A First Genome Survey and Genomic SSR Marker Analysis of Trematomus loennbergii Regan, 1913.

Trematomus loennbergii Regan, 1913, is an evolutionarily important marine fish species distributed in the Antarctic Ocean. However, its genome has not been studied to date. In the present study, whole genome sequencing was performed using next-generation sequencing (NGS) technology to characterize its genome and develop genomic microsatellite markers. The 25-mer frequency distribution was estimated to be the best, and the genome size was predicted to be 815,042,992 bp. The heterozygosity, average rate of read duplication, and sequencing error rates were 0.536%, 0.724%, and 0.292%, respectively. These data were used to analyze microsatellite markers, and a total of 2,264,647 repeat motifs were identified. The most frequent repeat motif was di-nucleotide with 87.00% frequency, followed by tri-nucleotide (10.45%), tetra-nucleotide (1.94%), penta-nucleotide (0.34%), and hexa-nucleotide (0.27%). The AC repeat motif was the most abundant motif among di-nucleotides and among all repeat motifs. Among microsatellite markers, 181 markers were selected and PCR technology was used to validate several markers. A total of 15 markers produced only one band. In summary, these results provide a good basis for further studies, including evolutionary biology studies and population genetics of Antarctic fish species.

Genome of the Southern Giant Petrel Assembled Using Third-Generation DNA Sequencing and Linked Reads Reveals Evolutionary Traits of Southern Avian.

The southern giant petrel Macronectes giganteus, a large seabird of the southern oceans, is one of only two members of the genus Macronectes and is the largest species in the order Procellariiformes. Although these two families account for the vast majority of the avian fauna inhabiting the Antarctic and sub-Antarctic regions, studies on the status of some populations and the associated genetic data are currently extremely limited. In this study, we assembled the genome of M. giganteus by integrating Pacific Biosciences single-molecule real-time sequencing and the Chromium system developed by 10x Genomics. The final M. giganteus genome assembly was 1.248 Gb in size with a scaffold N50 length of 27.4 Mb and a longest scaffold length of 120.4 Mb. The M. giganteus genome contains 14,993 predicted protein-coding genes and has 11.06% repeat sequences. Estimated historical effective population size analysis indicated that the southern giant petrel underwent a severe reduction in effective population size during a period coinciding with the early Pleistocene. The availability of this newly sequenced genome will facilitate more effective genetic monitoring of threatened species. Furthermore, the genome will provide a valuable resource for gene functional studies and further comparative genomic studies on the life history and ecological traits of specific avian species.

Genome survey and microsatellite motif identification of Pogonophryne albipinna.

The genus Pogonophryne is a speciose group that includes 28 species inhabiting the coastal or deep waters of the Antarctic Southern Ocean. The genus has been divided into five species groups, among which the P. albipinna group is the most deep-living group and is characterized by a lack of spots on the top of the head. Here, we carried out genome survey sequencing of P. albipinna using the Illumina HiSeq platform to estimate the genomic characteristics and identify genome-wide microsatellite motifs. The genome size was predicted to be ∼883.8 Mb by K-mer analysis (K = 25), and the heterozygosity and repeat ratio were 0.289 and 39.03%, respectively. The genome sequences were assembled into 571624 contigs, covering a total length of ∼819.3 Mb with an N50 of 2867 bp. A total of 2217422 simple sequence repeat (SSR) motifs were identified from the assembly data, and the number of repeats decreased as the length and number of repeats increased. These data will provide a useful foundation for the development of new molecular markers for the P. albipinna group as well as for further whole-genome sequencing of P. albipinna.

The complete mitochondrial genome of Trematomus loennbergii (Perciformes, Nototheniidae).

The complete mitochondrial genome of Trematomus loennbergii was studied using NGS technology with PacBio platform. The mitochondrial genome size was 19,374bp and it had 13 protein-coding genes, 22 tRNAs and 2 rRNAs. There were 4 types of stop codons which were TAA, TAG, AGG and T(AA) but start codon type was only one (ATG). The contents of GC were 44.09% and AT contents were 55.91%. To conduct phylogenetic analysis, 12 species in 3 families were used. The result suggested that T. loennbergii was close to Pagothenia borchgrevinki in Nototheniidae. This study would provide a fundamental data for molecular evolution of T. loennbergii.

Antimicrobial Resistance Profile of Acinetobacter spp. Isolates from Retail Meat Samples under Campylobacter-Selective Conditions.

