References of "Fillet, Marianne"
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See detailSeparation of human, bovine, and porcine insulins, three very closely related proteins, by micellar electrokinetic chromatography.
Lamalle, Caroline ULg; Roland, Diane ULg; Crommen, Jacques ULg et al

in Electrophoresis (2015), 36(19), 2504-6

Human, bovine, and porcine insulins are small proteins with very closely related amino acid sequences, which makes their separation challenging. In this study, we took advantage of the high-resolution ... [more ▼]

Human, bovine, and porcine insulins are small proteins with very closely related amino acid sequences, which makes their separation challenging. In this study, we took advantage of the high-resolution power of CE, and more particularly of micellar electrokinetic chromatography, to separate those biomolecules. Among several surfactants, perfluorooctanoic acid ammonium salt was selected. Then, using a design of experiments approach, the optimal BGE composition was found to consist of 50 mM ammonium acetate pH 9.0, 65 mM perfluorooctanoic acid ammonium salt, and 4% MeOH. The three insulins could be separated within 12 min with a satisfactory resolution. This method could be useful to detect possible counterfeit pharmaceutical formulations. Indeed, it would be easy to determine if human insulin was replaced by bovine or porcine insulin. [less ▲]

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See detailNew biomarkers for primary mitral regurgitation.
Deroyer, Céline ULg; Magne, Julien; Moonen, Marie ULg et al

in Clinical proteomics (2015), 12

BACKGROUND: Mitral regurgitation is a frequent valvular heart disease affecting around 2.5 % of the population with prevalence directly related to aging. Degeneration of mitral valve is broadly considered ... [more ▼]

BACKGROUND: Mitral regurgitation is a frequent valvular heart disease affecting around 2.5 % of the population with prevalence directly related to aging. Degeneration of mitral valve is broadly considered as a passive ongoing pathophysiological process and little is known about its physiological deregulation. The purpose of this study was to highlight new biomarkers of mitral regurgitation in order to decipher the underlying pathological mechanism as well as to allow the diagnosis and the monitoring of the disease. RESULTS: Modulation of various blood proteins expression was examined in patients suffering from different grades of mitral regurgitation (mild, moderate and severe) compared to healthy controls. To this end, several routine clinical assays and the multi analyte profile technology targeting 184 proteins were used. High-density lipoprotein, apolipoprotein-A1, haptoglobin and haptoglobin-alpha2 chain levels significantly decreased proportionally to the degree of mitral regurgitation when compared to controls. High-density lipoprotein and apolipoprotein-A1 levels were associated with effective regurgitant orifice area and regurgitant volume. Apolipoprotein-A1 was an independent predictor of severe mitral regurgitation. Moreover, with ordinal logistic regression, apolipoprotein-A1 remained the only independent factor associated with mitral regurgitation. In addition, myxomatous mitral valves were studied by immunocytochemistry. We observed an increase of LC3, the marker of autophagy, in myxomatous mitral valves compared with healthy mitral valves. CONCLUSION: These potential biomarkers of mitral regurgitation highlighted different cellular processes that could be modified in myxomatous degenerescence: reverse cholesterol transport, antioxidant properties and autophagy. [less ▲]

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See detailMicrofluidic analytical techniques in drug discovery
Fillet, Marianne ULg

Conference (2015)

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See detailHepcidin determination in dried blood by microfluidic LC-MS/MS: comparison of DBS and volumetric absorptive microsampling for matrix effect and recovery.
Houbart, Virginie ULg; COBRAIVILLE, Gaël ULg; Servais, Anne-Catherine ULg et al

in Bioanalysis (2015), 7(21), 2789-99

BACKGROUND: Dried blood analysis experiences a growing interest due to practical, ethical and financial advantages compared with classical wet plasma or serum analysis. Besides classical DBS, new ... [more ▼]

