Search

Palmitoylethanolamide (PEA, N -hexadecanoylethanolamide) is an endogenous compound belonging to the family of N -acylethanolamines. PEA has anti-inflammatory and analgesic properties and is very well tolerated in humans. In the present article, the basal pharmacology of PEA is reviewed. In terms of its pharmacokinetic properties, most work has been undertaken upon designing formulations for its absorption and upon characterising the enzymes involved in its metabolism, but little is known about its bioavailability, tissue distribution, and excretion pathways. PEA exerts most of its biological effects in the body secondary to the activation of peroxisome proliferator-activated receptor-α (PPAR-α), but PPAR-α-independent pathways involving other receptors (Transient Receptor Potential Vanilloid 1 (TRPV1), GPR55) have also been identified. Given the potential clinical utility of PEA, not least for the treatment of pain where there is a clear need for new well-tolerated drugs, we conclude that the gaps in our knowledge, in particular those relating to the pharmacokinetic properties of the compound, need to be filled.

Camptothecin (CPT) is an important topoisomerase I enzyme (Topo I) targeting anti-cancer drug, but its oral administration is limited by poor bioavailability and severe side effects. In this study, a redox sensitive CPT prodrug loaded solid lipid nanoparticles (SLN) system for oral delivery was developed. First of all, CPT-palmitic acid conjugate via a cleavable disulfide bond linker (CPT-SS-PA) was synthesized and encapsulated into SLN. The drug release of SLN was evaluated in neutral environment, simulated gastrointestinal fluid and reductive solution. The results indicated that CPT-SS-PA SLN maintained chemical structural stability in simulated physiological environment but exhibited quick reduction-response release of CPT in the presence of dithiothreitol. Furthermore, in vitro cytotoxicity of CPT-SS-PA SLN was tested against cancer cell lines, and the cellular uptake behavior for oral delivery was checked by confocal laser scanning microscopy (CLSM) using Caco-2 cells model. From the data, CPT-SS-PA SLN revealed high anti-cancer activity and enhanced Caco-2 cell absorption. Finally, the oral bioavailability and intestinal safety of CPT-SS-PA SLN were preliminary evaluated by in vivo pharmacokinetic and histopathological study, respectively. This study demonstrated that CPT-SS-PA SLN could be developed as an effective CPT oral delivery system due to its enhanced oral bioavailability and reduced intestinal side effect., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Scope: Fatty acids (FAs) may affect endothelial cell (EC) function, influencing atherogenesis and inflammatory processes. Palmitoleic acid (POA) has been described as an anti-inflammatory FA. However, its effects on ECs are underexplored. This study compares the effects of POA with those of palmitic acid (PA) and oleic acid (OA) on EC inflammatory responses., Methods and Results: EAHy926 cells (EC lineage) are exposed to PA, OA, or POA, and stimulated with tumor necrosis factor (TNF)-α. Associated with the FA's own incorporation, PA induces a twofold increase in arachidonic acid, while POA increases the amount of cis-vaccenic acid. PA, but not OA, enhances the production of IL-6 and IL-8 in response to TNF-α. In contrast, POA decreases production of monocyte chemotactic protein (MCP)-1, IL-6, and IL-8 compared to PA. TNF-α increases surface intercellular adhesion molecule-1 expression previously decreased by POA. TNF-α stimulation increases the expression of NFκB, cyclooxygenase (COX)-2, MCP-1, and IL-6 genes and reduces the expression of peroxisome proliferator-activated receptor (PPAR)-α gene. PA enhances the expression of MCP-1, IL-6, and COX-2 genes, while POA downregulates these genes, decreases expression of NFκB, and upregulates PPAR-α gene expression., Conclusion: POA has anti-inflammatory effects on ECs stimulated with TNF-α and may counter endothelial dysfunction., (© 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

This work evaluates the potential of a 68 Ga labeled long chain 16C fatty acid for cardiac metabolic imaging. For radiolabeling with 68 Ga, hexadecanedioic acid was coupled with the chelator p-NH 2 -Bn-NOTA. Under the optimized conditions, NOTA-hexadecanoic acid could be radiolabeled with 68 Ga in ≥95% yields. In biodistribution studies carried out in Swiss mice, 68 Ga-NOTA-hexadecanoic acid showed low myocardial uptake at 2 min p.i. (3.7 ± 1.3%ID/g). While 68 Ga-NOTA-hexadecanoic acid cleared rapidly from non-target organs such as blood, lungs, intestine and kidney, wash out from liver was slow. Radio-HPLC analyses of myocardial extracts of rats injected with 68 Ga-NOTA-hexadecanoic acid confirmed its metabolic transformation in the myocardium., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

We evaluated the effects of ACEA (selective cannabinoid (CB) 1 receptor agonist), WIN 55,212-2 mesylate (WIN; non-selective CB 1 and CB 2 receptor agonist) and N-palmitoylethanolamine (PEA; an endogenous fatty acid of ethanolamide) in DBA/2 mice, a genetic model of reflex audiogenic epilepsy. PEA, ACEA or WIN intraperitoneal (i.p.) administration decreased the severity of tonic-clonic seizures. We also studied the effects of PEA, WIN or ACEA after co-administration with NIDA-41020 (CB 1 receptor antagonist) or GW6471 (PPAR-α antagonist) and compared the effects of WIN, ACEA and PEA in order to clarify their mechanisms of action. PEA has anticonvulsant features in DBA/2 mice mainly through PPAR-α and likely indirectly on CB 1 receptors, whereas ACEA and WIN act through CB 1 receptors. The co-administration of ineffective doses of ACEA, PEA and WIN with some antiepileptic drugs (AEDs) was examined in order to identify potential pharmacological interactions in DBA/2 mice. We found that PEA, ACEA and WIN co-administration potentiated the efficacy of carbamazepine, diazepam, felbamate, gabapentin, phenobarbital, topiramate and valproate and PEA only also that of oxcarbazepine and lamotrigine whereas, their co-administration with levetiracetam and phenytoin did not have effects. PEA, ACEA or WIN administration did not significantly influence the total plasma and brain levels of AEDs; therefore, it can be concluded that the observed potentiation was only of pharmacodynamic nature. In conclusion, PEA, ACEA and WIN show anticonvulsant effects in DBA/2 mice and potentiate the effects several AEDs suggesting a possible therapeutic relevance of these drugs and their mechanisms of action., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Palmitoylethanolamide (PEA) has been suggested to have useful analgesic properties and to be devoid of unwanted effects. Here, we have examined critically this contention, and discussed available data concerning the pharmacokinetics of PEA and its formulation. Sixteen clinical trials, six case reports/pilot studies and a meta-analysis of PEA as an analgesic have been published in the literature. For treatment times up to 49 days, the current clinical data argue against serious adverse drug reactions (ADRs) at an incidence of 1/200 or greater. For treatment lasting more than 60 days, the number of patients is insufficient to rule out a frequency of ADRs of less than 1/100. The six published randomized clinical trials are of variable quality. Presentation of data without information on data spread and nonreporting of data at times other than the final measurement were among issues that were identified. Further, there are no head-to-head clinical comparisons of unmicronized vs. micronized formulations of PEA, and so evidence for superiority of one formulation over the other is currently lacking. Nevertheless, the available clinical data support the contention that PEA has analgesic actions and motivate further study of this compound, particularly with respect to head-to-head comparisons of unmicronized vs. micronized formulations of PEA and comparisons with currently recommended treatments., (© 2016 The Authors. British Journal of Clinical Pharmacology published by John Wiley & Sons Ltd on behalf of The British Pharmacological Society.)

