Copper (Cu) is an essential trace element associated with various physiological processes in the human body. In humans, more than 90% of Cu in serum is tightly bound to ceruloplasmin (Cp). The remaining Cu (exchangeable Cu) is loosely bound to human serum albumin (HSA) and, to a lesser extent, to alpha-2-macroglobulin (α2M) and low molecular mass (LMM) amino acids.

Elevated concentrations of Cu and Cp have been detected in the serum of patients with various cancers, including breast, prostate, lung, bile duct, bladder, colorectal, thyroid, and others.

In this study, we quantified different Cu species in serum samples of patients with hepatobiliary carcinoma collected before chemotherapy and after the last round of chemotherapy in a group of cancer patients and in the control group of healthy individuals. We performed Cu speciation using conjoint liquid chromatography (CLC) on convective interaction media (CIM) monolithic disks coupled with inductively coupled plasma mass spectrometry (ICP-MS). The CLC monolithic column consisted of two immunoaffinity (antiHSA) disks and one anion-exchange diethylaminoethyl (DEAE) disk. This setup enabled the separation of Cu bound to Cp, human serum albumin (HSA), and low molecular mass species (LMM). Separated Cu species were quantified using post-column isotope dilution inductively coupled plasma mass spectrometry (ID-ICP-MS).