Curcumin nanoformulations

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Review Curcumin nanoformulations: A review of pharmaceutical properties and preclinical studies and clinical data related to cancer treatment Ornchuma Naksuriya a , b , Siriporn Okonogi a , Raymond M. Schiffelers c , Wim E. Hennink b , * a Department of Pharmaceutical Sciences, Faculty of Pharmacy, Chiang Mai University, Suthep Rd, Mueang, Chiang Mai 50200, Thailand b Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Universiteitsweg 99, Utrecht 3805 TB, The Netherlands c Department of Clinical Chemistry and Hematology, University Medical Center Utrecht, Utrecht, The Netherlands article info Article history: Received 28 November 2013 Accepted 22 December 2013 Available online 15 January 2014 Keywords: Curcumin Cancer Nanoformulation Drug delivery Nanomedicine Clinical studies abstract Curcumin, a natural yellow phenolic compound, is present in many kinds of herbs, particularly in Cur- cuma longa Linn. (turmeric). It is a natural antioxidant and has shown many pharmacological activities such as anti-inflammatory, anti-microbial, anti-cancer, and anti-Alzheimer in both preclinical and clinical studies. Moreover, curcumin has hepatoprotective, nephroprotective, cardioprotective, neuroprotective, hypoglycemic, antirheumatic, and antidiabetic activities and it also suppresses thrombosis and protects against myocardial infarction. Particularly, curcumin has demonstrated efficacy as an anticancer agent, but a limiting factor is its extremely low aqueous solubility which hampers its use as therapeutic agent. Therefore, many technologies have been developed and applied to overcome this limitation. In this re- view, we summarize the recent works on the design and development of nano-sized delivery systems for curcumin, including liposomes, polymeric nanoparticles and micelles, conjugates, peptide carriers, cy- clodextrins, solid dispersions, lipid nanoparticles and emulsions. Efficacy studies of curcumin nano- formulations using cancer cell lines and in vivo models as well as up-to-date human clinical trials are also discussed. Ó 2014 Elsevier Ltd. All rights reserved. 1. Introduction Curcumin is a natural yellow colored phenolic antioxidant and was first extracted in an impure form by Vogel et al. [1]. The structure of curcumin was elucidated and it was synthesized by Milobedeska et al. and Lampe et al., respectively [2,3]. Many different plant species synthesize curcumin and the commercial product (such as from SigmaeAldrich) is isolated from the rhizome of Curcuma longa Linn. in which it is present in relatively high concentrations. The chemical structure of curcumin is shown in Fig. 1. It should be mentioned that the commercially available curcumin products also contain structurally related compounds (w17% demethoxycurcumin, and 3% bisdemethoxycurcumin). Sandur et al. reported that the potency for the suppression of tumor necrosis factor (TNF)-induced nuclear factor-kappaB (NF- k B) acti- vation ranked curcumin > desmethoxycurcumin > bisdesme- thoxycurcumin suggesting a critical role of the methoxy groups on the phenyl rings [4]. Moreover, curcumin has the highest car- dioprotective, neuroprotective and antidiabetic activities of the three curcuminoids shown in Fig. 1 [5e7]. Interestingly, the mixture of curcuminoids has increased nematocidal activity as compared to the individual compounds, suggesting a synergistic effect [8]. For many centuries, curcumin in its crude form has been used as spice and dietary supplement as well as component of many traditional Asian medicines [9]. In recent studies, it has been shown that curcumin exhibits a wide range of pharmacological activities against many chronic diseases including type II diabetes, rheuma- toid arthritis, multiple sclerosis, Alzheimer’s disease and athero- sclerosis. It also inhibits platelet aggregation, suppresses thrombosis and inhibits human immunodeficiency virus (HIV) replication. Further, curcumin enhances wound healing and pro- tects against liver injury, cataract formation, pulmonary toxicity and fibrosis [10e20]. Finally, the anti-cancer activity of curcumin has been extensively investigated and it has been suggested as a potential agent for both prevention and treatment of a great variety of different cancers, including gastrointestinal, melanoma, genito- urinary, breast, lung, hematological, head and neck, neurological and sarcoma [20e23]. At a molecular level, curcumin not only in- hibits cell proliferation and metastasis, but also induces apoptosis by modulating several pro-inflammatory factors (e.g. interleukin (IL)-1, IL-1 b , IL-12, tumor necrosis factor (TNF)- a and interferon * Corresponding author. Tel.: þ31 30 253 6964; fax: þ31 30 251 7839. E-mail address: W.E.Hennink@uu.nl (W.E. Hennink). Contents lists available at ScienceDirect Biomaterials journal homepage: www.elsevier.com/locate/biomaterials 0142-9612/$ e see front matter Ó 2014 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.biomaterials.2013.12.090 Biomaterials 35 (2014) 3365e3383 (INF)- g ), growth factors (e.g. epidermal growth factor (EGF), he- patic growth factor (HGF) and platelet-derived growth factor (PDGF)), receptors (e.g. epidermal growth factor receptor (EGFR), human epidermal growth factor receptor (HER)-2, IL-8R and Fas-R), transcription factors (e.g. signal transducer and activator of tran- scription (STAT) 3, nuclear factor (NF)- k B, Wilms’ tumor (WT-1) and peroxisome proliferator-activated receptor (PPAR) g ) and protein kinases, e.g. extracellular signal-regulated kinases (ERK), mitogen-activated protein kinases (MAPK), protein kinase A (PKA) B (PKB) and C (PKC) [20e25]. An overview of the different indications for which curcumin has been investigated is shown in Fig. 2. It has been suggested that, because of its many pleiotropic properties, curcumin can be more effective than single pathway targeted anticancer drugs [26,27]. Many preclinical studies have demonstrated that curcumin has anti-inflammatory and anticancer activity [27e30]. In a recent clinical study it appeared that oral administration of curcumin was well tolerated at doses of 12 g/day which indicates that curcumin is safe [31]. Curcumin can freely pass through cellular membranes due to its lipophilicity (log P ¼ 2.5) [32]. It should however be mentioned that curcumin has a very low aqueous solubility of only 0.6 m g/ml and is susceptible to degradation particularly under alkaline conditions [33e35]. These characteristics are the cause for its very low bioavailability resulting in suboptimal blood concen- trations to achieve therapeutic effects [21,34e36]. For instance, in a study in rats reported by Yang et al. a maximum serum concen- tration