Anti-hepatitis activities in the broth of Ganoderma lucidum supplemented with a Chinese herbal medicine.

The anti-hepatitis B virus activity and hepatoprotective activity of a liquid fermentation broth of Ganoderma lucidum were investigated. The cultured broth was supplemented with aqueous extract of Radix Sophorae flavescentis, a kind of Chinese herbal medicine. Our results indicated that the cultured broth had effects of anti-hepatitis B virus activity in vitro and protected mice from liver damage in vivo. Our results also indicated that the co-fermentation broth of Ganoderma lucidum in the presence of aqueous extract of Radix Sophorae flavescentis has better medicinal effects than simply mixing these two ingredients together, suggesting a potential novel way to prepare Chinese herbal mixtures.

[In vitro inhibitory efects on HBsAg and HBeAg secretion of 3 new components produced by Ganoderma lucidum in the medium contained Radix sophorae flavescentis extract].

In order to enhance the medical effects of Ganoderma lucidum submergedly cultured broth, the aqueous extract of Radix sophorae flavescentis, a traditional Chinese medicine, was added into the cultivation medium of G. lucidum. The organic acids were extracted with ethanol, chloroform, 5% NaHCO3 and chloroform in turn from the cultured broth of Ganoderma lucidum which cultivation medium contained Radix sophorae flavescentis extract. Six new components were separated from the organic acids with preparative HPLC. Their inhibitory effects on HBsAg and HBeAg secretion of HBV DNA transferred HepG2 cell (2.2.15 cell) were investigated. The results indicate that 3 components of the six have significant inhibitory effects on the antigen secretion.

[The process of Chinese medicine by submerged fermentation of pharmaceutical fungi].

The progress of Chinese medicine modernization is slow in china, and one of the reason is the dated process method. The fermentation technology of pharmaceutical fungi is more and more perfect. To process Chinese medicine by pharmaceutical fungi could enhance the effective density of active ingredient in Chinese medicine, and the fungi could produce a variety of active secondary metabolite, moreover, some compounds in Chinese medicine could be bio-transformed to produce new compound by pharmaceutical fungi in the fermentation. So process Chinese medicine by pharmaceutical fungi could produce de novo Chinese medicine preparations which include a variety of bioactive compounds, and it is suitable to produce in a large scale and convenient to control the quality. To process Chinese medicine by submerged fermentation of pharmaceutical fungi provides a new process method for Chinese medicine.

The Influence of Different Physicochemical Parameters of the Solvent On The Activity and Selectivity of Lipase.

The asymmetric esterification of octanoic acid with racemic 2-octanol catalyzed by a lipase from Pseudomonas sp. was investigated in several typical solvents. It was found that the catalytic activity and the enantioselectivity of the enzyme were governed by different physicochemical parameters of the solvent employed. While the former depended on the hydrophobicity (lgP) of the solvent, by contrast the latter was a function of the dielectric constant and the dipole moment. A mechanistic model for the binding site of the enzyme was postulated to rationalize this phenomenon based on the results of the kinetic studies of the reaction in some representative solvents.

[Effects of extracts of Chinese medicines on Ganoderma lucidum in submerged culture].

Effects of water and ethanol extracts of 10 Chinese medicines, such as Astragalus membranaceus, Coix lachryma-jobi, etc., on biomass and exopolysaccharide of Ganoderma lucidum were studied by submerged culture. The results showed: water extracts of all medicines can improve the culture of G. lucidum except of A. membranaceus, ethanol extracts of C. lachryma-jobi, Dioscorea opposita, Codonopsis pilosula, and Achyranthes bidentata( < 187.5g Medicine/L substrate) can also increase the biomass of G. lucidum, but the ethanol extracts of Angelica sinensis, Dendrobium nobile check the growth of G. lucidum. The production of exopolysa-ccharide can be improved by all the Chinese medicines and their dosage used in this experiment, Although A. sinensis, D. Nobile check the growth of G. lucidum, they could stimulate the secretion of exopolysaccharide in lower dosage. It is concluded that some Chinese medicines, such as C. lachryma-jobi, D. opposita, C. pilosula, etc. can be processed by the fermentation of G. lucidum, and bio-active compound can be produced by adding appropriate Chinese medicine in the substrate to culture G. lucidum.

[Relationship between mycelium morphology and laccase production of Pleurotus ferulae in submerged cultivation].

