Effect of feeding protected cholesterol on ruminant milk fat secretion

Feeding 1–2 g/day of cholesterol protected against ruminal hydrogenation caused a 20–30% drop in the secretion of milk fat by goats and cows. The effect was observed with goats fed conventional rations or with goats and cows fed rations supplemented with protected lipids, but was not observed with cows fed conventional rations, or when unprotected cholesterol and protected β-sitosterol was fed to these animals. The results suggest that this depression in milk fat is due to a decreased uptake of plasma triacylglycerol fatty acids by the mammary gland, induced by dietary cholesterol.     

Glycerides,waxes and sterols in ovaries ofAscaris lumbricoides (Nematoda)

Glycerides (65%), ascaroside esters (33%), waxes (1.5%) and sterols (0.26%) accounted for essentially all the neutral lipids inA. lumbricoides ovaries. Nineteen per cent by weight of the triglycerides contained only long chain fatty acids. Nearly all the remaining triglycerides contained 1 mol volatile acid. Mono- and diglycerides, free fatty acids and triglycerides containing 2 mol volatile acids were present in very small amounts. Mole percentages of glyceride volatile acids were α-methylavaleric (70), α-methylbutyric (23),n-valeric (ca. 7), and traces of acetic, propionic, isobutyric andn-butyric. Mole percentages of the alcoholic component of the waxes were 96% 1-octadecanol, and 4% of its 16, 17 and 19 carbon homologs. The acid components were α-methylbutyric (75 mol%), acetic (16 mol%), propionic andn-valeric (each 4 mol%), and traces ofn-butyric and α-methylvaleric. Sterols (42 wt% cholesterol, 30% cholestanol, 9.5% campestanol, 8.2% stigmastanol, 6.6% β-sitosterol, and 3.8% campesterol) were essentially the same as those found in the whole worm, except that no esterified sterols were present.     

Isolation and identification of cholesterol α-oxide and other minor sterols in human serum

The isolation and identification of cholesterol α-oxide, coprostanol, β-sitosterol, cholest-4-en-3-one and cholesta-4, 6-dien-3-one from human serum are reported. Compounds were isolated by thin layer chromatography and were identified by gas liquid chromatography and gas chromatography-mass spectrometry (GC-MS). Data for standard sterols are also reported. The possible origins of these minor components and the significance of their presence are discussed. Throughout the paper the following nomenclature is used: cholesterol α-oxide, cholesterol-5α, 6α-epoxide; coprostanol, 5β-cholestan-3β-ol; cholestanol, 5α-cholestan-3β-ol; dihydrolanosterol, 5α-lanost-8(9)-en-3β-ol; β-Sitosterol, 24β-ethylcholest-5-en-3β-ol; cholesterol, cholest-5-en-3β-ol; coprostanone, 5β-cholestan-3-one.     

Neutral lipid and fatty acid composition of earthworms (Lumbricus terrestris)

Earthworms (Lumbricus terrestris) were extracted with chloroform-methanol (2∶1) and examined primarily for neutral lipids and fatty acids. TLC showed spots for sterols, hydrocarbons, free fatty acids, phospholipids and pigments but none for glycerides (tri-, di- or mono). Saponification of the crude lipid extract yielded 32% fatty acids, 25% unsaponifiables and 43% unidentified. The lipid contained 3% hydrocarbon and 16% sterols. GLC of the hydrocarbons showed at least 13 components. GLC of the sterol fraction showed peaks corresponding to cholesterol (the major component), γ-sitosterol, β-sitosterol, stigmasterol, campesterol, and ergosterol. GLC showed that at least 38 fatty acids were present, with 18∶1, 18∶2, 18∶0, 20∶1 and 17∶0 predominanting. Abstracted in part from the Ph.D. dissertation of J. Cerbulis, Rutgers, The State University, 1966.     

