g emﬁbrozil encapsulation and release from microspheres and

8/12/2019 G emfibrozil encapsulation and release from microspheres and

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European Journal of Pharmaceutical Sciences 17 (2002) 207216

www.elsevier.nl/locate/ejps

Gemfibrozil encapsulation and release from microspheres and

macromolecular conjugatesa a , a a b* Anita Martinac , Jelena Filipovic-Grcic , Monika Barbaric , Branka Zorc , Dario Voinovich ,

aIvan Jalsenjak

aFaculty of Pharmacy and Biochemistry, University of Zagreb, A. Kovacica 1, 10 000 Zagreb, Croatia

bDepartment of Pharmaceutical Sciences, Faculty of Pharmacy, University of Trieste, P. le Europa 1, 34127 Trieste, Italy

Received 28 February 2002; received in revised form 5 September 2002; accepted 11 September 2002

Abstract

The purpose of this study was to evaluate and compare the ability of the macromolecular conjugates and microspheres to modify the

release rate of gemfibrozil (Gem). Gem was covalently linked to two similar polymers: poly[ a,b-(N-2-hydroxyethyl-DL-aspartamide)]

(PHEA) and poly[a,b-(N-3-hydroxypropyl-DL-aspartamide)] (PHPA) by an ester linkage. The polymerdrug conjugates obtained

(PHEAG(13) and PHPAG) differ in weight-average molecular weight, length of spacer and Gem content. Microspheres, composed of

chitosans of different molecular weight alone or as a mixture with (2-hydroxypropyl)methylcellulose (HPMC), PHEA or PHPA and with

different theoretical polymer /drug ratio (2:1 and 3:1, w/ w) were prepared by spray drying. The microparticulate systems were

morphologically characterised by scanning electron microscopy, particle size analysis and Gem content was determined. In vitro

dissolution tests were performed to evaluate the feasibility of conjugates and microspheres in modulating Gem release. The results

obtained show that microspheres are always suitable to modulate Gem release and that the best conditions are achieved by microspheres

composed of the low molecular weight chitosan (CL) combined with PHPA or HPMC with either 2:1 or 3:1 (w/w) polymer/drug ratio.

The PHEAG conjugates exhibited rapid Gem release within less than 2 h, while the PHPAG conjugate showed sustained Gem release

profiles over a 10-h period.

2002 Elsevier Science B.V. All rights reserved.

Keywords:Chitosan microspheres; Macromolecular conjugates; Gemfibrozil; Sustained release; Poly[a,b-(N-2-hydroxyethyl-DL-aspartamide)]; Poly[a,b-

(N-3-hydroxypropyl-DL-aspartamide)]

1. Introduction oral administration and its short plasma half-life requires

relatively frequent dosing. Some gastrointestinal symptoms

Gemfibrozil is the lipid-regulating agent, which is and rash were observed as side effects of gemfibrozil

generically classified as a fibric acid derivative. It appears treatment. In order to improve its pharmacokinetics and

to be most useful in the treatment of lipoprotein disorders bioavailability, aliphatic and aromatic gemfibrozil esters

characterised by elevation of very-low-density (VLD) (Piccoli et al., 1994), benzamides (Sircar and Holmes,

lipoproteins and plasma triglycerides, since it lowers 1983), nicotinic acid (Hoefle, 1981) and 3-ethoxy deriva-

triglycerides and both total and VLD-cholesterol, while tives (Wang et al., 1996) were synthesised.increasing high-density-lipoprotein (HDL)-cholesterol The aim of this work was to investigate the feasibility of

levels (Todd and Ward, 1988). It is rapidly absorbed after controlled delivery systems in order to optimise the

therapeutic properties of gemfibrozil and to lower its side

effects. For this purpose two different types of deliveryAbbreviations: C, chitosan; C80, chitosan food grade 80; C90, chitosan

systems were evaluated: macromolecular drug conjugatesfood grade 90; CH, chitosan of high molecular weight; CL, chitosan oflow molecular weight; CM, chitosan of medium molecular weight; Gem, and microspheres.gemfibrozil; HPMC, (2-hydroxypropyl)methylcellulose; PHEA, poly[a,b- Macromolecular drug carrier systems in which drugs are(N-2-hydroxyethyl-DL-aspartamide)]; PHPA, poly[a,b-(N-3-hydroxy- covalently linked to polymers have been largely studiedpropyl-DL-aspartamide)]

and suggested as an effective way to prolong the pharma-*Corresponding author. Tel.: 1385-1-461-2608; fax: 1385-1-461-

cological activity, minimize unfavourable side effects and2691. E-mail address:[email protected](J. Filipovic-Grcic). toxicity, decrease the required dose and increase the

0928-0987/02/$ see front matter 2002 Elsevier Science B.V. All rights reserved.

