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Biomedical subjects

E B Hansen

Publications and source records attributed to E B Hansen.

At least 19 recordsLinked to original sources
PubMed2001

Cementless porous-coated total knee arthroplasty: 10-year results in a consecutive series.

We report the results of 114 AGC 2000 porous-coated, cementless total knee arthroplasties (TKA) performed consecutively in 102 patients during the period 1984-1986. After 10 years, 58 TKAs in 52 patients were evaluated with patient assessment, Hospital for Special Surgery knee score, weight-bearing radiographs done under fluoroscopic control, and survivorship analysis. All dropouts within the first 9 years were patients dying with a functioning TKA except 1 revision secondary to a supracondylar fracture after 8.5 years. Of the patients, 53 (92%) were satisfied or very satisfied with their TKA, and 55 (95%) of the knees were rated good or excellent. There was no pain in 53 knees, and the median knee flexion was 110 degrees. Six radiolucencies >1 mm were found beneath parts of the tibial component, and 5 radiolucencies were seen beneath the femoral component. None had progressed compared with the 5-year follow-up, and in all cases trabeculae could be seen reaching the prosthetic component. No migrations had occurred since the 5-year follow-up. No obvious joint space reduction was seen. Osteolysis presenting as an isolated cyst was found in 1 knee in the lateral tibial condyle and was not progressive. Two tibial components had been revised because of aseptic loosening and 1 because of septic loosening, all within the first 3 years. No femoral or patellar components were revised. The cumulative prosthesis survival rate after 10 to 11 years was 97%. When pain and radiographic loosening also were considered, the success rate was 87%. Cementless insertion of a nonmodular, porous-coated TKA resulted in a long-term durable bone-prosthesis interface. The flat-on-flat articulation did not result in catastrophic polyethylene wear or osteolysis within the first 10 years.

Arthroplasty, Replacement, Knee↗
PubMed2000

Analysis of erythromycin by liquid chromatography/mass spectrometry using involatile mobile phases with a novel atmospheric pressure ionization source.

A critical limitation of electrospray ionization (ESI) liquid chromatography/mass spectrometry (LC/MS) sources is the susceptibility to blockage of interface orifices due to the deposition of involatile components from the sample and/or mobile phase. These components, including salts, buffers, and ion-pairing agents, can be essential to the performance of the chosen analytical method. We report here the performance enhancements provided by a novel atmospheric pressure ionization (API) source in the analysis of erythromycin A (ERY) using mobile phases that contain involatile components. The enhanced robustness of the new source is derived from the use of a continuous flow of aqueous solvent at the sampling cone orifice that maintains unobstructed ion transmission. The ESI mass spectral responses measured for ERY, using an LC separation that incorporates 10 mM sodium phosphate with and without 10 mM octane sulfonate, were monitored by repeated injections over 13-15 h total analysis time. Minimal effects on ESI mass spectral responses (integrated peak area) or chromatographic performance (peak shape, retention time) were observed during these studies. In the absence of the aqueous cleaning flow, complete loss of mass spectral responses and total blocking of the sampling cone was observed in less than 30 min. Responses for ERY spiked into chicken and beef liver, and catfish muscle at or below the regulatory level of interest (100 ppb), were quantified by internal standard calibration using this procedure. These results demonstrate the ability of a novel API-MS ion source to perform analyses that require the use of involatile mobile phase additives.

Animals↗
PubMed2000

Metabolism of the ethanolamine-type antihistamine diphenhydramine (Benadryl) by the fungus Cunninghamella elegans.

