Characterization of the interaction of Trimethoprime with Hydroxypropyl-B-cyclodextrin in solution

GARNERO,C.; ZOPPI, A; LONGHI, M. R.

Porto Alegre, Brasil

Congreso; XX Congreso Panamericano de Farmacia y XIV Congreso de la Federacion Farmaceutica Sudamericana; 2010

1. Introduction Trimethoprime is a bacteriostatic antibiotic, characterized by a very low aqueous solubility, sometimes prescribed for other uses in combination with some drugs, mainly used in the prophylaxis and treatment of urinary tract and certain types of pneumonia. Cyclodextrins (CDs) are macrocyclic oligosaccharides with six, seven or eight D-glucose units called alfa-cyclodextrin, beta-cyclodextrin and gamma-cyclodextrin, respectively. The cavities of CDs are relatively hydrophobic, while the external faces are hydrophilic. The most important structural feature of CDs is their capacity of forming stable complexes. The complex formation provides a way to increase the solubility, stability and bioavailability of drugs. 2. Objetive The present study is focused on the investigation of the complexation between trimethoprime and hydroxypropyl-B-cyclodextrin (HP-B-CD) in solution. 3. Materials and methods The characterization has been performed using solubility analysis, nuclear magnetic resonance techniques (1H-NMR, 1D-ROESY and DOSY), and conductivity measurements. All the experiments were performed with analytical grade chemicals and solvents. The solubility measurements were performed according to the method of Higuchi and Connors. The critical concentration for the aggregate formation was determined by measuring the specific conductivity change as a function of concentration, using a Malvern Zetasizer 3000. All Nuclear Magnetic Resonance (NMR) studies were performed on a Bruker Avance II High Resolution Spectrometer, equipped with a Broad Band Inverse probe (BBI) and a Variable Temperature Unit (VTU). NMR spectra were taken in deuterium oxide (D2O, deuterium content 99.9 %) from Aldrich USA. 4. Results Solubility study revealed that the trimethoprime solubility was significantly increased upon complexation with HP-B-CD. Phase solubility diagram and diffusion ordered spectroscopy (DOSY) techniques were used to determine the apparent stability constant (K1:1) at 25 C. The K1:1  value of 60.4 +/- 0.2 M-1 was estimated from the linear section of the solubility diagram, while a K1:1 of 341.5 M-1 was determined employing DOSY experiments. The differences in the K1:1 values obtained may be caused by the differences in the experimental conditions. In addition, conductivity experiments showed the presence of aggregates that permit to explain the AN type profile for the solubility isotherm. NMR experiments provided evidence of trimethoprime:HP-B-CD molecular interaction in solution. The observed spatial proximities between protons of HP-B-CD and trimethoprime molecules, the variations undergone by protons in the 1H-RMN spectra and the evidence of binding between differently sized species determinated by DOSY experiments, strongly suggest the inclusion of trimethoprime into the HP-B-CD cavity. 5. Conclusion Physicochemical characterization of the binary system trimethoprime:HP-B-CD demonstrated the formation of an inclusion complex. The results reported here revealed that the solubility of trimethoprime was enhanced in presence of HP-B-CD. At low HP-B-CD concentrations the formation of an inclusion complex produced an increase in the solubility; whereas at concentrations higher than the critical one were formed aggregates capable of solubilise the trimethoprime through the formation of noninclusion complexes. The NMR results evidenced that the interaction was an inclusion phenomenon since the modifications obtained for trimethoprime and HP-B-CD signals.