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Drug delivery systems have centred on non-biodegradable but biocompatible polymers in the past. However, their role as an implant is severely limited because it requires surgical removal after it has served its purpose. This led to the development and investigation of biodegradable polyesters in drug delivery systems (1, 2). Polyester microspheres have been extensively studied in the scientific and pharmaceutical sector. Polyesters include poly(lactic-co-glycolic acid) (PLGA), polylactic acid (PLA) and poly(caprolactone)(1, 3). The application of PLGA and PLA microspheres as drug delivery vehicles are rapidly expanding due to its biodegradable and biocompatible properties(4). They are also approved by the U.S. Food and Drug Administration as they are free from biological by-products and pathogens(4, 5). Microspheres consist of a porous inner matrix and variable surface, from smooth and permeable to uneven and nonporous. The encapsulated drug of choice is distributed throughout the internal matrix. Typically, the size range of microspheres is between 1 to 500 Âµm in diameter(3).
Chemical and Physical Characteristics of PLGA
PLGA is synthesized by ring-opening co-polymerisation of two monomers, lactic acid and glycolic acid which are linked via ester bonds (Figure 1)(6). The physicochemical properties of PLGA are influenced by the molar ratio and the sequential arrangements of the glycolide and lactide monomers. Lactic acid exists in both D and L stereoisomers. The L isomer occurs in vivo whereas D,L-lactic acid is usually used in synthesis. The disordered conformation of D,L-lactic chains is due to L-lactic acid, and further addition of glycolic acid renders the molecules even more disoriented(3).
(C3H4O2)x(C2H2O2)y + 2H2O Catalyst PLGA
Figure 1: Synthesis of Poly(lactic-co-glycolic acid)(3).
Amorphous vs Crystalline State
Poly(glycolic acid) and poly(L-lactic acid) homopolymers are crystalline, whereas poly(D,L-lactic acid) and PLGA consisting of both D,L-lactide and less than 85% glycolide are amorphous(1, 3). Crystalline solids have definite shapes and its units are arranged in an orderly behaviour. In contrast, the amorphous state is defined by random and unorganised arrangements of molecules(7). The amorphous or crystalline nature of solids influences its therapeutic properties. The crystalline homopolymers degrades slower than the less crystallised copolymers. Thus, PLGA hydrolysis increases as the amorphous content of the copolymer increases(5). Furthermore, the half-life of these polymers can be increased by the addition of a more hydrophobic comonomer, such as poly(caprolactone), as it lengthens the degradation period by lowering its water uptake. Generally, a 50:50 mol ratio is employed in microsphere fabrication. However, different ratios may be used where other factors are more important than the degradation rate(3).
Thermal analysis has been increasingly employed in the pharmaceutical sector for quality control of characterisation and identification of compounds, moisture content, amorphous content, stability and compatibility with excipients. It is a useful technique to investigate structural changes in polymers by measuring heat changes during transitions(8). Common applications in microsphere technology include differential scanning calorimetry (DSC) and thermogravimetric analysis (TGA). Generally, these methods involve heating a sample under controlled conditions while observing the chemical and physical changes that occur(7, 9).
i) Differential scanning calorimetry (DSC)
DSC is the most widely used method of thermal analysis within the pharmaceutical field. With regard to polyester microspheres, DSC is mainly used to measure glass transition of polymers, determination of possible plasticisation through the use of excipients, physical aging and gamma radiation effects(10). Generally, DSC is used to measure the heat flow into and out of a system. It compares the difference between the energy acquired or released by a sample and a suitable reference as a function of temperature or time, while they are subjected to a controlled temperature increase(7). Hence, any factor affecting the heat flow characteristics within the DSC will affect the quality of the data produced. The extent of heat flow is governed by the thermal resistance of the system, and the temperature difference between the reference and sample can be measured by:
dQ/dt = (TR -TS )/ R (Equation 1)
where dQ/dt is the heat flow, TR and TS are the reference and sample temperature, and R is the thermal resistance between sample and reference. Thus, it can be seen that the main factors which affects the measured heat flow in DSC is the difference between TR and TS , R, heating rate, heat capacity and kinetic events. These factors must be taken into account and controlled in order to obtain reliable data which is a representative of the sample's properties(11).
Moreover, DSC is useful in measuring thermal transitions such as glass transition temperature, melt temperature and degradation or decomposition temperature(12, 13). Crystal changes and amorphous behaviour can also be detected by DSC at below room temperature(11). Furthermore, protein stability in solution can be assessed through repeated thermal scans by measuring the midpoint temperature of denaturation, area and shape analysis of heat adsorption peak(2). Advantages of using DSC in experiments include small sample size requirements, wide temperature ranges and rapidity of measurement(11).
