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[Ultrastructure and morphogenesis of ceroid pigment. II. Late changes of lysosomes in Kupffer cells of rat liver after phagocytosis of unsaturated lipids (author's transl)].

Wistar rats were injected intravenously with cod liver oil emulsion. The lipid droplets ere phagocytized by Kupffer cells and stored in lysosomes. The transformation of these lipid-containing lysosomes into ceroid pigment granules was studied electron-microscopically and cytochemically for a period of 12 weeks after the injection. The lipid droplets enclosed in lysosomes show an increasing and continous condensation from the periphery towards the center due to oxidation and polymerization of unsaturated fatty acids. During the first week almost the total amount of the stored lipids is transformed into an amorphous, highly electron-dense material which disintegrates into cloddy and globular fragments during the following time. The fragments are embedded in a fine granular, slightly electron-dense matrix showing a marked activity of acid phosphatase. The lysosomal structures which contain remnants of condensed oxidized and polymerized lipids are the electron-microscopic equivalent of the granules as seen by light microscopy. These lipids, which have been changed in their molecular structure, cannot be hydrolized by lysosomal enzymes. They remain as an indigestible material, as a waste product in lysosomal residual bodies. Both lipofuscin and ceroid are lysosomal structures containing oxidized and polymerized lipids. The differences between these lipogenous pigments are due to their different formal and causal genesis. Lipofuscin develops in parenchymal and muscle cells by autophagocytosis and by subsequent oxidation and polymerization of segregated membrane lipids. Ceroid is formed in macrophages by heterophagocytosis of unsaturated lipid material which is also oxidized and polymerized.

Acid Phosphatase↗

Growth and dietary utilisation in goldfish (Carassius auratus Linn.) fed diets formulated with various local agro-produces.

During a 60 d feeding trial, goldfish (Carassius auratus) measuring 1.66+/-0.02 g by weight and 4.2+/-0.02 mm by length were fed diets containing 23.34%, 26.21%, 29.30%, 32.24% and 42.53% crude protein in Feeds I through V respectively. The four formulated-diets (Feeds I, II, IV and V) contained groundnut oil-cake, wheat bran, soybean meal, fish meal, cod liver oil, vegetable oil and vitamin-mineral mixture with tapioca as binder and a commercially-available diet (Feed III) procured from the market. All the feeds were dispensed twice daily at 4% body weight. Results revealed that the growth (in terms of net weight gain and specific growth rate) and dietary utilisation (in terms of feed conversion ratio, protein retention efficiency and energy retention efficiency) were observed to be significantly greater (p < or = 0.05) in Feed V which contained 42.53% protein followed by Feeds IV, I, II and III, and 7.58%, 6.11%, 6.63%, 7.86% and 7.91% lipid in Feeds I through V, describing clearly the role of a combination of protein and lipid in the diet for growth and feed utilisation. Lowest growth and dietary utilisation was observed in Feed III (the commercial diet). The alkaline and acid phosphatase activities in liver, gill and intestine as also the RNA:DNA ratio in the muscle tissues were greatest in fish fed Feed V, which indicated best growth. It was thus concluded that a protein content of 40-45% in the diet of young goldfish could significantly enhance growth and dietary utilisation.

Animal Feed↗

Effects of glucagon and insulin on plasma glucose, triglyceride, and triglyceride-rich lipoprotein concentrations in laying hens fed diets containing different types of fats.

