The Journal of the Korean Society for Microbiology
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v.35
no.2
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pp.97-108
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2000
Two-dimensional gel electrophoresis (2-DE) is an essential tool of proteomics to analyse the entire set of proteins of an organism and its variation between organisms. Helicobacter pylori was tried to identify differences between strains. As the first step, whole H. pylori was lysed using high concentration urea contained lysis buffer [9.5 M Urea, 4% CHAPS, 35 mM Tris, 65 mM DTT, 0.01% SDS and 0.5% Ampholite (Bio-Rad, pH 3-10)]. The extract ($10\;{\mu}g$) was rehydrated to commercially available immobilised pH gradient (IPG) strips, then the proteins were separated according to their charges as the first dimensional separation. The IPG strips were placed on Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis (SDS-PAGE) to separate according to molecular mass of the proteins as the second dimension. The separated protein spots were visualised by silver staining in order to compare different expression of proteins between strains. Approximately 120 spots were identified in each mini-protein electrophoresised gel, furthermore about 65 to 75 spots were regarded as identical proteins in terms of pI value and molecular weight between strains used. In addition, distinct differences were found between strains, such as 219-1, Y7 and Y14, CH150. Two representative strains were examined using strips which had pH range from 4 to 7. This strips showed a number of isoforms which were considered large spots on pH range 3-10. Furthermore, the rest of spots on pH 4-7 IPG strips appeared very distinctive compared to broad range IPG strips. 2-DE seems to be an excellent tool for analysing and identifying variations between H. pylori strains.
An efficient method of plant regeneration from Acanthopanax chiisanensis somatic embryos was developed. Cotyledonary somatic embryos were obtained in liquid Murashige and Skoog (MS) medium from embryogenic cell suspension cultures. They were desiccated for 0 to 72 hr and then cultured on MS medium containing NAA, BA, GA$_3$, (0-0.5mg/L). The highest multiple shoots formation (100%) was obtained from 72 hr desiccated somatic embryos on ifs medium with 0.5mg/L NAA+0.5mg/L BA or 0.5 mg/L NAA+0.5mg/L BA+0.5mg/L GA$_3$ after 6 weeks culture. Plant conversion from multiple shoots was not high. The highest plant conversion from multiple shoots was obtained on 1/3MS medium with 1.0mg/L GA$_3$. Converted plantlets were transferred to ex vitro condition and the highest survival rate (70%) of the plantlets was obtained on plastic pots containing vermiculite and sand. These results indicate that micropropagation procedure can be applied for an efficient mass propagation of Acanthopanax chiisanensis.
The purpose of this study was to investigate the dietary habits and daily intake of nutrients in college students. This survey was conducted using a self-administered questionaire. The average heights and weights were 173.5 cm and 72.3 kg of male students and 161.8 cm and 57.2 kg of female students. The average of BMI was $24.2kg/m^2$ of male and $21.9kg/m^2$ of female, and the value of male students was higher than the value of female students. The response to the daily meals was 54.6% for '$2{\sim}3$ times/week'. The regularity of mealtime was 41.7% for irregular and the frequency eating after nine was 45.7% for '5-6 times/week', respectively. The repast was 72.2% for 'overeating and little eating' and was a significant difference of male and female students (p<0.05). The eating rate was higher '$10{\sim}20min$'. As for breakfast food eaten, skipping breakfast was 23.6% for 'no/week' and female students were higher than male students (p<0.05). The frequency of snacks was 36.0% for 'nothing' of males students and 34.8% for '3-4 times/week' of female students (p<0.05). The type of snack was a significant difference of males and females students (p<0.01), and was the highest 75.0% for carbonated drinks of males and 37.5% for snacks of females. The eating due to stress solution was a significant difference of male and female students (p<0.01), and was the highest 23.0% for 'frequency' of males and 44.7% for 'sometime' of females. As for food intake of male and female students, the meat intake was 66.7% for 'everything of male and female students. The fish intake was 68.1 % for '1-2times/week'. The milk, milk products, eggs and beans were each 40.3%, 58.3%, 56.9%, 47.2% for '1-2 times/week' (p<0.05). The fat intake was 55.6% for '$1{\sim}2$ times/week'. The average consumption of energy was 58% of male and 67% of female of estimated energy requirement (EER). Their mean ratio of carbohydrate: protein: fat was 57 : 15 : 28 of all subjects. The mean intakes of vitamin C and folic acid were 70% and 51% of males and 62% and 52% of females of recommended intake (RI). The mean intakes of Ca, P, Fe and Na were 71%, 140%, 146% of males and 72%, 122%, 76% of female of RI and 273% of males and 233% of females of adequate intake (AI). Therefore, nutritional education is necessary for college students to establish physicall and mentall optimal health conditions though nutritional intervention.