In this study, the relationship between mycelium morphology and laccase production was studied. The results indicated that the morphology of P. ferulae pellets was changed when glass beads were added. Laccase production showed higher with spherical mycelium than with filamentous or flocculent mycelium. In addition, the spherical mycelium with a diameter of 0.2-0.4 mm highly affected laccase production. Effect of the composition of culture medium on pellets was investigated and results indicated that various concentrations of glucose, corn meal and wheat bran were important to the formation of pellets in diameter of 0.2-0.4 mm. Besides nutrients, the addition of non-nutritional substrates influenced the distribution of P. ferulae pellets. However, the production of laccase was not promoted by non-nutritional substrates.

[Effects of medicinal insect Catharsius molossus on submerged fermentation and in vivo anti-hepatoma activity of Ganoderma lucidum].

We studied the effects of Catharsius molossus (a Chinese medicinal insect) on the cell growth, fermentation kinetics of key bioactive substances and anti-cancer activity of Ganoderma lucidum in submerged fermentation. The results showed that C. molossus at all the tested concentrations had no stimulatory effect on the cell growth. However, addition of C. molossus at 5 g/L lead to significant effects on the fermentation kinetics of polysaccharides and triterpenoids of G lucidum, and at 7th day in fermentation process, the yields of polysaccharides and triterpenoids reached 2.81 g/L and 539.0 mg/L, respectively, while they were 2.25 g/L and 428.2 mg/L in control. In vivo anti-cancer studies showed that the inhibitory rates of control fermented G lucidum (CFG) and a combination of water extract from C. molossus and CFG on the developed tumor (Heps) in mice were 41.61% and 42.24%, respectively. Moreover, the inhibitory rate of the G lucidum fermented with C. molossus (GFC) reached 57.21%, which was enhanced 37.49%, compared to the inhibitory rate of the control fermented G lucidum. These results suggest that supplementation of C. molossus in submerged fermentation of G lucidum lead to a significant enhancement of the anti-cancer activity of cultured G lucidum.

[Effects of medicinal insect, Catharsius molossus on biosynthesis of polysaccharides from Ganoderma lucidum in submerged culture].

We studied the effects of several medicinal insects on biosynthesis of polysaccharides from Ganoderma lucidum in submerged culture. The results showed that the medicinal insect, Catharsius molossus at 5 g/L significantly promoted the biosynthesis of intracellular polysaccharides (IPS) and extracellular polysaccharides (EPS) of G. lucidum, and compared with control, IPS and EPS yields markedly enhanced from (1.93 +/- 0.09) g/L to (2.41 +/- 0.12) g/L and (520.3 +/- 20.2) mg/L to (608.9 +/- 20.2) mg/L, respectively (P < 0.05). Both IPS and EPS consisted of five kinds of components, and IPS-1 and EPS-1 were the major components of IPS and EPS, respectively. Further separation studies showed that IPS-1 was made up of three single compounds, while EPS-1 was made up of two single compounds. There were no new components in both IPS and EPS obtained from G lucidum in submerged culture by the addition of the insect, C. molossus, suggesting the biosynthetic pathways of the major components of IPS and EPS had not been changed.

Enhancement of polysaccharides production in Ganoderma lucidum by the addition of ethyl acetate extracts from Eupolyphaga sinensis and Catharsius molossus.

To screen stimulators from Chinese medicinal insects for mycelial growth and polysaccharides production of Ganoderma lucidum, G. lucidum was inoculated into the media with and without supplementation of medicinal insect extracts. The ethyl acetate extract of Eupolyphaga sinensis at 55 mg l(-1) lead to significant increase in both biomass and intracellular polysaccharides (IPS) concentration from 8.53 +/- 0.41 to 14.16 +/- 0.43 and 1.28 +/- 0.09 to 2.13 +/- 0.11 g l(-1), respectively. In addition, the ethyl acetate extract of Catharsius molossus at 55 mg l(-1) significantly enhanced extracellular polysaccharides (EPS) production; the EPS yield increased from 350.9 +/- 14.1 to 475.1 +/- 15.3 mg l(-1). There were no new components in the two types of polysaccharides obtained by the addition of the insect extracts.

[Purification and characteristic of proteinase inhibitor GLPIA2 from Ganoderma lucidum by submerged fermentation].

