Delayed secretory pathway contributions to VLDL-triglycerides from plasma NEFA, diet, and de novo lipogenesis in humans

Newly synthesized triglyceride (TG) may exit the liver immediately as VLDL-TG or be stored and secreted after a delay. We quantified the contributions from plasma NEFA, diet, and de novo lipogenesis (DNL) to VLDL-TG via immediate and delayed pathways in five lean, normolipidemic subjects; six obese, hypertriglyceridemic (HPTG) nondiabetics; and six obese, HPTG diabetics. Intravenous [(2)H(31)]palmitate and [1-(13)C(1)] acetate and oral [(2)H(35)]stearate were administered for 30 h preceding an overnight fast. [1,2,3,4-(13)C(4)]palmitate was infused during the subsequent 12 h fast. Contributions from plasma NEFA via the immediate pathway were 64 +/- 15, 33 +/- 6, and 58 +/- 2% in control, HPTG, and diabetic HPTG, respectively. Delayed pool fractional contributions were as follows: dietary FA, 2.0 +/- 0.9, 2.5 +/- 1, and 12 +/- 2%; DNL, 3 +/- 0.3, 14 +/- 3, and 13 +/- 4%; delayed NEFA, 15 +/- 4, 20 +/- 4, and 30 +/- 3%. VLDL-TG production rates and absolute input rates from the delayed pool were significantly higher in HPTG and diabetic HPTG than in controls. In conclusion, we provide direct kinetic evidence for a hepatic TG storage pool in humans and document its metabolic sources. The turnover time and sources of this pool differ in diabetic HPTG and nondiabetic HPTG, with potential therapeutic implications.     

Metabolism of very low density lipoproteins in genetically lean or fat lines of chicken

Metabolism of very low density lipoproteins (VLDL) has been compared in fat (FL) and lean (LL) lines of chicken. When refed after fasting, plasma triglyceride concentration reached a significantly higher plateau in FL, although their feed consumption was lower than in LL. Newly synthesized VLDL were studied using anti-lipoprotein lipase antibodies. VLDL triglyceride (TG) concentrations were increased by antibody injection and reached a higher concentration in FL plasma than in LL. Newly synthesized VLDL exhibited a similar lipid composition. Fatty acid profiles were also similar when birds ingested a very low fat diet. Comparison of in vitro affinity of lipoprotein lipase and VLDL from both genotypes did not reveal any difference in Km and Vmax. [14C]labelled VLDL from fat or lean donors were prepared and were injected into chickens from both genotypes. Fractional rate constants did not differ between lines. However, as plasma VLDL-TG pools were very different, plasma turnover was higher in FL than in LL. About 3-fold more VLDL-TG were incorporated in abdominal fat of FL than in LL. Difference in fattening between both genotypes seem to be due to both increased VLDL secretion and VLDL removal from plasma without difference in VLDL characteristics.     

Measuring VLDL-triglyceride turnover in humans using ex vivo-prepared VLDL tracer

There has been more interest in VLDL-triglyceride (TG) kinetics during the last decade. Unfortunately, robust measurement methods are elaborate and not readily available. Here, we describe a method using unique, ex vivo labeling of the fatty acid moiety of VLDL-TG followed by intravenous bolus infusion in the same person. We found that plasma disappearance of ex vivo-labeled VLDL-TG was comparable to that of in vivo-labeled VLDL-TG and that turnover rates can be safely estimated from the log linear decay of VLDL-TG specific activity. We found minor labeling of the plasma FFA (oleate) pool, which was largely attributable to coinfusion of free [14C]triolein; VLDL-TG did not contribute substantially to the plasma FFA pool. The plasma decay curve of VLDL-TG was not affected by the presence of tracer in the FFA pool, provided that the data from 2 h after the VLDL tracer bolus infusion was used. The FFA contamination problem was circumvented by minor modification of the VLDL-TG tracer preparation. The approach we describe should expand the opportunity to study processes that cannot be assessed if the FFA precursor pool is labeled. This method for VLDL-TG tracer preparation can allow measurement of VLDL turnover, tissue uptake of VLDL-TG, and oxidation of VLDL-TG.     

The influence of three days or three weeks of force feeding on the transport of plasma lipids in young female chickens (Gallus gallus L.).

