Identification of Cordyceps cicadae and Tolypocladium dujiaolongae based on ITS sequences and chemical pattern recognition method / 中国中药杂志

Based on ITS sequences, the molecular identification of Cordyceps cicadae and Tolypocladium dujiaolongae was carried out, and high-performance liquid chromatography(HPLC) fingerprint combined with chemical pattern recognition method was established to differentiate C. cicadae from its adulterant T. dujiaolongae. The genomic DNA from 10 batches of C. cicadae and five batches of T. dujiaolongae was extracted, and ITS sequences were amplified by PCR and sequenced. The stable differential sites of these two species were compared and the phylogenetic tree was constructed via MEGA 7.0. HPLC was used to establish the fingerprints of C. cicadae and T. dujiaolongae, and similarity evaluation, cluster analysis(CA), principal component analysis(PCA), and partial least squares discriminant analysis(PLS-DA) were applied to investigate the chemical pattern recognition. The result showed that the sources of these two species were different, and there were 115 stable differential sites in ITS sequences of C. cicadae and T. dujiao-longae. The phylogenetic tree could distinguish them effectively. HPLC fingerprints of 18 batches of C. cicadae and 5 batches of T. dujiaolongae were established. The results of CA, PCA, and PLS-DA were consistent, which could distinguish them well, indicating that there were great differences in chemical components between C. cicadae and T. dujiaolongae. The results of PLS-DA showed that six components such as uridine, guanosine, adenosine, and N~6-(2-hydroxyethyl) adenosine were the main differential markers of the two species. ITS sequences and HPLC fingerprint combined with the chemical pattern recognition method can serve as the identification and differentiation methods for C. cicadae and T. dujiaolongae.

Chemical Constituents from Mycelia and Spores of Fungus Cordyceps cicadae / 中草药·英文版

Objective To isolate and identify the chemical constituents from mycelia and spores of the fungus Cordyceps cicadae, respectively. Methods The chemical constituents were isolated and purified by repeated silica gel, Sephadex LH-20, and reversed phase HPLC. The structures of these compounds were elucidated on the basis of extensive spectroscopic analysis, including 1D and 2D NMR. Results Nine known sterols such as ergosterol (1), ergosterol peroxide (2), 9,11-dehydroergosterol peroxide (3), 3β,5α,9α-trihydroxy-(22E,24R)-ergosta-7,22-dien-6-one (4), 3β,5α,9α,14α-tetrahydroxy-(22E, 24R)-ergosta-7,22-dien-6-one (5), 5α,6α-epoxy-(22E,24R)-ergosta-8(14),22-diene-3β,7α-diol (6), 3β,5α,6β-(22E,24R)-ergosta-7,22-dien-3,5,6-triol (7), 3β,5α,6α-6-methoxyergosta-(22E,24R)-7,22-diene-3,5-diol (8), 4-hydroxy-17R-methylincisterol (9), together with a resorcinol derivative, 5-n-nonadecylresorcinol (10), a cyclodesipeptide, beauvericin (11), and a nucleoside, N6-(2-hydroxyethyl)adenosine (12) were successively isolated from the cultivated C. cicadae mycelia and spores. Conclusion Compounds 3–10 are reported for the first time from the title sample, beauvericin exhibits significant cytotoxicity against human leukemia cell line and human lung cancer cell line.

Characterizations of a new Cordyceps cicadae isolate and production of adenosine and cordycepin

Cordyceps is a fastidious pathogenic fungus infecting insects, and recent years have witnessed rapid progress in its medical properties. In this study, a wild isolate, C. cicadae MP12, was characterized through in vitro cultivation and its nuclear small-subunit (SSU) ribosomal DNA (rDNA) data. In vitro culture of C. cicadae MP12 was established by growing its fruiting bodies in a solid matrix. C. cicadae MP12 was inoculated into Cryptotympana atrata cicada pupae for in vivo culture, where the fungi developed its fruiting body as well. The contents of adenosine and cordycepin in dried fruiting bodies after culture were 1421.45µg/g and 1398.12 µg/g, respectively. Therefore, the established cultures from this study could be used for the production of various medically important metabolic substances.