Brain Penetration of Colistin in Mice Assessed by a Novel High-Performance Liquid Chromatographic Technique

Abstract

A sensitive and reliable liquid chromatographic method was developed and validated for the determination of colistin concentrations in mouse brain homogenate. With a mobile phase consisting of acetonitrile-tetrahydrofuran-water (50:25:25 [vol/vol]) at a flow rate of 1 ml/min, a linear correlation between peak area and colistin concentration was observed over the concentration range of 93.8 to 3,000 ng/g in brain tissue (R 2 > 0.994). Intra-and interday coefficients of variation were 5.1 to 8.3% and 5.8 to 8.5%, respectively, and the recovery ranged from 85% to 94%. This assay was then utilized to determine the amount of colistin that permeated the blood-brain barrier over a 2-h period following bolus intravenous administration of colistin sulfate to mice. After a single dose of 5 mg/kg of body weight to mice, brain homogenate concentrations of colistin were very low, relative to plasma colistin concentrations, suggesting that colistin permeability across the healthy blood-brain barrier is negligible during this experimental period.Colistin (also known as polymyxin E) is a cationic polypeptide antibiotic and is bactericidal against multidrug-resistant gram-negative pathogens (8). Like other members of the polymyxin family, colistin consists of two major components (colistins A and B), with colistins A and B differing only in the fatty acid side chain. In studies conducted several decades ago, colistin was shown to result in adverse effects after intramuscular or intravenous administration to patients (6, 14); nephrotoxicity and neurotoxicity were the most commonly observed adverse effects. It is possible that the prevalence of such toxicity may have been exaggerated due to a lack of understanding of colistin pharmacology and the use of inappropriate doses (22). Over the last few years, however, resistance to many other antibiotics and limited development of new antibiotics have resulted in an increasing use of colistin, administered in the form of its inactive prodrug, colistin methanesulfonate (CMS) (1), for the treatment of infections caused by gram-negative pathogens, such as Pseudomonas aeruginosa and Acinetobacter baumannii (5). With its increased use, interest in the pharmacology of this old antibiotic has been rekindled and optimization of dosage regimens is therefore urgently required in order to maximize its efficacy while minimizing potential toxicity.Gram-negative bacterial species are increasingly reported to cause severe central nervous system (CNS) infections, such as meningitis (11,24,26) and ventriculitis (7, 30), which have a high mortality rate if untreated or treated inappropriately. There have been several clinical case reports of successful treatment of meningitis and ventriculitis by parenteral administration of CMS, either alone or in combination with other antibacterials (11,12,18), resulting in eradication of the gramnegative pathogens from the cerebrospinal fluid (CSF). These studies suggest that CMS, or colistin that is generated from CMS in vivo, has the potential to permeate the blood...

Impact of P-Glycoprotein Inhibition and Lipopolysaccharide Administration on Blood-Brain Barrier Transport of Colistin in Mice

“…Many of these substrates share some common physicochemical properties such as a positive charge at physiological pH and a molecular mass of Ͼ400 Da (7). Given that the primary amines of colistin possess a pK a value of approximately 10 and that colistin has a high molecular weight (19), it is plausible that the low brain uptake of colistin observed in our previous study (16) may also be due, in part, to P-gpmediated efflux.…”

“…Two hours after the last dose, plasma and brain samples were collected and stored at Ϫ20°C until analysis. Concentrations of colistin in brain homogenate and in plasma samples were measured using HPLC (16), and a brain-to-plasma concentration (B/P) ratio of colistin was calculated at each postdose time point.…”

“…It has been clinically demonstrated that intravenous administration of colistin methanesulfonate, an inactive prodrug of colistin (5), leads to detectable levels of colistin in cerebrospinal fluid (CSF) (1,15,21); however, this is an indicator of blood-CSF barrier penetration and not BBB penetration. We have previously demonstrated that the brain uptake of colistin following a single intravenous dose to mice was negligible (16), suggesting minimal penetration across an intact BBB. However, the mechanisms governing the low BBB penetration of colistin remain unknown, and, furthermore, it is unclear as to whether brain accumulation of colistin occurs with repeated doses (as is used clinically), given that our previous study assessed colistin brain penetration after only a single intravenous dose (16).…”

“…We have previously demonstrated that the brain uptake of colistin following a single intravenous dose to mice was negligible (16), suggesting minimal penetration across an intact BBB. However, the mechanisms governing the low BBB penetration of colistin remain unknown, and, furthermore, it is unclear as to whether brain accumulation of colistin occurs with repeated doses (as is used clinically), given that our previous study assessed colistin brain penetration after only a single intravenous dose (16).…”

Melatonin Attenuates Colistin-Induced Nephrotoxicity in Rats

“…The concentrations of colistin in plasma, urine, and kidneys were determined by high-pressure liquid chromatography (HPLC) (18,24). Minor modifications for pretreatment of kidney samples involved homogenization (Kinematica polytron PT-DA 3007/2EC homogenizer; Kinematica, Luzernerstrasse, Switzerland) in 5 ml of 20 mM ice-cold HEPES buffer (pH 7.2) per g of tissue, followed by ultrasonication for 10 s (Sonics Vibracell VCX500).…”