GLYCOSIDES IN Viola Tricolor L.

CHAPTER 2: INTRODUCTION
Todifferentiatebetweentheisomers,twobasicmethodshavebeendescribed.
On one hand, they can be distinguished by their product ion spectra, through
the analysis of the Y1, Y0, and Z1 ions. The presence ofthe Z1 fragment implies
that one sugar unit is linked to another sugar and not directly to the aglycone,
therefore indicates an O- or C,O-diglycosyl structure [96, 98, 106]. In addition,
differences were observed in the relative abundancesofthe Y −
1
and Y−
0
ions for
thetwoO-glycosyl isomerforms. Fordi-O-glycosides,theY −
1 ionwasreported
tobethebasepeak(100%relativeabundance),andtheY −
0 ofmediumintensity
(about 30% relative abundance), whilst in the spectra of O-diglycosides the Y −
0
ion exhibited the highest abundance (Fig. 2.8) [106]. Analogously, for di-C,O-
glycosides, the Y −
0
case of C,O-diglycosides, the Y −
0
theZ −
1
observably exhibited the highest abundance, whereas in the
showed less than 10 % relative intensity, and
fragmentwasfoundtobethebasepeak[96]. Theothermethodisbased
on the presence of characteristic radical ions. The product ion spectra of di-
O-glycosides contained [Y1–H] −• radical ions and a [Y0–2H] − fragment. In the
spectra of O-diglycosides only the [Y0–H] −• radical ion was observed, the [Y1–
H] −• ionsweremissing,asitwasdemonstratedforflavonolglycosides(Fig.2.9a
and b) [104]. Similarly, the [Y1–H] −• ion was typically observed upon cleavage
oftheglycosidicbondbetweentheaglyconepartandtheO-linkedglycan,thus
only forthe di-C,O-glycosyl flavoneisomer [113].
Apparently,thedeterminationofthesugarunitdistributionwouldbeeasier
inthecaseofglycosideswithdifferentsugarunits. Asafirstapproximation,the
product ion spectra of di-O-glycosides would contain two Y1 + ions at different
m/z values, whereas in the spectra of O-diglycosides only one Y1 + fragment
would be observed, corresponding to the loss of the external sugar unit. In
the product ion spectra of O-diglycosides, however, a second Y1 + ion was also
reported–usuallylabeledasY ∗ –,originatingfromthelossoftheinternalsugar
residue making spectra interpretation misleading[108].
Glycan sequence Thisstructural characteristic isonly relevantfor glycosides
withdifferentsugarunitsanddeterminationoftheglycansequenceseemsself-
explanatory [98]. For example, in the product ion spectra of O-diglycosides,
theY1(externalsugar) andY0 fragmentswereexpected. However,asitwasdemon-
strated on the example of rutin (quercetin-3-O-rhamnosyl(1→6)glucoside), the
negative ion MS/MS spectra of (1→6) diglycosides often did not show signif-
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