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The antiepileptic drugs phenobarbital and phenytoin were determined in serum by enzyme immunoassay (Emit, Syva Corp.) and gas-liquid chromatography. The Emit assays were mechanized by the use of an Eppendorf analyzer 5010. The precision of the Emit system was sufficient (coefficient of variation within series 6-13% and from day to day 8-15% with various calibrators and control sera). Moreover the Emit method is rapid, specific and easy to perform. The procedure requires only 10 microliter of serum per determination. A disadvantage however is the high cost of the reagents. A comparison of the results obtained by Emit and gas-liquid chromatography in a series of about 50 patients showed a good correlation between both methods (correlation coefficient r = 0.968 for phenobarbital and 0.978 for phenytoin).

An evaluation of enzyme immunoassays for determination of thyroxine in serum (EMIT ABA thyroxine assay, Syva Corp., ENZYMUN assay thyroxine, Boehringer Mannheim) and of thyroxine binding index (ENZYMUN assay TBI1), Boehringer Mannheim) is presented. The precision of the enzyme immunoassays was adequate (coefficients of variation ranged from day to day with EMIT from 3--11% and with ENZYMUN from 4--11%). Both assays are specific and easy to perform. About 20 unknown samples can be analyzed in duplicate by EMIT within 60 minutes and by ENZYMUN within 250 minutes. A comparison of the results obtained by enzyme immunoassays and radioimmunoassay in a series of about 100 patients showed a good correlation between both methods. The precision of the ENZYMUN TBI assay was adequate (coefficient of variation from day to day 4.9%) and the thyroxine/TBI-ratio correlated well with the thyroxine/TBG-ratio.

Recently hydrazonopropionate derivatives have been recognized as a new group of compounds with strong hypoglycemic effects in various laboratory animals. The hypoglycemia probably has several causes of which an inhibition of gluconeogenesis and of intestinal glucose uptake could be identified. In the present report, the influence of phenylethylhydrazonopropionate, cyclohexylethylhydrazonopropionate, methylcinnamylhydrazonopropionate and their corresponding hydrazine derivatives on the glucose uptake of rat and guinea pig jejunum has been compared with the well known effects caused by phenformin. All substances tested inhibited the jejunal glucose uptake, phenelzine, methylcinnamylhydrazine and cyclohexylhydrazonopropionate more effective, the other compounds less effective than phenformin. The hydrazine derivatives appeared more effective than the hydrazone compounds used. The cellular ATP/ADP ratio was not influenced by the hydrazonopropionate compounds.

Pharmacokinetics of methotrexate (MTX) was studied in 34 patients (age 1-25 years, median 12 years) predominantly with primary brain tumors and osteosarcoma, who received a total of 64 high-dose infusions (12 g/m2/4 h, maximum dose 20 g), followed by leucovorin rescue (COSS 82). Serum samples were collected over a period of at least 72 h after the end of infusion and MTX was measured by enzyme immunoassay (EMIT). The data were fitted to a biexponential equation using a nonlinear regression analysis. The concentration-time decay of MTX in serum observed in 29/34 patients receiving 4 x 15 mg/m2/d p.o. leucovorin up to 5 days was biphasic with mean half-lives (+/- SD) of 2.42 +/- 0.45 h for t1/2 alpha and 19.9 +/- 7.6 h for t1/2 beta. The steady-state volume of distribution (Vss) was 0.56 +/- 0.18 l/kg and the total body clearance (CL) 71 +/- 20 ml/min/m2 (mean +/- SD). Peak serum concentrations ranged from 674-1778 mumol/l (mean +/- SD, 1201 +/- 293 mumol/l). In 5/34 patients who received a prolonged leucovorin rescue due to a delayed MTX elimination t1/2 alpha was greater than 3.1 h. The data of this study suggest that patients with MTX serum concentrations of less than or equal to 6.3 mumol/l at 24 h, less than or equal to 0.77 mumol/l at 48 h, and less than or equal to 0.33 mumol/l at 72 h after the end of infusion, and a t1/2 alpha of less than or equal to 3.1 h (97.5th percentiles) are at low risk of toxicity.(ABSTRACT TRUNCATED AT 250 WORDS)

