Non-ketotic C6-C10-dicarboxylic aciduria: biochemical investigations of two cases

Two boys, who are not related, with hypoglycemia and C6-C10-dicarboxylic aciduria were investigated. Besides substantial amounts of adipic, suberic and sebacic acids, the urinary metabolic profile of organic acids contained 5-OH-caproic acid and caproylglycine. During acute attacks the concentrations of adipic, suberic and sebacic acids were 300--530, 160--200 and 35--200 micrograms/mg creatinine, respectively, and the excretions of 5-OH-caproic acid and caproylglycine were 75--330 and 41--260 micrograms/mg creatinine, respectively. It is argued that the biosynthesis of adipic acid passes through an omega-oxidation, that the production of 5-OH-caproic acid is caused by an omega-1-oxidation, and that caproylglycine formation passes through a glycine-N-acylase catalysed conjugation of accumulated caproic acid in the patients. Suberic acid and sebacic acid are in the same way omega-oxidation products of accumulated caprylic acid and capric acid, respectively. From the excretion pattern presented it is hypothesized that the patients suffer from a defect in the dehydrogenation of fatty acids in the beta-oxidation pathway. The biological significance of the findings is discussed.     

Urinary C6-C12 dicarboxylic acylcarnitines in Reye's syndrome

C6-C12 dicarboxylic acylcarnitines have been identified for the first time in urine from a 2-year-old girl presenting with Reye's syndrome. The acylcarnitines were extracted by ion-exchange chromatography and analysed, both underivatised and as methyl esters using high-resolution fast-atom-bombardment mass spectrometry and B/E-linked scanning. The acylcarnitines were quantified by capillary gas chromatography of the acids extracted after hydrolysis of the acylcarnitine esters. Dodecandioylcarnitine was present in the highest concentration (35.9 mmol/mol creatinine) which exceeded the urinary free dodecandioic acid concentration. The adipic, suberic and sebacic acylcarnitine concentrations were less than 10% of the respective free acid concentrations. It is possible that beta-oxidation of dicarboxylic acids is partially inhibited in Reye's syndrome leading to accumulation of precursor dodecandioyl CoA which is metabolised to dodecandioylcarnitine. The accumulation of these metabolic intermediates may be significant in the pathogenesis of Reye's syndrome.     

In vitro fibroblast studies in a patient with C6-C10-dicarboxylic aciduria: evidence for a defect in general acyl-CoA dehydrogenase

Fatty acid oxidation studies were performed on cultured fibroblasts from a patients who had suffered from a Reye's syndrome-like episode, and who excreted adipic acid, suberic acid, sebacic acid, 5-OH-hexanoic acid, and hexanoylglycine. Assays using fatty acids of different chain length on both intact and disrupted cells indicated that the patient had a defect of the general (medium-chain) acyl-CoA dehydrogenase apoenzyme.     

Glutaric aciduria type II: report on a previously undescribed metabolic disorder.

A report is given on a hitherto undescribed metabolic disorder, characterized clinically by fatal neonatal acidosis, hypoglycemia and a strong 'sweaty-feet' odour. Biochemical features were a massive urinary excretion of glutaric and lactic acids. Isobutyric, isovaleric and alpha-methylbutyric acids were also greatly increased, followed by adipic, ethylmalonic, alpha-hydroxybutyric, n-butyric, beta-hydroxybutyric, sebacic, suberic, propionic, alpha-hydroxyisovaleric and hexanoic acids. The serum level of glutaric acid was highly elevated. In the serum there were also abnormal levels of lactic, alpha-hydroxybutyric, adipic, suberic, p-hydroxyphenyllactic, myristic, hexadecenoic, palmitic, oleic and stearic acids. Plasma lysine and valine were also elevated. Degradation of 14C-labelled glutaric acid and 14C-labelled branched-chain amino acids, alpha-ketoisovaleric and alpha-ketoisocaproic acids in intact fibroblasts was decreased, whereas that of pyruvic acid was normal. The defect was tentatively supposed to be localized at the level of the metabolism of a range of acyl-CoA compounds. The name glutaric aciduria 'type II' is proposed for the patient's disease.     

Dicarboxylic aciduria: the response to fasting.

