Production of luteoskyrin and isolation of a new metabolite, pibasterol, from Penicillium islandicum Sopp

Production of luteoskyrin, a hepatotoxin synthesized by Penicillium islandicum Sopp., was studied with various fermentation methods. Best results were obtained in static fermentations on glutinous rice at 30 degrees C. The isolated yield of pure luteoskyrin was approximately 400 mg per kg of rice. Also produced were skyrin, islandicin, iridoskyrin, rubroskyrin, chrysophanol, mannitol, and erythritol. A new metabolite, which we call pibasterol, was also isolated.     

Production of luteoskyrin and isolation of a new metabolite, pibasterol, from Penicillium islandicum Sopp.

Production of luteoskyrin, a hepatotoxin synthesized by Penicillium islandicum Sopp., was studied with various fermentation methods. Best results were obtained in static fermentations on glutinous rice at 30 degrees C. The isolated yield of pure luteoskyrin was approximately 400 mg per kg of rice. Also produced were skyrin, islandicin, iridoskyrin, rubroskyrin, chrysophanol, mannitol, and erythritol. A new metabolite, which we call pibasterol, was also isolated.     

Production of Luteoskyrin, a Hepatotoxic Pigment, by Penicillium islandicum Sopp

Various factors affecting the yields of luteoskyrin, a hepatotoxic mycotoxin, and related pigments in the liquid medium were studied. Maximal yields of luteoskyrin (0.13% by isolation) and of other pigments were attained in the late phase of the cultivation. The yield of the pigment was increased by supplying malt extract, malonic acid, glutamic acid, or asparagine. A useful material for preparation of (14)C-labeled luteoskyrin was 2-(14)C-malonate.     

The fungus Penicillium variabile sopp 1912 isolated from permafrost deposits as a producer of rugulovasines

It has been established that relict fungi Penicillium variabile Sopp can synthesize clavine alkaloids, rugulovasines A and B, which are revealed in this species for the first time. Submerged cultivation of the strain-producer revealed several microcycles of conidia formation. The synthesis of alkaloids was also of a cyclic character. The synchronism beween the cycles of rugulovasine biosynthesis and conidia formation was revealed. Zinc ions stimulated fungal growth but had a negative effect on the biosynthesis of rugulovasines.     

Production of cyclochlorotine and a new metabolite, simatoxin, by Penicillium islandicum Sopp.

Production of the hepatotoxin cyclochlorotine by Penicillium islandicum was studied under a variety of fermentation conditions. The best system for production was agitated red wheat. A thin-layer chromatographic method was developed for the detection of this cyclic polypeptide. These experiments have resulted in the isolation of a new toxic metabolite, which we call simatoxin.     

Effects of butylated hydroxyanisole,butylated hydroxytoluene and tertiary butylhydroquinone on growth and luteoskyrin production byPenicillium islandicum

Butylated hydroxyanisole (BHA), Butylated hydroxytoluene (BHT) and tertiary butylhydroquinone (TBHQ) alone in cultural media were tested for the inhibition of growth and luteoskyrin production by two toxigenic strains ofPenicillium islandicum UST-11 andP. islandicum HLT-6. In potato dextrose agar, the concentrations of BHA and TBHQ from 0.2 mg/disc, BHT from 5.0 mg/disc did affect the growth of both tested strains, but the initial concentrations of these antioxidants to reduced luteoskyrin production by UST-11 strain were BHA 0.5 mg/disc, BHT 1.0 mg/disc and TBHQ 0.4 mg/disc, while for HLT-6, BHA 0.4 mg/disc, BHT and TBHQ were 0.2 mg/disc, respectively. In grainy and powdery rice media, the effects of BHA, BHT and TBHQ on luteoskyrin production byP. islandicum UST-11 and HLT-6 were clearly demonstrated. The efficiency of the inhibitory effect was not only closely related to the concentration of antioxidants, but also completely inhibited the luteoskyrin production at a concentration of 200 mg/kg or higher. Also, the antioxidants at a concentration higher than 20 mg/kg reduced significantly the growth and luteoskyrin production by both strains ofP. islandicum.     

