RESUMEN
Amino acids with various functions are abundant in living organisms and foods. Recent advances in analytical technology show that trace amounts of D-amino acids exist in living organisms and foods. In addition, studies show that these amino acids are involved in various physiological functions that differ from those of L-amino acids. Thus, a technique for analyzing DL-amino acids is required. However, the simultaneous separation and highly sensitive detection of DL-amino acids are complicated; therefore, highly sensitive analytical methods that can rapidly separate and identify compounds are required. We previously developed our original chiral resolution labeling reagents for the separation and highly sensitive detection of DL-amino acids. Here, we developed a simple method for the rapid separation and highly sensitive detection of DL-amino acids in various foods and beverages by liquid chromatography-mass spectrometry (LC-MS) using an octadecyl (C18) column after labeling with 1-fluoro-2,4-dinitrophenyl-5-D-leucine-N,N-dimethylethylenediamineamide (D-FDLDA; enantiomeric excess > 99.9 %). In addition, we synthesized a stable isotope (13C6)-labeled D-FDLDA (13C6-D-FDLDA) and established an analytical method that can accurately identify the peak of each DL-amino acid. MS sensitivity of DL-amino acids labeled with our labeling reagent was higher than that of conventional labeling reagents (Marfey's reagents). The labeling reagent was neither desorbed from each DL-amino acid nor degraded for at least 1 week at 4 °C. Furthermore, we determined the DL-amino acid contents in foods and beverages using the proposed method, and differences in the total amino acid content and D/L ratio in each food and beverage were observed.
Asunto(s)
Aminoácidos , Bebidas , Análisis de los Alimentos , Aminoácidos/análisis , Aminoácidos/química , Bebidas/análisis , Análisis de los Alimentos/métodos , Estereoisomerismo , Límite de Detección , Reproducibilidad de los Resultados , Cromatografía Liquida/métodos , Leucina/química , Leucina/análisis , Espectrometría de Masas/métodos , Modelos LinealesRESUMEN
Recently, amino acid derivatives gradually gained attention, but studies on N-lactoyl-leucine (Lac-Leu) and N-lactoyl-isoleucine (Lac-Ile) are limited. This study aims to explore the contributions of Lac-Leu and Lac-Ile to soy sauce. Lac-Leu and Lac-Ile were synthesized via enzymatic synthesis method catalyzed by Tgase. The mixed solutions containing Lac-Leu were found to have greater taste improvement than those containing Lac-Ile. Sensory evaluation indicated the sour, bitter, and astringent taste of Lac-Leu in water as well as its kokumi, astringent, and umami-enhancing taste in MSG solution. The taste threshold and umami-enhancing threshold of Lac-Leu measured by TDA and cTDA, respectively, were 0.08 mg/mL and 0.16 mg/mL. Molecular docking of Lac-Leu and Lac-Ile with the kokumi receptor CaSR and the umami receptors T1R1 and T1R3 indicated that Lac-Leu had higher affinities with receptors than Lac-Ile. These findings demonstrated the underlying contribution Lac-Leu made to soy sauce, indicating its potential to improve the flavor quality of soy sauce.
Asunto(s)
Aromatizantes , Leucina , Alimentos de Soja , Espectrometría de Masas en Tándem , Gusto , Alimentos de Soja/análisis , Humanos , Leucina/química , Leucina/análisis , Aromatizantes/química , Cromatografía Líquida de Alta Presión , Simulación del Acoplamiento Molecular , Adulto , Masculino , Femenino , Receptores Acoplados a Proteínas G/química , Receptores Acoplados a Proteínas G/metabolismo , Cromatografía Líquida con Espectrometría de MasasRESUMEN
Understanding the nutritional content of protein supplements is crucial for optimal nutritional planning among athletes and other people. Distribution of macronutrients and aminograms in the main products available in the national Chilean market remains unknown. A descriptive cross-sectional study was conducted to identify the main protein supplements available in the Chilean market. Information on macronutrients and aminograms from the nutritional labels of each product was extracted. The analysis considered the content per portion and per 100 g. Cluster analysis models and graphical representations were explored. Eighty protein shakes were assessed in the Santiago de Chile market. The median protein dosage was 32 g (range from 25 to 52), and the median energy value stood at 390 kcal (range from 312 to 514). The median protein content per 100 g of product was found to be 75 g (range from 42.5 to 97.2). The combined median concentration of amino acids was 4749.75 mg. Among these, the essential amino acid L-Tryptophan exhibited the lowest concentration at 1591.50 mg, while the conditional amino acid L-Glutamine had the highest median concentration at 17,336 mg. There was a significant prevalence of animal-derived products, placing specific emphasis on protein supplements that feature elevated levels of the amino acids L-Glutamine and L-Leucine.
