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Effect of boiling on flavonoid contents of GLVs
Among the water extracts, the contents of epicatechin in amaranth, cowpea and pumpkin, rutin in pumpkin and spinach, as well as myricetin, quercetin, kaempferol, luteolin and apigenin in pumpkin decreased significantly (p < 0.05) after boiling. Among the 75% acetone extracts, epicatechin contents of amaranth, jute mallow and cowpea, as well as rutin contents of all GLVs also decreased after boiling. The content of total flavonoids of water extracts from boiled cowpea, pumpkin and spinach was less than that of raw samples. The 75% acetone extracts of all GLVs also exhibited a decrease in total flavonoids after boiling. A decrease in the flavonoid constituents of most GLVs after boiling may have resulted from leaching of vegetable flavonoids into the boiling water (Ruiz-Rodriguez et al., 2008). Boiling results in oxidation, thermal degradation, leaching and other events that led to lower levels of phenolics in processed food compared to fresh (Kalt, 2005). Salawu et al. (2009) found that extracts from blanched samples of jute mallow were highly viscous and showed very low concentration of phenolic constituents, forcing these researchers not to quantitatively evaluate the phenolic fractions in the extract. Gil et al. (1999) also reported leaching of approximately half of the flavonoids from spinach into boiling water. Boiling has also been reported to have induced flavonol losses of up to 20% and 40% in onions and asparagus, respectively (Makris and Rossiter, 2001). Price et al. (1998) reported significant leaching of flavonol glycosides into cooking water, which was also dependent on the conjugates. The decrease in phenolic compounds in broccoli inflorescences was also attributed to both leaching into the cooking water and thermal degradation by Vallejo et al. (2003).
The results of this study are in agreement with the study of Puupponen-Pimiä et al. (2003) who found that the effects of processing on flavonols were highly plant species-dependent. Because GLVs contain various kinds of polyphenol compounds, some polyphenols may protect others from decomposition during boiling. Buchner et al. (2006) studied the effect of thermal processing on rutin and quercetin and found that quercetin had the most intense degradation probably because these compounds split through different breakdown patterns, which could be explained by the protocatechuic acid being one of the cleavage reaction product of quercetin and the C-3 diglycosilation of rutin. In the present study, protocatechuic acid was not analysed and therefore the breakdown products of these flavonols in the extracts are not known. Murakami et al. (2004) reported that chlorogenic acid shielded rutin from decomposition when the two phenolic compounds were heated together. This was attributed to chlorogenic acid being an ester and therefore more easily decomposed than rutin. In contrast to the study of Murakami et al. (2004), chlorogenic acid decreased after boiling in water extracts of amaranth, cowpea, pumpkin and spinach as well as acetone extracts of amaranth (results not shown), but only water extracts of amaranth exhibited an increase in rutin after boiling.
Effect of extraction solvent
The variation in presence and content of flavonoids found in the two extraction solvents clearly show that it may not always be possible to extract multiple forms of phenolic compounds of interest with a single extraction solvent or solvent mixture. Luthria (2006) recommended the use of multiple solvent mixtures to extract different forms of compounds with varying polarities of conjugated mixtures. Polarity matching between the flavonoids of interest and extraction solvent is critical for optimum extraction as has been shown in this study.
Protection of erythrocytes by African GLVs against oxidative damage
Among raw samples, all GLV extracts with exception of raw jute mallow, significantly protected erythrocytes from AAPH-induced haemolysis (Figure 2.3.1). Extracts of raw jute mallow contributed to the damage of erythrocytes membrane and therefore offered no protection against haemolysis, while extracts from raw pumpkin offered the highest protection against haemolysis than the other extracts. GLVs are consumed after boiling, therefore it was appropriate to evaluate the effects of boiling on the efficiencies of these extracts to prevent haemolysis. Among extracts from boiled GLVs, amaranth offered highest protection against haemolysis, while jute mallow and pumpkin offered least protection. There was no difference in protection against haemolysis between raw and cooked amaranth extracts, although the results from the total antioxidant activity assays suggested that boiling reduced antioxidant activity in this vegetable (subsection 2.1.4.2). Furthermore, in agreement with the results of total antioxidant activity, extracts of raw pumpkin and spinach inhibited more haemolysis than when cooked.
Protection of plasmid DNA by African GLVs against oxidative damage
Figue 2.3.2 shows the electrophoretic pattern of pBR 322 plasmid DNA after AAPH-induced oxidative damage in the presence of GLV extracts. The absence of a band in Lane 2 is due to oxidative effects of AAPH that caused extensive damage and little of the original form of DNA. In other lanes (7 to 26), AAPH oxidation of plasmid DNA resulted in cleavage of supercoiled DNA to give the prominent open-circular form of DNA. Most of the plasmid DNA treated with African GLVs was retained in the supercoiled circular form (Lanes 8 to 22). This figure shows only two bands present in the agarose gel, indicating that the DNA was cut only once and converted to the open-circular form.
CHAPTER 1: INTRODUCTION
1.1 Statement of the Problem
1.2 Literature Review
1.2.1 African green leafy vegetables
1.2.2 Nutritional composition of African GLVs
1.2.3 Chemistry of plant phenolics
1.2.4 Phenolic compounds present in GLVs
1.2.5 Antioxidant properties of GLVs phenolics
1.2.6 Antioxidant mechanisms and structure-activity relationship of plant phenolics
1.2.7 Free radicals and oxidative stress
1.2.8 Evidence for health-promoting effects of fruits and vegetables
1.2.9 Health-promoting effects of some flavonoid-rich foods
1.2.10 Health-promoting effects of plant phenolics with particular reference to GLVs
1.2.11 Dietary intake of flavonoids
1.2.12 Bioavailability of phenolics
1.2.13 Effects of cooking on phenolic content and antioxidant activity
1.2.14 Analytical methodology for the determination of antioxidant content and activity28
1.2.15 Conclusions
1.3 Hypotheses
1.4 Objectives
CHAPTER 2: RESEARCH
2.1 Raw and cooked African green leafy vegetables have greater antioxidant content and activity than spinach
2.1.1 Abstract
2.1.2 Introduction
2.1.3 Materials and Methods
2.1.4 Results and Discussion
2.1.5 Conclusion
2.1.6 References
2.2 Comparative determination of flavonoids of African green leafy vegetables and spinach byhigh-performance liquid chromatography
2.2.1 Abstract
2.2.2 Introduction
2.2.3 Materials and Methods
2.2.4 Results and Discussion
2.2.5 Conclusion
2.2.6 References
2.3 Protective effects of African green leafy vegetables against AAPH-induced oxidative damage
2.3.1 Abstract
2.3.2 Introduction
2.3.3 Materials and Methods
2.3.4 Results and Discussion
2.3.5 Conclusion
2.3.6 References
CHAPTER 3: GENERAL DISCUSSION
3.1 Methodologies
3.2 Research Findings
3.3 African GLVs may reduce chronic diseases of lifestyle
CHAPTER 4: CONCLUSIONS AND RECOMMENDATIONS
