Cholesteryl Laurate Research Articles

The Role of Molecular Packing in Dictating the Miscibility of Some Cholesteryl n-Alkanoates at Interfaces.

Cholesteryl n-alkanoates of saturated fatty acids and their mixtures are widely studied in different physical states and also due to their significance in biology. Here, we address the miscibility of some homologues of cholesteryl n-alkanoates at interfaces, which are known to exhibit different (cholesteryl octanoate, ChC8, and cholesteryl stearate, ChC18) or the same (cholesteryl nonanoate, ChC9, and cholesteryl laurate, ChC12) molecular packing in bulk. Surface manometry and Brewster angle microscopy studies on ChC8 (cholesteryl-cholesteryl interaction, referred to as m-i packing)/ChC9 (cholesteryl-chain interaction, referred to as m-ii packing) and also on ChC18 (chain-chain interactions, referred to as the crystalline bilayer)/ChC9 mixtures reveal phase separation at the air-water (A-W) interface plausibly due to the difference in the molecular packing. In contrast, ChC12/ChC9 (both m-ii packing) mixtures form a homogeneous phase and exhibit a higher collapse pressure (almost twice) than that of ChC9 indicating higher stability. At the air-solid (A-S) interface, the height profiles extracted from the surface topography images using an atomic force microscope yielded thicknesses of 3.6 ± 0.1 and 5.6 ± 0.1 nm for ChC18/ChC9 mixtures (at 0.66 and 0.5 mole fractions (MF)) corresponding to individual assembly, whereas a uniform thickness of 3.5 ± 0.2 nm is obtained for the case of ChC12/ChC9 mixtures (at 0.2, 0.5, and 0.8 MF) corresponding to m-ii packing. Ellipsometry studies reveal that the desorption temperature increases with the mole fraction of ChC9 and attains a maximum at 406.8 ± 4.8 K for 0.4 MF of ChC9, beyond which it decreases. Raman spectroscopy studies are carried out for ChC12/ChC9 mixtures in the homogeneous phase and in the collapsed state. Here, the dependency of peak positions on different physical states was assessed. Our studies offer new insights into the compatibility of molecular packing influencing the phase behavior and may be of relevance to tear film studies and on the formation of crystals in atherosclerosis.

Self-assembly and molecular packing in cholesteryl esters at interfaces.

To understand the self-assembly and molecular packing in cholesteryl esters relevant to biological processes, we have studied them at the air-water and air-solid interfaces. Our phase and thickness studies employing imaging ellipsometry and atomic force microscopy along with surface manometry show that the molecular packing of cholesteryl esters at interfaces can be related to Craven's model of packing, given for bulk. At the air-water interface, following Craven's model, cholesteryl nonanoate and cholesteryl laurate exhibit a fluidic bilayer phase. Interestingly, we find the fluidic bilayer phase of cholesteryl laurate to be unstable and it switches to a crystalline bilayer phase. However, according to Craven, only cholesteryl esters with longer chain lengths starting from cholesteryl tridecanoate should show the crystalline bilayer phase. The thickness behavior of different phases was also studied by transferring the films onto a silicon substrate by using the Langmuir-Blodgett technique. Texture studies show that cholesterol, cholesteryl acetate, cholesteryl nonanoate, cholesteryl laurate, and cholesteryl myristate exhibit homogeneous films with large size domains, whereas cholesteryl palmitate and cholesteryl stearate exhibit less homogeneous films with smaller size domains. We suggest that such an assembly of molecules can be related to their molecular structures. Simulation studies may confirm such a relation.

Optical and electro-optical characterizations of thermal stability of induced smectic phases

ABSTRACTIn the present work, our investigation is to study the optical and electric properties of the binary mixture of heptyl-oxy-benzilidene-p-n-heptyl-aniline and cholesteryl laurate, which exhibit very interesting liquid crystalline Cho–TGB–SmA–SmC-ReSmA–SmE phases sequentially when the specimen is cooled from its isotropic phase. These phases have been characterized by using microscopic and optical anisotropic technique. Temperature variations of optical anisotropy and X-ray studies have been discussed. An electro-optical measurement has also been discussed.

