Implementing stepwise solvent elution in sequential injection chromatography for fluorimetric determination of intracellular free amino acids in the microalgae Tetraselmis gracilis

The concept of sequential injection chromatography (SIC) was exploited to automate the fluorimetric determination of amino acids after pre-column derivatization with ophthaldialdehyde (OPA) in presence of 2-mercaptoethanol (2MCE) using a reverse phase monolithic C(18) stationary phase. The method is low-priced and based on five steps of isocratic elutions. The first step employs the mixture methanol: tetrahydrofuran: 10 mmol L(-1) phosphate buffer (pH 7.2) at the volumetric ratio of 8:1:91; the other steps use methanol: 10 mmol L-1 phosphate buffer (pH 7.2) at volumetric ratios of 20:80, 35:65, SO:SO and 65:35. At a flow rate of 10 mu L s(-1) a 25 mm long-column was able to separate aspartic acid (Asp), glutamic acid (Glu), asparagine (Asn), serine (Ser), glutamine (Gln), glycine (Gly), threonine (Thr), citruline (Ctr), arginine (Arg), alanine (Ala), tyrosine (Tyr), phenylalanine (Phe), ornithine (Orn) and lysine (Lys) with resolution >1.2 as well as methionine (Met) and valine (Val) with resolution of 0.6. Under these conditions isoleucine (Ile) and leucine (Leu) co-eluted. The entire cycle of amino acids derivatization, chromatographic separation and column conditioning at the end of separation lasted 25 min. At a flow rate of 40 mu L s(-1) such time was reduced to 10 min at the cost of resolution worsening for the pairs Ctr/Arg and Orn/Lys. The detection limits varied from 0.092 mu mol L(-1) for Tyr to 0.51 mu mol L(-1) for Orn. The method was successfully applied to the determination of intracellular free amino acids in the green alga Tetraselmis gracilis during a period of seven days of cultivation. Samples spiked with known amounts of amino acids resulted in recoveries between 94 and 112%. (C) 2008 Elsevier B.V. All rights reserved.

Rapid method for the determination. of organic acids in wine by capillary electrophoresis with indirect UV detection

A capillary electrophoresis method for organic acids in wine was developed and validated. The optimal electrolyte consisted of 10 mmol/L 3,5-dinitrobenzoic acid (DNB) at pH 3.6 containing 0.2 mmol/L cetyltrimethylammonium bromide as flow reverser. DNB was chosen because it has an effective mobility similar to the organic acids under investigation, good buffering capacity at pH 3.6, and good chromophoric characteristics for indirect UV-absorbance detection at 254 nm. Sample preparation involved dilution and filtration. The method showed good performance characteristics: Linearity at 6 to 285 mg/L (r > 0.99); detection and quantification limits of 0.64 to 1.55 and 2.12 to 5.15 mg/L, respectively; separation time of less than 5.5 min. Coefficients of variation for ten injections were less than 5% and recoveries varied from 95% to 102%. Application to 23 samples of Brazilian wine confirmed good repeatability and demonstrated wide variation in the organic acid concentrations. (C) 2008 Elsevier Ltd. All rights reserved.

Amino acids profile of sugar cane spirit (cachaca), rum, and whisky

An analytical procedure for the separation and quantification of 20 amino acids in cachacas has been developed involving C18 solid phase cleanup, derivatization with o-phthalaldehyde/2-mercaptoethanol, and reverse phase liquid chromatography with fluorescence detection. The detection limit was between 0.0050 (Cys) and 0.25 (Ser) mg L-1, whereas the recovery index varies from 69.5 (Lys) to 100 (Tyr)%. Relative standard deviations vary from 1.39 (Trp) to 13.4 (Glu)% and from 3.08 (Glu) to 13.5 (His) for the repeatability and intermediate precision, respectively. From the quantitative profile of amino acids in 41 cachacas, 5 turns, and 12 whisky samples, the following order of amino acids in significant quantities is observed: Gly = Ser < Cys < Ile < His < Pro = Asp < Asn < Tyr for cachaca; Phe < Glu = Gln = Val = Ala < His = Gly Thr = Arg = Tyr < Asn Ser = Lys = Pro < Cys = Asp for rum; and Ala = Asn < Trp < Gln = His = Met = Ile = Cys < Thr < Asp Leu < Phe = Lys < Ser = Gly = Tyr = Val < Glu = Pro < Arg for whisky samples. (C) 2007 Elsevier Ltd. All rights reserved.

