Phospholipid Determination

Chen PS, Toribara TY (1956) Anal Chem 28 1756 1.3.3 Phospholipid Determination... 

Let us consider first lipid-lipid interaction. Urry et al, showed the existence of a positive CD band at 218 m/x and a negative CD band at about 192 m/z in phosphatidyl choline and phosphatidyl ethanolamine dissolved in trifluoroethanol (86). The 192-m/z band was not characterized in detail, but the 218-m/z band is of such position and shape that the addition of lipid and protein CD bands could produce a composite CD band, and hence an ORD Cotton effect, which is red shifted. As noted by Urry, the 218-m/z CD extremum of lecithin must arise from n — 7T transitions in the fatty acid ester groups. Although the optical activities of solutions of deproteinized membrane phospholipids determined at the same concentration as in the intact membrane are negligibly small, in membranes an ordered array of lipids could greatly enhance rotation. Such an effect could yield information on the nature of lipid-lipid association. This can be tested experimentally. Halobacterium cutirubrum offers a unique system since Kates has shown that the lipids in this extreme halophile contain ether bonds rather than ester bonds (43, 44), Hence, the n — tt transition essential to the CD band at 218 m/z in phospholipids does not exist. Nevertheless, we found that the ORD... 

As discussed in Section 3.7, when these phospholipids are mixed with water, they assemble in an arrangement called a lipid bilayer. The ionic heads of the phospholipid are oriented on the outside and the nonpolar tails on the inside. The identity of the fatty acids in the phospholipid determines the rigidity of this bilayer. When the fatty acids are saturated, they pack well in the interior of the lipid bilayer. and the membrane is quite rigid. When there are many unsaturated fatty acids, the nonpolar tails cannot pack as well and the bilayer is more fluid. Thus, important characteristics of this lipid bilayer are determined by the three-dimensional stmcture of the molecules that comprise it. 

Phospholipid determination is done by the phosphomolybde-num blue method (15). All measurements (calibration-i-unknown samples) are done in triplicate (see Note 18). The following protocol is followed ... 

Meers, R and Mealy, T, Phospholipid determinants for annexin V binding sites and the role of tryptophan 187, Biochemistry, 33[19], 5829, 1994. 

Long chains of water-insoluble lipids are constructed from building blocks called fatty acids, which are also stowed away as a source of fuel in fat cells. The chemical bonds linking fatty acid units together in glycolipids and phospholipids determine how rigid or floppy the lipid chains will turn... 

S. Brown, H. A. Dey, S. K. Caprioli, R. M., Spatial and temporal alterations of phospholipids determined by mass spectrometry during mouse embryo implantation , J. Lipid Res. 2009, 50, 2290-2298. 

Phospholipids occur in cells mostly as components of cellular membranes in which they are in close structural relation to proteins. This interaction is an important factor in relation to the problem of phospholipid preservation during dehydration and embedding of tissues. Data on total phospholipid retention, following labeling with fatty acids are given by Kom and Weisman (1966) in the amoeba and by Dermer (1968) in rat intestine. The former have noted better retention of phospholipids than of neutral lipids, while the latter reported that both were retained to the same extent Very little loss of phospholipids, determined chemically, from erythrocyte stroma fixed with glutaraldehyde and osmium tetroxide in the presence of Ca ions was reported by Mitchell (1969). The loss increased from 2 to 7% if the stroma was stored for 7-10 days prior to fixation. 

Phospholipids. Phospholipids, components of every cell membrane, are active determinants of membrane permeabiUty. They are sources of energy, components of certain enzyme systems, and involved in Hpid transport in plasma. Because of their polar nature, phosphoUpids can act as emulsifying agents (42). The stmcture of most phosphoUpids resembles that of triglycerides except that one fatty acid radical has been replaced by a radical derived from phosphoric acid and a nitrogen base, eg, choline or serine. 

HJ. Kisner, C.W. Brown, GJ. Kavamos, "Simultaneous Determination of Triglycerides, Phospholipids, and Cholesteryl Esters by Infrared Spectrometry", Anal. Chem. 1982 (54) 1479-1485. 

Ueno, M., Tanford, C., and Reynolds, J. A. (1984). Phospholipid vesicle formation using nonionic detergents with low monomer solubility. Kinetic factors determine vesicle size and polydis-persity. Biochemistry, 3070-3076. 

Using PTLC six major fractions of lipids (phospholipids, free sterols, free fatty acids, triacylglycerols, methyl esters, and sterol esters) were separated from the skin lipids of chicken to smdy the penetration responses of Schistosoma cercaria and Austrobilharzia variglandis [79a]. To determine the structure of nontoxic lipids in lipopolysaccharides of Salmonella typhimurium, monophosphoryl lipids were separated from these lipids using PTLC. The separated fractions were used in FAB-MS to determine [3-hydroxymyristic acid, lauric acid, and 3-hydroxymyristic acids [79b]. 

N. Ragendran, O. Matsuda. N. Imamura, and Y. Urushigawa, Variation in microbial biomass and community structure in sediments of eutrophic bays as determined by phospholipid ester-linked fatty acids, Appl. Environ. Microbiol. 58 562 (1992). 

B. H. Baird and D. C. White, Bioma.ss and eommunily strueture of the abyssal microbiota determined from the ester-linked phospholipids recovered from Venezuela Basin and Puerto Rico trench. sediment. Mar. Geol. 58 2 7 (1985). 

One major interest in vibrational surface spectroscopy is the ability to directly probe lipid layers. Similarly to the previous case, the structure of the alkyl chains of phospholipids is readily determined from the ratio of the magnitude of the CH2 and CH3 symmetrical stretching modes [136,137]. At the D2O-CCI4 interface, a layer of... 

From a practical point of view, the potential drop across the inner layer A 02 must be determined by fitting the experimental data to the equations derived from this theoretical approach, which led to some controversy about its value [53,54,56,57]. For the sake of simplicity, and also because recent studies of the ITIES structure do not confirm the presence of an inner layer [51,58], we neglect the finite size of the transferring ion and take X2 = X2 = 0 and A 02 = 0. This is equivalent to accepting that the potential difference Afl02 — A 0 is not modified by the presence of the phospholipids. 

As an example of suspected endocrine disruptors (EDs), studies of the estrogenic action of bisphenol A (BPA) have been in progress in medical, physiological, and biological fields. In this situation, physicochemical approaches are required to get the structural information of BPA trapped in biomembranes. Most recently, we have determined the site and the orientation of BPA trapped in phospholipid vesicles by NMR, using the HCS rule [47]. In particular, we have succeeded in monitoring the NMR spectral changes of phospholipid vesicles, which are induced by the BPA delivery. 

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