Concentration measurements of multiple analytes in human sera by near-infrared laser Raman spectroscopy.

Our primary goal in this study is to demonstrate that near-infrared Raman spectroscopy is feasible as a rapid and reagentless analytic method for clinical diagnostics. Raman spectra were collected on human sera by use of a 785-nm excitation laser and a single-stage holographic spectrometer. A partial-least-squares method was used to predict the analyte concentrations of interest. The prediction errors of total protein, albumin, triglyceride, and glucose in human sera ranged from 1.0% to 10%, which are highly acceptable for clinical diagnosis, of their mean physiological levels. For investigating the potential application of near-infrared Raman spectroscopy in screening of therapeutical drugs and substances of abuse the concentrations of acetaminophen, ethanol, and codeine in water solution were measured in the same fashion. The errors of the Raman tests for acetaminophen and ethanol are lower than their toxic levels in human serum, and the sensitivity for detection of codeine fails to reach its toxic level.

Determinants of low serum concentrations of salicylates in patients with Kawasaki disease.

The mechanisms leading to the previously reported difficulties in achieving therapeutic serum concentrations of salicylates in Kawasaki disease were studied in eight children, once during the acute (febrile) phase and again during the nonfebrile (subacute) phase of the disease. Salicylate bioavailability was impaired during the acute phase of the disease (47.7% +/- 6.6%), and increased significantly thereafter to 75.1% +/- 9.3%. During the febrile phase there was a significant correlation between salicylate bioavailability and steady-state serum concentrations. Salicylate renal clearance was significantly higher during the febrile phase (14.45 +/- 2.5 mL/kg.h), compared with the nonfebrile phase (7 +/- 1.6 mL/kg.h, P less than 0.05). The change in salicylate clearance could be explained by decreased protein binding in the acute phase (82.5% +/- 1.9%) with substantially more free salicylates caused by significantly lower serum albumin concentrations. Changes in urine metabolites during the acute and subacute phases were consistent with the changes in dose administered (100 mg/kg in the acute phase vs 10 mg/kg in the subacute phase). The pattern of metabolites excreted in the urine of children with Kawasaki disease receiving 100 mg/kg was similar to that in children with juvenile rheumatoid arthritis receiving the same dose.

Corticosteroids-salicylate interaction in a case of juvenile rheumatoid arthritis.

In an 11-year-old child with juvenile rheumatoid arthritis (JRA), the addition of prednisone caused a significant decrease in salicylate serum concentrations. A pharmacokinetic assessment suggested that these changes were not the result of altered compliance or impaired absorption of salicylate but rather an increase in salicylate clearance induced by the corticosteroid.

Delayed digoxin toxicity following discontinuance in acute renal failure.

A digitalized 75-year-old patient with postoperative renal failure demonstrated a progressive rise in serum digoxin concentration, peaking at 3.4 nmol X L-1 three days following discontinuance of the drug. This was accompanied by cardiac bradyarrhythmias. Although the serum digoxin concentration had already started to climb from a therapeutic level prior to the discontinuance of the drug, the unabated and substantial rise was consistent with a dramatic decrease in the apparent volume of distribution of digoxin accompanying acute renal failure. Serum digoxin levels were determined with fluorescence polarization immunoassay, which has an improved specificity when compared to the commonly used radioimmunoassays for digoxin.

Valproic acid disposition in rabbits during chronic treatment with Escherichia coli endotoxin.

The disposition of valproic acid (VPA) in rabbits was studied after chronic treatment with Escherichia coli endotoxin. Endotoxin (1-2 micrograms/kg) was administered daily to 10 male New Zealand white rabbits for 5 days. On day 5, 50 mg/kg of VPA was given iv during the time of the peak febrile response. Blood samples were drawn at appropriate time intervals and analyzed for free and total VPA levels as well as plasma proteins and free fatty acids. The data were compared with similar control experiments performed 2 weeks before and after endotoxin treatment. Pharmacokinetic analysis indicated that the changes in free VPA clearance after endotoxin were related to the change in the febrile response during chronic treatment (r = 0.77; p less than 0.05); that is, animals which developed tolerance to the febrile response showed elevated drug clearance, whereas nontolerant animals showed decreased clearance of VPA.

Pharmacokinetics of enteric-coated valproic acid.