N-Palmitoylethanolamide showed great therapeutic potential in the treatment of inflammation and pain but its unfavourable pharmacokinetics properties will hinder its use in the clinical practice. A nanotechnology-based formulation was developed to enhance the probability of N-palmitoylethanolamide therapeutic success, especially in skin disease management. Lipid nanoparticles were produced and characterized to evaluate their mean size, ζ-potential, thermal behaviour, and morphology. The ability of N-palmitoylethanolamide to diffuse across the epidermis as well as anti-inflammatory and analgesic effects were investigated. Particles had a mean size of about 150 nm and a ζ-potential of -40 mV. DSC data confirmed the solid state of the matrix and the embedding of N-palmitoylethanolamide while electron microscopy have evidenced a peculiar internal structure (i.e., low-electrondense spherical objects within the matrix) that can be reliably ascribed to the presence of oil nanocompartments. Lipid nanoparticles increased N-palmitoylethanolamide percutaneous diffusion and prolonged the anti-inflammatory and analgesic effects in vivo. Lipid nanoparticles seem a good nanotechnology-based strategy to bring N-palmitoylethanolamide to clinics., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Inflammation is fundamentally a protective cellular response aimed at removing injurious stimuli and initiating the healing process. However, when prolonged, it can override the bounds of physiological control and becomes destructive. Inflammation is a key element in the pathobiology of chronic pain, neurodegenerative diseases, stroke, spinal cord injury, and neuropsychiatric disorders. Glia, key players in such nervous system disorders, are not only capable of expressing a pro-inflammatory phenotype but respond also to inflammatory signals released from cells of immune origin such as mast cells. Chronic inflammatory processes may be counteracted by a program of resolution that includes the production of lipid mediators endowed with the capacity to switch off inflammation. These naturally occurring lipid signaling molecules include the N-acylethanolamines, N-arachidonoylethanolamine (an endocannabinoid), and its congener N-palmitoylethanolamine (palmitoylethanolamide or PEA). PEA may play a role in maintaining cellular homeostasis when faced with external stressors provoking, for example, inflammation. PEA is efficacious in mast cell-mediated models of neurogenic inflammation and neuropathic pain and is neuroprotective in models of stroke, spinal cord injury, traumatic brain injury, and Parkinson disease. PEA in micronized/ultramicronized form shows superior oral efficacy in inflammatory pain models when compared to naïve PEA. Intriguingly, while PEA has no antioxidant effects per se, its co-ultramicronization with the flavonoid luteolin is more efficacious than either molecule alone. Inhibiting or modulating the enzymatic breakdown of PEA represents a complementary therapeutic approach to treat neuroinflammation. This review is intended to discuss the role of mast cells and glia in neuroinflammation and strategies to modulate their activation based on leveraging natural mechanisms with the capacity for self-defense against inflammation.

Background and Purpose: Palmitoylethanolamide (PEA) acts via several targets, including cannabinoid CB1 and CB2 receptors, transient receptor potential vanilloid type-1 (TRPV1) ion channels, peroxisome proliferator-activated receptor alpha (PPAR α) and orphan G protein-coupled receptor 55 (GRR55), all involved in the control of intestinal inflammation. Here, we investigated the effect of PEA in a murine model of colitis., Experimental Approach: Colitis was induced in mice by intracolonic administration of dinitrobenzenesulfonic acid (DNBS). Inflammation was assessed by evaluating inflammatory markers/parameters and by histology; intestinal permeability by a fluorescent method; colonic cell proliferation by immunohistochemistry; PEA and endocannabinoid levels by liquid chromatography mass spectrometry; receptor and enzyme mRNA expression by quantitative RT-PCR., Key Results: DNBS administration caused inflammatory damage, increased colonic levels of PEA and endocannabinoids, down-regulation of mRNA for TRPV1 and GPR55 but no changes in mRNA for CB1 , CB2 and PPARα. Exogenous PEA (i.p. and/or p.o., 1 mg·kg(-1) ) attenuated inflammation and intestinal permeability, stimulated colonic cell proliferation, and increased colonic TRPV1 and CB1 receptor expression. The anti-inflammatory effect of PEA was attenuated or abolished by CB2 receptor, GPR55 or PPARα antagonists and further increased by the TRPV1 antagonist capsazepine., Conclusions and Implications: PEA improves murine experimental colitis, the effect being mediated by CB2 receptors, GPR55 and PPARα, and modulated by TRPV1 channels., (© 2014 The British Pharmacological Society.)

In order to quantify the relative bioavailability of glycidol from glycidyl fatty acid esters in vivo, glycidyl palmitoyl ester and glycidol were orally applied to rats in equimolar doses. The time courses of the amounts of glycidol binding to hemoglobin as well as the excretion of 2,3-dihydroxypropyl mercapturic acids were determined. The results indicate that glycidol is released from the glycidyl ester by hydrolysis and rapidly distributed in the organism. In relation to glycidol, there was only a small timely delay in the binding to hemoglobin for the glycidol moiety released from the ester which may be certainly attributed to enzymatic hydrolysis. In both cases, however, an analogous plateau was observed representing similar amounts of hemoglobin binding. With regard to the urinary excretion of mercapturic acids, also similar amounts of dihydroxypropyl mercapturic acids could be detected. In an ADME test using a virtual double tag (³H, ¹⁴C) of glycidyl palmitoyl ester, a diverging isotope distribution was detected. The kinetics of the ¹⁴C-activity reflected the kinetics of free glycidol released after hydrolysis of the palmitoyl ester. In view of this experimental data obtained in rats, it is at present justified for the purpose of risk assessment to assume complete hydrolysis of the glycidyl ester in the gastrointestinal tract. Therefore, assessment of human exposure to glycidyl fatty acid ester should be regarded as an exposure to the same molar quantity of glycidol.

N-Palmitoylethanolamine (NAE 16:0) is an endogenous lipid signaling molecule that has limited water solubility, and its action is short-lived due to its rapid metabolism. This poses a problem for use in vivo as oral administration requires a high concentration for significant levels to reach target tissues, and injection of the compound in a dimethyl sulfoxide- or ethanol-based vehicle is usually not desirable during long-term treatment. A depot injection of NAE 16:0 was successfully emulsified in sterile corn oil (10 mg/kg) and administered in young DBA/2 mice in order to elevate baseline levels of NAE 16:0 in target tissues. NAE 16:0 levels were increased in various tissues, particularly in the retina, 24 and 48 hours following injections. Increases ranged between 22% and 215% (above basal levels) in blood serum, heart, brain, and retina and induced an entourage effect by increasing levels of other 18 carbon N-Acylethanolamines (NAEs), which ranged between 31% and 117% above baseline. These results indicate that NAE 16:0 can be used as a depot preparation, avoiding the use of inadequate vehicles, and can provide the basis for designing tissue-specific dosing regimens for therapies involving NAEs and related compounds.

Introduction: Hepatocellular carcinoma is the most common form of primary hepatic carcinoma. A new N(2)S(2) tetradentate ligand, N-[2-(triphenylmethyl)thioethyl]-3-aza-19-ethyloxycarbonyl-3-[2-(triphenylmethyl)thioethyl]octadecanoate (H(3)MN-16ET), was introduced and labeled with (188)Re to create (188)Re-MN-16ET in the Lipiodol phase. The potential of (188)Re-MN-16ET/Lipiodol for hepatoma therapy was evaluated in a hepatocellular carcinoma animal model of Sprague-Dawley rats implanted with the N1S1 cell line., Methods: Synthesis of H(3)MN-16ET was described, and characterization was identified by infrared, nuclear magnetic resonance and mass spectra. We compared the effects of transchelating agents (glucoheptonate or tartaric acid) and a reducing agent (stannous chloride) on the complexing of (188)Re-perrhenate and H(3)MN-16ET. Twenty-four rats implanted with hepatoma were injected with 3.7 MBq/0.1 ml of (188)Re-MN-16ET/Lipiodol or (188)Re-MN-16ET via transcatheter arterial embolization. Biodistribution experiments and single-photon emission computed tomography imaging were performed to investigate tumor accumulation., Results: H(3)MN-16ET was proved to easily conjugate with the Re isotope and showed good solubility in Lipiodol. The radiochemical purity of (188)Re-MN-16ET/Lipiodol with 10 mg tartaric acid and stannous chloride was shown to be more than 90%. The major distribution sites of (188)Re-MN-16ET in Sprague-Dawley rats were hepatoma and the liver. However, the radioactivity at the tumor site postadministered with (188)Re-MN-16ET was quickly decreased from 9.15±0.23 (at 1 h) to 2.71%±0.18% of injected dose/g (at 48 h). The biodistribution and micro-single-photon emission computed tomography/computed tomography image data showed that (188)Re-MN-16ET/Lipiodol was selectively retained at the tumor site, with 11.55±1.44, 13.16±1.46 and 10.67%±0.95% of injected dose/g at 1, 24 and 48 h postinjection, respectively. The radioactivity in normal liver tissue was high but significantly lower than that of the tumors., Conclusion: H(3)MN-16ET is a suitable tetradentate ligand for (188)Re labeling. From the animal data, we suggest that (188)Re-MN-16ET/Lipiodol has the potential to be a therapeutic radiopharmaceutical for hepatoma treatment., (Copyright © 2011. Published by Elsevier Inc.)

