• Journal of Microbiology and Biotechnology
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• 제25권9호
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• pp.1537-1541
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• 2015

Previous detection methods for Citrobacter are considered time consuming and laborious. In this study, we have developed a rapid and accurate detection method for Citrobacter species in pork products, using matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry (MS). A total of 35 Citrobacter strains were isolated from 30 pork products and identified by both MALDI-TOF MS and 16S rRNA gene sequencing approaches. All isolates were identified to the species level by the MALDI-TOF MS, while 16S rRNA gene sequencing results could not discriminate them clearly. These results confirmed that MALDITOF MS is a more accurate and rapid detection method for the identification of Citrobacter species.

• 한국동물위생학회지
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• 제41권2호
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• pp.105-110
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• 2018

A total of 41 Salmonella (S.) strains were isolated from pigs suffered with severe watery diarrhea and were tried to identify by both matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) and polymerase chain reaction (PCR) analysis. Fibrinous exudate and ulceration in the large intestine were prevalent in gross observation, and variable degrees of enteritis were observed in the histology of large intestines. Subsequent polymerase chain reaction (PCR) analyses demonstrated that 41 strains were identified as S. Typhimurium (39 strains), though 2 stains were failed to identify. Further identification was performed using both direct smear and protein extraction method by MALDI-TOF MS analyses. In terms of extraction methods, 100% (41/41) of isolates were identified to species level of S. spp. Whereas only 43.9% (18/41) were identified to species level using the direct method. These results thus suggest that rapid and accurate diagnosis of porcine salmonellosis can be guaranteed by MALDI-TOF MS combined with protein extraction method.

• 한국동물위생학회지
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• 제39권4호
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• pp.247-251
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• 2016

Salmonella is one of the most common bacteria that causes heavy losses in swine industry and major causative pathogen of food poisoning in public health. Various methods for the identification of Salmonella such as Gram staining, agglutination test, enzyme-linked immunosorbent assay (ELISA), polymerase chain reaction (PCR) have been used. Several studies have demonstrated that Matrix Assisted Laser Desorption Ionization Time of Flight (MALDI TOF) Mass Spectrometry (MS) identification is an efficient and inexpensive method for the rapid and routine identification of isolated bacteria. In this study, MALDI TOF MS could provide rapid, accurate identification of Salmonella spp. from swine compared with end point PCR and real time PCR.

• Journal of Microbiology and Biotechnology
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• 제27권6호
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• pp.1128-1137
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• 2017

The aim of this study was to explore the accuracy and feasibility of matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) in identifying bacteria from environmental sources, as compared with rpoA gene sequencing, and to evaluate the occurrence of bacteria of the genus Enterococcus in wild birds. In addition, a phyloproteomic analysis of certain Enterococcus species with spectral relationships was performed. The enterococci were isolated from 25 species of wild birds in central Europe (Poland). Proteomic (MALDI-TOF MS) and genomic (rpoA gene sequencing) methods were used to identify all the isolates. Using MALDI-TOF MS, all 54 (100%) isolates were identified as Enterococcus spp. Among these, 51 (94.4%) isolates were identified to the species level (log(score) ${\geq}2.0$), and three isolates (5.6%) were identified at a level of probable genus identification (log(score) 1.88-1.927). Phylogenetic analysis based on rpoA sequences confirmed that all enterococci had been correctly identified. Enterococcus faecalis was the most prevalent enterococcal species (50%) and Enterococcus faecium (33.3%) the second most frequent species, followed by Enterococcus hirae (9.3%), Enterococcus durans (3.7%), and Enterococcus casseliflavus (3.7%). The phyloproteomic analysis of the spectral profiles of the isolates showed that MALDI-TOF MS is able to differentiate among similar species of the genus Enterococcus.

• Journal of Microbiology and Biotechnology
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• 제29권4호
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• pp.548-557
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• 2019

Staphylococcus species have a ubiquitous habitat in a wide range of foods, thus the ability to identify staphylococci at the species level is critical in the food industry. In this study, we performed rapid identification of Staphylococcus species using Matrix-Assisted Laser Desorption/Ionization Time-of-Flight mass spectrometry (MALDI-TOF MS). MALDI-TOF MS was evaluated for the identification of Staphylococcus reference strains (n = 19) and isolates (n = 96) from various foods with consideration for the impact of sample preparation methods and incubation period. Additionally, the spectra of isolated Staphylococcus strains were analyzed using principal component analysis (PCA) and a main spectra profile (MSP)-based dendrogram. MALDI-TOF MS accurately identified Staphylococcus reference strains and isolated strains: the highest performance was by the EX method (83.3~89.5% accuracy) at species level identification (EDT, 70.3~78.9% accuracy; DT, less than 46.3~63.2% accuracy) of 24-h cultured colonies. Identification results at the genus level were 100% accurate at EDT, EX sample preparation and 24-h incubation time. On the other hand, the DT method showed relatively low identification accuracy in all extraction methods and incubation times. The analyzed spectra and MSP-based dendrogram showed that the isolated Staphylococcus strains were characterized at the species level. The performance analysis of MALDI-TOF MS shows the method has the potential ability to discriminate between Staphylococcus species from foods in Korea. This study provides valuable information that MALDI-TOF MS can be applied to monitor microbial populations and pathogenic bacteria in the food industry thereby contributing to food safety.

