Journal of Antimicrobial Chemotherapy (2003) 51, 585–591

DOI: 10.1093/jac/dkg105
Advance Access publication 28 January 2003

Antibacterial poly(D,L-lactic acid) coating of medical implants using a

biodegradable drug delivery technology

Hans Gollwitzer1,2*, Karim Ibrahim1, Henriette Meyer1, Wolfram Mittelmeier2, Raymonde Busch3
and Axel Stemberger1

1Institut
für Experimentelle Onkologie und Therapieforschung; 2Klinik und Poliklinik für Orthopädie und
Sportorthopädie; 3Institut für Medizinische Statistik und Epidemiologie, der Technischen Universität München,
Ismaninger Strasse 22, 81675 Munich, Germany

Received 4 September 2002; returned 16 October 2002; revised 26 November 2002; accepted 26 November 2002

Objectives: Biomaterial-associated bacterial infections present common and challenging com-

plications with medical implants. The purpose of this study was to determine the antibacterial
properties of a low molecular weight biodegradable poly(D,L-lactic acid) coating with integrated
antibiotics gentamicin and teicoplanin.
Methods: Coating of Kirschner-wires was carried out by a solvent casting technique under
aseptic conditions with and without incorporated antibiotics. Release kinetics of gentamicin and
teicoplanin were studied in phosphate-buffered saline. Initial bacterial adhesion of Staphylo-
coccus epidermidis on coated and bare implants was determined by radiolabelling and counts
of detached viable organisms.
Results: The incorporated antibiotics showed a continuous release over a period of at least 96 h
with an initial peak of release in the first 6 h. Attachment of non-viable microorganisms, detected
by radiolabelled bacteria, was increased significantly by the polymer coatings (P < 0.05). In
contrast, the number of viable bacteria was reduced by the pure polymer (P < 0.01) and further by
the polymer–antibiotic combinations (P < 0.05).
Conclusions: Poly(D,L-lactic acid) coating of implants could offer new perspectives in preventing
biomaterial-associated infections. Combinations with other drugs to formulate custom-tailored
implant surfaces are feasible.

Keywords: polylactide, PDLLA, drug release, Staphylococcus, biomaterial

Introduction stay, increased morbidity and mortality, and serious economic

sequelae being common consequences.5
Since the first applications of biomaterials in medicine, It is possible that the risk of infection can be reduced by an
infections and deficient tissue-integration represent the most antiseptic surface coating for medical implants. The purpose
important complications, which still limit the unrestricted use of this study was therefore to determine the anti-infective
of biomaterials in humans. Implant-associated infections properties of a new biodegradable coating for medical
account for nearly 50% of the estimated 2 million nosocomial implants with regard to Staphylococcus epidermidis, one of the
infections in the United States per year,1 and infection rates up most important pathogens in biomaterial-associated infec-
to 100% are reported for certain implants, like external fix- tions.6,7 This coating is based on a polymer of low molecular
ation pins.2–4 Treatment of these infections is associated with weight poly(D,L-lactic acid) (PDLLA) and can be combined
high complication rates and places an enormous burden on with drugs like antibiotics or growth factors to establish a
both the patient and healthcare providers; prolonged hospital locally acting drug-delivery system.
..................................................................................................................................................................................................................................................................

*Corresponding author. Tel/Fax: +49-89-4140-7242; E-mail: h.gollwitzer@lrz.tu-muenchen.de

...................................................................................................................................................................................................................................................................

