The increase in human plasma antioxidant capacity after apple consumption is due to the metabolic effect of fructose on urate, not apple-derived antioxidant flavonoids
Introduction
Regular consumption of fruits and vegetables lowers the risk of cardiovascular diseases, certain types of cancer, and other chronic diseases [1], [2]. These beneficial effects of fruits and vegetables have been attributed in part to their high content of flavonoids, the intake of which also is inversely associated with the incidence of many chronic diseases [3], [4], [5]. Apples are one of the main sources of flavonoids in the Western diet, together with tea, wine, onions, and chocolate [4], [5], [6]. An increased intake of apples has been correlated with a decreased risk of heart disease and type 2 diabetes [3]. In addition, apple consumption has been associated with a decreased incidence of thrombotic stroke [7].
The mechanism(s) by which flavonoids in fruits and vegetables may lower chronic disease risk remains to be fully elucidated. Most flavonoids have antioxidant properties, and extracts and juices of fruits and vegetables exhibit substantial antioxidant capacity in vitro [8], [9], [10]. Therefore, it is conceivable that the health benefits of flavonoid-rich foods are related to the antioxidant protection of biological macromolecules, such as lipids, proteins, and DNA. However, this notion remains controversial. Although some studies have failed to show a short- or long-term antioxidant effect in humans of consumption of fruits, vegetables, or flavonoids [11], [12], [13], other studies have reported such effects, in particular an acute increase in the antioxidant capacity of plasma [14], [15], [16]. Interestingly, flavonoids are poorly absorbed by humans [17], and the increase in plasma antioxidant capacity observed after consumption of flavonoid-rich foods often greatly exceeds the increase in plasma flavonoids [18], [19], [20].
In addition, we recently found that apple extracts added to human plasma in vitro significantly protected endogenous urate, α-tocopherol, and lipids from free radical-mediated oxidation, but such antioxidant effects were not observed in plasma ex vivo after apple consumption by healthy subjects [21]. These data suggest that apple-derived flavonoids are not absorbed in sufficient amounts to significantly contribute to the antioxidant defenses in plasma in vivo.
In follow-up to these observations [21], we investigated the effects of apple consumption in humans on plasma antioxidant levels and total antioxidant capacity. We found a transient increase in plasma antioxidant capacity after apple consumption, which was explained by a transient increase in plasma urate. Furthermore, we discovered that very similar increases in plasma antioxidant capacity and urate could be achieved by consumption of fructose.
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Materials
Ascorbic acid, uric acid, ferric chloride (FeCl3), diethylenetriaminepentaacetic acid (DTPA), and ascorbate oxidase were purchased from Sigma (St. Louis, MO, USA). 2,4,6-Tri(2-pyridil)-s-triazine (TPTZ) was purchased from Fluka (Milwaukee, WI, USA). Chelex Resin 100 was purchased from Bio-Rad (Hercules, CA, USA). Water used in experiments was deionized (Milli Q) and additionally treated overnight with Chelex and then filtered. All solvents used were of high-performance liquid chromatography
Effect of apple consumption on plasma FRAP and antioxidants in humans
Plasma was obtained from six healthy subjects before and up to 6 h after consumption of five apples, and FRAP, ascorbate, and urate were determined. Plasma FRAP was 445 ± 35 μM trolox equivalents (TE) at baseline. After apple consumption, FRAP increased significantly (p < .0001, repeated-measures ANOVA), with a 12% increase (54.6 ± 8.7 μM TE) after 1 h (Fig. 1A). To assess the contribution of ascorbate, FRAP was analyzed after incubation of plasma with ascorbate oxidase. Baseline FRAPAO (363 ±
Discussion
In this in vivo study, we investigated the short-term effect of consuming five Red Delicious apples on plasma antioxidant capacity in six healthy subjects. One hour after apple consumption, a large, statistically significant increase in plasma antioxidant capacity was observed, as assessed by FRAP. This increase in FRAP could have been due to the absorption and uptake into the circulation of antioxidant flavonoids from apples. However, our subsequent analyses showed that the antioxidant
Acknowledgements
This work was supported by a grant from the Washington Tree Fruit Research Commission (Wenatchee, WA, USA). The authors thank Leslee Lucas and her staff (Student Health Services, Oregon State University, Corvallis, OR, USA) for their assistance in drawing blood, and the volunteers of the Linus Pauling Institute who participated in this study.
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