Rapid and reliable authentication of Artemisiae Annuae Herba and Artemisiae Iwayomogii Herba using ITS-based SCAR markers

Artemisia is commonly used in traditional East Asian medicine and has high industrial demand due to its bioactive compounds, including artemisinin and its derivatives such as artesunate and artemether. In Korea, two key herbal medicines derived from Artemisia are Artemisiae Annuae Herba (AAH) and Artemisiae Iwayomogii Herba (AIH). However, the diversity of the genus and the morphological similarities among these species complicate accurate identification, increasing the risk of adulteration. This study aimed to develop a rapid and reliable molecular authentication method for AAH and AIH using sequence-characterized amplified region (SCAR) assays based on variations in internal transcribed spacer (ITS) sequences. We collected 71 authenticated specimens representing 17 Artemisia species in Korea. Three DNA barcode regions (rDNA-ITS, psb A– trn H, and trn L–F) were analyzed, with rDNA-ITS showing superior discriminatory power. We designed species-specific SCAR primers for 10 Artemisia species, which exhibited high specificity and sensitivity, with a detection limit of 10 pg (0.1%). Additionally, we verified the species identities of commercial products (seven AAH and five AIH) using both SCAR-PCR and DNA barcoding. Only SCAR-PCR detected adulteration, identifying two of the seven AAH products containing the adulterant species A. sieversiana . This study presents a reliable SCAR-based authentication method for AAH and AIH, thereby enhancing the accuracy and efficiency of species identification and supporting the standardization and safety of these Artemisia -derived herbal products for industrial applications. • rDNA-ITS-based SCAR-PCR assay was developed for Artemisia species authentication. • The assay accurately discriminated between AAH and AIH with 10 pg sensitivity. • Twelve commercial samples were tested, revealing adulteration in AAH products. • The method offers a rapid and reliable alternative to conventional DNA barcoding. • This system supports routine quality control of Artemisia -derived herbal industries.