The urine of a child who presented with an episode of a disease resembling Reye's syndrome was found to contain large quantities of the dicarboxylic acids adipic and suberic acids, as well as the glycine conjugate of suberic acid, suberyl glycine. A variety of other dicarboxylic acids, both saturated and unsaturated, were also found in the urine at the time of the attack. It was found that the excretion of these unusual metabolites could be markedly increased by fasting for periods of greater than 10 h. These results indicate that the patient may have a defect in fatty acid oxidation which becomes clinically significant during periods of prolonged fasting.     

General (medium-chain) acyl-CoA dehydrogenase deficiency (non-ketotic dicarboxylic aciduria): quantitative urinary excretion pattern of 23 biologically significant organic acids in three cases

Urinary analysis of the pattern of 23 organic acid metabolites derived from fatty acids in three patients with general (medium-chain) acyl-CoA dehydrogenase deficiency was performed. Although there exist quantitative differences in the excreted amounts of the different metabolites in the three patients the qualitative picture was the same. The excretion of adipic, suberic and sebacic acids was substantial, whereas that of dodecanedioic acid was within or just above control limit. The monounsaturated C6-C10-dicarboxylic acid excretion was only marginally or not increased. 5-OH-hexanoic acid and hexanoylglycine were excreted in excessive amounts, whereas 7-OH-octanoic acid, 9-OH-decanoic acid, octanoylglycine and decanoylglycine were excreted in limited amounts. The excreted amounts of 6-OH-hexanoic, 8-OH-octanoic and 10-OH-decanoic acids were not or only marginally elevated compared to controls. In one of the patients the excretion of ethylmalonic and methylsuccinic acids was enhanced, whereas the excretion of these two acids in the two other patients was comparable to that in controls. The urinary excretion of hexanoic, octanoic, decanoic and dodecanoic acids was just a little above the control limit, whereas the esterified hexanoic and octanoic acids were excreted in appreciable amounts. It is argued that the microsomal omega- and omega-1-oxidation systems are involved in the dicarboxylic and omega-1-OH-monocarboxylic acids formation at C10 and C12 level and that the C8-C6-dicarboxylic and omega-1-OH-monocarboxylic acids are formed from higher chained acids by beta-oxidation in both mitochondria and peroxisomes.     

Bile acids and their sulphated and glucuronidated derivatives in bile, plasma, and urine of children with intrahepatic cholestasis: effects of phenobarbital treatment

Abstract. Bile acids and their sulphated and glucuronidated derivatives were studied in three children with persistent intrahepatic cholestasis, two children with intrahepatic biliary hypoplasia, and four healthy children. In children with cholestasis, biliary bile acids consisted of 11(±0–3) % 3 β-hydroxy-delta-5-cholenoic acid, 2-1(± 0–6) % lithocholic acid, 2-2(± 11) % deoxy-cholic acid, 5–8(±2-2) % ursodeoxycholic acid, 39-1(± 1 -4) % chenodeoxycholic acid, 0–5(± 0 2) % hyo-cholic acid, and 49-3(± 3 0) % cholic acid. Of these bile acids 121 (±l 9) % were sulphated and 4–5 (±0 6) % were glucuronidated. In healthy children, biliary bile acids consisted of 0–7 (±0–4) % lithocholic acid, 3–4 (±0 8.) % deoxycholic acid, 0–1 (±0 1) % ursodeoxycholic acid, 32-7 (±6 9) % chenodeoxycholic acid, and 631 (±7 1) % cholic acid. Of these bile acids, 0–6±0 1 % were sulphated and 0–2 ±0 1% were glucuronidated (mean ± SEM). In the urine of healthy children, 3-3(±0 6) mg/24 h bile acids (1–5±0 3 mg sulphates and 0–1 ±0 1 mg glucuronides) were excreted, in the urine of children with cholestasis 61-4 (± 10 2) mg/24 h (30 2 ±7 1 mg sulphates and 5 6 ±1 2 mg glucuronides) were excreted. Thus in children with cholestasis the amounts of sulphated and glucuronidated bile acids are greater than in healthy controls. Substantial amounts of sulphated and glucuronidated bile acids are excreted in bile and urine of these patients. Phenobarbitone treatment in the five children with cholestasis led to a reduction of serum bile acids from 90 4 (± 13 2) μg/ml to 39 3(±3 6) μ//ml, a relative increase of bile acid glucuronides in bile from 45 (±0 6)% to 8 l(±0 6)%, a slight alteration of the bile acid sulphates in bile from 121(±l 9) % to 111 (± 1 2)% and no alteration of the bile acid spectrum. Urinary excretion of bile acids decreased from 61 4 (± 10 2) mg/24 h to 34 7(±3 0) mg/24 h. Phenobarbitone treatment of children with cholestasis thus induced glucuronidation of bile acids but had no significant effect on sulphation or on formation of individual bile acids.     

