Institut Kirchhoff Berlin GmbH

Determining pyrollizidine alkaloids using online SPE-HPLC-MS/MS

Date: March 2016


Pyrrolizidine alkaloids (PA) are secondary plant ingredients that are formed in a multitude of plant types found all over the world to protect against predators. Over 600 of these compounds are currently known. They chiefly appear in the plant genera boraginaceae, asteraceae und fabaceae. Contamination of animal products such as honey (enters via nectar from plants containing PAs) is also known.

In recent years, PAs have become the focus of interest in the press, thanks mainly to the results of countless analyses of teas and herbal teas being published.

Over the last few years, our institute has analysed a large number of samples for PAs (predominantly herbal tea, tea and honey, but also pharmaceutical products). In doing so, we set great value on critical questioning of our analysis procedures, professional dialogue with other institutions working in the same area and optimising our analysis procedures.

So in February 2014, we organised a status seminar with the title: ‘Tea and honey – Old foods with new challenges – Pyrrolizidine alkaloids between scandalisation and trivialisation’. It featured presentations by well-known speakers from the German Federal Institute for Risk Assessment (BfR) as well as from the worlds of food monitoring, research institutions, industry and laboratories.

We also contributed poster submissions on the analysis of PAs to the German Food Chemists’ Days in 2014 and 2015 (see download area). We will also contribute a poster on this topic to this year’s Austrian Food Chemists’ Day.

Toxicological classification

Due its potential to endanger health, 1.2 unsaturated PA in particular in foodstuffs and pharmaceutical products can be classified as posing a risk to health. In higher doses, it can lead to acute liver damage. If PAs are ingested over a longer period of time, different types of organ damage (especially liver damage) can be expected.

Legal classification

There is currently no maximum PA content level in foodstuffs.

EFSA’s CONTAM panel decided that, due to the genotoxic and carcinogenic effect of 1.2 unsaturated PA, it would not be appropriate to state a TDI value. According to the margin of exposure (MoE) approach, a target value/reference value was set for a tolerable daily maximum intake of 0.007 µg/kg body weight.

In medicinal products for internal use, a maximum daily dose of 1 µg PA for maximum 6 weeks a year applies, although not during pregnancy and lactation. If daily exposure is less than 1 µg, the time-based use limit does not apply.

For determining the PA exposure of teas and herbal teas, the BfR recommends stating the total sum of the content of 21 PAs that are especially relevant. An extension of the analysis criteria is desirable as soon as the relevant standards become commercially available.


Online SPE-Module with cartridge changer
Online SPE-Module with cartridge changer

At the Institut Kirchhoff Berlin GmbH, pyrrolizidine alkaloids are analysed using a further development of the BfR method ‘Determining pyrrolizidine alkaloids (PA) in plant matter using SPE-LC-MS/MS’, which covers 28 analytes. These 28 analytes include the 21 PAs with particular relevance to teas and herbal teas, which are recommended by the BfR for establishing total content.

In order to process larger series of samples cost-efficiently and quickly, the Institut Kirchhoff Berlin GmbH replaces time-consuming manual work steps (purifying using solid-phase extraction and the resulting concentration of the extract) with an online SPE procedure. The online SPE system is integrated into the LC-MS/MS analysis system. All components are controlled via a standardised software interface. In the framework of validation, we were able to demonstrate the parity of this efficient procedure with established offline SPE.

Depending on the question at hand, instead of or as well as the 28 PAs covered by the BfR method, it can be a good idea to determine a total PA content that takes into account a great deal more individual compounds. We have therefore also established a PA cumulative method.

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