Studies on the reactions of daptomycin and aldehydes

The objectives of this project were to determine the reaction pathways of daptomycin in the presence of glyceraldehyde in acidic solutions, to quantitate the kinetics of the major pathways, and to employ this reaction to develop a daptomycin drug delivery system; i.e. daptomycin-activated dextran 40 conjugates. The macromolecular conjugates were targeted to deliver daptomycin to the infected fibrin clot of bacterial endocarditis caused by gram-positive bacteria.; In the presence of glyceraldehyde, daptomycin formed 2 major products separable by RP-HPLC. The products were identified using UV spectroscopy, fluorimetry, mass spectrometry, and 2D-1H NMR to be imine and anilide derivatives. The reaction scheme was proposed. Carbinolamine was believed to be a common intermediate in formation pathways of both products. The carbinolamine intermediate underwent either acid catalyzed dehydration resulting in imine formation or intramolecular hydrogen bonding and bond cleavage giving rise to anilide formation. The reaction of daptomycin and glyceraldehyde was first-order with respect to daptomycin and glyceraldehyde concentration. In a pH range of 1–7, the imine formation rate was pH dependent with a maximum rate at pH of ∼3–4. The observed pH dependence was consistent with the pH dependence of typical amine-aldehyde reactions.; Daptomycin was coupled to activated dextran 40 in the pH range of 2 to 7. Periodate oxidation was employed to chemically modify dextran hydroxyl groups to aldehydes. The number of aldehyde groups substituted depended on periodate concentration but was independent of reaction duration or dextran molecular weight. Reaction samples were separated according to molecular size on a Sephadex G-100 column using 10% acetonitrile/water as a mobile phase. The UV absorption maxima and fluorescence intensities of high and low molecular weight fractions were consistent with that of imine and anilide products. Therefore, the reaction between daptomycin and activated dextran 40 followed the proposed reaction scheme for daptomycin and glyceraldehyde. Fibrinogen affinity of daptomycin conjugates was relatively higher than that of daptomycin using surface plasmon resonance (SPR). Less than 9% of daptomycin was released from the conjugates probably due to the presence of excess aldehydes remaining in the activated dextran.