Hermann Wisser

Urinary excretion of N-acetyl-β-d-glucosaminidase and alanine aminopeptidase in patients receiving amikacin or cis-platinum

Urinary excretion of alanine aminopeptidase (EC 3.4.11.2) and N-acetyl-beta-D-glucosaminidase (EC 3.2.1.30) was determined for 25-70 days in five patients receiving cis-platinum and for 8-53 days in six patients receiving amikacin. This study was performed to investigate if the excretion of urinary enzymes represents a sensitive parameter for the early detection of toxic kidney damage. The determination of N-acetyl-beta-D-glucosaminidase was carried out by the method of Knoll et al. [13]. The procedure of Mondorf et al. [14] for the estimation of alanine aminopeptidase activity was adapted to the Gemsaec Fast-Analyzer. In both patient groups an increase in the excretion of the two enzyme activities could be demonstrated. In patients receiving amikacin, the excretion of alanine aminopeptidase was always higher than that of N-acetyl-beta-D-glucosaminidase, whereas in three patients receiving cis-platinum it was the opposite. In two cis-platinum patients the excretion of both enzymes was of the same size. The changes during amikacin therapy seem to be reversible, whereas in four cis-platinum patients these changes seemed to be partly irreversible. Serum creatinine concentration was less sensitive than the urinary enzyme excretion for detection of kidney damage.

Soluble transferrin receptor and zinc protoporphyrin – competitors or efficient partners?

Abstract:  Objectives: Soluble transferrin receptor (sTfR) and zinc protoporphyrin (ZPP) are both parameters of iron deficient erythropoiesis (IDE), the sTfR measurement is commonly regarded to be the more sensitive test. sTfR also reflects erythropoietic activity, it is increased in enhanced erythropoiesis. Methods: We investigated the diagnostic accuracy of sTfR in assessment of iron deficiency (ID) and compared it with ZPP. The study was performed on 174 subjects, in which ID has been precisely staged. Results: Individuals without ID and patients with storage iron depletion only, had normal sTfR values. Patients classified as IDE and patients with iron deficiency anemia had significantly increased sTfR. There was a good correlation between sTfR and hemoglobin (r = −0.86; P < 0.0001) and between sTfR and ZPP (r = 0.86; P < 0.0001). When diagnosing ID, ZPP was the more sensitive test. In mildly developed IDE associated with ZPP‐ratios between 40 and 70 μmol/mol heme, the sTfR concentration was elevated in only 25% of the cases. Reliably elevated sTfR values were observed only in more advanced IDE, associated with ZPP > 70 μmol/mol heme. Conclusions: ZPP is not inferior to sTfR when diagnosing IDE. Given the good correlation between sTfR and ZPP and because ZPP is uninfluenced by the erythropoietic activity, sTfR and ZPP are not competitors, rather efficient partners in diagnosing anemias. By measuring ZPP and sTfR simultaneously, the diagnostic uncertainty inherent in each of them individually can be eliminated. In particular, the simultaneous determination of ZPP and sTfR enhances the diagnostic power of sTfR in assessment of the erythropoietic activity.

Blood Loss from Laboratory Tests

BACKGROUND Laboratory tests can be an important source of blood loss in hospitals, especially for newborns and patients in intensive care. The aim of this study was to quantify blood loss for laboratory diagnostic tests in a large number of patients in a teaching hospital. METHODS We estimated blood loss by multiplying the number and volumes of sampling tubes collected from 2654 adult inpatients. We compared the number of tests per patient for all inpatients and intensive care unit patients during the first period and again in the same time period 1 year later when cumulative blood-loss volumes were being reported to physicians and educational information had been given to decrease blood loss from laboratory tests. RESULTS For 95% of the patients, blood loss during hospitalization was <196 mL. The largest proportion of the blood samples was used for clinical chemical tests (median, 45%), followed by hematologic (median, 26%) and coagulation (median, 17%) tests. In the surgical and cardiovascular surgical intensive care units, however, blood gas analyses accounted for 19-34% (medians) of the use. For 5% of the patients, all undergoing intensive care, blood loss was >200 mL and for 0.7% was >600 mL during their hospital stay. Such high blood losses were observed in patients with long-term ventilation, coagulation disorders, and repeated surgery. The largest median blood loss was in patients undergoing cardiovascular surgery (median, 201 mL). The mean number of tests was 44 per inpatient before cumulative blood loss was being reported and 46 when it was being reported. CONCLUSIONS Blood loss from laboratory diagnostic testing is not likely to pose a problem for most hospitalized patients. Blood loss is greater in intensive care patients and after cardiovascular surgical procedures. Reporting of the cumulative individual blood loss did not decrease blood loss for laboratory testing.