Effect of 7-methylated bile acids and bile alcohols on cholesterol metabolism in hamsters

The effect of 7-methyl substituted bile acid and bile alcohol analogues on cholesterol metabolism was studied in the hamster. Animals were fed chow plus 0.1% cholesterol supplemented with 0.1% of one of the following steroids: chenodeoxycholic acid, 7-methyl-chenodeoxy-cholic acid, 7β-methyl-24-nor-5β-cholestane-3α,7α,25-triol, cholic acid, 7-methyl-cholic acid, or 7β-methyl-24-nor-5β-cholestane-3α,7α,12α,25-tetrol. Cholesterol absorption was determined from fecal analysis after feeding of radiolabeled cholesterol and β-sitosterol. Of the six compounds studied, chenodeoxycholic acid and 7-methyl-chenodeoxycholic acid decreased intestinal cholesterol absorption (17% and 31% decrease, respectively). Only 7-methyl-chenodeoxycholic acid decreased serum cholesterol concentration (29% decrease), but there were no analogous changes of liver and biliary cholesterol concentration and cholesterol saturation of bile. Total fecal neutral sterol excretion was increased in the groups fed chenodeoxycholic acid and 7-methyl-chenodeoxycholic acid. In addition, the production of coprostanol was increased in both groups. These data suggest that 7-methyl-chenodeoxycholic acid resembles chenodeoxycholic acid in its effect on cholesterol metabolism and may be a potential candidate for further studies of its gallstone-dissolving properties.     

Effect of chitosan feeding on intestinal bile acid metabolism in rats

The effect of chitosan feeding (for 21 days) on intestinal bile acids was studied in male rats. Serum cholesterol levels in rats fed a commercial diet low in cholesterol were decreased by chitosan supplementation. Chitosan inhibited the transformation of cholesterol to coprostanol without causing a qualitative change in fecal excretion of these neutral sterols. Increased fiber consumption did not increase fecal excretion of bile acids, but caused a marked change in fecal bile acid composition. Litcholic acid increased sigificantly, deoxycholic acid increased to a leasser extent, whereas hyodeoxycholic acid and the 6β-isomer and 5-epimeric 3α-hydroxy-6-keto-cholanoic acid(s) decreased. The pH in the cecum and colon became elevated by chitosan feeding which affected the conversion of primary bile acids to secondary bile acids in the large intestine. In the cecum, chitosan feeding increased the concentration of α-,β-, and ω-muricholic acids, and lithocholic acid. However, the levels of hyodeoxycholic acid and its 6β-isomer, of monohydroxy-monoketo-cholanoic acids, and of 3α, 6ξ, 7ξ-trihydroxy-cholanoic acid decreased. The data suggest that chitosan feeding affects the metabolism of intestinal bile acids in rats.     

Differential uptake of cholesterol and plant sterols by rat erythrocytes in vitro

The in vitro uptake of radioactively labeled cholesterol and the plant sterol β-sitosterol has been examined in rat erythrocytes. From mixed micellar solutions containing egg yolk phospholipid and sodium taurocholate, the erythrocytes showed a nonlinear uptake of the two sterols. The uptake leveled off after about 45 min with the attainment of a 1∶1 total sterol-to-phospholipid ratio within the cell membrane, as determined on a mass basis. From soltuions containing egg yolk phospholipid, or purified egg yolk phosphatidylcholine, a preference for cholesterol over the plant sterol was observed, increasing with time from a cholesterol/β-sitosterol uptake ratio of unity (the media ratio) to a maximum of 2 after a 60-min incubation. Correction of the data for nonspecifically bound sterol increased the ratio to a maximum of 5 at the 30-min time point. The increase in the cholesterol/β-sitosterol uptake ratio with time, following an initial nonspecific association, showed that penetration of the plasma membrane by the sterol was required for the selectivity to be expressed. The presence of lysophosphatidylcholine or bovine serum albumin did not exert any noticeable influence over the extent of selectivity of absorption. Replacement of the egg yolk phospholipid with synthetic dipalmitoyl-phosphatidylcholine led to a loss of the sterol selectivity. No evidence was found to support a selective extraction of sterol from the erythrocyte membrane to account for the observed effects, nor was there any sign of a mass accumulation of phospholipid during the incubation. It is suggested that the media phospholipid influences the membrane permeability toward cholesterol and β-sitosterol. Presented in part at the 72nd annual meeting of the American Oil Chemists' Society, New Orleans, LA, May 1981.     