The current standard for testing tetrodotoxin (TTX) in foodstuffs is the mouse bioassay (MBA) in Korea as in many other countries. However, this test suffers from potential ethical concerns over the use of live animals. In addition, the mouse bioassay does not test for a specific toxin thus a sample resulting in mouse incapacitation would need further confirmatory testing to determine the exact source toxin (e.g., TTX, STX, brevotoxin, etc.). Furthermore, though the time of death is proportional to toxicity in this assay, the dynamic range for this proportional relationship is small thus many samples must be diluted and new mice be injected to yield a result that falls within the quantitative dynamic range. Therefore, in recent years, there have been many efforts in this field to develop alternative assays. High performance liquid chromatography (HPLC) coupled with mass spectrometry (MS) has been emerged as one of the most promising options. A LC-MS-MS method involves solid-phase extraction (SPE) and followed by analysis using an electrospray in the positive ionization mode and multiple reactions monitoring (MRM). To adopt LC-MS-MS method as alternative standard for testing TTX, we performed a validation study for the quantification of TTX in puffer fish. This LC-MS-MS method showed good sensitivity as limits of detection (LOD) of $0.03{\sim}0.08{\mu}g/g$ and limits of quantification (LOQ) of $0.10{\sim}0.25{\mu}g/g$. The linearity ($r^2$) of tetrodotoxin were 0.9986~0.9997, the recovery were 80.9~103.0% and the relative standard deviations (RSD) were 4.3~13.0%. The correlation coefficient between the mouse bioassay and LC/MS/MS method was higher than 0.95.
Kim, Jong-Hwan;Kwon, Young Sang;Shin, Min-Chul;Kim, Su Jong;Seo, Jong-Su
Analytical Science and Technology
/
v.29
no.5
/
pp.234-241
/
2016
The approach presented in this article refers to the bioanalytical method validation for the detection and quantitative determination of arsenic species including arsenite (As(III)), arsenate (As(V)), dimethylarsinic acid (DMA) and monomethylarsonic acid (MMA) in dog plasma by high-performance liquid chromatography inductively coupled plasma mass spectrometry (HPLC-ICP/MS). The arsenic species were separated using an agilent As speciation column by a mobile phase of 2 mM sodium phosphate monobasic, 0.2 mM ethylenediaminetetraacetic acid disodium salt dehydrate, 10 mM sodium acetate, 3 mM sodium nitrate and 1 % ethyl alcohol at pH 11 (adjusted with 1M NaOH). The method validation experiment was obtained selectivity, linearity, accuracy, precision, matrix effect, recovery, system suitability, dilution integrity and various stabilities. All calibration curves showed good linearity (R2>0.999) within test ranges. The lower limit of quantitation (LLOQ) was 5 ng/mL for As(III), As(V) and DMA, and 20 ng/mL for MMA. The system suitability and dilution values were within 6.5 % and 7.7 %. Subsequently, the developed and validated HPLC-ICP/MS method was also successfully applied to determine the arsenic speciation in dog plasma samples, and the recoveries for the spiked samples were in the range of 91.5–102.2 %. Therefore, this method could be applied to the evaluation of arsenic exposure, health effect assessment and other bio-monitoring studies in biological samples.
Total phenolic content was the highest in 60% ethanol extracts at 21.91 mg/g, and inhibitory activity against tyrosinase of 60% ethanol extracts was higher than ethanol extracts of other concentration. The inhibitory compounds against tyrosinase from Persimmon leaves were purified using Sephadex LH-20, MCI-gel CHP-20 column chromatography with gradient elution. Two purified compounds were isolated as a result. The chemical structures of each compound were determined and identified using $^1H$-NMR and $^{13}C$-NMR, FAB-Mass. The compounds were confirmed as (+)-gallocatechin and prodelphinidin B-3. The tyrosinase inhibitory activities of purified (+)-gallocatechin and prodelphinidin B-3 were 29.5 and 40.2%, respectively. The inhibitory activities of (+)-gallocatechin and prodelphinidin B-3 against melanin biosynthesis in melanoma cell were 32.5 and 46.7%. The safety of essence with tyrosinase inhibitory compounds from persimmon leaves was also assessed by various safety profiles. First, changes in pH (4.90~4.95) and viscosity (23,000~26,000 cP) was not detected for 60 days. Essence also showed stability against temperature and light for 60 days. All these findings suggest that extracts from persimmon leaves have a great potential as a cosmetical ingredient with a potent whitening effect.
