Biomedical subjects
Publications and source records attributed to G Spiteller.
A simple procedure to distinguish between urinary acids originating from nutrition and beverages and those produced from compounds stored in the body is described. Acids originating from nutrition disappear in urine samples after a few days of a zero diet, whereas those produced from body compounds do not. The zero diet can be substituted by a diet of peeled rice and water.
In the present study the effects on endogenous hormones of application of topical corticosteroids are investigated. Radioimmunological assays of the hormones cortisol, adrenocorticotrophic hormone (ACTH) and alpha-melanocyte stimulating hormone (alpha-MSH) before, during and after a large-area treatment with topical corticosteroids (30 g/d 0.025% fluocinolone acetonide ointment for 9 days) are performed in eight patients suffering from extensive inflammatory dermatoses (7 cases of eczema, 1 case of psoriasis). Cortisol was measured in plasma daily under treatment. In addition, gas chromatographic and mass spectrometric determinations of urocorticoids and uroandrogens are carried out in 24-h urine. A distinct, treatment-induced suppression of the endogenous steroid production was significant (-42.7%, p less than or equal to 0.05) within the first four days and diminished (-24.4%, p less than or equal to 0.10) after nine days' steroid therapy. The suppression of the plasma ACTH was also significant (-23.1%, p less than or equal to 0.05). The alpha-MSH level did not show any alterations. The measured urocorticoids showed an average fall of -38.9% (p less than or equal to 0.05). It is noteworthy that despite continued steroid treatment the marked initial suppression of the endogenous steroid production diminishes. It is possible that a therapy-induced "normalization" of the epidermis brings about a diminished steroid influx and thus a lower systemic load. Under the therapy conditions chosen here, the androgen metabolites in the urine did not show any significant fluctuations.
Capillary gas chromatography of amino acid derivatives obtained from biological material shows a large number of previously unknown constituents. The profiles of amino acids obtained from urine of healthy individuals and haemofiltrate of uraemic patients indicate that haemofiltration removes some amino acids to a considerably higher extent from uraemic patients than the kidney does from healthy persons. For instance, if haemofiltration is required three times a week, an approximately ten-fold amount of the essential amino acid methionine and a forty-fold amount of the essential amino acid leucine is lost compared to their excretion in urine by a healthy individual over the same period.
3,5-Dihydroxyphenylpropionic acid, 3,5-dihydroxycinnamic acid and 2,3-dihydroxycinnamic acid were detected for the first time to be components of human urine. In the course of this investigation all constitutional isomers of dihydroxy-benzoic, -phenylpropionic, -phenylacetic and -cinnamic acid were synthesized. Mass spectra and retention indices of methyl and trimethylsilyl (TMS) derivatives were determined. In contrast to many other substituted aromatic compounds the mass spectra of methyl and TMS derivatives of dihydroxy aromatic acids often allow a firm distinction to be made between constitutional isomers: TMS derivatives of aromatic acids containing two hydroxy groups located in the ortho position to each other can be recognized by ions resulting from a primary cleavage reaction mainly in the side chain or ester group, followed by loss of tetramethylsilane. In methyl derivatives of 1,2,3-trisubstituted isomers, methoxy groups are lost much more easily from the ions corresponding to the benzylic cleavage than in other isomers. Methyl derivatives of dihydroxycinnamic acids containing at least one methoxy group in the ortho position to the side chain are characterized by a fragmentation reaction, corresponding to the loss of dimethyl ether. TMS and methyl derivatives of 3,5-dihydroxy aromatic acids show unique structure-specific fragmentation reactions.
When alpha, beta-unsaturated acids and alpha-keto acids react with diazomethane not only are the corresponding methylates produced, but also diazomethane is added to the C = C double bond or to the oxo group. The gas chromatographic and mass spectral behaviour of these undesired products and some further artefacts produced in the hot inlet lines of a gas chromatograph are described. The mass spectra and retention indices allowed the structural assignment of several "unknown" compounds found previously in the methylated acid fraction of urine. A detailed analysis of the reaction of alpha-oxo acids with diazomethane revealed that, besides the already known oxirane methyl esters, homologous esters are also produced by an insertion reaction.
A two-step ultrafiltration method combined with anion-exchange chromatography is described for the separation of lipophilic and strongly lipophobic acids occurring in human blood, serum and plasma. After treatment with diazomethane, the acid fractions are separated further by gas chromatography. The acids were identified by their mass spectra. Profiles obtained from the strongly acidic fraction of blood samples of healthy individuals are characterized by main peaks corresponding to organic phosphoric acid esters. The peaks are absent in plasma and serum.
A mixture of long-chain furan fatty acids was prepared as methyl esters from testes lipids of Northern pike (Esox lucius). Upon feeding these esters to rats, dicarboxylic acids, which still contained the furan structure, were found in the urine. The first phase of a rapid but incomplete catabolism is beta-oxidation of the proximal chain of the furan fatty acids. It proceeds to a distance of three carbon atoms from the ring. omega-Oxidation of the terminal alkyl chain, followed by alpha-oxidation gives rise to a second alkylcarboxyl chain with five carbon atoms or less. The ring methyl substituents of the precursor acids seem to be more resistant to oxidation than the alkyl substituent with three or five carbon atoms. The urinary catabolites from furan fatty acids in the rat are similar to furan acids found in human urine, but only one of the structures occurs in both sources.