Asunto(s)
Proteínas en la Dieta , Suplementos Dietéticos , Valor Nutritivo , Chile , Estudios Transversales , Proteínas en la Dieta/análisis , Humanos , Aminoácidos/análisis , Etiquetado de Alimentos , Triptófano/análisis , Nutrientes/análisis , Leucina/análisis , Ingestión de Energía , Glutamina/análisisRESUMEN
BACKGROUND: Recent research has been reported that N-acetyl-leucine content is significantly reduced in the saliva of diabetic patients, but no reports of detection in human nails have been found. This study aims to develop a novel method for the chiral separation of N-acetyl-DL-leucine (Ac-DL-Leu) in human fingernails to investigate the differences between healthy volunteers (HVs), prediabetes (PDs) and diabetic patients (DPs), and to verify its effectiveness in early warning of diabetes. METHOD: Chiral resolution was performed using DBD-Apy pre-column derivatization on a C18 column (2.1 × 150 mm, 1.9 µm) at 40 °C, and detected by UPLC-ESI-MS/MS. RESULTS: The resolution and the limit of detection (LOD) of Ac-DL-Leu were 1.75 and 1.50 fmol, respectively. The linear range of Ac-DL-Leu was 10-2000 fmol and the determination coefficient (R2) was above 0.9997. The recovery of Ac-DL-Leu in human nails was 96.92-105.69 %. The contents of Ac-D-Leu and Ac-L-Leu were analyzed in 18 HVs, 13 PDs and 16 DPs fingernails. The results showed that their contents were significantly lower in DPs than in PDs and HVs (p < 0.0001). CONCLUSIONS: A method for evaluating the effectiveness of Ac-DL-Leu enantiomers in human fingernails as a biomarker for diabetes was firstly developed.
Asunto(s)
Diabetes Mellitus , Espectrometría de Masas en Tándem , Humanos , Espectrometría de Masas en Tándem/métodos , Leucina/análisis , Espectrometría de Masa por Ionización de Electrospray/métodos , Uñas/química , Cromatografía Líquida de Alta Presión/métodos , Diabetes Mellitus/diagnóstico , EstereoisomerismoRESUMEN
This study aimed to determine the association between the seed coat color of two chia seed genotypes for their composition, protein content, amino acid, and fatty acid profiles. The optimal pH for protein isolation for both genotypes (BCPI and WCPI) was 10, based on protein purity and solubility. Fatty acid profiling indicated, overall, 18 different fatty acids higher in BCPI10 with linolenic acid domination (â¼66%) followed by linoleic acid (â¼19%) and oleic acid (â¼6%), contributing PUFAs (â¼86%). Optimized protein isolates, black (BCPI10) and white (WCPI10) chia, had shown purity, L*-value, solubility, and yields of 90.65%, 75.86%, 77.75%, 11.30%, and 90.00%, 77.83%, 76.07%, 10.69%, respectively. BCPI10 depicted higher EAA (33.19 g/100 g N) and EEA indices (57.676%) compared to WCPI10 (32.14 g/100 g N) and 56.360%, respectively. Amino acid profiling indicated higher, PER, TAA, TEAA, TNEAA, TAAA, TBA, acidic AA values for BCPI10, and higher leucine/isoleucine ratio for WCPI10 having leucine and sulfur amino acids as limiting amino acids. BCPI10 had higher sulfur-containing amino acid contents, as the main contributor to the albumin a water-soluble fraction, leading to its higher in vitro digestibility (71.97%) than WCPI10 (67.70%). Both isolates exhibited good WHC and OHC of 3.18, 2.39 and 3.00, 2.20, respectively. Both protein isolates had similar ∆Td (°C) values with some variation in FTIR spectrum from 1000 cm-1 to 1651 cm-1 having more peak intensity for BCPI10. SDS-PAGE indicated bands at 150 kDa, representing globulin and mild bands at 25-33 kDa for glutelin and albumin. A significant (p < 0.05) variation reported in this study for protein and lipid profiles of both genotype attributes to genetic differences between the seeds. PRACTICAL APPLICATION: Based on the nutritional profile, both chia seed isolates (black and white) are suitable for consumption with an edge for black seed when supplemented with their limiting amino acids. The high values of the functional properties and structural characteristics combined with high nutritional values make the chia protein isolate an excellent source of raw material for various food formulations. Fatty acid profile of the oils from the genotypes showed the presence of high amounts of unsaturated fatty acids, especially the PUFAs with more number of fatty acids in black chia seed. The excellent lipid profile of chia seed oil indicates the benefit of using chia seed oil as a source of essential fatty acids in the human diet for optimal health.
