Comparing micellar, hemolytic, and antibacterial properties of di- and tricarboxyl dendritic amphiphiles.

Homologous dicarboxyl dendritic amphiphiles-RCONHC(CH(3))(CH(2)CH(2)COOH)(2), 4(n); and ROCONHC(CH(3))(CH(2)CH(2)COOH)(2), 5(n), where R=n-C(n)H(2)(n)(+1) and n=13-22 carbon atoms-were synthesized. Critical micelle concentrations (CMCs) in aqueous triethanolamine solutions and at pH 7.4 were measured along with hemolytic activity (effective concentrations, EC(10)) in phosphate-buffered saline (PBS). LogCMC showed a linear dependence on chain length (n); the longest chain in each series had the lowest CMC-in triethanolamine: 4(21), 180µM and 5(22), 74µM and at pH 7.4: 4(21), 78µM and 5(22), 33µM. These two series, 4(n) and 5(n), and three series of homologous tricarboxyl dendritic amphiphiles-RCONHC(CH(2)CH(2)COOH)(3), 1(n); ROCONHC(CH(2)CH(2)COOH)(3), 2(n); RNHCONHC(CH(2)CH(2)COOH)(3), 3(n), where R=n-C(n)H(2)(n)(+1) and n=13-22 carbon atoms-were tested for growth inhibition of Staphylococcus aureus strain ATCC 6358 and methicillin-resistant S. aureus (MRSA) strain ATCC 43330 by microdilution in 0.1-strength brain heart infusion broth (BHIB). Amphiphiles 4(19), 4(21), 5(18), and 5(20) showed the strongest antibacterial activity (2.2-3.4µg/mL) against S. aureus (vancomycin, MIC=0.25µg/mL). These four plus 1(21), 2(20), 2(22), and 3(20) exhibited the strongest antibacterial activity (1.7-6.8µg/mL) against MRSA (vancomycin, MIC=0.25µg/mL). The MICs of these amphiphiles against six clinical MRSA were similar to those against the ATCC strain. In PBS, EC(10)s of the most active homologues ranged from 7 to 18µg/mL and 18 to 220µg/mL for di- and tricarboxyl dendritic amphiphiles, respectively. To assess the potential safety of using dendritic amphiphiles as drugs, measurements of micellar and hemolytic properties were conducted in the same medium (full-strength BHIB) that was used for antibacterial activity. The CMCs (9-36µg/mL, ∼18-72µM) of ten amphiphiles were measured by microdilution (log2 progression) with dye-covered beads. The EC(10)s were similar to those in PBS. The MICs of most amphiphiles (14-72µg/mL) and vancomycin (1.1-2.2µg/mL) against both S. aureus and MRSA increased significantly compared to the MICs measured in 0.1-strength BHIB. The one exception, 5(18), had an MIC against S. aureus of 1.1µg/mL compared to vancomycin (2.2µg/mL). With CMC (9-18µg/mL) and EC(10) (16µg/mL) values higher than the MIC, 5(18) was discovered as a lead for further development.

Comparing anti-HIV, antibacterial, antifungal, micellar, and cytotoxic properties of tricarboxylato dendritic amphiphiles.

Three series of homologous dendritic amphiphiles--RCONHC(CH(2)CH(2)COOH)(3), 1(n); ROCONHC(CH(2)CH(2)COOH)(3), 2(n); RNHCONHC(CH(2)CH(2)COOH)(3), 3(n), where R = n-C(n)H(2n+1) and n = 13-22 carbon atoms--were assayed for their potential to serve as antimicrobial components in a topical vaginal formulation. Comparing epithelial cytotoxicities to the ability of these homologues to inhibit HIV, Neisseria gonorrhoeae, and Candida albicans provided a measure of their prophylactic/therapeutic potential. Measurements of the ability to inhibit Lactobacillus plantarum, a beneficial bacterium in the vagina, and critical micelle concentrations (CMCs), an indicator of the potential detergency of these amphiphiles, provided additional assessments of safety. Several amphiphiles from each homologous series had modest anti-HIV activity (EC(50) = 110-130 microM). Amphiphile 2(18) had the best anti-Neisseria activity (MIC =65 microM), while 1(19) and 1(21) had MICs against C. albicans of 16 and 7.7 microM, respectively. Two measures of safety showed promise as all compounds had relatively low cytotoxic activity (EC(50) = 210-940 microM) against epithelial cells and low activity against L. plantarum, 1(n), 2(n), and 3(n) had MICs490, 1300, and 940 microM, respectively. CMCs measured in aqueous triethanolamine and in aqueous potassium hydroxide showed linear dependences on chain length. As expected, the longest chain in each series had the lowest CMC-in triethanolamine: 1(21), 1500 microM; 2(22), 320 microM; 3(22), 340 microM, and in potassium hydroxide: 1(21), 130 microM; 3(22), 40 microM. The CMC in triethanolamine adjusted to pH 7.4 was 400 microM for 1(21) and 3900 microM for 3(16). The promising antifungal activity, low activity against L. plantarum, relatively high CMCs, and modest epithelial cytotoxicity in addition to their anti-Neisseria properties warrant further design studies with dendritic amphiphiles to improve their safety indices to produce suitable candidates for antimicrobial vaginal products.

Synthesis, critical micelle concentrations, and antimycobacterial properties of homologous, dendritic amphiphiles. Probing intrinsic activity and the "cutoff" effect.

