Plasma amino acids and nitrogen retention of human subjects who consumed isonitrogenous diets containing rice and wheat or their constituent amino acids with and without additional lysine

Six diets furnished 6.0 g of nitrogen to adult human subjects as follows: a combination of rice and wheat; mixtures in which either 100 or 50% of the amino acids in rice and wheat were replaced by their constituent amino acids; and similar diets in which lysine was increased from 1.2 to 1.8 g. Nitrogen retention was greater (P less than 0.01) and concentrations of several amino acids in plasma were lower when the cereals were supplemented with lysine than when all other diets were fed. Data obtained with mixtures of amino acids apparently cannot always be used interchangeably with that from cereal-based diets, even when amino acid content is the same. Changes in plasma amino acids are a useful indicator of utilization of dietary amono acids when nitrogen balance also is determined.     

Plasma amino acids in anorexia nervosa

OBJECTIVE: To evaluate the amino acid profile in a group of adolescents with anorexia nervosa, and to apply alternative ways of presenting and assessing results, so as to increase the information available for understanding the metabolic abnormalities developed in these patients. DESIGN: Plasma amino acid concentrations of a random group of patients with anorexia nervosa compared with values obtained from a 'healthy' adolescent population. SETTING: The study was performed at the tertiary children's Hospital Sant Joan de Deu. SUBJECTS: Female adolescents (n = 92, age: 15+/-1.8 y) at diagnosis of anorexia nervosa. Reference values for amino acids were obtained from apparently healthy adolescents (by history and analytical data) who underwent presurgical analysis for minor operations. Interventions: Plasma amino acid concentrations were measured by ion exchange chromatography. Basic laboratory analysis, carnitine and IGF-I were also determined. RESULTS: In anorexic patients plasma concentrations of taurine, asparagine, glutamine, glycine, methionine, phenylalanine, ornithine, and histidine were significantly higher than reference values (Mann-Whitney, P < 0.01-0.0001), whereas arginine and cystine were lower than our reference values (P < 0.0001). Relative amino acid values (the molar fraction of the patient medians relative to control medians) were plotted. The ratios of some amino acids were significantly greater than those obtained from the reference population: Phe/Tyr (P < 0.001), Met/Cys (P < 0.0001), and Gly/Val (P < 0.01). CONCLUSIONS: A trend to hyperaminoacidemia is a common feature in anorexia nervosa. Although absolute amino acid values cannot play a significant role in the assessment of nutritional status in this condition, the calculation of some ratios (Phe/Tyr, Met/Cys and Gly/Val) and the graphical representation of relative values may be useful. The plasma amino acid profile in anorexia nervosa is different from those of other severe malnutrition states, showing a marasmic pattern of balanced protein-energy undernutrition. Cystine and arginine may be considered limiting amino acids in this disease, and the consequences of their deficient concentrations for oxidative damage should be further evaluated.     

Plasma and urinary amino acids and selected sulfur metabolites in young men fed a diet devoid of methionine and cystine

A preliminary investigation was conducted to explore the use of plasma methionine and cystine for determining human sulfur amino acid requirements. Measurements of urinary methionine, cystine, taurine, and inorganic sulfate were included. After a 3-day control period, three young men were fed for 8 days a diet containing a purified -l-amino acid mixture, patterned after egg protein but devoid of methionine and cystine. Fasting plasma methionine and cystine levels showed little decrease during the 8-day period. Urinary cystine and taurine responses were inconsistent among the subjects. Urinary methionine and inorganic sulfate levels decreased markedly within a few days after feeding of the experimental diet, and may be useful criteria for determining human sulfur amino acid requirements.     