12 patients with obstructive airway diseases were given a single oral dose of 400 mg resp. 600 mg Cholintheophyllinate (Euspirax) and 350 mg Theophyllin-Aethylendiamine (Euphyllin retard). In an intra-individual comparative study the theophylline-serum-concentrations and the bronchospasmolytic effects of both drugs have been checked over a period of 12 hours. A very good resorption and the highest mean blood levels of 9,13 +/- 2,88 mg/1 resp. 12,11 +/- 2,55 mg/1 resulted from Cholintheophyllinate 2 hours after application. The effect on the lung function parameters was most expressed 2 to 4 hours after application of this drug and correlated well with the theophylline-blood-levels. The higher dose of Euspirax was followed more frequently by side effects. 2, 4 and 12 hours after application of Euphyllin retard the serum-concentrations were strikingly low. The mean values were 2,04 +/- 1,22 mg/l, 3,00 +/- 2,18 mg/l resp. 3,38 +/- 2,01 mg/l. In spite of the low blood levels a significant improvement of the lung function parameters could be noticed after 4 to 6 hours. The qualities of both drugs as to their bioavailability and effect on airway obstruction shall be examined by means of another comparative study.

The properties of six adsorbents for Haemoperfusion were investigated in vitro: Amberlite XAD-2; Amberlite XAD-4; Haemocol; collodion coated charcoal; albumine-collodion encapsulated charcoal and activated carbon fibers. Carbromal and diazepam are most quickly removed from aqueous solutions by Amberlite XAD-2 and XAD-4. Of the charcoal containing materials, Haemocol eliminates most rapidly carbromal and diazepam; while activated carbon fibers remove phenobarbital faster. When heparinized blood is perfused, a remarkable reduction in platelet count is observed. The drop is most pronounced with Amberlite XAD-2 and XAD-4, but even when using charcoal preparations the platelet depletion can reach 10% of the original number over 5 hours. Haemolysis is most pronounced with Amberlite XAD-2, XAD-4 and Haemocol. As the efficiency of the adsorbents towards the various drugs differ and as their adverse effects also vary, the decision for a particular material will have to be made according to the individual case.

Four different homogeneous non-isotopic immunoassays for the determination of total phenytoin in serum were evaluated and compared with a gaschromatographic method (GC) described by W. R. Külpmann & M. Oellerich [1981), J. Clin. Chem. Clin. Biochem. 19, 249-258) enzyme multiplied immunoassay technique (EMIT), fluorescence polarization immunoassay (FPIA), nephelometric inhibition immunoassay (NIIA) and substrate labeled fluorescent immunoassay (SLFIA). The between-days coefficients of variation in the medium therapeutic range were 4.0% (n = 29) with EMIT, 4.6% (n = 15) with FPIA, 7.8% (n = 10) with NIIA, 2.8% (n = 12) with SLFIA and 5.7% (n = 15) with GC. The recovery in spiked serum samples (phenytoin concentration: 11.9-99.1 mumol/l) was 98-101% with EMIT, 97-107% with FPIA, 102-110% with NIIA, 94-97% with SLFIA and 97-104% with GC. All of the tested immunoassays and GC yielded comparable results. The NIIA showed a somewhat lower correlation. In samples from an uraemic patient, however, great deviations from GC values were obtained with EMIT (bias: +22 to +85%) NIIA (+68 to +114%) and SLFIA (+48 to +52%). Only the results of FPIA were in good agreement with those of GC (bias: +1 to -7%). All the immunoassays showed a cross-reaction with 5-(4-hydroxyphenyl)-5-phenylhydantoin, which was most expressed with SLFIA and NIIA. The detectability of the immunoassays was adequate to allow precise measurements within the therapeutic range. After ultrafiltration of the serum, free phenytoin was measured by EMIT, FPIA and capillary gas chromatography.(ABSTRACT TRUNCATED AT 250 WORDS)