Plasma free and conjugated catecholamines in diagnosis and localisation of pheochromocytoma

In six patients urinary excretion of vanillylmandelic acid and catecholamines (CA) could establish the diagnosis of pheochromocytoma. Free norepinephrine (NE) in plasma was within the normal range in two patients and plasma free epinephrine (E) was only marginally elevated in one of them. The degree of CA conjugation was not altered and scattered as in controls and was therefore not complementary to the usual determination of plasma free CA. The intermittent nature of CA secretion by the tumour could be demonstrated by multiple blood samplings during a 48-h study period in two patients, e.g. normal plasma values might be associated with pheochromocytoma if measurements are made during a trough. Thus a single peripheral CA determination cannot be of discriminative value in the diagnosis of pheochromocytoma unless it shows marked elevation. Ten patients subjected to intracardial measurements and five patients suspected of having a pheochromocytoma underwent venous catheterisation to determine their free and conjugated plasma CA. In controls CA values near the orifices of adrenal veins differed enormously and partly overlapped with corresponding levels of patients with pheochromocytoma. In one patient with surgically proven left adrenal tumour highest concentrations of CA were measured in the vena cava superior. These high CA concentrations, caused by paroxysmal release of CA by the tumour arouse suspicion of an additional, ectopic tumour. Because venous catheterisation cannot be relied on implicitly we propose computed tomographic scanning as a first step in localisation of a pheochromocytoma.

Free and conjugated catecholamines in human plasma during physical exercise

1. To investigate changes of free and sulfoconjugated catecholamines in response to alterations in sympatho‐adrenal activity, free and conjugated noradrenaline, adrenaline and dopamine were determined radioenzymatically in plasma of 49 subjects.

Urinary protein profiling by high-performance gel permeation chromatography

We describe a new application of high-performance aqueous gel permeation chromatography for the analysis of human proteinuria. Separations of urinary proteins from normal subjects and patients with renal impairment were performed with TSK G 3000 SW columns. The effects of pH and ionic strength of the eluent on the separation of urinary proteins were investigated. Albumins were selectively separated from urine by affinity chromatography on Blue Sepharose CL-6B. According to the results of clinical investigations, urinary protein pattern derived from gel permeation chromatography revealed a good prediction of the site of renal involvement. Predominant excretion of proteins with lower molecular weight than albumin correlated with tubular damage. Albumin and higher molecular weight protein patterns wer associated with glomerular disease. Absorbance measurements of the eluent at 280 nm were used for quantitative determination of total urinary protein. Gel permeation chromatography was compared to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and the resulting protein patterns are in good agreement.

Influence of Food Intake on Concentrations of Plasma Catecholamines and Cortisol

Plasma concentrations of epinephrine, norepinephrine, dopamine and cortisol were determined in four healthy volunteers--resting completely in bed--before and after food intake. Meals were provided at 17.00 h on the first day and at 12.00 h on the second day of the test. Blood samples were drawn every 15 minutes. Epinephrine, norepinephrine, dopamine, and cortisol showed a clear dependence on food intake. After the meals plasma concentrations of the four analytes showed rises, which were more pronounced after lunch at 12.00 h than after dinner at 17.00 h. An increase of epinephrine was observed shortly before meal-times; this could be caused by physiological adaptation to the usual rhythm of food intake and reflex response to expectation of the meal.

Identification of the "major" polymorphic carbocysteine metabolite as S-(carboxymethylthio)-L-cysteine.

