The Development of New Methodology for Determination of Vincristine (VCR) in Human Serum Using LC-MS/MS-Based Method for Medical Diagnostics.

In this article, we have presented the development and validation of a rapid and sensitive reversed-phase liquid chromatography with tandem mass spectrometry (LC-MS/MS) method for the determination of vincristine (VCR) in patient serum samples. Chromatographic separation was achieved on a Kinetex® (Singapore) column using a mobile phase consisting of 25 mM acetic acid and 0.3% formic acid (A) and methanol (B) in a gradient elution mode at a flow rate of 0.3 mL/min. The VCR and internal standard (vinblastine) were monitored using the multiple reaction monitoring mode under positive electrospray ionization. The lower limit of quantification (LLOQ) was 0.67 ng/mL, and the upper limit of quantification (ULOQ) was 250 ng/mL for VCR. The calculated values of LOD and LOQ for VCR were 0.075 and 0.228 ng/mL, respectively. The calibration curve was linear over the VCR concentration range of 1.0−250 ng/mL in serum. The intra- and inter-day precision and precision were within the generally accepted criteria for the bioanalytical method (<15%). The method was successfully applied to the analysis of serum samples in clinical practice.

Neurotoxicological profile of the hallucinogenic compound 25I-NBOMe.

4-Iodo-2,5-dimethoxy-N-(2-methoxybenzyl)phenethylamine (25I-NBOMe) is a new psychoactive substance with strong hallucinogenic properties. Our previous data reported increased release of dopamine, serotonin, and glutamate after acute injections and a tolerance development in the neurotransmitters release and rats' behavior after chronic treatment with 25I-NBOMe. The recreational use of 25I-NBOMe is associated with severe intoxication and deaths in humans. There is no data about 25I-NBOMe in vivo toxicity towards the brain tissue. In this article 25I-NBOMe-crossing through the blood-brain barrier (BBB), the impact on DNA damage, apoptosis induction, and changes in the number of cortical and hippocampal cells were studied. The presence of 25I-NBOMe in several brain regions shortly after the drug administration and its accumulation after multiple injections was found. The DNA damage was detected 72 h after the chronic treatment. On the contrary, at the same time point apoptotic signal was not identified. A decrease in the number of glial but not in neural cells in the frontal (FC) and medial prefrontal cortex (mPFC) was observed. The obtained data indicate that 25I-NBOMe passes easily across the BBB and accumulates in the brain tissue. Observed oxidative DNA damage may lead to the glial cells' death.

A fully validated HPLC-UV method for determination of sulthiame in human serum/plasma samples.

Sulthiame is an old antiepileptic medicine with controversial history, whose effectiveness and safety in use have been stated in some current studies. However, there is still a need for further clinical examinations for confirmation of its usefulness and tolerability in monotherapy and add-on therapy for epilepsy of various etiologies. A fully validated RP HPLC-UV method for determination of sulthiame in serum/plasma samples using desethylatrazine as the internal standard was developed. The biological fluid was prepared for analysis by a simple precipitation method with acetonitrile. The following validation parameters of the method were determined: selectivity/specificity, linearity range (0.2-50.0 µl/ml, R2 > 0.9999), limits of detection (0.19 µl/ml) and quantification (0.58 µl/ml), precision (intra-day CV 1.06% and inter-day CV 1.25%), extraction recovery (~100%), accuracy (bias, -4.61-0.80%), carryover and ruggedness. Moreover, the stability of the medicine in plasma samples under different storage conditions was also tested. The usability of the method for clinical examinations was checked by analysis of serum samples originating from 19 patients treated with sulthiame. The proposed method is appropriate for determination of sulthiame in serum/plasma samples for drug monitoring purposes, as well as for pharmacokinetic studies.

Sample preparation and determination of pesticides in fat-containing foods.

Pesticides are still a very important factor in food plant cultivation. The lipophilic character of most pesticides can lead to their accumulation in fat, which can have harmful effects on humans and animals. The aim of this review is to present current knowledge about the isolation and determination of pesticides in fatty commodities. The following techniques for isolation are discussed: solvent partitioning, solid phase and dispersive solid phase, QuEChERS-based procedures, matrix solid-phase dispersion, gel permeation chromatography, Soxhlet extraction, accelerated solvent extraction, and microextraction techniques. Chromatographic methods predominate in the analysis of pesticide residues in fatty matrices and, for this reason, the review focuses on these methods, particularly those combined with mass spectrometry. Analytical chemists still face the challenge - in the determination of pesticide residues in fatty matrices - of developing simpler and quicker procedures that consume less organic solvents.

The Potential of Graphene as an Adsorbent for Five Pesticides from Different Classes in Rape Oil Samples Using Dispersive Solid-Phase Extraction.

Isolation conditions for five pesticides (metazachlor, tebuconazole, λ-cyhalothrin, chlorpyrifos, and deltamethrin) from rape oil samples were examined using the dispersive solid-phase graphene extraction technique. To determine the optimal extraction conditions, a number of experimental factors (amount of graphene, amount of salt, type and volume of the desorbing solvent, desorption time with and without sonication energy, and temperature during desorption) were studied. The compounds of interest were separated and detected by an HPLC-UV employing a Kinetex XB-C18 column and a mobile phase consisting of acetonitrile and water flowing in a gradient mode. The optimized extraction conditions were: the amount of graphene 15 mg, desorbing solvent (acetonitrile) 5 mL, time desorption 10 min at 40°C, and amount of NaCl 1 g. The detection limit for metazachlor, tebuconazole, λ-cyhalothrin, and chlorpyrifos was 62.5 ng·g-1, and for deltamethrin, it was 500 ng·g-1. The obtained results lead to the conclusion that graphene may be successfully used for the isolation of the five pesticides from rape oil. However, their determination at low concentration levels, as they occur in real oil samples, requires the employment of appropriately highly sensitive analytical methods, as well as a more suitable graphene form (e.g., magnetically modified graphene).

