Gram-negative bacteria tend to be more resistant to lipophilic and amphiphilic inhibitors than gram-positive bacteria. Such inhibitors include dyes, detergents, free fatty acids, antibiotics, and other chemotherapeutic agents. This property is used in the selective enrichment of gram-negative bacteria, especially of members of the family Enterobacteriaceae, for example with MacConkey agar (containing crystal violet and bile salts), EMB agar (containing dyes), and deoxycholate agar (containing sodium deoxycholate). The fact that Escherichia coli K-12 can grow in the presence of 1% sodium dodecyl sulfate has been rediscovered many times. Many lipophilic antibiotics, such as penicillin G, erythromycin, fusidic acid, and rifamycin SV, are much less active against most gram-negative bacteria. In fact, a survey of recently reported antibiotics of natural origin showed that, among those compounds that showed activity against gram-positive bacteria, more than 90% lacked activity at a useful level against E. coli (51). This intrinsic resistance of gram-negative bacteria has often been attributed entirely to the presence of the outer membrane barrier. This barrier does contribute to the resistance, as the narrow porin channels slow down the penetration of even small hydrophilic solutes, and the low ffuidity of the lipopolysaccharide leaffet decreases the rate of transmembrane diffusion of lipophilic solutes (38, 40). However, the outer membrane barrier cannot be the whole explanation, even with species such as Pseudomonas aeruginosa which produces an outer membrane of exceptionally low permeability (1, 57). This is seen from the fact that equilibration across the outer membrane is achieved very rapidly, in part because the surface-to-volume ratio is very large in a small bacterial cell. Thus, the periplasmic concentrations of many antibiotics are expected to reach 50% of their external concentrations in 10 to 30 s in P. aeruginosa and in a much shorter time period in E. coli (34). Additional mechanisms are therefore needed to explain the level of intrinsic resistance. With the earlier β-lactam compounds, this second contributing factor is the hydrolysis by the periplasmic β-lactamases that are encoded by chromosomal genes in many gram-negative bacteria, and the levels of resistance can be explained quantitatively, in many cases, by the synergy between the outer membrane barrier and β-lactamase (37). However, with dyes, detergents, other classes of antibiotics, and even the β-lactams developed more recently that are not hydrolyzed easily by common β-lactamases, the second factor was completely unknown (34). Recent studies showed that multipledrug efffux pumps, many with unusually broad specificities, play a major role in the intrinsic resistance of gram-negative bacteria.