysis based on graphene oxide (GO) and DNase I signal amplification. GO was adopted as a fluorescence quencher against ROX-modified aptamer and a protectant for the aptamer from cleavaging by DNase I for subsequent target cycling and signal amplification detection. This proposed sensing strategy exhibited a good linearity for FB1 determination in the dynamic range from 0.5 to 20 ng mL-1 with a good correlation of R2 = 0.995. Its detection of limit was established at 0.15 ng mL-1 (S/N = 3). The specific analysis indicated that this aptasensor was selective for FB1 other than other mycotoxins. In addition, the practical application in real samples of this aptasensor for the detection of FB1 was investigated. The sensing platform proposed here was useful for a potential application in the field of food safety for mycotoxins analysis.
(5) Articles highlights and future perspective
By using the novel aptamer-based biosensors, we developed several approaches for the detection of the most toxic mycotoxins for food safety. In addition, these sensing strategies could be applied for more hazard factors determination by simple replacement of the specific aptamers. More importantly, though the practical applicability, feasibility, and accuracy of these proposed aptasensors were investigated and evaluated through the analysis of the spiked samples experiments, the future’s researches were needed for a validation of these aptasensors with real contaminated samples to determine the performances such as limit of detection and quantification, precision, trueness, accuracy, etc. Through the method validation, these aptasensors will be widely used for the detection of mycotoxins. In addition, future direction will focus on the simplification of analytic principle and devices and the combination of novel aptamers with new materials and techniques to improve the analytical performance and market practicality of aptasensors.
