Measurement of hydroxychloroquine in blood from SLE patients using LC-HRMS-evaluation of whole blood, plasma, and serum as sample matrices
Measurement of hydroxychloroquine in blood from SLE patients using LC-HRMS-evaluation of whole blood, plasma, and serum as sample matrices
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DOI：
10.1186/s13075-020-02211-1
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发表时间：
2020-06-01
影响因子：
4.9
通讯作者：
Kultima, Kim
中科院分区：
文献类型：
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作者：
Carlsson, Henrik;Hjorton, Karin;Kultima, Kim
背景羟氯喹（HCQ）是治疗系统性红斑狼疮（SLE）、类风湿性关节炎（RA）和其他炎性风湿性疾病的标准治疗药物，并可能用于COVID-19患者的治疗。用于治疗药物监测（TDM）的HCQ测定可在全血（WB）、血清和血浆中进行。在SLE患者中，WB、血清和血浆HCQ水平的直接比较以前没有报道。我们描述了一种使用液相色谱-高分辨质谱（LC-HRMS）测定人血中HCQ的方法，并比较了三种样品基质的适用性。方法采用高效液相色谱-高分辨质谱法（LC-HRMS）测定26例SLE患者全血、血清和血浆中HCQ的含量。使用质控样品和患者样品的重复制备和测量，评价了该方法在三种基质中的重现性。通过与两种先前报道的提取方法进行比较，进一步评价了所开发的方法用于血清中HCQ测量的性能。结果该方法具有较高的准确度和精密度。在所有三种基质（WB、血清和血浆）中观察到SLE患者的HCQ浓度范围很大。WB中的平均水平约为血清和血浆中水平的两倍（813 ng/mL，分别为436 ng/mL和362 ng/mL）。加标质控品显示所有基质的重现性均较高（变异系数，CV，约5%），而在患者样本中，仅使用WB作为基质发现了同样高的精密度（CV 3%）。血清和血浆的CV分别为14%和39%。用于血清样品的两种替代方法未显示出精密度的改善。结论建立了人血中HCQ的LC-HRMS测定方法。在样品重现性方面，发现全血是上级样品基质。因此，当监测患者的HCQ治疗效果时，应使用全血样本进行HCQ分析。该测定法在临床上用于监测患者的HCQ水平。
Background Hydroxychloroquine (HCQ) is the standard of care in the treatment of systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), and other inflammatory rheumatic diseases and potentially for the treatment in COVID-19 patients. Determination of HCQ for therapeutic drug monitoring (TDM) can be performed in whole blood (WB), serum, and plasma. Direct comparisons of WB, serum, and plasma levels of HCQ in patients with SLE have not previously been reported. We describe a method for the determination of HCQ in human blood using liquid chromatography-high-resolution mass spectrometry (LC-HRMS) and compare the suitability of the three sample matrices. Methods A method for the determination of HCQ in human blood using LC-HRMS was developed, validated, and applied for the determination of HCQ levels in WB, serum, and plasma from 26 SLE patients. The reproducibility of the method, in the three matrices, was evaluated using quality control samples and repeated preparations and measurements of patient samples. The performance of the developed method for HCQ measurement in serum was further evaluated by comparison with two previously reported extraction methods. Results The performance of the presented method demonstrated high accuracy and precision. A large range of HCQ concentrations was observed for the SLE patients in all three matrices (WB, serum, and plasma). The mean levels in WB were approximately two-fold the levels in serum and plasma (813 ng/mL compared to 436 ng/mL and 362 ng/mL, respectively). Spiked quality controls showed high reproducibility for all matrices (coefficient of variation, CV, approx. 5%), whereas in patient samples, equally high-precision was only found using WB as the matrix (CV 3%). The CV for serum and plasma was 14% and 39%, respectively. Two alternative methods applied to serum samples did not demonstrate improved precision. Conclusions A LC-HRMS method for the measurement of HCQ in human blood was developed and validated. Whole blood was found to be the superior sample matrix in terms of sample reproducibility. Thus, whole blood samples should be used for HCQ analysis when patients are monitored for HCQ treatment effects. The assay is in clinical use to monitor levels of HCQ in patients.