BACKGROUND: Dried blood analysis experiences a growing interest due to practical, ethical and financial advantages compared with classical wet plasma or serum analysis. Besides classical DBS, new alternatives are commercialized as volumetric absorptive microsampling (VAMS) that are expected to overcome hematocrit influence. RESULTS: The feasibility of hepcidin (a peptide hormone) extraction and determination from DBS and VAMS blood sampling was investigated. Experimental design was used to determine the optimal extraction conditions. Matrix effect and extraction recovery were studied and a special attention was paid to phospholipid removal. CONCLUSION: The data suggest that the combination of VAMS and phospholipid removal plates provides low matrix effect and high sensitivity, and constitutes an easy and promising protocol for hepcidin analysis. [less ▲]

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See detailSimultaneous determination of insulin and its analogues in pharmaceutical formulations by micellar electrokinetic chromatography
Lamalle, Caroline ULg; Servais, Anne-Catherine ULg; RADERMECKER, Régis ULg et al

in Journal of Pharmaceutical & Biomedical Analysis (2015)

A simple and efficient MEKC method was developed to simultaneously determine human insulin, its five analogues, the main degradation products and the excipients usually present in injection formulations ... [more ▼]

A simple and efficient MEKC method was developed to simultaneously determine human insulin, its five analogues, the main degradation products and the excipients usually present in injection formulations. A very fast method with a total analysis time of 3 min was then successfully validated for the analysis of human insulin and the quality control of different commercial formulations was carried out. [less ▲]

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See detailTSC2 and 14-3-3 proteins down-regulate a RIP3 dependent PDT-induced Necroptosis in Glioblastoma
Fettweis, Grégory ULg; Coupienne, Isabelle; Fillet, Marianne ULg et al

Poster (2014, October)

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See detailComprehensive plasma profiling for the characterization of graft-versus-host disease biomarkers
De Bock, Muriel; BEGUIN, Yves ULg; Leprince, Pierre ULg et al

in Talanta (2014), 125

Acute graft-versus-host disease (aGVHD) remains a life-threatening complication of hematopoietic stem cell transplantation (HSCT), limiting its application. To optimize management of aGVHD and reduce ... [more ▼]

Acute graft-versus-host disease (aGVHD) remains a life-threatening complication of hematopoietic stem cell transplantation (HSCT), limiting its application. To optimize management of aGVHD and reduce therapy-related toxicity, early specific markers are needed. The main objective of this study was thus to uncover diagnostic biomarkers comparing plasma protein profiles of patients at the time of acute GVHD diagnosis and of patients undergoing HSCT without aGVHD. Additional analysis of samples taken 15 days before aGVHD diagnosis was also performed to evaluate the potential of the newly discovered biomarkers for early diagnosis. To extract a maximum of information from plasma samples, we used three complementary proteomic approaches, namely 2D-DIGE, SELDI-TOF-MS and 2D-LC-MSE. We identified and confirmed by means of a independent techniques, the differential expression of several proteins indicating significantly increased inflammation response and disturbance in the coagulation cascade. The variation of these proteins was already observed 15 days before GVHD diagnosis, suggesting the potential early detection of the disease before symptoms appearance. Finally, logistic regression analysis determines a composite biomarker panel comprising fibrinogen, fragment of fibrinogen beta chain, SAA, prothrombin fragments, apolipoprotein A1 and hepcidin that optimally discriminated patients with and without GVHD. The area under the receiver operating characteristic curve distinguishing these 2 groups was 0.95. [less ▲]

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See detailGalectin-3: a new promising cardiac biomarker in sports endurance?
LE GOFF, Caroline ULg; Devaux, Séverine; BREVERS, Eric ULg et al

in Cardiovascular Research (2014, July), 103(Supplement 1), 255

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See detailFluoxetine and norfluoxetine quantitation in rat serum by LC-chip-MS/MS
Houbart, Virginie ULg; Charlier, Thierry; Pawluski, Jodi et al

Poster (2014, June)

