The aims of this study were to investigate the effects of adlay testa (AT) on Cu(2+)-treated low-density lipoprotein (LDL) oxidation, 2,2'-diphenyl-1-picrylhydrazyl (DPPH)-scavenging capacity, and lipopolysaccharide (LPS)-induced inflammation in RAW 264.7 macrophages and determine its active components. The AT ethanolic extract (ATE) was partitioned into four fractions by various solvents as follows: n-hexane (ATE-Hex), ethyl acetate (ATE-Ea), n-butanol (ATE-Bu), and water (ATE-H(2)O). ATE-Ea and ATE-Bu were further fractionated into ATE-Ea-a-ATE-Ea-h and ATE-Bu-A-ATE-Bu-F, respectively, by column chromatography. Results showed that ATE-Ea, ATE-Bu, ATE-Ea-e, and ATE-Bu-C expressed antiradical, antioxidative, and anti-inflammatory activities with respect to the DPPH-scavenging capacity, LDL protection effect, and nitric oxide (NO) inhibitory activity. Inflammation was further modulated by ATE-Ea, ATE-Bu, ATE-Ea-e, and ATE-Bu-C through downregulating the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase 2 (COX-2) proteins. The following components were found in ATE-Ea-e and ATE-Bu-C after purification and high-performance liquid chromatographic analysis: chlorogenic acid (CGA), vanillic acid (VA), caffeic acid (CA), p-coumaric acid (PCA), ferulic acid (FA), and 2-O-beta-glucopyranosyl-7-methoxy-4((2)H)-benzoxazin-3-one (GMBO). Results showed that CGA, CA, and FA were the major components responsible for the antioxidative and anti-inflammatory activities of ATE-Ea-e and ATE-Bu-C. Subsequently, each gram of ATE-Bu-C had 30.3 mg of CGA, 9.02 mg of CA, and 189 mg of GMBO, while each gram of ATE-Ea-e had 1.31 mg of VA, 3.89 mg of PCA, and 47.6 microg of FA. In conclusion, ATE has antioxidative and anti-inflammatory activities, and its effects are partially related to its phenolic components. Thus, ATE has the potential to be developed as a functional food targeting chronic diseases.

We investigated the effects of adlay seed hull (AH) extracts on the lipopolysaccharide-induced inflammatory response in RAW 264.7 macrophages. An AH ethanol extract (AHE) was partitioned into ethyl acetate, n-butanol, and water fractions. Silica gel chromatography of the ethyl acetate fraction yielded 15 subfractions: AHE-Ea-A to AHE-Ea-O. Subfractions AHE-Ea-J, AHE-Ea-K, and AHE-Ea-M had anti-inflammatory activities, as they counteracted the increased cellular production of nitric oxide and prostaglandin E2 induced by lipopolysaccharide by down-regulating inducible nitric oxide synthase and cyclooxygenase 2 expression. Eriodictyol (1), the ceramide (2S,3S,4R)-2-[(2'R)-2'-hydroxytetracosanoyl-amino]-1,3,4-octadecanetriol (2), and p-coumaric acid (3) were found in the subfractions, and the first two compounds appeared to be primarily responsible for the anti-inflammatory activity. This is the first time that eriodictyol (1) and this ceramide (2) have been found in AH, and the anti-inflammatory properties of the AHE-Ea fraction can be attributed, at least in part, to the presence of these two compounds.

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