Threonine and methionine are limiting amino acids for protein synthesis in patients with AIDS

This study was conducted to identify the most rate-limiting amino acids for whole-body protein synthesis in acquired immunodeficiency syndrome (AIDS) patients. We postulated that an essential amino acid that would be rate limiting in AIDS should have a low basal plasma concentration and should remain at a low level during amino acid infusion. Seven male AIDS patients (median age 37 y, CD4 cell count: 76 mm-3) without any clinically active opportunistic infection during the month before the experiment were infused intravenously with a complete amino acid-glucose mixture for 2.5 h. Eight healthy volunteers were used as controls. Before the infusion, the concentrations of most free essential amino acids (methionine, threonine, histidine, isoleucine, leucine and tryptophan) were significantly lower (P < 0.05) in AIDS patients than in controls. Most plasma free essential amino acids increased significantly during infusion. However, the absolute increase above basal levels for threonine, valine, lysine, (P < 0.05) and methionine (P < 0.073) was smaller in AIDS patients than in control subjects. Thus, threonine and possibly methionine may be rate limiting for whole-body protein synthesis in AIDS patients, suggesting that there are selective amino acid requirements in patients with AIDS.     

Utilization of ileal digestible amino acids by growing pigs: methionine

An experiment was conducted to determine the utilization of ileal digestible methionine by growing pigs. Three methionine-deficient diets (0.09 g ileal digestible methionine/MJ digestible energy (DE)) were formulated using cottonseed meal, meat-and-bone meal and soya-bean meal respectively as the only source of methionine in the diet. An additional three diets were formulated with supplements of methionine to confirm that methionine was limiting in the first three diets. The growth performance and retention of methionine by pigs given the six diets over the 20-45 kg growth phase was then determined. Growth rates (g/d) of pigs given the three diets formulated to 0.09 g ileal digestible methionine/MJ DE were significantly different (P < 0.01): cottonseed meal 411, meat-and-bone meal 442, soya-bean meal 496 (SED 24.6). The response of pigs to the addition of methionine confirmed that methionine was limiting in these diets. Crude protein (N x 6.25) deposited by the pigs (g/d) was significantly higher (P < 0.05) for those given soya-bean meal (61) and meat-and-bone meal (57) relative to cottonseed meal (47; SED 3.3). The proportion of ileal digestible methionine retained by pigs given the three protein concentrates was: cottonseed meal 0.39, meat-and-bone meal 0.45, soya-bean meal 0.47 (SED 0.019). These results indicate that values for the ileal digestibility of methionine in protein concentrates do not reflect the proportion of methionine that can be utilized by the pig. It appears that, with heat-processed meals, a considerable proportion of the methionine is absorbed in a form(s) that is (are) inefficiently utilized.     

Initiation of protein synthesis in mammalian cells with codons other than AUG and amino acids other than methionine

Protein synthesis is initiated universally with the amino acid methionine. In Escherichia coli, studies with anticodon sequence mutants of the initiator methionine tRNA have shown that protein synthesis can be initiated with several other amino acids. In eukaryotic systems, however, a yeast initiator tRNA aminoacylated with isoleucine was found to be inactive in initiation in mammalian cell extracts. This finding raised the question of whether methionine is the only amino acid capable of initiation of protein synthesis in eukaryotes. In this work, we studied the activities, in initiation, of four different anticodon sequence mutants of human initiator tRNA in mammalian COS1 cells, using reporter genes carrying mutations in the initiation codon that are complementary to the tRNA anticodons. The mutant tRNAs used are aminoacylated with glutamine, methionine, and valine. Our results show that in the presence of the corresponding mutant initiator tRNAs, AGG and GUC can initiate protein synthesis in COS1 cells with methionine and valine, respectively. CAG initiates protein synthesis with glutamine but extremely poorly, whereas UAG could not be used to initiate protein synthesis with glutamine. We discuss the potential applications of the mutant initiator tRNA-dependent initiation of protein synthesis with codons other than AUG for studying the many interesting aspects of protein synthesis initiation in mammalian cells.     