The influence of dietary fat supplementations differing in the ratio of n-6 to n-3 polyunsaturated fatty acids (PUFA) on the effects of glucagon and insulin on plasma glucose, triglyceride (TG), and TG-rich lipoprotein concentrations was investigated in laying hens. Birds were fed either a low-fat control diet (LF) or diets supplemented with 4% pumpkin seed oil (PO; rich in n-6 PUFA) or 4% cod liver oil (CO; rich in n-3 PUFA). After 4 wk feeding of the experimental diets, hens were implanted with wing vein catheters and injected with porcine glucagon (20 microg/kg BW) and porcine insulin (0.5 IU/kg BW), 2 to 5 h after oviposition. Plasma glucose, TG, and TG-rich lipoprotein concentrations were determined from 10 min pre-injection to 60 min post-injection. PO diet resulted in a prolonged plasma glucose response to glucagon administration and altered hypoglycemic response to insulin. However, CO diet did not influence plasma glucose response to either glucagon or insulin administration compared to LF diet. The effects of glucagon and insulin on plasma TG and TG-rich lipoproteins were similar for all diets regardless of the amount or type of fat. The results suggest that feeding dietary fats with high n-6 to n-3 PUFA ratio alters the glucagon and insulin sensitivity of plasma glucose in laying hens. Fats rich in n-3 PUFA seem to have no influence on the plasma glucose response to glucagon and insulin.

Animals↗

Different changes of n-6 and n-3 fatty acids in adipose tissue from spontaneously hypertensive (SHR) and normotensive rats after diets supplemented with linolenic or eicosapentaenoic acids.

In spontaneously hypertensive (SHR) and normotensive rats of the Wistar-Kyoto (WKY) and Wistar-Schönwalde (WSCHOE) strain, diets supplemented with n-3 fatty acids of different chain length (alpha-linolenic acid, LNA--C 18:3, n-3 with linseed oil and eicosapentaenoic acid, EPA--C 20:5, n-3 with cod liver oil) were fed over a period of 22 weeks. After the LNA-rich diet, among the long-chain n-3 fatty acids EPA in epididymal adipose tissue remained unchanged, whereas docosapentaenoic (DPA) and docosahexaenoic acids (DHA) fell. The n-6 fatty acids linoleic (LA) and arachidonic acid (AA) both appeared decreased. After the EPA-rich diet, all n-3 fatty acids, i.e. not only EPA, DPA and DHA, but also LNA were augmented when compared with controls fed commercially available pellets. Among the n-6 fatty acids LA was extremely depressed, whereas AA appeared increased. The p/s-ratio was elevated after the LNA-rich diet, but decreased after the EPA-rich diet. The data indicate a differential effect of dietary n-3 fatty acids of different chain length on the supply of other n-3 fatty acids, of LA and AA as well as on the p/s-ratio in adipose tissue of rats. Blood pressure was not influenced by either diet in either SHR or in both normotensive strains of rats.

Adipose Tissue↗

n-3 Essential fatty acids decrease weight gain in genetically obese mice.

1. Lean (ln/ln) and obese (ob/ob) mice were given diets containing a fat source of 100 g evening primrose (Oenothera biennis) oil (fatty acids 18:2n-6, 18:3n-6; EPO) or 100 g cod liver oil (20:5n-3, 22:6n-3; CLO)/kg diet. 2. Weight gain was lower in the ob/ob mice fed on CLO, an effect unrelated to food intake. 3. In the ob/ob mice fed on CLO, thromboxane synthesis by clotting platelets was reduced compared with that in ob/ob mice fed on EPO. 4. The ob/ob CLO-fed mice had lower arachidonic acid but higher levels of n-3 fatty acids in liver, brown adipose tissue and white adipose tissue. 5. The n-3 fatty acids in CLO therefore replaced the n-6 fatty acids in tissue lipids and reduced synthesis of '2 series' prostaglandins in addition to causing lower weight gain in the CLO-fed ob/ob mice.

Animals↗

[The effect of dietary fish oil on bacterial growth in vivo.].

OBJECTIVE: Epidemiological studies have indicated that high intake of w-3 fatty acids influence various diseases such as cardiovascular diseases and autoimmune disorders. These fatty acids are essential in the diet since the body can not form them de novo. Fish oil is rich in w-3 fatty acids but the w-3 content of vegetable oil is low. The research group has shown increased survival of mice fed cod liver oil enriched diet versus mice fed corn oil enriched diet when infected with Klebsiella pneumoniae intramuscularly. In the present study we investigated the effect of dietary fish oil on bacterial growth in vivo. MATERIAL AND METHODS: Mice were fed fish oil enriched diet and a control group was fed corn oil enriched diet for six weeks and then the mice were infected with Klebsiella pneumoniae intramuscularly. The mice were sacrificed at various time intervals and bacteria were counted in blood and in the infected muscle. RESULTS: The bacteria count in blood and tissue was not significantly different between the two groups although a trend was noted towards more growth in the control group. CONCLUSIONS: We conclude that fish oil does not significantly affect bacterial growth in vivo. Hopefully, future research will reveal the pathophysiological effect of fish oil.

