High-resolution mass spectrometry is a powerful tool in clinical analysis but remains less explored due to its lower dynamic range and sensitivity compared to triple quadrupoles. Glycated hemoglobin (HbA1c) is the current gold standard biomarker to monitor the control of diabetes, representing long-term plasma glycemic levels. Due to its clinical importance, several methods have been developed for HbA1c quantification, using different principles; however, the results obtained with these techniques may differ according to the method adopted. Hence, there is a great need to standardize the current methods to quantify glycated hemoglobin. A new UPLC-QToF-MS method was fully validated and tested to quantify HbA1c in human samples. The peptides VHLTPE m/z 695.373 and gly-VHLTPE m/z 857.426, obtained via Glu-C digestion, were the selected peptides for quantification of HbA1c (mmol/mol). Chromatographic separation was obtained in a C18 column, maintained at 40 °C. The mobile phase was composed of water and acetonitrile, both containing 0.02% TFA and 0.1% acetic acid, and eluted in gradient mode. The method was fully validated, being considered linear in the range of 25-107 mmol/mol of HbA1c, and was sensitive, selective, precise, accurate, and free of matrix and carryover effects. The method was successfully applied to real samples, reaching about 90% agreement with reference method results, providing accurate and precise information on peptide mass, without laborious sample preparation. These results support the use of HRMS to improve the quality of quantitative results of HbA1c in health services and demonstrate a possible application of peptide investigation for clinical analysis in the near future.

中文翻译：

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通过LC-QToF-MS量化糖化血红蛋白的新验证方法：HRMS是否能够量化临床样品？

高分辨率质谱法是临床分析中的强大工具，但与三重四极杆相比，由于其较低的动态范围和灵敏度，因此仍未得到足够的研究。糖化血红蛋白（HbA1c）是目前监测糖尿病控制的金标准生物标志物，代表长期血浆血糖水平。由于其临床重要性，已经开发了几种使用不同原理进行HbA1c定量的方法。但是，根据所采用的方法，使用这些技术获得的结果可能会有所不同。因此，非常需要标准化当前定量糖化血红蛋白的方法。一种新的UPLC-QToF-MS方法已得到充分验证和测试，可以定量分析人类样品中的HbA1c。通过Glu-C消化获得的肽VHLTPE m / z 695.373和gly-VHLTPE m / z 857.426，是用于定量HbA1c（mmol / mol）的所选肽。在保持在40°C的C18色谱柱中进行色谱分离。流动相由水和乙腈组成，水和乙腈均含有0.02％TFA和0.1％乙酸，并以梯度模式洗脱。该方法经过充分验证，被认为在25-107 mmol / mol HbA1c范围内是线性的，并且灵敏，选择性，精确，准确，并且没有基质和残留效应。该方法已成功应用于实际样品，与参考方法结果达到约90％的一致性，可提供有关肽质量的准确信息，而无需费力的样品制备。