Acinetobacter strains are widely present in the environment. Some antimicrobial-resistant strains of this genus have been implicated in infections acquired in hospitals. Genetic similarities have been reported between Acinetobacter strains in nosocomial infections and those isolated from foods. However, the antimicrobial resistance of Acinetobacter strains in foods, such as meat, remains unclear. This study initially aimed to isolate Campylobacter strains; instead, strains of the genus Acinetobacter were isolated from meat products, and their antimicrobial resistance was investigated. In total, 58 Acinetobacter strains were isolated from 381 meat samples. Of these, 32 strains (38.6%) were from beef, 22 (26.5%) from pork, and 4 (4.8%) from duck meat. Antimicrobial susceptibility tests revealed that 12 strains were resistant to more than one antimicrobial agent, whereas two strains were multidrug-resistant; both strains were resistant to colistin. Cephalosporin antimicrobials showed high minimal inhibitory concentration against Acinetobacter strains. Resfinder analysis showed that one colistin-resistant strain carried mcr-4.3; this plasmid type was not confirmed, even when analyzed with PlasmidFinder. Analysis of the contig harboring mcr-4.3 using BLAST confirmed that this contig was related to mcr-4.3 of Acinetobacter baumannii. The increase in antimicrobial resistance in food production environments increases the resistance rate of Acinetobacter strains present in meat, inhibits the isolation of Campylobacter strains, and acts as a medium for the transmission of antimicrobial resistance in the environment. Therefore, further investigations are warranted to prevent the spread of antimicrobial resistance in food products.

The complete mitochondrial genome of Eaton's skate, Bathyraja eatonii (Rajiformes, Arhynchobatidae).

The complete mitochondrial genome of Eaton's skate Bathyraja eatonii was studied using the long-read technology, PacBio Sequel System. The complete mitochondrial genome form of B. eatonii was 16,698 bp and it's comprised of 13 protein-coding genes, 22 tRNA and 2 rRNA. The base composition of B. eatonii is analyzed 31.94% for A, 33.94% for T, 13.49% for G, 20.64% for C, the result of GC content was 33.94%. Phylogenetic analysis showed that B. eatonii was closely related to Bathyraja meridionalis in Arhynchobatidae family, and this first mitochondrial genome of Antarctic skate would provide fundamental information to the evolutional relationship of Antarctic fishes.

Characterization of oxazolidinone and phenicol resistance genes in non-clinical enterococcal isolates from Korea.

OBJECTIVES: To investigate the distribution and genetic characteristics of linezolid-resistant enterococci. METHODS: Enterococcus faecalis and Enterococcus faecium strains were isolated from pigs, equipment, grounds, and employees of 19 Korean swine farms in 2017. Antimicrobial susceptibility testing was then performed and linezolid resistance genes were detected via PCR. For genetic epidemiological characterization, multilocus sequence typing and whole-genome sequencing data were analysed. RESULTS: Twenty-eightE. faecalis and five E. faecium strains were isolated from 1026 samples obtained from the 19 farms. Ten sequence types were identified among the E. faecalis strains, of which ST256 (42.9%) and ST86 (25%) were the most abundant. The oxazolidinone and phenicol resistance genes poxtA, optrA, and fexA were detected in isolates of E. faecalis (100%, 85.7%, and 67.9%, respectively) and E. faecium (100%, 60%, and 80%, respectively). The minimum inhibitory concentrations of linezolid in these isolates ranged from 2 mg/L to 12 mg/L. The whole-genome sequencing data indicated that fexA was located upstream of poxtA. CONCLUSIONS: This is the first study to report the detection of poxtA in isolates that were both susceptible and resistant to linezolid in Korea. These results demonstrate the importance of antimicrobial resistance monitoring programmes, including regular antimicrobial susceptibility testing and resistance gene expression analysis, to facilitate the control of the spread of antibiotic resistance in non-clinical settings in Korea.

Topical application of autophagy-activating peptide improved skin barrier function and reduced acne symptoms in acne-prone skin.