Two strains of the filamentous fungus Cunninghamella elegans (ATCC 9245 and ATCC 36112) were grown in Sabouraud dextrose broth and screened for the ability to metabolize the ethanolamine-type antihistamine diphenhydramine. Based on the amount of parent drug recovered after 7 days incubation, both C. elegans strains metabolized approximately 74% of the diphenhydramine, 58% of this being identified as organic extractable metabolites. The organic extractable metabolites were isolated by reversed-phase high-performance liquid chromatography and identified by analyzing their mass and nuclear magnetic resonance spectra. Desorption chemical ionization mass spectrometry (DCIMS) with deuterated ammonia was used to differentiate possible isobaric diphenhydramine metabolites and to probe the mechanisms of ion formation under ammonia DCIMS conditions. C. elegans transformed diphenhydramine by demethylation, oxidation, and N-acetylation. The major metabolites observed were diphenhydramine-N-oxide (3%), N-desmethyldiphenhydramine (30%), N-acetyldidesmethyldiphenhydramine (13%), and N-acetyl-N-desmethyldiphenhydramine (12%). These compounds are known mammalian metabolites of diphenhydramine and may be useful for further toxicological studies.

Chromatography, High Pressure Liquid↗
PubMed1998

Nonaqueous capillary electrophoretic separation and thermo-optical absorbance detection of five tricyclic antidepressants and metabolism of amitriptyline by Cunninghamella elegans.

We developed a technique based on nonaqueous capillary electrophoresis and laser-based thermo-optical absorbance detection to assay five antidepressants with similar structures and mass-to-charge ratios. A mixture of methanol and acetonitrile with ammonium acetate was essential to achieve baseline resolution of these compounds. We investigated the effects of ammonium acetate concentration, temperature, applied voltage, and capillary length on separation efficiency. The nonaqueous capillary electrophoresis and laser-based thermo-optical absorbance detection technique was used to study the metabolism of amitriptyline by Cunninghamella elegans. Sample preparation procedures were simplified for fast screening of the parent drug and its metabolites. Reproducible electropherograms were obtained from replicate cultures of C. elegans growing in the presence of amitriptyline.

Amitriptyline↗
PubMed1998

On-line nonaqueous capillary electrophoresis and electrospray mass spectrometry of tricyclic antidepressants and metabolic profiling of amitriptyline by Cunninghamella elegans.

An on-line nonaqueous capillary electrophoresis-electrospray mass spectrometry (ESI-MS) technique was developed using a commercial ion spray interface. The nonaqueous capillary electrophoresis ESI-MS system was used to profile tricyclic antidepressants of similar structures and mass-to-charge ratios. We found that pure methanol can be used as a sheath liquid to obtain stable ion spray from nonaqueous capillary electrophoresis. The flow rate of the coaxial nebulizing gas affected baseline signals, separation efficiency, and migration times. Other nonaqueous capillary electrophoresis operating conditions and electrospray parameters were optimized for enhanced baseline separation and high sensitivity detection. The effect of sample stacking on separation and detection was evaluated. The calculated detection limits were approximately 3 pg injected onto the capillary. ESI mass spectra of tricyclic antidepressants from a single quadrupole MS were obtained and elucidated. The information was used to propose fragmentation pathways of the tricyclic antidepressants. The method was also used to analyze the metabolites of amitriptyline produced by the fungus Cunninghamella elegans. Sixteen metabolites were detected and most of them were tentatively identified as demethylated and/or hydroxylated, and/or N-oxidized products.

Amitriptyline↗
PubMed1998

Isolation of nitrofurantoin-resistant mutants of nitroreductase-producing Clostridium sp. strains from the human intestinal tract.

Five spontaneous nitrofurantoin-resistant mutants (one each of Clostridium leptum, Clostridium paraputrificum, two other Clostridium spp. strains from the human intestinal microflora, and Clostridium perfringens ATCC 3626) were selected by growth on a nitrofurantoin-containing medium. All of the Clostridium wild-type and mutant strains produced nitroreductase, as was shown by the conversion of 4-nitrobenzoic acid to 4-aminobenzoic acid. High-performance liquid chromatography (HPLC) analysis of the mutants during incubation with 50 microg of nitrofurantoin per ml showed the gradual disappearance of the nitrofurantoin peak. The nitrofurantoin peak also disappeared when cell-free supernatants instead of cultures of each of the resistant and wild-type bacteria were used, but it persisted if the cell-free supernatants had been inactivated by heat. At least two of the mutants converted nitrofurantoin to metabolites without antibacterial activity, as was shown by a bioassay with a nitrofurantoin-susceptible Bacillus sp. strain. Nitrofurantoin at a high concentration (50 microg/ml) continued to exert some toxicity, even on the resistant strains, as was evident from the longer lag phases. This study indicates that Clostridium strains can develop resistance to nitrofurantoin while retaining the ability to produce nitroreductase; the mutants metabolized nitrofurantoin to compounds without antibacterial activity.

