Aflatoxin M1 Research Articles

SERS and MRS signals engineered dual-mode aptasensor for simultaneous distinguishment of aflatoxin subtypes

The accurate monitoring of aflatoxin subtypes is vitally important for food safety. Herein, a dual-mode aptasensor with surface-enhanced Raman scattering (SERS) and magnetic relaxation switching (MRS) signals is developed for the detection of aflatoxin B1, B2 and M1 (i.e. AFB1, AFB2 and AFM1). Au-Ag Janus NPs and Au-mushroom NPs are prepared and show intense and non-interfering SERS peaks without the additional modification of Raman molecules, and are utilized as SERS nanotags for the distinguishment of AFB1 and AFB2. Fe3O4@Au NPs functionalized by AFM1 aptamers are applied as MRS nanoprobes for the monitoring of AFM1. Aptamers engineered SERS nanotags and MRS nanoprobes are assembled, and show strong SERS performances and high transverse relaxation time (T2). AFB1, AFB2 and AFM1 induce the separation of SERS nanotags from the assemblies and the dispersion of Fe3O4@Au NPs, resulting in the decrease of SERS signals at 1278 cm−1 and 1000 cm−1 as well as the reduction of T2 values. The dual-mode but three kinds of detection signals don’t interfere with each other and exhibit a significant linear relationship with the concentration of targets. This platform provides a high throughput monitoring strategy for the simultaneous analysis of different subtypes of mycotoxin.

Fate of aflatoxin M1 from milk to typical Italian cheeses: Validation of an HPLC method based on aqueous buffer extraction and immune-affinity clean up with limited use of organic solvents

Aflatoxins are mycotoxins produced by several species of Aspergillus fungi which can be found in food and feed. When lactating animals are exposed to feedstuff contaminated by aflatoxin B1, they can excrete its hydroxylated form, aflatoxin M1 (AFM1), into milk. In EU, the AFM1 maximum limit in milk is 0.050 μg kg−1.In this work, we validated a method based on extraction of AFM1 from hard and soft cheese by a sodium citrate solution without the use of organic solvents, and analysis by HPLC-fluorescence detection after purification by immuno-affinity.The method proved to be fit-for-purpose and was successfully verified by participating to proficiency tests.Finally, starting from milk artificially contaminated with AFM1 we prepared four different types of Italian cheese to evaluate AFM1 transfer from milk to cheese and calculate concentration factors occurring during cheese-making processes. We observed an increase of AFM1 concentration in hard cheeses compared to soft cheeses.

Prevalence and concentration of aflatoxin M1 and ochratoxin A in cheese: a global systematic review and meta-analysis and probabilistic risk assessment.

Exposure to mycotoxins such as aflatoxins can endanger human health, especially infants and children. In this study, an attempt was made to retrieved studies related to the concentration of aflatoxin M1 (AFM1) and ochratoxin A (OTA). Search was performed in international databases such as Embase, PubMed, Scopus, and Web of Science for the period 1 January 2010 to 20 February 2023. Then, the pooled concentration in the defined subgroups was calculated using meta-analysis and the health risk assessment was conducted by margin of exposure (MOEs). Thirty-one scientific papers with 34 data reports (Sample size=2,277) were included in our study. The lowest and highest prevalence of AFM1 in cheese was related to El Salvador (12.18 %) and Serbia (100.00 %). The pooled prevalence of AFM1 was 49.85 %, 95 %CI (37.93-61.78 %). The lowest and highest prevalence of OTAin cheese was related to Türkiye (6.67 %) and Italy (44.21 %). The pooled prevalence of OTA was 35.64 %, 95 %CI (17.16-56.44 %). Health risk of AFM1 revealed that except Pakistan and Iran, MOE in the other countries was lower than 10,000 for adults and also except Pakistan, MOE for other countries was lower than 10,000 for children. Health risk of OTA revealed that except Greece, MOE in the other countries was higher than 10,000 for adults and also except Germany and Greece, MOE for other countries was higher than 10,000 for children. Therefore, it is recommended toconduct control plans to reduce the concentration of mycotoxins in cheese, especially AFM1.

Regulated and Emerging Mycotoxins in Bulk Raw Milk: What Is the Human Risk?

