One new cyclopeptide was isolated from the ethyl acetate fraction of the 75% EtOH extract of Selaginella tamariscina by various column chromatography methods(HP-20, polyamide and semi-preparative HPLC). Its structure was identified as selapeptin A(1) by extensive spectroscopic analysis(HR-ESI-MS, 1 D and 2 D NMR). Compound 1 was evaluated for cytotoxic activities by MTT assay. It showed potent cytotoxic activity against B16 F10 with the inhibition rate of 51.57%±4.34% at 40 μmol·L~(-1) while had no impacts on MDA-MB-231 and MDA-MB-468 at 100 μmol·L~(-1).
Chromatography, High Pressure Liquid ; Magnetic Resonance Spectroscopy ; Molecular Structure ; Peptides, Cyclic/pharmacology* ; Selaginellaceae/chemistry*

Lycophytes and seed plants constitute the typical vascular plants. Lycophytes have been thought to have no paleo-polyploidization although the event is known to be critical for the fast expansion of seed plants. Here, genomic analyses including the homologous gene dot plot analysis detected multiple paleo-polyploidization events, with one occurring approximately 13-15 million years ago (MYA) and another about 125-142 MYA, during the evolution of the genome of Selaginella moellendorffii, a model lycophyte. In addition, comparative analysis of reconstructed ancestral genomes of lycophytes and angiosperms suggested that lycophytes were affected by more paleo-polyploidization events than seed plants. Results from the present genomic analyses indicate that paleo-polyploidization has contributed to the successful establishment of both lineages-lycophytes and seed plants-of vascular plants.
Evolution, Molecular ; Genome, Plant ; Genomics ; Phylogeny ; Polyploidy ; Selaginellaceae/genetics*

We investigated the anti-inflammatory effect of Pyunkang-tang extract (PGT), a complex herbal extract based on traditional Chinese medicine that is used in Korea for controlling diverse pulmonary diseases, on cigarette smoke-induced pulmonary pathology in a rat model of chronic obstructive pulmonary disease (COPD). The constituents of PGT were Lonicerae japonica, Liriope platyphylla, Adenophora triphilla, Xantium strumarinum, Selaginella tamariscina and Rehmannia glutinosa. Rats were exposed by inhalation to a mixture of cigarette smoke extract (CSE) and sulfur dioxide for three weeks to induce COPD-like pulmonary inflammation. PGT was administered orally to rats and pathological changes to the pulmonary system were examined in each group of animals through measurement of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) levels in bronchoalveolar lavage fluid (BALF) at 21 days post-CSE treatment. The effect of PGT on the hypersecretion of pulmonary mucin in rats was assessed by quantification of the amount of mucus secreted and by examining histopathologic changes in tracheal epithelium. Confluent NCI-H292 cells were pretreated with PGT for 30 min and then stimulated with CSE plus PMA (phorbol 12-myristate 13-acetate), for 24 h. The MUC5AC mucin gene expression was measured by RT-PCR. Production of MUC5AC mucin protein was measured by ELISA. The results were as follows: (1) PGT inhibited CSE-induced pulmonary inflammation as shown by decreased TNF-α and IL-6 levels in BALF; (2) PGT inhibited the hypersecretion of pulmonary mucin and normalized the increased amount of mucosubstances in goblet cells of the CSE-induced COPD rat model; (3) PGT inhibited CSE-induced MUC5AC mucin production and gene expression in vitro in NCI-H292 cells, a human airway epithelial cell line. These results suggest that PGT might regulate the inflammatory aspects of COPD in a rat model.
Animals ; Bronchoalveolar Lavage Fluid ; Campanulaceae ; Enzyme-Linked Immunosorbent Assay ; Epithelial Cells ; Epithelium ; Gene Expression ; Goblet Cells ; Humans ; In Vitro Techniques ; Inflammation ; Inhalation ; Interleukin-6 ; Korea ; Lonicera ; Lung Diseases ; Medicine, Chinese Traditional ; Models, Animal ; Mucins ; Mucus ; Necrosis ; Pathology ; Pneumonia ; Pulmonary Disease, Chronic Obstructive ; Rats ; Rehmannia ; Selaginellaceae ; Smoke ; Sulfur Dioxide ; Tobacco Products

