Oxidized plant sterols in human serum and lipid infusions as measured by combined gas-liquid chromatography-mass spectrometry.

Some oxidized forms of cholesterol (oxysterols) are thought to be atherogenic and cytotoxic. Because plant sterols are structurally related to cholesterol, we examined whether oxidized plant sterols (oxyphytosterols) could be identified in human serum and soy-based lipid emulsions. We first prepared both deuterated and nondeuterated reference compounds. We then analyzed by gas-liquid chromatography-mass spectrometry the oxyphytosterol concentrations in serum from patients with phytosterolemia or cerebrotendinous xanthomatosis, in a pool serum and in two lipid emulsions. 7-Ketositosterol, 7 beta-hydroxysitosterol, 5 alpha, 6 alpha-epoxysitosterol, 3 beta,5 alpha,6 beta-sitostanetriol, and probably also 7 alpha-hydroxysitosterol were present in markedly elevated concentrations in serum from phytosterolemic patients only. Also, campesterol oxidation products such as 7 alpha-hydroxycampesterol and 7 beta-hydroxycampesterol were found. Interestingly, sitosterol was oxidized for approximately 1.4% in phytosterolemic serum, which is rather high compared with the approximate 0.01% oxidatively modified cholesterol normally seen in human serum. The same oxyphytosterols were also found in two lipid emulsions in which the ratio of oxidized sitosterol to sitosterol varied between 0.038 and 0.041. In conclusion, we have shown that oxidized forms of plant sterols are present in serum from phytosterolemic patients and two frequently used soy-based lipid emulsions. Currently, it is unknown whether oxyphytosterols affect health, as has been suggested for oxysterols. However, 7 beta-hydroxycholesterol may be one of the more harmful oxysterols, and both sitosterol and campesterol were oxidized into 7 beta-hydroxysitosterol and 7 beta-hydroxycampesterol. The relevance of these findings therefore deserves further exploration.

Nocardia abscessus sp. nov.

Chemotaxonomic and 16S rRNA gene sequence analyses of four bacterial strains isolated from clinical material clearly demonstrated that these bacteria belong to the genus Nocardia. DNA-DNA hybridization data as well as the physiological characteristics of the isolates indicated that they are closely related and belong to a single species that differs from previously described members of the genus. The name Nocardia abscessus sp. nov. is proposed for these organisms represented by strain IMMIB D-1592T (= DSM 44432T). Strain IMMIB D-1592T exhibits 56.8 and 60.0% DNA-DNA relatedness to Nocardia asteroides ATCC 19247T and Nocardia paucivorans DSM 44386T, respectively.

Nocardia paucivorans sp. nov.

Chemotaxonomic and 16S rDNA sequence analyses of an isolate from the sputa and bronchial secretions of a patient with chronic lung disease clearly demonstrated that it belongs to the genus Nocardia. DNA-DNA hybridization data, as well as the biochemical characteristics of the isolate, indicate that it belongs to a new species that differs from previously described members of the genus Nocardia. The name Nocardia paucivorans sp. nov. is proposed for this isolate and is represented by strain IMMIB D-1632T (= DSM 44386T).

Tsukamurella tyrosinosolvens sp. nov.

Chemotaxonomic and 16S ribosomal DNA sequence analyses of four bacterial isolates from blood cultures from patients with cardiac pacemaker implants and sputa of patients with chronic lung infections clearly demonstrated that these bacteria belong to the genus Tsukamurella. DNA-DNA hybridization data, as well as the physiological characteristics of the isolates, indicate that they are closely related and belong to a single species that differs from previously described members of the genus Tsukamurella. The name Tsukamurella tyrosinosolvens sp. nov. is proposed for these isolates, and the new species is represented by strain IMMIB D-1397T (= DSM 44234T). Strain IMMIB D-1397T exhibits 53.4, 53.5, and 54.7% DNA-DNA relatedness to Tsukamurella paurometabola DSM 20162T, Tsukamurella inchonensis DSM 44067T, and Tsukamurella pulmonis DSM 44142T, respectively.

Detection and quantitation of amphetamine and methamphetamine: electron impact and chemical ionization with ammonia--comparative investigation on Shimadzu QP 5000 GC-MS system.

