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INRA, Unité de Nutrition Humaine, Equipe Stress Métabolique et Micronutriments, Centre de Clermont-Ferrand/Theix Saint-Genès Champanelle, FRANCE
Address reprint requests to: A. Mazur, Unité de Nutrition Humaine, Equipe Stress Métabolique et Micronutriments, Institut National de Recherche Agronomique, Centre de Clermont-Ferrand/Theix, 63122 Saint-Genès Champanelle, FRANCE. E-mail: mazur{at}clermont.inra.fr
| ABSTRACT |
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Methods: We investigated the effects of diets containing i) complex dietary carbohydrates and antioxidant micronutrients (potato Solanum tuberosum L.), ii) complex carbohydrates (starch) and iii) a simple carbohydrate (sucrose) on lipid metabolism and antioxidant status in rats.
Results: An increase in short chain fatty acid (SCFA) pools was observed in the cecum of rats fed a potato-based diet, resulting from an increase in all SCFAs, especially propionate (+360%, P < 0.0001). Feeding rats a potato-based diet for 3 weeks led to a decrease in cholesterol (–37%, potato vs. control and –32%, potato vs. sucrose) and triglycerides (–31%, potato vs. control and –43%, potato vs. sucrose) concentrations in triglyceride-rich lipoproteins (TGRLP) fractions. The antioxidant status was decreased by sucrose consumption and improved by potato consumption.
Conclusions: Our present results suggest that consumption of complex carbohydrates (provided as cooked potatoes), in combination with different antioxidant micronutrients, may enhance the antioxidant defences and improve lipid metabolism, when compared with starch (complex carbohydrates) and to sucrose consumption (source of simple sugar). These effects limit oxidative stress and reduce the risk of developing the associated degenerative diseases, including cardiovascular disease, and could have potential in cardiovascular disease prevention.
Key words: potato, sucrose, carbohydrates, cardiovascular disease, antioxidant status
| INTRODUCTION |
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D-fructose, when joined with a molecule of glucose, is a component of the disaccharide sucrose. Although there is little evidence that modest amounts of fructose have detrimental effects on carbohydrate and lipid metabolism, higher levels of fructose have been associated with several metabolic disorders in humans and in laboratory animals [3,4]. High-sucrose and high-fructose diets have been used in animal models to induce well-characterized metabolic dysfunctions. These animal models exhibit many of the symptoms of an early stage metabolic syndrome (syndrome X), a disorder that includes cardiovascular risk factors, insulin resistance, dyslipidemia and hypertension [5–7].
McDonald [8] suggests that the dietary sucrose, as opposed to complex carbohydrates, may have a differential effect on net oxidative stress and that these differences are reflected in the accumulation of advanced glycation products. Although the mechanism accounting for this increased oxidative damage has yet to be elucidated, it appears that the fructose moiety of the sucrose molecule plays a larger role than the glucose moiety. A high-fructose diet stimulates de novo lipogenesis, it increases the hepatic VLDL secretion and decreases the peripheral triglyceride clearance [9]. A diet rich in sucrose could alter cellular metabolism via several pathways and thereby increase oxidative stress. Indeed, current evidence suggests that dietary sucrose, as opposed to other complex carbohydrates, may have a differential effect on net oxidative stress and that the difference is reflected in the accumulation of advanced glycation products (Maillard reaction) [8]. Moreover, it has been shown that rats fed a high sucrose diet had altered heart antioxidant enzyme activity (Cu-Zn-SOD) and stress-related gene expression patterns [10]. Thus, the increased oxidative stress and the associated lipid peroxidation could be due to oxygen free radical production [11] and/or decreased protection from non-enzymatic or enzymatic antioxidants [12].
We hypothesized that the different carbohydrate compositions of starch, potato and sucrose may have different effects on lipid metabolism and antioxidant status in rats, potentially by impacting TG clearance and from the supply of antioxidant micronutrients provided by potato. To address this hypothesis, we compared the health effects of replacing sucrose with potato in the diet of rats. Among plant foods, potato (Solanum tuberosum L.) was thought to have protective effects. Indeed, the potato tuber is a source of fiber (especially when potato is consumed with skin) and antioxidant molecules, such as vitamin C [13] and E, carotenoids (principally lutein) [14] and phenolic acids (mainly caffeic and chlorogenic acids) [15,16]. These antioxidants are able to efficiently scavenge superoxides and peroxyl radicals, and in concert with the endogenous defense systems, they limit oxidative stress. This reduces the risk of developing degenerative diseases [17,18] such as cardiovascular disease, possibly by protecting lipoproteins from peroxidation [19–21].