Preferential Efflux of Phytosterols over Cholesterol from Macrophages

Phytosterols are structurally similar to cholesterol. Increased dietary intake of phytosterols effectively lowers LDL-cholesterol. Since phytosterols are incorporated in a growing number of foods and some of the ingested phytosterols reach the circulation, accumulation of phytosterols in foam-cell-prone cells such as macrophages might occur. Therefore we examined the influx and efflux of phytosterols by human THP-1 macrophages. The influx rates of methyl-β-cyclodextrin delivered phytosterols did not significantly differ from that of cholesterol (~3.8 pmol/min per mg cellular protein), neither did the total influx of oxidised LDL delivered phytosterols differ from that of cholesterol. The efflux of β-sitosterol and sitostanol from preloaded THP-1 cells to HDL was more efficient than the efflux of campesterol and cholesterol (rate constants of 0.41 ± 0.04/h, 0.62 ± 0.08/h, 0.23 ± 0.05/h and 0.29 ± 0.03/h, respectively). The efflux of β-sitosterol was not associated with a dominant transfer to ApoA-I, nor did ABCA1 induction-promoted cholesterol efflux to the level observed for β-sitosterol. Our data show that THP-1 macrophages take up phytosterols, but have efficient mechanisms to remove phytosterols from their cellular compartments. Consequently, it is less likely that macrophages preferentially accumulate phytosterols over cholesterol and hence promote foam-cell formation in vivo.     

Phytosterol Ester Constituents Affect Micellar Cholesterol Solubility in Model Bile

Plant sterols and stanols (phytosterols) and their esters are nutraceuticals that lower LDL cholesterol, but the mechanisms of action are not fully understood. We hypothesized that intact esters and simulated hydrolysis products of esters (phytosterols and fatty acids in equal ratios) would differentially affect the solubility of cholesterol in model bile mixed micelles in vitro. Sodium salts of glycine- and taurine-conjugated bile acids were sonicated with phosphatidylcholine and either sterol esters or combinations of sterols and fatty acids to determine the amount of cholesterol solubilized into micelles. Intact sterol esters did not solubilize into micelles, nor did they alter cholesterol solubility. However, free sterols and fatty acids altered cholesterol solubility independently (no interaction effect). Equal contents of cholesterol and either campesterol, stigmasterol, sitosterol, or stigmastanol (sitostanol) decreased cholesterol solubility in micelles by approximately 50% compared to no phytosterol present, with stigmasterol performing slightly better than sitosterol. Phytosterols competed with cholesterol in a dose-dependent manner, demonstrating a 1:1 M substitution of phytosterol for cholesterol in micelle preparations. Unsaturated fatty acids increased the micelle solubility of sterols as compared with saturated or no fatty acids. No differences were detected in the size of the model micelles. Together, these data indicate that stigmasterol combined with saturated fatty acids may be more effective at lowering cholesterol micelle solubility in vivo.     

A diet containing myristoleic plus palmitoleic acids elevates plasma cholesterol in young growing swine

The objective of this study was to test the effect of a novel fatty acid mixture, enriched with myristoleic and palmitoleic acids, on plasma lipoprotein cholesterol concentrations. Weanling pigs were assigned to one of six groups and each group received a diet differing in fatty acid composition. Diets were fed for 35 days and contained 10 g added cornstarch/100 g (to provide baseline data) or 10 g added fatty acids/100 g. For those diets containing added fatty acids, extracted lipids contained 36% myristoleic plus palmitoleic acid combined (14∶1/16∶1 diet), 52% palmitic acid (16∶0 diet), 51% stearic acid (18∶0 diet), 47% oleic acid (18∶1 diet), or 38% linoleic acid (18∶2 diet). Witht the exception of the cornstarch diet, all diets contained approximately 30% myristic acid. There were no significant differences in weight gain across treatment groups (P=0.22). All diets caused a significant increase in triglycerides and in total, low density lipoprotein, high density lipoprotein, and very low density lipoprotein cholesterol. The increase in total plasma cholesterol from pretreatment values was greatest in pigs fed the 14∶1/16∶1 and 18∶1 diets. However, the increase in low density lipoprotein cholesterol from the pretreatment concentration was greatest in the 14∶1/16∶1-fed pigs. Increases in very low density lipoprotein cholesterol above pretreatment concentrations were lowest in 16∶0-fed pigs and greatest in 18∶1-fed pigs. Dietary fatty acids elicited changes in plasma fatty acids which generally were reflective of the diets, although the 18∶0 diet did not alter plasma fatty acid concentrations and the 16∶0 diet increased plasma 16∶0 only at the end of the study. These results demonstrated that the combination of myristoleic plus palmitoleic acids increased plasma cholesterol in young pigs, suggesting that fatty acid chain length, rather than degree of unsaturation, is primarily responsible for the effects of fatty acids on circulating lipoprotein cholesterol concentrations.     