1. Ten-week old broiler females were force-fed (FF) for 3 days or 3 weeks. 2. Control livers were lighter in weight and contained less total lipid, neutral lipid and phospholipid than either FF group, which did not differ. 3. Radioactivity incorporated into liver neutral lipid fractions from 1-[14C]acetate injection was greater in birds FF 3 weeks than controls. Those FF 3 days were intermediate. In all groups, the triglyceride fraction contained 90-94% of isolated radioactivity, the cholesterol fraction 4-8% and the cholesterol ester fraction 1-2%. 4. Plasma lipids were elevated in the birds FF for 3 weeks, but not in those FF 3 days. After injection of 1-[14C]acetate, plasma lipid specific radioactivities were not different between the 3 groups at 20 and 60 min post injection, but were greater in the controls at 120 min.     

Effects of exercise on VLDL-triglyceride oxidation and turnover

Lipids are important substrates for oxidation at rest and during exercise. Aerobic exercise mediates a delayed onset decrease in total and VLDL-triglyceride (TG) plasma concentration. However, the acute effects of exercise on VLDL-TG oxidation and turnover remain unclear. Here, we studied the acute effects of 90 min of moderate-intensity exercise in healthy women and men. VLDL-TG kinetics were assessed using a primed constant infusion of ex vivo labeled [1-(14)C]triolein VLDL-TG. Fractional VLDL-TG-derived fatty acid oxidation was measured from (14)CO(2) specific activity in expired air. VLDL-TG concentration was unaltered during exercise and early recovery, whereas non-VLDL-TG concentration decreased significantly.VLDL-TG secretion rate decreased significantly during exercise and remained suppressed during recovery. Total VLDL-TG oxidation rate was unaffected by exercise. However, the contribution of VLDL-TG oxidation to total energy expenditure fell from 14 ± 9% at rest to 3 ± 4% during exercise. We conclude that VLDL-TG fatty acids are quantitatively important oxidative substrates under basal postabsorptive conditions but remain unaffected during 90-min moderate-intensity exercise and, thus, become relatively less important during exercise. Lower VLDL secretion rate during exercise may contribute to the decrease in TG concentrations during and after exercise.     

Effects of ethanol, fructose, and ethanol plus fructose infusions on plasma glucose concentration and glucose turnover in monkeys (Macaca fascicularis) as measured by [6-3H]glucose

Six adult, female, cynomolgus monkeys were fasted for 64 hr and then continuously infused with [6-3H]glucose to determine the rates of glucose turnover and clearance while they were also being infused with ethanol (110 mumol/min/kg), 1,3-butanediol (110 mumol/min/kg), fructose (30 mumol/min/kg) or ethanol plus fructose (110 and 30 mumol/min/kg) respectively. Both ethanol and 1,3-butanediol infusions decreased the glucose turnover rate (the steady-state input-output rate from the plasma glucose pool) and the plasma glucose concentration by halving the glucose production rate. In contrast, fructose infusions increased the glucose turnover rate and glucose concentration by increasing the glucose production rate by 20%. The plasma clearance rate of glucose was lowest when the animals were infused with ethanol plus fructose; this suggests that acetate from ethanol oxidation may have a glucose-sparing effect if normoglycemia is maintained.     

Energy expenditure, insulin, and VLDL-triglyceride production in humans

Hypertriglyceridemia is considered a cardiovascular risk factor in diabetic and nondiabetic subjects. In this study, we aimed to determine potential regulators of very low density lipoprotein-triglyceride (TG) production. VLDL-TG kinetics were measured in 13 men and 12 women [body mass index [mean (range)]: 24.8 (20.2-35.6) kg/m(2)]. VLDL-TG production was assessed from the plasma decay of a bolus injection of ex vivo labeled VLDL particles ([1-(14)C]triolein-VLDL-TG). Similar VLDL-TG production (micromol/min) was found in men and women. VLDL-TG production was not significantly correlated with palmitate flux ([9,10-(3)H]palmitate) (r = 0.09, P = 0.67) or palmitate concentration (r = -0.29, P = 0.2) but was correlated significantly with fasting insulin concentration (r = 0.46, P < 0.05) and resting energy expenditure (REE) (r = 0.45, P < 0.05). The latter correlation improved when adjusted for sex. The best multivariate model with VLDL-TG production as the dependent variable and REE, body composition, hormones, and substrate levels as independent variables included fasting insulin (P = 0.02) and REE (P = 0.02) (r(2) = 0.32, P < 0.001). We conclude that VLDL kinetics are similar in men and women and that REE and plasma insulin are significant independent predictors of VLDL-TG production. FFA availability and body fat distribution are unrelated to VLDL production. We suggest that REE plays a greater role in VLDL-TG production than previously anticipated. REE and insulin should be taken into account when VLDL-TG production comparisons between groups are made.     