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See detailQuantitation of hepcidin in dried blood spots by microfluidic LC-chip-MS/MS
Houbart, Virginie ULg; Servais, Anne-Catherine ULg; Fillet, Marianne ULg

Conference (2014, June)

Microfluidic LC on a chip coupled to tandem mass spectrometry has been shown to be a sensitive tool for the quantitative analysis of small molecules and peptides. A major advantage of microfluidic ... [more ▼]

Microfluidic LC on a chip coupled to tandem mass spectrometry has been shown to be a sensitive tool for the quantitative analysis of small molecules and peptides. A major advantage of microfluidic separation techniques is the small requirements in terms of sample volume. This feature is particularly valuable for applications in the context of studies on small laboratory animals like rodents, or less-invasive clinical routine tests in patients, especially for paediatric applications. We previously developed a quantitation method for hepcidin, a peptide biomarker, in human plasma using µ-SPE (miniaturised solid-phase extraction) as sample preparation technique. The developed method allowed the quantitation of hepcidin using only 50 µL plasma with an excellent sensitivity. However, this efficient technique is quite expensive and time-consuming. To further reduce sample volume needs and simplify sample preparation, we developed a preparation method based on dried blood spot (DBS) analysis. In this case, a precise volume of blood (< 20 µL) is collected with a micro-capillary, dispensed on a paper card and dried. The obtained blood spot is then punched from the card, extracted in an appropriate solvent, and analysed by microfluidic LC-chip coupled to mass spectrometry. Method development was assisted by experimental design to optimise blood deposition and extraction conditions to ensure maximal analyte recovery while reducing interference co-extraction. Different ways of internal standard addition were also evaluated. In this study, the results obtained using µ-SPE and DBS are compared in terms of ease-of-use, throughput, sample requirements, cost, sensitivity and robustness. [less ▲]

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See detailDéveloppement de méthodes séparatives pour détecter la contrefaçon de molécules biosynthétiques comme l'insuline et les GHRP
Lamalle, Caroline ULg; Baptiste, Emeline; Marini Djang'Eing'A, Roland ULg et al

in Spectra Analyse (2014), 43

Counterfeiting is a widespread problem in the world. The medicines, like insulin or GHRP, need a strict quality control. Capillary electrophoresis and liquid chromatography methods were developed to ... [more ▼]

Counterfeiting is a widespread problem in the world. The medicines, like insulin or GHRP, need a strict quality control. Capillary electrophoresis and liquid chromatography methods were developed to analyze these peptides. The human insulin and its different analogues (lispro, aspart, glulisin, glargin and detemir) were separated by MEKC within 15 minutes. The GHRP-2 and -6 were separated by HPLC also in 15 minutes. Several samples of GHRP-6 were analyzed and non-compliances were reported. These analytical approaches seem to be promising to fight against the counterfeiting of such medicines. [less ▲]

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See detailCardiac troponins and natriuretic peptides in runners: useful for cardiac risk screening ?
LE GOFF, Caroline ULg; Kaux, Jean-François ULg; Fillet, Marianne ULg et al

in British Journal of Sports Medicine (2014, April), 48(7), 173

Background Cardiac troponins (cTn) are considered as the best biomarkers for detection of myocardial cell injury and NT-proBNP as the best for the cardiac insufficiency. Objective Our aim was to compare ... [more ▼]