Reaction of osmium reagents with amino acids and proteins. Reactivity of amino acid residues and peptide bond cleavage

We report a study of the relative reactivity of the common amino acids and of their residues in lysozyme with osmium tetroxide, the osmium tetroxide-pyridine reagent, and with the oxo-osmium(VI)-pyridine reagent. With free amino acids, the osmium(VIII) reagents are most reactive with Met, Cys, His, Thr, Ser, Trp, Lys, and Pro; the osmium(VI) reagent only reacts significantly with His, Met, Cys, Thr, and Ser. In lysozyme, only Cys, Met, and Trp react extensively with the osmium(VIII) reagents; with the osmium(VI) reagent, Cys and Met are most reactive. We also note evidence both for cross-linking of proteins and for peptide bond cleavage, which appears to have considerable specificity for tryptophanyl residues.     

Plasma amino acid patterns during supplemental intravenous nutrition of low-birth-weight infants

In 42 low-birth-weight infants (smaller than 1,200 gm), we have compared the effects of intravenous nutrition supplement versus conventional feedings on growth, morbidity, mortality, and plasma amino acid patterns. Despite similar total caloric intake in INS and control groups, weight gain was greater in the INS infants. The overall mortality rate did not differ in the two groups. Nonsurviving infants receiving INS lived longer (mean equal to 30 days) than nonsurviving CON infants (mean equal to 5 days). Complications were equally frequent in both groups except that hyperglycemia occurred more often in infants receiving INS. The plasma aminogram of the LBW infant is described and compared to those of the full-term infant and adult. Hypoaminoacidemia was present at birth in the LBW infants, concentrations of glutamine, alamine, glycine, histidine, and ornithine being significantly (P smaller than 0.05) below FT values. During INS, elevations of threonine, serine, and methionine above FT values occurred. Glutamine remained subnormal despite INS. Recommendations for an INS solution more suitable for use in LBW infants are presented.     

Plasma amino acids determined by liquid chromatography within 17 minutes

We present an HPLC method for the determination of amino acids in plasma. The method is based on automated precolumn derivatization of amino acids with o-phthalaldehyde, separation of the derivatives by reversed-phase chromatography, and quantification by fluorescence detection. Complete separation was achieved within 12 min. Total analysis time, including derivatization, chromatography, and reequilibration of the column, was 17 min. The assay was linear from 5 to 800 mumol/L for all amino acids. Recovery of amino acids added to plasma samples was 96-106%, except for tryptophan (89%). Within-run precision (CV) was 1.8-6.4%, and between-run precision was 2.1-7.2%. The method can be used for determining primary amino acids in plasma and cerebrospinal fluid. The simple sample preparation and short analysis time make the method particularly suitable for routine analysis of large series of samples.     

Apparent ileal digestibilities of amino acids in newly weaned pigs fed diets with protease-treated soybean meal

Apparent ileal digestibilities of amino acids were determined in pigs fed cornstarch-based diets with untreated or protease-treated soybean meal as the protein source. Sixteen pigs were fitted with a modified post valve T-cecum cannula on d 14, 15, and 16 after birth and then returned to their sows until d 20 when they were weaned. Twelve of the pigs were selected for the study, which was conducted according to a two-period balanced change-over design. Treatments consisted of soybean meal that was 1) untreated (SBM), 2) processed by incubation (1:2 wt/vol distilled water adjusted to pH 4.5, for 16 h at 50 degrees C; CI-SBM), 3) sprayed with protease (supplied at 1 microL/g of soybean meal; PS-SBM), and 4) processed by incubation, as for CI-SBM, with protease in the water at the rate of 1 microL/g of soybean meal (PI-SBM). Each period consisted of 5 d of adaptation to diets followed by three 8-h collection periods (total of 24 h) that alternated with 8-h periods in which digesta were not collected. Apparent CP digestibilities were similar (P > .05) at 70.4, 72.4, 65.2, and 70.3% for the SBM, CI-SBM, PS-SBM, and PI-SBM diets, respectively. Corresponding amino acid digestibilities were also similar (P > .05), ranging from 62.5, 67.5, 57.9, and 65.0% for alanine to 83.5, 83.4, 78.7, and 84.7% for arginine. Apparent digestibilities were less (P < .05) for Period 1 (on d 7 after weaning) than for Period 2 (on d 16 after weaning). In conclusion, protease treatment of soybean meal had no effect on ileal digestibilities of CP and amino acids in newly weaned pigs.     