Determination of amino acids by capillary electrophoresis-electrospray ionization-mass spectrometry: An evaluation of different protein hydrolysis procedures

In this work, a CE equipment, online hyphenated to an IT MS analyzer by a linear sheath liquid interface promoting ESI, was used to develop a method for quantitative determination of amino acids. Under appropriate conditions (BGE composition, 0.8% HCOOH, 20% CH(3)OH; sheath liquid composition, 0.8% HCOOH, 60% methanol; V(ESI), +4.50 W), analytical curves of all amino acids from 3 to 80 mg/L were recorded presenting acceptable linearity (r > 0.99). LODs in the range of 16-172 mu mol/L were obtained. BSA, a model protein, was submitted to different hydrolysis procedures (classical acid and basic, and catalyzed by the H(+) form of a cation exchanger resin) and its amino acid profiles determined. In general, the resin-mediated hydrolysis yields were overall similar or better than those obtained by classical acid or basic hydrolysis. The resulting experimental-to-theoretical BSA concentration ratios served as correction factors for the quantitation of amino acids in Brazil nut resin generated hydrolysates.

Reactivity of Beer Bitter Acids Toward the 1-Hydroxyethyl Radical as Probed by Spin-Trapping Electron Paramagnetic Resonance (EPR) and Electrospray Ionization-Tandem Mass Spectrometry (ESI-MS/MS)

The iso-alpha-acids or isohumulones are the major contributors to the bitter taste of beer, and it is well-recognized that they are degraded during beer aging. In particular, the trans-isohumulones seem to be less stable than the cis-isohumulones. The major radical identified in beer is the 1-hydroxyethyl radical; however, the reactivity between this radical and the isohumulones has not been reported until now. Therefore, we studied the reactivity of isohumulones toward the 1-hydroxyethyl radical through a competitive kinetic approach. It was observed that both cis- and trans-isohumulones and dihydroisohumulones are decomposed in the presence of 1-hydroxyethyl radicals, while the reactivities are comparable. On the other hand, the tetrahydroisohumulones did not react with 1-hydroxyethyl radicals. The apparent second-order rate constants for the reactions between the 1-hydroxyethyl radical and these compounds were determined by electron paramagnetic resonance (EPR) spectroscopy and electrospray ionization-tandem mass spectrometry [ESI(+)-MS/MS]. It follows that degradation of beer bitter acids is highly influenced by the presence of 1-hydroxyethyl radicals. The reaction products were detected by liquid chromatography electrospray ionization-ion trap-tandem mass spectrometry (LC-ESI-IT-MS/MS), and the formation of oxidized derivatives of the isohumulones was confirmed. These data help to understand the mechanism of beer degradation upon aging.

An insight into the beginning stage of the aqueous ring opening metathesis polymerization of exo,exo-7-oxabicyclo[2.2.1]hept-5-ene-2,3-dicarboxylic acid with Ru(III) compounds: the consumption of Ru(III) species in the presence of carboxylic acids monitored by ESR spectroscopy

The presence of paramagnetic species in the aqueous ring opening metathesis polymerizations of the exo,exo-7-oxabicyclo[2.2.1]hept-5-ene-2,3-dicarboxylic acid monomer with RuCl(3) and K(2)[RuCl(5)H(2)O] compounds was studied using ESR techniques. It was observed that the intensities of the Ru(III) signals in the ESR spectra decrease on the time scale of the induction period so that the ROMP can take place. The intensity of the Ru(III) signal almost disappeared 50 min after reacting with K(2)[RuCl(5)H(2)O] and after 100 mm in the case of RuCl(3). Reactions of the cis-[Ru(NH(3))(4)(H(2)O)(2)](tfms)(3) and [Ru(NH(3))(5)H(2)O](tfms)(3) complexes with the monomer and different organic compounds representing the organic functions in the monomer (furan, norbornene, but-2-ene-1,4-diol and formic, acetic, oxalic and maleic acids) were also monitored by ESR and UV/vis spectra. It was deduced that the organic acids provide the disappearance of the Ru(III) signal. The proton NMR relaxation times of the residual water in D(2)O for reactions with oxalic acid suggested that the presence of paramagnetic ions in the solution decreases along with