A rapid new ultrafiltration technique (EMIT FreeLevel System I) for the routine measurement of unbound phenytoin concentrations was evaluated. The precision of this procedure was sufficient (between-days coefficient of variation, 11.7%). The results obtained by the ultrafiltration technique for the percentage of free phenytoin in samples from about 40 non-uraemic patients treated with this drug were in good agreement with those determined by ultracentrifugation and equilibrium dialysis (ultrafiltration vs ultracentrifugation, y = 0.94x + 0.60%; ultrafiltration vs equilibrium dialysis, y = 1.02x - 0.60%). The mean value of the results obtained by ultracentrifugation was significantly about 8% lower than that observed with equilibrium dialysis, apparently due to a sedimentation of free phenytoin during ultracentrifugation. With the methods used in our study, the mean values of percentage phenytoin bound to serum proteins obtained in samples from non-uraemic patients ranged from 91.8 to 92.8%. All 3 methods yielded similar binding curves for phenytoin, with spiked human pool sera containing albumin concentrations between 19 and 45 g/L. A rise of the unbound phenytoin fraction was observed with increasing total concentrations of the drug and a decrease of the albumin concentration. With samples from non-uraemic patients (n = 203), a rather good correlation was found between free and total phenytoin concentrations (r = 0.91). In a number of patients (n = 11), free phenytoin concentrations correlated better than total phenytoin concentrations with the clinical status. Patients with free phenytoin concentrations of less than or equal to 2.1 micrograms/ml and total phenytoin concentrations above the therapeutic range did not show signs of toxicity. From the results of our study it is concluded that the EMIT FreeLevel ultrafiltration technique is very well-suited for a rapid and reliable separation of unbound phenytoin. In patients with altered protein binding or an unusual clinical response, free phenytoin determinations appear to be necessary for proper interpretation of total phenytoin levels and rational dosage adjustment.

A method for rapid assessment of hepatic function in liver donors based on the formation of the lignocaine metabolite monoethylglycinexylidide (MEGX), was used in a prospective study of 69 donor-recipient pairs. The probability of graft survival over 120 days was significantly higher for livers from donors with MEGX test values above 90 micrograms/l than for those from donors with MEGX values of 90 micrograms/l or below. Other liver function tests (bilirubin, prothrombin time, activity of aminotransferases, glutamate dehydrogenase, and cholinesterase, indocyanine green clearance, and galactose elimination capacity) were inefficient at predicting early outcome of transplantation. For a 20-day graft survival, the MEGX test showed prognostic sensitivity of 73% and specificity of 78%. These findings suggest that the MEGX formation test could be valuable for selection of donor organs.

An evaluation of a heterogeneous enzyme immunoassay for determination of triiodothyronine in serum (Enzymun-Test T3, Boehringer Mannheim) is presented. The enzyme immunoassay was compared with the laboratory routine radioimmunoassay. The precision of both assays was satisfactory at triiodothyronine concentrations between 1.0 and 8.0 nmol/l (coefficients of variation from day to day less than or equal to 10%). Average recoveries of triiodothyronine in pool sera ranged with the Enzymun-Test T3 from 96--104% and with the radioimmunoassay from 88--111%. A comparison of the results obtained by Enzymun-Test T3 and the radioimmunoassay in a series of 103 patients showed a good correlation between both methods. L-thyroxine did not cause a relevant cross-reaction in the enzyme immunoassay. About 20 unknown samples can be analyzed in triplicate by Enzymun-Test T3 within 260 minutes.

A homogeneous enzyme immunoassay for the determination of methotrexate in serum (EMIT, Syva Corp.) was fully mechanized by the use of an Eppendorf analyzer 5010. Coefficients of variation from day to day were in the range 2--16%. A comparison of the results obtained by EMIT and a radioimmunoassay (Methotrexate 125 I Radioimmunoassay Kit, Diagnostic Biochemistry Inc.) in a series of 50 samples from patients showed a good correlation between both methods. The specificity of the EMIT assay appeared to be adequate. The time required for the determination of about 20 patient samples by this procedure was only about 80 minutes. From our clinical experimence the EMIT assay appears to be well suited for routine monitoring of methotrexate serum concentrations above 0.1 mumol/1 during intrathecal and high dose methotrexate therapy.

In a double-blind, multiple-dose, cross-over study the therapeutic effects of an S-R theophylline preparation given once each night (mean 11.2 mg/kg/day) were compared to standard treatment with an S-R aminophylline preparation every 12 h (mean 10.3 mg theophylline/kg/day) in 9 patients with nocturnal asthma. On Day 5 of each treatment period, serum theophylline concentrations and PEF were measured every 2 h over a 24 h period. With the twice-daily regimen peak theophylline concentrations were lower at night than during the day (mean 9.4 +/- 0.9 vs 11.3 +/- 1.0 mg/1). With the single administration, serum theophylline concentrations were considerably higher at night (mean C max 16.3 +/- 1.4 mg/1). In spite of the high theophylline peak to trough difference (mean 12.0 +/- 2.1 mg/1) during the once daily regimen, the circadian variations of PEF were significantly reduced. Thus, greater airways' stability can be obtained by appropriate timing of theophylline administration.

