Plasma sorbitol dehydrogenase, glutamate dehydrogenase, and alkaline phosphatase as potential indicators of liver intoxication in grey mullet (Mugil auratus Risso)
Plasma sorbitol dehydrogenase, glutamate dehydrogenase, and alkaline phosphatase as potential indicators of liver intoxication in grey mullet (Mugil auratus Risso)
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血浆山梨醇脱氢酶、谷氨酸脱氢酶和碱性磷酸酶作为灰鲻鱼 (Mugil auratus Risso) 肝中毒的潜在指标
DOI：
10.1007/bf00191250
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发表时间：
1993
影响因子：
2.7
通讯作者：
M. Krajnović
中科院分区：
文献类型：
--
作者：
B. Ozretić;M. Krajnović
材料和方法使用平均体重205g（·SD）的灰鲻鱼（Mugil auratus Risso）作为试验动物。鱼在连续流海水（盐度 37.2• 0.4~，温度 18• 0.5 o C）的曝气池（250 L）中适应至少 2 周。每天喂鱼至饱，然后除去剩余的食物。根据先前的经验（Krajnovi6-Ozreti6 和 Ozreti6 1987），每组 10 条灰鲻鱼腹膜内注射。（ip）单次剂量如下： 2 mL CClA kg”= 或 200 mg 苯酚 kg” 1' 将 10 条鱼暴露在海水连续流动系统中 300~ g NaCN L” 24 小时，并配有剂量器首先将苯酚溶解在蒸馏水中，而将 30 只未经处理的鲻鱼作为对照进行注射，在注射后 24 小时测定所有参数，此时观察到血液酶活性的最大变化（Krajnovi6-Ozreti6 和 Ozreti6 1987）。血液保持冰冻，并在冷冻离心机中分离血浆（2000 g，10 分钟）。GLDH (EC 1.4. 1.2) 和 SDH (EC 1.1. 1.14) 活性采用动态 UV 方法（Schmidt 1974；Gerlach 和 Wiby 1974）和 AP（EC 3.1.1.14）测定。 3.1) 根据 Klaus 和 Schutt (1974) 的方法，灰鲻鱼的 SDH 和 GLDH 测定条件（例如 pH 和底物浓度）的优化与哺乳动物研究中使用的条件非常相似。还在肝脏、心脏、肾脏、鳃、白色和红色肌肉的提取物中用 Polytron 研磨机对 10 份冷 0.2 M 的组织样品进行均质化。将含 5 mM 巯基乙醇的磷酸钠缓冲液在 17000 g 下离心 30 分钟，然后将上清液立即用于酶测定。总血浆蛋白通过双缩脲反应测定（Weichselbaum 1946）。使用 Sigma（美国圣路易斯）生产的商业诊断测试通过分光光度法估算血浆白蛋白。 Boehringer（德国曼海姆）生化检测试剂盒
MATERIALS AND METHODSGrey mullet, Mugil auratus Risso average weight 205 g (• SD) were used as test animals. Fish were acclimated at least 2 wk in aerated basins (250 L) with a continuous flow of sea water (salinity 37.2• 0.4~ and temperature 18• 0.5 o C). Fish were fed daily to satiation and the remaining food was removed. Following previous experience (Krajnovi6-Ozreti6 and Ozreti6 1987), ten grey mullets per group were injected intraperitoneally.(ip) with a single dos~ of the following: 2 mL CClA kg"= or 200 mg of phenol kg" 1'Ten fish were exposed fo# 24 hr to 300~ g NaCN L" in a sea water continuous flow system with a doser for cyanide. Phenol was first dissolved in distilledwater, while CC14 was injectedwithoutdiluti on. A separate group of thirty untreated mullet served as control. All parameters were determined 24 hr after the injection, the time when the maximum changes in blood enzyme activities were observed (Krajnovi6-Ozreti6 and Ozreti6 1987). Blood samples were taken by cardiac puncture. Blood sampling and ip injections were performed without the use of anesthetics, Heparin was used as anticoagulant. Blood was kept iced and plasma was separated (10 min at 2000 g) in a refrigerated centrifuge.GLDH (EC 1.4. 1.2) and SDH (EC 1.1. 1.14) activities were determined with the kinetic UV method (Schmidt 1974; Gerlach and Wiby 1974) and AP (EC 3.1. 3.1) according to the method of Klaus and Schutt (1974). Optimization of SDH and GLDH assay conditions (eg pH and substrata concentration) for grey mullet revealed to be very similar to those used in mammalian studies. The activity of the same enzymes was also measured in the extracts of liver, heart, kidneys, gills, white and red muscle. Tissue samples were homogenized with a Polytron grinder in 10 parts of a cold 0.2 M Na-phosphate buffer (pH 7.4) in 20% glycerol with 5 mM mercaptoethanol. Homogenatee were centrifuged at 17000 g for 30 min, and the supernatant was immediately used for enzyme assay. For plasma, the enzyme activity was calculated in relation to the unit volume or to the unit weight for body t~ ssues, a. Rd it was expressed as International Units (U L'" and U g, respectively). Total plasma proteins were determined with Biuret reaction (Weichselbaum 1946). Plasma albumin was estimated by spectrophotometry, using a commercial diagnostic test produced by Sigma (St. Louis, USA). Boehringer (Mannheim, Germany) biochemical test kits were