Characterization of azo coupling adducts of benzenediazonium ions with aromatic amino acids in peptides and proteins

A synthetic peptide, VLSPADKTNWGHEYRMF(cmC)QIG, was reacted with 4-chlorobenzenediazonium hexafluorophosphate as a model for reactions of aromatic diazonium ions with proteins. At a ratio of diazonium ion to peptide of 0.8:1, three products could be seen by reversed-phase HPLC. Electrospray mass spectrometric analysis of the isolated products revealed that two of the products had the same mass of 2648 Da, being 138 Da higher than the parent peptide and corresponding to the addition of a 4-chlorobenzenediazo group. The third isolated product had a mass of 2787 Da which corresponded to the addition of two 4-chlorobenzenediazo groups (276 Da). Digestion of the monoadducted intact peptides with trypsin or endoproteinase Glu-C and HPLC separation of adduct oligopeptides followed by sequencing with electrospray ionization tandem mass spectrometry showed unambiguously that histidine and tyrosine residues were the major sites of modification. Incubation of human serum albumin with 4-chlorobenzenediazonium hexafluorophosphate at molar ratios of 1:1, 1:2, and 1:10 resulted in adduct formation as detected by shifts in the HPLC retention time of the protein and also by an increase in mass as determined by electrospray mass spectrometry.     

The intake of proteins and essential amino acids in top-competing women athletes]

PURPOSE: The literature on diagnostic peritoneal lavage in the assessment of blunt abdominal trauma reflects an ongoing controversy. Therefore we conducted a prospective evaluation of the diagnostic management of blunt abdominal trauma used at our clinic, in which this procedure plays a substantial role. During the years 1993 and 1994 a total of 75 patients could be included in the study. The study population consisted of all patients with a diagnosis of blunt abdominal trauma. In addition, all trauma patients who were unresponsive on admission to the emergency receiving unit underwent the same program of diagnostic work-up. This group included polytraumatized patients, patients with craniocerebral injuries and all those who had been intubated prior to admission. Patients with stable vital signs were evaluated first by sonography of the abdomen, whereas those showing signs of hypovolemic shock received a diagnostic peritoneal lavage as the first evaluation of abdominal trauma. In order to assess the relative value of the two diagnostic methods, all patients who had had ultrasound as their first examination subsequently also underwent peritoneal lavage. RESULTS: 37 patients (49%) had lavage evidence of intraperitoneal bleeding. Of these 22 (29% of the total) subsequently underwent emergency laparotomy with lesions requiring surgical treatment found in 21 (95%). Only in one patient (1.3% of the study population) laparotomy failed to reveal a lesion requiring surgical correction. The accuracy of peritoneal lavage findings as an indication for laparotomy was 99%, compared to 82% for ultrasonography used as a initial diagnostic procedure. Diagnostic peritoneal lavage is quick, safe and almost independent of the experience of the investigating physician. It can be performed during other diagnostic procedures and can be repeated at will. If beyond macroscopical evaluation the lavage fluid is assessed chemically, even duodenal and pancreatic lesions as well as injuries to other hollow viscera can be suspected. With a sensitivity of 100% and a specificity of 98%, diagnostic peritoneal lavage is an extremely reliable diagnostic tool. It should be used as the initial diagnostic procedure in all hypovolemic and/or unresponsive patients suspected of having suffered blunt abdominal trauma. In conscious patients with stable vital signs, ultrasonography can be used for initial diagnosis. It should, however, be complemented by subsequent peritoneal lavage whenever the clinical course gives rise to suspicion.     