Development of a (99m)Tc-fatty acid analogue is of interest, as (99m)Tc is logistically advantageous over the cyclotron-produced (11)C and (123)I. Synthesis of a 16 carbon fatty acid derivative and its radiolabeling with the novel [(99m)TcN(PNP)](2+) core is described here. Hexadecanedioic acid was conjugated to cysteine in an overall yield of 55%. This ligand could be labeled with (99m)Tc via the [(99m)TcN(PNP)](2+) core, in 80% yield, as a mixture of two isomers (syn and anti). The major isomer isolated by HPLC was used for bioevaluation studies in swiss mice and compared with radioiodinated iodophenyl pentadecanoic acid (IPPA), an established agent for myocardial metabolic imaging. (99m)Tc-labeled complex cleared faster from the non-target organs, namely, liver, lungs, and blood compared to that of [(125)I]-IPPA. However, the complex exhibited lower uptake and faster washout from the myocardium as compared to [(125)I]-IPPA.

Marinobactins A-E are a suite of amphiphilic siderophores which have a common peptidic head group that coordinates Fe(III), and a fatty acid which varies in length and saturation. As a result of the amphiphilic properties of these siderophores it is difficult to study siderophore-mediated uptake of iron, because the amphiphilic siderophores partition indiscriminately in microbial and other membranes. An alternative method to distinguish amphiphilic siderophore partitioning versus siderophore-mediated active uptake for Fe(III)-marinobactin E has been developed. In addition, a new member of the marinobactin family of siderophores is also reported, marinobactin F, which has a C(18) fatty acid with one double bond and which is substantially more hydrophobic that marinobactins A-E.

Carnitine-dependent fatty acid import into mitochondria and beta-oxidation seem to be impaired in tumor cells. In the present study we show that a supply of palmitoylcarnitine together with L-carnitine potently induces apoptosis in HT-29 human colon cancer cells as a consequence of accelerated fatty acid oxidation. Caspase-3-like activities, measured by the cleavage rate of a fluorogenic tetrapeptide substrate and nuclear fragmentation determined after DNA labeling in fixed cells by fluorescence microscopy, served as indicators of apoptosis. Neither L-carnitine nor palmitoylcarnitine alone were able to increase caspase-3-like activities and DNA fragmentation, but when provided together, apoptosis occurred. That exogenous carnitine was indeed able to enhance fatty acid uptake into mitochondria was demonstrated by an increased influx of a fluorescent palmitic acid analog. Enhanced fatty acid availability in mitochondria led to an increased generation of O*2-, as detected by a O*2- -sensitive fluorogenic dye, indicating oxidation of delivered substrates. Benzoquinone, an O*2- scavenger, blocked O*2- generation and prevented apoptosis as initiated by the combination of palmitoylcarnitine and carnitine. The lack of effect of the ceramide synthesis inhibitor fumonisin on palmitoylcarnitine/carnitine-induced apoptosis further supports the notion that apoptotic cell death is specifically due to fatty acid oxidation. In contrast to HT-29 cells, nontransformed human colonocytes did not respond to exogenous palmitoylcarnitine/carnitine and no apoptosis was observed. In conclusion, our studies provide evidence that a limited mitochondrial fatty acid import in human colon cancer cells prevents high rates of mitochondrial O*2- production and protects colon cancer cells from apoptosis that can be overcome by an exogenous carnitine supply.

Regional distribution of exogenous N-palmitoylethanolamine in the rat brain was investigated in the study. Possible protective and adaptive effect of N-stearoylethanolamine under 2 Gy whole-body X-irradiation and changes of brain lipid composition were also studied. It was found that after per os administration to rats N-([9,10-3H]-palmitoyl)-ethanolamine was primarily accumulated in hypothalamus, pituitary and adrenal glands and the label amount in brain was 0.95% of the oral dose. Quantities of palmitic acid in total brain phospholipids and plasmalogen form of phosphatidylcholine were increased; free cholesterol and diacyl form of phosphatidylcholine were decreased in 2 weeks after irradiation. 11-OH-corticosteroid level in the blood of exposed rats was decreased in comparison with control animals. N-stearoylethanolamine pre-treatment prevented from increasing the plasmalogen form of phosphatidylcholine and decreasing its diacyl form and restored 11-OH-corticosteroid level in the blood of irradiated rats. Recovering of brain free cholesterol level was observed when N-stearoylethanolamine was post-treated. So, the accumulation of N-([9,10-3H]-palmitoyl)ethanolamine in brain indicates its penetration through blood-brain barrier and suggests the possible role of saturated N-acylethanolamines in brain functioning, particularly, in stress response regulation of the organism by hypothalamus-pituitary-adrenal system. N-stearoylethanolamine treatment of irradiated rats causes protective effect concerning the of irradiation induced changes in the brain lipid composition and in 11-OH-corticosteroid level and modifies phospholipid fatty acid composition.

Familial hypobetalipoproteinemia (FHBL) subjects may develop fatty liver. Liver fat was assessed in 21 FHBL with six different apolipoprotein B (apoB) truncations (apoB-4 to apoB-89) and 14 controls by magnetic resonance spectroscopy (MRS). Liver fat percentages were 16.7 +/- 11.5 and 3.3 +/- 2.9 (mean +/- SD) (P = 0.001). Liver fat percentage was positively correlated with body mass index, waist circumference, and areas under the insulin curves of 2 h glucose tolerance tests, suggesting that obesity may affect the severity of liver fat accumulation in both groups. Despite 5-fold differences in liver fat percentage, mean values for obesity and insulin indexes were similar. Thus, for similar degrees of obesity, FHBL subjects have more hepatic fat. VLDL-triglyceride (TG)-fatty acids arise from plasma and nonplasma sources (liver and splanchnic tissues). To assess the relative contributions of each, [2H2]palmitate was infused over 12 h in 13 FHBL subjects and 11 controls. Isotopic enrichment of plasma free palmitate and VLDL-TG-palmitate was determined by mass spectrometry. Non-plasma sources contributed 51 +/- 15% in FHBL and 37 +/- 13% in controls (P = 0.02). Correlations of liver fat percentage and percent VLDL-TG-palmitate from liver were r = 0.89 (P = 0.0001) for FHBL subjects and r = 0.69 (P = 0.01) for controls. Thus, apoB truncation-producing mutations result in fatty liver and in altered assembly of VLDL-TG.