of 0.36 Æ 0.05 m g/ml after an intravenous injection of 10 mg/ kg was reached, whereas 500 mg/kg orally administered curcumin gave a maximum plasma concentration of 0.06 Æ 0.01 m g/ml, indicating that oral bioavailability was only 1% [37]. Similarly, Shoba et al. showed a maximum serum concentration of 1.35 Æ 0.23 m g/ml at 1 h after administration of an oral dose of 2 g/ kg to rats, whereas healthy man volunteers (weighing 50e75 kg) receiving a single dose of 2 g curcumin (4 capsules of 500 mg each) showed an extremely low serum concentration of 0.006 Æ 0.005 m g/ml at 1 h [38]. An obvious approach to improve the poor biopharmaceutical properties of curcumin is to improve its aqueous solubility using nanocarriers. Nanocarriers have a small size (typically 10e100 nm) and can, besides for solubilization, also be used for the targeted delivery of drugs [39e44]. Nanocarriers can improve the circulation time of the loaded therapeutic agent and may improve its accumulation at the pathological site exploiting the so-called ‘enhance permeation and retention (EPR) effect’ [45e48]. During the last decades, various types of nano- carriers, such as polymeric micelles and nanoparticles, liposomes, conjugates, peptide carriers etc., for drug delivery/targeting have been investigated and some systems have reached clinical evalua- tions and applications [49e52]. Many studies, as summarized in the next sections, have shown that nanocarriers are suitable for increasing curcumin’s bioavailability and its targeted delivery to tumors and other sites of disease. This review focuses on the design and development, the evaluation in preclinical and clinical trials of curcumin nanoformulations, particularly focused on cancer ther- apy. In the next section, different curcumin nanoformulations are discussed with emphasis on their pharmaceutical properties. In the final section of this review the results of curcumin nano- formulations in preclinical studies and clinical evaluations are summarized and discussed. 2. Curcumin nanoformulations The nanoformulations discussed in this section primarily aim to achieve increased solubilization of curcumin, but at the same time protect curcumin against inactivation by hydrolysis. The formula- tion should be efficiently prepared and loaded and should retain curcumin for the required time period. Some formulations are aimed for a prolonged release of curcumin, while others have additional mechanisms for cellular delivery or intracellular release. 2.1. Liposomes Liposomes consist of one or more phospholipid bilayers sur- rounding an aqueous core. Both lipophilic compounds/drugs (sol- ubilized in the liposomal bilayer) and hydrophilic compounds (soluble in the aqueous core) can be loaded into liposomes. Different types of liposomes for targeted drug delivery have been developed and some systems have reached clinical practice [53e56]. Many liposomal curcumin formulations have been developed in recent years (Table 1) and a few studies are highlighted. Karewicz et al. prepared curcumin loaded liposomes composed of egg yolk phosphatidyl choline (EYPC), dihexyl phosphate (DHP), and cholesterol prepared by the film evaporation technique [57]. Because of its lipophilicity, curcumin is solubilized in the lipophilic bilayer. By using fluorescent probes, the authors showed that it was indeed located at the hydrophobic acyl side chain and positioned closely to the glycerol groups. It was shown that curcumin loaded into the EYPC/DPH/cholesterol liposomal bilayer stabilizes the system proportionally to its content. In a follow up study, the li- posomes were coated with the cationic lipid/polymer conjugate N- dodecyl chitosan-N-[(2-hydroxy-3-trimethylamine) propyl] (HPTMA) chloride. The obtained liposomes with a size of 73 nm were able to bind to and penetrate cells due to their cationic nature. These coated liposomes released their content in a sustained manner in about 10 h. Further, the formulations showed a slightly better cell killing activity than free curcumin, likely due to the improved cellular internalization of the cationic liposomes [58]. Re et al. developed curcumin loaded liposomes composed of bovine brain sphingomyelin, cholesterol, and 1,2-stearoyl-sn-glyc- ero-3-phosphoethanolamine-N-[maleimide(poly(ethylene glycol)- 2000)] and surface functionalized with the apolipoprotein E (ApoE) peptide as targeting ligand. The liposomes were prepared by the fi lm evaporation technique and non-incorporated curcumin Fig. 1. Chemical structures of curcumin (A), demethoxycurcumin (B) and bisdeme- thoxycurcumin (C). O. Naksuriya et al. / Biomaterials 35 (2014) 3365e33833366 was removed by size-exclusion chromatography using a PD-10 column. The recovery of lipids was about 90% and the liposomes had a mean size of w130 nm. It was shown these ApoE-liposomes enhanced the transport of their curcumin payload through RBE4 brain capillary endothelial cells making these nanocarriers inter- esting for brain targeting [59]. In another study, a cationic lipo- someepolyethylene glycol (PEG)epolyethylenimine (PEI) complex (lipoePEGePEI complex, LPPC) was used for the encapsulation of curcumin. Morphological analysis by transmission electron micro- scopy (TEM) showed a spherical shape of the liposomal nano- particle with hair like projections on the surface likely originating from PEG and PEI. The size of curcumin loaded LPPC was w260 nm and the encapsulation efficiency of curcumin was 45%. In vitro, these LPPC released curcumin within 120 h [60]. 2.2. Polymeric nanoparticles Different polymers, particularly biodegradable ones, have been used for the preparation of curcumin loaded nanoparticles [65] . PLGA (poly(D,L-lactic-co-glycolic) is widely used for drug delivery purposes due to its biocompatibility and biodegradability [66e72]. Shaikh et al. reported on curcumin loaded PLGA nanospheres pre- pared byemulsion-evaporation method usingPVA as surfactant.The obtained particles had a size of 264 nm and 77% entrapment effi- ciency resulting in 15% loading capacity of curcumin. The particles showed a biphasic release pattern characterized by a relatively rapid initial release of about 24% of the loading in 24 h followed by sus- tained releaseof about 20%ofthe loading duringthe next 20days. An in vivo study in rats revealed that the curcumin loaded PLGA nano- spheres improved the oral bioavailability of curcumin at least 9 fold when compared to curcumin administered with piperine. The latter compound was co-administered to improve curcumin availability as it inhibits curcumin inactivation by hepatic and intestinal glucur- onidation [73]. Yallapu et al. encapsulated curcumin in PLGA nano- particles by a nanoprecipitation method using poly(vinyl)alcohol (PVA) and poly( L -lysine) as stabilizers (nano-CUR 1e6) [74].Itwas found that the size ofthenanoparticles decreased from 560 to76 nm with increasing PVA concentration. Further, the particles had a neutral zeta-potential, although for poly( L -lysine) coated nano- particle a positive zeta-potential is expected. The absence of charge on the particle surfaces might be ascribed by the improper way the measurements were done (in distilled water with no pH control), whereas a low ionic strength buffer is preferred [75] or even the absence of the polylysine coating. The nanoparticles showed after a small burst of around 20% of the loading a sustained release of cur- cumin for 25 days (Fig. 3). The particles prepared with the highest concentration of PVA showed the slowest release and the authors hypothesized that the surface adsorbed PVA acts as a barrier and consequently controls the release rate. The Nano-CUR6 formulation that released 64% of the loaded curcumin in 25 days was selected for further in vitro studies (outcome discussed in section 3) [74]. Ghosh et al. developed curcumin-loaded PLGA nanoparticles (Nano Cur) for the treatment of diethylnitrosamine (DEN) induced hepatocellular carcinoma (HCC) in rats. Nano Cur was prepared by emulsion- diffusion-evaporation method and atomic force microscopy (AFM) showed that the particles had an average diameter of 14 nm. The optical density of Nano Cur was measured at l max of 422 nm to calculate the encapsulation efficiency which was 78%. Fourier transform infrared (FTIR)analysis revealed that there were no strong interactions between curcumin and the polymer matrix, but no Fig. 2. Indications for which curcumin has been investigated. O. Naksuriya et al. / Biomaterials 35 (2014) 3365e3383 3367 release data were reported. It was also not commented why the particles had a small size, but probably the strong surfactant (didodecyldimethylammonium bromide) in combination with the high-speed homogenizer that was used to produce the emulsions might be an explanation [76]. Anand et al. prepared curcumin- loaded PLGA nanospheres using a nanoprecipitation method and polyethylene glycol (PEG)-5000 as stabilizer. Curcumin was almost quantitatively entrapped in particles of 81 nm. However, no in vitro release data were reported [77]. Polylactic-co-glycolic acid (PLGA) and PLGAepolyethylene glycol (PLGAePEG) nanoparticles contain- ing curcumin were obtained by a single-emulsion solvent evapora- tion technique [78]. The encapsulation efficiency was over 70% and particles with asize w150 nm were formed.ThePLGAePEG particles released 21% of curcumin in 24 h, followed by a sustained release to 57% of the loading over 9 days. On the other hand, the PLGA particles showed a continuous release of 40% of the loading in 9 days. The authors hypothesized that the faster release of curcumin from the PLGAePEG nanoparticles is attributed the higher water-absorbing capacity of this matrix compared to PLGA only [78]. NIPAAm, N-vi- nyl-2-pyrrolidone, poly(ethyleneglycol) monoacrylate and N,N 0 ,- methylene bis acrylamide were copolymerized in water resulting in crosslinked nanoparticles with a size of 50 nm. An aqueous disper- sion of thesenanoparticleswas vortexed with asolutionof curcumin in CH 3 Cl and a very high 90% entrapment efficiency was obtained. The loaded nanoparticles released 40% of their content in 24 h [79]. Natural polymers have also been used to prepare curcumin nanomedicines. Liu et al. developed curcumin loaded chitosan/ poly(Ɛ-caprolactone) (chitosan/PCL) nanoparticles by a precipita- tion method. The mean diameter of the obtained nanoparticles was between 220 and 360 nm whereas the encapsulation efficiency and loading of curcumin were 71 and 4%, respectively. The curcumin chitosan/PCL nanoparticles released 68% of their content over 5 days in a sustained manner [80]. Rejinold et al. described chitosan-g- poly(N-isopropylacrylamide) for the development curcumin- loaded nanoparticles [81]. This polymer is temperature sensitive because of the presence of the pNIPAAm grafts [82,83]. Below the lower critical solution temperature (LCST) (38  C), chitosan-g- poly(N-isopropylacrylamide) was fully soluble in water whereas the polymer solution became turbid above the LCST. Particles of Table 1 Some recent curcumin liposomal formulations. Formulation Entrapment efficiency (%) Size (nm) Release kinetics Status of investigation Observations Reference Curcumin loaded liposomes coated with N-dodecyl chitosan-HPTMA chloride Not reported 73 >80% in 10 h In vitro Non-toxic for murine fibroblasts (NIH3T3) whereas toxic for murine melanoma (B16F10) cells. [58] Curcumin loaded liposomes coupled with the ApoE peptide Not reported 132 Not reported In vitro Increased accumulation of curcumin in RBE4 cell brain capillary endothelial cells. [59] Curcumin loaded lipoe PEGePEI complex 45 269 90% in 120 h In vitro/vivo The cytotoxic activity of the nanoformulation was higher than free curcumin on both curcumin-sensitive cells and curcumin-resistant cells. 60e90% inhibition of tumor growth in mice inocolated with CT-26 or B16F10 cells. [60] Curcumin loaded silica- coated flexible liposomes 91 157 Not reported In vivo Increased 3.3-fold bioavailability compared with curcumin loaded liposomes in mice through gavage administration. [61] Curcumin-conjugated nanoliposomes Not reported 207 Not reported In vivo Down regulated the secretion of amyloid peptide (A b ) and partially prevented A b induced toxicity in mouse model of Alzheimer disease. [62] Curcumin loaded soybean phosphati-dylcholine liposomes Not reported 176 37% in 48 h In vivo Decreased parasitemia and increase survival of Plasmodium berghei infected mice (anti-malarial therapy). [63] Curcumin loaded egg phosphatidyl-choline liposomes Not reported Not reported Not reported In vivo Exhibited cytoprotection for renal ischemiaereperfusion injury. [64] Fig. 3. Curcumin release profiles of nano-CUR 1e6. The red circle indicates the burst release. Nano-Cur 1 to 6 were prepared with different concentrations of PVA in the aqueous phase (0e1% w/v). Reprinted from Journal of Colloid and Interface Science, Vol. 351/1, M.M. Yallapu, B.K. Gupta, M. Jaggi, S.C. Chauhan, Fabrication of curcumin encapsulated PLGA nanoparticles for improved therapeutic effects in metastatic cancer cells, pp. 19e29, Copyright (2010), with permi ssion from Elsevier. