[High performance liquid chromatographic determination of reducing sugars in fruit juices with laser resonance Raman...

A liquid chromatographic method using semi-microcolumn separation combined with laser resonance Raman detection was developed for the determination of monosaccharides in fruit juices. Reducing sugars (galactose, glucose, mannose, arabinose, xylose and ribose) were derivatized with 4-dimethylaminoazobenzene-4'-sulfonyl (DABSYL) hydrazine. The derivatives of the monosaccharides exhibited strong resonance Raman scattering at 1136 cm(-1) when Ar+ laser emission line 488.0 nm was used. A semi-microcolumn Inertsil ODS-2 (250 mm x 1.5 mm i.d.) was used to the separation of the monosaccharide derivatives. The saccharides eluted within 25 min. However the excess DABSYL hydrazine was retained strongly with a retention time of about 60 min. In order to reduce the longer analysis time, the excess DABSYL hydrazine was removed by adding glyoxylic acid and then Na2CO3 aqueous solution when the derivation reaction was finished. As a result, the analysis time was reduced to about a half of its initial run time. The sensitivity of resonance Raman detection greater two orders of magnitude than refractive index (RI) detection. The detection limit of glucose is 10 ng (5.5 pmol). The high selectivity of Raman detection came from the facts that only the interested compounds in a given sample was derived with the Raman labeling reagent and that only the characteristic Raman bands of the derivatives were selectively detected. In this work, in spite of the coexisting large quantities of organic acids in fruit juice, the determined values of glucose agreed well with those obtained by RI detection without any derivatization procedure.