Mycotoxins are abiotic hazards whose contamination occurs at the pre- and post-harvest stages of the maize value chain, with animal exposure through contaminated feed leading to their excretion into milk. Currently, only aflatoxin M1 is regulated in milk products. Since feed materials and complete feed present a multi-mycotoxin composition and are the main mycotoxin source into milk, it is important to recognize the occurrence of multiple toxins and their co-occurrence in this highly consumed food product. The aim of this study was to determine the content of regulated and emerging mycotoxins in milk samples, which allowed for evaluating the occurrence and co-occurrence patterns of different mycotoxins known to contaminate feed materials and complete animal feed. Human exposure considering the occurrence patterns obtained was also estimated. Aflatoxins, fumonisins, zearalenone, and emerging mycotoxins were among the mycotoxins found to be present in the 100 samples analyzed. Concentrations ranged from 0.006 to 16.3 μg L-1, with no sample exceeding the AFM1 maximum level. Though several mycotoxins were detected, no exceeding values were observed considering the TDI or PMTDI. It can be concluded that the observed exposure does not pose a health risk to milk consumers, though it is important to recognize vulnerable age groups.

Evaluation of the pH effect on complex formation between bovine β-lactoglobulin and aflatoxin M1: a molecular dynamic simulation and molecular docking study

The aim of this work was to evaluate interaction between aflatoxin M1 (AFM1) and structural models of β-lactoglobulin (β-LG) at pH 4.0 and 6.5. This information would provide an explanation of the variability in AFM1 during cheese production. Once β-LG models were optimized using molecular dynamic (MD) simulation, it was found that a region of the Calyx cavity underwent conformational changes, at the E-F loop, from the closed conformation at pH 6.5 to the open at pH 4.0. No differences in Site C conformation were observed at both pH. The binding free energy (ΔGb ) of the β-LG-AFM1 complexes at the different pHs were determined by molecular docking. The ΔGb values obtained for the Calyx cavity showed that at pH 4.0 there is a more stable complex formation compared to pH 6.5 with values of −42.6 and −32.0 kJ mol−1, respectively. On the contrary, in the complexes formed in Site C at both pH´s there were no differences. Likewise, the ΔGb in the dimer interface was evaluated, obtaining a value of −29.3 kJ mol−1, like those obtained at Site C. In addition, by the MD simulations of the β-LG-AFM1 complexes, it was observed that at acidic pH the binding of AFM1 with β-LG is more stable. In conclusion, the computational tools showed that the most stable complex was formed at the Calyx cavity at pH 4.0. This suggests that during cheese production using acidic coagulation, the whey proteins show higher affinity toward AFM1 which may explain the observed variability of mycotoxin. Communicated by Ramaswamy H. Sarma

A systematic review to introduce the most effective postbiotics derived from probiotics for aflatoxin detoxification in vitro

The purpose of this study was to gather information about the percentage of aflatoxin decontamination by postbiotics and to find the most effective postbiotic. In this review article, studies were collected from databases. All the articles related to experimental studies were included in the study. It appeared that the postbiotics derived from Lentilactobacillus kefiri, Lentilactobacillus kefiri, Lacticaseibacillus rhamnosum, Lacticaseibacillus rhamnosum and Pediococcus pentosaceus probiotic bacteria were the most effective postbiotics with 97.22%, 95.27%, 86.2%, 81.4% and 91% inhibitory effect against aflatoxins B1, B2, G1, G2 and M1, respectively. Therefore, postbiotics could be used as safe anti-aflatoxin agents in food products.