Urinary albumin determination by the immediate bromcresol green method

It has been demonstrated recently that the reaction of serum samples with bromcresol green (BCG) reagent proceeds in two steps. Albumin is responsible for the immediate reaction while other serum proteins produce the slow reaction.

In this paper the immediate BCG reaction has been used for the determination of urinary albumin concentration in patients with proteinuria by a slightly modified method with a primary pH adjustment of the urine and the use of a urine blank.

Comparison of the immediate BCG method (γ) with Laurell “rocket” technique (χ) gave the following equation: γ = 17.2 +1.006χ (n = 98; r = 0.99) mg/1. The coefficient of variation (within-day), C.V. (%), ranged between 0.9 and 2.7% depending on the albumin concentration. It is thus possible to carry out rapid, accurate and precise albumin determinations in urine samples using this simple method.     

Metabolism and pharmacokinetics of medium chain fatty acids after oral administration of royal jelly to healthy subjects

The unique fatty acids in royal jelly (RJ), 10-hydroxy-2-decenoic acid and 10-hydroxydecanoic acid are expected to be associated with many health benefits, but little is known on the pharmacokinetics and metabolism. The aim of this study is to confirm the metabolism and pharmacokinetics of RJ fatty acids in humans. Twelve volunteers received RJ capsules or enzyme treated RJ (ETRJ) capsules (800 mg). The other group received two doses of ETRJ tablets (800 mg and 1600 mg). Plasma samples were collected up to 12 h after the RJ intake and urine samples were collected within 24 h after ETRJ tablet consumption. The samples were analyzed by LC/MS/MS. A multivariate analysis of the RJ dose plasma samples detected 2-decenedioic acid (2-DA), sebacic acid (SA), and 3-hydroxysebacic acid (3-HSA) with significantly different intensities (P < 0.05) before and after RJ intake. The area under the concentration (AUC) of 2-DA, SA, and 3-HSA was 2500.05 ± 569.58, 322.57 ± 137.36, and 242.98 ± 58.36 ng h mL⁻¹, respectively. By enzyme treatment, the AUC of 2-DA, SA, and 3-HSA was significantly increased (P < 0.05). The values of AUC and urinary excretion of these metabolites were dose-dependent. The major RJ fatty acids were metabolized to dicarboxylate, absorbed into the circulation and their absorption increased by enzyme treatment. This study provides useful information that will support studies aimed at clarifying the identity of bioactive RJ constituents and their biological effect, and further the development of RJ.

The royal jelly fatty acids were metabolized to dicarboxylic acids, absorbed into the circulation and then excreted in the urine.     

Effet chez l'homme de l'interruption partielle du cycle entéro-hépatique sur les concentrations biliaires du cholestérol et des sels biliairesEffect of the partial interruption of the enterohepatic cycle on the biliary concentration of cholesterol and bile salts

The concentration of bile cholesterol (Sperry and Webb), cholic and dihydroxy-cholanic acids (reversed phase partition chromatography) was studied in 14 cholecystectomized patients with a T-tube in the common bile duct, during 9 to 23 days (9 patients with gallstones, 5 without stones). The patients were classified in:

Group 1: daily volume of drained bile 230 ml;

Group 2: daily volume of drained bile 450 ml. The age, sex distribution, duration of drainage, and daily alimentary intake were not statistically different between the two groups.

The bile cholesterol concentration (14 patients, 196 daily bile samples), the total bile salts concentration and the ratio dihydroxycholanic acids/cholic acid (8 patients, 110 daily bile samples) were very highly significantly lower in group 2 (p < 0.0005). The dihydroxycholanic acid decrease is due to a decrease of desoxycholic acid.

The ratio cholesterol/bile salts (8 patients, 186 bile samples) was very significantly higher (p < 0.0005) in group 2.

In group 1, the bile cholesterol concentration (9 patients, 119 daily bile samples) was very significantly higher (p < 0.0005) in patients with gallstones than in patients without gallstones.

In group 2, the hepatic synthesis of bile salts was increased.     