585
© 2003 The British Society for Antimicrobial Chemotherapy
 H. Gollwitzer et al.
Materials and methods
The polymer coating
The commercially available Resomer R203 is a polymer of
PDLLA with a molecular weight of 29 000 Da and was pur-
chased from Boehringer Ingelheim (Ingelheim, Germany).
The polymer is a racemic mixture of the D- and L-enantiomers
of lactic acid and serves as a biodegradable coating for med-
ical implants.
Implants were coated with PDLLA by a solvent casting
technique. In brief, the drug-carrier was dissolved in ethyl-
acetate (Sigma–Aldrich AG, Deisenhofen, Germany) at a
concentration of 133.3 mg/mL. The coating solution was main-
tained on dry ice to prevent evaporation of the organic solvent
and a subsequent increase in the polymer concentration.
To create a local drug delivery system, 5% (w/w) of the
antibiotics gentamicin sulphate (COM Pharma, Hamburg,
Germany) and/or teicoplanin (Aventis, Frankfurt, Germany)
were added to the polymer solution. The implants were coated
by two dip-coating procedures to achieve a dense and regular
polymer coating. All coating steps were carried out under
aseptic conditions in a laminar air-flow.
Medical implants
Commercially available Kirschner-wires (K-wires) of stain-
less steel (ISO 5832-1; Synthes GmbH & Co. KG, Umkirch,
Germany) and titanium alloy (TiAl6V4, ISO 5832-11 and ISO
5832-3; Synthes GmbH & Co. KG) were studied. The wires
were cut into lengths, cleaned and sterilized by autoclaving.
The wires used for microbiological and antibiotic release
studies and their abbreviations are shown in Table 1.
Bacterial strains
A clinical isolate of a biofilm-forming strain of S. epidermidis
(strain SE 183) was used for the in vitro studies. The test strain
was susceptible to both gentamicin and teicoplanin (genta-
micin MIC 2 mg/L, teicoplanin MIC 0.5 mg/L). Biofilm for-
mation was demonstrated by qualitative assessment with the
Table 1. Sample groups and surface coating of stainless steel (S) and titanium alloy (T) for the
in vitro tests
Surface coating
Bare K-wires without coating (control group)
K-wires coated with PDLLA
K-wires coated with PDLLA and gentamicin (5.0% w/w)
K-wires coated with PDLLA and teicoplanin (5.0% w/w)
K-wires coated with PDLLA and gentamicin (1.7% w/w)
and teicoplanin (3.3% w/w)
586
tube assay previously described by Christensen et al.8 Stock
cultures of the isolate were lyophilized or stored at –70°C.
Preparation of bacteria
S. epidermidis (SE 183) was cultured to late logarithmic
growth phase on blood agar plates at 37°C for 18 h before test-
ing. Bacterial cells were then resuspended in normal saline
and adjusted to 5 × 107 cfu/mL by visual comparison with a
0.5 McFarland standard. This suspension was diluted with
normal saline to an inoculum of 2.5 × 105 cfu/mL.
Antibiotic release
Drug release from antibiotic- and polymer-coated K-wires
(n = 4; size 1.5, length 150 mm; stainless steel) was studied in
10 mL of phosphate-buffered saline (PBS) at 37°C (pH 7.4).
Samples were coated with the PDLLA polymer (133.3 mg/mL
ethylacetate) including gentamicin or teicoplanin (5% w/w).
PDLLA-coated samples without antibiotics served as a con-
trol. To assess antibiotic stability during the test period, PBS
of the control groups was supplemented with the gentamicin
and teicoplanin. Aliquots of 500 µL were taken at the time
points 10 min, 1, 6, 24 and 96 h from the PBS and assayed for
gentamicin and teicoplanin. Antibiotic concentrations were
determined by the fluorescence polarization immunoassays
TDx/TDxFLx Gentamicin assay (Abbott Laboratories, Abbott
Park, IL, USA) and Innofluor Teicoplanin assay system
(Opus Diagnostics, Fort Lee, NJ, USA).
Total bacterial adhesion
The total amount of live and dead adhering bacteria was
studied in a radiolabelling experiment, as described by Chris-
tensen et al.9 S. epidermidis was grown in Mueller–Hinton
broth (Oxoid GmbH, Wesel, Germany) substituted with
[3H]thymidine. After 24 h of incubation on a rotary shaker
(100 rpm), inoculum suspensions were prepared as described
earlier. Coated and bare K-wires of stainless steel and
titanium alloy (n = 9; size 1.8, length 12 mm) were incubated
with the radiolabelled bacterial cell suspension for 2 h at 37°C