Newkome-type, 1-->3 C-branched dendrons make an excellent headgroup for amphiphiles with ultralong, saturated, linear alkyl chains. Synthesis of a homologous series of five such amphiphiles from 14 to 22 carbons-RNHCONHC(CH2CH2CO2H)3, R = n-CnH2n+1, n = 14, 16, 18, 20, 22-proceeds readily. These amphiphiles are soluble in aqueous solutions of triethanolamine. Surface-tension measurements on this homologous series reveal an unusually gradual decrease in log critical micelle concentration (CMC) as the chain length increases. In fact, the tetradecyl homologue does not appear to form micelles. Further, measurements of minimal inhibitory concentration (MIC) by broth microdilution against Mycobacterium smegmatis as a function of the initial cell density provide a direct measure of the intrinsic activity (MIC0) of each homologue. The hexadecyl homologue is the most active at inhibiting growth with an MIC0 equal to 3.5 x 10-5 M, which is 100-fold below the CMC.

Synchrotron SAX and WAX diffraction study of a hydrated very long-chain, dendritic amphiphile + DPPC mixture.

The tri-headed anionic dendritic amphiphile, 4-(2-carboxyethyl)-4-[(icosyloxycarbonyl)amino]heptanedioic acid (3CCb20), forms mixed aggregates with dipalmitoylphosphatidylcholine (DPPC) in excess water at 3CCb20:DPPC = 0.91:1 molar ratio. On heating, these mixed aggregates transform into fluid bilayers stacked in the liquid crystalline lamellar L(alpha) phase at about 40 degrees C. This phase transition and the microstructure of 3CCb20 + DPPC aggregates were studied with small- and wide-angle synchrotron X-ray diffraction. The ability of 3CCb20 to solubilize solidlike lipid bilayers could contribute to the antimicrobial activities of 3CCb20, including its anti-HIV activity.

Comparison of (+)- and (-)-hemipalmitoylcarnitinium as inhibitors of hepatic mitochondrial carnitine palmitoyltransferases in diabetic rats.

The syntheses of (R)- and (S)-norcarnitine ethyl esters are described starting with an optimized, chiral chemical reduction of ethyl 4-chloroacetoacetate followed by azide substitution, reduction, and dimethylation. The reaction of (R)- and (S)-norcarnitine ethyl esters with 1-bromoheptadecan-2-one gives (+)- and (-)-6-[(methoxycarbonyl)methyl]-2-pentadecyl-4,4-dimethylmorpholinium bromide, respectively, which hydrolyzes to (+)- and (-)-6-(carboxylatomethyl)-2-pentadecyl-4,4-dimethylmorpholinium (hemipalmitoylcarnitinium, (+)- and (-)-HPC), respectively, upon treatment with a hydroxide resin. (+)- and (-)-HPC are reversible active-site directed inhibitors of hepatic mitochondrial CPTs. Both stereoisomers inhibit CPT I and CPT II in control and streptozotocin diabetic rat to the same extent (Imax=100%). Using intact mitochondria (CPT I), I50values for (-)-HPC and (+)-HPC were 15.5 microM and 47.5 microM, respectively. The I50 values for CPT II were 6.7 microM and 38.5 microM for (-)-HPC and (+)-HPC, respectively. The mode of inhibition was uncompetitive for CPT I with respect to acyl-CoA. The apparent K(i) for (-)-HPC is about 5 microM. These data suggest that (-)-HPC may be useful for further evaluation as an antidiabetic agent.

Antibacterial activities of dendritic amphiphiles against nontuberculous mycobacteria.

The anti-mycobacterial activities of nine series of dicarboxyl and tricarboxyl dendritic amphiphiles with one alkyl, two alkyl, and cholestanyl tails against Mycobacterium abscessus, Mycobacterium avium, Mycobacterium chelonae, Mycobacterium marinum and Mycobacterium smegmatis have been measured. The dendritic amphiphiles overcame the limited aqueous solubility of natural long-chain fatty acids, alcohols, and amines to enable profiling the susceptibilities of the different mycobacterial species to the physicochemical properties of these amphiphiles. Several dendritic amphiphiles showed strong anti-mycobacterial activity with high critical micelle concentrations and low hemolytic activities thereby offering platforms for the development of antibiotics of higher activity against nontuberculous mycobacteria.

Antimicrobial activity of long-chain, water-soluble, dendritic tricarboxylato amphiphiles.

OBJECTIVES: To measure the antimicrobial activities of three series of homologous, dendritic tricarboxylato (three-headed) amphiphiles against a battery of bacteria and fungi. METHODS: Three series of homologous dendritic amphiphiles were synthesized containing C13 to C22 fatty chains. Susceptibility of Escherichia coli, Klebsiella pneumoniae, Lactobacillus plantarum, Micrococcus luteus, Staphylococcus aureus, methicillin-resistant S. aureus (MRSA), Mycobacterium smegmatis, Saccharomyces cerevisiae, Candida albicans, Cryptococcus neoformans and Aspergillus niger to the amphiphiles was measured by broth microdilution and reported as the MIC. RESULTS: Several amphiphiles from each homologous series, designed and constructed to overcome the low solubility of saturated long-chain fatty acids, had antimicrobial activity against MRSA (MIC = 36 mg/L), C. albicans (MIC = 4.4 mg/L), S. cerevisiae (MIC = 1.1 mg/L) and M. smegmatis (MIC = 8.9 mg/L). These amphiphiles had considerably better antimicrobial activities than the corresponding saturated fatty acids. Alkyl chain length influenced the values of MIC; longer chains (C18-C22) were generally more antimicrobial, but there was no uniform pattern among the microorganisms tested. CONCLUSIONS: As the antimicrobial activity of the amphiphiles increased with increasing chain length, it is anticipated that maximum activity was not reached with these series. Thus, the identification of the optimal chain length would provide a target compound for development of low-cost, topical microbicides and anti-infectives. Further, these series of dendritic amphiphiles with the very long chains can be used as new water-soluble probes for elucidation of membrane structure and for identification of novel targets for antimicrobial design.