A proteinase inhibitor GLPIA2 was purified to homogeneity from Ganoderma lucidum by submerged fermentation. The purification was carried out by ethanol fractional precipitation (30%-80%), gel-filtration on Superdex 200 column (30 cm x 1.1 cm i.d.) and anion exchange on Source 30Q column (10 cm x 1.6 cm i.d.). The gel chromatographic conditions were as follows: 50 mmol/L sodium phosphate as mobile phase with a flow rate of 1 mL/min with the effluent collection of 1 mL/tube and detection at 280 nm. The anion exchange chromatographic conditions were as follows: 50 mmol/L Tris-HCl (pH 8.8) containing different amounts of NaCl as mobile phase with a flow rate of 2 mL/min with the effluent collection of 5 mL/tube and detection at both 215 nm and 280 nm. Two active fractions named GLPIA1 and GLPIA2 corresponding to proteinase A inhibitory activities were pooled and lyophilized. GLPIA2 only has the absorption at 215 nm. The relative molecular mass of the inhibitor was 15,000 as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The amino acid composition of GLPIA2 was analyzed by high performance liquid chromatography. The chromatographic conditions were as follows: C18 column (125 mm x 4.0 mm i.d.) with column temperature of 40 degrees C; a mixture of 20 mmol/L sodium acetate-methanol-acetonitrile as mobile phase with a flow rate of 1 mL/min and detection at 338 nm. The results indicate that GLPIA2 is rich in acidic amino acid (Glu) and low in aromatic amino acids (Phe and Tyr). The interaction of some proteinases with GLPIA2 was investigated. The inhibitors are more potent against pepsin and yeast proteinase A than other proteinases.

Enhancement of mycelial growth and polysaccharide production in Ganoderma lucidum (the Chinese medicinal fungus, 'Lingzhi') by the addition of ethanol.

Methanol, ethanol, 1-propanol and 2-propanol, at 1.5% (v/v), enhanced the growth and polysaccharide production of Ganoderma lucidum. Ethanol was the most effective at 1.5% (v/v) for increasing the biomass production, however, the maximal polysaccharide concentration was produced with 2% (v/v) ethanol in the medium. There was no new polysaccharide component produced by the addition of ethanol.

[Enzymatic synthesis of monoglycerides in microaqueous media by using lipase from Pseudomonus fluorescens].

Enzymatic synthesis of monoglycerides by glycerolysis of oil and fats in microaqueous solvent-free media was investigated by using lipase from pseudomonus fluorescens (PFL). Initial eutectic point(IEP) was substituted for melt point of oil and fats in Critical Temperature Theory. By investigating the glycerolysis under different IEP, it is showed that there is a relationship between composition of the oils and the yield of monoglycerides: Y = -0.0006 X3 + 0.0592 X2 - 0.8909 X + 26.753(13% < X < 76.5%), here X is the contents(W/W) of saturated fatty acid residue (C16 + C18) in the oils, Y is the yield of monoglycerides at 40 degrees C. The optimum isothermal reaction conditions for a system which IEP is 40 degrees C are: 40 degrees C, 3%-4.5% (W/W) water in glycerol, dosage of lipase is 500 u/g oil when the mole ratio of glycerol to oil is 2.5:1. The highest yield of monoglycerides is 81.4% in 48 h by means of programming temperature reaction.

[Synthesis of partial glycerol caprates by using lipase in nonaqueous media].

Enzymatic esterification by reacting caparic acid with glycerol in solvent-free system was studied. Lipases from Pseudomonas fluoresces(PFL), Mucor miehei(MML) and Candida antarictica(CAL) possessed good catalytic activity. The optimal reaction conditions to convert capric acid with CAL are: 60 degrees C, 20-100 u of CAL per gram capric acid, 12% (W/W) of initial water content in glycerol. CAL does not express its 1,3-specificity in final product. Mechanical fraying denatured CAL partly. 96.4% of catalytic activity of CAL recovered after 5 batches of reaction. Extraction with sodium carbonate solution can decrease acid value of product from 9.8 mg KOH/g to 0.68 mg KOH/g. Applying the enzymatic esterification in open system, under vacuum or dehydrating with molecular sieves all dehydrate effectively. Molar ratio of reactants does not influence the total conversion of capric acid but influences the yield of monoglyceride. With certain protocols, the reaction period could be shortened dramatically; conversion of capric acid reached 96.9% in 5 h.