The objective of this study was to examine the ocular absorption behavior of an amphiphilic prodrug after instillation onto the cornea of rabbits. A micellar solution of O-palmitoyl tilisolol (PalTL), an amphiphilic prodrug, was prepared. After instillation of tilisolol (TL) and PalTL, the drug concentrations in the tear fluid, cornea, aqueous humor, iris-ciliary body, vitreous body, and blood were measured. In addition, in situ ocular absorption behavior was also evaluated. After instillation of TL, the concentration of TL in the tear fluid quickly decreased. After instillation of PalTL, prolonged retention and high concentrations of PalTL in tear fluid and the cornea were observed. In addition, more prolonged retention of the TL concentration after instillation of PalTL than after instillation of TL was observed in the cornea, aqueous humor, and iris-ciliary body. In situ experiments demonstrated that PalTL was mainly absorbed by the corneal route and the improvement effects of PalTL under in vivo conditions was due to an enhanced transit time of PalTL in ocular tissues. PalTL, an amphiphilic prodrug, exhibited increased retention in the precorneal area compared with the parent drug, TL, resulted in improved ocular absorption of the parent drug., (Copyright 2001 Wiley-Liss, Inc. and the American Pharmaceutical Association)

Background: Referred hyperalgesia to a somatopically appropriate superficial site is a cardinal symptom of visceral inflammatory pain and has been demonstrated after turpentine-induced urinary bladder inflammation in the rat. The authors examined the effect of the endocannabinoids anandamide and palmitoylethanolamide on the referred hyperalgesia associated with this model., Methods: After measurement of baseline limb withdrawal latencies to a noxious heat stimulus, the bladders of 50 female Wistar rats were inflamed by intravesical administration of 0.5 ml 50% turpentine. Ten or 25 mg/kg of anandamide or palmitoylethanolamide or vehicle were administered immediately before introduction of turpentine. Antagonists to both the cannabinoid CB1 and CB2 receptors were coadministered with the higher dose of endocannabinoids. Latencies were recorded 2, 4, 6, 8, and 24 h after removal of turpentine. The difference between forelimb and hind limb withdrawal latencies was plotted against time, and areas under these curves were compared., Results: Inflammation of the urinary bladder was associated with a relative thermal hyperalgesia referred to the hind limb. Anandamide and palmitoylethanolamide attenuated this referred hyperalgesia at doses of 10 and 25 mg/kg. The CB1 receptor antagonist SR141716A reduced the antihyperalgesic effect of anandamide, but the CB2 antagonist SR144528 did not. Coadministration of SR141716A with palmitoylethanolamide did not affect the antihyperalgesic effect but was reduced by SR144528., Conclusions: Anandamide (via CB1 receptors) and palmitoylethanolamide (putatively via CB2 receptors) attenuated a referred hyperalgesia in a dose-dependent fashion. CB1 and CB2 receptors are strategically situated to influence the nerve growth factor-driven referred hyperalgesia associated with inflammation of the urinary bladder. These data implicate cannabinoids as a novel treatment for vesical pain.

N-([1-14C]-palmitoyl)-ethanolamine distribution was studied in the rat tissues. The following sequence of the label inclusion into tissues by the way of decreasing the radio activity: adrenal >> diaphragm > spleen > kidney > testis > lung > liver > heart > brain > plasma > erythrocytes was obtained.

beta-Methyl-substituted free fatty acids (FFAs) have been developed for myocardial single-photon emission tomography (SPET) imaging, but little is known about their kinetics in ischaemic conditions. The aim of this study was to determine the changes in the myocardial kinetics of a beta-methyl-branched FFA, [123I]16-iodo-3-methyl-hexadecanoic acid (MIHA), under ischaemic conditions. The kinetics of MIHA were analysed: (a) using a blood-perfused isolated heart model subjected to moderate ischaemia (50% flow reduction) and (b) in patients who had an exercise thallium-201 SPET defect corresponding to either necrotic (n = 13) or chronically ischaemic and viable (n = 15) myocardium, and who underwent two consecutive SPET studies after MIHA injection. In animals, the myocardial early retention fraction of MIHA, but not its clearance rate, was dependent on coronary flow, the early retention fraction being higher in ischaemic than in normoxic conditions (0.24 +/- 0.10 vs 0.14 +/- 0.04, P = 0.004). In the patient SPET studies, the uptake of MIHA calculated in ischaemic and viable areas (G1: 74% +/- 9% of maximal left ventricular value) was different from that calculated in necrotic (G2: 59% +/- 7%, P < 0.001) or normal (G3: 88 +/- 6%, P < 0.001) areas. By contrast, MIHA-clearance calculated between the two consecutive SPET studies was not different in G1, G2 and G3. Unlike in the case of other FFAs, the myocardial clearance of MIHA is not decreased by ischaemia. However, the early retention of MIHA is increased in the case of a moderate reduction in coronary flow, a property which might help in the detection of viability in chronically ischaemic myocardium.

A 2-year-old female was well until 12 months of age when she was found to be anemic and had dilated cardiomyopathy. Total plasma carnitine was 6 microM and acylcarnitine analysis while receiving carnitine supplement revealed an increase in the four-carbon species. Urine organic acids were normal. In vitro analysis of the mitochondrial pathways for beta oxidation, and leucine, valine, and isoleucine metabolism was performed in fibroblasts using stable isotope-labeled precursors to these pathways followed by acylcarnitine analysis by tandem mass spectrometry. 16-2H3-palmitate was metabolized normally down to the level of butyryl-CoA thus excluding SCAD deficiency. 13C6-leucine and 13C6-isoleucine were also metabolized normally. 13C5-valine incubation revealed a significant increase in 13C4-isobutyrylcarnitine without any incorporation into propionylcarnitine as is observed normally. These same precursors were also evaluated in fibroblasts with proven ETF-QO deficiency in which acyl-CoA dehydrogenase deficiencies in each of these pathways was clearly identified. These results indicate that in the human, there is an isobutyryl-CoA dehydrogenase which exists as a separate enzyme serving only the valine pathway in addition to the 2-methyl branched-chain dehydrogenase which serves both the valine and the isoleucine pathways in both rat and human., (Copyright 1998 Academic Press.)

Background: Intestinal transplantation is now used for patients with severe malabsorption, however, little data exists quantifying the ability of the graft to absorb fat. This study tested the hypothesis that intestinal transplantation would not affect the lymphatic or venous uptake of fatty acids., Methods: A syngeneic rat model of intestinal transplantation (SIT) with caval drainage of the graft was used. Control animals underwent intestinal division and reanastomosis (n=15 in each group). The animals were followed for 6 weeks, and fat absorption in vivo was quantified. The animals were anesthetized, sampling catheters were placed in the jugular and superior mesenteric veins and in the mesenteric lymphatic duct, and a feeding tube was passed into the duodenum. Animals were allowed to recover, and a steady-state duodenal infusion of lauric (C12:0) and palmitic (C16:0) fatty acid emulsion was begun. A radiolabeled pulse of lauric (C12:0) and palmitic (C16:0) fatty acid was then given, and the subsequent appearance in the lymphatic and venous systems was quantified., Results: In vivo absorption of dietary fat was preserved, but after transplantation the mesenteric lymphatic flow and cumulative lymphatic appearance of both labels was significantly reduced (flow reduced from 4.8+/-1.1 in controls to 1.0+/-0.29 ml/hr in transplant animals, whereas lauric acid absorption was 33+/-11.4% in controls vs. 7.5+/-2.5% in transplant animals). There was a modest increase in the jugular venous appearance of the fatty acids (2.0+/-1.1% in transplant animals vs. 0.75+/-0.55% in controls for lauric acid; P<0.05 for all comparisons). Absorption of lauric and palmitic acids was very similar, and there was no preferential absorption detected in the portal venous system. Dye studies demonstrated lymphatic recannulization around the vascular anastomosis, into the retroperitoneum., Conclusions: These results suggest that in this model of SIT, fat absorption via the mesenteric duct is reduced, but that compensatory collaterals form into the retroperitoneal lymphatics. There was no evidence of any significant increase in portal venous uptake of fatty acids after SIT, nor of preferential absorption of medium-chain fatty acids. These results may have implications for patients after SIT.

