Willy E. Lambert

Matrix Effect in Bio-Analysis of Illicit Drugs with LC-MS/MS: Influence of Ionization Type, Sample Preparation, and Biofluid

The purpose of the present work was to evaluate the synergistic effect of ionization type, sample preparation technique, and bio-fluid on the presence of matrix effect in quantitative liquid chromatography (LC)-MS/MS analysis of illicit drugs by post-column infusion experiments with morphine (10-µg/mL solution). Three bio-fluids (urine, oral fluid, and plasma) were pretreated with four sample preparation procedures [direct injection, dilution, protein precipitation, solid-phase extraction (SPE)] and analyzed by both LC-electrospray ionization (ESI)-MS/MS and LC-atmospheric pressure chemical ionization (APCI)-MS/MS. Our results indicated that both ionization types showed matrix effect, but ESI was more susceptible than APCI. Sample preparation could reduce (clean up) or magnify (pre-concentrate) matrix effect. Residual matrix components were specific to each bio-fluid and interfered at different time points in the chromatogram. We evaluated matrix effect in an early stage of method development and combined optimal ionization type and sample preparation technique for each bio-fluid. Simple dilution of urine was sufficient to allow for the analysis of the analytes of interest by LC-APCI-MS/MS. Acetonitrile protein precipitation provided both sample clean up and concentration for oral fluid analysis, while SPE was necessary for extensive clean up of plasma prior to LC-APCI-MS/MS.

Therapeutic drug monitoring of old and newer anti-epileptic drugs

Abstract The aim of the present paper is to provide information concerning the setting up and interpretation of therapeutic drug monitoring (TDM) for anti-epileptic drugs. The potential value of TDM for these drugs (including carbamazepine, clobazam, clonazepam, ethosuximide, felbamate, gabapentin, lamotrigine, levetiracetam, oxcarbazepine, pheneturide, phenobarbital, phenytoin, primidone, tiagabine, topiramate, valproic acid, vigabatrin and zonisamide) is discussed in relation to their mode of action, drug interactions and their pharmacokinetic properties. The review is based upon available literature data and on observations from our clinical practice. Up until approximately 15 years ago anti-epileptic therapeutics were restricted to a very few drugs that were developed in the first half of the 20th century. Unfortunately, many patients were refractory to these drugs and a new generation of drugs has been developed, mostly as add-on therapy. Although the efficacy of the newer drugs is no better, there is an apparent improvement in drug tolerance, combined with a diminished potential for adverse drug interactions. All new anticonvulsant drugs have undergone extensive clinical studies, but information on the relationship between plasma concentrations and effects is scarce for many of these drugs. Wide ranges in concentrations have been published for seizure control and toxicity. Few studies have been undertaken to establish the concentration-effect relationship. This review shows that TDM may be helpful for a number of these newer drugs.

Comparison of Matrix Effects in HPLC-MS/MS and UPLC-MS/MS Analysis of Nine Basic Pharmaceuticals in Surface Waters

When developing an LC-MS/MS-method matrix effects are a major issue. The effect of co-eluting compounds arising from the matrix can result in signal enhancement or suppression. During method development much attention should be paid to diminishing matrix effects as much as possible. The present work evaluates matrix effects from aqueous environmental samples in the simultaneous analysis of a group of 9 specific pharmaceuticals with HPLC-ESI/MS/MS and UPLC-ESI/MS/MS: flubendazole, propiconazole, pipamperone, cinnarizine, ketoconazole, miconazole, rabeprazole, itraconazole and domperidone. When HPLC-MS/MS is used, matrix effects are substantial and can not be compensated for with analogue internal standards. For different surface water samples different matrix effects are found. For accurate quantification the standard addition approach is necessary. Due to the better resolution and more narrow peaks in UPLC, analytes will co-elute less with interferences during ionisation, so matrix effects could be lower, or even eliminated. If matrix effects are eliminated with this technique, the standard addition method for quantification can be omitted and the overall method will be simplified. Results show that matrix effects are almost eliminated if internal standards (structural analogues) are used. Instead of the time-consuming and labour-intensive standard addition method, with UPLC the internal standardization can be used for quantification and the overall method is substantially simplified.

