N, N-dimethyltryptamine forms oxygenated metabolites via CYP2D6 - an in vitro investigation.

N, N-dimethyltryptamine (DMT) is a psychedelic compound that has shown potential in the treatment of depression. Aside from the primary role of monoamine oxidase A (MAO-A) in DMT metabolism, the metabolic pathways are poorly understood. Increasing this understanding is an essential aspect of ensuring safe and efficacious use of DMT.This work aimed to investigate the cytochrome 450 (CYP) mediated metabolism of DMT by incubating DMT with recombinant human CYP enzymes and human liver microsomes (HLM) followed by analysis using high-resolution mass spectrometry for metabolite identification.DMT was rapidly metabolised by CYP2D6, while stable with all other investigated CYP enzymes. The metabolism of DMT in HLM was reduced after inclusion of harmine and SKF-525A whereas quinidine did not affect the metabolic rate, likely due to MAO-A residues present in HLM. Analysis of the CYP2D6 incubates showed formation of mono-, di- and tri-oxygenated metabolites, likely as a result of hydroxylation on the indole core.More research is needed to investigate the role of this metabolic pathway in vivo and any pharmacological activity of the proposed metabolites. Our findings may impact on safety issues following intake of ayahuasca in slow CYP2D6 metabolizers or with concomitant use of CYP2D6 inhibitors.

Estimating Analytical Errors of Glomerular Filtration Rate Measurement.

BACKGROUND: Few studies are available on how to optimize time points for sampling and how to estimate effects of analytical uncertainty when glomerular filtration rate (GFR) is calculated. METHODS: We explored the underlying regression mathematics of how analytical variation of a kidney filtration marker affects 1-compartment, slope-and-intercept GFR calculations, using 2 or 3 time points following a bolus injection, and used this to examine the results from 731 routine 3-point iohexol plasma clearance measurements. RESULTS: GFR calculations inflated analytical uncertainty if the time points were taken too late after the bolus injection and too close after each other. The uncertainty in GFR calculation was, however, the same as the analytical uncertainty if optimal time points were used. The middle of the 3 samples was of little value. The first sample should be taken as early as possible after the distribution phase. Sampling before the patient specific half-life of the kidney filtration marker resulted in an exponential error inflation whereas no error inflation was seen when sampling occurred later than 2 half-lives. Theoretical GFR uncertainty could be lowered 2.6-fold if individually optimized time points for sampling had been used in our 731 clearance measurements. Using Taylor expansions to approximate the moments of transformed random variables, the uncertainty of an individual GFR measurement could be calculated in a simple enough way to be applicable by laboratory software. CONCLUSIONS: We provide a theoretical foundation to select patient-optimal time points that may both limit errors and allow calculation of GFR uncertainty.

Long-term Transplant Function After Thrombolytic Treatment Ex Vivo of Donated Kidneys Retrieved 4 to 5 H After Circulatory Death.

BACKGROUND: Using a novel thrombolytic technique, we present long-term transplant function, measured by creatinine and iohexol clearance, after utilizing kidneys from porcine donors with uncontrolled donation after circulatory deaths, with 4.5-5 h of warm ischemia. METHODS: Pigs in the study group were subjected to simulated circulatory death. After 2 h, ice slush was inserted into the abdomen and 4.5 h after death, the kidneys were retrieved. Lys-plasminogen, antithrombin-III, and alteplase were injected through the renal arteries on the back table. Subsequent ex vivo perfusion was continued for 3 h at 15°C, followed by 3 h with red blood cells at 32°C, and then transplanted into pigs as an autologous graft as only renal support. Living-donor recipient pigs that did not receive ex vivo perfusion, and unilateral nephrectomized pigs served as the controls. RESULTS: Pigs in the study group (n = 13), surviving 10 d or more were included, of which 7 survived for 3 mo. Four animals in the living-donor group (n = 6) and all 5 nephrectomized animals survived for 3 mo. Creatinine levels in the plasma and urine, neutrophil gelatinase-associated lipocalin levels, Kidney Injury Marker-1 expression, and iohexol clearance at 3 mo did not differ significantly between the study and living-donor groups. Histology and transmission electron microscopy after 3 mo showed negligible fibrosis and no other damage. CONCLUSIONS: The present method salvages kidneys from extended unontrolled donation after circulatory death using thrombolytic treatment while preserving histology and enabling transplantation after ex vivo reconditioning, with clinically acceptable late function after 3 mo, as measured by creatinine and iohexol clearance.

Unexpected benzimidazole ring formation from a quinoneimide species in the presence of ammonium acetate as supporting electrolyte used in the coupling of electrochemistry with mass spectrometry.

RATIONALE: Electrochemistry (EC) coupled to mass spectrometry (MS) has been used to study different phase-I reactions. Despite of the versatility of EC/MS, the effect of the nature of the supporting electrolyte on the formation of oxidation products has seldom been discussed during EC/MS experiments. Here, we present a comparison of two different supporting electrolytes and their effect on the identification of unstable intermediate oxidation species is discussed. METHODS: The oxidation of acebutolol was performed with a coulometric cell in the presence of two supporting electrolytes namely ammonium acetate and lithium acetate. Ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC/QTOFMS) using a binary gradient (water/acetonitrile) with positive electrospray ionization was used to identify the oxidation products in the presence and absence of glutathione. Chemical structure elucidations of the oxidation products were performed by high-resolution mass spectrometry (HRMS) and were also supported by nuclear magnetic resonance (NMR) measurements. RESULTS: From the electrochemical study and HRMS measurements, we demonstrate that the quinoneimide species resulting from the oxidative hydrolyses of acebutolol gives a benzimidazole ring product in the presence of ammonium acetate. Through the example of the oxidation of acebutolol, a correlation between the supporting electrolyte nature and oxidation product formation was established. The obtained results were supported by quantum mechanical calculations. CONCLUSIONS: We present here evidence of the side reactions induced by the presence of ammonia as supporting electrolyte during EC/MS measurements. Acebutolol was used as a model to postulate an uncommon and unexpected side reaction leading to benzimidazole ring formation. The findings may help to understand the identification of the intermediate species in the oxidative degradation process.

Voltammetry coupled to mass spectrometry in the presence of isotope 18O labeled water for the prediction of oxidative transformation pathways of activated aromatic ethers: acebutolol.

The coupling between an electrochemical cell (EC) and a mass spectrometer (MS) is a useful screening tool (EC-MS) to study the oxidative transformation pathways of various electroactive species. For that purpose, we showed that the EC-MS method, carried out in the presence and absence of isotope (18)O labeled water leads not only to a fast identification of oxidation products but also leads to a fast elucidation of the mechanism pathway reaction. We examined herein the case of the electrochemical hydrolysis of activated aromatic ether. Acebutolol (ß-blockers) was selected herein as model of activated aromatic ether, and its electrochemical oxidation was examined in both the presence and absence of isotope (18)O labeled water. To elucidate electrochemical hydrolysis pathway reaction: O-dealkylation or O-dealkoxylation, our approach was used to prove its applicability. The electrochemical oxidation mechanism was then elucidated showing an O-dealkoxylation reaction. In addition, density functional theory (DFT) calculations fully support the experimental conclusions.