The Influence of Measurement Parameters on the Specific Rotation of Amino Acids

The effects of solute and hydrochloric acid concentrations on optical rotation were studied using 20 naturally occuring amino acids.

There appeared to be no common factor among the amino acids as far as the inclination of optical rotation was concerned. Lutz-Jirgenson’s rule could be applied to few amino acids in the cationic form. Therefore, in the determination of the optical rotation, the concentration of the solute, nature of solvent and temperature must be rigorously controlled. The optical conditions of measurement and the specific rotation of 20 amino acids were recommended based on this work.     

苯环取代苯丙氨酸的合成及拆分

以苯胺类化合物为起始原料 ,经重氮化、丙烯酸加成、氨解等反应合成了 7种消旋苯环取代苯丙氨酸类氨基酸 ,并采用α 糜蛋白酶或枯草杆菌蛋白酶对所有氨基酸进行了拆分 ,其结构分别通过红外光谱、核磁共振、元素分析、熔点、旋光等分析得到确证。该法可以较大规模合成此类氨基酸。     

Effects of Citrus aurantifolia concentrated extract on the spontaneous proliferation of MDA-MB-453 and RPMI-8866 tumor cell lines.

The in vitro effects of concentrated lime juice (CLJ) extract on the spontaneous proliferation of a human breast carcinoma cell line (MDA-MB-453) and a human lymphoblastoid B cell line (RPMI-8866) were investigated. CLJ extract was prepared by freeze-drying fresh fruit juice and dialyzing the concentrated extract against phosphate buffered saline in order to deplete low molecular weight micronutrients such as flavonoids as well as adjusting the pH of the extract to the physiological range. The effects of different concentrations of the CLJ extract on the spontaneous proliferative responses of the cell lines were determined by 3H-thymidine incorporation after 24 hrs of incubation. CLJ extract had no significant effect on MDA-MB-453 cell line, however, using the concentrations of 125, 250, and 500 microg/ml of CLJ extract a significant inhibition of the spontaneous proliferation of RPMI-8866 cell line was detected (P < 0.05). Due to the protein nature of the biologically active macromolecules of the CLJ extract (Gharagozloo and Ghaderi, 2001), it is reasonable to assume that the protein components of the CLJ extract may have anti-proliferative effects on tumor cell lines.     

The effect of cortisol on the synthesis of rat plasma albumin, fibrinogen and transferrin

A decrease of absolute synthesis of albumin, no change in that of fibrinogen and an increased fractional synthesis of transferrin were observed 3h after intraperitoneal administration of a pharmacological dose of 5 mg of cortisol to 220g rats in the post-absorptive state and previously kept on a diet with 40% protein. The concentration in liver of total free amino acids was practically unchanged at this time. Intraperitoneal administration of a mixture of amino acids with the cortisol raised this concentration and was accompanied by an almost complete de-repression of the synthesis of albumin, with no real effect on that of fibrinogen. In considerable contrast, in rats studied at 24h after intraperitoneal administration of cortisol, and who had been fed once in the interim (but who had received no amino acids intraperitoneally), there was a marked increase in the absolute synthesis of albumin and fibrinogen, with an increase in fractional synthesis that was less proportionately but still very significant and which included transferrin. The amino acid concentrations had risen above the supplemented values at 3h but not as much proportionately as the fractional synthesis rates, and of course not as much as the absolute synthesis rates, of albumin and fibrinogen. These time-dependent effects of cortisol suggest to us that our studies resolve the apparently conflicting results of the effect of cortisol on the synthesis of albumin reported by others.     

Breeding experiments and genome‐wide association analysis elucidate two genetically different forms of non‐syndromic congenital cleft lip and jaw in Vorderwald × Montbéliarde cattle

Non‐syndromic congenital cleft lip and jaw (CLJ) is a condition reported in Vorderwald × Montbéliarde cattle. The objective of the present study was to perform a genome‐wide association study (GWAS) for 10 CLJ‐affected and 50 unaffected Vorderwald × Montbéliarde cattle using the bovine Illumina high density bead chip to identify loci for this condition. Phenotypic classification of CLJ was based on a detailed recording of orofacial structures using computed tomography. A breeding experiment among CLJ‐affected Vorderwald × Montbéliarde cattle and CLJ‐affected Vorderwald × Montbéliarde cattle with unaffected Holsteins confirmed recessive inheritance and different loci for bilateral or left‐sided versus right‐sided CLJ. The GWAS for the five cases with right‐sided CLJ gave a genome‐wide signal on bovine chromosome (BTA) 29 at 16 Mb. For the four left‐sided and one bilateral CLJ case, a genome‐wide significant association was identified on BTA4 at 32 Mb. Two different loci are very likely to be involved in CLJ in Vorderwald × Montbéliarde cattle because experimental matings among affected cows and bulls with different types of CLJ did not result in CLJ‐affected progeny, and in addition, two different loci were also found through GWAS and mapped on two different bovine chromosomes. Validation in 346 Vorderwald × Montbéliarde cattle for the highly associated SNPs on BTA4 and 29 gave ratios of 33/346 (0.095, BTA4) and 6/346 (0.017, BTA29) homozygous mutant genotypes. Further studies should elucidate the responsible mutations underlying the different types of CLJ in Vorderwald × Montbéliarde cattle.     

