Kai Cai, Jinsha Ba, Jie Zhang, Ruijuan Zhao, Yunfei Ma, Yechun Lin, Changyun Zhang, Decheng Li, Weichang Gao, Kesu Wei
Design of experiments with central composite design was employed to achieve the ideal extraction conditions, with a shaking count of 70 times and a static immersion time of 36 min. The matrix effect in the aqueous tobacco extract showed a suppression effect with relative quantitative errors below 5.0%, and the test strip's enzymatic reaction exhibited high specificity for glucose. The developed method showed a good linear relationship over the range of 2.00-90.0 mg/L (y = 0.9952× + 0.0253, R 2 = 0.9991). The average recoveries ranged from 94.1% to 96.0% for flue-cured tobacco and 95.0% to 97.5% for fresh tobacco leaves, with repeatability RSDs ≤ 3.86% and ≤ 4.51%, reproducibility RSDs ≤ 5.00% and ≤ 5.98%, respectively. Compared with the traditional derivatization GC-MS method, the linear regression slope between the two methods was 0.9781 (r = 0.9997 and p < 0.01), and the overall AGREEprep score of test strip with reflectometry method was 0.78, further confirming both the excellent accuracy and greenness. Finally, this method was successfully applied to evaluate glucose variation among different cultivars and growth stages, demonstrating its practical applicability.
INTRODUCTION: Glucose in tobacco leaves acts as a vital chemical precursor that thermally degrades and reacts with amino acids during combustion and curing to enrich aldehydes, ketones, and nitrogen-containing heterocyclic aroma compounds, thereby shaping flavor and masking harshness. However, current analytical methods for glucose quantification are limitations of high complexity, poor portability, and low eco-friendliness.
METHODS: A facile approach for on-site method for glucose determination in both flue-cured and fresh tobacco leaves, based on aqueous extraction followed by test-strip reflectometric analysis, was developed.
RESULTS: Design of experiments with central composite design was employed to achieve the ideal extraction conditions, with a shaking count of 70 times and a static immersion time of 36 min. The matrix effect in the aqueous tobacco extract showed a suppression effect with relative quantitative errors below 5.0%, and the test strip's enzymatic reaction exhibited high specificity for glucose. The developed method showed a good linear relationship over the range of 2.00-90.0 mg/L (y = 0.9952× + 0.0253, R 2 = 0.9991). The average recoveries ranged from 94.1% to 96.0% for flue-cured tobacco and 95.0% to 97.5% for fresh tobacco leaves, with repeatability RSDs ≤ 3.86% and ≤ 4.51%, reproducibility RSDs ≤ 5.00% and ≤ 5.98%, respectively. Compared with the traditional derivatization GC-MS method, the linear regression slope between the two methods was 0.9781 (r = 0.9997 and p < 0.01), and the overall AGREEprep score of test strip with reflectometry method was 0.78, further confirming both the excellent accuracy and greenness. Finally, this method was successfully applied to evaluate glucose variation among different cultivars and growth stages, demonstrating its practical applicability.
DISCUSSION: This study developed a simple, portable, and green method for the determination of glucose content in different tobacco matrices, which could provide scientific reference data for tobacco quality evaluation and physiological growth monitoring.