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Biomedical subjects

R L Bertholf

Publications and source records attributed to R L Bertholf.

33 records · Page 2Linked to original sources

Serum and lymphocyte, aluminum and nickel in chronic renal failure.

In the past decade, aluminum has been recognized as a toxic metal in patients with chronic renal failure. It is, however, possible that other trace metals, such as nickel, may also have toxic actions in these patients. The plasma concentration of a metal, such as aluminum or nickel, may not provide a valid index of either tissue content or total body burden. In the study reported here, the aluminum and nickel content of lymphocytes was measured and compared with plasma concentrations in normal controls and patients with chronic renal failure. The findings suggest that lymphocytes may be of value as a nucleated 'tissue' for the assessment of trace metal status.

Adult↗

Aluminium toxicity in chronic renal insufficiency.

Aluminium is a ubiquitous element in the environment and has been demonstrated to be toxic, especially in individuals with impaired renal function. Not much is known about the biochemistry of aluminium and the mechanisms of its toxic effects. Most of the interest in aluminium has been in the clinical setting of the haemodialysis unit. Here aluminium toxicity occurs due to contamination of dialysis solutions, and treatment of the patients with aluminium-containing phosphate binding gels. Aluminium has been shown to be the major contributor to the dialysis encephalopathy syndrome and an osteomalacic component of dialysis osteodystrophy. Other clinical disturbances associated with aluminium toxicity are a microcytic anaemia and metastatic extraskeletal calcification. Aluminium overload can be treated effectively by chelation therapy with desferrioxamine and haemodialysis. Aluminium is readily transferred from the dialysate to the patient's bloodstream during haemodialysis. Once transferred, the aluminium is tightly bound to non-dialysable plasma constituents. Very low concentrations of dialysate aluminium in the range of 10-15 micrograms/l are recommended to guard against toxic effects. Very few studies have been directed towards the separation of the various plasma species which bind aluminium. Gel filtration chromatography has been used to identify five major fractions, one of which is of low molecular weight and the others appear to be protein-aluminium complexes. Recommendations on aluminium monitoring have been published and provide 'safe' and toxic concentrations. Also, the frequency of monitoring has been addressed. Major problems exist with the analytical methods for measuring aluminium which result from inaccurate techniques and contamination difficulties. The most widely used analytical technique is electrothermal atomic absorption spectrometry which can provide reliable measurements in the hands of a careful analyst.

Aluminum↗

Evaluation of equilibrium gel filtration chromatography for the study of protein binding of aluminum in normal and uremic sera.

Normal and uremic serum samples were chromatographed on a gel filtration column and fractions were analyzed for protein and aluminum content. Whereas much of the aluminum in the serum samples eluted with protein, a significant fraction appeared to be associated with low molecular weight species. Analysis of column fractions for aluminum binding by a competitive binding assay indicated no aluminum-binding serum constituents in these later eluting fractions. Uremic serum appeared, however, to contain aluminum-binding constituents of a lower molecular weight than in normal serum.

Aluminum↗

Quantitative study of aluminum binding to human serum albumin and transferrin by a chelex competitive binding assay.

Binding of aluminum to human serum albumin and transferrin was investigated using a competitive binding assay incorporating a cation exchange resin, chelex. Both albumin and transferrin were found to produce linear Scatchard plots of aluminum binding data over the aluminum and protein concentration ranges found in humans. Binding constants measured for albumin and transferrin were 1.96 and 0.515 microM, respectively.

Aluminum↗

Electrothermal atomic absorption spectrometric determination of aluminum in serum with a new technique for protein precipitation.

We have studied the electrothermal atomic absorption measurement of aluminum in serum samples by direct analysis with standard additions and the use of a matrix modifier. We have also measured aluminum by analysis of the supernate after pretreatment of serum with concentrated nitric acid to precipitate proteins, and have compared the results obtained by these two techniques. Both appear to eliminate interferences arising from the serum matrix. We quantified the aluminum by utilizing a stabilized temperature (L'vov) platform. Within-run precision (CV) for the standard-additions method was 16.6% (means = 6.5 micrograms/L) and 6.0% (means = 86.8 micrograms/L) and for the protein precipitation method was 10.1% (means = 10.9 micrograms/L) and 4.2% (means = 88.5 micrograms/L). Linearity of the standard curve extended from 0 to 120 micrograms/L for the standard-additions method and from 0 to 100 micrograms/L for the protein precipitation method. Samples from 38 patients were analyzed by both techniques, and linear regression analysis yielded the equation y (protein precipitation) = 1.02 X (standard additions) + 2.04.

