In the preceding paper (1) we developed the partial structure (I) for the macrolide antibiotic lucensomycin from Streptomyces lucensis. A close similarity was apparent between lucensomycin and the structure suggested by Ceder and co-workers (2) for pimaricin, a metabolite of S. natalensis. Apart from the definitive placement of some minor structural features, lucensomycin appeared to be a higher homologue of pimaricin in which a n-butyl group replaced the methyl group present at C(R5) in the latter. Mass spectrometry of trimethylsilyl derivatives, a procedure developed by two of us (3) for the determination of molecular formulae of polyhydroxylic compounds of high molecular weight, has recently (5) necessitated revision of the molecular formula of pimaricin from C33H47NO14 (2) to C33H47NO13, with a corresponding alteration in the structural formula to (II, R = CH3). At this stage of the lucensomycin work, we determined the mass spectra of some TMS derivatives of lucensomycin for comparison with the corresponding pimaricin spectra. N-chloroacetyllucensomycin and N-acetyldodecahydrolucensomycin (in which the epoxide ring has been opened by hydrogenolysis) afforded TMS derivatives, the mass spectra of which indicated molecular formulae C56H102ClNO14Si6 (m+ 1215) and C59H123NO14Si7 (m+ 1265) respectively. These represent penta- and hexa-TMS ethers of N-acyl TMS esters, and necessitate that lucensomycin itself has the molecular formula C36H53NO13 containing only five hydroxyl groups. The inaccuracy of analytical data (4) from which the previously accepted C36H53NO14 formula was calculated is probably due to solvation of the macrolide crystals (cf. 3,5). With this revision of the molecular formula of lucensomycin, it is now necessary to accommodate only one additional hydroxyl group together with the epoxide in the partial structure (I). Moreover, the close similarity of fragmentation patterns present in the above-mentioned spectra to those of TMS-N-acetyl derivatives of pimaricin and dodecahydropimaricin indicates that, apart from possible stereochemical differences, lucensomycin is the n-butyl homologue of the revised structure (II: R = CH3) for pimaricin. Together with the additional chemical evidence summarized below, these facts permit us to assign to lucensomycin the structure (II: R = n-C4H9). Biogenetically lucensomycin carries an extra propionate unit in the aglycone compared to pimaricin.