Formation of C19 11-hydroxysteroids by porcine Leydig cells

1992 ◽  
Vol 70 (2) ◽  
pp. 174-176 ◽  
Author(s):  
J. I. Raeside ◽  
R. L. Renaud ◽  
M. W. Khalil
Keyword(s):  
Leydig Cells ◽  
High Performance ◽  
Solid Phase ◽  
Reversed Phase ◽  
Mature Male ◽  
Gas Chromatography Mass Spectrometry ◽  
Steroid Biosynthesis ◽  
Testicular Vein ◽  
Authentic Standard ◽  
Porcine Testis

In a previous study, we reported the presence of 11β-hydroxyandrostenedione and 11β-hydroxytestosterone in testicular vein blood from mature male pigs. Since C19 steroids with an oxygen function at C11 have not been recorded as products of steroid biosynthesis in normal mammalian testes, we have examined their possible production in purified preparations of porcine Leydig cells. Both androstenedione and cortisol were added as substrates in studies using cell incubations of Leydig cells from mature boars (> 8 months old). Steroids were recovered from media by solid-phase extraction and separated by reversed-phase high performance liquid chromatography. Peaks corresponding to retention times of authentic standard steroids were seen for both 11β-hydroxyandrostenedione and 11β-hydroxytestosterone from each substrate. Generally, lesser amounts of C19 11-oxosteroids were noted also. Definitive confirmation was made by gas chromatography – mass spectrometry for 11β-hydroxyandrostenedione in the media.Key words: 11β-hydroxylation, androgens, Leydig cells, porcine, testis.

Secretion of 19-hydroxyandrostenedione and 19-hydroxytestosterone by porcine Leydig cells in vitro and in vivo

1993 ◽  
Vol 137 (2) ◽  
pp. 281-289 ◽  
Author(s):  
J. I. Raeside ◽  
R. L. Renaud ◽  
R. M. Friendship ◽  
M. W. Khalil
Keyword(s):  
Leydig Cells ◽  
High Performance ◽  
Solid Phase ◽  
Gas Chromatography Mass Spectrometry ◽  
Isolation And Identification ◽  
Testicular Vein ◽  
Secretory Products ◽  
Adrenal Secretion

ABSTRACT 19-Hydroxytestosterone and 19-hydroxyandrostenedione have been identified as secretory products of the testes in the mature male domestic pig. Their isolation and identification were made by reverse-phase high-performance liquid chromatography and capillary gas chromatography-mass spectrometry (CGC-MS) of extracts from testicular vein blood and media of incubations with Leydig cells. Blood was collected from veins on the surface of the testes of anaesthetized boars. Collagenase-dispersed Percoll-purified cells (> 90% pure) were incubated (20 × 106 cells/flask) with androstenedione (8·75 μmol/l) or [3H]androstenedione (5 × 106 c.p.m.) for < 60 min. Steroids were recovered from plasma or media by solid-phase extraction and the unconjugated fractions chromatographed isocratically in two solvent systems (acetonitrile: water, 37:63 (v/v) and methanol: water, 70:30 (v/v)) before CGC-MS analysis. 19-Hydroxy-testosterone was present in greater quantities than 19-hydroxyandrostenedione in testicular vein blood; it was also seen as a quantitatively significant metabolite of unlabelled and radioactive androstenedione in the incubation studies. The demonstration of the secretion of 19-hydroxyandrogens from porcine testes thus raises questions concerning the physiological significance of a testicular, rather than an adrenal, secretion of these compounds. Journal of Endocrinology (1993) 137, 281–289

scholarly journals Metabolic Evaluation of Urine from Patients Diagnosed with High Grade (HG) Bladder Cancer by SPME-LC-MS Method

Molecules ◽  
2021 ◽  
Vol 26 (8) ◽  
pp. 2194
Author(s):  
Kamil Łuczykowski ◽  
Natalia Warmuzińska ◽  
Sylwia Operacz ◽  
Iga Stryjak ◽  
Joanna Bogusiewicz ◽  
...  
Keyword(s):  
Thin Film ◽  
Bladder Cancer ◽  
High Performance ◽  
Biomarker Discovery ◽  
Solid Phase ◽  
Biological Fluids ◽  
Reversed Phase ◽  
Control Group ◽  
Metabolic Evaluation ◽  
Metabolomic Profiling