Chylomicron metabolism in an animal model for hyperlipoproteinemia type I.

Mink homozygous for the mutation Pro214Leu in lipoprotein lipase (LPL) had only traces of LPL activity but amounts of LPL protein in their tissues similar to those of normal mink. In normal mink, lymph chylomicrons from rats given [3H]retinol (incorporated into retinyl esters, providing a core label) and [14C]oleic acid (incorporated mainly in triglycerides (TG)) were rapidly cleared from the circulation. In the homozygous mink, clearance was much retarded. The ratio of TG to core label in plasma did not decrease and much less [14C]oleic acid appeared in plasma. Still, half of the labeled material disappeared from the circulating blood within 30;-40 min and the calculated total turnover of TG in the hypertriglyceridemic mink was almost as large as in normal mink. The core label was distributed to the same tissues in hypertriglyceridemic mink as in normal mink. Half to two-thirds of the cleared core label was in the liver. The large difference was that in the hypertriglyceridemic mink, TG label (about 40% of the total amount removed) followed the core label to the liver and there was no preferential uptake of TG over core label in adipose or muscle tissue. In normal mink, only small amounts of TG label (<10%) appeared in the liver, while most was in adipose and muscle tissues. Apolipoprotein B-48 dominated in the accumulated TG-rich lipoproteins in blood of hypertriglyceridemic mink, even in fasted animals.     

Chylomicron remnant cholesteryl esters as the major constituent of very low density lipoproteins in plasma of cholesterol-fed rabbits.

Feeding rabbits 500 mg of cholesterol daily for 4 to 15 days greatly increased the concentration of esterified cholesterol in lipoproteins of d less than 1.006 g/ml. The origin of hypercholesterolemic very low density lipoproteins was investigated by monitoring the degradation of labeled lymph chyomicrons administered to normal and cholesterol-fed rabbits. Chylomicrons were labeled in vivo by feeding either 1) [3H]cholesterol and [14C]oleic acid or 2) [14C]cholesterol and [3H]retinyl acetate. After intravenous injection of labeled chylomicrons to recipient rabbits, [14C]triglyceride hydrolysis was equally rapid in normal and cholesterol-fed animals. Normal rabbits rapidly removed from plasma both labeled cholesteryl and retinyl esters, whereas cholesterol-fed rabbits retained nearly 50% of doubly labeled remnants in plasma 25 min after chylomicron injection. Ultracentrifugal separation of plasma into subfractions of very low density lipoproteins showed that chylomicron remnants in cholesterol-fed animals are found among all subclasses of very low density lipoproteins. Analysis of cholesteryl ester specific activity-time curves for the very low density lipoproteins subfraction from hypercholesterolemic plasma showed that nearly all esterified cholesterol in large very low density lipoproteins and approximately 30% of esterified cholesterol in small very low density lipoproteins was derived from chylomicron degradation. Apparently, nearly two-thirds of the esterified cholesterol in total very low density lipoproteins from moderately hypercholesterolemic rabbits is of dietary origin.     

