Cholesterol Journal Articles

Record 4421 to 4440

Direct comparison of a dietary portfolio of cholesterol-lowering foods with a statin in hypercholesterolemic participants
Jenkins, D. J., C. W. Kendall, et al. (2005), Am J Clin Nutr 81(2): 380-7.
Abstract: BACKGROUND: 3-Hydroxy-3-methyl-glutaryl-coenzyme A (HMG-CoA) reductase inhibitors reduce serum cholesterol and are increasingly advocated in primary prevention to achieve reductions in LDL cholesterol. Newer dietary approaches combining cholesterol-lowering foods may offer another option, but these approaches have not been compared directly with statins in the same persons. OBJECTIVE: The objective was to compare, in the same subjects, the cholesterol-lowering potential of a dietary portfolio with that of a statin. DESIGN: Thirty-four hyperlipidemic participants underwent all three 1-mo treatments in random order as outpatients: a very-low-saturated-fat diet (control diet), the same diet plus 20 mg lovastatin (statin diet), and a diet high in plant sterols (1.0 g/1000 kcal), soy-protein foods (including soy milks and soy burgers, 21.4 g/1000 kcal), almonds (14 g/1000 kcal), and viscous fibers from oats, barley, psyllium, and the vegetables okra and eggplant (10 g/1000 kcal) (portfolio diets). Fasting blood samples were obtained at 0, 2, and 4 wk. RESULTS: LDL-cholesterol concentrations decreased by 8.5+/-1.9%, 33.3+/-1.9%, and 29.6+/-1.3% after 4 wk of the control, statin, and portfolio diets, respectively. Although the absolute difference between the statin and the portfolio treatments was significant at 4 wk (P=0.013), 9 participants (26%) achieved their lowest LDL-cholesterol concentrations with the portfolio diet. Moreover, the statin (n=27) and the portfolio (n=24) diets did not differ significantly (P=0.288) in their ability to reduce LDL cholesterol below the 3.4-mmol/L primary prevention cutoff. CONCLUSIONS: Dietary combinations may not differ in potency from first-generation statins in achieving current lipid goals for primary prevention. They may, therefore, bridge the treatment gap between current therapeutic diets and newer statins.

Direct correlation between cholesterol synthesis and hepatic secretion of apolipoprotein B-100 in normolipidemic subjects
Watts, G. F., R. Naoumova, et al. (1995), Metabolism 44(8): 1052-7.
Abstract: The regulation of apolipoprotein B-100 (apo B) metabolism in man is not fully understood. In vitro studies suggest a key role for the hepatic availability of cholesterol substrate. We therefore examined whether there was a direct association between plasma mevalonic acid (MVA) concentration (an index of in vivo cholesterol synthesis) and hepatic secretion of very-low-density lipoprotein (VLDL) apo B in eight normolipidemic, healthy adult subjects. Hepatic secretion of VLDL apo B was estimated by endogenous labeling of apo B with an 8-hour primed, constant infusion of 1-13C-leucine. Isotopic enrichment of VLDL apo B was measured by gas chromatography-mass spectrometry (GCMS), from which the fractional secretion rate (FSR) was derived by a modified monoexponential function. Plasma concentration of MVA was measured by gas chromatography-electron-capture mass spectrometry in blood samples taken at 9 AM. The absolute secretion rate (ASR) of VLDL apo B (mean +/- SD) was 9.7 +/- 2.6 mg/kg/d, and MVA concentration was 5.0 +/- 2.5 ng/mL. There was a highly significant positive correlation between ASR of VLDL apoB and plasma MVA (r =.88, P =.004), which persisted after adjusting for apo E phenotype. The findings suggest that in vivo cholesterol synthesis is a determinant of hepatic secretion of apo B in normolipidemic subjects.

