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A search for novel insecticides has yielded the milbemycins, a new family of macrolide antibiotics with insecticidal and acaricidal activity. They are produced in submerged cultures of Streptomyces hygroscopicus subsp. aureolacrimosus. Fermentation studies on the strain were conducted in shaken flasks and 30-liter jar fermentors.From the culture broth 13 milbemycins were purified to homogeneity by column and thin-layer chromatography on silica gel and alumina. Physico-chemical data, such as mass spectra, UV and IR absorption spectra, optical rotations and melting points of the milbemycins are described.In a search for novel insecticides, we discovered that the culture broth of a Streptomyces was active against acarus, harmful agricultural and horticultural insects such as aphids and larvae of insects of the order Lepidoptera.1) The Streptomyces is a new subspecies and is designated S. hygroscopicus subsp.aureolacrimosus. Taxonomic studies will be reported elsewhere.From the culture broth thirteen milbemycins were isolated and purified to homogeneity by column and thin-layer chromatography on silica gel and alumina. As reported briefly,2,3) the milbemycins have the 16-membered macrolide structure shown in Fig. 1.Subsequently the avermectins, isolated from the culture broth of Streptomyces avermitilis, were found to have structures closely related to the milbemycin as, differing only in being substituted at the C-13 position by an oleandosyl oleandosyloxy moiety.4,5)In this paper we describe the fermentative production, isolation and physico-chemical properties of the milbemycins. Materials and Methods Fermentation studiesCultures were grown in Erlenmeyer flasks on a rotary shaker (220 rpm, 7 cm) at 28°C. Inocula were grown for 2 days in 500-m1 Erlenmeyer flasks containing 50 ml of seed medium with the following ingredients (in g/1,000 ml water): glucose 20, soybean meal 10, corn-steep liquor 5 and NaCl 3. The pH of the medium was adjusted to 6.8 before autoclaving at 120°C for 20 minutes.Production cultures in 100-ml Erlenmeyer flasks, each containing 30 ml of the medium, were inoculated with 1 ml of the seed culture.Fermentations with the Streptomyces were also conducted * The antibiotics were initially designated as B-41 .

Dentin powders from bovine incisors were treated with 10% NaClO solution. Differential thermal analysis (DTA) indicated the removal of organic material from the dentin sample following the treatment, since the exothermic reaction at 320 degrees C had disappeared. X-ray diffraction studies revealed a change in the crystallinity of the dentin crystals and the formation of calcite after the treatment. Infrared absorption analysis showed that the band due to carbonate ions was weakened after the treatment, while atomic absorption spectroscopic analysis showed that magnesium ions had been dissolved from the dentin sample. The a-axis lengths of treated and heated dentin samples differed from those of untreated and heated samples. Whitlockite was always found in the untreated/heated samples, whereas it was absent in the treated/heated samples. The unit cell dimensions of the whitlockite indicated the partial substitution of magnesium for calcium. Magnesium ions seemed to be more effective than carbonate ions in forming whitlockite. These results showed that some magnesium and carbonate ions were removed from the dentin crystal structure upon NaClO treatment, while at the same time organic materials were removed from the dentin sample. It was suggested that crystals in the NaClO-treated dentin were similar to enamel crystals from a crystallographic viewpoint.

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We investigated the effect of interleukin 6 (IL-6) on the migration of rabbit corneal epithelium in vitro and on the attachment of dissociated corneal epithelial cells to a fibronectin matrix. When corneal blocks were cultured with IL-6 for 24 hours, the length of the path of epithelial migration over exposed corneal stroma increased significantly (p less than 0.005 at the concentration of 10 ng/ml) in proportion to the concentrations of IL-6 (0.1-10.0 ng/ml). The addition of antiserum against fibronectin or of GRGDSP abolished the stimulatory effect of IL-6 on epithelial migration. When corneal epithelial cells were cultured with various concentrations of IL-6, suspended, and plated on wells coated with fibronectin (10 micrograms/ml), the number of cells attached to the wells increased in a dose-dependent manner. The presence of antibody against fibronectin or of GRGDSP during the attachment assay decreased the number of cells attached to the fibronectin matrix, regardless of the fact that the cells had been cultured with IL-6 or not. IL-6 stimulated the attachment of corneal epithelial cells to collagen type IV and to laminin matrices. However, the presence of GRGDSP did not affect the cell attachment to collagen type IV and to laminin. These findings strongly indicate that IL-6 stimulates epithelial migration in the cornea by a fibronectin-dependent mechanism, presumably the increased expression of fibronectin receptors.