The Elution of Vancomycin and Tobramycin from Calcium Sulfate/ Calcium Phosphate Composite Pellets
+1MacDougall J; 1Blum B; 1Burgess A; 1Morris L
+1Wright Medical Technology, Memphis, TN
Senior author [email protected]
Calcium sulfate and calcium phosphate have long been recognized as
safe and effective bone graft materials, and several publications have
suggested the potential for local antibiotic delivery due to their
resorbable nature [1, 2, 3, 4]. Calcium sulfate exhibits in vivo resorption
rates around six to eight weeks while some calcium phosphate materials
have been known to resorb over many months to years. In an effort to
better match the rate at which bone growth occurs, a calcium sulfate and
calcium phosphate composite bone graft substitute that cures in situ and
resorbs in vivo over three to six months has been developed (PRODENSE® Injectable Graft, Wright Medical). The composite material
has demonstrated the ability to regenerate stronger and denser bone at a
rate faster than autograft or calcium sulfate in the canine proximal
humerus model[5]. Due to these resorption and bone healing qualities,
interest in its use to locally deliver antibiotics has developed. Compared
to systemic administration of antibiotics, local delivery allows for high
levels of antibiotic at the affected site while serum levels remain low. In
an effort to better understand the release of antibiotics from the calcium
sulfate/calcium phosphate (CS/CP) composite graft, the elution and
dissolution profiles of tobramycin and vancomycin were examined.
(paired t-test, p ≤ 0.05). The vancomycin loaded pellets exhibited a
statistically significant difference in mass remaining on all four days in
comparison to the plain pellets. When comparing the vancomycin
loaded and tobramycin loaded CS/CP pellets, there was no statistically
significant difference between the groups after Day 1 (paired t-test, p ≥
0.05).
10000
Average Antibiotic Concentration
(ug/ml/g)
Introduction
1000
100
CS/CP Pellets w ith
4% Vancomycin
10
CS/CP Pellets w ith
4% Tobramycin
Sulfate
1
0
1
2
3
5
7
10
Days
Materials and Methods
Elution Testing
To examine the elution characteristics of the tobramycin and
vancomycin from the CS/CP, eight pellets were added to a bottle
containing 20ml of phosphate buffered saline (PBS) and kept in a 37°C
water bath (n=3 bottles for each antibiotic). Aliquots of eluant, 2mL,
were drawn at days 1, 2, 3, 5, 7, and 10 and measured for antibiotic
concentration using a fluorescence polarization immunoassay (FPIA)
technique (TDxFLx® system, Abbot Laboratories, Abbot Park, IL). The
entire 20mL of PBS was replaced at each time interval.
Accelerated Dissolution Testing
Antibiotic loaded CS/CP pellets along with plain CS/CP pellets were
individually placed in fritted glass thimbles. The thimbles were placed
in bottles containing 200mL of deionized water and kept in a waterbath
at 37°C (n=5 for each group). Every 24 hours, the pellets were dried and
weighed in the thimbles to determine the percent mass remaining. The
deionized water was changed after each time period.
Results
The elution profiles of vancomycin and tobramycin from the CS/CP
composite pellets are illustrated in Figure 1. The tobramycin and
vancomycin curves both exhibited an initial peak in concentration on
Day 1 (Vancomycin 1233µg/ml/g, Tobramycin 1243µg/ml/g) followed
by a subsequent decline over the remaining time period. The pellets
containing vancomycin exhibited an extended elution profile when
compared to the tobramycin profile. While vancomycin concentrations
were present above 10µg/ml/g at day 10, the tobramycin levels were less
than 1µg/ml/g. However, both were present above the respective
minimum inhibitory concentrations for Staphylococcus aureus out to
day 10 (ATCC 29213: Vancomycin 0.5-2.0µg/ml; Tobramycin 0.121.0µg/ml). As can be seen in Figure 2, the accelerated dissolution
profile of the antibiotic loaded pellets during this initial 4 day time
period is slightly decreased in comparison to plain pellets with all groups
still retaining approximately 55-65% of their original mass at Day 4.
There was a statistically significant difference in mass remaining
between plain pellets and tobramycin loaded pellets on Days 1 and 2
Figure 1. Antibiotic elution from CS/CP Pellets
Mass Remaining (%)
The CS/CP composite material containing either no antibiotics, 4%
Tobramycin Sulfate (Alpharma API, Copenhagan, Denmark), or 4%
Vancomycin (Hospira Inc. Lake Forest, IL) by weight was cast on a
silicone mat to form cylindrical pellets approximately 4.8mm in
diameter, 3.2mm in height, and 115mg in weight. The pellets were
allowed to air dry overnight before being oven dried at 40°C for at least
5 hours.
100
95
90
85
CS/CP Pellets
80
75
70
65
60
55
50
CS/CP Pellets w ith
4% Vancomycin
CS/CP Pellets w ith
4% Tobramycin
Sulfate
1
2
3
4
Days in vitro
Figure 2. Accelerated dissolution of CS/CP Pellets both with and
without antibiotics
Discussion
The elution curves of tobramycin and vancomycin from the CS/CP
composite graft demonstrate that the majority of the antibiotic was
released within the first few days, with a large initial peak in
concentration during the first 24 hours and a decline in concentration
over the remaining time-points. This decline was more prominent in the
group containing tobramycin. The presence of either antibiotic may
have slightly increased the rate at which the pellets dissolve compared to
pellets without antibiotics; however, the differences may be more
attributable to variability in the test. Even so, the similarity of the
dissolution rates between pellets with tobramycin and vancomycin
suggests the differences in antibiotic elution between the two groups
may be related to the antibiotic incorporation into the material during the
dual setting reactions of the calcium sulfate and calcium phosphate.
Based on the preliminary in vitro data, tobramycin and vancomycin may
be successfully incorporated into the CS/CP composite material.
References
[1] M.E. Shirtliff, CORR, No. 401, 2002
[2] J Lazarettos, CORR, No. 423, 2004
[3] T.M. Turner, CORR, No 437, 2005
[4] J.C. Wenke, JBJS, Vol. 88-B(8), 2006
[5] R.M. Urban, CORR, No. 459, 2007
Poster No. 551 • 55th Annual Meeting of the Orthopaedic Research Society