PII : S0928-0987(02)00190-2
mailto:[email protected]:[email protected]


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208 A. Martinac et al. /European Journal of Pharmaceutical Sciences 17 (2002) 207216

solubility of the drug, as well as alter the body distribution (Japan), and gemfibrozil and L-aspartic acid were from

and ensure adequate drug delivery to target cells or tissues Aldrich (USA). All other chemicals used were of analytical

(Duncan and Spreafico, 1994). grade and purchased from Kemika (Croatia).

Microparticulate systems have great potential, being

able to convert poorly soluble, poorly absorbable and2.2. Preparation of polymers and polymerGemlabile biologically active substances into promising drugs.conjugatesPolymers used for conjugation of gemfibrozil in this

work were PHEA and PHPA. PHEA is an especiallyThe outline of the procedure for the preparation of theinteresting and promising drug carrier since it is water

polymerGem conjugates (Scheme 1) was the same as thesoluble, non-toxic, no antigenic, and biodegradable whenmethod described previously (Lovrek et al., 2000). Theexposed to a complex set of enzymes (Neri et al., 1973;synthesis of the PHEAG (5) and PHPAG (6) wasDrobnk et al., 1979). Many pharmacologically activeessentially esterification of the polyhydroxyl polymersagents, bearing carboxylic, amino or hydroxyl groups havePHEA (2) and PHPA (3) by the azole activated Gem,been covalently linked to PHEA (see, for example, Zorc etGem-Bt (4). In short, a solution of the PHEA (2) or PHPAal., 1993; Giammona et al., 1994, 1995, 1998).(3), Gem-Bt (4), and triethylamine (TEA) in DMF wasAlso, we attempted to develop drug-loaded micro-stirred at room temperature (3 days). The reaction mixturespheres based on chitosans and mixtures of chitosans andwas evaporated under reduced pressure. The sticky residueHPMC, PHEA or PHPA in order to investigate thewas then triturated with cyclohexane, acetone and ether ininfluence of the polymeric composition of the micro-

order to remove benzotriazole, amine and eventuallyspheres on drug content and on drug release.unbound 4, since these compounds were soluble in theChitosan is a natural, non-toxic, biodegradable, biocom-used solvents and conjugates 5 or 6 were not. The finalpatible and mucoadhesive polysaccharide. Its ability to beloose products PHEAG (5) or PHPAG (6) were filteredmade into solutions, films, fibres, beads as well as micro-off.spheres has lead to many pharmaceutical applications

The PHEA (2) was prepared by thermal polycondensa-(Kas, 1997; Kotze et al., 1999). Chitosan itself reducestion of L-aspartic acid in the presence of phosphoric acidblood cholesterol levels and thus it could be expected thatand subsequent aminolysis of polysuccinimide (PSI (1))entrapment of gemfibrozil into chitosan microspheres

with ethanolamine (Neri et al., 1973; Jakopovic et al.,could potentiate that effect (Furda, 2000).1996; Lovrek et al., 2000). PHEA weight-average molecu-Although many polymers are used in the pharmaceuticallar masses were 31 000, 56 000 and 61 000 for PHEA(1),formulations, the most widely utilised are the cellulosePHEA(3) and PHEA(2), respectively (Table 1), and theyderivatives. The use of hydrophilic cellulose ether such aswere determined by the viscosimetric method (Antoni etHPMC, has played an important role in the development ofal., 1974). In order to prolong the distance between mainsustained release drug delivery systems. HPMC can takechain and hydroxyl side groups, aminolysis of PSI (1) wasup and retain large amounts of water, which influences thecarried out with 3-hydroxypropylamine as well. In thisphysical and chemical properties of polymer and drugway a new hydroxy-functionalised polyaspartamide poly-release profile (Nokhodchi and Rubinstein, 2001).mer PHPA (3) was prepared. It could be considered that itsMicrospheres were produced by spray drying, which is a

M was very close to 61 000,M of PHEA(2), since bothrapid high-yield technique that is applicable at industrial w w

polymers were derived from the same PSI fraction ( 1) andscale. The ability of macromolecular conjugates andPSI was aminolysed under analogous conditions. Themicrospheres to modify the release rate of gemfibrozilaverage molecular masses of PSI and PHEA, determinedwere evaluated and compared.according to the MarkHouwink equation for PSI [h] 5

22 0.761.32 3 10 3M (Vlasak et al., 1979) and for PHEA

w23 0.87

2. Materials and methods [h] 5 2.32 3 10 3M (Neri et al., 1973), respectively,w

revealed that both polymers had practically the same2.1. Reagents and chemicals polymerisation degree.

