■酮体(KBs)是备用葡萄糖的重要能源,为心肌提供主要能量,有氧运动时的骨骼肌,和大脑在分解代谢期间。KBs之间的水平和关系是代谢健康和疾病的关键指标。然而,在分离异构KBs方面的挑战和对样品稳定性的担忧以前限制了他们的临床测量。
■开发了一种新颖的基于6.5分钟液相色谱-质谱的测定法,能够精确测量α-,β-和γ-羟基丁酸酯,β-羟基异丁酸酯,和乙酰乙酸酯。通过研究提取效率,该方法在人血清和血浆样品中得到了充分验证。矩阵效应,准确度,recovery,精度内和精度间,线性度定量下限(LLOQ),结转,特异性,稳定性,还有更多.从107个正常样本中,建立了所有分析物和β-羟基丁酸酯/乙酰乙酸酯比率的参考范围.
■将所有五种分析物进行色谱充分分离。对于所有KB,观察到80-120%的提取效率。使用10个随机患者样本,通过加标和恢复来评估准确性,所有KBs的平均回收率为85-115%,变异系数≤3%。日内和日间不精确的变异系数<5%,和总的不精确性是<10%。没有观察到显著的干扰。样品在冰上保持稳定长达6小时或在室温下2小时。
■所开发的方法具有高度的灵敏度和鲁棒性。它已被验证用于人血清和血浆,克服稳定性问题,提供可靠和有效的酮体定量估计。
UNASSIGNED: Ketone bodies (KBs) serve as important energy sources that spare glucose, providing the primary energy for cardiac muscle, skeletal muscle during aerobic exercise, and the brain during periods of catabolism. The levels and relationships between the KBs are critical indicators of metabolic health and disease. However, challenges in separating isomeric KBs and concerns about sample stability have previously limited their clinical measurement.
UNASSIGNED: A novel 6.5-minute liquid chromatography-mass spectrometry-based assay was developed, enabling the precise measurement of alpha-, beta- and gamma-hydroxybutyrate, beta-hydroxyisobutyrate, and
acetoacetate. This method was fully validated for human serum and plasma samples by investigating extraction efficiency, matrix effects, accuracy, recovery, intra- and inter-precision, linearity, lower limit of quantitation (LLOQ), carryover, specificity, stability, and more. From 107 normal samples, reference ranges were established for all analytes and the beta-hydroxybutyrate/
acetoacetate ratio.
UNASSIGNED: All five analytes were adequately separated chromatographically. An extraction efficiency between 80 and 120 % was observed for all KBs. Accuracy was evaluated through spike and recovery using 10 random patient samples, with an average recovery of 85-115 % for all KBs and a coefficient of variation of ≤ 3 %. Coefficients of variation for intra- and inter-day imprecision were < 5 %, and the total imprecision was < 10 %. No significant interferences were observed. Specimens remained stable for up to 6 h on ice or 2 h at room temperature.
UNASSIGNED: The developed method is highly sensitive and robust. It has been validated for use with human serum and plasma, overcoming stability concerns and providing a reliable and efficient quantitative estimation of ketone bodies.