BACKGROUND: Recent studies about the important roles of autophagy signaling in sebaceous lipogenesis and epidermal differentiation suggest potential benefits of autophagy activation in acne. AIMS: To investigate the effects of an autophagy activator on acne-prone skin. METHODS: Autophagy signaling in human immortalized SZ95 sebocytes, normal human epidermal keratinocytes, and 3D reconstituted skin was examined. Effects of an autophagy-activating peptide on sebaceous lipogenesis were measured by fluorescence microscopic analysis. The clinical efficacy in acne-prone skin was evaluated through an eight-week, double-blind, randomized, vehicle-controlled study. Changes in skin surface lipid compositions were further analyzed. RESULTS: In cultured sebocytes and keratinocytes, the investigated autophagy-activating peptide increased LC3-II expression, indicating a stimulation of autophagy signaling. Testosterone and linoleic acid treatment induced lipogenesis in cultured sebocytes and is further inhibited by the autophagy activator peptide treatment. Increased expression of differentiation marker proteins in cultured keratinocytes was also observed by autophagy-activating peptide. In clinical study, reduction of closed comedones and the amount of skin surface lipids as well as of trans-epidermal water loss (TEWL) were observed in acne-prone skin after autophagy-activating peptide application. In addition, reduction of squalene and increase in cholesterol were observed after an 8-week application. CONCLUSIONS: Topical application of an autophagy activator downregulated sebaceous lipogenesis and improved the skin barrier function. Considering the important roles of sebum and skin barrier function in acne pathogenesis, autophagy activation might represent a new therapeutic option in early forms of acne.

Anti-Inflammatory, Barrier-Protective, and Antiwrinkle Properties of Agastache rugosa Kuntze in Human Epidermal Keratinocytes.

This work aimed to assess the skin-beneficial properties of Agastache rugosa Kuntze, an herbal medication used to treat different types of disorders in traditional folk medicine. The total phenolic compounds and total antiradical, nitrite scavenging, superoxide scavenging, antielastase, and antihyaluronidase activities of a hot water extract of A. rugosa Kuntze leaves (ARE) were spectrophotometrically determined. Intracellular reactive oxygen species (ROS) was fluorometrically quantitated using 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA). Inducible nitric oxide synthase (iNOS) and filaggrin were evaluated using Western analysis. Real-time quantitative RT-PCR was used to measure filaggrin mRNA. Caspase-14 activity was determined using a fluorogenic substrate. ARE contained the total phenolic content of 38.9 mg gallic acid equivalent/g extract and exhibited 2,2'-diphenyl-1-picrylhydrazyl (DPPH) radical, superoxide radical, and nitrite scavenging activities with the SC50 values of 2.9, 1.4, and 1.7 mg/mL, respectively. ARE exerted suppressive activities on nitric oxide (NO) and ROS levels elevated by lipopolysaccharide (LPS) or tumor necrosis factor-α (TNF-α) in HaCaT keratinocytes. It attenuated the LPS-stimulated expression of iNOS. ARE augmented the UV-B-reduced filaggrin expression on both protein and mRNA levels and was capable of upregulating the UV-B-reduced caspase-14 activity. ARE inhibited in vitro elastase and hyaluronidase activities associated with the wrinkling process. ARE, at the concentrations used, did not interfere with the viability of HaCaT keratinocytes. These findings preliminarily imply that the leaves of A. rugosa possess desirable cosmetic potentials, such as anti-inflammatory, barrier protective, and antiwrinkle activities, which infers their skin healing potentials.

Emergence of Transferable mcr-9 Gene-Carrying Colistin-Resistant Salmonella enterica Dessau ST14 Isolated from Retail Chicken Meat in Korea.

Colistin is an important antibiotic currently used to manage infections caused by multidrug-resistant pathogens in both humans and livestock animals. A new mobile colistin-resistance (mcr-9) gene was recently discovered; this discovery highlighted the need for rigorous monitoring of bacterial resistance against colistin. Salmonella is one of the major pathogens responsible for foodborne illnesses; however, there is minimal information regarding the presence of mcr genes in foodborne Salmonella strains. The aim of this study was to investigate the presence of mcr genes among 178 Salmonella strains isolated from chicken meat in Korea. Antimicrobial susceptibility was measured using the broth microdilution method. Bioinformatics characterization of colistin-resistant strains and genetic environment of the mcr-9 gene were analyzed using next-generation sequencing. Transferability of the mcr-9 carrying colistin-resistant Salmonella strain was tested using broth-mating conjugation. Thirteen of the 178 Salmonella isolates showed colistin resistance, but only one strain, Salmonella Dessau ST14 (KUFSE-SAL043) from a traditional chicken market in Korea, carried an mcr family gene, mcr-9. This strain also carried other acquired antimicrobial resistance genes such as blaTEM-1B, qnrS1, and aac(6')-Iaa. Only the IncX1 plasmid replicon type was detected in this strain. In the strain KUFSE-SAL043, the mcr-9 gene was located between two insertion sequences, IS903B and IS26, followed by the downstream regulatory genes qseB-like and qseC-like, which were located between IS1R and ΔIS1R. Conjugation tests revealed that the mcr-9 gene was successfully transferred to Escherichia coli J53 at a mean frequency of 2.03 × 10-7. This is the first report of a transferable mcr-9 gene in Salmonella isolated from chicken meat in Korea, highlighting the possibility of transfer of colistin resistance. Therefore, the wide use of colistin should be reconsidered, and a One Health perspective should be adopted to monitor the antimicrobial resistance of Enterobacteriaceae strains in humans, livestock, and the environment.