The origin of the molecular interaction between amino acids and gold nanoparticles: A theoretical and experimental investigation

This study describes the synthesis of novel biological hybrid materials, where 3D structures were obtained using gold nanoparticles (AuNps) and methionine (Met) in a one-step procedure in aqueous media. The type of nanostructure can be controlled by tuning the intermolecular interactions between Met and AuNp, which strongly depends on the pH used for the synthesis. Computational simulation using the density-functional theory (DFT) showed that the AuNp - Met 3D structures are formed upon reorientation of Met molecules so that the backbone amine groups interact via H-bonds. These findings were experimentally confirmed using FTIR and UV-vis spectroscopy. Crown Copyright (C) 2008 Published by Elsevier B. V. All rights reserved.

Amino acids and endurance exercise

Provides information on a study which examined the role of amino acids during endurance exercise and the implications for sports nutrition and performance. Description of amino acid utilization during exercise; Function of glutamine; Cardiovascular function of L-arginine.

α,ω-Bis(trichlorostannyl)alkanes as Bis(monodentate) lewis acids toward halide ions

The reactions between X₃Sn(CH₂)_nSnX₃ (X = Cl, Br; _n = 1, 3, 4, 8) and halide in solution are described. ¹¹⁹Sn NMR spectroscopy (CD₂Cl₂) indicates formation of five-coordinate dinuclear 1:2 adducts, [X₄Sn(CH₂)_nSnX₄]^2-, with chloride and bromide. A trigonal-bipyramidal geometry has been confirmed in the solid state for the compound having X = Cl and _n = 4. In contrast, fluoride reacts with X₃Sn(CH₂)_nSnX₃ to give six-coordinate tin species, [F₅Sn(CH₂)_nSnF₅].^4- There was no evidence indicating that the tin atoms could act as a bidentate acid.

Selective determination of amino acids using flow injection analysis coupled with chemiluminescence detection

The determination of the amino acids proline, histidine, tyrosine, arginine, phenylalanine and tryptophan using flow injection analysis (FIA) with chemiluminescence detection is described. Proline was the only amino acid to exhibit chemiluminescence with the tris(2,2-bipyridyl)ruthenium(III) reaction at pH 10. While, histidine was found to selectively enhance the reaction of luminol with Mn(II) salts in a basic medium. Acidic potassium permanganate chemiluminescence was able to selectively determine tyrosine at pH 6.75. Low pressure separations using a C18 guard column allowed the simultaneous determination of tyrosine and tryptophan or phenylalanine and tryptophan with acidic potassium permanganate and copper(II)–amino acid–hydrogen peroxide chemiluminescence, respectively. Precision for each method was less than 3.9% (R.S.D.) for five replicates of a standard (1×10−5 M) and the detection limits ranged between 4×10−9 and 7×10−6 M. Preliminary investigations revealed that the methodology developed was able to selectively determine the individual amino acids in an equimolar mixture of the 20 naturally occurring amino acids.

Strategies and methods for the attachment of amino acids and peptides to chiral [n]polynorbornane templates

A versatile synthesis of amino acid and peptide functionalised [n]polynorbornane scaffolds is described. The frameworks are constructed using the stereoselective and regioselective cycloaddition of suitably functionalised chiral cyclobutene epoxides with similar norbornenes. The strategies employed allow a range of topologies to be accessed and a number of regioselectively addressable linkage points to be accommodated.

