Tetracycline release from composite cryogels.

Tache, Alina Alexandra ; Stoica-Guzun, Anicuta ; Stroescu, Marta 等

1. INTRODUCTION

Polyvinyl alcohol (PVA) is a hydrophilic polymer capable of forming, in aqueous solutions and under certain conditions, no covalent spatial networks, also known as cryogels. In other words, cryogels are physically cross linked PVA gels prepared using a special method. This method, called freeze-thawing method, is a relatively new one, still being under intensively investigations concerning the freezing temperature, duration of freezing and thawing rate, concentration of polymers and other factors which can influence the final characteristics of the cryogel. Regarding the efficiency, this method requires minimum of energy and time resources, producing less residual solvent, while the main product is easy to scale up. The freezing-thawing method consists of repeated treatments of freezing and melting of the aqueous PVA solutions, in a range from -20[degrees]C up to room temperature (Yang et al., 2008).

PVA hydro gels prepared using freeze/thaw cycles have good mechanical strength, are stable at room temperature, biocompatible, non-toxic and free of initiators and cross-linkers. The main disadvantages of these cryogels are their opaque appearance, limited swelling capacity and thermal stability. In order to improve the PVA cryogels' properties, they are often combined with other polymers like poly (acrylic) acid or even with inorganic materials like hydroxyapatite.

The PVA cryogels or PVA composites cryogels have many biomedical applications as matrices for cell immobilization and for controlled release of drugs (Lozinsky et al., 2003; Nugent et al., 2007). The hydrophilic nature of the cryogel makes it different from the non-hydrophilic matrices regarding the release of the encapsulated component. The most common drug tested for controlled release is insulin.

The aim of this paper is to present experimental data concerning drug release from PVA cryogels and PVA-bacterial cellulose (BC) composites cryogels. Tetracycline was used as the model drug.

2. PREPARATION OF CRYOGELS

In the process of obtaining the cryogels, an aqueous solution of PVA (10% w/w) has been used, together with tetracycline solution of different concentrations and wet biocellulose fibrils (5.5% w/w). The polymer's molecular weight was 60 kDa and the saponification value was 98-99%. The culture medium used to obtain BC microfibrils in an air lift reactor was a modified Hestrin & Schramn (MHS) medium, composed of 2.0% (w/v) glucose, 0.6% (w/v) yeast extract, 0.8% (v/v) lactate, 0.27%(w/v) [Na.sub.2]HP[O.sub.4] and 0.115% (w/v) citric acid. After 7 days, the obtained microfibrils were purified by boiling them in a 0.5 M aqueous solution of NaOH for 30 minutes. Afterwards, the BC microfibrils were washed several times with deionized water until neutral pH of water.

[FIGURE 1 OMITTED]

The aqueous solutions of PVA and different contents of tetracycline and BC were poured into cylindrical moulds.

The cryogels were obtained using three freeze-thawing cycles. Each cycle involved lowering the temperature to -20[degrees]C and maintaining it for 6 hours. Then, the temperature was raised to 25[degrees]C. Each freeze-thawing cycle lasted 12 hours.

Two types of cryogels were made. One is made from PVA and tetracycline (1) and another was obtained from PVA, BC and tetracycline (2). The resultant cryogels used in this work have a cylindrical form, with 14 mm in diameter. Cylinders of different heights were sliced from these cryogels and subsequently studied for drug release. Figure 1 presents some photographs of PVA cryogels used in this work.

3. IN VITRO RELEASE STUDY

In vitro release studies were performed in a vessel under magnetic stirring. A certain quantity of cryogel cylinders having approximately the same height was contacted with a known volume of demineralised water (same ratio solid/liquid).

The tetracycline content was analyzed in the liquid phase at 365 nm using an UV-VIS spectrophotometer CENTRA 6 GBS-Scientific (Australia).

4. EXPERIMENTAL DATA ANALYSIS

The released data was analyzed by applying several release models proposed in literature. The most common mechanism of drug release from cryogels is passive diffusion, but the mechanism of release from cryogels can be classified in: diffusion-controlled, swelling-controlled and chemically-controlled. Diffusion-controlled mechanism is dependent on the mesh sizes within the matrix of the gel; in the case of swelling-controlled mechanism, swelling is considered to be the controlling step for the release behaviour. The chemically-controlled release is determined by chemical reactions occurring within the matrix gel (Hamidi et al., 2008).

