Optimization of Curcumin Nanocrystals as Promising Strategy for Nose-to-Brain Delivery Application

Abstract

Intranasal (IN) drug delivery is recognized to be an innovative strategy to deliver drugs to the Central Nervous System. One of the main limitations of IN dosing is the low volume of drug that can be administered. Accordingly, two requirements are necessary: the drug should be active at a low dosage, and the drug solubility in water must be high enough to accommodate the required dose. Drug nanocrystals may overcome these limitations; thus, curcumin was selected as a model drug to prepare nanocrystals for potential IN administration. With this aim, we designed curcumin nanocrystals (NCs) by using Box Behnken design. A total of 51 formulations were prepared by the sonoprecipitation method. Once we assessed the influence of the independent variables on nanocrystals' mean diameter, the formulation was optimized based on the desirability function. The optimized formulation was characterized from a physico-chemical point of view to evaluate the mean size, zeta potential, polidispersity index, pH, osmolarity, morphology, thermotropic behavior and the degree of crystallinity. Finally, the cellular uptake of curcumin and curcumin NCs was evaluated on Olfactory Ensheathing Cells (OECs). Our results showed that the OECs efficiently took up the NCs compared to the free curcumin, showing that NCs can ameliorate drug permeability.

Keywords: Box Behnken design; amorphism; bottom-up; crystallinity; experimental design; natural compound; olfactory ensheathing cells; response surface methodology; sonoprecipitation.

Figure 1

3D surface of the effect of (A) X1 = A: curcumin concentration (mg/mL) versus X2 = B: surfactant concentration (% w/v), (B) X1 = A: curcumin concentration (mg/mL) versus X2 = C: solvent to antisolvent (S/A) ratio (V/V) (C) X1 = B: surfactant concentration (% w/v) versus X2 = C: S/A ratio (V/V), on the nanocrystals’ (NCs) size using Poloxamer 188. 3D surface of the effect of (D) X1 = A: curcumin concentration (mg/mL) versus X2 = B: surfactant concentration (% w/v), (E) X1 = A: curcumin concentration (mg/mL) versus X2 = C: S/A ratio (V/V) (F) X1 = B: surfactant concentration (% w/v) versus X2 = C: S/A ratio (V/V), on the NC size using PVP. 3D surface of the effect of (G) X1 = A: curcumin conc. (mg/mL) versus X2 = B: surfactant concentration (% w/v), (H) X1 = A: curcumin conc. (mg/mL) versus X2 = C: S/A ratio (V/V) (I) X1 = B: surfactant concentration (% w/v) versus X2 = C: S/A ratio (V/V), on the NC size using Polysorbate 80.

Figure 2

Contour plot of the effect of (A) X1 = A: curcumin concentration (mg/mL) versus X2 = B: surfactant concentration (% w/v), (B) X1 = A: curcumin concentration (mg/mL) versus X2 = C: S/A ratio (V/V) (C) X1 = B: surfactant concentration (% w/v) X2 = C: versus S/A ratio (V/V), on the NC size using poloxamer 188. 3D surface of the effect of (D) X1 = A: curcumin concentration (mg/mL) versus X2 = B: surfactant concentration (% w/v), (E) X1 = A: curcumin concentration (mg/mL) versus X2 = C: S/A ratio (V/V) (F) X1 = B: surfactant concentration (% w/v) versus X2 = C: S/A ratio (V/V), on the NC size using PVP. 3D surface of the effect of (G) X1 = A: curcumin conc. (mg/mL) versus X2 = B: surfactant concentration (% w/v), (H) X1 = A: curcumin conc. (mg/mL) versus X2 = C: S/A ratio (V/V) (I) X1 = B: surfactant concentration (% w/v) versus X2 = C: S/A ratio (V/V), on the NC size using Polysorbate 80.

Figure 3

Mean size and polydispersity index (PDI) of NCs before and after freeze-drying without and with mannitol as cryoprotectant. BF = before freeze-drying; AF = after freeze-drying; 5% Mann = 5% (w/V) mannitol; 10% Mann = 10% (w/V) mannitol. Tukey’s test for NC size and PDI. The * symbol denotes statistical significance difference for size; The # symbol denotes statistical significance difference for PDI. Significance was defined as **** p < 0.0001.

Figure 4

(A) Differential scanning calorimetric (DSC) thermograms of curcumin NCs with and without mannitol, curcumin (Curc), Poloxamer 188 (P188), mannitol (Mann) and the physical mixture (p.m.) of curcumin-mannitol and curcumin-poloxamer 188; and curcumin-poloxamer 188-mannitol. (B) XRPD spectra of the samples. Each sample was reported with the specific max intensity; (C) FT-IR analysis of all samples tested. Details of named samples: curcumin (Curc); mannitol (Mann); poloxamer 188 (P188); physical mix curcumin and poloxamer 188 (p.m. Curc-P188); physical mix curcumin, mannitol and poloxamer 188 (p.m. Curc-Mann-P188); physical mix curcumin and mannitol (p.m. Curc-Mann); nanocrystals (NCs); nanocrystals with mannitol (NCs Mann).

Figure 5

Scanning electron microscopy of different samples (X5000). Details of named samples: curcumin (Curc); mannitol (Mann); poloxamer 188 (P188); physical mix curcumin and poloxamer 188 (p.m. Curc-P188); physical mix curcumin, mannitol and poloxamer 188 (p.m. Curc-Mann-P188); physical mix curcumin and mannitol (p.m. Curc-Mann); nanocrystals (NCs); nanocrystals with mannitol (NCs Mann).

Figure 6

Stability study of lyophilized NCs stored at room temperature and analyzed by photon correlation spectroscopy (PCS): A) mean size and PDI: B) zeta potential (ZP) and C) DSC analysis up to 6 months. Tukey’s test for NC size and ZP. The black * symbol indicates statistical significance difference vs. T0 group; The red * symbol indicates statistical significance difference vs. T1 group; The blue * symbol indicates statistical significance difference vs. T3 group. Significance was defined as *** p < 0.001 **** p< 0.0001.

Figure 7

The internalization and uptake of curcumin (A): 0.1 µM; 0.5 µM; 5 µM; and Curcumin NCs (B): 0.1 µM; 0.5 µM; 5 µM into OECs. Scale bar: 30 µm.