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.) O. Naksuriya et al. / Biomaterials 35 (2014) 3365e33833368 chitosan-g-poly(N-isopropylacrylamide) with a size of 180e220 nm were formed by ionic crosslinking using pentasodium tripoly phosphate (TPP)in the presence of curcumin addedto the mixtureas a solution in ethanol. Only 5% of the loaded amount of curcumin was released below the LCST in 35 h, whereas above this temperature 100% drug release was observed within 35 h. The authors hypoth- esized that below the LCST hydrogen bonds exist between the phenolic hydroxyl groups of curcumin and the amide groups of the pNIPAAm blocks that retain curcumin in the polymer matrix. Above this temperature the interpolymer interactions dominate and as a consequence curcumin-polymer interactions are weakened, which in turn results in release of the active [81]. In a recent study of Anitha et al.,curcumin-loaded nanoparticlesof dextran sulfate and chitosan were prepared by coacervation method resulting in spherically shaped and stable nanoparticles of 200 e220 nm which are hold together by electrostatic interaction between the two oppositely charged polymers and the curcumin encapsulation efficiency and loading capacity were 74 and 5%, respectively [84]. The drug release pattern was characterized by a burst release in the first 3 h followed by a sustained release of curcumin that reached 70% of the loaded amount within 120 h. The release was faster at pH 5 than at pH 7 due to the protonation of the amine groups of chitosan at low pH resulting in swelling of the polymer matrix [84]. 2.3. Polymeric micelles Polymeric micelles are composed of amphiphilic block co- polymers that spontaneously form micelles with a size ranging between 20 and 100 nm in aqueous solution above the critical micellar concentration (CMC). The hydrophobic core can accom- modate hydrophobic drugs and therefore polymeric micelles have been extensively used for solubilization and targeted delivery of drugs [85e92]. Song et al. loaded curcumin into micelles of amphiphilic methoxy poly(ethylene glycol)-b-poly(Ɛ-capro- lactone-co-p-dioxanone) by a solid dispersion method [93]. These micelles had a small size (30 nm) with a narrow size distribution, whereas the entrapment efficiency was more than 95% and the loading capacity was 12%. The micelles slowly released w80% of their content without a burst in 300 h [93]. A poly( D , L -lactide-co- glycolide)-b-poly(ethylene glycol)-b-poly( D , L -lactide-co-glycolide) (PLGAePEGePLGA) triblock copolymer was synthesized by ring- opening polymerization of D , L -lactide using PEG as macroinitiator [94]. Curcumin loaded triblock copolymer micelles were prepared by a dialysis method and it was shown that the CMC at room temperature was 2.8 Â 10 À2 mg/ml. The drug loading capacity and entrapment efficiency were 4 and 70%, respectively. TEM analysis showed that the micelles were spherically shaped and had a size of 26 nm which was con firmed by dynamic light scattering mea- surements [94]. No release data were reported, but these nano- particles were evaluated in vivo (discussed in Section 4). Zhao et al. used a central composite design to optimize a formulation of mixed micelles composed of Pluronics P123 and F68 [95]. The average size of the mixed micelles was 6 8 nm, and the encapsulation efficiency and loading capacity for curcumin were 87% and 7%, respectively. It was shown that 50% of the loaded curcumin was released from the micelles in 72 h demonstrating that this formulation had sustained release properties [95]. Samanta et al. conducted a molecular dy- namics study of curcumin with pluronic block copolymers and they concluded that the hydrophobic PPO chains cover the curcumin molecule leaving the hydrophilic PEO chains exposed, resulting in solvation of curcumin in water [96]. Gong et al. reported on the encapsulation of curcumin in monomethyl poly(ethylene glycol)- poly(Ɛ-caprolactone) (MPEGePCL) micelles by a one-step solid dispersion method [97]. Micelles with a mean diameter of 27 nm were obtained that were well dispersible in water after freeze- drying. The encapsulation efficiency and drug loading capacity were 99 and 15%, respectively. The release study was done by dialysis method using phosphate buffered saline (PBS) and 0.5% of tween 80 as external medium, and these micelles released about 58% of the loading in 14 days [97]. Ma et al. loaded curcumin in micelles of different PEO-PCL block copolymers by a cosolvent evaporation technique [98]. It was reported that the PEO 5000 - PCL 24500 showed the highest solubilization capacity whereas PEO 5000 -PCL 13000 had the best drug retention capacity resulting in the slowest release kinetics. The authors also found that the release was faster in the presence of HSA which is probably due to the high affinity of curcumin for HSA [98]. 2.4. Conjugates Conjugation of curcumin to small molecules (particularly amino acids) and as well as to both natural and synthetic hydrophilic polymers has been exploited to increase its aqueous solubility. Several amino acids among which proline, glycine, leucine, isoleucine, alanine, phenylalanine, phenyl glycine, valine, serine and cysteine were coupled to curcumin [99]. These conjugates were synthesized in dry dioxane using e.g. N,N 0 -dicyclohex- ylcarbodiimide (DCC) as coupling agent, and (4-dimethylamino- pyridine (DMAP) and triethylamine (TEA)) as catalysts, and purified by column chromatography. These amino acid conjugations increased curcumin’s aqueous solubility to 1e10 mg/ml [99]. Manju et al. reported on the conjugation of hyaluronic acid and curcumin both dissolved in a water/DMSO mixture using DCC and DMAP as coupling agent and catalyst, respectively [100]. Although hyal- uronic acid is very well soluble in water, the conjugates were amphiphilic due to the hydrophobic curcumin groups and as a result they self-assembled into particles with a size between 300 and 600 nm and a negative zeta-potential (À25 to À75 mV). It was found that curcumin conjugated to hyaluronic acid remained intact for 90% once incubated in aqueous solution at pH 7.4 for 8 h whereas free curcumin showed 60% degradation within 25 min [100]. Tang et al. conjugated curcumin to two short oligo(ethylene glycol) chains via b -thioester bonds that are labile in the presence of intracellular glutathione and esterases (Curc-OEG; Fig. 4B (top)) [101]. These Curc-OEG conjugates contained 25% by weight cur- cumin and formed micelles with a size of 37 nm that released less than 12% of the conjugated amount of curcumin by hydrolysis in 24 h at pH 7.4 and 5.0 indicating a good stability of this system in PBS. On the other hand, Fig. 4B (bottom) shows that more than 25% of the conjugated curcumin at pH 7.4 and 35% at pH 5.0 was released within 10 h in a medium containing reduced glutathione (GSH) and more than 80% of Curc-OEG hydrolyzed within 2 h at pH 7.4 in medium containing 30 U esterase. The authors argued that Curc-OEG will be stable in the blood circulation and release cur- cumin once in the cell catalyzed by a combination of