Major Carotenoids in Tobacco Laminas: Identification and Quantification by HPLC with Photodiode Array Detection

Abstract

The carotenoid compositions in various tobacco laminas are determined by high-performance liquid chromatography (HPLC) with the photodiode array detection (DAD). The carotenoids are extracted from powdered tobacco laminas by cold acetone containing 0.1 % butylated hydroxytoluene (BHT). A mobile phase of CH₃CN-H₂O (90:10, v/v) (A) and CH₃COOC₂H₅ (100 %) (B) with following gradient elution is developed: 100 % A in the beginning, maintained for 20 min, decrease to 50 % A in 50 min, maintain for 5 min and return to 100 % A in 1 h. A total of 11 carotenoids including some isomers in tobacco laminas are resolved within 55 min by using a ZorBAX SB-C₁₈ column with the flow-rate at 1.0 mL/min and detection at 445 nm. The limits of detection (LODs) of carotenoids varies from 6.0 to 14.0 ng/mL. The relative standard deviations (RSD) are from 2.69 to 3.63 % and the recovery ranges from 89.6 to 96.3 %. The method is successfully applied for the quantification of carotenoids in green, fresh fluecured and aging tobacco laminas and the analysis results show that total carotenoids in tobacco laminas range from 49.66 to 820.1 μg/g. Lutein and β-carotene are the most representative carotenoids in flue-cured and aging tobacco laminas because other carotenoids degrade greatly during flue-curing and aging periods. Furthermore, it worth to mention that zeaxanthin is not found in green tobacco laminas while presented in fresh flue-cured and aging tobacco laminas, luteoxanthin as a new carotenoid is first tentatively identified in the present work and α-carotene is not found in any significant amount in all those tobacco laminas.