AOKI Takafumi

KREMENIK Michael J

MIURA Suzuka

OYANAGI Eri

TERAMOTO Fusako

WATANABE Chihiro

YANO Hiromi

YOKOGAWA Yuka

Kawasaki University of Medical Welfare Institutional Repository

175Kawasaki Journal of Medical Welfare Vol. 26,  No. 2,  2021 175-180＊1 Graduate School of Health Science and Technology, Kawasaki University of Medical Welfare＊2 Department of Health and Sports Science, Faculty of Health Science and Technology,     Kawasaki University of Medical Welfare, Kurashiki, 701-0193, Japan    E-Mail: yanohiro@mw.kawasaki-m.ac.jp＊3 Department of Clinical Nutrition, Faculty of Health Science and Technology, Kawasaki University of Medical WelfareEffect of Partially Hydrolyzed Guar-gum Treatment on Fecal pH in Mice Fed with a High-fat-dietHiromi YANO＊1,2, Takafumi AOKI＊1,3, Yuka YOKOGAWA＊1,Chihiro WATANABE＊1, Suzuka MIURA＊1, Fusako TERAMOTO＊1,3,Michael J. KREMENIK＊2 and Eri OYANAGI＊2(Accepted November 16, 2020)Key words: dietary fiber, PHGG, obesity, glucose tolerance, gut microbiotaAbstract　To clarify the supplemental effect of partially hydrolyzed guar-gum (PHGG), which is a soluble-dietary fiber, on glucose metabolism and fecal pH in mice fed with a high-fat-diet (HFD), 4-week-old male C57BL/6J mice (n=24) were randomly divided into 4 groups: HFD-0%PHGG (HFD), HFD-1.0%PHGG (G1HFD), HFD-5.0%PHGG (G5HFD) and HFD-7.5%PHGG (G7.5HFD). After the experimental period of 10 weeks, a glucose tolerance test and measurement of fecal pH were carried out. PHGG treatment under HFD feeding did not show improvement in obesity measurements or glucose tolerance. However, both G5HFD and G7.5HFD groups accelerated the decrease of fecal pH (p<0.01). These findings suggest that a study using an over 5% PHGG treatment might activate the change in fermentation level of gut microbiota in HFD mice, although PHGG-treatment of less than 10 weeks may not contribute enough to prohibit HFD-induced obesity and diabetes.1. Introduction　It is known that the ingestion of dietary fiber may lead to numerous health benefits for the host1). Guar-gum is one of these dietary fibers, and is classified among soluble fibers. In order to make guar-gum more palatable, hydrolyzed guar-gum (partially hydrolyzed guar-gum: PHGG) was produced resulting in a low viscosity/nonviscous type of fiber. Although PHGG is well fermented by gut microbiota, PHGG intake might not contribute to the improvement of glycemic control2). In fact, Aoki et al.3) showed that there was no difference between the blood glucose area under the curve in HFD and PHGG+HFDC mice when the glucose tolerance test was performed. However, that study used a 5% contained PHGG diet, so a study using a different dose effect of PHGG intake and its reults on glucose metabolism and obesity remains unclear. Furthermore, it is unclear whether the PHGG administered dose-dependently induces the fermentative action via gut microbiota. In this study, we investigated the supplemental effect of PHGG on HFD fed-induced obesity, glucose tolerance and fecal pH via gut microbiota.Short Report176 Hiromi Yano et al.2. Methods2.1 Animals　In this study, 4-week-old male C57BL/6J mice (n=24) were housed 4 mice to a cage under a controlled environment (22 ± 1 ◦C, 12:12-h light-dark cycle) and were randomly divided into 4 groups: HFD, G1HFD, G5HFD and G7.5HFD. The mice were given access to food and drinking water ad libitum. The experiment involving the mice, including the procedures performed, was approved by the Institutional Animal Care and Use Committee of Kawasaki University of Medical Welfare (No.18-017).2.2 Diet 　The mice were fed HFD (D12492Gpx10, Research Diets, New Brunswick, NJ) containing 60 % fat, 20% protein and 20% carbohydrates for 10 weeks. The diet was cellulose free and was administered under three conditions: 1.0% (G1), 5.0% (G5) and 7.5 % (G7.5) PHGG (Sun fiber®, Taiyo Kagaku Co., Ltd, Yokkaichi, Japan) (Table 1). Table 1　Dietary compositionContents HFD G1HFD G5HFD G7.5HFDkcal kcal kcal kcalProtein 20 20 20 19Carbohydrate 20 20 22 23Fat 60 60 59 58Total 100 100 100 100kcal/g 5.6 5.6 5.4 5.3g % g % g % g %Protein 28.0 27.7 26.4 25.6  Casein 27.6 27.3 26.0 25.2  L-Cystine 0.4 0.4 0.4 0.4Carbohydrate 32.2 33.0 36.2 