| MATERIALS AND METHODS |
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Animals, about 180 g each, were housed in pairs, at 22°C with a 12 hour light-dark cycle (light from 8:00 to 20:00h) and access to food from 16:00 to 8:00h. The rats were randomly divided into three groups (8 per group) and fed a control diet, a 30% sucrose diet or a 30% potato diet (Table 1) ad libitum for 3 weeks. The diets were supplemented with antioxidants using mineral and vitamin supplements. However, in order to enhance oxidative stress, the vitamin supplement did not contain
-tocopherol. Therefore, in the control and sucrose diets, vitamin E was exclusively provided by corn oil, whereas the potato diet provided traces of vitamin E from the potatoes, in addition to vitamin E from corn oil. Potatoes were purchased from a local supplier "Jardin de Limagne". The potatoes were steamed and mashed with skin and accounted for 30% (dry matter) of the total diet. Food consumption and body weight were recorded twice a week.
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Rats were anesthetized during the post-absorptive period (between 08.00 a.m. and 10.00 a.m.), when the cecal fermentation is still active, by intraperitoneal sodium pentobarbital injection (40mg/kg of body weight). Blood was drawn from the abdominal aorta into heparinized tubes and centrifuged at 12,000 g for 2 min. Plasma samples were stored at 4°C for lipid and lipoprotein analysis, or immediately frozen and stored at –80°C for antioxidant assays.
After blood sampling, the cecum (wall with contents) was removed and weighed. The cecal wall was flushed clean with water, dried and weighed (cecal wall weight). Samples of cecal contents were collected and immediately frozen at –20°C. Supernatants for the short-chain fatty acid (SCFA) analysis were obtained by centrifugation at 20,000 g for 10 min. at 4°C.
The livers were freeze-clamped and stored at –80°C for lipid analysis and the peroxidation assay. Hearts were rapidly washed in physiological serum and immediately stored at –80°C for the lipid peroxidation assay.
Analytical Procedures
SCFA were measured in aliquots of cecal supernatants by gas-chromatography as previously described [22].
The plasma total cholesterol concentrations were determined enzymatically using a kit purchased from BioMerieux (Charbonnières-les-bains, France). Plasma triglyceride concentrations were determined using a kit from Biotrol (Paris, France). Liver and heart lipids were extracted with chloroform/methanol (2:1, v/v) as previously described [23].
Plasma lipoproteins were separated on a potassium bromide density gradient by ultracentrifugation (100,000 g for 24 hours at 15°C) using 2 ml plasma samples. The gradient was divided into 24 fractions of 500µL and the cholesterol and triglyceride contents of each fraction were determined as described above for the plasma samples. The results were expressed for pools with d < 1.040 kg/L (mainly triglyceride-rich lipoprotein: TGRLP, with a minor contribution of LDL) and d > 1.040 kg/L fraction (essentially HDL).
Vitamin E was analyzed by HPLC-UV [24]. Briefly, vitamin E was extracted twice from plasma after addition of
-tocoacetate (internal standard) by 2x2 volumes of hexane. The separation was carried out on a Vydac TP54 (250 x 4.6mm; Hesperia, CA) and a Nucleosil column (150 x 4.6mm; Interchim, Montluçon, France) in series. Elution was performed with methanol at a constant flow of 2 mL/min.
The ferric reducing ability of the plasma (FRAP) was determined using 100µL plasma samples diluted 1:2 and the tripyridyltriazine complexes formed with reduced ferrous ions were measured by spectrometry at 596 nm [25].
The levels of TBARS (Thiobarbituric Acid Reactive Species) in the urine samples were measured by a procedure modified from Lee et al. [26], reading absorbance at 532 nm. The quantity of TBARS is proportionate to the amount of MDA (Malondialdehyde), a lipid peroxidation product generated by the oxidation of membrane lipids by reactive oxygen species. MDA reacts with TBA (Thiobarbituric Acid) to form a 1:2 MDA-TBA adduct that absorbs at 532 nm. Data were normalized to urine creatinine concentrations. The results were determined as nmol/mg creatinine excreted, creatinine was measured with a kit purchased from BioMerieux (Charbonnières-les-bains, France).
MDA was determined in heart and liver homogenates by measuring the formation of TBARS upon induction of oxidation by a mixture of 2 mmol/L FeSO4 and 50 mmol/L of ascorbic acid, for 30 min at 37°C, in an oxygen-free medium [27].
