The Anti-oxidative and Anti-inflammatory Activities of Adlay (Coix lachryma-jobi L. var. ma-yuen Stapf) Seed Extract and Its Active Components and Effect on Anti-atherogenic Risk Factors
Date Issued
2010
Date
2010
Author(s)
Huang, Din-Wen
Abstract
The aims of the first part of this study were to investigate the effects of adlay seed extracts on the copper ion-mediated low density lipoprotein (LDL) oxidation and lipopolysaccharide (LPS)-induced inflammatory response in RAW 264.7 macrophages. Adlay hull ethanolic extract (AHE) was sequentially partitioned into ethyl acetate, n-butanol, and water-soluble fractions (AHE-Ea, AHE-Bu and AHE-H2O, respectively). AHE-Ea was further separated into AHE-Ea-A to AHE-Ea-O by column chromatography (CC). AHE-Ea and its subfractions, AHE-Ea-J, AHE-Ea-K, and AHE-Ea-M inhibited the nitric oxide (NO) and prostaglandin E2 (PGE2) formation through down-regulating inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) protein expression. Eriodictyol (1), a 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 and the ceramide have been found in adlay seed hulls, and the anti-inflammatory properties of the AHE-Ea fraction can be attributed, at least in part, to the presence of these two compounds.
The adlay testa ethanolic extract (ATE) was sequentially separated into n-hexane, ethyl acetate, n-butanol and water-soluble fractions (ATE-Hex, ATE-Ea, ATE-Bu and ATE-H2O, respectively). ATE-Ea and ATE-Bu were further separated into ATE-Ea-a to ATE-Ea-h and ATE-Bu-A to ATE-Bu-F respectively by CC. Results showed that ATE-Ea, ATE-Bu, ATE-Ea-e, and ATE-Bu-C expressed anti-oxidative and anti-inflammatory activities through the DPPH-radical scavenger, LDL oxidation inhibition, and NO inhibitory activity. Inflammation was further modulated by fractions of ATE-Ea and ATE-Bu, and their subfractions, ATE-Ea-e and ATE-Bu-C through downregulating the expression of iNOS and COX-2 proteins. Chlorogenic acid (CGA), vanillic acid (VA), caffeic acid (CA), syringic acid (SA), 4-hydroxyacetophenone (4HA), p-coumaric acid (PCA), ferulic acid (FA), and 2-O-β-glucopyranosyl-7-methoxy-4 (2H)-benzoxazin-3-one (GMBO), were analyzed or isolated from the active fractions of ATE by CC. Among them, CGA, CA, and FA were the major components responsible for the anti-oxidative and anti-inflammatory activities of ATE. Subsequently, HPLC analysis showed that each gram of ATE-Bu-C had 30.3 mg CGA, 9.02 mg CA, and 189 mg GMBO, while each gram of ATE-Ea-e had 1.31 mg VA, 3.89 mg PCA, and 47.6 micro-g FA. In brief, ATE had anti-oxidative and anti-inflammatory activities, and its effects were partially related to its phenolic components.
The adlay bran ethanolic extract (ABE) and polished adlay ethanolic extract (PAE) were sequentially separated into n-hexane, ethyl acetate, n-butanol and water-soluble fractions, respectively (ABE-Hex, ABE-Ea, ABE-Bu and ABE-H2O, respectively; PAE-Hex, PAE-Ea, PAE-Bu and PAE-H2O, respectively). Results showed that，ABE-Ea and PAE-Ea inhibited the NO formation through down-regulating iNOS and COX-2 protein expression. However, the effects of both fractions of ABE and PAE on prevention of LDL oxidation were worse than adlay testa.
The second part of this study, ATE, ATE-Ea, and the self-made mixture of phenolic acid components were further fed the C57BL/6 apo E-deficient (apoE-/-) mice with high cholesterol diet (HCD) for four weeks to investigate the anti-atherogenic risk factors effects of ATE-Ea in vivo and compared with wild type C57BL/6 mice. Results showed that when the HCD was supplied to the C57BL/6 apo EapoE-/- mice with or without the various samples for 4 weeks, the serum total cholesterol concentrations (TC) were all significantly decreased in test groups. The antioxidative enzymes in liver, glutathione reductase (GRd), glutathione S-transferase (GST) and catalase (CAT) were all up-regulated by high-dose ATE-Ea group resulting in the increasing of glutathione (GSH) and the decreasing of TBARS. The liver GRd is also significantly raised by self-made mixture of phenolic acid components group (p < 0.01). The expression of intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), monocyte chemoattractant protein-1 (MCP-1), and the subunit of NADPH oxidase (NOX4 and p22phox) in the aorta were all inhibited, however, the anti-oxidative enzyme superoxide dismutase-1 (SOD-1) and SOD-2 were both up-regulated by ATE-Ea. Although the expression of inflammatory-related proteins (ICAM-1 and VCAM-1) of knockout mice aorta were not influenced by self-made mixture of phenolic acid components, the anti-oxidative enzymes were up-regulated (SOD-1 and SOD-2) and the oxidative stress-related proteins (NOX4與p22phox) were down-regulated by self-made mixture of phenolic acid components.
In conclusion, ATE had anti-oxidative, and all of the adlay seed extracts had anti-inflammatory activities, and the effects of AHE and ATE were both partially related to its components, and in vivo observations, together with the functions of ATE-Ea: (1) To lower the cholesterol level in the serum, (2) to up-regulate the antioxidative enzymes in the liver, and (3) to modulate the expression of inflammation and oxidative stress-related protein of aorta, in the pathogenesis of HCD-induced atherosclerosis in C57BL/6 apoE-/- mice.
Subjects
Adlay
antioxidative
anti-inflammatory
LDL
NO
iNOS
COX-2
atherosclerosis
C57BL/6 apoE deficient mice
File(s)![Thumbnail Image]()
Loading...
Name
ntu-99-D94641007-1.pdf
Size
23.53 KB
Format
Adobe PDF
Checksum
(MD5):01709593d5565300661fd5b91a796c4a