Retinoic acid: an endogenous compound of human blood. Unequivocal demonstration of endogenous retinoic acid in normal physiological conditions.

Abstract The occurrence of retinoic acid (vitamin A acid) as a normal constituent of the vitamin A reserve from the body is demonstrated. Improved methodologies based on selected peak monitoring (spm) and high-performance liquid chromatography (hplc) are used for the detection of retinoic acid in EDTA-plasma. After extensive cleanup by double-phase extraction and chromatography on Sephadex LH-20, retinoic acid is determined as its methyl derivative by spm. A different approach using double-phase extraction combined with a preextraction step and hplc is used to confirm the findings of the spm experiments. Both techniques proved the presence of retinoic acid in human plasma at a concentration of 1 to 3 ng/ml.

Dried blood spots in toxicology: from the cradle to the grave?

About a century after its first described application by Ivar Bang, the potential of sampling via dried blood spots (DBS) as an alternative for classical venous blood sampling is increasingly recognized. Perhaps best known is the use of DBS in newborn screening programs, ignited by the hallmark paper by Guthrie and Susi half a century ago. However, it is only recently that both academia and industry have recognized the many advantages that DBS sampling may offer for bioanalytical purposes, as reflected by the strong increase in published reports during the last few years. Currently, major DBS applications include newborn screening for metabolic disorders, epidemiological surveys (e.g. HIV monitoring), therapeutic drug monitoring (TDM), as well as toxicology. In this review, we provide a comprehensive overview of the distinct subdisciplines of toxicology for which DBS sampling has been applied. DBS sampling for toxicological evaluation has been performed from birth until autopsy, aiming at the assessment of therapeutic drugs, drugs of abuse, environmental contaminants, toxins, as well as (trace) elements, with applications situated in fields as toxicokinetics, epidemiology and environmental and forensic toxicology. We discuss the strengths and limitations of DBS in the different subdisciplines and provide future prospects for the use of this promising sampling technique in toxicology.

Micronutrient levels in HIV-1-infected children.

DesignProspective study. SettingThe study was performed on HIV-1-infected children at the Paediatric Haematology, and Oncology Unit of Toulouse Hospital, France. PatientsTwenty-one children, suffering from HIV-1 infection, and 21 control subjects of similar age (2–9 years) were included in the study. In the HIV-1-infected children, two subgroups were considered according to stage (non-AIDS or AIDS), based on the Centers for Disease Control, and Prevention 1987 criteria. ResultsThe first statistically significant deficiencies occurred at non-AIDS stage, and were confirmed at AIDS stage: P<0.05 for lycopene, retinol, tocopherol, and P< 0.001 for transthyretin, and serum albumin. Levels of copper (40%), and long-chain polyunsaturated fatty acids (21%) were higher in the non-AIDS group than the controls. ConclusionBiological impairing of the micronutrient levels was observed in the non-AIDS stage without clinical sign. This information is useful in delineating eventual, and well considered nutritional intervention strategies that may improve the clinical status of HIV-1-infected children, and perhaps alter the course of their disease.

Hair ethyl glucuronide levels as a marker for alcohol use and abuse: A review of the current state of the art

BACKGROUND Ethyl glucuronide (EtG) is a minor alcohol metabolite that has been proposed as a stable marker in hair to detect and quantify alcohol consumption over long time periods. METHODS We provide an outline of currently available techniques for EtG hair sample analysis and highlight the pitfalls related to data interpretation. The literature of EtG analysis has been reviewed from January 1980 up to August 2013. In addition, we present an overview of the clinical and forensic studies which have used EtG quantification in hair as a marker for alcohol consumption/abstinence and we provide suggestions for future research. RESULTS EtG is a stable marker in hair that can be used to detect and quantify alcohol consumption over long time periods. This alcohol metabolite remains in hair after complete elimination of alcohol. Currently, there are three main analytical techniques used to quantify EtG in hair: gas chromatography-mass spectrometry (GC-MS), gas chromatography-tandem mass spectrometry (GC-MS/MS), and liquid chromatography-tandem mass spectrometry (LC-MS/MS). No standardized protocols are yet available for the analysis of EtG levels in hair samples, and the current protocols vary in sample preparation and extraction procedures. Variables such as hair length, cosmetic treatment, gender, and pathophysiological conditions influence the final results and should be taken into account. CONCLUSIONS EtG quantification in hair is a useful tool for the objective detection of alcohol consumption over extended time periods, but care should be taken when interpreting the results.

