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Impregnazione_tc_riassunto_sjf - english

Elena Bertola
Impregnation of a natural surgically implantable porous structure for the controlled
release of antibiotics

The spongy bone tissue is commonly used in medicine for dental implantations as support
for the osseous regeneration. One of the possible problems of such implantations after the
surgery is the growth of infections in the treated zone.
In this paper we study possibilities of transporting and depositing antibiotics all over the
surface in the inner part of the micro and nano-porous structure in order to reduce risks of
infection. In order to estimate the efficiency of the methods, different depositions have
been realized while measuring release sizes under physiological conditions.
Hydroxyapatite (HA) has been used as the osseous regeneration material, which allows
osteoconduction. Studied antibiotics are Tetracycline (TC) and Amoxicillin (AC), both of
them used with and without release-retarding potential. Release capacity at pH 7.4 has
been analyzed in order to make simulation conditions as realistic as possible.
The Hydroxyapatite matrix is prepared through supercritical extraction (CO2) and
deproteination with H2O2 at 700°C form an equine humerus bone.
At first interaction is studied between HA and antibiotic through the contact with TC
dissolved in N-methylpyrrolidone (NMP) on a milled HA column and the subsequent
desorption of the HA by NMP flushing, while measuring desorbed TC with UV
spectroscopy at 342 nm.
Impregnation is realized with the help of a supercritical fluid extraction plant (20 mL
extraction vessel) modified with HPLC pump for liquid solvent, in order to get a SAS
process (Solvent-Antisolvent-Supercritical). HA cubes are in contact with TC or AC
through a TC or AC supercritical solution in NMP 0.3 mg/mL (solvent solution) in which
supercritical CO2 is solved at 70 bar (Vss/Vsas = 60/40) in order to obtain a supercritical
solvent solution.
Increased pressure of the SCCO2 until 250 bar permit the precipitation of TC or AC thanks
to the antisolvent effect of the SCCO2. Finally NMP removal is realized by pure CO2 flow
at 250 bar.
The SAS process is first simulated at room conditions using liquid NMP as solvent and
liquid pentane as antisolvent.
TC deposition is verified through fluorescence at 366 nm and through desorption with NMP
and UV spectroscopy of the desorbed solution at 342 nm, whereas the AC one is tested
with SEM-EDX.
During release tests pieces of spongy bone impregnated with tetracycline are immersed in
physiologic buffer solution (NaCl/KCL/Na2HPO4/ pH 7.4).
Resulting solution is analyzed at intervals of approximately 10 min through a UV
spectrophotometer at 361 nm.
__________________________________________________________________Elena Bertola, Liceo Lugano 2 Results and discussion
Tests on HA ground permit to verify the good interaction between AI and TC: a slow flow
of 0.03mg/mL NMP-TC solution for each HA gram during 10 min produce a deposition of
0.002 mg/g (about 0.3µg/m2). Impregnation rate improves to 0.04 mg/g by using pentane
as antisolvent.
The SAS process with supercritical NMP-CO2 provides strong fluorescent HA cubes even
on internal surfaces, which indicates a TC deposition with elevated capillarity.
This fact is confirmed by SEM-EDX analysis on samples that were impregnated with
amoxicillin following the same process applied to the TC.
TC amount tests through NMP desorbtion of impregnated HA samples give loads values of
5 mg/g (50 µg/m2). In physiological solution, measured release is at most 1.0 mg / g,
corresponding to 20% of TC has been filed with the SAS process. This value is reached
quickly (after a few hours) and will hardly change even after several days.
Obtained impregnation level is more than 100 times higher than the one that can be achieved by simple contact of the porous tissue with liquid solutions. Impregnation rate may be further increased by higher TC concentration in NMP solution. Deposition occurs in the internal micro and nano-porous structure as well as in the external surface. Most of the antibiotic (80%) is held firmly by the impregnated structure after several days of contact with the physiological solution. Results should now be assessed in medical field. __________________________________________________________________Elena Bertola, Liceo Lugano 2


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