Lipid monitoring system

On the other hand, an increase in the flow rate of the enzyme solution in our lipid monitoring system decreases the peak intensity as well as the reaction time, since the lipid components in a post column effluent are diluted with the enzyme solution, and the reaction time is inversely proportional to the sum of the flow rate of the eluent and that of the enzyme solution. Thus, a slower flow rate of the enzyme solution is better provided the concentration of enzyme reagent is sufficient for complete reaction in the flow diagram. 

Reproducibility of our HPLC method for lipoprotein analysis is excellent. The elution volume Is reproducible within 0.01 ml for each lipoprotein peak in the case of the individual lipoproteins or serum lipid monitoring system. Reproducibility of the concentration of lipid in each lipoprotein class obtained as described in Section 5 is very high. The standard deviation from the mean value for each lipoprotein fraction was 0.1 to 0.4 mg/dl, and the coefficient of variation (Cv) is less than 2 % for the concentration of PL as presented in Table 3. 

The ability of flavonoids to enhance the resistance to oxidation and to terminate free-radical chain reactions in lipophilic systems can be monitored using low-density lipoproteins (LDL) as a model (Rice-Evans et al., 1996). The LDL oxidation is initiated either by copper or by a peroxyl radical [2,2 azobis(2-amidinopropane hydrochloride) (AAPH)] (Abuja et al., 1998). Hexanal liberated from the decomposition of oxidized n-6 polysaturated fatty acids in LDL may be determined by static headspace gas chromatography (Frankel and Meyer, 1998). Also, bleaching of P-carotene (Velioglu et al., 1998 Fukumoto and Mazza, 2000) and the tracing by HPLC (Fukumoto and Mazza, 2000) of malonaldehyde formed in lipid emulsion systems in the presence of iron (Tsuda et al., 1994) have been used to measure antioxidants in lipophilic systems. 

IV and V) according to the definition by Fredrickson et al. (40). The elution patterns of three lipid components using the G5000PW+G5000PW column system present the characteristic feature for each type as shown in Fig. 25. Thus, these HPLC patterns obtained from whole serum in our lipid monitoring technique are very useful for diagnosis of the type for hyperlipidemia. 

In transfection experiments, lipid L2 and DOTAP were used to form lipoplexes and their ability to transfect HepG2 cells were assessed. The lipoplexes were formed around expression plasmids encoding firefly luciferase and a strong promoter to monitor transfection efficiency. Lipoplexes were incubated with cells for 4 h and luciferase luminescence was quantitated. DNA/L2 complexes showed low luciferase expression however, mixtures of L2 and DOPE complexed with DNA showed improvement and matched that of DOTAP. Further optimization of the lipid/DNA ratio yielded complexes with similar transfection efficiency of other cationic lipid delivery systems. To address the problem of serum proteins degrading lipoplexes, the experiment was repeated for L2 with cells complimented with 25% fetal calf serum. After 24 h, luciferase expression was monitored and it was found that all fiuorinated lipoplexes retained or increased their transfection efficiency, compared to DOTAP whose performance saw a 30-40% reduction. 

The EO of Crithmum maritimum (=Cachrys maritima, Apiaceae, rock samphire) comprises limonene and y-terpinene with an amount of 22.3% and 22.9%, respectively, as the major components. Two different test methods (TBA assay and a micellar model system where the antioxidative activity in different stages of the oxidative process of the lipid matrix was monitored) were used. Both assays explain the very high activity of this EO. In the TBA assay BHT and a-tocopherol were used as positive standards and the oil showed a better capacity than those substances. Comparable results were obtained by the micellar method system where the EO acts as a protector of the oxidation of linoleic acid and inhibits the formation of conjugated dienes (Ruberto et al., 2000). The modification of LDL by an oxidative process for instance can lead to atherosclerosis. Natural antioxidants such as P-carotene, ascorbic acid, a-tocopherol, EOs, and so on are able to protect LDL against this oxidative modification. y-Terpinene proved itself to be the strongest inhibitor of all used authentic compounds for the formation of TBARS in the Cu -induced lipid oxidation system (Grassmann et al, 2003). So, the addition of y-terpinene to food can possibly stop the oxidative modification of LDL and reduce the atherosclerosis risk. 

