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Latin American applied research

versión On-line ISSN 1851-8796

Lat. Am. appl. res. vol.45 no.2 Bahía Blanca abr. 2015

 

A novel enteric polysacharide based on cellulose for sodium diclofenac encapsultaion

V. León, O. López, C. M. García, J. Rieumont§ and E. Bordallo

Dto de Desarrollo, Inst. Cubano de Invest. de los Derivados de la Caña de Azúcar ( ICIDCA). Cuba. vleon69@yahoo.com
Centro de Investigación y Desarrollo de Medicamentos (CIDEM). La Habana, Cuba. caridadgp@infomed.sld.cu
§ Dto. de Polímeros, Fac. de Química, Univ. de La Habana, Cuba.

Abstract— The polymers that respond to chemical changes, such as pH, are used in the modified drugs release. This paper presents the microencapsulation of diclofenac sodium, it has the disadvantage to produce gastric irritation, therefore it is suitable for encapsulation.
As coating was used carboxymethylcellulose (CMC) grafted with benzyl chloride. The microencapsulation method used was spray drying. It was obtained encapsulation efficiency to 90 % and a 57 % yield. Particles were characterized by FTIR, X rays diffraction and SEM.
In vitro dissolution test allowed to evaluate release of the active in simulated gastric and intestinal juice. The results showed excellent response in acid medium giving 2.7 %, result that confirmed the enteric behavior. The kinetic release showed that Peppas model was the best to match the experimental points. Diffusional exponent indicates that the drug transport had non-fickian behavior. The difference between Peppas and Schott models is discussed.

Keywords— Cellulose; Enteric Polymer; Microencapsulation; Sodium Diclofenac

I. INTRODUCTION

Great attention has been devoted to the use of oral-delivery systems of the so-called type "enteric polymers", since these polymers are essentially insoluble in the gastric juice and may be used to impart enteric solubility to the encapsulated drug serving as a drug target device (Vachon and Nairn, 1995;Morishita et al., 1993; Mooustafine et al., 2005 a, b; Raffin et al., 2007; Oosegi et al., 2008)

These polymers are sensitive to pH changes and are able to protect the drug from the degradation action of the enzymes and gastric fluid, that is very acid (pH=1-2) (Moustafine et al., 2006; Haznedar and Dortunc, 2004; Liu et al., 2009), and also to avoid side effects such as gastritis for example in patients that consume the drug daily (Vachon and Nairn, 1995).

The drug sodium diclofenac [sodium phenyl acetate (o-(2.6-dichloro-phenyl) amino] (Fig. 1) presents these drawbacks and by this reason its encapsulation is necessary (Kouchak and Atyabi, 2004; Sánchez et al., 2001).


Fig. 1: Molecular structure of sodium diclofenac

The behavior of the enteric polymers is dependent on protonation state: at higher pH the carboxylic groups become ionized, changing its conformation and expanding it due to the repulsion between the negative charges of the carboxylates. At lower pH the carboxylic groups are not ionized. The conformations are them so closed that the copolymer can precipitate.

This process is mimicking the pH change that accounts in the intestinal track from the stomach to the intestine.

In fact an enteric polymer should have in its structure a hydrophilic moiety and another hydrophobic one. This is the case for the polymer carboxymethylcellulose (CMC) grafted with benzyl chloride obtained by us in a previous communication (León et al., 2013) that posseses an adequate hydrophobic- hydrophilic balance to be pH dependent.

Another CMC derivative obtained by esterification has been reported to possess such characteristics (Kunal et al., 2005).

The aim of the present paper is to present the details of the diclofenac microencapsulated by spray drying with the polymer carboxymethylcellulose (CMC) grafted with benzyl chloride, its characterization by FTIR, SEM and X-ray diffraction, its behavior as an enteric copolymer during pH changes and the release mechanism of drug.

II. METHODS

A. Materials
The polymer Carboxymethylbenzyl cellulose (GS 0.8 (carboxyl) and 0.1 (benzyl)) used was the obtained in the ICIDCA (León et al., 2013). The drug used was sodium diclofenac without further purification. All other chemicals were of analytical grade.