English Abstract↗

Effects of dietary marine oils and olive oil on fatty acid composition, platelet membrane fluidity, platelet responses, and serum lipids in healthy humans.

The influence of various dietary marine oils and olive oil on fatty acid composition of serum and platelets and effects on platelets and serum lipids were investigated as part of an extensive study of the effects of these oils on parameters associated with cardiovascular/thrombotic diseases. Healthy volunteers (266) consumed 15 mL/d of cod liver oil (CLO); whale blubber oil (refined or unrefined); mixtures of seal blubber oil and CLO; or olive oil/CLO for 12 wk. In the CLO, seal oil/CLO, and whale oil groups, serum levels of eicosapentaenoic acid (EPA) were increased. In platelets, EPA was increased in the CLO, seal/CLO, and olive oil/CLO groups. The localization of n-3 polyunsaturated fatty acids in the triacylglycerols did not seem to influence their absorption. Intake of oleic acid is poorly reflected in serum and platelets. No significant differences in triacylglycerols (TG), total cholesterol, or high density lipoprotein cholesterol were observed, even though TG were reduced in the CLO, CLO/seal oil, and whale oil groups. Mean platelet volume increased significantly in both whale oil groups and the CLO/olive oil group. Platelet count was significantly reduced in the refined whale oil group only. Lipopolysaccharide-stimulated blood tended to generate less thromboxane B2 in CLO, CLO/seal, and CLO/olive groups. The whale oils tended to reduce in vivo release of beta-thromboglobulin. In conclusion, intake of various marine oils causes changes in platelet membranes that are favorably antithrombotic. The combination of CLO and olive oil may produce better effects than these oils given separately. The changes in platelet function are directly associated with alterations of fatty acid composition in platelet membranes.

Adolescent↗

Modification of lipoprotein patterns and retardation of atherogenesis by a fish oil supplement to a hyperlipidemic diet for swine.

We have studied the effect of addition of 30 ml cod liver oil (FO) daily to a highly atherogenic butter (BT) diet for swine on lesion development in the coronary arteries and aorta, plasma lipoprotein (LP) patterns, plasma levels of thiobarbituric acid-reactive substances (TBARS) and on tritiated thymidine-labeling indices ([3H]TdR LI) of smooth muscle cells (SMC) and monocyte/macrophages (M/M phi) in the atherosclerotic lesions. Seventeen male Yorkshire swine (11.1 +/- 0.4 kg) were divided into 3 groups: BT (n = 6), BT + FO (n = 6) and mash (n = 5). They were fed the respective diets for 4 months. Terminally, fasting plasma was obtained and cholesterol contents were determined in various fractions of lipoproteins separated by density gradient ultracentrifugation, Pevikon block electrophoresis and immunoelectrophoresis. Apoprotein (B, A-I, E and C) contents of the plasma and lipoprotein fractions were determined by polyacrylamide gel electrophoresis and densitometry of gels stained with Coomassie blue. Swine were injected intramuscularly with 0.5 mCi/kg of [3H]TdR 2 h before death. The aorta and coronary arteries were perfusion fixed in situ under anesthesia. Samples were obtained for microscopic morphometry, autoradiography and immunohistochemistry from distal abdominal aorta, thoracic aorta, and proximal coronary arteries; left main (LM), left anterior descending (LAD), left circumflex (LCX), right main (RM), and right coronary artery (RCA). On the BT diet without FO there was extensive atherosclerotic (AS) lesion development, which was drastically reduced by the addition of FO to the BT diet in all sites by from 71 to 94%. The overall plasma cholesterol (CH) levels were reduced only modestly by the FO (816 +/- 64 to 629 +/- 14 mg/dl) but the distribution of CH in the various lipoprotein classes was remarkably altered. The CH in the large lipoprotein molecules containing both B and E apoproteins was reduced from 488 +/- 84 to 204 +/- 17 mg/dl by the FO with an almost corresponding increase in the conventional LDL molecules containing apo B only (158 +/- 29 to 344 +/- 15 mg/dl). We offer the hypothesis that the large apo B,E containing molecules are much more atherogenic than the smaller apo B containing molecules. This hypothesis is supported by a highly significant correlation between extent of lesion development in all arterial sites and plasma levels of CH in apo B,E containing lipoproteins. Plasma TBARS were elevated by the BT + FO diet but seemed to have no significant effect on the lesions.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

