Minjie Zhao

Iodinated α-tocopherol nano-emulsions as non-toxic contrast agents for preclinical X-ray imaging

Micro-computed tomography (micro-CT) is an emerging imaging modality, due to the low cost of the imagers as well as their efficiency in establishing high-resolution (1-100 μm) three-dimensional images of small laboratory animals and facilitating rapid, structural and functional in vivo visualization. However use of a contrast agent is absolutely necessary when imaging soft tissues. The main limitation of micro-CT is the low efficiency and toxicity of the commercially available blood pool contrast agents. This study proposes new, efficient and non-toxic contrast agents for micro-CT imaging. This formulation consists of iodinated vitamin E (α-tocopheryl 2,3,5-triiodobenzoate) as an oily phase, formulated as liquid nano-emulsion droplets (by low-energy nano-emulsification), surrounded by a hairy PEG layer to confer stealth properties. The originality and strength of these new contrast agents lie not only in their outstanding contrasting properties, biocompatibility and low toxicity, but also in the simplicity of their fabrication: one-step synthesis of highly iodinated oil (iodine constitutes 41.7% of the oil molecule weight) and its spontaneous emulsification. After i.v. administration in mice (8.5% of blood volume), the product shows stealth properties towards the immune system and thus acts as an efficient blood pool contrast agent (t(1/2) = 9.0 h), exhibiting blood clearance following mono-exponential decay. A gradual accumulation predominantly due to hepatocyte uptake is observed and measured in the liver, establishing a strong hepatic contrast, persistent for more than four months. To summarize, in the current range of available or developed contrast agents for preclinical X-ray imaging, this agent appears to be one of the most efficient.

Microencapsulation of nanoemulsions: novel Trojan particles for bioactive lipid molecule delivery

Background Nanoemulsions consist of very stable nanodroplets of oil dispersed in an aqueous phase, typically below 300 nm in size. They can be used to obtain a very fine, homogeneous dispersion of lipophilic compounds in water, thus facilitating their handling and use in nanomedicine. However, the drawback is that they are suspended in an aqueous media. This study proposes a novel technique for drying lipid nanoemulsion suspensions to create so-called Trojan particles, ie, polymer microparticles (around 2 μm) which very homogeneously “entrap” the nano-oil droplets (around 150 nm) in their core. Methods Microencapsulation of the nanoemulsions was performed using a spray-drying process and resulted in a dried powder of microparticles. By using a low-energy nanoemulsification method and relatively gentle spray-drying, the process was well suited to sensitive molecules. The model lipophilic molecule tested was vitamin E acetate, encapsulated at around 20% in dried powder. Results We showed that the presence of nanoemulsions in solution before spray-drying had a significant impact on microparticle size, distribution, and morphology. However, the process itself did not destroy the oil nanodroplets, which could easily be redispersed when the powder was put back in contact with water. High-performance liquid chromatography follow-up of the integrity of the vitamin E acetate showed that the molecules were intact throughout the process, as well as when conserved in their dried form. Conclusion This study proposes a novel technique using a spray-drying process to microencapsulate nanoemulsions. The multiscale object formed, so-called Trojan microparticles, were shown to successfully encapsulate, protect, and release the lipid nanodroplets.

A new microfluidic setup for precise control of the polymer nanoprecipitation process and lipophilic drug encapsulation

A new microfluidic setup with impact-jet micromixers was built up and applied to control the nanoprecipitation process for generating polymeric nanoparticles and encapsulating a lipophilic drug. In contrast to a conventional nanoprecipitation processes in “bulk” phase, the microfluidic approach not only allows a continuous and controlled production of nanoparticles, but can also be used to manipulate and modify the nanoparticle size and the drug loading, by fine tuning the processing parameters. We developed a micromixer-assisted setup that can efficiently produce PMMA nanospheres, with a particle size of about 100 nm, and a narrow size distribution. Moreover, this setup enables a flow rate of the polymer phase as high as 1 mL min−1, opening the possibilities of large-scale production. The obtained nanoparticles can encapsulate high levels of a lipophilic drug (ketoprofen) and release it over 4 h. Finally, the solvent and non-solvent flow rates can be used to adjust the physicochemical and encapsulating/release properties of these nanosystems, opening new possibilities for nanoparticle production by nanoprecipitation.

