Absorption and metabolic fate of dietary3H-squalene in the rat

The absorption and metabolic fate of dietary squalene were investigated on the rat by administering a single oral dose of³H-squalene and¹⁴C-cholesterol. Experiments on rats with cannulated thoracic duct revealed that³H-squalene was, like¹⁴C-cholesterol, absorbed through the lymphatic vessels and that ca. 20% of absorbed³H-squalene was cyclized to sterols during the transit through the intestinal wall. Feces contained³H-sterols, indicating that newly synthesized mucosal sterols had been secreted into the gut lumen. In intact animals,³H-squalene appeared in the circulation more rapidly than¹⁴C-cholesterol and did not persist to any significant extent in the squalene-rich adipose and muscle tissues. The increase in dietary squalene load (8–48 mg) decreased the absorption percentage of³H-squalene (45–26%) but did not affect the absorption of¹⁴C-cholesterol (47%). Determination of fecal steroids revealed that during the first days absorbed³H-squalene was eliminated to a significantly higher extent than¹⁴C-cholesterol as fecal bile acids (34% vs 11%). The experiments indicate that the rat intestine has a marked capacity for absorbing dietary squalene and that the absorbed squalene is preferentially converted to bile acids in the liver.     

Incubation of human fecal homogenates with 4-14C-cholesterol

Fresh fecal homogenates from nine subjects consuming solid diets have been incubated for seven days at 37 C with 4-¹⁴C-cholesterol. One series of incubations was conducted under nitrogen, another under air. The extent of bacterial modification of cholesterol to known fecal metabolites varied considerably among the subjects, as expected, but when present such bacteria were shown to be highly active in the conditions used. Production of¹⁴CO₂ was essentially zero in all incubations. Recovery of added¹⁴C from the incubated homogenates following extraction with chloroformmethanol (2∶1 v/v) and evaporation to dryness was quantitative in all cases. About 4% of the labeled cholesterol added appeared to be present in acidic components following incubation. It is concluded that in the incubation system used vigorous bacterial conversion of 4-¹⁴C-cholesterol to metabolites known to be produced in the human intestine could occur readily, in either aerobic or anaerobic conditions. However, CO₂ or other small, relatively volatile fragments labeled with¹⁴C could not be detected. Part of this work was carried out at the Institute for Metabolic Research, Oakland, Calif.     

The incorporation of 2-aminoethylphosphonic acid into rat liver diacylglyceroaminoethylphosphonate

Rats intravenously administered (¹⁴C) 2-aminoethylphosphonic acid (AEP) incorporated ca. 16% of the total injected compound into liver lipids. Thin layer chromatography and selective chemical and enzymatic hydrolysis of the labeled lipids demonstrated that essentially all of the radioactivity was in one compound, diacylglycerol-AEP, the phosphonate analog of phosphatidylethanolamine. Lipids from kidneys, heart, skeletal muscle, adipose, pancreas and brain were examined and found to contain less than 2% collectively of the total injected radioactivity. The residues from the tissues contained ca. 3.2% of the total injected (¹⁴C) AEP.     

Untersuchungen zur Organverteilung von Gallensäuren bei gesunden Ratten

Investigations on the Kinetics of Bile Acids in Normal Rats The kinetic of bile acids has been determined in normal rats after administration of ¹⁴C marked Cholic acid and ¹⁴C marked chenodesoxycholic acid. Cholic acid was administered intravenously to 30 rats to evaluate the distribution into organs after 2, 5, 10, 15 and 20 minutes. In 16 animals the biliary elimination has been assessed. In a further 6 animals a total body autoradiography has been performed after intravenous administration of ¹⁴C chenodesoxycholic acid. Already 10 minutes after i.v. administration of ¹⁴C cholic acid more than 50 % of the dose has passed into the intestine. At that time only 12% of the total activity remained in the liver with a conspicuous activity enrichment in the kidneys. The examinations with biliary drainage confirmed the fast flow off of the cholic acid with only little change in the temporal course with increasing cholic acid concentration. After administration of chenodesoxycholic acid the same kinetic has been found as for cholic acid, as confirmed by autoradiography. However, an additional enrichment of the nuclide in the CNS could be detected. These examinations show that the bile acid pool in normal rats is located in the small intestine, the liver only serves for rapid transfer for the bile acids and does not have a depot function.     

