Development of a mass spectrometry method for the determination of a melanoma biomarker, 5-S-cysteinyldopa, in human plasma using solid phase extraction for sample clean-up

Abstract

5-S-cysteinyldopa is a well-known pigment intermediate and analysis of its plasma concentration is interesting for the early diagnosis, as well as for evaluation of treatment and follow-up of malignant melanoma. A determination method of 5-SCD in human plasma was developed using solid phase extraction (SPE) on disposable cartridges and liquid chromatography electrospray mass spectrometry (LC-ESI-MS-MS). Compound's sensitivity to light and oxidation requires the addition of anti-oxidative agents (AO), to work in acidic media at 4• C and to avoid light exposure of samples since blood collection. Different solid phases involving covalent binding to phenylboronic groups or dual retention mechanisms were evaluated and extraction cartridges containing both hydrophobic and strong cation exchange functionalities were finally selected. The LC separation of 5-SCD from endogenous catecholamines was achieved by gradient elution on a C18 stationary phase. 5-SCD was detected by multiple reaction monitoring (MRM) performed on ES(+) generated ions. Finally, the method was prevalidated in the lower ng/ml range. Good results with respect to accuracy, trueness and precision were obtained.

MFSD12 mediates the import of cysteine into melanosomes and lysosomes

“…The cysteinyldopa extraction protocol was adapted from previously described methods for measuring cysteinyldopas in serum 37 . Cells were plated under the same conditions used for the MelanoIP protocols.…”

Proteomics: An Indispensable Tool for Novel Biomarker Identification in Melanoma

“…In contrast, a targeted mode enables to identify peptide of interest in clinical samples. A version of targeted MS was specifically developed to detect ng/ml range of plasma concentrations of pigment intermediate 5-S-cysteinyldopain which is generally utilized for early diagnosis, evaluation of treatment as well as malignant melanoma progression [25]. MS is a great tool to analyze complex protein samples.…”

Selective Solid-Phase Extraction of Catecholamines and Metanephrines From Serum Using a New Molecularly Imprinted Polymer

“…A similar approach was attempted by Martin et al through an SPE cartridge containing silica-bonded phenylboronic groups. [18] Although high recoveries were obtained by the boronic complexation of the analytes, these catecholamine selective extractions suffer sometimes from poor repeatability. Moreover, boronic complexes are unstable in the case of O-methylated catechols (metanephrines).…”

“…Another approach targeted amino group of neurotransmitter molecules using either weak cation-exchange [11] or mixed-mode strong cationexchange [18] SPE supports. Recoveries higher than 90% were reported for most of the metanephrines although this method was not selective for neurotransmitters; indeed, other cationic molecules (drugs) were extracted.…”