During the course of screening for inhibitors of trehalase, we found the pseudodisaccharide termed trehazolin1 (1) in a culture broth of Micromonospora, strain SANK 62390n. This paper describes the isolation, structure determination, and inhibitory activity of trehazolin. One loopful of the producing organism was aseptically transferred from an agar slant into baffled 500-ml Erlenmeyer flasks each containing 80ml of a primary seed medium.This mediumwas composed of glucose 1%, glycerol 1%, oatmeal 0.5%, sucrose 1%, soybean meal 2%, Casamino acids 0.5%, pressed yeast 1%, CaCO3 0.1%, and Disfoam CB-442 (Nippon Yushi Co.) 0.01%. After formulation, the pH was adjusted to 7.0 with aq NaOH, and the mediumwas sterilized. The incubation was performed on a rotary shaker at 220rpmat 28°C for 216 hours. Forty ml of the primary seed culture were aseptically transferred into baffled 2-liter Erlenmeyer flasks each containing 800ml of a secondary seed/production medium which was composed of glucose 2%, soluble starch 1 %, pressed yeast 0.9%, meat extract 0.5%, Polypepton 0.5%, NaCl 0.5%, CaCO3 0.3%, and Disfoam CB-442 0.01%. The medium was then adjusted to pH 7.2
Three metabolites were isolated from the culture broth of an actinomycete strain identified as Streptomycesplatensis SANK601 91 , that induce the production of colony-stimulating factors (CSFs) by stromal cell line KM-102at ED50concentrations from 40 to 200 ng/ml. The compounds induced quantities of granulocyte CSF (G-CSF) and granulocyte-macrophage CSF (GM-CSF) comparable to those induced by interleukin-1 , a strong CSFinducer. These metabolites were called leustroducsins (A, B and C) and were later found to be structurally related to phoslactomycins. This is the first report of CSFinducing activity by members of the phoslactomycin class. 1503Recent studies have demonstrated the application of colony-stimulating factors (CSFs) in clinical use1}. These substances have been used to recover the peripheral blood leukocytes in leukopenia patients caused by cancer-chemotherapy, radiotherapy and bone marrowtransplantation. WhenCSFs were administered to patients, restorations ofleukocyte counts occurred. However,the in vivo role ofendogenous CSFs is not precisely understood. In particular, little is known about the regulatory mechanism for CSF production or about the relationship between normal blood cell production and endogenous CSFs. It is well knownthat bone marrow stromal cells play an important role in hematopoiesis. Regulation of CSF production by bone marrow stromal cells may be one of the key elements responsible for the control of hematopoiesis in vivo2). Therefore, substances that affect the regulation of CSF production by stromal cells are of potential interest, so we worked to develop screening methods for CSF inducers.In the previous paper3), we described the development of a new screening method for CSF inducers using human bone marrow stromal cell line KM-102. Using this screening method, one strain of actinomycetes, Streptomyces platensis SANK60191, was found to produce novel microbial metabolites that we provisionally namedleustroducsins. Structure determination studies revealed that they are congeners of phoslactomycins. In this paper we report the taxonomy of the producing organism, and also describe the fermentation and biological properties of leustroducsins (LSNs), A, B and C (Fig. 1). The isolation, physicochemical properties and structural elucidation of the metabolites are reported in the accompanying paper4).
A strain of actinomycetes identified as Streptomyces flavidovirens produced new antibiotics, mureidomycins (MRD's) A~D, specifically active against Pseudomonas aeruginosa. They were isolated from the culture filtrate by successive columnchromatographies such as Amberlite XAD-2and CG-50, Whatman DE-52 and Toyopearl HW-40. They were amphoteric white powders and soluble in methanol and water. Their molecular weights and molecular formulae in parentheses were 840 (CS8H48N8O12S), 842 (CsgHsoNgO^S), 897 (C^B^NgO^S) and 899 (C^H^NgO^S), respectively. m-Tyrosine and two unknown substances were detected by aminoacid analyses as their common constituents. MRD's A and C contained uracil but MRD'sB and D dihydrouracil instead of uracil.In the course of our screening program for new antibiotics with spheroplast forming activity, one strain1* of actinomycete, identified as Streptomyces flayidovirens SANK60486, was found to produce new antibiotics with specific activity against Pseudomonasaeruginosa.In this paper, we report the taxonomy of the producing organism, the fermentation, isolation and physico-chemical properties of mureidomycins (MRD's) A^B1^. The structural elucidation and biological properties of the antibiotics will be described in the accompanyingpapers3>4). Materials and Methods Taxonomic StudiesThe producing organism, strain SANK60486, was isolated from a soil sample collected at Miyake Island, Tokyo, Japan. Methods adopted by Waksman5)and by the International Streptomyces Project (ISP)6) were used for the studies of morphological and taxonomic characterizations, carbohydrate utilization and taxonomic identification.The procedure of Becker et alP was used for the preparation of cells and chromatographic detection of the isomers of diaminopimelic acid. Fermentati onA loopful amount of the culture of strain SANK 60486 was inoculated into a 500-ml Erlenmeyer flask which contained 80 ml of the medium consisting of glucose 3 %, pressed yeast 1 %, soybean meal 3%, CaCO3 0.4%, MgSO4-7H2O 0.2% and Nissan Disfoam CB-442 0.01 %. The inoculated flasks were incubated on a rotary shaker (220 rpm) at 22°C for 48 hours. Then a 2-ml aliquot of the culture was transferred into 2-liter flask containing 500 ml of the same medium. After 24 hours of incubation on a rotary shaker, 750 ml of this second seed culture was added to a 30-liter jar fermentor con-å "å Present address:
In the course of our screening for compounds which inhibit Rb protein phosphorylation at the cell level and cause cell cycle arrest at the Gl phase1}, reblastatin (Fig. 1) was found as a minor component of geldanamycin2~4) from the culture of Streptomyces hygroscopicus subsp. hygroscopicus SANK 61995 deposited as FERM BP-5140. One loopful of the producing strain was inoculated into 80ml of the seed medium consisting of glycefol 2.5%, glucose 2.5%, pressed yeast 1%, soybean meal 1%, CaCO3 0.5%, KH2PO4 0.05%, MgSO4-7H2O 0.05% and CB-442 (Nihon Yushi Co.) 0.005%, pH 7.0, in a 500 ml-Erlenmeyer flask. The seed culture was carried out for 4 days at 28°C on a rotary shaker (210 rpm). One milliliter of the culture was added to 80ml of a mediumconsisting of the same composition as the seed mediumin a 500ml-Erlenmeyer flask and the fermentation was carried out for 4 days at 28°C on a rotary shaker (210 rpm). An equal amount of acetone was added to the harvested culture (800 ml) after the pH was adjusted to 7.0 and the active substance was extracted for one hour by stirring. The
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