In two double-blind, multiple-dose cross-over studies the therapeutic effects of SR theophylline preparations given once each night (mean 11.2 mg/kg per day) versus twice daily in equal doses (mean 10.3 mg/kg per day) (study I) and SR-terbutaline in equal doses (mean 0.25 mg/kg per day) versus SR theophylline in unequally divided daily doses (mean 5.3 mg/kg morning dose, 10.6 mg/kg evening dose) study II) were compared in 19 patients with nocturnal asthma. At the end of each treatment period drug serum concentrations and PEFR were measured every 2 hr over a 24-hr period. With the twice-daily, equally divided regimen, serum theophylline concentrations were lower at night than during the day (mean 9.4 +/- 0.9 versus 11.3 +/- 1.0 mg/l). With the single evening administration, serum theophylline concentrations were considerably higher at night (Cmax 16.3 +/- 1.4 mg/l) and the circadian variation of PEFR was significantly reduced. PEFR was higher during night and early morning (283 +/- 14 versus 217 +/- 11 l/min, P less than 0.005). During daytime in study II, PEFR values were slightly higher with theophylline than terbutaline. There was no significant difference in peak flow between either treatment during the night and early morning. However, additional use of inhaled beta-2-mimetics because of asthmatic attacks occurred more often during terbutaline (79 times in 8/10 patients) than theophylline treatment (29 times in 5/10 patients). Symptom scores, number of attacks and side-effects clearly favor the theophylline regimen. We conclude that for patients with nocturnal asthma a once-nightly dose of SR theophylline can be sufficient for stabilization of the airways.

EMIT assays for the determination of phenytoin, methotrexate, disopyramide, digoxin and thyroxine were adapted to the Cobas Bio centrifugal analyzer and compared with the corresponding laboratory routine procedures. Evaluation of the data from the Cobas Bio by 5 different mathematical models showed that the four-parameter logit model correlated best with the comparison procedures and the originally recommended calculation model for the reagent lots used in our study. The precision of the EMIT phenytoin, methotrexate and disopyramide assays was in most cases very good (between-days coefficient) of variation 1.6-7.5%). A lower precision was observed with the EMIT digoxin assay (between-days coefficients of variation 9.2-16.8%) and at low concentrations also with the EMIT thyroxine assay (3.1-21.4%). Calibration curves of the EMIT phenytoin, methotrexate assays were stable for at least one hour. The results from the determination of phenytoin, methotrexate and disopyramide in patient samples by use of the Cobas Bio were in good agreement with those values obtained with the EMIT/LAB. The data determined with the EMIT digoxin assay adapted to the Cobas Bio correlated better with those of a radioimmunoassay than the values measured with the EMIT/LAB system. The results of thyroxin determinations with EMIT by use of the Cobas Bio and the original procedure with an ABA-100 were in good agreement and on average about 12% lower than those measured by radioimmunoassay. The Cobas Bio allows rapid determination with EMIT and a reduction in direct costs of up to 85%.

Theophylline concentrations were determined by fluorescence polarization immunoassay (FPIA) in saliva, serum and serum water of healthy volunteers and outpatients after administration of single theophylline doses, and after the administration of several doses in order to establish a steady state. The FPIA allowed rapid and reliable theophylline determinations in saliva and serum water (between-days coefficients of variation: less than 3%; recovery: 95-03%). Salivary theophylline concentrations measured by FPIA in 30 samples agreed well with those determined by HPLC. Furthermore the results obtained by ultrafiltration for the concentration of unbound theophylline in serum water were in good agreement with those determined by ultracentrifugation. The binding of theophylline by serum protein rose by about 25%, when the pH of the samples was increased from 7.0 to 8.0. After adjusting the pH to 7.4, average values for theophylline binding to proteins at 25 degrees C ranged from 48.5 to 52.2% in serum samples from outpatients and healthy adults. Salivary theophylline concentrations correlated well with total and free serum theophylline concentrations in healthy adults and outpatients (r = 0.90-0.98). The theophylline concentration in saliva was on average about 20-30% higher than the unbound theophylline concentration in serum water. The saliva/serum concentration ratio of theophylline showed some intersubject variation (0.68 +/- 0.08; range: 0.50-0.85). Using the mean saliva/serum concentration ratio of the patient group, steady state serum theophylline concentrations were predicted from salivary levels with a mean error of 7.6% (range: 0.0-26.8%). The salivary theophylline concentration appears to be a suitable parameter for assessment of compliance, for identification of patients with inappropriate dosage, and for consequent dosage adjustment.(ABSTRACT TRUNCATED AT 250 WORDS)

