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PubMed1980

The radiolysis of aqueous ammonium cyanide: compounds of interest to chemical evolution studies.

Oxygen-free aqueous solutions of NH4CN (0.1 M, pH9) were exposed to gamma rays from a 60Co source, the mixture of nonvolatile products was fractionated, and the fractions were analyzed. The procedures were chosen to make effective investigations of radiolytic products, and to minimize the contributions of chemical changes which are known to occur in aqueous solution in the absence of ionizing radiation. It has been found that the main constituents are: urea, 25.9%; an oligomer, very likely oligoimine (18.4%); and several fractions (about 50%) which release amino acids on hydrolysis. These fractions differ considerably, as shown by amino acid assay, enzymatic digestion, IR spectra, and biuret reaction. All these tests were found to be positive for two fractions; in two further fractions the enzymatic cleavage was absent, but other tests were positive. Negative enzymatic and biuret tests, and no bands characteristic of amide or peptide, were found for a fraction whose hydrolysate consisted of 55% glycine. Although most of the isolated materials were found to be composite, the results of the analyses were sufficient for getting a reliable over-all picture of the chemical action of the ionizing radiation. The role of free radicals in reactions leading to the formations of radiolytic products was considered.

Amino Acids↗
PubMed1976

Improved method for determining serum protein concentrations in the far ultraviolet.

We examined the feasibility of measuring absorbance in the far ultraviolet for serum protein determination with a medium-priced, double-beam spectrophotometer. The method has been simplified by using an automatic dilutor to dilute the serum, and a flow-through cell and recorder in conjunction with the spectrophotometer. Because it is only necessary to dilute the serum and determine its absorbance, the procedure is quite rapid. As compared to the biuret reaction, interferences from hemolysis have been about halved. Interferences owing to light scattering by turbid sera have also been appreciably decreased by use of an anionic detegent (sodium dodecyl sulfate) as a serum diluent. Comparisons of the present method to the biuret and Kjeldahl methods indicate that it is relatively accurate, and is practical for routine use.

Blood Proteins↗
PubMed1991

Continuous aqueous phase extraction of proteins: automated on-line monitoring of fumarase activity and protein concentration.

Automated assay techniques are described for on-line measurements of fumarase activity and total protein concentration, including in-line sample dilution and sample multiplexing during continuous aqueous phase extraction. Fumarase was determined by following the conversion of L-malate to fumurate at a wavelength of 250 nm, while the protein assay was based on the Biuret reaction. Actual assay times of 2 and 4 min were achieved for the fumarase and protein measurements, respectively, with an effective measurement cycle time of 2 min. Standard deviations of c. 3.2 and 2% of the measured values were calculated for the enzyme and protein values, respectively. The assay system was coupled to a computer to allow on-line data visualization and storage.

Automation↗
PubMed2006

Observation of the complex formation between Cu(II) and protein by capillary electrophoretic system incorporating an UV/CL dual detector.

We investigated the complex formation between Cu(II) and human serum albumin (HSA) through a biuret reaction by use of capillary electrophoretic system incorporating an ultra-violet absorption (UV) and chemiluminescence (CL) dual detector. Cu(II)-tartrate complex and Cu(II)-human serum albumin complex were detected by UV detection (282 nm) with on-capillary, followed by CL detection (luminol-hydrogen peroxide CL reaction) with end-capillary. We examined the effects of the reaction time and temperature on the UV and CL responses. On the basis of the obtained results we considered the Cu(II)-human serum albumin complex formation processes and its catalytic activity for the CL reaction. The system easily, rapidly, and simultaneously produced useful information concerning the complex formation of Cu(II) and human serum albumin due to the presence of the both detectors.

Copper↗
PubMed1989

Electrochemical detection of peptides.

A general method of wide applicability for the determination of peptides is described. Peptides longer than dipeptides react in the classical biuret reaction with Cu(II) to yield electroactive Cu(II)-peptide complexes that can be oxidized to the corresponding Cu(III) complexes. This allows the sensitive electrochemical detection of peptides following their separation by reversed-phase liquid chromatography. The reaction chemistry, which is reversible, allows for the determination of peptides that lack an electroactive group or a primary amine. Selectivity for a model peptide is 10(3)-10(4) over nonelectroactive amino acids.

Electrochemistry↗
PubMed1999

In patients with cirrhosis, serum albumin determination should be carried out by immunonephelometry rather than by protein electrophoresis.

