Heat stress (HS) negatively affects meat quality by affecting material and energy metabolism, and exploring the mechanism underlying the muscle response to chronic HS in finishing pigs is important for the global pork industry. This study investigated changes in the metabolic profiles of the longissimus dorsi (LD) muscle of finishing pigs under high temperature using ultra-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight mass spectrometry (UPLC-ESI-QTOF-MS) and multivariate data analysis (MDA). Castrated male DLY pigs (Duroc × Landrance × Yorkshire pigs, n = 24) from 8 litters were divided into three treatment groups: constant optimal ambient temperature at 22 °C and ad libitum feeding (CR, n = 8); constant high ambient temperature at 30 °C and ad libitum feeding (HS, n = 8); and constant optimal ambient temperature 22 °C and pair-feeding to the control pigs (PF, n = 8). The metabolic profile data from LD muscle samples were analyzed by MDA and external search engines. Nine differential metabolites (L-carnosine, acetylcholine, inosinic acid, L-carnitine, L-anserine, L-α-glycerylphosphorylcholine, acetylcarnitine, thiamine triphosphate, and adenosine thiamine diphosphate) were involved in antioxidant function, lipid metabolism, and cell signal transduction, which may decrease post mortem meat quality and play important roles in anti-HS. Four metabolites (L-carnosine, acetylcholine, inosinic acid, and L-carnitine) were verified, and it was indicated that the muscle L-carnitine content was significantly lower in HS than in CR (p < 0.01). The results show that constant HS affects the metabolites in the LD muscle and leads to coordinated changes in the endogenous antioxidant defense and meat quality of finishing pigs. These metabonomics results provide a basis for researching nutritional strategies to reduce the negative effects of heat stress on livestock and present new insights for further research.
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