Evaluation of Gentamicin Release of PMMA Cements Using Different Methods: HPLC, Elution and Inhibition Zone Testing

Abstract

(1) Background: There is an ongoing discussion on the elution efficacy of antibiotic-impregnated cements. Our experiments were intended to clarify if there are differences in the antibiotic elution of HPLC compared with inhibition zone testing using eluates or PMMA discs. (2) Materials and Methods: Two cement brands with different concentrations of the active ingredient were tested in antimicrobial Kirby-Bauer (disc diffusion) assays. Cement platelets were directly applied on the agar plates and their zone of inhibition was measured. In parallel, the platelets were incubated in phosphate buffered saline (PBS) and at distinct points of time transferred into new buffer. At these time points, 50 µL of the bone cement eluates was used for zone of inhibition testing. Standard gentamicin sulfate solutions served as a control in the same test setup. To verify the microbiological investigations, the antibiotic content of the eluates was also measured via high-performance liquid chromatography (HPLC). (3) Results: The experiments with cement eluates showed better differentiable results than the direct application of the cement discs. The results were also comparable to investigations with HPLC and gentamicin sulfate standard solutions. (4) Conclusions: The results of elution rates are influenced by the test system and the period of observation chosen. The microbial test systems reflect the results of HPLC to the same degree and give evidence of the efficacy of the antibiotics. The HPLC tests on eluates were more suitable in representing differences in release characteristics.

Keywords: HPLC; PMMA cement; antibiotic elution; area zone; eluates and discs; gentamicin.

Figure 1

Testing method agar diffusion: Cement discs were applied directly on agar plates streaked with S. aureus agar. Every 24 h, the discs were transferred onto new agar plates. The diameter of inhibition area is in millimeters.

Figure 2

Testing method agar diffusion: Cement discs were applied directly on agar plates streaked with S. epidermidis agar. Every 24 h, the discs were transferred onto new agar plates. The diameter of inhibition area is in millimeters.

Figure 3

Linear regression line. The black line is the regression line. Diameter of inhibition zone in mm for Palacos R+G (green) and CMW 1G (red) in 24 h.

Figure 4

Testing eluates: Diameter of Inhibition Areas of Palacos R+G (0.5 g) and CMW 1G (1.0 g) cements over 14 days, y-axis shows the diameters of inhibition areas [mm], x-axis represents the time scale [hours]. Values of the measured diameters differ significantly at the desired time points.

Figure 5

Testing elution with eluates: Accumulative elution of GS out of cement discs.