B. Encapsulation of the active ingredient. Preparation of microparticles (MP)
The microparticles were obtained using the spray drying method. To a dispersion of polymer of 30% in distilled water the drug was added in a drug/polymer ratio of 1/3 and homogenized. The dispersion was fed into a laboratory spray drier Büchi (Model 191 B) with the drying air flow and parallel feed. The following parameters remained constant: inlet temperature 120 °C, outlet temperature 80 °C, air flow rate 600 L/h and drying air flow rate 60 m3/ h respectively.

C. Characterization of microparticles: encapsulation efficiency and yield
The encapsulation efficiency, drug loading and yield were determined by the Eqs. 1-3, respectively.

(1)
(2)
(3)

where Enc_eff is the encapsulation efficiency, cal_mass is the calculated mass of drug in microparticles, theo_mass is the theoretical mass of drug in microparticles, drug_load is the drug loading, mass_drug is the mass of drug in microparticles, mass_pol is the mass of polymer, am_mic_obt is the amount of particles obtained and am_mic_exp is the amount of particles expected.

D. FTIR determinations
Fourier transform infrared measuremerents (FT-IR) were taken at room temperature using Bruker IFS 60v. Spectra were recorded in the range 4000-400 cm-1 and samples were prepared in KBr pellets.

E. Diffraction patterns
X-ray diffractograms were recorded with universal X-ray diffractometer, model URD6 flag Carl Zeiss Jena. The sweep was performed at a power and speed 40 kV/20 mA, 3°/min.

F. Size distribution
The size distribution was determined by optical microscopy and image analysis, using Motic Images Plus 2.0 ML software and counting 500 particles with a magnification of 50 times.

G. Scanning Electron Microscopy
Scanning electron microscopy (SEM) was used to analyze the morphology of the microparticles. Samples were adhered to a carbon ribbon and covered by sputtering with palladium (layer of 5 nm) and observed using a FESEM Field Emission ULTRA Plus Zeiss, Germany.

To examine the transformation that takes place in the morphology of the microparticles when they reach the intestinal juice, the sample was contacted with sodium phosphate buffer, pH 6.8 for 45 minutes at 37 °C to simulate the process. After this time, vacuum dried and proceeding similarly to the above explained.

H. In-vitro dissolution studies
For the in vitro evaluation, the microparticles were subjected to dissolution tests according to USP 25 in the two stages to simulate the gastric fluids (hydrochloric acid aqueous solution, pH 1.2) and intestinal (phosphate buffer, pH 6.8). The test was carried out using the apparatus 1 (100 rpm, 500 mL) of a dissolution equipment (SR8 Dissolution Test Station Hanson Research) at 37 ± 0.05 °C. The simulated gastric and intestinal fluids were used in sequence (2h each) to simulate the microparticle transit from stomach to intestine. Firstly at time intervals of 15 minutes and lately at 30 minutes, the samples were withdrawn with a syringe and replaced with an equal volume of fresh medium.

The active was quantified by high resolution liquid chromatography using a chromatograph (KNAUER) with UV/VIS detector (KNAUER) set at 254 nm. The column was RP-8, 10 µm (250 x 4 mm), the mobile phase consisted of sodium hydrogen phosphate solution, pH 2.5: methanol (30:70). The mobile phase was eluted at a flow rate of 1.0 mL/min.

The amount of drug in each case was determined by the quantity of that which is released in each medium at each time. The calculations were performed using the Eq. 4:

(4)

where m(L) is the mass of diclofenac released from the MP, m(P) is the mass of diclofenac in the MP and (released)% is the percentage of diclofenac released from the MP

III. RESULTS AND DISCUSSION

A. Encapsulation data
The encapsulation using the spray drying method gave an encapsulation efficiency of 90 %, a drug loading of 24 % and 57 % yield of microparticles.