Optical Studies on Smectic-Isotropic Phase Transition Behavior of Two Thermotropic Liquid Crystals

ABSTRACTThe binary mixture of two compounds, viz., Cholesteryl laurate (CL) and diethyl 4, 4′-azoxy benzoate shows the cholesteric and smectic phases sequentially when the specimen is cooled from its isotropic phase. Refractive indices, birefringence, and optical transmittance have been measured by the optical technique. With the help of measured data, the macroscopic ordered parameter has been discussed. The temperature dependence of these parameters has also been discussed.

Abstract 113: Phospholipid Physical State Determines the Efficiency of Cholesteryl Ester Transfer by Cholesteryl Ester Transfer Protein between Discoidal High Density Lipoproteins

The transfer of cholesteryl ester by recombinant cholesteryl ester transfer protein (CETP) between reconstituted discoidal HDL was studied. Particles contained apolipoprotein A-I, unsaturated palmitoyllinoleoylphosphatidylcholine (PLPC), palmitoyloleoylphosphatidylcholine (POPC) or saturated dipalmitoylphosphatidylcholine (DPPC) and cholesteryl ester up to 3 mol% as cholesteryl 1-pyrenedecanoate (CPD) or cholesteryl laurate (CL) in donor and acceptor rHDL, respectively. The concentration dependencies of excimerization upward deviated from a linear dependence, being more curvilinear for DPPC. Separately for DPPC complexes, monomer fluorescence compared to excimer was more efficiently quenched by acrylamide. The nonradiative energy transfer between apoA-I tryptophan residues and CPD molecules was more efficient for PLPC compared to DPPC complexes. Data fulfilled the quenching sphere-of-action model, the sphere with adjacent probe molecules being larger and more distant from apoA-I tryptophan residues in DPPC complexes. CETP-catalyzed cholesteryl ester exchange between donor and acceptor HDL followed by probe excimerization was characterized by a heterogeneous kinetics; the fast exchanging CPD pool was much higher in a case of POPC compared to DPPC complexes. Probe fraction accessible to CETP increased with temperature, suggesting a more homogeneous probe distribution. The noncompetitive inhibition of probe transfer by acceptor particles seems to satisfy the zero off-rate compatible with probe tunneling. The values of V max (0.063 μM·min -1 ) and k cat (0.42 s -1 ) together with a similarity of K m (0.9 μM CPD) and K I (2.8 μM CL) values for POPC-containing rHDL suggest the efficient cholesteryl ester transfer between nascent HDL with unsaturated phosphatidylcholine in vivo opposite to much less efficiency with HDL with saturated phosphatidylcholine. The physical state of phospholipid matrix may underlie CETP activity with discoidal HDL through self-association and location of cholesteryl ester in the bilayer, the availability of cholesteryl ester to cholesterol-binding site in apoA-I structure and the binding of apoA-I-interacting cholesteryl ester to CETP. We are grateful to Dr. A. Tall for providing the recombinant CETP.

Cholesteryl ester diffusion, location and self-association constraints determine CETP activity with discoidal HDL: Excimer probe study

The transfer of cholesteryl ester by recombinant cholesteryl ester transfer protein (CETP) between reconstituted discoidal high-density lipoprotein (rHDL) was studied. Particles contained apolipoprotein A-I, unsaturated POPC or saturated DPPC and cholesteryl ester as cholesteryl 1-pyrenedecanoate (CPD) or cholesteryl laurate (CL) in donor and acceptor rHDL, respectively. Probe dynamics fulfilled the quenching sphere-of-action model. The cholesteryl ester exchange between donor and acceptor particles was characterized by a heterogeneous kinetics; the fast exchanging CPD pool was much higher in a case of POPC compared to DPPC complexes. Probe fraction accessible to CETP increased with temperature, suggesting a more homogeneous probe distribution. Noncompetitive inhibition of probe transfer by acceptor particles was observed. The values of Vmax (0.063μMmin−1) and catalytic rate constant kcat (0.42s−1) together with a similarity of Km (0.9μM CPD) and KI (2.8μM CL) values for POPC-containing rHDL suggest the efficient cholesteryl ester transfer between nascent HDL with unsaturated phosphatidylcholine in vivo. The phospholipid matrix in discoidal HDL may underlie CETP activity through the self-association, diffusivity and location of cholesteryl ester in the bilayer, the accessibility of cholesteryl ester to cholesterol-binding site in apoA-I structure and the binding of cholesteryl ester, positionable by apoA-I, to CETP.

