Application of oxidized multi-walled carbon nanotubes and zeolite nanoparticles for simultaneous preconcentration of codeine and tramadol in saliva prior to HPLC determination

In this work, a dispersive micro-solid phase extraction technique along with high-performance liquid chromatography-UV detection was developed for simultaneous preconcentraion and determination of trace levels of codeine and tramadol in human saliva. This method is based on the adsorption of codeine and tramadol on a mixture of oxidized multi-walled carbon nanotubes and zeolite Y nanoparticles with 1:1 ratio as an efficient nanosorbent. Various analytical parameters influencing the adsorption step including the amount of adsorbent, the pH of the sample solution, the temperature, the stirring rate, the contact time of the sample solution, and the adsorption capacity were investigated. Based on the results, 10 mg adsorbent, sample solutions with pH = 7.6, temperature of 25 degrees C, stirring rate 750 rpm and contact time 15 min, in the adsorption step shows the best results for both drugs. Then the effective parameters on the analyte desorption stage such as the type of desorption solution, pH of the desorption solution, desorption time and desorption volume were investigated. Studies have shown that water/methanol (50:50 v/v) as desorption solution, pH = 2.0, desorption time of 5 min and desorption volume of 2 ml gives the best results.Chromatographic separation was performed on a RP-Shim-pack CLC-ODS-C18 column (250 mm x 4.6 mm, 5 mu m) with isocratic mode. The mobile phase contained of aceto-nitrile:phosphate buffer (18:82, v/v) at pH = 4.5 and the flow rate was 1 ml.min- 1. The wavelength of UV detector was adjusted at 210 nm and 198 nm for codeine and tramadol, respectively.Under optimum conditions, the extraction efficiencies of 98.5% and 99.2% were achieved for codeine and tramadol respectively. Enrichment factor of 13, detection limit of 0.3 mu g L-1, relative standard deviation of 4.07 for codeine; and an enrichment factor of 15, a detection limit of 0.15 mu g L-1, and standard deviation of 2.06 for tramadol were calculated. The linear range of the procedure for each drug was 1.0 to 1000 mu g L-1. This method was successfully applied for the analysis of codeine and tramadol in saliva samples.