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Related Experiment Videos

New high-performance liquid chromatographic method for amphotericin B analysis using an internal standard

M P Lambros1, S A Abbas, D W Bourne

  • 1University of Oklahoma Health Science Center, College of Pharmacy, Oklahoma City 73190, USA.

Journal of Chromatography. B, Biomedical Applications
|October 11, 1996
PubMed
Summary

A new, reliable High-Performance Liquid Chromatography (HPLC) method accurately measures amphotericin B (AmB) in biological samples. This assay is crucial for therapeutic drug monitoring and understanding AmB distribution in the body.

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Area of Science:

  • Analytical Chemistry
  • Pharmacology
  • Biochemistry

Background:

  • Amphotericin B (AmB) is a vital antifungal medication.
  • Accurate quantification of AmB in biological matrices is essential for effective treatment and safety.
  • Existing analytical methods may lack the required sensitivity, reproducibility, or applicability to various tissues.

Purpose of the Study:

  • To develop and validate a simple, reproducible High-Performance Liquid Chromatography (HPLC) method for quantifying amphotericin B (AmB) in serum, lung, and liver tissues.
  • To establish an assay suitable for therapeutic drug monitoring and pharmacokinetic studies.

Main Methods:

  • Developed an isocratic elution method using a C18 reversed-phase column with a mobile phase of acetonitrile-10 mM acetate buffer (pH 4.0) at a 37:63 (v/v) ratio.

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  • Utilized natamycin as an internal standard for extraction and analysis from serum, lung, and liver samples.
  • Employed a dual-detector HPLC system set at 303 nm for natamycin and 383 nm for AmB detection.

    • Main Results:

    • Achieved a limit of quantification of 0.1 microgram/ml with high recovery efficiency (96%-70%).
    • Demonstrated excellent reproducibility with within-day coefficients of variation < 8% and between-day variability < 7.7% for serum, lung, and liver.
    • Established assay linearity within the range of 1-40 micrograms/ml, with an R-squared value of 0.99.

    Conclusions:

    • The developed HPLC method is simple, reproducible, and accurate for the analysis of amphotericin B in serum, lung, and liver.
    • This validated assay provides a reliable tool for therapeutic drug monitoring of amphotericin B.
    • The method's robustness supports its application in pharmacokinetic and toxicological studies involving amphotericin B.