Modification of Ceiba pentandra cellulose for drug release applications

2.1 Materials

Kapok fruits were collected from Merida, Mexico, and their fibers were manually removed from the shells and seeds. Benzene, ethanol, sodium hypophosphite (SHP), butane-1,2,3,4-tetracarboxylic acid (BTCA), chlorhexidine acetate (CA), and phosphate-buffered saline (PBS) media were supplied by Sigma Aldrich Co. and were used as received.

2.2 Pretreatment

Kapok fibers are highly hydrophobic due to the presence of natural oils and waxes. The oily components and other impurities were removed by the TAPPI standard method T 204 (17) using a Soxhlet extractor with ethanol–benzene (1:2) for 4 h. Then, the extracted fibers were washed with deionized water and dried at 105°C.

2.3 Cellulose cross-linking

Five grams of the fibers (raw and pretreated) was dispersed for 5 min in an aqueous solution containing BTCA and SHP (50 wt% to BTCA) at different fiber:BTCA ratios (Table 1). The fibers were ground with a blender and dried in an oven at 102°C until constant weight was achieved. After drying, the fibers were cross-linked in an oven at 180°C for 5 min and then cooled to room temperature. Finally, the samples were washed with deionized water to remove the unreacted chemicals and then dried in a vacuum oven at 65°C until constant weight was achieved.

2.4 Chlorhexidine loading

Five hundred milligrams of the cross-linked fibers were immersed in 100 mL of distilled water using a magnetic stirrer for 30 min. CHX (8 and 16 wt% to fiber mass) was dissolved in 15 mL of distilled water using an ultrasonic bath for 30 min at room temperature and then mixed with the immersed fibers using a magnetic stirrer at room temperature for 24 h (the pH was adjusted to 4.1 with H₃PO₄). Finally, the fibers were filtered, washed with deionized water, and dried in a vacuum oven at 60°C until constant weight was achieved (18,19).

2.5 Drug release studies

In vitro studies on CHX release from the kapok fibers were carried out by placing 30 mg of fibers in 40 mL of PBS media (pH = 7.4) and slow magnetic stirring for 24 h at 37°C. Five milliliters of the dissolution medium was withdrawn at regular time intervals and replaced with a fresh solution medium to maintain sink conditions. The withdrawn aliquots were centrifuged, and the CA content in the supernatants was determined by UV spectrophotometer (λ = 264 nm) (18,20,21). The tests were performed in duplicate for each fiber samples.

2.6 Characterization

3.1 Scanning electron microscopy

SEM micrographs of the cross-linked fibers (raw and pretreated) are shown in Figure 1. The fiber shows a cylindrical shape, with a hollow structure and smooth thin walls. The typical diameter is ∼18 μm and the length was over a few centimeters. The fiber structure was preserved even after the solvent pretreatment and cross-linking process. Some fibers appear to be open or torn (marked in circles) as a result of the grinding process; however, the broken fibers were fewer than the intact fibers.

Figure 1

SEM micrographs of kapok fibers crosslinked with BTCA at different ratios: (a) raw fiber 20:1; (b) pretreated fiber 20:1; (c) raw fiber 5:1 and (d) pretreated fiber 5:1.

3.2 Fourier transform infrared spectroscopy

After the ethanol–benzene treatment, the average extractives content of the fibers was determined to be 2 wt% (five measurements were performed). This result is comparable to that reported by Hori et al. (3), who determined the extractives content as 2.2% and 1.8% using the same methodology; on the other hand, Draman et al. (22) and Abdullah et al. (23) reported 5.3% and 3% of wax content using chloroform. The same authors stated that the wax consists of fatty acids, long-chain alkenes, aldehydes, ketones, and small portions of alcohols and esters.

