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51.
偶联酶催化分光光度法测定黄嘌呤 总被引:2,自引:0,他引:2
研究了以黄嘌呤氧化酶-辣根过氧化物酶-苯酚-4-氨基安替比林反应为显色体系测定不同样液中黄嘌呤浓度的新方法。确定该测定方法的最佳反应条件为:黄嘌呤氧化酶(XO)0·32U/mL,辣根过氧化物酶(HRP)7·0U/mL,4-氨基安替比林(AAP)1mmol/L,苯酚(PA)6mmol/L溶于100mmol/LTris-HCl缓冲液(pH8·4);反应温度为37℃,保温时间为20min;检测波长为508nm。本方法测定黄嘌呤浓度的线性范围为0·2~10·0mmol/L,线性关系良好(R=0·9978),检测限为0·05mmol/L。方法操作简单易行,测定结果准确可靠,可有效应用于普通实验室和常规临床血液生化检测。 相似文献
52.
Peter Brodelius Kjell Nilsson Klaus Mosbach 《Applied biochemistry and biotechnology》1981,6(4):293-307
Whole cells ofTrigonopsis variabilis were immobilized by entrapment in Ca2+-alginate and used for the production of α-keto acids from the corresponding D-amino acids. The D-amino acid oxidase within
the immobilized cells has a broad substrate specificity. Hydrogen peroxide formed in the enzymatic reaction was efficiently
hydrolyzed by manganese oxide co-immobilized with the cells. The amino acid oxidase activity was assayed with a new method
based on reversed-phase HPLC. Oxygen requirements, bead size, concentration of cells in the beads, flow rate, and other factors
were investigated in a “ trickle-bed ” reactor. 相似文献
53.
QinZHAO RuoYUAN ChangLiMO YaQinCHAI XiaZHONG 《中国化学快报》2004,15(2):208-211
Naphthol green B was used, for the first time, as a new mediator in an amperometric glucose biosensor. It is a good mediator, promoting electron transfer from glucose oxidase to graphite electrode. The biosensor shows high sensitivity to glucose at low potential with response time of 30 seconds. The linear range is from 1.5 to 18μmol/L glucose with detection limit of 0.5μmol/L glucose. 相似文献
54.
New compounds obtained by enzymatic oxidation of phloridzin 总被引:1,自引:0,他引:1
Oxidation of phloridzin was studied in a model system in the presence of apple polyphenol oxidase. In addition to 3-hydroxy phloridzin, two major oxidation products were purified by reversed phase HPLC at the semi-preparative scale. Their structures were elucidated by UV, ESI-MSn and NMR spectroscopies. The first compound was a colourless product, whose novel structure strongly differs from its precursor showing a biphenyl moiety and a propionic acid chain. The second product was an oxidised form of the first one and corresponded to a stable yellow pigment with two isomeric forms. A mechanism of formation of these products, which implied successive oxidation and nucleophilic addition steps was proposed. 相似文献
55.
在有无5 mmol·L-1 CuSO4存在的两种情况下,运用荧光光谱研究了烟草多酚氧化酶在盐酸胍诱导下的变性和复性平衡。烟草多酚氧化酶在6.0 mol·L-1盐酸胍变性30 min(25 ℃)即完全失活。荧光光谱结果表明:铜离子能够提高烟草多酚氧化酶的结构稳定性和抗盐酸胍变性的能力,进而影响烟草多酚氧化酶在盐酸胍诱导下的变性和复性过程。没有外源铜存在的情况下,盐酸胍诱导的烟草多酚氧化酶变性和复性是一个可逆的二态过程;在5 mmol·L-1 CuSO4存在的条件下,由于结合了Cu2+的酶的中间态稳定性增加,显示特征荧光,结果显示,外源铜存在时,盐酸胍诱导的烟草多酚氧化酶变性和复性是一个可逆的三态过程。根据相应模型,进行了热力学计算。上述实验结果得到酶活性测定的进一步证实,在6 mol·L-1盐酸胍中放置5,10,15,20,25,30 min,烟草多酚氧化酶的剩余活性分别为18.7%,11.7%,8.9%,6.6%,3.6%,0.06%,但在5 mmol·L-1 CuSO4存在的条件下,剩余活性则分别为60.3%,44.6%,42.5%,40.2%,25.6%,25.3%。 相似文献
56.
Purification and reversible immobilization of d-amino acid oxidase from Trigonopsis variabilis could be simultaneously accomplished by hydrophobic interaction on Phenyl Sepharose CL-4B in the presence of 50 mM pyrophosphate buffer (pH 8.5). The presence of a high salt concentration of 2M, which is generally required for the hydrophobic interactions, was not essential for the hydrophobic immobilization. The
enzyme in free as well as immobilized form was optimally active between pH 7.0 and 9.0. The immobilized preparation could
be reused in a batch process for the conversion of d-amino acids to α-keto acids. When the activity of the preparation dropped below practical limits, the gel could be regenerated
by water wash and recharged with fresh crude extract from yeast. 相似文献
57.