The drug content in polymerGem conjugates was

Different molecular weight chitosans (CH, CM and CL) estimated by UV spectroscopy using the molar absorption

were purchased from Fluka (Switzerland): CH (M coefficient for Gem e 51866 l/mol/cm (in 96% EtOH,w 276

24600 000; deacetylation degree 83%), CM (M 400 000; c54.99310 M). The weight percentage of Gem in

w

deacetylation degree 83.5%), and CL (M 150 000; PHEAG(13) was in the range from 29 to 53%, and inw

deacetylation degree 87.4%). Chitosan food grade 90 PHPAG, 20% (Table 1). The drug content depended on

(C90) and 80 (C80) were obtained from Syntapharm the molar ratio of the reactant4 and monomer units of the

(Germany) and were used without further purification: C90 corresponding polymer 1, 2or 3.

(deacetylation degree 91%), C80 (deacetylation degree The proof that Gem was covalently bound in the

84%). HPMC was purchased from Shin-Etsu Chemicals synthesised polymerdrug conjugates could be found in


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A. Martinac et al. /European Journal of Pharmaceutical Sciences 17 (2002) 207216 209

Scheme 1. Procedure for the preparation of the polymerGem conjugates.

PolymerGem conjugates produced and their mainTable 1characteristics are listed in Table 1.Preparation and characteristics of PHEAG and PHPAG conjugates

Conjugate Polymer M of Gem contentw

2.3. Preparation of microspherespolymer (wt%)

PHEA(1)G PHEA(1) 31 000 53Microspheres were prepared by spray drying (Buchi 190PHEA(2)G PHEA(2) 61 000 34

mini spray drier, Switzerland). The drying conditions werePHEA(3)G PHEA(3) 56 000 29PHPAG PHPA 61 000 20 as follows: flow rate of 0.25 l /h, inlet air temperature of

120 8C and outlet air temperature of 75 8C.

Microspheres containing Gem with different polymericIR- and UV-spectra. The IR-spectra of5and 6showed an composition were prepared (Tables 24).

21ester carbonyl band at 1725 cm . All prepared conjugates

absorbed UV-light in the same absorption ranges as Gem, 2.3.1. Chitosan microspheres with Gemwhereas PHEA and PHPA themselves had no UV-absorp- Different types of chitosans or mixtures of chitosanstion at these wavelengths. (Table 2) at fixed concentration (1%, w/v) were solubil-

Table 2

Main characteristics of chitosan microspheres with Gem

Chitosan Viscosity of chitosan Theoretical polymer/ drug ratio (w/w)a

solution (mPa/ s)2:1 3:1

Mean diameter Drug content Entrapment Mean diameter Drug content Entrapmentb b

(mm) (wt%) efficiency (wt%) (mm) (wt%) efficiency (wt%)

CL 100 2.360.9 2263 67610 2.661.1 1863 73611

CM 200 2.660.9 2463 73610 2.661.2 1662 6369

CH1CL 2.260.8 2764 80612 2.461.1 2263 89612

CH 400 2.661.1 2562 7567 2.661.0 1562 6267

C90 70 2.360.9 2564 76612 2.360.9 1862 7166

C80 190 2.661.2 3362 9866 2.461.0 1564 61615

a1% solution in 1% acetic acid; according to Certificate of Analysis provided from the producer.

bEntrapment efficiency5theoretical drug content/actual drug content3100. Values are mean6S.D. (n 53).


