Preterm prelabour rupture of membranes (PPROM) complicated by intra-amniotic inflammation (IAI) represents a substantial portion of preterm birth cases. Currently, IAI is frequently defined as amniotic fluid IL-6 concentration above 2600 pg/mL However, the amniotic fluid IL-6 concentration was never correlated with the global response of other proinflammatory proteins to the ongoing IAI. In this cross-sectional study, protein quantification was performed using mass spectrometry (MS) analysis with isobaric peptide labeling followed by target quantification of selected proinflammatory proteins. Levels of amniotic fluid proteins determined by MS were put into the correlation with IL-6 concentration determined by ECLIA. In total, 925 proteins were efficiently quantified in all samples and differential expression analysis revealed 378 proteins upregulated towards IL-6 concentration above 10 000 pg/mL. Four proteins were selected to verify the achieved results (LCN2, MMP8, MPO, and S100A12). ROC curves were designed and the IL-6 concentration of 10 000 pg/mL was determined as cutoff value for global IAI response.