Background Cardiac troponins (cTn) are considered as the best biomarkers for detection of myocardial cell injury and NT-proBNP as the best for the cardiac insufficiency. Objective Our aim was to compare cTnT and NT-proBNP levels before and after the stress tests, in sportive subjects. Design Prospective, cohort study. Setting Amateur marathon runners and ultrarunners. Patients 28 subjects (26 men, 42.5±11 yrs) were enrolled. Interventions Subjects ran the Maasmarathon (42.195 kilometers) and 33 subjects (33 men, 45.7±9.3 yrs) ran the Ultratour of Liège (Belgium; 67 km). All subjects gave their informed consent. We took blood sample before (T0), just after (T1) and 3 hours after the race (T3). Main outcome measurements cTnT concentrations were measured by high sensitive methods (hsTnT, Roche Diagnostics) on heparin plasma. The NT-proBNP was also determined with the kit Roche on heparin plasma. All statistical analyses were performed using Medcalc version 8.1 for Windows. P-value <.01 was regarded as statistically significant. Results A significant difference between hsTnT concentrations at T0 and T1 (P<.001), and between T0 and T3 (P<.001) for NT-proBNP have been observed, but not between T1 and T3. This observation appeared only after a strenuous exercise. However, up to now this type of exercise is not reproducible easily in a laboratory. Moreover, nobody knows if these observations would have cardiac consequences at long terms. Conclusion Measurement of cardiac troponins by high sensitive methods allows detecting significant release of biomarkers from the heart during exercise. The value of NT-proBNP are also significant but less than TnThs. We think that the TnThs could be an interesting tool in the future to help sport medicine to detect risk of developing a cardiac problem in the future or a sudden death. [less ▲]

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See detailTwo biomarkers for the screening of cardiac risk among runners ?
LE GOFF, Caroline ULg; Kaux, Jean-François ULg; Fillet, Marianne ULg et al

in British Journal of Sports Medicine (2014, April), 48(7), 174

Background Heart-type fatty acid-binding protein (H-FABP) is a low molecular weight protein involved in the intracellular uptake and buffering of long chain fatty in the myocardium. Troponin T is a ... [more ▼]

Background Heart-type fatty acid-binding protein (H-FABP) is a low molecular weight protein involved in the intracellular uptake and buffering of long chain fatty in the myocardium. Troponin T is a component of the contractile apparatus of the striated musculature. Both are early markers for acute coronary syndrome. Objective The aim of our study was to compare the results obtained with the H-FABP and the highly sensitive cardiac troponins (hsTnT) and to test their cardiospecificity in healthy runners. Design Prospective, cohort study. Setting Amateur marathon runners. Patients 23 runners (marathon) were enrolled. Interventions We drowned blood samples at three times: just before (T0), just after (T1), and three hours after the end of the race (T3). Main outcome measurements H-FABP and hs-TnT were performed according to the manufacturer's instructions. A linear regression was calculated to observe if there is any correlation between the two biomarkers. Values above the 95th percentile for H-FABP (2.5 ng/mL) and the 99th percentile for hsTnT (14 ng/L) were considered as positive. Results At T0, none of the subjects were positive for hsTnT but 35% were positive for H-FABP; at T1, 83% for hsTnT and 100% for H-FABP; at T3, 83% for hsTnT and 96% for H-FABP. At T0, the regression equation was H-FABP T0=3.9454–0.1001×hsTnT T0; at T1: H-FABP T1=51.838–1.7026×hsTnT T1; at T3: H-FABP T3=47.977–1.6193×hsTnT T3. No correlation was observed between the 2 biomarkers. Conclusion We observed a significant increase of H-FABP and hsTnT in runners. These markers are independent to each other. These values could biologically correspond to a heart ischemia. These biomarkers could be helpful for the screening of cardiac risk among runners. [less ▲]

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See detailAvancées récentes en métabolomique et protéomique
Fillet, Marianne ULg

Conference (2014)

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See detailMicellar electrokinetic chromatography against the counterfeiting of insulin formulations
Lamalle, Caroline ULg; Servais, Anne-Catherine ULg; Crommen, Jacques ULg et al

Poster (2014)

Insulin plays an important role in the homeostasis of blood glucose concentration. A deficiency of this hormone causes diabetes, which can be treated by subcutaneous injection of synthetic insulin ... [more ▼]

