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The avermectins, a family of new anthelmintic agents, were isolated from the mycelia of Streptomyces avermitilis. Four closely related major components and four homologous minor components were separated from the complex. Solvent extraction, solvent partition, and adsorption methods were used to isolate and purify the complex; novel partition chromatography systems using Sephadex LH-20 were used to separate the components. A reverse-phase high-pressure liquid chromatography assay for the quantitative determination of all components was used extensively to monitor the purification methods.The avernectin complex, a family of new anthelmintic agents, is produced by fermentation of Streptomyces avermitilis (NRRL 8165), as described in a separate publication (1).The complex contains four closely related major components, Ala, A2a, Bia, and B2a, in varying proportions and four minor components, Alb, A2b, Blb, and B2b, each of which is a lower homolog ofthe corresponding major component. The structures and physicochemical properties of all components are the subject of a separate publication (G. Soc., in press).In this report, the isolation of the complex from broth, the separation of all of its components, and the chromatographic properties of each will be described. For the sake of simplicity, mixtures of homologs will be designated by omis- B2a, 6.74; A2b, 7.22; A2s, 8.00; B,I, 8.32; B,I., 9.70; Al,b, 11.06; and A,,, 11.78. The UV monitor was set at 246 nm to improve the resolution of a major impurity which exhibited a UV maximum at a lower wavelength. Although the UV maximum for avermectins occurred at 245 nm, sensitivity was only slightly reduced at 246 nm, whereas the sensitivity for the impurity was reduced substantially. The UV spectrum of B21 is given in Fig. 1 and is characteristic of all components. Recordings of high-pressure liquid chromatography (HPLC) assays of standard and unknown solutions are given in Fig. 2. RESULTSIsolation of avermectin complex. Approximately 8,000 liters of fermentation broth was filtered, and the cake was washed with water. The wet cake was then slurried in 3,000 liters of acetone, filtered, and washed with 400 liters of 80% acetone. The cake was discarded, and the combined extract and wash were concentrated to 800 liters. The concentrate was adjusted to pH 5 with dilute HCl and then extracted twice with 800-liter portions of methylene chloride.

The isolation of difficidin (1) and oxydifficidin (2) from fermentation broth of Bacillus subtilis ATCC39320 and the physico-chemical characterization of these labile antibiotics are described. Thestructures of the compounds represent a newclass of antibiotics, characterized as highly unsaturated 22-membered macrolide phosphates. Difficidin and oxydifficidin undergo reversible thermal isomerization to 3 and 4 respectively. Biological evaluation of the isomers is presented.The preceding paper1} describes the discovery of two novel antibiotics, diffiddin (1) and oxydifficidin (2), as products of the fermentation of Bacillus subtilis strains ATCC39320 and ATCC39374.The compounds exhibit good antibacterial activity against both aerobic and anaerobic organisms.The mode of action of difficidin is addressed in the following the paper2). During isolation work on difficidin and oxydifficidin, minor antibacterial components 3 and 4 were also isolated from fermentation broth. This paper presents the isolation and physico-chemical properties of compounds 1~4. Experimental data show that 3 and 4 arise from thermal isomerization of difficidin and oxydifficidin respectively. The in vitro and in vivo antibacterial potencies of 3 and 4 are compared to those of difficidin and oxydifficidin.Scheme 1.

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could be due to worker contamination through contact with previously used equipment or to exposure to contaminated surfaces in the general work environment.The developed procedure provides a rapid, reproducible, and sensitive method for monitoring occupational exposure to fenitrothion. The method should be useful for monitoring exposure to organophosphorus pesticides in general and is currently being employed in our laboratory for assessing exposure to azinphos-methyl.