*2.5. Antimicrobial Activity against Gram-Negative Species*

The activity of the compounds was determined in the Gram-negative species of the ESKAPEE panel, *Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa,* and *Escherichia coli* (Table 6). None of the tested compounds displayed any antimicrobial activity in the Gram-negative species up to a concentration of 128 μg/mL. However, when the outer membrane was permeabilized using the membrane permeabilizer polymyxin-B-nonapeptide (PMBN), MICs as low as 0.5 μg/mL were observed, suggesting an intracellular target. With the exception of **6** + **7**, all compounds tested displayed a good activity in *A. baumannii* and *E. coli* strains and in *P. aeruginosa* strain PAO1. The *P. aeruginosa* strain NCTC 13437 is a near-pan drug resistant strain, and this was reflected in the MICs to compounds **6** + **7**, **10** + **11**, **11**, and **12**, although **8** had an MIC of 8 μg/mL in the presence of PMBN. None of the compounds displayed activity in the two *K. pneumoniae* strains, even in the presence of PMBN, suggesting the target/pathway might be missing or modified in this species.


**6.** Gram-negative MIC Values (in μg/mL) of compounds **6**–**8** and **10**–**12** in the presence of the membrane permeabilizer polymyxin-B-nonapeptide at 30

**Table**