The relative absorption of fatty acids in brown trout (Salmo trutta) fed a commercial extruded pellet coated with different lipid sources

The objective of the present study was to investigate the fatty acid absorption capabilities of brown trout (Salmo trutta) fed commercial extruded diets. Five commercial extruded pellets, different only in the lipid sources used for fat coating, were tested on juvenile brown trout for 45 days. The trout were reared in fresh water at 14.6 ± 0.4° C and 7.7 ±
0.3 mg/l, temperature and dissolved oxygen, respectively. The tested lipid sources were fish oil, canola oil, oleine oil, swine fat and poultry fat. After the adaptation period faeces were collected by gently stripping from naesthetized fish. Fatty acid analysis was performed on experimental diets and on collected faeces to evaluate the relative absorption capabilities of the trout digestive system with respect to each detected fatty acid. The use of the relative absorption efficiency (rAE) was opted to evaluate the intrinsic capability of each fatty acid to be absorbed. Brown trout showed a
specific preferential order of absorption of the fatty acids, preferring shorter over longer chain fatty acids and preferring the more unsaturated to the more saturated fatty acids. The fatty acid that showed the best relative absorbability was the C18:4n-3 (rAE = 5.14 ± 0.72), which has a fairly short carbon chain, but at the same time a high unsaturation level, followed by the C18:3n-3 (rAE = 3.38 ± 0.30). The fatty acid that showed the worst relative absorbability (rAE = 0.21 ± 0.02) was C24:1n-9.

Validation of a FFQ to estimate the intake of PUFA using plasma phospholipid fatty acids and weighed foods records

Due to the growing knowledge about the role of specific fatty acids in health and disease, dietary intake measurements of individual fatty acids or classes of fatty acids are becoming increasingly important. The objective of this study was to evaluate the ability of the Nambour FFQ to estimate intakes of specific fatty acids, particularly PUFA. The study population was a sub-sample of adult participants in a randomised controlled trial of [beta]-carotene and sunscreen in the prevention of skin cancer (n 43). Dietary intake was assessed by a self-administered FFQ and a weighed food record (WFR). Non-fasting blood samples were collected and analysed for plasma phospholipid fatty acids. Median intakes on the FFQ were generally higher than the WFR except for the n-3 PUFA groups, where the FFQ estimated higher intakes. Correlations between the FFQ and WFR were moderate (r 0–32-0-59) except for trans fatty acids (r 0–03). Correlations between each of the dietary assessment methods and the plasma phospholipids were poor for all fatty acids other than the PUFA. Using the methods of triads approach, the FFQ validity coefficients for total n-3 fatty acids, total long chain n-3 fatty acids, EPA, arachidonic acid, docosapentaenoic acid and DHA were 0–50, 0–63, 0–45 and 0–62 and 0–62, respectively. For most fatty acids, the FFQ adequately estimates group mean fatty acid intakes and can adequately rank individuals; however, the ability of this FFQ to estimate trans fatty acids was poor.

Retention of essential amino acids during extrusion of protein and reducing sugars

This research investigates the retention of essential amino acid profiles of products during the extrusion of proteins and reducing sugars. Animal proteins (egg and milk protein at 10 and 30% levels) and reducing sugars (fructose and galactose at 0, 2, and 8% levels), with pregelatinized wheat flour, were extruded at 110 and 125 °C product temperatures and feed moistures of 19 and 23.5% for egg protein and 13.75 and 16% for milk protein. The nutritional property analyzed was essential amino acid retention, and sugar retention was also considered to understand the relationship of sugars with retention of amino acids. Lysine showed the lowest retention (up to 40%) of all the essential amino acids. Retention of other essential amino acids varied from 80 to 100% in most situations. Apart from lysine,  tryptophan, threonine, and methionine were found to be significantly changed (P < 0.05) with processing conditions. Increased protein and sugar levels resulted in a significant degradation of lysine. Greater lysine retention was found at a lower temperature and higher feed moisture. Results of sugar retention also showed similar patterns. The products made from fructose had greater lysine retention than products made from galactose with any type of protein. The outcomes of this research suggested that the combination of milk protein and fructose at a lower temperature and higher feed moisture is most favorable for developing high-protein extruded products.