Study on the Chemical Components of the Endophytic Strain Penicillium crustosum in Cordyceps sinensis
Ophiocordyceps sinensis is a complex of the subphylum of the linear fungus Ophiocordyceps and the larvae of its host bat moth family insects. It is a special species distributed in the high-altitude areas of the Qinghai Tibet Plateau. It has a flat nature, sweet taste, and is beneficial to the lungs and kidneys. It has the effects of resolving phlegm, stopping bleeding, nourishing the lungs and kidneys. Due to the unique therapeutic effects of Cordyceps sinensis, the market demand has increased, and natural cordyceps has been overexploited, leading to increasingly scarce resources and attracting widespread attention. Through long-term coevolution, endophytic bacteria in traditional Chinese medicine can produce active substances that are the same or similar to those in the host plant. For example, previous studies have reported the development of Bailing capsules from the mycelium of Cordyceps sinensis isolated from Cordyceps sinensis, Jinshuibao capsules from the fermented mycelium of Batworm Moth Penicillium, and Zhiling capsules from Batworm Moth. Therefore, endophytic bacteria in Cordyceps sinensis have great research potential.

Penicillium crustosum is a fungus belonging to the genus Penicillium in the Discellaceae family of the order Hemimycetes. Literature review: The study of secondary metabolites in P. crustosum focuses on liquid fermentation, and a series of alkaloid compounds including thiodiketopiperazines, benzodiazepines, indole diterpenes, quinolines, and novel polyketide compounds with clavitol units and gamma butyrolactone units have been isolated. He et al. isolated three new thiodiketopiperazines and two analogues from the secondary metabolites of P. crustosum. These two compounds can promote gastrointestinal motility in zebrafish treated with Nile red at doses>8 μ M and>16 μ M, respectively. Further interference with atropine suggests that the compounds may exert their effects by acting on the cholinergic nervous system; Liu et al. isolated a new polyene compound, a new diketopiperazine, and three known compounds from P. crustosum. The new diketopiperazine F has cytotoxic effects on K562 cells (human chronic myeloid leukemia cells) with an IC50 value of 12.7 μ M; Feng et al. isolated a pair of new enantiomers, an analog, and a new polyketide compound from their liquid fermentation products. The analog showed a slight inhibitory effect on MV4-11 (human myeloid monocytic leukemia cells) cells with an IC50 value of 36.23 μ M. However, there is relatively little research on the solid fermentation products of P. crustosum rice. In order to further enrich the chemical information of Penicillium crustosum metabolites and compare the effects of different fermentation methods on their secondary metabolites, and to reasonably develop the resources of Cordyceps sinensis, this paper uses various chromatographic separation techniques and modern spectroscopic techniques to cultivate the endophytic fungus Penicillium crustosum in Cordyceps sinensis using rice solid fermentation and study the chemical composition of its metabolites.

This article isolated 16 compounds from the rice solid fermentation product of the endophytic fungus P. crustosum of Cordyceps sinensis, mainly including benzodiazepines, indole diterpenes, and quinolone alkaloids. Among them, compounds 1 and 2 are new natural products, and compounds 1-4, 6, 9, 12-14, and 16 were isolated from this fungus for the first time. Similar to the results obtained by the research team in the early stage. Previous reports have shown that alkaloids have a wide range of pharmacological activities, such as anti-tumor, anti-inflammatory, and analgesic effects. This article conducted preliminary antibacterial tests on the isolated compounds, but none of them showed significant inhibitory activity. It is speculated that the selection of test strains is limited or that these compounds have other biological activities. The literature shows that compound 4 has a strong inhibitory effect on influenza neuraminidase, with an IC50 value of 5.02 μ M; Compound 7 has inhibitory effects on Balanus amphoteride and Bugula neritina, and exhibits anti fouling and antibacterial activity, with EC50 values of 6.2 and 2.1 μ g/mL, respectively. This suggests that we can explore the biological activity of such compounds from other perspectives, providing reference for subsequent experiments.

There is a significant difference in the types of metabolites between liquid culture of Penicillium crustosum and solid culture of rice. The liquid culture products contain more thiodiketone piperazine compounds, which have been reported to have strong toxicity due to the presence of disulfide bonds. However, the alkaloids in rice solid fermentation products do not have sulfur atoms, possibly due to the absence of relevant elements in the culture medium. Therefore, when studying the secondary metabolites of endophytic bacteria in traditional Chinese medicine, novel bioactive compounds can be obtained by changing the cultivation method.