Amino acid analysis concentration

Analysis of the blood of a catatonic football fan revealed large concentrations of a psychotoxic octapeptide. Amino acid analysis of this octapeptide gave the following results ... 

Since it is possible to differentiate well-preserved from badly preserved collagen through amino acid analysis and gel electrophoresis, it is also possible to determine which bone samples are likely to give erroneous isotopic ratios. At least for 8 C, it should be possible to estimate the in vivo isotopic signature by correcting the changed amino acid concentrations of the collagen extract. This way, a reasonable approach to the reconstruction of pale-odiet should be possible. 

Furthermore, the distribution of this peptide is compatible with PCP receptor densities, with highest concentrations of both ligand and receptor in areas that could be relevant to the psychotomimetic properties of PCP. Amino acid analysis of an aliquot of the purified fraction shows the peptide to contain at least 26 residues. Because the peptide is blocked at the N-terminus, enzyme fragments have been generated and purified over HPLC for sequence determination. 

Houpert, Y., Tarallo, P., and Siest, G., Amino acid analysis by ion-exchange chromatography using a lithium elution gradient. Influence of methanol concentration and sample pH, /. Chromatogr., 115, 33, 1975. 

Determination of the extinction coefficient is a relatively straightforward task. The target protein is diluted to give live different concentrations. These samples are then divided into two aliquots. Amino acid analysis (AAA) accurately determines the protein concentration of one set of samples at the live concentrations, and the absorbance at 280 nm (A280) is measured for the other set of samples. The slope of a plot of A2s0 vs. protein concentration by AAA yields the extinction coefficient. 

The fourth category of protein assay is amino acid analysis. This method is the most accurate and robust method for determination of protein concentration, but is appropriate only for pure proteins. In addition, it is relatively slow and requires specialized instrumentation and knowledge of the target protein s theoretical amino acid composition. 

The phospholipid concentration in a (proteo)liposome suspension can be determined by phosphorus analysis (73) and the protein concentration by automated amino acid analysis or by a calibrated colorimetric protein assay... 

As described for lEC, elution is done by a stepwise or a continuous change of buffer composition. The mildest elution buffer is an aqueous buffer with low ionic strength, e.g., 20 mm Tris-HCl. If it is not successful, desorb with a chaotropic solvent, e.g., 2 M potassium rhodanide (thiocyanate), 2.5 M guanidinium hydrochloride, up to 7 M urea, or with increasing concentrations of methanol or acetonitrile. Especially the use of rhodanide or urea may be accompanied by a chemical modification of amino acid side chains, which disturbs amino acid analysis. 

Compound 29 (2.44 g, 1.4 mmol) was dissolved in TFA (50 mL) at —10 °C and the soln allowed to react for 1 h at rt. The soln was concentrated to a residue, which was then triturated with 5.2 M HC1 in dioxane (0.4 mL, 2.1 mmol) and the residue precipitated with Et20 to give a powder. The product was washed with Et20 and dried. The entire product and 41 (3.0 g, 1.47 mmol) were dissolved in DMF (100 mL), and HODhbt (0.24 g, 1.47 mmol) was then added. EDC (free base, 0.27 mL, 1.47 mmol) was added to the soln with stirring at —10 °C, and the mixture allowed to react at rt for 8h. The product was precipitated by adding an excess of H20, the precipitate collected by filtration, washed successively with MeOH, EtOAc, and hexane, and then dried yield 4.5 g (87%) TLC (Merck Kieselgel 60F-254) R 0.41 (CHC13/ 80% aq AcOH/TFE 6 1 1). Formation of the desired product 42 was confirmed by amino acid analysis. 

The reduced form of Na+, K+-ATPase inhibitor-I (10) was obtained by treatment of the protected peptide synthesized by the soln procedure with HF, followed by reaction with Hg(OAc)2. After purification of the crude product on Sephadex G-25, the reduced peptide (110 mg) was dissolved in 0.1 M NH4OAc buffer (1L, pH 7.8) at a peptide concentration of 0.018 mM and then stirred at rt. After 24 h, the major peak in the HPLC, which coeluted with the natural product, corresponded to 55% of the product distribution. The mixture was acidified to pH 3 with AcOH and 10 was purified by RP-HPLC. When the oxidation was carried out in the presence redox reagents at a peptide/GSH/GSSG ratio of 1 100 10, after 24 h the major oxidation product increased to 69%. The mixture was acidified with AcOH and the product (10) isolated by preparative HPLC yield 20%. The product was characterized by MALDI-TOF-MS and amino acid analysis a combination of enzymatic peptide mapping and synthetic approaches were applied to assign the cystine connectivities. 