Noninvasive urinary organic acids test to assess biochemical and nutritional individuality in autistic children

ObjectivesQuantitative organic acid testing can give information about potential problems, especially with energy production, neurotransmitter metabolism, intestinal dysbiosis and nutritional individuality which is very important in autistic children. The aim of this study was to find out potential differences between the levels of organic acids in the urine of autistic and non-autistic children.Design and methodsThe organic acids in the urine were determined by capillary gas chromatography/mass spectrometry (GC/MS). All overnight urine samples were collected from 35 autistic children and 36 neurologically normal children as healthy controls (4–10 years).ResultsSignificant differences were found between the autistic children and the control group in organic acids: 2-oxoglutaric, isocitric, citric, 4-hydroxybenzoic, 4-hydroxyphenylacetic, hippuric, adipic, suberic (all with p < 0.05).ConclusionOrganic acids test can be used to assess an individual need for nutrient and biochemical abnormalities, especially important for autistic children.     

Renal hypouricaemia in insulin treated diabetes mellitus

Uric acid metabolism was investigated in 69 insulin-treated male diabetic outpatients and in 23 healthy male subjects, because of a reported coincidence between diabetes and gout. All subjects had normal serum creatinine concentrations and none received diuretic treatments.

Compared with normal, the diabetics had significantly lower mean serum uric acid concentrations (0.34 ± 0.08 (SD) mmol/l versus 0.23 ± 0.06 mmol/1, p < 0.001). 17% of the diabetic patients had serum concentrations below the normal mean—2 SD. In contrast, the diabetic patients had a 42% increase in renal uric acid excretion rate (p <0.01), and an 83% increase in the ratio of uric acid clearance/creatinine clearance (p < 0.001). These indices of renal uric acid excretion were both positively correlated to fasting blood glucose levels (r = 0.57, p < 0.001, and r = 0.50, p < 0.001, respectively), to the degree of glycosuria (r = 0.73, p < 0.001, and r = 0.63, p < 0.001, respectively), and to the magnitude of water diuresis (r = 0.60, p < 0.001, and r = 0.39, p < 0.01, respectively).

The hypouricaemia observed in these insulin-dependent diabetic male subjects may probably be caused by the increased renal excretion of uric acid in the presence of hyperglycaemia. The study gave no evidence of increased serum uric acid concentrations in insulin-dependent diabetics. It is therefore likely that any coincidence between gout and diabetes derives from other coexisting serum uric acid raising factors.     

Identification of ethylmalonic acid in urine of two patients with the vomitting sickness of Jamaica.

Large amounts of ethylmalonic acid have been identified in urines from two patients with the vomitting sickness of Jamaica. The amounts were 178 and 882 mug per mg creatinine which are 70 and 350 times, respectively, over control values. Other short and medium chain dicarboxylic acids including glutaric and adipic acids and those with eight and ten carbon chain, saturated and cis-unsaturated, were also detected in large quantities as in the case of hypoglycin treated rats; urine. However, the large increase of urinary ethylmalonic acid in these two human cases is in a sharp contrast to the findings in hypoglycin treated rats in which urinary ethylmalonic acid increased only 3 times over control. It appears that ethylmalonic acid is produced in the cases with the vomiting sickness of Jamaica by carboxylation of n-butyryl-CoA which is not oxidized further due to the inhibition by hypoglycin A. In case of hypoglycin-treated rats, n-butyryl-CoA is mainly conjugated with glycine or deacylated to free butyric acid.     

Aliphatic C6−C14 dicarboxylic acids in urine from an infant with fatal congenital lactic acidosis

A newborn male infant with a severe metabolic acidosis who died one day after birth was found to have high levels of lactic acid in his urine, together with a series of dicarboxylic acids with 6 to 14 carbon atoms chain length. Adipic and suberic acids which occurred in a concentration of ~ 0.05 mmoles/1 were also present in the urine of normal newborns. The concentration in the urine of dicarboxylic acids with 10, 12 and 14 carbon atoms was 0.1, 0.5 and 0.2 mmoles/1. These acids were saturated, monoenic and dienic. The monounsaturated acids were mainly of the cis-Δ₅ configuration. It is suggested that the findings could be consistent with a defect in the normal β-oxidation of fatty acids located at the acyl-CoA dehydrogenase step.     

Development and Progression of Renal Insufficiency With and Without Albuminuria in Adults With Type 1 Diabetes in the Diabetes Control and Complications Trial and the Epidemiology of Diabetes Interventions and Complications Study

OBJECTIVE

This multicenter study examined the impact of albumin excretion rate (AER) on the course of estimated glomerular filtration rate (eGFR) and the incidence of sustained eGFR <60 ml/min/1.73 m² in type 1 diabetes up to year 14 of the Epidemiology of Diabetes Interventions and Complications (EDIC) study (mean duration of 19 years in the Diabetes Control and Complications Trial [DCCT]/EDIC).

