The establishment of TaqMan qPCR method for the identification and quantification of Amynthas aspergillum originated medicinal products

Le résumé fourni par la source

“Guang Dilong” (GDL, Amynthas aspergillum ), a pharmacopeial “DiLong” (DL, Pheretima ) source in traditional Chinese medicine, faces widespread adulteration of various earthworm species. To enable specific and precise quantification of GDL in dried and highly processed medicinal products, a TaqMan qPCR assay targeting species-specific (including the newly classified subgroup of GDL) and earthworm-universal sequences with short amplicons (<100 bp) was established. The assay was validated for specificity (7 species tested), sensitivity (LOD = 0.0001 ng/μL), amplification efficiency (100.16 %), and repeatability (inter-day precision CV < 1.6 %). Two relative quantification approaches (ΔCT and ΔΔCT) were compared for accuracy under different conditions. The ΔCT method offers a simple workflow suitable for matrices with high homogeneity, while the ΔΔCT method employs dynamic normalization to achieve enhanced robustness and accuracy in complex and processed samples. Quantitative deviation remained within ±18 % for formula granule and decoctions. For Xiaohuoluo pills, GDL content ranged from 0.07 % to 62.42 %, showing highly consistency with droplet digital PCR assay ( r = 0.9988， P < 0.0001). This is the first study to identify and quantify the origin of GDL in both decoction pieces and downstream medicinal products, including highly processed forms and those with complex compositions, and it demonstrates the strong applicable potential for authenticity verification of GDL across the entire product chain.

Ce résumé expose les affirmations des auteurs. BNTIC ne l’interprète pas comme une validation indépendante des résultats.