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Biomedical subjects

E Gelpi

Publications and source records attributed to E Gelpi.

At least 73 records · Page 4Linked to original sources

Increased synthesis of systemic prostacyclin in cirrhotic patients.

Urinary excretion of two prostacyclin metabolites was investigated in 48 subjects: 8 controls and 40 cirrhotics (9 without ascites, 22 with ascites and preserved renal function, and 9 with functional renal failure). Urinary 6-keto-prostaglandin F1 alpha (6-keto-PGF1 alpha), believed to reflect renal prostacyclin production, was significantly increased in patients without ascites and in ascitic patients with preserved renal function, but cirrhotics with renal failure showed rates similar to controls. Excretion of 2,3-dinor-6-keto-PGF1 alpha (PGI-M), the major urinary metabolite of systemic prostacyclin, was increased in all groups of patients, including those with renal failure. A single dose of sulindac, a renal-sparing prostaglandin synthesis inhibitor, reduced PGI-M but not 6-keto-PGF1 alpha in 5 cirrhotic patients. This would be consistent with the predicted renal origin of the latter and the systemic origin of the former. Ascitic patients with high urinary excretion of PGI-M (above the median value) showed significantly lower mean arterial pressure and higher plasma renin activity and aldosterone than patients with excretion below the median. Urinary 6-keto-PGF1 alpha was higher in patients with low PGI-M. Finally, creatinine clearance corrected excretion of PGI-M, as an estimation of relative plasma levels correlates both with plasma renin activity and plasma aldosterone in the 31 subjects who presented with ascites. It is suggested that enhanced synthesis of systemic prostacyclin may influence hemodynamic changes in patients with liver cirrhosis. Overproduction of systemic prostacyclin in the absence of increased renal prostacyclin synthesis appears to be characteristic of patients with functional renal failure.

6-Ketoprostaglandin F1 alpha↗

Study of leucine-enkephalin in rat brain by a new high-performance liquid chromatographic method.

A specific method is presented for the assay of leucine-enkephalin (Leu-Enk) and its metabolites by reversed-phase high-performance liquid chromatography on a muBondapak C18 column with a mobile phase of methanol-water and 0.005 M tetrabutylammonium phosphate. The four substances are resolved by a linear program from 8 to 70% methanol in 25 min. Detection is achieved by monitoring the absorbance at 280 nm. Time of analysis can be reduced by means of a new high-speed liquid chromatography package. This method allows the study of the effect of Phe-Ala on the cerebral metabolism of Leu-Enk. For this purpose, membrane preparations from rat striatum were incubated in the presence of [3H]Leu-Enk with different concentrations of Phe-Ala from 10(-7) to 10(-3) M during 1 h at 37 degrees C. Collected eluates were counted by liquid scintillation. The results suggest the presence of three membrane enzymes which generate the three metabolites, Tyr, Tyr-Gly-Gly and Tyr-Gly, in order of abundance. Maximum inhibition of [3H]Leu-Enk degradation is obtained at a concentration of 10(-3) M Phe-Ala.

Animals↗

Quantitative profiling of the metabolic cascade of arachidonic acid by capillary gas chromatography mass spectrometry.

The analytical approach described allows for the rapid screening and concurrent quantitative determination of all of the major metabolites of the cyclooxygenase pathway of arachidonic acid, including the primary prostaglandins PGE2, PGD2 and PGF2 alpha in addition to the stable end-products of short-lived prostacyclin and thromboxane A2 (6-keto-PGF1 alpha and TXB2, respectively), generated by incubating arachidonic acid with mouse peritoneal macrophage cells. After derivatization into the corresponding methyl ester-methoxime-TMS derivative, the extracted endogenous compounds were analysed by combined high efficiency glass capillary column chromatography and selected ion monitoring. Comparatively lower detection limits can be achieved with the methyl ester-butylboronate-TMS derivatives (e.g. 10 pg for 6-keto-PGF1 alpha). A brief account is also presented on a new type of mixed methyl ester-pentafluorobenzyloxime-butylboronate-TMS derivative of TXB2 and 5-keto-PGF1 alpha.

Animals↗

Prostaglandins E and F, and 19-hydroxylated E and F (series I and II) in semen of fertile men. Gas and liquid chromatographic separation with selected ion detection.