Ternary complexes of palladium (II) with levamisole and L-amino acids

The complexes of general formula [(LMS)2Pd(amino acid)]Cl with LMS = levamisole, and amino acid = L-alanine, L-phenylglycine, L-phenylalanine, L-valine, L-methionine, and L-proline, were synthesized by the interaction of [(LMS)2PdCl2] with the sodium salts of L-amino acids. The newly synthesized complexes are characterized by elemental analysis, conductivity, magnetic susceptibility, optical rotation measurements, and UV-Vis, IR and 13C NMR spectral data. Levamisole is coordinated to palladium via the N-7 nitrogen and the amino acids through the amino nitrogen and carboxylate oxygen, except for L-methionine which binds the metal via nitrogen and sulfur atoms. Optically active [(LMS)2Pd(amino acid)]Cl complexes are obtained when L-amino acids or D,L-amino acids are used for the synthesis of these complexes. L-Methionine and L-proline complexes induce new cell forms in Baker's yeast (Saccharomyces cerevisiae) cells.     

Common vetch varietal differences in hay nutritive value,ruminal fermentation,nutrient digestibility and performance of fattening lambs

In some lower rainfall regions of the world (300–750 mm), common vetch (Vicia sativa L., hereafter referred to as vetch) has been shown to have yields competitive with alternate crops and provide high-quality hay for ruminant diets, but there are few studies of vetch performance as a livestock feed, or of vetch varietal differences in livestock feeding value. This study evaluated vetch varietal differences in hay nutritive value, ruminal fermentation properties, nutrient digestibility, nitrogen retention and animal performance in fattening lambs consuming a diet comprising 20% vetch. Fifty male Hu lambs with an initial BW of 17.7 ± 0.27 kg and 2–3 months of age were assigned randomly into five groups of 10 lambs, and each allocated one of five dietary treatments for 67 days (10 days of adaptation and a 57 day experimental period). All diets contained 30% maize stover, 50% concentrate, and with a different forage source (on an as-fed basis): 20% alfalfa hay (Control), 20% vetch 333A (C333A) hay, 20% vetch Lanjian No. 1 (CLJ1) hay, 20% vetch Lanjian No. 2 (CLJ2) hay, or 20% vetch Lanjian No. 3 (CLJ3) hay. Hay CLJ3 had greater contents of ash, CP, ether extract, in vitro organic matter digestibility and metabolizable energy, and lower cell wall contents (P < 0.05) than those of C333A and CLJ1 hays, but similar to the CLJ2 hay (P > 0.05). Compared to the Control diet, the CLJ2 and CLJ3 diets resulted in greater (P < 0.05) final BW, average daily gain, total tract apparent digestibility of CP and NDF, and nitrogen balance, both when expressed as g/day and relative to nitrogen intake, while animal performance when fed diets with C333A or CLJ1 diet did not differ (P > 0.05) from the Control diet. No differences (P > 0.05) were observed between dietary treatments with respect to average daily feed intake, ruminal pH, total volatile fatty acid contents and molar proportions of acetate, butyrate, valerate, isobutyrate, and isovalerate, or total tract apparent digestibility of DM, organic matter, and ADF. The feed efficiency of tested feeds ranked CLJ3 = CLJ2 > CLJ1 > Control with C333A intermediate between CLJ1 and Control. In summary, considering hay quality, nutrient digestibility and animal weight gain, performance as a ruminant feed of hays from recently released vetch cultivars Lanjian No. 2 and Lanjian No. 3 was superior to the older cultivar C333A and the alfalfa control.     

