Plant Huperzia Serrata Research Articles

Secondary metabolites from Penicillium sp. HS-11, a fungal endophyte of Huperzia serrata

Three new sorbicillinoids sorbicatechols E-G (1–3), along with seven known compounds 4–10, were obtained from the ethanol extract of Penicillium sp. HS-11, a fungal endophyte of the medicinal plant Huperzia serrata. The structures of 1–3 were established by detailed interpretation of the spectroscopic data and their absolute configurations were established by comparative analyses of the ECD spectra. Sorbicatechol G (3) represented the first hybrid sorbicillinoid bearing a tetralone skeleton. In the in-vitro bioassay, trichodimerol (5) exhibited moderate inhibitory activity against the Escherichia coli β-glucuronidase (EcGUS) with an IC50 value of 92.0 ± 9.4 μM.

Ginkgolides and Huperzine A for complementary treatment of Alzheimer's disease.

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by gradual deterioration of cognitive function, memory, and inability to perform daily, social, or occupational activities. Its etiology is associated with the accumulation of β-amyloid peptides, phosphorylated tau protein, and neuroinflammatory and oxidative processes in the brain. Currently, there is no successful pharmacological treatment for AD. The few approved drugs are mainly aimed at treating the symptoms; however, due to the increasing discovery of etiopathological factors, there are great efforts to find new multifunctional molecules to slow down the course of this neurodegenerative disease. The commercial Ginkgo biloba formulation EGb 761® and Huperzine A, an alkaloid present in the plant Huperzia serrata, have shown in clinical trials to possess cholinergic and neuroprotective activities, including improvement in cognition, activities of daily living, and neuropsychiatric symptoms in AD patients. The purpose of this review is to expose the positive results of intervention with EGb 761® and Huperzine in patients with mild to moderate AD in the last 10 years, highlighting the pharmacological functions that justify their use in AD therapy.

Identification of a Huperzine A-producing endophytic fungus from Phlegmariurus taxifolius

Highly prized huperzine A (Hup A), a natural alkaloid formerly isolated from the Chinese medicinal plant Huperzia serrata, has been widely used for the treatment of Alzheimer disease, inspiring us to search for endophytic fungi that produce this compound. In this study, we obtained the C17 fungus isolate from the Mexican club moss Phlegmariurus taxifolius, which produced a yield of 3.2μg/g Hup A in mycelial dry weight, when cultured in potato dextrose broth medium. The C17 isolate was identified as belonging to the genus Fusarium with reference to the colony´s morphological characteristics and the presence of macroconidia and microconidia structures; and this was confirmed by DNA-barcoding analysis, by amplifying and sequencing the ribosomal internal transcribed spacer (rITS).

A Nasal Temperature and pH Dual-Responsive In Situ Gel Delivery System Based on Microemulsion of Huperzine A: Formulation, Evaluation, and In Vivo Pharmacokinetic Study.

Huperzine A (hup A), extracted from the Chinese medicinal plant Huperzia serrata, is a reversible and highly selective second-generation acetylcholine esterase (AchE) inhibitor for treating Alzheimer's disease (AD), but it suffers from low bioavailability in the brain. This study aimed to develop a nasal temperature and pH dual-responsive in situ gel delivery system based on microemulsion of hup A (hup A-M-TPISG). The optimal formulation was obtained by central composite design and response surface methodology. The optimized mucoadhesive formulation, hup A-M-TPISG, was composed of pluronic F127 (20.80%), pluronic F68 (2.8%), and chitosan (0.88%) as the gel matrix, which could gelatinize under physiological conditions (29-34°C, pH6.5) because of its temperature and pH responsiveness. The optimized hup A-M-TPISG formulation was further evaluated by in vitro release and in vivo pharmacokinetic studies via microdialysis. The in vitro release study showed continuous and steady drug release from hup A-M-TPISG, which was in accordance with the first-order model. Moreover, the pharmacokinetic results revealed that the optimized formulation for nasal administration, with convenient administration and improved patient compliance, could achieve similar brain-targeting properties as intravenous administration. In conclusion, the hup A-M-TPISG for intranasal administration, as an effective and safe vehicle, could enhance the absorption of hup A in vivo and would be a promising noninvasive alternative for partially improving brain-targeting therapy.

