6 9 Vol. 6 No. 9 2015 9 Journal of Food Safety and Quality Sep., 2015 1, 陈 金 华 2 1,, 谭 斌 2, 黄 建 安 1*, 刘 仲 华 (1., 410128; 2., 410128) 1, 2 摘 要 : 目 的 方 法, (black tea, BT), (malondialdehyde, MDA) (superoxide dismutase, SOD) (glutathione peroxidase, GSH-Px) (alanine transaminase, ALT) (aspartate transaminase, AST), 结 果, ALT AST MDA, SOD GSH-Px, 结 论, 关 键 词 : ; ; ; Protective effects of black tea on mice liver under high fat diet CHEN Jin-Hua 1, 2, TAN Bin 1, 2, HUANG Jian-An 1*, LIU Zhong-Hua 1, 2 (1. Key Lab of Tea Science of Ministry of Education, Hunan Agricultural University, Changsha 410128, China; 2. National Research Center of Engineering & Technology for Utilization of Functional Ingredients from Botanicals, Hunan Agricultural University, Changsha 410128, China) ABSTRACT: Objective To study the protective effect of black tea (BT) on mice liver under a high fat diet. Methods The mice were fed with BT of different types and quantities under the condition of high fat diet. Liver index and content of liver malondialdehyde (MDA), liver superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), serum glutamic-pyruvic transaminase (ALT) and glutamic-oxalacetic transaminase (AST) were measured, and the histopathological changes in liver tissues were observed with unaided eyes and under an optical microscope as well. Results The content of serum ALT, AST, liver MDA and liver index in the black tea feeding group were lower than the high hyperlipoidemia model group, while liver SOD activity and GSH-Px activity were higher, and the liver lesions were reduced. Conclusion It indicates that BT has positive effect on protecting and repairing mice liver with high fat diet. The mechanisms may be that black tea has effects on ameliorating oxidative stress and reducing lipid peroxidation product. KEY WORDS: black tea; high fat diet; mice; liver 基 金 项 目 : (CARS-23-C) (2013FJ4222) (13C421) Fund: Supported by the National Tea Industry Technology System Project (CARS-23-C), the Provincial Science and Technology Projects (2013FJ4222) and the Scientific Research Fund of Hunan Provincial Education Department (13C421) * 通 讯 作 者 :,, E-mail: jian 7513@sina.com *Corresponding author: HUANG Jian-An, Professor, Key Lab of Tea Science of Ministry of Education, Hunan Agricultural University, Changsha 410128, China. E-mail: jian 7513@sina.com
