Identification of Flavonoid–4'–Glucosides
by Beta–Glucosidase
Digestion
Extracts from the albedo of 'Kao Panne' pummelo had high proportions of
naringin–4'–glucoside and narirutin–4'–glucoside, high
overall flavanone content, and simple HPLC profiles. In order to correctly
identify the flavonoid–4'–glucosides in these samples, beta–glucosidase
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. Beta–glucosidase
was added to one aliquot of this mixture to a final concentration of 1 mg/mL. A
parallel sample was prepared except that beta–glucosidase 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
for HPLC.
The percentage gain of the naringin peak and the concomitant loss of the
naringin–4'–glucoside peak in the glucosidase-treated sample were
compared with the total percentage of naringin and the putative naringin–4'–glucoside
peak in the untreated sample. If all other percentages remained constant, the
loss of the naringin–4'–glucoside peak and the gain in the naringin
peak that agreed with the percentage of naringin–4'–glucoside in the
untreated sample was considered as evidence for naringin–4'–
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 narirutin–4'–glucoside.
United States Department of
Agriculture
Agricultural Research
Service
The material on this page is in the public domain.
Original posting: April 1, 1999. ΓΏ |
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