GSH and esterase [101]. In a recent study, three curcumin molecules were covalently linked to the distal end of a block copolymer of methoxy poly(ethylene glycol) (mPEG) and PLA via a tris (hydroxyl methyl) aminomethane (Tris) spacer (mPEGePLAeTriseCur) (Fig. 5) [102]. Also, a block copolymer of (mPEG) and PLA to which one molecule of curcumin was coupled was synthesized. Micelles with a size from 60 to 100 nm were prepared by a dialysis method and they con- tained both conjugated and solubilized curcumin with a high loading (up to 20%; only 2% loading for mPEGePLA micelles). The release was studied using a Franz cell and PBS (pH 7.4) containing 5% sodium dodecyl sulfate as the acceptor medium. It was found that the release of curcumin was due to a combination of diffusion of physically loaded curcumin and hydrolysis of the ester bond that connects curcumin and the polymer (Fig. 5). The authors reported that mPEGePLAeCur and mPEGePLAeTriseCur showed a rapid O. Naksuriya et al. / Biomaterials 35 (2014) 3365e3383 3369 release of curcumin during the first 12 h which then leveled off. The authors argued that the release was controlled by hydrolysis of the ester bond connecting the active and the polymer but that simul- taneously degradation of released curcumin occurred resulting in a steady state concentration of the compound. However, no convincing data were presented to substantiate this explanation [102]. Wichitnithad et al. coupled curcumin via different carboxylic ester spacers to mPEG 2000. The authors reported a log-linear release of curcumin in time for all conjugates tested. Further, it was shown that as compared to the half-life of free curcumin (0.56 h at pH 7.4 and 37  C), PEG bound curcumin had a substantial better stability (t 1/2 is w3to13h)[103]. 2.5. Peptide/protein carriers Beta casein, an amphiphilic polypeptide with molecular mass of 24,650 Da, spontaneously forms micelles (CMC at 37  Cis 8m M ). When curcumin was loaded in the hydrophobic core of these casein micelles, its solubility increased 2500 fold [104]. However, no data regarding size and release properties were re- ported. Nanoparticles of cross-linked human serum albumin (HSA) have shown good biocompatibility and have been used for drug delivery purposes [105,106]. Kim et al. presented curcumin-HSA nanoparticles that were prepared by homogenization of a mixture of HSA in water and curcumin in chloroform [107]. The mean size of curcumin-loaded HSA particles was 135 nm and the loading capacity was 7.2%. The authors speculated that the parti- cles were formed by crosslinking of albumin molecules via disul- fide exchange due to heating associated with cavitation produced by the high-pressure homogenizer. Curcumin was likely solubi- lized in hydrophobic cavities of albumin [108,109] resulting in a 300 fold increase in solubility. However, the release characteristics of the nanoparticles were not investigated [107]. 2.6. Cyclodextrins Cyclodextrins are cyclic oligosaccharides with a hydrophilic outer surface and a lipophilic cavity that can solubilize hydrophobic drugs and other small hydrophobic compounds such as curcumin [110,111]. Yadav et al. used 2-hydroxypropyl- g -cyclodextrin (HP g CD) to complex curcumin by a pH shift method. Curcumin was dissolved in an alkaline solution containing HP g CD and subse- quently the pH was adjusted to 6.0 [112]. Due to this pH change curcumin becomes hydrophobic and consequently partitioned in Fig. 4. Synthesis of curcumin amino acid conjugates (A). Reprinted from Food Chemistry, Vol. 120/2, K. Parvathy, P. Negi, P. Srinivas, Curcumineamino acid conjugates: Synthesis, antioxidant and antimutagenic attributes, pp. 523e530, Copyright (2010), with permission from Elsevier. (B) top; chemical structure of Curc-OEG, middle; synthesis of Curc-OEG, bottom; degree of hydrolysis of Curc-OEG at different conditions (B). Reproduced from Nanomedicine, Volume 5, Issue 6, pp. 855e865 with permission of Future Medicine Ltd. O. Naksuriya et al. / Biomaterials 35 (2014) 3365e33833370 the hydrophobic cavity of the CD. Yallapu et al. developed a b - cyclodextrin ( b -CD)-curcumin inclusion complex by solvent evap- oration method. b -Cyclodextrin (CD) was dissolved in deionized water and varying amounts of curcumin in acetone were added while stirring overnight to evaporate acetone. Then, b -cyclodextrin ( b -CD)-curcumin inclusion complexes were recovered by freeze drying. Analysis showed that 1e2 curcumin molecules were encapsulated per b -CD cavity [113]. The same group synthesized poly( b -CD) (molecular weight from 2900 to 4100 Da) which was subsequently loaded with 5e10% of curcumin. Poly( b -CD)/curcu- min self-assembled formulations were prepared by drop-wise precipitation method [114]. TEM analysis showed that a curcu- min/poly( b -CD) inclusion complex (loading: 10e30%) self assem- bled into nanoparticles with a size of 250 nm. An in vitro stability study was performed in PBS and it was noted that >70% of the loaded curcumin was retained in the nanoparticles during 72 h of incubation at pH 7.4 and 37  C, demonstrating a good compatibility of curcumin and its carrier [114]. 2.7. Solid dispersions Solid dispersions are dispersions of a drug/compound (either molecularly dissolved in amorphous or (semi) crystalline form) in an inert matrix [115,116]. Solid dispersions are prepared by melt method or solvent evaporation technique and used to enhance the solubility and dissolution rate of poorly water-soluble drugs [117e 120]. Lyophilized 2-hydroxypropyl- b -cyclodextrin (HP- b -CD)-cur- cumin co-precipitates were prepared by a solid dispersion method [121].HP- b -CD and curcumin (molar ratios from 0.5 to 2.8) were dissolved in methanol and converted into an amorphous co- precipitate which was subsequently lyophilized. The lyophilisates had a porous structure that showed enhanced hydration and dissolution. It was further shown that solutions of the curcumin solid dispersions showed a pronounced decrease in curcumin concentration up to 90% of the loaded amount af ter storage for 168 h, indicating that supersaturated curcumin solutions were formed upon dissolution of the lyophilisates. These HP- b -CD-cur- cumin co-precipitates significantly inactivated Escherichia coli after exposure to blue light (400e500 nm), most likely caused by the photosensitizing activity of curcumin [121]. Seo et al. reported on curcumin-polyethylene glycol-15-hydroxystearate (Solutol Ò HS15) solid dispersions which were prepared by a solvent evaporation method and it was shown that the solubility of curcumin increased to 560 m g/ml. Upon incubation in buffer, 90% the loaded amount of curcumin released/dissolved within 1 h [122]. 