Comparison of physiologically functional compounds in Sika deer Cervus nippon meats obtained from different regions in Japan.

In Japan, the promotion of effective use of many wild deer as food resource has been conducted. However, they are not necessarily utilized effectively. Thus, we focused physiologically functional compounds to find characteristics of Sika deer meats (commercially available) obtained from different regions such as Hokkaido, Wakayama, Tokushima, and Miyazaki prefectures in Japan, making it a valuable resource for future studies and applications. The amount of carnosine, anserine, and balenine in muscle of deer from Wakayama prefecture was significantly lower than that in muscle of deer from other prefectures. The differences of amount of imidazole dipeptides in different prefectures seems to be caused by feed, rearing environment, and breed. The amount of carnitine in deer meat from Hokkaido was significantly lower than that in muscle of deer from other prefectures, while the amount of acetyl-carnitine in deer meat from Miyazaki prefectures was significantly higher than that from other prefectures. The amounts of glutamine, ornithine, and 3-methylhistidine in muscles of deer from Wakayama prefectures were significantly higher than those in muscle of deer from other prefectures. These results might be caused by differences in feeding habits, habitat, the muscle types, and subspecies of deer obtained from four regions in Japan.

Comparative analysis of hepatic transcriptomes and metabolomes of Changshun green-shell laying hens based on different green eggshell color intensities.

The eggshell color of avian species is an important trait that is predominantly determined by the pigments biliverdin and protoporphyrin. Various factors affect eggshell pigment deposition and coloration; however, the underlying mechanisms remain unclear. We analyzed the hepatic transcriptomes and metabolomes of Changshun green-shell hens laying dark green and light green eggs to investigate the potential role of the liver in regulating the intensity of the green eggshell color. In total, 350 differentially expressed genes and 211 differentially altered metabolites were identified. Gene set enrichment analysis revealed that the enriched pathways and Gene Ontology (GO) terms were mainly associated with energy, immunity, and nutrient metabolism. Metabolite set enrichment analysis revealed that the enriched pathways were mainly associated with amino acid, vitamin, bile acid, and lipid metabolism. Moreover, gene-metabolite interaction network analysis revealed 1 subnetwork. Most genes and metabolites in this subnetwork were determined to be related to melanin metabolism and transport. In conclusion, our results suggest that hepatic melanin metabolism and transport are critical for eggshell coloration. Six candidate genes (CDKN2B, DDC, PYCR1, ABCG5, SLC3A1, and P2RX2) and 7 candidate metabolites (serotonin, 5-hydroxyindoleacetic acid, ornithine, acetylcholine, L-tryptophan, D-ornithine, and ADP) were suggested to play important roles in this process. Meanwhile, this study suggests that changes in hepatic energy metabolism, immune status, antioxidation activity, nutrient availability, and bile acid synthesis can impair eggshell coloration.

Effective mRNA Protection by Poly(l-ornithine) Synergizes with Endosomal Escape Functionality of a Charge-Conversion Polymer toward Maximizing mRNA Introduction Efficiency.

For efficient delivery of messenger (m)RNA, delivery carriers need two major functions: protecting mRNA from nucleases and translocating mRNA from endolysosomes to the cytoplasm. Herein, these two complementary functionalities are integrated into a single polyplex by fine-tuning the catiomer chemical structure and incorporating the endosomal escape modality. The effect of the methylene spacer length on the catiomer side chain is evaluated by comparing poly(l-lysine) (PLL) with a tetramethylene spacer and poly(L-ornithine) (PLO) with a trimethylene spacer. Noteworthily, the nuclease stability of the mRNA/catiomer polyplexes is largely affected by the difference in one methylene group, with PLO/mRNA polyplex showing enhanced stability compared to PLL/mRNA polyplex. To introduce the endosomal escape function, the PLO/mRNA polyplex is wrapped with a charge-conversion polymer (CCP), which is negatively charged at extracellular pH but turns positive at endosomal acidic pH to disrupt the endosomal membrane. Compared to the parent PLO/mRNA polyplex, CCP facilitated the endosomal escape of the polyplex in cultured cells to improve the protein expression efficiency from mRNA by approximately 80-fold. Collectively, this system synergizes the protective effect of PLO against nucleases and the endosomal escape capability of CCP in mRNA delivery.