Bladder cancer (BC) is a common malignancy of the urinary system and a leading cause of death worldwide. In this work, untargeted metabolomic profiling of biological fluids is presented as a non-invasive tool for bladder cancer biomarker discovery as a first step towards developing superior methods for detection, treatment, and prevention well as to further our current understanding of this disease. In this study, urine samples from 24 healthy volunteers and 24 BC patients were subjected to metabolomic profiling using high throughput solid-phase microextraction (SPME) in thin-film format and reversed-phase high-performance liquid chromatography coupled with a Q Exactive Focus Orbitrap mass spectrometer. The chemometric analysis enabled the selection of metabolites contributing to the observed separation of BC patients from the control group. Relevant differences were demonstrated for phenylalanine metabolism compounds, i.e., benzoic acid, hippuric acid, and 4-hydroxycinnamic acid. Furthermore, compounds involved in the metabolism of histidine, beta-alanine, and glycerophospholipids were also identified. Thin-film SPME can be efficiently used as an alternative approach to other traditional urine sample preparation methods, demonstrating the SPME technique as a simple and efficient tool for urinary metabolomics research. Moreover, this study’s results may support a better understanding of bladder cancer development and progression mechanisms.

scholarly journals Determination of Flumorph Residues in Vegetables, Soil, and Natural Water by Solid-Phase Extraction Cleanup and High-Performance Liquid Chromatography with UV Detection

2008 ◽  
Vol 91 (6) ◽  
pp. 1459-1466 ◽  
Author(s):  
Ji-Ye Hu ◽  
Yu-Chao Zhang ◽  
Hai Yan
Keyword(s):  
Solid Phase Extraction ◽  
Natural Water ◽  
High Performance ◽  
Solid Phase ◽  
Residue Analysis ◽  
Reversed Phase ◽  
Uv Detection ◽  
Phase Extraction ◽  
Limit Of Quantification

Abstract A method for high-performance liquid chromatographic (HPLC) determination of flumorph residues in cucumber, tomato, soil, and natural water was developed and validated. Primary secondary amine or octadecylsilyl (C18) solid-phase extraction cartridges were used for sample preparation. Reversed-phase HPLC with UV detection was used for separation and quantification of the pesticide. The combined cleanup and chromatographic method steps were sensitive and reliable for simultaneous determination of residues of the 2 isomers of flumorph in the studied samples. This method is characterized by recovery &gt;97.9, coefficient of variation &lt;6.2, and limit of quantification of 0.01 mg/kg, in agreement with directives for method validation in residue analysis. Flumorph residues in the samples were further confirmed by HPLC/mass spectrometry. The proposed method is fast, easy to perform, and could be used for monitoring of pesticide residues.

scholarly journals Multiresidue Analytical Method for the Determination of Eight Penicillin Antibiotics in Muscle Tissue by Ion-Pair Reversed-Phase HPLC after Precolumn Derivatization

1999 ◽  
Vol 82 (5) ◽  
pp. 1083-1095 ◽  
Author(s):  
Eric Verdon ◽  
Pierrick Couëdor
Keyword(s):  
Muscle Tissue ◽  
Phosphate Buffer ◽  
High Performance ◽  
Solid Phase ◽  
Sodium Thiosulfate ◽  
Reversed Phase ◽  
Extraction Column ◽  
Precolumn Derivatization ◽  
Solution Ph

Abstract A high-performance liquid chromatographic multiresidue method was developed for the determination of 8 penicillin compounds (benzylpenicillin, phenoxymethylpenicillin, ampicillin, amoxicillin, nafcillin, oxacillin, cloxacillin, and dicloxacillin) at trace levels in muscle tissue. This method involves extraction of the penicillins with phosphate buffer pH 9 followed by cleanup and concentration on a C18 solid¯phase extraction column and reaction with benzoic anhydride at 50°C for 5 min and with 1,2,4-triazole and mercury(II) chloride solution pH 9 at 65°C for 10 min. The derivatized compounds are eluted on a C18 column with a mobile phase containing acetonitrile and phosphate buffer (pH 6; 0.1 mol/L) loaded with sodium thiosulfate and ion-pairing tetrabutylammonium hydrogenosulphate. The method detection limit is approximately 3-11 μg/kg and the limit of determination was evaluated down to 25 μg/kg in line with the criteria of the EU decision No. 93/256/EEC.