The effects of 1,3-dioleoyl-2-palmitoyl glycerol (OPO) on lymph chylomicron transport, composition and size in rats were investigated in comparison with 1,2-dioleoyl-3-palmitoyl glycerol (OOP). The OPO and OOP were prepared by enzymatic transesterification reactions. The concentrations of OPO and OOP in the preparations were 65.7 g/100 g, and the composition of fatty acids was similar for each. The OPO preparation contained triacylglycerols with 76.6% of the palmitic acid in the sn-2 position, whereas 100% of the oleic acid was esterified to the sn-2 position in the OOP preparation. Rats were infused with lipid emulsion containing 150 g/L of OPO or OOP via a stomach catheter. Lymph was collected through the mesenteric lymphatic trunk at 1-h intervals for 12 h. Collected lymph chylomicrons were analyzed for triacylglycerol, fatty acids, apolipoprotein A-I and particle size. The maximum transport rates of triacylglycerols in the OPO group were higher than those in the OOP group. The overall absorption of triacylglycerols, palmitic acid and oleic acid in the OPO group was also higher than that in the OOP group. In the chylomicrons, 60-70% of the fatty acids at the sn-2 position of the infused triacylglycerol was transported at the original position. The transport rates of dioleoyl-palmitoyl glycerol in the OPO group were higher than those in the OOP group. The transport rates of apolipoprotein A-I did not differ between groups, whereas the mean diameter of the chylomicrons in the OPO group was larger than that in the OOP group. These results indicate that OPO is absorbed and transported more effectively than OOP.

Anandamide (arachidonoylethanolamide, AnNH) and palmitoylethanolamide (PEA) have been proposed as the physiological ligands, respectively, of central and peripheral cannabinoid receptors. Both of these receptors are expressed in immune cells, including macrophages and mast cells/basophils, where immunomodulatory and/or anti-inflammatory actions of AnNH and PEA have been recently reported. We now provide biochemical grounds to these actions by showing that the biosynthesis, uptake, and degradation of AnNH and PEA occur in leukocytes. On stimulation with ionomycin, J774 macrophages and RBL-2H3 basophils produced AnNH and PEA, probably through the hydrolysis of the corresponding N-acylphosphatidylethanolamines, also found among endogenous phospholipids. Immunological challenge of RBL-2H3 cells also caused AnNH and PEA release. The chemical structure and the amounts of AnNH and PEA produced upon ionomycin stimulation were determined by means of double radiolabeling experiments and isotope dilution gas chromatography/electron impact mass spectrometry. Both cell lines rapidly sequestered the two amides from the culture medium through temperature-dependent, saturable and chemically inactivable mechanisms. Once uptaken by basophils, AnNH and PEA compete for the same inactivating enzyme which catalyzes their hydrolysis to ethanolamine. This enzyme was found in both microsomal and 10,000 x g fractions of RBL cell homogenates, and exhibited similar inhibition and temperature/pH dependence profiles but a significantly higher affinity for PEA with respect to neuronal "anandamide amidohydrolase." The finding of biosynthetic and inactivating mechanisms for AnNH and PEA in macrophages and basophils supports the previously proposed role as local modulators of immune/inflammatory reactions for these two long chain acylethanolamides.

5-Methyl-17-[18F]fluoroheptadecanoic acid (5-MFHA) has been proposed as a new myocardial imaging tracer for positron emission tomography (PET), containing methyl-branching at the odd-numbered position, except the 3-position. To compare the site of methyl-branching of fatty acids on the contribution to myocardial imaging, the biological evaluation of 5-MFHA-using accumulation studies, metabolic studies, and PET studies of the heart muscle-was investigated. In the comparative biodistribution studies for 16-[18F]fluoropalmitic acid (FPA), 3-methyl-17-[18F]fluoroheptadecanoic acid (3-MFHA) and 5-MFHA, the initial myocardial uptake of 5-MFHA (2.64 [%dose/g tissue]) was relatively high between those of FPA and 3-MFHA (3.45 and 1.58, respectively), and the washout from myocardium of 5-MFHA was midway between those of FPA and 3-MFHA. In the lipid analysis studies, 5-MFHA was mainly metabolized to triglycerides in the myocardium, and its metabolic pattern was similar to that of straight-chain fatty acids (FPA). In the PET studies using 5-MFHA in canines, good myocardial images were obtained for up to 30 min after injection.

This study determined the effect of dietary (dipalmitoyl) triacylglycerol (TAG) structure on the absorption and metabolism of fat in the presence or absence of cholesterol. Sprague-Dawley male rats weighing approximately 180 g were fed diets containing 10 g/100 g diet 1,3-dipalmitoyl-2-oleoyl-glycerol-rich oil (POP) or 1(3),2-dipalmitoyl-3(1)-oleoyl-glycerol-rich oil (PPO), either with or without cholesterol supplementation (0.5 g/100 g diet), for 2 wk. There were no differences in the growth of rats fed the various diets. The apparent absorption efficiency of dietary fat especially the apparent absorption efficiency of palmitic acid, was significantly lower in the POP groups than in the groups fed the PPO diets. The excretion of cholesterol in feces of rats fed the POP diet with cholesterol was significantly greater than those fed the PPO diet containing cholesterol. The excretion of total bile acids in the rats fed cholesterol was significantly greater than in those fed the diets without cholesterol. Liver cholesterol was significantly greater in rats fed cholesterol and was not affected by the TAG structure. Plasma cholesterol was significantly lower in the POP dietary groups than in the PPO groups. These results suggest that the absorption and metabolism of fat depend on the positional isomer of the fatty acid.

To determine if the sex differences in clearance of long chain fatty acids by the liver observed in the rat are relevant to humans, the authors isolated hepatocytes from human adult males and females (five per group) and measured the initial (unidirectional) clearance of [3H]-palmitate from buffer containing albumin. The clearance was significantly higher (about twofold) in hepatocytes from females because of a higher permeability of the plasma membrane to the fatty acid. The livers had been perfused with University of Wisconsin (UW) solution and stored for 12-16 h before isolation of the cells. The magnitudes of the clearances in humans were similar to those in the rat when the livers were stored similarly, but lower than in cells isolated from fresh rat liver. When hepatocytes isolated from fresh rat liver were stored in UW solution, clearance of [3H]-palmitate was unaffected. Thus, hepatocytes prepared from intact liver stored for several hours in UW solution do not have as good preservation of function as hepatocytes isolated from fresh liver and then stored in UW solution.

Unlabelled: We determined regional incorporation coefficients (k*) of plasma [1-11C]palmitate into stable brain lipids of anesthetized monkeys with PET., Methods: Carbon-11-palmitate was injected intravenously in untreated animals and in animals pretreated with methyl palmoxirate (MEP), an inhibitor of beta-oxidation of palmitate in the brain and periphery. Plasma radioactivity was followed, and brain radioactivity was determined at various times using PET. A least-squares method was used to fit the data to an operational equation to obtain regional values of k* and of cerebral blood volume (Vb) in individual experiments., Results: MEP significantly decreased the integral of plasma [11C]CO2 following 11C-palmitate infusion. Mean values of k* in monkeys not given MEP were 4.9, 4.2, 4.9, 4.0 and 2.9 x 10(-5) ml/sec.g for the temporal, frontal, parietal and occipital cortices and white matter, respectively. With the exception of k* in white matter, which was increased by MEP, k* in the other brain regions was not significantly changed by MEP. The Vb ranged from 0.035 ml/g to 0.048 ml/g in gray matter regions and equaled 0.022 ml/g in white matter., Conclusion: PET can be used to determine regional incorporation coefficients of 11C-palmitate into the primate brain in vivo. Combined with MEP, 11C-palmitate could be used with PET to examine regional brain phospholipid metabolism in humans in normal and pathological conditions.