38.2  Corn Starch 0.0 0.0 0.0 0.0  Maltodextrin 17.3 17.1 16.3 15.7  Sucrose 14.9 14.7 14.0 13.6  Cellulose 0.0 0.0 0.0 0.0  PHGG 0.0 1.0 5.0 7.5Fat 37.3 36.9 35.1 34.0  Soybean Oil 3.5 3.4 3.3 3.1  Lard 33.8 33.4 31.9 30.92.3 Glucose tolerance test　After the mice fasted for 5 hours blood samples were collected from the tail vein, and then blood glucose levels were measured using the glucose monitoring device Accu-Chek (Roche, Basel, Schweiz) immediately prior to glucose administration, and at 15, 30, 60 and 120 minutes after glucose administration (2 g/kg, i.p.). Each mouse was lightly anesthetized with the inhalant Isoflurane prior to the glucose administration4).2.4 Fecal pH measurements　At the 6th and 10th week, fecal materials were collected from mice that were as fresh as possible. The materials were diluted 2-3 folds (w/v) in distilled water and were homogenized by the homogenizer pestle. After calibrating the pH meter (twin pH B-212, Horiba Ltd, Kyoto, Japan), the pH of these diluted samples 177Alteration of Fecal pH by PHGGwas measured5).2.5 Measurements of body mass, food intake, cecum contents and epididymal adipose tissue　Body weight and food intake were recorded for the final week. The food intake was expressed as total weekly intake. Two days after the experimental period the mice were euthanized under Isoflurane anesthesia and then the cecum contents and epididymal adipose tissue were collected and weighed.2.6 Statistical analysis　The statistical analyses were performed using the IBM SPSS Statistics 23.0 for Windows software program. The data were analyzed using the one-way ANOVA, and then a post-hoc test was performed using the Bonferroni’s test. P values of <0.05 were considered to be indicators of statistical significance.3. Results and discussion 3.1 Effect of PHGG on obesity and glucose metabolism　Although body weight was different in the G5HFD and G7.5HFD groups (Figure 1a, p<0.05), the G1HFD, G5HFD and G7.5HFD groups did not show any attenuation in body mass gain compared with HFD mice. Since food intake did not differentiate among all groups (Figure 1b), it was suggested that PHGG was ingested in mice in a dose-dependent manner. Nevertheless, we did not observe the inhibition of weight gain by PHGG. Furthermore, adipose tissue mass in HFD mice also was not affected by PHGG intake (Figure 1c). Hence, at the least, our results suggest a small obesity-preventing effect of PHGG. Recently, it was reported that combined with 3% cellulose, 7% guar gum intake did not attenuate body mass gain and adipose tissue Figure 1　 The effect of PHGG treatment on body weight (a), food intake (b), adipose tissue weight (c) and cecum contents (d) in HFD mice. Food intake showed as energy intake (kcal) per body weight at final week. The values were expressed as the mean ± S.E.M. *: p<0.05 and **: p < 0.01.178 Hiromi Yano et al.mass in HFD mice6). It is the considered opinion of this research team that this report contains similar results to our own. On the other hand, cecum contents were increased in a PHGG dose-dependent manner. However, no difference between the cecum content in G5HFD and G7.5HFD groups was observed (Figure 1d). Indigestible carbohydrate supplementation increases cecum content3,7). This alteration is caused by indigestibility with increased water-holding capacity. In addition, it was reported that a significant and dose-dependent increase in cecum weight was observed by galactooligosaccharides treatment, and that this was accompanied by an alteration of gut microbiota7). Moreover, polyphenol, n-3 polyunsaturated fatty acids and HFD which influences gut microbiota also affects cecum content weight, although they do not have water-holding capacity8,9). Therefore, changes in cecum content weight is one of the beneficial indexes of gut microbiota alteration. This result suggests that starting at 5% PHGG treatment might activate the change in fermentation levels of gut microbiota in HFD mice.　