Selaginella tamariscina is a typical resuscitation medicinal plant with extreme drought tolerance. Trehalose plays an important role in the resurrection process, and the trehalose-6-phosphate synthase(TPS) is the key enzyme to synthesize trehalose in plants. In this study, the sequence of TPS was obtained by splicing from the transcriptome data of S. tamariscina. After the synthesis of cDNA based on the template of total RNA, the sequence was cloned by RT-PCR for verification and then analyzed by bioinformatics methods. The results indicated that the full-length coding sequence of StTPS was 2 799 bp (GenBank accession no. MH155231), and the encoded protein contained 932 amino acids. StTPS could be located in the chloroplastid according to subcellular localization prediction. There were two conserved domains belonging to glycogen phosphorylase glycosyltransferase (GPGTF) family but no signal peptide or transmembrane domain in StTPS. The expression of StTPS was determined by qRT-PCR and the variation of trehalose content was measured by HPLC-ELSD during the resurrection process of S. tamariscina. Meanwhile, the correlation between them was analyzed. The results showed that both the expression level of StTPS and the trehalose content increased associated with the extension of dehydration time, and declined associated with the extension of rehydration time which proved a significant positive correlation between the StTPS expression level and the trehalose content. The results suggested that the StTPS probably plays a central role in recovery process in S. tamariscina.
Amino Acid Sequence ; DNA, Complementary ; Glucosyltransferases ; Selaginellaceae ; Trehalose

Glutathione S-transferase (GST) is important in plants to resist various stresses. In this study, two Phi GST genes (SmGSTF1 and SmGSTF2) were cloned from Selaginella moellendorffii. SmGSTF1 and SmGSTF2 genes encode proteins of 215 amino acid residues. Gene expression analysis showed that the two genes were expressed in roots, stems and leaves. The recombinant SmGSTF1 and SmGSTF2 proteins were overexpressed in Escherichia coli, and purified by Ni-affinity chromatography. SmGSTF1 and SmGSTF2 had the catalytic activity towards 1-Chloro-2,4-Dieitrobenzene, 4-Chloro-7-nitro-1,2,3-benzoxadiazole (NBD-Cl), and 4-Nitrobenzyl chloride substrates. SmGSTF1 also had the activity towards Fluorodifen and Cumyl hydroperoxide (Cum-OOH), whereas SmGSTF2 not. The enzyme kinetics analysis showed that SmGSTF1 and SmGSTF2 had high affinity towards glutathione, and low affinity towards 1-Chloro-2, 4-Dieitrobenzene. The enzymatic activity of SmGSTF1 and SmGSTF2 had high catalytic activity between pH 7 and 8.5, and between 45 and 55 °C. SmGSTF1 and SmGSTF2 may have an important role in the resistance of Selaginella moellendorfii against stress.
Amino Acid Sequence ; Cloning, Molecular ; Escherichia coli ; Glutathione Transferase ; genetics ; metabolism ; Plant Proteins ; genetics ; metabolism ; Selaginellaceae ; enzymology

Abstract: A new selaginellin derivative named as selaginellin S (1) was isolated from the whole plants of Selaginella pulvinata (Hook. et Grev.) Maxim. (Selaginellaceae), together with a known one (selaginellin M, 2). Compounds 1 and 2 were separated and purified by silica gel and Sephadex LH-20 column chromatography. Their structures were determined on the basis of extensive spectroscopic analysis including IR, MS, 1D and 2D NMR experiments, as well as ECD calculations. Compound 1 is a key intermidiant in the biosynthesis pathway of selaginellins. Compound 2 is first reported in this plant.
Biphenyl Compounds ; chemistry ; isolation & purification ; Cyclohexanones ; chemistry ; isolation & purification ; Molecular Structure ; Selaginellaceae ; chemistry

In the current study, nine flavonoids were isolated and purified from 75% ethanol extract of Selaginella uncinata (Desv.) Spring by column chromatographic techniques over macroporous resin, polyamide, silica gel, Sephadex LH-20 and pre-HPLC. On the basis of their physico-chemical properties and spectroscopic data analyses, these compounds were elucidated as cirsimarin (1), nepitrin (2), apigenin-6-C-α-L-arabinopyranosyl-8-C-β-D-glucopyranoside (3), apigenin-6-C-β-D-glucopyranosyl-8-C-α-L-arabinopyranoside (4), apigenin-7-O-β-D-glucopyranoside (5), 2,3-dihydroamentoflavone (6), 4'-O-methylamentoflavone (7), 2,3-dihydro-4'-O-methyl-amentoflavone (8), and 2,3,2",3"-tetrahydron-4'-O-methyl-robustaflavone (9). Compounds 1-5 belong to flavonoid glycosides and were isolated from the genus Selaginella for the first time.
Flavonoids ; analysis ; Selaginellaceae ; chemistry