Fatty acid ethyl esters (FAEEs), esterification products of ethanol and fatty acids, have been found selectively in the organs damaged by ethanol abuse, and on that basis have been implicated as contributors to ethanol-induced organ damage. To directly assess the cytotoxic potential of FAEEs with intact cells in a physiological system, solubility must be achieved for these highly nonpolar lipids in aqueous medium. After ethanol ingestion, FAEEs can be found within low-density lipoproteins (LDLs). Therefore, to achieve solubility with FAEEs bound to a naturally occurring lipid carrier, we developed a method for FAEE solubilization and delivery to cells in culture. We synthesized radiolabeled FAEEs and incorporated them into human LDL particles that bind to LDL receptors and deliver FAEEs to intact cells. Ethyl palmitate and ethyl oleate were incorporated into LDLs yielding molar ratios of FAEEs to LDLs of 2,153 +/- 249 and 4,208 +/- 403, respectively. LDL reconstituted with FAEE had the same electrophoretic mobility on agarose gel electrophoresis as native LDL, indicating that the reconstituted LDL (rLDL) was not oxidatively modified. Quantitative analysis of the solubilization of FAEEs in aqueous medium was investigated by adding FAEEs to tissue culture medium either directly or reconstituted in LDL at a concentration of 27 microM. The percentage of FAEE quantitated was 40.0 +/- 2.5% and 89.3 +/- 0.6% for FAEEs added directly and in rLDLs, respectively. After sterile filtration of these two media, the percentage of FAEE that remained was 11.8 +/- 1.3% (direct addition) and 74.9 +/- 1.3% (addition within rLDL), further demonstrating that the LDL particle did solubilize the FAEE.(ABSTRACT TRUNCATED AT 250 WORDS)

Bovine serum albumin (albumin) was modified by treatment with nitric oxide (NO) to form S-nitrosoalbumin. Analysis of the reduced sulfhydryl groups showed that more than 99% of the albumin was converted to S-nitrosoalbumin. Using a 1:1 molar ratio of protein:palmitate, the unbound palmitate fraction in the presence of S-nitrosoalbumin was determined to be greater (28%) than in the presence of albumin as determined by heptane: water partitioning. NO degradation products neither affected the palmitate heptane:water partition ratio in the absence of binding protein nor the hepatocyte uptake of [3H]palmitic acid. The equilibrium association constants (Ka) for albumin-palmitate and S-nitrosoalbumin-palmitate complexes were determined using the stepwise equilibrium model. The Ka for the first and second palmitate binding sites were (4.6 +/- 1.2) x 10(8) M-1 and (3.3 +/- 0.5) x 10(7) M-1 and (3.1 +/- 0.9) x 10(8) M-1 and (1.3 +/- 0.8) x 10(8) M-1 for albumin and S-nitrosoalbumin, respectively. Thus, the increased unbound fraction of palmitate in the presence of S-nitrosoalbumin was apparently due to a decreased binding affinity at the first high-affinity binding site. Palmitate uptake by hepatocyte suspensions was 27% higher in the presence of S-nitrosoalbumin as compared with albumin. This increase paralleled the increased unbound palmitate fraction. When the albumin concentration was adjusted to account for the increased unbound fraction, there was no difference in the palmitate uptake rates between albumin and S-nitrosoalbumin. Our findings indicate that under conditions where NO concentrations are high (e.g. cirrhosis) and extensive S-nitrosylation of serum albumin occurs, the decreased ligand binding ability of S-nitrosoalbumin may be an important consideration when modeling drug uptake in pathological states.

Translational diffusion of a fluorescent sterol probe was measured in the plasma membranes of protoplasts isolated from cortical cells of the primary root of maize seedlings. The apparent lateral diffusion coefficient was typically observed to be nearly insensitive to temperature, while the mobile fraction increased with increasing temperature. These fluorescence photobleaching recovery (FPR) measurements were compared with the electron paramagnetic resonance (EPR) spectra of the methyl ester of 13-doxyl palmitic acid in membranes of corn root tissue in situ. The complex spectra observed with this probe were analyzed as weighted sums of simpler spectra of various order parameters and rotational correlation times. The reconstituted spectra calculated from the model show that EPR also detects a mobile (less ordered, fluid) fraction, distinguished by the order parameter S = 0.1 to 0.2, which becomes more abundant as temperature increases and is qualitatively comparable to the mobile fraction determined by the FPR method. The observed results on the mobile fractions and the diffusion rates for translational (FPR) as well as rotational (EPR) motions are interpreted in terms of membrane organization, thus providing information on the population and structural patterns of the coexisting domains with a special emphasis on the response of the membrane to temperature changes.

Milk fatty acids contain 20-30% palmitic acid (16:0), with approximately 70% of the 16:0 esterified to the sn-2 position of the milk triacylglycerol. Formulae containing vegetable and oleo oils contain different amounts of 16:0, but all have unsaturated fatty acids esterified to the triacylglycerol sn-2 position. Intraluminal triacylglycerol hydrolysis by endogenous lipases produces sn-2 monoacylglycerols and free fatty acids, which are absorbed and re-esterified in the enterocyte for secretion to plasma. The extent of absorption and re-esterification of sn-2 monoacylglycerols from milk or formula fats in infants is unknown. This was studied by feeding piglets sow milk or formulae containing similar total saturated fat, 18:1, 18:2(n-6) and 18:3(n-3) with unsaturated fatty acids at the sn-2 position or with rearranged triacylglycerols containing approximately 30% 16:0, with 70% 16:0 in fatty acids at the sn-2 position. Feeding milk or 16:0 on the sn-2 position of formula with rearranged triacylglycerols resulted in higher 16:0 esterified to the plasma triacylglycerol sn-2 position and in cholesteryl esters than feeding formulae with 0.4 to 4.4% 16:0 in the sn-2 position fatty acids. The absorption of 16:0 as monopalmitin from milk and any metabolic importance in human infants has yet to be determined.

The human serum albumin (HSA)-dependent unbound clearance (Clu) of [3H]palmitic acid (PA) by hepatocyte suspensions isolated from immature and mature male and female and pregnant female rats was studied. The Clu values obtained experimentally were compared with the predictions of a noncompartmental diffusion-reaction (Bass-Pond) theory for the cellular uptake of protein-bound ligands. In all groups, as the concentration of HSA (Ca) was increased, there was a striking increase in Clu. These enhancement factors were predicted by the theory. Adult females had higher Clu values at high Ca values than males or immature females. Furthermore, at high Ca values, Clu in pregnant animals was twice as high as in the nonpregnant animals and four times as high as in the aged-matched males. The absolute values of Clu obtained experimentally in both pregnant and nonpregnant females exceeded the maximal predictions of the theory, using reasonable values of all of the parameters. Thus, according to current data on the physicochemical characteristics of the uptake system, the study demonstrates that some specialized process exists to facilitate hepatocellular uptake of fatty acid from albumin, and that it is potentiated by the female sex hormones.

The clearance ratio test has been used to determine whether hepatocytes isolated from female rat livers facilitate the dissociation rate of albumin-palmitate complexes. This test requires knowledge of the clearance rate and unbound palmitate fraction at two protein concentrations. The unbound fractions have been estimated using the heptane-water partition ratio method. These fractions were determined to deviate from the expected linear relationship because of radiolabeled impurities that were still present after purification of the [3H]palmitic acid. With the use of experimentally determined partition ratio values, the palmitate clearance ratios were statistically greater than those predicted by the diffusion-reaction model at albumin concentration combinations above 99 microM/49 microM. Experimental clearance rates also exceeded those predicted by the diffusion-reaction model, but only at albumin concentrations greater than 198 microM. These data support the proposal that hepatocytes facilitate the dissociation of albumin-palmitate complexes at high albumin concentrations.

