• Journal of Microbiology and Biotechnology
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• v.27 no.9
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• pp.1593-1601
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• 2017

Vibrio species are generally recognized as pathogens predominant in seafood along coastal areas. The food industry has sought to develop efficient microbial detection methods. Owing to the limits of conventional methods, this study aimed to establish a rapid identification method for Vibrio isolated from Korea, based on matrix-assisted laser-desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Four different preparation procedures were compared to determine the appropriate means to pretreat Vibrio species, using 17 isolates and five reference strains. Extended direct transfer and full formic acid extraction methods using bacterial colonies on agar plates revealed very low identification rates. Formic acid and trifluoroacetic acid (TFA) extractions using bacterial broth cultures were also performed. All Vibrio isolates and reference strains prepared by TFA extraction were successfully identified to the species level (17/22, 77.3%) and to the genus level (5/22, 22.7%). Thus, TFA extraction was considered the most appropriate method to pretreat Vibrio species for MALDI-TOF MS. The remaining 33 isolates and two reference strains were prepared by TFA extraction and analyzed by MALDI-TOF MS. Overall, 50 isolates were identified to the species level (40/50, 80%) and to the genus level (10/50, 20%). All isolates were identified as 43 V. alginolyticus, six V. parahaemolyticus, and one V. vulnificus species. V. alginolyticus and V. parahaemolyticus were isolated from fish offal (87.5% and 12.5%, respectively), seawater (91.3%, 8.7%), and shellfish (62.5%, 37.5%), whereas V. alginolyticus and V. vulnificus were isolated from sediment (90.9% and 9.1%, respectively). This study established a reliable system of MALDI-TOF MS preparation and analysis for Vibrio identification.

• Journal of Microbiology and Biotechnology
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• v.25 no.11
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• pp.1954-1959
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• 2015

Bacillus subtilis HM1 was isolated from the rhizosphere region of halophytes for its antifungal activity against Colletotrichum acutatum, the causative agent of anthracnose. Treatment of postharvest apples with the cell culture or with a cell-free culture supernatant reduced disease severity 80.7% and 69.4%, respectively. Both treatments also exhibited antifungal activity against various phytopathogenic fungi in vitro. The antifungal substances were purified and analyzed by acid precipitation, gel filtration, high-performance liquid chromatography, and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Three compounds were identified as fengycin, iturin, and surfactin. The MALDI-TOF/TOF mass spectrum revealed the presence of cyclized fengycin homologs A and B, which were distinguishable on the basis of the presence of either alanine or valine, respectively, at position 6 of the peptide sequence. In addition, the cyclized structure of fengycin was shown to play a critical role in antifungal activity.

• Bulletin of the Korean Chemical Society
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• v.34 no.6
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• pp.1708-1714
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• 2013

Polyhexamethylene guanidine (PHMG) polymers used as an active ingredient in an antibacterial humidifier disinfectant were reported to cause harm to the human health when inhaled, although physical contact with this material was known to present low toxicity to humans. It is therefore necessary to develop an optimal analysis method which enables detection and analysis of PHMG polymers. MALDI-TOF investigations of PHMG are performed with a variety of matrices, and it is found that CHCA and 2,5-DHB are excellent matrices which well reflects the polymer population even at high mass. For the provided PHMG sample, the number-average ($M_n$) and weight-average ($M_w$) molecular masses were determined to be 744.8 and 810.7, respectively, when the CHCA was used as a matrix. The rank of the matrices in terms of averaged molecular weight was CHCA ~2,5-DHB > 5-NSA > DHAP, THAP > ATT > IAA ~ super-DHB ~ HABA. In addition, PSD of the PHMG oligomer ions exhibited a few unique fragmenation characteristics. The formation of a- and c-type fragments was the major fragmentation pathway, and the 25-Da loss peaks generally accompanied a- and c-type fragments.

• Korean Journal of Clinical Laboratory Science
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• v.52 no.3
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• pp.221-228
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• 2020

Ochrobactrum anthropi is a non-fermentative oxidative gram-negative bacillus that produces oxidase. Distinguishing a mixed culture with non-fermenting bacteria having a similar appearance and oxidase-positive is difficult, and there is a limit to accurate identification with a biochemical identification system. This paper proposes that the Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry Platform (MALDI-TOF) method is useful for classifying bacteria that are difficult to identify using biochemical testing methods. As a result of analyzing five cases of O. anthropi examined using MicroScan, it took 6.5 days to the final report, which was 3.5 days more than the 3.0 days of E. coli. The pus sample in patient 5 was a mixed infection with Achromobacter xylosoxidans, and it took 11.3 days because of multiple subculture and retests. Four patients were over 60 years old with an underlying disease, and the possibility of opportunistic and nosocomial infections could not be excluded. Among them, samples collected after 92 days of hospitalization were resistant to imipenem and meropenem. Therefore, an examination using the MALDI-TOF method will be useful for the rapid and adequate treatment of patients with difficult identification, such as O. anthropi.