Growth, D-glucose utilization and malolactic fermentation by Leuconostoc œnos strains in 18 media deficient in one amino acid

Six Leuconostoc œnos strains were used to study the effect of the deficiency of one amino acid on growth, heterofermentative pathway and malolactic fermentation. All strains had an absolute requirement for four amino acids (isoleucine, glutamic acid, tryptophan and arginine) and needed six other amino acids (valine, methionine, cysteine, leucine, aspartic acid and histidine) for optimum growth. Each deficiency in one amino acid had a particular effect on D-glucose utilization. Overproduction and underproduction of D-lactic acid were observed. The rate of L-malic acid consumption in media deficient in one amino acid was lower than in the complete medium with all amino acids.
Although some deficiencies (glycine, phenylalanine, proline or tyrosine) had no influence on the growth, they noticeably limited the malolactic fermentation.     

Direct specific rotation measurements of amino acids,dipeptides, and tripeptides by laser‐based polarimetry

This paper addresses direct HPLC detection of amino acids, di and tripeptides via laser‐based polarimetry. Mass detection limits typically range from 0.5 to 50 μg injected on‐column, corresponding to concentration levels from 0.1 to 1 mg/ml. These levels are 1 to 2 orders of magnitude lower than measurements with conventional polarimetry, implying that concentration‐dependent phenomena (e.g., solute dimerization) are minimized. This is the first published report of the specific rotation of the di and tripeptides of Ala and Phe. The relationship between pH and specific rotation of selected amino acids, peptides, and lysozyme is described. Chirality 11:187–194, 1999. © 1999 Wiley‐Liss, Inc.     

Synthesis, determination of the absolute configuration of tonkinelin, and inhibitory action with bovine heart mitochondrial complex I

The first synthesis of two possible diastereomers of tonkinelin was achieved. By comparison of the optical rotation of two candidates of tonkinelin and the natural compound, it is suggested that the absolute configuration of natural tonkinelin is likely to be (17S,18S). The inhibitory activity of these compounds was examined with bovine heart mitochondrial NADH-ubiquinone oxidoreductase. These compounds showed remarkably weak inhibitory activity compared to ordinary acetogenins such as bullatacin.     

Synthesis of biodegradative threonine dehydratase in Escherichia coli: role of amino acids, electron acceptors, and certain intermediary metabolites

The specific activity of inducible biodegradative threonine dehydratase (EC 4.2.1.16) in Escherichia coli K-12 increased significantly when the standard tryptone-yeast extract medium or a synthetic mixture of 18 L-amino acids was supplemented with 10 mM KNO3 or 50 mM fumarate and with 4 mM cyclic AMP. In absolute terms, almost four times as much enzyme was produced in the amino acid medium as in the tryptone-yeast extract medium. Enzyme induction in the amino acid medium was sensitive to catabolite repression by glucose, gluconate, glycerol, and pyruvate. An analysis of amino acid requirements for enzyme induction showed that a combination of only four amino acids, threonine, serine, valine, and isoleucine, produced high levels of threonine dehydratase provided that both fumarate and cyclic AMP were present. Immunochemical data revealed that the enzyme synthesized in the presence of these four amino acids was indistinguishable from that produced in the tryptone-yeast extract or the medium with 18 amino acids. We interpret these results to mean that not the amino acids themselves but some metabolites derived anaerobically in reactions involving an electron acceptor may function as putative regulatory molecule(s) in the anaerobic induction of this enzyme.     

Stereoselective synthesis of chaulmoogric acid and related fatty acid from 2-(±)-cyclopentenecarboxylic acid by Bacillus subtilis (ATCC 7059)

2-(±)-Cyclopentenecarboxylic acid added to the culture medium is incorporated into two new fatty acids by the growing cells of Bacillus subtilis (ATCC 7059). The new fatty acids, amounting to 24% of the total cellular fatty acids, are identified as hydrocarpic [11-(2′-cyclopentenyl)-hendecanoic] and chaulmoogric [13-(2′-cyclopentenyl)-tridecanoic] by gas-liquid chromatography and mass spectrometry. These C₁₆ and C₁₈ fatty acids are optically active, levorotatory, with the specific rotation of ?50.4° as mixture, thus the optical purity of approximately 80%. This indicates that the optical rotation of these bacterial fatty acids are opposite with that of the fatty acids from plant oils.     

