Ion suppression; a critical review on causes, evaluation, prevention and applications.

The consequences of matrix effects in mass spectrometry analysis are a major issue of concern to analytical chemists. The identification of any ion suppressing (or enhancing) agents caused by sample matrix, solvent or LC-MS system components should be quantified and measures should be taken to eliminate or reduce the problem. Taking account of ion suppression should form part of the optimisation and validation of any quantitative LC-MS method. For example the US Food and Drug Administration has included the evaluation of matrix effects in its "Guidance for Industry on Bioanalytical Method Validation" (F.D.A. Department of Health and Human Services, Guidance for industry on bioanalytical method validation, Fed. Regist. 66 (100) 2001). If ion suppression is not assessed and corrected in an analytical method, the sensitivity of the LC-MS method can be seriously undermined, and it is possible that the target analyte may be undetected even when using very sensitive instrumentation. Sample analysis may be further complicated in cases where there are large sample-to-sample matrix variations (e.g. blood samples from different people can sometimes vary in certain matrix components, shellfish tissue samples sourced from different regions where different phytoplankton food sources are present, etc) and therefore exhibit varying ion-suppression effects. Although it is widely agreed that there is no generic method to overcome ion suppression, the purpose of this review is to: provide an overview of how ion suppression occurs, outline the methodologies used to assess and quantify the impact of ion suppression, discuss the various corrective actions that have been used to eliminate ion suppression in sample analysis, that is to say the deployment of techniques that eliminate or reduce the components in the sample matrix that cause ion suppression. This review article aims to collect together the latest information on the causes of ion suppression in LC-MS analysis and to consider the efficacy of common approaches to eliminate or reduce the problem using relevant examples published in the literature.

Development of a nano-electrospray MSn method for the analysis of serotonin and related compounds in urine using a LTQ-orbitrap mass spectrometer.

Serotonin, its key metabolite hydroxyindole acetic acid (5-HIAA) and dopamine have been shown to be potential biomarkers whose levels in serum and urine can be correlated with certain psychiatric and physiological disorders and illness, including depression, schizophrenia, anxiety and dementia. Recently we have published elsewhere that 5-HIAA has been identified as a potential biomarker for Attention Deficit Hyperactivity/Hyperkinetic Disorder (AD-HKD). This study describes a versatile and validated method for the analysis of these three compounds in urine using a nanoelectrospray-MS(n) method interfaced with an LTQ Orbitrap mass spectrometer. No chromatographic separation is required prior to nanoelectrospray infusion. Good linear calibrations were obtained for analytes in urine (with serotonin and dopamine giving R(2)=0.9999 and 5-HIAA having a lower R(2) value of 0.9955). Acceptable intraday repeatability was achieved for all analytes with RSD values (n=5) ranging from 4.4% to 6.2% (57, 65 and 52 nmol/L for serotonin, dopamine and 5-HIAA respectively) to 2.1-8.1% (2837, 3268, 2618 nmol/L for serotonin, dopamine and 5-HIAA respectively). Excellent limits of detection (LOD) and limits of quantitation (LOQ) were achieved with spiked samples for all compounds; with LODs of 9-12.9 nmol/L and LOQs of 27.2-57.7 nmol/L for analytes in urine. An appropriate sample clean-up procedure for urine was developed to ensure highest recovery and reproducibility on analysis.

Development of an LC-MS/MS method for the analysis of serotonin and related compounds in urine and the identification of a potential biomarker for attention deficit hyperactivity/hyperkinetic disorder.

Serotonin is a major neurotransmitter and affects various functions both in the brain and in the rest of the body. It has been demonstrated that altered serotinergic function is implicated in various psychiatric disorders including depression and schizophrenia. Serotonin has also been implicated along with dopamine in attention deficit-hyperkinetic disorder (AD-HKD). This study provides a versatile validated method for the analysis of serotonin, hydroxyindole acetic acid and dopamine in urine using LC-MS/MS. This method was then used to quantify these analytes in a test group of 17 children diagnosed with severe AD-HKD. This group was compared to a matched control group to investigate the possibility that one of these compounds may be a potential biomarker for this condition. The developed method provided good linear calibration curves for the multiplex assay of analytes in urine (0.05-3.27 nmol/L; R(2) ≥ 0.9977). Acceptable inter-day repeatability was achieved for all analytes with RSD values (n = 9) ranging from 1.1% to 9.3% over a concentration range of 0.11-3.27 µmol/L in urine. Excellent limits of detection (LOD) and limits of quantitation (LOQ) were achieved with LODs of 8.8-18.2 nmol/L and the LOQs of 29.4-55.7 nmol/L for analytes in urine. Recoveries were in the ranges of 98-104%, 100-106% and 91-107% for serotonin, 5-HIAA and dopamine, respectively. An appropriate sample clean-up procedure for urine was developed to ensure efficient recovery and reproducibility on analysis. Evaluation of matrix effects was also carried out and the influence of ion suppression on analytical results reported. Confirmatory analysis was carried out on a linear trap quadrupole-Orbitrap mass spectrometer to obtain high mass accuracy data of the target analytes in the clinical samples.