Direct determination of lipoprotein(a) cholesterol by ultracentrifugation and agarose gel electrophoresis with enzymatic staining for cholesterol
Nauck, M., K. Winkler, et al. (1995), Clin Chem 41(5): 731-8.
Abstract: Lipoprotein(a) (Lp(a), a low-density lipoprotein (LDL)-like particle, contains in addition to LDL a specific protein component, apolipoprotein(a) apo(a). Conventionally, Lp(a) has been measured by immunological methods that distinguish between Lp(a) and LDL by dealing with apo(a) as an antigen. We describe a new method to determine Lp(a) on the basis of its cholesterol content. Very-low-density lipoproteins were removed from serum by preparative ultracentrifugation at a density of 1.006 kg/L. The infranate was subjected to agarose gel electrophoresis to separate Lp(a) and LDL. Lp(a) cholesterol was then determined by direct enzymatic staining for cholesterol. On electrophoresis of the > 1.006 kg/L (bottom) fraction, Lp(a) migrates to the pre-beta position, regardless of the genetic apo(a) isoform. The interassay CVs of Lp(a) cholesterol determinations ranged from 6.9% to 11.5%, and the results correlated well with the Lp(a) concentrations measured by immunonephelometry (r = 0.937). There was an inverse relation between the molecular mass of the genetically determined apo(a) isoforms and Lp(a) cholesterol concentrations. Patients with angiographically proven coronary artery disease (CAD) had significantly more Lp(a) cholesterol than healthy controls did. The ratio of Lp(a) cholesterol to immunologically determined Lp(a) tended to be lower in CAD patients, suggesting that Lp(a) particles contained less cholesterol than apo(a). In addition, the new method allows determination of LDL cholesterol without contamination by Lp(a).

Direct effect of an acyl-CoA:cholesterol acyltransferase inhibitor, F-1394, on atherosclerosis in apolipoprotein E and low density lipoprotein receptor double knockout mice
Chiwata, T., K. Aragane, et al. (2001), Br J Pharmacol 133(7): 1005-12.
Abstract: The acyl-CoA:cholesterol acyltransferase (ACAT) enzyme is thought to be responsible for foam cell formation and the subsequent progression of atherosclerosis. The apolipoprotein E and low density lipoprotein receptor double knockout (apoE/LDLr-DKO) mouse is an animal model that develops severe hyperlipidaemia and atherosclerosis. Here we have examined the effect of oral administration of an ACAT inhibitor, F-1394, on atherosclerosis in apoE/LDLr-DKO mice fed a regular chow diet. In en face analysis, a dose of 10, 30, or 100 mg kg(-1) day(-1) F-1394 for 10 weeks reduced the extent of lesions visible in the aorta by 24, 28 and 38%, respectively, as detected by staining with oil red O, without affecting serum cholesterol level in these mice. At the highest dose 100 mg kg(-1) day(-1) of F-1394, the reduction was statistically significant. For quantitative analysis of the cellular and non-cellular components comprising the lesions at the aortic sinus, the effects of an oral dose of 100 mg kg(-1) day(-1) F-1394 for 15 weeks were studied. There was a significant reduction (31.9%) in the oil-red O-stained area in cross-sections of the aortic sinus. In addition, the neointimal area, as well as levels of ACAT-1 protein tended to be decreased (15.2 and 25.8%, respectively, not significant). However, the areas containing macrophages, smooth muscle cells, and collagen were not affected by F-1394. In vitro, F-1394 attenuated foam cell formation in mouse peritoneal macrophages. These results indicate that ACAT may be primarily responsible for lipid accumulation in atherosclerotic lesions, and that its inhibition diminishes the lipid deposition via a direct effect on macrophages in the arterial wall.