There are two types of conventional DSC instruments, which are heat flux and power compensation. Recently, a newer instrument was introduced called modulated temperature DSC (MTDSC) as an extension to the conventional types. This method involves the application of a perturbation to the heating program of a conventional DSC combined with a mathematical procedure to distinguish different types of sample behaviour. Basically, the general approach to this method is to apply a sinusoidal variation of temperature superimposed on the usual linear heating signal. Although the underlying heating process may be equivalent to that of a conventional DSC run, the sample temperature is oscillated in a sinusoidal manner. Hence, resulting in the heat flow being modulated (Figure 2)(8, 11).
Figure 2: Schematic presentation of temperature as a function for conventional DSC and MTDSC. (Adapted from Duncan QMC, Mike R, Thermal analysis of pharmaceuticals,CRC Press/Taylor & Francis; 2007.)
Thermal properties of PLGA are affected by in vitro degradation process. The main factors influencing this are the molecular weight and fluid uptake. As the polymer undergoes degradation, its molecular weight decreases as well. The effects of degradation on DSC thermograms are illustrated in the first DSC scan (Figure 2) and second scan (Figure 3). The differences obtained are due to physical aging of the polymer and fluid uptake. From both figures, it can be seen that glass transitions appeared less distinguished and at a lower temperature as the aging time increases. Thus, at elevated temperatures, the stability of polymers decreases with increasing aging time, mostly due to a reduction of molecular weight(12).
Figure 2: DSC Thermogram on effects of degradation (First scan)(12). Figure 3: Thermogram on effects of degradation (second scan).
ii) Thermogravimetric Analysis (TGA)
TGA is used to measure the difference in weight of a polymer while it is heated. Chemical and physical processes which occur during heating can be measured. Furthermore, TGA can be used to measure thermodynamics quantities as well as study the thermal behaviour of liquid reactants and gas-solid reactions(11). A vacuum recording balance with a sensitivity of 0.1Âµg is employed to record the polymer weight under pressures of 10-4 mm to 1 atm. TGA can also be used to determine polymer stability and decomposition kinetics. To determine whether the desolvation is attributed to water or residual solvents from chemical processing, Karl Fisher analysis is used.
Karl Fisher analysis is a potentiometric titration method to determine the amount of water associated with a solid material. This method is utilised in pharmaceutical applications to study the humidity effects in solids undergoing water sorption from air, as well as in quality control efforts to demonstrate the amount of water associated in different polymers(7). Consequently, TGA also proves to be a simple method to measure weight loss which is more rapid and convenient than Karl Fisher analysis. However, great care is necessary with respect to the assumptions made concerning the material loss is water, as they may be decomposition of the substrate material or volatisation of other residual solvents. Additionally, when an amorphous material is being studied, it is vital to know its water content when measuring the Tg.
Glass Transition Temperature (Tg)
Tg occurs in amorphous solids and can be described as a transition in the heat capacity of polymers during heating. Below the Tg, the amorphous polymer chains exist in a 'glassy and brittle' state while becoming 'rubbery' at temperatures above it(7). They differ from crystalline solids as they tend to flow when subjected to sufficient pressure over a period of time. The mechanical strength is affected and therefore is reduced above the Tg. Similarly, the internal matrix of the PLGA microsphere is also dependant on the Tg, and as water enters the microsphere, Tg is decreased, hence initiating a rapid increase in chain mobility, water uptake and drug release(14). If the microspheres are exposed to temperatures above the glass trasition temperature (Tg) of the amorphous region, then the degradation rate will increase as well. Tg of PLGA copolymers rises with increasing molecular weight and lactide content. Thus, PLGA with molecular weight of 14 kDa with a 50:50 mol ratio has a Tg around body temperature, while a higher molecular weight PLGA has a Tg of 40-50Â°C(12). On the contrary, crystalline solids have definite melting points, hence passing sharply from the solid state to liquid state. The melting point of crystals can be defined as the temperature at which solid and liquid are in equilibrium(7).
Degradation and erosion of polymers
Degradation and erosion occurs in all polymers. Generally, the two methods which polymers degrade by are surface erosion and bulk erosion (Figure 4)(15). In the former, the degradation rate is faster than the water ingress into the matrix, hence eroding only at the surface(16). The slow water uptake results in heterogeneous dispersion throughout the matrix, consequently causing a reduction in diameter size. This is attributed to the autocatalytic action of an increased amount of carboxylic acid end groups in the microsphere core(14). In the latter, water rapidly penetrates the whole microsphere before surface erosion occurs. Water disperses homogeneously throughout the matrix, such that the original size of the microsphere is maintained for a longer period, while erosion begins from within. Studies demonstrated that PLGA microspheres only undergo surface erosion above pH 13 or is of an extremely large size(14). Therefore, it is generally accepted that polyester microspheres degrade via bulk erosion at physiological pH. Various techniques such as gel permeation chromatography (GPC), differential scanning calorimetry(DSC) and scanning electron microscopy(SEM)were used during the this period to demonstrate the degradation process(14).
