Extraction, characterization and in vitro testing of flavonoids rich fractions obtained from Actinidia macrosperma fruit

Abstract

In this PhD study, phenolic compounds extracted from Actinidia macrosperma L. fruit grown in New Zealand, were separated and isolated by series of chromatographic procedures followed by identification on high performance liquid chromatography-tandem mass spectrometry (HPLC-ESI-MS/MS). Single factor experimental design was adopted to optimise the extraction conditions and their phenolic profiles, antioxidant and antihypertensive activities were compared to commercial kiwifruit varieties. Among the extracts tested, 70% aqueous acetone by steeping the A. macrosperma fruit variety showed the highest values of total phenolic (TP) content (823.1±14.4 mg gallic acid equivalent/100 g DW), total flavonoid (TFO) content (170.9±1.9 mg catechin equivalent/100 g DW), total flavanol (TFA) content (82.6±0.6 mg catechin equivalent/100 g DW) and antioxidant capacity (5.1±0.1 mmol Trolox equivalent (TE)/100 g DW and 8.3±0.1 mmol Fe (II) equivalent/100 g DW for DPPH and FRAP assays, respectively). The qualitative and quantitative analysis of each defatted crude extract on high pertformance liquid chromatography-diode array detection (HPLC-DAD) and high pertformance liquid chromatography-tandem mass spectrometry (HPLC-ESI-MS/MS) revealed that 70% aqueous acetone extract from the A. macrosperma fruit contains many potential antioxidant flavonoids compared to commercial kiwifruit varieties namely, Actinidia deliciosa cv Hayward, Actinidia deliciosa cv Sweetgreen, Actinidia chinensis cv Sungold and Actinidia chinensis cv Gold. Quercetin-3-O-galactoside was identified as the most abundant flavonoid among the other flavonoids present in A. macrosperma fruit. Previously undiscovered eleven flavonoids from the A. macrosperma fruit were tentatively identified by using LC-ESI/MS/MS after a series of purification steps including liquid-liquid partitioning, silica gel flash column chromatography, size exclusion chromatography on Sephadex LH-20, and semi-preparative HPLC. It could be concluded that the optimized extraction conditions along with the HPLCESI- MS/MS technique were effectively performed to identify eleven flavonoid compounds, not previously reported from Actinidia macrosperma fruit. This is the first study regarding the phenolic composition and antioxidant properties of new commercial kiwifruit varieties, Actinidia deliciosa cv Sweetgreen, Actinidia chinensis cv Sungold and non-commercial variety, Actinidia macrosperma cultivated in New Zealand. Finally, the effect of Actinidia macrosperma fruit on inhibition of angiotensin converting enzyme (ACE) has not been previously reported and could therefore be recorded as a novel biological activity.