The interaction of rubroskyrin, a modified bis-anthraquinone pigment fromPenicillium islandicum Sopp, with respiratory chain of liver mitochondria

Summary  Rubroskyrin, a modified bisanthraquinone pigment from an yellow rice moldPenicillium islandicum Sopp, was examined for its redox-interaction with the mitochondrial respiratory chain by using rat liver submitochondrial particles (SMP) and was compared with luteoskyrin and rugulosin. Rubroskyrin showed a redox interaction with the NAD-linked respiratory chain of SMP, promoting NADH oxidase in the presence of rotenone, a specific inhibitor to coupling site I of the respiratory chain. Rubroskyrin-mediated NADH oxidase was not inhibited by antimycin A and cyanide, inhibitors to coupling sites II and III, respectively, indicating a generation of an electron transport shunt from a rotenone-insensitive site of NADH dehydrogenase (complex I) to dissolved oxygen. An electrontransport shunt to cytochromec oxidase from complex I was also observed in the experiment with cytochromec and antimycin A. Rubroskyrin did not interact with succinate-linked respiratory chain. Such enzymatic redox response which generates electron transport shunt was not detected for luteoskyrin and rugulosin in the present study.     

Hepatotoxicity of cyclochlorotine--a cyclic peptide produced by Penicillium islandicum]

Toxicological investigations of "Yellow Rice" which have been contaminated with various Penicillium species have shown that Penicillium islandicum apart from well-known hepatotoxic mycotoxin, lyteoskyrin synthesizes cyclochlorotine (chloropeptide, CP), which is also hepatotoxic. Isolation of chloropeptide and its chemical characteristics, acute toxicity and biochemical alterations, tissue distribution and excretion, deformation of hepatic cells, and interaction with cytoskeleton filaments are described. It is shown that the hepatotoxicity of CP is inactivated by dehalogenation of CP by the cytochrome P-450 system.     

Production of the Hepatotoxic Chlorine-Containing Peptide by Penicillium islandicum Sopp

Twenty-six strains of Penicillium islandicum Sopp were tested for production of a chlorine-containing peptide (Cl-peptide), which is a hepatotoxin. Highest levels of the mycotoxin were produced in a modification of Wickerham medium. The mycotoxin was isolated by adsorption on charcoal and, after washing the charcoal with acetone-water (1:1), was eluted into n-butanol. Further purification was accomplished by gel filtration. Maximum yields were 10 to 20 mg of toxin per liter of culture filtrate.     

Development of monoclonal-antibody-ELISA and -DIP-STICK immunoassays for Penicillium islandicum in rice grains

Monoclonal antibodies (mAbs) were raised against Penicillium islandicum, a fungus commonly isolated from stored rice grains in South-East Asia. Mice were immunized by a direct, simple method; fresh cell-free surface washings from a solid agar slant culture were injected directly into the peritoneum without prior concentration. Hybridoma supernatants were screened by ELISA. Most of the mAbs raised cross-reacted with other storage fungi and/or uninfected rice grains but there were species-specific. One of these, PI01, was used to develop ELISA and DIP-STICK assays for the detection of P. islandicum in individual grains. All inoculated grains and approximately 90% of grains in natural infected samples from Indonesia tested positively, by ELISA, for P. islandicum. This result and those obtained for discoloured grains from both Indonesia and the Philippines, 32% and 14% respectively, are higher than those obtained by direct plating of surface-sterilized grains. Heat and periodate treatment of the PI01 antigen and binding on Western blots indicate that it is a glycoprotein of Mr greater than 90,000. Hyphae of all ages stained uniformly by immunofluorescence using the PI01 antibody but mature conidia stained only weakly.