The increase of fungal resistance to drugs, such as azole family, gave rise to the development of new antifungals. In this context, echinocandins emerged as a promising alternative for antifungal ...therapies. Following the commercialization of caspofungin in 2001, echinocandins became the first-line therapy for invasive candidiasis in different patient populations. The quantification of these drugs has gained importance since pharmacokinetic/pharmacodynamic and resistance studies are a paramount concern. This fact has led us to exhaustively examine the methodologies used for the analysis of echinocandins in biological fluids, which are mainly based on LC coupled to different detection techniques. In this review, we summarize the analytical methods for the quantification of echinocandins focusing on sample treatment, chromatographic separation and detection methods.
Neonatal sepsis is a major cause of infant mortality in developing countries because of delayed injectable treatment, making it urgent to develop noninjectable formulations that can reduce treatment ...delays in resource-limited settings. Ceftriaxone, available only for injection, needs absorption enhancers to achieve adequate bioavailability via nonparenteral administration. This article presents all available data on the nonparenteral absorption of ceftriaxone in humans and animals, including unpublished work carried out by F. Hoffmann-La Roche (Roche) in the 1980s and new data from preclinical studies with rabbits, and discusses the importance of these data for the development of noninjectable formulations for noninvasive treatment. The combined results indicate that the rectal absorption of ceftriaxone is feasible and likely to lead to a bioavailable formulation that can reduce treatment delays in neonatal sepsis. A bile salt, chenodeoxycholate sodium salt (Na-CDC), used as an absorption enhancer at a 125-mg dose, together with a 500-mg dose of ceftriaxone provided 24% rectal absorption of ceftriaxone and a maximal plasma concentration of 21 µg/ml with good tolerance in human subjects. The rabbit model developed can also be used to screen for the bioavailability of other formulations before assessment in humans.
A simple method using gas chromatography–mass spectrometry was applied to analyse structures of ceramides. Identification of trimethylsilylated ceramides were obtained in short analysis times ...(derivatization of ceramides in 30 min at room temperature and 20 min gas chromatography mass spectrometry run) even for complex mixtures. For example in ceramide Type III, 18 peaks were observed which represent 27 various structures. The coeluted compounds were ceramides containing the same functional groups and the same carbon number but with a different distribution on the two alkyl chains of the molecule. They were accurately differentiated by mass spectrometry. Therefore, 83 structures of trimethylsilylated ceramides were identified in 11 different commercial mixtures. For 52 structures of these, mass spectral data were not described in the literature, neither full mass spectra nor characteristic fragments.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
For the last few decades, many efforts have been made in developing cell culture methods in order to overcome the biological limitations of the conventional two-dimensional culture. This paradigm ...shift is driven by a large amount of new hydrogel-based systems for three-dimensional culture, among other systems, since they are known to mimic some living tissue properties. One class of hydrogel precursors has received interest in the field of biomaterials, low-molecular-weight gelators (LMWGs). In comparison to polymer gels, LMWG gels are formed by weak interactions upon an external trigger between the molecular subunits, giving them the ability to reverse the gelation, thus showing potential for many applications of practical interest. This study presents the use of the nucleoside derivative subclass of LMWGs, which are glyco-nucleo-bola-amphiphiles, as a proof of concept of a 3D cell culture scaffold. Physicochemical characterization was performed in order to reach the optimal features to fulfill the requirements of the cell culture microenvironment, in terms of the mechanical properties, architecture, molecular diffusion, porosity, and experimental practicality. The retained conditions were tested by culturing glioblastoma cells for over a month. The cell viability, proliferation, and spatial organization showed during the experiments demonstrate the proof of concept of nucleoside-derived LMWGs as a soft 3D cell culture scaffold. One of the hydrogels tested permits cell proliferation and spheroidal organization over the entire culture time. These systems offer many advantages as they consume very few matters within the optimal range of viscoelasticity for cell culture, and the thermoreversibility of these hydrogels permits their use with few instruments. The LMWG-based scaffold for the 3D cell culture presented in this study unlocked the ability to grow spheroids from patient cells to reach personalized therapies by dramatically reducing the variability of the lattice used.
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IJS, KILJ, NUK, PNG, UL, UM
Ceftriaxone, a third generation cephalosporin, has a wide antibacterial spectrum that has good CNS penetration, which makes it potentially suitable for initial treatment of severe neonatal pediatric ...infections providing suitable formulation. We evaluated its physicochemical and technical characteristics to assess its potential for development as a non-parenteral dosage form. As ceftriaxone is marked only for injectable use, these data are not available. Using HPLC and Karl Fischer titration, sensitivity of ceftriaxone to water, feasibility and impact of pharmaceutical processes and compatibility with common pharmaceutical excipients were assessed. X-ray diffraction studies gave deeper insight into the mechanisms involved in degradation. Chemometrical analysis of near infrared spectra enabled classification of ceftriaxone powder according to exposure conditions or processes applied. The results showed that ceftriaxone was not highly hygroscopic, could be processed in all climatic zones, but should be packaged protected against humidity. Controlling water presence in formulation was shown critical, as ceftriaxone degraded in the presence of water content above 2.4% w/w. To improve flowability, a critical parameter for dry dosage form development, granulation (wet and dry techniques, providing complete drying and moderate force compaction respectively) was shown feasible. Compression with moderate forces was possible, but grinding and high compression forces significantly affected long term ceftriaxone stability and should be avoided. Based on these results, development of ceftriaxone non-parenteral solid or liquid non-aqueous forms appears feasible.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK, ZRSKP
•An RP-HPLC/DAD method was developed for paracetamol quantification in cell culture fluid.•Sample and separation methods were performed in green conditions.•Application of paracetamol in BBB model ...allows determining its transcellular passage.