Mechanism of suckling-rat hypercholesteremia. II. Cholesterol biosynthesis and cholic acid turnover studies

Cholesterol biosynthesis from acetate-2-¹⁴C by the livers of suckling rats, which is known to be relatively slow, was increased 2–3-fold within 24 hours after severing the bile duct. Cholesterol synthesis by sham-operated litter mates showed no change under similar treatment. Mevalonate biosynthesis from acetate-2-¹⁴C in vitro by recombined liver microsomal supernatant (105,000×g) fractions from suckling rats also was only 10% of that of comparable recombined fractions from normal controls (young adult rats which were consuming colony diet). Activity was not improved by combining either the microsomal or supernatant fraction from suckling rat livers with the complementary fraction from normal adult livers. On the other hand, activity was restored to 100% when microsomes from livers of duct-served suckling rats were combined with the supernatant fraction from normal controls. Likewise, recombined liver fractions prepared from adult rats fed synthetic diets exhibited low activity for mevalonate biosynthesis. Activity was restored by bile duct cannulation, but inhibited when cholic acid was infused into the cannulated animal. Therefore, surgical procedures which interrupt the enterohepatic recirculation of bile components lead to a restoration of cholesterol biosynthesis and, at least in the adult animal where cannulation studies are practicable, this effect can be reversed readily by bile acid infusion. A slow rate of fecal excretion of¹⁴C-cholic acid was observed in suckling rats and rats fed synthetic diets, apparently reflecting an efficient enterohepatic recirculation of bile salts. The data suggest that under these dietary conditions bile salt retention either directly or indirectly influences hepatic synthesis of cholesterol. Presented in part before Federation of American Societies for Experimental Biology, Fed. Proc.24, 1078, 1965 (abstract). and in part at the AOCS Meeting, Los Angeles, April 1966. Journal Paper No.2835, Purdue University Agricultural Experiment Station.     

Synthesis of 14C-labelled FD & C blue no. 1 (brilliant blue FCF) and its intestinal absorption and metabolic fate in rats

[Methylene-¹⁴C]FD & C Blue No. 1 was synthesized in eight steps from barium [¹⁴C]carbonate. Female Sprague-Dawley rats were given a single dose (0·27 mg; 1·74 μCi) of the ^14C-labelled colouring by gavage. In bile-duct ligated rats, intestinal absorption of FD & C Blue No.1 (estimated from urinary ¹⁴C excretion, expired ¹⁴CO₂ and residual radioactivity in internal organs and tissues 96 hr after oral administration) averaged 2·05% of the dose. mean faecal excretion was 97·28% and the total recovery of administered radioactivity was 99·38%. Intestinal absorption [¹⁴C]FD & C Blue No. 1 in intact rats averaged only 0·27% (91·% recovery), while biliary excretion in bile-duct cannulated animals averaged 1·32% of the dose. Thin-layer chromatography of urine and bile samples revealed that about 95% of excreted radioactivity was unaltered FD & C Blue No. 1 and that about 5% was an unidentified metabolite or degradation product of FD & C Blue No. 1. The results show that Fd & C Blue No.1 is poorly absorbed from the gastro-intestinal tract, and undergoes subsequent rapid and complete biliary excretion.     