Gas chromatographic-mass spectrometric (GC-MS) analyses of amphetamine and methamphetamine with chemical ionization (CI) (NH3) and electron impact (EI) were compared after solid-phase extraction from urine. The optimal CI (NH3) conditions for amphetamines were selected; the CI mass spectra of amphetamine, methamphetamine, methylenedioxyamphetamine (MDA), methylenedioxymethamphetamine (MDMA), and methylenedioxyethylamphetamine (MDEA) were obtained. Amphetamine and methamphetamine were monitored as [M+H]+ ions in SIM mode. Under these conditions the GC-MS limits of detection and quantitation were determined for amphetamine (2.1 and 5.0 mg/L, respectively) and for methamphetamine (0.9 and 2.0 mg/L, respectively). The heptafluorobutanoyl (HFB) derivatives were used to increase the sensitivity of the GC-MS analysis of amphetamines. Under CI (NH3) conditions these compounds form mainly the cluster ions [M+NH4]+. For HFB-derivatives the limits of detection and quantitation for CI (NH3) were 95 and 170 micrograms/L, respectively, for amphetamine (m/z 349) and 90 and 160 micrograms/L, respectively, for methamphetamine (m/z 363). For EI they were 10 and 30 micrograms/L, respectively, for amphetamine (m/z 240) and 9 and 27 micrograms/L, respectively, for methamphetamine (m/z 254). The GC-MS-CI (NH3) technique was found to be suitable for confirmation of amphetamine and methamphetamine in urine samples in addition to the routine GC-MS-EI method.

Rapid chemotaxonomic diagnosis of human tuberculosis.

65 different clinical specimens from patients suspected of being infected with Mycobacterium tuberculosis were examined by three different diagnostic methods. Two of these methods were the conventional microscopic and cultural examinations. The third, a modern chemotaxonomical method is based upon the detection of tuberculostearic acid by GC-MS analysis using selected ion monitoring (GC-MS/SIM). Comparison of the results of the GC-MS analysis with those of the conventional methods has indicated that tuberculostearic acid analysis can be used for diagnosing tuberculosis under diagnostic routine conditions. The GC-MS method is rapid, usually providing results within 20 hours or less.

Tsukamurella pulmonis sp. nov.

Chemotaxonomic and 16S ribosomal DNA sequence analyses of an isolate from the sputum of a patient with a mycobacterial lung infection clearly delineated a new species of the genus Tsukamurella. This new species can be defined on the basis of genotypic and phenotypic data. The name Tsukamurella pulmonis sp. nov. is proposed for this organism; the type strain is IMMIB D-1321T (= DSM 44142T). This isolate shows 44.2 and 36.2% DNA relatedness to Tsukamurella paurometabola DSM 20162T (T = type strain) and Tsukamurella inchonensis DSM 44067T, respectively.

Tsukamurella inchonensis sp. nov.

Chemotaxonomic and genomic 16S ribosomal DNA sequence analyses of two isolates obtained from two different clinical materials clearly delineated a new species of the genus Tsukamurella. This new species can be identified by its 16S ribosomal DNA similarity values, as well as its physiological characteristics. The name Tsukamurella inchonensis sp. nov. is proposed for these isolates, which are represented by strain IMMIB D-771T (= DSM 44067T) (T = type strain). This strain exhibits only 45% DNA relatedness to Tsukamurella paurometabola.

Lentzea gen. nov., a new genus of the order Actinomycetales.

We describe a new genus of mesophilic actinomycetes, for which we propose the name Lentzea. The strains of this genus form abundant aerial hyphae that fragment into rod-shaped elements. Whole-cell hydrolysates contain the meso isomer of diaminopimelic acid and no characteristic sugar (wall chemotype III). The phospholipid pattern type is type PII (phosphatidylethanolamine is the characteristic phospholipid); the major menaquinone is MK-9. The fatty acid profile comprises saturated, unsaturated, and branched-chain fatty acids of the iso and anteiso types in addition to tuberculostearic acid (10Me-C18:0). A 16S ribosomal DNA sequence analysis revealed that the genus Lentzea is phylogenically related to the genera Actinosynnema, Saccharothrix, and Kutzneria. The type species of this genus is Lentzea albidocapillata sp. nov.; the type strain of this species is strain IMMIB D-958 (= DSM 44073).

Cellular fatty acid methyl ester profiles as a tool in the differentiation of members of the genus Mycobacterium.

