Bacterial cell wall synthessis; lipopolysaccharides and phospholipids in the outer cell membrane of gram-negative bacteria
Cytoplasmic membrane of Gram-negative bacteria (via self-promoted uptake and membrane disruption) [1]

Colistins work like detergents to kill gram-negative bacteria. This mechanism includes competitively displacing divalent cations (calcium and magnesium) from the negatively charged phosphate groups of membrane lipids, as well as interaction with the lipopolysaccharides and phospholipids of the outer membrane through electrostatic interference[1]. Colistin, also known as polymyxin E, is a good treatment for infections brought on by gram-negative bacteria that are resistant to many drugs because of its quick bacterial killing, limited spectrum of action, and concomitant delayed development of resistance. Commercially, colistin comes in two forms: colistin methanesulfonate (sodium) for parenteral use and colistin (sulfate), primarily for topical use[2].

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Colistin sulfate (polymyxin E) and its methosulfate derivative colimycin are cationic lipopeptides that act on the cytoplasmic membrane of Gram-negative bacteria. Polymyxin B (structurally similar) has a net charge of +5 and exhibits Gram-negative selectivity. The proposed mechanism involves self-promoted uptake across the outer membrane followed by disruption of the cytoplasmic membrane barrier. [1]

Following intraperitoneal instillation, slow and sustained CMS conversion leads to high concentrations of colistin in rat ELF[3]. Piglets are frequently given colonistin, however both under- and overdosing are common. The fecal microbiota of piglets is not significantly disrupted by colistin overdoses or underdoses, and chromosomal resistance in the predominant E. coli population is seldom selected for[4].

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Colistin methanesulfonate (CMS), the inactive prodrug of colistin, is administered by inhalation for the management of respiratory infections. However, limited pharmacokinetic data are available for CMS and colistin following pulmonary delivery. This study investigates the pharmacokinetics of CMS and colistin following intravenous (i.v.) and intratracheal (i.t.) administration in rats and determines the targeting advantage after direct delivery into the lungs. In addition to plasma, bronchoalveolar lavage (BAL) fluid was collected to quantify drug concentrations in lung epithelial lining fluid (ELF). The resulting data were analyzed using a population modeling approach in S-ADAPT. A three-compartment model described the disposition of both compounds in plasma following i.v. administration. The estimated mean clearance from the central compartment was 0.122 liters/h for CMS and 0.0657 liters/h for colistin. Conversion of CMS to colistin from all three compartments was required to fit the plasma data. The fraction of the i.v. dose converted to colistin in the systemic circulation was 0.0255. Two BAL fluid compartments were required to reflect drug kinetics in the ELF after i.t. dosing. A slow conversion of CMS (mean conversion time [MCTCMS] = 3.48 h) in the lungs contributed to high and sustained concentrations of colistin in ELF. The fraction of the CMS dose converted to colistin in ELF (fm,ELF = 0.226) was higher than the corresponding fractional conversion in plasma after i.v. administration. In conclusion, pulmonary administration of CMS achieves high and sustained exposures of colistin in lungs for targeting respiratory infections[3].

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Colimycin (methosulfate derivative of colistin) has been successfully used in an aerosol formulation against Pseudomonas aeruginosa lung infections in cystic fibrosis patients. [1]

MIC determination.[2]
MICs were determined by both broth macrodilution and microdilution in cation-adjusted Mueller-Hinton broth ccording to NCCLS standards (16). Strains were considered resistant to colistin and colistin methanesulfonate if the MICs were ≥32 mg/liter.

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Time-kill kinetics.[2]
The time-kill kinetics of four strains, ATCC 27853 and three clinical isolates, two of which were mucoid, were examined. The clinical isolates were selected in order to have a range of MICs within the susceptible category. The MICs of colistin and colistin methanesulfonate, respectively, for the four strains were as follows: ATCC 27853, 4 and 16 mg/liter; 18982, 4 and 8 mg/liter; 19056, 1 and 8 mg/liter; and 20223, 4 and 16 mg/liter. Colistin and colistin methanesulfonate were added to a logarithmic-phase broth culture of approximately 106 CFU/ml to yield concentrations of 0, 0.5, 1, 2, 4, 8, 16, 32, and 64 times the MIC for the strain under study. Subcultures for viable counts were performed on nutrient agar at 0, 5, 10, 15, 20, 25, 30, 45, and 60 min and 2, 3, 4, and 24 h after antibiotic addition. Viable counts were determined after 24 h of incubation of subcultures at 37°C.

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PAE.[2]
The in vitro PAE was determined by the standard in vitro method for two of the three clinical strains noted above and the ATCC strain with both agents. For each experiment, P. aeruginosa (≈106 CFU/ml) in logarithmic phase growth was exposed for 15 min (for colistin) or 1 h (for colistin methanesulfonate) in Mueller-Hinton broth to the antibiotics at concentrations of 0.5, 1, 2, 4, 8, and 16 times the MIC. Fifteen minutes of exposure was used for colistin due to its very rapid bactericidal effect, to ensure that there were adequate numbers of bacteria for sampling at the end of the exposure interval. Antibiotic was removed by twice centrifuging at 3,000 × g for 10 min, decanting the supernatant, and resuspending in prewarmed broth. Viable counts were performed at 0, 1, 2, 3, 4, 5, 6, and 24 h on nutrient agar. A growth control was performed in the same fashion but without exposure to antibiotic. The colonies were counted after 24 h of incubation at 37°C. PAE was determined by comparing regrowth of treated and growth control cultures, using the standard formula of the time for the control culture to increase 10-fold subtracted from the time for the treated culture to do the same.

