Population pharmacokinetics of amphotericin B lipid complex in neonates.

The pharmacokinetics of amphotericin B lipid complex (ABLC) were investigated in neonates with invasive candidiasis enrolled in a phase II multicenter trial. Sparse blood (153 samples; 1 to 9 per patient, 1 to 254 h after the dose) and random urine and cerebrospinal fluid (CSF) samples of 28 neonates (median weight [WT], 1.06 kg; range, 0.48 to 4.9 kg; median gestational age, 27 weeks; range, 24 to 41 weeks) were analyzed. Patients received intravenous ABLC at 2.5 (n = 15) or 5 (n = 13) mg/kg of body weight once a day over 1 or 2 h, respectively, for a median of 21 days (range, 4 to 47 days). Concentrations of amphotericin B were quantified as total drug by high-performance liquid chromatography. Blood data for time after dose (TAD) of <24 h fitted best to a one-compartment model with an additive-error model for residual variability, WT0.75 (where 0.75 is an exponent) as a covariate of clearance (CL), and WT as a covariate of volume of distribution (V). Prior amphotericin B, postnatal age, and gestational age did not further improve the model. The final model equations were CL (liters/h) = 0.399 x WT(0.75) (interindividual variability, 35%) and V (liters) = 10.5 x WT (interindividual variability, 43%). Noncompartmental analysis of pooled data with a TAD of >24 h revealed a terminal half-life of 395 h. Mean concentrations in the urine after 1, 2, and 3 weeks ranged from 0.082 to 0.430 microg/ml, and those in CSF ranged from undetectable to 0.074 microg/ml. The disposition of ABLC in neonates was similar to that observed in other age groups: weight was the only factor that influenced clearance. Based on these results and previously published safety and efficacy data, we recommend a daily dosage between 2.5 and 5.0 mg/kg for treatment of invasive Candida infections in neonates.

Efficacy of a single intravenous dose of amphotericin B for Candida urinary tract infections: further favorable experience.

Studies in experimental animals and humans have shown that Amphotericin B (AmB) persists in urine for days to weeks after a single IV dose in levels that should inhibit candidal organisms and thereby obviate the need for frequent dosing. Including data from four previously described patients, we have now treated a total of 11 patients (12 episodes) with Candida urinary tract infections with single-dose AmB (six, Candida albicans; two, C. tropicalis; four, other nonalbicans Candida). The duration of candiduria prior to entry ranged from 18 to 180 days. Predisposing conditions included renal transplantation (1), diabetes mellitus (8), genitourinary stones (1) or anomalies (4), catheterization (2), and antibacterial therapy (11). A single patient was intolerant of AmB. Out of 11 evaluable candiduric episodes, eight resolved. Failure occurred in one patient with a chronic indwelling bladder catheter and in the allograft recipient. The data suggest that the sustained urinary excretion of AmB may permit successful single- or paucidose therapy of Candida urinary tract infections in some patients with a minimum of toxicity.

Pharmacokinetics and tissue distribution of a new partricin A derivative (N-dimethylaminoacetyl-partricin A 2-dimethylaminoethylamide diaspartate) in mice.

A single-dose trial in mice (1.25 mg/kg SPA-S-753 or 1 mg/kg amphotericin B [AmB] by intravenous route) was performed to study the pharmacokinetics, tissue distribution, and urinary excretion of a new polyene, SPA-S-753 (N-dimethylaminoacetyl-partricin A 2-dimethylaminoethylamide diaspartate), in comparison with AmB. Antibiotic concentrations were determined by microbiological assay (agar diffusion method). The elimination half-lives in serum were 15.1 and 19.8 h, respectively, for SPA-S-753 and AmB; the area under the curve from 0 to infinity values were 49.3 for SPA-S-753 and 23.6 microg. h/mL for AmB, because of the higher serum levels of SPA-S-753 found just after administration. The tissue concentrations of SPA-S-753 were lower than those of AmB in liver and lungs but higher in the kidneys. The urine concentrations of SPA-S-753 and the percent of the administered dose recovered from the urine were quite low in mice, whereas those of AmB were higher.

Pharmacokinetics, excretion, and mass balance of 14C after administration of 14C-cholesterol-labeled AmBisome to healthy volunteers.