Statistical Analysis
Statistical analysis was performed using the GraphPad Instat software package (GraphPad Inc., San Diego, CA, USA). Results were expressed as mean values with standard errors. All data were subjected to one-way ANOVA, followed by the Student-Newman-Keuls test to determine differences (P < 0.05) among the dietary groups.
| RESULTS |
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+53%) and to hypertrophy of the cecal wall itself (+26%). Rats fed the potato-based diet had a significantly higher SCFA cecum pool (Fig. 1), resulting in an increase in of all SCFAs (+170%) (especially of propionate, + 360%). The cecal enlargement was associated with a significant acidification of the cecal contents (the pH was lowered to 6.34 ± 0.16 in rats fed the potato-based diet, compared to 6.74 ± 0.10 and to 7.01 ± 0.03 in rats fed the control and sucrose-based diets, respectively, P < 0.05).
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Hepatic triglyceride levels were significantly affected by potato consumption (–16%, P < 0.05). There was no difference in heart cholesterol concentrations among the groups, whereas the triglyceride levels in heart tissues were significantly lower in rats fed the potato-based diet when compared with rats fed the control and sucrose diets, –20% and –24% (P < 0.05) (Table 3), respectively.
Plasma, Urine and Tissue Antioxidant Capacity
The FRAP value, which reflects the antioxidative capacity of plasma, was decreased in sucrose-fed rats, compared to control rats (–17%, P < 0.05, control vs. sucrose), whereas potato consumption tended to slightly increased the FRAP value (Table 4).
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We also observed that the potato-based diet led to a 45% and a 32% increase in vitamin E plasma concentrations when compared to the sucrose-fed and the control rats, respectively. The vitamin E/TG ratio was almost two-fold higher in rats fed the potato diet when compared to those on the control diet (Fig. 3).
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| DISCUSSION |
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Only a few studies have investigated the impact of potato consumption on rats [32] and humans [33]. However, potato, one of the most consumed vegetables in the world, is a good source of complex carbohydrates.
The aim of this work was to compare the impact of different carbohydrates (purified starch, simple carbohydrates and complex carbohydrates provided by a complex plant food containing antioxidant micronutrients) on lipid metabolism and on antioxidant status in rats. The rats were under a dietary condition allowing the development of a significant hypercholesterolemia, without inducing fatty liver (supplementation of the diet with 0.2% cholesterol), and the macronutrient supply was relatively well-equilibrated. The potatoes were eaten with skin to retain most of the tubers fiber and antioxidant micronutrients, thereby exacerbating the impact on lipid metabolism and the antioxidant status.
Our study shows that under these experimental conditions, potato consumption exerts a significant cholesterol-lowering effect on plasma and liver, compared with the control and sucrose groups. In the metabolic syndrome, the TG-RLP composition is modified, VLDLs are larger and richer in triglycerides, which could decrease their rate of catabolism. A decrease in the cholesterol content (–37%) in the potentially atherogenic lipoproteins (VLDL and LDL) was observed in the potato group. This effect can be considered beneficial in cardiovascular disease prevention and it can be attributed to the fiber fraction of the potatoes. Indeed, Ullrich et al. [34] reported similar effects with high-fiber diets, and fibers are known to affect the lipoprotein profile in cholesterol-fed rats [23]. Fibers could also exert indirect effects on the cholesterol metabolism by their fermentation in the large intestine, leading to a production of short-chain fatty acids, such as propionate, which may be involved in the control of hepatic cholesterol synthesis [35]. An increase in all SCFA levels was observed in the potato-fed rats, especially in the acetate and propionate levels. Studies on isolated hepatocytes have demonstrated that propionate could inhibit cholesterol biosynthesis from acetate [36]. Nevertheless, propionate's impact on liver cholesterol metabolism is most likely less effective than the direct effect of fiber on digestive cholesterol absorption and the indirect effect of fibers on cholesterol conversion to bile acids.