Aluminum↗

Measurement of plasma prostaglandin E2 using capillary gas chromatography negative ion chemical ionization mass spectrometry.

A stable isotope dilution assay for the measurement of plasma prostaglandin E2 (PGE2) employing capillary column gas chromatography-negative ion chemical ionization mass spectrometry (GC-NICIMS) is described. PGE2 was extracted from plasma using C18 and silica SEP-PAKS. Further purification and separation was accomplished by thin layer chromatography. The prostaglandin was analyzed as its pentafluorobenzyl ester-methoxime-trimethyl-silyl ether, using fragment ions at m/e 524 (protium) and m/e 528 (deuterium) for quantitation. The mean plasma concentration of PGE2 determined in 8 healthy volunteers was 2.8 +/- 2.0 pg/ml.

Adult↗

Improved determination of aluminum in serum by electrothermal atomic absorption spectrophotometry.

This method for determining aluminum in human serum involves electrothermal atomic absorption spectrophotometry. A longer drying time allows less pre-dilution of the sample, and the method also includes a flush cycle after atomization. Standard-addition methodology is used to eliminate matrix effects and the need for a standard curve. We used this method on serum from 50 normal persons and from 34 patients with chronic renal failure who were on long-term intermittent hemodialysis. The mean normal serum aluminum concentration was 6.1 micrograms/L (CV 2.7%), and values for the patients ranged from 13 to 475 micrograms/L.

Aluminum↗

Lactate dehydrogenase isoenzyme 1 with the centrifugal analyzer after immunochemical removal of other isoenzymes.

We describe a centrifugal analyzer method for measuring the LD-1 isoenzyme of lactate dehydrogenase (EC 1.1.1.27) in serum, after immunochemical precipitation of the other four isoenzymes. Enzymic activity was measured kinetically at 30 degrees C with the pyruvate-to-lactate assay. The method for LD-1 was linear to 1000 U/L. The precision (CV) of the assay was 1.0--2.2% within-run and 3.1--4.5% day-to-day. The reference interval was 26--73 U/L (n = 51), corresponding to 21--35% of total LD activity.

Antigen-Antibody Complex↗

Use of urine cotinine to validate smoking self-reports in U.S. Navy recruits.

Cotinine analysis of urine has been used by many researchers to determine validity of smoking self-reports. This technique is easy and inexpensive, but has not been used previously in military smoking studies. This study incorporated a random validation of self-reported smoking by U.S. Navy recruits participating in a smoking relapse program (N = 496). Results of cotinine analysis indicate only a 1% misrepresentation of actual smoking status. These results suggest smoking self-reports from U.S. Navy recruits are very good indicators of actual smoking status.

Adult↗

Detection of cocaine and its metabolites in amniotic fluid and umbilical cord tissue.

The increased use of cocaine by women of child-bearing age has left many health care scientists searching for improved methods of detecting prenatal cocaine exposure. To that end, a study of the determination of cocaine and its metabolites in amniotic fluid and umbilical cord tissue was undertaken. Amniotic fluid (n = 32) and umbilical cord tissue (n = 70) specimens were collected from pregnant subjects admitted to labor and delivery at Shands Hospital at the University of Florida (Gainesville, FL). Subjects were interviewed regarding drug use during each trimester. Subjects reporting cocaine use were designated as target subjects, and those denying use were control subjects. The specimens were subjected to solid-phase extraction and analyzed for cocaine and its metabolites by gas chromatography-mass spectrometry. Cocaine analytes (predominantly benzoylecgonine) were detected in 28.1 and 18.5% of the amniotic fluid and umbilical cord tissue specimens, respectively. Other cocaine analytes frequently detected included ecgonine methyl ester and m-hydroxy-benzoylecgonine in amniotic fluid specimens and ecgonine methyl ester, norcocaine, and m-hydroxybenzoylecgonine in umbilical cord tissue specimens. This study has shown that cocaine and its metabolites are readily detected in specimens of maternal and fetal origin.

Adolescent↗

Ionized calcium buffering in the transfused anhepatic patient: Ab initio calculations of calcium ion concentrations.