• Journal of Microbiology and Biotechnology
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• v.28 no.7
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• pp.1112-1121
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• 2018

Jeotgal is a Korean traditional fermented seafood with a high concentration of salt. In this study, we isolated lactic acid bacteria (LAB) from galchi (Trichiurus lepturus, hairtail) and myeolchi (Engraulis japonicas, anchovy) jeotgal on MRS agar and MRS agar containing 5% NaCl (MRS agar+5% NaCl), and identified them by using 16S rRNA gene sequencing and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) as culture-dependent methods. We also performed polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) as a culture-independent method to identify bacterial communities. Five samples of galchi-jeotgal and seven samples of myeolchi-jeotgal were collected from different regions in Korea. A total of 327 and 395 colonies were isolated from the galchi- and myeolchi-jeotgal samples, respectively. 16S rRNA gene sequencing and MALDI-TOF MS revealed that the genus Pediococcus was predominant on MRS agar, and Tetragenococcus halophilus on MRS agar+5% NaCl. PCR-DGGE revealed that T. halophilus, Tetragenococcus muriaticus, and Lactobacillus sakei were predominant in both types of jeotgal. T. halophilus was detected in all samples. Even though the same species were identified by both culture-dependent and -independent methods, many species identified by the culture-dependent methods were not in the bacterial list identified by the culture-independent methods. The distribution of bacteria in galchi-jeotgal was more diverse than in myeolchi-jeotgal. The diverse LAB in galchi- and myeolchi-jeotgals can be further studied as candidates for starter cultures to produce fermented foods.

• Bulletin of the Korean Chemical Society
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• v.24 no.9
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• pp.1308-1312
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• 2003

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was used to observe folic acid and its derivatives such as tetrahydrofolate and 5-methyltetrahydrofolate in a vitamin tablet and in foods. Folic acid in a vitamin tablet was determined using angiotensin I as an internal reference. Tetrahydrofolic acid, 5-methyltetrahydrofolic acid, and an oxygenated folate were observed from a human blood sample using graphite plate. The results show that these mass spectrometric methods are useful for quickly obtaining a profile of folates.

• Proceedings of the Korean Vacuum Society Conference
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• 2013.02a
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• pp.113-114
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• 2013

Time-of-flight secondary ion mass spectrometry (TOF-SIMS) imaging is a powerful technique for producing chemical images of small biomolecules (ex. metabolites, lipids, peptides) "as received" because of its high molecular specificity, high surface sensitivity, and submicron spatial resolution. In addition, matrix-assisted laser desorption and ionization time-of-flight (MALDI-TOF) imaging is an essential technique for producing chemical images of large biomolecules (ex. genes and proteins). For this talk, we will show that label-free mass imaging technique can be a platform technology for biomedical studies such as early detection/diagnostics, accurate histologic diagnosis, prediction of clinical outcome, stem cell therapy, biosensors, nanomedicine and drug screening [1-7].

• Journal of Life Science
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• v.15 no.6 s.73
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• pp.1005-1012
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• 2005

Endophytic Bacillus sp. CY22 was previously isolated from the interior of balloon flower root and showed strong antifungal activity against phytopathogenic fungi such as Rhizoctonia solnni, Fusarium oxysporum, and Phythium ultimum. Many Bacillus strains produce antifungal compound such as iturin, fengycin, and mycosubtilin. We isolated and identified antifungal compound from cell supernatant of the endophytic strain. By the MALDI-TOF mass result, the antifungal compound was similar to the known antifungal lipopeptide iturin. It was found that the purified iturin had three isoforms with protonated masses of m/z 1,043.39, 1,057.42, and 1,071.42 and different structures in combination with $Na^{+}$ ion using MALDI-TOF MS. The ita22 gene, which transacylase gene is associated with production of antifungal iturin, had an open reading frame (ORF) of 1,200 bp encoding 400 amino acids. Results of deduced amino acids sequence homology search, Ita22 was homologous with FenF (BAB69697) of Bacillus subtilis 168.

• Journal of Life Science
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• v.8 no.6
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• pp.654-659
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• 1998

The 2B subunit of N-methyl-D-aspartate (NMDA) receptors (NR2B) is the major phosphotyrosine-containing pro-tein in the postsynaptic density (PSD). In order to identify the site for tyrosine phosphorylation on NR2B, a mass spectrometry was applied on tryptic and endolys-C peptides. The NR2B subunit was isolated from N-octyl glucoside (NOG)-insoluble PSD fraction through SDS-PAGE and electroelution. The eluted protein was confirmed to be NR2B and phosphorylated on tyrosine by its cognate antibody and phosphotyrosine-specific antibody. By matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry of the peptides generated by digesting the eluted NR2B with trysin or endolys-C, a potential site for tyrosine phosphorylation could be identified as Tyr-1304.

• Bulletin of the Korean Chemical Society
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• v.28 no.9
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• pp.1499-1509
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• 2007

Proteomic analyses of synaptic vesicle fraction from rat brain have been performed for the better understanding of vesicle regulation and signal transmission. Two different approaches were applied to identify proteins in synaptic vesicle fraction. First, the isolated synaptic vesicle proteins were treated with trypsin, and the resulting peptides were analyzed using a high-pressure capillary reversed phase liquid chromatography/tandem mass spectrometry (cRPLC/MS/MS). Alternatively, proteins were separated by two-dimensional gel electrophoresis (2DE) and identified by matrix-assisted laser desorption ionization mass spectrometry (MALDI-TOF/MS). Total 18 and 52 proteins were identified from cRPLC/MS-MS and 2DE-MALDI-TOF-MS analysis, respectively. Among them only 2 proteins were identified by both methods. Of the proteins identified, 70% were soluble proteins and 30% were membrane proteins. They were categorized by their functions in vesicle trafficking and biogenesis, energy metabolism, signal transduction, transport and unknown functions. Among them, 27 proteins were not previously reported as synaptic proteins. The cellular functions of unknown proteins were estimated from the analysis of domain structure, expression profile and predicted interaction partners.