Enhancement of Palmarumycins C(12) and C(13) production in liquid culture of endophytic fungus Berkleasmium sp. Dzf12 after treatments with metal ions.
ABSTRACT: The influences of eight metal ions (i.e., Na+, Ca2+, Ag+, Co2+, Cu2+, Al3+, Zn2+, and Mn4+) on mycelia growth and palmarumycins C(12) and C(13) production in liquid culture of the endophytic fungus Berkleasmium sp. Dzf12 were investigated. Three metal ions, Ca2+, Cu2+ and Al3+ were exhibited as the most effective to enhance mycelia growth and palmarumycin production. When calcium ion (Ca2+) was applied to the medium at 10.0 mmol/L on day 3, copper ion (Cu2+) to the medium at 1.0 mmol/L on day 3, aluminum ion (Al3+) to the medium at 2.0 mmol/L on day 6, the maximal yields of palmarumycins C(12) plus C(13) were obtained as 137.57 mg/L, 146.28 mg/L and 156.77 mg/L, which were 3.94-fold, 4.19-fold and 4.49-fold in comparison with that (34.91 mg/L) of the control, respectively. Al3+ favored palmarumycin C(12) production when its concentration was higher than 4 mmol/L. Ca2+ had an improving effect on mycelia growth of Berkleasmium sp. Dzf12. The combination effects of Ca2+, Cu2+ and Al3+ on palmarumycin C(13) production were further studied by employing a statistical method based on the central composite design (CCD) and response surface methodology (RSM). By solving the quadratic regression equation between palmarumycin C(13) and three metal ions, the optimal concentrations of Ca2+, Cu2+ and Al3+ in medium for palmarumycin C(13) production were determined as 7.58, 1.36 and 2.05 mmol/L, respectively. Under the optimum conditions, the predicted maximum palmarumycin C(13) yield reached 208.49 mg/L. By optimizing the combination of Ca2+, Cu2+ and Al3+ in medium, palmarumycin C(13) yield was increased to 203.85 mg/L, which was 6.00-fold in comparison with that (33.98 mg/L) in the original basal medium. The results indicate that appropriate metal ions (i.e., Ca2+, Cu2+ and Al3+) could enhance palmarumycin production. Application of the metal ions should be an effective strategy for palmarumycin production in liquid culture of the endophytic fungus Berkleasmium sp. Dzf12.
Project description:The endophytic fungus Berkleasmium sp. Dzf12, isolated from Dioscorea zingiberensis, was found to produce palmarumycins C12 and C13 which possess a great variety of biological activities. Seven biocompatible water-immiscible organic solvents including n-dodecane, n-hexadecane, 1-hexadecene, liquid paraffin, dibutyl phthalate, butyl oleate and oleic acid were evaluated to improve palmarumycins C12 and C13 production in suspension culture of Berkleasmium sp. Dzf12. Among the chosen solvents both butyl oleate and liquid paraffin were the most effective to improve palmarumycins C12 and C13 production. The addition of dibutyl phthalate, butyl oleate and oleic acid to the cultures of Berkleasmium sp. Dzf12 significantly enhanced palmarumycin C12 production by adsorbing palmarumycin C12 into the organic phase. When butyl oleate was fed at 5% (v/v) in medium at the beginning of fermentation (day 0), the highest palmarumycin C12 yield (191.6 mg/L) was achieved, about a 34.87-fold increase in comparison with the control (5.3 mg/L). n-Dodecane, 1-hexadecene and liquid paraffin had a great influence on the production of palmarumycin C13. When liquid paraffin was added at 10% (v/v) in medium on day 3 of fermentation, the palmarumycin C13 yield reached a maximum value (134.1 mg/L), which was 4.35-fold that of the control (30.8 mg/L). Application of the aqueous-organic solvent system should be a simple and efficient process strategy for enhancing palmarumycin C12 and C13 production in liquid cultures of the endophytic fungus Berkleasmium sp. Dzf12.
Project description:Three crude oligosaccharides were respectively prepared by acid hydrolysis of three polysaccharides, which were water-extracted polysaccharide (WEP), sodium hydroxide-extracted polysaccharide (SEP) and acid-extracted polysaccharide (AEP) from the rhizomes of Dioscorea zingiberensis. Among the three oligosaccharides, the crude oligosaccharide prepared by acid hydrolysis of WEP was found to be the most efficient elicitor to enhance the production of palmarumycins C(12) and C(13) in liquid culture of endophytic fungus Berkleasmium sp. Dzf12. When OW was applied to the medium at 300 mg/L on day 3 of culture, the maximal yields of palmarumycin C(12) (87.96 mg/L) and palmarumycin C(13) (422.28 mg/L) were achieved on day 15 of culture, which were 9.83 and 3.24-fold in comparison with those (8.95 and 130.43 mg/L) of control, respectively. The results indicate that addition of the oligosaccharides from the host plant D. zingiberensis should be an effective strategy for enhancing production of palmarumycins C(12) and C(13) in liquid culture of endophytic fungus Berkleasmium sp. Dzf12.