The neutral steroid fractions in the urine of eleven patients suffering from various forms of liver disease with cholestasis and of ten healthy individuals were studied by glass capillary gas chromatography-mass spectrometry. The steroid conjugates in urine were enzymatically solvolysed, the liberated steroids extracted and transformed into the trimethylsilylether for measurements. The excretion rates of androstane and pregnane metabolites of patients with liver disease were far lower than those of healthy persons. The main compounds in the urine of the former were the bile alcohols 27 - nor - 3 alpha, 7 alpha, 12 alpha, 24 xi, 25 xi - pentahydroxy - 5 beta - cholestane and 3 alpha, 7 alpha, 12 alpha, 25 xi, 26 - pentahydroxy - 5 beta - cholestane. Our data suggest a correlation between the excretion rates of these bile alcohols and the serum levels of bilirubin. While the excretion rate of the two bile alcohols in the urine of healthy individuals was approximately 0.24 mg/24 h (0.6 mumol/24 h) a patient with a serum bilirubin of 841 mumol/l excreted 4 mg/24 h (9 mumol/24 h). The accumulation of bile alcohols described in this study possibly indicates alternative pathways of cholic acid formation in liver disease.
2-Mercapto-1-methyl-5-methylmercapto-imidazole (13) was found in urine samples of man and rat after intake of thiamazole (1). It is assumed that the metabolite is produced via a N-oxidation intermediate enabling a nucleophilic attack at carbon-5 in the thiazole ring.
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1. G.l.c.--mass spectral analysis of t.l.c. fractions of urine samples of patients treated with 5-(2-chloroethyl)-4-methylthiazole (clomethiazole), has revealed two minor metabolites, each with two sulphur atoms. 2. Their structures were found to be 2-methylthio-clomethiazole and 5-acetyl-4-methyl-2-mercapto-thiazole, formed by thiomethylation and thiohydroxylation, respectively, of the original compound. 3. The structures of six other minor metabolites resulting from side-chain degradation have been elucidated. 4. The occurrence of metabolites with substituents at position 2 of the heterocyclic nucleus is assumed to be initiated by oxidative attack at the nitrogen, followed by nucleophilic substitution in position 2.
Besides 6-hydroxychlorzoxazon II, the main metabolite of 5-chlorobenzoxazolin-2-one (chlorzoxazone I, Paraflex) two other compounds were found to be metabolites of chlorzoxazone: 5-chloro-2,4-dihydroxyacetanilide XXII is produced from 6-hydroxychlorzoxazone II by ring cleavage and acetylation of the nitrogen. More interesting is the formation of the 6-hydroxybenzoxazolone XX. This compound is formally produced by substitution of Cl for hydrogen and hydroxylation of a neighbour ring-carbon atom.
A sequence of chromatographic methods (thin-layer chromatography, high-performance liquid chromatography and glass capillary gas chromatography) was used to separate the acid fraction of human urine. The power of this method to separate and detect previously unknown compounds and the elucidation of their final structure with mass spectrometry is exemplified by the identification of N-acetyl-2-aminooctanoic acid as a metabolic compound in the urine of healthy individuals. In addition, the conjugate of glycine with indolepropionic acid, N-formylanthranilic acid, succinoylphenylalanine, delta-hydroxyvaleric acid, beta-hydroxycapric acid, 3-hydroxyadipic acid, and higher homologues were detected in a polar fraction of human urine.
The organic acid fraction of hemofiltrates was investigated in the form of methylates by glass capillary gas chromatography-mass spectrometry. The pattern obtained is similar to that of urinary organic acid methylates from healthy individuals. A marked difference was noted for N-phenylacetyl-alpha-aminoglutarimide, present in hemofiltrate at levels 50-100 times higher than those in urine. Analysis of hemofiltrate samples taken at different times during a hemofiltration with post-dilution technique revealed that the hemofiltrate concentration of most compounds was drastically reduced the course of the hemofiltration treatment. Compared to the other compounds, the reduction in hemofiltrate concentration of N-phenylacetyl-alpha-aminoglutarimide was extremely rapid.
Urine steroid profiles of healthy individuals can be divided into two groups according to greatly different excretion rates of dehydroepiandrosterone (DHEA). About 80% of the population show an excretion of DHEA in urine of just above the detection limit or less of the main androgens androsterone (A) and etiocholanolone (E). This excretion is only enhanced in psychological stress situations. The remaining 20% excrete DHEA in roughly equal amounts as A and E. While the relation of excreted steroids is rather constant, the absolute amounts may vary greatly. In contrast to the behaviour of all other steroids DHEA excretion is not in relation to other steroids. The group of "high DHEA" producing individuals in particular shows drastic changes in the excretion during a day: the DHEA excretion rapidly rises from morning until afternoon and then drops to rather low values in the resting period during the night. A recognizable DHEA production seems to be closely related to the waking period.
Extrelut columns commonly used to isolate metabolites from biological fluids often contain fatty acids, phthalates, alcohols and especially resin acids which occassionally cause great analytical problems. Therefore purification before use of Extrelut columns is strongly recommended.
The steroid fraction of hemofiltrate contains a large number of unknown steroids, which were separated by a combination of different chromatographic procedures and finally characterized by gas chromatography mass spectrometry. Besides different isomeric 3,16,17-trihydroxyandrostanes in a rather polar fraction a 3,16,17-trihydroxy-11-ketoandrostane was detected, obviously the first naturally occurring representative of 3,11,16,17-oxygen-tetrasubstituted androstanes.
The acidic compounds of urine were separated by thin-layer chromatography in eight fractions. Each fraction was investigated separately by the combination glass capillary gas chromatography-mass spectrometry. About 500 compounds were detected, 2/5 of these could be characterized by their mass spectra. Retention data and key fragments of the mass spectra were tabulated. Many of the detected compounds are still unknown.