Mireille Thomassin

Comparison of quantitative high performance thin layer chromatography and the high performance liquid chromatography of parabens

A method is described for the densitometric determination of the p-hydroxybenzoic esters and p-hydroxybenzoic acid in mixtures or in drugs. This method is compared with the one used in high performance liquid chromatography (HPLC). The calibration curves were linear in interval 0.250-3.60 mumol ml-1 per 200 nl per spot. The limit of detection and the relative standard deviation (RSD) are higher than in HPLC (RSD is 6% in HPTLC. 3% in HPLC; limit of detection about 40 pmol in HPTLC and 25 pmol in HPLC) but HPTLC quantitative determination of parabens in drugs is faster.

Triazinic herbicide determination by gas chromatography-mass spectrometry in breast milk

A solid-phase extraction procedure using a graphitized carbon black cartridge for extraction and cleaning of a series of five triazines (atrazine, deethylatrazine, deisopropylatrazine, ametryne and prometryne) from breast milk samples was developed. Using a chemometric methodology, the optimisation of both the analysis time and the triazinic herbicide separation by gas chromatography-mass spectrometry (GC-MS) was then carried out with only 18 experiments. Detection and quantification limits for 1ml breast milk sample were, respectively, 0.3 and 1 ppb for each studied compound. The variation coefficients were less than 5% over the concentration range from 1 to 100 ppb. The accuracy was between 98.63 and 104.62% for each triazinic herbicide. The recovery was between 58.64 and 63.22% for the concentration range from 1 to 100 ppb for each triazinic herbicide. The assay was successfully applied to the analysis of several breast milk samples.

Analysis of the progesterone displacement of its human serum albumin binding site by β-estradiol using biochromatographic approaches: effect of two salt modifiers

The mechanisms of (i) the binding of two sex-hormones (i.e. progesterone and beta-estradiol) to human serum albumin (HSA) and (ii) the progesterone displacement of its HSA binding cavity by beta-estradiol were studied by biochromatography using three different methods. In the first time, zonal elution method was used to prove the direct competition effect between the two sex-hormone. In the second time, the competition effect between beta-estradiol and progesterone to bound on the same HSA site was analysed by the competitive bi-Langmuir approach. Finally, the thermodynamic data of these two binding processes were studied. The Gibbs free energy value (Delta(approximately)G degrees) of the displacement equilibrium was negative demonstrating that beta-estradiol displaced progesterone of its HSA binding cavity. Moreover, the effect of two chloride modifiers (i.e. Na(+), Mg(2+)) on these two binding processes were analysed. Results showed that in the salt biological concentration ranges, the Mg(2+) cation enhanced strongly the bioavailable progesterone, whereas the Na(+) cation interacted slowly on the progesterone displacement of its HSA binding site by beta-estradiol. This study showed that it must be useful to carry out more in vivo test on the magnesium supplementation effect for women who suffer from estrogen dominance syndrome.

Role of the magnesium cation on antihypertensive molecule-human serum albumin binding: affinity chromatography approach.

The role of the Mg2+ cation on antihypertensive molecule binding on human serum albumin (HSA) was studied by affinity chromatography. The thermodynamic data corresponding to this binding were determined for a wide range of Mg2+ concentrations (c). For the nifedipine molecule, an increase in the Mg2+ concentration produced a decrease in binding due to a decrease in the electrostatic interactions. For verapamil and diltiazem, which have the highest solvent accessible surface area, the solute binding on HSA was divided into two Mg2+ concentration regions. For a low c value below c(c) (approximately 1.6 mmol/l), the binding dependence with c was similar to that of nifedipine. For c above c(c) the hydrophobic effect created in the bulk solvent associated with a decrease in the van der Waals interactions between the solute molecule and the HSA implied a decrease in its binding. These results showed that for patients with hypertension, an Mg2+ supplementation during treatment with these antihypertensive molecules can increase the active pharmacological molecule concentration.

Chiral discrimination of dansyl-amino-acid enantiomers on teicoplanin phase: sucrose-perchlorate anion dependence

Abstract The chiral recognition mechanism for a series of d , l -dansyl-amino-acids (test solutes) on a teicoplanin stationary phase was investigated in reversed phase liquid chromatography (RPLC). The effect of both a surface tension modifier (sucrose) and a chaotropic agent (perchlorate anion) on the enantiomeric separation was studied by varying their concentration, c , in the mobile phase. The thermodynamic data supported the fact that the sucrose molecule acted only on the hydrophobic part of the interaction teicoplanin/dansyl-amino-acid and not on the specific chiral part. It was demonstrated that the enhancement of the separation factor observed as the perchlorate salt concentration increased in the mobile phase was enthalpically controlled owing to stereoselective bonding interactions. Such behavior was used to optimize the chromatographic conditions for separation of dansyl-amino-acids on teicoplanin.