HPLC. The crude product was dissolved in 0.1% aq TFA (10 mL), filtered, and purified by preparative HPLC [Preppak, Cartridge YMC ODS column (4.8 x 30 cm), gradient elution MeCN/0.1% TFA/H20 from 20 to 50% MeCN in 90 min (flow rate of 50mL-min-1)]. The fractions containing pure product were combined, concentrated, and lyophilized yield 22.4mg (44%) the peptide was characterized by HPLC (purity 98%), amino acid analysis, and FAB-MS. 

A soln of reduced glutathione (H-yGlu-Cys-Gly-OH 5.2 mg, 0.017mmol) and 2 (6.9mg, 0.029 mmol) in 50% aq TFA (1 mL) was kept at rt for 24 h. The mixture was concentrated and the residue dissolved in 10% aq AcOH (1 mL). The soln was then applied to a Sephadex G-25 SF column (3 x 54 cm), equilibrated, and eluted with 10% aq AcOH. The product was located in the effluent by spectrophotometry, the UV absorption being similar to that of tryptathionine. The relevant fractions were combined and lyophilized. The product was further purified by a second gel filtration yield 5 mg (58%). Amino acid analysis of an acid hydrolyzate with TosOH 10 gave Glu 1.00, Gly 0.98, Cys 0.70, oxindolylalanine 1.08, and traces of cystine. 

Hyperammonemia has occurred during parenteral nutrition as a component of therapy for renal insufficiency (905). The hyperammonemia presented as a change in mental status, developing about 3 weeks after initiation of parenteral nutrition therapy in most cases the episodes are of increasing duration and paroxysmal. In three of the patients, serum amino acid analysis in the acute phase showed reduced concentrations of ornithine and citrulline (the respective substrate and product of condensation with carbamyl phosphate at its entry into the urea cycle). Concentrations of arginine, the precursor to ornithine, were raised. 

A 61-year-old man with epilepsy had altered consciousness after his dose of valproate was increased because of poor seizure control. Electroencephalography showed triphasic waves and high-amplitude delta-wave activity with frontal predominance. Although serum aspartate transaminase and alanine transaminase were normal, the serum ammonium concentration was high at 960 ng/ml (reference range 30-470). Serum amino acid analysis showed multiple minor abnormalities. Valproate was withdrawn. He improved within 4 days... 

SEC and anionic HPLC-Q-ICP-MS can be used to determine the approximate molecular weight of Se-containing proteins and identify fractions for seleno-amino acid analysis. The coupling of anion and cation HPLC and Q-ICP-MS also allows selenoamino acids in hydrolyzed protein extracts to be measured. However, the lack of knowledge about species, low Se concentrations, and lack of detection power make quantification of Se species difficult. The concurrent use of HPLC-MS-MS is needed to confirm the identity of Se species, but at present the low concentrations of Se in extracts preclude its use. It is essential that CRMs are used to confirm the validity of results regarding total Se and Se species. 

Perhaps the most promising method for understanding the degradation mechanism for woolen archaeological textiles is the systematic examination of specimens by amino acid analysis. The study of the changes in concentration and composition of the amino acids present in the wool substrate for archaeological and modern specimens is currently under investigation. 

Both of these peptides exhibited considerable sequence identity with the locust adipokinetic hormone (Lom-AKH-I) and the crustacean red pigment-concentrating hormone (RPCH), as had been suggested by chromatographic behavior, N-terminal analysis, and amino acid analysis (3,4). Indeed, both peptides were shown to be far more potent as stimulators of hyperglycaemia than as myotropins (3). 

Barium hydroxide hydrolysis followed by amino acid analysis revealed a sulfated Tyr residue. Subsequently, the myotropic nature of PSK was demonstrated on the isolated hindgut preparation of P. americana where the threshold of activity concentration was determined to be 0.25 nM, virtually the same as the threshold concentration (0.22 nM) of LSK on the isolated L. maderae hindgut (13). In addition to PSK, the non-sulfated form of LSK-II was isolated and structurally characterized from the P. americana cc extracts, but the biologically active sulfated form was not found (19). 

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