Potential functionality and digestibility of oryzanol as determined using in vitro cell culture models

A mouse lymphatic endothelial cell (SVEC4-10) model and a human intestinal cell (C2BBe1) model in vitro were developed and capable to be used to study antioxidant activity, hypocholesterolemic capability and digestibility of γ-oryzanol. The critical and vital parameters in developing these cell models were the emulsion preparation of hydrophobic compounds for cell models, the consistent management of cell culture, and the selection of cell viability detection methods compatible with the cell lines and the test substances. The results showed that, in some situations, γ-oryzanol could present a more effective antioxidant activity than α-tocopherol, in terms of reducing tert-butyl hydroperoxide promoted oxidative damage on cellular mitochondrial activity. After 1-hour oxidation, cell viability was 81.8% when incubated with γ-oryzanol, compared to 54.5% with the control and 74.6% with α-tocopherol. The three major components of γ-oryzanol, cycloartenyl ferulate, 24-methylene cycloartanyl ferulate and campesteryl ferulate, generally had higher antioxidant activity than γ-oryzanol and among them, 24-methylene cycloartanyl ferulate was found to be relatively more effective and could be more powerful than α-tocopherol. A synergistic antioxidant activity among γ-oryzanol, ferulic acid and α-tocopherol was also found. With regard to the hypocholesterolemic capacity of γ-oryzanol, the results suggested that the intact γ-oryzanol was poorly absorbed by intestinal cells in vitro. Therefore, it was speculated that the effect of γ-oryzanol might take place in the lumen of gastrointestinal tract, possibly by means of reducing the micellar solubility of cholesterol and cholesteryl esters and inhibiting cholesterol esterase-facilitated cholesteryl ester hydrolysis. The results showed that preincubation of cholesterol and γ-oryzanol for six hours significantly reduced the cholesterol uptake into cells. γ-Oryzanol also showed a trend towards inhibition of cholesteryl esterase, which is responsible for hydrolyzing cholesteryl esters to free cholesterol before uptake. The results of in vitro digestibility studies of γ-oryzanol showed that the ester bond of γ-oryzanol, especially campesteryl ferulate and sitosteryl ferulate, was broken down by cholesterol esterase and produced triterpene alcohols or sterols and ferulic acid, which were further degraded. The degradation of γ-oryzanol in the stomach may undergo a different pathway because sterols or triterpene alcohols were also not found after peptic digestion.