天然產(chǎn)物化學(xué)英文版副本匯總

1、Myoung Jin Son, Catherine W. Rico, Seok Hyun Nam, and Mi Young KangAbstract: The effects of oryzanol and ferulic acid on the glucose metabolism of high-fat-fed mice were investigated. Male C57BL/6N mice were randomly divided into 4 groups: NC group fed with normal control diet; HF group fed with hig

2、h-fat (17%) diet; HF-O group fed with high-fat diet supplemented with 0.5% oryzanol; and HF-FA group fed with high-fat diet supplemented with 0.5% ferulic acid. All animals were allowed free access to the experimental diets and water for 7 wk. At the end of the experimental period, the HF-O and HF-F

3、A groups exhibited significantly lower blood glucose level and glucose-6-phosphatase (G6pase) and phosphoenolpyruvate carboxykinase (PEPCK) activities, and higher glycogen and insulin concentrations and glucokinase (GK) activity compared with NC and HF groups. The results of this study illustrate th

4、at both oryzanol and ferulic acid could reduce the risk of high-fat diet-induced hyperglycemia via regulation of insulin secretion and hepatic glucose-regulating enzyme activities.Keywords: diabetes, ferulic acid, high-fat-fed mice, hypoglycemic effect, oryzanolIntroductionChronic consumption of a h

5、igh-fat diet has been associated with the development of obesity and type 2 diabetes mellitus (Hill and others 1992; Bray and others 2004). Scientific studies have shown that excessive intake of dietary fat results in increased body weight and poor glucose regulation (Alsaif and Duwaihy2004; Petro a

6、nd others 2004; Messier and others 2007). Diabetes is characterized by hyperglycemia that results in the generation of free radicals leading to oxidative stress (West 2000). Due to changes in lifestyle patterns, particularly poor eating habit and sedentary lifestyle, the incidence of diabetes has ra

7、pidly increased in epidemic proportions. Around 171 million cases of diabetes worldwide were reported in 2001 and it was projected that by 2030, 366 million people will have diabetes (Wild and others 2004). With this increasing global prevalence of diabetes, the need for therapeutic measures against

8、 the disease has become stronger and more urgent. A wide range of oral medicines are currently being used for treating diabetes. However, various adverse effects and high rates of secondary failures have been associated with the available antidiabetic medicines (Inzucchi 2002). Thus, finding natural

9、 drugs with hypoglycemic activity has now become the focus of scientists and researchers. At present, there is a considerable public and scientific interest in utilizing phytochemicals for the treatment and prevention of various diseases. Naturally occurring phenolic compounds, such as oryzanol and

10、ferulic acid, are known to have strong antioxidant activities (Wang and others 2002; Srinivasan and others 2007). Oryzanol is a mixture of ferulic acid (4-hydroxy-3-methoxycinnamic acid) esters with phytosterols (Lerma-Garcia and others 2009) and primarily extracted from rice bran. Ferulic acid is c

11、ommonly found in fruits and vegetables, including banana, broccoli, rice bran, and citrus fruits (Zhao and Moghadasian 2008). Both oryzanol and ferulic acid possess several physiological proper ties, such as reduction of serum cholesterol levels (Wilson and others 2007), inhibition of tumor promotio

12、n (Yasukawa and others 1998), and protective action against liver injury (Choti-markorn and Ushio 2008). Oxidative stress is regarded as the key factor in the development of diabetes and its associated health disorders. The high-fat diet fed C 57BL/6 mouse model has long been used by researchers in

13、investigating the pathophysiology of impaired glucose tolerance and type 2 diabetes and for the development of new treatments (Surwit and others 1988; Surwit and other s 1991; Schreyer and others 1998; Winzell and Ahren 2004). Since diabetes is a free radical mediated disease, the strong antioxidant

14、 activity of oryzanol and ferulic acid may be useful in preventing the development of diabetic hyperglycemia under a high-fat diet. There are limited reports on the physiological functions of these phenolic compounds in relation to glucose metabolism in animal models. Thus, this study was conducted

15、to investigate the effects of dietary feeding of oryzanol and ferulic acid on the glucose metabolism in high-fat-fed C57BL/6 mice.1. Materials and Methods1.1 Animals and dietsTwenty-four male C57BL/6N mice of 4 wk of age, weighing 12 g, were obtained from Orient Inc. (Seoul, Korea). They were indivi

16、dually housed in stainless steel cages in a room maintained at 25C with 50% relative humidity and 12/12 h light/dark cycle and fed with a pelletized chow diet for 2 wk after arrival. The mice were then randomly divided into 4 dietary groups (n = 6). The 1st and 2nd groups were fed with a normal and

17、high-fat (17%, w/w) diets, respectively, while the other 2 groups were fed with high-fat diet supplemented with either 0.5% oryzanol or 0.5% ferulic acid (98% pure, Tsuno, Osaka, Japan). The composition of the experimental diet (Table 1) was based on the AIN-76 semisynthetic diet. The mice were fed