Biotransformation of Huperzine A by Irpex lacteus-A fungal endophyte of Huperzia serrata

The biotransformation of huperzine A (hupA), one of the characteristic bioactive constituents of the medicinal plant Huperzia serrata, by a fungal endophyte of the host plant was studied. Two previously undescribed compounds 1–2, along with a known analog 8α,15α-epoxyhuperzine A (3), were isolated and identified. The structures of all the isolates were established by spectroscopic methods including NMR, MS, IR, and UV spectra. In particular, the absolute configurations of 1 and 2 were elucidated by CD spectra comparison and theoretic NOE strength calculation. In the LPS-induced neuro-inflammation injury assay, 1–3 exhibited moderate neuroprotective activity by increasing the viability of U251 cell lines with EC50 values of 35.3 ± 0.9, 32.1 ± 0.9, and 50.3 ± 0.8 nM, respectively.

Antibacterial diketopiperazines from an endophytic fungus Bionectria sp. Y1085.

Two new diketopiperazines (1, 2), one new polyprenol (3), together with 19 known compounds (4-22) were obtained from the EtOAc extract of Bionectria sp. Y1085, an endophytic fungus isolated from the plant Huperzia serrata. Their structures were elucidated by extensive NMR and MS analysis. Bionectin D (1) is a rare diketopiperazine with a single methylthio substitution at the α-carbon of cyclized amino acid residue. The antibacterial activity of compounds was assayed against Escherichia coli, Staphylococcus aureus, and Salmonella typhimurium ATCC 6539, and some metabolites (1, 2, 10, 11, and 14) exhibited evident antibacterial activity.

Metabolites from the endophytic fungus colletotrichum sp. F168

An endophytic fungus, Colletotrichum sp. F168, was isolated from plant Huperzia serrata (Thunb. ex Murray) Trev. and was subjected to phytochemical investigation. Nine compounds including two new structures were obtained from SDA solid fermentation products of strain F168, which were elucidated by extensive spectroscopic analyses, including 1 D- and 2 D-NMR, and HR-MS experiments. The acetylcholinesterase inhibitory activity of two new compounds 1-2 was tested in vitro. Two new compounds didn’t show evident acetylcholinesterase inhibitory activity.

Biotransformation of Huperzine B by a Fungal Endophyte of Huperzia serrata.

The biotransformation of huperzine B (hupB), one of the characteristic bioactive constituents of the medicinal plant Huperzia serrata, by a fungal endophyte of the host plant was studied. One new compound, 8α,15α-epoxyhuperzine B (1), along with two known oxygenated hupB analogs, 16-hydroxyhuperzine B (2) and carinatumin B (3), was isolated and identified. The structures of all the isolates were deduced by spectroscopic methods including NMR, MS, IR, and UV spectra. The known compounds 2 and 3 were obtained from a microbial source for the first time. To the best of our knowledge, it is the first report on the microbial transformation of hupB and would facilitate further structural modification of hupB by chemo-enzymatic method. In the LPS-induced neuro-inflammation injury assay, 8α,15α-epoxyhuperzine B (1) exhibited moderate neuroprotective activity by increasing the viability of U251 cell lines with an EC50 of 40.1 nm.

Allobranchiibius huperziae gen. nov., sp. nov., a member of Dermacoccaceae isolated from the root of a medicinal plant Huperzia serrata (Thunb.).

A Gram-stain-positive, non-spore-forming actinobacterial strain, designated CPCC 204077T, was isolated from the surface-sterilized root of a medicinal plant Huperzia serrata (Thunb.) collected from Sichuan Province, south-west China. The peptidoglycan type of strain CPCC 204077T was detected as A4α with an l-Lys-l-Ser-d-Asp interpeptide bridge. Galactose, glucose, rhamnose and ribose were the sugar compositions in the whole-cell hydrolysates. MK-8(H4) was the only menaquinone. The polar lipid profile consisted of diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, phosphatidylinositol, one unidentified phospholipid and one unidentified glycolipid. The major fatty acid was iso-C16 : 0. The genomic DNA G+C content was 71.0 mol%. The phylogenetic tree based on 16S rRNA gene sequences showed that strain CPCC 204077T stood for a distinct lineage within the family Dermacoccaceaealongside the genera Branchiibius, Demetria and Barrientosiimonas, with the highest 16S rRNA gene sequence similarities to Branchiibius hedensis Mer 29717T (95.0 %), Calidifontibacter indicus PC IW02T (95.0 %), Barrientosiimonas humi 39T (94.9 %) and Demetria terragena HKI 0089T (94.7 %), and less than 94.7 % sequence similarities to all other species. Signature nucleotides in the 16S rRNA sequence showed that the strain contained the Dermacoccaceaefamily-specific 16S rRNA signature nucleotides and a genus-specific diagnostic nucleotide signature pattern. Combining the genotypic and phenotypic analyses, we propose that strain CPCC 204077T represents a novel species of a new genus in the family Dermacoccaceae with the name Allobranchiibius huperziae gen. nov., sp. nov. Strain CPCC 204077T (=NBRC 110719T=DSM 29531T) is the type strain of the type species.

