Simple detection of residual enrofloxacin in meat products using microparticles and biochips.

A simple and sensitive method for detecting enrofloxacin, a major veterinary fluoroquinolone, was developed. Monoclonal antibody specific for enrofloxacin was immobilised on a chip and fluorescent dye-labelled microparticles were covalently bound to the enrofloxacin molecules. Enrofloxacin in solution competes with the microparticle-immobilised enrofloxacin (enroMPs) to bind to the antibody on the chip. The presence of enrofloxacin was verified by detecting the fluorescence of enrofloxacin-bound microparticles. Under optimum conditions, a high dynamic range was achieved at enrofloxacin concentrations ranging from 1 to 1000 µg kg(-1). The limits of detection and quantification for standard solutions were 5 and 20 µg kg(-1) respectively, which are markedly lower than the maximum residue limit. Using simple extraction methods, recoveries from fortified beef, pork and chicken samples were 43.4-62.3%. This novel method also enabled approximate quantification of enrofloxacin concentration: the enroMP signal intensity decreased with increasing enrofloxacin concentration. Because of its sensitivity, specificity, simplicity and rapidity, the method described herein will facilitate the detection and approximate quantification of enrofloxacin residues in foods in a high-throughput manner.

FMN-Based Fluorescent Proteins as Heavy Metal Sensors Against Mercury Ions.

Bacterial light-oxygen-voltage-sensing photoreceptor-derived flavin mononucleotide (FMN)- based fluorescent proteins act as a promising distinct class of fluorescent proteins utilized for various biomedical and biotechnological applications. The key property of its independency towards oxygen for its chromophore maturation has greatly helped this protein to outperform the other fluorescent proteins such as GFP and DsRed for anaerobic applications. Here, we describe the feasibility of FMN-containing fluorescent protein FbFP as a metal-sensing probe by measuring the fluorescence emission changes of a protein with respect to the concentration of metal ions. In the present study, we demonstrated the mercury-sensing ability of FbFP protein and the possible amino acids responsible for metal binding. A ratiometric approach was employed here in order to exploit the fluorescence changes observed at two different emission maxima with respect to Hg(2+) at micromolar concentration. The engineered variant FbFPC56I showed high sensitivity towards Hg(2+) and followed a good linear relationship from 0.1 to 3 µM of Hg(2+). Thus, further engineering with a rational approach would enable the FbFP to be developed as a novel and highly selective and sensitive biosensor for other toxic heavy metal ions as well.

Temperature sensing using red fluorescent protein.

Genetically encoded fluorescent proteins are extensively utilized for labeling and imaging proteins, organelles, cell tissues, and whole organisms. In this study, we explored the feasibility of mRFP1 and its variants for measuring intracellular temperature. A linear relationship was observed between the temperature and fluorescence intensity of mRFP1 and its variants. Temperature sensitivities of E. coli expressing mRFP1, mRFP-P63A and mRFP-P63A[(4R)-FP] were -1.27%, -1.26% and -0.77%/°C, respectively. Finally, we demonstrated the potentiality of mRFP1 and its variants as an in vivo temperature sensor.

Detection of enrofloxacin and its metabolite ciprofloxacin using gold nanoparticles and laser desorption/ionization time-of-flight mass spectrometry.

This paper describes a new, simple, and sensitive method for detecting two fluoroquinolones: enrofloxacin and its metabolite ciprofloxacin, which are widely used as drugs for humans and animals. We utilized gold nanoparticles (AuNPs) and laser desorption/ionization time-of-flight (LDI-TOF) mass spectrometry (MS) with a matrix-free format. An antibody for the drug was immobilized on a chip based on self-assembled monolayers (SAMs) on gold, and AuNPs were decorated with the drug along with a large excess of small molecules, called amplification tags (Am-tags). In this strategy, target drugs in solution bound to the antibody on the chip compete with the AuNP-immobilized drugs. The presence of targets was verified by the amplified LDI-TOF MS signals of Am-tags on AuNPs.

Structural properties of polyphenols causing cell cycle arrest at G1 phase in HCT116 human colorectal cancer cell lines.

Plant-derived polyphenols are being tested as chemopreventive agents; some polyphenols arrest the cell cycle at G1 phase, whereas others inhibit cell cycle proliferation at G2/M phase. Therefore, polyphenols have been proposed to inhibit cell cycle progression at different phases via distinct mechanisms. Indeed, our previous studies showed that small structural differences in polyphenols cause large differences in their biological activities; however, the details of the structural properties causing G1 cell cycle arrest remain unknown. In this study, we prepared 27 polyphenols, including eight different scaffolds, to gain insight into the structural conditions that arrest the cell cycle at G1 phase in a quantitative structure-activity relationship study. We used cell cycle profiles to determine the biophores responsible for G1 cell cycle arrest and believe that the biophores identified in this study will help design polyphenols that cause G1 cell cycle arrest.

