飲食油脂與大蒜精油對大鼠抗氧化酵素及生物轉換酵素活性之影響

本實驗主要目的是探討飲食油脂種類及含量與大蒜精油(garlic oil, GO)對大鼠肝臟抗氧化酵素活性與生物轉換酵素活性及表現之影響。實驗以五週齡雄性SD大鼠為對象，分別餵食三種不同油脂飲食：低玉米油(5 % corn oil)、高玉米油(23.5% corn oil)和高魚油飲食(20.5% fish oil + 3% corn oil)，同時各組大鼠也利用口灌方式分別灌食0, 30, 80及200mg/kg bw大蒜精油(載劑玉米油1 ml/kg)，每週灌食3次，為期6週。 結果顯示：魚油組肝臟磷脂質中EPA、DHA脂肪酸比例顯著大於玉米油組，而花生四烯酸比例則顯著低於玉米油組(P<0.05)。高玉米油及高魚油組大鼠肝臟/體重比顯著高於低玉米油組(P<0.05)，肺臟、心臟及腎臟體重比則在各飲食油脂組間均相似；然而脾臟/體重比則顯著受到大蒜精油的影響(P<0.05)，隨大蒜精油劑量增加而增加。血脂質部分，高玉米油組比低玉米油組及高魚油組有較高的HDL-C (P<0.05)；而LDL-C、三酸甘油酯及總膽固醇量則是低玉米油組最高，高玉米油組次之，高魚油組最低(P<0.05)。進一步分析肝臟總脂質含量，發現高玉米油組的肝臟總脂質量明顯高於低玉米油組及高魚油組(P<0.05)。 低玉米油組和高魚油組相較高玉米油組有較高還原型GSH及總GSH 濃度(P<0.05)；肝臟內生性TBARS濃度，魚油組則顯著高於玉米油組(P<0.05)，且大蒜精油劑量與飲食油脂對肝臟脂質過氧化程度存有交互作用(P<0.05)。各飲食組中，glutathione peroxidase (GPx)活性均與大蒜精油劑量呈現負相關，相反的，glutathione reductase (GRd)、glutathione S-transferase (GST)及placental glutathione S-transferase (PGST)等酵素活性則與大蒜精油劑量呈正相關；GRd、PGST與superoxide dismutase (SOD)等酵素活性以高魚油組最高(P<0.05)，除SOD活性外，飲食油脂與大蒜精油劑量並無交互作用。利用氧化物誘發肝臟均質液脂質過氧化時，發現大蒜精油可明顯降低50μM FeSO4誘發TBARS生成濃度(P<0.05)。在紅血球中，還原態GSH濃度、總GSH濃度及GSH / GSSG比值均隨大蒜精油劑量增加而增加(P<0.05 )；魚油組GSSG濃度也較低玉米油組及高玉米油組高(P<0.05)；紅血球GSH相關抗氧化酵素活性顯示，除了高玉米油組GPx活性顯著大於高魚油組及低玉米油組(P<0.05)外，GRd及GST活性均不受飲食油脂影響；GRd活性均隨大蒜精油劑量增加而減少(P<0.05)；而GST活性，則是隨著大蒜精油劑量增加而遞增。 魚油組pentoxyresorufin O-depentylase (PROD)、ethroxyresorufin O-deethylase (EROD)活性顯著高於高玉米油組及低玉米油(P<0.05)；N-nitrosodimethylamine demethylase (NDMAD)活性則是高魚油組顯著高於低玉米油組(P<0.05)；大蒜精油效應上，肝臟PROD與EROD活性顯示出隨著大蒜精油劑量增加而增加的趨勢，相反的，NDMAD活性則是隨著大蒜精油劑量增加而減少。CYP 2B1、1A1、2E1、3A1及PGST蛋白質含量均以高魚油組表現最多，除了CYP 3A1蛋白質外，其餘3種細胞色素蛋白表現也以高玉米油組高於低玉米油組，然而GST Ya、Yb、Yc蛋白質則以高玉米油組最顯著；除飲食油脂外，大蒜精油也明顯調控生物轉換酵素蛋白質表現，各飲食組中GST Ya、Yb、Yc、PGST及CYP 1A1、2B1、3A1蛋白質表現均隨大蒜精油劑量增加而增加；相反的，CYP 2E1蛋白質表現則是隨大蒜精油增加而減少。肝臟解毒酵素mRNA表現與蛋白質表現相似，PGST、CYP 2B1、1A1和3A1 mRNA表現量均隨大蒜精油劑量增加而遞增，油脂效應也是以高魚油組表現最多，高玉米油組次之，低玉米油組最低。 以上實驗結果得知，飲食油脂種類及含量與大蒜精油確實可影響肝臟解毒酵素的表現，且魚油對肝臟PGST、CYP 2B1、1A1和3A1蛋白質及mRNA表現與大蒜精油有增加作用。大蒜精油可經由調控肝臟抗氧化酵素活性而增加抗氧化能力。

The main objective of this study was to investigate the combined modulatory effect of source and amount of dietary fat and garlic oil on rat hepatic antioxidant and biotransformation enzyme activities and expression. Sprague-Dawley rats were fed either a low corn oil diet (LCO, 5% corn oil), high corn oil diet (HCO, 23.5% corn oil), or high fish oil diet (HFO, 3.5% corn oil + 20% fish oil) and received 0, 30, 80, 200 mg/kg body weight of garlic oil (corn oil as a vehicle, 1ml/kg bw) three times per week for 6 weeks. Results showed that eicosapentaenoic acid and docosahexaenoic acid levels in hepatic phospholipids were higher in the fish oil-fed group than in the corn-oil fed groups (P<0.05). In contrast, the corn oil-fed groups had greater hepatic phospholipid arachidonic acid levels (P<0.05). Rats fed the HCO or HFO diet had significantly greater relative liver weights than rats fed the LCO diet (P<0.05). The relative lung, heart and kidney weights were not changed by dietary lipid and garlic oil. But the spleen weight was significantly changed by garlic oil (P<0.05). In each dietary lipid group, garlic oil dose-dependently increased the relative spleen weights. HDL-C in rats fed the HCO diet was significantly higher than rats fed the LCO or HFO diet (P<0.05). LDL-C, triacylglycerol and total cholesterol in rats fed the LCO diet were significantly greater than rats fed the HCO diet, and rats fed the HCO diet had significantly greater than rats fed the HFO diet (P<0.05). Rats fed the HCO diet had significantly greater hepatic total lipids than rats fed the LCO or HFO diet (P<0.05). Rats fed the LCO or HFO diet had significantly greater hepatic GSH and total GSH levels than rats