Journal
MOLECULES
Volume 26, Issue 3, Pages -Publisher
MDPI
DOI: 10.3390/molecules26030665
Keywords
pharmaco-toxicological investigation; sample derivatization; screening of extraction conditions
Funding
- Cassa di Risparmio di Citta di Castello Foundation, Citta di Castello, Italy
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This study described an optimized method for extracting AMK from murine whole blood, achieving nearly quantitative (95%) extraction of the drug by using RP-HPLC-UV detection and derivatization with 1-fluoro-2,4-dinitrobenzene. The extraction yield was significantly improved by the addition of 5% ammonium hydroxide in a hydro-organic solution with acetonitrile.
Amikacin (Amk) analysis and quantitation, for pharmacokinetics studies and other types of investigations, is conventionally performed after extraction from plasma. No report exists so far regarding drug extraction from whole blood (WB). This can represent an issue since quantification in plasma does not account for drug partitioning to the blood cell compartment, significantly underrating the drug fraction reaching the blood circulation. In the present work, the optimization of an extraction method of Amk from murine WB has been described. The extraction yield was measured by RP-HPLC-UV after derivatization with 1-fluoro-2,4-dinitrobenzene, which produced an appreciably stable derivative with a favorable UV/vis absorption. Several extraction conditions were tested: spiked Amk disulfate solution/acetonitrile/WB ratio; presence of organic acids and/or ammonium hydroxide and/or ammonium acetate in the extraction mixture; re-dissolution of the supernatant in water after a drying process under vacuum; treatment of the supernatant with a solution of inorganic salts. The use of 5% (by volume) of ammonium hydroxide in a hydro-organic solution with acetonitrile, allowed the almost quantitative (95%) extraction of the drug from WB.
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