Hydrolysis of long-chain, n-3 fatty acid enriched chylomicrons by cardiac lipoprotein lipase.

The hydrolysis of chylomicrons enriched in long-chain n-3 fatty acids by cardiac lipoprotein lipase was studied. In 60 min, 24.8% of the triacylglycerol fatty acids were released as free fatty acids. The fatty acids were hydrolyzed at different rates. DHA (docosahexaenoic acid, 22:6n-3) and EPA (eicosapentaenoic acid, 20:5n-3) were released at rates significantly less than average. Stearic acid (18:0), 20:1n-9, and alpha-linolenic acid (18:3n-3) were released significantly faster than average. There was no relationship between the rate of release of a fatty acid and the number of carbons or the number of double bonds. Lipoprotein lipase selectively hydrolyzes the fatty acids of chylomicron triacylglycerols. This selectively will result in remnants that are relatively depleted in 18:0, 20:1, and 18:3 and relatively enriched in 20:5 and 22:6.

Differential accumulation and release of long-chain n-3 fatty acids from liver, muscle, and adipose tissue triacylglycerols.

Eicosapentaenoic acid (EPA, 20:5n-3) is less efficiently accumulated in tissue triacylglycerols (TAGs) during fish oil feeding than docosahexaneoic acid (DHA, 22:6n-3) or docosapentaenoic acid (DPA, 22:5n-3), and EPA is preferentially released from the TAG of isolated adipocytes in vitro and adipose tissue in vivo during fasting compared with DHA or DPA. It is not known if this preferential release occurs in vivo under nonfasting conditions or if it is limited to adipose tissue. Accordingly, we have carried out experiments to study the turnover of EPA, DHA, and DPA in the TAG of adipose tissue, liver, and skeletal muscle. Weanling rats were fed diets containing fish oil for 6 weeks and then switched to diets containing only corn oil as the dietary fat for 8 weeks. The fatty acid composition and mass in epididymal fat pads, omental fat, liver, and soleus muscle TAGs were determined weekly for the first 10 weeks and at weeks 12 and 14. Subsequent to the change to the corn oil diet, EPA (20:5n-3), DPA (22:5n-3), and DHA (22:6n-3), which had accumulated during fish oil feeding, were lost from the tissue TAG pools of each tissue examined. After 8 weeks on the corn oil diet, less than 10% of the accumulated EPA, DPA, and DHA remained in the liver and muscle. The loss of EPA, DPA, and DHA from epididymal fat pad was slower. In each tissue, EPA was lost more rapidly than DPA or DHA. This selective loss of EPA relative to DHA or DPA may explain the previously reported underrepresentation of EPA compared with DHA or DPA in tissue TAG.

Fasting-induced selective mobilization of brown adipose tissue fatty acids.

This study was conducted to determine whether there is selective mobilization of fatty acids from brown adipose tissue. Rats were fed a fish-oil diet and then fasted for 0, 7, or 10 days followed by analysis of the fatty acid content and composition of triacylglycerols (TAG) and phospholipid (PL) in interscapular brown adipose tissue (IBAT). Fatty acids were selectively lost from IBAT triacylglycerols, the mobilization following the same structural rules as those previously demonstrated for white adipose tissue. Fractional mobilization increased with unsaturation at a given chain length and tended to decrease with chain length at a given unsaturation. However, linoleic acid (18:2 n-6) was mobilized significantly less than predicted by these structural rules. In IBAT phospholipid, fatty acids were also selectively lost but there was no such relationship between the fractional mobilization of a fatty acid and its structure. The fatty acids of the n-6 series were exceptional in their behavior because they displayed below average fractional mobilization. In fact, linoleic and arachidonic acids actually increased their mass in IBAT phospholipid during the fast. It is concluded that, in IBAT, fatty acids are selectively mobilized during a fast, and that fasting-induced remodeling of the fatty acid composition leads to the selective retention of linoleate in the case of TAG and all the n-6 fatty acids in the case of PL.

Thiamine, riboflavin, folate, and vitamin B12 status of low birth weight infants receiving parenteral and enteral nutrition.

