Addressing the need for biomarker liquid chromatography/mass spectrometry assays: A protocol for effective method development for the bioanalysis of endogenous compounds in cerebrospinal fluid

Abstract

Route 206 and Province Line Road, Lawrenceville, NJ 08543, USA RATIONALE: Research on disorders of the central nervous system (CNS) has shown that an imbalance in the levels of specific endogenous neurotransmitters may underlie certain CNS diseases. These alterations in neurotransmitter levels may provide insight into pathophysiology, but can also serve as disease and pharmacodynamic biomarkers. To measure these potential biomarkers in vivo, the relevant sample matrix is cerebrospinal fluid (CSF), which is in equilibrium with the brain's interstitial fluid and circulates through the ventricular system of the brain and spinal cord. Accurate analysis of these potential biomarkers can be challenging due to low CSF sample volume, low analyte levels, and potential interferences from other endogenous compounds. METHODS: A protocol has been established for effective method development of bioanalytical assays for endogenous compounds in CSF. Database searches and standard-addition experiments are employed to qualify sample preparation and specificity of the detection thus evaluating accuracy and precision. RESULTS: This protocol was applied to the study of the histaminergic neurotransmitter system and the analysis of histamine and its metabolite 1-methylhistamine in rat CSF. CONCLUSIONS: The protocol resulted in a specific and sensitive novel method utilizing pre-column derivatization ultra high performance liquid chromatography/tandem mass spectrometry (UHPLC/MS/MS), which is also capable of separating an endogenous interfering compound, identified as taurine, from the analytes of interest.

Bioanalysis of (1R,4R,5S,6R)-4-Amino-2-Oxabicyclo[3.1.0]Hexane-4,6-Dicarboxylic Acid (Ly379268) in Rat Plasma Using Derivatization Liquid Chromatography/Mass Spectrometry

“…Employing a generic HILIC method resulted in a MS response of 20,000 AU intensity; however, peak splitting was observed ( Figure 2B). In order to improve both the chromatographic behavior and MS detection limits in LC/MS-based bioanalysis, chemical derivatization of analytes has been widely used (i.e., d-LC/MS) [7,[9][10][11][12]. The d-LC/MS analysis of LY379268 (2500 nM, derivatized with PFB-Cl) showed a dramatically increased MS response of 600,000 AU intensity and also improved LC retention ( Figure 2C).…”

“…To address these issues, derivatization of the drug analyte has been utilized to improve the chromatographic behavior and MS detection limits. Employing a protocol using precolumn derivatization coupled to LC/MS (derivatization-LC/MS [d-LC/MS]) [7], an assay was developed for the sensitive and specific bioanalysis of LY379268 in rat plasma.…”

Bioanalysis of acetylcarnitine in cerebrospinal fluid by HILIC–mass spectrometry

“…A relevant sample matrix for studying metabolites in neurological diseases is cerebrospinal fluid (CSF), which is in equilibrium with the brain's interstitial fluid and circulates through the ventricular system of the brain and spinal cord. Neurochemicals reflecting the pathophysiology of the brain are thus present in CSF and are amenable to in vivo analysis (Benitex et al, ).…”

External Influences on Invertebrate Brain Histamine and Related Compounds via an Automated Derivatization Method for Capillary Electrophoresis

“…This kind of approach has been classically used on brain samples for 30 years, mainly on microdialysates from laboratory mammals − or microdialysates from crayfish hemolymph . More recently, a method using benzoyl chloride or acetaldehyde- d 4 derivatization and HPLC with mass spectrometry (MS) detection allowed the HA monitoring in rat brain microdialysates, or rat artificial CSF . Hydrophilic liquid chromatography (HILIC) method was also coupled to tandem MS for HA microdialysate determination .…”

“…CE coupled to laser-induced fluorescence detection (CE-LIFD) could be an alternative way to determine the contents of brain HA. In fact, this amine, derivatized with naphthalene-2,3-dicarboxaldehyde (NDA), has been successfully separated with that approach in beer samples or tobacco leaf extracts. , …”