Amphotericin B (AmB) in small unilamellar liposomes (AmBisome) provides higher plasma concentrations and greater safety than the conventional deoxycholate formulation. The authors compared the disposition of the liposome's drug and cholesterol components by measuring AmB and radioactivity in plasma, urine, and feces for 1 week after a single 2-hour infusion of 14C-cholesterol-labeled AmBisome (2 mg/kg, 1 microgCi/kg) in healthy adults (4 males, 1 female). The plasma profile of 14C-cholesterol differed from that of AmB, lacking an initial rapid disappearance phase, having a lower total clearance, and having a volume of distribution (0.13 L/kg) close to that of the plasma compartment. The biphasic disappearance and long plasma half-life (147 h) of 14C-cholesterol were similar to those of other low-clearance liposomes. This and the low clearance of 14C-cholesterol from the plasma compartment suggest that it served as a liposome marker. The plasma drug-lipid ratio fell during the study, showing that AmB was cleared from plasma more rapidly than cholesterol or liposomes and suggesting that the composition of the liposomes changed over time. 14C-radioactivity was recovered mainly in the feces (9.5% of dose), consistent with the catabolism of cholesterol to bile salts. Combined fecal and renal clearances were < 18% of total clearance, suggesting that most of the liposomal drug and lipid remained in the body 1 week after dosing. Thus, AmBisome remains in the circulation for an extended period of time while releasing AmB, resulting in its markedly altered pharmacokinetic and safety profiles.

Rapid determination of amphotericin B in serum and urine by third-order derivative spectrophotometry.

A derivative spectrophotometric method for rapid monitoring of amphotericin B in serum and urine down to 30 ng/mliters is described. Samples are treated with acetonitrile, and amphotericin B is directly quantified in the crude extracts on the basis of the intensity of the peak that appears at 402 nm when the normal absorption spectrum is submitted to third-order derivative processing. Accuracy data suggested recoveries in the range of 84.3-94.9% for serum and 85.6-93.4% for urine. The precision of the method was better than 11.3% for serum and 9.2% for urine when samples contained as low as 29.6 ng/mliters of amphotericin B. Ease of applicability, short analysis time, low cost, and reliability are the main advantages of the method.

High-performance liquid chromatographic analysis of amphotericin B in plasma, blood, urine and tissues for pharmacokinetic and tissue distribution studies.

A sensitive and reproducible high-performance liquid chromatographic method was developed to assay ampherotericin B in plasma, blood, urine and various tissue samples. Amphotericin B was isolated from each sample matrix by solid-phase extraction (Bond-Elut). The eluate from Bond-Elut containing amphotericin B was injected onto a reversed-phase C18 column (Waters, mu Bondpak, 10 microns, 300 mm x 3.9 mm I.D.) with a mobile phase of 45% acetonitrile in 2.5 mM Na2EDTA at 1 ml/min. Detection of amphotericin B was by ultraviolet absorption at 382 nm. Blood and tissues were homogenized and extracted with methanol prior to Bond-Elut extraction. The extraction efficiencies of amphotericin B from plasma, blood and tissues were approximately 90, 70 and 75%, respectively. The sensitivity of the assay was less than or equal to 5 ng/ml for plasma, less than or equal to 25 ng/ml for blood, 2.5 ng/ml for urine and 50 ng/g for tissues. The linearity of the assay method was up to 2.5 micrograms/ml for plasma, 5 micrograms/ml for blood, 500 ng/ml for urine and 500 micrograms/g for tissues. The assay was reproducible with an intra-day coefficient of variation (C.V., n = 3) of less than 5% in general for plasma, blood and tissues. The inter-day C.V. of the assay was less than 5% for plasma (n = 5), less than 10% for blood (n = 4) and less than 5% for tissues (n = 3). The overall variability in the urine assay was generally less than 10%. This method has demonstrated significant improvement in the sensitivity and reproducibility in assaying amphotericin B in plasma and especially in blood, urine and tissues. We have employed this assay to compare the pharmacokinetic and tissue distribution profiles of amphotericin B in rats and dogs following administration of Fungizone and ABCD (amphotericin B-cholesteryl sulfate colloidal dispersion), a lipid-based dosage form. In addition, the assay method for plasma and urine samples can also be applied to pharmacokinetics studies of amphotericin B in man.

Fungicidal effect of amphotericin B in urine: in vitro study to assess feasibility of bladder washout for localization of site of candiduria.

An in vitro study was performed to determine the optimum amphotericin B concentration and exposure time required to kill various strains of Candida albicans in urine. This is a preliminary study to assess the feasibility of using amphotericin B bladder washout for localization of the site of candiduria. In broth kinetic killing studies, amphotericin B at a concentration of greater than 100 micrograms/ml produced almost complete killing of 5 x 10(5) CFU of C. albicans per ml within 1.5 to 2 h. In urine studies (with various pH values, osmolalities, and electrolyte concentrations), amphotericin B at a concentration of 200 micrograms/ml with a 2-h exposure time decreased fungal counts of 21 strains of C. albicans from 5 x 10(6) to less than 200 CFU/ml. Bladder washout with greater than or equal to 200 micrograms of amphotericin B per ml and a dwell time of 2 h can therefore sufficiently sterilize the bladder of yeasts and may be a useful localization test.

High-pressure liquid chromatographic method for determination of Sch 28191 in biological fluids.

