Progress in Nutrition 2016; Vol. 18, N. 2: 152-160 © Mattioli 1885 Original Article Anti-obese effects of chitosan and psyllium husk containing vitamin C in Sprague-Dawley (SD) rats fed a high fat diet Eun Young Jung1†, Yang Hee Hong2†, Un Jae Chang3, Hyung Joo Suh4 1Department of Home Economics Education, Jeonju University, Jeonju 560-759, Korea; 2Department of Beauty Art, Suwon Women’s University, Suwon 441-748, Korea; 3Department of Food and Nutrition, Dongduk Women’s University, Seoul 136-714, Korea; 4Department of Public Health Science, Graduate School, Korea University, Seoul 07249, Republic of Korea - E-mail: [email protected]; †Eun Young Jung and Yang Hee Hong contributed equally to this study Summary. We investigated whether vitamin C would enhance the anti-obese effects of chitosan and psyllium husk. The study was carried out with male Sprague-Dawley (SD) rats for 6 wk using the following diet groups. Normal diet (control), high fat diet (F-control), 5% chitosan (chitosan-1), 5% chitosan containing 1% vitamin C (chitosan-2), 10% psyllium husk (psyllium-1), and 10% psyllium husk containing 1% vitamin C (psyllium-2). Chitosan-2 (1.5%) showed a significant (p<0.05) decrease in epididymal fat pad weight as compared to F- control (2.3%) and chitosan-1 (2.1%). The epididymal fat pad weight of psyllium-2 (1.9%) was significantly (p<0.05) lower than that of F-control (2.3%). In conclusion, additions of vitamin C to chitosan and psyllium husk may be useful for reducing body fat. Key words: chitosan, psyllium husk, vitamin C, body weight, body fat Introduction models have indicated the efficacy of dietary fiber in regulating body weight (3, 4). Moreover, a number of Obesity, characterized by excess accumulation of randomized studies have described the positive effects body fat, is closely related to the development of diabe- of foods rich in dietary fiber on obesity (5, 6). tes, dyslipidaemia, arteriosclerosis, hypertension, and Psyllium (Plantago psyllium) husk is a dietary fiber other metabolic diseases (1). Obesity has become an rich in soluble components (7). Psyllium husk is safe and important public health problem that demands urgent well tolerated when used as an adjunct to a low-fat diet. therapeutic attention and interventional approaches. Although by itself it produces a modest effect in lowering The essential emphasis in the management of obesity body weight, consistent results in clinical trials and meta- is to mitigate modifiable risk factors such as physi- analyses have encouraged physicians to recommend its cal inactivity and an inappropriate atherogenic diet use in combination with dietary interventions (8, 9). through lifestyle changes (2). Although the ideal di- Although chitosan is not derived from plants, it etary pattern for obese patients is yet to be ascertained, shares the characteristics of dietary fiber such as psyl- there are growing opinions among nutritionists and lium husk by being a polysaccharide that is indigestible scientists about the benefits of increased dietary in- by mammalian digestive enzymes. Chitosan is the dea- takes of fruits, legumes, vegetables, and cereals, which cetylated form of chitin, an aminopolysaccharide found are considered important sources of dietary fiber. in the exoskeleton of arthropods and certain fungi (10). Dietary fiber appears to be particularly beneficial Several studies have shown chitosan to have anti-obese with regard to its putative role in weight control. A effects in animal models by strong inhibition of fat di- large number of studies in humans and experimental gestion (11). Chitosan is dissolved in the stomach and Anti-obese effects of chitosan and psyllium husk containing vitamin C in sprague-dawley (SD) rats fed a high fat diet 153 then changes to a gelled form, entrapping fat in the in- under conditions of controlled temperature (24±1°C), testine. Kanauchi et al., (12) reported that inhibition of relative humidity (55%), and lighting (dark /light 12/12 fat digestion by chitosan was improved in the presence h) in a room with low background noise. In the prelimi- of vitamin C. nary period, they were given ad libitum access to water Therefore, the addition of vitamin C to chitosan and diet. After an adaptation period, they were divided might be more effective than chitosan alone in the in- into 6 groups (8 rats/group): normal diet (control), high hibition of fat digestion. We hypothesized that vita- fat diet (F-control), high fat diet with 5% chitosan (chi- min C might also improve the hypolipidemic effect of tosan-1), high fat diet with 5% chitosan containing 