{"title":"液相色谱-串联质谱法同时测定尿中n - τ-甲基组氨酸、n -乙酰- n - τ-甲基组氨酸和肌酐的建立和验证。","authors":"Daichi Ijiri, Yoshio Nishimura, Ayumi Katafuchi, Saki Shimamoto, Miyu Kamimura, Ichiro Oshima, Namiko Nakamura, Shozo Tomonaga, Shinya Ishinhara, Akira Ohtsuka","doi":"10.1016/j.tjnut.2025.09.024","DOIUrl":null,"url":null,"abstract":"<p><strong>Background: </strong>N<sup>τ</sup>-Methylhistidine released endogenously from muscle into urine is an index of muscle protein breakdown, and part of N<sup>τ</sup>-methylhistidine it is excreted as the acetylated derivative, N-acetyl-N<sup>τ</sup>-methylhistidine).</p><p><strong>Objective: </strong>To measure the urinary concentrations of N<sup>τ</sup>-methylhistidine and N-acetyl-N<sup>τ</sup>-methylhistidine, a quantitative method was established using liquid chromatography-tandem mass spectrometry (LC-MS/MS) operated in Q1 single-ion monitoring (SIM) mode and multiple reaction monitoring (MRM) mode with stable-isotope dilution analysis.</p><p><strong>Methods: </strong>We synthesized N-acetyl-N<sup>τ</sup>-methylhistidine and its isotopic analog for stable-isotope dilution analysis. Then, we validated a method for quantifying urinary concentrations of intact N<sup>τ</sup>-methylhistidine, N-acetyl-N<sup>τ</sup>-methylhistidine, and creatinine to precisely determine their concentrations in rats, humans, cattle, and dogs. Creatinine correction was applied to normalize for urine volume.</p><p><strong>Results: </strong>For both SIM and MRM analysis, the acceptable linear ranges of detection were 5 pmol/mL to 10 nmol/mL with r<sup>2</sup> = 1.000 in SIM mode and MRM mode. The LC-MS/MS methods operated in both SIM and MRM transitions detected changes in the urinary excretion levels of N<sup>τ</sup>-methylhistidine and N-acetyl-N<sup>τ</sup>-methylhistidine in response to starvation and re-feeding of rats. The methods also detected urinary concentrations of N<sup>τ</sup>-methylhistidine and N-acetyl-N<sup>τ</sup>-methylhistidine in cattle, humans, and dogs.</p><p><strong>Conclusions: </strong>These results suggest that the LC-MS/MS method operated in SIM mode and MRM mode can be used to measure urinary concentrations of N<sup>τ</sup>-methylhistidine and N-acetyl-N<sup>τ</sup>-methylhistidine.</p>","PeriodicalId":16620,"journal":{"name":"Journal of Nutrition","volume":" ","pages":""},"PeriodicalIF":3.8000,"publicationDate":"2025-09-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Development and Validation of a Liquid Chromatography-Tandem Mass Spectrometry Method for the Simultaneous Quantification of N<sup>τ</sup>-Methylhistidine, N-Acetyl-N<sup>τ</sup>-Methylhistidine, and Creatinine in Urine.\",\"authors\":\"Daichi Ijiri, Yoshio Nishimura, Ayumi Katafuchi, Saki Shimamoto, Miyu Kamimura, Ichiro Oshima, Namiko Nakamura, Shozo Tomonaga, Shinya Ishinhara, Akira Ohtsuka\",\"doi\":\"10.1016/j.tjnut.2025.09.024\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><strong>Background: </strong>N<sup>τ</sup>-Methylhistidine released endogenously from muscle into urine is an index of muscle protein breakdown, and part of N<sup>τ</sup>-methylhistidine it is excreted as the acetylated derivative, N-acetyl-N<sup>τ</sup>-methylhistidine).</p><p><strong>Objective: </strong>To measure the urinary concentrations of N<sup>τ</sup>-methylhistidine and N-acetyl-N<sup>τ</sup>-methylhistidine, a quantitative method was established using liquid chromatography-tandem mass spectrometry (LC-MS/MS) operated in Q1 single-ion monitoring (SIM) mode and multiple reaction monitoring (MRM) mode with stable-isotope dilution analysis.</p><p><strong>Methods: </strong>We synthesized N-acetyl-N<sup>τ</sup>-methylhistidine and its isotopic analog for stable-isotope dilution analysis. Then, we validated a method for quantifying urinary concentrations of intact N<sup>τ</sup>-methylhistidine, N-acetyl-N<sup>τ</sup>-methylhistidine, and creatinine to precisely determine their concentrations in rats, humans, cattle, and dogs. Creatinine correction was applied to normalize for urine volume.