Determination of polyamines and related compounds in saliva via in situ derivatization and microextraction by packed sorbents coupled to GC-MS.

Here we show the determination of different polyamines (putrescine, cadaverine, spermidine) and related compounds (gamma-aminobutyric acid and l-ornithine) in saliva samples. These compounds are known to be biomarkers for several diseases. We have optimised an in situ derivatization process using ethyl chloroformate, an automated microextraction by packed sorbent and the determination of the corresponding products using a programmed temperature vaporizer coupled to a gas chromatograph - mass spectrometer. After finding that saliva matrix has an effect on the analysis, quantitation was performed using the one-point standard additions method and normalization to IS. This allows the detection of the analytes in the range of µg/L within a matrix obtained by a non-invasive procedure. The method has been successfully validated and it has been used in the determination of these compounds in six saliva samples finding that putrescine and cadaverine present the highest concentrations in the subject diagnosed with rheumatoid arthritis. For ornithine and spermidine, the highest concentrations were found for male subjects, especially heavy smokers. All concentrations found for the compounds were in good agreement with data found in bibliography.

Staphylococcus aureus exhibits heterogeneous siderophore production within the vertebrate host.

Siderophores, iron-scavenging small molecules, are fundamental to bacterial nutrient metal acquisition and enable pathogens to overcome challenges imposed by nutritional immunity. Multimodal imaging mass spectrometry allows visualization of host-pathogen iron competition, by mapping siderophores within infected tissue. We have observed heterogeneous distributions of Staphylococcus aureus siderophores across infectious foci, challenging the paradigm that the vertebrate host is a uniformly iron-depleted environment to invading microbes.

Production of sheep milk cheese with high γ-aminobutyric acid and ornithine concentration and with reduced biogenic amines level using autochthonous lactic acid bacteria strains.

Consumer demand for health-promoting foods is generating the need to develop biofunctional dairy products. Lactic acid bacteria are employed in cheese-making and some of them are able to produce beneficial compounds on human health such as γ-aminobutyric acid (GABA) and ornithine but also to synthetize biogenic amines. The aim was to investigate the effect of four selected autochthonous co-cultures on the free amino acid profile, with special emphasis on GABA and ornithine, and on the biogenic amine content of pasteurized sheep milk cheese during ripening. High average concentrations of GABA (1296.75 mg/kg cheese) and ornithine (2355.76 mg/kg cheese) were found in all the cheese batches at 240 days of ripening. Batch 2, manufactured with the co-culture containing autochthonous Lactococcus lactis strains as starter and Lactobacillus plantarum TAUL1588 as adjunct, showed 2.37 fold reduced biogenic amines concentration with respect to the batch 1 made with the starter during the ripening time. The microstructure and microbiological counts of cheeses were affected (P ≤ 0.001) by the ripening time, without appreciating differences (P ≥ 0.05) in the physico-chemical composition between batches. This study could be a good approach to the development of functional sheep milk cheese.

Screening of medium constituents for clavulanic acid production by Streptomyces clavuligerus

ABSTRACT Clavulanic acid is a β-lactam compound with potent inhibitory activity against β-lactamases. Studies have shown that certain amino acids play essential roles in CA biosynthesis. However, quantitative evaluations of the effects of these amino acids are still needed in order to improve CA production. Here, we report a study of the nutritional requirements of Streptomyces clavuligerus for CA production. Firstly, the influence of the primary nitrogen source and the salts composition was investigated. Subsequently, soybean protein isolate was supplemented with arginine (0.0-3.20 g L-1), threonine (0.0-1.44 g L-1), ornithine (0.0-4.08 g L-1), and glutamate (0.0-8.16 g L-1), according to a two-level central composite rotatable design. A medium containing ferrous sulfate yielded CA production of 437 mg L-1, while a formulation without this salt produced only 41 mg L-1 of CA. This substantial difference suggested that Fe2+ is important for CA biosynthesis. The experimental design showed that glutamate and ornithine negatively influenced CA production while arginine and threonine had no influence. The soybean protein isolate provided sufficient C5 precursor for CA biosynthesis, so that supplementation was unnecessary. Screening of medium components, together with experimental design tools, could be a valuable way of enhancing CA titers and reducing the process costs.

