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제안 방법
- 8 mg GAE/g MFE, respectively (data not shown). Antioxidant activities of MFH and MFE regarding DPPH scavenging, hydrogen peroxide scavenging, ferrous ion chelating, and reducing power were evaluated. As shown in Fig.
- Unsaturated fatty acids in cell membranes are one of the major targets of free radical-mediated oxidation. In order to investigate the effect of MFH and MFE on the inhibition of cell membrane lipid peroxidation, DPPP, a sensitive fluorescence probe, was used to determine the lipid hydroperoxide levels of RAW264.7 cells exposed to AAPH. DPPP itself is not fluorescent, but DPPP oxide the resulting product of reaction with hydroperoxide, is fluorescent with a high fluorescence yield.
- However, MFH exhibited toxicity when incubation for 48 hr at the tested concentrations. Therefore, based on this result, non-toxic concentrations and incubation time of MFH and MFE were used for all experiments.
- However, its biological and pharmacological activities are still poorly understood. Therefore, in this study, we prepared two kinds of mulberry fruit extracts using hot water and 80% ethanol, and their total phenolic content, and antioxidant activity in non-cellular and cellular system and AChE inhibitory activity were investigated.
대상 데이터
- All chemicals including 2,2-diphenyl-1-picrylhydrazyl (DPPH), gallic acid, ethylenediaminetetraacetic acid (EDTA), FolinCiocalteau’s phenol reagent, H2O2, 2,2-azino-bis(3- ethylbenzthiazoline)-6-sulfonic acid (ABTS), 2,2-azobis- (2-amidinopropane)-hydrochloride (AAPH), thio-barbituric acid (TBA), trichloroacetic acid (TCA), peroxidase, potassium ferricyanide, electric-eel AChE, ACh, 5,5'- dithiobis[2-nitrobenzoic acid] (DTNB), and ferrozine were purchased from Sigma-Aldrich (St. Louis, MO, USA).
- All experiments were performed in triplicate. The data are presented as mean±standard error (SE).
- Absorbance was measured at 720 nm using microplate reader (ELx 808TM; BioTek, Winooski, VT, USA). All tests were performed in triplicate. Gallic acid was used as a standard.
- RAW264.7 cells were obtained from the American Type Culture Collection (Manassas, VA, USA) and were grown in Dulbecco’s modified Eagle medium (DMEM) containing 10% fetal bovine serum, 100 U/mL of penicillin, and 100 µg/mL of streptomycin, and maintained at 37℃ under a humidified atmosphere with 5% CO2.
데이터처리
- Statistical significance was determined by Student’s t-test.
이론/모형
- Hydrogen peroxide scavenging activity was determined according to the method of Müller (11).
- TPCs were determined by the method of Singleton et al. (9). Briefly, 40 µL of MFH or MFE (1 mg/mL) was mixed with 200 µL FolinCiocalteau reagent and 1,160 µL of distilled water for 3 min, and then followed by 600 µL 20% sodium carbonate (Na2CO3).
- The AChE inhibition assay was conducted via the spectrophotometric method developed by Ellman et al. (17). ACh was employed as a substrate to assay the inhibition of AChE.
- The DPPH scavenging activities of MFH and MFE were measured according to the slightly modified method of Blois (10). DPPH solution (1.
- After incubation, 3 mM AAPH in PBS was added and DPPP oxide fluorescence intensity was measured after 6 hr (λexcitation=361 nm, λemission=380 nm) using a GENios® microplate reader. The fluorescence values were normalized to cell numbers using the MTT cell viability assay.
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