Project description:Two spirobisnaphthalenes, namely palmarumycins C2 and C3, were isolated from cultures of the endophytic fungus Berkleasmium sp. Dzf12 after treatment with 1-hexadecene. After addition of 1-hexadecene at 10% to the medium on day 6 of culture, the maximal yields of palmarumycins C2 and C3 were obtained as 0.40 g/L and 1.19 g/L, which were 40.00 fold and 59.50 fold higher, respectively, in comparison with those of the control (0.01 g/L and 0.02 g/L). The results indicated that addition of 1-hexadecene can be an effective strategy for enhancing the production of palmarumycins C2 and C3 in liquid culture of endophytic fungus Berkleasmium sp. Dzf12. Palmarumycin C3 exhibited stronger antimicrobial and antioxidant activities than palmarumycin C2.
Project description:High-speed counter-current chromatography (HSCCC) was applied for the first time for the preparative separation of spirobisnaphthalenes from a crude extract of the endophytic fungus Berkleasmium sp. Dzf12, associated with the medicinal plant Dioscorea zingiberensis. Six spirobisnaphthalenes were successfully separated by HSCCC with a two-phase solvent system composed of n-hexane-chloroform-methanol-water (1.5:3.0:2.5:2.0, v/v). About 18.0 mg of diepoxin ? (1), 245.7 mg of palmarumycin C13 (2), 42.4 mg of palmarumycin C16 (3), 42.2 mg of palmarumycin C15 (4), 32.6 mg of diepoxin ? (5), and 22.3 mg of diepoxin ? (6) with purities of 56.82, 71.39, 76.57, 75.86, 91.01 and 82.48%, respectively, as determined by high-performance liquid chromatography (HPLC), were obtained from 500 mg of the crude extract in a one-step elution within 7 h of separation procedure by HSCCC. The purified spirobisnaphthalenes were further structurally characterized by means of physicochemical and spectrometric analysis.
Project description:Berkleasmium sp. Dzf12, an endophytic fungus from Dioscorea zingiberensis, is a high producer of spirobisnaphthalenes with various bioactivities. The exopolysaccharide (EPS) produced by this fungus also shows excellent antioxidant activity. In this study, the experimental designs based on statistics were employed to evaluate and optimize the medium for EPS production in liquid culture of Berkleasmium sp. Dzf12. For increasing EPS yield, the concentrations of glucose, peptone, KH(2)PO(4), MgSO(4)·7H(2)O and FeSO(4)·7H(2)O in medium were optimized using response surface methodology (RSM). Both the fractional factorial design (FFD) and central composite design (CCD) were applied to optimize the main factors which significantly affected EPS production. The concentrations of glucose, peptone and MgSO(4)·7H(2)O were found to be the main effective factors for EPS production by FFD experimental analysis. Based on the further CCD optimization and RSM analysis, a quadratic polynomial regression equation was derived from the EPS yield and three variables. Statistical analysis showed the polynomial regression model was in good agreement with the experimental results with the determination coefficient (adj-R(2)) as 0.9434. By solving the quadratic regression equation, the optimal concentrations of glucose, peptone and MgSO(4)·7H(2)O for EPS production were determined as 63.80, 20.76 and 2.74 g/L, respectively. Under the optimum conditions, the predicted EPS yield reached the maximum (13.22 g/L). Verification experiment confirmed the validity with the actual EPS yield as 13.97 g/L, which was 6.29-fold in comparison with that (2.22 g/L) in the original basal medium. The results provide the support data for EPS production in large scale and also speed up the application of Berkleasmium sp. Dzf12.
Project description:This study was to examine the effects of yeast extract (YE) and its fractions (YE1 and YE2) on the growth and diepoxin ζ (a spirobisnaphthalene with a diversity of bioactivities) production in liquid culture of Berkleasmium-like endophytic fungus Dzf12 from Dioscorea zingiberensis. When YE was applied to the liquid medium at 10 g/L on day 3 of culture, the diepoxin ζ production was most effectively enhanced 3.2-fold (378.70 mg/L versus 120.09 mg/L in control) after another 10 days culture. Feeding with 15 g/L of YE on day 9, the mycelia biomass reached 16.44 g/L, about 2.3-fold in comparison with the control (7.15 g/L). The polysaccharide fraction (YE1) was mainly responsible for stimulating diepoxin ζ accumulation, and the non-polysaccharide fraction (YE2) was mainly responsible for promoting mycelia growth. The results showed that the diepoxin ζ production in liquid culture of endophyte Dzf12 could be effectively enhanced by YE and its fractions.