Formation and turnover of triglyceride-rich vesicles in the chick liver cell. Effects of cAMP and carnitine on triglyceride mobilization and conversion to ketones

Refractile cytoplasmic vesicles are formed in less than 10 h when chick liver cell monolayers are incubated with serum-free medium containing 0.9 mM oleate. These vesicles are identical in microscopic appearance to those formed in monolayers by de novo fatty acid synthesis (Tarlow, D. M., Watkins, P. A., Reed, R. E., Miller, R. S., Zwergel, E. E., and Lane, M. D. (1977) J. Cell Biol. 73, 332-353), but require about one-seventh the incubation time to achieve comparable size. After release from the cells by lysis in hypotonic medium, the vesicles can be isolated by flotation at 27,000 X g. Electron microscopy reveals that the isolated vesicles are rimmed by a membrane. Analysis of vesicles isolated from cells labeled with [14C]oleate or [14C]acetate showed that greater than 95% of their 14C content was in the form of triglyceride and that most cellular [14C]triglyceride was contained in the triglyceride-rich vesicles. Exposure of cells to dibytyryl-cAMP after removal of oleate from the medium caused the disappearance of triglyceride-rich vesicles within 36 h. In the absence of cyclic nucleotide, the vesicles persist. Consistent with this morphological change, dibutyryl-cAMP caused a 5.5-fold activation of the apparent rate of mobilization of cellular [14C]triglyceride from cells previously labeled with [14C]oleate. L-(--)-Carnitine alone had no effect; however, when added with dibutyryl-cAMP, cellular triglyceride mobilization was activated 7.4-fold. Although [14C]triglyceride was the principal 14C-labeled product secreted in the absence of cyclic nucleotide and comprised 90% of the total, [14C]acetoacetate and [14C] beta-hydroxybutyrate became major products when cells were treated with dibutyryl-cAMP. Thus, dibytyryl-cAMP activated ketogenesis from cellular [14C]triglyceride by 200-fold and when added with L-(--)-carnitine, by 400-fold. Cells containing triglyceride-rich vesicles labeled with [2-glyceryl-3H]triglyceride were generated by incubation with medium containing [2-3H]glycerol. A comparison of the rates of loss of cellular [1-oleoyl-14C- and [2-glyceryl-3H]triglyceride revealed that substantial re-esterification, i.e. recycling, of 14C-fatty acid released by lipolysis occurred. Under conditions where recycling of 3H label ws minimal, it was determined that 15% of the cellular [2-glyceryl-3H]triglyceride was secreted "en bloc," i.e. without prior lipolysis. En bloc secretion was not affected by dibutyryl-cAMP. The rate of lipolysis of vesicle-associated [2-glyceryl-3H]triglyceride was increased 2.2-fold in the presence of dibutyryl-cAmP. Chloroquine markedly inhibited the dibutyryl-cAMP-dependent lipolysis suggesting the participation of lysosomes in the mobilization of triglyceride-rich vesicles. Mechanisms are presented which could account for the effects of cAMP and carnitine on the turnover of vesicle triglyceride both at the level of lipolysis and the utilization of the released fatty acids by mitochondria...     

Increased concentration of plasma cholesteryl ester transfer protein in nephrotic syndrome: role in dyslipidemia.

Hyperlipidemia is a prominent feature of the nephrotic syndrome. Lipoprotein abnormalities include increased very low and low density lipoprotein (VLDL and LDL) cholesterol and variable reductions in high density lipoprotein (HDL) cholesterol. We hypothesized that plasma cholesteryl ester transfer protein (CETP), which influences the distribution of cholesteryl esters among the lipoproteins, might contribute to lipoprotein abnormalities in nephrotic syndrome. Plasma CETP, apolipoprotein and lipoprotein concentrations were measured in 14 consecutive untreated and 7 treated nephrotic patients, 5 patients with primary hypertriglyceridemia, and 18 normolipidemic controls. Patients with nephrotic syndrome displayed increased plasma concentrations of apoB, VLDL, and LDL cholesterol. The VLDL was enriched with cholesteryl ester (CE), shown by a CE/triglyceride (TG) ratio approximately twice that in normolipidemic or hypertriglyceridemic controls (P < 0.001). Plasma CETP concentration was increased in patients with untreated nephrotic syndrome compared to controls (3.6 vs. 2.3 mg/l, P < 0.001), and was positively correlated with the CE concentration in VLDL (r = 0.69, P = 0.004) and with plasma apoB concentration (r = 0.68, P = 0.007). Treatment with corticosteroids resulted in normalization of plasma CETP and of the CE/TG ratio in VLDL. An inverse correlation between plasma CETP and HDL cholesterol was observed in hypertriglyceridemic nephrotic syndrome patients (r = -0.67, P = 0.03). The dyslipidemia of nephrotic syndrome includes increased levels of apoB-lipoproteins and VLDL that are unusually enriched in CE and likely to be atherogenic. Increased plasma CETP probably plays a significant role in the enrichment of VLDL with CE, and may also contribute to increased concentrations of apoB-lipoproteins and decreased HDL cholesterol in some patients.     

