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Oxygen free radicals and prostaglandins are implicated in the pathophysiology of acute pancreatitis, although their mechanisms of action remain unclear. We have studied the effect of administration of exogenous 16,16-dimethyl prostaglandin E2 and superoxide dismutase on oxygen free radical production in acute pancreatitis. For this purpose, five experimental rat groups were studied: group I, control; group II, sodium taurocholate-induced acute pancreatitis; group III, same as group II but with previous administration of 16,16-dimethyl prostaglandin E2; group IV, same as group II but with previous administration of indomethacin; and group V, same as group II but with previous administration of superoxide dismutase. In sodium taurocholate-treated rats, xanthine dehydrogenase is completely converted to xanthine oxidase within the first 5 min with subsequent oxygen free radical production while in 16,16-dimethyl prostaglandin E2-treated rats this enzyme transformation does not occur. In the superoxide dismutase-treated group xanthine oxidase activation is partially prevented. These data suggest that xanthine oxidase is the main source of oxygen free radicals, which contribute to extending the cellular damage in sodium taurocholate-induced acute pancreatitis.

1 Polychlorinated dioxins (PCDDs) and furans (PCDFs) are known to produce a wide range of toxic effects. 2 PCDDs and PCDFs are typical contaminants of chlorinated phenols, and pentachlorophenol and related compounds have been shown to be widely distributed among selected oil samples taken from the 1981 Spanish toxic oil epidemic. 3 Six control and eight case oil samples were analysed using GC/MS for PCDDs and PCDFs. Only small concentrations, normally below 1 ng g-1, of the higher chlorinated PCDDs and PCDFs were detected. There were no statistical differences between the case and control oils. 4 These levels seem to be too low to elicit toxic effects, although they could be enough to potentiate the toxicity of other xenobiotics present in the oils. However, it is uncertain whether the levels of these compounds measured in 1990 reflect the levels present when the oils were consumed in 1981, or whether or not the levels measured in crude oils are representative of fried oils.

Eicosanoid production by uterine strips and by embryos obtained from normal and diabetic rats at day 10 of pregnancy was studied. It was found that the release of 6-keto-PGF1 alpha (representing PGI2 synthesis) and of LTB4 was less in preparations from diabetic animals than in controls. The production of TXB2 (indicating the formation of TXA2) by uterine tissue obtained from diabetic rats was almost double that of controls. The synthesis and release of eicosanoids when tissues were incubated in glucose-containing solution or in glucose-free medium were similar, with the exception of LTB4, which was diminished with uterine strips from diabetic rats. The mean number of embryos in control pregnant rats (12.4 +/- 0.5) and in diabetic mothers (10.1 +/- 1.3) was not significantly different, but in 4 of the 14 diabetic rats studied, all of their embryos were resorbed. Although embryos released large amounts of PGF2 and PGE2, and small amounts of 6-keto-PGF1 alpha, TXB2 and LTB4, the amounts of each eicosanoid in control and diabetic groups were similar. The present results indicate that the diabetic state, which induces alterations in uterine eicosanoid production, do not influence arachidonic metabolism in their corresponding embryos.

Several studies have reported that prostanoids are involved in many of the physiopathological mechanisms underlying acute pancreatitis but their precise role in this disease remains to be established. The objective of this work is to evaluate the variation of local tissue production of prostanoids and lipoxygenase metabolites of arachidonic acid in acute pancreas inflammation induced by intraductal administration of 3.5% sodium taurocholate (0.1 ml/100 mg body weight) in rats. Pancreatic tissue levels of leukotriene B4 (LTB4), 15 hydroxyeicosatetraenoic acid (15-HETE), 6-keto prostaglandin F1 alpha (6-keto PGF1 alpha), thromboxane B2 (TXB2) and prostaglandin E2 (PGE2) were determined by HPLC-RIA techniques at 5 and 60 minutes after induction of acute pancreatitis (AP). Prostanoids increased significantly at 5 minutes and LTB4 and 15-HETE at 60 minutes. These data confirm that the prostanoid imbalance could be considered as an early specific response of the pancreas to the inflammatory events characteristic of induced AP while the altered levels of the lipoxygenase products (LTB4 and 15-HETE) would be more of a nonspecific organ response associated to the high cellular infiltration rate and necrosis observed in the late phases of acute pancreatitis.