Quantification of lowered cholesterol oxidation in guinea pigs with latent vitamin C deficiency

Fifty-two male guinea pigs fed on a scorbutigenic diet were divided into a control group (10 mg ascorbicacid per animal per day) and a group with latent vitamin C deficiency (2 weeks on the scorbutigenic diet only, followed by a maintaining dose of 0.5 mg ascorbic acid per animal per day). After 13 weeks, 26-¹⁴C-cholesterol was administered intraperitoneally to all the animals, in which the¹⁴C excretion in the expired CO₂ and the urine and cholesterol specific activity in the blood serum and liver were then studied at intervals of 24 hr and 1, 3, 5, 7, 9 and 11 weeks. The ascorbic acid concentration in the liver and spleen of the control animals was five times higher than in vitain C-deficient animals. The total cholesterol concentration in serum and liver was significantly higher in the vitamin C-deficient guinea pigs. A two-pool analysis of the disappearance curves of serum cholesterol specific activity showed that the size of the cholesterol pool A (blood and tissues with rapid cholesterol exchange) was greater in the vitamin C-deficient animals. The rate of the transformation of cholesterol to bile acids was estimated as the ratio of¹⁴CO₂ expired to liver cholesterol specific activity. Latent vitamin C deficiency caused significant slowing down of this process (controls: 11.8±0.6; vitamin C deficiency: 8.3±0.4 mg/24 r/500 g w/w). A significant correlation between the liver ascorbic acid concentration and the rate of cholesterol transformation to bile acids was found. The results demonstrate that ascorbic acid is necessary for a normal course of cholesterol catabolism. In latent vitamin C deficiency, the rate of cholesterol catabolism slows down and cholesterol consequently accumulates in the blood and liver of vitamin C-deficient guinea pigs.     

Metabolic fate of gossypol: the metabolism of14C-gossypol in rats

Balance studies designed to obtain information concerning the metabolic fate of gossypol in rats were carried out utilizing two groups of animals. One was fed a basal diet and the other the same diet plus 500 ppm of iron as FeSO₄. Single doses of 5 mg each of¹⁴C-labeled gossypol (spec. act. 19.8 μCi/mmole) were administered. The animals were maintained in metabolic cages and killed after various periods of time. The data indicate that gossypol was poorly absorbed from the gastrointestinal tract and rapidly eliminated from the animal body. Although the main route for gossypol elimination from the animal was by fecal excretion in both treatments, the percentage of the total activity eliminated via the feces varied and depended on the level of iron supplied in the diet. The results demonstrate that gossypol was least excreted via urine and that urinary excretion of radioactivity was diminished by iron supplementation to the diet. Most of the radioactivity retained was found in the contents of different parts of the gastrointestinal tract. Tissues, the liver, muscle, kidney and blood had the highest radioactivity, with the liver having the highest specific activity. The data also demonstrate that addition of iron to the ration diminishes¹⁴C radioactivity in the animal body. This effect might be attributed to the formation of chelates that could not be absorbed through the small intestine. Catalysis of the decarbonylation of gossypol by iron also appears to be a factor. Deceased March 9, 1969.     

Arylsulfonate esters as hypocholesteremic agents: III. Mechanism of action studies

The mechanism responsible for the hypocholestermic action of arylsulfonate esters of long chain fatty alcohols has been studied with rats fed either normocholesteremic or hypercholesteremic (1% cholesterol plus 0.5% glycocholate) diets. Linoleyl tosylate is more effective in lowering plasma and liver cholesterol levels of rats on the hypercholesteremic diet than several other hypocholesteremic agents tested. Linoleyl tosylate does not redistribute cholesterol to extrahepatic tissues nor inhibit hepatic cholesterol biosynthesis. Linoleyl tosylate is not effective in counteracting Tritoninduced hypercholesteremia nor in lowering plasma cholesterol levels of the suckling rat. Linoleyl tosylate increases the fecal elimination of dietary [4-¹⁴C] cholesterol and prevents its accumulation in blood and liver. Oleyl p-(n-decyl) benzene sulfonate also prevents the apparent absorption of [26-¹⁴C] cholesterol from the gastrointestinal tract. Linoleyl tosylate increases the fecal excretion of neurtal sterols but not of bile acids. The results indicate that the arylsulfonate esters of long chain fatty alcohols lower body cholesterol levels by inhibiting cholesterol absorption from the gastrointestinal tract. Exactly how absorption is inhibited is not clear, but linoleyl tosylate was found to stimulate the activity of cholesteryl esterase prepared from the intestinal mucosa. Published as Journal Paper No. 6698 Agricultural Experiment Station, Purdue University, Lafayette, IN.     