B. Evaluation of the polymer/drug interaction
Most of the microencapsulation processes involve an intimate mixing between the polymer and the active ingredient, so that any physicochemical interactions can further influence the therapeutic efficiency of the dosage form. Accordingly, it is convenient to characterize the physical state of the polymer and the active principle separately and also in the microparticle, and evaluating the possible existence of interactions.

The polymer contains hydroxyl groups and carboxymethyl capable of interacting with other molecules through the formation of hydrogen bonds or by electrostatic interactions. Sodium diclofenac shows equally groups capable of interacting with the polymer chains, mainly by the amino groups of the structure. Consequently, it may cause the formation of hydrogen bonds between the amino groups in the diclofenac and hydroxyl and carboxyl groups in the derivative. These interactions can cause a decrease in the electron density of the oxygen atom, resulting signals in the spectrum of the microparticles less intense at the same wavelength.

The spectrum of sodium diclofenac (Fig. 2) shows bands at 3340 cm-1 which correspond to the signal secondary amine 1570 cm-1 characteristic of carboxyl -C=O and 745 cm-1 due to C-Cl bond. In the physical mixture and the encapsulated can be seen that the carboxyl bands is little shifted (Fig 2) if it to compare with polymer.


Fig. 2. FTIR spectra of polymer, diclofenac, physical mixing and encapsulated.

According to the characteristics of the spectra it can be stated the eventual existence of weak interactions polymer/drug microparticles or physical mixing.

X-ray difractogram studies are reported in Fig.3. The diffraction pattern of sodium diclofenac shows a cristalline typically profile, with well defined main peaks at positions 13.3, 14.1, 21.7, 26.1, 31.4 and 36.5 °. In the case of diffraction for the physical mixture of the drug and the polymer it can be seen some peaks of low intensity corresponding to diclofenac present in the mixture. Their presence proves that the active ingredient is dispersed in the polymer maintaining its crystallinity. However, when the drug was loaded into polymer in the form of microparticles, the intensity of each peak markedly decreased. This is a sign of the active inclusion into the polymeric material by the encapsulation method that turns the drug amorphous.


Fig. 3. X-ray difractograms of polymer, diclofenac, physical mixing and encapsulated

C. Size distribution
Figure 4 presents typical pictures of the microparticles obtained by optical microscopy. From the photomicrographs obtained for microparticles produced it was possible to determine the microparticles size distribution. The mean diameters obtained for the microparticles were 7,17 ± 2,89 μm. Figure 4 shows that this particular size distribution is normal Gaussian distribution with size between 1 y 14 mm.


Fig. 4. Frequency histogram represents the particles size distribution

D. Scanning Electron Microscopy (SEM)
Figure 5 (a) shows SEM images of microparticles. As can be seen, microparticles show a smooth surface and spheroidal morphology. The absence of an ideal spherical morphology can be probably attributed to the drying process that can cause invaginations in the particles. The particles tend to agglomerate, probably due to existence attractive electrostatic forces.


Fig. 5. Scanning Electron Microscopy. (a) Microparticles (b) Microparticles after of contact with buffer phosphate during 45 min at 37oC

The micrograph of Fig. 5 (b) confirms the expected behavior of the polymer, by putting it in contact with the buffer of pH 6.8. In this way the material starts to swell as a result of the carboxyl group ionization, the swelling and rupture of the particle under these conditions show that the coating offered to the active polymer loses its function, allowing the release as an enteric polymer does.

E. In vitro release studies
In drug release studies, HCl aqueous solution (pH=1.2) is the typical medium to mimic gastric fluid while phosphate buffered solution (PBS, pH=6.8) is the typical medium to mimic intestinal fluid. And therefore the release mediums with HCl and PBS are used for the present study. The percent drug release from microparticles in different release medium is shown in Fig. 6.


Fig. 6. Percentage of drug release from MP in PBS (pH=6.8) and HCl (pH=1.2) aqueous solution medium

The release profile of the drug shows a pH-dependent behavior according to the results published by Zhang et al. (2012). In fluids simulating gastric content (pH=1.2), during the first 120 min (stomach transit time), diclofenac released was only 2.7 %. This result is

very satisfactory considering that the USP establishes acceptance criteria for this case not exceed 10 %.