Anti-Infective Agents, Urinary↗
PubMed1997

Confirmation of malachite green, gentian violet and their leuco analogs in catfish and trout tissue by high-performance liquid chromatography utilizing electrochemistry with ultraviolet-visible diode array detection and fluorescence detection.

A sensitive analytical procedure for the confirmation of residues of malachite green (MG), gentian violet (GV) and their leuco analogs (LMG and LGV) in catfish and trout tissue at 10 ng/g is described. Frozen (-20 degrees C) fish fillets were cut into small pieces and homogenized in Waring blendors. The compounds of interest were extracted from 20-g amounts of homogenized fish tissue with acetonitrile-buffer, partitioned against methylene chloride, and isolated with tandem neutral alumina and propylsulfonic acid cation-exchange solid-phase extraction cartridges. Samples of 100 microl (0.8 g equiv.) were chromatographed isocratically in 10 min using an acetonitrile-buffer mobile phase on a short-chain deactivated (SCD) reversed-phase column (150x4.6 mm I.D.) in-line with a post-column oxidation coulometric electrochemical cell (EC), a UV-Vis diode array detector and a fluorescence detector.

Animals↗
PubMed1997

Fungal transformations of antihistamines: metabolism of cyproheptadine hydrochloride by Cunninghamella elegans.

1. Metabolites formed during incubation of the antihistamine cyproheptadine hydrochloride with the zygomycete fungus Cunninghamella elegans in liquid culture were determined. The metabolites were isolated by hple and identified by mass spectrometric and proton nmr spectroscopic analysis. Two C elegans strains, ATCC 9245 and ATCC 36112, were screened and both produced essentially identical metabolites. 2. Within 72 h cyproheptadine was extensively biotransformed to at least eight oxidative phase-I metabolites primarily via aromatic hydroxylation metabolic pathways. Cyproheptadine was biotransformed predominantly to 2-hydroxycyproheptadine. Other metabolites identified were 1- and 3-hydroxycyproheptadine, cyproheptadine 10,11-epoxide, N-desmethylcyproheptadine, N-desmethyl-2-hydroxycyproheptadine, cyproheptadine N-oxide, and 2-hydroxycyproheptadine N-oxide. Although a minor fungal metabolite, cyproheptadine 10,11-epoxide represents the first stable epoxide isolated from the microbial biotransformation of drugs. 3. The enzymatic mechanism for the formation of the major fungal metabolite, 2-hydroxycyproheptadine, was investigated. The oxygen atom was derived from molecular oxygen as determined from 18O-labelling experiments. The formation of 2-hydroxycyproheptadine was inhibited 35, 70 and 97% by cytochrome P450 inhibitors metyrapone, proadifen and 1-aminobenzotriazole respectively. Cytochrome P450 was detected in the microsomal fractions of C. elegans. In addition, 2-hydroxylase activity was found in cell-free extracts of C. elegans. This activity was inhibited by proadifen and CO, and was inducible by naphthalene. These results are consistent with the fungal epoxidation and hydroxylation reactions being catalysed by cytochrome P450 monooxygenases. 4. The effects of types of media on the biotransformation of cyproheptadine were investigated. It appears that the glucose level significantly affects the biotransformation rates of cyproheptadine; however it did not change the relative ratios between metabolites produced.