Analytical methodologies for the determination of benzodiazepines in biological samples.

Benzodiazepine drugs belong to important and most widely used medicaments. They demonstrate such therapeutic properties as anxiolytic, sedative, somnifacient, anticonvulsant, diastolic and muscle relaxant effects. However, despite the fact that benzodiazepines possess high therapeutic index and are considered to be relatively safe, their use can be dangerous when: (1) co-administered with alcohol, (2) co-administered with other medicaments like sedatives, antidepressants, neuroleptics or morphine like substances, (3) driving under their influence, (4) using benzodiazepines non-therapeutically as drugs of abuse or in drug-facilitated crimes. For these reasons benzodiazepines are still studied and determined in a variety of biological materials. In this article, sample preparation techniques which have been applied in analysis of benzodiazepine drugs in biological samples have been reviewed and presented. The next part of the article is focused on a review of analytical methods which have been employed for pharmacological, toxicological or forensic study of this group of drugs in the biological matrices. The review was preceded by a description of the physicochemical properties of the selected benzodiazepines and two, very often coexisting in the same analyzed samples, sedative-hypnotic drugs.

Sequential cloud-point extraction for toxicological screening analysis of medicaments in human plasma by high pressure liquid chromatography with diode array detector.

A complex extraction system with the use of cloud-point extraction technique (CPE) was developed for sequential isolation of basic and acidic/neutral medicaments from human plasma/serum, screened by HPLC/DAD method. Eight model drugs (paracetamol, promazine, chlorpromazine, amitriptyline, salicyclic acid, opipramol, alprazolam and carbamazepine) were chosen for the study of optimal CPE conditions. The CPE technique consists in partition of an aqueous sample with addition of a surfactant into two phases: micelle-rich phase with the isolated compounds and water phase containing a surfactant below the critical micellar concentration, mainly under influence of temperature change. The proposed extraction system consists of two chief steps: isolation of basic compounds (from pH 12) and then isolation of acidic/neutral compounds (from pH 6) using surfactant Triton X-114 as the extraction medium. Extraction recovery varied from 25.2 to 107.9% with intra-day and inter-day precision (RSD %) ranged 0.88-1087 and 5.32-17.96, respectively. The limits of detection for the studied medicaments at λ 254nm corresponded to therapeutic or low toxic plasma concentration levels. Usefulness of the proposed CPE-HPLC/DAD method for toxicological drug screening was tested via its application to analysis of two serum samples taken from patients suspected of drug overdosing.

Capillary electrophoresis screening method for six tricyclic antidepressants in human serum.

A capillary electrophoretic (CE) method for screening of six tricyclic antidepressants: amitriptyline, nortriptyline, imipramine, desipramine, doxepin and nordoxepin, in human serum, has been developed. The drugs were separated in a bare fused silica capillary (50 microm i.d.) using the background electrolyte: 50 mM CAPSO (pH 9.54) in methanol/water with KCI addition. For increasing the method sensitivity, the sample concentration in the capillary (sample stacking) using pressure and electrokinetic injection has been applied. The standard addition method was used for calibration of the developed analytical procedure. The precision of the identification parameter (the relative migration time) and the quantitative parameter (the relative peak area) was within the range of 0.05-1.65 and 0.73-6.7 (RSD %), respectively. The detection limits were found to be 30 ng/mL for desipramine, 62.5 ng/mL for nortriptyline, and 50 ng/mL for remaining analytes.

[Hemostatic and fibrinolysis markers in the early phase of acute ischaemic stroke]. / Markery aktywacji wczesnej fazy krzepniecia i fibrynolizy u chorych z zawalem mózgu w najwczesniejszym okresie choroby.

Stroke is the most frequent cause of focal brain damage. It seems that some coagulation and fibrinolysis abnormalities play an important role in this disease. We measured coagulation parameters in 39 patients with acute ischaemic stroke within 1 week of onset of the stroke (1st, 3rd and 7th day), including thrombin-antithrombin complex (TAT), prothrombin fragment F1+2 (F1+2), fibrin degradation products (D-dimer) and fibrinogen (FBG). Marker levels in stroke were compared with controls (n=25). A significant elevation of plasma concentration of TAT, F1+2 and fibrinogen was observed in all days of measurements (p < 0.001). D-dimer level was significantly elevated in 30.7% of patients at the 1st day and in 23.0% of patients at the 3rd day of stroke onset. Marker utility in acute stroke awaits a larger study.

HPLC/DAD Screening Method for Selected Psychotropic Drugs in Blood.

The influence of experimental conditions on gradient high-performance liquid chromatography/diode array detector separation and identification of 13 psychotropic drugs belonging to two groups--phenothiazines and tricyclic antidepressants--was examined. The main interaction effects of three experimental factors were determined according to a 2 3 factorial design, and the optimum conditions of analysis were searched for. The degree to which the chromatographic peaks overlap is taken into account in a proposed criterion for separation quality. The screening analysis proposed was tested on whole blood samples spiked with mixtures of the examined drugs. The method was characterized by such validation parameters as relative retention times, absorbance ratios at two wavelengths, detection limits, linearity ranges, and extraction recoveries. The method proved to be a suitable tool for the identification of psychotropic drugs tested for forensic purposes.