A high-pressure liquid chromatographic method has been developed for the measurement of N-D-ornithyl amphotericin B methyl ester (Sch 28191) in biological fluid. The method involves protein precipitation with methanol, followed by separation of the supernatant on a reverse-phase column and quantitation by absorbance at 405 nm. This technique resulted in a recovery of 97%. There was a good linear relationship between the peak height ratio and Sch 28191 concentrations ranging from 0.015 to 20 micrograms/ml. In addition, this method was specific for Sch 28191 since all of its analogs tested did not interfere with the assay. The method was reproducible with a lower limit of quantitation of 0.015 microgram/ml. Serum levels obtained from this method were in good agreement with those obtained from a microbiological assay only when drug concentrations were higher than 1.5 microgram/ml. The high-pressure liquid chromatographic method is useful in monitoring serum and urine drug levels in animals and should prove to be useful for pharmacokinetic studies of the drug with therapeutic doses in humans.

Elimination of amphotericin B in impaired renal function.

The influence of impaired renal function on the steady-state plasma clearance of amphotericin B was determined in seven patients with creatinine clearances ranging from zero to normal. Contrary to previous reports, steady-state plasma concentrations of total drug were lower in uremic patients than in patients with normal renal function. Total plasma clearance of amphotericin B ranged from 16.7 to 39.9 ml/min, correlated directly with the plasma creatinine concentration, and correlated inversely with the creatinine clearance. Urinary excretion of unchanged drug accounted for less than 10% of the dose. In 10 healthy subjects, mean percent of amphotericin B unbound in plasma was 3.55 +/- 0.32 (SD). Binding was determined in a further group of 10 uremic patients. Mean unbound percent (4.15 +/- 0.73, SD) was higher than in the healthy subjects, and the binding ratio (molar concentration of bound to unbound drug) correlated weakly with the creatinine clearance. This suggests that plasma clearance of unbound amphotericin B and, therefore, steady-state plasma concentrations of unbound drug are not affected by renal impairment, and that dosage requirements will be overestimated if based on measurements of total drug plasma concentration.

Excretion pathways of amphotericin B.

The role of the biliary system in excretion of amphotericin B was explored in a dog model that allowed either external diversion of all bile or complete biliary obstruction. In dogs with biliary diversion, which were given a single dose of amphotericin B intravenously, excretion of amphotericin B in the bile lasted for seven to 10 days and accounted for only 3% +/- 2% (mean +/- SD) of the dose, whereas excretion in the urine was prolonged (23--35 days) and greater (21% +/- 5% of the dose); the stool contained no amphotericin B. However, bile salt depletion may have depressed biliary excretion of amphotericin B: in a dog with an intact biliary system, 19% of the dose was excreted in the stool over 11 days. In dogs given amphotericin B daily, serum levels were 19% +/- 3% higher during periods of biliary obstruction than during periods of free bile flow (P less than 0.05). Thus, excretion of amphotericin B in the bile (less than or equal to 19% of the dose) and in the urine (21% of the dose) accounted for a minority of total drug clearance. Nevertheless, prolonged excretion of amphotericin B by these routes after a single dose suggests that infrequent doses of amphotericin B may provide effective treatment for certain forms of fungal infection.

Quantitation of amphotericin B with use of high-pressure liquid chromatography.

A chemical method for determination of concentrations of amphotericin B in serum and cerebrospinal fluid (CSF) is described. After extraction with methanol, the antibiotic was separated by reverse-phase, high-pressure liquid chromatography and quantitated by absorption at 405 nm. The lower limit of detection of this assay was 0.02 microng/ml. Relative standard deviations of less than 3.6% were noted for multiple determinations of sera containing 0.20 and 1.00 microng of amphotericin B/ml. No interfering peaks were found in extracts of serum or CSF from normal humans or in extracts of serum from patients treated with other drugs and antimicrobial agents, including 5-fluorocytosine. Comparison of the method with microbiological assays showed correlation coefficients of 0.90 and 0.83 for serum and CSF determinations, respectively. This chemical assay is very rapid (less than 30 min), sensitive, accurate, and specific and appears to be suitable for routine clinical use.

Bioassay for hamycin and amphotericin B in serum and other biological fluids.

A bioassay suitable for measuring concentrations of the polyene antifungal agents hamycin and amphotericin B in biological fluids is described. By using Paecilomyces varioti as the indicator organism, sensitivity of the bioassay was found to be in the range of 0.01 to 0.02 mug/ml. A linear dose-response curve was obtained with amphotericin B; the curve for hamycin was curvilinear. In a series of assays, hamycin serum levels in the range of 0.01 to 3.5 mug/ml were measured; with amphotericin B, serum levels in the range of 0.015 to 0.175 mug/ml were measured in patients receiving orthodox intravenous medication and as high as 9.0 mug/ml in one patient treated with extraordinarily high doses of the drug.