UPLC-MS Analysis, Quantification of Compounds, and Comparison of Bioactivity of Methanol Extract and Its Fractions from Qiai (Artemisia argyi Lévl. et Van.).

The Artemisia argyi Lévl. et Van. growing in the surrounding areas of Qichun County in China are called Qiai (QA). Qiai is a crop that can be used both as food and in traditional folk medicine. However, detailed qualitative and quantitative analyses of its compounds remain scarce. The process of identifying chemical structures in complex natural products can be streamlined by combining UPLC-Q-TOF/MS data with the UNIFI information management platform and its embedded Traditional Medicine Library. For the first time, 68 compounds in QA were reported by the method in this study. The method of simultaneous quantification of 14 active components in QA using UPLC-TQ-MS/MS was reported for the first time. Following a screening of the activity of QA 70% methanol total extract and its three fractions (petroleum ether, ethyl acetate, and water), it was discovered that the ethyl acetate fraction enriched with flavonoids such as eupatilin and jaceosidin had the strongest anti-inflammatory activity, while the water fraction enriched with chlorogenic acid derivatives such as 3,5-di-O-caffeoylquinic acid had the strongest antioxidant and antibacterial activity. The results provided the theoretical basis for the use of QA in the food and pharmaceutical industries.

Full-Length Transcriptomic Sequencing and Temporal Transcriptome Expression Profiling Analyses Offer Insights into Terpenoid Biosynthesis in Artemisia argyi.

Artemisiae argyi Folium is a traditional herbal medicine used for moxibustion heat therapy in China. The volatile oils in A.argyi leaves are closely related to its medicinal value. Records suggest that the levels of these terpenoids components within the leaves vary as a function of harvest time, with June being the optimal time for A. argyi harvesting, owing to the high levels of active ingredients during this month. However, the molecular mechanisms governing terpenoid biosynthesis and the time-dependent changes in this activity remain unclear. In this study, GC-MS analysis revealed that volatile oil levels varied across four different harvest months (April, May, June, and July) in A. argyi leaves, and the primarily terpenoids components (including both monoterpenes and sesquiterpenes) reached peak levels in early June. Through single-molecule real-time (SMRT) sequencing, corrected by Illumina RNA-sequencing (RNA-Seq), 44 full-length transcripts potentially involved in terpenoid biosynthesis were identified in this study. Differentially expressed genes (DEGs) exhibiting time-dependent expression patterns were divided into 12 coexpression clusters. Integrated chemical and transcriptomic analyses revealed distinct time-specific transcriptomic patterns associated with terpenoid biosynthesis. Subsequent hierarchical clustering and correlation analyses ultimately identified six transcripts that were closely linked to the production of these two types of terpenoid within A. argyi leaves, revealing that the structural diversity of terpenoid is related to the generation of the diverse terpene skeletons by prenyltransferase (TPS) family of enzymes. These findings can guide further studies of the molecular mechanisms underlying the quality of A. argyi leaves, aiding in the selection of optimal timing for harvests of A. argyi.

A biflavonoid-rich extract from Selaginella moellendorffii Hieron. induces apoptosis via STAT3 and Akt/NF-κB signalling pathways in laryngeal carcinoma.

Selaginella moellendorffii Hieron. (SM), a perennial evergreen plant, has been used in the treatment of acute infectious hepatitis, thoracic and hypochondriac lumbar contusions, systemic oedema and thrombocytopaenia. However, the role of a biflavonoid-rich extract from SM (SM-BFRE) in anti-larynx cancer has rarely been reported. In this study, the in vitro and in vivo anti-laryngeal cancer activity and potential mechanisms of SM-BFRE were investigated. An off-line semipreparative liquid chromatography-nuclear magnetic resonance protocol was carried out to determine six biflavonoids from SM-BFRE. In vitro, MTT assay revealed that SM-BFRE inhibited the proliferation of laryngeal carcinoma cells. A wound healing assay indicated that SM-BFRE suppressed the migration of laryngeal cancer cells. Hoechst 33 258 and Annexin V-FITC/PI double staining assays were performed and verified that SM-BFRE induced apoptosis in laryngeal carcinoma cells. The Hep-2 bearing nude mouse model confirmed that SM-BFRE also exhibited anticancer effect in vivo. In addition, Western blot analysis demonstrated that SM-BFRE exerted its anti-laryngeal cancer effect by activating the mitochondrial apoptotic pathway and inhibiting STAT3 and Akt/NF-κB signalling pathways. All results suggested that SM-BFRE could be considered as a potential chemotherapeutic drug for laryngeal cancer.

