Sensitive and Specific Photometric Determination of Mannitol in Human Serum
Sensitive and Specific Photometric Determination of Mannitol in Human Serum
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DOI：
10.1515/cclm.2003.162
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发表时间：
2003-01
期刊：
影响因子：
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通讯作者：
H. Graefe;B. Gütschow;H. Gehring;L. Dibbelt
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文献类型：
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作者：
H. Graefe;B. Gütschow;H. Gehring;L. Dibbelt
甘露醇是一种具有药理活性的多元醇，常存在于微创手术中用于冲洗暴露组织的液体中。由于这种多元醇通常不会在人血浆中检测到任何显著程度，因此可将其用作手术期间含甘露醇冲洗液吸收的实验室标志物。为此，我们开发了一种可在近患者环境中进行的人血液或血清中甘露醇的光度测定法。用三氯乙酸对样品进行脱蛋白后，将上清液与NAD+和市售甘露醇2-脱氢酶制剂混合，并在pH 7.8和37 ℃下孵育30至60分钟。在孵育期结束时，适当稀释溶液，并在340 nm处用光度法测定甘露醇氧化形成的NADH浓度。本法对血清甘露醇的检测限为0.05mmol/l，线性范围约为3 mmol/l。在分析物浓度为0.48、1.38和3.48 mmol/l时，试验间变异系数分别为12.1、6.7和4.9%。血清样品中甘露醇的分析回收率接近100%。27多元醇，单糖和低聚糖测试，没有表现出可测量的底物活性，只有D-果糖显着抑制甘露醇的氧化在样品浓度高于10 mmol/l;酶促反应，但是，强烈影响EDTA。通过分析24例接受经尿道前列腺切除术患者血清样本中的甘露醇，证明了该测定作为检测和定量术中吸收冲洗液的常规诊断工具的适用性。
Abstract Mannitol is an osmotically active polyalcohol often present in fluids used for irrigation of exposed tissue during minimal invasive surgery. Since this polyol normally is not detected in human plasma to any significant extent, it may be used as a laboratory marker of absorption of mannitol-containing irrigative fluids during surgery. For this aim, we developed a photometric assay of mannitol in human blood or serum that may be performed in a near-patient setting. Following deproteinization of the sample with trichloroacetic acid, the supernatant is mixed with NAD+ and a commercially available preparation of mannitol 2-dehydrogenase and is incubated at pH7.8 and at 37 °C for 30 to 60 minutes. At the end of the incubation period the solution is appropriately diluted and the concentration of NADH formed by oxidation of mannitol is determined photometrically at 340 nm. The limit of detection of serum mannitol with this assay is 0.05 mmol/l, the linear range of measurement extends to about 3 mmol/l. At analyte concentrations of 0.48, 1.38 and 3.48 mmol/l, coefficients of inter-assay variation of 12.1, 6.7 and 4.9%, respectively, were obtained. The analytical recovery of mannitol added to serum samples was close to 100%. Of 27 polyalcohols, monosaccharides and oligosaccharides tested, none exhibited a measurable substrate activity and only D-fructose significantly inhibited the oxidation of mannitol at sample concentrations above 10 mmol/l; the enzymatic reaction, however, was strongly affected by EDTA. The suitability of the assay as a routine diagnostic tool for detection and quantification of intraoperatively absorbed irrigation fluid was demonstrated by analyzing mannitol in serum samples obtained from 24 patients undergoing transurethral prostatectomy.