1% psyllium husk, which has similar activities as chitosan. vitamin C (chitosan-2), high fat diet with 10% psyllium Based on these considerations, we investigated whether husk (psyllium-1), high fat diet with 10% psyllium husk vitamin C enhanced the anti-obese effects of chitosan containing 1% vitamin C (psyllium-2) (Table 1). and psyllium husk on body weight gain and fat loss in male Sprague-Dawley (SD) rats fed a high fat diet. We Measurements of body weight, food intake, organ weights, also determined the antioxidant activities of chitosan plasma lipids, and fat pads and psyllium husk mixtures containing vitamin C, a free radical scavenger with strong antioxidant properties. Food intake and body weight were monitored every 2 d for 6 wk. The food was removed for the last 12 h of the experimental term, and the rats were an- Material and methods esthetized with ethyl ether, sacrificed, and dissected. Blood was collected from the heart with a heparin- Animals and diets ized syringe. Plasma was separated by centrifugation at 3,000×g for 15 min at 4 °C and then stored at -70 °C The experimental protocol was reviewed and ap- until analysis. The levels of triacylglycerol, total choles- proved by the Korea University Animal Care Com- terol, and HDL-cholesterol in plasma were measured mittee. The study was carried out with male Sprague- using a FUJI DRI-CHEM 3500 (Fuji Photo Film Dawley rats aged 6 wk (Central Lab Animal Inc., Seoul, Co., Tokyo, Japan). LDL-cholesterol was estimated Korea) for 6 wk. The rats were acclimatized for 5 days using the method validated by Friedewald et al., (13). Table 1. Composition of the experimental diets for rats (g/100 g diet) Ingredient Control F-control Chitosan-1 Chitosan-2 Psyllium-1 Psyllium-2 Casein 19.6 26.2 26.2 26.2 26.2 26.2 Corn starch 56.6 27.6 27.6 27.6 22.6 22.6 Cellulose 5 5 0 0 0 5 Sucrose 9.95 9.95 9.95 9 9.95 9 Lard 0 22.4 22.4 22.4 22.4 22.4 Soybean oil 4 4 4 4 4 4 Mineral mixturea 3.5 3.5 3.5 3.5 3.5 3.5 Vitamin mixtureb 1 1 1 1 1 1 L-methionine 0.3 0.3 0.3 0.3 0.3 0.3 Chitosan 0 0 5 5 0 0 Pyllium husk 0 0 0 0 10 10 Vitamin C 0.05 0.05 0.05 1 0.05 1 a 4 Mineral mixture(g/kg diet): CaPO4·2H2O,145.6; KH2PO4, 257.2; NaH2PO , 93.5; NaCl, 46.6; calcium lactate, 350.9; ferric citrate, 31.8; b MgSO4, 71.7; ZnCO3, 1.1; MnSO4·4H2O, 1.2; CuSO4·5H2O, 0.3; KI, 0.1. Vitamin mixture: ICN Vitamin mixture (No 904654, 1999). 154 E.Y. Jung, Y.H. Hong, U.J. Chang, H.J. Suh The liver, kidney, spleen, ileum, jejunum, duode- ugation at 2,000×g for 10 min at 4°C. GSH in the su- num, colon, cecum, epididymal fat, and perirenal fat pernatant was assayed as follows: 100 μL of plasma or were removed and the wet weights were measured. The liver tissue, 800 μL of 0.3 mmol Na2HPO4 and 100 μL liver was washed in 0.15 mol NaCl solution, blotted of 0.04% 5,5’-dithiobis 2-nitrobenzoic acid (DTNB) onto filter paper, weighed, and homogenized in 4.5 mL in 0.1% sodium citrate. The absorbance of DTNB was potassium phosphate buffer (pH 7.0). The homogen- monitored at 412 nm for 5 min. A standard curve of ates were centrifuged at 13,000×g for 4 min at 4°C and GSH was prepared and the sensitivity of measure- the supernatant was stored at -70°C until analysis. ments was determined to be between 1 and 100 μL. 2,2-Azinobis(3-ethylbenzthiazoline-6-sulfonate) Statistical analysis (ABTS) radical cation scavenging activity All statistical analyses were performed using the The scavenging activity on ABTS radical cation Statistical Package for Social Sciences (SPSS) ver. was measured according to the method of Re et al., 12.0 (IL, USA). The differences among groups were (14) with some modifications. ABTS radical cation was evaluated statistically by one-way analysis of variance generated by adding 7 mmol ABTS to 2.45 mmol po- (ANOVA) and Duncan’s multiple tests. All data were tassium persulfate solution and the mixture was left to two-sided with a 5% significance level and were re- stand overnight in the dark at room temperature. The ported as means±standard deviations (SD). ABTS radical cation solution was diluted with distilled water to obtain an absorbance of 1.4-1.5 at 414 nm. The diluted ABTS radical cation solution (1 mL) was Results and discussion added to 50 μL of plasma or liver tissue. After 90 min, the absorbance was measured at 414 nm. ABTS radical Body weight gain, food intake and food efficiency ratio cation scavenging activity was expressed as ascorbic acid equivalent antioxidant activity (AEAC), and defined as Body weight gain, food intake and food efficiency mg of ascorbic acid equivalents per 100 g of sample (15).