Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0021051 (immunodeficiency)
 71,517 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Phosphorothioate oligodeoxynucleotides (S-ODNs) have potential as anti-viral agents and are being investigated for the chemotherapy of AIDS. A high-performance liquid chromatographic method is described for the analysis, in urine and plasma, of a 28-unit deoxycytidine homopolymer (S-dC28) and a 28-unit S-ODN "antisense" to the rev gene of the human immunodeficiency virus. This method employs ion-pairing HPLC with a polymeric column. Tetrabutylammonium is used as the ion-pairing agent in a mobile phase of acetonitrile in pH 7.0 phosphate buffer. Analysis of the S-ODNs is relatively rapid (20 min) and sensitive (20 nm) and is accomplished by a gradient elution (22.5-30.0% acetonitrile) followed by ultraviolet (266 or 272 nm) absorption detection. This method is likely applicable, with appropriate modifications, to all S-ODNs of similar molecular weight regardless of sequence. The S-ODNs bind very strongly to plasma proteins but are readily prepared for analysis by a phenol extraction procedure. In a preliminary pharmacokinetic study in mice with S-dC28, very rapid elimination of the oligomer from plasma was observed (half-time, 11.6 min). Estimates for the apparent volume of distribution and total body clearance were 3 ml and 0.2 ml/min, respectively. It appears that the majority of the oligomer is eliminated by renal clearance (glomerular filtration), a property likely shared by all S-ODNs of similar molecular mass.
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PMID:High-performance liquid chromatographic analysis of phosphorothioate analogues of oligodeoxynucleotides in biological fluids. 208 59

We describe here a one step HPLC technique for purifying the four gag proteins (p26, p15, p11 and p9) and two env glycoproteins (gp90 and gp45) from purified equine infectious anemia virus (EIAV), a member of the lentivirus subfamily of retroviruses. The purification procedure employs a reverse-phase phenyl Radial-pak cartridge contained in a high pressure radial compression chamber in which a shallow, multistep acetonitrile gradient is applied at ambient temperatures. The purified proteins are recovered at an efficiency of 60-70%. Moreover, the isolated components retain their antigenicity and are suitable for a variety of biochemical analyses including protein sequencing. The purification of EIAV gp90 and gp45 represents the first successful isolation of a lentivirus glycoprotein from purified virus preparations. The availability of these separated proteins permitted direct protein sequencing which confirmed the previously reported env gene sequence and provides important antigens for the development of diagnostic immunoassays and subunit vaccines. The procedures described appear applicable to other lentiviruses, including human immunodeficiency virus (HIV), and perhaps to hydrophobic membrane proteins in general.
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PMID:Lentivirus antigen purification and characterization: isolation of equine infectious anemia virus gag and env proteins in one step by reverse phase HPLC and application to human immunodeficiency virus glycoproteins. 283 62

[D-Ala1]peptide T amide is a metabolically stable and more potent analogue of peptide T, a proposed inhibitor of human immunodeficiency viral infectivity of human T-cell lymphocytes. The peptide was synthesized by solid-phase methods to provide amounts of several grams. The product was purified by chromatography on a 25 cm x 2 in. column of DuPont Zorbax Pro-10 C8 (10 micron) packing. Sample loads of 100-450 mg were chromatographed isocratically in 0.1% trifluoroacetic acid and 5% acetonitrile at a flow-rate of 110 ml/min. Under these conditions, the pure peptide fraction was eluted reproducibly between 15 and 22 min. After solvent removal and lyophilization, the recovery of pure peptide was 50% by weight.
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PMID:Reversed-phase preparative chromatography of [D-Ala1]-peptide T amide. 320 39

A sensitive high-performance liquid chromatography (HPLC) assay has been developed to simultaneously determine levels of the anti human immunodeficiency virus agent, zidovudine (AZT), and its major metabolite (the 5'-O-glucuronide) in serum. Samples were first mixed with an internal standard (a stereoisomer of AZT), then prepared for analysis using solid-phase extraction columns and chromatographed using a reversed-phase analytical column. Isocratic elution with a mobile phase of 15% acetonitrile, buffered to pH 2.70 with ammonium phosphate, gave good resolution of the three analytes and endogenous serum components. The HPLC analysis time required per sample was 34 min and analyte recoveries were reproducibly high (greater than 93%). Replicate analyses of prepared standards gave satisfactory precision and accuracy, with coefficients of variation less than 15% and deviations from expected concentrations less than 10%.
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PMID:Simultaneous quantification of zidovudine and its glucuronide in serum by high-performance liquid chromatography. 323 23

