Development and validation of a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to quantify...

Protoporphyrin IX (PPIX) is a photoreactive heme precursor and is a key pathogenic driver in erythropoietic protoporphyria (EPP) and X-linked protoporphyria (XLP). In both EPP and XLP, excess PPIX in plasma circulates to the skin and hepatobiliary system, resulting in acute, painful cutaneous photosensitivity and in some patients, gallstones, or hepatic failure. A key distinguishing feature between EPP and XLP is proportions of PPIX that become metalated with zinc (Zn-PPIX) or remain metal-free. Thus, ability to accurately measure both PPIX and Zn-PPIX in plasma is especially useful for assessing emerging therapies for EPP and XLP. This manuscript presents a fully validated, sensitive liquid chromatography-tandem mass spectrometry method for the quantification of PPIX and Zn-PPIX across a dynamic range in human plasma, which is especially suited for measuring drug-induced effects on plasma PPIX and Zn-PPIX levels. It also presents the first reported accurate measurements of plasma PPIX in healthy individuals and begins to establish a preliminary baseline range for this major form of PPIX in plasma of individuals without protoporphyria, which can facilitate future development of therapies for EPP and XLP.