Transformation of (sup14)C-Lignin-Labeled Cell Walls of Wheat by Syntrophococcus sucromutans, Eubacterium oxidoreducens, and Neocallimastix frontalis

Wheat cell walls, saponified or not, labeled with [U-(sup14)C]phenylalanine or [O-methyl-(sup14)C]sinapate were fermented by Neocallimastix frontalis or Syntrophococcus sucromutans plus Eubacterium oxidoreducens or a mixed culture. Phenolics were less solubilized but more transformed by bacteria than by the fungus, and mineralization was slight. S. sucromutans O-demethylated [O-methyl-(sup14)C]syringyl lignins, yielding labeled acetate.     

Transmucosal triglyceride transport rates in proximal and distal rat intestine in vivo.

Transmucosal transport rates for triolein in proximal and distal intestine were compared in unanesthetized rats. Emulsified [1-14-C] triolein together with bile and pancreatic juice from donor rats was infused for 6 hr into either the duodenum or the midpoint of the small intestine at such a rate that absorption was essentially complete in both regions of the intestine. Lymph was collected from the thoracic duct during triolein infusion and for an additional 6-hr period. The decrease in the rate of lymphatic output of labeled fat was found to follow a simple exponential function in all animals. This rate of decrease (decay rate) was used to calculate the half-times of lipid turnover through the intestinal wall and the fractional output rates. Distal intestine transported lipid 40% more slowly than proximal intestine, and the difference was associated with a greater accumulation of triglyceride in the distal intestinal wall. Chylomicron synthesis and/or release is the rate-limiting step for distal lymphatic fat transport in vivo, whereas fat uptake from the lumen is rate limiting for proximal intestine.     

Effects of fish oil on VLDL triglyceride kinetics in humans

Dietary n-3 fatty acids (FAs) found in fish oils markedly lower plasma triglyceride (TG) and very low density lipoprotein (VLDL) levels in both normal and hypertriglyceridemic subjects. The present study examined the mechanism of this effect. Ten subjects with widely different plasma triglyceride levels (82 to 1002 mg/dl) were fed metabolically controlled diets containing 20% fat. The control diet contained a blend of cocoa butter and peanut oil (P/S = 0.8). The test diet contained fish oil (P/S = 1.1) and provided 10-17 g of n-3 FAs per day (depending on calorie intake). After 3 to 5 weeks of each diet, the kinetics of VLDL-TG were determined over a 48-h period after the injection of [3H]glycerol. The fish oil diet reduced the VLDL-TG synthetic rate from 23 +/- 14.3 (mean +/- SD) to 12.6 +/- 7.5 mg/h per kg ideal weight (P less than 0.005) and increased the fractional catabolic rate (FCR) for VLDL-TG from 0.23 +/- 0.12 to 0.38 +/- 0.16 h -1 (P less than 0.005). At the same time, there was a 66% reduction of plasma triglyceride levels, resulting largely from a 78% decrease in VLDL-TG levels (398 +/- 317 to 87 +/- 77 mg/dl; P less than 0.005). There was a strong correlation (r = 0.83; P less than 0.01) between the change in synthetic rates and pool sizes, but there was no correlation (r = 0.24; NS) between changes in FCRs and pool sizes. The VLDL cholesterol: triglyceride ratio increased during the n-3 diet suggesting that smaller VLDL particles were present. These particles would be expected to leave the VLDL fraction more rapidly than larger particles producing a higher FCR. We conclude that the hypotriglyceridemic effect of fish oil appears to be caused primarily by an inhibition of very low density lipoprotein-triglyceride synthesis, but an additional, independent effect upon VLDL catabolism cannot be ruled out.     