Asunto(s)
Aminoácidos Sulfúricos , Salvia , Albúminas , Aminoácidos Sulfúricos/análisis , Ácidos Grasos/análisis , Ácidos Grasos Insaturados/análisis , Genotipo , Glútenes/análisis , Humanos , Isoleucina/análisis , Leucina/análisis , Ácidos Linoleicos/análisis , Aceites/análisis , Ácidos Oléicos/análisis , Salvia/química , Salvia/genética , Salvia hispanica , Semillas/química , Azufre/análisis , Agua/análisis , Ácido alfa-Linolénico/análisisRESUMEN
Plum (Prunus spp.) is an economically and nutritionally important stone fruit that is grown worldwide. Gummosis disease (GD) is one of the most common limiting factors that adversely affects the yield and quality of stone fruits such as plum. Elucidating plum fruit metabolomics responses is essential to develop sustainable agricultural practices to combat GD in the future. Herein, an ultra-high-performance liquid chromatography coupled to mass-spectrometry (UHPLC-MS) pseudo-targeted metabolomic profiling was first performed to elucidate the overall metabolic alterations in Asian plum (Prunus salicina Lindl.) fruit in response to GD. The most pivotal differential metabolites, including certain amino acids and proanthocyanidins, in GD and control groups were identified by combining multivariate data analysis with strict statistical criteria. Metabolic pathway enrichment analysis showed that GD induced a series of coordinated defence responses and reprogramming of various metabolic pathways, including glucosinolate biosynthesis, 2-oxocarboxylic acid metabolism, valine, leucine and isoleucine degradation, and isoquinoline alkaloid biosynthesis pathways. Using UHPLC-MS-based pseudo-targeted metabolomic profiling, we systematically evaluated overall metabolic modifications in Asian plum fruits in response to GD for the first time. The identified metabolic pathway alterations helped to better understand the internal relationships and related metabolic networks.
Asunto(s)
Alcaloides , Proantocianidinas , Prunus domestica , Alcaloides/análisis , Cromatografía Líquida de Alta Presión , Frutas/química , Glucosinolatos/análisis , Isoleucina/análisis , Isoquinolinas/análisis , Leucina/análisis , Espectrometría de Masas , Redes y Vías Metabólicas , Proantocianidinas/análisis , Prunus domestica/metabolismo , Valina/análisisRESUMEN
Protein de novo sequencing based on tandem mass spectrometry is a crucial technology that enables the identification of peptides without searching databases and assembling unknown sequence proteins, especially for monoclonal antibodies (mAbs). However, the discrimination of leucine (Leu) and isoleucine (Ile) residues in the target protein sequence is still challenging. Herein, we developed an accurate method by continuous digestion with MS3-based fragmentation and multiple spectra integration (evaluated by combined verification score, CVS) to distinguish Leu and Ile residues. Continuous digestion promotes the diversity of peptides in order to expose more Leu and Ile at the N-terminal. CVS integrates multiple MS3 spectra to reduce the interference from noise and co-fragmented ions and improve accuracy. This method successfully resolved all 75 Leu/Ile in bovine serum albumin, especially 3 consecutive Leu/Ile. We further applied the method to analyze trastuzumab and 67 out of the 68 Leu/Ile from the light chain and heavy chain were accurately discriminated, demonstrating the great potential in mAbs sequencing.