Michel F Thomas F Hamman S Saint-Aman E Bucher C Pierre JL 《Chemistry (Weinheim an der Bergstrasse, Germany)》2004,10(17):4115-4125
Galactose oxidase (GO) is an enzyme that catalyzes two-electron oxidations. Its active site contains a copper atom coordinated to a tyrosyl radical, the biogenesis of which requires copper and dioxygen. We have recently studied the properties of electrochemically generated mononuclear Cu(II)-phenoxyl radical systems as model compounds of GO. We present here the solution chemistry of these ligands under various copper and dioxygen statuses: N(3)O ligands first chelate Cu(II), leading, in the presence of base, to [Cu(II)(ligand)(CH(3)CN)](+) complexes (ortho-tert-butylated ligands) or [(Cu(II))(2)(ligand)(2)](2+) complexes (ortho-methoxylated ligands). Excess copper(II) then oxidizes the complex to the corresponding mononuclear Cu(II)-phenoxyl radical species. N(2)O(2) tripodal ligands, in the presence of copper(II), afford directly a copper(II)-phenoxyl radical species. Addition of more than two molar equivalents of copper(II) affords a Cu(II)-bis(phenoxyl) diradical species. The donor set of the ligand directs the reaction towards comproportionation for ligands possessing an N(3)O donor set, while disproportionation is observed for ligands possessing an N(2)O(2) donor set. These results are discussed in the light of recent results concerning the self-processing of GO. A path involving copper(II) disproportionation is proposed for oxidation of the cross-linked tyrosinate of GO, supporting the fact that both copper(I) and copper(II) activate the enzyme. 相似文献
58.
Leitner Christian Hess Johann Galhaup Christiane Ludwig Roland Nidetzky Bernd Kulbe Klaus D. Haltrich Dietmar 《Applied biochemistry and biotechnology》2002,98(1-9):497-507
The wood-degrading fungus Trametes multicolor secretes several laccase isoforms when grown on a simple medium containing copper in the millimolar range for stimulating
laccase synthesis. The main isoenzyme laccase II was purified to apparent homogeneity from the culture supernatant by using
anion-exchange chromatography and gel filtration. Laccase II is a monomeric glycoprotein with a molecular mass of 63 kDa as
determined by sodium dodecylsulfate polyacrylamide gel electrophoresis, contains 18% glycosylation, and has a pI of 3.0. It oxidizes a variety of phenolic substrates as well as ferrocyanide and iodide. The pH optimum depends on the substrate
employed and shows a bell-shaped pH activity profile with an optimum of 4.0 to 5.0 for the phenolic substrates, while the
nonphenolic substrates ferrocyanide and 2,2′-azino-bis(3-ethylbenzthiazoline-6-sulfonate) show a monotonic pH profile with
a rate decreasing with increasing pH. 相似文献
59.
Zhihuang Chen 《Talanta》2007,72(4):1410-1415
In this paper, the electrochemiluminescence (ECL) behavior of bis-[3,4,6-trichloro-2-(pentyloxycarbonyl)-phenyl] oxalate (BTPPO) at glassy carbon electrode (GCE) in phosphate buffer solution in the presence of hydrogen peroxide has been investigated when linear sweep voltammetry was applied. The optimum chemical conditions and electrochemical parameters for this ECL system have been investigated in detail. Under the optimum conditions, it was found that the concentration of BTPPO was linear with the ECL intensity in the range of 3.0 × l0−6 to 3.0 × 10−4 mol/L, and the detection limit (S/N = 3) for BTPPO was 1.0 × 10−7 mol/L. The possible mechanism for ECL of BTPPO at the GCE in the presence of hydrogen peroxide was also discussed. Furthermore, based on the fact that glucose oxidase can react with glucose to produce hydrogen peroxide, a new ECL sensing system of BTPPO has been developed for detection of glucose. The enhanced ECL intensity has a linear relationship with the concentration of glucose in the range of 1.0 × l0−4 to 1.0 × 10−3 mol/L, and the detection limit for glucose is found to be 5.0 × 10−5 mol/L (S/N = 3). 相似文献
60.
《Analytical letters》2012,45(3):215-221
Abstract An enzymatic method for estimating the age of biological specimens is described. The method is based on the slow racemization of L-aspartic acid to the D isomer. Conditions for protein extraction, enzymatic hydrolysis, pretreatment with stereospecific enzymes and assaying are discussed. The results obtained by this method are compared with the more conventional radiocarbon technique and its application to Holocene climatology is discussed. 相似文献