Improvement of Total Lipid and Glycerophospholipid Recoveries from Various Food Matrices Using Pressurized Liquid Extraction

The extraction of three major phospholipid (PL) classes contained in soybean, egg yolk, calf brain, and ox liver was investigated by means of two methods. The PL amounts were evaluated. A new method, based on pressurized liquid extraction (PLE), was applied for total lipids (TL), including PL, extraction and compared with a standard liquid extraction method, a modified Folch method. The three PL classes (phosphatidylethanolamine (PE), phosphatidylinositol (PI), and phosphatidylcholine (PC)) that were recovered in the obtained TL extracts were quantified using HPLC with an evaporative light-scattering detector (ELSD). Using the PLE method, a single extraction allowed a recovery of more than 94% of TL and 96% of each PL class. Two successive extractions could achieve a total recovery of the three studied PL classes. With the modified Folch method, 77-83% of TL, 80-91% of PE, 82-94% of PC, and no more than 78% of PI could be achieved from various food matrices after one extraction. Four successive extractions were necessary to recover the whole TL content and each PL class. Results indicate that PLE is a rapid and efficient lipid extraction system for the broad range of plant and animal tissues.

Oxidative Stability at High Temperatures of Oleyol and Linoleoyl Residues in the Forms of Phosphatidylcholines and Triacylglycerols

An investigation was carried out into the stability of fatty acyl groups to heat-induced oxidative changes as affected by their chemical environment. The behavior of oleic and linoleic acyl groups when esterified in triacylglycerols (TAGs) and phosphatidylcholines (PCs) was evaluated. The monitoring of the oxidative degradation using liquid chromatography-mass spectrometry (LC-MS) showed that fatty acyl groups are less likely to be oxidized when in the form of PCs than when in the form of TAGs. In addition, oxidation products from PCs were more stable than those from TAGs. This finding strengthens the idea that the choline group in PCs increases the stability of fatty acyl groups to oxidation in comparison to TAGs.

Design, characterisation, and bioefficiency of insulin-chitosan nanoparticles after stabilisation by freeze-drying or cross-linking.

Insulin delivery by oral route would be ideal, but has no effect, due to the harsh conditions of the gastrointestinal tract. Protection of insulin using encapsulation in self-assembled particles is a promising approach. However, the lack of stability of this kind of particles in biological environments induces a low bioavailability of encapsulated insulin after oral administration. The objective of this work was to evaluate the effect of two stabilisation strategies alone or combined, freeze-drying and cross-linking, on insulin-loaded chitosan NPs, and to determine their bioefficiency in vitro and in vivo. NPs were prepared by complex coacervation between insulin and chitosan, stabilised either by cross linking with sodium tripolyphosphate solution (TPP), by freeze-drying or both treatments. In vitro bioefficiency NP uptake was evaluated by flow cytometry on epithelial models (Caco-2/RevHT29MTX (mucus secreting cells)). In vivo, NPs were injected via catheter in the peritoneum or duodenum on insulinopenic rats. Freeze-drying increased in size and charge (+15% vs control 412 ± 7 nm; + 36 ± 0.3 mV) in comparison with cross linking which decreased NP size (-25%) without impacting the NP charge. When combined the consecutive treatments reduced NPs size and increased charges as compared to standard level. Freeze drying is necessary to prevent the destruction of NP in intestinal environment in comparison with no freeze dryed one where 60% of NP were destroyed after 2h. Additionally freeze drying combined with cross linking treatments improved bioefficiency of NP with uptake in cell increased when mucus is present. Combination of both treatment showed a protection of insulin in vivo, with a reduction of glycemia when NPs were administrated. This work showed that the combination of freeze drying and cross linking treatment is necessary to stabilize (freeze-drying) and increase bioefficiency (cross-linking) of self assembled NP in the delivery of insulin in vitro and in vivo.