Direct electrospray tandem mass spectrometry of the unstable hydroperoxy bishemiacetal product derived from cholesterol ozonolysis
Pulfer, M. K., K. Harrison, et al. (2004), J Am Soc Mass Spectrom 15(2): 194-202.
Abstract: Cholesterol is the most abundant neutral lipid in the epithelial lining fluid of the lower airways of the lung also known as pulmonary surfactant and a potential target for reaction with ambient ozone when inspired into the human lung. The isolated double bond of cholesterol has been shown to be susceptible to attack by ozone, but the major reaction product from cholesterol ozonolysis had been remarkably difficult to structurally characterize. Recently, NMR and X-ray crystallography have been used to suggest the formation of a hydroperoxy, hydroxy hemiacetal product, using various derivatives and models of cholesterol to stabilize this chemically reactive product. Electrospray ionization mass spectrometry was used to study the somewhat unstable ozonolysis product of cholesterol which was found to display unique ionization and fragmentation properties when collisionally activated. The electron-deficient carbon atoms of this highly oxygenated product permitted covalent attachment of an acetate anion during negative ion electrospray ionization, leading to the formation of abundant adduct ions at m/z 511. Surprisingly, positive ions were not readily formed. Collision induced dissociation of the adduct anion yielded a major ion at m/z 477, corresponding to the loss of hydrogen peroxide. The most abundant fragment ion following collisional activation was observed at m/z 93, resulting from a complex rearrangement subsequent to the attack of the hydroperoxide anion on the carbon center of the acetate adduct. Based on the interpretation of the tandem mass spectral data, the major cholesterol ozonization product was characterized as a hydroperoxy, hydroxy hemiacetal derivative, which was consistent with the NMR and X-ray crystallographic studies which were carried out on the more stable methyl ether derivative.

Direct evidence for cholesterol crystalline domains in biological membranes: role in human pathobiology
Preston Mason, R., T. N. Tulenko, et al. (2003), Biochim Biophys Acta 1610(2): 198-207.
Abstract: This review will discuss the use of small-angle X-ray diffraction approaches to study the organization of lipids in plasma membranes derived from two distinct mammalian cell types: arterial smooth muscle cells and ocular lens fiber cells. These studies indicate that cholesterol at an elevated concentration can self-associate and form immiscible domains in the plasma membrane, a phenomenon that contributes to both physiologic and pathologic cellular processes, depending on tissue source. In plasma membrane samples isolated from atherosclerotic smooth muscle cells, the formation of sterol-rich domains is associated with loss of normal cell function, including ion transport activity and control of cell replication. Analysis of meridional diffraction patterns from intact and reconstituted plasma membrane samples indicates the presence of an immiscible cholesterol domain with a unit cell periodicity of 34 A, consistent with a cholesterol monohydrate tail-to-tail bilayer, under disease conditions. These cholesterol domains were observed in smooth muscle cells enriched with cholesterol in vitro as well as from cells obtained ex vivo from an animal model of atherosclerosis. By contrast, well-defined cholesterol domains appear to be essential to the normal physiology of fiber cell plasma membranes of the human ocular lens. The organization of cholesterol into separate domains underlies the role of lens fiber cell plasma membranes in maintaining lens transparency. These domains may also interfere with cataractogenic aggregation of soluble lens proteins at the membrane surface. Taken together, these analyses provide examples of both physiologic and pathologic roles that sterol-rich domains may have in mammalian plasma membranes. These findings support a model of the membrane in which cholesterol aggregates into structurally distinct regions that regulate the function of the cell membrane.