Homeostatic control of ionized calcium can be volatile during liver transplantation, particularly during the anhepatic stage. Recently, an opportunity arose to evaluate an 11-year-old girl who developed persistent ionic hypocalcemia during a prolonged anhepatic period subsequent to the failure and removal of the graft. The patient was remarkable for having survived a 34-hour anhepatic interval before a second and successful orthotopic liver transplant. Ionic hypocalcemia (ionized calcium less than 1 mmol per L) coexisted with significant hypercalcemia (total calcium greater than 5 mmol per L) during this anhepatic interval. The discrepancy was due to high concentrations of citrate, which accumulated from the multiple transfusions of citrated blood, and the inability to metabolize citrate in the anhepatic state. Using a mathematical model to solve for free calcium ion concentration in the presence of multiple ligands, it is demonstrated that prolonged hypercitricemia markedly alters the calcium ion buffering properties of blood, and these changes must be recognized in order to prevent adverse clinical consequences of ionic hypocalcemia.

Blood Transfusion↗

Measurement of lipid peroxidation products in rabbit brain and organs (response to aluminum exposure).

A method was developed for measuring the concentration of lipid peroxidation products in rabbit brain, heart, lung, liver, and kidney tissue. Specimens were homogenized in cold buffer, acidified, and heated to near boiling in the presence of thiobarbituric acid in order to form the malondialdehyde-thiobarbiturate adduct. After centrifugation, the supernatant was injected onto a reversed-phase high pressure liquid chromatography (HPLC) column, and the effluent was monitored for absorbance at 532 nm. Absorbances were compared to a standard curve constructed from absorbance data for tetraethoxypropane standards, which yield stoichiometric amounts of the malondialdehyde-thiobarbiturate adduct. Results were expressed as nmol of adduct per gram (dry weight) of tissue. Hippocampus had significantly greater concentrations of lipid peroxidation products (79.0 +/- 15.7 nmol per g) than did brainstem (52.1 +/- 13.8 nmol per g), but there was no significant increase in lipid peroxidation in aluminum treated rabbit brains when compared with controls. Aluminum intoxication appeared, however, to stimulate lipid peroxidation in heart, lung, liver, and kidney. Aluminum accumulation in brain and organ tissue of treated rabbits was confirmed by atomic absorption spectrophotometry of an acid digest of the homogenate. These results are in contrast to previous studies which demonstrated an increase in lipid peroxidation products in rat brains following oral administration of aluminum hydroxide.

Aluminum↗

Measurement of cocaine and metabolites in urine, meconium, and diapers by gas chromatography/mass spectrometry.

Analytical methods were evaluated for measuring cocaine (CO), benzoylecgonine (BE), and ecgonine methyl ester (EME) in urine and methanolic extracts from meconium and diapers by isotope dilution gas chromatography/mass spectrometry (GC/MS). Volatile derivatives of the extracted drugs were generated before GC/MS analysis. Methanolic extracts from meconium and diapers were reconstituted in drug-free urine and treated as above. The limit of detection for the GC/MS method was calculated to be approximately 11 ng per mL. Within-run coefficients of variation (CVs) for urinary CO, BE, and EME were 5.7, 5.3, and 11.4 percent, respectively (N = 10); corresponding CVs for meconium 6.4, 10.7, and 21.9 percent (N = 8). Quantitative results were linear from 25 to 10,000 ng per g of meconium and 25 to 5,000 ng per mL of urine. Day-to-day precision varied from eight percent (CV) for BE in refrigerated or frozen urine to 34 percent for EME in refrigerated meconium. Recoveries of CO, BE, and EME from urine were 63, 19, and 42 percent, respectively; corresponding recoveries from meconium were 64, 21, and 25 percent. Cocaine and metabolites were extracted from wet but meconium-free diapers into methanol, which was evaporated before reconstituting in drug-free urine and extraction on a solid phase column. The CO, BE, and EME were detected in previously drug-free meconium after portions were deposited in a diaper which was wet with drug-positive urine. Unless precautions are taken to prevent extracorporeal contamination of meconium with urine, concentrations of CO and metabolites in meconium may be substantially augmented by contamination with urine. Analysis by GC/MS of CO and metabolites extracted from diapers provides an attractive alternative to collection of urine, which is difficult and may cause discomfort for the neonate.

Cocaine↗

The expanding role of molecular biology in clinical chemistry.

Technical improvements in the application of molecular biology methods to detection and identification of specific nucleic acid sequences have resulted in more widespread incorporation of these techniques into clinical laboratory services. Gene amplification techniques have been refined and automated, making possible the rapid and economical detection of attomolar and smaller quantities of genetic material. Of particular benefit to clinical chemistry applications is the remarkable specificity of a DNA probe for its complementary sequence. Applications of molecular biology techniques in clinical chemistry include diagnosis of infectious, neoplastic, genetic, and hematological diseases. In addition, the use of oligonucleotides as molecular recognition probes may provide new analytical strategies for a wide range of diagnostic applications that currently rely on antibodies.

Automation↗