Use of stable isotopically labeled tracers to measure very low density lipoprotein-triglyceride turnover

Tracer methods for VLDL-TG kinetics vary in their ability to account for the effect of tracer recycling, which can influence the calculation of VLDL-TG fractional catabolic rates (FCRs). We evaluated a novel approach, involving stable isotopically labeled glycerol or palmitate tracers in conjunction with compartmental modeling, for measuring VLDL-TG kinetics in normolipidemic human subjects. When administered as a bolus simultaneously, both tracers provided identical VLDL-TG FCRs when the data were analyzed by a compartmental model that accounted for hepatic lipid tracer recycling, but not by non-compartmental analysis. The model-derived FCR was greater than that determined using a non-compartmental approach, and was 2- to 3-fold higher than that usually reported by using a bolus of radioactive [3H]glycerol. When palmitate tracer was given as a constant infusion, VLDL-TG turnover appeared 5-fold slower, because tracer recycling through hepatic lipid pools could not be resolved with the infusion protocol. We conclude that accounting for tracer recycling, particularly the contribution of hepatic glycerolipid pools, is essential to accurately measure VLDL-TG kinetics, and that bolus injection of stable isotopically labeled glycerol or palmitate tracers in conjunction with compartmental modeling analysis offers a reliable approach for measuring VLDL-TG kinetics.     

Lipid metabolism. Evidence of a δ-oxidation pathway for saturated fatty acids

1. Specific radioactivities of milk triglyceride fatty acids and gamma- and delta-hydroxy fatty acids were measured after the intramammary infusion of [1-(14)C]acetate, delta-hydroxy[1-(14)C]laurate and [1-(14)C]laurate as their sodium salts into fed lactating goats. 2. Net incorporations of the radioactive tracer into the total milk lipids were comparable, being 16, 17 and 21% of the label infused respectively. 3. The specific radioactivities of the C(4)-C(8) fatty acids after [1-(14)C]acetate infusion were lower than those of the C(10)-C(14) fatty acids. 4. After delta-hydroxy[1-(14)C]laurate administration the milk triglyceride fatty acids were labelled and their specific radioactivities were characterized by decreasing values with increasing chain length of the fatty acids, implicating C(4) unit incorporation. 5. The gamma- and delta-hydroxy fatty acids isolated after [1-(14)C]laurate infusion were highly labelled and the milk triglyceride fatty acids, other than laurate, exhibited a labelling pattern similar to that of the fatty acids derived from the radioactive delta-hydroxy fatty acid. 6. Evidence is presented for the existence of saturated fatty acid delta-oxidation in the mammary gland, in which the gamma- and delta-hydroxy fatty acids are active intermediates.     

N tau methyl histidine excretion and [U-14C]amino acid oxidation in fully chickens from two lines selected for high and low body fat contents

Protein turnover and amino acid oxidation were examined in two lines of domestic broiler chicken selected for high (fat) and low (lean) plasma very low density lipoprotein levels. Protein turnover was assessed by comparing N tau methyl histidine excretion in two lines. No differences between the fat and lean birds were found. Oxidation of [U-14C]amino acids in vivo was measured in birds of both lines which had been fed ad lib. The lean birds oxidized significantly less of the administered [U-14C]amino acids to 14CO2 than their fat counterparts during the first 90 min following administration. Lean birds also excreted significantly less 14C than fat birds from [U-14C]amino acids.     

Turnover and tissue distribution of 125-I-labeled low density lipoprotein in swine and dogs.

Swine plasma low density lipoprotein (LDL) isolated ultracentrifugally (d 1.019-1.063) was labeled with 125-I, dialyzed, and reisolated by centrifugation at d 1.063. Over 96% of the radioactivity was shown to be associated with the apoprotein. After reinjection into the donor animal, disapperance of 125-I was followed for up to 122 hr. At all time intervals examined, over 95% of the total plasma 125-I was recovered in LDL (D 1.006-1.063), i.e., there was apparently no transfer of radioactivity to high density or very low density lipoproteins. The disappearance curve was biexponential, with half-lives of 0.83 plus or minus 0.06 and 22.5 plus or minus 1.7 hr for the first and second phases, respectively (13 studies). The mean calculated fractional catabolic rate was 0.041 plus or minus 0.003 hr-minus 1. Similar results were obtained in three dogs using autologous LDL of density 1.020-1.050; fractional catabolic rates were 0.031, 0.031, and 0.029 hr-minus 1. Tissue distribution of 125-I was determined in swine killed at various time intervals after [125-I]LDL injection with corrections for radioactivity in trapped plasma. Of the tissues examined, the liver showed by far the highest concentration. Total hepatic radioactivity, expressed as a percentage of total plasma radioactivity, was rather constant and independent of the time of killing from 3 to 122 hr (15.8 plus or minus 1.9%). The total extravascular LDL pool calculated from analysis of the plasma disappearance curves was about 20-30% of the size of the plasma LDL pool. These data are consistent with the conclusion that the liver accounts for a very large fraction of the total extravascular LDL pool. These data are consistent with the conclusion that the liver accounts for a very large fraction of the total extravascular LDL pool and that it is infairly rapid equilibrium with the plasma pool. To what extent the liver is involved in irreversible degradation cannot be inferred from these findings.     

