Understanding oxygenate transformations during biomass valorization and bio-oil upgrade processes is critical in developing viable, sustainable, and competitive biorefineries. This study introduces a comprehensive analytical platform to clarify the intricate quantitative composition and chemistry of bio-oils across hydrodeoxygenation stages. This platform provides a detailed composition resolution at the molecular and quasi-quantitative levels by integrating data from multiple analytical techniques. The approach involves validating the results using interapparatus relationships and innate compounds as internal standards, merging them into quasi-quantitative data sets representing the entire mixture. A crucial focus of this platform is applying high-resolution mass spectrometry under various ionization conditions to elucidate high molecular weight structures, particularly those with varying oxygen-containing functional groups. This work offers insight into the complex chemical transformations in biorefinery.