RESEARCH DESIGN AND METHODS

Urinary albumin measurements from 4-h urine collections were obtained from participants annually during the DCCT and every other year during the EDIC study, and serum creatinine was measured annually in both the DCCT and EDIC study. GFR was estimated from serum creatinine using the abbreviated Modification of Diet in Renal Disease equation.

RESULTS

A total of 89 of 1,439 subjects developed an eGFR <60 ml/min/1.73 m² (stage 3 chronic kidney disease on two or more successive occasions (sustained) during the DCCT/EDIC study (cumulative incidence 11.4%). Of these, 20 (24%) had AER <30 mg/24 h at all prior evaluations, 14 (16%) had developed microalbuminuria (AER 30–300 mg/24 h) before they reached stage 3 chronic kidney disease, and 54 (61%) had macroalbuminuria (AER >300 mg/24 h) before they reached stage 3 chronic kidney disease. Macroalbuminuria is associated with a markedly increased rate of fall in eGFR (5.7%/year vs. 1.2%/year with AER <30 mg/24 h, P < 0.0001) and risk of eGFR <60 ml/min/1.73 m² (adjusted hazard ratio 15.3, P < 0.0001), whereas microalbuminuria had weaker and less consistent effects on eGFR.

CONCLUSIONS

Macroalbuminuria was a strong predictor of eGFR loss and risk of developing sustained eGFR <60 ml/min/1.73 m². However, screening with AER alone would have missed 24% of cases of sustained impaired eGFR.It has generally been thought that increases in urine albumin excretion rate (AER) precede a fall in glomerular filtration rate (GFR) in patients developing diabetic chronic kidney disease (1). Some large studies in patients with type 2 diabetes (2–4) and a few smaller studies in individuals with type 1 diabetes (5–9), however, have demonstrated that a substantial proportion of diabetic individuals with decreased GFR levels do not have increased AER.In this article, we examine the effects of prior and current levels of AER on the rate of decline in estimated GFR (eGFR) and on the risk of decreased levels of eGFR (<60 ml/min/1.73 m²) in subjects with type 1 diabetes in the Diabetes Control and Complications Trial (DCCT) and/or the follow-up Epidemiology of Diabetes Interventions and Complications (EDIC) study.     

Pattern of aliphatic dicarboxylic acids in uremic serum including a new organic acid, 2,4-dimethyladipic acid.

(1) 2,4-Dimethyladipic acid was first identified in normal human urine using gas chromatography-mass spectrometry. Urinary excretion of 2,4-dimethyladipic acid in 7 healthy adults ranged from 4.9 mumol to 14 mumol per 24 h. (2) Succinic acid, adipic acid, 3-methyladipic acid, 2,4-dimethyladipic acid, pimelic acid and azelaic acid were identified in the ultrafiltrate of the blood obtained from a chronic uremic patient using a hemodialyzer. (3) Levels of succinic acid, adipic acid, 3-methyladipic acid, 2,4-dimethyladipic acid, pimelic acid and azelaic acid in uremic serum were determined using a mass fragmentographic technique. Concentration of succinic acid in uremic serum was comparable to that in normal serum, whereas concentrations of adipic acid, 3-methyladipic acid, 2,4-dimethyladipic acid, pimelic acid and azelaic acid were highly elevated in uremic serum.     

Structure of some aliphatic dicarboxylic acids found in the urine of an infant with congenital lactic acidosis.

Gas chromatography/mass spectrometry was used to identify a series of acids in urine and serum from a child who died 26 h after birth in severe metabolic acidosis with high lactate excretion. cis-5-Decene-1, 10-dioic acid and cis-5-dodencene-1, 12-dioic acid were synthesized and used as references. The following acids were found: hexane-1,6-dioic acid, octane-1,8-dioic acid, decane-1,10-dioic acid, dodecane-1,12-dioic acid, cis-5-decene-1,10-dioic acid, cis-5-dodecen-1,12-dioic acid, cis-5-tetradecene-1,14-dioic acid, trans-3-decene-1,10-dioic acid, and trans-3-dodecene-1,12-dioic acid. The concentration of C6 to C14 acids in the patient's urine was 3.7 mol/mol of creatinine; it was less than 0.2 mol/mol of creatinine in eight normal newborns and approximately 0.1 mol/mol of creatinine in a case of fructose-1,6-biphosphatase deficiency with lactic acidosis. 5-cis-Dodecenedioic acid was present in highest concentration: 1 mol/mol of creatinine in urine and 61 mumol/liter in serum. We propose that impaired beta-oxidation, probably at the acyl-CoA-dehydrogenase step, resulted in the formation of the observed acids. The parents were consanguineous, and a sibling died with the same clinical picture, which suggests a genetic defect.     