Biotransformation↗
PubMed1996

Cementless total knee arthroplasty in rheumatoid arthritis. A report on 51 AGC knees followed for 54 months.

Fifty-one primary consecutive cementless AGC 2000 (Anatomically Graduated Components, Biomet, Warsaw, IN) total knee arthroplasties were performed during 1985 through 1990 in 40 patients with rheumatoid arthritis. Forty-one knees (32 patients) were available for clinical and radiologic follow-up analysis after 24 to 76 months (median, 54 months). There was no pain in 33 knees and mild pain in the rest. Median range of motion was 110 degrees (range, 50 degrees-130 degrees). Median knee score was 90 (range, 71-97), and all knees were rated good or excellent. Radiolucencies greater than 1 mm were found under five tibial components, but no obvious migrations were seen. One tibial component was revised due to aseptic loosening. The cumulative success rate after 4 to 5 years was 97% (lower limit of 95% confidence interval, 91.8%). The medium-term results are considered to compare favorably with reported cemented series.

Adult↗
PubMed1996

Fungal biotransformation of the antihistamine azatadine by Cunninghamella elegans.

The metabolism of the antihistamine azatadine by the zygomycete fungus Cunninghamella elegans ATCC 9245 was investigated. Within 72 h from the addition of the drug to 48-h-old cultures grown in Sabouraud dextrose broth, 95% of azatadine was biotransformed. Two major metabolites, 7-hydroxyazatadine (25%) and 8-hydroxyazatadine (50%), and two minor metabolites, N-desmethylazatadine and 9-hydroxyazatadine, were isolated by high-performance liquid chromatography and characterized by mass spectrometric and proton nuclear magnetic resonance spectroscopic analyses.

Biotransformation↗
PubMed1996

Cloning and transcriptional analysis of two threonine biosynthetic genes from Lactococcus lactis MG1614.

Two genes, hom and thrB, involved in threonine biosynthesis in Lactococcus lactis MG1614, were cloned and sequenced. These genes, which encode homoserine dehydrogenase and homoserine kinase, were initially identified by the homology of their gene products with known homoserine dehydrogenases and homoserine kinases from other organisms. The identification was supported by construction of a mutant containing a deletion in hom and thrB that was unable to grow in a defined medium lacking threonine. Transcriptional analysis showed that the two genes were located in a bicistronic operon with the order 5' hom-thrB 3' and that transcription started 66 bp upstream of the translational start codon of the hom gene. A putative -10 promoter region (TATAAT) was located 6 bp upstream of the transcriptional start point, but no putative -35 region was identified. A DNA fragment covering 155 bp upstream of the hom translational start site was functional in pAK80, an L. lactis promoter probe vector. In addition, transcriptional studies showed no threonine-dependent regulation of hom-thrB transcription.

Amino Acid Sequence↗
PubMed1995

High-performance liquid chromatographic analysis of the antituberculosis drugs aconiazide and isoniazid.

Reversed-phase HPLC methods are described for determining the stability and concentration certification of the antituberculosis prodrug aconiazide (ACON) in aqueous dosing solution and for assessing the concentrations of ACON and isoniazid (INH) in plasma from ACON-treated male and female Fischer-344 rats. ACON was analyzed in plasma by direct injection; it was separated on a 250 x 4.6 mm I.D. 5 microns C18 column using a 40% aqueous methanol mobile phase containing 5 g/l ammonium formate, and detected at 313 nm. INH was determined in the plasma of treated rats after a two-step precipitation of plasma proteins; it was separated on a 250 mm x 4.6 mm I.D. 5 microns CN column, eluted with 5% aqueous isopropanol containing 5 g/l ammonium formate, and detected with an electrochemical detector at +0.8 V. These methods allow a simple, rapid, and reliable determination of ACON and INH in plasma down to 0.1 micrograms/ml.

Animals↗
PubMed1995

Isolation of Lactococcus lactis nonsense suppressors and construction of a food-grade cloning vector.

