In vivo determination of triglyceride secretion using radioactive glycerol in rats fed different dietary saturated fats

Our objective was to determine the relative rates ofin vivo triglyceride (TG) secretion and the composition of very low density lipoproteins (VLDL) in rats fed different dietary saturated fats. Male Sprague-Dawley rats (150–200 g) were fed diets containing 16% corn oil, or 14% butterfat, 14% beef tallow, 14% olive oil, or 14% coconut oil plus 2% corn oil for 5 wk. Changes in plasma TG specific radioactivity were determined in individual, unanesthetized fasted rats after injection of 100 μCi [2-³H]glycerol. Nonlinear regression analysis using a 2-compartment model was used to determine the fractional rate constant for TG turnover in plasma. The plasma TG pool was 33–40% larger with beef tallow than with corn, olive or coconut oil feeding (p<0.05), and 20% larger with beef tallow than with butterfat feeding. The rate of TG secretion into plasma (mg/min/100 g body weight) was 60% higher in animals fed beef tallow than corn or coconut oil (p<0.05) and 26–33% higher in animals fed beef tallow than olive oil or butterfat. Differences in VLDL composition (% wt) were also noted. Our data suggest that greater TG secretion is the primary factor contributing to the larger TG pool with ingestion of beef tallow relative to butterfat, corn or coconut oil. These results suggest that different dietary saturated fats have unique effects on TG metabolism in rats. Presented in part at the 1990 meeting of the Federation of American Societies for Experimental Biology in Washington, D.C. (see ref. 1).     

Comparison of Supercritical Fluid and Hexane Extraction Methods in Extracting Kenaf (<Emphasis Type="Italic">Hibiscus cannabinus</Emphasis>) Seed Oil Lipids

The objective of this study was to investigate and compare fatty acids, tocopherols and sterols of kenaf seed oil extracted by supercritical carbon dioxide and traditional solvent methods. Fatty acids, tocopherols and sterols were determined in the extracted oils as functions of the pressure (400 bar, 600 bar), temperature (40 °C, 80 °C) and CO₂ flow rate (25 g/min) using a 1-L extraction vessel. Gas chromatography was used to characterize fatty acids and sterols of the obtained oils while tocopherols were quantified by HPLC. No differences were found in the fatty acid compositions of the various oil extracts and the main components were found to be linoleic (38%), oleic (35%), palmitic (20%) and stearic acid (3%). Extraction of tocopherols using high pressure (600 bar/40 °C, 600 bar/80 °C) gave higher total tocopherols (88.20 and 85.57 mg/100 g oil, respectively) when compared with hexane extraction which gave yield of 62.38 mg/100 g oil. Extraction of kenaf seed oil using supercritical fluid extraction at high temperature (80 °C) gave higher amounts of sterols when compared with hexane extraction.     

Particle size effects on supercritical CO<Subscript>2</Subscript> extraction of oil-containing seeds

Rosehip seeds were milled, sieved, and extracted with 26.3 g/g substrate/h of supercritical carbon dioxide (CO₂) at 40°C and 300 bar. The extraction kinetics were characterized by an initial solubility-controlled period (8.78 g oil/kg CO₂ at 40°C and 300 bar), followed by a transition period to a final mass transfer-controlled process. The integral yield of oil approached an asymptotic value that was dependent on the particle size of the substrate: 57.1 g oil/kg dry oil-free substrate (large particles), 171.0 g/kg (medium-size particles), or 391.5 g/kg (small particles). Based on gravimetric determinations and microscopic analysis, our size-classification process segregated seed parts having different oil contents. Particles ≥0.85 mm were mainly composed of tough, lignified testa fragments devoid of oil, whereas particles ≤0.425 mm contained mostly brittle, oil-rich germ fragments. The segregation of seed in fractions with different oil contents may be a common occurrence in supercritical extraction experiments, especially for seeds with thick and/or hard testa and small germ, whose fractions can be separated by sieving.     