Influence of dietary fat composition on intestinal absorption in the rat

Omega-3 fatty acids influence the function of the intestinal brush border membrane. For example, the omega-3 fatty acid eicosapentaenoic acid (20∶5ω3) has an antiabsorptive effect on jejunal uptake of glucose. This study was undertaken to determine whether the effect of feeding α-linolenic acid (18∶3ω3) or EPA plus docosahexaenoic acid (22∶6ω3) on intestinal absorption of nutrients was influenced by the major source of dietary lipid, hydrogenated beef tallow or safflower oil. Thein vitro intestinal uptake of glucose, fatty acids and cholesterol was examined in rats fed isocaloric diets for 2 weeks: beef tallow, beef tallow + linolenic acid, beef tallow + eicosapentaenoic acid/docosahexaenoic acid, safflower oil, safflower oil + linolenic acid, or safflower oil + eicosapentaenic acid/docosahexaenoic acid. Eicosapentaenoic acid/docosahexaenoic acid reduced jejunal uptake of 10 and 20 mM glucose only when fed with beef tallow, and not when fed with safflower oil. Linolenic acid had no effect on glucose uptake, regardless of whether it was fed with beef tallow or safflower oil. The jejunal uptake a long-chain fatty acids (18∶0, 18∶2ω6, 18∶3ω3, 20∶4ω6, 20∶5ω3 and 22∶6ω3) and cholesterol was lower in salfflower oil than with beef tallow. When eicosapentaenoic acid/docosahexaenoic acid was given with beef tallow (but not with safflower oil), there was lower uptake of 18∶0, 20∶5ω3 and cholesterol. The demonstration of the inhibitory effect of linolenic acid or eicosapentaenoic acid/docosahexaenoic acid on cholesterol uptake required the feeding of a saturated fatty acid diet (beef tallow). These changes in uptake were not explained by differences in the animals’ food intake, body weight gain or intestinal weight. Feeding safflower oil was associated with an approximately 25% increase in the jejunal and ileal mucosal surface area, but this increase was prevented by combining linolenic acid or eicosapentaenoic acid/docosahexaenoic acid with safflower oil. Different inhibitory patterns were observed when mixtures of fatty acids were present together in the incubation medium, rather than in the diet: for example, when 18∶0 was in the incubation medium with 20∶4ω6, the uptake of 20∶4ω6 was reduced, whereas the uptake was unaffected by 18∶2ω6 or 20∶5ω3. Thus, (1) the inhibitory effect of eicosapentaenoic acid/docosahexaenoic acid on jejunal uptake of glucose, fatty acids and cholesterol was influenced by the major dietary lipid, saturated (beef tallow) or polyunsaturated fatty acid (safflower oil); and (2) different omega-3 fatty acids (linolenic acid versus eicosapentaenoic acid/docosahexaenoic acid) have a variable influence on the intestinal absorption of nutrients.     

Mitochondrial metabolism of (D,L)-threo-9, 10-dibromo palmitic acid

Bromination of palmitoleic or palmitelaidic acid proceeds bytrans addition and yields dibrominated products which cannot undergo β-oxidation when incubated with mitochondria isolated from hamster brown adipose tissue. These mitochondria were selected because they have a high capacity for oxidation of C₁₆ fatty acids and because they are readily uncoupled by an excess of free fatty acids of this chain length. The only metabolites which could be recovered from the incubation mixtures were dibromopalmitoylcarnitine and dibromopalmitoyl CoA. Free fatty acid was also recovered. Addition of synthetic carnitine or CoA esters of brominated fatty acids did not interfere with subsequent oxidation of palmitoylcarnitine. Addition of the free brominated fatty acids did significantly increase the rate of oxidation of subsequent additions of palmitoylcarnitine, as did other known synthetic uncouplers. These results are consistent with observations by others that feeding brominated oils leads to brominated fatty acid incorporation into tissue lipids, and indicate why this is so. They also provide a possible explanation for the hepatic damage noted in feeding experiments.     