The inter-relationships between albuminuria, plasma albumin concentration and indices of glycaemic control in non-insulin-dependent diabetes mellitus

We studied the relationship between albuminuria (measured as albumin/creatinine ratio (alb/Cr) in a random urine sample) and measures of glycaemic control (fructosamine, HbA₁ and glucose) in 470 patients with non-insulin-dependent diabetes mellitus (NIDDM). Albumin excretion was in the microalbuminuric range (alb/Cr ratio > 5.4–40.3) in 112 (23.8%) and in the macroalbuminuric range (alb/Cr ratio > 40.3 mg/mmol) in 89 patients (18.9%). Fourteen percent (n = 67) of patients had a normal plasma HbA₁ (≤8.5%) while 27% (n = 127) had a normal plasma fructosamine concentration (≤ 2.2 mmol/l). Using stepwise multiple regression analysis, plasma fructosamine concentration was found to be independently and negatively associated with urine albumin/creatinine ratio (B = 0.24, P < 0.006) in the macroalbuminuric group. Further analysis of the relationship between plasma albumin concentration and indices of glycaemic control showed that plasma albumin concentration correlated negatively with random plasma glucose concentration in the normoalbuminuric patients (r = −0.16, P = 0.008) but not in microalbuminuric or macroalbuminuric groups. HbA₁ was not correlated with plasma albumin concentration. Our results indicate that albuminuria has an effect on the plasma fructosamine concentration which is independent of plasma albumin concentration.     

The diagnosis of 21-hydroxylase deficiency in a prematurely born infant on the basis of the urinary steroid excretion pattern

The urine of a 6-day-old prematurely born female infant (birth weight 1060 g) suspected of having a 21-OH deficiency showed no steroid abnormalities on capillary GLC analysis. Using GC-MS tetrahydrocortisone (THE) and also 3, 17-dihydroxy-5β-pregnane-20-one (17-OH-Polone) were absent, but two androstanetriolone peaks were observed. In the urine collected on day 9 THE was absent, but a large amount of 3, 11β-dihydroxy-5-androstane-17-one (11-HA) was found by GC-MS to be contaminated by a small amount of 17-OH-Polone. The next urine specimen collected on the 22nd day while the child received cortisol therapeutically showed the characteristic steroid profile for the diagnosis 21-OH deficiency, large peaks of 17-OH-Polone, pregnanetriol (P₃) and 11-keto-pregnanetriol (11-keto-P₃). Over the next few weeks two other compounds were found to have been excreted in relatively large amounts, 3ξ, 16ξ, 17ξ, 20ξ-pregnanetetrol (16-OH-P₃) and surprisingly also a 21-hydroxylated compound, namely 3β, 20, 21-trihydroxy-5-pregnene. These same two compounds were also found in the urine of another infant with suspected 21-OH deficiency.

The urinary steroid excretion patterns characteristic for 21-OH deficiency are dependent on the maturity and age of .the infant. In the prematurely born infant androstanetriolones appear in the urine before 17-OH-Polone. The occurrence of these different steroid excretion patterns is tentatively explained.     

A semiautomated method for the determination of 11-deoxy-17-oxo-steroids in urine

A semiautomated method is described for the determination of total 11-deoxy-17-oxo-steroids (11-DOOS: androsterone, etiocholanolone plus dehydroepiandrosterone) in urine.

Urinary conjugates are manually extracted on a XAD-2 resin, hydrolysed by (β-glucuronidase and “solvolysed” in acid ethyl-acetate according to Burnstein-Lieberman. The free 11-DOOS are extracted automatically with iso-octane and estimated colorimetrically by the Zimmerman reaction in an Auto-Analyzer II system.

The method was evaluated by investigation of its precision, accuracy, sensitivity and specificity. It was found to be satisfactory for the rapid and reliable screening of large numbers of urine samples.