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Table 3

Main characteristics of HPMC and C/HPMC microspheres with Gem

Polymers Theoretical polymer/drug ratio (w/w)

2:1 3:1

Mean diameter Drug content Entrapment Mean diameter Drug content Entrapmenta a

(mm) (wt%) efficiency (wt%) (mm) (wt%) efficiency (wt%)

HPMC 3.661.3 3264 96611 3.861.4 2561 10064C80 /HPMC 2.861.7 3364 99612 3.061.5 1964 74615

C90 /HPMC 2.961.5 3064 90611 3.161.6 2163 84612

CL/HPMC 3.361.5 3362 9866 3.261.3 2562 99.567

aEntrapment efficiency5theoretical drug content/actual drug content3100. Values are mean6S.D. (n 53).

ized in 0.5% acetic acid solution. In order to obtain the concentration of 1% (w/ v) was solubilized in 0.5% acetic

microspheres with different theoretical polymer/drug ratio acid solution. Polymers, PHEA(13) or PHPA, were

Gem was dissolved at different concentrations (2 and 3%, dissolved in purified water at different concentrations

w/ v) in ethanol and added to chitosan solution in a 1:6 (Table 4). Gem was dissolved in ethanol (2 or 3%, w/ v).

(v/ v) ratio. Table 2 lists all the batches of Gem-loaded Different mixtures were prepared by varying PHEA used,

chitosan microspheres produced. while the CL/G ratio was kept constant (2:1, w/w). For

the preparation of the CL/PHPA microspheres two mix-2.3.2. HPMC microspheres with Gem tures were prepared by varying CL/ G ratio (2:1 and 3:1,

HPMC was dissolved in a mixture of ethanol and water w/w) (Table 4). As for the PHEA/G and PHPA/G ratios

(2:3, v/ v). The polymer concentration was 1% (w/v). In the same ratios were used as present in the PHEAG and

order to obtain the microspheres with different theoretical PHEAG conjugates (Table 1). The mixtures were spray-

polymer /drug ratio Gem was dissolved at different con- dried under the conditions described in Section 2.3.

centrations (2 and 3%, w/v) in ethanol and added to Properties of the microspheres prepared are given in Table

HPMC solution in a 1:6 (v/ v) ratio. The mixtures were 4.

spray-dried under the conditions described above. The

characteristics of microspheres prepared are given in Table2.4. Encapsulation efficiency determination

3.

The drug content of the microspheres was determined2.3.3. C/HPMC microspheres with Gem

spectrophotometrically (l5276 nm; Ultrospec Plus, Phar-For the preparation of the C/ HPMC microspheresmacia LKB) after digestion of microspheres with 0.1 M

chitosan and the HPMC solutions were prepared as de-HCl. The microspheres prepared using HPMC were di-

scribed above at 1% (w/ v) concentration. The type ofgested with the mixture of 0.1 M HCl and ethanol.

chitosan varied between the preparations while the C/Preliminary studies showed that the presence of dissolved

HPMC ratio was kept constant (1:1, w/w). The ethanolicpolymer did not interfere with the Gem absorbance at 276

solution of Gem (2 or 3%, w/v) was mixed with solutionnm. Each determination was carried out in triplicate.

of polymers in 1:6 (v/v) ratio. Mixtures were spray-dried

under the conditions described in Section 2.3. The micro-

spheres obtained and their characteristics are given in 2.5. Particle size distribution

Table 3.

The microscopical image analysis technique for de-

2.3.4. CL/PHEA and CL/PHPA microspheres with Gem termination of particle size distribution was used. Micro-

The chitosan of low molecular weight (CL) at fixed sphere sizes and distribution were determined with

Table 4

Preparation and the main characteristics of CL/ PHEA and CL/ PHPA microspheres with Gem

Microsphere sample Polymer Concentration of CL/polymer CL/G Mean diameter Drug content Entrapmenta

polymer solution (%) ratio (w/w) (w/w) (mm) (wt%) efficiency (wt%)

CL/PHEA(1) PHEA(1) 4.36 2.3:1 2:1 2.461.0 2362 87611

CL/PHEA(2) PHEA(2) 9.54 1.04:1 2:1 2.360.9 1463 71611

CL/PHEA(3) PHEA(3) 12.20 1:1.22 2:1 2.561.1 1562 83611

CL/PHPA(2:1) PHPA 20.38 1:2 2:1 2.761.2 962 66615

CL/PHPA(3:1) PHPA 20.38 1:1.36 3:1 2.661.1 762 58612

aEntrapment efficiency5theoretical drug content/actual drug content3100. Values are mean6S.D. (n 53).


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Olympus BH-2 microscope, equipped with a computer- conjugates with the polyaspartamide polymers PHEA and

controlled image analysis system (OptomaxV, Cambridge). PHPA in phosphate buffer.