PECULIARITIES OF THE RETENTION MECHANISM OF CIRCULAR AND LINEAR DNA FRAGMENTS USING NON-EQUILIBRIUM CHROMATOGRAPHY

Two different methods have been used to investigate the retention mechanism of a series of DNA fragments in non equilibrium chromatography (NEC) over a range of column temperatures (T) and with different mobile phase flow-rates (F). The first approach was the separate study of each factor affecting the retention mechanism; the second method was the simultaneous variation of all these factors. The apparent relative retention time of each DNA fragment was used as a retention marker. The data obtained showed that the retention mechanism of the DNA fragment was dependent on the F and T values, and also on the DNA form, i.e., linear or circular form. For low F values, the retention mechanism was based on the classical hydrodynamic regime (HDC) specific to the circular form. On the contrary, for the linear forms, their retention mechanism was based on a slalom chromatographic process (SC) specifically for high F values at low column temperature. This confirms that the SC and HDC modes are interconnected and that the HDC⇄SC transition exists and was clearly visualized. In addition, using a simplex process connected to the chemometric methodology, the optimal conditions for F and T were calculated to obtain the more efficient separation of the linear DNA forms in a minimum analysis time.

A biochromatographic framework to evaluate the calcium effect on the antihypertensive molecule-human serum albumin binding.

The Ca(2+) cation effect on the antihypertensive molecule binding on human serum albumin (HSA) was studied by biochromatography. The thermodynamic parameters corresponding to this binding were determined for a wide range of Ca(2+) concentration (x). For the two antihypertensive molecules under study, their binding to HSA can be divided into two Ca(2+) cation concentration regions due to a HSA phase transition. This result was confirmed by an enthalpy-entropy investigation. For a low x value (below x(c)=1.6 mmol l(-1)), the HSA cavity was in an ordered solid-like state leading to an increase in the interactions between the antihypertensive drugs and the HSA cavity and consequently, a solute-HSA affinity increase. For x above x(c), the HSA cavity was in a disordered solid-like state, implying a decrease in the antihypertensive drug-HSA binding.

Magnesium effect on the acetylcholinesterase inhibition mechanism: A molecular chromatographic approach

The acetylcholinesterase enzyme (AChE) was immobilized on a chromatographic support to study the effect of magnesium on the binding mechanism of five AChE inhibitors (donepezil, tacrine, galanthamine, physostigmine and huperzine). The determination of the enthalpy and entropy changes of this binding at different magnesium concentration values suggested that van der Waals interactions and hydrogen bonds predominated the donepezil and tacrine association to AChE. As well, hydrophobic and electrostatic forces seemed to be the major interactions controlling the huperzine, galanthamine and physostigmine association with AChE. In addition, it appeared that magnesium cation increased the binding affinity of galanthamine and physostigmine to the active site gorge of AChE. A comparison of the inhibitors hydrophobicity to their relative bound percentage with AChE showed an affinity enhanced with the increase in the molecule hydrophobicity and confirmed that the hydrophobic forces played an important role in the AChEI-AChE binding process. This novel biochromatographic column could be useful to find a specific inhibitor for this enzyme and so open new perspectives to be investigated.

Association mechanism of four acetylcholinesterase inhibitors (AChEIs) with human serum albumin: a biochromatographic approach.

In this work, the interaction of a series of acetylcholinesterase inhibitors (AChEIs; donepezil, galanthamine, huperzine and neostigmine) with human serum albumin (HSA) immobilized on porous silica particles was studied using a biochromatographic approach. For all the tested AChEI molecules, linear retention plots were observed at all temperatures. An analysis of the thermodynamics (i.e. enthalpy (DeltaH degrees ), entropy ((S degrees *)) of the interaction of the AChEI molecules with the immobilized human serum albumin was also carried out. The (H degrees and (S degrees * values for donepezil, galanthamine and neostigmine, were negative due to van der Waals interactions and hydrogen bonding which govern this association with albumin. Whereas the positive values of (H degrees and (S degrees * of huperzine binding on HSA indicated a predominance of hydrophobic interactions. The association of AChEIs with HSA was increased linearly with pH. A comparative thermodynamic study with benzodiazepine molecules was also done to determine the potential binding site of these drugs on HSA.

A novel cyclic hexapeptide as chiral selector: Application for HPLC separation of a series of dansyl amino and arylalkanoic acids.

In this paper, the synthesis of a cyclic hexapeptide molecule was presented and evaluated for the enantiomer separation of a series of dansyl amino and arylalkanoic acids using high performance liquid chromatography (HPLC). It was clearly vizualized that this chiral selector allowed the separation of a great number of enantiomer pairs. The influences of the size and the hydrogen bonding donor (HBD) parameter of the organic modifier (OM) (THF (HBD=0.00), propan-2-ol (HBD=0.33), methanol (HBD=0.43)) added in the mobile phase were also investigated on both the enantiomer-chiral selector association and enantioseparation.