Self-assembled monolayers of cholesterol and cholesteryl esters on graphite.

The molecular arrangements of self-assembled monolayers (SAMs) of cholesterol, cholesteryl laurate, and cholesteryl stearate adsorbed on a graphite surface were studied using scanning tunneling microscopy (STM) at the liquid-solid interface. The STM images of the SAMs showed two-dimensional periodic arrays of bright regions that corresponded to the sterol rings. However, individual sterol rings could not be observed in the bright regions in the STM images of the cholesterol monolayers. Nevertheless, by comparing the STM images and the crystallographic data, it is concluded that the cholesterol molecules are arranged in pairs oriented head-to-head owing to the hydrogen bonds between the hydroxyl groups. These dimers, in turn, are oriented parallel to each other, owing to the interactions between the sterol rings. The STM images of cholesteryl ester monolayers had molecular resolution and showed pairs of cholesteryl ester molecules oriented in an antiparallel manner, with their fatty acid chains located in the central regions. Furthermore, the fatty acid chains of cholesteryl stearate were observed to be oriented in the (1120) zigzag direction of the graphite lattice, whereas those of cholesteryl laurate were oriented in the (1010) armchair direction. These observations reveal that the interactions between the fatty acid chains affect the structure of the SAMs. The molecular arrangements also depend on the lengths of the fatty acid chains of the cholesterol esters and hence on the interactions between the alkyl chains and the graphite surface. The self-assembly at the liquid-solid interface is therefore controlled by the interactions between sterol rings, between alkyl chains, and between alkyl chains and the substrate.

High-temperature GC-MS-based serum cholesterol signatures may reveal sex differences in vasospastic angina

Alterations of cholesterol metabolism are responsible for vasospastic angina and atherosclerosis. To comprehensively evaluate cholesterol metabolism, 18 sterols, including cholesterol, 6 cholesteryl esters (CEs), 3 cholesterol precursors, and 8 hydroxycholesterols (OHCs), were simultaneously analyzed using hybrid solid-phase extraction (SPE) purification coupled to high-temperature gas chromatography-mass spectrometry (HTGC-MS). Methanol-based hybrid SPE increased the selective extraction, and HTGC resulted in a good chromatographic resolution for the separation of lipophilic compounds. The limits of quantification of cholesterol and CEs ranged from 0.2 to 10.0 μg/ml, while OHCs and cholesterol precursors ranged from 0.01 to 0.10 μg/ml. Linearity as the correlation coefficient was higher than 0.99 with the exception of cholesteryl laurate, myristate, oleate, and linoleate (r2 > 0.98). The precision (% coefficient of variation) and accuracy (% bias) ranged from 1.1 to 9.8% and from 75.9 to 125.1%, respectively. The overall recoveries of CEs ranged from 26.1 to 64.0%, and the recoveries of other sterols ranged from 83.8 to 129.3%. The cholesterol signatures showed sex differences in patients with vasospastic angina and may associate with 24-reductases. This technique can be useful for making clinical diagnoses and for an increased understanding of the pathophysiology of vasospastic angina.

Characterization of Codon-Optimized Recombinant Candida rugosa Lipase 5 (LIP5)

Recombinant Candida rugosa lipase 5 (LIP5) has been functionally expressed along with other isoforms in our laboratory. However, the characterization and codon optimization of LIP5 have not been done. In this work, we characterized, codon-optimized and compared LIP5 with commercial lipase. LIP5 activity on hydrolysis of p-nitrophenyl (p-NP) butyrate was optimal at 55 °C as compared with 37 °C of the commercial lipase. Several assays were also performed to determine the substrate specificity of LIP5. p-NP butyrate (C(4)), butyryl-CoA (C(4)), cholesteryl laurate (C(12)), and N-carbobenzoxy-l-tyrosine-p-nitrophenyl ester (l-NBTNPE) were found as preferred substrates of LIP5. Interestingly, LIP5 specificity on hydrolysis of amino acid-derivative substrates was shown to be the highest among any lipase isoforms, but it had very weak preference on hydrolyzing triacylglycerol substrates. LIP5 also displays a pH-dependent maximum activity of a lipase but an esterase substrate preference in general. The characterization of LIP5 along with that of LIP1-LIP4 previously identified shows that each lipase isoform has a distinct substrate preference and catalytic activity.