Asunto(s)
Isoleucina , Análisis de Secuencia de Proteína , Secuencia de Aminoácidos , Anticuerpos Monoclonales/química , Digestión , Isoleucina/análisis , Leucina/análisis , Péptidos/química , Análisis de Secuencia de Proteína/métodosRESUMEN
BACKGROUND: Branched-chain amino acids (BCAAs: isoleucine, leucine, and valine) and aromatic amino acids (AAAs: phenylalanine and tyrosine) are hypothesized to influence early-life obesity risk. OBJECTIVE: To assess HM free amino acid (AA) concentrations and infant intakes of HM AAs from women with obesity (OB) compared to those with normal weight (NW) and determine the relationships between HM AA consumption and infant growth. METHODS: HM samples were collected at 0.5 (n = 151), 2 (n = 129), and 6 (n = 93) months postpartum from mothers with NW (body mass index [BMI] = 18.5-24.9 kg/m2 ) and OB (BMI > 30 kg/m2 ). HM AAs were quantified via mass spectrometry. Infant HM intake, anthropometrics and body composition were assessed. Linear mixed-effects models (LMEM) examined the relationships between maternal BMI and HM AA intakes, and HM AA intake and infant growth over the first 6 months postpartum after adjusting for maternal and infant characteristics. RESULTS: Maternal BMI was positively associated with infant intakes of isoleucine, leucine, and AAAs across timepoints. HM AA intakes were positively associated with weight-for-length z-score, fat mass index, and fat-free mass index in infants (p < 0.05). CONCLUSIONS: Maternal BMI led to differences in HM AA composition, which was associated with infant body composition.
Asunto(s)
Isoleucina , Leche Humana , Índice de Masa Corporal , Lactancia Materna , Femenino , Humanos , Lactante , Isoleucina/análisis , Leucina/análisis , Leche Humana/metabolismo , Obesidad/epidemiología , Obesidad/metabolismoRESUMEN
The aim was to evaluate the amino acid compositions of three commonly consumed leafy vegetables (Solanum aethiopicum, Amaranthus hybridus, and Telfairia occidentalis) in Abakaliki, Ebonyi State. Leafy vegetables are important protective foods and beneficial for the maintenance of healthy living and prevention of diseases. The fresh leaves of A. hybridus, T. occidentalis, and S. aethiopicum were air-dried under room temperature for 1 week. The dried samples were further milled into a fine powder using a mechanical grinder and were stored in an air-tight plastic container. Amino acid content was determined using an applied Bio-system (phenylthiohydantoin, PTH) amino acid analyzer. Among amino acids determined in the vegetables, glutamic acid had the highest value with 12.59, 11.20, and 11.96 g/100 g protein, which was followed closely by leucine with 9.81, 7.94, 9.28 g/100 g protein, and aspartic acid with 8.99, 8.62, and 9.74 g/100 g protein in S. aethiopicum, A. hybridus, and T. occidentalis, respectively on dry weight bases. The leaf that contained the highest total amino acid (TAA) was S. aethiopicum with 88.69 g/100 g protein followed by T. occidentalis with 80.39 g/100 g protein while A. hybridus being the lowest, had 73.38 g/100 g protein. The limiting essential amino acid was tryptophan with 1.98 g/100 g protein while leucine with 9.0 g/100 g protein was the most abundant TAA. The percentage concentration of different groups of amino acid in vegetables revealed that total essential amino acid (TEAA) had 54.85%, total non-essential amino acid (TNEAA) had 48.27%, total neutral amino acid (TNAA) had 22.24%, total acidic amino acid (TAAA) had 32.48%, total basic amino acid (TBAA) had 11.53%, total aromatic amino acid (TArAA) had 11.89% while total sulfur amino acid (TSAA) had 3.94%. The results indicate that the vegetables studied are rich in essential amino acids and could serve as a good source of quality protein. Therefore, they could be recommended as food supplements, especially when animal proteins become more expensive as a source of protein.