Milk fatty acids consist of about 20-25% palmitic acid (16:0), with about 70% of 16:0 esterified to the sn-2 position of the milk triacylglycerols. Hydrolysis of dietary triacylglycerols by endogeneous lipases produces sn-2 monoacylglycerols and free fatty acids, which are absorbed, reesterified, and then secreted into plasma. Unesterified 16:0 is not well absorbed and readily forms soaps with calcium in the intestine. The positioning of 16:0 at the sn-2 position of milk triacylglycerols could explain the high coefficient of absorption of milk fat. However, the milk lipase, bile salt-stimulated lipase, has been suggested to complete the hydrolysis of milk fat to free fatty acids and glycerol. These studies determined whether 16:0 is absorbed from human milk as sn-2 monopalmitin by comparison of the plasma triacylglycerol total and sn-2 position fatty acid composition between breast-fed and formula-fed term gestation infants. The human milk and formula had 21.0 and 22.3% of 16:0, respectively, with 54.2 and 4.8% 16:0 in the fatty acids esterified to the 2 position. The plasma triacylglycerol total fatty acids had 26.0 +/- 0.6 and 26.2 +/- 0.6% of 16:0, and the sn-2 position fatty acids had 23.3 +/- 3.3 and 7.4 +/- 0.7% of 16:0 in the three-month-old exclusively breast-fed (n = 17) and formula-fed (n = 18) infants, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

In order to diagnose patients with medium-chain acyl-CoA dehydrogenase deficiency with a noninvasive diagnostic technique such as positron emission tomography, we have developed a synthesis of [omega-11C]palmitic acid. The radiochemical synthesis was achieved by coupling an alkylfuran Grignard reagent (7) with [11C]methyl iodide, followed by rapid oxidative cleavage of the furan ring to the carboxylate using ruthenium tetraoxide. Tissue biodistribution studies in rats comparing [omega-11C]palmitic acid and [1-11C]palmitic acid show that the %ID/g and %ID/organ in the heart tissue after administration of [omega-11C]palmitic acid is approximately 50% greater than after administration of [1-11C]palmitic acid, due to the diminished metabolism of the [omega-11C]palmitic acid. These studies show as well, low uptake in nontarget tissues (blood, lung, kidney, and muscle). PET images of a dog heart obtained after administration of [omega-11C]-and [1-11C]palmitic acid show virtually identical uptake and distribution in the myocardium. The differing cardiac washout of labeled palmitates measured by dynamic PET studies may allow diagnosis of disorders in cardiac fatty acid metabolism.

Several studies have reported that parenteral lipid emulsions containing medium-chain triglycerides (MCT) and structured lipids (SL) are better utilized than those containing long-chain triglycerides (LCT). The objective of this study was to test the hypothesis that parenteral LCT require more extensive modification via hydrolysis and reesterification (triglyceride-free fatty acid [TG-FFA] recycling) for effective utilization, whereas MCT and SL do not. As an index of TG-FFA cycling activity, we measured glycerol and palmitate kinetics in rats (204 to 243 g) fed parenterally one of three isocaloric (250 kcal/kg/d) isonitrogenous (1.5 g N/kg/d) diets with half of the nonprotein energy from glucose and the rest from either LCT, LCT plus MCT, or SL for 5 days. Two experiments were performed. On day 5, rats were given a 7-to 8-hour infusion of either 5H2 Glycerol and 1-14C Palmitate bound to albumin to measure palmitate and glycerol kinetics (experiment 1), or U-13C glucose to determine the proportion of endogenous glycerol production derived from glucose (experiment 2). Data are presented as means +/- SEM. Endogenous glycerol production was significantly higher with LCT (11.33 +/- 2.89 mmol/kg/h) than with SL (2.91 +/- 0.62 mmol/kg/h). The value for the physical mixture of LCT plus MCT (5.46 +/- 1.29 mmol/kg/h) fell midway between that for LCT and SL (P = NS). There were no significant differences in palmitate kinetics or oxidation. The increased glycerol production is due to the mobilization of endogenous triglyceride and is consistent with a higher rate of TG-FFA cycling being involved in the metabolism of LCT than of SL.(ABSTRACT TRUNCATED AT 250 WORDS)

This paper investigates the incorporation of intravenously (i.v.) administered radiolabelled fatty acids--[9,10(3)-H]palmitate (3H-PA), [1-14C]arachidonate (14C-AA) and [1-14C]docosahexaenoate (14C-DHA)--into intracerebrally implanted tumours in awake Fischer-344 rats. A suspension of Walker 256 carcinosarcoma tumour cells (1 x 10(6) cells) was implanted into the right cerebral hemisphere of 8- to 9-week-old rats. Seven days after implantation, the awake rat was infused i.v. for 5 min with 3H-PA (6.4 mCi/kg), 14C-AA (170 microCi/kg) or 14C-DHA (100 microCi/kg). Twenty minutes after the start of infusion, the rat was killed and coronal brain sections were obtained for quantitative autoradiography and histology. Each fatty acid showed well-demarcated incorporation into tumour tissue. Areas of necrosis or haemorrhage showed no or small levels of incorporation. The ratios of incorporation into the tumour to incorporation into contralateral brain regions were 2.8-5.5 for 3H-PA, 2.1-3.3 for 14C-AA and 1.5-2.2 for 14C-DHA. The mean ratios differed significantly between the fatty acids (P < 0.01). 3H-PA was not incorporated into necrotic tumours despite the presence of an open blood-tumour barrier, indicated by extravasated horseradish peroxidase. The incorporation rate constant of 3H-PA was similar for small intracerebral and large extracerebral tumours. The results show that 3H-PA, 14C-AA and 14C-DHA are incorporated more readily into tumour tissue than into brain, and that the increase is primarily due to increased utilization of fatty acids by tumour cells and not due to a high blood-tumour permeability. The relative increases in rates of incorporation for the different fatty acids may be related to lipid composition of the tumour and to the requirement of and specific role of these fatty acids in tumour cell growth and division.

Radioiodine labelled 17-iodo-heptadecanoic acid (IHA) is used for non-invasive study of myocardial metabolism in coronary heart disease and cardiomyopathy. Yet in the interpretation of in vivo myocardial tracer kinetics, it is controversial whether the intracellular degradation of IHA or the removal of iodide across cellular membranes is the rate-limiting step in iodide release from the myocardium. In five patients undergoing coronary sinus catheterization, a mixture of about 40 kBq of [123I] NaI was injected into the left coronary artery. During the following 15-min period, frequent blood samples were taken from the aorta and the coronary sinus. In the aqueous phase of the venous blood, 14CO2 and inorganic 131I appeared nearly in parallel, with a peak time of 4-5 min. Moreover, as shown by the AV difference, there was no significant back diffusion of IHA and no significant non-specific deiodination detectable over the period of observation. There was myocardial retention of inorganic iodide (123I) injected into the left coronary artery. The data strongly support the premise that lipid turnover through beta-oxidation is the rate-limiting step in the externally measured release of iodide after IHA injection, provided that recirculating inorganic radioactive iodide is corrected for. In addition, 15 volunteers were studied using [11C]palmitic acid and [123I]IHA using PET and dynamic planar camera scintigraphy with iodide correction. There was no significant difference between the mean values of the elimination half-times, and also no significant correlation between half-times of both fatty acids for single individuals.

These studies were undertaken to measure the kinetic constants that characterize the disappearance of a representative free fatty acid (FFA) from the plasma of fetal and newborn sheep. A bolus of albumin-complexed [14C]palmitic acid was infused intravenously, and during the next 8 min, 24 arterial samples were collected to characterize the disappearance curve. Palmitic acid disappearance from plasma was well described by a double-exponential model. When birth was simulated in utero, kinetic values were not changed by cooling. However, after intrauterine ventilation with O2, the volume of distribution of the FFA increased 29%, its plasma clearance rate decreased 26%, and its apparent half-life in the plasma lengthened from 0.8 to 1.2 min (all P < 0.01, n = 8). After umbilical cord occlusion, plasma clearance rate decreased a further 19% and half-life lengthened to 1.4 min. About 60% of the increase in FFA concentration during simulated birth is explained by increased release from adipose stores, and 40% is explained by decreased clearance. Further experiments tested the influence of FFA concentrations themselves. After infusion of unlabeled FFA, clearance of the tracer decreased 23% (P < 0.05, n = 5), a result consistent with a saturable membrane transporter of FFAs.

