Parent exposure or metabolite behavior?Should the method focus on parent antifungal levels, or also evaluate oxidative metabolites, CYP-related products, and parent–metabolite relationships?
Is tissue distribution needed?Does the research model require tissue homogenates, skin-related matrices, lung-associated samples, or infection-relevant compartments?
Are formulation effects important?Could lipid-associated delivery, amphipathic structure, or formulation behavior affect exposure, recovery, or matrix compatibility?
Is CYP behavior part of the study?Do azole antifungals require CYP-related metabolism support, interaction-sensitive exposure comparison, or metabolite profiling?
Does the analyte challenge the matrix?Could protein binding, low solubility, adsorption, or matrix-dependent recovery affect extraction strategy, calibration, or reproducibility?
Which fungal readouts matter?Should exposure be paired with ergosterol biosynthesis indicators, β-glucan markers, membrane integrity, fungal burden, or host-response markers?