Five adult epileptic patients received 1,000 mg of valproic acid (Depakene) in both the regular and the enteric-coated form. Serum valproic acid levels were determined at suitable intervals after drug administration. Pharmacokinetic parameters were equivalent for both preparations except for an absorption lag with the enteric-coated form. The relative bioavailability of the two compounds was similar across the group of patients, although there were marked differences between individual subjects. Close supervision of valproic acid serum levels is suggested after a change in drug formulation.

Characterization of proteins associated with nuclear ribonucleoprotein particles by two-dimensional polyacrylamide gel electrophoresis.

Rat liver nuclear ribonucleoprotein particles were prepared by two different methods and defined as 40S ribonucleoprotein (40S RNP) and heterogeneous nuclear ribonucleoprotein (HnRNP) particles. The RNP particles were either solubilized in 8 M urea--6 mM 2-mercaptoethanol--20 mM glycine--20 mM Tris--HCl (pH 8.4) or subjected to removal of RNA by phenol extraction prior to solubilizing the proteins in the urea buffer. The proteins associated with 40S RNP and HnRNP were heterogeneous and very similar in their electrophoretic patterns when analyzed by two-dimensional PAGE, except a protein with molecular weight of 62 000 and an isoelectric point (pI) of 6.2 was present only in HnRNP particles. At least 12 major and 22 minor components could be identified in both preparations. The major proteins were found at pI values varying from 6.0 to 8.5 and with molecular weights from 32 000 to 42 000, and a group of proteins with molecular weight approximately 65 000 were more prominent in HnRNP than in 40S RNP. The other components were found mainly at pI ranges from 5.0 to 6.5 with molecular weights from 43 000 to 65 000. The phenol method extracted essentially all proteins associated with either 40S RNP and HnRNP, but was less effective in extracting a group of proteins with pI values from 5.0 to 5.5 and more efficient for proteins with pI values from 7.5 to 8.5. When chromatin proteins isolated by phenol extraction were compared with HnRNP particle proteins isolated by the same method, the electrophoretic mobilities of the HnRNP particle proteins were found to be identical with a fraction nonhistone chromatin proteins. The 40S RNP particles were further purified by metrizamide isopycnic density gradient centrifugation. The electrophoretic patterns of these proteins were very similar to those prepared by sucrose density gradient centrifugation. Therefore, we concluded that the proteins of RNP particles constituted part of the chromatin proteins.

Quantitative determination of thiazides in urine by a sensitive colorimetric method.

Chloriothiazide and hydrochlorothiazide may be determined in urine by the Bratton-Marshall reaction after extracting with ethyl acetate and treating the extract with Florisil. Such treatment reduces the background color of the urine sample and eliminates interferences. The modified procedure is thus sensitive enough to determine both drugs in 24 hour urine specimens from patients receiving therapeutic doses orally.

Fractionation of nucleolar proteins by two-dimensional gel electrphoresis.

Isolation of nucleolar proteins was obtained by dissociation in the presence of urea-guanidine hydrochloride, followed by high-speed centrifugation to remove nucleic acids. At least 31 fractions of nucleolar proteins were detected by isoelectrofocusing gel electrophoresis in pH range 3.5-10. Following two-dimensional gel electrophoresis on sodium dodecyl sulfate-polyacrylamide slab gels, more than 100 components of nucleolar proteins were identifieid. Two-thirds of nucleolar proteins were located in the pH range 5-8 following isoelectrofocusing. The molecular weights of these classes of proteins were shown to be mostly 30000-70000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.

Isolation of nuclear acidic proteins from rat tissues. Characterization of acetylated liver nuclear acidic proteins.

Nuclear acidic proteins isolated from rat brain, heart, kidney and liver showed similar, complex patterns on electrophoresis in sodium dodecyl sulphate-polyacrylamide gels. The contamination of nuclear acidic proteins by nuclear-membrane acidic proteins was found to the extent of 11%. Incorporation of [(3)H]acetate into the various nuclear acidic proteins in vivo, which were fractionated by polyacrylamide-gel electrophoresis, differed from tissue to tissue. Hydrolysis of these acetylated nuclear acidic proteins with 6m-HCl at 110 degrees C released 70% of the radioactivity, which indicated that labile acetyl groups had been incorporated into these proteins. Analysis of [(3)H]acetate-labelled nuclear acidic proteins revealed two acetylated amino acid residues, N(2)-acetylserine and N(2)-acetyl-lysine. The significance of the role played by nuclear acidic proteins in relation to gene regulation is discussed.