Thirty infants were randomly assigned to receive either 3 mL of MVI-Pediatric supplement (PAR3 group, parenterally fed) or 2 mL (PAR2 group, parenterally fed). For the first week, 100% received total parenteral nutrition (TPN), 50% by the second, and less than 33% by the third. Eighteen control infants received enteral feeds of infant formula. Baseline (before TPN) and subsequent weekly blood samples, dietary data, and 24-hour urine collections were obtained. The adequacies of thiamine and riboflavin were assessed by the thiamine pyrophosphate effect and erythrocyte glutathione reductase activity, respectively. Urinary thiamine and riboflavin levels were measured by fluorometry. Plasma folate, red blood cell folate, urinary folate, and plasma vitamin B12 concentrations were determined by radioassay. No differences between groups were observed in thiamine pyrophosphate effect, erythrocyte glutathione reductase activity, urinary B1 or B2, or red blood cell folate levels at any time. Plasma folate differed (p less than .05) among the PAR3 group (24 +/- 7 ng/mL), and both the PAR2 (13 +/- 5 ng/mL) and enterally fed (ENT) groups (16 +/- 3 ng/mL) before the initiation of feeds, at week 1 (PAR3 = 32 +/- 15 ng/mL; PAR2 = 18 +/- 4 ng/mL; ENT = 19 +/- 9, ng/mL) and between the PAR3 (30 +/- 16 ng/mL) and PAR2 (16 +/- 4 ng/mL) infants at week 2. Plasma vitamin B12 levels differed among the ENT groups (551 +/- 287 pg/mL) and both the parenteral groups (PAR2 = 841 +/- 405 pg/mL; PAR3 = 924 +/- 424 pg/mL) at week 1 and between the ENT (530 +/- 238 pg/mL) and PAR3 (999 +/- 425 pg/mL) groups at week 2.(ABSTRACT TRUNCATED AT 250 WORDS)

Net renal arginine flux in rats is not affected by dietary arginine or dietary protein intake.

Kidneys of adult animals serve as a major biosynthetic source of arginine. Recently, we demonstrated that kidneys of rats infused with citrulline responded to the elevated plasma citrulline concentrations by increasing citrulline uptake and producing greater quantities of arginine. The objective of the present study was to determine the effects of feeding adult male rats different levels of arginine or protein on renal arginine synthesis. Feeding arginine (0, 0.5 and 2.0%) for 1 wk increased circulating plasma concentrations of arginine 2.5-fold (196 mumol/L) in the 2.0% arginine group compared with the 0 and 0.5% arginine groups (73 and 81 mumol/L, respectively). However, the plasma citrulline concentration was unchanged. The renal uptake of citrulline and release of arginine were similar in all the three groups fed different levels of arginine. Feeding diets containing 5, 12 and 50% protein for 1 wk did not alter the circulating plasma concentrations of either citrulline or arginine, and there were no significant differences in the renal release of arginine. These results suggest that renal arginine synthesis is independent of dietary arginine or protein intake.

Intestinal absorption of fish oil in rats previously adapted to diets containing fish oil or corn oil.

A study was conducted to evaluate whether the composition of previous dietary fat affects the absorption and composition of lymph obtained after a meal of fish oil. Adult male Sprague-Dawley rats were fed diets containing either corn oil or fish oil (MaxEPA) for 2 weeks. They were then given intraduodenally a bolus of an emulsion of 0.5 ml of fish oil plus 0.5 ml of 20 mM sodium taurocholate. Intestinal lymph was collected from a cannula in the main intestinal lymph trunk for various times after oil administration. Rats proportion of the test dose fo fish oil than those fed corn oil. There was an effect of previous diet on the fatty acid composition of the lymph. Rats fed fish oil had a higher percentage of eicosapentaenoic and docosahexaenoic acids in the lymph lipids than those fed corn oil while those fed corn oil had a higher percentage of linoleic acid. These results rule out decreased intestinal absorption as a mechanism for the hypotriacylglycerolemic effect of dietary fish oils. They also indicate a significant contribution of endogenous lipids to the fatty acids in lymph.

Cellular and subcellular localization of enzymes of arginine metabolism in rat kidney.