18、for 7 wk and allowed free access to food and water during the experimental period. The body weight gain was measured weekly. At the end of the experimental period, the mice were anaesthetized with 60-L Ketamine-HCl following a 12 h fast and sacrificed. Blood samples were collected and centrifuged at

19、 1000 g for 15 min at 4C to obtain the plasma. The livers were removed, rinsed with physiological saline, and stored at 70C until analysis. The current study protocol was approved by the Ethics Committee of Kyungpook Natl. Univ. for anima studies.1.2 Measurement of blood glucose levelThe blood gluco

20、se level in mice was measured using Accu-Chek Active Blood Glucose Test Strips (Roche Diagnostics GmbH, Germany). Blood samples were drawn from the tail vein of the mice before and after 3 and 7 wk of feeding the animals with experimental diets.1.3 Determination of glycogen and insulin levelsThe gly

21、cogen concentration in liver was determined using the method described by Seifter and others (1950)。 Fresh liver (100 mg) was mixed with 30% KOH and heated at 100C for 30 min. The mixture was then added with 1.5 mL ethanol (95%) and kept over night at 4C. The pellet was mixed with 4 mL distilled wat

22、er. A 500 L of the mixture was added with 0.2% anthrone (in 95% H2SO4) and the absorbance of the sample solution was measured at 620 nm. The results were calculated on the basis of a standard calibration curve of glucose. The insulin content was measured using enzyme-linked immunosorbent assay (ELIS

23、A) kits (TMB Mouse Insulin ELISA kit, Sibayagi, Japan).1.4 Measurement of hepatic glucose-regulating enzyme activitiesThe hepatic enzyme source was prepared according to the method developed by Hulcher and Oleson (1973). The glucokinase (GK) activity was determined based from the method of Davidson

24、and Arion (1987) with slight modification.A 0.98 mL of the reaction mixture containing 50 mM Hepes-NaOH(pH 7.4), 100 mM KCl, 7.5 m M MgCl2, 2.5 mM dithioerythritol, 10 mg/mL albumin, 10 mM glucose, 4 units of glucose-6-phosphate ( G6pase) dehydrogenase, 50 mM NAD+, and 10 L cytosol was preincubated

25、at 37C for 10 min. The reaction was initiated with the addition of 10 L of 5 mM ATP and the mixture was incubated at 37C for 10 min. The G6pase activity was measured using the method described by Alegre and others (1988). The reaction mixture contained 765 L of 131.58 mM Hepes-NaOH (pH 6.5), 100 L o

26、f 18 mM EDTA ( pH 6.5), 100 L of 265 mM G6pase, 10 L of 0.2 M NADP+, 0.6 IU/mL mutarotase, and 0.6 IU/mL glucose dehydrogenase. the mixture was added with 5 L microsome and incubated at 37C for 4 min. The change in absorbance at 340 nm was measured. The phosphoenolpyruvate carboxykinase (PEPCK) acti

27、vity was determined based from the method developed by Bentle and Lardy (1976).The reaction mixture consisted of 72.92 mM sodium Hepes (pH 7.0), 10 mM dithiothreitol, 500 mM NaHCO3, 10 mM MnCl2, 25 mM NADH, 100 mM IDP, 200 mM PEP, 7.2 unit of malic dehydrogenase, and 10L cytosol. The enzyme activity

28、 was determined based from the decrease in the absorbance of the mixture at 350 nm at 25C 350nm。1.5 Statistical analysisAll data are presented as the mean SE.The data were evaluated by 1-way ANOVA using a Statistical Package for Social Sciences software program (SPSS Inc., Chicago, Ill., U.S.A.) and

29、 the differences between the means we reassessed using Duncans multiple range test. Statistical significance was considered at P 98%的純,參會(huì)者,大阪,日本)。實(shí)驗(yàn)飲食的組成(表1)是基于ain - 76半合成的飲食。7周的老鼠,允許自由獲取食物和水在實(shí)驗(yàn)期間。每周身體體重測量。在實(shí)驗(yàn)周期結(jié)束后,小鼠麻醉與60-L Ketamine-HCl 12 h后快速和犧牲。血液樣本收集和離心機(jī)在1000 g15分鐘4C獲得等離子體。肝臟被移除,用生理鹽水沖洗,并存儲(chǔ)在70C

30、直到分析。當(dāng)前的研究協(xié)議是變異性慶北的經(jīng)倫理委員會(huì)批準(zhǔn)。大學(xué)生命研究。小鼠的血糖水平是衡量使用Accu-Chek活躍的血糖測試條(德國羅氏診斷GmbH)。血液樣本取自小鼠尾靜脈的前后3和7周的飲食喂養(yǎng)的動(dòng)物實(shí)驗(yàn)。肝臟的糖原含量是決定使用Seifter描述的方法和其他人(1950)。新鮮肝臟(100毫克)與30% KOH和混合加熱30分鐘的100C。然后添加1.5毫升乙醇混合物(95%)和保持在晚上4C。顆粒與4毫升蒸餾水混合。500L混合物添加0.2%蒽酮硫酸(95%)和樣品溶液的吸光度測量在620海里。計(jì)算結(jié)果的基礎(chǔ)上,一個(gè)標(biāo)準(zhǔn)的校準(zhǔn)曲線的葡萄糖。胰島素含量測定采用酶聯(lián)免疫吸附試驗(yàn)(ELIS