Figure 2 shows the FTIR spectra of the raw and pretreated kapok fibers. The intensity of the band at 3,340 cm⁻¹, corresponding to –OH groups of cellulose, is higher for pretreated fibers (spectra were normalized to an absorbance of 1 at 1,037 cm⁻¹ wavenumber); –OH groups could be more exposed due to the removal of wax and oil from the fiber surface. The bands around 2,980 and 2,830 cm⁻¹ are related to the asymmetric and symmetric stretching vibrations of the C–H bonds of cellulose (24,25); the band at 1,732 cm⁻¹ corresponds to the ester bonds of the carbonyl groups in the hemicellulose and lignin (26,27). The peaks at 1,505 and 1,594 cm⁻¹ correspond to C═C aromatic skeletal vibration of lignin. The band at 1,425 cm⁻¹ is attributed to the bending vibrations of C–H from cellulose and lignin. The peaks at 1,370, 1,161, and 1,035 cm⁻¹ correspond to cellulose, hemicellulose, and lignin, respectively; the first one for C–H bending vibration, the second one for C–O–C asymmetric stretching, and the third one for stretching of the C–O bonded to the aromatic ring of lignin (22,28,29). Finally, the band at 899 cm⁻¹ corresponds to the presence of β-glycosidic linkage between glucose units in cellulose (30).

Figure 2

FTIR spectra of raw and pretreated fibers.

During cross-linking process (Figure 3), ester bonds were formed among cellulose –OH groups and carboxylic groups from the cross-linking agent; as a result, there is a decrease in hydroxyl-to-ester ratio (Abs 3,340 cm⁻¹)/(Abs 1,732 cm⁻¹) from 2.215 to 1.753 for raw fibers and from 2.388 to 1.786 for pretreated fibers, i.e., the hydroxyl bands tend to decrease and ester bands tend to increase. In this sense, the fibers cross-linked with higher concentration of cross-linking agent (i.e., 5:1 fiber:BTCA ratio) exhibited –OH bands at 3,340 cm⁻¹ with relative lower intensity than those using lower concentration of BTCA. Similarly, higher concentration of BTCA produced higher intensity bands at 1,732 cm⁻¹, indicating a correlation between the concentration of cross-linking agent and the cross-linking degree.

Figure 3

FTIR spectra for all fibers crosslinked at different BTCA ratios.

3.3 X-ray photoelectron spectrometry

The surface elemental compositions of all kapok fibers are summarized in Table 2. The analysis of the survey spectra indicates that the fibers consist mainly of carbon (C1s) and oxygen (O1s). In general, oxygen content increased with the concentration of the cross-linking agent, mainly attributed to the introduction of O-containing groups. The pretreated sample exhibited higher oxygen content than the raw sample since the oils and waxes were removed.

A high-resolution scan of the XPS spectra of C1s showed a decomposition in four component peaks around 284.7, 286.1, 287.5, and 289.2 eV, resulting from C1 (carbon atoms bonded to carbon (C–C) and carbon atoms bonded to hydrogen atom (C–H)), C2 (carbon–oxygen bonds of alcohols and ethers (C–O)), C3 (carbon atoms bonded to two non-carbonyl oxygen (O–C–O) or to a single carbonyl oxygen atom (C═O)), and C4 (carbon atoms bonded to a carbonyl and a non-carbonyl oxygen (O–C═O)) (31,32).

In general, C2 and C3 carbon atoms decrease with the cross-linker content since OH groups were consumed during the esterification reaction. In this sense, the results indicate that the C4 carbon atom contribution increased with the cross-linker content regardless of the pretreatment process; the highest contribution was observed in the pretreated sample cross-linked with a 5:1 fiber:BTCA ratio, indicating that new ester bonds were formed. The FTIR results show that the sample with a higher degree of cross-linking is the pretreated fiber modified with a 5:1 fiber–BTCA ratio.

3.4 Elemental analysis

The elemental analysis results are presented in Table 3; it was noted that the nitrogen content increases in the pretreated fibers loaded with CHX. The low N values and the highly hydrophobic nature of the kapok fiber could prevent the CHX diffusion into the hollow structure of the fiber. In the solvent pretreated fibers, the hydrophobicity of the fibers is reduced, allowing CHX loading more efficiently.