In-line and off-line trapping of lipids extracted from chicken liver by supercritical carbon dioxide

This supercritical fluid extraction study determined the retentive properties of neutral alumina sorbent as an in-line trap for lipids in the dynamic state over a pressure range of 490–680 bar and temperatures of 40 and 80°C. Lipids were extracted from a chicken liver matrix using supercritical carbon dioxide over a 40-min period at a flow rate of 3 L/min (expanded gas), then were quantified by high-performance liquid chromatography using an evaporative light-scattering detector. Approximately 30 and 18%, respectively, of the total extracted lipids were trapped on the in-line alumina sorbent bed at 40°C as the operating pressure increased from 490 to 680 bar, while the remaining lipids were trapped off-line after CO₂ decompression. The major lipid classes trapped in-line were fatty acids and cholesterol, whereas only minor amounts of the less polar lipid classes such as sterol esters and triacylglycerols were retained. At 80°C and 680 bar, less than 1.5% of the extracted total lipids was trapped in-line, indicating the lack of adsorptive selectivity for lipids by alumina under these conditions.     

Pressurized liquid extraction of polar and nonpolar lipids in corn and oats with hexane,methylene chloride,isopropanol, and ethanol

Samples of freshly ground corn kernels and freshly ground rolled oats were extracted via pressurized liquid extraction (accelerated solvent extraction) using four different organic solvents [hexane, methylene chloride (also known as dichloromethane), isopropanol, and ethanol] at two temperatures (40 and 100°C). Lipid yields varied from 2.9 to 5.9 wt% for ground corn and from 5.5 to 6.7 wt% for ground oats. With ground corn, more lipid was extracted as solvent polarity was increased, and for each individual solvent, more lipid was extracted at 100°C than at 40°C. With ground oats, the same temperature effects was observed, but the solvent polarity effect was more complex. For both corn and oats, methylene chloride extracted the highest levels of each of the nonpolar lipid classes. In general, for both corn and oats, icnreasing solvent polarity resulted in increasing yields of polar lipids, and for each solvent, more of each lipid class was extracted at 100°C, than at 40°C. Among the lipids in corn extracts, the phytosterols may be the most valuable, and total phytosterols ranged from about 0.6 wt% in the hot ethanol extracts to about 2.1 wt% in the hot hexane and methylene chloride extracts. Total phytosterols in all oat extracts were about 0.1 wt%. Digalactosyldiacylglycerol was the most abundant polar lipid in the oat extracts; its levels ranged from 1.6 wt% in the cold hexane extracts to 4.3 wt% in the hot ethanol extracts.     

The Fractional Extraction of Lipids and Cholesterol from Dried Egg Yolk Using Supercritical Carbon Dioxide

Results from extraction of cholesterol and other lipid components from dried egg yolk using supercritical carbon dioxide at the range of temperature from 40°C to 60°C and pressure between 150 bar und 350 bar for 2.5 hours and 2.7 kg CO₂ consumption is described in this paper. The solubility of lipids and cholesterol increased with the increase of pressure at a constant temperature of 50°C, while at a constant pressure, more cholesterol was removed at 45°C than that at other temperatures. Nearly 60 percent cholesterol was removed at 45°C and 250 bar. Lipids were more efficiently extracted at 60°C than at 40°C at 250 and 350 bar, however, a decrease in the total extracted lipids was observed with the increase in temperature at 150 bar. The removed total lipids from dried egg yolk at 250 bar/55°C was over 80 %.     

Selective distribution of saturated fatty acids into the monoglyceride fraction during enzymatic glycerolysis

Four triglyceride fats and oils (beef tallow, lard, rapeseed oil and soybean oil) were reacted with glycerol while using lipase as the catalyst. For all fats examined, at reaction temperatures above the critical temperature (T_c), the fatty acid compositions of the monoglyceride (MG) and diglyceride (DG) fractions and of the original fat were similar. A relatively low yield of MG was obtained (20–30 wt%). When the reaction was carried out with beef tallow or lard at a temperature below the T_c (40°C), the concentration of saturated fatty acids in the MG fraction was 2 to 4 times greater than that in the DG fraction. Correspondingly, the concentration of unsaturated fatty acids in the DG fraction was more than two times greater than that in the MG fraction. At 5°C, a similar trend was observed for rapeseed oil and soybean oil. Direct analysis of partial glycerides during glycerolysis by high-temperature gas-liquid chromatography showed that below T_c the content of C16 MG increased relatively more than C18 MG. C36 DG and C54 TG were apparently resistant to glycerolysis. Preferential distribution of saturated fatty acids into the MG fraction was accompanied by a high yield of monoglyceride (45–70 wt%) and solidification of the reaction mixture. It is concluded that during glycerolysis below T_c, preferential crystallization occurs for MGs that contain a saturated fatty acid.     

