Comparison of capillary electrophoretic and liquid chromatographic determination of hypoxanthine and xanthine for the diagnosis of xanthinuria.

A capillary electrophoretic (CE) method for the determination of hypoxanthine and xanthine in urine was developed to diagnose xanthinuria. The linearity was excellent up to 200 mumol l-1 for the two compounds and the limit of quantitation was 2 mumol l-1. A comparison o the results obtained using CE was made with those obtained by the high-performance liquid chromatographic (HPLC) technique described previously. With regard to specificity, sensitivity and reproducibility, the results are similar but CE is more rapid than HPLC.

Liquid-chromatographic study of purine metabolism abnormalities in purine nucleoside phosphorylase deficiency.

Using HPLC methods, we measured the concentrations of nucleosides and nucleotides for a patient with no purine nucleoside phosphorylase (PNP; EC 2.4.2.1) enzymatic activity. Concentrations of inosine and guanosine were abnormally high in urine and plasma, whereas guanosine diphosphate (GDP) and guanosine triphosphate (GTP) concentrations in erythrocytes were depleted. The unusual presence of deoxyribonucleosides (deoxyinosine and deoxyguanosine) and deoxyribonucleotides (dGDP and dGTP) was also notable. Thus, HPLC represents an accurate and useful tool for the study of purine metabolic disorders.

Capillary electrophoretic analysis of DNA restriction fragments and PCR products for polymorphism and mutation studies in cystic fibrosis and Gaucher's disease.

Two methods are described for the analysis of DNA restriction fragments and PCR products in studies on polymorphism and mutation in cystic fibrosis and Guacher's disease, based on capillary electrophoresis. In one CE system, a Beckman kit for producing a chemical gel (polymerized within the capillary) is used for single-stranded DNA fragments from 10 to 300 bases in size. Its performance was demonstrated on the separation of a mixture of polydeoxyadenylic acids p(dA)40-60 at 30 degrees C. Electrokinetic injection was used (5-7 kV for 5-20 s), the applied field being 300 V cm-1 for an effective length of 7, 20 or 30 cm and 100 microns i.d., with Tris-borate buffer containing urea. Typical electropherograms are presented for the analysis of CF mutation delta F508 in PCR products from homozygous and heterozygous individuals, illustrating the resolution of two complementary single strands (95b and 95b) of a DNA fragment. DNA fragments differing in size by only one base could also be resolved, as shown for the 105b and 106b fragments obtained from a heterozygote for 3905 insT CF mutation, with a run time of ca-45 min. If discrimination were only required between fragments differing by two or more bases, run times could be reduced by 6 when using a capillary length of only 7 cm x 100 microns i.d.(ABSTRACT TRUNCATED AT 250 WORDS)

Comparison of red cell transfusion and polyethylene glycol-modified adenosine deaminase therapy in an adenosine deaminase-deficient child: measurement of erythrocyte deoxyadenosine triphosphate as a useful tool.

The effect of red cell transfusion and polyethylene glycol-modified adenosine deaminase therapy on biochemical abnormalities, clinical status, and immunologic function in an adenosine deaminase-deficient child was investigated. After red cell transfusions, erythrocyte deoxyadenosine triphosphate (dATP) concentrations decreased about 95% and were closely related to adenosine deaminase activities; deoxyadenosine diphosphate concentrations decreased only approximately 30%. The evolution of dATP levels was also closely related to the improvement in clinical status of the patient. However, immune function was not restored. After polyethylene glycol-modified adenosine deaminase therapy, the concentration of erythrocyte dATP decreased to undetectable levels correlated with an increase of T lymphocyte counts and an increase of lymphocyte responses to mitogens. Immune functions were restored only when dATP levels were below 15 mumols/L. It appears that red cell transfusion therapy is not sufficiently effective to reduce and maintain erythrocyte dATP levels at values compatible with normal immune function. On the contrary, polyethylene glycol-modified adenosine deaminase therapy is a suitable treatment to reduce dATP levels to near undetectable values, allowing the immune function to be restored, dATP measurement is a very useful tool for monitoring and evaluating the degree of efficiency of therapy in adenosine deaminase deficiency.

Diagnosis of alcaptonuria: rapid analysis of homogentisic acid by HPLC.

The clinical and biochemical features of five cases of alcaptonuria were reported. The concentration of homogentisic acid was determined in urine and also in plasma using a rapid, sensitive and specific HPLC method. In all five cases, the concentrations of homogentisic acid were elevated in urine rising up to 46.5 mmol/24 h. In plasma, homogentisic acid levels ranged from 33 to 38 mumols/l. In healthy individuals, homogentisic acid was not detectable in plasma and urine. The method described represents a very useful and suitable analytical tool for the diagnosis of alcaptonuria.

Homogentisic acid determined in biological fluids by HPLC.

In this rapid, specific, and sensitive high-performance liquid-chromatographic method of analysis for homogentisic acid in biological fluids, homogentisic acid is separated on a column of Nucleosil CN. This method, which we applied to the diagnosis of three cases of alcaptonuria, represents a suitable analytical tool for the diagnosis of alcaptonuria.

[Use of thiopental in man. Determination of this drug and its metabolites in plasma and urine by liquid phase chromatography and mass spectrometry]. / Utilisation du thiopental chez l'homme. Détermination de ce médicament et de ses métabolites dans le plasma et les urines par chromatographie en phase liquide et spectrométrie de masse.

Thiopental is an anaesthetic drug which is currently used for cerebral resuscitation. In this last indication the drug is administrated at high doses over a period of several days. Under clinical trials thiopental and two metabolites namely 5 ethyl-5 (1' methyl-3' hydroxy-butyl) 2 thiobarbituric acid and 5 ethyl-5 (1' methyl-3' carboxy-propyl) 2 thiobarbituric acid have been determined by high performance liquid chromatography and mass spectrometry. In human plasma high concentrations of thiopental and 5 ethyl-5 (1' methyl-3' hydroxy-butyl) 2 thiobarbituric acid and a lower concentration of 5 ethyl-5 (1' methyl-3' carboxy-propyl) 2 thiobarbituric acid have been found. In urine samples these metabolites are excreted in large and approximately equal quantities, whereas small amounts of thiopental were recovered.