The National Cancer Institute is pursuing preclinical development of michellamine B (MB), a novel dimeric polyhydroxylated naphthalene-tetrahydroisoquinoline alkaloid isolated from Ancistrocladus abbreviatus, as an anti-human immunodeficiency virus (HIV) agent. MB protects human lymphoid cells from the cytopathic effects of both HIV-1 and HIV-2 in vitro. A specific, sensitive, and convenient method for assaying the compound in biological fluids has been developed. Samples were prepared for analysis by initial treatment with dilute trichloroacetic acid followed by thorough mixing with a solution of the internal standard (alpha-naphthoflavone) in acetonitrile to denature macromolecules. The supernatant afforded by centrifugation, upon dilution with the aqueous component of the liquid chromatographic eluent, was loaded onto a 4-microns Nova-Pak phenyl column (3.9 mm x 15 cm). Chromatography was performed at ambient temperature using an isocratic mobile phase composed of 10 mM octyl sodium sulfate and 15 microM tetrabutylammonium hydrogen sulfate in acetonitrile/0.05 M ammonium formate buffer, pH 4.0 (46/54, v/v), at a flow rate of 0.6 ml/min. The intense native fluorescence of MB, which exhibited excitation and emission maxima in the mobile phase at 232 and 393 nm, respectively, provided a highly sensitive and selective means of detection. Mean values of the retention times for the drug and internal standard determined over 11 months were 10.71 +/- 0.53 and 13.14 +/- 0.52 min, respectively (SD, n = 52). Employing a sample volume of 50 microliters, the lowest concentration of MB included in the standard curves of mouse, dog, and human plasma, 10 ng/ml (11.4 nM), was quantified with coefficients of variation less than 10%.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Determination of michellamine B in biological fluids by high-performance liquid chromatography with fluorescence detection. 813 66

Proteolytic experiments in conjunction with 1H-NMR spectroscopy show that the Nef (negative factor) protein from human immunodeficiency virus type 1 probably consists of two main domains, the N-terminal anchor domain at amino acid positions 2-65 and the C-terminal core domain at positions 66-206. The N-terminal domain is likely to be located at the surface of the protein, while the C-terminal domain has a compactly folded core and is stable in the absence of the anchor domain. It is conceivable that the core domain represents a functional domain of the Nef protein, activated after the removal of the membrane anchor by the human-immunodeficiency-virus protease or cellular proteases. Nef is stable at pH 5-12 and denatures at 317-322 K. The Nef protein remains in its native conformation in dimethyl-sulfoxide/water mixtures up to 35% (by vol.), and in acetonitrile/water up to 14% (by vol.). Nef refolds spontaneously after denaturation with urea or guanidinium hydrochloride. The 1H-NMR parameters and pKa values of five of the nine histidine residues and one of the seven tyrosine residues were determined and were found in four cases to be typical for residues which are not located in the interior of the protein.
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PMID:Stability and proteolytic domains of Nef protein from human immunodeficiency virus (HIV) type 1. 817 61

The dissociation of dimeric reverse transcriptase (RT) of the human immunodeficiency virus (HIV) types 1 and 2 has been investigated using acetonitrile as a dissociating agent. The equilibrium transitions were monitored by combining different approaches (fluorescence spectroscopy, polymerase activity assay, and size-exclusion HPLC). The dissociation of RT induced a complete loss of polymerase activity and a 25% increase of the intrinsic fluorescence. It is fully reversible, and the midpoints of the equilibrium transition curves are dependent on the concentration of the enzyme used, suggesting a two-state transition model for the dissociation of RT in which dimers are in equilibrium with folded monomers. For both RTs, the heterodimeric form is more stable against dissociating agents and different pH than the corresponding homodimeric form. Moreover, heterodimeric HIV-2 RT exhibits a higher stability than HIV-1 RT, with a free energy of dissociation of 12.1 kcal/mol at pH 6.5 and 25 degrees C, instead of 10 kcal/mol for HIV-1 RT. The binding of a primer/template induces a marked conformational change in both RTs, shown by the lower accessibility of the tryptophans to quenchers and the increase in tryptophan heterogeneity, and stabilized the dimeric form of both RTs (10-100-fold). The central role of hydrophobic interactions in dimer formation has been revealed by the 30% increase of exposure of the tryptophan cluster to quenchers upon dissociation of RT and the binding of 4 equiv of 1-anilino-8-naphthalenesulfonate to the dissociated enzymes.
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PMID:Conformational stability of dimeric HIV-1 and HIV-2 reverse transcriptases. 884 59

