Phospholipids, analyzers

Langmuir-Blodgett films (LB) and self assembled monolayers (SAM) deposited on metal surfaces have been studied by SERS spectroscopy in several investigations. For example, mono- and bilayers of phospholipids and cholesterol deposited on a rutile prism with a silver coating have been analyzed in contact with water. The study showed that in these models of biological membranes the second layer modified the fluidity of the first monolayer, and revealed the conformation of the polar head close to the silver [4.300]. 

An excellent example of PLC applications in the indirect coupling version is provided by the works of Miwa et al. [12]. These researchers separated eight phospholipid standards and platelet phospholipids from the other lipids on a silica gel plate. The mobile phase was composed of methylacetate-propanol-chloro-form-methanol-0.2% (w/v) potassium chloride (25 30 20 10 10, v/v). After detection with iodine vapor (Figure 9.2), each phospholipid class was scraped off and extracted with 5 ml of methanol. The solvent was removed under a stream of nitrogen, and the fatty acids of each phospholipid class were analyzed (as their hydrazides) by HPLC. The aim of this study was to establish a standardized... 

Preparative TLC may be applied to cleanup selected compound fractions separated from geochemical samples by such methods as HPLC, as Aries et al. [113] has described. To analyze phospholipids and nonphospholipids in sediments, organic matter was extracted and extracts LC-fractionated to obtain polar fractions. At the... 

Black, G. E. Snyder, A. Heroux, K. Chemotaxonomic differentiation between the Bacillus cereus group and Bacillus subtilis by phospholipid extracts analyzed with electospray tandem mass spectrometry. J. Microbiol. Meth. 1997, 28, 187-190. 

Historically, the target analytes in clinical mass spectrometric applications were small, volatile compounds that could be analyzed by GC-MS (see Chapter 4). With time, new chemical preparation techniques and derivatization schemes broadened the scope of these metabolites to include fatty acids, amino acids, intermediates of glucose oxidation, phospholipids, steroids, neurogenic amines, nucleic acids, etc. The molecular weights (molar masses) after derivatization were less than 1000 Da, a mass range easily within the limits of most conventional mass spectrometers. 

Six male Wistar rats inhaled HCN at 55 ppm for 30 min (Bhattacharya et al. 1984). HCN was generated by reaction of KCN with sulfuric acid and circulated through the chamber at the rate of 1 L/min. The rats were fitted with a lung mechanics analyzer (Buxco Electronic Inc.), and changes in air flow, transthoracic pressure, tidal volume, compliance, resistance, respiratory rate, and minute volume were determined every 10 min. Animals were sacrificed immediately following the exposure, and lungs were excised and analyzed for phospholipids (surfactant). 

A newer approach for lipid analysis is electrospray ionization tandem mass spectrometry (ESI-MS/MS) (Welti et al., 2002). This method requires limited sample preparation and sample size to identify and quantify lipids. Fauconnier et al. (2003) used ESI-MS/MS to analyze phospholipid and galactolipid levels during aging of potato tubers. 

Kitchen (1977) has analyzed the fatty acids in the phospholipids isolated from the MFGM, finding more unsaturated and less saturated acids than in the membrane TG. These findings are not unexpected. 

Phospholipid-derived fatty acids are often used to identify bacteria by capillary GC analysis after liquid solvent extraction, concentration steps, and chemical derivatization to their methyl esters. Our initial investigations attempted to extract the intact phospholipids, but no significant recoveries were achieved using pure C02. Even if SFE conditions were developed that could extract intact phospholipids, an additional derivatization step would be required before GC analysis of the fatty acid components. For these reasons, chemical derivatization/SFE was investigated in an effort to eliminate the lengthy conventional liquid solvent extractions as well as to combine (and shorten) the extraction and derivatization steps. The derivatization/SFE procedure was performed on samples of whole bacteria using 0.5 mL of 1.5% TMPA in methanol. The static derivatization step was performed for 10 minutes at 80°C and 400 atm C02, followed by dynamic SFE for 15 minutes at a flow rate of ca. 0.5 mL/min of the pressurized C02. Extracts were collected in ca. 3 mL of methanol and immediately analyzed by capillary GC without any further sample preparation. 

Consequently, a more objective way to measure the habitual intake of milk fat would be the fatty acid composition of adipose tissue. However, this is not routinely performed in larger cohort studies, due to cost and that the procedure is invasive and less tolerated by study participants. Analysis of plasma fatty acid composition is thus a more feasible option for examination to determine dairy intake in the study population. While some groups have separated plasma into its constituent phospholipids and cholesterol esters to analyze serum 15 0 and 17 0 as markers of dairy intake (Smedman et al., 1999), Baylin et al. (2005) found that plasma that was not separated into its constituent cholesteryl ester, phospholipids, and triacylglycerols was still able to reflect habitual dairy intakes comparably to adipose tissue. Thus, whole plasma is an acceptable alternative to fractionated plasma in the absence of adipose tissue for analysis to reflect habitual dairy intakes and may be a cost effective option for consideration when conducting future intervention studies to assess the affect of dairy products on health outcomes. 

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