Characterization of Arginine Catabolism by Lactic Acid Bacteria Isolated from Kimchi.

Kimchi fermentation depends on diverse lactic acid bacteria, which convert raw materials into numerous metabolites that contribute to the taste of food. Amino acids and saccharides are important primary metabolites. Arginine is nearly exhausted during kimchi fermentation, whereas the concentrations of other amino acids are reported not to increase or decrease dramatically. These phenomena could imply that arginine is an important nutritional component among the amino acids during kimchi fermentation. In this study, we investigated the arginine-catabolism pathway of seven lactic acid bacteria isolated from kimchi and evaluated the products of arginine catabolism (citrulline and ornithine) associated with the bacteria. The arginine content dramatically decreased in cultures of Lactobacillus brevis and Weissella confusa from 300 µg/mL of arginine to 0.14 ± 0.19 and 1.3 ± 0.01 µg/mL, respectively, after 6 h of cultivation. Citrulline and ornithine production by L. brevis and W. confusa showed a pattern that was consistent with arginine catabolism. Interestingly, Pediococcus pentosaceus, Lactobacillus plantarum, Leuconostoc mesenteroides, and Leuconostoc lactis did not show increased citrulline levels after arginine was added. The ornithine contents were higher in all bacteria except for L. lactis after adding arginine to the culture. These results were consistent with the absence of the arginine deiminase gene among the lactic acid bacteria. Arginine consumption and ornithine production were monitored and compared with lactic acid bacteria by metagenomics analysis, which showed that the increment of ornithine production correlated positively with lactic acid bacteria growth.

Explorative investigation of the anti-glycative effect of a rapeseed by-product extract.

The formation of advanced glycation end-products (AGEs) in biological systems is increased during hyperglycaemia due to higher levels of circulating glucose and carbonyl reactive species. AGEs are causative factors of common chronic diseases. Since synthetic AGE-inhibitors exert unwanted side effects and polyphenols act as potent antiglycative agents, vegetables (fruits, seeds and related by-products) are good candidates when searching for natural inhibitors. The aim of this research is to explore the suitability of a polyphenol-rich rapeseed cake extract (RCext) to decrease the formation of AGEs in an in vitro model. Different phenols, amino acids, carbohydrates, organic acids and fatty acids were identified in the RCext by GC-MS. The results confirm a high concentration of polyphenols (73.85 ± 0.64 and 86.85 ± 2.08 mg of gallic acid equivalents per g of RCext spray dried and freeze dried, respectively) which is correlated with the antioxidant capacity and anti-glycative activity in a dose dependent manner. Rapeseed cake extract (3.7 mg mL-1) significantly reduced the formation of free fluorescent AGEs and pentosidine up to 34.85%. The anti-glycative activity of the extract is likely to be due to the high concentration of sinapinic acid (0.108 ± 0.0043 mg g-1) in its metabolic profile, and the mechanism of action is mediated by methylglyoxal trapping. The results show promising potential for using rapeseed cake extract as a food supplement to ameliorate the formation of AGEs. Rapeseed cake extract should therefore be considered a potential candidate for the prevention of glycation-associated complications of age-related pathologies.

Expanding the Coverage of the Metabolome with Nitrogen-Based NMR.