Liquid chromatography with mass spectrometry and NMR spectroscopy based discovery of cytotoxic principles from Daphne tangutica Maxim.

An ethyl acetate extract from the barks of the ethnic Chinese medicine Daphne tangutica Maxim. exhibited antihepatocellular carcinoma activity against HepG2 and Hep3B cell lines. By using high-performance liquid chromatography based activity profiling in combination with offline liquid chromatography with mass spectrometry and NMR analysis, we rapidly identified ten major components of the extract, including seven active principles, coumarins (1-4) and biscoumarins (7, 8, 10), along with three inactive flavonoids (5, 6, 9). This study demonstrated that our combined protocol can be used as an important strategy for chemical profiling, dereplication as well as the identification of bioactive compounds from herbal medicines.

Development and Characterization of Polymorphic Microsatellite Markers for Sedum sarmentosum (Crassulaceae) and Their Cross-Species Transferability.

Sedum sarmentosum is an important Chinese medicinal herb that exhibits anti-inflammatory, anti-angiogenic and anti-nociceptive properties. However, little is known about its genetic background. The first set of 14 microsatellite markers were isolated and characterized for S. sarmentosum using an SSR-enriched library. Fourteen polymorphic microsatellite markers were acquired with satisfactory amplifications and a polymorphic pattern in 48 S. sarmentosum individuals. The number of alleles ranged from 3 to 15. The observed and expected heterozygosities varied from 0.0833 to 0.8750 and 0.2168 to 0.9063, respectively. Two loci showed significant departure from the Hardy-Weinberg equilibrium. Cross-species amplification was carried out in other Sedum species. High rates of cross-species amplification were observed. The transferability value ranged from 85.7% in S. lineare to 64.3% in S. ellacombianum. These markers will be valuable for studying the genetic variation, population structure and germplasm characterization of S. sarmentosum and related Sedum species.

[Study on Identification of Three Medicinal Plant Leaves from Elaeagnus Genus by Infrared Spectroscopy].

OBJECTIVE: To study and identify the three species of dry medicinal plant leaves trom Elaeagnus genus (E. pungens, E. lanceolata and E. henryi) by Infrared Spectroscopy(IR). METHODS: Fourier transform infrared spectroscopy and second derivative infrared spectroscopy were used to study and compare the characteristics of leaves of three Elaeagnus medicinal plants. RESULTS: The IR spectra and second derivative infrared spectra of the three Elaeagnus plants leaves were similar on the whole, the intensity or ratio of intensity of some absorption peaks still had certain distinctions, and the differences of the second derivative infrared spectra were more obvious. There were only slight differences between large and small leaf type of samples of Elaeagnus lanceolata; the differences of the plant leaves of one species collected in different harvest periods were far smaller than those of others belonging to the same genus. CONCLUSION: IR can be relatively reliably used for identification of the three Elaeagnus leaves.

[HPLC characteristic fingerprints of sedi linearis herba and sedi herba].

OBJECTIVE: To study HPLC characteristic fingerprint of Sedum lineare from different harvest periods, and to compare with its related species Sedum sarmentosum. METHODS: The HPLC fingerprints of Sedum lineare from different collecting periods were established and compared with Sedum sarmentosum by the same detection method. RESULTS: Hyperin, isoquercitrin and astragaloside were identified from the HPLC fingerprint of Sedum lineare. The fingerprint of Sedum lineare growing in the same area but different environment were basically identical; while there were remarkable differences of Sedum lineare growing in the same place but from different harvest periods, with the area of most common peaks changing from little to great, and slightly different peak number. The HPLC fingerprint of the two Sedum species had four common peaks, but could be distinguished from each other. The optimal harvest period of these two species should be full-bloom stage. CONCLUSION: The established method can provide reference for identification and quality analysis of Sedum lineare.

A Biflavonoid-Rich Extract from Selaginella doederleinii Hieron. against Throat Carcinoma via Akt/Bad and IKKß/NF-κB/COX-2 Pathways.