fed the HCO diet. The endogenous TBARS level in the liver tissues, however, was significantly higher in the HFO-fed group than the LCO- or HCO-fed group (P<0.05). There was also an interaction between dietary fat and garlic oil on the hepatic TBARS levels. In each dietary group, garlic oil dose-dependently decreased glutathione peroxidase (GPx) activity (P<0.05). Regardless of dietary lipid, garlic oil caused a dose-dependent increase of hepatic glutathione reductase (GRd), glutathione S-transferase (GST), placental form of glutathione S-transferase (PGST) and superoxide dismutase (SOD) activities. Moreover, the hepatic GRd, PGST and SOD activities were highest in rats fed the HFO diet (P<0.05). In contrast to the lack of interaction noted on hepatic glutathione-related enzyme activities, garlic oil interacted with dietary fat on the SOD activity. The lipid peroxidation was also studied in the presence of exogenous oxidants. Ferrous sulfate (50μM)-induced lipid peroxidation was significantly suppressed by garlic oil (P<0.05). In the red blood cells, total GSH, GSH/GSSG ratio and GRd activity were increased dose-dependently by garlic oil (P<0.05). Rats fed the fish oil diet had significantly higher GSSG level than rats fed the corn oil diets (P<0.05). GSH-related enzyme activity assays revealed that, with the exception of HCO-fed group had significantly greater GPx activity than HFO- or LCO- fed group (P<0.05), GRd and GST activities were not affected by dietary lipids. Garlic oil caused a dose-dependent decrease of GRd activity (P<0.05). In contrast, garlic oil tended to dose-dependently increased GST activity. Regarding to the biotransformation enzymes in liver tissues, rats fed the fish oil diet had greater pentoxyresorufin O-depentylase (PROD), ethroxy- resorufin O-deethylase (EROD) activities than rats fed the corn oil diets (P<0.05), and rats fed the HFO diet had greater N-nitrosodimethylamine demethylase (NDMAD) activity than those fed the LCO diet (P<0.05). In all dietary fat groups, garlic oil tended to dose-dependently increase PROD and EROD activities in low corn oil group. However, NDMAD activity in all dietary lipid groups was dose-dependently decreased by garlic oil. Immunoblot assay showed that rats fed the HFO diet had the highest CYP 2B1, 1A1, 2E1, 3A1 and PGST proteins levels among threr dietary lipids groups. Besides the CYP 3A1 protein level, the protein levels of CYP 1A1, 2B1 and 2E1 in the HCO-fed group was higher than the LCO-fed group. To GST isozyme expression, HCO increased GST Ya, Yb, Yc protein levels as compared wuth LCO or HFO. It was also noted that garlic oil caused a dose-dependent increase with GST Ya, Yb, Yc, PGST and CYP 1A1, 2B1, 3A1 protein levels. On the contrary, garlic oil caused a dose-dependent decrease of CYP 2E1 pretein level. The induction of the hepatic CYP mRNA expression were consistent with the peotein levles. The mRNA expression of PGST, CYP 2B1, 1A1 and 3A1 were dose-dependently increased in the presence of garlic oil. These mRNA expression were highest in rats fed the HFO diet. In addition to the effect of fish oil, the mRNA expression of CYP enzymes in rats fed the HCO diet was higher than rats fed the LCO diet. In conclusion, results suggest that the source and amount of dietary lipids and garlic oil are potentiate in modulation of the biotransformation enzymes. Moreover, fish oil and garlic oil independently regulated the expression of PGST, CYP 2B1, 1A1 and 3A1 protein and mRNA. Garlic oil increased antioxidant capacity by modulating the hepatic antioxidant enzyme activities.