Abstract:
Chromatographic techniques can be made more environmentally friendly, i.e., “greener”,
by a number of different strategies. One approach is to replace the toxic mobile phase
solvents such as acetonitrile with greener alternatives. Acetone, in particular, has proven
to be a suitable alternative to acetonitrile, since the two solvents have similar
physicochemical properties, including solubility, miscibility and viscosity properties.
However, due to acetone’s high ultraviolet (UV) cut–off wavelength (330 nm), it normally
cannot be used as a mobile phase solvent when performing high performance liquid
chromatography (HPLC) analysis with UV detection. In this study, a reversed-phase
HPLC–UV method using acetone-containing mobile phase was developed for the
determination of reducing sugars in apple juice and two indigenous fruits, Berchemia
discolor and Hyphaene petersiana. Pre-column derivatisation of analytes via reductive
amination with p-aminobenzoic acid ethyl ester (ABEE) was performed to enable
photometric detection at 307 nm. The method was directly compared to a method that
utilised acetonitrile-containing mobile phase. Although the detection wavelength of the
ABEE derivatives is below the UV cut–off wavelength of acetone, it is high enough above
acetone’s absorbance maximum (~280 nm) to enable satisfactory detection of the
derivatives. Hence, the method compared well with the acetonitrile method, providing
similar resolution and selectivity, as well as sufficient sensitivity to facilitate the
quantitation of glucose and fructose in all the fruits and juice investigated in this study.
Although the method was only validated in terms of precision, linearity, limit of detection
and quantitation but no other aspects such as accuracy, it shows potential to be used as a
greener alternative for sugar analysis for laboratories that only have access to HPLC–UV
instruments