Isotopically labeling a metabolite and tracing its metabolic fate has provided invaluable insights about the role of metabolism in human diseases in addition to a variety of other issues. 13C-labeled metabolite tracers or unlabeled 1H-based NMR experiments are currently the most common application of NMR to metabolomics studies. Unfortunately, the coverage of the metabolome has been consequently limited to the most abundant carbon-containing metabolites. To expand the coverage of the metabolome and enhance the impact of metabolomics studies, we present a protocol for 15N-labeled metabolite tracer experiments that may also be combined with routine 13C tracer experiments to simultaneously detect both 15N- and 13C-labeled metabolites in metabolic samples. A database consisting of 2D 1H-15N HSQC natural-abundance spectra of 50 nitrogen-containing metabolites are also presented to facilitate the assignment of 15N-labeled metabolites. The methodology is demonstrated by labeling Escherichia coli and Staphylococcus aureus metabolomes with 15N1-ammonium chloride, 15N4-arginine, and 13C2-acetate. Efficient 15N and 13C metabolite labeling and identification were achieved utilizing standard cell culture and sample preparation protocols.

Screening of medium constituents for clavulanic acid production by Streptomyces clavuligerus.

Clavulanic acid is a ß-lactam compound with potent inhibitory activity against ß-lactamases. Studies have shown that certain amino acids play essential roles in CA biosynthesis. However, quantitative evaluations of the effects of these amino acids are still needed in order to improve CA production. Here, we report a study of the nutritional requirements of Streptomyces clavuligerus for CA production. Firstly, the influence of the primary nitrogen source and the salts composition was investigated. Subsequently, soybean protein isolate was supplemented with arginine (0.0-3.20gL-1), threonine (0.0-1.44gL-1), ornithine (0.0-4.08gL-1), and glutamate (0.0-8.16gL-1), according to a two-level central composite rotatable design. A medium containing ferrous sulfate yielded CA production of 437mgL-1, while a formulation without this salt produced only 41mgL-1 of CA. This substantial difference suggested that Fe2+ is important for CA biosynthesis. The experimental design showed that glutamate and ornithine negatively influenced CA production while arginine and threonine had no influence. The soybean protein isolate provided sufficient C5 precursor for CA biosynthesis, so that supplementation was unnecessary. Screening of medium components, together with experimental design tools, could be a valuable way of enhancing CA titers and reducing the process costs.

DAF in diabetic patients is subject to glycation/inactivation at its active site residues.

Decay accelerating factor (DAF or CD55) is a cell associated C3 and C5 convertase regulator originally described in terms of protection of self-cells from systemic complement but now known to modulate adaptive T cell responses. It is expressed on all cell types. We investigated whether nonenzymatic glycation could impair its function and potentially be relevant to complications of diabetes mellitus and other conditions that result in nonenzymatic glycation including cancer, Alzheimer's disease, and aging. Immunoblots of affinity-purified DAF from erythrocytes of patients with diabetes showed pentosidine, glyoxal-AGEs, carboxymethyllysine, and argpyrimidine. HPLC/MS analyses of glucose modified DAF localized the sites of AGE modifications to K125 adjacent to K126, K127 at the junction of CCPs2-3 and spatially near R96, and R100, all identified as being critical for DAF's function. Functional analyses of glucose or ribose treated DAF protein showed profound loss of its regulatory activity. The data argue that de-regulated activation of systemic complement and de-regulated activation of T cells and leukocytes could result from non-enzymatic glycation of DAF.

UV-B induced biosynthesis of a novel sunscreen compound in solar radiation and desiccation tolerant cyanobacteria.

