Comparison Between Methyl and Trimethylsilyl Ester Derivatives in the Separation and GC Quantification of Triterpene Acids in Eugenia brasiliensis Leaf Extract

Abstract

This study details a method to characterize the triterpene acid-rich extract obtained from the defatted leaves of Eugenia brasiliensis (Myrtaceae) via extraction with 2 % NaOH in ethanol at room temperature. The crude extract (yield 2.35 %) was submitted to analysis by gas chromatography coupled to mass spectrometry (GC–MS) confirming ursolic acid as its major compound. The optimal conditions for the separation of oleanolic, betulinic and ursolic acids were assayed by GC with flame ionization detection (GC–FID) using two different columns (DB-5 and DB-17HT) and by applying two distinct derivatizing protocols. The use of a DB-17HT column led to the best results, with a shorter runtime and a better resolution (Rs) between the oleanolic and betulinic signals for both the bis-trimethylsilyl (Rs 2.84) and methyl ester derivatives (Rs 2.47). A DB-5 column also gave satisfactory results for the TMS ester, with a runtime of 30 min and Rs 2.14. Ursolic acid in the crude extract was quantified by comparison to two individual standard curves determined using commercial ursolic as its TMS derivative on the DB-5 column and its methyl ester on the DB-17HT column. Good linearity was achieved in both cases (r2 = 0.9776 and 0.9953, respectively), and the amounts of ursolic acid in the extracts were calculated to be 144.7 and 147.9 mg·g−1, respectively. These results showed no significant differences when compared using Tukey’s HSD test. Total triterpene acids amounted to 0.52 % in E. brasiliensis dry leaves.