Lethariella cladonioides (Nyl.) Krog (EE) proved to be a prolific source of polyphenols. UHPLC-Q-TOF-MS/MS was used to identify 38 polyphenolic compounds in L. cladonioides and determined the fragmentation pathways of its unique compounds (Vermicularin). The chosen polyphenols were utilized in the COSMO-RS method to tailor the most appropriate deep eutectic solvent (DES) for L. cladonioides . The Materials Studio program was used to construct virtual cluster molecules from 34 types of DESs and the 8 chosen polyphenols. The 17 DESs were configured and used to extract the polyphenols. The results indicated that choline chloride-ethanol amine exhibited outstanding dissolution effects. Furthermore, ultrasound-assisted DES extraction process parameters were optimized using response surface methodology, including ultrasound time, ultrasound power, and temperature and the optimization significantly increased the total phenol content to 41.17 ± 0.24 mg GAE/g. In the study, 38 polyphenol components in L. cladonioides were analyzed by mass spectrometry and ChCl-Eth was successfully screened out as the optimal extraction solvent based on the modeling, providing new ideas for the application of molecular simulation in the field of polyphenol extraction.
Microreactor technology has demonstrated significant advantages in natural product extraction due to its superior mass transfer
efficiency. This study optimized the extraction of essential oil from L. cladonioides leaves using a continuous flow microreactor
through Box-Behnken experimental design. An extraction yield of 1.81% was achieved under optimal conditions (74 μm powder,
43°C, 2.5 h, ethanol solvent). GC-MS analysis identified 29 compounds accounting for 97.59% of total content, predominantly
terpenoids (91.33%), with α-pinene (24.88%), β-pinene (19.65%), and p-mentha-1,8-diene (16.97%) as major constituents. Eight
compounds, previously unreported, were detected in this study. The essential oil demonstrated potent antibacterial activity against
Gram positive (S. aureus MIC = 1.4 μL/mL), Gram negative (E. coli MIC = 1.5 μL/mL) bacteria, and fungi (C. albicans MIC
= 1.7 μL/mL), exhibiting superior antifungal efficacy compared to amoxicillin. Antioxidant assays revealed moderate DPPH
radical scavenging capacity (IC50 = 37.69 μg/mL) and β-carotene/linoleic acid system inhibition (IC50 = 0.77 mg/mL). Antibacterial
mechanisms involved disruption of cell membrane integrity, as evidenced by extracellular nucleic acid leakage. Cytotoxicity testing
confirmed safety for HEK293T cells at concentrations ≤ 50 μg/mL. These findings demonstrate that microreactor extracted L.
cladonioides essential oil represents a natural antimicrobial and antioxidant agent with low cytotoxicity, demonstrating promising
applications.
Keywords: Lethariella cladonioides (L. cladonioides), Essential oil, Composition identification, Antibacterial activity, Continuous
flow microreactor
Lethariella , a folk herbal tea originating from southwest China, has been consumed for centuries as a traditional beverage. However, the distinct chemical characteristics and sensory properties of Lethariella remain poorly understood. In this study, the two most commonly found Lethariella species, Lethariella zahlbruckneri (LZ) and Lethariella cladonioides (LC), were collected for detailed analysis. After brewing, the infusion of LZ exhibited a taste profile that was more astringent, bitter, and thicker, whereas LC liquor was characterized by a sweeter taste profile, coupled with a stronger lingering sweet aftertaste. This variation in taste between LZ and LC infusions can potentially be ascribed to the higher levels of phenolic compounds present in LZ in comparison to LC, along with the lower content of sugars in LZ. The aroma intensity of LC is more robust, with its relative contents of key volatile compounds being significantly higher compared to those in LZ.
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