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Methods: The fresh aerial part of green Salicornia herbacea L. was extracted with methanol and the extraction was sequentially fractionated using hexane, chloroform, ethyl acetate and butanol. 25µM acrolein was added to the A549 human lung carcinoma and HepG2 human hepatoma cells for one hour, followed by washing with PBS. Cells were incubated in new media with fractionation extracts of Salicornia herbacea L. at various concentrations (0 to 100 µg/mL), and cell viability was estimated by MTT assay. Inhibitory rates against acrolein-induced cytotoxicity were calculated. Results: In vitro, exposure of A549 human lung carcinoma and HepG2 human hepatoma cells to acrolein resulted in time- and concentration-dependent cell death. Among the fraction extracts tested in the present study, the fraction of Salicornia herbacea L. extracted with butanol resulted in complete recovery of the acrolein-induced cytotoxicity in both A549 and HepG2 cells at 100 µg/mL concentration. Significance: The present work suggests that butanol fractions of Salicornia herbacea L. might be used to reduce the cytotoxicity due to the acrolein produced during food processing. Further study is needed to evaluate various functional effects of Salicornia herbacea L., against various cells. P2-71 Establishment and Validation of an Analytical Method for Detection of Zearalenone in Medical Herbs by HPLC DSC NEEMA KASSIM, Kyeongyeol Kim, Liu Qing, Won B. Shim and Duck H. Chung Gyeongsang National University, Division of Applied Life Science (BK 21 program), Jinju, Korea, South Introduction: Fusarium species have been known to be causative agents of tricothecene mycotoxins. F. graminearum, F. culmorum and F. crookwellense produce a life threatening estrogenic toxin called zearalenone in foods, and feeds as well as in medical herbs. Purpose: For the quantitative detection of zearalenone in medical herbs, this study established and validated an analytical method using high performance liquid chromatography fluorescence detection (HPLC-FLD) after immunoaffinity column clean up. Methods: Two clean up procedures based on Solid Phase Extraction (SPE) and immunoaffinity columns after extraction with acetonitrile/methanol water at different concentrations and different dilution buffers (distilled water, phosphate buffer saline with / without Tween 20 and phosphate buffer pH. 8.0) were compared. Samples were extracted with NaCl, 80% MeOH, diluted with 2% Tween 20 in phosphate buffer saline (PBST), and cleaned up through SPE cartridges and immunoaffinity columns prior to HPLC analysis. For validation, medical herb samples (Gwal-loo-in, Doo-choong, Bokboon-ja, Sook-ji hwang, Shin-E) were artificially contaminated with zearalenone at 100, 200 and 500 ng/g, to evaluate precision and accuracy. Specificity, range, limits of detection (LOD) and quantification (LOQ) were determined on chromatograms of zearalenone artificially contaminated samples. Linearity (R2) of the standard curve (5, 10, 20, 50, 100, 200, 500, 1000 ng/g) was also evaluated. Results: Immunoaffinity columns after extraction with 80% MeOH and dilution with 2% Tween 20 in PBS, provided precise, accurate and consistent results in comparison to SPE cartridges. Linearity (R2) of the standard curve was 0.9995, and precision (relative standard deviation: 0.39 to 4.40) and accuracy (recovery: 76.59 to 105.95 %) were in the acceptable range of 70 to 120%. LOD and LOQ were 5 and 8 ng/g respectively, with an analytical detectable range from 5 to 2000 ng/g. Significance: Since there is insufficient information on the natural occurrence of zearalenone in medical herbs, the established validated method may be a useful basis for zearalenone surveillance studies. P2-72 Study on Control of Aspergillus spp. and Aflatoxin B1 in Feed by Gamma Irradiation DSC BO R. NAM, Won B. Shim, Kyeongyeol Kim, Jae H. Kim, Ju W. Lee, Myung W. Byun and Duck H. Chung Gyeongsang National University, Division of Applied Life Science (BK 21 program), Jinju, Korea, South Introduction: Aspergillus spp. are widespread in Korea and all over the world on feeds and foods. They have been known to produce aflatoxins, which are mutagenic and carcinogenic to humans. Purpose: The objective of this study was to determine the optimum gamma-irradiation dose for reduction and control of Aspergillus spp. and aflatoxin B1 in feeds. Methods: The suspension of fungal spore (107 spores/ml) was prepared in 0.85% NaCl containing 0.1% Tween 80 by a cheese cloth method and exposed to 0, 0.5, 1, 2 and 3 kGy of gamma-irradiation, followed by spreading on potato dextrose agar (PDA). The optimum gamma-irradiation dose to reduce fungal growth was estimated by checking fungal colony and size on PDA and AFB1 producibility in culture media. AFB1 level in culture media were tested by direct competitiveenzyme linked immunosorbent assay (DC-ELISA). To confirm growth inhibition of fungi on feeds by gamma-irradiation, the feeds artificially inoculated with Aspergillus spp. (107 spore/5 g) were prepared and exposed, with radiation ranging from 0 to 3 kGy. Additionally, aqueous solutions of AFB1 (0~5 ng/ml) were irradiated up to 5 kGy and then the AFB1 level was determined by DC-ELISA. Results: Gamma-irradiation has effectively inhibited fungal growth. In case of fungal spores in aqueous solution, fungal growth appeared on PDA when it was exposed to ≤1 kGy of gamma-irradiation. However, no fungal growth was observed with irradiation ≥ 2 kGy. With ≥ 2 kGy of gamma-irradiation fungal spores artificially inoculated in feed were completely inactivated. The effectiveness of gamma-irradiation to aqueous solution and feed was the same. The fungal spores irradiated with ≥ 2 kGy produced no AFB1 (< 0.6 ng/ml), whereas ≥1 0 ng/ml of AFB1 was detected in fungal spores cultivated without gamma irradiation. AFB1 level at 5 ng/ml was reduced approximately by 85% using a minimal irradiation dose (2 kGy). The results indicate that Aspergillus spp. and AFB1 on feed could be controlled by 2 kGy of gamma-irradiation. Significance: Gamma-irradiation has proved to be an effective and safe treatment that could be used as a method to reduce fungal growth and AFB1 in feeds. 130 — Abstract Book Poster
IAFP 2009 Program Book and Abstract Book
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