</p><p><strong>Results: </strong>For both SIM and MRM analysis, the acceptable linear ranges of detection were 5 pmol/mL to 10 nmol/mL with r<sup>2</sup> = 1.000 in SIM mode and MRM mode. The LC-MS/MS methods operated in both SIM and MRM transitions detected changes in the urinary excretion levels of N<sup>τ</sup>-methylhistidine and N-acetyl-N<sup>τ</sup>-methylhistidine in response to starvation and re-feeding of rats. The methods also detected urinary concentrations of N<sup>τ</sup>-methylhistidine and N-acetyl-N<sup>τ</sup>-methylhistidine in cattle, humans, and dogs.</p><p><strong>Conclusions: </strong>These results suggest that the LC-MS/MS method operated in SIM mode and MRM mode can be used to measure urinary concentrations of N<sup>τ</sup>-methylhistidine and N-acetyl-N<sup>τ</sup>-methylhistidine.</p>\",\"PeriodicalId\":16620,\"journal\":{\"name\":\"Journal of Nutrition\",\"volume\":\" \",\"pages\":\"\"},\"PeriodicalIF\":3.8000,\"publicationDate\":\"2025-09-23\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Journal of Nutrition\",\"FirstCategoryId\":\"3\",\"ListUrlMain\":\"https://doi.org/10.1016/j.tjnut.2025.09.024\",\"RegionNum\":3,\"RegionCategory\":\"医学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q2\",\"JCRName\":\"NUTRITION & DIETETICS\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of Nutrition","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.1016/j.tjnut.2025.09.024","RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"NUTRITION & DIETETICS","Score":null,"Total":0}
引用次数: 0
摘要
背景：n τ-甲基组氨酸从肌肉内源性释放到尿液中是肌肉蛋白分解的指标，部分n τ-甲基组氨酸作为乙酰化衍生物（n -乙酰化- n τ-甲基组氨酸）排出体外。目的：建立液相色谱-串联质谱（LC-MS/MS）联用Q1单离子监测（SIM）模式和多反应监测（MRM）模式，结合稳定同位素稀释分析，测定尿中n - τ-甲基组氨酸和n -乙酰- n -甲基组氨酸浓度的定量方法。方法：合成n -乙酰基- n τ-甲基组氨酸及其同位素类似物进行稳定同位素稀释分析。然后，我们验证了一种定量尿中完整n -甲基组氨酸、n -乙酰- n -甲基组氨酸和肌酐浓度的方法，以精确测定它们在大鼠、人、牛和狗中的浓度。应用肌酐校正使尿量正常化。结果：对于SIM和MRM分析，在SIM模式和MRM模式下，可接受的线性检测范围为5 pmol/mL至10 nmol/mL， r2 = 1.000。LC-MS/MS方法在SIM和MRM转换中检测了大鼠在饥饿和再喂养时尿中n -甲基组氨酸和n -乙酰- n -甲基组氨酸水平的变化。该方法还检测了牛、人类和狗尿液中n - τ-甲基组氨酸和n -乙酰- n -甲基组氨酸的浓度。结论：LC-MS/MS方法在SIM模式和MRM模式下可用于测定尿中n - τ-甲基组氨酸和n -乙酰- n -甲基组氨酸的浓度。
Development and Validation of a Liquid Chromatography-Tandem Mass Spectrometry Method for the Simultaneous Quantification of Nτ-Methylhistidine, N-Acetyl-Nτ-Methylhistidine, and Creatinine in Urine.
Background: Nτ-Methylhistidine released endogenously from muscle into urine is an index of muscle protein breakdown, and part of Nτ-methylhistidine it is excreted as the acetylated derivative, N-acetyl-Nτ-methylhistidine).
Objective: To measure the urinary concentrations of Nτ-methylhistidine and N-acetyl-Nτ-methylhistidine, a quantitative method was established using liquid chromatography-tandem mass spectrometry (LC-MS/MS) operated in Q1 single-ion monitoring (SIM) mode and multiple reaction monitoring (MRM) mode with stable-isotope dilution analysis.
Methods: We synthesized N-acetyl-Nτ-methylhistidine and its isotopic analog for stable-isotope dilution analysis. Then, we validated a method for quantifying urinary concentrations of intact Nτ-methylhistidine, N-acetyl-Nτ-methylhistidine, and creatinine to precisely determine their concentrations in rats, humans, cattle, and dogs. Creatinine correction was applied to normalize for urine volume.
Results: For both SIM and MRM analysis, the acceptable linear ranges of detection were 5 pmol/mL to 10 nmol/mL with r2 = 1.000 in SIM mode and MRM mode. The LC-MS/MS methods operated in both SIM and MRM transitions detected changes in the urinary excretion levels of Nτ-methylhistidine and N-acetyl-Nτ-methylhistidine in response to starvation and re-feeding of rats. The methods also detected urinary concentrations of Nτ-methylhistidine and N-acetyl-Nτ-methylhistidine in cattle, humans, and dogs.
Conclusions: These results suggest that the LC-MS/MS method operated in SIM mode and MRM mode can be used to measure urinary concentrations of Nτ-methylhistidine and N-acetyl-Nτ-methylhistidine.
期刊介绍：
The Journal of Nutrition (JN/J Nutr) publishes peer-reviewed original research papers covering all aspects of experimental nutrition in humans and other animal species; special articles such as reviews and biographies of prominent nutrition scientists; and issues, opinions, and commentaries on controversial issues in nutrition. Supplements are frequently published to provide extended discussion of topics of special interest.
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