Comparative metabolism of aflatoxin B1 in mouse, rat and human primary hepatocytes using HPLC–MS/MS

Aflatoxin B1 (AFB1) is a highly hepatotoxic and carcinogenic mycotoxin produced by Aspergillus species. The compound is mainly metabolized in the liver and its metabolism varies between species. The present study quantified relevant AFB1- metabolites formed by mouse, rat, and human primary hepatocytes after treatment with 1 µM and 10 µM AFB1. The use of liquid chromatographic separation coupled with tandem mass spectrometric detection enabled the selective and sensitive determination of phase I and phase II metabolites of AFB1 over incubation times of up to 24 h. The binding of AFB1 to macromolecules was also considered. The fastest metabolism of AFB1 was observed in mouse hepatocytes which formed aflatoxin P1 as a major metabolite and also its glucuronidated form, while AFP1 occurred only in traces in the other species. Aflatoxin M1 was formed in all species and was, together with aflatoxin Q1 and aflatoxicol, the main metabolite in human cells. Effective epoxidation led to high amounts of DNA adducts already 30 min post-treatment, especially in rat hepatocytes. Lower levels of DNA adducts and fast DNA repair were found in mouse hepatocytes. Also, protein adducts arising from reactive intermediates were formed rapidly in all three species. Detoxification via glutathione conjugation and subsequent formation of the N-acetylcysteine derivative appeared to be similar in mice and in rats and strongly differed from human hepatocytes which did not form these metabolites at all. The use of qualitative reference material of a multitude of metabolites and the comparison of hepatocyte metabolism in three species using advanced methods enabled considerations on toxification and detoxification mechanisms of AFB1. In addition to glutathione conjugation, phase I metabolism is strongly involved in the detoxification of AFB1.

Aflatoxin M1 decreases the expression of genes encoding tight junction proteins and influences the intestinal epithelial integrity.

Aflatoxin M1 (AFM1) is a mycotoxin that is commonly found as a milk contaminant, and its presence in milk has been linked to cytotoxicity. The present study aimed to evaluate the acute cytotoxic effects of AFM1 on intestinal Caco-2 cells. Initially, we checked the morphology and viability of Caco-2 cells after treatment with different concentrations of AFM1 (5ng/L, 50ng/L, 250ng/L, 500ng/L, 1000ng/L, and 2000ng/L) for different time intervals (6h, 12h, and 24h). It was found that AFM1 did not show any effect on cell morphology, but 10% decrease in viability above 1000ng/L after 12h. Furthermore, DCFDA assay showed increased ROS production after 6h treatments. qPCR analysis showed an increased expression of epithelial-specific cytoskeleton marker genes, Cytokeratin, Villin, Vimentin, and JAM-1, and a decreased expression of tight junction protein genes, Claudin-1, Occludin, and ZO-1. Similarly, we found an increased expression of Cyp1a1 transcript with an increasing AFM1 concentration and incubation time. This gene expression analysis showed AFM1 can cause disruption of tight junctions between intestinal cells, which was further confirmed by a transwell experiment. In conclusion, consumption of AFM1-contaminated milk does not show any effect on cells morphology and viability but decreases the expression of intestinal barrier transcripts that may lead to the disruption of intestinal barrier function and leaky gut.

Occurrence of Aflatoxin M1 in yogurt samples found in markets in Kosovo during spring 2023

Aflatoxin M1 (AFM1), a toxic byproduct of aflatoxin B1 (AFB1) produced by Aspergillus fungi, is a carcinogenic mycotoxin that can contaminate various agricultural commodities. It can be transferred from AFB1-contaminated feed to milk and dairy products, including yogurt, posing a potential health risk to consumers. In spring 2023, a total of 74 yogurt samples were collected from the largest food suppliers in Kosovo for analysis, including samples produced in Kosovo and seven other countries: Albania, North Macedonia, Bosnia and Herzegovina, Slovenia, Greece, Italy, and Germany. A rapid and sensitive analytical method, Enzyme-linked immunosorbent assay (ELISA), was used for the analysis. The results of the study highlight discernible differences in the maximum tolerable levels of AFM1 between the countries. More specifically, yogurt samples from Slovenia and Germany had lower levels than those from other countries. Additionally, the median levels of AFM1 in samples from Slovenia and Germany were significantly lower. The mean concentrations of AFM1 in yogurt samples from Kosovo and other countries were 0.071 μg/kg and 0.080 μg/kg, respectively. Out of all samples, 66 (89%) exceeded the maximum tolerable limit of 0.05 μg/kg. Among the exporting countries, Albania had the highest median AFM1 level of 0.085 μg/kg and the highest maximum level of 0.195 μg/kg. Slovenia had the lowest median AFM1 level, while Germany had the lowest maximum AFM1 level. All samples from Albania, Greece, and Bosnia and Herzegovina exceeded the maximum tolerable limit. High prevalence was also observed in samples from Kosovo, North Macedonia, and Slovenia. Considering the average daily consumption of about 250 grams of yogurt, and the total median value of Aflatoxin M1 concentration (0.071 μg/kg), the estimated daily intake was calculated to be 0.017 μg. These findings highlight the importance of monitoring and enforcing regulatory limits to ensure yogurt safety and to protect public health. Efforts should be focused on mitigating AFM1 contamination and implementing measures to minimise its presence in dairy products, especially in regions where levels exceed the established limits.