Transaminase of branched chain amino acids. XI. Leucine (methionine) transaminase of rat liver mitochondria

An aminotransferase (transaminase) which is active for leucine and methionine, but not for valine or isoleucine, was purified from rat liver mitochondria. The purified preparation appeared homogeneous on polyacrylamide disc gel electrophoresis. Its molecular weight was shown to be 55 000 by gel filtration. It differed from enzyme II (leucine aminotransferase, EC 2.6.1.6) in the supernatant fraction, another transaminase which is also specific for leucine and methionine, in molecular weight, Km values for substrates, electrophoretic mobility, chromatographic behavior and heat stability. From comparison with related transaminases it was concluded to be a new enzyme and named mitochondrial leucine (methionine) transaminase.     

Plasma and erythrocyte alpha-tocopherol and plasma retinol concentrations in term infants fed formula enriched with long-chain polyunsaturated fatty acids

OBJECTIVE: To assess the effect of long-chain polyunsaturated fatty acids (LCPUFA)- and vitamin E-supplemented formula feeding on erythrocyte and plasma alpha-tocopherol (VE), and plasma retinol (VA) concentrations in neonates and to compare these values with those found in infants feeding on infant formula without LCPUFA or breast milk SETTING: University Hospital of Granada, Spain. SUBJECTS: 49 full-term infants. DESIGN AND INTERVENTION: Subjects who chose not to breast feed were fed either (i) unsupplemented infant formula (F) or (ii) infant formula supplemented with LCPUFA and vitamin E (FL). Alpha-tocopherol and retinol were measured at 7 days, 1 month and 3 months. RESULTS: Plasma and erythrocyte VE concentrations and plasma VE/total lipids ratio increased significantly in all groups at 1 month of life (P < 0.05), but did not change significantly between 1 month and 3 months in any group (P > 0.05). Erythrocyte VE and VA retinol concentrations were higher in infants fed an infant formula than in breast milk-fed infants at 1 month of life (P < 0.05). Finally, there were no significant differences in plasma or erythrocyte VE levels, plasma VA or plasma VE/total lipid ratio between any groups at 3 months of life (P > 0.05). CONCLUSION: Infants fed on LCPUFA- and vitamin E-supplemented infant formula for 3 months have similar vitamin E and A status to infants fed on breast milk or infant formula without LCPUFA supplementation.     

Allergy to extensively hydrolyzed cow milk proteins in infants: identification and treatment with an amino acid-based formula

OBJECTIVE: To evaluate acetowhite changes of the cervix and vulva as a predictor of human papillomavirus (HPV) infection. METHODS: In this population-based study all women aged 19, 21, 23, and 25 years and registered as living in a primary health care area within the city of Ume?, Sweden were eligible for inclusion. Each participant underwent a gynecologic examination with sampling of epithelial cells for HPV-DNA detection and Papanicolaou smear. Colposcopy was performed 5 minutes after application of 5% acetic acid. A two-step polymerase chain reaction (PCR) technique was employed for HPV-DNA detection. RESULTS: Colposcopy and sampling of epithelial cells could be performed in 535 women. The sensitivity of detection of HPV infection by the acetowhitening of the cervix was 22% (95% confidence interval [CI] 18%, 26%). The specificity of detection of HPV infection by the acetowhitening of the cervix was 90% (95% CI 87%, 93%). The sensitivity of detection of HPV infection by cytology was 13% (95% CI 10%, 16%), and the specificity was 99% (95% CI 98%, 100%). The combination of acetowhitening and cytology did not improve the diagnostic value. CONCLUSION: Acetowhitening of the cervix and vulva has low sensitivity as a predictor of HPV infections as determined by PCR.     