under static conditions. Individual wires were removed with
sterile forceps, washed three times with sterile normal saline
Stainless steel Titanium alloy
S1 T1
S2 T2
S3 T3
S4 T4
S5 T5
 Antibacterial poly(D,L-lactic acid) coating
for 15 s and air-dried. After transfer to scintillation vials con-
taining scintillation fluid, counts per minute were measured in
a β-counter (1219 Rackbeta; LKB Wallac, Bromma, Sweden).
Adhesion of viable bacteria
Adhesion of viable bacteria was evaluated in a bacterial
adhesion assay. Coated and bare K-wires of stainless steel and
titanium alloy (n = 10; size 1.8, length 12 mm) were immersed
in 2 mL of the bacterial suspension (2.5 × 105 cfu/mL normal
saline) and incubated for 2 h at 37°C under static conditions.
After washing in normal saline, the K-wires were placed in
vials containing 2 mL of trypsin solution (1% w/w) and
sonicated (Sonorex RK255H, 50 kHz; Bandelin Electronic,
Berlin, Germany) for 15 min to remove the adhering micro-
organisms. Serial dilutions of each trypsin solution were
plated on blood agar plates for quantification of viable organ-
isms. Blood agar plates were incubated for 48 h at 37°C, and
the cfu were counted visually.
Complete detachment of the adhering microorganisms by
sonication was verified through scanning electron micro-
scopy (SEM).
Morphological analysis—SEM
Representative K-wire specimens from the in vitro studies
were prepared for SEM as follows. Specimens were fixed
in an aqueous solution of 1% (v/v) glutaraldehyde (Serva,
Heidelberg, Germany) in a buffer of sodium phosphate
(dibasic) monohydrate (Merck KGaA, Darmstadt, Germany)
and sodium hydroxide (Merck KGaA) for a minimum of 16 h.
The buffer contained 18.8 g/L sodium phosphate (dibasic)
monohydrate and 4.3 g/L sodium hydroxide. Dehydration
was carried out in an ascending ethanol series (50–70–80–90–
100%) and samples were dried in a critical point dryer (CPD
030; BAL-TEC AG, Balzers, Liechtenstein) with carbon di-
oxide. Once coated with gold palladium (Polaron Autocoating
Unit SEM 5200; Quorum Technologies, Newhaven, UK)
each specimen was examined through a low vacuum SEM
(JEOL 5900; JEOL Germany GmbH, Eching, Germany).
Figure 1. Elution profiles of gentamicin (left-hand panel) and teicoplanin (right-hand panel) into PBS from individual stainless steel K-wires with
PDLLA coatings containing either gentamicin or teicoplanin. In each panel the different lines show the release kinetics of the four samples tested.
587
Calculations and statistical methods
Data from bacterial adherence studies were compared for
statistical significance using non-parametric methods and the
method of closed testing procedure,10 with P < 0.05 consid-
ered significant (Kruskal–Wallis and Mann–Whitney tests).
Results
Morphological analysis
After the dip-coating procedure, a stable and regular PDLLA
coating could be observed through SEM. Complete detach-
ment of adhering bacteria was observed after sonication.
Antibiotic release
The concentrations of the released antibiotics gentamicin and
teicoplanin in the elution buffer are shown in Figure 1. A
continuous drug release could be demonstrated for at least
96 h for both antibiotics. After an initial peak of release in the
first few hours, a slow and continuous drug release could be
observed for the rest of the testing interval. Gentamicin
showed a pronounced initial peak of release in the first hour.
Teicoplanin release, in contrast, was delayed compared with
gentamicin, the initial peak of release lasted ∼6 h and the
remaining teicoplanin was released to the PBS with kinetics
similar to that of gentamicin. Both gentamicin and teicoplanin
showed a marked variation of initial release between indi-
vidual wires (between 3.87 and 10.58 mg/L for gentamicin
and between 2.71 and 15.11 mg/L for teicoplanin after 6 h).
Antibiotic concentrations of the control groups remained
constant throughout the entire study interval and gentamicin
and teicoplanin were stable in PBS in control experiments for
at least 96 h.
Total bacterial adhesion
The total biomass of bacteria adhering to the wires was
measured using the radioactively labelled bacteria method. In
 H. Gollwitzer et al.

