Soo Hyun Chung

Synthesis of conjugated linoleic acid by human-derived Bifidobacterium breve LMC 017: utilization as a functional starter culture for milk fermentation.

This study was performed to discover bifidobacteria isolated from human intestines that optimally convert linoleic acid (LA) to conjugated linoleic acid (CLA) and to optimize the culture conditions of milk fermentation. One hundred and fifty neonatal bifidobacteria were screened for CLA-producing ability, and Bifidobacterium breve LMC 017 was selected as it showed about 90% conversion of free LA in MRS broth. The selected strain showed resistance at 0.5% LA in microaerophillic conditions. When monolinolein (LA 90%) was used as a substrate for CLA production, the conversion rate was lower compared to free LA, but the growth rate was unaffected during the milk fermentation. There was no significant difference in CLA production between aerobic and anaerobic conditions, and little decline in CLA was shown after the maximal CLA level had been reached. CLA production increased by 80% with 24 h of incubation in milk containing additional skim milk (5%), where the proteins may have facilitated the production of CLA by enhancing the interaction of substrate with the bacteria. CLA production did not decline after 9 h of fermentation and an additional 12 weeks of storage with other commercial starters. This demonstrates the possibility of using this strain as a costarter in the production of CLA-enriched yogurt.

Characterization of conjugated linoleic acid production by Bifidobacterium breve LMC 520.

This study was performed to characterize the CLA production ability of a bacterial strain, Bifidobacterium breve LMC 520, which can actively convert linoleic acid (LA) to cis-9,trans-11 conjugated linoleic acid (CLA), a major isomer derived from microbial enzymatic conversion. The culture conditions were optimized to improve CLA production under the aerobic conditions. B. breve LMC 520 was tested with different amounts of LA in varied culture conditions, such as air, additives, and pH. A maximal level of CLA production (up to 90% of substrate) was obtained after 24 h of incubation in culture medium containing 1 mM LA at pH 5.5 and under anaerobic conditions. There was no decline in the CLA level with prolonged incubation until 48 h. When the effect of pre-incubation with LA on CLA production was tested, there was no significant difference between the CLA-producing activity of pre-incubated and untreated bacteria at the third passage but there was a significant reduction in CLA production by the pre-incubated cells after the fourth passage. These results demonstrate that the CLA-producing activity of B. breve LMC 520 could be maximized by numerous environmental factors. The data also indicate its potential for increasing CLA accumulation in dairy products when B. breve LMC 520 is used as a functional starter culture.

Behaviour of Aspergillus flavus and Fusarium graminearum on rice as affected by degree of milling, temperature, and relative humidity during storage

We investigated the survival and growth patterns of Aspergillus flavus and Fusarium graminearum, as well as mycotoxin production, on Korean rice as affected by the degree of milling (rough, brown, and white rice) and storage conditions (21 °C/85% relative humidity [RH], 21 °C/97% RH, and 30 °C/85% RH). When rice was stored at 21 °C/85% RH, the population of A. flavus remained constant and aflatoxin was not produced, regardless of the degree of milling. At 21 °C/97% RH and 30 °C/85% RH, the populations of A. flavus increased significantly (P ≤ 0.05) and aflatoxins were produced. The highest population of A. flavus and highest amount of aflatoxin B1 were observed on brown rice stored at 21 °C/97% RH. For F. graminearum, when stored at 85% RH, the populations were reduced to less than a detectable level (5 CFU/g of rice) within 120 days and no deoxynivalenol (DON) was produced, regardless of the degree of milling and storage temperature. However, at 21 °C/97% RH, the population of F. graminearum increased significantly (P ≤ 0.05) and DON was produced on all types of rice. Findings from this study provide insights concerning storage conditions necessary to prevent growth and mycotoxin production by A. flavus and F. graminearum on Korean rice with different degrees of milling.

The proportion of non-aflatoxigenic strains of the Aspergillus flavus/oryzae complex from meju by analyses of the aflatoxin biosynthetic genes