Effects of a single ingestion of sodium taurocholate on esterified cholesterol concentration in liver and cholesterol turnover in the rat

The overall response of the rat’s cholesterol metabolism to a single ingestion of taurocholate (80 mg) was studied with the isotopic equilibrium method. The bile acid production, measured by the daily¹⁴CO₂ output of rats in isotopic equilibrium of [26-¹⁴C]-cholesterol, initially decreased and then increased. Conversely, the hepatic concentration of esterified cholesterol first increased and then decreased. Moreover, the ingestion of taurocholate increasing the intestinal absorption coefficient of dietary cholesterol increased the abosprtion and decreased the fecal excretion and the intestinal biosynthesis of cholesterol. The balance of these last effects is an excess cholesterol inflow. The classical hypothesis of negative feedback regulation of bile acid production fails to explain the observed biphasic effect of taurocholate. This compound, when its origin is exogenous, appears to stimulate the storage of esterified cholesterol in the liver, at the expense of bile acid synthesis. This accumulation rate takes into account not only the decrease in cholesterol transformation into bile acids but also the excess inflow of cholesterol. As the exogenous taurocholate was eliminated from the body, cholesteryl ester hydrolysis occurred and provided a supplementary source of free cholesterol for bile acid synthesis.     

Influence of dietary fat on bile acid secretion of rats after portal injection of3H-cholesterol and [4-14C] cholesteryl esters

Labeled cholesterol and its esters were injected via the portal vein into bile duct-cannulated rats, subsequent to a 7 week regimen of either 5% safflower oil or 5% beef tallow in a hypercholesterolemic diet. Analysis of bile collected over a 6 hr period from the safflower group showed 8.6% higher output of bile acids, 13.6% higher conversion of³H-cholesterol to bile acids and 40% higher conversion of [4-¹⁴C]cholesteryl oleate to bile acids than bile collected from the tallow group. During the 1st hr conversion of both oleyl and linoleyl esters of¹⁴C-cholesterol to bile acids was much slower than conversion of the free³H-cholesterol to bile acids, thus eliminating these esters as preferred substrate for bile acid formation. However at 6 hr two-thirds of the injected¹⁴C of oleyl ester was recovered in the liver, and about half of this was in the form of free cholesterol. Thus cholesterol ester hydrolase, though inhibited by dietary cholesterol, evidently did not impose limitations on formation of free cholesterol for subsequent oxidation reactions. Specific radioactivities were of doubtful significance because of uncertainities as to “active” pool size. The data suggest that dietary linoleate exerts its hypocholesterolemic effect in some manner other than ester formation and that its point of action involves stimulation of cholesterol oxidation to bile acids. Journal Paper No. 4938 EAS, Purdue University.     

Origins of the cholesterol in milk

Studies were conducted to investigate the origin of milk cholesterol in the ruminant. In the first experiment, [1-¹⁴C] sodium acetate was infused into one side of the udder of a lactating goat via the teat canal whereas in the second, [1,2-³H] cholesterol was injected intravenously and concurrently with a [¹⁴C] acetate intrammamary infusion. In both experiments, blood and milk samples were collected at intervals for 6 days postinjection. Maximum unesterified cholesterol specific activity (sp act) in whole milk appeared at 78 hr after intravenous injections of³H cholesterol and within 3–7 hr after infusion of [¹⁴C]acetate. Virtually all the tritium in milk was associated with unesterified cholesterol. The sp act of¹⁴C-labeled cholesterol was only 20% of gland-synthesized decanoic acid. Decanoic acid is known to be completely synthesized in the mammary gland, and, like cholesterol, acetate is its precursor. The results indicate that, although some milk cholesterol is synthesized in the mammary gland, it is derived principally from serum cholesterol. The data show also that serum cholesterol equilibrates with membrane cholesterol of the lactating cell prior to its secretion in milk.     