Cholestanol and plant sterols in pigeon skin

Cholestanol (4.6%) and plant sterols (0.2%) have been demonstrated, for the first time, in avian skin by argentation and gas liquid chromatography. In contrast to results of previous studies with rat and human skin, cholestanol represented a significant amount in the pigeon. The proportion of campesterol was always higher than that of β-sitosterol.     

Sterol lipids in finger millet(Eleusine coracana)

Neutral lipids, glycolipids, and phospholipids (1.3%, 0.25%, and 0.10% of seed weight) were isolated from the total lipids (chloroform-methanol) of finger millet seeds(Eleusine coracana), and four sterol-containing lipids further isolated from neutral and glycolipids by preparative column and thin layer chromatography. On seed weight, these comprised: free sterols (S) 0.091%, sterol esters (SE) 0.013%, sterol glycosides (SG) 0.025%, acyl sterol glycosides (ASG) 0.020%, and total 0.149%. The major fatty acids, totaling 85-90%, were the same in both esterified sterols, but the proportions varied: 16:0, 18:1, and 18:2 comprising 24, 49, and 17% in SE (calculated iodine value 75) and 43, 36, and 7% in ASG (calculated iodine value 46). All four sterol lipids contained 80-84% of β-sitosterol, the remainder being stigmasterol. The only sugar in SG and ASG was D-glucose. It is deduced that the major representative species are: SE, β-sitosterol oleate/palmitate; SG, β-D-glucopyranosyl-(l → 3)-β-sitosterol; and ASG, 6-0-palmitoyl-β-D-glucopyranosyl-(l → 3)-β-sitosterol.     

Effects of cholestanol feeding and cholestyramine treatment on the tissue sterols in the rabbit

Rabbits were fed diets enriched with cholestanol or cholestereol (3.5 g/wk) for 4–12 weeks. During cholestanol feeding, the concentration of cholestanol in blood serum, liver, heart and aorta increased 15–30 times. In serum and liver, the concentration of cholesterol also increased. Cholestanol-fed rabbits developed inflammatory changes in the liver, with proliferation of small bile ducts. Liver tests were only slightly abnormal. Morphological atherosclerosis of the aorta was only occasionally seen in rabbits receiving cholestanol for eight weeks or less. During cholesterol feeding, the amounts of cholesterol in different tissues increased dramatically, most in the aorta. Morphological atherosclerosis in the aorta was found in all rabbits fed cholesterol-enriched diets for more than four weeks. Brain cholestanol was doubled in rabbits fed cholestanol for eight weeks, whereas brain sterols did not change significantly during cholesterol feeding. After an additional regression period with cholestyramine for eight weeks, the increased content of cholestanol in the brain was unchanged in cholestanol-fed rabbits. These observations are discussed in relation to the cholestanolosis of the brain that develops in the rare inherited human disease cerebrotendinous xanthomatosis.     

Utilization of various sterols by lecithin-cholesterol acyltransferase as acyl acceptors

Highly purified lecithin-cholesterol acyltransferase of human plasma was used to study the utilization of various sterols as the acyl acceptor. The esterification of sterols was facilitated by the presence of a 3β-hydroxyl group and thetrans configuration of the A/B rings, as was evident from the lack of acceptor activity of all 3α-hydroxy sterols tested and coprostanol. Cholesterol analogs in which the side chain is modified, such as campesterol, β-sitosterol, desmosterol and stigmasterol, were less effective than cholesterol as acyl acceptors. However, androstan-3β-o1, which completely lacks the side chain, was found to be more active than cholesterol. The transfer of the acyl group to all effective sterols required the presence of the cofactor peptide apolipoprotein A-I.     

