Identification of Flavonoid4'Glucosides
Extracts from the albedo of 'Kao Panne' pummelo had high proportions of
naringin4'glucoside and narirutin4'glucoside, high
overall flavanone content, and simple HPLC profiles. In order to correctly
identify the flavonoid4'glucosides in these samples, betaglucosidase
was added to prepared extracts to selectively remove the 4'glucose groups
before examination by HPLC. Samples of 'Kao Panne' albedo were ground in liquid
nitrogen, after which 50 mL of a 200-mM NaOAc, pH 5 solution was added. Betaglucosidase
was added to one aliquot of this mixture to a final concentration of 1 mg/mL. A
parallel sample was prepared except that betaglucosidase was not added.
Both mixtures were incubated at room temperature overnight. A trichloroacetic
acid (Sigma Chemical Co., St. Louis, MO*)
solution was added to obtain a 25-percent mixture to precipitate proteins. The
resulting mixture was centrifuged and filtered as described above to prepare it
The percentage gain of the naringin peak and the concomitant loss of the
naringin4'glucoside peak in the glucosidase-treated sample were
compared with the total percentage of naringin and the putative naringin4'glucoside
peak in the untreated sample. If all other percentages remained constant, the
loss of the naringin4'glucoside peak and the gain in the naringin
peak that agreed with the percentage of naringin4'glucoside in the
untreated sample was considered as evidence for naringin4'
glucoside. Once the identity of the 4'glucoside peaks were confirmed in
this manner, peak assignments were then made using the original
methanol:dimethylsufoxide extract HPLC runs.
The same procedure was followed on 'Cutter' sweet orange for similar reasons
in the identification of narirutin4'glucoside.
United States Department of
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Original posting: April 1, 1999.ÿ