Direct evidence for immiscible cholesterol domains in human ocular lens fiber cell plasma membranes
Jacob, R. F., R. J. Cenedella, et al. (1999), J Biol Chem 274(44): 31613-8.
Abstract: The molecular structure of human ocular lens fiber cell plasma membranes was examined directly using small angle x-ray diffraction approaches. A distinct biochemical feature of these membranes is their high relative levels of free cholesterol; the mole ratio of cholesterol to phospholipid (C/P) measured in these membranes ranges from 1 to 4. The organization of cholesterol in this membrane system is not well understood, however. In this study, the structure of plasma membrane samples isolated from nuclear (3.3 C/P) and cortical (2.4 C/P) regions of human lenses was evaluated with x-ray diffraction approaches. Meridional diffraction patterns obtained from the oriented membrane samples demonstrated the presence of an immiscible cholesterol domain with a unit cell periodicity of 34.0 A, consistent with a cholesterol monohydrate bilayer. The dimensions of the sterol-rich domains remained constant over a broad range of temperatures (5-20 degrees C) and relative humidity levels (31-97%). In contrast, dimensions of the surrounding sterol-poor phase were significantly affected by experimental conditions. Similar structural features were observed in membranes reconstituted from fiber cell plasma membrane lipid extracts. The results of this study indicate that the lens fiber cell plasma membrane is a complex structure consisting of separate sterol-rich and -poor domains. Maintenance of these separate domains may be required for the normal function of lens fiber cell plasma membrane and may interfere with the cataractogenic aggregation of soluble lens proteins at the membrane surface.

Direct G protein activation reverses impaired CCK signaling in human gallbladders with cholesterol stones
Yu, P., Q. Chen, et al. (1995), Am J Physiol 269(5 Pt 1): G659-65.
Abstract: Human gallbladders were used to investigate the mechanisms of the impaired contraction induced by cholecystokinin (CCK) associated with cholesterol stones. Single muscle cells were isolated enzymatically with collagenase. Inositol 1,4,5-trisphosphate was measured by high-performance liquid chromatography. Diacylglycerol was assayed by thin-layer chromatography. CCK stimulation showed decreased muscle contraction and production of inositol 1,4,5-trisphosphate and diacylglycerol in gallbladders with cholesterol stones compared with those with pigment stones. Exogenous calmodulin induced maximal contraction of 22.4 +/- 0.5 and 21.0 +/- 0.6% in gallbladders with cholesterol and pigment stones, respectively. Similar findings were observed with a synthetic diacylglycerol analogue. Two G protein activators, aluminum fluoride and guanosine 5'-O-(3-thiotriphosphate), evoked similar responses in these two types of gallbladders, with maximal contractions of 21.3 +/- 0.4 and 23.3 +/- 0.5%, respectively, in those with cholesterol stones and 20.9 +/- 0.8 and 22.6 +/- 0.4%, respectively, in those with pigment stones. These results suggest that receptor-dependent ligands like CCK cannot fully activate the intracellular pathways, which, however, can be fully stimulated by circumventing receptors with G protein activators or second messengers. After G protein activation, the pathways appear to be functionally intact. The defect might then reside in the receptor or in the interaction between receptors and G proteins.

Direct measure of the low-density fractions of serum cholesterol
Purdie, N., L. H. Murphy, et al. (1991), Anal Chem 63(24): 2947-51.
Abstract: Data on total cholesterol (TC) and its distribution among the three solubilizing lipid fractions in human serum have been obtained from three independent laboratories, and binary linear correlations between TC and each of the various fractions are compared. Two sources used the approved double-enzymatic multistep Allain-Trinder reaction with absorption detection. In the third method, which is entirely new, a nonenzymatic chromogenic reaction and circular dichroism (CD) detection were used. TC results from all three sources are in excellent agreement. HDL-C values measured by both enzymatic methods also agree in their correlations with TC but these are quite different from the correlation observed between HDL-C and TC values obtained by the new nonenzymatic procedure. Reasons are given which suggest that the nonenzymatic method is more accurate for the measurement of the low-density lipid fractions and why health risk determinations that are based upon calculated values for this variable should be deemphasized until a more dependable procedure is approved for use.

Direct measurement of HDL cholesterol preferable to precipitation method
Okamoto, Y., S. Tanaka, et al. (1995), Clin Chem 41(12 Pt 1): 1784.