Factors affecting the electrical resistivity of soybean oil

The electrical resistivity of soybean oil that had been purified to remove polar constituents was determined, and the effect of measuring conditions and the addition of polar constituents (free fatty acids, phospholipids, monoglyceride, α-to-copherol, β-sitosterol, β-carotene, peroxides, and water) on resistivity was investigated. For reproducible resistivity measurements, voltages in excess of 50 volts and charging times greater than 120 s were necessary. As temperature was increased linearly, the resistivity of the oil decreased logarithmically. For making comparisons, a temperature of 24°C, a potential of 50 volts, and 120 s charging times were chosen. All polar constituents decreased the resistivity of the purified soybean oil, but water, phospholipids, and monoglycerides had the greatest effects. Water increased the resistivity-lowering effects of all other constituents except for free fatty acids, which were affected by water only slightly. The synergistic effect of water was much greater for phospholipids and monoglyceride than for other constituents.     

Studies on fixed oil of seeds ofSida acuta burm

The fixed oil of the seeds ofS. acuta growing around the University of Saugar campus was studied for its physico-chemical characteristics and fatty acid composition. The percentage of fixed oil was found to be higher than, and its composition differed from, that reported earlier. The saponifiable fraction was found to have oleic, linoleic, palmitic, stearic, myristic and plamitoleic acids, whereas the unsaponifiable fraction contained β-amyrin, β-sitosterol and an unknown waxy nonsteroidal substance.     

Effects of different medium-chain fatty acids on intestinal absorption of structured triacylglycerols

To study the effect of the chain length of medium-chain fatty acids on the intestinal absorption of long-chain fatty acids, we examined the lymphatic transport of fat following administration of five purified structured triacylglycerols (STAG) containing different medium-chain fatty acids in the sn-1, 3 positions and long-chain fatty acids in the sn-2 position in a rat model. Significant amounts of medium-chain fatty acids were found in lymph samples after intragastric administration of 1,3-dioctanoyl-2-linoleyl-sn-glycerol (8∶0/18∶2/8∶0), 1,3-didecanoyl-2-linoleyl-sn-glycerol, and 1,3-didodecanoyl-2-linoleyl-sn-glycerol. The accumulated lymphatic transport of medium-chain fatty acids increased with increasing carbon chain length. The recoveries of caprylic acid (8∶0), capric acid (10∶0), and lauric acid (12∶0) were 7.3±0.9, 26.3±2.4, and 81.7±6.9%, respectively. No significant differences were observed for the maximal intestinal absorption of linoleic acid (18∶2n−6) when the chain length of medium-chain fatty acids at the primary positions was varied, and the absorption of 18∶2 and oleic acid (18∶1) from 8∶0/18∶2/8∶0 and 1,3-dioctanoyl-2-oleyl-sn-glycerol was similar. We conclude that the chain length of the medium-chain fatty acids in the primary positions of STAG does not affect the maximal intestinal absorption of long-chain fatty acids in the sn-2 position in the applied rat model, whereas the distribution of fatty acids between the lymphatics and the portal vein reflects the chain length of the fatty acids. Presented in part at the 3rd ISSFAL Conference, Lyon, France, June 1–5, 1998.     

The Spectrum of Plant and Animal Sterols in Different Oil-Derived Intravenous Emulsions

Intravenous lipid constituents have different effects on various biological processes. Some of these effects are protective, while others are potentially adverse. Phytosterols, in particular, seem to be implicated with parenteral nutrition-associated cholestasis. The aim of this study is to determine the amount of plant and animal sterols present in lipid formulations derived from different oil sources. To this end, animal (cholesterol) and plant (β-sitosterol, campesterol, and stigmasterol) sterols in seven different commercially available intravenous lipid emulsions (ILEs) were quantified by capillary gas chromatography after performing a lipid extraction procedure. The two major constituents of the lipid emulsions were cholesterol (range 14–57% of total lipids) and β-sitosterol (range 24–55%), followed by campesterol (range 8–18%) and stigmasterol (range 5–16%). The fish oil-derived formulation was an exception, as it contained only cholesterol. The mean values of the different sterols were statistically different across ILEs (P = 0.0000). A large percentage of pairwise comparisons were also statistically significant (P = 0.000), most notably for cholesterol and stigmasterol (14 out of 21 for both), followed by campesterol (12 out 21) and β-sitosterol (11 out 21). In conclusion, most ILEs combined significant amounts of phytosterols and cholesterol. However, their phytosterols:cholesterol ratios were reversed compared to the normal human diet.