The small-molecule sunscreen compounds, mycosporine-like amino acids (MAAs), have strong ultraviolet (UV) absorption and can protect cyanobacteria against UV-B damage. However, the molecular mechanism underlying UV-B signaling and MAA chemical diversity remain largely unclear. Here, we identified a five-gene cluster for MAA biosynthesis in the solar radiation and desiccation tolerant cyanobacterium Nostoc flagelliforme. A LuxR family protein OrrA was identified as a positive UV-B responsive regulator binding to the promoter region of this gene cluster. OrrA functions as an activator mediating the UV-B induced MAA biosynthesis. Overexpression of orrA strengthened its UV-B tolerance during desiccation, and enhanced the photosynthetic recovery upon rehydration. Heterologous expression of this gene cluster in Anabaena PCC 7120 produces the same MAA as that in field samples of N. flagelliforme. The MAA structure is assigned as mycosporine-2-(4-deoxygadusolyl-ornithine) with a molecular weight of 756 Da, the structurally unique MAA compound reported to date. This MAA was catalyzed by mysD-mysC2-mysC1 encoding proteins from 4-deoxygadusol, which was synthesized through the catalysis of mysA-mysB products. Thus, we elucidated the transcriptional mechanism for a novel type MAA biosynthesis in solar radiation and desiccation tolerant cyanobacteria, which shed light on the identification of other components for UV-B signaling in cyanobacteria.

Simultaneous Quantitation of Advanced Glycation End Products in Soy Sauce and Beer by Liquid Chromatography-Tandem Mass Spectrometry without Ion-Pair Reagents and Derivatization.

The aim of this study was to develop a simple and sensitive method to analyze several advanced glycation end products (AGEs) simultaneously using liquid chromatography-tandem mass spectrometry (LC-MS/MS), and to apply this method to the quantitation of AGEs in brown-colored foods. The developed method enabled to separate and quantitate simultaneously seven AGEs, and was applied to the determination of free AGEs contained in various kinds of soy sauce and beer. The major AGEs in soy sauce and beer were Nε-carboxymethyllysine (CML), Nε-carboxyethyllysine (CEL), and Nδ-(5-hydro-5-methyl-4-imidazolon-2-yl)ornithine (MG-H1). Using the developed LC-MS/MS method, recovery test on soy sauce and beer samples showed the recovery values of 85.3-103.9% for CML, 95.9-107.4% for CEL, and 69.5-123.2% for MG-H1. In particular, it is the first report that free CML, CEL, and MG-H1 were present in beer. Furthermore, long-term storage and heating process of soy sauce increased CML and MG-H1.

Free and Protein-Bound Maillard Reaction Products in Beer: Method Development and a Survey of Different Beer Types.

The Maillard reaction is important for beer color and flavor, but little is known about the occurrence of individual glycated amino acids in beer. Therefore, seven Maillard reaction products (MRPs), namely, fructosyllysine, maltulosyllysine, pyrraline, formyline, maltosine, MG-H1, and argpyrimidine, were synthesized and quantitated in different types of beer (Pilsner, dark, bock, wheat, and nonalcoholic beers) by HPLC-ESI-MS/MS in the multiple reaction monitoring mode through application of the standard addition method. Free MRPs were analyzed directly. A high molecular weight fraction was isolated by dialysis and hydrolyzed enzymatically prior to analysis. Maltulosyllysine was quantitated for the first time in food. The most important free MRPs in beer are fructosyllysine (6.8-27.0 mg/L) and maltulosyllysine (3.7-21.8 mg/L). Beer contains comparatively high amounts of late-stage free MRPs such as pyrraline (0.2-1.6 mg/L) and MG-H1 (0.3-2.5 mg/L). Minor amounts of formyline (4-230 µg/L), maltosine (6-56 µg/L), and argpyrimidine (0.1-4.1 µg/L) were quantitated. Maltulosyllysine was the most significant protein-bound MRP, but both maltulosyllysine and fructosyllysine represent only 15-60% of the total protein-bound lysine-derived Amadori products. Differences in the patterns of protein-bound and free individual MRPs and the ratios between them were identified, which indicate differences in their chemical, biochemical, and microbiological stabilities during the brewing process.

An Analysis of Responses to Defibrotide in the Pulmonary Vascular Bed of the Cat.