The influence of 2-(3-methyl-cinnamyl-hydrazono)-propionate on the utilization of various substrates in isolated rat hemidiaphragms was investigated in comparison with other hypoglycaemic compounds. The effect of 2-(3-methyl-cinnamyl-hydrazono)-propionate was concentration-dependent. At a concentration of 0.5 mmol/l 2-(3-methyl-cinnamyl-hydrazono)-propionate, glucose utilization increased from 0.276 +/- 0.043 mumol.g-1.l-1 to 0.894 +/- 0.303 mumol.g-1.l-1 (p less than 0.05). Pyruvate and lactate utilization were stimulated to a lesser extent, while acetate utilization remained nearly constant. At a concentration of 2 mmol/l 2-(3-methyl-cinnamyl-hydrazono)-propionate, the oxidation of palmitate decreased from 0.214 +/- 0.017 mumol.g-1.l-1 to 0.060 +/- 0.005 mumol.g-1.l-1, while the oxidation of octanoate was not decreased. These findings point to a stimulation of the glycolytic flux by inhibition of long-chain fatty acid oxidation.

The main principles of heterogeneous and homogeneous enzyme-immunoassays are reviewed. Furthermore, an overview is given of enzymes currently used as labels and methods applied for conjugate preparation. The reliability and practicability of enzyme-immunoassays are discussed. Special consideration is given to possible interferences, the detection limits and the mechanization of these assays. A range of curve-fitting methods for evaluation of the results is listed. Future prospects for the further development of enzyme-immunoassay and application of this technique are discussed.

A novel quantitative liver function test is described which is based on monoethylglycinexylidide (MEGX) formation after lidocaine bolus injection. Following the administration of small single doses of lidocaine hydrochloride (1 mg/kg), monoethylglycinexylidide serum concentration-time curves were determined by a novel highly sensitive fluorescence polarisation immunoassay (FPIA) in healthy volunteers, liver donors and patients with liver cirrhosis. The FPIA allowed rapid and reliable monoethylglycinexylidide determinations in serum and urine (between-days coefficient of variation: less than 10.3%, recovery: 80-113%). Monoethylglycinexylidide concentrations measured by FPIA in 32 serum samples from patients correlated well those determined by HPLC. The monoethylglycinexylidide concentration in serum determined 15 min after a lidocaine bolus injection proved to be a highly sensitive and specific indicator of hepatic dysfunction. Average monoethylglycinexylidide concentrations in serum obtained 15 min after lidocaine injection were substantially lower in patients with liver cirrhosis than in healthy volunteers. The average monoethylglycinexylidide concentrations in serum were also substantially lower in liver donors with ballooning or fatty changes of hepatocytes than in donors without relevant alterations of liver histology. By means of monoethylglycinexylidide formation in the liver donors, primary function of the transplanted liver was correctly predicted in 32/37 cases and initial non-function in 4/6 cases.

Sodium nitroprusside was found to be a very strong inhibitor of the glucose formation from lactate in the perfused guinea pig liver. Hepatic cell respiration and oxidative phosphorylation appeared to be simultaneously suppressed by this compound. The pattern of some hepatic metabolite concentrations was similarly affected by nitroprusside as described for biguanides and pyridinium chloride derivatives. Cyanide had to be added to the perfusion medium in a tenfold higher concentration to achieve a comparable inhibition of gluconeogenesis from lactate.

