HPLC-DAD and HPLC-ESI-MS analyses of Tiliae flos and its preparations.

In the present study extensive HPLC-DAD, HPLC-ESI-MS and NMR analyses were undertaken in the aqueous preparations (decoctions, infusions) and tinctures of Tilia platyphyllos Scop inflorescences. The aim of this work was to examine in depth the qualitative and quantitative profile of the investigated preparations, which find until today wide applications in pharmaceutical and cosmetic industry, and to propose a validated method for their quality control. An HPLC-DAD-ESI-MS method was developed and optimised for the quantitative determination of the constituents. Marker constituents of Tiliae flos are the flavonoids, while the volatile content is also used for the quality control. However, the analyses of the non-volatile fraction gave complex chromatographic fingerprints containing simple phenolics and low molecular weight procyanidins. The use of different HPLC columns permitted a good separation of the constituents and enabled their quantitation, while HPLC-MS analyses permitted the detection of procyanidin oligomers. Overall, 31 constituents were detected and identified. Extensive preparative chromatographic investigations and 2D-NMR analyses allowed the characterisation of procyanidins as epicatechin derivatives. Finally, the HPLC method was validated and complied with ICH guidelines. This is the first report of detailed analysis of the chemical composition of Tiliae flos.

Stability study and lyophilization of vitamin E-loaded nanocapsules prepared by membrane contactor.

In this research, we studied the accelerated stability of vitamin E-loaded nanocapsules (NCs) prepared by the nanoprecipitation method. Vitamin E-loaded NCs were optimized firstly at the laboratory scale and then scaled up using the membrane contactor technique. The optimum conditions of the membrane contactor preparation (pilot scale) produced vitamin E-loaded NCs with an average size of 253 nm, polydispersity index 0.19 and a zeta potential -16 mV. The average size, polydispersity index and zeta potential values were 185 nm, 0.12 and -15 mV, respectively for the NCs prepared at laboratory scale. No significant changes were noticed in these values after 3 and 6 months of storage at high temperature (40±2 °C) and relative humidity (75±5%) in spite of vitamin E sensitivity to light, heat and oxygen. The entrapment efficiency of NCs prepared at pilot scale was 97% at the beginning of the stability study, and became (95%, 59%) after 3 and 6 months of storage, respectively. These values at lab-scale were (98%, 96%, and 89%) at time zero and after 3 and 6 months of storage, respectively. This confirms the ability of vitamin E encapsulation to preserve its stability, which is one major goal of our work. Lyophilization of the optimized formula at lab-scale was also performed. Four types of cryoprotectants were tested (poly(vinyl pyrrolidone), sucrose, mannitol, and glucose). Freeze-dried NCs prepared with sucrose were found acceptable. The other lyophilized NCs obtained at different conditions presented large aggregates.

Preparation of vitamin E loaded nanocapsules by the nanoprecipitation method: from laboratory scale to large scale using a membrane contactor.

Vitamin E or α-tocopherol is widely used as a strong antioxidant in many medical and cosmetic applications, but is rapidly degraded, because of its light, heat and oxygen sensitivity. In this study, we applied the nanoprecipitation method to prepare vitamin E-loaded nanocapsules, at laboratory-scale and pilot-scale. We scaled-up the preparation of nanocapsule with the membrane contactor technique. The effect of several formulation variables on the vitamin E-loaded nanocapsules properties (mean diameter, zeta potential, and drug entrapment efficiency) was investigated. The optimized formulation at laboratory-scale and pilot-scale lead to the preparation of vitamin E-loaded nanocapsules with mean diameter of 165 and 172 nm, respectively, and a high encapsulation efficiency (98% and 97%, respectively).

Epigenetic plasticity of hTERT gene promoter determines retinoid capacity to repress telomerase in maturation-resistant acute promyelocytic leukemia cells.

The expression of hTERT gene, encoding the catalytic subunit of telomerase, is a feature of most cancer cells. Changes in the chromatin environment of its promoter and binding of transcriptional factors have been reported in differentiating cells when its transcription is repressed. However, it is not clear whether these changes are directly involved in this repression or only linked to differentiation. In a maturation-resistant acute promyelocytic leukemia (APL) cell line (NB4-LR1), we have previously identified a new pathway of retinoid-induced hTERT repression independent of differentiation. Using a variant of this cell line (NB4-LR1(SFD)), which resists to this repression, we show that although distinct patterns of histone modifications and transcription factor binding at the proximal domain of hTERT gene promoter could concur to modulate its expression, this region is not sufficient to the on/off switch of hTERT by retinoids. DNA methylation analysis of the hTERT promoter led to the identification of two distinct functional domains, a proximal one, fully unmethylated in both cell lines, and a distal one, significantly methylated in NB4-LR1(SFD) cells, whose methylation was further re-enforced by retinoid treatment. Interestingly, we showed that the binding to this distal domain of a known hTERT repressor, WT1, was defective only in NB4-LR1(SFD) cells. We propose that epigenetic modifications targeting this distal region could modulate the binding of hTERT repressors and account either for hTERT reactivation and resistance to retinoid-induced hTERT downregulation.

Spectrophotometric Determination of Alfuzosin HCl in Pharmaceutical Formulations with some Sulphonephthalein Dyes.

Bromocresol purple (BCP), bromophenol blue (BPB) and bromothymol blue (BTB) were used to determine alfuzosin hydrochloride either in pure form or in pharmaceutical formulations. Alfuzosin was extracted as an ion-pair complex from sample solution containing KCl-HCl buffer pH2.2, 2.4 and 2.6 into CHCl3 and the absorbance was measured at 407, 413 and 412nm with use of the cited reagents, respectively. The analytical parameters and their effects on the reported systems are investigated. The reactions were extremely rapid at room temperature and the absorbance values remains unchanged up to 24 h. Beer's law was obeyed in the concentration ranges 1.20-38.3, 0.85-46.0 and 0.63-34.0 µg/ml and detection limits were 0.28, 0.24 and 0.18 µg/ml with BCP, BPB and BTB, respectively. Recoveries were 98.80-101.33%. Interferences of the other ingredients and excipients were not observed. The proposed method is simple, fast and sensitive, and the first reported extractive method for the determination of alfuzosin in commercial tablets.