THE INFLUENCE OF NEW D-SECO-ESTRONE DERIVATIVES ON THE BEHAVIOR OF THE CHOLESTERIC LIQUID CRYSTALS BINARY MIXTURES

We have studied the influence of the new chiral non-mesogenic seco-estrone derivatives 1-7, synthesized in several synthetic steps starting from estrone, onto the physical characteristics of the binary mixtures of cholesteric liquid crystals. We have examined the phase transitions of the mixture of cholesteryl laurate and cholesteryl enantate with the added chiral non-mesogenic additives 1-7 (45%-45%-10%; Mix.1-Mix.7, respectively). A considerable shift of the I → Ch phase transition temperature, as well as of the temperature of the SmA * phase formation, was observed. X-ray diffraction data enabled the determination of the molecular structures of the compounds 3, 5, 6 and 7, the crystal lattice parameters of the new chiral additives and their molecular length (ℓ) and width (d*), as well as some parameters of the mixtures Mix.1-Mix.6: the thickness of smectic layers and the longitudinal spacing distance in the cholesteric phase (d) and the average distance between the long axes of neighbouring molecules (D). Using molecular-mechanic calculations, on the basis of solved molecular structures, the most probably conformation of the other compounds (1, 2 and 4) used in this work is determined. We have also investigated the influence of these chiral additives onto the pitch of the cholesteric helix of a binary mixture of cholesteric liquid crystals.

Monolayers and 3D Films of Cholesteryl Derivatives at the Air−Water Interface

Cholesterol and its derivatives have drawn much attention because of their relevance to biology. They have also been studied at the air−water (A−W) interface. Here, we have systematically investigated the two-dimensional (2D) and three-dimensional (3D) phases of cholesteryl derivatives at the A−W interface employing surface manometry, epifluorescence, Brewster-angle and reflection microscopy techniques. We found that the short-chain ester cholesteryl acetate forms a stable monolayer. The higher homologues cholesteryl heptanoate and cholesteryl octanoate do not form monolayers; they yield 3D crystallites at very large area per molecule. Interestingly, we found cholesteryl nonanoate to be in the crossover regime in the homologous series of cholesteryl esters. It spontaneously forms a fluidlike bilayer at the A−W interface. The higher homologue cholesteryl laurate forms an unstable bilayer phase. The long-chain esters cholesteryl myristate, cholesteryl palmitate, and cholesteryl stearate exhibit crystalline ...

Tricarbocyanine Cholesteryl Laurates Labeled LDL: New Near Infrared Fluorescent Probes (NIRFs) for Monitoring Tumors and Gene Therapy of Familial hypercholesterolemia.

AbstractFor Abstract see ChemInform Abstract in Full Text.

Multiple mutagenesis of non-universal serine codons of the Candida rugosa LIP2 gene and biochemical characterization of purified recombinant LIP2 lipase overexpressed in Pichia pastoris.

The 17 non-universal serine codons (CTG) in the Candida rugosa LIP2 gene have been converted into universal serine codons (TCT) by overlap extension PCR-based multiple site-directed mutagenesis. An active recombinant LIP2 lipase was overexpressed in Pichia pastoris and secreted into the culture medium. The recombinant LIP2 showed distinguishing catalytic activities when compared with recombinant LIP4 and commercial C. rugosa lipase. The purified enzyme showed optimum activity at pH 7 and a broad temperature optimum in the range 30-50 degrees C. The enzyme retained 80% of residual activity after being heated at 70 degrees C for 10 min. Recombinant LIP2 demonstrated high esterase activity towards long-chain (C12-C16) p-nitrophenyl esters. Tributyrin was the preferred substrate among all triacylglycerols tested for lipolysis. Among cholesteryl esters, LIP2 showed highest lipolytic activity towards cholesteryl laurate. The esterification of myristic acid with alcohols of various chain lengths showed that the long-chain n-octadecanol (C18) was the preferred substrate. In contrast, the esterification of n-propanol with fatty acids of various chain lengths showed that the short-chain butyric acid was the best substrate. From comparative modelling analysis, it appears that several amino acid substitutions resulting in greater hydrophobicity in the substrate-binding site might play an important role in the substrate specificity of LIP2.

