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Combination of Injection Volume Calibration by Creatinine and MS Signals’ Normalization to Overcome Urine Variability in LC-MS-Based Metabolomics Studies
journal contribution
posted on 2013-08-20, 00:00 authored by Yanhua Chen, Guoqing Shen, Ruiping Zhang, Jiuming He, Yi Zhang, Jing Xu, Wei Yang, Xiaoguang Chen, Yongmei Song, Zeper AblizIt is essential to choose one preprocessing
method for liquid chromatography–mass
spectrometry (LC-MS)-based metabolomics studies of urine samples in
order to overcome their variability. However, the commonly used normalization
methods do not substantially reduce the high variabilities arising
from differences in urine concentration, especially for signal saturation
(abundant metabolites exceed the dynamic range of the instrumentation)
or missing values. Herein, a simple preacquisition strategy based
on differential injection volumes calibrated by creatinine (to reduce
the concentration differences between the samples), combined with
normalization to “total useful MS signals” or “all
MS signals”, is proposed to overcome urine variabilities. This
strategy was first systematically compared with other popular normalization
methods by application to serially diluted urine samples. Then, the
method has been verified using rat urine samples of pre- and postinoculation
of Walker 256 carcinoma cells. The results showed that the calibration
of injection volumes based on creatinine values could effectively
eliminate intragroup differences caused by variations in the concentrations
of urinary metabolites, thus giving better parallelism and clustering
effects. In addition, peak area normalization could further eliminate
intraclass differences. Therefore, the strategy of combining peak
area normalization with calibration of injection volumes of urine
samples based on their creatinine values is effective for solving
problems associated with urinary metabolomics.