Effect of divalent cations and proteases on skin sulfhydryl oxidase activity

Crude skin sulfhydryl oxidase was independently isolated from the living cell layer, the granular cell layer, and the lower living cell layer (spinous and basal) of cow snout. The properties of this enzyme were subsequendy investigated. The addition of 1 mM CuCl₂ and FeCl₂ stimulated the enzyme activity to 216% and 166% of the initial activity, respectively. Neither CaCl₂ nor MgCl₂ had much effect on the activity, and ZnCl₂ inhibited it. Diethyldithiocarbamate (DEDTC), a copper ion chelating agent, inhibited the activity in a dose and pre-incubation time-dependent manner: however, other divalent cation chelators such as EDTA, EGTA, o-phenanthrolin and α,α'-dipyridyl did not have any effect on the enzyme's behavior. From these findings, it was determined that Cu²⁺ was essential for the activity of skin sulfhydryl oxidase. This enzyme was stimulated to 130–150% of its initial activity by treatment with 1 mg/ml trypsin, chymotrypsin or urokinase but was not affected by plasmin, elastase or cathepsin D. Trypsin treatment enhanced the activity in a dose and treatment-time dependent manner. Skin sulfhydryl oxidase from the lower living cell layer (spinous and basal) was more highly activated by trypsin treatment than was that from the granular cell layer. These findings suggest that this enzyme may be activated by some kind of serine proteases during the keratinocytes autolysis process in the granular cell layer.     

铜离子络合法测定血浆中司帕沙星的含量

目的 建立铜离子络合法测定血浆中司帕沙星含量.方法 以适量的铜离子与司帕沙星作用,生成稳定的络合物,增强被测药物的灵敏度,采用紫外分光光度法,在297 nm处测定血浆中司帕沙星的含量.结果 司帕沙星在0.6-10 μg/ml范围内线性良好,r=0.991 8,检测限为0.015 9 μg/ml,回收率为98.82%-112.5%.高、中、低三浓度的日内精密度的RSD分别为0.67%,1.15%,7.54%,日间精密度分别为0.51%,2.75%,5.94%.均符合生物样品测定的要求.结论 本法具有快速、简便、准确等特点,可用于临床司帕沙星的血药浓度检测.     

Strain-Promoted Catalyst-Free Click Chemistry for Rapid Construction of 64Cu-Labeled PET Imaging Probes

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Electrocatalytic effects in the electrochemical reduction of pyrazine on single crystal IB metal electrodes

Due to the presence of the second nitrogen atom, pyrazine has an increased affinity for electrons and lower energies of the empty orbitals than pyridine. For this reason the electrochemical reduction of pyrazine takes place at considerably less negative potentials than that of pyridine. The reduction of pyrazine on Cu(111), Ag(111) and Au(111) electrodes in neutral aqueous media has been found to take place in one step as previously reported for Hg. The potential region and the current densities associated with this process were, however, observed to depend significantly on the electrode nature. Pyrazine was also found to be involved in a chemical interaction with the hydrogen atoms resulting in the water electro-reduction process occurring on Au(111) and Cu(111). The specific interaction of water with the single crystal electrodes turned out to be an important factor for developing the reduction of the organic molecule, both chemically and electrochemically.     

0～3岁儿童全血微量元素的检测及意义

目的检测了解0～3岁健康儿童全血铜、锌、钙、镁、铁5种微量元素水平,为制订儿童营养膳食结构及健康检查提供指导和参考。方法采用原子吸收光谱法测定2 000例0～3岁儿童全血中铜、锌、钙、镁、铁5种微量元素并对检测结果进行分析。结果 0～3岁儿童分为三组,三组儿童同种微量元素比较水平差异无统计学意义（P〉0.05）,其中锌缺乏率为54.75%,铁缺乏率为23.55%,铜缺乏率为0.8%,钙缺乏率为0.45%,镁缺乏率为0.1%。结论 0～3岁儿童全血中锌和铁缺乏较为严重,铜、钙、镁缺乏较低,建议加大宣传力度,调整饮食结构,重视儿童健康发育,定期健康体检,争取做到早发现、早预防、早治疗,保障儿童健康成长。     

Preventive effect of melatonin on the progression of alpha-naphthylisothiocyanate-induced acute liver injury in rats

