To establish a molecular method for the authentication of spices that can be used under different conditions for qualitative or semi-quantitative analyses.

The real-time polymerase chain reaction (PCR) assay was developed based on SYBR GREEN I. Specific primers were successfully designed for the detection of star anise, fennel, pepper and prickly ash.

This method was able to specifically detect star anise, fennel, pepper and prickly ash with good linearity being found between Ct values and standard volume fraction in the range of 0.001%–10%. The limit of detection (LOD) was 1% for star anise standard, and 0.001% for fennel, pepper and prickly ash standards. The developed method was used to detect five actual samples. Star anise, fennel, pepper and prickly ash were not detected in sample 1; Star anise, fennel and pepper were detected in samples 2, 3 and 5; Star anise, fennel, pepper and prickly ash were detected in sample 4.

This method has high sensitivity and good repeatability, and can be applied to the detection of real samples.