Solid phase extraction of Aflatoxin M1 in milk samples by graphene oxide-poly-L-lysine nanocomposite followed by HPLC

ABSTRACT An efficient, novel and green Graphene Oxide-Poly-L-Lysine (GO-PLL) adsorbent was successfully synthesised and used for solid-phase extraction of aflatoxin M1 (AFM1) in liquid milk samples followed by high performance liquid chromatography (HPLC) equipped with fluorescence detector (FLD). The synthesised adsorbent was characterised by Fourier Transform Infrared Spectrophotometry (FT-IR), Field Emission Scanning Electron Microscopy (FESEM), Energy-dispersive X-ray spectroscopy (EDX) and X-Ray diffraction (XRD) analysis. Different parameters affecting the recovery percentage of AFM1 including pH of sample solution, amounts of adsorbent, contact time and desorbent solution (types and volume) were completely investigated and optimised. It was found that, with an enrichment factor of 15.2, the calibration curve is linear in the range of 0.05–4.00 ng ml−1 AFM1 and the limit of detection (LOD) and limit of quantification (LOQ) values were 0.019 and 0.063 ng mL−1, respectively. The relative standard deviations (RSDs) ranged from 4.5% to 5.7% and 4.2% to 6.7% for intra-day and inter-day variations, respectively. Finally, the proposed solid-phase extraction method was successfully applied for determination of trace levels of AFM1 in different milk samples.

Incidence of Aflatoxin M1 in milk powder with special reference to the effect of Gamma irradiation and Ultra violet rays on its level and their further effects on the examined samples

Incidence of Aflatoxin M1 in milk powder with special reference to the effect of Gamma irradiation and Ultra violet rays on its level and their further effects on the examined samples

Impact of Seasonal Variations (Aflatoxin M1 & Heavy metals) on Quality of Marketed Milk

This study thoroughly examined the safety and quality of commercially accessible milk in light of the growing concerns about food safety and its effects on public health. Objective: To monitor the amount of aflatoxin and heavy metals found in commercially available milk from different cities in central Punjab, Pakistan. Method: The collection of milk samples was done from three major cities in central Punjab, including Lahore, Faisalabad, and Jhang, to measure the contamination of Aflatoxin M1 and heavy metals from 2018-2019. Results: The maximum concentration of AFM1 (0.38-1.65µg/L) was observed in the semi-flush season from Lahore, followed by Faisalabad (0.37-1.63 µg/L) and Jhang (0.35-1.62µg/L) whereas, the minimum concentration of AFM1 (0.15-0.46µg/L) was observed during the lean season in the milk samples procured from Jhang, followed by Faisalabad (0.17-0.47µg/L) and Lahore (0.18-0.49µg/L). The levels of lead and cadmium in milk samples fluctuated seasonally, with Lahore having the highest contamination (Pb: 0.062ng/L, cadmium: 0.037ng/L), followed by Faisalabad and Jhang. Milk sample %ages exceeded the national and international aflatoxin M1 (AFM1) maximum residual limits (MRL). Conclusion: All milk samples from different milking seasons were above the EU's maximum residual level of 0.05µg/L. They were still within the Pakistan Standard and Quality Control Authority's (10µg/L) recommended range.