Tricarbocyanine cholesteryl laurates labeled LDL: new near infrared fluorescent probes (NIRFs) for monitoring tumors and gene therapy of familial hypercholesterolemia

For monitoring low-density lipoprotein receptors (LDLr) in tumors and in livers of patients with familial hypercholesterolemia (FH) treated with gene therapy, a series of tricarbocyanine cholesteryl laurates were synthesized with the cholesteryl laurate moiety serving as the lipid-chelating anchor for low-density lipoprotein (LDL). One of these conjugates, TCL17, was successfully used to label LDL to give a new NIRF, TCL17-LDL. Ex vivo biological studies on an LDLr overexpressing tumor model, human hepatoblastoma G 2 (HepG 2), confirmed that this NIRF were internalized selectively by the tumor and detected with high sensitivity by a low-temperature 3-D redox scanner.

Analysis of steryl esters in cocoa butter by on-line liquid chromatography–gas chromatography

On-line liquid chromatography–gas chromatography (LC–GC) has been applied to the analysis of steryl esters in cocoa butter. Separation of the steryl esters was achieved after on-line transfer to capillary GC. HPLC removes the large amount of triglycerides and pre-separates the components of interest, thus avoiding time-consuming sample preparation prior to GC analysis. The identities of the compounds were confirmed by GC–MS investigation of the collected HPLC fraction and by comparison of the mass spectra (chemical ionization using ammonia as ionization gas) to those of synthesized reference compounds. Using cholesteryl laurate as internal standard, steryl esters were quantified in commercial cocoa butter samples, the detection limit being 3 mg/kg and the quantification limit 10 mg/kg, respectively. Only slight differences in percentage distributions of steryl esters depending on the geographical origin of the material were observed. The patterns were shown to remain unchanged after deodorization. The method described might be a valuable tool for authenticity assessment of cocoa butter.

Cholesteryl esters on the body surfaces of the camel tick, Hyalomma dromedarii (Koch, 1844) and the brown dog tick, Rhipicephalus sanguineus (Latreille, 1806)

Cholesteryl esters were found to constitute a major component of the lipids coating the body cuticle of females of the camel tick, Hyalomma dromedarii and the brown dog tick, Rhipicephalus sanguineus. One or more cholesteryl esters, alone or in combination, have been shown to serve as the mounting sex pheromone of several species of ixodid ticks. Consequently, knowledge of these compounds is important for an understanding of the mating behavior of these ticks. Based on thin layer chromatography, cholesterol and cholesteryl esters were the most abundant neutral lipids found on the body surfaces of fed females of these two species. Analysis using HPLC demonstrated significant quantities of the following compounds, tentatively identified as cholesteryl esters (expressed in micrograms per female equivalent), in H. dromedarii: Cholesteryl acetate 18.2; cholesteryl laurate, 6.8; cholesteryl linoleate, 24.8; cholesteryl oleate, 12.9; cholesteryl palmitate, 0.3; and cholesteryl stearate 1.7. In contrast, the same method revealed only 3 cholesteryl esters in extracts of females of R. sanguineus: Cholesteryl acetate, 2.0; cholesteryl linoleate, 8.5; and cholesteryl oleate, 3.0. In both species, two unidentified peaks, with the spectral characteristics of cholesteryl esters, were also observed. Identification of the cholesteryl esters was confirmed: by (1) positive bioassay results with conspecific (H. dromedarii) males and heterospecific (Dermacentor variabilis) males; (2) similarity of ultraviolet spectra between identified sample peaks and authentic standards; and (3) demonstration of cholesterol and the corresponding free fatty acid following enzymatic digestion of each of the HPLC-separated fractions containing the different cholesteryl esters. Comparisons with the cholesteryl ester composition of the mounting sex pheromone of other metastriate Ixodidae are discussed. These findings, along with studies reported previously, suggest that differences in the mounting sex pheromones of ixodid ticks are an important factor in minimizing heterospecific matings in nature.

