Conversion of Exogenous Cholesterol into Glycoalkaloids in Potato Shoots, Using Two Methods for Sterol Solubilisation

Erik V Petersson,Rikard Tröger,Paresh C Dutta,Nurun Nahar,Anders Broberg,Paul Dahlin,Folke Sitbon,Lisbeth Jonsson

doi:10.1371/journal.pone.0082955

Abstract

Steroidal glycoalkaloids (SGA) are toxic secondary metabolites naturally occurring in the potato, as well as in certain other Solanaceous plant species, such as tomato, eggplant and pepper. To investigate the steroidal origin of SGA biosynthesis, cut potato shoots were fed cholesterol labelled with deuterium (D) in the sterol ring structure (D5- or D6-labelled), or side chain (D7-labelled), and analysed after three or five weeks. The labelled cholesterol and presence of D-labelled SGA were analysed by GC-MS and LC-MS/MS, respectively. When feeding D-labelled cholesterol solubilised in Tween-80, labelled cholesterol in free form became present in both leaves and stems, although the major part was recovered as steryl esters. Minor amounts of D-labelled SGA (α-solanine and α-chaconine) were identified in cholesterol-treated shoots, but not in blank controls, or in shoots fed D6-27-hydroxycholesterol. Solubilising the labelled cholesterol in methyl-β-cyclodextrin instead of Tween-80 increased the levels of labelled SGA up to 100-fold, and about 1 mole% of the labelled cholesterol was recovered as labelled SGA in potato leaves. Both side chain and ring structure D labels were retained in SGA, showing that the entire cholesterol molecule is converted to SGA. However, feeding side chain D7-labelled cholesterol resulted in D5-labelled SGA, indicating that two hydrogen atoms were released during formation of the SGA nitrogen-containing ring system. Feeding with D7-sitosterol did not produce any labelled SGA, indicating that cholesterol is a specific SGA precursor. In conclusion, we have demonstrated a superior performance of methyl-β-cyclodextrin for delivery of cholesterol in plant tissue feeding experiments, and given firm evidence for cholesterol as a specific sterol precursor of SGA in potato.

Highlights

Sterols are major components of cellular membranes in eukaryotic organisms
About 80% of D5-cholesterol was recovered after saponification of the sterol fraction, indicating that the major part was stored as steryl esters (Table 2)
We demonstrate here that the choice of solubilisation method has a profound impact on the conversion of cholesterol into steroidal glycoalkaloids (SGA) in cut potato shoots

Summary

Introduction

Cholesterol is the main sterol, whereas plants contain a mixture of sterols with varying length and saturation of the side chain [1]. It was believed that plants did not contain cholesterol, but since the finding of cholesterol in potato and Dioscorea spiculiflora [2], cholesterol has been found in most plant species, usually as a minor sterol. In some genera, e.g. the Solanaceae and Liliaceae, cholesterol constitutes a significant part of total sterols [3,4]. Besides having a role in membrane structure and function, it has been proposed that plant cholesterol is a precursor of specific steroidal secondary metabolites, such as sapogenins and steroidal glycoalkaloids (SGA) [5]. The SGA are toxic [6,7,8,9], and food authorities in many countries have established a maximum level of 200 mg total SGA/kg fresh weight (FW) in unpeeled potato tubers aimed for consumption [9,10]

Methods

Results

Conclusion