The cellular fatty acid profiles of 84 strains belonging to 53 different species of the genus Mycobacterium were determined by gas liquid chromatography-mass spectrometry (GCMS). Two main types and four subtypes of fatty acid profiles were recognizable. The first main type is the G type, nominally referring to M. gordonae, members of which lack tuberculostearic acid or other 10-methyl branched-chain fatty acids, but contain normal saturated and unsaturated fatty acids. This type is further subdivided into the G alpha subtype that is characterized by 2-methyl tetradecanoic acid (2-Me-C14:0) as the only 2-methyl branched fatty acid. Strains belonging to the second main type, the T type, nominally referring to tuberculosis, contain tuberculostearic acid and other 10-methyl branched acids in addition to the normal saturated and unsaturated ones. This type has been further subdivided into three subtypes: the T alpha subtype that does not contain any 2-methyl branched fatty acids; the T beta subtype that contains both 2-methyl tetradecanoic (2-Me-C14:0) and 2,4-dimethyl tetradecanoic (2,4-DMe-C14:0) acids as 2-methyl branched fatty acids; the T gamma subtype which contains 2-methyl dodecanoic (2-Me-C12:0), 2,4-dimethyl dodecanoic (2,4-DMe-C12:0) and 2,4-dimethyl tetradecanoic (2,4-DMe-C14:0) acids as 2-methyl branched-chain acids. Fatty acid analysis showed a great homogeneity within the genus and the profiles produced were not very helpful in distinguishing between members of the genus Mycobacterium except for the identification of M. gordonae, M. kansasii, and M. gastri.

A screening method for the rapid detection of barbiturates in serum by means of tandem mass spectrometry.

A mass spectrometric screening method for barbiturates in serum was developed. Under electron impact conditions barbiturates fragment by loss of HNCO. A 'constant neutral loss' scan of 43 u provides, therefore, an indication of the presence of members of this toxicologically relevant class of compounds. Subsequent identification is possible either by 'daughter' or by 'parent ion' scans. The detection limit for propallylonal, buto- and phenobarbital was determined as better than 1 micrograms ml-1 serum.

[A screening method for rapid detection of barbiturates in serum using tandem mass spectrometry]. / Eine Screening-Methode zum schnellen Nachweis von Barbituraten im Serum mittels Tandem-Massenspektrometrie.

A mass spectrometric screening method for the detection of barbiturates in serum will be presented. Fragmentation of barbiturates under EI conditions leads to a loss of HNCO. Therefore, "neutral loss scans" of 43 u allow the indication of possibly existing representatives of this class of compounds. Identification can be achieved by "daughter" or "parent ion scans". Thus, 10 micrograms/ml apro-, pento- and phenobarbital could be detected in serum.

[Possibilities of quality assurance (accuracy, precision) in forensic blood alcohol determination in compliance with the reduced safety increase of 0.1 g promille]. / Möglichkeiten einer Qualitätssicherung (Richtigkeit, Präzision) in der forensischen Blutalkoholuntersuchung bei Einhaltung des reduzierten Sicherheitszuschlags von 0,1 g Promille.

A lowering of thresholds in blood alcohol determination can be realized by critical reflection on efficiency of the techniques of measurement including calibration and internal and external quality control. A threshold value can be controlled by a technique of measurement if the prognosis interval of this technique is equal or smaller than the control interval. From date 1/7/1990 we measure blood probes like described. Blood measurement before this date must be proofed for accuracy. In cases of non compliance the old threshold value should be used.

Menaquinone patterns of Amycolatopsis species.

The menaquinones of representative strains of the actinomycete genus Amycolatopsis were examined by reverse phase thin-layer chromatography and mass spectrometry. Representatives of all seven validly described species contained various combinations of di-, tetra- and hexahydrogenated menaquinones with nine isoprene units as predominant isoprenologues. It seems likely that the variation in the predominant menaquinones merely reflects the stages from the growth cycle from which biomass was taken. The detection of major proportions of hydrogenated menaquinones with nine isoprene units serves to distinguish Amycolatopsis strains from most other actinomycetes, notably those belonging to related genera such as Amycolata and Pseudonocardia.

[Biotransformation of meclozine in the human body]. / Untersuchungen zur Biotransformation von Meclozin im menschlichen Körper.