The initial rapid release was common to all PHEAG

2.6. Scanning electron microscopy (SEM) samples. Drug release progressed in extent of more than

80% within 2 h. The dissolution rate of Gem from the

The shape and surface characteristics of the microparti- PHEAG(1) conjugate with the lowest drug content (29%)

cles were observed by scanning electron microscopy. The was the fastest. However, it could be concluded that

microspheres were sputter-coated with Au/ Pd using a molecular mass of polymers used for conjugation and drugvacuum evaporator (Edwards) and examined using a content did not significantly affect Gem release profile

scanning electron microscope (Philips 500, Eindhoven) at from the PHEAG conjugates.

10 kV accelerating voltage. In the case of the PHPA G conjugate the release was

slow and linear. On the first 50 min, the release of Gem

2.7. In vitro release of Gem from conjugates and was about 20%, and it took 12 h to release 96% of Gem.

microspheres This reduction in release rate could be explained by longer

distance between the main chain and hydroxyl side groups

In vitro release profiles of Gem from conjugates and and differences in polymer /drug ratio and lipophilicity.

microspheres were examined in phosphate buffer, pH 7.4.

The drug loaded microspheres containing 10 mg of Gem 3.2. Characterisation of chitosan microspheres with gem

were put into rotating basket (50 rpm) and placed in 250 and drug release

ml of the dissolution medium, thermostated at 37 8C. Therelease from conjugates was assessed through dialysis Five samples of chitosan of different molecular weight

membranes (Spectr/ Por membranes Mwco 1214 000), were used for the preparation of microspheres. The main

which were placed in continuously-stirred 250 ml volumes characteristics of chitosans used and microspheres pre-

of phosphate buffer at 37 8C. At scheduled time intervals, pared are shown in Table 2. The amount of chitosan varied

agitation was stopped, the samples (2 ml) were withdrawn among the preparations while the amount of Gem was kept

and replaced with fresh medium. The samples were filtered constant.

and assayed spectrophotometrically at 276 nm. All experi- The preparation method produced well-formed micro-

ments were carried out in triplicate and average values spheres with good morphological characteristics for all

were plotted. batches prepared as shown in Fig. 2.

Particle size analyses revealed that the microspheres

were characterised by not so narrow size distributions with

3. Results and discussion mean diameter ranging between 2.360.9 and 2.661.2 mm.

Chitosan molecular weight, polymeric composition and

3.1. In vitro release of Gem from PHEA and PHPA polymer /drug ratio in the microspheres did not influence

conjugates particle size characteristics.

The encapsulation efficiencies were always very high,

Fig. 1 shows the dissolution profiles of Gem from its between 61 and 98% (Table 2). In the case of theoretical

Fig. 1. The release profiles of gemfibrozil from PHEAG and PHPAG

conjugates: (s) PHEA(1)G; (h) PHEA(2)G; (^) PHEA(3)G; (d) Fig. 2. SEM micrograph of CL microspheres with polymer/ drug ratio 3:1

PHPAG. (w/w).


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polymer /drug ratio (3:1, w/ w), the highest entrapment, density (having degree of deacetylation 87%) from other

89%, was obtained when the mixture of CL and CH types of chitosan used. Consequently, it differed in Gem

chitosans was used (1:1, w /w) for encapsulation, followed affinity.

by the CL chitosan with 73% encapsulation efficiency. The The release profiles of Gem from the chitosan micro-

microspheres prepared with theoretical polymer/drug ratio spheres with different polymeric composition in vitro are

2:1 (w/ w) showed higher entrapment of Gem than the shown in Fig. 3.

microspheres prepared with theoretical polymer/drug ratio All batches of Gem loaded microspheres showed the

3:1 for the same type of chitosan used. Exception was, most significant differences in drug release in the first 4 hagain, the microspheres prepared with the CL chitosan and completely released the drug in 12 h.

alone and the mixture of CL and CH chitosans, which A better control of drug release was obtained with the

entrapped less Gem as polymer /drug ratio decreased. microspheres made of the polymer/ drug ratio 3:1 (w/ w)

These results indicated that no direct correlation could be than with the microspheres made of the polymer/drug ratio

drawn between the viscosities of chitosans used for 2:1 (w/ w), except for microspheres made of the mixture of

microencapsulation and entrapment efficiency obtained, CH and CL chitosans (Fig. 3d). For CH microspheres,

and suggested that the CL probably differed in cationic Gem release profiles did not seem critically dependent on

Fig. 3. The release profiles of Gem from chitosan microspheres made of theoretical polymer/ drug ratio 2:1 (w/ w) (s) and 3:1 (w/w) (j), using different

types of chitosan: (a) CL, (b) CM, (c) CH, (d) CH1CL, (e) C80, and (f) C90.