Low concentrations of prostaglandins (PG) could be related to male clinical infertility althought relevant experimental data are scarce. The aim of this work is to establish reliable seminal PG levels in fertile men by rigorous sample control, to prevent degradation, and by rapid and simple extraction and assay procedures. Single semen samples from healthy fertile men were immediately centrifuged (within 30 min of ejaculation) adding PGF2 alpha D4 to the seminal plasma as internal standard. The samples were next ultrafiltered and the PGs in the ultrafiltrate were derivatized with a mixture of N,O-bis(trimethylsilyl)trifluoroacetamide and piperidine (1:1) at 60 degrees for 30 min. Optimum gas-liquid chromatographic separation of all of the peaks of interest was achieved on 4 m x 1/4 in. I.D. Dexsil 300 packed columns at 280 degrees. The detection and quantitation of all the peaks of interest depends on the selected ion monitoring of specific masses. The values obtained (in micrograms/ml, range in parentheses) were: PGEs, 63.5 +/- 49.3 (9--164); PGFs, 2.6 +/- 1.92 (0.95--6.63); 19-OH PGEs, 592.6 +/- 312.5 (142.1--1047); and 19-OH PGFs, 12.66 +/- 5.21 (4--19). Individual values for members of both series I and II are also presented. The sample collection and extraction procedures were further checked by high-performance liquid chromatography on a muPorasil column, with individual isolation and collection of all of the PGs, including the 19-OH PGs not previously separted by liquid chromatography.

Adult↗

Derivative and graphical procedures for correction of irrelevant UV spectrophotometric absorption in changeable matrixes.

The application of direct zero-order UV spectrophotometric and graphical or derivative background correction methods to selected pharmaceutical preparations shows their relative advantages in different situations. The assay of the active components in a changeable matrix is a problem with formulations having a limited shelflife. Although the standard three-point correction procedures can provide accurate data, there are practical problems if the irrelevant absorption band is such that only the derivative approach gives enough resolving power for reliable qualitative and quantitative determinations.

Chemistry, Pharmaceutical↗

New derivatives of prostaglandin A1 and specific detection of prostaglandin A's and 190hydroxylated prostaglandin A's in human semen.

The derivatization of prostaglandins of the A series with 1:1 mixtures of bis-(trimethylsilyl)trifluoroacetamide and nitrogen-containing non-aromatic heterocyclics such as piperidine, pyrrolidine, morpholine and hexamethylenimine (1--4 h at 60--70 degrees C) gives new types of derivatives, designated as 11-heterocycle, 9-enol PGA (TMS)3. These derivatives show very simplified and characteristic mass spectral patterns strikingly dominated by a common [M-173]+ fragment ion and easily detectable by selected ion monitoring. This feature allows the concurrent analytical detection of both prostaglandin A's and 19-hydroxy prostaglandin A's in biological samples. In this case 2 ml samples of human semen were extracted by direct ultrafiltration on a Pellicon membrane with a nominal molecular weight limit of 1000. The prostaglandins in the approximately or equal to 1.6 ml of ultrafiltrate thus obtained were recovered in ethyl acetate, derivatized as indicated above and detected by monitoring of the corresponding [M-173]+ ions.

Azepines↗

Direct gas chromatograph-mass spectrometer connection of glass capillary columns for the analysis of serotonin and metabolites by selective ion monitoring.

A direct connection system that requires no modifications to either the gas chromatograph or the mass spectrometer is described for the coupling of glass capillary columns to a commercial mass spectrometer. Vacuum-tight connections can be readily achieved with standard 1/16-in. fittings and septum discs. The system can be operated with column outlet helium flow-rates of 1.5-2 ml/min at ion source pressures of 2-5 - 10(-5) mmHg. Although an injector splitter of our own design has been used, the initial test results obtained with a splitless injection method based on the use of a solids injection syringe together with a packed column injector assembly indicate the feasibility of developing a totally splitless and connection system that is simple to use. The system has been applied to the detection of the pentafluoropropionyl derivatives of serotonin, methoxytryptamine, methoxytryptophol and 5-hydroxytryptophol by selective ion monitoring techniques. Column performances under both sets of conditions (gas-liquid chromatography alone and combined with mass spectrometry) are compared and the evaluation of direct versus indirect coupling showed yields of the order of 50% through the molecular separator.

Chromatography, Gas↗

Profiles of prostaglandins A, B, E and F (series I and II) obtained by gas chromatography with multiple-ion detection.

A new method is presented for the simultaneous one-step derivatization of prostaglandins A1, A2, B1, B2, E1, E2, F1alpha and F2alpha with N,O-bis(trimethylsilyl)trifluoroacetamide-piperidine. The procedure provides stable and reproducible trimethylsilyl derivatives that show good chromatographic properties on OV-17 and OV-225 columns, and precludes any significant degree of interconversion of the prostaglandins. The prostaglandins F, B and E are determined as the corresponding tetrakis(trimethylsilyl), bis(trimethylsilyl) and 9-enol-tetrakis(trimethylsilyl) derivatives, respectively. The enolization of prostaglandins E proceeds quantitatively only if piperidine is added to the acetamide. This mixture also produces the novel derivative 9-piperidyl-tris(trimethylsilyl)prostaglandin A. Evidence is also presented on the quantitative injection losses due to the type of syringe used (1micronl or 10 micronl). A modified extraction procedure has been applied to samples of human seminal fluid. The procedure, which involves the centrifugation of the samples, allows the recovery of all of the prostaglandins in one fraction, which is then derivatized and screened by combined gas-liquid chromatography-multiple-ion detection techniques in order to identify the individual compounds.

