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A practical TLC-direct bioassay method was established for screening and evaluating antioxidant activity. The method was based on series of reactions. Irradiation of linoleic acid with UV 254 nm produced lipid peroxides. Upon heating, the peroxides were converted into malondialdehyde, which combined with thiobarbituric acid to a red pigment with a maximum absorption at 535 nm. Following a TLC separation, substance with anti-lipid peroxidation activity showed white spots/zones against red background on the TLC plate. The proposed method was established by the dot blot test, and validated with respect to selectivity, sensitivity, linearity, precision, and recovery after optimizing key experimental parameters. In an exemplary screening for antioxidant constituents from Perilla fruits, two active zones were discovered and tentatively characterized as salviaflaside and rosmarinic acid by TLC-interface-ESI-MS, respectively. Further verification by HPTLC-bioassay-guided-isolation was carried out and in addition to salviaflaside and rosmarinic acid, also luteolin 5- O -β- d -glucopyranoside and 3′-dehydroxyl-rosmarinic acid-3- O -β-D-glucoside were discovered to show prominent anti-lipid peroxidation activity. [Display omitted] • A practical TLC-bioassay method was established based on anti-lipid peroxidation. • A combined technique TLC-bioassay with TLC-MS was used to quickly identify antioxidants. • Four compounds with anti-lipid peroxidation were discovered and isolated from Perilla fruits. [ABSTRACT FROM AUTHOR]

Abstract: Radix Linderae, the dry roots of Lindera aggregata (Sims) Kosterm (L. strychnifolia Vill), has been long-term used in traditional Chinese medicine for treating various diseases, and alkaloids are believed to be the main active components. Previously, we reported that the total alkaloids from Radix Linderae (TARL) could effectively alleviate inflammation and protect joints from destruction in mouse collagen-induced arthritis, an animal model of human rheumatoid arthritis (RA). To get insight into the underlying mechanisms of TARL, the present study was performed to investigate the effects of TARL on the activation of macrophages and resultant production of inflammatory mediators. In vitro, TARL concentration-dependently prevented the production of nitric oxide, interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α), as well as the expressions of iNOS, IL-1β and TNF-α mRNA in RAW 264.7 cells stimulated by lipopolysaccharid (LPS). However, it showed little effect on the production of interleukin-6 (IL-6) and the expression of IL-6 mRNA. Signal transduction studies showed that TARL significantly down-regulated the phosphorylation of extracellular signal-regulated kinase (ERK) and p38 MAP kinase rather than c-jun NH2-terminal kinase (JNK). Additionally, TARL prominently decreased LPS-induced activation of IKKα and phosphorylation of p65 on serine 276, but had little impact on the phosphorylation and degradation of IκBα. In summary, our results demonstrate that TARL exhibits inhibitory effects on the production of inflammatory mediators from macrophages via blocking NF-κB and MAPKs signaling pathways. The findings provide a plausible explanation for the therapeutic efficiency of TARL on the inflammation and joint destruction in RA. [Copyright &y& Elsevier]