Defibrotide is a polydisperse mixture of single-stranded oligonucleotides with many pharmacologic properties and multiple actions on the vascular endothelium. Responses to defibrotide and other vasodepressor agents were evaluated in the pulmonary vascular bed of the cat under conditions of controlled pulmonary blood flow and constant left atrial pressure. Lobar arterial pressure was increased to a high steady level with the thromboxane A2 analog U-46619. Under increased-tone conditions, defibrotide caused dose-dependent decreases in lobar arterial pressure without altering systemic arterial and left atrial pressures. Responses to defibrotide were significantly attenuated after the administration of the cyclooxygenase inhibitor sodium meclofenamate. Responses to defibrotide were also significantly attenuated after the administration of both the adenosine 1 and 2 receptor antagonists 8-cyclopentyl-1,3-dimethylxanthine and 8-(3-chlorostyryl)caffeine. Responses to defibrotide were not altered after the administration of the vascular selective adenosine triphosphate-sensitive potassium channel blocker U-37883A, or after the administration of the nitric oxide synthase inhibitor L-N-(1-iminoethyl)-ornithine. These data show that defibrotide has significant vasodepressor activity in the pulmonary vascular bed of the cat. They also suggest that pulmonary vasodilator responses to defibrotide are partially dependent on both the activation of the cyclooxygenase enzyme and adenosine 1 and 2 receptor pathways and independent of the activation of adenosine triphosphate-sensitive potassium channels or the synthesis of nitric oxide in the pulmonary vascular bed of the cat.

Profiling of ornithine lipids in bacterial extracts of Rhodobacter sphaeroides by reversed-phase liquid chromatography with electrospray ionization and multistage mass spectrometry (RPLC-ESI-MS(n)).

Ornithine lipids (OLs), a sub-group of the large (and of emerging interest) family of lipoamino acids of bacterial origin, contain a 3-hydroxy fatty acyl chain linked via an amide bond to the α-amino group of ornithine and via an ester bond to a second fatty acyl chain. OLs in extracts of Rhodobacter sphaeroides (R. sphaeroides) were investigated by high-performance reversed phase liquid chromatography (RPLC) with electrospray ionization mass spectrometry (ESI-MS) in negative ion mode using a linear ion trap (LIT). The presence of OLs bearing both saturated (i.e, 16:0, 17:0, 18:0, 19:0 and 20:0) and unsaturated chains (i.e., 18:1, 19:1, 19:2 and 20:1) was ascertained and their identification, even for isomeric, low abundance and partially co-eluting species, was achieved by low-energy collision induced dissociation (CID) multistage mass spectrometry (MS(n), n = 2-4). OLs signatures found in two R. sphaeroides strains, i.e., wild type 2.4.1 and mutant R26, were examined and up to 16 and 17 different OL species were successfully identified, respectively. OLs in both bacterial strains were characterized by several combinations of fatty chains on ester-linked and amide-linked 3-OH fatty acids. Multistage MS spectra of monoenoic amide-linked 3-OH acyl chains, allowed the identification of positional isomer of OL containing 18:1 (i.e. 9-octadecenoic) and 20:1 (i.e. 11-eicosenoic) fatty acids. The most abundant OL ([M-H](-) at m/z 717.5) in R. sphaeroides R26 was identified as OL 3-OH 20:1/19:1 (i.e., 3-OH-eicosenoic acid amide-linked to ornithine and esterified to a nonadecenoic chain containing a cyclopropane ring). An unusual OL (m/z 689.5 for the [M-H](-) ion), most likely containing a cyclopropene ester-linked acyl chain (i.e., OL 3-OH 18:0/19:2), was retrieved only in the carotenoidless mutant strain R26. Based on the biosynthetic pathways already known for cyclopropa(e)ne ring-including acyl chains, a plausible explanation was invoked for the enzymatic generation of this ester-linked chain in R. sphaeroides.

Mesorhizobium soli sp. nov., a novel species isolated from the rhizosphere of Robinia pseudoacacia L. in South Korea by using a modified culture method.