Rat kidneys extract citrulline derived from the intestinal metabolism of glutamine and convert it stoichiometrically into arginine. This pathway constitutes the major endogenous source of arginine. We investigated the localization of enzymes of arginine synthesis, argininosuccinate synthase and lyase, and of breakdown, arginase and ornithine aminotransferase, in five regions of rat kidney, in cortical tubule fractions and in subcellular fractions of cortex. Argininosuccinate synthase and lyase were found almost exclusively in cortex. Arginase and ornithine aminotransferase were found in inner cortex and outer medulla. Since cortical tissue primarily consists of proximal convoluted and straight tubules, distal tubules and glomeruli, we prepared cortical tubule fragments by collagenase digestion of cortices and fractionated them on a Percoll gradient. Argininosuccinate synthase and lyase were found to be markedly enriched in proximal convoluted tubules, whereas less than 10% of arginase and ornithine aminotransferase, were recovered in this fraction. Arginine production from citrulline was also enriched in proximal convoluted tubules. Subcellular fractionation of kidney cortex revealed that argininosuccinate synthase and lyase are cytosolic. We therefore conclude that arginine synthesis occurs in the cytoplasm of the cells of the proximal convoluted tubule.

The 1990 Borden Award Lecture. Dietary regulation of fatty acid and triglyceride metabolism.

The level of circulating triacylglycerols is determined by the balance between their delivery into the plasma and their removal from it. Plasma triacylglycerols are derived either from dietary fat as chylomicrons or from endogenous hepatic synthesis as very low density lipoproteins. Their removal occurs through the action of lipoprotein lipase after which the fatty acids are either stored in adipose tissue or oxidized, primarily in skeletal muscle and heart. The composition of the diet has been shown to influence many of these processes. Hepatic fatty acid synthesis and triacylglycerol secretion are affected by the quantity and composition of dietary fat, carbohydrate, and protein. Polyunsaturated but not saturated fats reduce hepatic fatty acid synthesis by decreasing the amount of the lipogenic enzymes needed for de novo fatty acid synthesis. Dietary fish oils are particularly effective at reducing both fatty acid synthesis and triacylglycerol secretion and as a result are hypotriacylglycerolemic, particularly in hypertriacylglycerolemic individuals. In addition, dietary fish oils can increase the oxidation of fatty acids and lead to increased activity of lipoprotein lipase in skeletal muscle and heart. It appears that the hypotriacylglycerolemic effect of dietary fish oils is mediated by effects on both synthesis and removal of circulating triacylglycerols.

Uric acid synthesis by avian exocrine pancreas.

1. Chicken pancreas has been shown to synthesize and secrete uric acid. Uric acid synthesis from xanthine in vitro by isolated pancreatic acinii is saturable and dependent on the activity of xanthine dehydrogenase. 2. Chicken pancreas is unable to synthesize uric acid de novo but the variety of substrates which support urate synthesis suggests that it occurs by the purine degradation pathway.

Renal arginine synthesis: studies in vitro and in vivo.

Renal arginine synthesis is a major endogenous source of arginine. Argininosuccinate lyase occurs almost exclusively in kidney cortex. In studies with isolated renal cortical tubules, we observed rapid rates of arginine synthesis from citrulline, provided a source of the N atom of the guanidino group of arginine was supplied. Aspartate, glutamate, or glutamine were effective, whereas glycine, alanine, serine, or NH4Cl were ineffective as this second substrate. Arginine synthesis as a function of citrulline concentration was determined and was found to be highly sensitive to citrulline concentrations in the physiological range (60 microM), suggesting that renal arginine synthesis in vivo could be regulated by circulating citrulline levels. Therefore, arginine synthesis by the kidney was investigated in vivo by measuring the net flux of citrulline and arginine in saline-infused (control group) and citrulline-infused rats. In normal animals, uptake of citrulline was 60.5 +/- 20.7 nmol.min-1.100 g body wt-1, and a similar arginine release was observed. Citrulline infusion that increased circulating citrulline levels fourfold resulted in a similar increase in renal citrulline uptake (224 +/- 33 nmol.min-1.100 g-1) and a similar increase in renal production of arginine. The results suggest that the availability of citrulline is a limiting factor for renal arginine synthesis in rats.