In a double blind, placebo controlled, crossover study the pharmacokinetics and acute effects of enprofylline and theophylline on airway reactivity during histamine challenge were investigated in 10 healthy volunteers. The pharmacokinetic parameters of enprofylline were (mean): elimination half-life 1.9 h, total body clearance 191.1 ml X kg-1. h-1, volume of distribution 0.481 X kg-1, and protein binding 49%. Bronchial reactivity in the histamine inhalation test was expressed as the concentration causing a 20% fall in FEV1.0 (PC20). Mean PC20 values were lowest after placebo and highest after theophylline with the enprofylline values in between. Only the difference in PC20S after placebo and theophylline was statistically significant (p less than 0.05). At the time of determination of the PC20, the serum concentration of enprofylline was between 16.5 and 11.8 mumol/l, and that of theophylline was between 78.3 and 61.1 mumol/l. Adverse actions of enprofylline were nausea (3/10) and cardiovascular reactions (2/10), whereas theophylline mainly caused restlessness (3/10) and tremor (2/10). Thus enprofylline, in one-fifth of the serum concentration of theophylline cannot be regarded as equipotent in terms of bronchoprotection.

In a double-blind cross-over study, the effects of slow release (S-R) terbutaline tablets (b.i.d. 0.25 mg/kg per day) and S-R theophylline (5.31 mg/kg morning and 10.62 mg/kg evening) were compared in eleven patients with nocturnal asthma. On day seven of each treatment period, drug serum concentrations and peak expiratory flow (PEF) were measured every 2h over a 24-h period. During daytime, terbutaline concentrations ranged from 1.6-14.1 (median 4.5) microgram/l and during the night from 2.1-18.7 (median 4.9) micron/l. Theophylline concentrations ranged from 3.9-24.3 (median 11.5) mg/l during the day and from 3.3-20.9 (median 10.4) mg/l at night. Nocturnal wheezing occurred during theophylline treatment in four patients 7 times and during terbutaline treatment in six patients 22 times. Daytime PEF values were 472 +/- 161 l/min during theophylline therapy versus 445 +/- 169 l/min during terbutaline therapy (p less than 0.05). In the night and early morning there was no significant difference between PEF values with the two treatment forms. During theophylline treatment, fewer inhalations of beta 2-sympathomimetics were used, and there were fewer side effects. One patient experienced severe asthmatic attacks during the terbutaline treatment period. The patients preferred theophylline for the treatment of nocturnal asthma.

An evaluation of an enzyme immunoassay for the determination of theophylline in serum (EMIT, Syva Corp.) is presented. The EMIT assay was fully mechanized by the use of an Eppendorf analyzer 5010. The precision of this enzyme immunoassay was satisfactory (coefficients of variation from day to day ranged from 5--6%). The EMIT assay proved to be specific, rapid and easy to perform. A comparison of the results obtained by EMIT and ultraviolet spectrophotometry in a series of 100 patients showed a good correlation between both methods.

A novel Bayesian drug dosing program (Abbott Pharmacokinetic Systems, Theophylline Program) was evaluated. The predictive accuracy of this method was assessed in 10 healthy volunteers receiving single intravenous test doses. Estimates for clearance and distribution volume were compared with those obtained from the area under the curve. The observed prediction error depended largely on sampling time. The deviations were lowest for the distribution volume during the first 60 min and for clearance at 12 hours after theophylline administration. Furthermore the Bayesian technique was prospectively evaluated in 10 hospitalized and 22 outpatients treated with sustained-release theophylline preparations (Uniphyllin, Bronchoretard, PulmiDur). Predictive precision and accuracy were adequate, if theophylline was given twice daily. The highest predictive accuracy was achieved in outpatients, if predictions were based on trough concentrations. In 19/22 outpatients prediction errors were within a clinically acceptable range (mean prediction error +/- standard deviation; -0.6 +/- 2.1 mg/l). Moreover in hospitalized patients (n = 5) with twice-daily maintenance regimens, concentration-time curves could mainly be predicted with sufficient accuracy. Hospitalized patients (n = 5) with once-daily dosing showed large fluctuations between peak and trough theophylline concentrations in serum. In these patients a reliable prediction of the concentration-time curves was not possible apparently due to non-linearity of theophylline kinetics. Relatively large prediction errors were found in one patient with acute viral respiratory illness and 3 patients with altered absorption. Despite certain limitations the clinical application of the Bayesian forecasting method tested appears to be promising.