N-phenyllinoleamide (NPLA) has been detected as extraneous compound in adulterated cooking oils associated with a unique epidemic disease known as the Toxic Oil Syndrome (TOS). In this communication we report on the action of NPLA on the endogenous cyclooxygenase and lipoxygenase arachidonate metabolism. Results show that mouse peritoneal macrophages (MPM) exposed to 1 mM NPLA for 2 h undergo significant increases of 6-keto prostaglandin F1a, prostaglandin E2, leukotriene B4, 12- and 15-hydroxyeicosatetraenoic acids. MPM prelabelled with 3H-AA showed an enhanced release when exposed to NPLA. Thus, it is concluded that NPLA potentiates AA release from cell membrane phospholipids and the subsequent cyclooxygenase and lipoxygenase oxidative metabolism of this precursor to various eicosanoids. This is in agreement with the implication of peroxidative process mediated by fatty acids anilides in TOS.

Systemic prostacyclin and thromboxane A2 production in rat experimental acute pancreatitis has been evaluated by measuring the urinary excretion of the 2,3-dinor 6-keto prostaglandin F1 alpha and 2,3-dinor thromboxane B2, respectively. Acute pancreatitis was induced by intraductal administration of 4.5% sodium taurocholate (0.1 ml/100 mg body weight) and intravenous cerulein perfusion (5 micrograms/kg/hr) for 6 hr, respectively. Urinary excretion of 2,3-dinor 6-keto prostaglandin F1 alpha and 2,3-dinor thromboxane B2 were much more important in sodium taurocholate- than in cerulein-induced acute pancreatitis. These data confirm an altered prostacyclin and thromboxane metabolism occurring in experimental acute pancreatitis. Phospholipase A2 activity and the effect of gabexate mesilate on the arachidonate metabolism were also evaluated.

N-phenyllinoleamide (NPLA), the anilide of linoleic acid, has been associated with the epidemiology of Toxic Oil Syndrome, but no data are available on its metabolism. On account of the similarity in chemical structure between the linoleic acid and NPLA, the aim of this study has been to investigate the oxidative metabolism of this xenobiotic by the human nasal polyp, a tissue with elevated 15-lipoxygenase activity. For this purpose, tissue homogenates have been incubated for 2 h with NPLA (0.1 mM) spiked with either N-(ring G-3H)PLA (0.2 microCi/ml) or N-P(1-14C)LA (0.05 microCi/ml). Gas chromatographic/mass spectrometric analysis of the high performance liquid radiochromatographic fractions shows that the 9,12,13-trihydroxy, 12,13-epoxy-11-hydroxy and 13-hydroxy NPLA derivatives are the major metabolites. These results revealed that NPLA metabolites are chemical structures related to the linoleic acid derivatives, some of which may show biological activity.

A commercial automated solid-phase extraction system for cyclooxygenase arachidonic acid metabolites in urine samples has been evaluated. Comparison of manual and automatic batch (36 samples) extraction procedures for tritium labelled prostanoids added as tracers to urine samples has shown equivalent results with recoveries greater than 90% for prostaglandins E2, F2alpha and 6-keto prostaglandin F1alpha as well as thromboxane B2. Analyte stability is not affected by the automated procedure, which uses less solvents and has a faster overall processing time than the manual method. The automated system has been applied to the extraction of prostanoids in urine samples from workers exposed to dichloroethane.