Project description:In an attempt to explore the biosynthetic potential of endosymbiotic fungi, the secondary metabolite profiles of the endophytic fungus, Anteaglonium sp. FL0768, cultured under a variety of conditions were investigated. In potato dextrose broth (PDB) medium, Anteaglonium sp. FL0768 produced the heptaketides, herbaridine A (1), herbarin (2), 1-hydroxydehydroherbarin (3), scorpinone (4), and the methylated hexaketide 9S,11R-(+)-ascosalitoxin (5). Incorporation of commonly used epigenetic modifiers, 5-azacytidine and suberoylanilide hydroxamic acid, into the PDB culture medium of this fungus had no effect on its secondary metabolite profile. However, the histone acetyl transferase inhibitor, anacardic acid, slightly affected the metabolite profile affording scorpinone (4) as the major metabolite together with 1-hydroxydehydroherbarin (3) and a different methylated hexaketide, ascochitine (6). Intriguingly, incorporaion of Cu2+ into the PDB medium enhanced production of metabolites and drastically affected the biosynthetic pathway resulting in the production of pentaketide dimers, palmarumycin CE4 (7), palmarumycin CP4 (8), and palmarumycin CP1 (9), in addition to ascochitine (6). The structure of the new metabolite 7 was established with the help of spectroscopic data and by MnO2 oxidation to the known pentaketide dimer, palmarumycin CP3 (10). Biosynthetic pathways to some metabolites in Anteaglonium sp. FL0768 are presented and possible effects of AA and Cu2+ on these pathways are discussed.
Project description:Aluminium is known to be toxic to cells from bone, brain and bone marrow but the molecular target(s) affected by Al3+ are not known. We show here that Al3+ disrupts the oscillatory free Ca2+ responses of hepatocytes exposed to the Ca2(+)-mobilizing agonist phenylephrine. Al3+ initially increases the frequency of the oscillations and later induces broad Ca2+ spikes lasting several minutes. These broad spikes persist after removal of both agonist and Al3+ from the medium. In the absence of agonist, Al3+ has no effect on free Ca2+. The data suggest that some component(s) of the receptor-phosphoinositide-Ca2+ signalling pathway might be the site at which Al3+ exerts toxic effects.
Project description:1. In electroporated L1210 cells, Ins(1,3,4,5)P4 causes Ca2+ release, owing to its conversion into Ins(1,4,5)P3, but this does not happen in cells permeabilized by digitonin treatment [Cullen, Irvine, Drøbak & Dawson (1989) Biochem. J. 259, 931-933]. 2. If the assay medium is subjected to electroporation by using a commercially available electroporation apparatus and then the cells are added and permeabilized with digitonin, the cells behave as if they had been electroporated. 3. Electroporation causes the release of high concentrations of Al3+ into the experimental medium, and addition of these concentrations of Al3+ into the experimental medium mimics the effect of electroporation on the conversion of Ins(1,3,4,5)P4 into Ins(1,4,5)P3. 4. It is concluded that the difference between electroporated and digitonin-permeabilized L1210 cells in this experimental system can be attributed to dissolution of Al3+ from the electroporation cuvette. Al3+ contamination may thus be a serious problem when using this apparatus.
Project description:The layered double hydroxides (LDHs) of Ca2+ and trivalent cations, Al3+ and Fe3+, are single-source precursors to generate supported CaO, which picks up CO2 from the gas phase in the temperature range 350-550 °C. The supports are ternary oxides, mayenite, and Ca2Fe2O5. The uptake capacity of the Fe3+-containing LDH at 1.9 mmol g-1 is two times the capacity of the Al3+-containing LDH. The product of CO2 uptake is calcite CaCO3. It is observed that the intercalated chloride ions reduce the thermal penalty by inducing the early decomposition of CaCO3. In the case of the chloride-intercalated LDHs of Ca2+ and Fe3+, the CaCO3 formed is completely decomposed at 900 °C. This is in contrast with the CaCO3 formed from bare CaO, which shows no sign of decomposition at 900 °C under similar conditions. This work shows that the hydrocalumite-like LDHs are candidate materials for CO2 mineralization.