The nitrogen content was used to quantify the amount of drug loaded into the cross-linked fibers. The nitrogen content was calculated, considering that a sample of pure CHX, with the chemical formula C₂₆H₃₈C_l2N₁₀O₄ and a molecular weight of 625.5 g/mol, contains 27.70 wt% of nitrogen content. The amount of chlorhexidine in the final products ranged from 3.59 to 7.99 wt%. The concentration of chlorhexidine in the commercially available products ranges from 0.5% to 2.0% either in solution or topical forms; the results of this study suggest the use of kapok fibers as a feasible material for their future development in pharmaceutics.

3.5 Thermogravimetric analysis

Figure 4a shows the TGA thermograms of the raw and pretreated fibers, revealing clearly three stages of weight loss: the first one from 50°C to 100°C, which is related to water from moisture, and the second one from 230°C to 360°C, which is ascribed to the thermal degradation of cellulose and hemicellulose (22,33). The last weight loss is above 360°C related to lignin, which is reported to have a thermal degradation interval from 200°C to 900°C (28). At 700°C, the residual mass of the raw fiber is 14.62% and that of the pretreated fiber is 18.28%.

Figure 4

(a) TGA, (b) DTG of raw and pretreated fibers and (c) deconvolution of DTG curves of raw fiber.

Figure 4b shows the DTG curves of the raw and pretreated fibers. It shows that the raw fibers not only exhibit a higher decomposition temperature than the pretreated fibers (357°C vs. 336°C, respectively) but also a shoulder around 290°C. These facts could be attributed to the elimination of oils and waxes during the solvent pretreatment, decreasing the thermal resistance (34). The deconvolution of the DTG curve (Figure 4c) reveals two overlapped decomposition peaks, corresponding to the degradation of the hemicellulose at 319°C and the degradation of the cellulose at 360°C.

Figure 5 shows the TGA thermograms of the cross-linked fibers with BTCA at different ratios. The temperature of maximum degradation peak decreases with the concentration of BTCA, suggesting that the cross-linking process reduces the thermal resistance. This agrees with the work reported by vom Stein et al. (35), who found that cellulose depolymerization can be catalyzed by carboxylic acids at temperatures above 160°C; also, it is possible that the cross-linking process reduces the hydrogen bonding of cellulose, diminishing the thermal stability of cellulose crystals.

Figure 5

DTG curves of crosslinked fibers.

3.6 Drug-release studies

The cumulative drug release from the cross-linked fibers is shown in Figure 6. In general, the cross-linked fibers showed a burst release within the first hours; then, a slower release was observed. The cross-linked raw fibers showed a burst release in the first 3 h, then the drug release was slower in the next 21 h. On the other hand, the pretreated cross-linked fibers exhibited a burst release within the first 2 h, followed by a slower drug release process. In both cases, the initial burst release is associated with the CHX on the fiber surface, which can be easily released into the buffer solution, and the second behavior could be associated with the diffusion of CHX from the hollow fiber structure. The cross-linked fibers with a higher amount of BTCA showed higher CHX release; higher degree of cross-linking leads to a more porous material, increasing the probability for adsorbing more drug.

Figure 6

Cumulative release of chlorhexidine from Ceiba pentandra s fibers. Raw fibers (top), pretreated fiber (bottom).

3.7 Release kinetics data analysis

Drug release kinetics was modeled using several models (Table 4), with the Korsmeyer–Peppas equation (36), with an n value of 0.25, which describes better the drug release kinetics. According to the model, the n value of 0.25 has been associated with the presence of pores in the polymeric matrix (37) and the high R² values indicate that the drug release depends on the diffusion of the drug from the porous materials obtained by the chemical cross-linking of the fibers. Table 4 summarizes the R² and rate constant k for chlorhexidine release from the cross-linked CP fibers. The rate constant k of the samples are comparable regardless of the amount of CHX loaded. The extracted fibers exhibit higher k values, suggesting that the hydrophobic nature of the raw fibers prevents the out diffusion of CHX to the buffer solution.