Complete genome sequence of Sphingobium sp. strain PAMC 28499 reveals a potential for degrading pectin with comparative genomics approach.

BACKGROUND: Spingobium sp. PAMC 28499 is isolated from the glaciers of Uganda. Uganda is a unique region where hot areas and glaciers coexist, with a variety of living creatures surviving, but the survey on them is very poor. The genetic character and complete genome information of Sphingobium strains help with environmental studies and the development of better to enzyme industry. OBJECTIVE: In this study, complete genome sequence of Spingobium sp. PAMC 28499 and comparative analysis of Spingobium species strains isolated from variety of the region. METHODS: Genome sequencing was performed using PacBio sequel single-molecule real-time (SMRT) sequencing technology. The predicted gene sequences were functionally annotated and gene prediction was carried out using the program NCBI non-redundant database. And using dbCAN2 and KEGG data base were degradation pathway predicted and protein prediction about carbohydrate active enzymes (CAZymes). RESULTS: The genome sequence has 64.5% GC content, 4432 coding protein coding genes, 61 tRNAs, and 12 rRNA operons. Its genome encodes a simple set of metabolic pathways relevant to pectin and its predicted degradation protein an unusual distribution of CAZymes with extracellular esterases and pectate lyases. CAZyme annotation analyses revealed 165 genes related to carbohydrate active, and especially we have found GH1, GH2, GH3, GH38, GH35, GH51, GH51, GH53, GH106, GH146, CE12, PL1 and PL11 such as known pectin degradation genes from Sphingobium yanoikuiae. These results confirmed that this Sphingobium sp. strain PAMC 28499 have similar patterns to RG I pectin-degrading pathway. CONCLUSION: In this study, isolated and sequenced the complete genome of Spingobium sp. PAMC 28499. Also, this strain has comparative genome analysis. Through the complete genome we can predict how this strain can store and produce energy in extreme environment. It can also provide bioengineered data by finding new genes that degradation the pectin.

Characterization of Extended-Spectrum ß-Lactamase-Producing Escherichia coli Isolated from Fresh Produce and Agricultural Environments in Korea.

ABSTRACT: This study was conducted to characterize Escherichia coli strains and evaluate the spread of antimicrobial resistance among these strains from fresh produce and farm environments in Korea. We then conducted phenotypic and genetic studies on antimicrobial-resistant isolates. We determined the genetic epidemiological characteristics of isolates that produced extended-spectrum ß-lactamase (ESBL) and confirmed plasmid transfer in isolates that carried blaCTX-M-type genes. E. coli strains were isolated from 8 samples of fresh produce and 152 samples from the farm environment collected from May 2014 to June 2016. Cephalosporin resistance was the most prevalent (61.8%) type of resistance among the isolates. Five ESBL-producing strains with high genetic homology with E. coli of human or livestock origin were identified. Lateral transfer of plasmids harboring blaCTX-M-type genes to transconjugants was successful. Two isolates from Chinese cabbage and from water samples collected from a nearby stream harbored the ISEcp1-blaCTX-M-55-orf477 operon and were confirmed as sequence type 1196 and the same type of plasmid replicon, suggesting that cross-contamination was highly likely. A high-risk clone of sequence type 69 (clonal complex 69) isolates was also recovered from the farm environment. This study provides genetic evidence that antimicrobial resistance factors in E. coli from farm environments originate in the clinic or in livestock, highlighting the fact that good agricultural practices in farming are important to inhibit the spread of antimicrobial resistance to bacteria on fresh produce.

The Genome Assembly and Annotation of the Southern Elephant Seal Mirounga leonina.