A Reversed Phase-High Performance Liquid Chromatography/Diode Array Detection method was developed and validated for paracetamol quantification in cell culture fluid from an in vitro Blood Brain Barrier model. The chromatographic method and sample preparation were developed using only aqueous solvents. The column was a XTerra RP18 150×4.6mm, 3.5μm with a guard column XTerra RP18 20×4.6mm, 3.5μm at 35°C and the mobile phase was composed by 100% formate buffer 20mM at pH 4 and flow rate was set at 1mL/min. The detection was at 242nm. The sample was injected at 10μL. Validation was performed using the accuracy profile approach. The analytical procedure was validated with the acceptance limits at ±10% over a range of concentration from 1 to 58mgL−1. The procedure was then used in routine to determine paracetamol concentration in a brain blood barrier in vitro model. Application of the Unither paracetamol formulation in Blood Brain Barrier model allowed the determination and comparison of the transcellular passage of paracetamol at 37°C and 4°C, that excludes paracellular or non specific leakage.
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Pharmaceutical development and manufacturing process optimization work was undertaken in order to propose a potential paediatric rectal formulation of azithromycin as an alternative to existing oral ...or injectable formulations. The target product profile was to be easy-to-use, cheap and stable in tropical conditions, with bioavailability comparable to oral forms, rapidly achieving and maintaining bactericidal concentrations. PEG solid solution suppositories were characterized in vitro using visual, HPLC, DSC, FTIR and XRD analyses. In vitro drug release and in vivo bioavailability were assessed; a study in rabbits compared the bioavailability of the optimized solid solution suppository to rectal solution and intra-venous product (as reference) and to the previous, non-optimized formulation (suspended azithromycin suppository). The bioavailability of azithromycin administered as solid solution suppositories relative to intra-venous was 43%, which compared well to the target of 38% (oral product in humans). The results of 3-month preliminary stability and feasibility studies were consistent with industrial production scale-up. This product has potential both as a classical antibiotic and as a product for use in severely ill children in rural areas. Industrial partners for further development are being sought.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
A rapid and reliable capillary zone electrophoresis method was developed and validated for the assay of oseltamivir phosphate in capsules. Separation was carried out in fused silica capillary (60.2
...cm total length and 10.0
cm effective length, 75
μm i.d.) by applying a potential of −15
kV at 25
°C. The selected electrophoretic buffer consisted of 50
mM sodium phosphate, pH 6.3 (direct UV detection, 226
nm). A short electrophoretic analysis time (less than 1.5
min) was obtained using the short end injection mode. The method was validated in terms of specificity, linearity, precision and accuracy. The RSD values were 0.94 and 0.98% for repeatability and intermediate precision, respectively. Recovery determinations allowed the calculation of a confidence interval from 98.64 to 100.26% with a relative standard deviation value of 0.38%. LOD and LOQ were estimated at 0.97 and 3.24
μg/mL, respectively. The validated method was successfully applied to the determination of oseltamivir in three commercially available capsules (Tamiflu
®, Saiflu
® and Flufy
®). The results were in good agreement with those obtained by a HPLC method previously developed in our laboratory. This method presents advantages including short run time, simple and rapid sample preparation and no use of non-aqueous solvent throughout the analysis.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
Retention with porous graphitic carbon was investigated with 25 structures of fatty acid methyl esters (FAMEs) with two different mobile phases: CH
3CN:CHCl
3 60:40 (v/v) and CH
3OH:CHCl
3 60:40 ...(v/v) with both 0.1% triethylamine (TEA) and an equimolar amount of HCOOH. Preliminary results showed that the use of TEA/HCOOH led to the response increase of saturated FAMEs with evaporative light scattering detection. No increase was observed for unsaturated one. These modifiers may slightly reduce the retention of FAMEs but did not significantly modify the separation factor with porous graphitic carbon. Thermodynamic parameters were calculated for each structure using Van’t Hoff plot measured over the temperature range from 10 to 50
°C, with the both mobile phase conditions. All the studied compounds were found linked by the same retention mechanism on porous graphitic carbon. Quantitative
in silico analysis of the retention using a molecular mechanics calculation demonstrated a good correlation between the retention factors and the molecular interaction energy values (
r
>
0.93). Especially the Van der Waals energy was predominant, and the contribution of electrostatic energy was negligible for the quantitative analysis of the retention. The results indicate that Van der Waals force, hydrophobic interaction, is predominant for the retention of FAMEs on this packing material. The relative retention for highly unsaturated homologues can be changed by the selection of the weak solvent CH
3CN or CH
3OH. Then isomers differing only in the position of the carbon double bond on the alkyl chain can be separated and their behaviour is summarised as the closer the carbon double bonds to the FAME polar head, the more the retention decreases. Finally, the more important the number of carbon double bonds in the alkyl chain is, the smaller the retention is.
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