Figure 2. Total biomass (median and inter-quartile ranges) of S. epidermidis adhering to either stainless steel and PDLLA-coated K-wires (n = 9;
left-hand panel) or titanium alloy and PDLLA-coated K-wires (n = 9; right-hand panel), as assessed by the binding of radiolabelled bacteria.
*, extremes (cases with values more than 3 box lengths from the upper or lower edge of the box); H, outliers (cases with values between 1.5 and 3 box
lengths from the upper or lower edge of the box).

Figure 3. Median viable counts of S. epidermidis adhering to either stainless steel and PDLLA-coated K-wires (n = 10; groups S1–S5 according to
Table 1; left-hand panel) or titanium alloy and PDLLA-coated K-wires (n = 10; groups T1–T5 according to Table 1; right-hand panel). *, extremes
(cases with values more than 3 box lengths from the upper or lower edge of the box); H, outliers (cases with values between 1.5 and 3 box lengths from
the upper or lower edge of the box).

this method, the number of counts per minute is directly pro-
portional to the number of implant-adhering microorganisms,
and independent of their viability (data not shown). The
median counts and inter-quartile ranges (along with outliers)
obtained for the different wires are shown in Figure 2. The
titanium alloy used showed a slightly lower biomass of S. epi-
dermidis adhering to the wire than stainless steel alloys,
although the results are not significantly different (P > 0.05).
However, for both types of wire a marked increase in bacterial
adhesion can be observed for all PDLLA-coated samples
compared with the bare specimens. For all of the titanium
alloy wires this increase was significant at the P < 0.05 level,
whereas for the stainless steel wires the increases were
statistically significant (P < 0.05) for all except the PDLLA/
gentamicin-coated (S3) wire.
With regard to the effect of antibiotic integration, genta-
micin showed a tendency (P = 0.05–0.1) to reduce bacterial
biomass after 2 h compared with pure PDLLA. Integration of
teicoplanin on the other hand significantly increased the
biomass of S. epidermidis compared with implants coated
with pure PDLLA and the PDLLA/gentamicin combination
(P < 0.05).
Adhesion of viable bacteria
The number of viable bacteria adhering to the stainless steel
and titanium alloy implants was evaluated in a quantitative
adhesion assay. Results are expressed in colony forming units
per agar dilution plate (Figure 3), this measurement being a
direct indicator of the number of viable counts adhering to the
implants.