Cholesterol is abundant in many membranes of eukaryotic cells, the total percentage reaching up to 50% of the total lipid content. The behavior of lipid-cholesterol mixtures has therefore attracted much attention Despite the vast amount of publications on the thennotropic behavior of phospholipid-cholesterol mixtures, the macroscopic description of lipid-cholesterol systems using phase diagrams is still very much debated, because different analytical methods such as NMR-, ESR-, fluorescence- and FT-IR-spectroscopy monitor changes in physicochemical behavior of lipid-cholesterol mixtures as a function of composition and temperature which cannot easily reconciled with observations using DSC [61-72]. 

A method which uses supercritical fluid/solid phase extraction/supercritical fluid chromatography (SE/SPE/SEC) has been developed for the analysis of trace constituents in complex matrices (67). By using this technique, extraction and clean-up are accomplished in one step using unmodified SC CO2. This step is monitored by a photodiode-array detector which allows fractionation. Eigure 10.14 shows a schematic representation of the SE/SPE/SEC set-up. This system allowed selective retention of the sample matrices while eluting and depositing the analytes of interest in the cryogenic trap. Application to the analysis of pesticides from lipid sample matrices have been reported. In this case, the lipids were completely separated from the pesticides. 

Whatever the technique used, it is important to note that (i) only an equivalent viscosity can be determined, (ii) the response of a probe may be different in solvents of the same viscosity but of different chemical nature and structure, (iii) the measured equivalent viscosity often depends on the probe and on the fluorescence technique. Nevertheless, the relative variations of the diffusion coefficient resulting from an external perturbation are generally much less dependent on the technique and on the nature of the probe. Therefore, the fluorescence techniques are very valuable in monitoring changes in fluidity upon an external perturbation such as temperature, pressure and addition of compounds (e.g. cholesterol added to lipid vesicles alcohols and oil added to micellar systems). 

The formation of polar metabolites from nonpolar materials may actually facilitate monitoring programs—in many cases the polar chemicals are highly concentrated in certain body fluids such as bile and urine. On the other hand, materials such as certain cyclodienes and polychlorinated biphenyls, which are very lipid soluble and resistant to metabolism, may accumulate and these chemicals may persist in the environment and may be transferred via the food chain to man. There is also interest in these biotransformation processes in lower organisms since the simplicity of these systems may lead to a better understanding of the phylogenetic development of xenobiotic metabolism. 

The major difficulty in analyzing OPPs in fatty samples has to do with the wide polarity range for both pesticides and lipids present in the matrix. Normal-phase HPLC is an adequate technique for cleaning up this type of sample using silica gel and modifiers with different polarity. In fact, an automated sample-cleanup system based on normal-phase HPLC using a silica gel column has been reported efficiently to clean up and fractionate chlorpyriphos, chlorpyriphos methyl, and their metabolites in molluscs. The system presents several advantages The procedure is fully automated, from the injection of the extract to the collection of fractions, which are injected directly into the GC system, and a diode array detector (DAD) allows online monitoring of the elution of lipids (68). 

It now remained for us to apply the Coward protocol to our system and complete the synthesis of lipid I. Thus, phosphate 26 [Scheme 10], prepared by reductive cleavage of the phosphodiester protective groups of 9 (H2, Pd/C in MeOH, followed by pyridine, 91% yield), was converted to the corresponding phosphoroimidazolidate, whose formation was readily monitored via mass spectrometry. Excess carbonyldiimidazole was quenched via addition of methanol. The lipid phosphate salt was then added in portions via syringe until complete consumption of the phosphoroimidazolidate intermediate was observed. Mass spectrometry also allowed us to monitor the appearance of the desired lipid-linked diphosphate product. When the reaction was judged to be complete, the reaction solution was carefully concentrated and the crude product was treated with sodium hydroxide in aqueous dioxane in order to achieve global deprotection. The crude product was purified by reverse-phase... 

Mid-IR has also been demonstrated for real-time concentration monitoring of a fermentation using a standard transmission cell, and the spectral response was similar, regardless of whether the broth was filtered or not.38 A PLS regression was used to perform quantitation of the substrate, the lactic acid bacterium, and the major metabolites. In another article describing quantitative mid-IR for fermentation studies, a model system was investigated under various fermentation conditions.39 The mid-IR provided insight into the relative concentrations of carbohydrates, nucleic acids, proteins and lipids in the host cells. Mid-IR has also been demonstrated for multi-component quantitation... 

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