Tahran, İran'da yüksek performanslı sıvı kromatografi ile süt ve süt ürünlerinde aflatoksin M1 içeriğinin değerlendirilmesi

Aflatoxin M1 (AFM1) is the most important aflatoxin in milk and dairy products, which is carcinogenic and hepatotoxic. This study aimed to evaluate the AFM1 content in the milk and distributed dairy products in Tehran. 75 samples, including 15 samples of raw milk, 15 samples of pasteurized milk, 15 samples of ultra-high temperature milk, 15 samples of pasteurized yogurt, and 15 samples of pasteurized cheese, were collected from October to December 2020 in Tehran by simple random sampling. The dietary exposure or estimated dietary intake (EDI) and hazard index (HI) were calculated for milk and dairy product consumers. The AFM1 content in the samples was determined by using high-performance liquid chromatography (HPLC) along with a fluorescence detector. AFM1 was observed in all samples. The values of AFM1 in all samples were higher than the acceptable range determined by the European Union. 100% of milk and yogurt samples and 82% of cheese samples exceeded the Iranian maximum limit (100 ng kg-1 in milk and yogurt, and 250 ng Kg-1 in cheese). Mean AFM1 content in raw milk samples, pasteurized milk samples, UHT milk samples, pasteurized yogurt samples, and pasteurized cheese samples were 337±17.7, 306±15.5, 305±17.4, 320±17.6, and 309±18.5 ng Kg-1, respectively. The highest value of HI was observed in children of Tehran, Iran. Based on the results, the aflatoxin content in milk and distributed dairy products in Tehran in the autumn is inconvenient. It is recommended that the aflatoxin levels should be measured at different times of the year, especially in raw milk, and feed monitoring is intensified for contamination with toxin-producing molds.

Biological reduction of aflatoxin M1 in dairy products using probiotic strains: a systematic review and meta-analysis

Abstract Aflatoxins are naturally occurring toxic substances produced mainly by species of the genus Aspergillus that can contaminate almost all foodstuffs. Apart from the harmful effects they have on human and animal health, they can be secreted unchanged in animal milk and cause contamination of milk and its products. Aflatoxin M1 (AFM1) is the major and most toxic type of aflatoxin after aflatoxin B1 (AFB1). The use of probiotic strains to reduce the amount of aflatoxin in milk and by-products has been observed in many studies. In this systematic review and meta-analysis, articles in PubMed, Scopus, Google Scholar and ISI Web of Science were searched to find eligible studies that reported reduction of AFM1 using probiotics in dairy products. The results were pooled using a random-effects model. In most studies, the efficiency of probiotics in milk has been tested by Lactobacillus strains. The results indicated that probiotic microorganisms could significantly reduce AFM1 by 55.76% (confidence interval (CI): 54.35%, 57.16%; I2 = 100%). Besides, the rank order of AFM1 reduction in dairy products based on probiotic strain subgroup was: Lactobacillus 51.99% (CI: 50.14%, 53.85%, I2 = 100%), Saccharomyces 67.36% (CI: 65.05%, 69.67%, I2 = 100%), Bifidobacterium 54.80% (CI: 54.18%, 55.43%, I2 = 99.9%), and 61.90% (CI: 53.80%, 70.00%, I2 = 100%) by a mix of strains. Considering the high binding potential of AFM1 to probiotic strains, these microorganisms can be recommended as a safe system to reduce AFM1 in dairy products.

Simultaneous Detection and Quantification of Aflatoxin M1, Eight Microcystin Congeners and Nodularin in Dairy Milk by LC-MS/MS

Dairy milk holds a prominent position as a widely consumed food, particularly among infants and children. However, it is crucial to address the presence of multiple natural toxic compounds that may co-occur in dairy milk to ensure its safety prior to consumption. Aflatoxin M1 (AFM1), an emerging mycotoxin of interest, is a potential contaminant in the milk of animals who ingest aflatoxin B1 (AFB1). The toxin is regulated in the European Union under Commission Regulation No 1881/2006. Unfortunately, there is a notable lack of data concerning the transfer of various emerging microbial contaminants into dairy milk and, therefore, their natural occurrences. In this study, a simple and sensitive LC-MS/MS method was developed and validated for the quantification of multiple cyanotoxins (microcystin congeners and nodularin) and AFM1 by the main analytical guidelines. Toxins are extracted with methanol 80%, followed by an SPE clean-up step before LC-MS/MS analysis. The LOQ was fixed at 1 µg/L for the nine cyanotoxins and 0.05 µg/L for AFM1. Recoveries were measured between 82.67% and 102%. To the best of our knowledge, there are no other LC-MS/MS methods available for the simultaneous quantification of cyanotoxins and mycotoxins in milk.