Solvent extraction of the oils of rubber,melon, pumpkin and oilbean seeds

Solvents of differing dielectric constant were used to extract oils from the seeds of: rubber [Hevea brasiliensis (Kunth) Muell. Arg.], melon [Colocynthis vulgaris Schrad], fluted pumpkin [Telfairia occidentalis Hook f.] and oilbean [Pentaclethra macrophylla Benth]. The aim was to examine the effect of solvent polarity on oil yield and oil properties. The oils were extracted under Soxhlet conditions with the following solvents: petroleum benzene (60–80°C), cyclohexane, isopropyl ether, ethyl acetate, tetrahydrofuran, propan-2-ol and acetone. The oils were characterized by acid number, iodine value and color intensity determinations. The oil yields of each seed in different solvents ranged as follows: 58.0–64.4% (pumpkin), 56.1–59.1% (melon), 40.6–48.8% (rubber) and 35.4–43.3% (oilbean). The equilibrium extracting capacity of each solvent was found to depend on two factors, namely, the nature of the oil and the polarity of the solvent. Both factors were found to determine the acid number, iodine value and color intensity of each oil.     

Solid phase enzymatic glycerolysis of beef tallow resulting in a high yield of monoglyceride

A mixture of mono-, di- and triglycerides was obtained when beef tallow was reacted with glycerol using lipase enzyme as a catalyst. The reaction was carried out batchwise in a small vessel with agitation by magnetic stirring. The yield of monoglyceride (MG) was greatly influenced by the reaction temperature—at higher temperatures (48–50°C) a yield of approximately 30% MG was obtained, while at lower temperatures (38–46°C) a yield of approximately 70% MG was obtained. A sharp transition was observed between the high and low yield equilibrium states. The temperature at which this transition occurred is called the critical temperature (T_c) and was found to be 46°C in the case of tallow. During the course of the reaction, when approximately 40% MG had been synthesized, the reaction mixture became solid but the reaction continued until approximately 70% MG had been synthesized. A yield of 70% MG also was obtained with tallow at 42°C when a glycerol/tallow mole ratio ranging from 1.5 to 2.5 was used. The free fatty acid content at equilibrium depended on the water concentration in the glycerol phase and varied from 0.5% to 11.0% when the water content ranged from 0.6% to 12.5%. Above 8% water content, the yield of MG was reduced. Of the commercially available lipases that were investigated, lipase fromPseudomonas fluorescens orChromobacterium viscosum resulted in the highest yield of MG.     

Extraction of lipid-grown bacterial cells by supercritical fluid and organic solvent to obtain pure medium chain-length polyhydroxyalkanoates

A simple two-step process was developed to extract and purify medium chain-length polyhydroxyalkanoates (MCL-PHA) from bacterial cells (Pseudomonas resinovorans) grown on lard and tallow. The process consists of supercritical fluid extraction (SFE) of the lyophilized cells with carbon dioxide to remove lipid impurities, followed by chloroform extraction of the cells to recover the MCL-PHA. SFE conditions were varied as to temperature (40–100°C), pressure (2000–9000 psi), and carbon dioxide flow rate (0.5–1.5 L/min, expanded gas). Lipid material, usually 2–4%, but in some cases as high as 11%, was extracted from the dried cells by SFE. A pressure range (5000–9000 psi, increased stepwise), a temperature of 60°C, and a carbon dioxide flow of 1.5 L/min were routinely used to extract the bacterial cells (4–5 g) after 3 h. Higher flow rates could shorten the extraction time even more. SFE did not extract MCL-PHA from the cells. Yield of MCL-PHA after chloroform extraction at room temperature was a maximum of 42.4% based on dry cell weight. The results show that the two-step process saves time, uses much less organic solvent, and produces a purer MCL-PHA biopolymer than previous extraction and purification methods. A more environmentally friendly clean-up procedure based on SFE and organic solvent recovery was developed to remove contaminating lipid materials from the fermentation biomass, allowing for the recovery of higher purity MCL-PHA that are suitable for more demanding applications.     