Chromatography, Gas↗

Gas chromatographic separation of histamine and its metabolites.

A new, rapid and simple gas-liquid chromatographic method is described for the simultaneous determination of histamine, 1-methylhistamine, 4-imidazoleacetic acid and 1-methyl-4-imidazoleacetic acid. These compounds are derivatized at 60 degrees in one fast 30-min reaction with bis(trimethylsilyl)acetamide-trimethylchlorosilane and are then separated on OV-17. No previous methylation of the acids is needed. A complete profile of the four metabolites can be obtained in 8 min with baseline resolution, and there are no tailing effects. The quantitative response curves are linear in the nanogram range, with detection limits of the order of 10 ng for the acids and 100 ng for the amines. The structures of these new trimethylsilyl derivatives have been verified by combined gas-liquid chromatography-mass spectrometry.

Chromatography, Gas↗

Mass spectrometric determination of new prostaglandin derivatives (series A and E).

The reaction of prostaglandins E1, E2, A1 and A2 with a 1:1 mixture of N,O-bis-trimethylsilyltrifluoroacetamide and piperidine favours the quantitative enolization of the 9-keto group of prostaglandins E, thus giving the corresponding tetrakis-TMS derivatives with molecular ions at m/e 642 and m/e 640 for prostaglandins E1 and E2, respectively, and also gives a new type of prostaglandin A-piperidyl-TMS derivative characterized by very strong base peaks at m/e 464 and 462, with molecular ions at m/e 637 and 635 for prostaglandins A1 and A2, respectively. The probable reaction mechanisms prevailing in both cases are discussed in accordance with the low and high resolution mass spectral data presented.

Chemical Phenomena↗

Adsorption of tryptophan metabolites from physiological fluids on XAD-2 and determination by single ion monitoring.

Endogenous tryptamine, 5-hydroxytryptamine, indoleacetic acid, 5-hydroxyindoleacetic and tryptophan have been recovered from urine and cerebro-spinal fluid by adsorption on XAD-2 resin (0.3 g). After adsorption of the sample on the resin, desorption with methanol provides a single fraction that contains all of these metabolites. The mass spectra of their pentafluoropropionyl derivatives show prominent ions at m/e 276 and 438 which are characteristic of indoles and 5-hydroxyindoles, respectively, a feature that allows the concurrent determination of all the components of each group by functional group analysis. A method has been developed to carry out single ion monitoring with the peak matching system of an Hitachi RMU-6H mass spectrometer. Identifications are based on the respective Kovats Indices and single ion monitoring of two characteristic ions per compound: tryptophan (m/e 276 and 347); tryptamine (m/e 276 and 289); indoleacetic acid (m/e 276 and 335); 5-hydroxytryptamine (m/e 438 and 451); 5-hydroxyindoleacetic acid (m/e 438 and 497). The method described illustrates the feasibility of assaying biogenic indoleamines and acidic metabolites, as well as their precursor amino acid on a single fraction in contrast to other standard fractionation methods. This is possible even if the mass spectrometer is not equipped with an alternating voltage accelerator provided that it has a peak matcher, although the lack of an alternating voltage accelerator requires two separate injections of the same sample, for quantification and identification; one for the indole profile and another for the 5-hydroxyindole profile. Both profiles can be verified by individual monitoring of the other confirmatory ions. With this method the use of a multiple ion detector would allow a simultaneous determination of all of these metabolites in one gas chromatograph mass spectrometer run.

Adsorption↗

Applications of gas chromatography and mass spectrometry in neurochemical studies: determination of indole amine profiles at the picogram level.

Work is presented on the application of sensitive and specific analytical methods for the simultaneous assay of tryptophan and its indolic metabolites (5-hydroxytryptophan, serotonin, 5-hydroxy-indole-acetic acid, tryptamine and indoleacetic acid). Due to their small concentrations in selected areas of the central nervous system, suitable derivatives have been prepared for separation by Gas Chromatography (GC) and detection by electron capture (ECD). Carboxylic acid groups have been methylated with BCl3-methanol and phenolic and amine groups have been acylated to pentafluoropropionyl derivatives. This type of derivatives allow a detectability limit of 5 picograms (5 x 10(-12) gr) if the corresponding reaction kinetics and yields are properly optimized. Response curves are presented. Identifications have been obtained by combined GC-MS and by the use of Kovats indeces in two GC stationary phases. The GC separation of the six compounds indicated above can be performed in less than 5 minutes with quantitative reproducibility. This work opens the possibility of performing comparative studies ("screening") of various physiological and pathological conditions by detecting any changes in the profile representative of a whole metabolic pathway, such as that of the indolic compounds in this case.

Chromatography, Gas↗