To describe the diffusional transport of the drug the following premises were used: at the surface of the dosage the drug concentration is constant; the release of the drug follows a first order kinetics (the release rate is proportional to its concentration); diffusion is isotropic (it does not depend on the spatial direction); the convective process is insignificant, thus it can be neglected; the release of the drug from the cylinders occurs only in axial direction (Siepmann et al., 2006). The model is based on Fick's second law of diffusion (considering only one dimension):

dc/dt = D x ([[partial derivative].sup.2]c/[partial derivative][x.sup.2]]) - kc (1)

where: c--concentration of drug in the cryogel; t--time; D--diffusion coefficient; x--spatial coordinate; k--first order rate constant.

The following initial conditions were used: c=0 for t=0, x [greater than or equal to] a; c = [c.sub.o] for t > 0, x = a and c = 0 for t > 0, x [right arrow] [infinity]; where a--half of the cylinder width and [c.sub.o]--drug concentration at the surface of the cylinder.

Considering non-steady state conditions (drug concentration variation with time and position), the following solution for the differential equation (1) can be derived:

[MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII] (2)

The drug concentration on the axial direction of the cylindrical form can be calculated using equation (2).

5. RESULTS AND DISCUSSIONS

Figure 2 presents tetracycline release for four cryogels with different compositions and different heights. All curves present an initial burst for the first two hours.

To analyze tetracycline release equation 2 was used. From our simulations, parameter k does not influence the results, and it was considered [10.sup.-5] m/s in all experiments. Thus, the only variable parameter remaining is the diffusion coefficient. Two diffusion coefficients were calculated, each for the two types of cryogel: [D.sub.(1)] = 0.95 x [10.sup.-9] [m.sup.2]/s for cryogels containing PVA and tetracycline and [D.sub.(2)] = 1.00 x [10.sup.-9] [m.sup.2]/s for the cryogels containing PVA-BC and tetracycline.

The experimental results and theoretical curves are presented in figure 3 for the cryogels having 5 mm height. A good agreement can be observed between experimental and predicted values obtained from equation (2).

Relatively higher values of the diffusion coefficients proove that the diffusion occurs mainly through the water phase. These values were obtained for cylinders with H < 5 mm, where H is the cylinder height. For cylinders with higher values of H, the agreement between theory and experiment is not conclusive. One possible explanation could be a swelling-controlled mechanism and even an erosion mechanism which were not taken into account in the proposed model.

[FIGURE 2 OMITTED]

[FIGURE 3 OMITTED]

6. CONCLUSIONS

An experimental study of tetracycline release from cryogels is presented. Two cryogel types were used, one containing only PVA and teracycline and the other, a composite one, containing also bacterial cellulose. A simple diffusion model was used to obtain diffusion coefficients of tetracycline from these cryogels in water. The values obtained only for the cryogels with the height smaller than 5 mm are relatively high and very close to the tetracycline diffusion coefficient in water (D = 3.83 x [10.sup.-9] [m.sup.2]/s). These values can be explained by the fact that the diffusion occurs mainly through the water which is contained in cryogels and which can also migrate from the continuous external phase.

There were no significant differences between the values of diffusion coefficients for the cryogels containing only PVA and tetracycline and those containing also bacterial cellulose. Further experiments indicated that bacterial cellulose has the potentioal to enhance mechanical and chemical resistance of cryogels. Nevertheless, more experiments are necessary to elucidate the interactions between polyvinyl alcohol and bacterial cellulose in composite cryogels.

The authors express their acknowledgements to the CNCSIS grants committee, as part of this research has been supported by the IDEI program, project code ID_1031, contract number 177/2007.

7. REFERENCES

Hamidi, M., Azadi, A., Rafiei, P., Hydrogel nanoparticles in drug delivery, Advanced Drug Delivery Reviews (2008), 60, 1638-1649

Lozinsky, V. I., Galaev, I. Yu., Plieva, F. M., Savina, I. N., Jungvid, H., Mattiasson, B., Polymeric cryogels as promising materials of biotechnological interest, Trends in Biotechnology (2003), 32, 445-451

Nugent, M., Higginbotham, C. L., Preparation of a novel freeze thawed poly (vinyl alcohol) composite hydrogel for drug delivery applications, European Journal of Pharmaceutics and Biopharmaceutics (2007), 67, 377-386

Siepmann, J., Siepmann, F., Florence, A.T., Local controlled drug delivery to the brain: Mathematical modeling of the underlying mass transport mechanisms, Review, International Journal of Pharmaceutics (2006), 314, 101-119

Yang, X., Liu, Q., Chen, X., Yu, F., Zhu, Z., Investigation of PVA/ws-chitosan hydrogels prepared by combination [gamma]-irradiation and freeze-thawing, Carbohydrates Polymers (2008), 73, 410-408