Calibrators and control samples for bilirubinometers.

The different matrix properties of neonatal serum and commercial control samples can lead to considerable errors in the calibration and control of bilirubinometers. These difficulties can be avoided by calibration with serum from healthy adults which is supplemented with unconjugated bilirubin. But this procedure is impracticable for most routine laboratories. Under certain preconditions, control samples, with bilirubin concentrations determined with correctly calibrated bilirubinometers or spectrophotometers, are also suitable as calibrators. This was established by determination of the bilirubin concentration of 16 different control samples, using both the reference method and correctly calibrated bilirubinometers or spectrophotometers in three or four specialist laboratories. This was also confirmed in several interlaboratory surveys, some involving up to 72 laboratories. The results of these investigations show that a control sample should be used for the calibration of a bilirubinometer only if it meets the following preconditions: 1. There should be no significant difference between the bilirubin values determined with the reference method and with a correctly calibrated spectrophotometer or bilirubinometer. 2. The bilirubin concentration should lie in the range 230-300 mumol/l. The photometric response of bilirubinometers has a limited linear range, so that analytical results greater than 300 mumol/l must be rated as basically unreliable.

Constitutional delay of growth and adolescence. Results of short-term and long-term treatment with GH.

During recent years numerous reports on the favourable results of short-term trials with GH in patients with constitutional delay of growth and adolescence (CDGA) have been published, but it has been unclear whether such treatment affects final height. In the present study, the results of long-term therapy with GH in replacement doses have been evaluated in 15 patients who were treated with GH for several years (three years on average). At the start of treatment, 10 of the children were prepubertal and 5 were in puberty. All patients were followed up until final height was reached. Mean final height of the 13 male patients was 170.0 +/- 4.4 cm, i.e. -1.58 SDS. In the two female patients, final height was 150.0 cm (-3.5 SDS) and 164.0 cm (-0.8 SDS), respectively. Adult height of the patients lagged behind target height by 5.4 +/- 3.2 cm (mean +/- SD). Measured adult height corresponded to adult height as predicted prior to treatment. In conclusion, GH treatment of patients with CDGA did not increase final height.

Calibration of direct reading photometers for the determination of neonatal bilirubin.

Blijenberg et al. (J. Clin. Chem. Clin. Biochem. 25, 737-741 (1987) (1)) proposed that the assigned values of calibrators for the determination of bilirubin with simple direct reading photometers (bilirubinometers or "bilimeters") should be determined on the same type of instrument. For this purpose, the instrument is calibrated with neonatal serum, whose bilirubin concentration is determined with a reference method unbiased by matrix effects. In the present study, this procedure was developed and tested. Bilirubin was determined in capillary plasma (n = 30) and venous serum (n = 31) of newborns. Measurements were made by direct spectrophotometry in 4 bilimeters, and by a micromodification of Vink's spectrophotometric method after dilution with caffeine reagent (Vink et al. Clin. Chem. 34, 67-70 (1988) (2)). When the bilimeters were calibrated with a commercial control serum, using the concentrations quoted by the manufacturer, the results were higher by 13% (plasma) and 19% (serum) than those obtained by Vink's method. These discrepancies practically disappeared when the bilimeter results were recalculated using a calibrator concentration determined according to Blijenberg. Application of the proposed method to other control sera gave assigned values up to 23% lower than those stated by the manufacturers. After calibration of the bilimeters with various neonatal sera (specimens drawn for therapeutic purposes and residues, obtained from the clinical laboratory), differences of about 10% were observed between the assigned values. It is therefore essential to use several sera. Minor differences were found between bilimeters.(ABSTRACT TRUNCATED AT 250 WORDS)

[Spontaneous secretion of growth hormone in deep nocturnal sleep. II. Studies on hypophyseal dwarfism and constitutional developmental delay]. / Spontansekretion des Wachstumshormons im nächtlichen Tiefschlaf. II. Untersuchungen bei hypophysärem Minderwuchs und bei konstitutioneller Entwicklungsverzögerung.

To evaluate the somatotropic function of the pituitary, the measurement of the spontaneous nocturnal secretion of GH is a rather suitable method. Whereas the provocation tests check the capacity of the gland after intensive stimulation, spontaneous secretion reflects the behaviour of the hypothalamohypophyseal system under everyday conditions. The investigation of 65 children with pituitary dwarfism showed in all cases a strong diminution of the GH secretion. Overlapping with the control group was hardly seen. The maxima measured during sleep were identical with those reached in the provocation tests. The subdivision in complete and partial GH deficiencies is more precise with measuring the spontaneous secretion than with provocation tests. Nocturnal spontaneous GH secretion was determined in 128 patients with constitutional delay of growth and adolescence. Also in these children provocation tests were performed simultaneously. Spontaneous GH secretion was found significantly diminished in all stages of puberty, compared to the controls (p less than or equal to 0.01). By contrast, the provocation tests showed no significant differences from controls. According to these results, the retarded growth of the patients is due to a relative GH deficiency. As is evident from the normal results of the provocation tests, no organic insufficiency of the pituitary is demonstrable. Rather a cybernetic disorder is responsible for the reduced hormone secretion.