Complete assignments of 1H and 13C NMR data for 21 naphthalenyl-phenyl-pyrazoline derivatives.

To find potent new chemotherapy drugs, we designed and synthesized a series of naphthochalcones bearing naphthalenyl-phenyl-pyrazoline moieties. The complete (1)H and (13)C NMR data for these compounds are reported here and can be used to identify further new naphthochalcones bearing the desired pyrazoline moieties.

Assessment of Korean consumer exposure to sodium saccharin, aspartame and stevioside.

The dietary intakes of sodium saccharin, aspartame and stevioside were estimated on the basis of food consumption data of the Korean consumer and the concentration of sweeteners in processed foods. Results were compared with the acceptable daily intake (ADI) of sweeteners. Among the 28 food categories for which the application of sodium saccharin, aspartame and stevioside is permitted in Korea, they were detected in 5, 12 and 13 categories, respectively. The estimated daily intake (EDI) of sodium saccharin and aspartame were high in infants and children, whereas the EDI of stevioside was high in adolescents and adults. The most highly consumed sweetener was aspartame, and the highest EDI/ADI ratio was found for sodium saccharin. The main food categories contributing to sweetener consumption were beverages, including alcoholic beverages. For most Korean consumers, the EDIs were no greater than 20% of their corresponding ADI; however, the EDI of sodium saccharin for conservative consumers aged 1-2 years reached 60% of their ADI.

Assessment of exposure of Korean consumers to acesulfame K and sucralose using a stepwise approach.

Using a stepwise assessment of the exposure of Korean consumers to acesulfame K and sucralose, theoretical maximum daily intakes of the sweeteners were calculated using the Budget screening method, which resulted in values greater than the acceptable daily intakes (ADIs). Accordingly, the daily intakes of the sweeteners based on food consumption data and concentrations determined by instrumental analysis of 605 food samples were estimated for the more refined approach. The estimated daily intakes (EDIs) of all ordinary consumers were lower than the ADI, which was considered safe. However, for infants and 95th percentile high-level consumers (especially those who choose sucralose-containing foods), the EDIs of sucralose were very close to and higher than the ADI. Therefore, the sucralose concentration in sweetened beverages should be reduced; this would benefit the health of both high-level consumers and infants.

The effects of ginsenoside Rg(3) on human Kv1.4 channel currents without the N-terminal rapid inactivation domain.

Kv1.4 channel belongs to the family of voltage-gated potassium channels that mediate transient and rapidly inactivating A-type currents and N-type inactivation. This N-type inactivation can be removed by the deletion of N-terminal domains, which exhibit non-inactivating currents and C-type inactivation. In our previous report, we demonstrated that 20(S)-ginsenoside Rg(3) (Rg(3)), one of the active ingredients of ginseng saponins, inhibits human Kv1.4 (hKv1.4) channel currents through the interaction with amino acids, including Lys (K) residue, which is known as K(+) activation and the extracellular tetraethylammonium (TEA) binding site. In the present study, we examined the effects of Rg(3) on hKv1.4 channel currents without the N-terminal rapid inactivation domain. We constructed hKv1.4Delta2-61 channels by N-terminal deletion of 2-61 amino acid residues. We investigated the effect of Rg(3) on hKv1.4Delta2-61 channel currents. We found that Rg(3) preferentially inhibited non-inactivating outward currents rather than peak outward currents of hKv1.4Delta2-61 channels. The mutation of K531 hKv1.4Delta2-61 to K531Y hKv1.4Delta2-61 and raising of extracellular [K(+)](o) abolished Rg(3) inhibitions on non-inactivating outward currents. Rg(3) treatment increased the C-type inactivation rate, but raising the extracellular [K(+)](o) reversed Rg(3) action. These results provide additional evidence that K531 residue also plays an important role in the Rg(3)-mediated non-inactivating current blockages and in Rg(3)-mediated increase of the C-type inactivation rate in hKv1.4Delta2-61 channels.

Development of predictive mathematical model for the growth kinetics of Staphylococcus aureus by response surface model.