Tissue distribution of lipogenesis in vivo in the common murre (Uria aalge) and the domestic chicken (Gallus domesticus).

1. Total body lipogenesis was similar in the murre and the chicken. 2. The liver contributes 10.4% to whole body lipogenesis in fed murres when measured in vivo using 3H2O. 3. The liver contributes 28.0% to whole body lipogenesis in the fed chicken. 4. The lower contribution of the liver in the murre may be a consequence of the high fat diet of the murre relative to the chicken.

Intestinal absorption and lymphatic transport of fish oil (MaxEPA) in the rat.

Adult male, chow-fed Sprague-Dawley rats were given intraduodenally a bolus of emulsion of 0.5 ml of fish oil (MaxEPA) or olive oil plus 0.5 ml of 20 mM sodium taurocholate. Intestinal lymph was collected from a cannula in the main intestinal lymph trunk for various times after oil administration. There was no difference in the absorption of either type of oil over 6 and 24 h, over which times about 40 and 70% of the administered dose was taken up. For MaxEPA, the flux of triacylglycerols remained at a basal level of 0.07 mumol/min for 30 min, after which it rose rapidly to a maximum of 0.87 mumol/min between 90 and 120 min. The flux was 0.4 mumol/min for the subsequent 4 h. After 30 min, the composition of the lymph triacylglycerols began to change to show the presence of large proportions of fatty acyl chains that were characteristic of fish oil, especially eicosapentaenoate (20:5(n-3] and docosahexaenoate (22:6(n-3]. The composition of the lymph remained fairly similar to that of the fish oil for up to 6 h, the last time point at which detailed analysis was done. The docosahexaenoate in the triacylglycerols of the fish oil was primarily in the sn-2 position of glycerol, whereas a more random distribution of eicosapentaenoate over all glycerol positions was found. The positional distribution of the acids in the lymph triacylglycerols was similar to that in the fish oil. There was no evidence of substantial chain elongation or shortening during absorption. The results indicate that fish oil is effectively absorbed from the rat intestine without substantial alteration in the acyl chains of the triacylglycerols.

The effect of dietary fish oil on muscle and adipose tissue lipoprotein lipase.

The activity of lipoprotein lipase (LPL) in the adipose tissue and skeletal muscle of rats fed glucose- or fructose-based diets containing fish oil, corn oil or tallow was examined. In addition, heart LPL activity was measured in rats fed a glucose-based diet containing either corn oil or fish oil. Adipose tissue LPL activity was unaffected by dietary fat. In both heart and skeletal muscle, LPL activity was higher in rats fed the fish oil diet. These results suggest that increased removal of triglyceride by muscle may contribute to the blood triglyceride lowering effect of dietary fish oil.

Hepatic fatty acid synthesis and triglyceride secretion in rats fed fructose- or glucose-based diets containing corn oil, tallow or marine oil.

The effects of three dietary fats, corn oil (CO), tallow (T) and marine oil (MO), on serum triglycerides, hepatic lipogenic enzyme activity and lipogenesis in vivo using 3H2O were measured in fed and fasted rats that had been consuming diets in which the carbohydrate was either glucose or fructose. Hepatic triglyceride secretion was also measured in fasted rats fed the same diets. In both the fed and fasted state, hepatic enzyme activity and lipogenesis in vivo were greater in fructose-fed rats than in glucose-fed rats and less in both CO- and MO-fed rats than in T-fed rats. In rats fed glucose, serum triglycerides were lower in fasted rats fed MO than in fasted rats fed CO. In rats fed glucose or fructose, hepatic triglyceride secretion was lower in rats fed MO than in those fed CO or T.

Gluconeogenesis in liver and kidney of common murre (Uria aalge).