Olestra formulation and the gastrointestinal tract

Olestra is a mixture of compounds comprising sucrose esterified with 6–8 long-chain fatty acids. It is not hydrolyzed by pancreatic lipase and as a result is not absorbed from the small intestine. Olestra in general has physical properties similar to those of a triacylglycerol with the same fatty acid composition. Foods made with olestra are virtually identical in taste and texture to those made with typical triacylglycerols. Olestra consumption does not generate hydrolytic products in the small intestine and, therefore, does not generate some of the signals that alter motility in the gastrointestinal tract. A reduction in gastroesophageal reflux with olestra, in contrast to triacylglycerols, is consistent with a lack of effect on stomach emptying. Unlike triacylglycerols that are absorbed in the proximal small intestine, olestra is distributed throughout the small intestine during transit and passes into the colon. In the colon, olestra's effects depend on its physical properties. Liquid nondigestible lipids result in separation of oil from the fecal matrix. Olestra formulations made with specific fatty acid compositions, particularly those containing a solid sucrose polyester component including behenic acid, possess appropriate rheology to hinder separation of oil from the rest of the fecal matrix, thereby reducing gastrointestinal symptoms.     

Arylsulfonate esters of fatty alcohols: IV. Effects on cholesterol catabolism

Hypercholesterolemic rats, fed 1% cholesterol and 0.5% glycocholate, were treated with arylsulfonates in various ways to observe the pattern of cholesterol elimination. Dietary linoleyl p-toluene-sulfonate (LTS) hastened return to normocholesterolemia and lowered hepatic cholesterol either with or without continued cholesterol feeding. LTS administered via the portal vein significantly lowered plasma cholesterol in 48 hr; ethyl linoleate and monoolein produced no lowering. LTS administered via the portal vein to glycocholate-infused rats increased the biliary excretions of label from [4-¹⁴C]cholesterol administered intracardially and also increased total bile acid excretion 21% without increased bile volume when compared to similar injection of ethyl linoleate. No change in biliary excretion of cholesterol was seen. Bile acid kinetics were studied by using isotopic dilution techniques. Cholate turnover was enhanced by feeding oleyl p-toluenesulfonate (OTS) and oleylp-(n-decyl)-benzenesulfonate (ODS) as suggested by a 16–35% decrease in half-life in both normal and hypercholesterolemic rats. Rats consuming a grain-based colony diet had a 54% increase in cholate synthesis when OTS was included in the diet. The composition of bile was changed when either OTS or ODS was fed; an increase in chenodeoxycholate was noted. This change was gradual with OTS but rapid with ODS and paralleled enhanced decay of chenodeoxycholate specific radioactivity in response to treatment. ODS and OTS also increased¹⁴CO₂ expiration from oral [26-¹⁴C] cholesterol in hypercholesterolemic rats. Dietary OTS and ODS elevated hepatic free cholesterol in hypercholesterolemic rats; ODS also elevated plasma free cholesterol and increased cholesteryl ester hydrolase activity in the liver. The data suggest that arylsulfonates stimulate cholesterol catabolism, in addition to the reported inhibition of cholesterol absorption [Lipids 12:819 (1977)]. Published as Journal Paper No. 6699, A.E.S., Purdue University.     

The gastrointestinal handling and metabolism of [1-13C]palmitic acid in healthy women

