In the food sector, liposome systems are utilized to boost absorptivity, regulate active-molecule release, and enhance food quality. The efficacy of liposomes containing methyl jasmonate (0.1 mM) mixed with vitamin B12 (2 μM) (MeJA+Vit B12 liposome) on the alleviation of chilling injury (CI) in ‘Queen’ pineapples by using peduncle infiltration was investigated. The treatment of MeJA+Vit B12 liposome was compared with the treatments of MeJA, Vit B12 and MeJA+Vit B12 and untreated fruits (control) during refrigerated storage at 13°C for 14 d. The treatments of MeJA, Vit B12 and MeJA+Vit B12 liposome could alleviate CI symptoms (translucency and browning in tissue adjacent to the core). Among these treatments, MeJA+Vit B12 liposome inhibited the CI symptom more than other treatments. MeJA+Vit B12 liposome retarded the increments of tissue electrolyte leakage, malondialdehyde content, lipoxygenase activity, polyphenol oxidase activity, and total phenols content of tissue contiguous to the core of the pineapples compared to untreated fruits. The treatment also enhanced the ascorbic acid content and antioxidant activities (ferric reducing antioxidant power and free radical scavenging activity) of tissue adjacent to the core. Therefore, MeJA+Vit B12 liposome peduncle infiltration is a feasible novel approach to alleviate the CI incidence of ‘Queen’ pineapples during refrigeration.