Jatrophihabitans huperziae sp. nov., an endophytic actinobacterium isolated from surface-sterilized tissue of the medicinal plant Huperzia serrata (Thunb.).

A novel endophytic actinobacterium, designated strain CPCC 204076T, was isolated from surface-sterilized tissue of the medicinal plant Huperzia serrata (Thunb.) collected from Sichuan Province, south-west China. The taxonomic position of the isolate was investigated by a polyphasic approach. The strainwas aerobic, Gram-stain-positive, non-motile, non-spore-forming and rod-shaped. Growth was observed at 10-37 °C, at pH 5.0-10.0 and with 0-3.0 % (w/v) NaCl. The polar lipid fraction consisted of diphosphatidylglycerol, a phospholipid, an aminolipid, a glycolipid, an aminophospholipid and phosphatidylinositol. The cell wall contained meso-diaminopimelic acid as the diagnostic diamino acid and the peptidoglycan was of type A4γ. The menaquinone system consisted of MK-9(H4) and MK-8(H4). The major cellular fatty acids (>10 %) were iso-C16 : 0 and anteiso-C17 : 0. The genomic DNA G+C content of strain CPCC 204076T was found to be 71.9 mol%. Phylogenetic analysis based on 16S rRNA gene sequences revealed that CPCC 204076T belongs to the genus Jatrophihabitans with highest sequence similarity to Jatrophihabitans endophyticus DSM 45627T (96.5 %), Jatrophihabitans soli DSM 45908T (96.5 %) and Jatrophihabitans fulvus JCM 30448T (96.1 %), and much lower similarities (<95.0 %) to other available 16S rRNA gene sequences from validly described pure cultures. However, DNA-DNA hybridyzation values between strain CPCC 204076T and the three recognized Jatrophihabitans species were 31±3.1 % (J. endophyticus DSM 45627T), 33±2.9 % (J. soli DSM 45908T) and 37±1.7 % (J. fulvus JCM 30448T), which were all far below the recommended cut-off value of 70 %. The phenotypic and genomic characteristics distinctly indicated that strain CPCC 204076T represents a novel species of the genus Jatrophihabitans, for which the name Jatrophihabitans huperziae sp. nov. is proposed. The type strain is CPCC 204076T (I13A-01604) (=DSM 46866T=NBRC 110718T).

De novo RNA sequencing and transcriptome analysis of Colletotrichum gloeosporioides ES026 reveal genes related to biosynthesis of huperzine A.

Huperzine A is important in the treatment of Alzheimer’s disease. There are major challenges for the mass production of huperzine A from plants due to the limited number of huperzine-A-producing plants, as well as the low content of huperzine A in these plants. Various endophytic fungi produce huperzine A. Colletotrichum gloeosporioides ES026 was previously isolated from a huperzine-A-producing plant Huperzia serrata, and this fungus also produces huperzine A. In this study, de novo RNA sequencing of C. gloeosporioides ES026 was carried out with an Illumina HiSeq2000. A total of 4,324,299,051 bp from 50,442,617 high-quality sequence reads of ES026 were obtained. These raw data were assembled into 24,998 unigenes, 40,536,684 residues and 19,790 genes. The majority of the unique sequences were assigned to corresponding putative functions based on BLAST searches of public databases. The molecular functions, biological processes and biochemical pathways of these unique sequences were determined using gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) assignments. A gene encoding copper amine oxidase (CAO) (unigene 9322) was annotated for the conversion of cadaverine to 5-aminopentanal in the biosynthesis of huperzine A. This gene was also detected in the root, stem and leaf of H. serrata. Furthermore, a close relationship was observed between expression of the CAO gene (unigene 9322) and quantity of crude huperzine A extracted from ES026. Therefore, CAO might be involved in the biosynthesis of huperzine A and it most likely plays a key role in regulating the content of huperzine A in ES026.

