Development of microdetermination of 3-series prostanoid and the study of the physiological significance in diseases

3系前列腺素微量测定方法的进展及其在疾病中的生理意义研究

• 批准号: 05454580
• 负责人: HISHINUMA Takanori
• 金额: $ 3.9万
• 依托单位: TOHOKU UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (B)
• 财政年份: 1993
• 资助国家: 日本
• 起止时间: 1993 至 1994
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-05454580/
• 关键词: prostaglandin; prostacycline; thromboxane; GC-MS; 11-dehydro-TXB2; 11-dehydro-TXB3; biodistribution; 11-dehydro-TXB_3; 6,15-diketo-13,14-dihydro-PGF_<1α>; GC; MS; SIM; モノクローナル抗体; 阻害ELISA; 3型PG

1.The microdetermination of 11-dehydro-thromboxane B3 (11-dehydro-TXB3) in human urine is described. We prepared [^<18>O] 11-dehydro-thromboxane B3 for use as an internal standard (IS). Samples to which was added [^<18>O] analogue were extracted by chromatographic sample preparation using Sep Pak tC18 and silica gel column. Conversion of the extracted11-dehydro-thromboxane B3 to 1-methyl ester (ME) -11-n-propylamide (PA) -methoxime-9,11,15-tris-dimethylisopropylsilyl (DMIPS) ether derivative was followed by gas chromatography/high resolution-selected ion monitoring (GC/HR-SIM). Interfering substances from the urine matrix were eliminated during GC/HR-SIM analysis using a MP-65HT column. Good linear response over the range of 10 pg-100 ng/tube was demonstrated. We were able to detect 11-dehydro-TXB3 in the range from 26 to 375 pg/ml of the human urine. The present method can be applied to the determination of 11-dehydro-TXB3 in the human urine2.The microanalysis of 2,3-dinor-6-keto-pros … More taglandin F1alpha (I) in human urine was developped. Samples to which was added [^2H4] -analogue as an internal standard were extracted by chromatographic sample preparation using Bond Elut C18, and silica gel. Conversion of the extracted I to 1-methyl ester-6-methoxime-9,11,15-tris-dimethy-lisopropylsilyl ether derivative was followed by gas chromatography/high resolution-selected ion monitoring (GC/HR-SIM). Inrerfering substances from the urine matrix were eliminated during GC/HR-SIM analysis.A method of the microdetermination of 2,3-dinor-DELTA^<17>-6-keto-PGF1alpha, a urinary metabolite of PGI3, is developped. An authentic 2,3-dinor-DELTA^<17>-6-keto-PGF1alpha was preparedfrom DELTA^<17>-6-keto-PGF1alpha incubated with homogenate of rat liver. [^<18>O] DELTA^<17>-6-keto-PGF1alpha was synthesized by repeating base-catalyzed hydrolysis of methyl ester derivatives in [^<18>O] water, to obtain an internal standard in gas chromatography/selected ion monitoring (GC/SIM) of DELTA^<17>-6-keto-PGF1alpha. Good linear response over the range of 10 pg-10ng was demonstrated. Chromatographic conditions using a MP-65HT column presented nearly baseline separation of DELTA^<17>-6-keto-PGF1alpha and 6-keto-PGF1alpha. We were able to detect DELTA^<17>-6-keto-PGF1alpha in the range from 6 to 26 pg/ml of the human plasma. The present method can be applied to the determination of DELTA^<17>-6-keto-PGF1alpha in the human urine and plasma. Less