The preventive effect of melatonin on the progression of alpha-naphthylisothiocyanate (ANIT)-induced acute liver injury with cholestasis was examined in rats treated once with the hepatotoxin [75 mg/kg body weight (BW), i.p.]. In rats treated with ANIT alone, liver injury with cholestasis occurred 24 hr after treatment and progressed at 48 hr, judging from the serum levels of hepatobiliary marker enzymes and components. Melatonin (10 or 100 mg/kg BW) was orally administered to the ANIT-treated rats, 24 hr after the hepatotoxin treatment at which time hepatic injury had already developed. The administered indoleamine prevented the progression of liver cell damage rather than biliary cell damage more effectively at the higher dose than at the lower dose. In rats treated with ANIT alone, the serum and hepatic concentrations of thiobarbituric acid reactive substances, an index of lipid peroxidation, and the hepatic activity of myeloperoxidase, an index of tissue neutrophil infiltration, increased 24 hr after treatment and further increased at 48 hr. In the liver of rats treated with ANIT alone, Cu,Zn-superoxide dismutase activity decreased 24 hr after treatment and was further reduced at 48 hr, although there was no change in Mn-superoxide dismutase activity. Catalase and Se-glutathione peroxidase activities also decreased at 48 hr, while reduced glutathione concentrations remained increased at 24 and 48 hr. The melatonin administered to the ANIT-treated rats attenuated the increases in serum and hepatic concentrations of thiobarbituric acid reactive substances and the decreases in hepatic activities of Cu,Zn-superoxide dismutase, catalase, and Se-glutathione peroxidase found at 48 hr after the hepatotoxin treatment more effectively at the higher dose than at the lower dose; on the other hand, melatonin treatment had no effect on the increases in hepatic myeloperoxidase activity and reduced glutathione concentration found at 48 h. These results indicate that orally administered melatonin at pharmacological doses prevents the progression of ANIT-induced acute liver injury, mainly liver cell damage, in rats, and suggest that the administered melatonin exerts these preventive effects through its direct and indirect antioxidant actions.     

Diagnostic value of the copper/zinc ratio in hepatocellular carcinoma: a case control study

Background: The aim of this study was to assess the accuracy of the copper/zinc ratio in the evaluation of a group of patients with hepatocellular carcinoma (HCC). Methods: A total of 105 patients were studied and separated into three groups: group I (n = 40), patients with HCC, group II (n = 25), patients with liver cirrhosis, and group III (n = 40), patients with benign digestive disease. Serum levels of copper and zinc were measured by atomic absorption spectrophotometry. Results: The serum levels of copper (μg/dl) in patients with HCC (97.4 ± 27.2; P < 0.05) were significantly higher than those in patients with liver cirrhosis (73.7 ± 17.5) or benign digestive disease (77.1 ± 20.8), and the serum levels of zinc (μg/dl) were significantly lower (71.6 ± 30.5; P < 0.05) than those in patients with benign digestive disease (81.7 ± 17.7 μg/dl) and were similar to those in cirrhotic patients (68.5 ± 17.1). The Cu/Zn ratio was also significantly higher in patients with HCC (1.52 ± 0.64; P < 0.05) than in patients with liver cirrhosis (1.06 ± 0.2) or patients with benign digestive disease (0.95 ± 0.39). Considering a cutoff value of 1.15, the sensitivity of the Cu/Zn ratio was 87.5%, with a specificity of 86.1%, a positive predictive value of 79.5%, and a negative predictive value of 91.8%. Conclusions: The Cu/Zn ratio was found to be significantly higher in patients with HCC compared with that in age and sex-matched controls, with a sensitivity of 87.5%; this ratio might be useful in the evaluation of suspected hepatocellular malignancy. Received: February 15, 2002 / Accepted: June 14, 2002 Acknowledgments. This work was partially supported by grant No. D-113-903903 from the Consejo Nacional de Ciencia y Tecnología, Mexico City, México. Reprint requests to: J.L. Poo, Centro de Investigación Farmacológica y Biotecnológica, Hospital Médica Sur, Puente de Piedra No 150, 14050, Mexico City, Mexico Editorial on page 104     