2.8. Miscellaneous nanoformulations Mohanty et al. prepared curcumin loaded nanoparticles composed of glycerol monooleate and Pluronic F127 [123]. The entrapment efficiency was around 90% and size of the nano- particulates was 192 nm with a high negative zeta potential (À32 mV) that ensured long term stability and avoided aggregation of the particles. When dispersed in buffer, these nanoparticles enhanced the stability of curcumin by protecting it against hydro- lysis [123]. Anuchapreeda et al. prepared a curcumin nanoemulsion Fig. 5. mPEGePLAeTriseCur: synthetic route and loading and release of curcumin. Reprinted from Pharmaceutical Research, Vol. 29/12, R. Yang, S. Zhang, D. Kong, X. Gao, Y. Zhao, Z. Wang, Biodegradable polymerecurcumin conjugate micelles enhance the loading and delivery of low-potency curcumin, pp. 3512e3525, Copyright (2012), with permission from Springer. O. Naksuriya et al. / Biomaterials 35 (2014) 3365e3383 3371 based on soybean oil, hydrogenated l- a -phosphatidyl choline from egg yolk and co-surfactants (tween 80 and polyoxyethylene hy- drogenated castor oil 60, Cremophor-HR30) with a mean particle diameter of 47e55 nm and with a concentration of curcumin of 0.9 mg/ml. This formulation was stable for 60 days at 4  C. Further, 25% of the loaded amount was released from these nanoemulsions in 72 h when dispersed in PBS, pH 7.4, containing 25% human serum [124]. In another study, curcumin-loaded lipid-core poly(Ɛ- caprolactone) nanocapsules coated with polysorbate 80 (C-LNCs) were prepared by interfacial deposition of preformed polymer. The particles had a mean size of 96 nm, a negative zeta potential of wÀ10 mV and showed 100% encapsulation efficiency [125]. These C-LNCs released 35% of the loaded amount within 2 h [125]. 3. In vitro studies of curcumin nanoformulations The cytotoxicity of curcumin nanoformulations has been stud- ied in many types of cancer cell lines. Interpretation of the rele- vance of the results is often difficult due to the prolonged exposure of cells to high static concentrations of curcumin (either in its free for or as nanoformulation) that however are not necessarily related to the concentrations achieved in vivo. Yallapu et al. demonstrated that the intracellular drug retention of Nano-CUR6 formulation was better than free curcumin (dis- solved in DMSO) due to the sustained release of the active. This formulation also increased the cellular uptake 2 and 6 fold in MDA- MB-231 metastatic breast cancer cells and A2780CP cisplatin resistant ovarian cancer cells, respectively, compared to free cur- cumin. The 50% inhibitory concentrations (IC 50 ) of Nano-CUR6 were 13.9 and 9.1 m M against A2780CP and MDA-MB-231 cells, respectively, whereas the IC 50 ’s of free curcumin were higher than Nano-CUR6 (15.2 m M and 16.4 m M against A2780CP and MDA-MB- 231 cells, respectively) [74]. Apoptosis induction of KBM-5 human chronic myeloid leukemia cells upon incubation with curcumin- loaded PEG-5000-PLGA nanoparticles was investigated by Anand et al. [77]. Curcumin-loaded PEG-5000-PLGA nanoparticles were more potent than free curcumin in inducing apoptosis which could be related to the higher intracellular curcumin concentration upon Fig. 6. Viability of different cancer cells after incubation with curcumin loaded PEG-5000-PLGA for 24 h. Reprinted from Biochemical Pharmacology, Vol. 79/3, P. Anand, H.B. Nair, B. Sung, A.B. Kunnumakkara, V.R. Yadav, R.R. Tekmal, B.B. Aggarwal, Design of curcumin-loaded PLGA nanoparticles formulation with enhanced cellular uptake, and increased bioactivity in vitro and superior bioavailability in vivo, pp. 330e338, Copyright (2010), with permission from Elsevier. O. Naksuriya et al. / Biomaterials 35 (2014) 3365e33833372 incubation with the nanoparticles due their excellent cellular internalization as compared to intracellular concentrations ob- tained after exposure to free curcumin. The uptake of curcumin- loaded PEG-5000-PLGA nanoparticles and free curcumin by KBM- 5 cells was investigated by fluorescence microscopy. PEG-5000- PLGA nanoparticles were taken up already after 5 min exposure and reached a maximum at 30 min. In contrast, the uptake of free curcumin could only be detected after 30 min incubation. The mechanism of cellular uptake of the nanoparticles was not inves- tigated, but they most likely entered the cells by endocytosis [126,127]. However, no differences in viability were observed after explore of the cells to either free curcumin or curcumin-loaded PEG-5000-PLGA nanoparticles (Fig. 6). The authors explained that apoptosis was examined after 24 h of incubation, whereas prolif- eration was examined at 72 h. [77]. Liu et al. reported that curcumin-loaded chitosan/polycarpolactone nanoparticles exhibi- ted cytotoxicity on HeLa cervical cancer cells and OCM-1 human choroidal melanoma cells to the same extent as free curcumin after 48 h incubation [80]. Furthermore, Wichitnithad et al. revealed that mPEG 2000ecurcumin conjugates had IC 50 values in the range of 3e6 m M against Caco-2 colon adenocarcinoma cells and IC 50 values in the range of 1e3 m M against KB oral epidermoid carcinoma, MCF7 breast adenocarcinoma, and NCIeH187 small lung carcinoma cells. mPEG 2000ecurcumin conjugates showed a potency comparable to free curcumin (IC 50 values in the range of 1e3 m M ) on all cancer cells used in this study [103]. These studies demonstrate that nanoparticle-encapsulation of curcumin is not always beneficial. This is also underlined by cytotoxicity studies of curcumin-loaded nanoemulsions on B16F10 mouse melanoma and leukemic cell lines (K562, Molt4, U937 and HL60) by Anuchapreeda et al. [124].It was shown that the 50% inhibitory concentration values (IC 50 ) ranged from 3.5 to 53.7 m M . On the other hand, free curcumin dis- solved in DMSO showed lower IC 50 in B16F10 cells and also in leukemic cell lines as compared to that of curcumin-loaded nano- emulsions. The authors argued that the lower activity of curcumin- loaded nanoemulsions was due to the incomplete release during 24e72 h (incubation time of the formulations with the cells). In the same study, it was shown that leukemic cell lines were less sensi- tive to curcumin both in its free form and as nanoemulsion than B16F10 cells. It was hypothesized by the authors that the pheno- type of B16F10 melanoma cells is responsible for this difference. However, in the same study it was shown that there were no dif- ferences in the IC 50 values of free and curcumin-loaded lipid nanoemulsion in four leukemic cell lines (K562, Molt4, U937 and HL60) [124]. It was found by Bisht et al. that the cytotoxicity of curcumin loaded micelles based on cross-linked random co- polymers of NIPAAm with N-vinyl-2-pyrrolidone and poly(- ethyleneglycol) monoacrylate (nanocurcumin) against pancreatic XPA-1 cells was lower than curcumin in its free form [79]. Dhule et al. developed curcumin loaded HP- g -cyclodextrin liposomes that showed 50% encapsulation efficiency with a size of 98 nm [128]. The cytotoxic activity of these liposomes against KHOS osteosar- coma and breast cancer MCF-7 cell lines (IC 50 ¼ 6 and 12 m g/ml, respectively) was higher than that of curcumin in DMSO (IC 50 ¼ 23 and 20 m g/ml, respectively). Interestingly, non-cancerous mesen- chymal stem cells and skin fibroblasts were unaffected by the nanoformulation but were affected by free curcumin, indicating an improved safety profile. Of note, a RFOS osteosarcoma cell line derived from an untreated osteosarcoma patient was resistant against free curcumin as well as its nanomedicine formulation. The authors argued that this resistance was because of the low curcu- min uptake (either in free form or as nanoformulation) because RFOS cells have a very slow growth rate and a low uptake capacity which is caused by the low metabolic activity of the cell [129].Ina recent study, it was shown that curcumin loaded Pluronic/ polycaprolactone (Pluronic/PCL) block copolymer micelles with a size of 196 nm released 60% of the loaded amount in 108 h [130]. The loaded Pluronic/PCL micelles were evaluated for their uptake by Caco 2 cells using fluorescence microscopy based on curcumin’s intrinsic fluorescence. Cells incubated with the curcumin-loaded micelles showed a higher fluorescence intensity than those incu- bated with free curcumin, demonstrating good cellular internali- zation of the micelles, that, as hypothesized by the authors, occurred via endocytosis. They argued that the insertion of the hydrophobic part of pluronic block copolymers into the lipid bi- layers of cell membranes resulted in a lower membrane micro- viscosity and internalization of micelles [130]. Park et al. loaded curcumin into nanoparticles based on the R7L10 peptide which is composed of a 7-arginine stretch and a 10-leucine stretch which were prepared by an oil-in-water (O/W) emulsion/solvent evapo- ration method [131]. The cationic arginine groups of these peptide micelles were used to make complexes with plasmid DNA. Inter- estingly, the authors found synergistic effects of curcumin on transfection (Fig. 7). The authors hypothesized that the hydro- phobic curcumin stabilizes the structure of the complexes by facilitating the formation of R7L10 micelles. These stable R7L10e curcumin plasmid complexes may show increased endocytosis and cellular internalization resulting in enhanced transfection. The R7L10ecurcumin plasmid complexes also showed anti- inflammatory activity by reducing the TNF- a levels of LPS- activated Raw264.7 macrophage cells. Moreover, it was shown that after intratracheal injection of this R7L10-curcumin formula- tion, a stronger decrease in TNF- a levels in lung tissue in an acute lung injury mouse model was observed than after administration of free curcumin whereas no liver toxicity was detected [131]. Yallapu investigated the hemocompatibility of various curcumin nano- formulations based on PLGA, b -cyclodextrin, cellulose, poly-N- isopropylacrylamide and polyamidoamine dendrimer [132].It was found that the curcumin dendrimer nanoparticles adsorbed more proteins than the other mentioned formulations and had higher lytic activity towards red blood cells, likely caused by the positive surface charge of the dendrimer particles [132]. Several studies have given evidence that drug-resistant cancer cells are sensitive to curcumin. Zhang et al. demonstrated that curcumin showed a similar cytotoxic effect against A549/DDP multidrug-resistant human lung adenocarcinoma cells compared to non-resistant cells [133]. The IC 50 of curcumin at 48 h was 16 m M for A549 cells and 18 m M for A549/DDP cells indicating that the multidrug resistant cells were still sensitive to curcumin. The au- thors also found that curcumin inhibited the expression of miR- 186*, a miRNA that targets caspase-10 mRNA. Inhibition of this miRNA resulted in increased apoptosis as a result of the increased caspase-10 activity in these cells [133]. Duan et al. co-encapsulated doxorubicin and curcumin in poly(butyl cyanoacrylate) nano- particles (CUR-DOX-PBCA-NPs, size 133 nm) prepared by emulsion polymerization and interfacial polymerization [134]. The results showed that CUReDOXePBCA-NPs inhibited the growth of multi- drug resistant human breast cancer cells (MCF-7/ADR) for 97% which was substantially higher than observed for cells incubated with a cocktail of free curcumin and doxorubicin (cell growth in- hibition was only 20%). It is important to notice that the adminis- tration of a nanoformulation loaded with both doxorubicin and curcumin achieved the strongest down-regulation of P-glycopro- tein activity, which is considered to be a major mechanism in multidrug resistance in MCF-7/ADR cells, compared to the combi- nation of free curcumin and doxorubicin. This higher cytotoxicity was ascribed by the authors to the high concentration of curcumin near the cell membrane that bound to P-glycoprotein resulting in inhibition of the dox efflux [134]. In another study, the cytotoxicity of cationic PEGePEI liposomes loaded with curcumin against O. Naksuriya et al. / Biomaterials 35 (2014) 3365e3383 3373 different cell lines, curcumin-resistant B16F10 murine melanoma cells and CT26 colorectal adenocarcinoma cells (obtained by continuously culturing the parental tumor cells in growth media containing 5 m M of curcumin) was investigated [60]. It was found that this liposome formulation, likely because of its rapid cellular internalization, was substantially more cytotoxic (IC 50 of w1 m M ) than free curcumin (IC 50 of w25 m M ). Also, these liposomes showed antitumor activity in tumor bearing mice after intravenous administration (Fig. 8) [60]. 