Dietary fish oil reduces microthrombi over atherosclerotic lesions in hyperlipidemic swine even in the absence of plasma cholesterol reduction.

We have investigated in swine the effect of fish oil additives to a butter-cholesterol hyperlipidemic diet (BT) on atherogenesis and thrombogenesis when average plasma cholesterol levels were kept similar in fish oil-treated and untreated BT groups. The studies included evaluation of lesion sizes and cell numbers, counts of adherent monocytes over lesions, and counts of platelet clumps (microthrombi) over lesions either attached directly to endothelium or to adherent monocytes. Anatomic sites studied for lesion development were the left anterior descending coronary artery (LAD), the distal 1/5 of the abdominal aorta, and a proximal portion of the thoracic aorta. Counts of attached monocytes and platelet clumps were made by scanning electron microscopy only for the LAD and expressed per mm2 of surface. The most striking new result was in regard to the platelet clumps. These were reduced by the fish oil from 996 +/- 295/mm2 in the untreated BT group to 313 +/- 59 and 364 +/- 105 in BT+cod liver oil and BT+menhaden oil groups, respectively. Most of the platelet clumps were adherent to attached monocytes in all groups and the number of attached monocytes were greatly reduced by the fish oil additive. Thus there were close relationships among platelet clumps, monocytes, and lesion endothelium. Numbers of attachments over nonlesion endothelium were much less than those over lesions in all dietary groups. The most surprising result was the lack of retardation of lesion growth by the fish oil additives in spite of the reduction in attached monocytes and platelet clumps. In previous studies where the high plasma cholesterol levels in the BT swine had been modestly reduced (about 25%) there had been a marked retardation of lesion growth. The current result suggests that plasma cholesterol is the major factor controlling lesion growth in this model through under milder conditions and longer observation periods other factors might become apparent.

Animals↗

Omega-3 fatty acids in Hawaii seafood.

Omega-3 fatty acids, the most potent of which are found in seafood, are of interest because of their effects on cardiovascular and inflammatory diseases and their possible effects on cancer. However, consumers in Hawaii wishing to increase their dietary omega-3 to omega-6 fatty acid ratio are faced with the difficulty that several types of seafood popular in Hawaii and aquacultured seafood new in the marketplace have unknown omega-3 fatty acid levels. The purpose of this work is to determine omega-3 fatty acid levels of selected seafood and fish oil capsules. Several seafoods and some over-the-counter fish oil capsules were sampled and analyzed. Aku eggs, aquacultured hamachi (yellowtail jack from Japan), one sample of turbot, and EPA Plus, Promega, and Omega-3 Super EPA capsules were found to contain high levels of omega-3 fatty acids. Levels were comparable to those in the fatty fishes such as salmon and mackerel. Butterfish, mahimahi eggs; other fish oil capsules (ProEPA and Omega 3) contained moderate levels of omega-3 fatty acids. Fish cakes, seaweed, several lean fishes, and cod liver oil capsules had small quantities of omega-3 fatty acids. It appeared that the omega-3 fatty acid content of aquacultured species studied was significantly higher than in wild caught species. There was a substantial difference between claimed and actual omega-3 fatty acid levels in commercially available fish oil capsules. These findings can help consumers when selecting types of seafood for their diet that are good sources of omega-3 fatty acids.

Fatty Acids, Omega-3↗

[Protective effect of alpha-linolenic acid in encephalomalacia in chickens].