Figure 4: Schematic illustration of the changes a polymer matrix undergoes during surface and bulk erosion(17).
i) Degradation Process
Degradation occurs via a hydrolytic mechanism and is described by random chain scission on the ester bond linkages which are cleaved to form oligomers and finally, monomers(14). During this state, the molecular weight of the polymer decreases while the mass of the microsphere remains unchanged(17). There are many factors modulating the degradation rate of lactide/glycolide copolymer microspheres and they are indicated in Table 1(5, 14).
Factors Affecting Degradation Rate of Polyester Microspheres
Molecular weight and molecular weight distribution
Glass transition temperature (glassy, rubbery)
Morphology (crystalline/ amorphous)
Stability of ester bond
The degradation rate of PLGA is affected by the ratio of hydrophilic polyglycolic acid to hydrophobic PLLA. Higher glycolide to lactide mole ratio in copolymers increases the rate of degradation due to ester bonds in neighbouring glycolic acid having high hydrolytic activity(4, 5, 15). The slower degradation of polylactic acid can be attributed to the steric effects of alkyl groups hindering the attack of water. However, studies showed that the fastest degrading system was 50 mol% glycolic acid and 50 mol% lactic acid. Large molecular weight distributions indicate relatively large numbers of carboxylic acid end groups, thus facilitating the autocatalytic degradation of polymers. Autocatalysis arises from the water uptake of polymers and from degradation which creates an increased amount of carboxylic acid end groups(17). Hence, wide ranges of molecular weight distribution accelerate the degradation rate(14).
However, studies illustrated that PDLA microspheres demonstrated different degradation characteristics relating to different polymer molecular weights. This was due to the modification in polymer morphology in the hydrated state. Lower molecular weight PDLA microspheres were rubbery at incubation temperatures and tend to degrade much faster than its higher molecular weight counterparts which remained in a glassy state at the same incubation condition(14).
In semicrystalline polyesters, the amorphous region degrades before the crystalline domain of the microsphere. During the degradation process, crystallinity of the polymer progressively increases, hence resulting in a highly crystalline material which has much more resistance to hydrolysis than the starting polymer. This is explained by an increase in mobility of the partially degraded polymer chains due to a higher degree of entanglement enabling the polymer chains to be aligned in an ordered crystalline manner. The rate of biodegradation is also influenced by the porosity of microspheres, particularly when the pore dimension is large enough to permit cellular migration into pores of the microsphere(5, 18). Consequently, end-capping the polymer with a lactic acid ethyl ester instead of a free COOH terminal delays degradation(19, 20).
ii) Erosion Process
The erosion process of polymers is much more complicated than degradation because it consists of many processes, such as degradation, swelling, dissolution and diffusion of monomers and oligomers and morphological changes(19, 20). Erosion of polyester microspheres designates material loss due to monomers and oligomers leaving the polymer. As water enters the polymer bulk, it begins to swell and polymer degradation is triggered, hence leading to the formation of monomers and oligomers. As the polymer gradually degrades, the microstructure of the bulk is altered through the formation of pores, via which the monomers and oligomers diffuses to the external medium(20). As erosion proceeds, the polymer becomes more porous, which can be detected by mercury intrusion porosimetry. Loss in mass of the microsphere is observed as monomers and oligomers are released(15). An increase of molecular weight of PLGA polymers enhances its resistance to erosion significantly for both end-capped as well as non-end capped PLA and PLGA. Nevertheless, despite increasing the duration of erosion, the general profile of bulk erosion remains unchanged(17, 21).
It is essential to recognise that degradation and erosion are two different concepts. Degradation is the loss in molecular weight of polymers while erosion is the loss in mass of the microsphere. Understanding of the degradation-erosion mechanism is important to control and predict the 'burst release' profile of drugs from microspheres(3).
PLGA microspheres are used therapeutically to bring about controlled release of drugs. However, a 'burst release' is often observed, where more than 50% of the protein is released within the first 24 hours from the microsphere(22). Generally, the burst release is thought to arise from solubilisation and diffusion of proteins being loosely associated within the porous network of the microsphere matrix(3). Subsequently, there is a remarkable reduction in the rate of protein release which continues for over 2 months(23). The rate of protein release is dependent on the molecular weight of the protein, microsphere size and degradation rate of the polymer. Higher copolymer concentration and molecular weight results in a smaller burst release due to the decrease in porosity of the microsphere.
In summary, the formulation of polyester microspheres is likely to be continually studied for drug delivery. Its biodegradable and biocompatible property has proved that PLGA will remain to be utilised in the pharmaceutical industry for years to come. However, the underlying mechanisms of its thermal properties and degradation mechanism need to be studied in greater detail to have a greater understanding on predicting its burst release profile.