Folates and Folic Acid: From Fundamental Research Toward Sustainable Health

Folates are of paramount importance in one-carbon metabolism of most organisms. Plants and microorganisms are able to synthesize folates de novo, making them the main dietary source for humans and animals, which are dependent on food or feed supplies for folates. Folate deficiency is an increasing problem in the developing, as well as in the developed regions of the world, affecting millions of people. Different strategies, such as food fortification and folic acid supplementation, remain far from accessible for the poor rural populations in developing countries. Increasing knowledge concerning folate biosynthesis, transport and catabolism does not only deepen our insight on the regulation of folate metabolism but also provides the keys towards folate enhancement through metabolic engineering in bacteria, as well as in plants. Recently, promising results were obtained using such an approach, but further fundamental research is a prerequisite to develop a practicable solution to fight folate deficiency. In parallel, progress in the development and improvement of folate analysis has been made. Here, we provide the state-of-the-art of folate biosynthesis, catabolism, and salvage. Finally, we report on progress in folate biofortification and discuss the agroeconomical aspect of biofortified crop plants.

Validation of a solid-phase extraction and liquid chromatography-electrospray tandem mass spectrometric method for the determination of nine basic pharmaceuticals in wastewater and surface water samples.

This paper reports the development and validation of a quantitative LC-electrospray (ESI)-MS/MS method for the simultaneous analysis of nine basic pharmaceuticals (flubendazole, pipamperone, cinnarizine, ketoconazole, miconazole, rabeprazole, itraconazole, domperidone and propiconazole) in environmental waters. Sample preparation consisted of solid-phase extraction on a Speedisk phenyl and a NH(2) solid-phase extraction tube for sample clean-up. Chromatography was performed on a pentafluorophenyl column in a total run time of 24min. Due to different matrix effects measured in different surface water samples, standard addition was the only method to perform accurate quantification. Limits of detection and quantification were in the range of <0.05-1ng/l and 0.05-10ng/l, respectively. The method showed good precision and accuracy. Recoveries were in the range of 60-100%. This method allows to identify and quantify these pharmaceuticals in wastewater and surface water and enables to perform comprehensive studies on the occurrence in and removal of these drugs from influent and effluent wastewaters and surface waters.

Relevant Issues in the Monitoring and the Toxicology of Antidepressants

This review provides specific information concerning the interpretation and usefulness of antidepressant drug monitoring. Fifteen antidepressants (ADs) were selected based on their importance in the 7 major markets (Japan, USA, France, United Kingdom, Italy, Spain, Germany) according to the Cognos Plus Study #11. Literature data were reviewed concerning monitoring of the tricyclic ADs amitriptyline, clomipramine, imipramine, and new generation antidepressants such as citalopram, escitalopram, fluoxetine, fluvoxamine, paroxetine, sertraline, venlafaxine, mianserin, mirtazapine, bupropion, and milnacipran. In addition, St.-Johns Wort was added as this natural antidepressant is very popular in Europe. The objectives of therapeutic drug monitoring (TDM) for old and new generation ADs differs, as older ADs have narrow therapeutic windows with higher risks of severe drug interactions, while the new ADs have a wide therapeutic range, but an unclear plasma concentration-effect relationship. Therefore, the purpose of TDM for new-generation ADs leads more to the monitoring of patient compliance and special patient groups such as the elderly, patients with liver and kidney impairment, patients with poor metabolism by CYP 450 and comedication with inhibitors and inducers of those enzymes.