1.本文报道了人尿中11-脱氢血栓烷B_3（11-dehydro-TXB_3）的微量测定。我们制备了[^<18>O] 11-脱氢-血栓烷B3用作内标（IS）。通过<18>使用Sep Pak tC 18和硅胶柱的色谱样品制备来提取向其中加入[2 O]类似物的样品。通过气相色谱/高分辨率选择离子监测（GC/HR-SIM）将提取的11-脱氢血栓烷B3转化为1-甲酯（ME）-11-正丙酰胺（PA）-甲肟-9，11，15-三-二甲基异丙基甲硅烷基（DMIPS）醚衍生物。在GC/HR-SIM分析过程中，使用MP-65 HT色谱柱消除尿液基质中的干扰物质。在10 pg-100 ng/管范围内线性良好。我们能够检测到11-脱氢-TXB_3的范围从26到375 pg/ml的人尿。本方法可用于人尿中11-去氢血栓素B_3的测定。2.2，3-二诺-6-酮基丙酸的微量分析 ...更多信息 本文报道了人尿中标记物F1 α（I）的测定。加入[^2H4] -类似物作为内标物的样品通过使用Bond Elut C18和硅胶的色谱样品制备进行提取。将萃取的I转化为1-甲酯-6-甲肟-9，11，15-三-二甲基-异丙基甲硅烷基醚衍生物，随后进行气相色谱/高分辨率选择离子监测（GC/HR-SIM）。采用GC/HR-SIM分析法消除尿基质中的干扰物质，建立了尿中前列腺素I3代谢产物2，3-二去甲-Δ ^<17>-6-酮-前列腺素F1 α的微量测定方法。用大鼠肝匀浆孵育DELTA^-6-keto-PGF 1 α，得到了一种真正的2，3-dinor-DELTA <17>^-6-keto-PGF 1 <17>α。通过<18><17>重复在[^ O]水中的甲酯衍生物的碱催化水解来合成[^ O] DELTA^-6-keto-PGF 1 α<18>，以获得DELTA^-6-keto-PGF 1 α的气相色谱/选择离子监测（GC/SIM）中的内标物<17>。线性范围为10 pg-10 ng。使用MP-65 HT柱的色谱条件显示DELTA^<17>-6-keto-PGF 1 α和6-keto-PGF 1 α几乎基线分离。我们能够<17>在6 - 26 pg/ml的人血浆中检测到DELTA^-6-keto-PGF 1 α。本方法可用于<17>人尿液和血浆中DELTA^-6-keto-PGF 1 α的测定。少

项目成果

石橋正兀: "Analysis of 11-dehydro-TXB_2 in human urine improved by use of GC/HR-SIM with ^<18>O_2-labelled annlogue as an internal standard." Biol.Mass.Spectrometr.23. 612-620 (1994)

Masaki Ishibashi：“通过使用 GC/HR-SIM 以^ 18 O_2 标记的环类似物作为内标，改进了人尿液中 11-脱氢-TXB_2 的分析。”612-620。 1994）

Masadaka Ishibasi: "Analysis of ll-dehydro thromboxane B_2 in human urive improved by use of GC/HR-SIM with ^<18>O-labelled analogue as an IS." Biochem Mass Spectrm. 23. 612-620 (1994)

Masadaka Ishibasi：“通过使用 GC/HR-SIM 以 18 O 标记类似物作为 IS 改进了人尿中 11-脱氢血栓素 B_2 的分析。”

Michinao Mizugaki: "Microanalysisi of 2,3-dinor-6-keto-PGFld in human urire using GC/HR-SIM" J.Chromatogr. 658. 11-19 (1994)

Michinao Mizugaki：“使用 GC/HR-SIM 对人尿中 2,3-dinor-6-keto-PGFld 进行微量分析”J.Chromatogr。

Takanori Hishinuma: "Microdeterwination of the PGB_1 in human plasma by GC-SIM using [^<18>O] PGB_1 as an internal standard" Prostaglandins. (in press). (1995)

Takanori Hishinuma：“使用[^ 18 O] PGB_1作为内标，通过GC-SIM对人血浆中的PGB_1进行微量测定”前列腺素。

水柿道直: "Microanalysis of 2,3-dinor-6-keto-PGF_<1α> in human urine using GC/HR-SIM" J.Chromatogr.658. 11-19 (1994)

Michinao Mizukaki：“使用 GC/HR-SIM 对人尿液中的 2,3-dinor-6-keto-PGF_<1α> 进行微量分析”J.Chromatogr.658 (1994)。