姜黄素的微生物转化产物体外抗低密度脂蛋白氧化作用

目的 利用红曲霉菌对姜黄素进行微生物转化,研究姜黄素微生物转化总产物对低密度脂蛋白氧化修饰的抑制作用,为进一步筛选新型高效的姜黄素类抗动脉硬化新药奠定基础.方法 利用红曲霉菌固体发酵法对姜黄素进行微生物转化,设立只加姜黄素不接种菌体的底物对照和只接种菌体不加入姜黄素的菌体对照,乙酸乙酯提取转化产物.从血浆中分离低密度脂蛋白,建立低密度脂蛋白在Cu2+诱导下的无细胞氧化体系和巨噬细胞氧化体系,姜黄素转化产物进行氧化干预,以低密度脂蛋白在琼脂糖凝胶电泳的迁移距离及硫代巴比妥酸反应物含量作为低密度脂蛋白氧化的指标.结果 姜黄素转化总产物在4～500 mg/L浓度范围内(相当于含姜黄素0.04～5 mg/L),能显著抑制Cu2+诱导下的低密度脂蛋白氧化,与菌体对照组相比差异有显著性(P<0.001);与底物对照比较在20 mg/L和4 mg/L浓度作用下差异有显著性(P<0.001).转化总产物作用小鼠巨噬细胞24 h能显著抑制低密度脂蛋白的氧化,其迁移距离比氧化对照组明显缩短(P<0.001);姜黄素对照组在5 mg/L时有明显的抑制作用;底物对照组和菌体对照组迁移距离与氧化对照组差异无显著性(P>0.05),基本无抑制作用.转化总产物在4～500 mg/L浓度范围内,能显著抑制小鼠巨噬细胞TBARS的产生,其抑制作用随着剂量的增大而增强.而底物对照组只在浓度为500 mg/L和100 mg/L(相当于含姜黄素5 mg/L和1 mg/L)、姜黄素对照组在浓度为5 mg/L和1mg/L时有一定的抑制,菌体对照组则无抑制作用.转化总产物对巨噬细胞氧化体系低密度脂蛋白氧化的抑制作用,与底物对照组、菌体对照组相比差异有显著性(P<0.001).结论 姜黄素经红曲霉菌转化后,对低密度脂蛋白的抗氧化作用显著增强,有望获得姜黄素类高效抗动脉硬化的新型化合物.     

Effect of melatonin on changes in hepatic antioxidant enzyme activities in rats treated with α-naphthylisothiocyanate

We have reported that melatonin protects against alpha-naphthylisothiocyanate (ANIT)-induced acute liver injury in rats by preventing enhanced lipid peroxidation. Herein, we examine the effect of melatonin on hepatic antioxidant enzyme activities in rats with a single i.p. injection of ANIT (75 mg/kg body weight) in order to clarify the protective mechanism of the indoleamine against ANIT-induced acute liver injury. Rats received a single oral administration of melatonin (10 or 100 mg/kg body weight) at 12 hr after ANIT treatment. Hepatic Cu,Zn-superoxide dismutase (Cu,Zn-SOD), Mn-superoxide dismutase (Mn-SOD), catalase (CAT), Se-glutathione peroxidase (Se-GSH-Px), glutathione reductase (GSSG-R), and glucose-6-phosphate dehydrogenase (G-6-PDH) activities and reduced glutathione (GSH) concentration were determined 12 and 24 hr after ANIT treatment. ANIT-treated rats showed decreases in hepatic Cu,Zn-SOD and GSSG-R activities at 24 hr after treatment, transient increases in hepatic CAT and Se-GSH-Px activities at 12 hr, and no changes in hepatic Mn-SOD and G-6-PDH activities at 12 or 24 hr. Only the high dose of melatonin attenuated the decrease in hepatic Cu,Zn-SOD activity, while both doses of the indoleamine almost completely attenuated the decrease in hepatic GSSG-R activity. Neither dose of melatonin affected hepatic CAT, Se-GSH-Px, and G-6-PDH activities. ANIT-treated rats showed an increase in hepatic GSH concentration at 24 hr after treatment. Neither dose of melatonin affected the increase in hepatic GSH concentration. These results indicate that orally administered melatonin prevents decreases in Cu,Zn-SOD and GSSG-R activities in the liver of ANIT-treated rats, and suggest that the indoleamine may protect against ANIT-induced acute liver injury by attenuating the disruption of hepatic antioxidant defense systems.