3638 6 1 引 言,,,,,,,,,,,, [1],,,,, [2-8],,, [9-15] (superoxide dismutase, SOD) (glutathione peroxidase, GSH-Px),, ; (malondialdehyde, MDA),, (alanine transaminase, ALT) (aspartate transaminase, AST),,,,,,, ;,,, MDA SOD GSH-Px ALT AST,, 2 材 料 与 方 法 2.1 材 料 与 设 备 2.1.1 主 要 仪 器 (TBA-40FR, ); (EG 1150C, Leica ); (TP1020, Leica ); (RM2235, Leica ); (DM2000, Leica ); (MIKRO 22R, Hettich ); (Modulyod-230, ); (BUCHI R-200, BUCHI ); (F6/10, FLUKO); (Thermo Scientific Varioskan Flash, Thermo ); (Eppdrof) 2.1.2 主 要 试 剂 ( ALT) (AST), ; (MDA) (SOD) (GSH-Px), 2.1.3 实 验 动 物 及 饲 养 条 件 SPF ICR, 30 g, 5,, : SCXK( )2011-0003 : 20~25, 45%~55%,,,, 1 d 1, 2 1 2.1.4 供 试 材 料 (2011, ), (2011, ), (2011, ), (2011, ), (2012, ), (2011, ), (2011, ) 2.1.5 动 物 饲 料, : SCXK( )2009-0004, : 24.3% 7.5% 8.6% 3.5% 0.7% 1.16% 7.5% [16] :, 78.8%;, 1%;, 10%;, 10%;, 0.2%
9, : 3639 2.1.6 阳 性 药 : (, ) 2.2 实 验 方 法 2.2.1 红 茶 粉 的 制 备, 1:10(m:V) (100 ), 30 min, ; 1:5 20 min, 10 min,,,, -20 12 h 24 h,,, -20 2.2.2 动 物 实 验 (1) ( 60 kg 10 g, 166.7 mg kg -1 d -1 ) 5 10( ) 20 [17] (60 kg) 10 mg kg -1 d -1, 9.1 [18], 91 mg kg -1 d -1 0.1 ml/10 g bw [19] (2) 3 d, 7 ( ), 24 10,,,,, 1, 35 d [17,19] 2.3 指 标 测 定 2.3.1 血 清 ALT AST 的 测 定 12 h( ),, 4 3000 r/min 10 min,, ALT AST 2.3.2 肝 匀 浆 制 备 及 MDA SOD GSH-PX 测 定,,,, 0.2 g,, 10%, 4 3000 r/min 10 min, -20 MDA SOD GSH-PX 2.3.3 小 鼠 肝 脏 的 组 织 病 理 切 片 制 备 及 观 察 0.5 g, 10%, ; HE 2.4 数 据 处 理 Excel 2010 SPSS 19.0, ± ( x ±s), (least significant difference, LSD), P 0.05 3 结 果 与 分 析 3.1 红 茶 对 小 鼠 肝 体 比 的 影 响 1,,,,,, (P 0.01),, 3.2 红 茶 对 小 鼠 MDA SOD 和 GSH-Px 的 影 响 SOD GSH-Px MDA 2 3 4, MDA, SOD GSH-Px,, SOD GSH-Px (P 0.01), MDA, (P 0.01), 3.3 红 茶 对 小 鼠 血 清 ALT 和 AST 的 影 响 ALT AST 1, ALT AST,,
3640 食品安全质量检测学报 Fig. 1 图 1 红茶对小鼠肝指数的影响 Effects of black tea (BT) on hepatic index of mice 注: CK: 正常对照组, M: 模型对照组, Y: 阳性药对照组 与模型对照组比较* P<0.05, ** P<0.01 图 2 3 4 与此相同 图 2 红茶对小鼠肝匀浆 SOD 的影响 Fig. 2 Effects of BT on liver SOD of mice 图 3 红茶对小鼠肝匀浆 GSH-Px 的影响 Fig. 3 Effects of BT on liver GSH-Px of mice 第6卷
9, : 3641 4 MDA Fig. 4 Effects of BT on liver MDA of mice,, ALT AST, (P 0.05) (P 0.01), 1 ALT AST,, ALT AST 3.4 红 茶 对 小 鼠 肝 组 织 病 理 形 态 学 的 影 响 3.4.1 小 鼠 肝 脏 肉 眼 观 察 结 果, ( 5),,,,, ;,,,, ;,,,,,,,,,,, 5,, :,, ;, 表 1 红 茶 对 小 鼠 血 清 ALT AST 水 平 的 影 响 ( x±s) Table 1 Effects of BT on ALT and AST in serum of mice n ALT (U/L) AST (U/L) 10 38.75±4.13 153.25±34.78 10 45.25±4.74 192.13±23.47 10 42.00±5.81 170.50±32.88 10 31.57±9.83 * 104.00±54.46 ** 10 34.00±8.62 * 130.50±46.47 * 10 32.60±15.57 * 