OBJECTIVE: Serum albumin is a key parameter for prognosis in cirrhosis. We compared levels of serum albumin determined by both protein electrophoresis and immunonephelometry, with special reference to the Child-Pugh classification. DESIGN AND METHODS: One hundred and thirty-one patients, including 39 with cirrhosis, were included prospectively during 2 months. The aetiology of cirrhosis was mainly alcoholism (67%) and hepatitis C virus (HCV) (18%). Serum albumin was determined simultaneously by electrophoresis (Hydrasys SEBIA following protein determination by the biuret reaction) and by immunonephelometry (BECKMAN Nephelometer). Values were compared by non-parametric tests. RESULTS: For the whole population, electrophoretic and immunonephelometric values correlated (p = 0.85; P < 0.0001), but electrophoresis significantly overestimated serum albumin by a median 1.6 g/l (P < 0.0001) with a large spread in values (range, -3.9 to 12.7). Median overestimation in cirrhosis was 2.6 g/l (P < 0.0001; range, -2.0 to 10.2) and 1.0 g/l (P < 0.0001; range, -3.9 to 12.7) in patients without cirrhosis (difference, P < 0.02). For 6/39 (15.4%) patients with cirrhosis, this overestimation led to an underestimation in the Child-Pugh classification. CONCLUSION: In our experience, electrophoresis can lead to serum albumin values which are significantly different compared to those obtained by immunonephelometry. This discrepancy may lead to an incorrect Child-Pugh classification. Therefore, in the follow-up of cirrhotic patients, serum albumin should be determined by immunonephelometry.

Adult↗
PubMed1988

Measurement of total serum protein by near-infrared reflectance spectroscopy.

This study describes the measurement of total protein in serum by near-infrared reflectance spectroscopy. With an algorithm, generated by the calibration procedure, the protein content of serum samples was calculated from absorbance data at various wavelengths in the near-infrared. A good correlation (r = 0.993) was found between near-infrared reflectance spectroscopy measurement of serum protein and analysis by the biuret reaction.

Blood Proteins↗
PubMed2000

Effect of cultivation mode on a bioprocess for chromium yeast biomass enrichment.

Defined cultivation media for yeast growth which contained 278.8 mM of glucose and 0.1 mM of chromium(III) added as K2Cr(SO4)2 x 12 H2O was used in batch and combined batch/fed-batch cultivation mode. In fed batch cultivation mode the rate of substrate addition remained constant during growth of yeast and corresponded to a growth rate of 0.25 h(-1). In both cases the growth and yeast activity was followed by on line measurement of optical density, pH and pO2 at 30 degrees C. At the end of the bioprocess the concentration of protein in yeast biomass was determined off line by the biuret reaction. Total and organically bound chromium was detected by ETA-AAS. Different cultivation modes affected the total cell protein concentration of yeast grown in media supplemented with chromium. In batch process the protein content represented 25.7% of dry yeast biomass, in contrast in the mixed bioprocess this value was 16.9% one the same period of time. The influence of cultivation mode on chromium uptake was seen in total chromium accumulation which reached 8.68 +/- 0.16 micromol g(-1) d.wt. in batch and 1.92 +/- 0.04 micromol of chromium g(-1) of dry yeast biomass in combined batch/fed-batch cultivation mode. The opposite was observed for organically bound chromium. The 60% of total accumulated chromium was organically bound during yeast growth in combined batch/fed-batch mode. When yeast was grown in batch mode this value attained 13.5%. Results suggested that a combined batch/fed-batch mode of cultivation was more effective over a batch system in chromium biotransformation to organically bound chromium, regardless of the lower protein ratio determined in the yeast biomass.

Biomass↗
PubMed1975

Improved biuret procedure for routine determination of urinary total proteins in clinical proteinuria.

This communication describes and evaluates an improved routine methodology for quantitating clinical proteinuria. Based on investigations of Piscator and of Savory et al., a modified Tsuchiya's reagent (ethanolic HCI-phosphotungstic acid) is used to precipitate proteins at 56 degrees C, followed by biuret spectrophotometry at 540 nm. The accuracy of the proposed procedure was assessed by comparisons with results obtained by using an ultrafiltration membrane that retains solutes with an average molecular weight in excess of 10 000 for separating of urinary proteins before they are measured with the biuret reaction. Precision of the method (coefficient of variation) is typically 2-3%.

Evaluation Studies as Topic↗
PubMed1975

[Physiological proteinuria. Data of acrylamide-SDS gel electrophoresis and other methods of qualitative and quantitative analysis].