Endophytes in the plant Huperzia serrata: fungal diversity and discovery of a new pentapeptide.

Endophytic fungi are an underexploited resource of natural products and have a capacity to produce diverse classes of plant-derived secondary metabolites. Here, we investigated the diversity of endophytic fungi from Huperzia serrata and the potential for discovering novel fungal natural products. One hundred and fifty-five endophytic fungi isolates obtained from H. serrata, belonging to four classes Dothideomycetes (47.3%), Sordariomycetes (36.8%), Eurotiomycetes (10.6%) and an undefined class (5.3%, Mucoraceae), were grouped into nine genera based on morphological and molecular identification. Colletotrichum, Cladosporium, Sordariomycetes and Guignardia were the dominant genera, whereas the remaining genera were infrequent groups. To our knowledge, the fungal genera Mucor and Neurospora were first reported in Huperzia plant. Among these endophytic fungi isolates, strain B14, belonging to Penicillium oxalicum, gave a gray precipitate with Dragendorff's reagent. A new pentapeptide was isolated from the culture of strain B14, and its chemical structure was elucidated on the basis of spectroscopic data from (1)H NMR, (13)C NMR and ESI-MS/MS. Taken together, H. serrata has a significant diversity of endophytic fungi that could be a rich resource for the discovery of new natural products.

Ceriponols A–K, tremulane sesquitepenes from Ceriporia lacerate HS-ZJUT-C13A, a fungal endophyte of Huperzia serrata

Tremulane sesquiterpenes ceriponols A-K, together with three known ones tremulenediol A, 11,12-dihydroxy-1-tremulen-5-one and conocenol B, were isolated from the cultures of Ceriporia lacerate, a fungal endophyte residing in the stems of the medicinal plant Huperzia serrata. Among these isolates, ceriponol B possessed an unprecedent 12-nortremulane skeleton. The structures of all isolates were elucidated by spectroscopic methods, including MS and NMR (1D and 2D) spectroscopic techniques. The cytotoxic activities of ceriponols A-K were evaluated against three human tumor cell lines (HeLa, HepG2, and SGC 7901). Ceriponols F and K exhibited moderate cytotoxicity against all the tested human cancer cell lines with IC50 values ranging from 32.3 ± 0.4 to 173.2 ± 1.5 μM, while ceriponol G showed slightly better cytotoxicity against a HeLa cell line.

Multihydroxylation of ursolic acid by Pestalotiopsis microspora isolated from the medicinal plant Huperzia serrata

The structural modification of ursolic acid by an endophytic fungus Pestalotiopsis microspora, isolated from medicinal plant Huperzia serrata was reported for the first time. The structure diversity was very important for the SAR study of ursolic acid and its derivatives. Incubation of ursolic acid 1 with P. microspora afforded four metabolites: 3-oxo-15α, 30-dihydroxy-urs-12-en-28-oic acid (2), 3β, 15α-dihydroxy-urs-12-en-28-oic acid (3), 3β, 15α, 30- trihydroxy-urs-12-en-28-oic acid (4) and 3,4-seco-ursan-4,30-dihydroxy-12-en-3,28-dioic acid (5). All products were new compounds and their structures elucidation was mainly based on the spectroscopic data.

Biotransformation of Ursolic Acid by an Endophytic Fungus from Medicinal Plant Huperzia serrata

Endophytic fungi were used not only for their producing bioactive products but also for their ability to transform natural compounds. An endophytic fungus, isolated from medicinal plant Huperzia serrata, was identified as Umbelopsis isabellina based on the internal transcribed spacer of ribosomal DNA (rDNA-ITS) region. It was used to transform ursolic acid (1), a pentacyclic triterpene. Incubation of ursolic acid with U. isabellina afforded three products, 3β-hydroxy-urs-11-en-28,13-lactone (2), 3β,7β-dihydroxy-urs-11-en-28,13-lactone (3), 1β,3β-dihydroxy-urs-11-en-28,13-lactone (4). Although product 2 was a known compound, it was first obtained by microbial transformation. Products 3 and 4 were new compounds. The structural elucidation of the three compounds was achieved mainly by the 1D- and 2D-NMR, MS, IR data. The endophytic fungus U. isabellina can hydroxyate the C12-C13 double bond at position 13 of ursolic acid 1 and form a five-member lactone effectively. In the meantime, this fungus can also introduce the hydroxyl group at C-1 or C-7 of ursolic acid 1.