The gastrointestinal handling and metabolism of [1-¹³C]palmitic acid given as the free fatty acid was examined in six healthy women by measuring the excretion of¹³C-label in stool and in breath as¹³CO₂. The gastrointestinal handling of [1-¹³C]palmitic acid was compared with the apparent absorption of dietary lipid by measuring lipid losses in stool. The variation both within and between subjects was determined by repeating the study in the same individuals on separate occasions. The time course for excretion of label in stool over the five-day study period followed a common pattern, with most of the label excreted over the first two days of the stool collection.¹³C-Label excreted in stool over the five-day study period was 14.3±9.8% of that administered and on repeating the trial was 31.6±24.7% (not significantly different due to variability); there was poor agreement within subjects. Lipid excreted in stool expressed as a percentage of ingested lipid was 5.2±4.4% in Trial 1 and 5.9±4.0% in Trial 2, and was the same in each individual on repeating the trial. There was no clear relationship between the excretion of¹³C-label and lipid in stool (Trial 1:R=−0.43,P>0.40; Trial 2:R=−0.02,P>0.97). On the first occasion, 22.0±4.5% of the administered label was excreted on breath over the 15-h study period and on repeating the trial was 15.8±9.5% (not significantly different) with poor repeatability in a given individual. There was an inverse relationship between the proportion of¹³C-label excreted in stool and that excreted on breath in Trial 1 (R=−0.80,P>0.06) with a weaker association observed in Trial 2 (R=−0.49,P>0.32). Correcting for differences in the apparent absorption of label reduced the variability in its excretion in breath observed between subjects, particularly in Trial 2. It is concluded that although there are differences in the gastrointestinal handling of [1-¹³C]palmitic acid both within and between healthy adults, the postprandial oxidation of absorbed substrate was similar. The assumptions underlying these observations need to be examined by characterizing the nature of¹³C-label in stool.     

Effects of different levels of dietarytrans-octadecenoate on steroid metabolism in rats

Rats were fed semipurified diets containing olive oil or partially hydrogenated corn oil at the 5 or 20% level for ca. 30 days. These fat diets contained the same amount of octadecenoate but differed in the geometry with respect to each fat level. Contents oft-18∶1 were 26% and 41% of total fatty acids, respectively. The linoleic acid content was also made equivalent (3.8 energy %). After feeding on cholesterol-free diets, rats ontrans fat, compared to those oncis fat, showed: (a) no changes in serum cholesterol and apolipoprotein levels, (b) no effects on the bile flow and concentrations of biliary cholesterol or bile acids, (c) a trend toward increased fecal excretion of neutral and acidic steroids, (d) a lesser extent of transformation of cholesterol to coprostanol in the gut, and (e) no changes in the composition of biliary and fecal bile acids. Observations (c) and (d) were more marked with a hightrans fat regimen. These observations, except for serum apolipoproteins and fecal steroid excretion, were practically reproducible even when rats were fed cholesterol-enriched diets. A preliminary part of this study was presented at the 74th annual AOCS meeting, Chicago, 1983.     

Probable sources of plasma cholesterol during phosphatide induced hypercholesterolemia

Rats in isotopic steady state with respect to 4-¹⁴C-cholesterol were infused intravenously with massive amounts of lecithin and also injected once with Na acetate³H. During the following 8 hr their plasma gained an average of 11.3 mg of cholesterol; the specific activity of¹⁴C-cholesterol fell in plasma while total¹⁴C-cholesterol and³H activity doubled as compared to controls. The specific activity of¹⁴C-cholesterol diminished in livers of rats receiving lecithin but not in controls. Specific activity of either isotope in cholesterol of intestine, lungs, muscle, skin and brains was the same in control and experimental groups. Total activity of¹⁴C and³H fell in cholesterol of liver. The results show that plasma accumulation of cholesterol during lecithin infusion derives from both cholesterol pre-existing prior to infusion and from that newly synthesized after the start of infusion and that about one third of this cholesterol of mixed origin is supplied from the liver. The authors speculatively suggest skin as a likely source for most of the remainder, with a small additional contribution from brain.     