The southern elephant seal Mirounga leonina is the largest phocid seal and one of the two species of elephant seals. They are listed as 'least concern' by the International Union for Conservation of Nature (IUCN) Red List of Threatened Species 2015. Here, we have assembled the reference genome for M. leonina using the 10× chromium sequencing platform. The final genome assembly of M. leonina was 2.42 Gb long, with a contig N50 length of 54 Mb and a maximum length of 111.6 Mb. The M. leonina genome contained 20,457 predicted protein-coding genes and possessed 41.51% repeated sequences. The completeness of the M. leonina genome was evaluated using benchmarking universal single-copy orthologous genes (BUSCOs): the assembly was highly complete, containing 95.6% of the core set of mammalian genes. The high-quality genomic information on M. leonina will be essential for further understanding of adaptive metabolism upon repeated breath-hold dives and the exploration of molecular mechanisms contributing to its unique biochemical and physiological characteristics. The southern elephant seal genome project was deposited at NCBI (National Center for Biotechnology Information) under BioProject number PRJNA587380.

The complete mitochondrial genome of Macrourus whitsoni (Gadiformes, Macrouridae).

The complete mitochondrial genome of Macrourus witsoni was determined in this study by the Long-read Technology, such as PacBio Sequel System. The Long-read Technology, which can sequence continuously the whole vertebrate mitochondrial genome, allows more accurate genomes to be completed. The circular form of its mitochondrial genome was 16,714bp, which contained 13 protein-coding genes, 22 tRNA, and 2 rRNA. The gene orders of M.witsoni was identical to that of the other species of Macrouridae family. Phylogenetic analysis indicated M. witsoni was mostly close to C.kishinouyei in the Macrouridae family.

The complete mitochondrial genome of the Antarctic marbled rockcod, Notothenia rossii (Perciformes, Nototheniidae).

The complete mitochondrial genome of Notothenia rossii was obtained using PacBio Sequel long-read sequencing platform. The mitogenome of N. rossii was circular form and 18,274 bp long, which consists of 13 protein-coding genes, 24 tRNAs, 2 rRNAs, and non-coding control region. Particularly, we found duplicated tRNAThr and tRNAPro in addition to the typical 22 tRNAs. The phylogenetic tree revealed that N. rossii was most closely related to N. coriiceps among species in the Nototheniidae clade within the suborder Notothenioidei.

Complete genome sequencing of Shigella sp. PAMC 28760: Identification of CAZyme genes and analysis of their potential role in glycogen metabolism for cold survival adaptation.

Shigella sp. PAMC 28760 (isolated from Himantormia sp. lichen in Antarctica) is a gram-negative, non-sporulating bacterium that has cellulolytic and amylolytic characteristics as well as glycogen metabolic pathways. In this study, we isolated S. sp. PAMC 28760 from Antarctic lichen, and present the complete genome sequence with annotations describing its unique features. The genome sequence has 58.85% GC content, 4,278 coding DNA sequences, 85 tRNAs, and 22 rRNA operons. 16S rRNA gene sequence analyses revealed strain PAMC 28760 as a potentially new species of genus Shigella, showing various differences from pathogenic bacteria reported previously. dbCAN2 analyses revealed 91 genes related to carbohydrate-metabolizing enzymes. S. sp. PAMC 28760 likely degrades polysaccharide starch to obtain glucose for energy conservation. This study provides a foundation for understanding Shigella survival adaptation mechanisms under extremely cold Antarctic conditions.

Mussel adhesive Protein-conjugated Vitronectin (fp-151-VT) Induces Anti-inflammatory Activity on LPS-stimulated Macrophages and UVB-irradiated Keratinocytes.

BACKGROUND: Skin inflammation and dermal injuries are a major clinical problem because current therapies are limited to treating established scars, and there is a poor understanding of healing mechanisms. Mussel adhesive proteins (MAPs) have great potential in many tissue engineering and biomedical applications. It has been successfully demonstrated that the redesigned hybrid type MAP (fp-151) can be utilized as a promising adhesive biomaterial. The aim of this study was to develop a novel recombinant protein using fp-151 and vitronectin (VT) and to elucidate the anti-inflammatory effects of this recombinant protein on macrophages and keratinocytes. METHODS: Lipopolysaccharide (LPS) was used to stimulate macrophages and UVB was used to stimulate keratinocytes. Inducible nitric oxide synthase (iNOS) and cyclooxygenase (COX)-2 were analyzed by Western Blot. Inflammatory cytokines and NO and ROS production were analyzed. RESULT: In macrophages stimulated by LPS, expression of the inflammatory factors iNOS, COX-2, and NO production increased, while the r-fp-151-VT-treated groups had suppressed expression of iNOS, COX-2, and NO production in a dose-dependent manner. In addition, keratinocytes stimulated by UVB and treated with r-fp-151-VT had reduced expression of iNOS and COX-2. Interestingly, in UVB-irradiated keratinocytes, inflammatory cytokines, such as interleukin (IL)-1b, IL-6, and tumor necrosis factor (TNF)-a, were significantly reduced by r-fp-151-VT treatment. CONCLUSIONS: These results suggest that the anti-inflammatory activity of r-fp-151-VT was more effective in keratinocytes, suggesting that it can be used as a therapeutic agent to treat skin inflammation.

