Simultaneous profiling of multiple neurochemical pathways from a single cerebrospinal fluid sample using GC/MS/MS with electron capture detection

Abstract

Biogenic amines and amino acids are widely characterized in the pathways representing neurotransmission. Although several analytical methodologies have been used to detect specific target molecules in relevant fluids such as cerebrospinal fluid (CSF), multiple assays must be used to survey the primary pathways involved. This article describes the development of a GC/MS/MS method capable of analyzing up to 43 analytes (representing 20 amino acids and more than seven neurochemical pathways) from a single 50 microl CSF sample. In this procedure, a CSF sample is first treated with acetonitrile to precipitate proteins. The dried sample is then derivatized with a mixture of 2,2,3,3,3-pentafluoro-1-propanol and pentafluoropropionic acetic anhydride to replace all active hydrogen atoms with fluorine-containing groups. Due to the concentration difference between amino acids and neurotransmitters, these two compound classes are analyzed in separate injections of the same derivatized extract. The total run time for each injection is approximately 15-20 min. An essential feature of the method is the use of argon as a reagent gas for electron capture chemical ionization (ECCI), as the use of the more traditional gas (methane) lacked sufficient durability to be considered for use with the present instrumentation. This article describes the development of this method including a detailed investigation of the chemical ionization conditions used. The resultant conditions allow for the profiling of biogenic amines (e.g. serotonin, norepinephrine, and dopamine) in the low picogram per milliliter range.

Effects of a novel mGlu2/3 receptor agonist prodrug, LY2140023 monohydrate, on central monoamine turnover as determined in human and rat cerebrospinal fluid

“…A previously described gas chromatography/tandem mass spectrometry (GC/MS/MS) assay (Eckstein et al 2008) using electron capture negative chemical ionization was adapted with minor modifications (described in the ESM) to profile selected metabolites in human CSF samples from the dopaminergic, noradrenergic, and serotonergic neurotransmitter pathways. Samples were also assayed for DA, NE, and 5-HT; however, of these, only DA was reliably measured in human CSF under the conditions used for analysis.…”

“…Linear dynamic ranges (based on CSF samples spiked with stable isotope-labeled compounds) exceeded two orders of magnitude in all cases. The stability of the analytes was controlled as described previously (Eckstein et al 2008).…”

The effects of multi-strain probiotics on liver disease

“…This made it difficult to be accurate with our recovery data. With argon as our chemical ionization gas [26], the signal remained stable and recovery and accuracy data was able to be determined. Our extraction method consistently recovers >90% (typically >95%) of added internal standard with an accuracy of >96% (Fig.…”

“…When operating with methane as the CI gas, re-tuning once the ion volume had become contaminated was not possible. However, based on the publication of Eckstein et al [26], we switched to argon as the CI gas and re-calibrating following numerous injections was possible. While calibrating returned some of the sensitivity, a complete change of the ion volume was still the only way to fully restore the instruments sensitivity.…”

“…25 The hygiene hypothesis provides a biologically plausible explanation for this trend that implicates a loss or diminished early childhood infectivity with an enhanced risk for later life GIT inflammatory problems such as autoimmune diseases. 26 With the increasing prevalence of GI diseases, and bacteria resistant to antibiotic medications, 27 multi-strain probiotics may significantly reduce the risk of developing GI diseases by promoting restoration of GIT commensal bacteria balance Probiotics are live microorganisms that when administered in adequate amounts confer health benefits to the host. 28 The mechanisms through which probiotics reduce inflammation are complex and diverse, and depend on a number of factors that include strain specificity, dosage, the presence and concentration of other bacteria, and the specific disease state of the individual.…”