Even in specific risk groups, the relation between exposure to organic solvents and chronic renal diseases remains controversial. Thus, in a collaborative European study, we assessed the renal effects of occupational exposure to perchloroethylene (PCE) in dry-cleaners compared with matched controls who were simultaneously examined. Single high and low molecular weight proteins, kidney-derived antigens and enzymes, and prostanoids were measured in urine. beta 2-microglobulin, creatinine, laminin fragments, and anti-glomerular basement membrane antibodies were also measured in serum. A canonical function based on 23 such variables correctly classified 93% of individuals as either PCE-exposed or controls; with 13 markers, group membership was identified in 87% of subjects. Increased high molecular weight protein in urine was frequently (17/50 vs 1/50, p less than 0.0001) associated with tubular alterations. Changes were consistent with diffuse abnormalities along the nephron in workers exposed to low levels of PCE (median 15 parts per million). Generalised membrane disturbances might account for the increased release of laminin fragments, fibronectin, and glycosaminoglycans, for high molecular weight proteinuria, and for the increased shedding of epithelial membrane components from tubular cells with different location along the nephron (brush-border antigens and Tamm-Horsfall glycoprotein). These findings of early renal changes indicate that solvent-exposed subjects, especially dry-cleaners, need to be monitored for the possible development of chronic renal diseases.

This study was carried out to investigate the proportion of the 6-keto prostaglandin F1 alpha (6-keto PGF1 alpha) and thromboxane B2 (TXB2) alteration that is due to ischemia in pancreas transplantation against the proportion due to reperfusion. For this purpose, Lewis rats were divided in three experimental groups: Group I = Control, Group II = Donor pancreas subjected to 15 minutes of cold ischemia, Group III = Same as group II but pancreas were transplanted to the recipient individual and then subjected to reperfusion. The results indicate that increases in pancreas 6-keto PGF1 alpha occur as a consequence of cold ischemia while TXB2 remains unchanged. When blood flow was restored, 6-keto PGF1 alpha remained unchanged compared to the ischemic group while pancreatic levels of TXB2 were significantly increased. These results suggest a different induction of prostanoid metabolism during ischemia and reperfusion in pancreatic tissue.

The total body production of prostacyclin was shown to be increased in cirrhotic patients, suggesting that its synthesis by blood vessels of the systemic circulation is enhanced. However, the mechanism by which the synthesis of systemic prostacyclin is stimulated is not known. The present study investigated the urinary excretion of 2,3-dinor-6-keto-PGF1 alpha, an index of total body prostacyclin synthesis, first, in cirrhotics with portal hypertension (n = 19) as compared with cirrhotics with reduced portal pressure after portacaval shunt surgery (n = 18) and with control noncirrhotic subjects (n = 11), and; second, in cirrhotics before and after intestinal decontamination by oral nonabsorbable antibiotics (n = 9 antibiotic treated patients, n = 10 control nontreated cirrhotics). Control noncirrhotic subjects showed lower urinary excretion of 2,3-dinor-6-keto-PGF1 alpha than both groups of cirrhotics (P less than 0.001). Interestingly, urinary excretion of 2,3-dinor-6-keto-PGF1 alpha was significantly higher in cirrhotics with portacaval shunt than in those with portal hypertension (P less than 0.01). The urinary excretion of 2,3-dinor-6-keto-PGF1 alpha decreased significantly after intestinal decontamination in the antibiotic-treated group (580.1 +/- 232.4 vs. 431.2 +/- 219.2 pg/mg creatinine; P less than 0.05) but not in nontreated patients (543.9 +/- 214.4 vs. 581.2 +/- 281.4 pg/mg creatinine; P = NS). These data suggest that the increased urinary excretion of 2,3-dinor-6-keto-PGF1 alpha observed in cirrhotics is not directly related to portal hypertension itself but to portal blood factors that bypass the liver. Some such factors may be of intestinal bacterial origin.