A rapid, sensitive and specific high-performance liquid chromatography (HPLC) procedure for the quantification of indinavir, a potent human immunodeficiency virus (HIV) protease inhibitor, in human plasma is described. Following C18 solid-phase extraction, indinavir was chromatographed on a reversed-phase C8 column using a simple binary mobile phase of phosphate buffer-acetonitrile (60:40, v/v). UV detection at 210 nm led to an adequate sensitivity without interference from endogenous matrix components. The limit of quantification was 25 ng/ml with a 0.1 ml plasma sample. The standard curve was linear across the range from 25 to 2500 ng/ml with an average recovery of 91.4%. The mean relative standard deviations for concentrations within the standard curve ranged between 1.4 and 9.7%. Quality control standards gave satisfactory intra- and inter-assay precision (R.S.D. from 3.5 to 15.8%) and accuracy within 15% of the nominal concentration. Sample handling experiments, including HIV heat inactivation, demonstrated analyte stability under expected handling processes. The assay is suitable for the analysis of samples from adult and pediatric patients infected with HIV.
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PMID:Rapid quantification of indinavir in human plasma by high-performance liquid chromatography with ultraviolet detection. 1005 96

Efavirenz (EFV, DMP-266) is a new antiretroviral agent belonging to the class of nonnucleoside reverse transcriptase inhibitors. It has recently been approved by the Food and Drug Administration in management of human immunodeficiency virus (HIV). Preliminary pharmacokinetic studies on EFV in healthy volunteers show that the drug may influence the metabolism of protease inhibitors. For the determination of EFV in human plasma, a validated and specific reverse-phase high-performance liquid chromatography (HPLC) method, with UV detection, was developed. We used 100 microL plasma sample for a liquid-liquid extraction with diethyl ether after basification. The mobile phase was a mixture of acetonitrile and water, pumped at a flow rate of 1.2 mL/min. Ultraviolet detection was carried out at a wavelength of 247 nm. Retention times for EFV and internal standard (IS) were 5.3 and 4.5 minutes, respectively, and there was no chromatographic interference from other commonly administered drugs. The limit of detection was 100 ng/mL. The described assay is a rapid and accurate method for measurement of EFV in plasma: the easy preparation and small sample size makes this assay highly suitable for pharmacokinetic studies and routine clinical analysis in patients with HIV. In addition, the reproducibility of the method is only moderately increased by including IS, so analyzing without IS may be an alternative.
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PMID:High-performance liquid chromatography method for analyzing the antiretroviral agent efavirenz in human plasma. 1036 51

The goal of this study was to develop a fast, inexpensive and quantitative method for serum determination of the human immunodeficiency virus protease inhibitors Crixivan (C), Viracept (V), Invirase (I) or Fortovase (F), and Norvir (N), using common conditions for isolation and analysis. The best separation procedure developed thus far involves uncoated silica capillary and a buffer containing formic acid and acetonitrile. This procedure allows us to analyze three drugs (C, V and I or F) in 15 min. Norvir requires different analytical conditions. These four drugs are isolated from patient sera with a mixture of ethyl acetate and hexane. Sensitivity of the capillary zone electrophoresis protocols is sufficient for the detection of these pharmacological agents at the lowest clinically relevant concentrations (0.1 microgram/ml).
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PMID:Determination of human immunodeficiency virus-1 protease inhibitors in patient serum using free solution capillary zone electrophoresis. 1048 50


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