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Biomedical subjects

A Safavi

Publications and source records attributed to A Safavi.

At least 19 recordsLinked to original sources
PubMed2005

Simultaneous spectrophotometric determination of Fe(III), Al(III) and Cu(II) by partial least-squares calibration method.

A spectrophotometric method for simultaneous determination of Fe(III), Al(III) and Cu(II) using Alizarin Red S as a chelating agent was developed. The parameters controlling behavior of the system were investigated and optimum conditions were selected. A partial least-squares multivariate calibration method was used for the analysis of ternary mixtures of Fe(III), Al(III) and Cu(II) over the range of 450-6000, 140-4000 and 450-15000 ng ml(-1), respectively. Absorbance data were taken between 400 and 800 nm. Applying this method to simultaneous determination of these metal ions in several synthetic alloy solutions with total relative standard error of less than 5% validated the proposed method.

Aluminum↗
PubMed2004

Flow-injection determination of isoniazid using sodium dichloroisocyanurate- and trichloroisocyanuric acid-luminol chemiluminescence systems.

A chemiluminescent (CL) method for the determination of isoniazid is described. The method is based on the CL generated during the oxidation of luminol by sodium dichloroisocyanurate (SDCC) and trichloroisocyanuric acid (TCCA) in alkaline medium. It was found that isoniazid greatly enhances this CL intensity when present in the luminol solution. Based on this observation, a new flow-injection CL method for the determination of isoniazid has been proposed in this paper. The detection limits were 2 and 3 ng ml(-1) isoniazid for the SDCC-luminol and TCCA-luminol CL systems, respectively. The relative CL intensity was linear with the isoniazid concentration in the range of 4-100 and 100-200 ng ml(-1) for the SDCC-luminol CL system, and 6-200 and 200-1000 ng ml(-1) for the TCCA-luminol CL system. The results obtained for the assay of pharmaceutical preparations compared well with those obtained by the official methods and demonstrated good accuracy and precision.

Chemistry Techniques, Analytical↗
PubMed2004

Cloud point extraction, preconcentration and simultaneous spectrophotometric determination of nickel and cobalt in water samples.

Cloud point extraction has been used for the preconcentration and simultaneous spectrophotometric determination of nickel and cobalt after the formation of a complex with 2-amino-cyclopentene-1-dithiocarboxylic acid (ACDA), and latter analysis by spectrophotometer using Triton X-114 as surfactant. The parameters affecting the separation phase and detection process were optimized. Under the optimum experimental conditions (i.e. pH=5, 0.07 mM ACDA, Triton X-114=0.25% (w/v)), calibration graphs were linear in the range of 20-500 and 20-200 microg l(-1) with detection limits of 10 and 7.5 microg l(-1) for Ni and Co, respectively. The method was applied to the determination of Ni and Co in natural and waste water samples with satisfactory results.

Carboxylic Acids↗
PubMed2004

Sensitive indirect spectrophotometric determination of isoniazid.

A simple, rapid, sensitive and accurate indirect spectrophotometric method for the microdetermination of isoniazid (INH) in pure form and pharmaceutical formulations is developed. The procedure is based on the reaction of copper(II) with isoniazid in the presence of neocuproine (NC). In the presence of neocuproine, copper(II) is reduced easily by isoniazid to a Cu(I)-neocuproine complex, which shows an absorption maximum at 454 nm. By measuring the absorbance of the complex at this wavelength, isoniazid can be determined in the range 0.3-3.5 microgml-1. This method was applied to the determination of isoniazid in pharmaceutical formulation and enabled the determination of the isoniazid in microgram quantities (0.3-3.5 microgml-1). The results obtained for the assay of pharmaceutical preparations compared well with those obtained by the official method and demonstrated good accuracy and precision.

Antitubercular Agents↗
PubMed2000

Kinetic spectrophotometric determination of traces of sulfide in nonionic micellar medium.

A sensitive kinetic spectrophotometric method for the determination of ng amounts of sulfide has been developed based on the reduction of Azure A by sulfide in the presence of Brij-35 at pH 7. The decrease in absorbance of Azure A at 600 nm is proportional to the concentration of sulfide over the range 25-1,400 ng mL(-1). The variables affecting the rate of the reaction were investigated and the optimum conditions were established. The method is simple, rapid, precise, sensitive, and widely applicable. The limit of detection is 17 ng mL(-1), and the relative standard deviation of seven determinations of 500 ng mL(-1) sulfide was 2.1%. The method was applied to the determination of sulfide in spring water.

Journal Article↗
PubMed1999

A catalytically active Jak2 is required for the angiotensin II-dependent activation of Fyn.

