International Journal of Biological Macromolecules 48 (2011) 98–105 Contents lists available at ScienceDirect International Journal of Biological Macromolecules journal homepage: www.elsevier.com/locate/ijbiomac 5-Fluorouracil loaded fibrinogen nanoparticles for cancer drug delivery applications N. Sanoj Rejinold, M. Muthunarayanan, K.P. Chennazhi, S.V. Nair, R. Jayakumar ∗ Amrita Centre for Nanosciences and Molecular Medicine, Amrita Institute of Medical Sciences and Research Centre, Amrita Vishwa Vidyapeetham University, Kochi 682041, India article info Article history: Received 17 September 2010 Received in revised form 5 October 2010 Accepted 6 October 2010 Available online 14 October 2010 Key words: Fibrinogen nanoparticles Coacervation method Loading efficiency Cellular uptake Cancer drug delivery abstract In this study, 5-flurouracil loaded fibrinogen nanoparticles (5-FU-FNPs) were prepared by two step coacervation method using calcium chloride as cross-linker. The prepared nanoparticles were charac- terized using DLS, SEM, AFM, FT-IR, TG/DTA and XRD studies. Particle size of 5-FU-FNPs was found to be 150–200 nm. The loading efficiency (LE) and in vitro drug release was studied using UV spectrophotome- ter. The LE of FNPs was found to be ∼90%. The cytotoxicity studies showed 5-FU-FNPs were toxic to MCF7, PC3 and KB cells while they are comparatively non toxic to L929 cells. Cellular uptake of Rhodamine 123 conjugated 5-FU-FNPs was also studied. Cell uptake studies demonstrated that the nanoparticles are inside the cells. These results indicated that FNPs could be useful for cancer drug delivery. © 2010 Elsevier B.V. All rights reserved. 1. Introduction Natural macromolecules proteins have gained interest as bio- materials owing to their inherent property of biodegradability, lack of toxicity and non antigenicty [1–3]. Colloidal systems based on proteins might be incredibly promising as they unite the advan- tages of nanoparticles with the advantage of covalent protein drug conjugates [4–6]. Protein nanoparticles due to their existence of charged groups may also be used as a matrix in which drugs can be physically entrapped [7–9]. Fibrinogen is the precursor of fibrin, the blood clot substance. It is a large, asymmetric molecule, which is highly elongated. The molecular weight is 340-kDa. Fibrinogen has commonly been used as a coagulant in the medical field, but little is known regarding its suitability as a carrier of antitumor drugs. Nanoparticles from fibrinogen should yield a novel biodegradable system for drug delivery [10]. Fibrinogen nanoparticle preparation by two-step coacervation method has been reported by our group in our own method earlier, using calcium chloride as cross linker instead of toxic glutraldehyde [11]. The major disadvantage of the emulsion methods for particle preparation includes the need of applying toxic organic solvents for the formulation, organic surfactants for the stabilization of the emulsion and removal of the oil residues from the formulation. ∗ Corresponding author. Tel.: +91 484 2801234; fax: +91 484 2802020. E-mail addresses: rjayakumar@aims.amrita.edu, jayakumar77@yahoo.com (R. Jayakumar). Therefore, as an alternative method for the preparation of protein nanoparticles, a desolvation process derived from the coacervation method is presented. In this method, particles in aqueous solution were formed by a coacervation process and later on will be stabi- lized by cross-linking agent such as glutaraldehyde [12]. In general, the coacervating agent induces some progressive modifications of protein tertiary structure to give hydrophobicity, which then form aggregates of dissolved protein (coacervates). These coacervates are usually unstable and have to be hardened or stabilized either by cross-linking mode via a physical or chemical method. The sta- bilized particles are in the micro or nano range, which is purely based on the concentration of the precursors, which depends on their size. 5-FU, a pyramidine analogue that interferes with thymidy- late synthesis has a broad spectrum of activity against solid tumors. Limitations are short biological half-life due to rapid metabolism, incomplete and non-uniform oral absorption due to rapid metabolism by dihydropyramidine dehydrogenase and non- selective action against healthy cells. To prolong the circulation time of 5-FU and increase its efficacy, its delivery has to be modified by incorporation into nanoparticulate carriers to reduce the 5-FU associated side effects and there by improve its therapeutic index [13]. The aim of our study is to develop a nano formulation for the 5-fluouracil drug to enhance its action against the cancer cells with minimal toxicity on the normal cells. The objective of our study is to prepare and characterize the 5-FU-FNPs for cancer drug deliv- ery applications. In addition, the in vitro drug release studies, cell uptake and cell cytotoxicity studies in detail for normal and cancer cell lines were investigated. 0141-8130/$ – see front matter © 2010 Elsevier B.V. All rights reserved. doi:10.1016/j.ijbiomac.2010.10.003