Delivering Tamoxifen within Solid Lipid Nanoparticles - Pharmaceutical Technology

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Delivering Tamoxifen within Solid Lipid Nanoparticles
The aim of this study was to prepare and characterize physiochemically and biologically tamoxifen-loaded SLNs to evaluate their effectiveness as a drug-delivery system to treat breast cancers.


Pharmaceutical Technology
Volume 35, Issue 4, pp. 74-79

Results


Table I: Particle sizes, particle-size distribution (PI), specific surface area (Aspec), and zeta potential of solid lipid nanoparticle formulations of tamoxifen.
All data were subjected to one-way analysis of variance followed by post hoc multiple comparison and Duncan tests after verifying that data were distributed normally. The authors used high-pressure homogenization to prepare SLNs because the matrix lipid composed of palm oil (i.e., a triglyceride mixture of natural, hydrogenated, and unbranched fatty-acid chains) was suitable for the incorporation of lipophilic drugs, such as tamoxifen (10). Soy lecithin was the most useful surfactant in SLN dispersions. SLNs and tamoxifen-loaded SLNs were characterized in vitro for particle size, particle-size distribution, and zeta potential (see Table I). In this study, the average size of tamoxifen-loaded SLNs was significantly larger than that of the free SLNs, and the surfaces of tamoxifen-loaded SLNs carried a positive charge.


Figure 1: Micrographs of tamoxifen-loaded solid lipid nanoparticles by transmission electron microscope (bar = 500 nm). (ALL FIGURES ARE COURTESY OF THE AUTHORS)
The transmission electron microscopy (TEM) image of tamoxifen-loaded SLNs is shown in Figure 1, where the particles have a round and uniform shape. The DSC thermogram and melting point of tamoxifen-loaded SLNs is shown in Figure 2. The melting point of the bulk lipid matrix was 58.88 C. Drug-free SLNs prepared using lecithin and oleyl alcohol had a melting point of 57.88 C, and incorporating tamoxifen into the SLNs reduced the melting point to 56.56 C. The WAXD pattern for the bulk lipid was different from that of the nanoparticle, showing relatively sharper peaks compared with SLN and tamoxifen-loaded SLNs (see Figure 3). The WAXD pattern also showed that the typical peak shape associated with free tamoxifen was absent in the tamoxifen-loaded SLN.


Figure 2: Thermograms of bulk lipid, solid lipid nanoparticles (SLNs), and tamoxifen-loaded SLNs (TAM-SLN). The thermograms were recorded within seven days of preparation (scan rate: 5 C/min).
The EE and DL of tamoxifen-loaded SLN were 89.98 1.5% and 17.99 1.9%, respectively (see Table I). The release profile of tamoxifen-loaded SLNs in human plasma is shown in Figure 4. The release rate remained low for the first 8 h, increasing by a mere 2% for that period. Immediately after 8 h, there was a sudden burst of drug release approaching 10% by 11 h after drug incorporation.


Figure 3: Wide-angle X-ray diffraction patterns of bulk lipid, solid lipid nanoparticle (SLN), tamoxifen (TAM), and TAM-loaded SLN.
The cytotoxicity test suggested that tamoxifen-loaded SLNs had an equally efficient cytotoxic effect on MCF-7 cells compared with free tamoxifen. The IC50 of tamoxifen-loaded SLNs on breast-cancer cell lines was generally lower than those for free tamoxifen (see Table II).

Discussion


Figure 4: Entrapment efficiency of three batches of tamoxifen-loaded solid lipid nanoparticles. Mean values are represented (n = 3).
Particle size is an important characteristic for pharmaceutical applications because it significantly affects in vitro and in vivo studies (11). When tamoxifen was incorporated into SLNs, the increase in particle size suggested that loaded tamoxifen was either adsorbed onto the particle surface or entangled in the aliphatic chains of triglycerides. Zeta potential is also an important factor when evaluating the stability of colloidal systems (12). In the presence of 1 mg of tamoxifen, some of the negative charges were neutralized by the complex formation, thus leading to a less negative or positive zeta potential (see Table I). The positive charge also might be raised by the tamoxifen amino group and by tamoxifen localization on the surface of SLNs (13). The TEM image shows that some particles were in the 40–100-nm diameter range (see Figure 1). This particle-size distribution could allow regional drainage if it is directed into or close to the primary tumor or surrounding tissues attacked by cancer cells (14).