Rats: Sterile 0.9% sodium chloride is used to prepare fresh dose solutions of colistin methanesulfonate (sodium) and colistin sulfate. Colistin methanesulfonate (CMS) or sulfate solutions are injected bolus-xstyle through the jugular vein cannula for the intravenous studies. The method for pulmonary administration is called intratracheal (i.t.) instillation. CMS is given intravenously to animals at doses of 14 mg/kg, 28 mg/kg, or 56 mg/kg of body weight. Colistin is given intraperitoneally (i.v.) to rats in an independent study at doses of 0.21 mg/kg, 0.41 mg/kg, or 0.62 mg/kg[3].

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Inhalation therapy with colistin has been applied in cystic fibrosis patients with chronic P. aeruginosa lung infections. The aerosol formulation was well tolerated. [1]

Colistin did not exhibit significant genotoxic activity or carcinogenic structural warnings, was poorly absorbed in the gastrointestinal tract, and no tumors or precancerous lesions were observed in repeated oral or parenteral administration studies in rats over a period of 26 weeks. Therefore, the Committee concluded that colistin compounds are unlikely to be carcinogenic. The relevant endpoint for risk assessment was determined to be disruption of the colonic colonization barrier through toxicity to the gut microbiota, with Escherichia coli being the most susceptible. In vitro studies have shown that the minimum inhibitory concentration (MIC50) against 50% of E. coli strains is 1 µg/ml colistin base. Based on the MIC50 value of E. coli, the Committee converted the upper limit of ADI to 0–7 µg/kg body weight/day (420 µg/p/d for a 60 kg adult) using the following formula: ADI (µg/kg bw/d) = (MIC50 (1 µg/ml) × colonic contents mass (220 g)) / (bioavailability (0.5) × safety factor (1) × body weight (60 kg)). The committee recommends maximum residue limits (MRLs) of 150 µg/kg for colistin A+B in the liver, muscle, and fat (including applicable skin and fat) of cattle, sheep, goats, pigs, chickens, turkeys, and rabbits; 200 µg/kg for kidneys; 300 µg/kg for eggs; and 50 µg/kg for cow and sheep milk. The recommended MRLs result in a total daily intake (TMDI) of 229 µg (55% of the acceptable daily intake). The calculated acceptable daily intake (EDI) is equivalent to 4% (chicken) to 9% (cattle) of the upper limit of the daily intake. The acceptable daily intake of 56.9 µg (14% of the daily intake) was calculated using the highest median among tissues and food-producing species.

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Oral LD50 in rats: 121 mg/kg, Antibiotics Yearbook, 7(61), 1959/1960
Intraperitoneal LD50 in rats: 10572 ug/kg, Antibiotics Yearbook, 7(61), 1959/1960
Subcutaneous LD50 in rats: 72200 ug/kg Lung, pleural or respiratory system: dyspnea; Lung, pleural or respiratory system: cyanosis. Clinical Reports, 13(7), 1979.
Oral LD50 in mice: 793 mg/kg. Pharmaceutical Research, 11(395), 1961.
Intraperitoneal LD50 in mice: 21800 μg/kg. Pharmaceutical Research, 11(395), 1961.

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The systemic toxicity of natural lipopeptides such as colistin is a concern. Chemical modifications (e.g., mesylate derivatives) are intended to reduce systemic toxicity. The lipid tail is a source of some of the toxicity; even deacylated derivatives (polymyxin B nonapeptide) are considered to be too toxic for systemic human use. [1]

[1]. Peptide antibiotics. Antimicrob Agents Chemother. 1999 Jun;43(6):1317-23.

[2]. In vitro pharmacodynamic properties of colistin and colistin methanesulfonate againstPseudomonas aeruginosa isolates from patients with cystic fibrosis. Antimicrob Agents Chemother. 2001 Mar;45(3):781-5.

[3]. Population pharmacokinetics of colistin methanesulfonate in rats: achieving sustained lung concentrations of colistin for targeting respiratory infections. Antimicrob Agents Chemother. 2013 Oct;57(10):5087-95.

[4]. Impact of two different colistin dosing strategies on healthy piglet fecal microbiota. Res Vet Sci. 2016 Aug;107:152-60.