The purpose of this study is to examine the "in vivo" release of 15-HETE and other arachidonic acid metabolites in nasal secretions following a challenge with "Dermatophagoides Pteronyssinus" in patients with allergic rhinitis and non-allergic controls. In addition, we examine the effects of a membrane stabilizer, such as sodium cromoglycate, on these metabolites. Thirteen allergic subjects and seven healthy controls are studied. 15-HETE, peptide leukotrienes, LTB4, PGD2, PGE2 and PGF2 alpha levels are evaluated before and after nasal challenge in sodium cromoglycate treated and untreated subjects. This study provides "in vivo" evidence that the pathophysiological responses to nasal antigen challenge could be related to the release of 15-HETE as well as other arachidonic acid metabolites, mainly arising from the lipoxygenase pathway.

Significant increases of TXB2 and PGE2 are reported to occur in pancreas transplantation. These increases are prevented with scavengers of oxygen-free radicals. In this communication, we report on changes of prostacyclin metabolites such as tissue 6-keto prostaglandin F1 alpha and urinary 2,3-dinor 6-keto prostaglandin F1 alpha in rats subjected to pancreas transplantation after different periods of organ cold preservation ischemia as well as the effect of superoxide dismutase (SOD) on these changes. For this purpose, male Lewis rats were classified as follows: Group I, Control; Group II, syngenic pancreas transplantation after 15 min of organ preservation in Collins solution at 4 degrees C; Group III, same as II but with 12 hours of organ preservation; Group IV, same as III, but with SOD pretreatment. Results have shown significant posttransplantation increases of both tissue 6-keto PGF1 alpha and urinary 2, 3 dinor 6-keto PGF1 alpha, the latter being a useful marker to evaluate systemic prostacyclin (PGI2) production by rat pancreas. This effect was prevented when the organ had been exposed to SOD during the period of cold preservation ischemia. These results confirm the implication of oxygen-free radicals (OFR) in the ischemia-reperfusion process associated to rat pancreas transplantation leading to enhanced arachidonic acid metabolism.

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.

Prostaglandins E2 and F2 alpha were estimated in nasal secretions from ten healthy volunteers by high performance liquid chromatography and radioimmunological assay. Prostaglandin concentrations determined in five consecutive nasal washes with saline at room temperature showed that the nasal mucosa was stimulated after instillation. A substantial increase of basal levels was associated with the second nasal lavage. In all volunteers aspirin treatment inhibited prostaglandin release.

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.

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.

The effects of predictable and unpredictable shock on concentrations of serotonin (5-hydroxytryptamine, 5-HT), 5-hydroxyindoleacetic acid (5-HIAA), tryptophan (TP) and noradrenaline (NA) have been studied in 7 regions of rat brain. Two separate experiments have been carried out determining these substances both at 30 min and 2 h after the stress session. Unpredictable shock depleted NA levels in all brain regions except the striatum. However, at 2 h poststress NA in these regions increased significantly in comparison with both controls and predictably shocked rats. Predictable shock also decreased NA in locus coeruleus, brainstem and hypothalamus, which was not observed 2 h later. Both predictable and unpredictable shock decreased 5-HT in brainstem and hypothalamus. At 2 h poststress, 5-HT levels in these regions were still decreased in predictably shocked rats, but had attained control values in unpredictably shocked rats. 5-HT metabolism expressed as the 5-HIAA/5-HT ratio, was significantly increased 30 min after predictable shock in all regions except the locus coeruleus and hippocampus. Unpredictable shock produced a much more marked increase in 5-HIAA/5-HT ratio. At 2 h poststress 5-HT metabolism returned to control values in most of the brain regions of predictably shocked animals, but it remained high after unpredictable shock. The activation of serotonergic metabolism following each type of shock is different according to the nucleus in which the 5-HT nerve endings originate. Only slight increases in tryptophan were observed after both types of shock. Our results suggest that unpredictable shock is perceived as a more anxiogenic situation and that under this condition both 5-HT and NA levels are more effectively normalized with time.