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the polymer/drug ratio employed (Fig. 3c). When micro-

spheres of hydrophilic polymers are immersed in water,

they swell and form a gel diffusion layer that hinders the

outward transport of the drug within the matrix, hence

producing a controlled release effect (Lim et al., 2000). As

the amount of polymer increases, the thickness of the

hydrogel layer increases as well and the drug diffusion is

more retarded. That can explain the slower release oflipophilic Gem from microspheres with higher theoretical

polymer/drug ratio. In addition, chitosan can bind lipo-

philic compounds, which could also affect the Gem release

profile (Furda, 2000).

When comparing the Gem release profiles from micro-

spheres composed of different molecular weight chitosans

and with the same theoretical polymer/drug ratio it couldFig. 4. The release profiles of Gem from HPMC microspheres made ofbe seen that CL and CM microspheres with theoreticaltheoretical polymer/ drug ratio: (s) 2:1, (j) 3:1 (w/w).polymer/drug ratio 2:1 and 3:1 (w/w) are characterised by

insignificantly different Gem dissolution profiles (Fig. 3a

and b).

The fastest Gem release was achieved with the micro- Due to the polymer swelling described previously, thespheres prepared using CH for both polymer/ drug ratios release of Gem was slower as polymer /drug ratio in-

employed. The C80 and C90 microspheres showed drug creased. The microspheres with higher drug content were

dissolution profiles ranging between the release profiles of expected to be more porous than those with low drug

the CH and CL microspheres (Fig. 3e and f). content, which might facilitate the release of residual drug

Fig. 3 also shows an initial burst (ranging between 10 from microspheres (Wan et al., 1994). This could explain

and 50% in 10 min) of Gem release from all batches of the significant difference in the percentage of the Gem

microspheres. This is most likely due to the presence of released from the HPMC microspheres differing in poly-

Gem on the surface of the microsphere and a certain mer/ drug ratio.

amount of Gem that was not entrapped at all. Its reduction

in crystallinity, caused by spray drying, enhanced its

dissolution rate (Moyano et al., 1995). The initial rapid 3.4. Characterisation of C/HPMC microspheres with

release may have a functional use in providing an initial Gem and drug release

dose during the drug delivery, minimising any lag period.

The CL (3:1, w/w) microspheres present the lowest burst The main characteristics of microspheres prepared are

effect and the most regular Gem dissolution profile. given in Table 3. The highest entrapment efficiency

(99.5%) was obtained for the CL /HPMC microspheres

with the theoretical polymer/ drug ratio 3:1 (w/ w), al-

3.3. Characterisation of HPMC microspheres with Gem though all microspheres exhibited high Gem entrapment

and drug release efficiency (7499%). The C/ HPMC microspheres ob-

tained were spherical in shape and could easily be re-

The main characteristics of microspheres obtained are suspended in water, like chitosan and the HPMC micro-

given in Table 3. The entrapment efficiency was very high spheres. As expected, the average sizes of the C /HPMC

(96 and 100%), giving the microspheres with 32 and 25% microspheres were between the sizes of C and HPMC

Gem content. The HPMC microspheres were larger than microspheres, ranging between 2.861.7 and 3.361.5 mm.

the chitosan microspheres, characterised by size distribu- The C/ HPMC microsphere dissolution profiles aretions with mean diameters of 3.661.3 and 3.861.4mm for shown in Fig. 5. About 80% of Gem was released from all

the microspheres made of the polymer /drug ratio 2:1 and C/HPMC microspheres within 4 h. The CL/ HPMC micro-

3:1 (w/ w), respectively. Polymer/ drug ratio did not in- spheres (drug contents of 33 and 25%) show similar

fluence particle size characteristics and Gem entrapment dissolution profiles (Fig. 5a). The Gem release was slower

efficiency. from the C80/HPMC and C90 /HPMC microspheres made

The HPMC microsphere dissolution profiles are shown of the polymer/ drug ratio 3:1 (w/ w) than from the C80/

in Fig. 4. HPMC and C90/HPMC microspheres made of the poly-

Gem release from the HPMC microspheres with polymer/ drug ratio 2:1 (w/ w) (Fig. 5b,c). This could be

mer/ drug ratio 2:1 (w/ w) was completed within 5 h while explained by polymer swelling, but at the same time by

only 65% of Gem from the HPMC microspheres with the higher drug content of the microspheres made of polymer /

polymer /drug ratio 3:1 (w/ w) was released in that period. drug ratio 2:1 (w/ w) and consequently higher porosity.