Encephalomacia is a vitamin E deficiency syndrome which affects the cerebellum of young chicks. The lesion includes degenerative alterations of cellular and fibrillar elements, apparently as the result of the ischaemia caused by thrombotic events in the microvascular system. A supply of linoleic acid, as fatty acid methyl esters prepared from safflower oil (Carthamus tinctorius), caused a high incidence of encephalomalacia. On the other hand, linseed oil esters, rich in alpha-linolenic acid, did not induce any symptoms and protected the chicks to a large extend against the development of signs produced by linoleic acid. Fatty acid esters of cod liver oil, rich in long-chain derivatives of alpha-linolenic acid, exerted a relatively weak protective effect. The analytical results show that a supply of alpha-linolenic acid led to an accumulation of eicosapentaenoic acid, 20:5 omega 3, and a reduced concentration of arachidonic acid in the phospholipds of liver and plasma. The results suggest that, under the conditions leading to encephalomalacia, the prostacyclin-thromboxane balance is shifted in direction of an excessive production of TXA2, causing thrombus formation in the capillaries of the cerebellum, alpha-linolenic acid, by modifying the PUFA profile, exerts a multiple action the main result of which appears to be an antithrombotic effect at the level of the microvascular system of the cerebellum.

Animals↗

Ultrasensitive determination of beta-carotene in fish oil-based supplementary drugs by HPLC-TLS.

This research study demonstrates that thermal lens spectrometry (TLS) combined with efficient isocratic HPLC separation provides an excellent analytical means to profile and quantify carotenes in fish body oils and possibly also in vegetable oils. In particular, a highly sensitive, selective, simple and rapid method for the determination of ultratraces of beta-carotene in fish oil-based supplementary drugs has been proposed. The analyte could be determined reliably and precisely in the presence of a complex matrix including other fat-soluble vitamins (A, D and E), polyunsaturated fatty acids, sterols and other pigments by matrix-matched calibration in cod liver oil. The suitability of the method is also evidenced by favourable analytical figures of merit for beta-carotene such as a linearity range of 1-120 ng ml(-1), LOD of 0.58 ng ml(-1), recovery of 101.4+/-3.3% and measurement repeatability of 4.1%.

Antioxidants↗

Iron trends in rat serum related to dietary fish oils under protein malnutrition.

The iron trend in the serum of male rats fed a low wheat gluten (5% level) diet containing several fish oils for 2 to 4 wk was investigated. The body weight in every experimental group decreased during the first 3 d and thereafter their body weights dropped gradually until the 4th week. At the end of the experimental period, the weights of the spleens from the rats in the cod liver oil (CLO) and sardine oil (SO) groups were significantly heavier than those of the corn oil (control: CO) group. The iron concentrations in the serum of all dietary groups at the 4th week had a tendency to decrease compared to that of the control (CO) group. Ferritin in serum in the 4th week was significantly increased compared to that of the 2nd week for both the CO and CLO groups. However, that of the SO group remained at a low level similar to that measured in the 2nd week. The unsaturated iron binding capacity (UIBC) of the SO group in the 2nd week was significantly higher than that of the control (CO) group. In the 2nd week, the non-heme iron content in the liver was almost equal to the standard value. In the 4th week, however, the non-heme iron in the liver tissues of all dietary groups was significantly increased compared to those of the corresponding groups in the 2nd week. These findings suggest that iron accumulates in the liver.

Animals↗

Determination of iodine values using 1,3-dibromo-5,5-dimethylhydantoin (DBH) and ethyl acetate as solvent. Analytical methods with DBH in respect to environmental and economical concern, part 18.

Iodine values (iodine numbers) of several fixed oils and lard can be determined in ethyl acetate, an easily biodegredable solvent, instead of chloroform according to PH. EUR. 2002. Iodine monobromide has been replaced by 1,3-dibromo-5,5-dimethylhydantoin (DBH) and potassium iodide (KI) and the reaction time was reduced to 5 min only. However, cod-liver oil and linseed oil require a reaction time of 30 min and a smaller weight of sample. Longer reaction times are also necessary for soya oil and wheat germ oil. Iodine values of linseed oil determined according to method A of PH. EUR. 2002, are dependent on the amount of sample, even in the range prescribed by the pharmacopoeia.

