Actaplanin B1

A bioautographic method is described that allows the identification and semiquantitation of residues of the glycopeptide antibiotic actaplanin in cow's milk at 0.01 ppm. The milk sample is precipitated with acetonitrile and the resultant pellet is extracted by a buffer. This extract is defatted, then chromatographed on an Amberlite resin column. Actaplanin is eluted with methanol-HCl. Purified extract is then chromatographed on a cellulose thin layer, developed in a methanol-chloroform-ammonium hydroxide mixture. This thin layer chromatography increased the sensitivity of the determination by concentrating the actaplanin components in a single spot. The antibiotic was then detected by bioautography, using Bacillus subtilis as a test organism. Parameters influencing the diffusion of the antibiotic (namely, the nature and concentration of the agar-agar), and the sensitivity of the test strain (namely, the nutritional value of the medium, and the addition of a synergistic inhibitor) have been optimized.

A solid-phase enzyme-receptor assay (SPERA) has been developed for glycopeptide antibiotics of the vancomycin class such as teicoplanin, vancomycin, ristocetin, avoparcin, actaplanin, A-47934, A-41030, and A-35512-B. The assay exploits the mechanism of most action of these antibiotics, which is based on their interaction with acyl-D-alanyl-D-alanine, a constituent of the walls of most growing bacterial cells. The antibiotics and enzyme-labeled teicoplanin compete for a synthetic analog of the biological receptor, albumin-epsilon-aminocaproyl-D-alanyl-D-alanine. The various antibiotics produced different competition curves, 50% displacement being obtained with antibiotic concentrations ranging from 0.04 to 4 mg/L, vancomycin and actaplanin being the weakest and strongest competitors, respectively. For teicoplanin in human serum the intra-assay CV was 7.2%, the interassay CV was 11.2%, and the analytical recovery 94%. Teicoplanin concentrations obtained by SPERA (chi) correlated well with those obtained by microbiological assay (y): y = 1.03 chi + 0.053 (r = 0.943; n = 60). We conclude that SPERA is a powerful tool for identification and quantitative detection of glycopeptide antibiotics, even in complex media.

Brahman (16), Brangus (32) and Simmental X Brahman (16) open heifers were assigned to four treatment levels and two body weight categories. The weight categories were light (250 kg) and heavy (306 kg). Treatment groups were 0, 600, 900 and 1,200 mg actaplanin.head-1.feeding-1 with feeding only on Monday and Friday. Heifers were group-fed .91 kg/head of supplement (95% soybean meal: 5% molasses) containing actaplanin. Heifers grazed eight Coastal bermudagrass pastures throughout the experiment (July 29 to November 15, 1982). After 75 d on experiment, an additional 1.36 kg of a commercial 14% crude protein supplement was fed on the days that actaplanin was not fed. All heifers were weighted and rotated among pastures at 14-d intervals (last weight period was 11 d) as an attempt to equalize forage availability among groups. Over the entire experiment (109 d), light-weight heifers that were fed actaplanin responded (P less than .01) with greater average daily gain (ADG; .37, .35, .21 kg/d; 600, 900, 1,200) compared with controls (0 mg; .18 kg/d). A trend in response (P less than .10) to actaplanin feeding occurred for heavy-weight heifers (.23, .38, .34 kg/d; 600, 900, 1,200 mg vs controls (.22 kg/d). For light-weight heifers, ADG decreased lineraly (P less than .01) as actaplanin increased from 600 to 1,200 mg, while ADG for heavy-weight heifers increased linearly (P less than .10).