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PHPA microspheres with Gem prepared by spray drying

are given in Table 4. While the CL/G ratio was kept

constant (2:1, w /w), different CL/ PHEA microspheres

were obtained by varying the type of PHEA used

(PHEA(13)) and combined with CL in that way that the

PHEA/ G ratios were the same as in corresponding PHEA

G conjugates. Two batches of the CL /PHPA microspheres

were prepared with the CL/G ratios of 2:1 and 3:1 (w/w).In both batches the PHPA/G ratio was the same as in the

corresponding PHPAG conjugate.

The entrapment efficiencies of the microspheres were

between 58 and 87% w/ w. The highest encapsulation

efficiency was obtained for the microspheres made of CL

and PHEA(1) giving the microspheres with highest Gem

content (23%, w /w). The CL/ PHEA(2) and CL/PHEA(3)

microspheres have the similar Gem content of 14 and 15%,

respectively, while the entrapment efficiency was lowest

for the CL/PHPA microspheres (66 and 58% for CL/G

ratio 2:1 and 3:1, respectively) resulting with low Gem

content (9 and 7% w/w) as shown in Table 4. This couldbe attributed to the relatively high ratio of PHPA when

compared to CL in the preparation.

Fig. 6 shows release profiles of Gem from the CL/

PHEA and CL/ PHPA microspheres in comparison with

release profiles of Gem from its PHEAG and PHPAG

conjugates. The drug content influenced the Gem release

rates from the CL/PHEA microspheres in such a way that

the release of Gem was faster as the drug content in-

creased. From the CL/PHPA microspheres made of the

CL/G ratio of 2:1 and 3:1 (w/w), the drug was released

after an initial burst (about 20% in 10 min), with an almost

constant rate for 8 h and the total release of about 60%.

As shown in Fig. 6ac release of Gem from the PHEA

G conjugates is faster than from the CL/PHEA micro-

spheres containing both, Gem and PHEA. In case of the

PHPAG conjugates and the CL/PHPA microspheres (Fig.

6d), the release profiles were similar with slightly slower

Gem release from the conjugates than from the micro-

spheres. This could be attributed to the difference in drug

content of the conjugates and the microspheres as well as

to the physico-chemical nature of these two delivery

systems. The Gem content of the PHEAG and PHPAG

conjugates was significantly (about two times) higher

compared with the CL/PHEA and CL/ PHPA microspheres

(Tables 1 and 4). In the PHEAG and PHPAG conju-gates, Gem is chemically bound to the polymer by an ester

Fig. 5. The release profiles of Gem from C/HPMC microspheres made of bond while it is physically entrapped into the matrix of thetheoretical polymer/drug ratio 2:1 (w/w) (s) and 3:1 (w/w) (j), and CL/PHEA and CL/PHPA microspheres. It appeared thatvarying the type of chitosan used: (a) CL/HPMC; (b) C80/HPMC; (c)

the ester bond in the PHEAG conjugates was hydrolyti-C90/HPMC.

cally very labile and subsequent Gem release faster than its

diffusion from the microspheres polymer matrix. PHPA

has longer distance (propyl group) between main polymer

3.5. Characterisation of CL/PHEA and CL/PHPA chain and hydroxyl side groups than PHEA, which seemed

microspheres with Gem and drug release to diminish hydrolysis of Gem from the corresponding

conjugate. Also, relatively high ratio of this hydrophilic

The main characteristics of the CL/ PHEA and CL/ polymer in the CL/ PHPA microspheres produced a more


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Fig. 6. The comparison of Gem release from CL/PHEA or CL/PHPA microspheres and PHEAG or PHPAG conjugates: (s) microspheres with: (a)

CL/PHEA(1), (b) CL/PHEA(2), (c) CL/PHEA(3), and (d) CL/PHPA polymers; (j) conjugates with: (a) PHEA(1), (b) PHEA(2), (c) PHEA(3), and (d)

PHPA.

hydrophilic matrix decreasing drugmatrix permeability References

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