Availability of amino acids from casein proteins subjected to heating under model conditions

Localization of a radiopharmaceutical agent in a "tumor" is best conceptualized in terms of the altered regional physiology attendant to the presence of the "tumor". Such localization should be expected to occur in association with other disease states characterized by similar altered regional physiology. Neoplasms, areas of inflammation, and certain phases of infarct development are characterized by increased permeability of their capillary beds to macromolecules. This is largely due to neovascularization and the large intercapillary pores associated with new growth of capillary beds in these circumstances. Often, total perfusion to such lesions is increased in comparison to surrounding normal tissue. Thus, in all three clinical conditions, the entry of macro-molecules into the interstitial fluid space from the intravascular space is increased above that seen in normal tissue. Moreover, with neoplasms and inflammatory processes, there may be a delay in new lymphatic vessel growth adding to the residence time of the macromolecules in the interstitial fluid space. In all three conditions, the increased macrophage activity associated with tissue necrosis may result in ingestion of the labeled macromolecule by the macrophage. Pinocytosis may result in ingestion of the labeled macromolecule by other cells in the lesion, or there may be specific receptor sites on the cell membrane for the macromolecule, which may lead to fixation of the labeled macromolecule on the cell surface and possible intracellular translocation of the label itself. Radiolabeled macromolecules such as albumin, fibrinogen, or gamma globulins, and radionuclides that bind to macromolecules such as radiogallium, radioindium, and other radioelements, exhibit localizing behavior in tumors, inflammatory lesions, and during certain stages of infarcts. In the case of radiogallium and radioindium, the binding macromolecule is transferrin, and it is known that some cells have specific receptor sites for transferrin-bound iron on the cell membrane. It is possible that certain cells within these lesions have cell membrane receptor sites for radiogallium- and radioindium-labeled transferrin, and the cell erroneously accepts these radioelements from the transferrin in lieu of iron in attempting to engage in heme enzyme synthesis. Another mechanism that may be operative in the localization of agents in neoplasms, infarcts, and inflammatory lesions may be the altered cell permeability found in many cells of such lesions. It is known that many agents, such as supravital dyes, are excluded from entering normal cells by the selective permeability of normal cell membranes. When cell membrane permeability is altered, such as can be seen in traumatized, dying, or dead cells, the normally excluded agent may penetrate the abnormal cell membrane and bind, and consequently accumulate in intracellular constituents.     

Aromatic amino acid transamination and methionine recycling in trypanosomatids

Although trypanosomatids are known to rapidly transaminate exogenous aromatic amino acids in vitro and in vivo, the physiological significance of this reaction is not understood. In postmitochondrial supernatants prepared from Trypanosoma brucei brucei and Crithidia fasciculata, we have found that aromatic amino acids were the preferred amino donors for the transamination of alpha-ketomethiobutyrate to methionine. Intact C. fasciculata grown in the presence of [15N]tyrosine were found to contain detectable [15N]methionine, demonstrating that this reaction occurs in situ in viable cells. This process is the final step in the recycling of methionine from methylthioadenosine, a product of decarboxylated S-adenosylmethionine from the polyamine synthetic pathway. Mammalian liver, in contrast, preferentially used glutamine for this reaction and utilized a narrower range of amino donors than seen with the trypanosomatids. Studies with methylthioadenosine showed that this compound was readily converted to methionine, demonstrating a fully functional methionine-recycling pathway in trypanosomatids.     

The prediction of the conformation of membrane proteins from the sequence of amino acids

The methods of Chou & Fasman [Biochemistry (1974) 13, 211-222, 222-245] and of Lim [J. Mol. Biol. (1974)88, 857-872, 873-894] for predicting secondary structure from amino acid sequence have been applied to five predominantly helical membrane-associated peptides. The predictions from the method of Lim (1974a,b) are consistent with the experimental observations, whereas those from Chou & Fasman (1974a,b), although not inconsistent with alpha-helix, favour a beta-structure for several very hydrophobic regions. The results may be rationalized in terms of the effect of the solvent on the conformation of a polypeptide.