The association of verapamil with halothane causes ischaemic-like myocardial dysfunction. Using an isolated rat heart model perfused with a radiolabelled fatty acid (123I-labelled iodohexadecenoic acid) as a sensitive marker of ischaemia this study investigated whether or not this dysfunction is of ischaemic origin. Hearts were perfused with a control solution or with solutions containing either 1% of halothane or 150 ng ml-1 of verapamil or the association of 0.75% halothane + 120 ng ml-1 verapamil. The ischaemic group was perfused at a reduced perfusion rate (-50%). Intracellular fate of IHA was assessed, and its esterification ratio computed. Ischaemia and the drugs induced a similar depression of haemodynamics. The esterification ratio in the ischaemic group was significantly higher (0.723 +/- 0.04) than in controls (0.0526 +/- 0.03) and than in the treated groups: halothane (0.533 +/- 0.06), verapamil (0.411 +/- 0.027) or the association halothane+verapamil (0.408 +/- 0.05), suggesting a non-ischaemic origin for the dysfunction caused by halothane-verapamil.

We have previously demonstrated that the arachidonic acid (20:4) incorporation into brain lipids differs according to the age of the animals used and the experimental conditions adopted. These differences led to a further investigation of arachidonic acid uptake in both aged and adult rat brains, its transformation into CoA derivatives, its incorporation into diacyl-glycerols and polar lipids, and finally its oxidation to CO2. These metabolic parameters were then compared with those obtained after using the saturated fatty acid palmitate (16:0). In both cases slices or mitochondria from different brain areas of 24-month-old and 4-month-old rats were examined. The results obtained indicate that the uptake of the fatty acids into cells is not modified by age. However, the successive metabolic transformations of the acids are altered to a considerable extent. In particular, in 24-month-old animals (compared with 4-month-old rats) there is a significant decrease of 20:4 in its incorporation into lipids as well as its oxidation to CO2, while arachidonoyl-CoA content increases by about 50%. This increased amount of CoA derivative, which has a potent detergent effect, may interfere with membrane structure and affect membrane physiological functions. Furthermore, because the free arachidonate pool is maintained in a dynamic equilibrium with its esterified forms, the final result may be a perturbation of this equilibrium.

Albumin-dependent uptake of unbound [3H]palmitic acid by hepatocytes isolated from female rat livers was studied and the experimental results compared with the predictions of a noncompartmental diffusion-reaction theory for the cellular uptake of protein-bound ligands. The outright theoretical predictions involve values for the parameters of the system, some newly measured (hepatocyte radii and the rate constant for the dissociation of palmitate-albumin complex) and some taken from the literature (diffusion coefficients and the equilibrium association constant for the palmitate-albumin complex). The measured unbound clearance of [3H]palmitic acid, defined as the initial uptake velocity divided by the unbound [3H]palmitic acid concentration in the medium, was enhanced 6.6-fold as the concentration of human serum albumin was increased from approximately 5 to 480 microM. This enhancement factor was predicted by the theory, according to which the enhancement reflects codiffusion of bound ligand across the unstirred layer adjacent to the cell membrane and, therefore, an increased delivery of unbound ligand to the cell surface. In contrast, the absolute magnitude of the unbound clearance was consistent with the theory only for the lowest published value for the equilibrium association constant, 15 microM-1. For higher published values (62 and 94 microM-1), the magnitude of the unbound clearance observed experimentally was severalfold higher than that predicted by the theory. If in fact the association constant exceeds 30 microM-1, the data would imply that an albumin-dependent facilitation mechanism exists which enhances the availability of palmitate to the cell over and above the enhancement predicted by the diffusion-reaction theory.

The contribution of free fatty acid oxidation to the elevation in energy expenditure after trauma has not been well characterized. Six control subjects and six traumatized patients were fasted for 48 hours and given a primed continuous infusion of (1-14C)palmitate to measure plasma palmitate and total free fatty acid kinetics. Traumatized patients had greater urinary nitrogen losses (20.8 vs. 9.3 g N per day) and a significantly greater ratio of measured to predicted resting energy expenditure (+36% vs. -6%) compared with controls. Individual and total plasma free fatty acid concentrations were similar for the two groups. The turnover and oxidation of plasma palmitate and total free fatty acids were not changed by multiple trauma. These results demonstrated that plasma free fatty acids and palmitate do not contribute to increased energy expenditure following trauma.

Labelled fatty acids have been proposed to explore cardiac metabolism. For the analysis of the external detection curve obtained with 16-iodo 9-hexadecenoic acid (IHA), we developed a mathematical 4-compartment model with compartments 0, 1, 2 and 3 representing vascular IHA, intracellular IHA, esterified forms and iodide, respectively. This model, used here for isolated rat hearts perfused in a recirculating system, is validated by an intracellular analysis, then tested in various metabolic conditions. Thus, the mathematical analysis of the external detection curve gives us numerical data on IHA metabolism, especially the distribution between degradation and storage. Our results confirm the suitability of IHA for assessing myocardial metabolism.

The incorporation of equimolar doses of [14C]-palmitic and [3H]-oleic acids into the lipid moiety of hepatocytes after muramyl dipeptide (MDP), adamantylamide dipeptide (AdDP) and levamisole (LM) administration were investigated. The utilization of [14C]-palmitic acid for the synthesis of neutral lipids and phospholipids of crude nuclear fraction increased significantly after MDP and AdDP administration and to a lesser extent after LM. While the incorporation of [3H]-oleic acid did not change significantly after MDP and AdDP, LM significantly increased the incorporation of this acid into the phospholipids of nuclear and microsomal fractions. The changes in the incorporation of saturated palmitic and unsaturated oleic acids suggest a possible influence on deacylation-reacylation mechanisms. These changes could alter the physical properties of membrane and affect certain membrane functions, including the activity of membrane bound enzymes and transport mechanisms.

Unlabelled: Forearm and systemic adipose tissue free fatty acid (FFA) release was measured in eight nonobese, six lower-body obese, and eight upper-body obese women under basal, hyperinsulinemic, and hypoinsulinemic conditions to determine whether forearm fat is regulated in a similar manner as whole body fat., Results: Adipose tissue palmitate release was greater from forearm than whole body (5.97 +/- 0.75 vs. 3.84 +/- 0.34 mumol.kg fat-1.min-1, respectively, P less than 0.005, n = 22 subjects). Systemic palmitate release, relative to fat mass, was significantly (P less than 0.01) greater in nonobese than upper-body obese, and upper-body obese than lower-body obese women, and forearm adipose tissue palmitate release followed the same pattern. Hyperinsulinemia suppressed systemic and forearm lipolysis to similar degrees, however, hypoinsulinemia consistently increased systemic palmitate flux without increasing forearm palmitate release. These results confirm the heterogeneity of adipose tissue in an in vivo model and emphasize the need to consider which adipose tissue depots are responsible for the differences in systemic FFA flux in obese and nonobese humans.

Newly synthesized 17-[18F]fluoro-3-methylheptadecanoic acid ([18F]BMHDA), 16-[18F]fluoropalmitic acid ([18F]PA) and 15-(p-[125I]iodophenyl)-3-R,S-methylpentadecanoic acid ([125I]BMIPP), fatty acid tracers, were examined for the possibility of tumor imaging using 10 tumor models in rats and mice. The highest tumor/muscle ratios with [18F]BMHDA were 2.3 using rat tumor AH109A and 1.9 using mouse tumor B16F1 (tumor accumulation 0.41 +/- 0.05, 4.29 +/- 0.77% ID/g, respectively). Tumor/muscle ratios with [125I]BMIPP and [18F]PA were lower than those with [18F]BMHDA. Labeled fatty acids seem to have a lower potential for tumor detection.