Acetates↗

Naturally occurring vitamin D3 in fish products analysed by HPLC, using vitamin D2 as an international standard.

A chemical method for the analysis of naturally occurring vitamin D is proposed. The unsaponifiable matter of oils and tissues is prepared, cholesterol is partly removed by double precipitation at low temperature in methanol. The vitamin D fraction is collected on an adsorption column by high performance liquid chromatography. The fraction is further purified and the vitamins D2 and D3 are separated on a partition column (reverse phase) by HPLC. Recovery was 89 to 93%, standard deviation 3%. The only vitamin D analogue found in fish oils, livers and fillets, was cholecalciferol (D3). Hence, ergocalciferol (D2) could be used as an internal standard. The provitamins ergosterol and 7-dehydrocholesterol, as well as the previtamins, were separated from the vitamin D-fraction on the adsorption column. Results in the range 0.050 to 134 microgram D3 per gram (2 to 5360 I.U. per gram) are given. One cod liver oil was analysed in a rat bioassay, giving supporting results.

Adsorption↗

Determination of acid values of fats and oils by flow injection analysis with electrochemical detection.

A new method using a flow injection system with electrochemical detection was developed to determine acid values of fats and oils. VK3 (2-methyl-1,4-naphthoquinone) solution, i.e., ethanol containing 3 mM VK3 and 38 mM LiClO4, was used as the carrier solution. Flow signals were monitored at -0.33 V vs. Ag/AgCl. For preparation of a sample solution, an oil sample was completely dissolved in VK3 solution, or fatty acids were extracted from the sample into this solution. Aliquots (5 microliters) of the sample solution were injected into the flow injection system. Acid values were determined based on flow signals for 14 samples and the results were found to be consistent with those by potentiometric titration. Relative standard deviation was less than 2%. Samples were processed at the rate of 60 h-1. The stability of fish and cod liver oils was followed by measuring acid values for 8 weeks. This method proved to be a simple and rapid means for acid value determination.

Acids↗

Fish oil, atherogenesis, and thrombogenesis.

Marine fish consumption is known to reduce mortality from ischemic heart disease. The use of fish oil as a dietary supplement, however, is not universally recommended. In large doses, fish oil reduces plasma cholesterol and triacylglycerol but increases low density lipoprotein (LDL) levels and the potential for free radical generation and bleeding. Moderate marine fish consumption is known to reduce mortality without altering commonly measured variables, i.e., plasma cholesterol levels, in vitro platelet aggregation, and bleeding times. In swine, we observed that monocyte adhesions and platelet clumps over the lesion surface of proximal left anterior descending (LAD) coronary arteries are markedly reduced when an atherogenic diet was supplemented with cod-liver oil, even when the cholesterol levels were equalized with the untreated group. These findings suggest that fish oil is hypothrombogenic. We developed an in vitro assay to delineate the mechanism whereby fish oil reduced monocyte-endothelial cell interactions in vivo. The effects of supplementing the culture medium with different fatty acids on adhesions between lipopolysaccharide (LPS) stimulated swine aortic endothelial cells (SAEC) and the human monocyte-like cell line, U937, was investigated in a 10 minute adhesion assay at 37 degrees C. Exposure of SAEC for 6 hours to media containing 50-200 microMs eicosapentaenoic (EPA), stearic, oleic, linoleic, and arachidonic acid, respectively, revealed that only EPA reduced U937-SAEC adhesion. Exposure of U937 to EPA also reduced adhesions. EPA was not effective when added to the SAEC more than 2 hours after they were stimulated with LPS. Exposure of human umbilical vein endothelial cells (HUVEC) to EPA reduced the expression of VCAM-1, ELAM-1, and ICAM-1 after 5 hours of stimulation with LPS. These results suggest that EPA may functionally impair the induction/expression of adhesion molecules.

Animals↗