A heterogeneous enzyme immunoassay for the determination of thyroxine binding globulin (TBG) was developed and assessed in clinical trials in 12 laboratories. The assay is based on the competition principle and employs plastic tubes coated with goat anti-TBG. CV's between 1.4-8.9% for intra-assay precision and 2.9-8.6% for inter-assay precision were found over the concentration range of 4-40 mg/l TBG. In comparative studies using highly purified TBG as standard, values with Enzymun-Test TBG were found to be on average 30% lower than those obtained by various TBG-RIAs. A broad-base study, to determine reference values, was carried out on a group of control persons 18 to 50 years old without previous history of thyroid disease. This study revealed a median of 14.33 mg/l TBG, with 95% of all values between 9.6 and 18.5 mg/l TBG. The median in women of 14.5 mg/l TBG was significantly higher than in men (13.4 mg/l TBG). TBG values in hyperthyroid patients were within the reference range while those in hypothyroid individuals were elevated. Highly elevated TBG values were seen in women receiving oestrogen (median: 22.2 mg/l TBG) and in pregnant women (median: 28.5 mg/l TBG). The T4/TBG ratios made it possible to distinguish between euthyroid, hyperthyroid and hypothyroic subjects (median: 4.9, 11.3 and 1.0, respectively). These ratios were significantly lower in pregnant women (median: 3.1) than in the control persons.

This paper summarises the results of the evaluation of a heterologous enzyme immunoassay in antibody coated tubes for the determination of triiodothyronine (T3) by a group of 11 laboratories. The assay procedure is analogous to an established heterologous thyroxine enzyme immunoassay. The evaluation demonstrated that 1. the specificity of the assay and its analytical range between 0.46 and 9.2 nmol/l meet diagnostic requirements, 2. the intra- and interassay precision was in accordance with that commonly found in radio immunoassays, 3. the recovery of added T3 was between 90 and 103%, 4. the results agreed well with those of self-established radio immunoassays and were comparable with commercial RIA kits, 5. there was no interference by hyperbilirubinaemia or by high concentrations of bile acids, but interference can occur in very haemolytic and lipaemic samples. The great advantage of T3 enzyme immunoassays lies in the absence of restrictions and any authorization needed for working with radioactive substances.

2-(3-Methyl-cinnamyl-hydrazono)-propionate (MCHP) is a new compound which effectively lowers the blood glucose level in guinea pigs. 0.04 mmol/l MCHP inhibited glucose formation from lactate, pyruvate and alanine, but not from dihydroxyacetone in the perfused guinea pig liver. In the presence of both hexanoic acid and alanine, 0.04 mmol/l MCHP did not effect hepatic gluconeogenesis. Gluconeogenesis was probably reduced by an inhibition of the pyruvate carboxylase: The pattern of hepatic metabolite concentrations indicated a block between pyruvate and the triose phosphates. The intrahepatic concentration of acetyl-CoA was consistently decreased. The decrease of the acetyl-CoA concentration could be explained by an influence of MCHP on the fatty acid oxidations, which occurred at the carnitine palmitoyl transferase (CPT) step. This hypothesis is supported by the fact that a medium chain fatty acid not requiring CPT reversed the inhibition of gluconeogenesis by MCHP. From all results reported it is concluded that transamination and oxidative phosphorylation were not effected to a degree relevant for the inhibition of gluconeogenesis by 0.04 mmol/l MCHP.

This paper describes the evaluation of a heterologous enzyme immunoassay for the determination of total thyroxine in serum by a group of seven clinical chemical laboratories. The test follows the principles of the enzyme linked immunosorbent assay (ELISA) and uses peroxidase as a marker. The evaluation of analytical reliability yielded the following results within the analytical range from 39 unto 322 nmol/l: 1. Within-batch precision ranged from 3.1 unto 10.4% (coefficient of variation) with single analyses. 2. Between-batch precision ranged from 3.7 unto 20.4% with single analyses. 3. Between-laboratories precision ranged from 5.4 unto 6.8%. 4. Pure thyroxine, added to serum or thyroxine-free serum, gave recoveries between 93 and 120%. 5. Analysis of control sera gave results essentially comparable to the assigned values based upon radioimmunoassays. 6. Analysis of 288 clinical sera gave slightly higher results by the enzyme immunoassay than by the analogous radioimmunoassay from the same manufacturer. 7. Comparison with other methods of analysis (radioimmunoassays, competitive protein ligand assays, hormonal iodine assay) yielded partly comparable, partly higher results. 8. Comparison with the homogenous enzyme immunoassay (EMIT) led to comparable results. 9. Interference due to hyperlipemia or hemolysis was not observed. 10. There might be an interference in hyperbilirubinaemic sera, due to an as yet unknown factor. With respect to practicability the ELISA-test compares favourably with the analogous solid phase radioimmunoassay. The main differences are the absence of radioactive material and a longer shelf-live of reagents. Following the manual procedure the time taken to perform the enzyme immunoassay is slightly longer than for the analogous radioimmunoassay.

