Elsevier

Il Farmaco

Volume 57, Issue 6, 22 May 2002, Pages 427-433
Il Farmaco

Gamma irradiation effects and EPR investigation on poly(lactide-co-glycolide) microspheres containing bupivacaine

https://doi.org/10.1016/S0014-827X(02)01220-XGet rights and content

Abstract

The effects of γ radiation on the stability of microspheres made of a polylactide-co-glycolide 50:50 copolymer (PLGA) and loaded with 40% bupivacaine (BU) were studied. The radiolysis mechanisms of BU and BU-loaded microspheres were investigated by using electronic paramagnetic resonance (EPR) analysis. Microspheres were prepared by means of a spray drying method. γ Irradiation was carried out in the open, at the dose of 25 kGy, by using a 60Co source. The stability of BU-loaded microspheres was evaluated over a 1-year period on the basis of drug content and dissolution profile. Non-irradiated microspheres were stable over the whole period under consideration. Immediately after irradiation the amount of BU released after 24 h from irradiated microspheres increased from 17 to 25%; in the following 3 months of storage it increased to about 35%, and then it kept constant for 1 year. Radicals generated by BU irradiation were identified by EPR analysis; the sensitivity to γ radiation of BU was about four times lower than that of PLGA. Furthermore, the EPR spectra of loaded microspheres showed that the relative abundance of BU radicals plus PLGA radicals was proportionate to the electronic fractions of the components; this implies that no spin transfer BU/PLGA had occurred during γ irradiation.

Introduction

Biodegradable polyesters based on glycolic acid (PGA) and lactic acid (PLA) and their copolymers poly-(d,l-lactide-co-glycolide) (PLGA) are often used as drug carriers for the development of parenteral drug delivery systems. As they are susceptible to heat sterilisation, ionising radiations are frequently used for final sterilisation or for sanitisation after aseptic preparation.

The investigations on the radiolytic pattern of these drug-delivery systems are useful in the evaluation of their safety.

The predominant effect due to γ radiations on PGA is thought to be chain scission within a radical mechanism where cage effects play a major role, and leads to a concentration of the radiation damage in amorphous regions [1]. Chain degradation leads to a faster loss of tensile strength and to an enhanced hydrolysis rate. A radiolysis mechanism mainly based on radical recombination, disproportionation and H abstraction reactions was postulated [1]. A similar behaviour was observed for PLA and PLGA with the exception of the enhanced sensitivity to radiation degradation, which is attributed to the presence of a tertiary carbon centre [1], [2]. The relative importance of chain scissions and cross linking and their effects on average molecular weights and polydispersity index were investigated by several authors adopting thermal, viscometric and chromatographic methods [3], [4], [5]. Conflicting conclusions were reached as regards the relative importance of random chain scissions versus end chain scission.

The radiation chemistry investigations were also extended to the polymer/drug interactions with interesting results pertaining to the nature and role of polymer interaction. Lalla et al. [6] described a decreased release of piroxicam from γ irradiated PLA microspheres. Volland et al. [7] showed the same pattern from γ irradiated PLA microspheres containing captopril. On the contrary, Yoshoka et al. [5] described an increased progesterone release from irradiated PLA microspheres in relation to the irradiation dose. Mohr et al. [8] showed accelerated kinetics of estradiol release by increasing irradiation doses, due to dose-dependent polymer degradation.

The influence of drug loading on polymer degradation is also discussed. PLA degradation was independent from methadone loading [9], but it was higher when prometazine loading increased [10]. Bittner et al. [11] showed that PLGA degradation rate slowed down following the incorporation of tetracycline in the microspheres.

Although the effects of ionising radiation on PLA and PLGA molecular weight and drug release were discussed by a number of papers, very few works were focused on the ageing of irradiated microparticulate systems [12], [13].

In this work, the research is focused (a) on the evaluation of the stability of PLGA microspheres loaded with bupivacaine (BU) and (b) on the elucidation of radical components production (release), of the radiolytic mechanisms and of their modifications as a consequence of drug–polymer molecular interactions, by matrix EPR spectroscopy.

Placebo and BU-loaded microspheres were prepared by means of a spray drying method.

Microspheres were irradiated by a 60Co source in the open, at the dose of 25 kGy. A minimum absorbed dose of 25 kGy is regarded as adequate for the purpose of sterilising pharmaceutical products without providing any biological validation [14].

The stability of non-irradiated and irradiated microspheres loaded with BU was evaluated for a period of more than 1 year, on the basis of their drug content and dissolution profile.

BU was selected because it is a local anaesthetic drug, usually administered by parenteral route for the regional control of major pain and regional anaesthesia, together with a consequent decreased systemic administration of narcotic drugs. In both regional control of major pain and regional anaesthesia, the development of prolonged drug delivery systems appears interesting in order to avoid repeated administration or infusion via indwelling catheters. LeCorre et al. [14] investigated the use of PLA microspheres for the controlled spinal delivery of BU; promising results were obtained in the biopharmaceutical and pharmacodynamic evaluation of the anaesthetic action in rabbit.

Section snippets

Materials

Polylactide-co-glycolide 50:50, (PLGA) Resomer® RG 503, inherent viscosity 0.39 dl/g, 34 000 Mw (Boehringer Ingelheim KG, Ingelheim am Rheim, G).

Bupivacaine hydrochloride (Sigma, St. Louis, MO, USA); bupivacaine base (BU) was obtained from bupivacaine hydrochloride following the method of LeCorre et al. [14].

l-Alanine; 1,1-diphenyl-2-picrylhydrazyl (DPPH) (Fluka, Milan, I).

Paraffin wax (Aldrich, Milan, I).

Unless specified, all other compounds were of analytical grade.

Preparation of microspheres

Microsphere preparation was

Morphology

Scanning photomicrographs of the drug-loaded microspheres before irradiation and after irradiation at 25 kGy are shown in Fig. 1a and b, respectively. BU-loaded microspheres had spherical shape and porous surface. Microsphere size, as determined by granulometric analysis, resulted in a range between 2 and 5 μm (Fig. 2). The γ irradiation did not seem to cause morphological changes.

Conclusions

The non-irradiated BU-loaded microspheres stored at 4 °C were stable over the period of 1 year.

γ Irradiation at the dose of 25 kGy caused a decrease of BU content of about 4% due to radiolytic decomposition of the active ingredient.

Immediately after irradiation the in vitro drug release increased and further increased in the following 90 days of storage; then it kept constant for 1 year. This behaviour suggested that a stabilisation of microsphere structure occurred 90 days after irradiation.

EPR

Acknowledgements

This research is supported by a grant for the project “Sterilization processes and their influence on drug properties”, Istituto Superiore di Sanità. The authors wish to thank Dr. P. Riccardi, Centro Grandi Strumenti, University of Pavia for SEM analyses.

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