Direct measurement of HDL cholesterol: method eliminating apolipoprotein E-rich particles
Okada, M., H. Matsui, et al. (2001), J Clin Lab Anal 15(4): 223-9.
Abstract: It has been reported that the existing direct method of high density lipoprotein (HDL) cholesterol measures particles enriched with apolipoprotein E (apoE). The aim of our study was to investigate a new analytical protocol to directly measure HDL cholesterol that eliminates apoE-rich particles. The interactions of four lipoproteins (HDL(3), HDL(2), LDL, and VLDL + chylomicron) with surfactants, divalent cations, sugars, and lectins were investigated. By analyzing sera, HDL(3), and HDL(2), we examined the relationships among the measurements obtained by our protocol, a precipitation method using heparin-MnCl(2), and a commercially available kit for this direct method. A significant difference was found between the direct method and the heparin-MnCl(2) method, but not between our protocol and the heparin-MnCl(2) method. Multiple regression analysis showed that the difference between the direct method and the heparin MnCl(2) method is dependent on sources of apoE-rich HDL. In conclusion, our protocol enables a direct measurement of HDL cholesterol that eliminates apoE-rich particles.

Direct measurement of high-density lipoprotein cholesterol by the reflotron assay with no manual precipitation step
Ng, R. H., K. M. Sparks, et al. (1991), Clin Chem 37(3): 435-7.
Abstract: We evaluated the quantification of high-density lipoprotein (HDL) cholesterol in plasma with the Boehringer Mannheim Reflotron reflectance photometric analyzer. The Reflotron is designed for testing in small to medium-size laboratories and physicians' offices. This HDL method does not require a manual precipitation step because the reagent tab contains dextran sulfate (Mr 50,000) and magnesium acetate. It takes 90 s to complete an analysis of 30 microL of plasma. Within-day standard deviations (SDs) were 0.02-0.04 mmol/L (6-16 mg/L). Total SDs over a three-month period were 0.03-0.06 mmol/L (11-23 mg/L). The Reflotron values averaged 0.02 mmol/L (6 mg/L) or 1.3% lower than the Hitachi 737 values; the standard error of the estimate (Sy.x) was 0.07 mmol/L (29 mg/L).

Direct measurement of high-density lipoprotein cholesterol in serum with polyethylene glycol-modified enzymes and sulfated alpha-cyclodextrin
Sugiuchi, H., Y. Uji, et al. (1995), Clin Chem 41(5): 717-23.
Abstract: We have developed an automated method for measuring high-density lipoprotein (HDL)-cholesterol in serum without prior separation, using polyethylene glycol (PEG)-modified enzymes and sulfated alpha-cyclodextrin. When cholesterol esterase and cholesterol oxidase enzymes were modified with PEG, they showed selective catalytic activities towards lipoprotein fractions, with the reactivity increasing in the order: low-density lipoprotein < very-low-density lipoprotein approximately chylomicron < HDL. In the presence of magnesium ions, alpha-cyclodextrin sulfate reduced the reactivity of cholesterol, especially in chylomicrons and very-low-density lipoprotein, without the need for precipitation of those lipoprotein fractions. The combination of PEG-modified enzymes with alpha-cyclodextrin sulfate provided selectivity for the determination of HDL-cholesterol in serum in the presence of a small amount of dextran sulfate without the need for precipitation of lipoprotein aggregates. The results of the HDL-cholesterol assayed in serum by this direct method correlated well with those obtained by precipitation-based methods and also that by an ultracentrifugation method.

Direct measurement of LDL cholesterol
Levinson, S. S. and J. J. Maciejko (1996), Clin Chem 42(5): 780-1.