Molecular identification of endophytic fungi from medicinal plant Huperzia serrata based on rDNA ITS analysis

Fifty-two endophytic fungi strains with different colony morphologies were isolated from stems, leaves and roots of Huperzia serrata (Thunb. ex Murray) Trevis. collected from Bawangling Reserve of Hainan Province in southern China. They were identified mainly based on rDNA ITS sequences and phylogenetic analysis. The results showed that all strains belonged to four classes, i.e. Sordariomycetes (92.31%), Dothideomycetes (3.85%), Pezizomycetes (1.92%) and Agaricomycetes (1.92%). Forty-seven strains were identified at the genus level, including Glomerella (Colletotrichum), Hypocrea (Trichoderma), Pleurostoma, Chaetomium, Coniochaeta (Lecythophora), Daldinia, Xylaria, Hypoxylon,Nodulisporium, Cazia and Phellinus. As to the other five strains, three were identified at the order level and two at the family level, indicating that a great diversity of fungi taxa exists in H. serrata. Most isolated strains belonged to the genus of Glomerella (Colletotrichum) and Hypoxylon, twenty-one from Glomerella and its anamorph Colletotrichum (42.3% of total isolated strains) and ten from Hypoxylon (19.2% of total isolated strains). Pleurostoma, Chaetomium, Coniochaeta (Lecythophora), Daldinia, Xylaria, Hypoxylon,Nodulisporium, Cazia and Phellinus were reported as endophytic fungi isolated from H. serrata for the first time.

Capillary electrophoresis determination of huperzine A: A new prospective anti-dementia drug

Huperzine A is an alkaloid isolated from the Chinese plant Huperzia serrata. The absorption spectra and electrophoretic behaviour of the compound were studied. The compound shows two absorption maxima (&lambda;max 230 and 310nm). The protonation constant was determined by spectrophotometry and capillary zone electrophoresis (CZE) and the recommended value is pK=7.70&plusmn;0.07. A CZE method for the determination of Huperzine A in pharmaceutical tablets and serum has been developed. The method is based on separation of the compound as a protonated species with electrophoretic mobility determined as 50.7 x 10-9 m2V-1s-1. The optimal separation conditions were: acetate buffer (pH 4.60; 20 mM) as a background electrolyte, separation voltage 20 kV, temperature 25 °C, hydrodynamic injection 10 s and detection at wavelengths 230 and 310 nm. The analyses were run in an uncoated fused-silica capillary with total length of 47.5 cm (39.0 cm to the detector) and 75 &mu;m I.D. The detection limit was found to be 70 &mu;g/l. The method of Huperzine A determination was applied for analysis of various pharmaceutical products. The results were found to be in good agreement with the declared amount. The precision of the method is &plusmn;2.30%. A method for the determination of HupA in serum was also developed with detection limit &asymp; 110 &mu;g/l, which is applicable for pharmacokinetics and therapeutic drug monitoring of this compound.

END RUN AROUND FDA?

The over-the-counter sale of huperzine A, a natural product with an activity similar to that of two commercial Alzheimer's therapeutics, is riling some Alzheimer's researchers and clinicians. The plant extract, considered to be a potential treatment for Alzheimer's disease, is being made commercially available in the U.S. without having undergone any human clinical trials in the U.S. Huperzine A, an alkaloid extracted from the leaves of the Chinese club moss plant Huperzia serrata, is already being sold over the Internet by PanoLife Products, Mountain View, Calif. And another company, Nutrapharm, New York City, plans to market it as a dietary supplement, or nutraceutical, at stores that sell vitamin and nutrition products. It will thus be widely accessible to Alzheimer's disease patients The release of the compound as a nutraceutical is totally legal and aboveboard. Under current law, natural products generally can be sold as over-thecounter dietary supplements without a prescription, providing they are...