We tested for residual pesticide levels in school foodservice agricultural products in Seoul, Korea from 2010 to 2012. A total of 316 samples of 23 different types of agricultural products were analyzed via gas chromatography-nitrogen phosphate detector (GC-NPD), an electron capture detector (GC-${\mu}ECD$), a mass spectrometry detector (GC-MSD), and a high performance liquid chromatography-ultraviolet detector (HPLC-UV). We used multi-analysis methods to analyze 185 different pesticide types. Among the selected agricultural products, residual pesticides were detected in 26 samples (8.2%), of which 6 samples (1.9%) exceeded the Korea Maximum Residue limits (MRLs). We detected pesticide residue in more than 65% of the Chwinamul, while 6 among 9 analyzed samples contained pesticide residue, and 1 sample exceeded the Korea MRLs. Among the 185 kinds of pesticides that we have tested, 18 were detected, while 7 of them were detected more than twice. Data obtained were then used for estimating the potential health risks associated with the exposures to these pesticides. The most critical commodity is carbofuran in the perilla leaves, which has contributed 3.8% to the hazard index (HI). These results showed that the detected pesticides could not be considered as a serious public health problem. Nevertheless, constant supervision is recommended.
The radical scavenging activity measurement system linked with liquid chromatography (LC) is a useful tool for identifying the radical scavenging active compound in a sample composed of numerous compounds such as plant extracts. Using this system, DPPH and ABTS radical scavenging activity were measured on extracts of Ulmus pumila cortex, which is known as an herbal medicine with antioxidant activity. Mass spectrometry (MS) was performed on the identified radical scavenging active compounds to identify the four components estimated to be procyanidin B2, procyanidin B3, catechin-7-O-β-D-apiofuranoside, and catechin-5-O-β-D-apiofuranoside, respectively. In order to compare the relative contents between extract samples, multiple reaction monitoring (MRM) mode analysis conditions were set for the four compounds in order to examine the possibility of comparing the content of radical scavenging active compounds in Ulmus pumila cortex extract using LC-MS/MS. As a result of the relative content comparison, it was found that the higher the ethanol concentration of the extraction solvent, the higher the content of radical scavenging active compounds. As with the results of measuring the radical scavenging activity of each extract, it was confirmed that the content difference of three of the compounds (all except the compound estimated as procyanidin B3) was not significantly observed in the extracts with an ethanol concentration of 50% or more.
The present study was conducted to development mass production methods for peanut sprouts that is considered as a field of blue ocean among the agricultural products. 'Jopyeong' was the best as a major cultivar for peanut sprouts production. The manual for the production of high-quality peanut sprouts is as following. Germination temperature appropriate for production of high-quality peanut sprouts was $27^{\circ}C$. Peanut sprouts at the growth stage of 8th day, and older plants with advanced growth showed deteriorated merchantable and eating quality. Resveratrol compound was not found in the seeds, but its highest amount was detected from 9-day old sprouts. The best water temperature applicable to high quality peanut sprout production was $25^{\circ}C$. The growth of peanut sprout was inhibited by the high temperatures above $35^{\circ}C$ and low temperatures below $15^{\circ}C$.
This study was conducted to investigate the rheological properties of protein-bound polysaccharide powders (SD-1, 2, 3) using ultrafiltration (UF) and spray drying (SD) process from Agaricus blazei Murill. The calculated weight-average molar mass (Mw) in the positions at 29.7 mL (for SD-1), and at 27.8 mL (for SD-2), and at 18.7 mL (for SD-3) was $8.2{\times}10^3,\;9.6{\times}10^4$, and $5.9{\times}10^6g/mol$, respectively. As concentration increased the solution showed higher pseudoplasticity where the pseudoplasticity decreased as temperature increased. The flow behaviors of spray dried powder solutions were more fitted to Herschel-Bulkley equation than Power law equation. Apparent viscosity of SD-2 was more temperature-dependent than that of SD-1 and 3. However, the SD-3 tended to be more concentration-dependent than SD-1 and 2 as temperature increasing.
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