Rapid and sensitive assay for fluconazole which uses gas chromatography with electron capture detection

Abstract

Fluconazole, an orally active antifungal agent, has been shown to be clinically beneficial for maintenance therapy of cryptococcal meningitis. A sensitive gas-liquid chromatographic assay with electron capture detection, which required only a single extraction step and precluded any pretreatment of the chromatographic column, was developed for fluconazole. The assay was linear from 0.1 to 20 p,g/ml, with a correlation coefficient of 0.999. The intraassay and interassay coefficients of variation were less than 9%o. The measured values on average were within 8% of the target values. The extraction recoveries ranged from 87 to 106%. Steady-state plasma fluconazole levels (mean + standard deviation) in three AIDS patients with cryptococcal meningitis receiving 200 mg of fluconazole per day ranged from 8.95 1.32 to 11.41 + 0.63 ,ug/ml and were within the expected range for this dosing rate, on the basis of previous studies. The ratio of fluconazole concentration in cerebrospinal fluid to fluconazole concentration in plasma in one patient receiving 400 mg/day was 0.73 at steady state and was consistent with published reports.Fluconazole is an orally active, triazole antifungal agent recently approved by the Food and Drug Administration for treatment of candidiasis and cryptococcal meningitis in patients with AIDS. Advantages of fluconazole are its water solubility, low level of protein binding, high bioavailability, long half-life, metabolic stability (7), and high cerebrospinal fluid (CSF) penetration (1, 3, 11). It can be given orally or intravenously and is usually well tolerated, with no major adverse reactions (2, 9, 10). Peak serum levels with chronically administered dosages of 50 and 400 mg/day typically result in fluconazole concentrations of 2 and 14 ,ug/ml, respectively (4). Maintenance therapy with an oral fluconazole dose of 100 mg (2) or 200 mg (2, 4) once daily has been shown to be highly effective in preventing recurrent cryptococcal infections.Previously published assays for fluconazole include bioassay (11), high-pressure liquid chromatography (3, 6), gas chromatography with electron capture (GC-EC) detection (12), and GC with nitrogen selective detector (5,8). GC-EC detection is a sensitive method; however, the published method requires three extraction steps and inactivation of the GC column to obtain good chromatographic resolution and better peak shapes. We report here a GC-EC detection method that requires only a single extraction step and no preinactivation treatment of the column and that results in excellent chromatographic resolution with sharp and symmetrical peaks.MATERUILS AND METHODS Materials and reagents. Fluconazole was obtained from Pfizer Central Research, Groton, Conn. The internal standard for this assay (4,4'-dibromobiphenyl) was purchased from Aldrich Chemical Co., Milwaukee, Wis. The structures of these compounds are shown in Fig. 1. Analytical-grade methanol, ethyl acetate, sodium hydroxide, and sodium chloride were purchased from Fisher Scientific, Pittsburgh, * Corresponding...

Novel Spectrophotometric Methods for the Determination of Fluconazole in the Presence of Its Oxidative Degradation Product

“…This can be summarized as the following: The literature survey reveals several analytical methods used for quantitative determination of FLZ in body fluids and pharmaceutical preparations. These methods include high-performance liquid chromatography [2][3][4][5][6][7][8] , liquid chromatography-tandem mass spectrometry [9][10][11] , gas chromatography [12][13][14][15] ,micellar electrokinetic capillary chromatography 16 and bioassay 17 .…”

Quantitative Determination of Fluconazole in Syrups by First Order Derivative Spectrophotometry

“…There are various analytical studies reported in literature for the determination of fluconazole in biological fluids by GC [1][2][3][4][5][6] GC-MS, [7] HPLC, [8][9][10][11][12][13][14][15][16][17][18] SFC [19] and by microbiological bioassays. There are various analytical studies reported in literature for the determination of fluconazole in biological fluids by GC [1][2][3][4][5][6] GC-MS, [7] HPLC, [8][9][10][11][12][13][14][15][16][17][18] SFC [19] and by microbiological bioassays.…”

Quantitative Determination of Fluconazole by Ultra-Performance Liquid Chromatography Tandem Mass Spectrometry (UPLC-MS/MS) in Human Plasma and its Application to a Pharmacokinetic Study

“…This potentially exposes individuals to organic solvents or requires additional washing time to clean the pump and column as a result of the use of buffers. Gas chromatography/mass spectrometry, liquid chromatography/mass spectrometry methods that are available are sen sitive, but the sample preparation is sometimes laborious [7][8][9][10]. There is no simple, specific, sensitive, and reproducible analytical method available to quantify FLA in human plasma.…”