Separation, detection, and quantification of hydroperoxides formed at side-chain and backbone sites on amino acids, peptides, and proteins

Hydroperoxides are major reaction products of radicals and singlet oxygen with amino acids, peptides, and proteins. However, there are few data on the distribution of hydroperoxides in biological samples and their sites of formation on peptides and proteins. In this study we show that normal-or reversed-phase gradient HPLC can be employed to separate hydroperoxides present in complex systems, with detection by postcolumn oxidation of ferrous xylenol orange to the ferric species and optical detection at 560 nm. The limit of detection (10-25 pmol) is comparable to chemiluminescence detection. This method has been used to separate and detect hydroperoxides, generated by hydroxyl radicals and singlet oxygen, on amino acids, peptides, proteins, plasma, and intact and lysed cells. In conjunction with EPR spin trapping and LC/MS/MS, we have obtained data on the sites of hydroperoxide formation. A unique fingerprint of hydroperoxides formed at alpha-carbon (backbone) positions has been identified; such backbone hydroperoxides are formed in significant yields only when the amino acid is part of a peptide or protein. Only side-chain hydroperoxides are detected with free amino acids. These data indicate that free amino acids are poor models of protein damage induced by radicals or other oxidants.     

Amino Acid Changes in a Genetic Strain of Epileptic Beagle Dogs

A neurochemical evaluation of beagle dogs with naturally occurring spontaneous generalized convulsive seizures was performed. Amino acid profiles of serum, cerebrospinal fluid (CSF), and biopsied cerebral cortex from epileptic dogs were compared with those from seizure-free siblings. No differences in absolute levels were noted. However, when levels were normalized as a percent of total free amino acids, seizures was performed. Amino acid profiles of serum, cerebrospinal fluid (CEF), and biopsied cerebral cortex from epileptic dogs were compared with those seizure-free siblings. No differences also the two groups differed in certain respects. Ten significant correlations between amino acid pairs appeared in epileptic dogs, but only one was seen in seizure-free animals. Seven of these ten correlations involved glutamate or taurine. It was noted that the highly correlated amino acids (taurine, glutamate, glycine, glutamine, alanine) all utilize sodium-dependent membrane transport processes. The sum of glutamate, aspartate, and glycine levels (competing sodium-dependent high-affinity systems) was significantly lower in epileptic beagles. Since this difference was noted in serum but not CSF or brain, it may indicate a diminished capacity of sodium-dependent high-affinity renal transport for acidic and certain small neutral amino acids.     

Sterospecific selection of nucleophilic compounds in the chymotrypsin-catalyzed formation of the peptide bonds]

The course of stereospecific selection of nucleophilic compounds was studied in the reaction of acyl-enzymes interaction with razemic substrate-like nucleophiles, e.g. amino acid esters, by measuring optical rotation or incorporation of labelled D-compounds. It was shown that the acyl-enzymes are not responsible for the stereospecific selection of substrate-like nucleophiles. Since stereospecific selection of nucleophiles occurs in some chymotrypsin-catalyzed reactions, such selection may be produced by chymotrypsin till the formation of an acyl-enzyme compound with the substrate at the enzyme-inhibitor stage (or the Michaelis complex) with nucleophilic compounds. Even under the optimal conditions no absolute stereospecific selection of nucleophiles occurred, as was observed in case of a substrate (a donor of the acyl amino acid residue), undergoing degradation. An essential role of a specific site of nucleophile binding in the reactions of chymotrypsin-catalyzed peptide bond formation, is emphasized.     

Synthesis and circular dichroism studies of N,N-bis(2-quinolylmethyl)amino acid Cu(II) complexes: determination of absolute configuration and enantiomeric excess by the exciton coupling method

We report a method to determine the absolute configuration of alpha-amino acids by exciton coupled circular dichroism (ECCD). Naturally occurring amino acids were successfully derivatized with 2-bromomethylquinoline. Complexation of these conformationally flexible ligands with Cu(II) salts yielded defined propeller-like structures. The direction of the twist (i.e., the relative orientation of the chromophores to each other) is governed by the asymmetric amino acid carbon center. The transition moments of the chromophores couple and yield a bisignate circular dichroism spectrum, the sign of which corresponds to the absolute configuration of the chiral center of the amino acid. Enantiomeric excess (e.e.) of amino acid derivatives is linearly related to the differential extinction coefficient Delta(epsilon) and can be assessed easily utilizing a standard curve. This efficient, sensitive technique requires low analyte concentrations, offers several advantages over established methods, and could be applied in medicinal, pharmaceutical, or chemical retail and manufacturing industry.     