588
 Antibacterial poly(D,L-lactic acid) coating
The PDLLA-coated implants significantly reduced adhe-
sion of viable staphylococci compared with bare K-wires
made from either titanium or stainless steel alloy (P < 0.01).
Combination of PDLLA with either gentamicin or teico-
planin or both antibiotics on the implant together reduced
viable counts to almost undetectable levels (P < 0.05).
Although there were no significant differences between
gentamicin and teicoplanin (P > 0.05), there was a slight
tendency towards less bacterial growth with gentamicin-
containing coatings, and also between the titanium alloy and
stainless steel (Figure 3).
Discussion
Bacterial adhesion to biomaterials and the capability of many
microorganisms to form biofilms on foreign bodies are well-
established steps in the pathogenesis of implant-associated
infections.11–13 In a biofilm, bacteria are protected from the
host immune defence11,14 and exhibit a marked but reversible
increase in antibiotic resistance.11,15,16 Even high local drug
concentrations beyond the MBC for planktonic microorganisms
do not completely eradicate bacteria located in biofilms.15,17
As a result, prevention of bacterial colonization and bio-
film formation is an important consideration and may be sup-
ported by the use of antiseptic surface coatings. In the present
work, a new antibacterial surface coating using a biodegrad-
able drug-delivery system was studied.
The chosen PDLLA coating enables the possibility of
covering medical implants with a biodegradable and biocom-
patible surface coating. This polymer is applied to implants by
a solvent casting technique that allows coating of alloys and
plastics with polished, irregular or porous surface materials.
Breakdown of the polylactic acids is based on hydrolytic
splitting of the polymer backbone over several months to
oligomers, and release of lactic acid, which is metabolized in
the citric acid cycle of the organism.18–20
Incorporation of antibacterials into this coating to give a
local drug-delivery system ensures high concentrations around
the implant for long periods of time and the risks and side-
effects for the host organism are minimized compared with
systemic drug application.21
In this study, elution fluids were not changed, and an elu-
tion model that is broadly applied in the literature, especially
with antibiotic loaded bone cement, was used.22,23 Powdered
teicoplanin and gentamicin do not dissolve very well in the
volatile organic solvents used, and the final coating on the
implants consisted of antibiotic particles in the polymer. This
leads to an initial peak of release and high variability between
experiments, caused by fine drug particles located at, and
washed out of the polymer coating surface, a process that is
not well-defined. On the other hand, after the initial peak,
subsequent release is very predictable and comparable for all
589
samples, being primarily the result of diffusion of incorpor-
ated antibiotics from deeper levels of the coating.
In this study, we observed significant differences between
the results of adhesion studies depending on whether total or
viable bacteria were assessed. The coating increased the total
amount of attached microorganisms but at the same time
significantly reduced the number of viable bacteria even with-
out antibiotics, suggesting that the bare PDLLA coating has
bactericidal action against adhering S. epidermidis SE 183.
Since most of the bacterial adhesion studies only employ one
type of test, either counting the total number of attached
microorganisms24–26 or counting only viable bacteria,27–29
results from these studies may be misleading. Since genta-
micin and teicoplanin are integrated into the polymer as fine
particles, drugs located at the surface can be washed out to
leave small irregularities in the polymer. As one of the most
important features in primary bacterial adherence is the
topography texture,30,31 this may promote the bacterial adhe-
sion seen in this study. Apart from morphological features
of the polymer, physiochemical characteristics like surface
charge influence bacterial adhesion as well. In the present
study, teicoplanin increased the total number of adhering micro-
organisms compared with gentamicin. Gallardo-Moreno et al.32
and Wilcox et al.33,34 have previously described promotion of
bacterial adhesion to implant surfaces by subinhibitory concen-
trations of glycopeptide antibiotics.32–34 Our results would sug-
gest that this effect may also occur at inhibitory concentrations
and highlight the importance of undertaking total as well as
viable bacterial counts.
PDLLA has excellent features with respect to implant coat-
ing, with high mechanical stability,35 good osteoinductive
potential36 and excellent biocompatibility in vivo.36 The
material also shows good anti-thrombogenic characteris-
tics.37 In this study we have been able to demonstrate that the
antibacterials gentamicin and teicoplanin can be incorporated
into the polymer to give local drug-delivery systems that
reduce bacterial adhesion in vitro. We conclude that a com-
bination of the antibiotic–polymer coating with other drugs to
create a multifunctional and custom-tailored surface is there-
fore possible and could offer new opportunities in the use of
established biomaterials.
Acknowledgements
We gratefully acknowledge the contributions of I. Kappstein
(Associate Professor of the Department of Hospital Epidemi-
ology, University Hospital, Technical University Munich,
Germany) for inspiring discussions, constructive micro-
biological advice and for reading the manuscript. We would
also like to thank V. Vatou and M. Chihaja for their assistance
in microbiological testing. Furthermore, we acknowledge the
contributions of D. Hall of the 1st Medical Clinic of the Tech-
nical University Munich for assistance in preparing the manu-
 H. Gollwitzer et al.
script. This study was supported by a grant from the ‘Bavarian
Research Cooperation for Biomaterials (FORBIOMAT)’.
The authors did not receive any payments or benefits from any
other research fund, foundation, educational institution or
other non-profit organization. No benefits in any form have
been received or will be received from a commercial party
related directly or indirectly to the subject of this article.
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