Michael Francis Dube

Enhancement of volatile aglycone recovery facilitated by acid hydrolysis of glucosides from Nicotiana flower species.

Four different Nicotiana flowers (Nicotiana alata (alata), Nicotiana sylvestris (Sy), Nicotiana suaveolens (Su), and Nicotiana tabacum cv. Flue-Cured (FC)) from farms in Virginia and North Carolina were harvested and promptly quenched with liquid nitrogen and hand-ground prior to analysis. Each Nicotiana flower was pre-extracted with hexane to remove unbound volatiles. Fifteen standard compounds that were thought to be in the pre-extract were employed to aid in GC-MS identification and quantification. Glucosides were then chromatographically isolated and next hydrolyzed via 2 M sulfuric acid for 24 h at 75 °C. For each flower, the products of hydrolysis were extracted in tandem with hexane and dichloromethane (DCM) prior to analysis by GC-MS. The mixture of hexane and DCM extracts of the flowers after hydrolysis were then analyzed for each of 15 external standards via GC-MS to determine the concentration of any isolated flower-derived aglycone. Quantitative results for each of the possible 15 free volatile compounds extracted before and after hydrolysis were compared. Benzyl alcohol, phenethyl alcohol, and cis-3-hexenol were found in all Nicotiana both before and after acid hydrolysis. Enormous increases in the mass of benzyl alcohol and phenethyl alcohol were obtained with all flowers as a result of acid hydrolysis. With selected Nicotiana flowers, significant increases were observed for eugenol and cinnamaldehyde. The significant increases observed in cinnamaldehyde and eugenol upon mild acid hydrolysis strongly indicate that this approach could be a viable alternative process for the production scale isolation of these important natural flavor compounds.

Synthesis of Pyrazines Using Sugar Derived from Tobacco Cellulose and Hydrolyzed Tobacco F1 Protein as an Amino Acid Source

Summary An array of pyrazines have been synthesized using sugars derived from tobacco cellulose (CDS), ammonium hydroxide, and hydrolyzed tobacco F1 protein as a source of free amino acids (isolated amino acids from F1 hydrolysate, from filtered F1 hydrolysate and from non-filtered F1 hydrolysate). All reactions were performed at 120 °C for 60 min using a 40-mL Parr reaction vessel. Results showed that the addition of hydrolyzed F1 protein as free amino acid source increased the number of pyrazines with branched alkyl chains (for example, 2-butyl-3-methyl pyrazine) compared to when no amino acids were added. However, using isolated amino acids from hydrolyzed F1 protein versus just hydrolyzed F1 protein (filtered or not filtered) did not make a difference in yield or type of branched pyrazines. When non-filtered hydrolyzed F1 protein was used, the solution was much more viscous and contained suspended solid material when compared to the use of filtered hydrolyzed F1 protein. Addition of threonine (THR) to the reaction mixture did not increase the yield of pyrazines but did slightly shift the distribution of pyrazines toward those with three and four carbons attached. Similar but not identical arrays of pyrazines were obtained when somewhat resembling reaction conditions were applied on a larger reaction scale (~1.5 L). A significant 50%-decrease in pyrazine yield was observed when the reaction temperature was reduced from 120 to 100 °C. No noticeable difference in the array of pyrazines from these two reactions was observed. In the majority of cases, the presence of free amino acids resulted in an increase in pyrazine yield coupled with a change in the qualitative array of pyrazines. These results clearly illustrate that sugar prepared from tobacco cellulose (glucose) can be used just like high fructose corn syrup to prepare flavor compounds via Amadori and Maillard reactions. The evidence highlights that hydrolyzed amino acids from F1 tobacco protein can be used via Maillard reactions to produce complementary arrays of pyrazine flavor compounds.