Asunto(s)
Amaranthus , Cucurbitaceae , Solanum , Amaranthus/química , Aminoácidos/análisis , Animales , Leucina/análisis , Nigeria , Hojas de la Planta/química , Verduras/químicaRESUMEN
Lactoferrin (LF) belongs to the family of transferrins having multifunctional roles associated with the immune system of animals. To follow the aims for this study was selected 20 sequences of LF from mammalian species to evaluate the chemical, biological, and structural properties. Bioinformatics approaches used programs such as MAFFT for sequence alignment; PartitionFinder and MrBayes for phylogenetic approaches; I-TASSER, PROCHECK, Molecular Operating Environment (MOE), SWISS Model server, Peptide DB and Expasy ProtParam to estimate the physicochemical properties, to model the protein and predicted secondary structures. A phylogenic analysis shows species with genetic similarities clustered by complexity and unique grouping between Capra hircus, Macaca mulatta, and Myotis lucifugus, since they presented more amino acids but not overall changes in the iron-binding sites or biological aspects. Structural deviations in these clusters obtained in LF from those species were found in residues 46 (position 406-450), that is part of alpha-helix, and 37 (position 295-331), that is part of the beta-sheets. Our predicted model can be used to investigate more about structural aspects of LF and be applied for medicinal research.
Asunto(s)
Lactoferrina/química , Alanina/análisis , Secuencia de Aminoácidos , Animales , Bases de Datos de Proteínas , Lactoferrina/metabolismo , Leucina/análisis , Modelos Moleculares , Filogenia , Conformación Proteica en Hélice alfa , Estructura Terciaria de Proteína , Especificidad de la Especie , Relación Estructura-ActividadRESUMEN
Casein nonphosphopeptide (CNPP), a byproduct formed during the preparation of casein phosphopeptide (CPP), is often discarded on a large scale. Although our previous studies have demonstrated the ameliorative effect of CNPP on muscle wasting disorders, its structure-function mechanism is still unclear. Therefore, considering the great influence of structural characteristics on function, this study aims to explain the potential mechanism by characterizing the physicochemical and functional properties of CNPP. The results of structural characterization indicated that CNPP was of low molecular weight and composed of the complete range of amino acids; it was particularly rich in leucine. Compared with casein, CNPP had a lower molecular size and total/free sulfhydryl content (reduced 2.44 and 2.02 µmol/g in CNPP, respectively). Additionally, Fourier transform infrared spectroscopic analysis revealed that enzymatic hydrolysis caused protein unfolding, and the content of ß-turns and random coils reached 50.20% and 10.67%, respectively. Fluorescence-dependent detection of CNPP indicated a reduction of spectral intensity and the occurrence of a red shift. The changes in the structure of CNPP significantly affected its functional characteristics. CNPP has better solubility, foaming, and digestion properties than those of casein and whey protein. Specifically, the foam stability and emulsification properties decreased in the order of casein > CNPP > whey protein. The present study can provide a substantial basis for future application of CNPP as a functional ingredient against sarcopenia.
Asunto(s)
Caseínas/química , Fosfopéptidos/química , Aminoácidos/análisis , Fenómenos Químicos , Emulsionantes/química , Industria de Alimentos , Hidrólisis , Leucina/análisis , Peso Molecular , Estructura Secundaria de Proteína , Desplegamiento Proteico , Solubilidad , Residuos , Proteína de Suero de Leche/químicaRESUMEN
Liquid chromatography coupled with tandem mass spectrometry was employed for the detection of pesticides (thiamethoxam, lambda-cyhalothrin, deltamethrin, and metalaxyl) and their metabolites in Raphanus sativus var. longipinnatus exposed to these compounds under experimental conditions. Metalaxyl (0.008 mg/kg), metalaxyl acid (0.009 mg/kg), and (+)-trans-chrysanthemic acid (0.098 mg/kg) were identified in the plants exposed to the individual pesticides and their metabolites. Non-targeted analysis revealed the presence of thiamethoxam, lambda-cyhalothrin, and deltamethrin metabolites in plants exposed to these substances, despite the fact that the pesticide concentrations were below the analytical method's limit of quantification (0.005-0.006 mg/kg). Based on the non-targeted screening, non-specific (leucine and tyramine) and specific (epinephrine, dopamine, tryptamine, and serotonin) markers of plant exposure to the mentioned stress-inducing compounds were detected. These findings prove that non-targeted analysis is an indispensable tool for determining plants' exposure to pesticides, even when the parent compound has been completely metabolized.