Icacina senegalensis (Icacinaceae), traditionally used for the treatment of malaria, inhibits in vitro Plasmodium falciparum growth without host cell toxicity

BackgroundWith the aim of discovering new natural active extracts against malaria parasites, Icacina senegalensis was selected after an ethnopharmacological survey conducted on plants used in traditional malaria treatment in Senegal.MethodsDifferent concentrations of the plant extract and fractions were tested on synchronized Plasmodium falciparum cultures at the ring stage using the parasite lactate dehydrogenase assay. Their haemolytic activity and in vitro cytoxicity were evaluated. The chromatographic profiles of active fractions were also established.ResultsThe plant extract and fractions revealed anti-plasmodial activity (IC50 < 5 μg/mL) with no toxicity (Selectivity indexes >10). The dichloromethane fraction showed stronger anti-plasmodial activity than the total extract.ConclusionAnti-plasmodial activity and toxicity of I. senegalensis are reported for the first time and showed promising results in malaria field research.

Liquid chromatography-tandem mass spectrometry for the determination of sphingomyelin species from calf brain, ox liver, egg yolk, and krill oil.

In this study, molecular species of sphingomyelin (SM) in egg yolk, calf brain, ox liver, and krill oil were investigated. Classes of phospholipids (PLs) were purified, identified, and quantified by normal phase semipreparative high-performance liquid chromatography (HPLC) combined with evaporative light scattering detectors (ELSD). For SM molecular species identification, pure SM collected through a flow splitter was loaded to HPLC-electrospray ionization-tandem mass spectrometry (LC-ESI-MS(2)), with 100% methanol containing 5 mM ammonium formate as mobile phase. In addition to classes of PLs, the used approach allowed the determination of profiles of SM species in egg yolk, ox liver, and calf brain, whereas krill oil turned out not to contain any SM. It also allowed the separation and identification of SM subclasses, as well as tentative identification of species with the same molecular mass, including isomers. The results showed that egg yolk contained the highest proportion of (d18:1-16:0)SM (94.1%). The major SM molecular species in ox liver were (d18:1-16:0)SM (25.5%), (d18:1-23:0)SM (19.7%), (d18:1-24:0)SM (13.2%), and (d18:1-22:0)SM (12.5%). Calf brain SM was rich in species such as (d18:1-18:0)SM (40.7%), (d18:1-24:1)SM (17.1%), and (d18:1-20:0)SM (10.8%).

Analysis of sitosteryl oleate esters in phytosterols esters enriched foods by HPLC–ESI-MS2

Phytosteryl esters (PE)-enriched spreads are marketed for eating and cooking purposes. Temperature and also light exposure are the major factors leading to the formation of PE oxides in food matrix. In this study a high-speed HPLC-MS(2) method was developed to analyze the major PE present in PE-enriched spreads: sitosteryl oleate (SO) and its oxidation products, by using synthesized compounds as standards. This analytical method was used to quantify seven SO oxides formed in PE-enriched spreads after heating at different temperatures for varying time periods and after prolonged exposure to sunlight. Quantification of remaining native SO was also performed after these different treatments. It was found that under specific heating conditions the decrease of the SO amount was much more important compared to the formation of SO oxides showing that many other products are formed. In contrast to heating, sunlight radiation did not result in the degradation of SO and very few oxides were formed.

Itaconic and Fumaric Acid Production from Biomass Hydrolysates by Aspergillus Strains.

Itaconic acid (IA) is a dicarboxylic acid included in the US Department of Energys (DOE) 2004 list of the most promising chemical platforms derived from sugars. IA is produced industrially using liquid-state fermentation (LSF) by Aspergillus terreus with glucose as the carbon source. To utilize IA production in renewable resource-based biorefinery, the present study investigated the use of lignocellulosic biomass as a carbon source for LSF. We also investigated the production of fumaric acid (FA), which is also on the DOEs list. FA is a primary metabolite, whereas IA is a secondary metabolite and requires the enzyme cis-aconitate decarboxylase for its production. Two lignocellulosic biomasses (wheat bran and corn cobs) were tested for fungal fermentation. Liquid hydrolysates obtained after acid or enzymatic treatment were used in LSF. We show that each treatment resulted in different concentrations of sugars, metals, or inhibitors. Furthermore, different acid yields (IA and FA) were obtained depending on which of the four Aspergillus strains tested were employed. The maximum FA yield was obtained when A. terreus was used for LSF of corn cob hydrolysate (1.9% total glucose); whereas an IA yield of 0.14% was obtained by LSF of corn cob hydrolysates by A. oryzae.