Synthesis and Physical Properties of Tallow-Oleic Estolide 2-Ethylhexyl Esters

Tallow-oleic estolide 2-ethylhexyl (2-EH) esters were synthesized in a perchloric acid catalyzed one-pot process from industrial 90% oleic and tallow fatty acids at various ratios, while varying the ratio of tallow and oleic fatty acids, with the esterification process incorporated into an in situ second step to provide a functional fluid. Their viscosities ranged 57–80 cSt at 40 °C and 10.8–14.0 cSt at 100 °C with viscosity index (VI) 169–185. The 100% tallow estolide 2-EH ester had modest low-temperature properties (pour point = −15 °C and cloud point = −14 °C), while the 50:50 mixture of oleic and tallow fatty acids produced an estolide that had better low-temperature properties (pour point = −21 °C and cloud point = −21 °C) without a large negative effect on the oxidative stability. The oxidative stability increased as the amount of saturation increased (rotating pressurized vessel oxidation test (RPVOT) × 165–274 min). The tallow-oleic estolide 2-EH esters have shown remarkably low evaporative losses of only 1% loss compared to a 15–17% loss for commercial materials of similar viscosity grade. Along with expected good biodegradability, these tallow-oleic estolide 2-EH esters had acceptable properties that should provide a specialty niche. Names are necessary to report factually on available data; however, the USDA neither guarantees nor warrants the standard of the product, and the use of the name by USDA implies no approval of the product to the exclusion of others that may also be suitable.     

Supercritical Carbon Dioxide Extraction and Characterization of Argentinean Chia Seed Oil

Extraction of chia seed oil was performed with supercritical carbon dioxide (SC-CO₂). To investigate the effects of pressure and temperature on the oil solubility and yield, two isobaric (250 and 450 bar) and two isothermal (40 and 60 °C) extraction conditions were selected. The global extraction yield of chia oil increased with pressure enhancement, but temperature had a little influence on it. The maximum oil recovery using SC-CO₂ at a mass flow rate of 8 kg/h was 97%, which was obtained at 60 °C, 450 bar for a 138-min extraction. The results showed that solubility changed from 4.8 g oil/kg CO₂ at 60 °C–250 bar to 28.8 g oil/kg CO₂ at 60 °C–450 bar. The final extract obtained by SC-CO₂ under different conditions and Soxhlet extraction contained mainly α-linolenic (64.9–65.6%) and linoleic (19.8–20.3%) acids. SC-CO₂ extraction is an interesting alternative methodology because it is possible to achieve a chia oil yield close to that obtained by conventional extraction with a similar fatty acid composition using an environmentally friendly process.     

Thermodynamic phase behavior of fluoropolymer mixtures with supercritical fluid solvents

Cloud-point experimental data were presented at temperatures up to 260 °C and pressures up to 2,500 bar for poly(vinylidene fluoride) (PVDF) [M_w=180,000 and 275,000] in CO₂, CHF₃, CHClF₂, and dimethyl ether (DME). Also, phase behavior curves were shown for the PVDF [M_w= 180,000] in CO₂ with 1-propanol and 1-pentanol. Poly(vinyl fluoride) does not dissolve in CO₂ even at 215 °C and 2,500 bar, while poly(vinylidene fluoride) does dissolve in CO₂ at 220 °C and pressures of 1,700 bar. To dissolve poly(vinylidene fluoride) in dimethyl ether, pressures in excess of 200 bar and temperatures in excess of 165 °C are needed. The cloud-point behavior between the ternary PVDF [M_w=180,000]-CO₂-DME mixture shows a temperature range of 100–195 °C and pressure range of 350–1,360 bar by adding the 11, 29, and 60 wt% DME in cosolvent. The phase behavior for PVDF [M_w=180,000]-CHF₃-9, 19, and 40 wt% CHClF₂ system shows a temperature at below 200 °C and pressure up to 1,700 bar. The effect of CHClF₂ as a cosolvent for PVDF [Mw=275,000] in CO₂ decreased with the pressure increasing 10, 26, 40 and 61 wt% of cosolvent (CHClF₂) contents. Also, the effect of 15 and 40 wt% DME for the PVDF [M_w=275,000]-CHF₃ mixture shows temperature ranges of 120–206 °C and pressures up to 1,570 bar.     