Nonsense suppressor strains of Lactococcus lactis were isolated using plasmids containing nonsense mutations or as revertants of a nonsense auxotrophic mutant. The nonsense suppressor gene was cloned from two suppressor strains and the DNA sequence determined. One suppressor is an ochre suppressor with an altered tRNA(gln) and the other an amber suppressor with an altered tRNA(ser). The nonsense suppressors allowed isolation of nonsense mutants of a lytic bacteriophage and suppressible auxotrophic mutants of L. lactis MG1363. A food-grade cloning vector based totally on DNA from Lactococcus and a synthetic polylinker with 11 unique restriction sites was constructed using the ochre suppressor as a selectable marker. Selection, following electroporation of a suppressible purine auxotroph, can be done on purine-free medium. The pepN gene from L. lactis Wg2 was subcloned resulting in a food-grade plasmid giving a four- to fivefold increase in lysine aminopeptidase activity.

Alleles↗
PubMed1995

Cloning and partial characterization of regulated promoters from Lactococcus lactis Tn917-lacZ integrants with the new promoter probe vector, pAK80.

Transposon Tn917-LTV1 was used to produce a collection of Lactococcus lactis strains with fusion of a promoterless lacZ gene to chromosomal loci. Screening 2,500 Tn917-LTV1 integrants revealed 222 that express beta-galactosidase on plates at 30 degrees C. Pulsed-field gel electrophoresis revealed Tn917-LTV1 insertions in at least 13 loci in 15 strains analyzed. Integrants in which beta-galactosidase expression was regulated by temperature or pH and/or arginine concentration were isolated. In most cases, the regulation observed on plates was reproducible in liquid medium. One integrant, PA170, produces beta-galactosidase at pH 5.2 but not at pH 7.0, produces more beta-galactosidase at 15 degrees C than at 30 degrees C, and has increased beta-galactosidase activity in the stationary phase. DNA fragments potentially carrying promoters from selected Lactococcus lactis integrants were cloned in Escherichia coli. A new promoter probe vector, pAK80, containing promoterless beta-galactosidase genes from Leuconostoc mesenteroides subsp. cremoris and the Lactococcus lactis subsp. lactis biovar diacetylactis citrate plasmid replication region was constructed, and the lactococcal fragments were inserted. Plasmid pAK80 was capable of detecting and discriminating even weak promoters in Lactococcus lactis. When inserted in pAK80, the promoter cloned from PA170 displayed a regulated expression of beta-galactosidase analogous to the regulation observed in PA170.

Amino Acid Sequence↗
PubMed1995

Fungal transformations of antihistamines: metabolism of brompheniramine, chlorpheniramine, and pheniramine to N-oxide and N-demethylated metabolites by the fungus Cunninghamella elegans.

1. Two strains of the filamentous fungus Cunninghamella elegans (ATCC 9245 and ATCC 36112) were screened for their ability to metabolize three alkylamine-type antihistamines; brompheniramine, chlorpheniramine and pheniramine. 2. Based on the amount of parent drug recovered after 168 h of incubation, C. elegans ATCC 9245 metabolized 60, 45 and 29% of brompheniramine, chlorpheniramine and pheniramine added respectively. The results from strain ATCC 36112 were essentially identical to those of strain ATCC 9245. 3. The metabolic products of N-oxidation and N-demethylation were isolated by reversed-phase hplc and identified by analysing their mass and proton nmr spectra. For all three antihistamines, the mono-N-demethylated metabolite was produced in the greatest amounts. The chloro- and bromo-substituents appeared not to affect the route of metabolism but did influence the relative amounts of metabolites produced. 4. Circular dichroism spectra of the metabolites and the unmetabolized parent antihistamines showed each to be a racemic mixture of the (+) and (-) optical isomers. In addition, comparison of the metabolism of racemic chlorpheniramine to that of optically pure (+) chlorpheniramine showed no significant differences in the ratios of metabolites produced. There was therefore no metabolic stereoselectivity observed by the fungal enzymes.

Biotransformation↗