Specific DNA identification of Pheretima in the Naoxintong capsule

Abstract

Background Pheretima is a minister drug in Naoxintong capsule (NXTC), a well-known traditional Chinese medicine (TCM) formula for the treatment of cardiovascular and cerebrovascular diseases. Owing to the loss of morphological and microscopic characteristics and the lack of recognized chemical marker, it is difficult to identify Pheretima in NXTC. This study aims to evaluate the feasibility of using DNA techniques to authenticate Pheretima, especially when it is processed into NXTC. Methods DNA was extracted from crude drugs of the genuine and adulterant species, as well as nine batches of NXTCs. Based on mitochondrial cytochrome c oxidase subunit I (COI) gene, specific primers were designed for two genera of genuine species, Metaphire and Amynthas, respectively. PCR amplification was performed with the designed primers on crude drugs of Pheretima and NXTCs. The purified PCR products were sequenced and the obtained sequences were identified to species level with top hit of similarity with BLAST against GenBank nucleotide database. Results Primers MF2R2 and AF3R1 could amplify specific DNA fragments with sizes around 230–250 bp, both in crude drugs and NXTC. With sequencing and the BLAST search, identities of the tested samples were found. Conclusion This study indicated that the molecular approach is effective for identifying Pheretima in NXTC. Therefore, DNA identification may contribute to the quality control and assurance of NXTC.

Naoxintong capsule for treating cardiovascular and cerebrovascular diseases: from bench to bedside

“…In order to fill this gap, our group used DNA barcoding molecular identification technology to design the specific primers of Hirudo, Scorpio, and Pheretima, which can be used to identify whether the sample of NXT contains the corresponding authentic animal drugs, greatly improving the quality control of NXT. Using DNA barcoding technology, we developed a method to authenticate zoological drugs including Hirudo, Scorpio, and Pheretima in NXT ( Zhu et al, 2019a ; b ; 2020 ). Specific primers were designed through mitochondrial cytochrome c oxidase subunit I (COI) gene sequence for genuine species, namely, Metaphire and Amynthas generas of Pheretima, Whitmania pigra and Hirudo nipponica of Hirudo, Mesobuthus martensii of Scorpio, respectively.…”

Establishment of an anti-inflammation-based bioassay for the quality control of the 13-component TCM formula (Lianhua Qingwen)

“… 2014 ; Zhu et al. 2019 ). The efficacy of TCM is often due to the overall effects of multiple components (Jiang et al.…”

Uncovering the mechanism of Naoxintong capsule against hypertension based on network analysis and in vitro experiments

“…The makeup of NXT can be found in the book “Yi Lin Cuo Gai”, comprising 16 kinds of traditional Chinese medicinal ingredients, including Hedysarum Multijugum Maxim (HMM), Radix Paeoniae Rubra (RPR), Radix Salviae (RS), Angelicae Sinensis Radix (ASR), Chuanxiong Rhizoma (CXR), Persicae Semen (PS), Carthami Flos (CF), Spatholobus Suberectus Dunn (SSD), Achyranthis Bidentatae Radix (ABR), Cinnamomi Ramulus (CR), and Ramulus Mori (RM) (Han et al., 2019; Zhu et al., 2019). This drug has the effect of replenishing qi, improving blood circulation and removing blood stasis, and it is primarily used to treat coronary heart disease, ischemic cerebrovascular disease, angina pectoris, and stroke (GBD 2016 Disease and Injury Incidence and Prevalence Collaborators, 2017; Yellon & Hausenloy, 2007).…”