Testing with growth hormone-releasing factor (GRF(1-29)NH2) and somatomedin C measurements for the evaluation of growth hormone deficiency.

Growth hormone (GH) responses to GRF (1 microgram/kg BW i.v.) were investigated. Comparison between GRF(1-40) and GRF(1-29)NH2 in 11 young adult volunteers gave identical results. One hundred and thirty-one children and adolescents (45 with idiopathic GHD) were tested with GRF (1-29)NH2. The maximal GH levels (max GH) in response to GRF during the 120 min test period were found suitable to characterize the response. In cases without GHD no correlation to age, sex and pubertal development was observed. A maximal GH level of above 10 ng/ml was found to be normal. In 3 out of 86 children without GHD (one with Turner syndrome; two with simple obesity) max GH fell short of 10 ng/ml, while 11 of 45 cases with GHD exceeded this margin. In GHD, max GH was inversely correlated with age. There was no difference in max GH between groups with or without perinatal pathology as a presumed cause of GHD. GH levels to GRF were positively correlated with maximal GH level during sleep in GHD, but not correlated with responses seen to insulin or arginine. The value of GRF testing for the confirmation of GHD is discussed in the light of other GH stimulatory tests and basal somatomedin C measurements. It is suggested that the combination of testing with GRF and the determination of a basal SmC level offers a safe and convenient way to diagnose GHD in clinically suspected cases, though in some cases further diagnostic tests may be needed.

Chlorinated hydrocarbons in adipose tissue of infants and toddlers: inventory and studies on their association with intake of mothers' milk.

Chlorinated hydrocarbon and polychlorinated biphenyl (PCB) concentrations were determined in adipose tissue from 34 infants, 14 children in the 2nd year of life, and 2 older children. The highest mean concentration detected during the first 2 years of life was for PCBs (0.67 ppm), followed by DDT (0.57 ppm), HCB (0.23 ppm), and HCH (0.15 ppm). Concentrations of HCB and PCB, which are especially characteristic of highly industrialised countries, were considerably higher in children of German mothers than in those of Turkish mothers. All single investigated values were lower than the mean values for adults in the Federal Republic of Germany, but many were still higher than mean concentrations for adults in other parts of the world. A breakdown into children with high mothers' milk intake and those with low intake showed a highly significant association with the quantity of mothers' milk consumed: the concentration of organohalogens in adipose tissue of children with high intake was significantly higher than in those with low intake. Two tasks urgently demand our attention: the development of further ways to reduce environmental sources of organohalogen contamination and the study of the possible pathogenetic effect of these organohalogens on the health of our children.

Sulphated bile acids in duodenal juice of healthy infants and children compared with sulphated bile acids in paediatric patients with various gastroenterological diseases.

The bile acid pattern in the first 10 minute fraction of intestinal juice after injection of pancreozymin (2 U/kg bw) was studied in 32 gastroenterologically healthy children and 26 paediatric patients with various diseases of the liver or intestine. The sulphated and non-sulphated portions of monohydroxy, dihydroxy, and trihydroxy bile acids were evaluated in relation to the total concentration of bile acids as well as the ratios of the individual acids to each other in esterified and unesterified form. In healthy patients, the extent of sulphation was not age dependent. The quantity of potentially toxic monohydroxy and dihydroxy bile acids did not determine the percentage of sulphation, which was induced only by the severity of cholestasis. Ursodeoxycholic bile acids were usually esterified with sulphuric acid to a remarkably high degree. Less sulphated bile acids were detected in patients with coeliac disease than in the control group.

A quantitative assay for the cytoplasmic androgen receptor using [3H]dihydrotestosterone in the presence of NAD+-nucleosidase.

In prostatic cytosol DHT1 is metabolized to 5 alpha-androstane-3 alpha (or beta), 17 alpha-diols with a half life of 2 h even at 4 degrees C. Thus, [3H]DHT appears to be a poor marker for a quantitative assessment of androgen receptors (AR). Methyltrienolone (R1881) seems to be advantageous as it is not metabolized. However, because of considerable binding to progestin receptors, assays using [3H]R1881 are not specific for AR in tissues containing progestin receptors. We, therefore, developed a specific assay for AR using [3H]DHT (14 nM) as marker, where metabolism of DHT is prevented by pre-incubation with NAD+-nucleosidase. The [3H]DHT-receptor complex is separated from free, SHBG-bound and unspecifically bound [3H]DHT by agar gel electrophoresis. The binding sites of high affinity and low capacity are characterized by suppression with unlabelled R1881 (2 microM) in a parallel assay. Under these conditions DHT and R1881 appear to have the same kinetics of association and dissociation. Weighted non-linear regression analysis of specific binding capacity at various ligand concentrations reveals that in rat prostatic cytosol the affinity of DHT (Kd = 0.405 +/- 0.0839 nM) is significantly higher (P less than 0.01) than that of R1881 (Kd = 1.25 +/- 0.271 nM).

