Obesity is associated with metabolic disorders. Fibroblast growth factor 21 (FGF21) has been recognized as important in metabolism. Glucosamine (GLN) has been demonstrated to perform diverse beneficial functions. This study aimed to reveal whether and how GLN would modulate FGF21 production in relation to metabolism. With in vivo model of normal diet (ND) and high-fat diet (HFD) mice receiving GLN injection and in vitro model of mouse AML12 liver cells and differentiated 3T3L1 adipocytes challenged with GLN, GLN appeared to improve the glucose metabolism in HFD and ND mice and to elevate FGF21 protein expression in HFD liver and to increase both FGF21 protein and mRNA levels in WAT from HFD and ND mice and it also upregulated FGF21 expression in both AML12 and differentiated 3T3L1 cells. By using inhibitors against various signaling pathways, p38, Alt, NF-kappa B, and PIGS appeared potentially involved in GLN-mediated FGF21 production in AML12 cells; GLN was able to mediate activation of NF-kappa B, p38 or PKA/CREB signaling. Our accumulated findings suggest that GLN may potentially improve the metabolic performance by inducing FGF21 production in liver and adipose tissues and such induction in liver cells may act in part due to GLN induction of the NF-kappa B, p38 and PKA pathways. (C) 2020 Elsevier Inc. All rights reserved.