Strains of the Aspergillus flavus/oryzae complex are frequently isolated from meju, a fermented soybean product, that is used as the starting material for ganjang (soy sauce) and doenjang (soybean paste) production. In this study, we examined the aflatoxin producing capacity of A. flavus/oryzae strains isolated from meju. 192 strains of A. flavus/oryzae were isolated from more than 100 meju samples collected from diverse regions of Korea from 2008 to 2011, and the norB-cypA, omtA, and aflR genes in the aflatoxin biosynthesis gene cluster were analyzed. We found that 178 strains (92.7%) belonged to non-aflatoxigenic group (Type I of norB-cypA, IB-L-B-, IC-AO, or IA-L-B- of omtA, and AO type of aflR), and 14 strains (7.3%) belonged to aflatoxin-producible group (Type II of norB-cypA, IC-L-B+/B- or IC-L-B+ of omtA, and AF type of aflR). Only 7 strains (3.6%) in the aflatoxin-producible group produced aflatoxins on Czapek yeast-extract medium. The aflatoxin-producing capability of A. flavus/oryzae strains from other sources in Korea were also investigated, and 92.9% (52/56) strains from air, 93.9% (31/33) strains from rice straw, 91.7% (11/12) strains from soybean, 81.3% (13/16) strains from corn, 82% (41/50) strains from peanut, and 73.2% (41/56) strains from arable soil were included in the non-aflatoxigenic group. The proportion of non-aflatoxigenicity of meju strains was similar to that of strains from soybean, air and rice straw, all of which have an effect on the fermentation of meju. The data suggest that meju does not have a preference for non-aflatoxigenic or aflatoxin-producible strains of A. flavus/oryzae from the environment of meju. The non-aflatoxigenic meju strains are proposed to be named A. oryzae, while the meju strains that can produce aflatoxins should be referred to A. flavus in this study.

Natural occurrence of type-B trichothecene mycotoxins in Korean cereal-based products

Type-B trichothecenes (deoxynivalenol (DON), nivalenol (NIV), fusarenone-X (FUS-X), 15-acetyldeoxynivalenol (15ADON), and 3-acetyldeoxynivalenol (3ADON)) were determined in 338 cereal-based products. Detection limit, quantification limit and mean recovery for five toxins were in the ranges 0.7–2.6 µg kg−1, 2.1–7.8 µg kg−1 and 73–110%, respectively. The range of occurrence and average level in samples were, respectively, 21–88% and 5.2–121.8 µg kg−1 for NIV, 10–96% and 1.7–109.5 µg kg−1 for DON, 2–39% and 0.4–3.6 µg kg−1 for FUS-X, 0–80% and 0–17.3 µg kg−1 for 15ADON, and 0–29% and 0–1.5 µg kg−1 for 3ADON. Regarding co-occurrence, 64% of samples had more than two type-B trichothecenes. The estimated daily intakes of NIV, DON, FUS-X, 15ADON, and 3ADON were 0.077, 0.048, 0.004, 0.006 and 0.002 µg kg−1 bw day−1, respectively. These results suggest that current exposure levels do not indicate the possibility of adverse effects, but consideration of the combined exposure of type-B trichothecenes may be required due to the high frequency of co-occurrence.

Utilization of Monolinolein as a Substrate for Conjugated Linoleic Acid Production by Bifidobacterium breve LMC 520 of Human Neonatal Origin

This study was designed to isolate bifidobacteria from human intestines that efficiently converts monolinolein, a monoglyceride form of linoleic acid, into conjugated linoleic acid (CLA), as well as to optimize culture conditions for improving CLA production during milk fermentation. Among 150 screened neonatal bifidobacteria, Bifidobacterium breve LMC 520 showed the highest CLA-producing ability and was tested with different types of fat substrates at various concentrations to determine the optimal conditions for CLA production. Monolinolein was tested as a substrate for CLA production. The incubation time optimized for CLA production was 24 h, and CLA production was proportionally increased with monolinolein concentration. The incubation of LMC 520 with commercial starter strains caused minimal reduction in CLA production. Our results demonstrate that the CLA-producing ability of B. breve LMC 520 could offer beneficial effects when utilized as a starter culture for the development of functional dairy products.

Simultaneous Determination of Multi-Mycotoxins in Cereal Grains Collected from South Korea by LC/MS/MS

An improved analytical method compared with conventional ones was developed for simultaneous determination of 13 mycotoxins (deoxynivalenol, nivalenol, 3-acetylnivalenol, aflatoxin B1, aflatoxin B2, aflatoxin G1, aflatoxin G2, fumonisin B1, fumonisin B2, T-2, HT-2, zearalenone, and ochratoxin A) in cereal grains by liquid chromatography-tandem mass spectrometry (LC/MS/MS) after a single immunoaffinity column clean-up. The method showed a good linearity, sensitivity, specificity, and accuracy in mycotoxin determination by LC/MS/MS. The levels of 13 mycotoxins in 5 types of commercial grains (brown rice, maize, millet, sorghum, and mixed cereal) from South Korea were determined in a total of 507 cereal grains. Mycotoxins produced from Fusarium sp. (fumonisins, deoxynivalenol, nivalenol, and zearalenone) were more frequently (more than 5%) and concurrently detected in all cereal grains along with higher mean levels (4.3–161.0 ng/g) in positive samples than other toxins such as aflatoxins and ochratoxin A (less than 9% and below 5.2 ng/g in positive samples) from other fungal species.