Recent work with interleukins has shown a convergence of tyrosine phosphorylation signal transduction cascades at the level of the Janus and Src families of tyrosine kinases. Here we demonstrate that activation of the seven-transmembrane AT(1) receptor by angiotensin II induces a physical association between Jak2 and Fyn, in vivo. This association requires the catalytic activity of Jak2 but not Fyn. Deletion studies indicate that the region of Jak2 that binds Fyn is located between amino acids 1 and 240. Studies of the Fyn SH2 and SH3 domains demonstrate that the SH2 domain plays the primary role in Jak2/Fyn association. Not surprisingly, this domain shows a marked preference for tyrosine-phosphorylated Jak2. Surface plasmon resonance estimated the dissociation equilibrium constant (K(d)) of this association to be 2.36 nM. Last, in vivo studies in vascular smooth muscle cells show that, in response to angiotensin II, Jak2 activation is required for Fyn activation and induction of the c-fos gene. The significance of these data is that Jak2, in addition to serving as a critical angiotensin II activated signal transduction kinase, also functions as a docking protein and participates in the activation of Fyn by providing phosphotyrosine residues that bind the SH2 domain of Fyn.

Angiotensin II↗
PubMed1998

Insulin-degrading enzyme regulates extracellular levels of amyloid beta-protein by degradation.

Excessive cerebral accumulation of the 42-residue amyloid beta-protein (Abeta) is an early and invariant step in the pathogenesis of Alzheimer's disease. Many studies have examined the cellular production of Abeta from its membrane-bound precursor, including the role of the presenilin proteins therein, but almost nothing is known about how Abeta is degraded and cleared following its secretion. We previously screened neuronal and nonneuronal cell lines for the production of proteases capable of degrading naturally secreted Abeta under biologically relevant conditions and concentrations. The major such protease identified was a metalloprotease released particularly by a microglial cell line, BV-2. We have now purified and characterized the protease and find that it is indistinguishable from insulin-degrading enzyme (IDE), a thiol metalloendopeptidase that degrades small peptides such as insulin, glucagon, and atrial natriuretic peptide. Degradation of both endogenous and synthetic Abeta at picomolar to nanomolar concentrations was completely inhibited by the competitive IDE substrate, insulin, and by two other IDE inhibitors. Immunodepletion of conditioned medium with an IDE antibody removed its Abeta-degrading activity. IDE was present in BV-2 cytosol, as expected, but was also released into the medium by intact, healthy cells. To confirm the extracellular occurrence of IDE in vivo, we identified intact IDE in human cerebrospinal fluid of both normal and Alzheimer subjects. In addition to its ability to degrade Abeta, IDE activity was unexpectedly found be associated with a time-dependent oligomerization of synthetic Abeta at physiological levels in the conditioned media of cultured cells; this process, which may be initiated by IDE-generated proteolytic fragments of Abeta, was prevented by three different IDE inhibitors. We conclude that a principal protease capable of down-regulating the levels of secreted Abeta extracellularly is IDE.

Alzheimer Disease↗
PubMed1998

Janus kinase 2 (Jak2) must be catalytically active to associate with the AT1 receptor in response to angiotensin II.

Angiotensin II evokes a variety of biological responses by binding to a seven transmembrane cell surface receptor termed AT1. Ligand binding to the AT1 receptor induces the physical association and activation of the intracellular kinase Jak2. To elucidate the mechanism of this association, COS-7 cells were co-transfected with the AT1 receptor and either wild type Jak2 or a catalytically inactive Jak2. AT1 receptor-Jak2 association was assessed in vitro by a GST-AT1 receptor fusion protein binding assay and in vivo by direct co-immunoprecipitation of the receptor-Jak2 complex. Both studies showed that Jak2 must be catalytically active to form a complex with the AT1 receptor, and that complex formation is associated with Jak2 tyrosine phosphorylation. These results were confirmed using the Jak2 specific inhibitor AG-490. We also found that over-expression of wild type Jak2 in COS-7 cells leads to in vivo complex formation of spontaneously autophosphorylated Jak2 with the AT1 receptor. No such complex formation was observed with a dominant negative Jak2. Thus, the physical association of Jak2 with the AT1 receptor is regulated by an angiotensin II mediated autophosphorylation event.

Angiotensin II↗
PubMed1997

Increased expression of an endopeptidase (gamma-EGE/IDE) hydrolyzing beta-endorphin during differentiation and maturation of bone marrow macrophages.