Insulin plays an important role in the homeostasis of blood glucose concentration. A deficiency of this hormone causes diabetes, which can be treated by subcutaneous injection of synthetic insulin. Besides human regular insulin, several modified analogues have been developed to accelerate (Lispro, Aspart, Glulisin) or delay (Glargin, Detemir) its absorption. Moreover, protamine is sometimes associated with human, Lispro or Aspart insulin to give a crystalline form, which delays the action of insulin, providing it with a prolonged absorption profile after injection. Diabetes is one of the most common metabolic diseases in the world; its prevalence increases continuously. A lot of patients are therefore concerned with the treatment, which is relatively expensive and requires a prescription. Some pharmaceutical formulations can sometimes be found without prescription on the parallel market but the risk of drug counterfeiting is then considerably increased. The poor quality of these drugs can lead to harmful consequences for the public health. It is therefore essential to develop a suitable method for the identification and quantification of human insulin and its analogues inside formulations. Ortner et al. [1] have already proposed micellar electrokinetic chromatography (MEKC) methods to detect simultaneously human insulin and its five analogues but no quantitative applications were presented. Furthermore, formulations containing protamine were not tested so we included them in our study. The first optimisation step involved the sample preparation procedure. An acidic sample solution (10 mM HCl) was finally selected to solubilise protamine and Glargin. Then the background electrolyte composition was investigated to separate the components present in the formulations. A basic buffer (50 mM ammonium acetate pH 9) was selected, providing an important and stable electroosmotic flow, a negative charge to the insulins and avoiding any adsorption to the capillary wall. The addition of sodium dodecylsulfate (SDS) and acetonitrile (ACN) was also found crucial for selectivity. With 50 mM SDS and 15% ACN the six insulins and the two major excipients (phenol and meta-cresol) were fully separated within 15 minutes. This method was then entirely validated for the human insulin and the quality control of related formulations was performed. The next step will be the validation and the quantification of the other analogues. [less ▲]

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See detailDevelopment and validation of a liquid chromatographic method for the stability study of a pharmaceutical formulation containing voriconazole using cellulose tris(4-chloro-3-methylphenylcarbamate) as chiral selector and polar organic mobile phases.
Servais, Anne-Catherine ULg; Moldovan, Radu-Cristian ULg; Farcas, Elena ULg et al

in Journal of chromatography. A (2014), 1363

The ophthalmic solution of voriconazole, i.e. (2R,3S)-2-(2,4-difluorophenyl)-3-(5-fluoropyrimidin-4-yl)-1-(1H-1,2,4-triazol-1-y l)butan-2-ol, made from an injection formulation which also contains ... [more ▼]

The ophthalmic solution of voriconazole, i.e. (2R,3S)-2-(2,4-difluorophenyl)-3-(5-fluoropyrimidin-4-yl)-1-(1H-1,2,4-triazol-1-y l)butan-2-ol, made from an injection formulation which also contains sulfobutylether-beta-cyclodextrin sodium salt as an excipient (Vfend((R))), is used for the treatment of fungal keratitis. A liquid chromatographic (LC) method using polar organic mobile phase and cellulose tris(4-chloro-3-methylphenylcarbamate) coated on silica as chiral stationary phase was successfully developed to evaluate the chiral stability of the ophthalmic solution. The percentage of methanol (MeOH) in the mobile phase containing acetonitrile (ACN) as the main solvent significantly influenced the retention and resolution of voriconazole and its enantiomer ((2S,3R)-2-(2,4-difluorophenyl)-3-(5-fluoropyrimidin-4-yl)-1-(1H-1,2,4-triazol-1- yl)butan-2-ol). The optimized mobile phase consisted of ACN/MeOH/diethylamine/trifluoroacetic acid (80/20/0.1/0.1; v/v/v/v). The method was found to be selective not only regarding the enantiomer of voriconazole but also regarding the specified impurities described in the monograph from the European Pharmacopoeia. The LC method was then fully validated applying the strategy based on total measurement error and accuracy profiles. Under the selected conditions, the determination of 0.1% of voriconazole enantiomer could be performed. Finally, a stability study of the ophthalmic solution was conducted using the validated LC method. [less ▲]

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