Reduced graphene oxide-zinc iron oxide nanomaterial as selective dispersive solid-phase extraction sorbent for extraction and enrichment of aflatoxins from milk combined with UHPLC-MS/MS analysis

Fungal species capable of producing aflatoxins frequently contaminate the feed provided to dairy animals, leading to the subsequent occurrence of aflatoxin contamination in milk. Milk is renowned for its exceptional nutritional composition, and the consumption of milk contaminated with aflatoxins by humans can engender a multitude of health complications. In this study, reduced graphene oxide zinc iron oxide (rGO-Zn/Fe2O4) nanomaterial was utilized as a dispersive solid phase extraction adsorbent to separate and enrich aflatoxins (aflatoxin B1, aflatoxin B2, aflatoxin G1, aflatoxin G2, and aflatoxin M1) from milk. The rGO-Zn/Fe2O4 nanomaterial was morphologically characterized by scanning electron microscopy, transmission electron microscopy, and X-ray diffraction, indicating that Zn/Fe2O4 nanoparticles were embedded on the surface of rGO. The acetonitrile/formic acid (99/1, v/v) combined with ultrasonic treatment for a duration of 40 min expressed an efficient extraction solvent. Employing the matrix-spiked technique, different parameters of the dispersive solid-phase extraction (d-SPE) method were meticulously optimized, yielding the following conditions: 2% acetonitrile as loading solvent, 15 mg of rGO-Zn/Fe2O4 as the adsorbent quantity, 15 min of ultrasonication as adsorption time, and n-hexane as the washing solvent. The desorption reagent was 5 mL acetonitrile, with a desorption time of 5 min. The combination of the dispersive solid-phase extraction (d-SPE) technique with validated UHPLC-MS/MS quantitative analysis demonstrated excellent selectivity, a robust linear relationship (R2 ≥ 0.994), remarkable sensitivity (limit of quantification ranging from 0.006 to 0.013 µg/L), and satisfactory recovery rates of 78%–104% at three different matrix-spiked levels. Moreover, the method exhibited precision with relative standard deviations (RSD) ranging from 2.4% to 10.8%. Following appropriate pretreatment procedures, the milk samples underwent rigorous aflatoxin analysis, leading to the identification of AFM1 presence in all of the examined milk samples (n = 9). However, the contamination levels detected were found to be within the permissible limits as defined by regulatory standards.

The coexistence of aflatoxin M1 and ochratoxin A induced intestinal barrier disruption via the regulation of key differentially expressed microRNAs and long non-coding RNAs in BALB/c mice

Food safety can be seriously threatened by the existence of both aflatoxin M1 (AFM1) and ochratoxin A (OTA) in milk and corresponding products. The importance of intestine integrity in preserving human health is widely understood in vitro, but the fundamental processes by which AFM1 and OTA cause disruption of the intestinal barrier are as yet unknown, especially in vivo. Based on the analysis of the whole transcriptome of BALB/c mice, the competing endogenous RNA (ceRNA) regulation network was obtained in the current study. Each of 12 mice were separated into five treatments: saline solution treatment, 1.0% DMSO vehicle control treatment, 3.0 mg/kg b.w. individual AFM1 treatment (AFM1), 3.0 mg/kg b.w. individual OTA treatment (OTA), and combined mycotoxins treatment (AFM1 +OTA). The study period lasted 28 days. The jejunum tissue was collected for the histological assessment and whole transcriptome analysis, and the whole blood was collected, and determination of serum biochemical indicators. The phenotypic results demonstrated that AFM1 and OTA caused intestinal barrier disruption via an increased apoptosis level and decreased expression of tight junction (TJ) proteins. The ceRNA network demonstrated that AFM1 and OTA induced cell apoptosis through activating the expression of DUSP9 and suppressing the expression of PLA2G2D, which were regulated by differentially expressed microRNAs (DEmiRNAs) (miR-124-y, miR-194-z, miR-224-x, and miR-452-x) and differentially expressed long non-coding RNAs (DElncRNAs) (FUT8 and GPR31C). And AFM1 and OTA decreased TJ proteins via inhibiting the expression of PAK6, which was regulated by several important DEmiRNAs and DElncRNAs. These DE RNAs in intestinal integrity were involved in MAPK and Ras signaling pathway. Overall, our findings expand the current knowledge regarding the potential mechanisms of intestinal integrity disruption brought on by AFM1 and OTA in vivo.