Colistin sulfate is a cyclic polypeptide antibiotic derived from Bacillus myxomyces. It consists of polymyxins E1 and E2 (or colistin A, B, and C), which act as cell membrane detergents. Colistin is less toxic than polymyxin B, but similar in other aspects; colistin mesylate is an oral formulation. This study investigated the in vitro pharmacodynamic characteristics of colistin and colistin mesylate by determining the minimum inhibitory concentration (MIC), bactericidal kinetics, and post-antibiotic effect (PAE) of colistin and colistin mesylate against Pseudomonas aeruginosa mucoid and non-mucoid strains isolated from cystic fibrosis patients. Twenty-three clinical isolates (including multidrug-resistant strains) and one standard strain were selected for MIC determination. Eleven of these strains were resistant; the MICs of these strains were all greater than 128 mg/L. For susceptible strains, the minimum inhibitory concentration (MIC) of colistin ranged from 1 to 4 mg/L, while that of colistin mesylate was significantly higher, ranging from 4 to 16 mg/L. This study investigated the time-kinetics of the drug using three strains at concentrations ranging from 0.5 to 64 times the MIC. Colistin exhibited extremely rapid bactericidal activity, completely eliminating bacteria within 5 minutes at the highest concentration; while colistin mesylate had a slower bactericidal rate, requiring a concentration 16 times the MIC to completely kill bacteria within 24 hours. At 16 times the MIC, colistin showed a significant post-exposure effect (PAE) against all three strains, lasting 2–3 hours after 15 minutes of exposure. Colistin mesylate had a shorter PAE within the tested concentration range. Even after adjusting for MIC differences, the overall bactericidal activity and PAE of colistin mesylate were lower than those of colistin. Our data suggest that for patients with cystic fibrosis, Pseudomonas aeruginosa infection may require higher doses of colistin mesylate (2 to 3 mg/kg body weight per 12 hours) to be effective. [2]

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Colistin is commonly used in piglets, but underdosing and overdosing are common. This study investigated the effects of such administration on the fecal microbiota. Piglets were treated with low-dose colistin by oral gavage for 5 days or with an overdose of colistin added to their feed for 14 days. The composition of the fecal microbiota was investigated using quantitative PCR, 16S rRNA sequencing, Enterobacteriaceae culture, and quantitative analysis of short-chain fatty acids (SCFAs). The mean concentrations of colistin in feces during the treatment period were 14.4 μg/g and 64.9 μg/g, respectively, in the low-dose and overdosing groups. Regardless of the piglet species or sampling date, the two major phyla were Firmicutes and Bacteroidetes, and the major families were Lactobacilliceae, Clostridiumlesales, Trichophyceae, and Ruminococciaceae. The main disturbance was a significant but transient decrease in the number of Escherichia coli during treatment; however, all isolated Escherichia coli were sensitive to colistin. In addition, colistin did not affect the production of short-chain fatty acids (SCFAs). These results suggest that neither insufficient nor excessive colistin administration leads to significant disruption of the fecal microbiota of piglets and rarely induces chromosomal resistance in the dominant Escherichia coli population. [4] Colistin sulfate is a non-ribosomally synthesized cationic lipopeptide antibiotic. It belongs to the polymyxin family and is clinically used to treat lung infections, especially in aerosol form. Its mechanism of action includes self-promoted absorption through the bacterial outer membrane and disruption of the cytoplasmic membrane. Due to toxicity issues, it is usually limited to local or aerosol administration rather than systemic administration. [1]

C52H98N16O13.2.5H2O4S

1400.64

1,266.73

C, 50.22; H, 8.11; N, 17.68; O, 21.46; S, 2.53

1264-72-8

Colistin;1066-17-7

White to off-white solid powder

1.3±0.1 g/cm3

1537.3±65.0 °C at 760 mmHg

200-220°C

883.5±34.3 °C

0.0±0.6 mmHg at 25°C

1.575

-3.68

CCC(C)CCCC(N[C@@H](CCN)C(N[C@H](C(N[C@H](C(N[C@@H](CCNC([C@H]([C@H](O)C)N1)=O)C(N[C@@H](CCN)C(N[C@H](CC(C)C)C(N[C@@H](CC(C)C)C(N[C@@H](CCN)C(N[C@@H](CCN)C1=O)=O)=O)=O)=O)=O)=O)CCN)=O)[C@H](O)C)=O)=O.O=S(O)(O)=O

ZJIWRHLZXQPFAD-FPSDIOKYSA-N

(R)-N-((S)-4-amino-1-(((2S,3R)-1-(((S)-4-amino-1-oxo-1-(((3S,6S,9S,12S,15R,18S,21S)-6,9,18-tris(2-aminoethyl)-3-((R)-1-hydroxyethyl)-12,15-diisobutyl-2,5,8,11,14,17,20-heptaoxo-1,4,7,10,13,16,19-heptaazacyclotricosan-21-yl)amino)butan-2-yl)amino)-3-hydroxy-1-oxobutan-2-yl)amino)-1-oxobutan-2-yl)-6-methyloctanamide sulfate

Belcomycin; Polymixin E; Colistin sulfate

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment, avoid exposure to moisture.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

H2O : ~50 mg/mL (~35.70 mM)

Solubility in Formulation 1: 50 mg/mL (35.70 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with sonication.

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 (Please use freshly prepared in vivo formulations for optimal results.)