Urinary Profile of Endogenous Gamma-Hydroxybutyric Acid and its Biomarker Metabolites in Healthy Korean Females: Determination of Age-Dependent and Intra-Individual Variability and Identification of Metabolites Correlated With Gamma-Hydroxybutyric Acid

Abstract

Gamma-hydroxybutyric acid (GHB), used as a therapeutic and an illegal anesthetic, is a human neurotransmitter produced during gamma-aminobutyric acid (GABA) biosynthesis and metabolism. Potential biomarker metabolites of GHB intoxication have been identified previously; however, reference concentrations have not been set due to the lack of clinical study data. Urinary profiling of endogenous GHB and its biomarker metabolites in urine samples (n = 472) of 206 healthy females was performed based on differences in age and time of sample collection using liquid chromatography-tandem mass spectrometry following validation studies. The unadjusted and creatinine-adjusted urinary concentrations ranges were obtained after urinary profiling. The creatinine-adjusted concentrations of glutamic and succinic acids and succinylcarnitine significantly increased, whereas that of glycolic acid significantly decreased with advancing age. Significant inter-day variation of GABA concentration and intra-day variation of 3,4-dihydroxybutyric acid and succinylcarnitine concentrations were observed. The urinary concentrations of 2,4-dihydroxybutyric acid, succinic acid, and 3,4-dihydroxybutyric acid showed the highest correlation with that of GHB. Data from this study suggest population reference limits to facilitate clinical and forensic decisions related to GHB intoxication and could be useful for identification of biomarkers following comparison with urinary profiles of GHB-administered populations.

New gamma‐hydroxybutyric acid (GHB) biomarkers: Development and validation of a liquid chromatography–tandem mass spectrometry method for the determination of GHB amino acid, carnitine, and fatty acid conjugates in urine

“…Concentrations determined for organic acids 2,4‐DHB, 3,4‐DHB, GA, and succinylcarnitine in GHB‐negative cases were similar to ranges reported earlier in larger sample cohorts 28,33 . An exception was observed for SA in the present samples with determined concentrations exceeding those of former studies.…”

“…T A B L E 3 Unadjusted and creatinine-normalized concentrations of GHB, GHB conjugates, and organic acids determined in authentic samples Concentrations determined for organic acids 2,4-DHB, 3,4-DHB, GA, and succinylcarnitine in GHB-negative cases were similar to ranges reported earlier in larger sample cohorts. 28,33 An exception was observed for SA in the present samples with determined concentrations exceeding those of former studies. The observed instability of SA in the other studies 28,30,32 but not in the present study, might explain these discrepancies.…”

“…Increased levels of DHB isomers and GA in (suspected) GHB intoxication cases and postmortem cases were observed, useful as additional markers for GHB interpretation 29–31 . Kim et al analytically expanded on this using LC–MS/MS after chemical derivatization with benzoyl chloride for the urinary analysis of not only 2,4‐DHB, 3,4‐DHB, GA, and SA but also GABA, glutamic acid and succinylcarnitine 32 and applied their method to characterize endogenous (GHB‐unrelated) reference ranges for these compounds 33 …”

“…[29][30][31] Kim et al analytically expanded on this using LC-MS/MS after chemical derivatization with benzoyl chloride for the urinary analysis of not only 2,4-DHB, 3,4-DHB, GA, and SA but also GABA, glutamic acid and succinylcarnitine 32 and applied their method to characterize endogenous (GHB-unrelated) reference ranges for these compounds. 33 Untargeted metabolome screening approaches on urine and serum samples from a controlled GHB administration study in humans identified a number of novel conjugates of GHB with carnitine, the amino acids (AA) glycine, glutamate, and taurine, the carbohydrate pentose, and some interesting compounds (U3, U4, and U16) unknown to date, next to changes in endogenous compounds. 23,25 Fatty acid (FA) esters and phospholipid metabolites, 34 well-known metabolites, for example, for ethanol, were recently also considered as possible GHB metabolites.…”

Interaction between γ-Hydroxybutyric Acid and Ethanol: A Review from Toxicokinetic and Toxicodynamic Perspectives

“…Moreover, it was previously reported that acute alcohol exposure alters the concentrations of dopamine, glutamic acid, and GABA in rats’ brains [ 62 , 63 ]. GHB is metabolized to 2,4- or 3,4-dihydroxybutyric acid or succinic acid which goes through the Krebs cycle [ 64 , 65 ] and then rapidly eliminated from the body in the form of carbon dioxide and water. GHB was detected only within 6 h in the blood and 12 h in urine after a single exposure to 60 or 50 mg/kg of GHB [ 16 , 66 , 67 ].…”