Supercritical fluid chromatographic determination of cholesterol and cholesteryl esters in serum on ODS-silica gel column

Cholesterol and cholesteryl esters in human serum were determined by supercritical fluid chromatography on an inert ODS-silica gel column using supercritical carbon dioxide as a mobile phase without a modifier. Chromatograms were obtained by monitoring the eluent simultaneously with an FID and UV detector at the wavelength of 190 nm. The retention behavior of cholesterol and cholesteryl esters was investigated in terms of the density of CO2 mobile phase. The separation mode was found to be reversed phase, as in liquid chromatography. The amounts of cholesterol and cholesteryl esters extracted from human serum reference material (NIST SRM 909) were determined individually using cholesteryl laurate as an internal standard to give good agreement of total cholesterol with the value certified by NIST.

Molecular interactions in binary solids: crystal structure of a cholesteryl ester solid solution.

The x-ray crystal structure analysis of a cholesteryl ester solid solution, cholesteryl undecanoate/cholesteryl laurate in a molar ratio 0.52/0.48, is described. The unit cell is monoclinic with a = 13.005(2) A, b = 9.005(1) A, c = 31.421(3) A, and beta = 90.82(1) degrees and the space group P2(1) with Z = 4 (two molecules per asymmetric unit). Thus, the d001 spacing is almost the value predicted by Vegard's law from the values for the pure compounds. The room-temperature crystal structure is very much like that of cholesteryl laurate monolayer I packing, in the form where the esterified fatty acid chains are fully extended, with no salient sign of conformational disorder seen in this binary solid. The final R factor for 6571 observed reflections is 0.097.

Effect of cholesteryl additive on poly(ethyl methacrylated)–poly(vinyl acetate) blend

AbstractThree different compositions (9 : 1, 7 : 3, and 1 : 1) of poly(ethyl methacrylate) (PEMA) and poly(vinyl acetate) (PVAc) are chosen to check the miscibility of a polymer pair. The 9 : 1 and 7 : 3 PEMA‐PVAc blends are immiscible and have shown two distinct loss peaks in dynamic mechanical analysis (DMA) studies, while 1 : 1 PEMA‐PVAc has shown a single loss peak. Immiscible composition 7 : 3 PEMA‐PVAc is selected to study the effect of cholesteryl additives on the miscibility of the polymers. The additives chosen for the present studies are cholesteryl chloride, cholesteryl caprylate, and cholesteryl laurate. The DMA studies showed that all the additives caused a merging of the two loss peaks into one. X‐ray diffraction (XRD), scanning electron microscopy (SEM), and Fourier transform infrared (FTIR) studies of the blend and blend with additive samples are utilized to get further information. The FTIR studies do not show any detectable change. The SEM micrographs of the blend and cholesteryl additive systems show a single‐phase ordered structure, and XRD data studies indicate the presence of small crystallites.

Dielectric and Viscoelastic Behavior of Poly-(n-Butyl Methacrylate) Containing Cholesteric Compounds

Abstract Dielectric and dynamic mechanical behavior of poly(n-butyl methacrylate) (PBMA) containing small concentrations of cholesteryl chloride (ChCl), cholesteryl caprylate (ChCp), and cholesteryl laurate (ChL) have been studied in the temperature range -20 to 80°C. Tan αmax (dielectric and mechanical) in all systems containing an additive shifts to lower temperature compared to that in pure PBMA. In the glassy region, the dynamic storage modulus E′ for all the PBMA plus additive systems is higher than that for pure PBMA. This behavior is similar to the β-peak suppression and the increase in tensile strength of poly(vinyl chloride) on addition of a small concentration of plasticizer. Using the WLF equation with C1 and C2 obtained from the data on pure PBMA, the reference temperature To for PBMA plus additive systems are determined for the best fit of the experimental points to the WLF curve and are in agreement with measured glass-transition temperatures. With respect to To of pure PBMA, the To of PBMA ...