By changing the medium it is rapidly observed a steep slope for the curve and the rate of drug release significantly increased. The percentage of diclofenac sodium released starts to increase, yielding 49 % (180 min) and 73 % (240 min).

At pH values above 6 the carboxyl groups are ionized, this causes repulsion between them, that result in conformations that occupy a greater volume. The material swells, allowing diffusion of sodium diclofenac from the polymer matrix into the dissolution medium, in agreement with that observed by scanning electron microscopy (see Fig. 5b).

It is known that the release profile of a drug using pristine CMC as matrix has not a pH dependent behavior as demonstrated by Cai et al. (2011). In our case, substitution of the side groups of the polymer with hydrophobic benzyl groups will provide the balance needed in order to reach the desired behavior.

The above results show that indeed the material serves to protect the active ingredient in acidic environment, enabling release with a change in environmental conditions.

F. Kinetic Models
Apparently the behavior for the zero order, Higuchi and the Peppas equation using all the experimental data did not match a straight line as Schott equation do giving a correlation coefficient of 0.992, (Fig. 7-10).


Fig. 7. % release versus time (min)


Fig. 8. % release versus square root of time (min)


Fig. 9. Plot of the Peppas equation


Fig.10. Plot of t/% versus time according second order Schott equation

However the experimental points for the first portion of the curves for the three models mentioned above gave good results as shown in Table 1. It is related to the fact that according to the first Fick law the flux (release in our case) is proportional to the concentration gradient and for the zero order to be observed it should be constant to the end of release but it do not happen. For the Higuchi equation the solution of the second Fick law equation gives an early approximation depending on t1/2 but the late approximation is of first order.

Table 1 Correlation coefficients, diffusional exponents and AIC values for the enteric release of sodium diclofenac at short time.

Linearity observed for the Schott plot can be the result of solvent uptake synchronization with the drug release. However a system completely ruled by the swelling should be related much more to a zero order that to a second order equation because swelling is a superior stage of chain relaxation.

The transport mechanism can be estimated, if it is ruled by diffusion or relaxation, as shown in Table 1. Correlation coefficients indicate good linearity for the experimental points at the beginning for the three systems but they are too much close each to other. The discrimination of mechanisms can be afforded with the statistical Akaike Information Criterium (Yamoaka et al., 1978; Serra et al, 2006). This criterium (AIC= N ln SSR + 2p where N is the number of experimental points, SSR is the sum of square residuals and p a is parameters numbers) gives lower values for the mechanism that better match the experimental points. Thus, Peppas model is more reliable and the diffusion exponent 0.72 indicates a non fickian behavior where diffusion and relaxation are operating.

Finally it can be appreciated in Fig. 10 that some bias of linearity is observed at the beginning in the Schott plot. The same feature has been reported in literature for the system (de Oliveira, et al., 2009) metronidazole encapsulated in Eudragit RLPO.

Another feature of the sodium diclofenac drug release is that does not attain the 100% keeping around 10-20% occluded by drug-polymer interaction or the physical crosslinkings of the polymer.

IV. CONCLUSIONS

In this study we confirmed the feasibility of use of spray drying for encapsulation of drugs in enteric polymers.

FTIR and X-ray diffraction studies were useful to confirm the effective loading of the sodium diclofenac into the polymer. The microparticles showed a highly sensitive response to the pH of the media, showing that they are able to release the drug preferentially at intestinal fluid.

Peppas equation was adequate to describe in a first approximation the non-fickian drug transport for the sodium diclofenac encapsulated in a matrix of benzylated CMC because it gives information about diffusion and relaxation. Schott equation even giving a good linearity can not explain the complex behavior of a swellable enteric copolymer.

ACKNOWLEDGEMENTS
The authors would like to thank Dr. Mazzimo Lazzari and MSc. Manuel Gómez of University of Santiago de Compostela for the support for the SEM analysis.

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Received: September 11, 2013.
Accepted: September 19, 2014.
Recommended by Subject Editor: María Luján Ferreira.