Since detection of the first stage of nephropathies requires an exact definition of physiological proteinuria, this has been sought in 97 healthy individuals. Measured by a modification of the biuret reaction, physiological proteinuria did not exceed 100 mug/min in the recumbent position (average of 24.5 mug/min) and 150 mg/24 h (average of 51 mg/24 h). No significant differences were seen with respect to age or sex. Qualitative analysis of urinary proteins was done by cellulose acetate electrophoresis, immunoelectrophoresis and polyacrylamide-SDS electrophoresis. This latter method has the great advantage of classifying the proteins according to their molecular weight, wtihout electrostatic interference. Its graphic representation delineates a normal zone allowing an objective distinction between physiological and pathological proteinurias. In the majority of cases, the proportion of albumin is between 25 and 55%. The orthostatic position increases proteinuria (average of 41 mug/min) with a tendency to a distribution which erroneously suggests a glomerulopathy. Accordingly, investigation for small changes in proteinuria should always be carried out on urine formed in the recumbent position.

Adult↗
PubMed2001

[Osmotic activity of plasma in plasmapheresis in patients with multiple myeloma].

AIM: To control safety and efficiency of therapeutic plasmapheresis (PA) by osmolality, colloido-osmotic pressure (COP), total protein concentration before and after the procedure in patients with paraproteinemic hemoblastosis. MATERIAL AND METHODS: 20 patients with multiple myeloma have undergone 42 PA procedures conducted by two techniques: continuous flow centrifugation on blood fractioners or intermittent centrifugation of blood in plastic containers. The removed plasma volume averaged 1/3 (group 1) or 2/3 of the plasma volume (group 2). The removed protein reached 62-197 g. Isotonic sodium chloride solution and/or reopolyglucin (20-60 g) replaced the removed plasm. Total protein concentration was measured colorimetrically in biuretic reaction, plasma osmolality--cryoscopically and COP--on Knauer osmometer. RESULTS: PA leads to a short decline in osmolality (97.0-99.1%), of total protein concentration (82.8-78.6%) and of COD (79.2% in replacement with saline and 90.2% in replacement with dextran). During recovery after the procedure plasma osmotic activity and protein concentration return to the baseline. CONCLUSION: In elimination of 1/3 of plasma volume and crystalloid infusion, hemodilution promotes release of abnormal proteins from the tissues into the circulation and thereafter removal them from the organism. In removal of 1/2 and more of plasma volume, COP demans correction made by administration of colloids, e.g. solution of low molecular dextran. There is a potential danger of COD lowering several hours after PA due to different speed of dextran elimination and mobilization of protein reserve.

Adult↗
PubMedDate not supplied

[Preparation and use of fish fillet infusion as a basic medium for culturing bacteria].

The authors present the first results on the utilization of fish infusion (IFP) as a basic medium for the cultivation of bacteria. The infusion was obtained from a common marine fish, corvina (Micropogonias furnieri) according to the technique used in the preparation of beef infusion broth. Streptococcus pyogenes, S. pneumoniae, Neisseria meningitidis, Campylobacter jejuni, Escherichia coli, Klebsiella pneumoniae, Serratia marcescens, Pseudomonas aeruginosa, Staphylococcus aureus and Bacillus subtilis were cultured in liquid and solid media prepared with IFP as well as in recommended standard media. Solid media used for cultivation of S. pyogenes, S. pneumoniae, N. meningitidis and C. jejuni were supplemented with 5% of defibrinated sheep blood and for the latter, substances recommended to increase aerotolerance were included in solid and liquid media. All of these strains grew on the media prepared with IFP except S. pneumoniae when cultured in IFP diluted 1:2 with a sodium chloride solution. Only S. pyogenes produced colonies smaller than those of the standard medium. No more differences were detect in the observation of colony morphology. The growth of E. coli, K. pneumoniae, S. marcescens, P. aeruginosa, S. aureus and B. subtilis was measured in liquid media after 8 hours. In solid media, the growth index was expressed by dividing the number of colonies produced in IFP-agar and Nutriente Agar by the number of colonies on Trypticase soy agar plates. Some differences were observed in colonial size and morphology when compared with those generated in standard media. The average value obtained from the analyses of total proteins by biuret reaction in twelve batches of IFP was 5.03 mg/ml. The experiments showed that culture media prepared with IFP supported the growth of bacterial strains used in this work. It suggests that fish infusion has promising conditions of being an alternative substrate for cultural purposes.