[Gas chromatographic determination of phenytoin, phenobarbital and primidone: flash-methylation after direct addition of trimethyl-phenyl-ammonium hydroxide to the ethyl acetate extract (author's transl)]. / Gaschromatographische Bestimmung von Phenytoin, Phenobarbital und Primidon: Methylierung im Einspritzblock nach direktem Zusatz von Trimethyl-phenyl-ammonium-hydroxid zum Ethylacetat-Extrakt.

25 microliters plasma, 25 microliters internal standard and 5 microliters sodium dihydrogenphosphate solution are extracted with 100 microliters ethyl acetate. 40 microliters of the extract are mixed with 10 microliters trimethyl-phenyl-ammoniumhydroxide (0.1 mol/l methanol). 3 microliters of this solution are injected for gas chromatography using a nitrogen-specific detector. Trimethyl-phenyl-ammoniumacetate appears to be a satisfactory methylating agent for these drugs in the injection port of the gas chromatograph.

Comparison of phenytoin determinations in plasma, plasma dialysate and saliva for control of antiepileptic therapy in children.

Phenytoin was determined in plasma or serum, dialysate from plasma or serum and saliva in 45 children and 7 adults by gas chromatography. 200 microliter of plasma were necessary for dialysis. Close correlations were found between phenytoin concentrations in the different media (r greater than or equal to 0.95). Since in some cases the levels in saliva were extremely high in comparison to plasma and dialysate, precautions are advisable when using phenytoin determinations in saliva for therapeutic decisions. Clinical data from 15 children with high phenytoin levels suggest that there is no advantage in determining the unbound fraction of the drug in plasma for the control of antiepileptic therapy.

[The application of a combined Xylose-lactose tolerance-test in children with suspected malabsorption (author's transl)]. / Xylose-Laktose-Belastungstests bei Kindern. Die Anwendung des kombinierten Xylose-Laktose-Belastungstests bei Kindern mit Verdacht auf Resorptionsstörungen

A combined xylose-lactose tolerance-test and a duodenal biopsy were performed in 68 children with suspected malabsorption-syndrome. The purpose of the present work was to assess the diagnostic value of xylose concentrations in blood at different times and to determine the additional discriminatory value to glucose levels. The contribution of the glucose rise to a calculated discriminant function is statistically significant but practically negligible and therefore does not justify its determination. A small-bowel biopsy is recommended if the concentration of xylose after 60 min is less than 26 mg/100 ml or the increment of xylose concentration above fasting level is 18 mg/100 ml or less.

[The difficulties to diagnose a clinical important defect of disaccharidases (author's transl)]. / Die Schwierigkeiten der Diagnostik eines klinisch-relevanten Disaccharidasen-Mangels

Duodenal mucosa of 100 infants and children was histologically investigated and the activities of dissaccharidases in the mucosa homogenate were determined. In addition, a combined xylose-lactose loading test was carried out and the behaviour of the stools under the test conditions was observed. The results were compared with regard to the clinical importance of an enzyme defect. The rise of xylose in blood gives no evidence for a defect of disaccharidases in the mucosa. The determination of the glucose maximum after a loading test with disaccharides seems rather to lead to false-normal results. The possiblity of a false-pathological interpretation by an exclusive determination of enzymes must be prevented by further gastroenterological examinations.

[Disaccharidases of the small intestine mucosa in infants and children. "Normal values", log normal distribution and age dependence]. / Disaccharidasen der Dünndarmschleimhaut bei Säuglingen und Kindern -- "Normalwerte", Lognormalverteilung und Altersabhängigkeit--

In 63 infants and children with a histological normal mucosa of the duodenum, without an isolated defect of enzyme and with a normal increase of xylose and glucose in serum after a combined xylose-lactose loading test the activities of disaccharidases were log normal distributed. The asymmetric distributions were transformed into symmetric ones and the geometric mean (x) as well as the range (+/- 2 s) of maltase, saccharase, isomaltase, lactase and trehalase were calculated. Only the activity of lactase shows a significant dependency on age. In the first year of age the lower limit (x -- 2 s) of this enzyme is much higher than later.

[Hereditary deficiency of isomaltase and saccharase responsible for a malabsorption syndrone (author's transl)]. / Hereditärer Isomaltase-Saccharase-Mangel der Dünndarmschleimhaut als Ursache für ein Malabsorptionssyndrom

Among 135 infants and children with a supposed malabsorption syndrome, a deficiency of isomaltase-saccharase of the duodenal mucosa was detected in 5 cases by measuring the disaccharidases directly in the mucosa homogenate. In one instance a deficiency of lactase was found in addition. In all patients the villi were of normal length, with an increased cell infiltration of the stroma detected in two cases. The loading tests with xylose-sucrose yielded a diminuished rise in the blood glucose level. Three of the patients were dwarfish, but only one showed an increased growth after the reduction of sucrose in the supplied diet. As a result of adaptation difficulties in the change of diet, one patient had to be treated with an additional saccharase substitution.