A response surface model was developed for predicting the growth rates of Staphylococcus aureus in tryptic soy broth (TSB) medium as a function of combined effects of temperature, pH, and NaCl. The TSB containing six different concentrations of NaCl (0, 2, 4, 6, 8, and 10%) was adjusted to an initial of six different pH levels (pH 4, 5, 6, 7, 8, 9, and 10) and incubated at 10, 20, 30, and 40 degrees C. In all experimental variables, the primary growth curves were well (r2=0.9000 to 0.9975) fitted to a Gompertz equation to obtain growth rates. The secondary response surface model for natural logarithm transformations of growth rates as a function of combined effects of temperature, pH, and NaCl was obtained by SAS's general linear analysis. The predicted growth rates of the S. aureus were generally decreased by basic (pH 9-10) or acidic (pH 5-6) conditions and higher NaCl concentrations. The response surface model was identified as an appropriate secondary model for growth rates on the basis of correlation coefficient (r=0.9703), determination coefficient (r2=0.9415), mean square error (MSE=0.0185), bias factor (B(f)=1.0216), and accuracy factor (A(f)=1.2583). Therefore, the developed secondary model proved reliable for predictions of the combined effect of temperature, NaCl, and pH on growth rates for S. aureus in TSB medium.

Natural occurrence of aflatoxin B1 in marketed foods and risk estimates of dietary exposure in Koreans.

Aflatoxin B1 (AFB1) is an unavoidable food contaminant. To evaluate the potential health risk of AFB1 to Koreans posed by food consumption, we determined the natural occurrence of AFB1 in food and estimated the excess risk for liver cancer through dietary exposure to AFB1. A total of 694 food samples collected from six different regions of South Korea were analyzed for their AFB, content. One hundred four of the 694 samples were found to give positive enzyme-linked immunosorbent assay (ELISA) readings for AFB1 and were further investigated with high-performance liquid chromatography. Thirty-two samples, including 2 maize samples, 3 soybean products, 20 peanut samples, nut samples, and their products, and 7 spices, were found to be contaminated with AFB1 (4.6% incidence), up to 48.6 microg kg(-1). The level of AFB1 contamination in 28 of the 32 food products was below 10 microg kg(-1), which is the legal tolerance limit in Korea. From data on daily food consumption, the exposure dose of AFB1 was estimated to be 6.42 x 10(-7) mg kg(-1) body weight (bw) day(-1). The major contributors to the dietary intake of AFB1 were soybean paste and soy sauce, which composed 91% of the total exposure to AFB1. The excess risk of liver cancer for those exposed to AFB1 through food intake was estimated to be 5.78 x 10(-6) for hepatitis B-negative individuals and 1.48 x 10(-4) for hepatitis B-positive individuals. These results suggest that special consideration is required to reduce the intake of AFB1 in hepatitis B-positive individuals.

Bactericidal effects of CaO (scallop-shell powder) on foodborne pathogenic bacteria.

This study was investigated the bactericidal effects of calcium oxide (CaO) on three common foodborne pathogenic bacteria: Escherichia coli, Listeria monocytogenes, and Salmonella typhimurium. Each bacteria level was determined in a CaO solution (0.01, 0.03, 0.05, 0.10, 0.15, and 0.20% [w/v]) exposed for either 15 sec, 30 sec, 1 min, 2 min, 3 min, 5 min, 10 min, or 30 min. All three bacteria were not greatly affected by CaO solutions at concentrations of 0.01 and 0.03%, however, the decline of E. coli (99%; 2.78 log10 CFU/mL), L. monocytogens (45%; 1.44 log10 CFU/mL), and S. typhimurium (70%; 2.08 log10 CFU/mL) was greatest when they were exposed to 0.05% CaO solution for 10 min. Moreover, the bactericidal action of CaO was maintained for at least 24 h of storage. The results of this study provide evidence that CaO, as a substitute for synthetic chemical substances has potential for use in the disinfection and sanitization of foods and food processing equipment.

Hypocholesterolemic property of Yucca schidigera and Quillaja saponaria extracts in human body.

This study was undertaken to observe the effects of the blend of partially purified Yucca schidigera and Quillaja saponaria extracts on cholesterol levels in the human's blood and gastrointestinal functions, and to determine if a new cholesterol-lowering drug can be developed by the further purification of the extracts. Ultrafiltration and sequential diafiltration increased the amounts of steroidal saponin in aqueous yucca extract and terpenoid saponin in aqueous quillaja extract from 9.3% and 21.4% to 17.2% and 61.8%, respectively. Taking 0.9 mg of the blend (6:4, v:v) of the resulting filtrates a day for 4 weeks resulted in the decreases in total and LDL cholesterol levels in blood plasma of hyper-cholesterolemic patients with enhancement in gastrointestinal symptoms of patients.