After oral administration of 1-[(4-chlorophenyl)-phenylmethyl]-4-[3-methylphenyl)-methyl]-piperazine (1, Meclozine) eleven compounds were isolated from human urine and faeces. The structural elucidation of the metabolites was accomplished by comparison of their spectral data with those of the synthetic reference compounds. The metabolites were identified as: Meclozine (1), N-[(4-chlorophenyl)-phenylmethyl]-piperazine (2), 3-(4-[(4-chlorophenyl)-phenylmethyl]-piperazino)-methyl-benzoic acid (3), 3-(4-[(4-chlorophenyl)-phenylmethyl]-piperazino)-methyl-benzamide] (4), 2-[3-(4-[(4-chlorophenyl)-phenylmethyl]-piperazino)-methyl-benzoyl ]-amino- ethanesulphonic acid (5), 3-(4-[(4-chlorophenyl)-3'-hydroxy-4'-methoxyphenylmethyl]-piper azino)- methyl-benzoic acid (6), 1-[(4-chlorophenyl)-phenylmethyl]-4-[(3-methylphenyl)-methyl]- piperazine-N4-oxide (7), 1-[(4-chlorophenyl)-phenylmethyl]-4-[(3-methylphenyl)-methyl]- piperazine-N,N'-dioxide (8), 3-methyl-benzoic acid (9), 3-methyl-hippuric acid (10) and 3-methyl-benzoic acid-glucuronide (11). The structure of compound 11 was confirmed after enzymatic cleavage and identification of the aglycon. A further metabolite was detected, but not identified.

Menaquinone composition in the classification and identification of aerobic actinomycetes.

Menaquinones were the only isoprenoid quinones found in 36 strains representing different species of the genera Nocardia, Mycobacterium, Rhodococcus, Amycolatopsis, Saccharothrix, Streptomyces, Nocardiopsis and Actinomadura. Dihydrogenated menaquinones with nine isoprene units [MK-9(H2)] were the main components isolated from Mycobacterium. Dihydrogenated and tetrahydrogenated menaquinones with eight isoprene units were the predominant compounds identified in typical Rhodococcus and Nocardia strains, respectively. "Nocardia phenotolerans" differed from all of the other Nocardia species included in the study, in that it contained the MK-9(H2) [MK-8(H2)] menaquinone system. Nocardioform bacteria lacking mycolic acids contained tetrahydrogenated menaquinones with nine isoprene units as the main component. The Streptomyces strains studied exhibited complex mixtures of partially saturated menaquinones with nine isoprene units with the hexa- and/or octahydrogenated components predominating. Actinomadurae contained major amounts of hexahydrogenated menaquinones with nine isoprene units. In contrast, the single Nocardiopsis strain examined possessed complex mixtures of menaquinones with ten isoprene units, the dihydrogenated components being main constituents.

[Isolation of chlorphenoxamine metabolites in human urine and their identification]. / Isolierung von Chlorphenoxamin-Metaboliten aus menschlichem Harn und ihre Identifizierung.

After the administration of chlorphenoxamine (2-[1-(4-chlorophenyl)-1-phenylethoxy]-N,N-dimethylethanamine++ +, Systral) (I) the following compounds have been detected in human urine. They were identified as chlorphenoxamine (I), N-demethyl-chlorphenoxamine (II), chlorphenoxamine-N-oxide (III), 1-(4-chlorophenyl)-l-phenylethanol (IV), 1-(4-chlorophenyl)-1-(4'-hydroxyphenyl)-ethanol (V), 1-(4-chlorophenyl)-1-(4'-hydroxyphenyl)-ethene (VI), 1-(4-chlorophenyl)-1-(4'-hydroxy-3'-methoxyphenyl)-ethanol (VII), 1-(4-chlorophenyl)-1-(4'-hydroxy-3'-methoxyphenyl)-ethene (VIII), 2-[1-(4-chlorophenyl)-1-(4'-hydroxyphenyl)-ethoxy]-N-methyl-ethanamine (IX) and 2-[1-(4-chlorophenyl)-1-(4'-hydroxy-3'-methoxyphenyl-ethoxy]- N-methylethanamine (X). The compounds IV, V, VI, VII, VIII, IX and X were also found to be excreted as conjugates. It cannot be excluded that the compounds VI and VIII are artefacts.

[Detection and identification of a new metabolite of fenethylline]. / Nachweis und Identifizierung eines neuen Metaboliten des Fenethyllin.

Fenetylline is metabolized in humans on two pathways. In addition to previously described degradation to amphetamine and 7-oxyethyltheophylline fenetylline undergoes moreover oxydative N-dealkylation to yield 7-aminoethyltheophylline and phenylacetone.