Using multi-toxin analysis to overcome challenges in mycotoxin analysis
by R-Biopharm, UK
Laboratories are typically presented with a wide variety of samples and include solid samples like cereals, which could be classified as an easy matrix. However, they may also consist of more complex samples like animal feed or spice mixtures. A number of samples may also be liquids like fruit juices and sauces.
Regardless of the format of the sample, all will contain components other than the analyte of interest and may be a mix of carbohydrates, sugars, fats, salts, proteins and pigments. All of these components can result in what is referred to as sample matrix. The matrix can have a considerable effect on the way the analysis is conducted and the quality of the results obtained; such effects are called matrix effects.
In order to obtain good, reliable results consistently the sample matrix should be removed. It is, therefore, necessary to ensure that a suitable sample extraction and clean-up method are employed.
The removal of any sample matrix will reduce any issues with:
- Blockages within the analytical detection system which can ultimately lead to downtime where no samples are analysed
- False positives, as positive samples are normally required to be re-analysed in order to confirm results. This leads to an increase in analysis time and potentially a reduction in overall margin
- Any results with high %RSD, which indicate poor accuracy and reproducibility and again would need to be re-analysed
- Poor sensitivity. Sensitivity is important to ensure you are complying with legislative levels and to ensure that you are not reporting false negatives.
There are several options available to laboratories analysing samples. The first is to use direct injection or dilute and shoot methods. These are typically used prior to LC-MS/MS detection, however, it should be noted that these methods are the most basic form of clean-up and, ultimately, will still result in some matrix effect being observed. These will need to be corrected for by using isotopic labelled standards and potentially also matrix matched standards.
Solid phase columns are also typically used prior to LC-MS/MS detection, however, again are considered as a basic form of clean-up and, as a result, are most commonly used for the analysis of simple commodities like certain cereal samples.
Alternatively, immunoaffinity clean-up is available. Although traditionally immunoaffinity columns were focused on single toxins, the market is changing and there are now various multi-toxin options available which allow you to clean-up any sample type, effectively removing all matrix components. This ultimately means that you have a clean solvent-based eluate and removes the need for the use of isotopic labelled and matrix matched standards, resulting in quicker and more cost-effective analysis.
Everyone is facing new challenges on a day to day occurrence and in general labs are facing the same challenges:
- Time pressure. This may be as a result of increased sample numbers or it may be that more and more samples are having to be analysed for increased numbers of mycotoxins
- Again, demands to increase sample throughput may be due to sample numbers or they may be to increase overall turnaround
- Staff and general costs may be increasing, therefore, harmonisation of methods could offer some benefits
- As more and more samples are having to be analysed for more than one mycotoxin, it makes sense to try and harmonise methods, reducing time spent on extractions and ultimately cost of consumables.
The use of multi-mycotoxin immunoaffinity columns present an opportunity to manage these challenges in the laboratory while still maintaining quality results. Typically, the aim for any busy laboratory is to implement as few methods as possible. Therefore, if analysing a wide variety of samples on a daily basis, the best option may be to consider multi-toxin clean-up options.
Current legislation also highlights that a multi-toxin approach could be advantageous, and many laboratories are also considering multi-toxin analysis as a way to reduce overhead costs. Mycotoxin regulations are complex, with different limits applied to specific commodities. Figure 1 illustrates a simplistic overview of the key toxins regulated in specific foodstuffs.
If you are analysing only nuts, nut products and peanuts on a daily basis, it makes no sense to analyse using a multi-toxin method. However, for dried fruit and spices there are levels for both aflatoxin and ochratoxin, so in this case a multi-method targeted at this combination may be of interest. Cereals present a different situation where the fusarium toxins are regulated along with aflatoxin and ochratoxin. Baby food and animal feed are also regulated for the majority of toxins and as a result a multi-toxin approach could offer many benefits.
Immunoaffinity columns can be used for any sample type and offer particular benefits with complex or coloured samples as they are the only form of clean-up that completely removes all co-extractives from the sample, resulting in clean eluates. In addition, due to the concentration effect, the columns are also particularly suitable for samples like baby food where detection at low legislative levels are required.
Multi-toxin analysis can ultimately reduce the time spent at the bench and allows more samples to be analysed on a daily basis, ultimately helping you to achieve many of the challenges encountered. In addition, the use of multi-toxin immunoaffinity columns can reduce costs as there are savings in:
- Storage of columns
- Transportation of products
- Overall waste disposal costs
- Consumable costs such as solvents and disposables used
- Labour as number of man hours taken to run analysis for numerous toxins is decreased.
Typically, immunoaffinity columns are used in official methods like AOAC and CEN methods as they have been successfully used in various collaborative trials where it has been widely demonstrated that consistent, reliable results can be achieved. Essentially, immunoaffinity columns are considered as high-quality products, however, they allow you to meet the increasingly stringent quality standards required.
However, it should be noted that not all immunoaffinity columns behave in the same way and performance and results very much depend on the solvent used during extraction, the overall clean-up method and the antibodies used in the immunoaffinity column of choice. The use of a good, reliable immunoaffinity column is vital in ensuring accurate results and can help to reduce the number of samples re-analysed due to poor chromatography and results.
R-Biopharm can offer a number of flexible options suit your requirements. We offer a range of innovative products and methodologies that not only improve and optimise your workflow but also allow you to maximise sample throughput. The products allow you to easily analyse for a number of mycotoxins simultaneously from a diverse range of commodities effectively whilst removing all matrix effects resulting in accurate and reliable results every time.