The presence and regulated expression of peptidase activity is a powerful mechanism with the potential to terminate or alter receptor recognition, cell membrane signal transduction, and physiological responses of immune cells to exogenous opioid peptides. In this study, the expression of an endopeptidase that hydrolyzes beta-endorphin to gamma-endorphin and other peptide products was investigated during in vitro differentiation and maturation of recombinant granulocyte-macrophage colony-stimulating factor (rGM-CSF) -derived, bone marrow-derived macrophages. In freshly isolated intact isolated mouse bone marrow cells the rate of beta-endorphin hydrolysis is undetectable (<0.1 nmol beta-endorphin hydrolyzed/h/10[6] cells). However, total intracellular beta-endorphin hydrolytic activity was increased significantly to 20.0 +/- 1.7 nmol/h/10(6) cells in the mature mouse macrophages derived in vitro by culture with rGM-CSF. rGM-CSF-derived macrophages expressed significantly higher levels of both protein and mRNA for the major beta-endorphin endopeptidase, gamma-endorphin-generating enzyme/insulin-degrading enzyme (gamma-EGE/IDE). Moreover, this enzymatic activity appears to be responsible for cleavage of exogenous beta-endorphin by intact rGM-CSF-derived macrophages or peritoneal macrophages to generate gamma-endorphin and other peptide products.

Animals↗
PubMed1996

Identification of gamma-endorphin-generating enzyme as insulin-degrading enzyme.

The EL-4 thymoma cell line contains a peptidase which converts beta-endorphin to beta-endorphin 1-17 (gamma-endorphin), beta-endorphin 1-18, and their corresponding C-terminal fragments. This enzyme was purified approximately 700-fold to a single band on an SDS-polyacrylamide gel (106 kDa) in 16% yield. Estimation of the native molecular weight by molecular sieve chromatography gave a value of approximately 220 kDa, indicating that this enzyme is a dimer. Peptide sequencing demonstrated this activity can be attributed to insulin degrading enzyme, a previously described member of the inverzincin family (Hooper, 1994). Kinetic studies with a number of peptide substrates indicate that the enzyme preferentially cleaves on the amino side of hydrophobic or basic residues. However, the substrate specificity is more complex since not all basic and hydrophobic residues in a peptide are cleaved. The enzyme exhibits a requirement for a P'2 residue. On the basis of kcat/K(m) values, insulin, growth hormone releasing factor, and beta-endorphin are nearly equivalent substrates for the enzyme; however, growth hormone releasing factor and beta-endorphin exhibit a 40-fold higher kcat, but a 10-fold decreased affinity relative to insulin. A role for insulin-degrading enzyme as both a beta-endorphin-processing and -inactivating enzyme is implicated from these studies.

Amino Acid Sequence↗
PubMed1996

Kinetic-spectrophotometric determination of sulfide by its reaction with resazurin.

A sensitive, rapid and selective method for the determination of sulfide based on its reaction with resazurin at pH 7 is described. The method is simple and has the great advantage of being free from interference of other sulfur anions and thus sulfide can be easily determined at trace levels in the presence of other sulfur compounds. The method has been applied to some synthetic samples and a spring water sample. The importance of sulfide compounds in biological processes has been widely demonstrated [1]. However, most of the methods used so far are either not sensitive enough or require complicated and expensive instruments; or are subjected to interferences from other sulfur compounds. Therefore, more sensitive, simple and selective methods are still required.

Journal Article↗
PubMed1996

Delayed primary repair of intrathoracic esophageal perforation: is it safe?

The management of intrathoracic esophageal perforation with delayed diagnosis is a subject of controversy. Because of the obvious advantages of primary repair as a simple single-stage operation, this technique was preferentially used to treat 18 of 22 consecutive patients with esophageal perforation. These patients were stratified into three groups according to the time interval between perforation and repair: group A, less than 6 hours, five patients (28%); group B, 6 to 24 hours, six patients (33%); and group C, more than 24 hours, seven patients (39%). Group A patients were older (p < 0.05) and group B had fewer iatrogenic perforations (B, 17%; A, 80%; C, 57%, p < 0.1). Additional tissue was used to buttress the repair site in all three groups (A, 3/5 patients, 60%; B, 4/6 patients, 67%; C, 6/7 patients, 86%; p = not significant). In seven patients (39%), a fundic wrap was used to reinforce the site of primary repair. The outcomes of the three groups were analyzed. Group A had the lowest proportion of postoperative leaks (A, 0/4 patients, 0%; B, 4/6 patients, 67%; C, 5/6 patients, 83%; p < 0.05) and postoperative morbidity (A, 2/5 patients, 40%; B, 6/6 patients, 100%; C, 6/7 patients, 86%; p < 0.1). However the increased incidence of leak and morbidity did not lead to an increase in mortality. One death occurred in each group, with an overall mortality of 17% (A, 1/5 patients, 20%; B, 1/6 patients, 17%; C, 1/7 patients, 14%; p = not significant). We conclude that in the era of advanced intensive care capabilities, primary repair of intrathoracic esophageal perforation can be safely accomplished in most patients regardless of the time interval between perforation and operation. Leakage at the suture site is common unless primary repair is carried out without delay. Postoperative leakage, however, is usually inconsequential and does not necessarily result in an adverse outcome.