Contribution of propionate to glucose synthesis in sheep

1. The production rate of propionate in the rumen and the entry rate of glucose into the body pool of glucose in sheep were measured by isotope-dilution methods. Propionate production rates were measured by using a continuous infusion of specifically labelled [(14)C]propionate. Glucose entry rates were estimated by using either a primed infusion or a continuous infusion of [U-(14)C]glucose. 2. The specific radioactivity of plasma glucose was constant between 4 and 9hr. after the commencement of intravenous infusion of [U-(14)C]glucose and between 1 and 3hr. when a primed infusion was used. 3. Infusion of [(14)C]propionate intraruminally resulted in a fairly constant specific radioactivity of rumen propionate between about 4 and 9hr. and of plasma glucose between 6 and 9hr. after the commencement of the infusion. Comparison of the mean specific radioactivities of glucose and propionate during these periods allowed estimates to be made of the contribution of propionate to glucose synthesis. 4. Comparisons of the specific radioactivities of plasma glucose and rumen propionate during intraruminal infusions of one of [1-(14)C]-, [2-(14)C]-, [3-(14)C]- and [U-(14)C]-propionate indicated considerable exchange of C-1 of propionate on conversion into glucose. The incorporation of C-2 and C-3 of propionate into glucose and lactate indicated that 54% of both the glucose and lactate synthesized arose from propionate carbon. 5. No differences were found for glucose entry rates measured either by a primed infusion or by a continuous infusion. The mean entry rate (+/-s.e.m.) of glucose estimated by using a continuous infusion into sheep was 0.33+/-0.03 (4) m-mole/min. and by using a primed infusion was 0.32+/-0.01 (4) m-mole/min. The mean propionate production rate was 1.24+/-0.03 (8) m-moles/min. The conversion of propionate into glucose was 0.36 m-mole/min., indicating that 32% of the propionate produced in the rumen is used for glucose synthesis. 6. It was indicated that a considerable amount of the propionate converted into glucose was first converted into lactate.     

Production of Specifically Labeled Compounds by Methanobacterium espanolae Grown on H(2)-CO(2) plus [C]Acetate

Methanobacterium espanolae, an acidiphilic methanogen, required acetate for maximal growth on H(2)-CO(2). In the presence of 5 to 15 mM acetate, at a growth pH of 5.5, the mu(max) was 0.05 h. M. espanolae consumed 12.3 mM acetate during 96 h of incubation at 35 degrees C with shaking at 100 rpm. At initial acetate levels of 2.5 to 10.0 mM, the amount of biomass produced was dependent on the amount of acetate in the medium. C nuclear magnetic resonance spectra of protein hydrolysates obtained from cultures grown on [1-C]- or [2-C]acetate indicated that an incomplete tricarboxylic acid pathway, operating in the reductive direction, was functional in this methanogen. The amino acids were labeled with a very high degree of specificity and at greater than 90% enrichment levels. Less than 2% label randomization occurred between positions primarily labeled from either the carboxyl or methyl group of acetate, and very little label was transferred to positions primarily labeled from CO(2). The labeling pattern of carbohydrates was typical for glucogenesis from pyruvate. This methanogen, by virtue of the properties described above and its ability to incorporate all of the available acetate (10 mM or lower) from the growth medium, has advantages over other microorganisms for use in the production of specifically labeled compounds.