Selaginella doederleinii Hieron. is a common pharmacological plant, and this folk herbal medicine and its complex preparations have been widely used for the treatment of throat carcinoma (TC) and several associated complications in traditional Chinese medicine. This study was aimed at investigating the specific anti-throat carcinoma impacts and potential mechanisms of a biflavonoid-rich extract from S. doederleinii (SD-BFRE). The phytochemical profiling of SD-BFRE was performed by HPLC-ESI-QTOF-MS and UPLC-PDA, and the detailed pharmacological effects and mechanisms were respectively evaluated in vitro and in vivo. MTT assay, the Transwell assay and flow cytometry were performed to evaluate the abilities of SD-BFRE on inhibiting cell infiltrative growth in TC cells (Hep-2 and FaDu) in in vitro experiments. In vivo experiments used Hep-2 tumor-bearing nude mice to evaluate the anti-TC effect of SD-BFRE. Western blotting was used to explore the potential apoptotic pathway of TC cells. Here, we found that SD-BFRE exhibited anti-proliferation and pro-apoptotic effects in TC cells. Mechanistic studies have identified that SD-BFRE can suppress the activity of IKKß and IκB-α kinase and then down-regulate the effector proteins of NF-κB/COX-2 signaling. Moreover, SD-BFRE induced apoptosis partly by regulating the Akt/Bad/caspase signaling pathway. Taken together, this study firstly demonstrated that SD-BFRE exerted its anti-TC effects by way of IKKß/NF-κB/COX-2 and Akt/Bad pathways and might represent a potential chemotherapeutic agent for throat carcinoma.

[Comparative identification research on different extracts of Dai Medicine "Pokou" and its processed product by FTIR].

OBJECTIVE: To study identification methods of different extracts of Dai Medicine "Pokou" (the rhizome of Homalomena gigantea Engl. ) and its processed product made by immersing it in water, and provide reference for identification of the drug in further researches and applications. METHODS: FTIR technique was used for identifying the features of different extracted parts of this crude drug and its processed product. RESULTS: Compared with the crude drug, the petroleum ether-extracted parts of processed product turned out to have no obvious distinction in the FTIR. There was a large difference in the ethyl acetate-extracted parts, and the n-butanol-extracted parts also had certain discrepancy. A preliminary analysis was made on the chemical fundamentals which caused the changes in the FTIR before and after the drug's processing. CONCLUSION: The results provide an infrared spectral identification method for the drug and its applications.

[Comparative identification on Dai medicine "Pokou" and its processing product].

OBJECTIVE: To research identification methods of the Dai Medicine "Pokou" (the rhizome of Homalomena gigantea) and its processing product, and provide basis for identification of the drug in further research and application. METHODS: Macroscopic, microscopic observation and TLC and FTIR techniques were used to authenticate this raw medicine and its processing product. RESULTS: There were certain differences in the macroscopic features. The TLC result and infrared spectra of the samples had also obvious differences. The methods for identification of this raw medicine and its processing product were established, The detailed tissue and powder of this medicine were drawn. CONCLUSION: The results provided the basis for identification of the medicine and establishment of its quality standard.

[Studies on chemical constituents from roots of Polygonum amplexicaule].

OBJECTIVE: To investigate the chemical constituents in the roots of Polygonum amplexicaule var. sinense. METHOD: The compounds were isolated by column chromatography on silica gel and Sephadex LH-20. The structures were identified by means of IR, 1H-NMR, 13C-NMR and MS analyses. RESULT: Eight compounds were isolated and identified as friedelin (1), beta-sitosterol (2), simiarenone (3), angelicin (4), psoralen (5), palmitic acid (6), (-)-epicatechin (7), and quercetin (8), respectively. CONCLUSION: All compounds were isolated from this species for the first time and compounds 1, 3-6 were obtained from this genus for the first time.

[Study on microscopic observation and TLC identification of Dai medicine "Guomaguo" (Spondias pinnata)].

OBJECTIVE: To study identification methods of Dai medicine" Guomaguo", the fruit of Spondias pinnata. METHODS: Characteristic, microscopic observation and TLC idertification were used to authenticate this crude drug. RESULTS: The characters of the cross section, powder and TLC of the drug were reported, and the relevant drawings of the tissue, powder and TLC of this ethnomedicine were drawn. CONCLUSION: These results can supply evidences for the identification of the ethnomedicine in its exploitation and utilization.

[Studies on identification of Sedum emarginatum].

OBJECTIVE: To establish methods for identification of the whole plant of Sedum emarginatum Migo. METHODS: Macroscopic and microscopic observation and FTIR technique were used to authenticate this crude drug, and the identification characteristics were studied. RESULTS: The stem cross section and the whole plant powder had some notable micro-characters. The infrared spectras of the samples collected in the different habitats and seasons were very consistent with each other. CONCLUSION: The results can be used as the evidence for identification of this ethnomedicine.

[Study on pharmacognostic identifcation of Cassia siamea].

