Partitioning of Seven Different Classes of Antibiotics into LPS Monolayers Supports Three Different Permeation Mechanisms through the Outer Bacterial Membrane

Abstract

The outer membrane (OM) of Gram-negative (G−) bacteria presents a barrier for many classes of antibacterial agents. Lipopolysaccharide (LPS), present in the outer leaflet of the OM, is stabilized by divalent cations and is considered to be the major impediment for antibacterial agent permeation. However, the actual affinities of major antibiotic classes toward LPS have not yet been determined. In the present work, we use Langmuir monolayers formed from E. coli Re and Rd types of LPS to record pressure–area isotherms in the presence of antimicrobial agents. Our observations suggest three general types of interactions. First, some antimicrobials demonstrated no measurable interactions with LPS. This lack of interaction in the case of cefsulodin, a third-generation cephalosporin antibiotic, correlates with its low efficacy against G– bacteria. Ampicillin and ciprofloxacin also show no interactions with LPS, but in contrast to cefsulodin, both exhibit good efficacy against G– bacteria, indicating permeation through common porins. Second, we observe substantial intercalation of the more hydrophobic antibiotics, novobiocin, rifampicin, azithromycin, and telithromycin, into relaxed LPS monolayers. These largely repartition back to the subphase with monolayer compression. We find that the hydrophobic area, charge, and dipole all show correlations with both the mole fraction of antibiotic retained in the monolayer at the monolayer–bilayer equivalence pressure and the efficacies of these antibiotics against G– bacteria. Third, amine-rich gentamicin and the cationic antimicrobial peptides polymyxin B and colistin show no hydrophobic insertion but are instead strongly driven into the polar LPS layer by electrostatic interactions in a pressure-independent manner. Their intercalation stably increases the area per molecule (by up to 20%), which indicates massive formation of defects in the LPS layer. These defects support a self-promoted permeation mechanism of these antibiotics through the OM, which explains the high efficacy and specificity of these antimicrobials against G– bacteria.

Mechanistic Understanding from Molecular Dynamics in Pharmaceutical Research 2: Lipid Membrane in Drug Design

“…Indocyanine green [144,170] Potential anticancer drug Curcumin [164,[227][228][229], Aplysiatoxin [230], Bryostatin [231], Phorbol [231], 12,13-dibutyrate [231], Prostratin [231] Antibiotics Imipenem [232], Doripenem [232], Ertapenem [232], Meropenem [232], Ciprofloxacin [233,234], Ciprofloxacin ternary copper complex [235], Daunorubicin [209], Idarubicin [209],…”

“…Levofloxacin [236][237][238], Clarithromycin [236], Isoniazid N -acylated derivatives [239], Rifampicin [234,240], Mangostin [241], Trimethoprim [242], Negamycin [243] Potential antibiotic Kanamycin A [133], nTZDpa and its derivatives [244], Cholic acid derived amphiphiles [245], γ-terpineol [246], Bithionol [247] Antimicrobial compound Chlorhexidine [248][249][250], Triclosan [251], Octenidine [250] Antiparasitic Praziquantel [252] Antiviral drugs Darunavir [253], Amantadine [254][255][256], Spiro[pyrrolidine-2,2 -adamantane] [254,255], 20,30-dideoxyadenosine (Didanosine) [242], Saffron [257] Antifungal drug Itraconazole, [184,186,187,258], Nystatin [259], Amphotericin B [260] Rheumatoid arthritis Lapatinib [213] Nonsteroidal antiinflammatory drugs, inhibitor of cyclooxygenase-1 and -2…”

Langmuir Monolayer Techniques for the Investigation of Model Bacterial Membranes and Antibiotic Biodegradation Mechanisms

“…The controlled density of monolayer packing provides the antibiotic molecules access to all parts of lipid molecules through a range of compression states and also allows estimation of the molecular insertion area. Cethuk et al [116] have applied the Langmuir monolayer method to confirm that the structure of the antibiotic has a significant impact on the incorporation mechanism of the compound in the model membrane. They have revealed that the hydrophobicity of the antibiotic is a desirable feature which helps in its insertion and retention inside the aliphatic slab of LPS over a range of pressures.…”

“…The measurements using the Langmuir technique have also confirmed that the polar/electrostatic mode is obviously more disruptive for the LPS layer, which explains why aminoglycosides and polymyxins are more effective and specific against Gram-negative bacteria. Moreover, based on the data obtained with the Langmuir technique, the authors of [116] have proposed some simple criteria to predict which compounds will cross the membrane. It has been suggested that if an antibiotic can hold a detectable mole fraction among LPS molecules during compression all the way to reach the collapse pressure, then it can be expected that this drug would likely permeate through the native bacterial membrane.…”

A Novel Lipid-Based MALDI-TOF Assay for the Rapid Detection of Colistin-Resistant Enterobacter Species

“…Colistin, or polymyxin E, is a cationic lipopeptide and “last-resort” antibiotic used for MDR Gram-negative infections, such as those producing extended-spectrum beta-lactamases and those resistant to carbapenems ( 6 ), which is an increasing concern in clinical isolates of Enterobacter species ( 7 ). Targeting the cell membrane, cationic colistin binds with anionic lipopolysaccharide (LPS) through electrostatic interactions ( 8 – 11 ). Colistin resistance is conferred by reduction of the electronegativity of the cell membrane through modification of the terminal phosphate residues of lipid A, the membrane anchor of LPS, thereby decreasing the binding affinity of colistin ( 12 ).…”