Novel Porphyrinic Covalent Organic Polymer with Polarity-Switchable Dual Wavelength for Accurate and Sensitive Photoelectrochemical Sensing.

Herein, we synthesized a novel porphyrinic covalent organic polymer (TPAPP-PTCA PCOP) for constructing a polarity-switchable dual-wavelength photoelectrochemical (PEC) biosensor with ferrocene (Fc) and hydrogen peroxide (H2O2) as regulator and amplifier simultaneously. Interestingly, this new PCOP possessed both n-type and p-type semiconductor characteristics, which thus enabled the appearance of a dual-polarity photocurrent at two different excitation wavelengths. Furthermore, Fc and H2O2 could readily switch the photocurrent of PCOP to the cathode and anode stemming from its efficient electron collection and donation function, respectively. Based on these, a PCOP-based PEC biosensor skillfully integrating dual wavelengths with reliable accuracy and polarity switch with high sensitivity was instituted. As a result, the developed PEC biosensor exhibited a low detection limit down to 0.089 pg mL-1 for the most powerful natural carcinogen aflatoxin M1 (AFM1) assay. Impressively, the target exhibited a completely opposite photocurrent difference to the interfering substances, and the linear correlation coefficient of the assay was improved compared to single-wavelength detection. The PEC sensing platform not only provided a basis for exploring multicharacteristic photoactive material but also innovatively developed the detection mode of the PEC biosensor.

Novel dual-recognition electrochemical biosensor for the sensitive detection of AFM1 in milk

Aflatoxin M1 (AFM1) is a highly carcinogenic and toxic compound that is commonly found in dairy foods such as milk. We developed an electrochemical MIP (molecularly imprinted polymer) and aptamer (Apt) based aptasensor (Apt-Au@PEIM/AFM1/MIP–Apt/AuNPs)/GCE) with dual-recognition signal amplification to detect AFM1 in milk. The MIP–Apt/AuNP/GCE sensor was prepared by synthesizing an MIP film through the electropolymerization of resorcinol on gold nanoparticles-modified electrodes. The dropwise addition of our cApt-Au@PEIM signal amplification probe could enhance the current signal response and improve the sensitivity of the sensor to detect AFM1. The AFM1 present in milk could prevent the probe from binding to the sensor, thereby drastically reducing the current signal. Under optimal experimental conditions, the sensor exhibits a linear range of 0.01–200 nM and a limit of detection of 0.07 nM (S/N = 3). The developed dual-recognition aptasensor would provide a novel and rapid method for specific detection of AFM1 in milk.

DETERMINATION OF LEVEL OF AFLAFLATOXIN IN RAW MILK AND PASTEURIZED MILK SAMPLES

Aflatoxins are toxic compounds produced mainly by different genera of fungi and transferred to milk through contamination which causes life health hazards. The objective of the current study was to govern its existence in different raw and branded liquid milk samples sold in different areas of District Lahore in Pakistan. For this purpose, 80 samples ,40 raw and 40 branded milk samples were obtained during the months of July and August and analyzed to determine the level of AFM1 in milk by using ELISA technique. Typically, the investigative measures go through the subsequent stages: sampling, extraction, determination and quantification. 50% of the samples were positive for Aflatoxin M1, with a range of 49.73-429.61 ppt. Contamination beyond EU permissible limits for raw milk samples was 67.5% with 52.75-429.60 ppt range and 203.42 ppt mean. Samples collected from Shalimar Tehsil were observed more contaminated than others. On the other hand, branded milk contamination beyond EU limit was 30% with range of 49.73-381.04 ppt and 138.53 ppt mean. Consequently, Greater level of AFM1 in raw milk is a communal health risk, but a checking and scrutiny program for Aflatoxin M1 control in milk industry should be established to avert well-being harms.