Crystal structure of hypothetical protein PA4202 from Pseudomonas aeruginosa PAO1 in complex with nitroethane as a nitroalkane substrate.

Nitroalkane oxidase (NAO) and nitronate monooxygenase (NMO) are two different types of nitroalkane oxidizing flavoenzymes identified in nature. A previous study suggested that the hypothetical protein PA4202 from Pseudomonas aeruginosa PAO1 is NMO and utilizes only anionic nitronates. However, the structural similarity between the PA4202 protein and Streptomyces ansochromogenes NAO has motivated investigation for what features of the two enzymes differentiate between the NAO and NMO activities. Herein, we report the crystal structure of PA4202 in a ternary complex with a neutral nitroethane (NE) and flavin mononucleotide (FMN) cofactor to elucidate the substrate recognition mechanism using a site-directed mutagenesis. The ternary complex structure indicates that the NE is bound with an orientation, which is poised for the proton transfer to H183 (which is the essential first catalytic step with nitroalkanes), and subsequent reactions with FMN. Moreover, a kinetic study reveals that the catalytic reactions of the wild type and H183 mutants PA4202s with nitroalkane substrates may yield the products of hydrogen peroxide and nitrite that are specified to NAO, although they show a low catalytic efficiency. Our results provide the first structure-based molecular insight into the substrate binding property of the hypothetical protein PA4202, including the interactions with neutral nitroalkanes as the substrate.

Crystal structure of oleate hydratase from Stenotrophomonas sp. KCTC 12332 reveals conformational plasticity surrounding the FAD binding site.

Unsaturated fatty acids are toxic to various bacteria, causing their death or growth inhibition. To prevent this toxicity, unsaturated fatty acids should be converted into saturated fatty acids via hydrogenation reaction, which is the complete reduction of double bonds on the carbon chain. In a recent report, we observed that Stenotrophomonas sp. KCTC 12332 exhibited a high biotransformation activity of oleic acid (OA) in 10-hydroxystearic acid and identified the gene encoding oleate hydratase (OhySt) by complete genomic analysis. In the present study, to further investigate the structural features of OhySt, the recombinant protein was expressed in Escherichia coli, and then purified and crystallized. Biochemical assay showed that OhySt produces 10-hydroxystearic acid in a flavin adenosine dinucleotide (FAD)-dependent manner, indicating that it requires FAD as a cofactor. The OhySt structure, which is determined in its apo state, allows for a structural comparison with the previously reported FAD bound structure of oleate hydratase. The comparison of structures indicates remarkable conformational change of the loop region surrounding the FAD molecule upon binding of FAD. This change forces one of the important catalytic residues into position for catalysis.

Simultaneous determination of ethyl carbamate and urea in Korean rice wine by ultra-performance liquid chromatography coupled with mass spectrometric detection.

In this study, a rapid method for simultaneous detection of ethyl carbamate (EC) and urea in Korean rice wine was developed. To achieve quantitative analysis of EC and urea, the conditions for Ultra-performance liquid chromatography (UPLC) separation and atmospheric-pressure chemical ionization tandem mass spectrometry (APCI-MS/MS) detection were first optimized. Under the established conditions, the detection limit, relative standard deviation and linear range were 2.83µg/L, 3.75-5.96%, and 0.01-10.0mg/L, respectively, for urea; the corresponding values were 0.17µg/L, 1.06-4.01%, and 1.0-50.0µg/L, respectively, for EC. The correlation between the contents of EC and its precursor urea was determined under specific pH (3.5 and 4.5) and temperature (4, 25, and 50°C) conditions using the developed method. As a result, EC content was increased with greater temperature and lower pH. In Korean rice wine, urea was detected 0.19-1.37mg/L and EC was detected 2.0-7.7µg/L. The method developed in this study, which has the advantages of simplified sample preparation, low detection limits, and good selectivity, was successfully applied for the rapid analysis of EC and urea.