Additionally, G1HFD, G5HFD and G7.5HFD groups did not show improvement in glucose tolerance compared with HFD mice (Figures 2a and b). It was already known that any beneficial effect, such as improved glycemic control, is abolished when guar-gum is hydrolyzed to a nonviscous form2). In fact, dietary PHGG did not significantly affect plasma glucose levels in a db/db mice10) or in one human study11). Therefore, our results suggest that glucose metabolism dose not improve PHGG intake regardless of the intake dose. Interestingly, Aoki et al.3) recommends that exercise has a complementary effect on insulin resistance in PHGG intake.Figure 2　 The effect of PHGG treatment on blood glucose concentration and the incremental area under the curve (AUC) of blood glucose during glucose tolerance tests in HFD mice. Time dependent blood glucose levels after glucose load (2 g/kg, i.p.) (a), and the corresponding incremental blood glucose area under the curves (BGAUC) (b). The values were expressed as the mean ± S.E.M.3.2 Effect of PHGG on fecal pH　In both the 6th and 10th weeks of the experimental period, the reduction of fecal pH depended on the dose of PHGG administration. However, there was no significant difference between the pH levels in G5 and G7.5 groups in both the 6th and 10th weeks (Figure 3). PHGG could elicit constipation relief and modulate gut microbiota, which shows the potential to act as a dietary fiber for constipation treatment12). PHGG treatment has been shown to reduce colonic mucosal damage in an animal colitis model13). This fermentable fiber is useful as a prebiotic, and it was reported as such in HFD-induced obese mice. At the least, we confirmed that cecum content weights were higher and the fecal pH was lower in mice fed with a HFD diet with PHGG than in those fed diets without PHGG. There was agreement with the results of previous studies, which suggested that the possible mediators of the effects of PHGG were the SCFAs produced by microbial 179Alteration of Fecal pH by PHGGFigure 3　 The effect of PHGG treatment on fecal pH in HFD mice. White column: HFD, plaid column: G1HFD, checkered column: G5HFD, and black column: G7.5HFD. The values were expressed as the mean ± S.E.M. *: p<0.05 and **: p < 0.01.fermentation of PHGG in the large intestine3,14). In addition, those effects were shown to occur at sufficient levels enough with the administration of 5% PHGG. It seems that this is the adequate concentration that should be taken in in vivo for PHGG research in the future. 　In conclusion, our findings suggest that over 5% PHGG supplementation might activate the fermentation level via gut microbiota in HFD mice, although PHGG alone may not contribute to improvement of HFD-induced diabetes.Acknowledgements　We are thankful that this study has been supported by two funding arms, the Interdepartmental Research Fund of Kawasaki University of Medical Welfare (to H.Y.) and Grant-in-Aids for Scientific Research (B-19H04213) (to H.Y.).COI　The authors declare no conflicts of interest are associated with the present study.References1. Slavin J : Fiber and prebiotics: Mechanisms and health benefits. Nutrients, 5, 1417-1435, 2013, https://doi.org/10.3390/nu5041417.2. McRorie JW Jr and McKeown NM : Understanding the physics of functional fibers in the gastrointestinal tract: An evidence-based approach to resolving enduring misconceptions about insoluble and soluble fiber. The Journal of the Academy of Nutrition and Dietetics, 117, 251-264, 2017.3. Aoki T, Oyanagi E, Watanabe C, Kobiki N, Miura S, Yokogawa Y, Kitamura H, Teramoto F, Kremenik MJ and Yano H : The effect of voluntary exercise on gut microbiota in partially hydrolyzed guar gum intake mice under high-fat diet feeding. Nutrients, 12, E2508, 2020, https://doi.org/10.3390/nu12092508.4. Bradley RL, Jeon JY, Liu FF and Maratos-Flier E : Voluntary exercise improves insulin sensitivity and adipose tissue inflammation in diet-induced obese mice. American Journal of Physiology Endocrinology and Metabolism, 295, E586-E594, 2008, https://doi.org/10.1152/ajpendo.00309.2007.5. 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Effect of Partially Hydrolyzed Guar-gum Treatment on Fecal pH in Mice Fed with a High-fat-diet

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Effect of Partially Hydrolyzed Guar-gum Treatment on Fecal pH in Mice Fed with a High-fat-diet

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