Direct measurement of low-density-lipoprotein cholesterol is more effective than total cholesterol for the purpose of lipoprotein screening
Okada, M. and R. Ishida (2001), Prev Med 32(3): 224-9.
Abstract: BACKGROUND: We have developed a new method for chemically measuring blood low-density-lipoprotein (LDL) cholesterol. In the present study, we simulated guidelines of the National Cholesterol Education Program (NCEP) using our LDL cholesterol measurements. METHODS: Blood samples were collected from 1,069 individuals (519 males, 550 females) who were referred to our laboratory at Niigata University Hospital for lipoprotein analysis. LDL cholesterol levels were determined according to our assay protocol, which has been published previously. Subjects were categorized by NCEP guidelines and identified "false positives" and "false negatives" on the basis of LDL cholesterol levels measured by our method. RESULTS: The sensitivity of the NCEP guidelines is 87.5% and the specificity is 87.1%, provided we assume that every individual has fewer than two risk factors for coronary heart disease. If we assume that every individual has two or more risk factors, the sensitivity and specificity of the guidelines are 99 and 56.8%, respectively. CONCLUSION: This study presents an opportunity to reevaluate guidelines for routine lipoprotein screening. The chance that individuals with higher LDL cholesterol and lower high-density-lipoprotein cholesterol levels in serum would be missed at initial classification should be zero. The chance that individuals with desirable lipid levels would undergo further lipoprotein analysis should be decreased. Since the new method can be implemented cost-effectively in routine lipoprotein screening, direct measurement of LDL cholesterol could replace total cholesterol.

Direct measurement of serum low-density lipoprotein cholesterol in patients with acute myocardial infarction on admission to the emergency room
Van Dis, F. J., L. M. Keilson, et al. (1996), Am J Cardiol 77(14): 1232-4.
Abstract: Measurement of low-density lipoprotein cholesterol during acute myocardial infarction in nonfasting patients on initial presentation to an emergency room by any of 3 methods (ultracentrifugation, immunoseparation, or the Friedewald estimate), identifies patients eligible for antilipemic interventions. Although slightly less sensitive, the conventional Friedewald estimate of low-density lipoprotein cholesterol levels provides clinicians good correlation with ultracentrifugation.

Direct method for the measurement of low-density lipoprotein cholesterol levels in patients with chronic renal disease: a comparative assessment
Akanji, A. O. (1998), Nephron 79(2): 154-61.
Abstract: BACKGROUND/AIM: This study was performed to comparatively evaluate the results obtained for low-density lipoprotein (LDL) cholesterol concentrations by either a newly described direct method or the Friedewald equation in subjects with and without chronic renal disease. METHODS: Fasting plasma was obtained from a total of 169 subjects, 105 with normal renal function (including 53 hyperlipidaemic) and 64 with chronic renal disease (nephrotic syndrome and/or chronic renal failure; including 40 hyperlipidaemic patients), and analyzed for LDL cholesterol using the Friedewald equation and a direct LDL assay method. RESULTS: The Friedewald equation and the direct LDL cholesterol assay correlated well with each other (r = 0.79-0.90 in all subjects with plasma triglyceride, TG, levels greater than or less than 4.0 mmol/l and with and without chronic renal disease and/or hyperlipidaemia, all p < 0.0001). The values for LDL cholesterol, however, tended to be higher with the direct measurement. This mean difference was trivial in hyperlipidaemic subjects with (8.5%) and without (7.1%) normal renal function (both p < 0.05), but could be clinically significant in those with TG >4.0 mmol/l (mean difference 18%, p < 0.001). Indeed, bias plots confirmed this observation of wider negative bias for Friedewald estimation in these moderately hypertriglyceridaemic subjects. CONCLUSION: For most routine laboratories the options immediately available for assessment of lipid levels are the Friedewald equation or the direct measurement. The Friedewald equation and the direct assay method for LDL cholesterol are about equally good for assessment of the LDL status in patients with chronic renal disease and plasma TG <4.0 mmol/l. Where there are restraints on laboratory budgets, it would appear appropriate that the more expensive direct assay method be restricted to cases in whom plasma TG >4.0 mmol/l or to patients who, for whatever reason, are unable to produce fasting samples.

