New DNA Tests Expose Spice Fraud Across Popular Herbs and Spices

By Pooja Toshniwal PahariaReviewed by Susha Cheriyedath, M.Sc.Apr 28 2026

A Europe-wide validation study shows that DNA-based qPCR methods can reliably detect and quantify adulterants in some of the world’s most widely used spices and herbs, strengthening the fight against food fraud.

Study: Interlaboratory validation of thirteen qPCR methods to quantify adulterants in culinary spices and herbs. Image Credit: alexkoral / Shutterstock

In a recent study published in the journal European Food Research and Technology, researchers at the European Commission, Joint Research Centre (JRC), Geel, Belgium, interlaboratory-validated 13 real-time deoxyribonucleic acid (DNA)-based quantitative polymerase chain reaction (qPCR) methods, selected from a previously in-house validated set of 30 methods, to detect and quantify key adulterants in six spices and herbs widely used in the daily diet.

These included paprika/chili, saffron, turmeric, oregano, black pepper, and cumin. They strengthened the evaluation with an interlaboratory comparison (ILC) involving 25 laboratories overall, with each method tested by 15 Europe-based laboratories.

The assays showed strong analytical performance, with repeatability ranging from 4% to 25%, reproducibility generally from 6% to 25% (one exception at 28.5%), and trueness bias between −11% and 27%.

The results support their potential inclusion in international technical standards and use in official control laboratories, supporting food authenticity and consumer protection.

Spice Adulteration and qPCR Authentication Background

Culinary spices and herbs are high-value commodities vulnerable to fraud through adulteration, substitution, and contamination with heavy metals, synthetic dyes, or cheaper plant materials.

These practices compromise authenticity and raise concerns for consumer health and trust. European Union (EU) monitoring systems, including the Coordinated Control Plan (EUCCP) and Rapid Alert System for Food and Feed (RASFF), highlight the need for robust authentication tools.

In response, the European Commission has promoted reliable, standards-aligned analytical methods. Although DNA-based approaches show promise, they have not yet undergone full interlaboratory validation for herbs and spices, leaving a key gap in standardized testing in routine food authentication.

Interlaboratory qPCR Validation Study Design

In this study, researchers interlaboratory-validated 13 qPCR methods that had previously met in-house validation criteria for detecting the five most common adulterants in spices and herbs, including paprika/chili, saffron, turmeric, oregano, black pepper, and cumin. Methods meeting predefined criteria were then advanced to an ILC involving 25 participating laboratories overall.

The team selected laboratories based on technical capacity, requiring experience with 96-well format qPCR systems and at least annual instrument calibration. Interested laboratories first completed a questionnaire assessing their technical expertise, analytical capability, and ability to manage the required sample workload within the timeframe.

Each selected laboratory received standardized DNA templates prepared centrally by the Joint Research Center–Geel (JRC-GEEL), ensuring uniformity across all analyses. These included five calibration samples with known mixing ratios, five blinded test samples with unknown adulterant concentrations, no-template controls (NTCs), primers, and qPCR master mix. Participants reported cycle threshold (Cq) values using a standardized reporting template within one month.

To generate the reference materials, the researchers prepared binary plant mixtures gravimetrically, followed by cetyltrimethylammonium bromide (CTAB)-based DNA extraction, semi-automated purification, and fluorometric quantification. They evaluated a range of adulterant–matrix combinations, such as maize in paprika, safflower in saffron, and rice in black pepper.

Before distribution, the researchers determined limits of detection (LOD) and limits of quantification (LOQ) in-house using serial dilutions and repeated qPCR runs across multiple days. Measurement uncertainties were then assessed from the interlaboratory performance data.

For the ILC, they analyzed the samples over two consecutive days under repeatability conditions to assess method performance. Key validation parameters included reproducibility, repeatability, and trueness.

qPCR Performance Across Participating Laboratories

Of the 48 laboratories that responded to the questionnaire, 25 were selected based on technical capability, instrument calibration status, experience in food fraud detection, and capacity to use multiple methods.

Each method was tested by 15 laboratories, with the full ILC conducted across two participation periods. The 13 qPCR methods demonstrated strong and consistent analytical performance across laboratories. Relative standard deviation of repeatability (RSDr) ranged from 4.0% to 25%, while relative reproducibility (RSDR) was generally between 6.0% and 25%, with one exception at 28.5% for paprika mixed with sunflower-derived press cake. Trueness bias remained within −11% to 27% across all methods, supporting overall analytical reliability.

Importantly, RSDr stayed below 25% across all tested binary mixtures, even at the lowest levels of adulteration (0.5–2.0 g per 100 g depending on the spice matrix). This met and, in many cases, exceeded the minimum performance requirements for genetically modified organisms (GMOs) and food testing. A lack of systematic trend between concentration levels and RSDr indicated stable performance across the full analytical range.

Reproducibility was also robust, with all but one value remaining below the 25% threshold. The exception was observed in paprika mixed with 1.0 g per 100 g sunflower-derived press cake (28.5%). However, this RSDR value remained below the acceptable limit of 35%, confirming overall interlaboratory consistency.

Trueness assessment further supported method reliability. In-house evaluations showed bias below 25% across all concentrations, a finding largely confirmed by the ILC results. Only one case marginally exceeded this threshold in oregano with 5.0 g per 100 g goosefoot (27%).

Implications for Official Spice Testing

The interlaboratory comparison confirms that the 13 qPCR methods are robust, reliable, and suitable for routine use in official food control laboratories. They consistently met international validation criteria and showed strong performance in detecting and quantifying adulterants in commonly used spices. However, they should be applied within a broader analytical framework rather than as standalone tools.

The authors also noted that DNA extraction and purification were not part of the ILC, and that biological variability in plant materials can affect DNA-based quantification. These methods can complement screening approaches like next-generation sequencing and digital PCR, as well as chemical and traceability-based investigations.

Moving forward, digital PCR and integrated weight-of-evidence strategies offer promising pathways to further strengthen food fraud detection.

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