Our results are consistent with previous data [5] and indicate that consumption of sucrose may lead to an increase in tissue and plasma triglyceride concentrations. In our study, sucrose made up 30% of the diet, which is a lower level than commonly used in this type of experimental study (60–65%). Combined with the short-term intake, this could explain the non-significant increase in plasma TG concentrations in rats fed the sucrose diet, compared with the control rats. When compared with the control group, the sucrose-fed rats showed an increase in TG in TGRLP (+20%). Compared with the potato-fed group, the sucrose-fed rats showed a larger increase in TG in TGRLP (+75%). In contrast to this, we observed a hypotriglyceridemic effect in the tissue homogenates, and to a lesser extent in plasma, from rats fed the potato-based diet. Considering the potential mechanisms of fiber action in the intestine [34,37], the hypotriglyceridemic effect may be related to an altered rate of fat and glucose absorption, resulting in reduced postprandial triglyceride levels in chylomicrons and VLDL [23]. The slight decrease in plasma triglycerides in potato-fed rats was accompanied by higher
-tocopherol plasma levels. Thus, the
-tocopherol/TG ratio was improved by the potato-based diet, suggesting an increased protection of lipids from free radical attack. Previous reports have indicated that potato contains high amounts of vitamin C (about 15 mg/100g of steamed potato, contributing to 25–30% of the RDA [Recommended Dietary Allowance] [38,39]) and other antioxidant micronutrients such as vitamin E, carotenoids (primarily lutein) [14] and phenolic acids (mainly caffeic and chlorogenic acids) [15,16]. Although rats can synthesize vitamin C, it has been reported that the consumption of lettuce (containing 0.8 mg vitamin C/g DW) induced a significant increase in the plasma antioxidant capacity and the ascorbic acid plasma concentrations, 2h and 4h after tube-feeding [40]. It is well-recognized that in addition to its interactions with superoxide and hydroxyl radicals, L-ascorbic acid has the ability to regenerate the activities of lipid-soluble antioxidants, including
-tocopherol and β-carotene, in in vitro models [39]. Vitamin E is transported with plasma lipoproteins and tocopherol is secreted by the liver in VLDL particles and protects lipoproteins by preventing their oxidation. If lipoproteins are exposed to oxidative conditions, the antioxidants scavenge the radicals,
-tocopherol is typically consumed first [41]. Thus, the regeneration of vitamin E may prevent lipoproteins from subsequent oxidative stress. Previous data have suggested that vitamin E is important in rendering LDLs resistant to oxidation, although other factors have been implicated as well [42]. An increase in the TG-rich lipoprotein levels and in the lipoproteins susceptibility to peroxidation are factors that may contribute to an increased risk of developing cardiovascular diseases. Compared with the potato-based diet, sucrose consumption resulted in a higher frequency of aortic atherosclerotic plaques in animal models, consistent with previous reports [43].
In addition to the effects on lipid metabolism, we observed a significant decrease in the plasma antioxidant capacity of sucrose-fed rats, measured by FRAP assay. We investigated the defense against lipid peroxidation in urine and tissues by measurement of the TBARS concentrations. This assay, although non-specific, is widely used as an indicator of the lipid peroxidation process and indirectly as a marker for oxidative stress [44]. In urine samples, this method is subject to interference. Indeed, in comparison with plasma and tissue homogenates, urine samples contain a more complex mix of aldehyde products. Compared to the control group, the urinary excretion of TBARS in the sucrose-fed rats tended to increase, suggesting an increased production of these substances from lipid peroxidation in vivo, whereas urinary excretion of TBARS in the potato-fed rats decreased. Rats fed the potato-based diet had lower susceptibility to lipid peroxidation in heart and liver tissues than those fed the control or sucrose-based diets. Hyperlipidemia and hyperglycemia have been associated with increased oxidative damage, affecting both lipoproteins and the antioxidant status [45]. Post-prandial increases in lipid and carbohydrate levels lead to increased oxidative stress, which has been associated with an increased risk for atherosclerosis and related disorders [46]. Our results indicate that consumption of the sucrose-based diet induced an increase in triglyceride levels. However, it has been demonstrated that sucrose-fed rats are insulin resistant [47,48], and that dietary sucrose may increase the accumulation of glycated proteins, which are associated with oxidative stress [8]. On the other hand, the rats fed the potato-based diet had decreased plasma and tissue lipid concentrations, which were associated with fewer oxidation products and an increased antioxidant capacity of the plasma.
In conclusion, we have demonstrated that potatoes may have beneficial health effects. The effects can potentially be attributed to differences in carbohydrate composition between potato, purified starch and sucrose, leading to an improved lipid metabolism in potato-fed rats. Both the complex carbohydrates and the antioxidant micronutrients, provided by the potato-based diet, contributed to the improved antioxidant status in rats.
It is of interest to reduce the consumption of simple sugars, often provided by the food industry, and to favor consumption of complex carbohydrates from whole grain bread, cereals, legumes and potatoes. Different potatoes cultivars have different antioxidant micronutrient contents, which may exert divers and interesting effects on the risk factors for developing cardiovascular disease. Furthermore, potatoes are rich in potassium which contributes to the prevention of hypertension [49,50].
Further studies are required in rats and in humans to investigate the impact of potato intake in attenuation of metabolic disorders, induced by consumption of simple carbohydrates, such as dyslipidemia, insulin resistance and hypertension.
| ACKNOWLEDGMENTS |
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| FOOTNOTES |
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Received February 13, 2006. Accepted December 8, 2006.
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