4. In vivo studies of curcumin nanoformulations: kinetics and efficacy The pharmacokinetic, biodistribution and therapeutic efficacy of different curcumin nanoformulations have been investigated in many animal studies in order to get insight into the potential value of these systems for the treatment of different diseases. It has been shown in many studies that oral or intravenous administration of curcumin nanoformulations resulted in a larger area under the concentrationetime curve (AUC) than after administration of cur- cumin in its free form. A more than 40 fold increase in the maximum concentration (C max ) and a 10 fold increase in AUC in mice were observed after an oral dose of 1 g/kg of a curcumin nano-emulsion composed of PEG 600 and Cremophor EL when compared with a suspension of curcumin in 1% methylcellulose [135]. Khalil et al. showed that curcumin loaded PLGA and curcumin loaded PLGAe PEG nanoparticles displayed better pharmacokinetics profiles compared to a curcumin aqueous dispersion after a single oral dose of 50 mg/kg in rats (Fig. 9) [78]. The mean half-lives of curcumin loaded PLGA and curcumin loaded PLGAePEG nanoparticles were 4 and 6 h, respectively, compared to a half-life of free curcumin of 1 h. It was also shown that for the same formulations the C max values were 2.9 and 7.9 fold, respectively, higher than free curcumin and the AUCs were 15.6 and 55.4 fold, respectively, higher than free curcumin. According to the authors, the better performance of the PLGAePEG nanoformulation was due to the more rapid release of curcumin from PLGAePEG nanoparticles than from PLGA nano- particles making the drug quicker available in blood. They also refered to the lack of interaction of the PEGylated systems with Fig. 7. Comparison of transfection activity of R7L10-curcumin/pDNA formulation with other carriers as measured by luciferase assay. Reprinted from Biomaterials, Vol. 33/27, J.H. Park, H.A. Kim, J.H. Park, M. Lee, Amphiphilic peptide carrier for the combined delivery of curcumin and plasmid DNA into the lungs, pp. 6542e6550, Copyright (2012), with permission from Elsevier. Fig. 8. The effect of curcumin/LipoePEGePEI complex (LPPC) on tumor growth in vivo. (A) Balb/c mice were inoculated with CT26 cells and subsequently treated with 2.1 mg/ kg of free curcumin, cationic PEGePEI liposomes (LPPC) or curcumin/LPPC; (B) C57BL/ 6J mice bearing B16F10 tumors were treated with 40 mg/kg of curcumin or curcumin/ LPPC. Reprinted from Nanomedicine : Nanotechnology, Biology and Medicine, Vol. 8/3, Y.L. Lin, Y.K. Liu, N.M. Tsai, J.H. Hsieh, C.H. Chen, C.M. Lin, K.W. Liao, A LipoePEGePEI complex for encapsulating curcumin that enhances its antitumor effects on curcumin- sensitive and curcumin-resistance cells, pp. 318e327, Copyright (2012), with permis- sion from Elsevier. O. Naksuriya et al. / Biomaterials 35 (2014) 3365e33833374 [...]... rats to evaluate the pharmacokinetics of curcumin and the tissue distribution of the nanoparticles The administration of this nanocurcumin formulation (dose: 10 mg/kg) resulted in a w1750 greater Cmax of curcumin (25.5 Æ 6 mg/ml) as compared to curcumin administered as DMSO/PBS (1:1) formulation The AUC of curcumin from nanocurcumin group was 51 Æ 14 h mg/ml, while curcumin after administration of the... release of the formed PEG/ curcumin hydrogels Release is due to hydrolysis of the carbonate bond between PEG and curcumin In the initial stage, the hydrolysis produced some free curcumin and various curcumin conjugates (curcumin conjugated PEG and DTE monomers/oligomers) Then, the conjugates were further hydrolyzed to yield free curcumin A small burst release was observed and the curcumin derived hydrogels... Pelletier J Curcumin- biological and medicinal properties J Pharmacol 1815;2 50e50 [2] Milobedeska J, Kostanecki V, Lampe V Structure of curcumin Ber Dtsch Chem Ges 1910;43:2163e70 [3] Lampe V, Milobedeska J Studien über curcumin Ber Dtsch Chem Ges 1913;46:2235e40 [4] Sandur SK, Pandey MK, Sung B, Ahn KS, Murakami A, Sethi G, et al Curcumin, demethoxycurcumin, bisdemethoxycurcumin, tetrahydrocurcumin and... systems for the formulation of curcumin because of their high loading capacity (15e20%) and small size of 10e100 nm Although nanoformulations of curcumin showed good safety profiles in both animals and humans, more attention should be given on the toxicity of nanoformulations of curcumin, particularly after (repeated) administration of high dose formulations High concentrations of curcumin may increase the... tumor growth in comparison with free curcumin Further, longer survival of mice receiving this dendrosomal curcumin formulation was observed compared to mice that received free curcumin (70 and 54 days, respectively) [150] Recently, Zanotto-Filho et al developed curcumin- loaded lipid nanocapsules for the treatment of gliomas [125] They showed that these curcumin nanoformulations, when administered intraperitoneally... curcumin nano-colloidal dispersion named THERACURMIN (curcumin dispersed in gum ghatti, glycerin and water; formulation prepared using a high pressure homogenizer) at Table 2 Examples of the application of curcumin nanoformulations for prevention and treatment of other diseases than cancer Formulation Dose of curcumin Route of administration Duration of study Animal Disease Observations Reference Curcumin. .. docetaxel resulted in significant reductions in tumor growth, 47% and 58%, respectively [188] As summarized in this paper, nanoformulations of curcumin have shown superior therapeutic effects compared to free curcumin It is therefore a logical step that in future (clinical) studies, curcumin nanoformulations are combined with classical cytostatic drugs such as paclitaxel and doxorubicin [134,189,190] These... 4.5 fold, respectively) compared to free curcumin In another study also supported that the sustained release of curcumin from the PLGAePEGePLGA micelles resulted in improved pharmacokinetic parameters [138] The biodistribution analysis showed that curcumin loaded PLGAePEGePLGA micelles yielded higher curcumin concentrations in lung, brain and kidney and lower curcumin concentrations in spleen and liver... toxicity of curcumin at doses up to 12 g per day [29,168,169] Because of its low bioavailability, in recent years also curcumin nanoformulations have been evaluated in clinical trials In a human pharmacokinetic study [171], a single 650 mg curcumin dose of orally administered solid lipid nanoparticles to healthy volunteers achieved a mean Cmax of 22 ng/ml, whereas no detectable plasma curcumin concentrations... the pharmacokinetic and biodistribution of curcumin loaded PLGAePEGePLGA micelles as well as that of a curcumin aqueous solution also containing 15% of dimethylacetamide, 45% PEG 400 and 40% dextrose [94] A single dose of 10 mg/kg of curcumin loaded PLGAePEGePLGA micelles and curcumin solution were intravenously injected into mice This study revealed that curcumin loaded PLGAePEGePLGA micelles substantially . this paper, nanoformulations of curcumin have shown superior thera- peutic effects compared to free curcumin. It is therefore a logical step that in future (clinical) studies, curcumin nanoformulations are. Studien über curcumin. Ber Dtsch Chem Ges 1913;46:2235e40. [4] Sandur SK, Pandey MK, Sung B, Ahn KS, Murakami A, Sethi G, et al. Curcumin, demethoxycurcumin, bisdemethoxycurcumin, tetrahydrocurcumin. evaluation in preclinical and clinical trials of curcumin nanoformulations, particularly focused on cancer ther- apy. In the next section, different curcumin nanoformulations are discussed with emphasis