Colloids and Surfaces B: Biointerfaces 135 (2015) 50–55 Contents lists available at ScienceDirect Colloids and Surfaces B: Biointerfaces journal homepage: www.elsevier.com/locate/colsurfb In-vitro assessment of cytotoxicity of halloysite nanotubes against HepG2, HCT116 and human peripheral blood lymphocytes Farrukh Rafiq Ahmed a,b , Muhammad Harris Shoaib a,∗ , Mudassar Azhar c , Soong Ho Um d , Rabia Ismail Yousuf a , Shahkamal Hashmi e , Ahsana Dar c a Department of Pharmaceutics, Faculty of Pharmacy, University of Karachi, Karachi 75270, Pakistan b Department of Pharmaceutics, Faculty of Pharmacy, Ziauddin University, Karachi 75600, Pakistan c Dr. Panjwani Center for Molecular Medicine and Drug Research, ICCBS, University of Karachi, Karachi 75600, Pakistan d School of Chemical Engineering and SKKU Advanced Institute of Nanotechnology (SAINT), Sungkyunkwan University, Suwon 440-746, South Korea e Dow International Medical College, Dow University of Health Sciences, Karachi 75600, Pakistan a r t i c l e i n f o Article history: Received 3 March 2015 Received in revised form 2 July 2015 Accepted 8 July 2015 Available online 17 July 2015 Keywords: Halloysite nanotubes HepG2 HCT116 Peripheral blood lymphocytes In-vitro cytotoxicity Pharmaceutical excipients Size distribution a b s t r a c t Halloysite is a clay mineral with chemical similarity to kaolin, a pharmaceutical ingredient. It consists of mainly aluminosilicate nanotubular particles in the size range of ∼200–1000 nm. Many studies have tried to empirically explore this novel clay for its potential in drug delivery systems but no work has yet studied its cytotoxicity from the perspective of oral drug delivery system. In this study, the halloysite nanotubes (HNTs) were subjected to size distribution analyses, which reveal more than 50% of nanotubes in the size range of 500 nm and rest mainly in the sub micrometer range. HNTs were then evaluated for in-vitro cytotoxicity against HCT116 (colorectal carcinoma) and HepG2 (hepatocellular carcinoma) cells which represent the earliest entry point and the first accumulating organ, respectively, for nanoparticles en-route to systemic circulation after oral delivery. Moreover, HNTs were tested for their cytogenetic toxicity against human peripheral blood lymphocytes. Both these results collectively indicated that HNTs are generally safe at practical concentrations of excipients for oral dosage forms. © 2015 Elsevier B.V. All rights reserved. 1. Introduction Halloysite is an aluminosilicate clay mineral mined universally in many countries across the world [1]. Proportionately, it is largely constituted of elongated tubular shaped particles of sizes from hun- dreds of nanometersto maximum few micrometers in length (see Fig. 1) [2]. The diameter of the particles also varies from tens to few hundreds of nanometers, primarily due to the varying num- ber of alternate octahedral alumina and tetrahedral silica layers in 1:1 ratio which tend to mismatch and formed into rolled tubu- lar structure during volcanic activity [2,3]. Halloysite has been used commercially in ceramics, pottery and traditional Chinese medicines for various ailments ranging from diarrhoea to hemosta- sis and help healing wounds [4,5]. In recent times it has also been studied extensively for possible applications in the area of drug ∗ Corresponding author. E-mail addresses: mhshoaib@uok.edu.pk, harrisshoaib2000@yahoo.com (M.H. Shoaib). delivery systems and nanobiomedicine [4,6–9]. Moreover, a recent study has tried to rationalize the anti-inflammatory activity of this natural material with some promising results [10]. Chemically, halloysite is similar to another clay mineral, kaolin, which is used pharmaceutically as an excipient for many prepa- rations and as an active agent for anti-diarrheal preparations. However, halloysite has not been approved yet by the FDA and other regulatory bodies due to concerns of its nanotubular parti- cles and their associated safety [3,11]. So far few studies have tried to explore its cytotoxicity with some promising results through mostly random in-vitro studies against some cancer cell lines for e.g., HeLa, MCF-7, NIH-3T3, and A549 etc. [12–16]. A very recent in-vivo toxicity study in a soil nematode Caenorhabditis elegans has also demonstrated high safety prospects of this novel clay even at high concentration of 1000 g/mL [17]. Various studies have shown drug delivery and biological potential of this nanotubular material through in-vitro experiments, yet none has examine cytotoxicity from the perspective of oral drug delivery systems [8,13,18–20]. In this study, halloysite nanotubes have been characterized for size distribution (length, diameter, and aspect ratio) before being http://dx.doi.org/10.1016/j.colsurfb.2015.07.021 0927-7765/© 2015 Elsevier B.V. All rights reserved.