J. of Supercritical Fluids 82 (2013) 206–212 Contents lists available at ScienceDirect The Journal of Supercritical Fluids jou rn al hom epage: www.elsevier.com/locate/supflu Encapsulation of perﬂuorocarbon gases into lipid-based carrier by PGSS S. Rodríguez-Rojo a,b , D. Deodato Lopes b,c , A.M.R.C. Alexandre b,c , H. Pereira c , I.D. Nogueira d , C.M.M. Duarte b,c,∗ a High Pressure Processes Group, Department of Chemical Engineering and Environmental Technology, Universidad de Valladolid, Dr Mergelina s/n, 47005 Valladolid, Spain b Instituto de Biologia Experimental e Tecnológica, Apartado 12, 2781-901 Oeiras, Portugal c Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, Avenida da Republica, 2780-157 Oeiras, Portugal d Instituto Superior Tecnico, Instituto de Ciencias e Engenharia de Materiais e Superﬁcies, Universidade Tecnica de Lisboa, P-1096 Lisbon, Portugal a r t i c l e i n f o Article history: Received 24 March 2013 Received in revised form 23 May 2013 Accepted 24 May 2013 Keywords: Perﬂuorocarbons Particles from Gas Saturated Solutions (PGSS) Gas-ﬁlled particles Microbubbles Ultrasound triggered delivery a b s t r a c t For the ﬁrst time, gas-ﬁlled microparticles were successfully prepared using a supercritical ﬂuid based technology. Low molecular weight perﬂuorcarbon (PFC) gases, C 3 F 8 or C 4 F 8 , have been encapsulated into Gelucire ® 50/13 (lipid-based carrier: polyethylene glycol glycerides), using PGSS ® (Particles from Gas Saturated Solution) technique. Particles were produced from the fast expansion of the melted lipid carrier saturated with a mixture of (CO 2 + PFC). The presence of the gas into the produced microparticles was veriﬁed by Nuclear Magnetic Resonance (NMR) analysis of ﬂuorine atom. The effect of carrier to PFC mass ratio and PFC structure on the entrapment efﬁciency of the PFC gas into the particles was evaluated at ﬁxed at 8.5 MPa and 353 K. These parameters were ﬁxed in a preliminary study according to the morphology, size and ﬂowability of the particles. The stability of encapsulated C 4 F 8 in microparticles showed to be higher than C 3 F 8 ; it was determined to be 2 h, at room conditions at the optimized carrier: PFC mass ratio of 30:1. © 2013 Elsevier B.V. All rights reserved. 1. Introduction Since the last two decades, there is an increasing interest in gas-containing particulate systems using lipid based carriers for medical purposes (as contrast agents for ultrasound imaging and diagnostics) and in pharmaceutical applications (related to tar- geted drug delivery). These particles are called microbubbles (MBs) or gas ﬁlled microparticles. First generation MBs were simple air bubbles without any stabilizing shell; in second generation MBs a stabilizing shell was introduced, however, as the gas core was air or N 2 , their half life is less than 5 min (very short) due to the fact that these gases are sparingly soluble in blood. In third generation MBs, the gas core is formed by perﬂuorocarbons which are chem- ically and physiologically inert and practically insoluble in water, increasing the half-life of MBs [1,2]. Currently, Gas-ﬁlled microparticles are being investigated because of their great potential for Ultrasound-assisted drug deliv- ery for small molecules, nucleic acids, proteins and genes [3]. ∗ Corresponding author at: Instituto de Tecnologia Química e Biológica, Universi- dade Nova de Lisboa, Avenida da Republica, 2780-157 Oeiras, Portugal. Tel.: +351 214469727. E-mail address: cduarte@itqb.unl.pt (C.M.M. Duarte). Gas-ﬁlled microparticles can be destroyed precisely on the target site upon ultrasound triggering. Moreover, ultrasounds showed to be able to transiently enhance permeability of several biological barriers, such as the blood–brain barrier [4] and cell membranes [5], thereby facilitating the delivery of bioactive substances into tissues and cells [3–7]. Clinically, gas-ﬁlled bubbles can be used as drug vehicles or co-administrated separately but simultaneously with other delivery formulations [8]. Most common gases used in the synthesis of MBs are low molecular weight perﬂuorocarbons, as octoﬂuoropropane (C 3 F 8 ), octoﬂuorocyclobutane (C 4 F 8 ) and decaﬂuorobutane (C 4 F 10 ). Besides, some liquid ﬂuoroalkanes are also used: dodecaﬂuoropen- tane (C 5 F 12 ), which became gas above 29.5 ◦ C at atmospheric pressure, and tetradecaﬂuorohexane (C 6 F 14 ) [1,2]. Up to date, engineering gas-ﬁlled microparticles still remains a challenging task. Commonly used methods for the manufacture of commercially available MBs require several steps in the formu- lation process. Firstly, dry or colloidal particles or liposomes are produce by conventional methods such as o/w or w/o/w emul- sion followed by freeze drying, spray-drying of a solution of the shell material to produce void particles and thin phospholipid ﬁlm hydration. If dry particles are formed, they are reconstituted with an appropriated physiological buffer. Afterwards, the suspension is placed into vials, and the remaining head-space of the vials is ﬁlled 0896-8446/$ – see front matter © 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.supﬂu.2013.05.015