scholarly journals Sensitive Mercury Speciation by Reversed-Phase Column High-Performance Liquid Chromatography with UV-Visible Detection After Solid-Phase Extraction Using 6-Mercaptopurine and Dithizone

2008 ◽  
Vol 91 (6) ◽  
pp. 1453-1458 ◽  
Author(s):  
Hamid Hashemi-Moghaddam ◽  
Mohamad Saber-Tehrani
Keyword(s):  
High Performance ◽  
Solid Phase ◽  
Reversed Phase ◽  
Phase Extraction ◽  
Organic Mercury ◽  
Tuna Fish ◽  
Rp Hplc ◽  
Uv Visible ◽  
Phase Column ◽  
Reversed Phase Column

Abstract A highly selective and sensitive method was developed for preconcentration of inorganic and organic mercury compounds followed by reversed-phase column high-performance liquid chromatography (RP-HPLC) with UV-visible detection. The method was based on the reaction of mercury with 6-mercaptopurine and solid-phase extraction (SPE) of the complex on an octadecylsilane (C18) cartridge. The complex was then treated with ammoniacal dithizone solution, and the complexes of inorganic and organic mercury with dithizone were eluted by methanol. The speciation analysis of methylmercury (MeHg), phenylmercury (PhHg), and inorganic Hg (II) was carried out by RP-HPLC. Some experimental variables that influence the SPE and derivatization, such as pH, chelating and derivatizing agent concentration, and surfactant addition, were investigated. The calibration graphs of MeHg, PhHg, and Hg (II) were linear [correlation coefficient (r) &gt; 0.999] from the detection limits (0.12, 0.16, and 0.14 ng) to 8.5, 6.0, and 6.7 ng Hg, respectively. By applying the SPE procedure, a 100-fold concentration of the sample was obtained. The procedure was applied to sea water and tuna fish samples. The method's accuracy was investigated by using tuna fish certified reference material BCR 464 and by spiking the samples with different amounts of MeHg, PhHg, and Hg (II). The average recoveries of MeHg, PhHg, and Hg (II) from spiked samples (0.12.0 g/L Hg) were 96 4, 98 3, and 104 4, respectively.

EVIDENCE FOR BIIIARY EXCRETION OF PHENPROCOUMON AND ITS METABOLITES IN HUMANS; IDENTIFICATION BY HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY AND GAS CHROMATOGRAPHY-MASS SPECTROMETRY

1987 ◽  
Author(s):  
J X de Vries ◽  
R Raedsch ◽  
A Stiehl ◽  
U Voelker ◽  
I Walter-Sack ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Gas Chromatography ◽  
High Performance Liquid Chromatography ◽  
Liquid Chromatography ◽  
High Performance ◽  
Gradient Elution ◽  
Reversed Phase ◽  
Gas Chromatography Mass Spectrometry ◽  
Phase Gradient ◽  
Chromatography Mass Spectrometry

Recently it has been shown that in man the oral couma-rin anticoagulant phenprocoumon is eliminated up to 60-70 % in urine and 30-40 % in faeces; in urine phenprocoumon (PH) and its metabolites 7-hydroxy-(7-OH),6-hydroxy-(6-OH) and 4'-hydroxy-(4'-OH) phenprocoumon are present mainly as conjugates. No data so far were available on the biliary excretion of these compounds.We examined bile obtained from four in-patients during PH treatment; bile samples were aspirated in the duodenum at the papilla during routine diagnostic endoscopy and immediately deep frozen before analysis. Samples were extracted both untreated as well as after hydrolysis with 6-glucuronidase/aryl sulfatase and separated by reversed phase gradient elution high performance liquid chromatography (HPLC) with fluorescence detection; for confirmation, the same extracts were methylated and analysed by gas chromatography-mass spectrometry (CG-MS) (J.X.de Vries et al J Chromatogr., 338 (1985) 325). PH, 7-OH, 6-OH and 4'-OH were identified by comparison with synthetic authentic samples'''''''

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