Simple high-performance liquid chromatography method for the simultaneous analysis of aflatoxins, ochratoxin A, and zearalenone in dried and ground red pepper.

Aflatoxins B1, B2, G1, G2 (AFB1, AFB2, AFG1, AFG2), ochratoxin A (OTA), and zearalenone (ZEA) in dried and ground red pepper (Capsicum annuum) were simultaneously analyzed with high-performance liquid chromatography coupled to fluorescence detection after post-column derivatization. The analytical method was validated for specificity, selectivity, linearity, limits of detection and quantification, recovery, precision, and measurement of uncertainty. The limits of detection and quantification were 0.10 and 0.25 μg/kg for AFB1, 0.04 and 0.06 μg/kg for AFB2, 0.14 and 0.50 μg/kg for AFG1, 0.05 and 0.10 μg/kg for AFG2, 0.12 and 0.45 μg/kg for OTA, and 4.00 and 13.25 μg/kg for ZEA, respectively. The average recoveries ranged from 80.4 to 98.5% for different concentrations of AFB1, AFB2, AFG1, AFG2, OTA, and ZEA in spiked samples. The measurement uncertainties were 0.64 to 1.62 μg/kg for AFB1, 0.24 to 0.45 μg/kg for AFB2, 0.79 to 2.19 μg/kg for AFG1, 0.32 to 0.61 μg/kg for AFG2, 0.81 to 2.31 μg/kg for OTA, and 8.48 to 26.25 μg/kg for ZEA. This method was successfully applied for the simultaneous determination of mycotoxins for 78 red peppers collected from Korean and Indian markets. Aflatoxins (sum of AFB1, AFB2, AFG1, and AFG2) were detected in 2% of nonpacked samples (n = 23) and 43% of packed samples (n = 55), at levels of 0.04 to 38.03 μg/kg. OTA was detected in 4% of nonpacked samples and 48% of packed samples, at levels of 0.15 to 56.30 μg/kg. ZEA was not detected in any samples. These findings indicate that the analytical method described here is suitable for the routine determination of the amounts of AFs, OTA, and ZEA in dried and ground red pepper.

Occurrence of aflatoxin and aflatoxigenic Aspergillus species in corn harvested in Korea

Sixty six corn samples freshly harvested in Korea were analyzed for the occurrence of aflatoxin and aflatoxigenic Aspergillus using chromatographic and multiplex polymerase chain reaction (PCR) methods. Aflatoxin and aflatoxigenic Aspergillus were detected in 13.6% (0.02 to 0.48 μg kg−1) and 3.0% of the corn samples, respectively. Aflatoxigenic Aspergillus isolates and A. flavus KCCM60330 showed high similarity (98–98.8%). These results suggest that occurrence of aflatoxin and aflatoxigenic Aspergillus in corn harvested from Korea is low.

Production of Conjugated Linoleic Acid (CLA) by Bifidobacterium breve LMC520 and Its Compatibility with CLA-Producing Rumen Bacteria

This study was performed to characterize the ability of an active Bifidobacterium strain to produce conjugated linoleic acid (CLA) and to test its possible utilization as a probiotic compatible to the ruminal condition. Bifidobacterium breve LMC520 can actively convert linoleic acid (LA) to cis-9,trans-11-CLA, which is a major isomer derived from microbial conversion. LMC520 showed reasonable tolerance under acidic conditions (pH 2.5 with 1% pepsin) and in the presence of oxgall (0-3%). The growth and CLA production of LMC520 were tested under ruminal conditions and compared with those of Butyrivibrio fibrisolvens A38, which is a major CLA producer in the rumen as an intermediate in the biohydrogenation (BH) process. LMC520 converted 15% of LA to CLA under ruminal conditions, which was 2 times higher activity than that of A38, and there was no decline in CLA level during prolonged incubation of 48 h. The BH activity of LMC520 was comparable to that of A38. When LMC520 was cocultured with A38, even with slight decrease of CLA due to high BH activity by A38, but the level of CLA was maintained by the high CLA-producing activity of LMC520. This comparative study shows the potential of this strain to be applied as a functional probiotic not only for humans but also for ruminants as well as to increase CLA production.