Distribution among tissues of intravenously administered sucrose octaoleate

The distribution of [¹⁴C]oleate label in rat tissues in the 6 hr after intravenous administration of sucrose octa-[¹⁴C]oleate (7.5 mg; SuO₈) was compared with that observed after administration of [¹⁴C]triolein. The [¹⁴C]-oleate label, whether injected as triolein emulsion, or as chylomicrons obtained from donor animals, rapidly cleared from the serum; only 10% or less remained in the serum 15 min after injection. Labeled SuO₈ disappeared less rapidly from the serum; about one-third of the dose was present after 15 min, and after 120 min 14% remained. In the liver, there was an initial greater accumulation of fatty acid label when and emulsion of either triolein or SuO₈ was given rather than the chylomicrons. The octaester continued to accumulate in the liver throughout the 6 hr of study, and 78% of the initial dose was present at that time. By contrast, although one-third of the triolein, as of SuO₈, was found in the liver shortly after injection, levels subsequently decreased; at 6 hr, 12% of the label remained associated with that organ. A small portion, up to 8% of the acid label, whether administered as chylomicrons or as a triolein emulsion, was found in the epididymal fat pads. Smaller amounts, usually 1% or less, of the [¹⁴C]oleate label were found in fat pads following the injection of labeled SuO₈. In a separate study, the levels of acid label in the liver and spleen were monitored for 21 days following the intravenous administration of [¹⁴C]SuO₈. There was an initial accumulation of approximately half of the injected lipid label in the liver and one-quarter in the spleen. By day 21, the level in the liver had decreased to one-third of that administered, while the level in spleen remained at one-quarter.     

Mechanisms Underlying Decreased Hepatic Triacylglycerol and Cholesterol by Dietary Bitter Melon Extract in the Rat

In these studies, we focused on finding the mechanism(s) underlying the bitter melon (Momordica charantia L.) methanol fraction (MF)-dependent reduction in the concentration of hepatic triacylglycerol (TAG) and cholesterol in the rat. Rats were fed diets containing low (5 %) fat for 2 weeks (experiment 1), or low (5 %) and high (15 %) fat for a longer period of 8 weeks (experiment 2). MF was supplemented at 1 % level in both experiments. After feeding, rats were sacrificed, and their livers were prepared as slices and hepatocytes, followed by incubation with [1(2)-¹⁴C] acetate or [1-¹⁴C] oleic acid (18:1 n-6). Under these conditions, we found that rats fed diets containing MF, as compared to those without MF, showed: (1) no adverse effects on food intake and growth, (2) a decreased hepatic TAG and total cholesterol, irrespective of the difference in dietary fat level or feeding period, and (3) a decreased incorporation of [1(2)-¹⁴C] acetate and [1-¹⁴C] oleic acid into TAG of liver slices and hepatocytes. MF-supplemented rats also showed no altered incorporation of labeled acetate into cholesterol and cholesterol ester, an increased fecal excretion of neutral steroids, but not of acidic steroids, and an enhanced mRNA abundance of carnitine palmitoylacyltransferase I, which is the rate-limiting enzyme for fatty acid oxidation. These results suggest that dietary MF decreases hepatic TAG synthesis while enhancing fatty acid oxidation, thereby reducing the concentration of hepatic TAG. The liver cholesterol-lowering effect of MF, however, is probably mediated through an increased fecal excretion of neutral steroids, without an effect on cholesterogenesis.     

Measurement of bile acid synthesis in man by release of14CO2 from [26-14C]cholesterol: Comparison to isotope dilution and assessment of optimum reference cholesterol specific activity

Bile acid synthesis can be measured as release of¹⁴CO₂ from [26-¹⁴C]cholesterol divided by cholesterol specific activity, but this method has not been validated in human subjects. We made twelve comparisons of this CO₂ method to standard isotope dilution in six normal subjects and found a mean discrepancy of 6%. Linear regression analysis of one value with respect to the other revealed a correlation coefficient of 0.83 (P<0.001), a Y-intercept close to zero (−4.98) and a slope close to 1 (1.06), suggesting good correspondence between the two methods. To assess the potential for error arising from use of serum cholesterol to estimate specific activity of cholesterol used for bile acid synthesis, we compared synthesis measured using serum free cholesterol specific activity to that measured using bile cholesterol specific activity, which is known to be near isotopic equilibrium with the precursor pool used for bile acid synthesis. Synthesis calculated in these two ways differed by less than 10%. The data indicate that the CO₂ method using either serum or bile cholesterol specific activity provides a valid estimate of bile acid synthesis in man.