31、A)試劑盒(三甲小鼠胰島素酶聯(lián)免疫試劑盒,Sibayagi,日本)。肝酶源制備根據(jù)方法由Hulcher和奧爾森(1973)。葡糖激酶(門將)活動(dòng)確定方法的基礎(chǔ)戴維森和Arion(1987)與輕微的修改。0.98毫升的反應(yīng)混合物50 mM Hepes-NaOH(pH值7.4),100毫米氯化鉀,MgCl2 7.5米,2.5毫米dithioerythritol,10毫克/毫升白蛋白,10毫米葡萄糖,4單位glucose-6-phosphate(G6pase)脫氫酶,NAD + 50毫米,和10L細(xì)胞溶質(zhì)在37 preincubatedC 10分鐘。反應(yīng)開始的10L 5毫米ATP和混合物在37孵化C

32、 10分鐘。G6pase活動(dòng)被全部用描述的方法測量等人(1988)。131.58毫米的反應(yīng)混合物含有765L Hepes-NaOH(pH值6.5),100L 18毫米EDTA(pH值6.5),100L 265毫米G6pase 10L輔酶ii + 0.2米,0.6國際單位/毫升mutarotase,0.6國際單位/毫升葡萄糖脫氫酶?；旌咸砑?L微粒體和孵化37C 4分鐘。在340納米測量吸光度的變化。磷酸烯醇丙酮酸的carboxykinase(PEPCK)活動(dòng)是基于確定的方法由Bentle和拉迪(1976)。72.92毫米鈉的反應(yīng)混合物由玫瑰(pH值7.0),10毫米二硫蘇糖醇,500毫米NaH

33、CO3,10毫米MnCl2,25毫米NADH IDP 100毫米,200毫米PEP,7.2單位的蘋果酸脫氫酶,和10l胞質(zhì)?；诿富钚詼y定的吸光度下降的混合物在350 nm 25C。在25C 350海里。所有數(shù)據(jù)提出了均值SE。方法進(jìn)行評估的數(shù)據(jù)方差分析使用社會(huì)科學(xué)統(tǒng)計(jì)軟件包軟件程序(SPSS Inc .)、芝加哥、生病。、美國)和之間的差異意味著我們重新使用鄧肯的多個(gè)測試范圍。統(tǒng)計(jì)顯著性被認(rèn)為是在P 0.05。2結(jié)果沒有顯著差異之前在動(dòng)物體重組與實(shí)驗(yàn)飲食喂養(yǎng)的老鼠(表2)。老鼠是恒定的日常食物攝取(3 g / d)在整個(gè)研究。實(shí)驗(yàn)周期的結(jié)束,然而,顯著增加在動(dòng)物喂食高脂肪的飲食(高頻組)相對

34、于控制的老鼠(NC組)。當(dāng)老鼠與高脂肪飲食補(bǔ)充谷維素(HF-O組)或阿魏酸(HF-FA組)也顯示更高的體重增加與NC組相比,他們的身體體重明顯低于高頻組。HF-O和HF-FA團(tuán)體之間,后者表現(xiàn)出較低的最終體重。之前最初的小鼠的血糖水平與實(shí)驗(yàn)飲食喂養(yǎng)各組之間沒有顯著差異(圖1)然而,高脂肪喂養(yǎng)導(dǎo)致顯著增加3周后小鼠的血糖水平。數(shù)控組還顯示葡萄糖含量的增加類似于高頻老鼠。最后工作,HF-O HF-FA老鼠表現(xiàn)出大幅降低葡萄糖水平較高頻和對照組。糖原和胰島素濃度遠(yuǎn)遠(yuǎn)高于HF-O和HF-FA老鼠比控制和高頻的(表3)。2.4肝glucose-regulating酶的活動(dòng)肝GK酶活性明顯高于在白鼠谷維素和阿魏酸比的控制和HF-fed(圖2)。高脂喂養(yǎng)G6pase活動(dòng)導(dǎo)致顯著增加(圖3)。然而,谷維素和阿魏酸的補(bǔ)充飲食抑制高度和規(guī)范化G6pase活動(dòng)。同樣,顯著降低PEPCK活動(dòng)觀察HF-O和HF-FA老鼠相比,數(shù)控和HFones(裝具)。3討論谷維素和阿魏酸的強(qiáng)抗氧化活動(dòng)已經(jīng)有據(jù)可查(王等2002;Srinivasan等2002)。自認(rèn)為氧化應(yīng)激是糖尿病發(fā)展的一個(gè)關(guān)鍵因素,谷維素和阿魏酸的影響在葡萄糖代謝high-fat-fed老鼠了。小鼠的體重顯著降低美聯(lián)儲(chǔ)谷維素和阿魏酸的相對于hig