Aflatoxins in hazelnuts - cleaned up with AflaCLEAN SMART
LCTech GmbH
Our Solutions for Your Laboratory: Sample Preparation for Food, Feed, Environmental and Toxicological Samples
Sample Preparation
The RASFF identified more than 240 complaints for nuts and nut products in 2021 that were due to mycotoxins. These mostly resulted in import bans or rejections at EU borders. Currently, nuts represent the largest fraction of food products rejected due to mycotoxins in the area of import at EU borders. Hazelnuts are frequently contaminated with aflatoxin G1/G2 but also B1.
AflaCLEAN SMART - Small + Fast + Economical = SMART
The AflaCLEAN SMART columns are available in a small 3 cm polypropylene format, which has significant handling and storage advantages over larger columns.
The advantages at a Glance:
The processing time for extraction, dilution, washing and elution is significantly reduced. For example, no column emptying is necessary and loading the sample and washing the column only takes a maximum of 4 minutes with a 10 mL sample feed and a flow rate of 3 mL/min. Since a maximum of only 400 μL is also used for elution, this reduces the processing time even further.
The functional mechanism is based on the principle of immunoaffinity chromatography with increased flow rates and an adapted, low elution volume to save processing time. The material in the column is coated with antibodies directed against aflatoxin B and G. The sample extract is loaded onto the column. When the sample extract is loaded, the aflatoxins B and G are retained in the column while the matrix components pass through the column. After a washing step, the aflatoxins B and G can be quantitatively eluted from the column by methanol and subsequently analysed by HPLC.
Manual processing with AflaCLEAN
Extract 20 g of homogenized hazelnut with 100 mL of 80/20 (v/v) methanol/ water. Add 2 g sodium chloride and 50 mL n-hexane during extraction to separate fats and oils.
After stirring for 10 min, filter the crude extract and optionally centrifuge at 3000 x g to ensure efficient phase separation between the methanolic extract and the n-hexane. Now dilute 3.5 mL of the n-hexane free extract with 21.5 mL of PBS buffer. Load 10 mL of this diluted sample onto the AflaCLEAN SMART column at 3 mL/min.
Then rinse the sample tube with 2 mL of deionized water and load this onto the column as well. After drying the column, elute the toxin through 400 μL methanol and measure an aliquot after dilution to run ratios in HPLC using fluorescence after photochemical derivatization by the UVE.
Automation with the FREESTYLE ThermELUTE? robotic system
The sample clean-up process can also be fully automated on a FREESTYLE system with TheremELUTE? module. Parameterization is performed via the software, the sample list is generated, and the sample introduction vessels and columns are placed in the instrument. After less than 30 minutes, the first result can be assessed using the chromatography software. After this first cycle, another result follows every 20 minutes, so a throughput of up to 120 samples in 24 hours can be achieved.
Automation with the FREESTYLE ThemELUTE? robotic system enables high sample throughput (500 samples/week). The system can be combined with a direct injection module with any HPLC or LC-MS system - the result is comprehensive automation with processing from raw extract to finished chomatogram without manual steps with convincing advantages:
Conclusion
Consistent recoveries in the range of 2 - 20 ppb (total toxin). Recoveries are reproducivle, fast, efficient and samples can be analyzed fully automated with good chromatographic performance.