Analysis of lipids from cooked beef by thin-layer chromatography with flame-ionization detection

Lipids were extracted from cooked ground beef with methylene chloride/methanol (2:1). The lipids were separated on a silicic acid column, into neutral and polar fractions by elution with methylene chloride, followed by methanol. These fractions were analyzed by Iatroscan thin-layer chromatography with flame-ionization detection instrumentation on Chromarods S-III (silica gelcoated quartz rods). Comparison of cooked beef stored for 0, 4 and 7 d at 4°C indicated that storage caused a decrease in total lipids, an increase in neutral lipids and a decrease in polar lipids, specifically in phosphatidylcholine. These changes in the lipid fraction were associated with meat flavor deterioration and an increase in lipid oxidation.     

Supercritical fluid extraction and chromatography of emulsifiers

Selected emulsifiers, which included acetylated monoglycerides, lactylated monoglycerides, hexaglycerol distearate, triglycerol mono/dioleate and decaglycerol decaoleate, were separated with capillary supercritical fluid chromatography on a 25% cyanopropyl stationary phase with a mobile phase of CO₂ at 100–150°C. In general, the density/pressure programs that produced the best separations were those with reduced pressure/density ramp rates, which encompassed the largest possible pressure or density range. Samples of acetylated monoglycerides were placed in a supercritical fluid extraction cell on a glass bead bed and extracted for 15 min at 50°C at 340, 408, 544 and 680 atm with CO₂. At 544 atm, 103.6 ± 1.0% of the emulsifiers was extracted. Acetylated monoglycerides were added during twin-screw extrusion of cornstarch (3% w/w emulsifier/corn starch). The acetylated monoglycerides were extracted from the cornstarch extrudate for 15 and 45 min, at 544 and 646 atm and 50–120°C with 0 and 5% added methanol. The percent acetylated monoglyceride extracted after 45 min at 120°C and 646 atm was 60.0 ± 5.7%, while the amount extracted during a 7.5-h Soxtec extraction ranged from 17.9 to 29.4%, depending upon the solvent used.     

Hydrogenation of gluconolactones in equilibrium with gluconic acid on ruthenium catalyst

Water solutions of glucono-δ-lactone and glucono-γ-lactone in equilibrium with gluconic acid were hydrogenated with molecular hydrogen in the presence of a commercial carbon-supported ruthenium catalyst (5% Ru/C, Engelhard Escat 40). Reactions were conducted batchwise on 20 wt% solutions under 100 bar pressure in the temperature range 80–140°C. Reaction rates were unexpectedly high at moderate temperatures (e.g., 0.58 mol h^-1 g _Ru ^-1 at 100°C) because lactones were probably the reactive species. The selectivity to sorbitol at total conversion was larger than 99% at temperatures lower than 100°C. Kinetic study was done at different temperatures, pressures and concentrations. Modelling of the reaction kinetics showed that the reaction followed a rate law corresponding either to the Langmuir–Hinshelwood mechanism without H₂ dissociation, or to the Horiuti–Polanyi mechanism. This revised version was published online in July 2006 with corrections to the Cover Date.     