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.

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.

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.

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.

We analysed the production of prostaglandin E2 (PGE2) by monocytes in 11 drug addicts with AIDS and opportunistic infections and in 13 controls. In three patients and six controls we isolated the PGE2 fraction from culture supernatants of purified monocytes (greater than 95%) incubated in medium containing (3H) arachidonic acid, using silicic acid columns. In two of three patients PGE2-fraction values (21.9 and 21.6 pmol/g protein) were significantly higher than controls (10 +/- 4). In eight AIDS patients and seven controls, PGE2 levels were determined by high-performance liquid chromatography and radioimmunoassay (HPLC and RIA). In three out of eight patients PGE2 levels were markedly higher (736, 419 and 208 pg/ml) than the mean + 2 s.d. values from controls (73 +/- 51). We tested the effect of PGE2 on the production of interleukin-2 by normal phytohemagglutinin-stimulated peripheral blood lymphocytes (PBL). There was a significant suppression of interleukin-2 production by PGE2 released from AIDS monocytes. PGE2 may be one of the factors involved in the severe immune dysfunction associated with AIDS.

19-hydroxy-prostaglandins and prostaglandins of the E series (19-OH PGEs) were estimated in the seminal plasma of asthenozoospermic patients (n = 15) and individuals affected by prostatitis (n = 10) and compared to controls (n = 13) and secretory azoospermic patients (n = 8). All of them were free from infections (except individuals affected by prostatitis), biochemical and ultrastructural problems. The results indicate that endogenous prostaglandin levels (19-OH PGEs and PGEs) bear no correlation either to motility or absence of spermatozoa. Significant increases of PGEs were observed in patients affected with prostatitis. Surprisingly PGE levels showed no correlation with the levels of 19-OH PGEs.

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.

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.

The effects of 1 h/day restraint in plastic tubes for 24 days on the levels of serotonin (5-HT), 5-hydroxyindoleacetic acid (5-HIAA), tryptophan (TP), and noradrenaline (NA) in six regions of rat brain 20 h after the last restraint period were investigated. The levels of 5-HT, 5-HIAA, and NA but not TP increased in several regions. The effects of 1 h of immobilization on both control and chronically restrained rats were also studied. Immobilization per se did not alter brain 5-HT, 5-HIAA, and TP levels, but decreased NA in the pons plus medulla oblongata and hypothalamus. However, immobilization after chronic restraint decreased 5-HT, increased 5-HIAA, and decreased NA in most brain regions in comparison with values for the chronically restrained rats. We suggest that chronic restraint leads to compensatory increases of brain 5-HT and NA synthesis and sensitizes both monoaminergic systems to an additional acute stress. These changes may affect coping with stress demands.

The early identification of fatty acid anilides in suspect oils directed attention to their possible role in the Spanish toxic oil syndrome. These anilides or their oxidized derivatives could have been spontaneously formed during the handling and/or storage of the oil. The exact cause of the intoxication is still unknown but free radical and peroxidative mechanisms have been implicated in its etiology. Epoxy-hydroxylated derivatives from linoleic acid anilide were obtained using a model of accelerated oxidation. Their mass spectral patterns agree with the trimethylsilyl ethers of N-phenyl-9,10-epoxy-11-hydroxy-12-octadecenamide and N-phenyl-12,13-epoxy-11-hydroxy-9-octadecenamide, respectively. These compounds were also identified is rapeseed oil samples supplemented with N-phenyl-linoleamide and submitted to the reported accelerated oxidation method.

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.

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.

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.