Aged↗
PubMed1995

Purification and characterization of a secreted arginine-specific dibasic cleaving enzyme from EL-4 cells.

A secreted dibasic cleaving peptidase capable of converting dynorphins into Leu-enkephalin-Arg6 was purified from the medium of EL-4 mouse thymoma cells. The enzyme is a novel metalloendopeptidase with a neutral pH optimum (6.9) and a molecular weight of approximately 130 000. The dibasic cleaving enzyme was completely inhibited in the presence of 20-50 mM amine buffers, 0.1 mM EDTA, 0.5 mM 1,10-phenanthroline, 0.5 mM N-ethylmaleimide, and 1mM DTNB. Unlike the Kex2 family of proteases, Ca2+ did not activate the endopeptidase, but high concentrations (1 mM) of metal ions such as Cu2+, Ni2+, Zn2+, and Co2+ completely inhibited the enzyme. Inhibition was not seen with 0.2 mM TLCK, 1 mM DTT, and 1 mM PMSF. The enzyme will cleave Arg-Arg and Arg-Lys bonds, but not Lys-Arg or Lys-Lys bonds in identical environments, and no aminopeptidase or carboxypeptidase activity was seen. The size of the substrate does not seem to be a determining factor, since dynorphin A(1-12) is cleaved at a rate similar to prodynorphin B(228-256) containing 29 amino acids. The identity of the residues on either side of the cleavage site influences the rate of processing, as noted by different rates of cleavage for the same size peptides dynorphin A(1-13) vs dynorphin A(1-9) vs beta-neoendorphin. The presence of proline in the P3' (alpha-neoendorphin), P4' (dynorphin A(1-11)), or P5' (bovine adrenal medulla dodecapeptide) position does not prevent cleavage, but neurotensin and its (1-11) fragment containing both P2 and P2' proline residues are not cleaved.

Amino Acid Sequence↗
PubMed1995

Degradation of dynorphin-related peptides by the puromycin-sensitive aminopeptidase and aminopeptidase M.

The degradation of dynorphin-related peptides by the puromycin-sensitive aminopeptidase and aminopeptidase M was examined using these peptides as alternate substrate inhibitors. Ki determinations showed that both aminopeptidases exhibit a higher affinity for longer dynorphin-related peptides, i.e., Ki for dynorphin A-17 = 23-30 nM with the Ki increasing to 25-50 microM for the enkephalin pentapeptides. Binding appears dependent not only on peptide length, but also on its sequence. With aminopeptidase M, as the peptide size increases from five to 10 amino acids, kcat remains relatively constant; however, as the peptide size increases beyond a decapeptide, kcat decreases significantly. With the puromycin-sensitive aminopeptidase, similar results were obtained except that kcat was greatest for the pentapeptide. Thus, if one considers kcat/Km as the relevant kinetic constant for estimating in vivo peptide hydrolysis, these results are consistent with the involvement of aminopeptidase M and the puromycin-sensitive aminopeptidase in the degradation of extended dynorphin-related peptides.

Amino Acid Sequence↗
PubMed1995

One-stage primary repair of distal esophageal perforation using fundic wrap.

Esophageal leak following primary repair of esophageal perforation is a serious complication that can lead to severe mediastinitis and sepsis. Complete diversion with esophageal exclusion or resection is designed to minimize further mediastinal contamination. However, this approach is not necessarily associated with less morbidity or mortality. Furthermore, a second stage operation is required to restore esophageal continuity. From 1986 to 1994, we performed a one-stage primary repair of the distal esophagus in seven patients with either iatrogenic (n = 5) or spontaneous (n = 2) perforations and reinforced the repair by a fundic wrap. One patient underwent an additional modified Heller myotomy for achalasia. Delay between perforation and operation was less than 6 hours in 3 patients, 6 to 24 hours in 2 patients, and greater than 24 hours in 2 patients. Only one patient (14%) developed a small esophageal leak that spontaneously resolved with adequate mediastinal drainage, intravenous antibiotics, and aggressive nutritional support. One patient (14%), whose repair was delayed by 12 hours, died postoperatively of profound sepsis. This patient was moribund from sepsis preoperatively, and postmortem examination of the esophagus revealed no evidence of esophageal leak. Esophageal continuity was maintained in all patients. The median length of stay was 21 days (range, 15-58 days). We conclude that primary reinforced repair of esophageal perforation using a fundic wrap is an effective method of treatment for distal esophageal perforation, even when the repair is delayed by more than 24 hours.

Adult↗