Bacteria↗
PubMed2003

[The expression of connective tissue growth factor in renal cortex of 5/6 nephrectomized rats and its modulation by fluvastatin].

OBJECTIVE: To investigate the connective tissue growth factor (CTGF) mRNA expression in the renal cortex of 5/6 nephrectomized rats and its modulation by fluvastatin, a 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor. METHODS: Twenty-four rats underwent operation 2 times: during the first operation 2/3 of the left kidney was resected, and the right kidney was resected completely one week after. The 24 5/6 nephrectomized rats were randomly divided into 2 groups: untreated 5/6 nephrectomized group (model group, n=12) and fluvastatin-treated 5/6 nephrectomized group (treatment group, fluvastatin was orally administered 7 mg.kg(-1).d(-1) for 13 weeks, n=12), and 6 sham-operated rats served as control (sham operation group). In the weeks 2, 4, 8, and 13 after the second operation metabolic cage was used to collect the 24-hour urine 2 times. Urine protein was examined by biuret reaction so as to calculate urinary protein excretion. By the end of experiment blood was collected to examine the serum cholesterol, triglyceride, urea nitrogen, and creatinine contents. The rats were killed and their kidneys taken out. The CTGF mRNA expression in the renal cortex was detected by RT-PCR. Immunohistochemistry was used to examine the expression of transforming growth factor-beta1 (TGF-beta1), type IV collagen and fibronectin in the glomeruli. Renal pathological changes and glomerular sclerosis index (GSI) were evaluated as well. RESULTS: At the end of the experiment, the mean urinary protein excretion in the model group was 305.4 mg/24 h, significantly higher than that in the treatment group (230.9 mg/24 h, P<0.01) and the sham operation group (5.6 mg/24 h, P<0.01) The serum urea nitrogen of the model group was (24.5 +/- 4.9) mmol/L, significantly higher than that of the treatment group [(15.8 +/- 3.9) mmol/L, P<0.05] and that of the sham-operated group (7.4 +/- 0.3 mmol/L, P<0.01). The serum creatinine (P<0.05) of the model group was 88 micromol/L +/- 14 micromol/L, significantly higher than that of the treatment group [(58 +/- 5) micromol/L, P<0.05)] and that of the sham-operated group [(54 +/- 5) micromol/L, P<0.05]. The creatinine clearance rate of the model group was (1.7 +/- 0.7) ml.min(-1).kg(-1), significantly lower than that of the treated group [(3.2 +/- 1.1) ml.min(-1).kg(-1), P<0.05] and that of the sham-operated group [(3.9 +/- 1.5) ml.min(-1).kg(-1), P<0.05]. The glomerular sclerosis index (GSI) in the model group was 41.8 +/- 11.5, significantly higher than that in the sham operation group (2.2 +/- 1.3, P<0.01) and the treatment group (23.4 +/- 6.1, P<0.05). The mean optical density of CTGF mRNA expression in the renal cortex of the model group was a 3 times that of the sham operation group, and the mean optical density of CTGF mRNA expression in the renal cortex of the treatment group was lower by 55.4% compared with that of the model group (P<0.01). The glomerular expressions of TGF-beta1, type IV collagen and fibronectin were significantly up-regulated in the model group in comparison with those in the sham operation group (all P<0.01). The glomerular protein expressions of TGF-beta1, type IV collagen and fibronectin were significantly weaker in the fluvastatin treatment group as compared with the model group (all P<0.01). CONCLUSION: CTGF mRNA expression is markedly upregulated in the renal cortex of 5/6 nephrectomized rats. Fluvastatin suppresses the increased CTGF mRNA expression in renal cortex and ameliorates the glomerular extracellular matrix accumulation.

Animals↗
PubMed1989

Congenital afibrinogenaemia.

A case of congenital afibrinogenaemia in a young female child is described. She had haemorrhagic tendency since birth in the form of markedly prolonged umbilical bleeding and easy bruising afterwards. Two of her brothers had bleeding tendencies, one died shortly after birth due to uncontrollable umbilical bleeding and other died at the age of 12 years from internal haemorrhage. The family study indicates the mode of inheritance to be probably autosomal recessive. The principal laboratory findings are complete non-coagulability of blood, grossly abnormal coagulation tests, zero ESR value, failure to detect fibrinogen by heat coagulation or chemical precipitation tests and biuret reaction and correction of thrombin time after fibrinogen infusion.

Afibrinogenemia↗
PubMed1989

An improved pyrogallol red-molybdate method for determining total urinary protein.

