Simultaneous quantitation of histamine and its major metabolite 1-methylhistamine in brain dialysates by using precolumn derivatization prior to HILIC-MS/MS analysis.

In nonclinical drug development targeting the central nervous system (CNS), the quantitative determination of extracellular brain concentrations of neurotransmitters is a key challenge. In some CNS disorders, the monitoring of the modified profile of neurotransmitter release such as that of histamine may explain the mechanism of action of the drug candidate. Microdialysis is a commonly used method for sampling extracellular levels of neurotransmitters/drug candidates in small laboratory animals. Detection and quantification of extracellular levels of neurotransmitters remain an analytical and technical challenge owing to the low concentrations of neurotransmitters collected, the small microdialysis sample size, and the high amount of inorganic salts. A precolumn derivatization strategy prior to hydrophilic interaction liquid chromatography (HILIC)-tandem mass spectrometry analysis is proposed to quantify histamine release after administration of a CNS research compound. Derivatization using propionic anhydride dissolved in organic solvent combined with the HILIC approach effectively eliminated three time-consuming steps, organic layer transfer, dry down, and reconstitution, all of which are required by traditional reversed-phase liquid chromatography. The formation of propionylated amides, performed under mild conditions, required no further sample cleanup. After a dual microdialysis probe implantation into the prefrontal cortex (neurotransmitters) and in the inferior vena cava of rat (drug candidate), microdialysate fractions were collected every 15 min for 8 h and stored frozen at -20 °C until analysis. The method was validated using 10 µL microdialysate, achieving low limits of quantitation of 83.4 and 84.5 pg.mL(-1) for histamine and 1-methylhistamine, respectively. These limits were suitable to assess kinetic release of neurotransmitters and are compatible with those obtained by microdialysis sampling. This method provided the required selectivity, sensitivity, accuracy, and precision to assess release kinetics of histamine and 1-methylhistamine in several hundred rat brain microdialysates after intravenous infusion of CNS drug candidates.

Stereoselective renal tubular secretion of levocetirizine and dextrocetirizine, the two enantiomers of the H1-antihistamine cetirizine.

Competition for uptake and/or efflux transporters can be responsible for drug interactions. Cetirizine is mainly eliminated unchanged in urine through both glomerular filtration and tubular secretion. The aim of this study was to investigate whether the eutomer, levocetirizine, and the distomer, dextrocetirizine, have a similar tubular secretion. The renal clearance associated with tubular secretion was calculated from the renal clearance of levocetirizine and dextrocetirizine obtained in a study in healthy volunteers. The values of the unbound fraction in plasma were obtained in an in vitro study of the binding of (14)C-cetirizine and (14)C-levocetirizine to human plasma proteins using equilibrium dialysis and chiral high-performance liquid chromatography (HPLC) with on-line liquid scintillation counting. The unbound fraction was 0.074 for levocetirizine and 0.141 for dextrocetirizine. The tubular secretion of dextrocetirizine (44.5 mL/min) is higher than that of levocetirizine (23.1 mL/min), which may have consequences for drug interactions at the renal level. The higher tubular secretion for dextrocetirizine may be due to the higher free fraction available for secretion or to a higher affinity for (a) renal transporter(s) mediating the secretion pathway.

In vivo tolerance assessment of skin after insertion of subcutaneous and cutaneous microdialysis probes in the rat.

The purpose of the study was to evaluate the trauma induced by insertion of the linear microdialysis probe in the subcutaneous and dermal tissue in the rat and to check if the microdialysis probe insertion affects transdermal drug delivery. Non-invasive bioengineering methods (TEWL, Laser Doppler Velocimeter, Chromameter) as well as histology were combined to characterize these effects. The results showed that the dermal and subcutaneous insertion of microdialysis probes did not change skin permeability, blood flow and color, confirming the safety of this technique. The probe depth did not influence the trauma. No significant physical damage after probe insertion was noticed. Thus, the present work validates the use of microdialysis in dermatopharmacokinetics studies after topical or systemic drug delivery.

Validation of subcutaneous microdialysis sampling for pharmacokinetic studies of flurbiprofen in the rat.

The objective of this study was to validate subcutaneous (sc) microdialysis sampling to study flurbiprofen pharmacokinetics and plasma protein binding in the awake freely moving rat. A linear microdialysis probe was manufactured using a Hemophane hollow fiber which was tested in vitro and in vivo for the recovery of flurbiprofen and naproxen used as retrodialysis marker. Flurbiprofen was administered intraperitoneally and intravenously at a dose of 20 mg/kg in rats. In both cases, conventional blood sampling and sc microdialysis sampling were simultaneously performed. The microdialysates were analyzed on-line by high-pressure liquid chromatography. Naproxen, which was shown to have a similar in vivo loss by retrodialysis as flurbiprofen (71.5 +/- 0.9% and 71.0 +/- 0.8% respectively, n = 3), was used to continuously monitor probe recovery. Concentration-dependent protein binding of flurbiprofen was demonstrated in vivo based on experiments with a simultaneous sc microdialysis and blood sampling. Values of unbound fraction were similar to those reported previously by intravenous microdialysis sampling, demonstrating that the sc unbound concentrations are very similar to those in the central compartment. There was no significant difference among pharmacokinetic parameters (AUC, CL, t(1/2z), Vd) for total or unbound flurbiprofen determined after intraperitoneal and intravenous administration. Subcutaneous microdialysis is a simple yet powerful tool to study the pharmacokinetics and the in vivo plasma protein binding of flurbiprofen in the awake unrestrained rat.