A Complete
Pipeline for Untargeted Urinary Volatolomic
Profiling with Sorptive Extraction and Dual Polar and Nonpolar Column
Methodologies Coupled with Gas Chromatography Time-of-Flight Mass
Spectrometry
posted on 2023-01-05, 13:04authored byQing Wen, Antonis Myridakis, Piers R. Boshier, Simone Zuffa, Ilaria Belluomo, Aaron G. Parker, Sung-Tong Chin, Stephanie Hakim, Sheraz R. Markar, George B. Hanna
Volatolomics offers an opportunity for noninvasive detection
and
monitoring of human disease. While gas chromatography–mass
spectrometry (GC–MS) remains the technique of choice for analyzing
volatile organic compounds (VOCs), barriers to wider adoption in clinical
practice still exist, including: sample preparation and introduction
techniques, VOC extraction, throughput, volatolome coverage, biological
interpretation, and quality control (QC). Therefore, we developed
a complete pipeline for untargeted urinary volatolomic profiling.
We optimized a novel extraction technique using HiSorb sorptive extraction,
which exhibited high analytical performance and throughput. We achieved
a broader VOC coverage by using HiSorb coupled with a set of complementary
chromatographic methods and time-of-flight mass spectrometry. Furthermore,
we developed a data preprocessing strategy by evaluating internal
standard normalization, batch correction, and we adopted strict QC
measures including removal of nonlinearly responding, irreproducible,
or contaminated metabolic features, ensuring the acquisition of high-quality
data. The applicability of this pipeline was evaluated in a clinical
cohort consisting of pancreatic ductal adenocarcinoma (PDAC) patients
(n = 28) and controls (n = 33),
identifying four urinary candidate biomarkers (2-pentanone, hexanal,
3-hexanone, and p-cymene), which can successfully
discriminate the cancer and noncancer subjects. This study presents
an optimized, high-throughput, and quality-controlled pipeline for
untargeted urinary volatolomic profiling. Use of the pipeline to discriminate
PDAC from control subjects provides proof of principal of its clinical
utility and potential for application in future biomarker discovery
studies.