Abstract The metabolism of the mucolytic agent carbocysteine ( S- carboxymethyl- l -cysteine , CMC) has attracted substantial interest during recent years [1]. It has been reported that sulphoxidation of CMC and its cysteinyl metabolites consitutes a major pathway of biotransformation. Moreover, it has been suggested that CMC sulphoxidation exhibits a genetic polymorphism expressed in two phenotypes in the population, the poor sulphoxidizers (PS) and the extensive sulphoxidizers (ES) [2]. This sulphoxidation polymorphism has been linked to certain idiopathic diseases (e.g. primary biliary cirrhosis) and the toxicity of sulphur-containing drugs (e.g. d -penicillamine, sodium aurothiomalate) [1]. However, more detailed metabolic studies on CMC and a carbon-13 labelled analogue ( S- carboxy -[ 13 C ] methyl- l -cysteine , 13C-CMC) have demonstrated the absence of significant amounts of any of the putative cysteinyl sulphoxide metabolites in the urine. These studies which were based on HPLC [3–5], 13C NMR [5,6], and gas chromatography/mass spectrometry (GC/MS) [7] demonstrated that, besides unchanged carbocysteine, thiodiglycolic acid (di-[carboxymethyl] sulphide, TDGA) and its sulphoxide (TDGA-SO) were the major biotransformation products of CMC in man [8]. Approximately 6–8 hr after administration of the drug, urinary excretion of an additional novel metabolite (1) was observed by TLC after sulphur-selective visualization [9]. This particular metabolite 1 of yet unknown structure was the only urinary component exhibiting polymorphic character present after oral administration of CMC [10]. About 7.5–10% of the two populations excreted only marginal amounts of 1, and a more than two-thousand fold difference in the urinary excretion of metabolite 1 has been estimated by TLC [10,11]. The objective of this study was to isolate and identify this metabolite.

The relationship of free and conjugated catecholamines in plasma and cerebrospinal fluid in cerebral and meningeal disease

The concentration of free and conjugated norepinephrine (NE), epinephrine (E) and dopamine (DA) were measured by a modified radioenzymatic assay in the plasma and in the cerebrospinal fluid (CSF) of 45 patients with normal and in 21 patients with disturbed blood-CSF barriers. In patients with an undisturbed blood-CSF barrier the free NE and E in CSF were 128±45 ng/l and 27±20 ng/l (mean values±S.E.), respectively, and represented about 50 % of the average plasma values. Mean DA was not different in plasma (47±22 ng/l) and in CSF (41±19 ng/l). Both in plasma and in CSF, considerable higher free catecholamine (CA) levels were measured in patients with dysfunction of the blood-CSF barrier. In one patient with bacterial meningitis twofold higher concentrations of free NE and DA in CSF as compared with plasma were detectable. Sulfate conjugates of catecholamines are predominant in plasma and CSF. The contribution of conjugated CA to total CA in plasma from patients with normal blood-CSF barrier averaged 69.7 %, 63.1 % and 98.1 % for NE, E and DA, respectively and was significantly lower in the CSF (p<0.001). In patients with disturbed blood-CSF barrier, the increases of conjugated CA were more pronounced in CSF than in plasma. Further, the contribution of conjugated NE and E to total NE and E in CSF was not only increased in patients with bacterial meningitis, but also in patients with renal insufficiency compared to the “control” patients (p<0.02 and p<0.001 resp.). Free and conjugated NE, E and DA in the plasma and CSF were related significantly (p<0.01 resp.) with stronger correlation for conjugated CA (p<0.001 resp.). These results together with findings in the literature, suggest that there is little or no rostral-caudal gradient in CSF CA conjugate concentrations and that even in patients with intact blood-CSF barrier plasma conjugated CA concentrations influence those in CSF. Thus only free CA levels in CSF may reflect the central adrenergic activity.

Determination of catecholamines in plasma by HPLC and amperometric detection. Comparison with a radioenzymatic method

The determination of norepinephrine and epinephrine in plasma by HPLC with amperometric detection was modified, giving detection limits of 25 ng/l for norepinephrine and epinephrine, respectively, using 1 ml plasma. In order to achieve this sensitivity, it was necessary to minimize the background noise by modification of instrumentation and specimen handling. Particularly important was the extra purification of the reagents, the application of micro-bore HPLC, the enzymatic cleavage of uric acid and temperature control of the amperometric cell and the amplifier. Comparison of the present method with the radioenzymatic determination of catecholamines resulted in coefficients of correlation of r = 0.924 and 0.919 for norepinephrine and epinephrine, resp. (n = 38). The concentrations of the 38 different samples used for the comparison were in the physiological range.