With ageing comes a deterioration of metabolic and physiological changes that often manifest themselves as age-related comorbidities. Calorie restriction (CR) is a robust intervention that can prevent and reverse such changes, resulting in reduced ageassociated disease and increased lifespan across a wide range of species. Moreover, a link between the extent of restriction and increased lifespan has also been established. Though widely studied, the mechanisms behind the beneficial effects of CR have yet to be fully understood. Consequently, I investigated metabolomic changes in the liver, plasma, brown adipose tissue (BAT) and cerebellum in five month old male C57BL/6 mice undergoing three months of either 10, 20, 30 or 40% CR, in addition to 12 hour and 24 hour ad libitum fed groups. Behavioural, physiological and molecular data was collected on each individual mouse and I used this information, in addition to my own metabolomic data to determine associations between phenotypic changes with graded CR. My results indicate that increasing CR resulted in greater numbers of significantly differentiated metabolites across all four tissues, and these were related to changes across sphingolipids, carnitines, bile acids, vitamins and amino acids. Metabolic remodelling in the liver indicated a shift from lipogenesis to lipolysis and changes in the plasma indicated an increase in absorption of vitamins from the stomach and colon. Changes in neurotransmitters and their precursors suggested activity and temperature driven BAT activation, in addition to an increase in antioxidant power, this was also seen in the cerebellum where metabolites associated with signalling in the hypothalamus were increased in a graded fashion with CR. In all tissues changes were linked with behaviours that accompany hunger signalling such as increased food anticipatory activity and reduced body temperature. Together, these changes reflect multi-tissue beneficial effects of CR, which may function to alleviate age-related comorbidities.