Asunto(s)
Cromatografía Líquida de Alta Presión , Metaboloma , Plaguicidas/farmacología , Raphanus/metabolismo , Espectrometría de Masas en Tándem , Epinefrina/análisis , Epinefrina/aislamiento & purificación , Leucina/análisis , Leucina/aislamiento & purificación , Nitrilos/farmacología , Piretrinas/farmacología , Raphanus/efectos de los fármacos , Extracción en Fase Sólida , Tiametoxam/farmacologíaRESUMEN
Various process intensification methods were proposed to improve the yield, quality, and safety of fermented products. Here, we report the enhancement of L-leucine production by Corynebacterium glutamicum CP using ultrasound-assisted fed-batch fermentation. Response surface methodology was employed to optimize the sonication conditions. At an ultrasonic power density of 94 W/L, frequency of 25 kHz, interval of 31 min, and duration of 37 s, C. glutamicum CP produced 52.89 g/L of L-leucine in 44 h, representing a 21.6% increase compared with the control. The production performance of L-leucine was also improved under ultrasonic treatment. Moreover, the effects of ultrasound treatment on the fermentation performance of L-leucine were studied in terms of cell morphology, cell membrane permeability, and enzyme activity. The results indicate that ultrasonication is an efficient method for the intensification of L-leucine production by C. glutamicum CP.
Asunto(s)
Técnicas de Cultivo Celular por Lotes , Corynebacterium glutamicum/metabolismo , Leucina/biosíntesis , Ultrasonido , Biomasa , Permeabilidad de la Membrana Celular , Corynebacterium glutamicum/citología , Corynebacterium glutamicum/ultraestructura , Fermentación , Glucosa/análisis , Leucina/análisis , Factores de TiempoRESUMEN
Herein, the fabrication of a fascinating multifunctional cyclodextrin (CD) chiral stationary phase and its chiral separation performance in capillary electrochromatography are proposed. A facile interfacial polymerization was used to anchor ethanediamine-ß-cyclodextrin (EDA-ß-CD) polymerized with trimesoyl chloride (TMC) and to form the chiral stationary phase (CSP) composite onto the surface wall of the capillary. The characters of prepared columns were confirmed by Fourier transform infrared spectroscopy (FT-IR), X-ray Photoelectron Spectrometer (XPS), scanning electron microscopy (SEM) and energy dispersive X-ray spectrometry (EDS). This novel CSP offers multi-typical interactions including hydrogen bonding, π-interaction, hydrophobic and electrostatic interaction as well as steric effects which contribute to prominent chiral recognition for Dansyl-DL-amino acids in CEC modes. The EDA-ß-CD modified column showed eminent enantioseparation performance towards five Dansyl-DL-amino acids (the DL-forms of valine, threonine, leucine, phenylalanine, serine). Besides, the prepared columns were perfectly reproducible and stable. The relative standard deviations of the enantiomer retention times for intra-day (n = 5), inter-day (n = 3) runs and column-to-columns (n = 3) are below 0.54%, 1.35% and 4.89%, individually. This innovative chiral stationary phase shows a broader application view and scope in chiral recognition domain.
Asunto(s)
Aminoácidos/análisis , Electrocromatografía Capilar/métodos , Compuestos de Dansilo/química , beta-Ciclodextrinas/química , Aminoácidos/aislamiento & purificación , Compuestos de Dansilo/análisis , Compuestos de Dansilo/aislamiento & purificación , Etilenodiaminas/química , Leucina/análogos & derivados , Leucina/análisis , Leucina/aislamiento & purificación , Fenilalanina/análisis , Fenilalanina/aislamiento & purificación , Reproducibilidad de los Resultados , Espectroscopía Infrarroja por Transformada de Fourier , EstereoisomerismoRESUMEN
A pathogenic bacterium has its own mechanisms for not only pathogenic attack but also exogenous invasion defense, in which the bacterial cell wall is the front line of attack and defense. We developed a biochemical lanthanide-encoding approach to quantify the uncanonical d-amino acid (d-X) that was edited in a small proportion into the terminal acyl-d-Ala-d-X of nascent peptidoglycan UDP-MurNAc-pentapeptides in the bacterial cell wall. This approach overcomes the difficulties regarding quantification and accuracy issues encountered by the popular optical imaging and traditional high-performance liquid chromatography-based methods. Newly synthesized azide-d-Leu and ketone-d-Met were used together with alkynyl-d-Ala for their metabolic assembly and then bioorthogonally encoded by the correspondingly fabricated DBCO-DOTA-Gd, H2NO-DOTA-Eu, and azide-DOTA-Sm tags. This approach allows direct quantification of the d-X in situ in the cell wall using 158Gd, 153Eu, and 154Sm species-unspecific isotope dilution inductively coupled plasma mass spectrometry, avoiding any tedious and complex "cell-broken" pretreatment procedures that might induce racemization of the d-X. The obtained site-specific and accurate in situ information about the d-X enables quantitative monitoring of the bacterial response when Staphylococcus aureus meets vancomycin, showing that the amounts of azide-d-Leu and ketone-d-Met assembled are more important after determining the structure- and composition-dependent bacterial antibiotic resistance mechanisms. In addition, we found that the combined use of vancomycin and d-Ala restores the efficacy of vancomycin and might be a wise and simple way to combat vancomycin intermediate-resistant S. aureus.