Fractionation of non-polar compounds of basil (Ocimum basilicum L.) by supercritical fluid extraction (SFE)

Fractionation by supercritical fluid extraction (SFE) was used to obtain various extracts from Ocimum basilicum L. (sweet basil). The extractions were done at temperature of 40 °C and 50 °C, and at different pressure, which was increased successively from 100 bar (150 bar, 200 bar) to 300 bar. The identification and quantification of the extract compounds was done using gas chromatography–mass spectrometry (GC–MS) and gas chromatography with flame-ionization detector (GC–FID) analyses. The yield of basil SFE extracts varies from 0.138 to 1.008% (w/w). The major components identified in the extracts were linalool, eugenol, α-bergamotene, germacrene D, γ-cadinene, δ-cadinene, β-selinene and spathulenol. The highest extraction yield, in fractionation process, of linalool (167.03 mg per 100 g of basil) was achieved at pressure of 100 bar and temperature 50 °C (solvent density 0.378 kg/m³).     

Viscosities of fatty acids and methylated fatty acids saturated with supercritical carbon dioxide

The viscosities of several types of lipids saturated with supercritical carbon dioxide (SC-CO₂) were measured with a high-pressure capillary viscometer. Oleic acid and linoleic acid were evaluated from 85 to 350 bar at 40 and 60°C. The more SC-CO₂-soluble methylated derivatives of these fatty acids were evaluated from 90 to 170 bar at 40 and 60°C. The complex mixture of anhydrous milk fat (AMF) was evaluated from 100–310 bar at 40°C. The viscosities of the methylated fatty acids saturated with SC-CO₂ decreased between 5 and 10 times when the pressure increased from 1 to 80 bar, followed by a further decrease by a factor of 2 to 3 when the pressure was increased from 80 to 180 bar. The viscosities of the fatty acids and AMF saturated with SC-CO₂ had viscosity reduction similar to the methylated fatty acids between 1 and 80 bar, but they decreased much less between 80 and 350 bar. At constant pressure, the viscosity of the fatty acids and AMF decreased with increasing temperature, whereas the viscosity of the methylated fatty acids increased with increasing temperature. The lipid/SC-CO₂ mixtures were Newtonian, and their viscosities were best interpreted by using the mass concentration of dissolved SC-CO₂ in the lipids and the pure component viscosities.     

Enzymic hydrolysis of animal fats in organic solvents at temperatures below their melting points

Lipase fromCandida rugosa catalyzed the hydrolysis of inedible beef tallow and pork lard (edible and inedible) in the presence of organic solvents at temperatures below the melting point of the fat. Reactions were carried out at 50% substrate with 180 lipase units per gram of fat in a two-liter reactor. In the presence of isooctane (5-10%) beef tallow yielded 94% hydrolysis in 24 hr both at 37° and 31°C. Edible pork lard yielded 97% hydrolysis under these conditions and at temperatures as low as 25°C, while inedible lard gave hydrolysis intermediate between the other two fats.     

Further improvements in the yield of monoglycerides during enzymatic glycerolysis of fats and oils

Three approaches were used in an effort to increase the yield of monoglycerides (MG) during the lipase catalyzed reaction of glycerol with triglyceride fats and oils: i) various commercially available lipases were screened for ability to catalyze MG synthesis; ii) mixtures of lipases were compared with single lipases; and iii) two-step temperature programming was applied during the reaction. Of these, temperature programming was found to be the most effective. With an initial temperature of 42°C for 8–16 hr followed by incubation at 5°C for up to 4 days, a yield of approximately 90 wt% MG was obtained from beef tallow, palm oil and palm stearin. When the second incubation temperature was greater than 5°C, the yield of MG was progressively lower with increasing temperature. In the case of screening of newly available commercial lipase preparations, lipases fromPseudomonas sp. were found to be most effective, giving a yield of approximately 70 wt% MG at 42°C from tallow. Lipases fromGeotrichum candidum, Penicillium camembertii (lipase G) andCandida rugosa were inactive. A mixture of lipases fromPenicillium camembertii andHumicola lanuginosa was found to be more effective than either enzyme alone, giving a yield of approximately 70 wt% MG using beef tallow or palm oil. A mixture ofPenicillium camembertii lipase with eitherPseudomonas fluorescens lipase orMucor miehei lipase was not more effective thanPseudomonas fluorescens orMucor miehei lipase alone.