Assignment of the absolute configuration of [n]-ladderanes by TD-DFT optical rotation calculations

In this study, we report theoretical specific rotation values for a series of cis-/trans-alkylated-[5]-ladderanes and cis-/trans-methylated-[n]-ladderanes. Using time-dependent density functional response theory optical rotation calculations, we can assign (+) and (-) optical rotation signs to trans-(S)-alkyl-[5]-ladderane and trans-(R)-alkyl-[5]-ladderane configurations, respectively. In order to qualitatively validate our absolute configuration predictions, we computed optical rotation values at three different levels of theory--B3LYP, RI-BP86, and Hartree-Fock--using the aug-cc-pVDZ basis set. We observe a novel rung-parity-controlled oscillatory optical rotatory phenomenon in our computations, which, to the best of our knowledge, has never been reported or observed before. Furthermore, this preliminary analysis of optical rotation properties in this class of compounds should facilitate the correct absolute stereochemical assignment of natural and synthetic ladderanes, such as the trans-isomer of pentacyclic C(20)-fatty acid methyl ester (pentacycloanammoxic methyl ester), without the need for derivatization, in particular for cases where NMR or X-ray crystal structures are not readily available.     

Deletion analysis of MotA and MotB, components of the force-generating unit in the flagellar motor of Salmonella

MotA and MotB are cytoplasmic membrane proteins that form the force-generating unit of the flagellar motor in Salmonella typhimurium and many other bacteria. Many missense mutations in both proteins are known to cause slow motor rotation (slow-motile phenotype) or no rotation at all (non-motile or paralysed phenotype). However, large stretches of sequence in the cytoplasmic regions of MotA and in the periplasmic region of MotB have failed to yield these types of mutations. In this study, we have investigated the effect of a series of 10-amino-acid deletions in these phenotypically silent regions. In the case of MotA, we found that only the C-terminal 5 amino acids were completely dispensable; an adjacent 10 amino acids were partially dispensable. In the cytoplasmic loop region of MotA, deletions made the protein unstable. For MotB, we found that two large segments of the periplasmic region were dispensable: the results with individual deletions showed that the first consisted of six deletions between the sole transmembrane span and the peptidoglycan binding motif, whereas the second consisted of four deletions at the C-terminus. We also found that deletions in the MotB cytoplasmic region at the N-terminus impaired motility but did not abolish it. Further investigations in MotB were carried out by combining dispensable deletion segments. The most extreme version of MotB that still retained some degree of function lacked a total of 99 amino acids in the periplasmic region, beginning immediately after the transmembrane span. These results indicate that the deleted regions in the MotA cytoplasmic loop region are essential for stability; they may or may not be directly involved in torque generation. Part of the MotA C-terminal cytoplasmic region is not essential for torque generation. MotB can be divided into three regions: an N-terminal region of about 30 amino acids in the cytoplasm, a transmembrane span and about 260 amino acids in the periplasm, including a peptidoglycan binding motif. In the periplasmic region, we suggest that the first of the two dispensable stretches in MotB may comprise part of a linker between the transmembrane span of MotB and its attachment point to the peptidoglycan layer, and that the length or specific sequence of much of that linker sequence is not critical. About 40 residues at the C-terminus are also unimportant.     

Evaluation of the concentration and enantiomeric purity of selected free amino acids in fermented malt beverages (beers)

Even though amino acids are important trace components in the brewing of beers, they have not been extensively evaluated in these beverages. Studies involving the enantiomeric composition of these amino acids are even less prevalent. A brief summary of the brewing process for malt beverages is given. The total concentration and enantiomeric composition of three amino acids (leucine, phenylalanine, and proline) were determined in 25 different beers. Proline tended to have the highest average absolute concentration and the lowest percentage of the D -enantiomer in most samples. In some cases the relative amounts of D -phenylalanine and D -leucine exceeded 10% of the individual amino acids. The enantiomeric composition of the amino acids in different beer samples did not vary as extensively as the absolute concentrations. The reason for the concentration differences between proline and the other amino acids is discussed. A knowledge of amino acid concentrations and enantiomeric compositions appears to be useful in characterizing specific beers and brewing processes. © 1996 Wiley-Liss, Inc.     

Thehalose mycolates from Nocardia asteroides,Nocardia farcinica,Gordona lentifragmenta and Gordona bronchialis

Isolation of glycolipids from Nocardia asteroides, N. farcinica, Gordona lentifragmenta and G. bronchialis, by column chromatography of lipid extracts on a 50% (w/w) mixture of silicic acid and silica gel H, is described. The isolated materials were partially characterized by infrared spectroscopy, optical rotation and refractive index measurements, and by identifying the products of alkaline hydrolysis. Analytical studies showed that the glycolipids released only trehalose in the aqueous phase while mycolic acids were the constituent fatty acids identified.The isolated lipids are trehalose esters in which the trehalose molecule is esterified with mycolic acids.