Phosphoenolpyruvate carboxykinase (PEPCK) in murre liver occurs in both cytoplasmic and mitochondrial forms. During a 3-day fast, hepatic PEPCK increases from 9.1 U/g with 19% cytosolic to 12.2 U/g with 35% cytosolic. The increase in activity is due almost entirely to increased cytosolic activity. PEPCK in murre kidney was present only in the mitochondrial compartment. Gluconeogenesis in vitro was determined in both hepatocytes and kidney tubules isolated from 3-day-fasted murres. In hepatocytes, lactate was the best substrate, but both pyruvate and alanine were good gluconeogenic substrates. This observation is consistent with the existence of a cytosolic form of PEPCK. In the kidney, glycerol was the best substrate but was only slightly better than lactate. Alanine and pyruvate were not as effective as gluconeogenic precursors, presumably because of the lack of cytosolic PEPCK. We propose that the major site of gluconeogenesis from amino acids in the murre is the liver, since this is a much larger organ than the kidney and has a cytosolic form of PEPCK necessary for gluconeogenesis from oxidized substrates.

Interaction of dietary carbohydrate and fat in the regulation of hepatic and extrahepatic lipogenesis in the rat.

1. In order to examine the interaction of dietary fat and carbohydrate in the regulation of lipid metabolism, we have studied hepatic and extrahepatic lipogenesis, and adipose tissue lipoprotein lipase (EC 3.1.1.34) in rats fed on one of the following diets: a fructose-based diet containing 0 (F0) or 150 g maize oil (F15)/kg, or a glucose-based diet containing 0 (G0) or 150 g maize oil (G15)/kg. 2. The rats were meal-fed on the diets for 2 weeks after which the activities of a number of hepatic 'lipogenic' enzymes were measured and the activity of epididymal-fat-pad lipoprotein lipase. The activities of the lipogenic enzymes were: F0 greater than G0 greater than G15 greater than F15. Lipoprotein lipase activity was F0 = G0 = F15 = G15. The percentage of total body fatty acid synthesis which occurred in the liver was F0 greater than G0 greater than F15 greater than G15. 3. We conclude that fructose-induced hypertriglyceridaemia is primarily a result of the increased hepatic synthesis rather than decreased adipose-tissue lipoprotein lipase activity.

Interorgan metabolism of amino acids, glucose, lactate, glycerol and uric acid in the domestic fowl (Gallus domesticus).

Arterial--venous differences for metabolites across liver, kidney and hindquarters were measured in fed or starved, artificially ventilated chickens. The results indicate that the liver takes up amino acids under both conditions. Urate and glucose are released by the liver in both the fed and the starved state. Lactate and amino acids are extracted from blood by the kidneys, and this increases in the starved chicken. Urate is removed from the circulation by the kidney in the fed and starved state and excreted. In the fed bird there is no significant arteriovenous difference of glucose across the kidney, but in the starved state the kidney releases glucose into the circulation. The hindquarters take up glucose in the fed but not in the starved state. The branched-chain amino acids valine and leucine were taken up by the hindquarters in the fed, but not the starved, chicken. Glycerol is released by the hindquarter of fed and starved chickens. In the starved state, alanine and glutamine represent 57% of the amino acids released by the hindquarter. Lactate is released by the hindquarter of starved chickens and represents the major gluconeogenic carbon source released by the hindquarter and taken up by kidney and liver. Although the liver is the major gluconeogenic organ in the starved chicken, the kidney accounts for approx. 30% of the glucose produced.

Use of alcohol oxidase to measure the methanol produced during the hydrolysis of D- and L-methyl-3-hydroxybutyric acid.

An enzymatic assay for the measurement of methanol has been developed. The assay uses alcohol oxidase and peroxidase coupled to the oxidation of 2,2'-azino-di-(3-ethyl)-benzthiazoline-6-sulfonic acid as the chromogen. The assay is linear up to 50 nmol of methanol in a 200-microliters sample and sensitive; 1.25 nmol of methanol in a 200-microliters sample can be measured. The assay is rapid and measurements can be made at any convenient time between 15 min and 4 h after initiation of the reaction. The assay shows highest activity with methanol but significant activity with other primary alcohols up to 1-butanol. Little activity is shown with secondary alcohols and diols. We have used this assay to follow the hydrolysis of the two isomers of the methyl ester of 3-hydroxybutyric acid.