Determination of plasma testosterone using a simple liquid chromatographic method.

Area of Science:

• Analytical Chemistry
• Biochemistry
• Endocrinology

Background:

• Accurate quantification of testosterone in human plasma is crucial for diagnosing and monitoring various endocrine disorders.
• Existing methods may lack the sensitivity, specificity, or efficiency required for routine clinical application.

Purpose of the Study:

• To develop and validate a simple, sensitive, and reliable high-performance liquid chromatographic (HPLC) method for determining testosterone concentrations in human plasma.

Main Methods:

• Testosterone and griseofulvin (internal standard) were extracted from plasma using a dichloromethane-2,2,4-trimethylpentane mixture.
• Separation was performed on a C(18) column with a mobile phase of sodium dihydrogenphosphate-acetonitrile-methanol (pH 3.1).
• Ultraviolet detection at 240 nm was employed for quantification.

Main Results:

• The method demonstrated a quantification limit of 1.6 ng/ml with high recoveries (>92% for testosterone, ~95% for griseofulvin).
• Excellent precision (RSD < 8%) and accuracy (96.0–106.0%) were achieved across the linear range (1.6–400 ng/ml).
• The method was successfully applied to analyze plasma testosterone levels in healthy male volunteers, yielding results within the normal range (3.1–8.4 ng/ml).

Conclusions:

• The developed HPLC method is simple, sensitive, and accurate for testosterone determination in human plasma.
• This validated method is suitable for routine analysis and clinical research applications.
• The assay provides reliable quantification of plasma testosterone, aiding in the assessment of hormonal status.