123.20±61.34 * 10 29.50±9.67 ** 121.50±55.89 ** 10 32.00±9.73 * 122.86±59.71 ** 10 34.40±14.69 119.80±54.62 ** 10 40.13±15.06 143.88±45.59 * 10 29.13±9.34 ** 122.13±42.57 ** 10 29.14±13.23 ** 118.14±67.85 ** 10 33.25±12.99 * 109.13±36.48 ** 10 23.25±10.36 ** 99.13±51.31 ** 10 25.43±7.09 ** 91.14±29.61 ** 10 35.88±9.73 153.38±52.47 10 29.00±14.56 ** 117.00±74.64 ** 10 35.43±14.64 148.00±54.77 10 29.00±6.63 ** 91.50±32.89 ** 10 35.57±13.05 114.14±41.97 ** 10 32.88±11.89 * 136.25±49.72 * 10 38.63±14.97 139.38±43.52 * 10 26.86±12.05 ** 96.14±59.57 ** 10 29.00±6.63 ** 82.67±31.60 ** : * P<0.05, ** P<0.01
3642 第6卷 食品安全质量检测学报 3.4.2 小鼠肝组织显微镜观察结果 小鼠肝组织 HE 染色后显微镜观察并拍照, 结果 如图 6 所示 正常对照组小鼠肝脏组织结构完整, 肝 小叶轮廓 中央静脉 肝细胞索及肝窦清晰, 肝细胞 索围绕中央静脉呈放射状排列, 肝细胞大小较一致, 细胞核清晰, 胞浆均匀(图 6-CK) 而饲喂 5 周高脂饲 料的模型组, 肝细胞组织结构异常, 肝小叶结构紊乱 且界限模糊, 肝窦变窄甚至消失, 肝细胞排列辐射状 模糊, 肝小叶周围肝细胞胞浆出现大量的圆形空泡, 造成严重脂肪变性(图 6-M) 相比于高脂模型组, 进食高脂饲料同时灌喂低 中 高剂量红茶能够梯度性的改善由高脂饲料引起的 肝组织病变(图 6), 肝组织切片观察显示, 低剂量组 肝组织有严重水样变, 中剂量组轻微水样变, 但均未 形成脂肪空泡, 高剂量组肝组织基本恢复正常, 与阳 性药效果相当, 肝小叶周围的肝细胞发生水泡的细 胞数量减少, 脂滴明显减少甚至消失, 肝细胞肿胀 变性的程度有所降低, 细胞间界限较清晰 说明红茶 对高脂饮食小鼠肝脂肪变性具有较强的修复作用, 有效减轻了小鼠的肝损伤 4 讨 论 研究结果提示, 高脂饮食可造成小鼠体内严重 的氧化与抗氧化的失衡, 使机体肝组织正常的抗氧 化能力明显受到损伤 而红茶能显著升高肝脏 SOD 与 GSH-Px 酶的活性, 与此同时还可以降低 MDA 的 含量,提高肝细胞的抗氧化能力, 有效清除机体自由 基, 调节和改善自由基代谢平衡, 改善肝脏脂质过氧 化状态, 减少脂质过氧化产生的毒素, 从而达到保护 肝脏以及修复已损伤的肝组织的功能 有研究表明, 茶多酚 茶黄素及茶多糖具有清除氧自由基和抗脂 质过氧化的能力 [20-22], 红茶在体内对肝组织氧化损 伤保护作用的机制很可能是与其有关 一方面红茶 Fig. 5 图 5 小鼠肝照片 Liver photos of mice in different groups 注: CK-空白对照组; M-高脂模型对照组; Y-阳性药组; JD-将军红 低剂量组; JZ-将军红中剂量组; JG-将军红高剂量组; SD-石门红茶 中的茶多酚与茶黄素含有酚性羟基, 为清除自由基 团提供了位点, 易氧化成醌类而提供 H+, H+可结合 自由基而将其还原为惰性化合物, 防止损伤肝脏; 红低剂量组; SZ-石门红茶中剂量组; SG-石门红茶高剂量组; QD- 另一方面, 红茶中含有较高的茶多糖, 该类物质具 祁门红茶低剂量组; QZ-祁门红茶中剂量组; QG-祁门红茶高剂量 有较好的抗氧化作用, 能清除多种自由基, 以防其 组; ND-宁红低剂量组; NZ-宁红中剂量组; NG-宁红高剂量组; FD- 对肝脏的损伤 凤庆红茶低剂量组; FZ-凤庆红茶中剂量组; FG-凤庆红茶高剂量 组; DD-大唐英红低剂量组; DZ-大唐英红中剂量组; DG-大唐英红 本研究发现, 高脂模型组小鼠血清 ALT 与 AST 高剂量组; XD-霞山红低剂量组; XZ-霞山红中剂量组; XG-霞山红 活力均有所升高, 说明高脂饮食对肝脏正常功能造 高剂量组 图 6 中相同 成了一定的影响 而红茶各剂量组均不同程度地降低
第9期 陈金华, 等: 红茶对高脂饮食小鼠肝脏的保护作用研究 Fig. 6 3643 图 6 小鼠肝组织形态学变化( 100) Changes of morphology of liver tissues in mice ( 100) 血清中 ALT 与 AST 活力, 这提示红茶能够保护并修 小鼠喂食高脂饲料后, 肝组织肉眼观察与肝脏 复和稳定肝细胞的质膜 线粒体膜的作用 与模型对 切片显微镜观察显示, 肝细胞及肝小叶结构出现了 照组比较, 灌胃各剂量红茶的小鼠肝脏指数显著降 异常, 而同时灌胃了不同剂量红茶后发现, 胞浆内 低, 这在一定程度上与红茶中一些酚类物质及其氧 脂滴减少或消失, 无明显炎细胞坏死或浸润, 其中 化产物的抗氧化以及抗炎作用有关 [20-22] 效地减轻了肝脏的水肿和病变症状, 从而也有 高剂量红茶组更接近正常对照组肝脏情况, 而低剂 量组相比高脂模型组, 肝组织病变程度有所改善,
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