Analysis of amino acids in tobacco by derivatization and dispersive liquid–liquid microextraction based on solidification of floating organic droplet method

• Derivatization and microextraction of amino acids were performed in one step.
• The derivatives of amino acids were extracted by 2-dodecanol which is less toxic.
• First report on the application of DLLME-SFO for analysis of amino acids in tobacco.
• Good recoveries and reproducibilities in a short analysis time.

A new one-step derivatization and microextraction technique was developed for determination of amino acids in tobacco samples. In the proposed method, amino acids were derivatized with isobutyl chloroformate (IBCF) in aqueous solution. The derivatives were extracted by dispersive liquid–liquid microextraction based on solidification of floating organic droplet (DLLME-SFO). Parameters affecting the extraction efficiency were investigated in detail. The optimum conditions were as follows: a mixture of 500 μL acetone, containing 40 μL 2-dodecanol, was rapidly injected by syringe into the 2 mL water sample. After centrifugation, the 2-dodecanol droplets were floated at the top of the tube. Then, the tube was cooled in an ice bath. After 5 min the solvent had solidified and was then transferred into a conical vial; it melted quickly at room temperature and it was injected into a gas chromatograph for analysis. Under the optimum conditions, the limits of detection were of the order of 0.12–2.82 μg/mL. The calibration curves showed good linearity over the investigated concentration range between 0.5 and 200 μg/mL with a coefficient of estimation (R2) > 0.9887 for GC–MS (SIM). The proposed method is an alternative approach to the quantification of amino acids in tobacco samples.