Table II: The half maximal inhibitory concentrations (IC50) of tamoxifen (TAM) and tamoxifen-loaded solid lipid nanoparticle (SLN) formulations on MCF-7 cells.
With regard to the melting point and crystallization behavior of SLNs, incorporating tamoxifen reduced the melting point from 57.88 to 56.56 C. Tamoxifen thus is probably in an amorphous state and not crystalline (see Figure 2). A substance with a less ordered crystal or amorphous state requires less energy to overcome lattice forces, and perfect crystalline substances require high melting enthalpy. Hence, the lipid-phase of SLN is less-ordered crystal than the bulk lipid (15). The shape of the bulk-lipid DSC thermogram was a sharp trough, whereas those of the drug-free SLNs and tamoxifen-loaded SLNs were broader (see Figure 2). The shape of the latter could be associated with the surfactant and the dispersion of lipid, and the DSC thermogram of tamoxifen-loaded SLNs displayed only one endothermic melting point (see Figure 2), also indicating dispersion (9). The WAXD pattern for the bulk lipid was different from that of the nanoparticle, showing relatively sharper peaks than for SLN (see Figure 3). In the tamoxifen-free and tamoxifen-loaded SLNs, less-ordered crystals were predominant, and the amorphous state may contribute to high drug-loading capacity (15). This study showed that the tamoxifen was fully entrapped into the SLN during preparation.


Figure 5: Drug loading of three batches of tamoxifen-loaded solid lipid nanoparticles (SLN). Mean values are represented (n = 3).
The EE of tamoxifen-loaded SLNs was quite high (i.e., 89.98%). The palm oil used to prepare the SLN dispersion produced the highest entrapment efficiency. Triglycerides with various fatty acids offer relatively better drug solubilization (16). In 2004, Wong et al. used doxorubicin, verapamil HCl, propranolol HCl, and quinidine sulfate in SLNs stabilized with tween 80, and showed that increasing drug concentration led to a significant increase (p < 0.05) in DL and significant decrease (p < 0.05) in EE (17). These effects were a result of reduced SLN dispersions and high solubility of the drug in high lipid concentrations. Other researchers showed a positive correlation between particle size and drug loading (18, 19).


Figure 6: Human plasma-concentration profile of tamoxifen-loaded solid lipid nanoparticles (SLN). Mean values are represented (n = 3) standard deviation.
Among the factors that support fast drug release from SLNs are a large surface area, small molecular size, low matrix viscosity, and short diffusion distance of the drug (20). In the authors' study, however, the release of tamoxifen from SLN, which contains a lipid matrix, was retarded. Characterization of tamoxifen-loaded SLNs suggested that tamoxifen was either encapsulated within the matrix and membrane of palm oil or entrapped in aliphatic chains. This situation could explain the burst release of the drug from SLNs after eight hours. Considering that SLN is solid at room temperature and that the incorporated drug is released relatively slowly, SLNs have potential as a sustained-release drug carrier. When tamoxifen was incorporated into the SLN carrier system, its antitumoral activity was maintained, suggesting that SLN is a good drug carrier. Tamoxifen-loaded SLNs showed an equally efficient cytotoxic activity against MCF-7 cells, compared with free tamoxifen, and the IC50 of tamoxifen-loaded SLNs was generally lower than that of free tamoxifen. This result indicates that tamoxifen's cytotoxicity may result from improved drug internalization through encapsulation into the SLN matrix and endocytosis (21). A previous study had a similar finding, with reduced MCF-7 cell viability in the presence of tamoxifen-loaded SLN (1). It seems that the improved cytotoxicity of the incorporated drug did not depend on the composition of the SLN. In fact, the IC50 value of drug-loaded SLNs composed of different materials was lower than that of the free drug solution (18).

Conclusion

The tamoxifen-loaded SLNs showed high entrapment efficiency and drug loading. Characterization of tamoxifen-loaded SLNs indicated that the drug was encapsulated within the membrane of palm oil or entrapped in aliphatic chains. The authors concluded that SLN can serve as a good drug carrier for the sustained released of tamoxifen.

Acknowledgment

The authors would like to thank Condea and Lipoid for their kind support with materials, and the Universiti Putra Malaysia for financial support and facilities.

Roghayeh Abbasalipourkabir* is a biochemistry lecturer, and Aref Salehzadeh is an associate professor of toxicology, both at Hamadan University of Medical Science, Hamadan, Iran, tel. +98 811 251 4227, fax +98 811 827 6299,
. Rasedee Abdullah is a professor of biochemistry at University Putra Malaysia.

*To whom all correspondence should be addressed.

Submitted: Apr. 9, 2010. Accepted: Nov. 22, 2010.


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