Dark, Firm and Dry (DFD) beef is a major meat quality defect that is produced by animals' subjected to pre-slaughter stress and is considered to be one of the main meat quality defects resulting in significant economic losses. Conflicting views have been reported surrounding the use of conventional markers of pre-slaughter stress and their association to meat quality and subsequently the prevalence of DFD beef remains. The overall aim of this research was to utilise targeted and untargeted analytical approaches to enhance the understanding of the physiological pre-slaughter stress response in cattle and its association to downstream meat quality. Two animal studies were conducted; one investigated the stress response of young bulls (n=298) reared and slaughter in a commercial setting and the second assessed the social disruption of young bulls (n=27) maintained under controlled experimental conditions. Studies demonstrated that conventional markers of pre-slaughter stress (urinary/blood cortisol and cortisone) were unable to differentiate between varying intensities of pre-slaughter stress and showed no clear association to meat quality (as measured by ultimate pH (pHu)) In contrast, untargeted high resolution-mass spectrometry based metabolomic profiling of bovine urine from corresponding animals revealed 15 new urinary metabolites (9 up- and 6 down-regulated) that discriminated between the different levels of pre-slaughter stress. Based on the development of a predictive model these urinary metabolites were shown to relate to carcass pHu, suggesting potential for use as a means of predicting downstream meat quality. Furthermore, metabolomic profiling of longissimus dorsi muscle tissue extracts revealed 6 up-regulated metabolites that differentiated between normal and high pHu meat. Findings from this research has revealed new metabolites that can distinguish between animals exposed to a combination of psychological and physiological stress, which will aid in mitigating against associated meat quality losses, thus leading to improved sustainability and profitability of the meat industry.