Direct observation of lipoprotein cholesterol ester degradation in lysosomes
Lusa, S., K. Tanhuanpaa, et al. (1998), Biochem J 332 (Pt 2): 451-7.
Abstract: We have investigated whether pyrene-labelled cholesterol esters (PyrnCEs) (n indicates the number of aliphatic carbons in the pyrene-chain) can be used to observe the degradation of low-density lipoprotein (LDL)-derived cholesterol esters (CEs) in the lysosomes of living cells. To select the optimal substrates, hydrolysis of the PyrnCE species by lysosomal acid lipase (LAL) in detergent/phospholipid micelles was compared. The rate of hydrolysis varied markedly depending on the length of the pyrenyl chain. Pyr10CE was clearly the best substrate, while Pyr4CE was practically unhydrolysed. Pyr10CE and 3Hcholesteryl linoleate, the major CE species in LDL, were hydrolysed equally by LAL when incorporated together into reconstituted LDL (rLDL) particles, thus indicating that Pyr10CE is a reliable reporter of the lysosomal degradation of native CEs. When rLDL particles containing Pyr4CE or Pyr10CE were incubated with fibroblasts, the accumulation of bright intracellular vesicular fluorescence was observed with the former fluorescent derivative, but not with the latter. However, when the cells were treated with chloroquine, an inhibitor of lysosomal hydrolysis, or when cells with defective LAL were employed, Pyr10CE also accumulated in vesicular structures. HPLC analysis of cellular lipid extracts fully supported these imaging results. It is concluded that PyrnCEs can be used to observe degradation of CEs directly in living cells. This should be particularly useful when exploring the mechanisms responsible for the accumulation of lipoprotein-derived CEs in complex systems such as the arterial intima.

Direct observation of the action of cholesterol oxidase in monolayers
Slotte, J. P. (1995), Biochim Biophys Acta 1259(2): 180-6.
Abstract: The oxidation of monolayer cholesterol by cholesterol oxidase has been visualized using monolayer fluorescence microscopy. A direct microscopic visualization was possible because the lateral distribution of a lipid fluorophore, tetramethylrhodamine (TRITC)-labeled phosphatidylethanolamine, was very different in a cholesterol containing monolayer as compared with a cholestenone monolayer. The lipid fluorophore was effectively excluded from the condensed cholesterol phase, but was readily miscible in the cholestenone phase. One could therefore observe the appearance of fluorophore rich cholestenone-domains in the cholesterol monolayer as a result of the cholesterol oxidase catalyzed oxidation reaction. The oxidation experiments were performed at 22 degrees C with a monolayer surface pressure of 5 mN/m (on 50 mM Tris-HCl buffer, containing 140 mM NaCl, pH 7.4). When 40 mU/ml of cholesterol oxidase was injected beneath the monolayer under observation, it appeared that the enzyme penetrated the cholesterol monolayer at random sites and initiated the oxidation reaction. Once the oxidation reaction had commenced, it progressed rapidly and converted the condensed (cholesterol-rich) phase into an expanded (cholestenone-rich) phase. When the oxidation of cholesterol in mixed cholesterol/dimyristoylphosphatidyl-choline monolayers was visualized, it was observed that the enzyme-catalyzed oxidation started from the expanded phases (domains with higher compressibility) and the reaction eventually led to the dissipation of the boundary line between expanded and condensed phases. With time all condensed phases were dissolved and the monolayer became uniformly fluorescent. The association of TRITC-labeled cholesterol oxidase with a non-fluorescent mixed cholesterol/dimyristoylphosphatidylcholine monolayer led to the penetration (or association) of the fluorescent cholesterol oxidase into expanded phases of the mixed monolayers. The monolayer lateral domain morphology was similar whether the fluorescent probe was TRITC-PE or TRITC-labeled enzyme. It is concluded that cholesterol oxidase associated with (or penetrated to some extent into) the expanded phases of a monolayer, and carried out its oxidation reaction in the expanded phase or at the interface between expanded and condensed phases.

Direct testing for low-density lipoprotein cholesterol
Cohen, J. D. (1995), Am J Cardiol 75(12): 831-2.

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