Optimization of coenzyme Q10 encapsulation in liposomes using supercritical carbon dioxide
Graphical abstract
Introduction
Coenzyme Q10 (CoQ10) is a benzoquinone, which possesses interesting health benefits, such as antioxidant [1], anti-cancer [2], anti-diabetic [3], cardioprotective [4] and neuroprotective properties [5]. Moreover, CoQ10 plays a fundamental role for the biological energy transfer in the body as a cofactor in the mitochondrial electron transport chain. Despite a certain amount being synthesized endogenously, approximately a quarter of the total CoQ10 in an adult human body must be ingested daily. Furthermore, tissue levels of CoQ10 drop progressively with increasing age and its deficiency has been also observed in various medical conditions [6]. As a dietary source, only some meats, fish and oils contain substantial amounts of CoQ10 [7]. Therefore, its exogenous supplementation to compensate the low daily intake or age-related decline is recommended.
The fortification of food products with CoQ10 is challenging due to its low water solubility as a consequence of its crystalline nature, highly hydrophobic polyisoprenoid tail and large molecular mass. Moreover, the bioaccesibility and bioavailability of CoQ10 is low and it is sensitive to high temperature, light and oxygen [8]. To overcome these limitations, different encapsulation approaches have been developed, and among those, liposomes have received growing attention.
Liposomes are vesicles composed of one or more phospholipid bilayers surrounding an aqueous core. Due to their non-toxicity, small size, biocompatibility and amphipathic character, liposomes have great potential as delivery systems for bioactive compounds. Conventional methods to produce liposomes, such as thin film hydration, reverse phase evaporation and detergent dialysis methods, present important drawbacks, including the use of excessive amounts of organic and toxic solvents, residues in the product, the degradation of bioactive compounds due to the use of high temperatures and the difficulty to control the particle size, which leads to heterogeneous size distribution and low reproducibility of the results [9,10]. To overcome these drawbacks, the use of supercritical CO2 (SCCO2) has become a green alternative. CO2 is inexpensive, inert, non-toxic, non-flammable, allows obtaining solvent-free products and its low critical temperature prevents or minimizes the degradation of bioactive compounds. Moreover, the solvent power and fast diffusion rates of SCCO2 can be exploited to create supersaturated systems and design of particle formation processes, which result in the precipitation of small particles and homogeneous products [11].
A number of studies have been reported on the production of liposomes containing CoQ10. In the majority of these studies, liposomes have been produced by conventional methods [[12], [13], [14], [15], [16], [17], [18], [19]] and only two studies have employed SCCO2, where the particles were produced by Rapid Expansion of Supercritical Solutions (RESS) [20] and Supercritical Anti-Solvent (SAS) [21] techniques. Nevertheless, in both of them, the use of organic solvents was necessary.
In the present study, a novel single step SCCO2 process, which did not require the use of organic solvents, buffers or surfactants, was applied for the preparation of CoQ10-loaded liposomes with soy lecithin for the first time. The objective was to investigate the effects of pressure, pressurization rate, temperature and CoQ10-to-lecithin ratio on the particle size, size distribution and bioactive loading. The influence of each variable on the responses measured was mathematically modeled and optimized using the response surface methodology (RSM). In addition, the encapsulation efficiency of CoQ10 and the morphology, phase transition and zeta-potential of particles were determined.
Section snippets
Materials
CoQ10 (98.34 % purity) was obtained from PureBulk (Roseburg, OR, USA). Soy lecithin was supplied by Fisher Scientific (Ottawa, ON, Canada). Anhydrous ethanol was purchased from Sigma-Aldrich (St. Louis, MO, USA). Ethyl acetate (99.9 % purity) was obtained from Fisher Scientific (Ottawa, ON, Canada). CO2 (99.9 % purity, < 3 ppm H2O) was purchased from Praxair Canada (Mississauga, ON, Canada). Mili-Q water was used in the experiments.
Preparation of the crude suspension
Soy lecithin (MW =677.92 g∙mol−1, as specified by the supplier)
Model fitting
The effects of pressure (10, 20 and 30 MPa), temperature (40, 50 and 60 °C), pressurization rate (6, 12 and 18 MPa min−1) and CoQ10-to-lecithin molar ratio (1, 9 and 17 mol%) on bioactive loading (BL), particle size (Ps) and polydispersity index (PdI) were investigated. Table 2 shows the design matrix and the experimental results obtained. BL values close to 10 % (run 8) were achieved. Ps and PdI values ranged, respectively, from 168 nm (run 29) to 258 nm (run 23) and from
Conclusions
Unlike the other supercritical processes reported until now for the encapsulation of CoQ10 in liposomes, a simpler single step method was employed in the present study, which did not require the use of organic solvents, additional surfactants or buffers. In addition, soy lecithin was used as the source of phospholipids, which is more economical and widely available compared to the purified phospholipids typically used in other studies.
The response surface model adequately described the selected
Author contributions
Dr. Feral Temelli has carried out the funding acquisition and supervision of the study, as well as the conceptualization, the methodology and the provision of the materials for the research.
Dr. David Villanueva-Bermejo has participated in the conceptualization and has performed the experiments and data analysis as well as writing the draft manuscript.
Both authors have participated in the interpretation of the results and the review and editing of the manuscript.
Declaration of Competing Interest
The authors declare no conflict of interest.
Acknowledgments
The financial support of the Natural Sciences and Engineering Research Council of Canada (NSERC) − Discovery Grants Program (RGPIN-2017-04384) is gratefully acknowledged. D. Villanueva-Bermejo gratefully acknowledges the post-doctoral fellowship provided by the Spanish Alfonso Martin Escudero Foundation.
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