• Bulletin of the Korean Chemical Society
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• 제34권9호
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• pp.2635-2639
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• 2013

The rapid and accurate identification of biological agents is a critical step in the case of bio-terror and biological warfare attacks. Recently, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry has been widely used for the identification of microorganisms. In this study, we describe a method for the rapid and accurate discrimination of Bacillus anthracis spores using MALDI-TOF MS. Our direct in-situ analysis of MALDI-TOF MS does not involve subsequent high-resolution mass analyses and sample preparation steps. This method allowed the detection of species-specific biomarkers from each Bacillus spores. Especially, B. anthracis spores had specific biomarker peaks at 2503, 3089, 3376, 6684, 6698, 6753, and 6840 m/z. Cluster and PCA analyses of the mass spectra of Bacillus spores revealed distinctively separated clusters and within-groups similarity. Therefore, we believe that this method is effective in the real-time identification of biological warfare agents such as B. anthracis as well as other microorganisms in the field.

• Journal of Ginseng Research
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• 제45권5호
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• pp.539-545
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• 2021

Background: Red ginseng polysaccharides (RGPs) have been acknowledged for their outstanding immunomodulation and anti-tumor activities. However, their studies are still limited by the complexity of their structural features, the absence of purification and enrichment methods, and the rarity of the analytical instruments that apply to the analysis of such macromolecules. Thus, this study is an attempt to establish a new mass spectrometry (MS)-based analysis procedure for RGPs. Methods: Saponin pre-excluded powder of RG (RG-SPEP, 10 mg) was treated with 200 µL of distilled water and centrifuged for 5 h at 1000 rpm and 85 ℃. Ethanol-based precipitation and centrifugation were applied to obtain RGPs from the heated extracts. Further, endo-carbohydrase treatments were performed to produce specific saccharide fragments. Solid-phase extraction (SPE) processes were implemented to purify and enrich the enzyme-treated RGPs, while matrix-assisted laser desorption/ionization time-of-flight/time-of-flight (MALDI-TOF/TOF) MS was employed for the partial structural analysis of the obtained RGPs. Results: Utilizing cellulase, porous graphitized carbon (PGC), hydrophilic interaction chromatography (HILIC), and MALDI-TOF/TOF MS, the neutral and acidic RGPs were qualitatively analyzed. Hex_n and Hex_n-18 (cellulose analogs) were determined to be novel neutral RGPs. Additionally, the [Unknown + Hex_n] species were also determined as new acidic RGPs. Furthermore, HexA_n (H) was determined as another form of the acidic RGPs. Conclusion: Compared to the previous methods of analysis, these unprecedented applications of HILIC-SPE and MALDI-TOF/TOF MS to analyze RGPs proved to be fairly effective for fractionating and detecting neutral and acidic components. This new procedure exhibits great potential as a specific tool for searching and determining various polysaccharides in many herbal medicines.

• 대한방사성의약품학회지
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• 제3권2호
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• pp.54-58
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• 2017

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry(MALDI-TOF MS) is one of the powerful methods that enable analysis of small molecules as well as large molecules up to about 500,000 Da without severe fragmentation. MALDI-TOF MS, thus, has been a very useful an analytical tool for the confirmation of synthetic molecules, probing PTMs, and identifying structures of a given protein. In recent nuclear medicine, MALDI-TOF MS liner ion mode helps researcher calculate the average number of chelator(or linkage) per an antibody conjugate, such as DOTA-(or DFO-) trastuzumab for labeling a medical radioisotope. This simple technique can be utilized to improve the labeling method and control the quality at the development of antibody-based radiopharmaceuticals, which is very effected to diagnosis and therapy for in vivo tumor cells, with radioisotopes like $^{89}Zr$, $^{64}Cu$, and 177Lu. To minimize the error, MALDI-TOF MS measurement is repeatedly performed for each sample in this study, and external calibration is carried out after data collection.

• KSBB Journal
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• 제31권4호
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• pp.263-269
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• 2016

For decades, the microorganisms in arctic soils have been newly discovered according to the climate change and global warming. In this study, the chemical structure of a lipid A molecule from Pseudomonas sp. strain PAMC 28615 which was newly discovered from arctic soils was characterized by mass spectrometric approaches such as matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) and MALDI multi-stage tandem mass spectrometry (MS). First, lipopolysaccharide (LPS) from Pseudomonas sp. strain PAMC 28615 was extracted and subsequently hydrolyzed to obtain the lipid A. The parent ion peak at m/z 1632 was determined by MALDI-TOF MS, which also can validate our lipid A purification method. For detailed structural determination, we performed the multiple-stage tandem mass analysis ($MS^4$) of the parent ion, and subsequently the abundant fragment ions in each MS stage are tested. The fragment ions in each MS stage were produced from the loss of phosphate groups and fatty acyl groups, which could be used to confirm the composition or the position of the lipid A components. Consequently, the mass spectrometry-based lipid A profiling method could provide the detail chemical structure of lipid A from the Pseudomonas sp. strain PAMC 28615 as an arctic bacterium from the frozen arctic soil.

• 생명과학회지
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• 제8권6호
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• pp.654-659
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• 1998

NR2B는 연접후 치밀질의 주요 tyrosine 인산화 단백질이다. 본 연구에서는 mass spectrometry 방법을 적용하여 NR2B의 tyrosine 인산화 위치를 동정하였다. NR2B를 N-octyl glucoside (NOG)에 용해되지 않는 PSD 분획으로부터 SDS-PAGE와 electroelution방법으로 분리하였다. 분리한 단백질을 NR2B와 phos-photyrosine에 특이한 항체로 조사한 결과 이들은 phosphotyrosine을 유지하고 있는 NR2B임이 확인되었다. 이 단백질을 trypsin 혹은 endolys-C 처리하고, matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry 방법으로 조사한 결과 Tyr-1304이 인산화됨을 확인하였다.