Asunto(s)
Antibacterianos/farmacología , Marcaje Isotópico/métodos , Elementos de la Serie de los Lantanoides/química , Staphylococcus aureus/efectos de los fármacos , Staphylococcus aureus/metabolismo , Vancomicina/farmacología , Alanina/análogos & derivados , Alanina/análisis , Alanina/farmacología , Europio/química , Gadolinio/química , Leucina/análogos & derivados , Leucina/análisis , Metionina/análogos & derivados , Metionina/análisis , Viabilidad Microbiana/efectos de los fármacos , Peptidoglicano/química , Peptidoglicano/metabolismo , Samario/química , Estereoisomerismo , Uridina Difosfato Ácido N-Acetilmurámico/análogos & derivados , Uridina Difosfato Ácido N-Acetilmurámico/química , Uridina Difosfato Ácido N-Acetilmurámico/metabolismoRESUMEN
The molecular recognition ability of tryptophan (Trp) for isomeric amino acids, such as leucine (Leu) and isoleucine (Ile), and isomeric amino acid-containing dipeptides, such as Leu-Gly, Ile-Gly, Gly-Leu, and Gly-Ile (where Gly denotes glycine), was investigated using a tandem mass spectrometer equipped with an electrospray ionization source and cold ion trap. The ultraviolet photodissociation spectra of the cold gas-phase clusters of Leu and Ile with Na+Trp in the wavelength range of 265-290 nm revealed that the relative intensities of Leu and Ile were only different in the wavelength range of 265-273 nm; however, no differences in the relative intensities were observed when the wavelength exceeded 274 nm. The molecular recognition ability of photoexcited Trp was used for the identification and quantification of Leu and Ile in dipeptides in solution. The mole fractions of Leu and Ile in dipeptides could be determined from the abundances observed in a single product ion spectrum of the cold gas-phase clusters of dipeptides with Na+Trp.
Asunto(s)
Dipéptidos/análisis , Dipéptidos/química , Isoleucina/análisis , Isoleucina/química , Leucina/análisis , Leucina/química , Triptófano/química , Glicina/química , Isomerismo , Procesos Fotoquímicos , Espectrometría de Masa por Ionización de Electrospray/métodos , Espectrofotometría Ultravioleta/métodos , Espectrometría de Masas en Tándem/métodosRESUMEN
Herein we propose, for the first time, a rapid method based on flow injection analysis, electrospray ionization-tandem mass spectrometry (FIA-ESI-MS/MS) and multivariate calibration for the determination of l-leucine, l-isoleucine and L-allo-isoleucine in saliva. As far as we know, multivariate calibration has never been applied to the data from this non-separative approach. The possibilities of its use were explored and the results obtained were compared with the corresponding ones when using univariate calibration. Partial least square regression (PLS1) multivariate calibration models were built for each analyte by analyzing different saliva samples, and were subsequently applied to the analysis of another set of samples which had not been used in any calibration step. For Leu, the model worked satisfactorily with root mean square errors in the prediction step of 17%. This error can be considered acceptable and is common in methodologies that do not include a separation step. Results were compared with those obtained when univariate calibration was used, using the m/z transition 132.1 â 43.0 as the quantitation variable. In this case, the obtained results were not acceptable, with RMSEP of 236%, due to the fact that saliva samples contained another compound, different to the target analytes, which also shared the same transition. Ile and aIle have the same fragmentation patterns, so quantification of the sum of both compounds was performed, with RMSEP of 14% using a PLS1 model. Similar results were obtained when a univariate calibration model using the m/z transition 132.1 â 69.0 was employed. However, the use of this transition should be carefully examined when other compounds present in the matrix contribute to the analytical signal. The method increases sample throughput more than one order of magnitude compared to the corresponding LC-ESI-MS/MS method and is especially suitable as screening. When abnormally high or low concentrations of the analytes studied are obtained, the use of the method that includes separation is recommended to confirm the results.