AIM: To investigate the chemical constituents of Selaginella sinensis (Desv.) Spring. METHODS: Chromatographic separations on Diaion HP-20, silica gel, and Sephadex LH-20 were used. The structures of the isolates were elucidated on the basis of spectroscopic analysis, as well as chemical methods. RESULTS: Eight compounds were obtained and their structures were identified as sinensioside A (1), syringaresinol-4- O-β-D-glucopyranoside (2), (+)-medioresinol-4-O-β-D-glucopyranoside (3), pinoresinol-4, 4'-di-O-β-D-glucopyranoside (4), quercetin (5), eucomic acid (6), shikimic acid (7), and 2, 3-dihydroamentoflavone (8). CONCLUSION Compound 1 is a new dihydrobenzofuran sesquilignan glycoside from Selaginella sinensis.
Benzofurans ; chemistry ; isolation & purification ; Furans ; chemistry ; isolation & purification ; Glucosides ; chemistry ; isolation & purification ; Lignans ; chemistry ; isolation & purification ; Molecular Structure ; Plant Extracts ; chemistry ; Plant Stems ; chemistry ; Quercetin ; chemistry ; isolation & purification ; Selaginellaceae ; chemistry

This study is to observe the protection effect of amentoflavone (AMT) in Selaginella tamariscina against TNF-alpha-induced vascular inflammation injury of endothelial cells. On the basis of TNF-alpha induced human umbilical vein endothelial cell, observe the influence of AMT on endothelial active factor, the contents of SOD and MDA, the protein expression of vascular endothelial adhesion molecules and inflammatory factor; study the effect of its common related signal pathways such as NF-kappaB; research the effect of AMT against TNF-a induced human umbilical vein endothelial cell injury by means of MTT, ELISA, Western blotting and the cell immunofluorescence. The results showed that AMT could increase the content of NO and decrease the levels of VCAM-1, E-selectin, IL-6, IL-8 and ET-1; enhance the activity of SOD, reduce the content of MDA; downregulate the protein expressions of VCAM-1, E-selectin, NF-kappaBp65 and up-regulate IkappaBalpha, attenuate the NF-kappaBp65 transfer to cell nucleus. AMT has the effect of protect vascular endothelial and maybe via the signal pathway of NF-kappaB to down-regulate the inflammation factor and oxidative damage factor of downstream.
Biflavonoids ; isolation & purification ; pharmacology ; Cell Cycle ; drug effects ; Cell Survival ; drug effects ; E-Selectin ; metabolism ; Endothelin-1 ; metabolism ; Human Umbilical Vein Endothelial Cells ; Humans ; I-kappa B Proteins ; metabolism ; Interleukin-6 ; metabolism ; Interleukin-8 ; metabolism ; Malondialdehyde ; metabolism ; NF-KappaB Inhibitor alpha ; Nitric Oxide ; metabolism ; Plants, Medicinal ; chemistry ; Protective Agents ; isolation & purification ; pharmacology ; Selaginellaceae ; chemistry ; Superoxide Dismutase ; metabolism ; Transcription Factor RelA ; metabolism ; Tumor Necrosis Factor-alpha ; adverse effects ; Vascular Cell Adhesion Molecule-1 ; metabolism

OBJECTIVETo establish a HPLC-DAD model for the simultaneous determination of two selaginellins (selaginellin and selaginellin B) and four biflavones (amentoflavone, sequoiaflavone, hinokiflavone and isocryptomerin) contained in 10 batches of Selaginella tamariscina and 12 batches of S. pulvinata produced in different areas.

METHODThe analysis was performed on a Waters Cosmosil C18 column (4.6 mm x 250 mm, 5 microm) eluted with acetonitril-0.1% formic acid as mobile phase in a linear gradient mode. The detection wavelength was set at 280, 337 nm. The flow rate was kept at 1.0 mL x min(-1), and the column temperature was 30 degrees C.

RESULTThe six active constituents showed significant different in content. Amentoflavone in S. tamariscina contains (5. 628-9. 184 mg x g(-1)) is more than that contained in S. pulvinata (0.823-7.131 mg x g(-1)), while selaginellin in S. pulvinata (0.123-0.593 mg x g(-1)) is more than that contained in S. tamariscina (0.067-0.133 mg x g(-1)). All the calibration curves showed good linear correlation coefficients (r > 0.9997) over the wide test ranges.

CONCLUSIONThe developed HPLC-DAD method is simple, sensitive and accurate and has the good repeatability in separation, which is available for the quality control of S. tamariscina and S. pulvinata.

Biflavonoids ; chemistry ; Biphenyl Compounds ; chemistry ; Chromatography, High Pressure Liquid ; methods ; Cyclohexanones ; chemistry ; Flavones ; chemistry ; Flavonoids ; chemistry ; Selaginellaceae ; chemistry