OBJECTIVE: To establish methods for identification of Dai medicine, the heartwoods and leaves of Cassia siamea. METHODS: Macroscopic, microscopic observation and TLC technique were used to anthenticate this crude drug, and the identification characteristics were studied. RESULTS: Macroscopic and microscopic identification methods and TLC characters of the ethnomedicine were re-ported, and the simple and detailed drawings of the transections of the heartwoods and leaves, as well as microscopic drawings of their powders were drawn in this paper. CONCLUSION: The results can serve as evidence for identification of the ethnomedicine in the utilization.

[RAPD analysis of 8 medicinal species of Selaginella].

OBJECTIVE: To identify 8 medicinal species of Selaginella with method of molecular marker and to explore their genetic relationship. METHODS: 8 primers were selected from 60 random primers to amplify the total DNA extracts of 17 samples with RAPD method and then the amplified were analyzed. RESULTS: A total of amplified 58 bands and RAPD map were obtained. Based on them, a Jaccard's genetic similarity matrix and a dendrogram for the 8 plants were established using SPSS 11.5 software. CONCLUSION: The method revealed distinct differences of different species and the variation of the same species of Selaginella from different habitats. It can provide with genetic proof for authentication of some species and units below species.

[Study on microscopic identification of Dai medicine "ge ga" (Zanthoylum armatum)].

The root, bark, leaf and fruit of Zanthoylum armatum are used as traditional crude drugs by Dai and some other minority nationalities in China, and the Dai drug name is "ge ga". This paper reported microscopic, physical and chemical characters of the bark and leaf The abbreviated and detailed diagrams of the transections of the bark and leaf, and the powder diagram of the bark were drawn in the study. The result showed that there were distinct microscopic characters of the transection and the powder, and these characters could be used as evidence for identification of the ethnomedicine in the exploitation and utilization.

[Study on HPLC fingerprint characteristics of Selaginella plants].

OBJECTIVE: To establish a method to analyze HPLC fingerprint characteristics of 10 plants from Selaginella. METHOD: HPLC was applied for establishment of fingerprints, which were used to evaluate and distinguish the different species of Selaginella. RESULT: The different species from Selaginella showed different HPLC fingerprint characteristic. The samples of the same species but collected in different period, different environment or different locations showed certain difference in fingerprints CONCLUSION: 2 important mutual fingerprint peaks were found in the 10 plants of Selaginella species and 5 peaks can be used as "main fingerprint peaks". The dates of these peaks can used for assessment of phylogenetic relation among species and evaluation of quality.

[Pharmacognostic identification on Dai medicine" guo sha" (Broussonetia papyrifera)].

The present paper reports descriptions and microscopical studies on Dai medicine" Guo sha" (Broussonetia papyrifera) , which is also the ethnomedicines of some minority nationalities in China. The diagram and the detailed drawing of the transection of the root, as well as microscopic drawing of its powder, were drawn in the studies. It was found that there are distinct microscopic characters of the transection and the powder which can serve as evidence for identification of the ethnomedicine.

Amplexicaule A exerts anti-tumor effects by inducing apoptosis in human breast cancer.

Chemotherapy is the main treatment for patients with breast cancer metastases, but natural alternatives have been receiving attention for their potential as novel anti-tumor reagents. Amplexicaule A (APA) is a flavonoid glucoside isolated from rhizomes of Polygonum amplexicaule D. Don var. sinense Forb (PADF). We found that APA has anti-tumor effects in a breast cancer xenograft mouse model and induces apoptosis in breast cancer cell lines. APA increased levels of cleaved caspase-3,-8,-9 and PARP, which resulted from suppression of MCL-1 and BCL-2 expression in the cells. APA also inactivated the Akt/mTOR pathway in breast cancer cells. Thus, APA exerts a strong anti-tumor effect on breast cancer cells, most likely through induction of apoptosis. Our study is the first to identify this novel anti-tumor compound and provides a new strategy for isolation and separation of single compounds from herbs.

[Investigations and studies on medicinal plants from Selaginellaceae in Hubei Province].

OBJECTIVE: To investigate the species, the distribution and the utilization of the medicinal plants from Selaginellaceae in Hubei Province. METHOD: Through field investigations and comparing the collected specimens and literatures, the classification and identification of the species in Hubei Province were studied. RESULT: The results indicated that 15 species of plants from Selaginellaceae in Hubei, including 14 medicinal and 1 newly recorded species existed. The distribution and use in folk medicine were investigated. And the morphological description of several species was appended. CONCLUSION: The results provided a basis for the exploitation and utilization of the medicinal plant resources of Selagingellaceae.