Identification and Quantification of Stilbenes in Fruits of Transgenic Tomato Plants (Lycopersicon esculentumMill.) by Reversed Phase HPLC with Photodiode Array and Mass Spectrometry Detection

Abstract

Reversed-phase high-performance liquid chromatography (RP-HPLC) with photodiode array (PDA) and mass spectrometry (MS) detection was employed to study the accumulation of stilbenes and other naturally occurring polyphenol intermediates of flavonoid pathway in tomato fruits of plants genetically modified to synthesize resveratrol. The transgenic tomato fruits were obtained by overexpression of a grapevine gene encoding the enzyme stilbene synthase in tomato plants (Lycopersicon esculentum Mill.). Stilbenes and flavonoids, either glycosylated or free, were simultaneosly identified by electrospray interface (ESI)-MS in negative ionization mode and were quantified by PDA detection at the wavelength corresponding to their maximum absorbance. The two detectors were coupled online with an HPLC system utilizing a narrow-bore C18 reversed-phase column, which was eluted by a multistep gradient of increasing concentration of acetonitrile in water containing 0.5% (v/v) formic acid. The results of these analysis revealed that the genetic modification of the tomato plants originated different levels of accumulation of four stilbenes (i.e., trans- and cis-piceid and trans- and cis-resveratrol) in their fruit depending on the stages of ripening. Either at immature or at mature stages of ripening the stilbenes were preferentially accumulated in the fruit peel as the glycosylated form. The highest amount of trans-piceid and trans-resveratrol were found in the peel of fruits harvested at mature stage of ripening. The variations in the levels of rutin, naringenin, and chlorogenic acid found in the samples extracted from the fruits of transgenic tomato plants, in comparison to that determined in the control lines, seemed to be related to the genetic transformation, whose effect on the flavonoid biosynthetic pathway needs to be elucidated by additional studies.

MS‐based analytical methodologies to characterize genetically modified crops

“…On the basis of the retention times and UV and MS data, the identification of resveratrol and its glycosilated forms was possible in one analysis. Results indicated differences in the levels of rutin, naringenin, and chlorogenic acid found in transgenic tomatoes in comparison to the control lines; those differences seem to be related to the genetic transformation (Nicoletti et al, 2007).…”

“…Hierarchical clustering of data revealed a closer correlation between GM/non-GM plants of the same variety than between conventional plants of different varieties. In a different study, Nicoletti et al (2007) concentrated on the LC-MS profiling of stilbenes, a specific class of polyphenols, in transgenic tomato. The GM tomato overexpressed a grapevine gene that encoded the enzyme stilbene synthase.…”

Profiling of Genetically Modified Organisms Using Omics Technologies

“…Similarly, LC-ESI-Q MS has been used to investigate GM tomato overexpressing a grapevine gene that encodes the enzyme stilbene synthase [90].…”

MS‐Based Methodologies for Transgenic Foods Development and Characterization

“…Hierarchical clustering of data revealed a closer correlation between GM/non-GM plants of the same variety than between unmodified plants of different cultivars. Also, an LC-MS method has been developed for the profiling of stilbenes, a specific class of polyphenols, in transgenic tomato overexpressing a grapevine gene that encoded the enzyme stilbene synthase (Nicoletti et al, 2007). The study was focused on the detection of potential alterations on the synthesis of other metabolites along the flavonoid's pathway.…”