The hydrocarbon composition of two algae, a golden-brown (Bot-ryococcus braunii) and a blue-green (Anacystis montana), has been investigated by gas chromatography-mass spectrometry. Both show distributions of aliphatic hydrocarbons of odd carbon numbers in the medium and high ranges of molecular weight, with maxima at n-C(17) and n-C(29) for B. braunii and n-C(17) and n-C(29) for A. montana. With the exception of the n-heptadecane of A. montana all the hydrocarbons are monoenes, dienes, or trienes. Since certain continental sediments and oils show similar distributions of alkanes with respect to carbon number, these organisms may be the precursors of the hydrocarbons in these formations.

The composition and nature of the fatty acids and hydrocarbons of Sarcina lutea were elucidated by gas chromatography and by combined gas chromatography-mass spectrometry. The distribution of fatty acids found in S. lutea showed two families of pairs, or dyads, of saturated monocarboxylic acids (C12-C18) with and without methyl branching. These pairs of fatty acids showed a pattern of iso and anteiso structures for C13, C15, and C17, and iso and normal structures for C12, C14, and C16. Only the C18 showed unsaturation. The distribution of hydrocarbons in the range C22-C29 showed two families of tetrads of unsaturated aliphatic hydrocarbons all showing methyl branching. Each tetrad was composed of four isomers identified as two iso olefins and two anteiso olefins. The only difference between the tetrads pertaining to different families was found in the relative gas chromatographic retention times of the last two components of each group.

Gas chromatography and combined gas chromatography-mass spectrometry have been used to study the fatty acids and hydrocarbons of a bacterium from the Pacific Ocean, Vibrio marinus, a freshwater blue-green alga, Anacystis nidulans, and algal mat communities from the Gulf of Mexico. Both types of microorganisms (bacteria and algae) showed relatively simple hydrocarbon and fatty acid patterns, the hydrocarbons predominating in the region of C-17 and the fatty acids in the range of C-14 to C-18. The patterns of V. marinus were more comparable to those of the algal populations than to patterns reported for other bacteria. An incomplete correlation between fatty acids and hydrocarbons in both types of organisms was observed, making it difficult to accept the concept that the biosynthesis of hydrocarbons follows a simple fatty acid decarboxylation process.

The nonpolar (acetone-soluble) lipids of the extremely halophilic bacterium, Halobacterium cutirubrum, were found to consist of red carotenoid pigments (43%) and squalenes (48%) with a small amount of a vitamin K-type quinone. The squalenes were shown by n.m.r. and mass spectra to consist of the fully isoprenoid squalene (S; C(30)H(50)), dihydrosqualene (S(2); C(30)H(52)), and tetrahydrosqualene (S(4); C(30)H(54)) in the ratio of 1.0:0.4:0.1. S(2) probably has one reduced internal isoprenoid group, and S(4) has one internal and one terminal reduced isoprenyl group. The vitamin K-type quinone was shown by n.m.r. and mass spectra to have a C(40) isoprenoid side chain, and is thus identified as menaquinone-8 (MK-8).

The Pueblito de Allende meteorite contains only 0.1-0.5 ppm of solvent-extractable carbon, most or all of which is due to terrestrial contamination. Determinations of the total carbon content gives values from 0.23 to 0.35 per cent. The presumed indigenous insoluble carbon is thus present in a ratio of 2300:1 relative to contaminant carbon. A sample of the meteorite was extracted with a deuterated solvent containing benzene:methanol (4:1 w/w) to remove contaminants and then was subjected to pyrolysis-gas chromatography-mass spectrometry to investigate the nature of the remaining carbon. The pyrolysis products (about 20 ppm) consisted chiefly of aromatic and substituted aromatic hydrocarbons. The pyrolysis results were confirmed by oxidative thermal analysis which showed that the bulk of the carbon present was a macromolecular structure and not graphite. This suggests that an insoluble nongraphitic condensed aromatic polymer is indigenous to the meteorite. The origin and precise chemical structure of this material has not been determined, but it bears a superficial resemblance to coal-like structures.

It appears unlikely that the organic material detected in the meteorite that fell in Mexico last year can have been introduced by contamination.