Published:Journal of Chromatographic Science,
ISSN 0021-9665 Volume
46, Number 9, October 2008, pp. 828-834
Application of an Experimental Design for the
Optimization and Validation of a New HPLC Method for the Determination
of Vancomycin in an Extemporaneous Ophthalmic Solution
Montse Enrique, Encarna García-Montoya, Montserrat
Miñarro, Anna Orriols, Joseph Ramon Ticó, Joseph Maria Suñé-Negre,
and Pilar Pérez-Lozano
Unit of Pharmaceutical Technology, Pharmacy and Pharmaceutical
Technology Department, School of Pharmacy, University of Barcelona, Avda
Joan XXIII s/n 08028. Barcelona, Spain
An experimental design has been used to develop
and optimize a new high-performance liquid chromatographic (HPLC)
method for the determination of Vancomycin in an extemporaneous
ophthalmic solution. After the preliminary studies and literature
review, the optimized method was carried out on a second generation
of a C18 reverse-phase column (Luna 150 ¥ 4.6 mm i.d., 5 µm
particle size) and using methanol as organic phase, a less toxic
solvent than acetonitrile, described in the extended literature.
The experimental design consisted of a Placket-Burman design
where six different variables were studied (flow rate, mL/min;
temperature, °C; pH mobile phase; % buffer solution; wavelength;
and injection volume) to obtain the best suitability parameters
(Capacity factor-K’, tailing factor, resolution, and theoretical
plates). After the optimization of the chromatographic conditions
and statistical treatment of the obtained results, the final
method uses a mixture of a buffer solution of water–phosphoric
acid (85%) (99.83:0.17, v/v) adjusted to pH 3.0 using triethylamine
and mixed with methanol (87:13, v/v). The separation is achieved
using a flow rate of 1.0 mL/min at 35°C. The UV detector
was operated at 280 nm. The validation study carried out, demonstrates
the viability of the method, obtaining a good selectivity, linearity,
precision, accuracy, and sensitivity.
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