Strain NHI-8(T) was isolated from a forest soil sample, collected in South Korea, by using a modified culture method. Comparative analysis of its nearly full-length 16S rRNA gene sequence showed that strain NHI-8(T) belongs to the genus Mesorhizobium and to be closely related to Mesorhizobium chacoense PR5(T) (97.32 %). The levels of DNA-DNA relatedness between strain NHI-8(T) and reference type strains of the genus Mesorhizobium were 32.28-53.65 %. SDS-PAGE of total soluble proteins and the sequences of the housekeeping genes recA, glnII, and atpD were also used to support the clade grouping in rhizobia. The new strain contained summed feature 8 (57.0 %), cyclo-C19:0ω8c (17.3 %), and C18:0 (11.0 %) as the major fatty acids, as in genus Mesorhizobium. The strain contained cardiolipin, phosphatidylglycerol, ornithine-containing lipid, phosphatidylethanolamine, phosphatidyl-N-dimethylethanolamine, and phosphatidylcholine. Morphological and physiological analyses were performed to compare the characteristics of our strain with those of the reference type strains. Based on the results, strain NHI-8(T) was determined to represent a novel member of the genus Mesorhizobium, and the name Mesorhizobium soli is proposed. The type strain is NHI-8(T) (=KEMB 9005-153(T) = KACC 17916(T) = JCM 19897(T)).

Phenotypes of Escherichia coli isolated from urine: Differences between extended-spectrum ß-lactamase producers and sensitive strains.

BACKGROUND: Escherichia coli is a frequent causative agent of urinary tract infections, and increasing resistance of E. coli to antimicrobials presents a growing challenge. METHODS: Here we compare phenotypes of extended-spectrum ß-lactamase (ESBL) producers (n = 220) with a control group of sensitive strains (non-ESBL producers; n = 150). For each strain, we assessed the presence of O25 antigen, hemolysis, biofilm production, sensitivity to antibiotics, and biochemical profile. RESULTS: Compared to the control group, ESBL producers were more frequently O25 positive (6.0% vs. 42.3%) and less frequently hemolytic (34.7% vs. 6.4%). Comparison of biofilm production in brain-heart infusion (BHI) and in BHI with 4% glucose supplementation showed that ESBL-positive strains produced biofilm in BHI with glucose less intensely than the control group (p < 0.05). Most ESBL producers were ciprofloxacin-resistant (91.8%). Biochemical analyses revealed that ESBL producers more frequently utilized inositol, ornithine, sorbitol, melibiose, and saccharose, whereas the control group more frequently used esculin, lysine, arginine, and dulcitol. The control group strains with O25 antigen were more commonly resistant to ciprofloxacin (p < 0.05). Pulsed-field gel electrophoresis results showed higher variability among the control group of sensitive strains. CONCLUSION: These findings suggest a potential to detect ESBL strains based on virulence factors and biochemical properties, which could be useful in shaping proper empiric antimicrobial therapy, and for initiating such therapy as soon as possible.

Sensitive determination of taurine, γ-aminobutyric acid and ornithine in wolfberry fruit and cortex lycii by HPLC with fluorescence detection and online mass spectrometry identification.

A new, simple and highly sensitive method for the determination of taurine, γ-aminobutyric acid and ornithine using high-performance liquid chromatography (HPLC) with fluorescence detection is described. Three non-protein amino acids were derivatized by a novel precolumn derivatization reagent 2-[2-(dibenzocarbazol)-ethoxy]ethyl chloroformate before injected. Optimum derivatization was obtained at 40°C for 5 min in the presence of sodium borate buffer (pH 9.0). Derivatives were sufficiently stable to be efficiently analyzed by HPLC without pretreatment. On a reversed-phase Hypersil BDS C8 column, the amino acids were separated in conjunction with a gradient elution with a good baseline resolution. The identification of derivatives was carried out by online postcolumn mass spectrometry with an electrospray ionization source in positive ion mode. Excellent linear responses were observed with the correlation coefficients of >0.9996, and instrument detection limits (at a signal to noise of 3 : 1) were in the range of 0.30-0.33 nmol/L. The proposed method is sensitive and reproducible for the precise determination of the amino acids from wolfberry fruit and cortex lycii samples.