We adapted the pyrogallol red-molybdate method for total urinary protein to the Cobas Bio centrifugal analyzer. The method is simple, rapid, sensitive, and inexpensive. Addition of 25 mg of sodium dodecyl sulfate per liter to the reagent modifies protein reactivities so that the chromogenicity of human gamma globulins is the same as that of albumin. Results by this method and a comparison method that included gel filtration and a modified biuret reaction correlated well (r = 0.951).

Adult↗
PubMed1987

Influence of ambient temperature on the Ektachem DT60 results for total bilirubin and total protein.

We evaluated the Kodak Ektachem DT60 analyzer with the DTE module in a two-month clinical trial before its introduction in the Netherlands. At ambient temperatures that differed from that at which the DT60 was calibrated, aberrant behavior for total bilirubin and total protein assays was observed. At subnormal temperatures the former gave higher results than expected, and the latter, lower results. We found a similar effect for total protein determined by the manual biuret reaction at different temperatures, when results were calculated from calibration at a fixed temperature. Total bilirubin assayed by the manual diazobilirubin method showed no such effect. Although the DT60 analyzer is equipped with a temperature-regulated incubator, we conclude that the manufactures's recommended room temperature range for these assays should be narrowed and the three-month calibration period adjusted according to external circumstances.

Autoanalysis↗
PubMedDate not supplied

[Effect of cryopreservation of thyroid parenchyma on iodoamino acid composition of thyroglobulin and its iodination].

The content and distribution of iodoamino acids in thyroglobulin of the cryopreserved thyrotoxically-changed thyroid parenchyma were studied, as was thyroglobulin iodination. Thyroid tissue obtained during operations of patients with diffuse-toxic goiter was investigated. The thyroid parenchyma was cryopreserved according to the method developed at the Institute of Cryobiology and Cryomedicine Problems, Academy of Sciences of the Ukrainian SSR. The tissues were stored for 4-6 months. Thyroglobulin was isolated by gel filtration of the thyroid saline extract through a column packed with Sephadex G-200. Thyroglobulin was iodinated with KI + I2 water solution, pH 9.2, at 37 C for 30 min. The amount of iodine added was 100 moles of I2 per protein mole. Protein concentration was determined by the biuret reaction. Thyroglobulin iodoamino acid composition was determined by direct spectrophotometry. Absorption spectra were measured by an EPS-3T recording spectrophotometer ("Hitachi", Japan). The processes of freezing (-196 degrees C) and thawing of the thyroid parenchyma were shown to induce no changes in the thyroglobulin iodoamino acid composition. Cryopreservation of the thyroid parenchyma considerably affected iodine incorporation and formation of iodoamino acids in the thyroglobulin during its in vitro iodination. It may be supposed that cryopreservation of the thyroid tissue affects the thyroglobulin conformational status, that results in increased iodination of this iodoprotein.

Culture Media↗
PubMed1980

In vitro K+-effect on ATP and phosphocreatine levels and on Na+ K+-atpase activity of mouse brain cells.

Cell suspensions were prepared form mouse brain cortices. Cells were incubated in a medium also containing 10 mmol.l-1 glucose and either 5 mmol l-1 K+ or 50 mmol.l-1 K+ concentration. In order to standardize individual experiments, the biuret reaction was modified for rapid determination of the protein in cell suspensions. The cellular reserves of energy-rich phosphates were determined in the course of 60 min of cell incubation with 5 mmol.l-1 K+ and following 30 min incubation of cells with 50 mmol.l-1 K+. The level of ATP was significantly elevated after 10-60 min of incubation with low K+, from 0.58 to 0.78 micromoles per 100 mg protein; the creatine phosphate content during the same interval was in the range 1.27-1.44 micromoles per 100 mg protein. A significant decrease of energy reserves in cells was observed if the extracellular concentration of K+ was increased. After 10 and 30 min of incubation, a decrease by 36.2% and 38.5% for creatine phosphate and 34.6% and 44.9% for ATP was found, respectively. Na+ K+-ATPase activity of cells incubated for 60 min with 5 mmol.l-1 K+ was expressed as 4.99 micromoles of liberated Pi per 100 mg protein.1 h. Enzyme activity was stimulated with 50 mmol.l-1 K+ by 24.3% and 25.7% after 10 and 30 min of cell incubation respectively. Stimulation of Na+ K+-ATPase activity of brain cortex cells was directly dependent on the actual presence of stimulating 50 mmol.l-1 K+ concentration.

Adenosine Triphosphatases↗