Preterm birth (PTB) is a critical global health issue, contributing to significant neonatal mortality and long-term complications. Almost half of the preterm births occur due to spontaneous preterm labour (PTL). Despite substantial research efforts, our knowledge of labour onset and progression leading to either preterm and term delivery is limited. With the intent of exploring more about the physiological processes of pregnancy and labour, we performed label-free shotgun proteomics on maternal urinary samples using two different modes of mass spectrometer – data- dependent acquisition (DDA) and data-independent acquisition (DIA). The goal was to assess what approach resulted in high quality and reproducible data by detecting and quantifying large number of proteins for a discovery study using urine samples. Comparative analysis of DDA and DIA demonstrated superior performance of DIA. DIA resulted in better data in terms of consistency of protein detection, retaining higher number of total proteins and uniquely detected proteins for downstream analysis and presence of fewer number of missing values. To investigate further, we expanded our discovery cohort and performed DIA proteomics. Proteomics data analysis revealed elevated levels of Defensin alpha 1 (DEFA1) and Kallikrein 1 (KLK1) in the Labour group while the Non Labour group showed elevated levels of Endosialin (CD248) and Leucine-rich alpha-2-glycoprotein (LRG1). To identify the proteomics changes with respect to gestational age, we compared Preterm Non Labour (PTNL) group with Term Non Labour (TNL) group. The PTNL group was found significantly enriched in Fatty acid binding protein 5 (FABP5) and Keratin type 1 cytoskeletal 16 (KRT16) while TNL group showed significant enrichment in Calmodulin (CALM1). Differential levels of aforementioned proteins seem to be associated with labour and changes with gestational age. Future studies are required to validate the observed trends in protein abundances in the study groups and to determine the potential of these proteins for prediction of preterm labour or preterm birth.