The rate of mitochondrial carnitine-carnitine exchange mediated by carnitine acylcarnitine translocase was measured by following the uptake of L-[methyl-14C]carnitine. It was demonstrated that the hypoglycaemic compound 2-(3-methyl-cinnamyl-hydrazono)-propionate causes a concentration-dependent decrease in the rate of the translocase-mediated transport of carnitine in guinea pig liver mitochondria. Apparent initial influx rates were decreased by 20% at 0.3 mmol/1 2-(3-methyl-cinnamyl-hydrazono)-propionate, 38% at 0.5 mmol/l, and 75% at 2.0 mmol/l of this compound. This finding may explain the previously observed inhibitory effects of this substance on long-chain fatty acid oxidation, ketone body production and gluconeogenesis.

Dosage prediction methods for optimization of aminoglycoside therapy were evaluated in 19 critically ill patients. Two different three-point methods and a novel Bayesian method were used. The predictive accuracy and precision of all tested methods were clinically satisfactory. The time needed for the necessary calculations was 10-15 min. After administration of predicted dosages, approximately 90% of the observed peak and trough aminoglycoside serum concentrations fell within the therapeutic range. Without systematic use of these prediction methods, only approximately 40% of tobramycin peak serum concentrations and 80% of trough serum concentrations were within the therapeutic range. Daily dosage adjustments particularly in patients with rapid changes of aminoglycoside clearance are greatly facilitated by use of these prediction methods. These findings suggest that all of the dosage prediction methods tested are well suited for optimization of aminoglycoside dosage regimens.

A gas chromatographic procedure for the simultaneous determination of carbamazepine, phenobarbital and phenytoin using SP 2510 DA as stationary phase is presented. The antiepileptic drugs are determined simultaneously without derivatisation under isothermal conditions by a flame ionisation detector. The gas chromatographic procedure can be easily mechanized. The coefficient of variation for the precision from day to day is 8.9% for carbamazepine, 7.0% for phenobarbital and 4.3% for phenytoin as calculated from single determinations. The deviations from the target value of spiked pool sera range from 4.3 to 9.5%. The gas chromatographic results of this method and of determinations by Dexsil 300 for primidone and SP 1000 for ethosuximide are compared with the corresponding enzyme immunoassays (EMIT). The precision of the gas chromatographic methods is somewhat better than EMIT. When patients' sera are analysed by both procedures the results show no clinically relevant differences. It can be concluded that the different methods are interchangeable and may be selected according to practical necessities.

The hydrazone-compound 2-(3-methyl-cinnamylhydrazono)-propionate (MCHP) significantly lowered the blood glucose concentration in fasted guinea pigs and rats. A significant decrease of blood glucose levels was observed in fasted guinea pigs already after an intraperitoneal injection of 20.5 mumol/kg MCHP, while much higher doses (about 1000 mumol/kg) were necessary to produce a hypoglycemic effect in the fasted rat. After oral administration MCHP (82.0 mumol/kg) significantly decreased the blood glucose concentration in guinea pigs. Furthermore MCHP caused a dose-dependent increase of plasma free fatty acid concentrations in guinea pigs and rats. In addition, MCHP decreased the concentrations of blood ketone bodies, plasma cholesterol and intrahepatic acetyl-coenzyme A in the guinea pig. All of these findings appear to be due to a reduced fatty acid utilization in the presence of MCHP resulting presumably in an intramitochondrial deficiency of acetyl-CoA. At hypoglycemic effective doses the intramitochondrial and cytoplasmatic redox ratios as well as the hepatic ATP/ADP ratio were not influenced by MCHP in fasted guinea pigs. Even at large doses (123 mumol/kg) MCHP decreased the activity of monoamino oxidase in guinea pigs only by less than 15%. Furthermore MCHP showed under our experimental conditions no relevant influence on the activity of various liver enzymes in plasma, the plasma concentration of creatinine, the plasma triglyceride-glycerol level and on the intrahepatic triglyceride-glycerol concentration of fasted guinea pigs. It is concluded that MCHP meets basic requirements for a potential oral antidiabetic agent.