Asunto(s)
Isoleucina/análisis , Leucina/análisis , Saliva/química , Calibración , Femenino , Voluntarios Sanos , Humanos , Análisis de los Mínimos Cuadrados , Masculino , Conformación Molecular , Análisis Multivariante , Espectrometría de Masa por Ionización de ElectrosprayRESUMEN
Aygün F, Kiykim E, Aktuglu-Zeybek Ç, Zubarioglu T, Cam H. Treatment of maple syrup urine disease with high flow hemodialysis in a neonate. Turk J Pediatr 2019; 61: 107-110. Continious renal replacement therapy (CRRT) is a well recognizied treatment of choice in acute renal failure, however CRRT became a preferred treatment of metabolic emergencies with high leucine and ammonia levels like Maple syrup urine disease (MSUD). MSUD is a rare metabolic disorder caused by deficiency in the activity of the branched-chain a-ketoacid dehydrogenase complex. The toxic accumulation of branched chain amino acids during acute metabolic decompensation is associated with the appearance of permanent neurological symptoms. Four patients were admitted to our pediatric intensive care department with complains of poor feeding, vomitting, irratibility and coma. Physical examination of the neonates were similar having stupor, hypotonia and depressed newborn reflexes. The leucine levels were between 930-4400 µmol/L. The diagnosis of MSUD was confirmed in all four. They were treated successfully with high flow CRRT having the rates were between 4120 ml/h/1.73m2 and 9830 ml/h/1.73m2. Early treatment is essential to prevent neurotoxicity and death. CRRT is a choice of treatment in metabolic crisis of MSUD. Herein, we report the successful treatment of acute metabolic decompensation of MSUD with CRRT in 4 neonates.
Asunto(s)
Terapia de Reemplazo Renal Continuo , Enfermedad de la Orina de Jarabe de Arce/terapia , Femenino , Humanos , Recién Nacido , Leucina/análisis , Masculino , Enfermedad de la Orina de Jarabe de Arce/diagnósticoRESUMEN
Three new linear peptides containing d-leucine, named whitmantides A-C (1-3), were isolated from the dried whole bodies of Whitmania pigra Whitman. Their structures with absolute configurations were elucidated by Edman degradation, mass spectrometry, Marfey's analysis, and solid-phase synthesis. It is the first time that peptides containing d-amino acid in leeches were discovered. Compounds 1-3 displayed neuroprotective activities against oxygen-glucose deprivation/reperfusion injury on Neuro-2a cells. In addition, ex vivo serum stability tests showed that 1-3 were resistant to protease degradation.
Asunto(s)
Sanguijuelas/química , Leucina/análisis , Fármacos Neuroprotectores/farmacología , Péptidos/química , Animales , Fármacos Neuroprotectores/químicaRESUMEN
This study developed a novel form of sufu. Salted egg white sufu (SEWS) was produced by fermenting steamed salted egg white with douchi koji and ripening at 25°C, 35°C, or 45°C for 19 days. The results show that the protease activity of the koji reduced pronouncedly during fermentation, whereas the one in the SEWS initially increased but subsequently decreased. The total nitrogen and amino nitrogen contents and degree of protein hydrolysis increased during fermentation and decreased during preripening. SEWS, particularly processed at 35°C, contained more free amino acids, notably glutamic acid and leucine, than steamed salted egg white did. After processing, SEWS had darker colors, particularly when manufactured at higher temperatures, and hardness and springiness decreased. The 35°C and 45°C SEWS had higher sensory acceptability. In conclusion, ripening at 35°C for 19 days is recommended for producing this novel animal protein-based sufu.