Alkenylacyl Phospholipids Research Articles

Influence of temperature on 31P NMR chemical shifts of phospholipids and their metabolites I. In chloroform–methanol–water

Spectral overlap of 31P NMR resonances and the lack of reproducibility in chemical shifts corresponding to phospholipids in organic solvents challenge the accuracy of band assignments and quantification. To alleviate these problems, the use of temperature coefficients is proposed. Changes in temperature enable the resolution of overlapped resonances and provide a facile approach for the computation of temperature coefficients. The coefficients were evaluated for various glycero- and sphingo-phospholipids. Their values suggest that differences in H-bonding between the phosphate and the head groups are responsible for the changes of chemical shift with temperature. Among parent phospholipids, and in addition to sphingomyelin, the smallest temperature coefficient values (closest to zero) were observed for phosphatidylcholine, phosphatidylglycerol, dihydrosphingomyelin, and cardiolipin. The highest values were exhibited by phospholipids with protonated head groups, such as phosphatidylserine and phosphatidylethanolamine. The lowest and, in fact, negative values were measured for phospholipids with an exposed phosphate group: phosphatidic acid, ceramide-1-phosphate, and dihydroceramide-1-phosphate. Diacyl, alkyl-acyl, and alkenyl-acyl phospholipids with the same head group exhibited comparable coefficients but differed slightly in chemical shifts. Compared to their parent glycerophospholipids, all lyso analogs had greater temperature coefficients, possibly due to the presence of an extra OH capable of forming a H-bond with the phosphate group.

Analysis of plasmenylethanolamines using electrospray tandem mass spectrometry and its application in screening for peroxisomal disorders.

Ether phospholipids are major membrane constituents in mammalian cells and can be divided into two subgroups, namely alkenyl-acyl phospholipids and alkyl-acyl phospholipids. The alkenyl-acyl phospholipids are predominantly present in the phosphatidylethanolamine fraction (1-O-alk-1'-enyl-2-acyl-sn-glycero-3-phosphatidylethanolamine), whereas the alkyl-acyl phospholipids are predominantly found in the phosphatidylcholine fraction (l-alkyl-2-acyl-sn-glycero-3-phosphatidylcholine) (Lee 1998). The glycerolipids with alk-1-enyl groups at the sn-1 position are also referred to as plasmalogens. Two essential steps in the biosynthesis of ether phospholipids take place in peroxisomes and are catalysed by acyl-CoA: dihydroxyacetone-phosphate acyltransferase (DHAP-AT, EC 2.3.1.42) and alkyldihydroxyacetone-phosphate synthase (alkyl-DHAP synthase, EC 2.5.1.26) (Lee 1998). In a number of peroxisomal disorders (e.g. peroxisomal biogenesis defects, rhizomelic chondrodysplasia punctata (RCDP) and isolated DHAP-AT or alkyl-DHAP synthase deficiency), ether phospholipid biosynthesis is (partially) deficient (Van den Bosch et al 1993; Wanders et al 1992; Wanders et al 1994). For diagnostic purposes, erythrocyte plasmalogens are usually measured indirectly after transmethylation, which converts the plasmalogens to their corresponding dimethyl acetals (DMAs), followed by gas chromatographic (GC) analysis (Bjorkhem et al 1986; Dacremont and Vincent 1995). The relative plasmalogen content is then calculated as a ratio of DMA versus the corresponding fatty acid methyl ester.

Distribution of arachidonic acid among phospholipid subclasses of lone star tick salivary glands

The subclass composition of choline- and ethanolamine-containing phospholipids was determined by analysis of acyl-linked fatty acids released by base hydrolysis of diradylglycerobenzoates formed from lone star tick salivary gland diacyl, alkylacyl and alkenylacyl phospholipids. The diacyl subclass comprises 87% of all choline-containing phospholipids, while the alkylacyl subclass comprises c. 9% and the alkenylacyl subclass c. 4%. The diacyl subclass comprises 72–77% of ethanolamine-containing phospholipids and about 14 and 13% of this subclass of phospholipid are alkylacyl and alkenylacyl lipids, respectively. Arachidonic acid (20:4) is the most abundant fatty acid (28% of all fatty acids) esterified in the alkylacyl form of phosphatidylcholine (PC) and it comprises 17% of the fatty acids in alkenylacyl-PC. The alkylacyl form of phosphatidylethanolamine (PE) is also rich in 20:4 (24%) while the alkenylacyl-PE subclass contains only 9% 20:4. Despite the relatively high amounts of 20:4 within the ether-linked phospholipids, the majority of the salivary gland 20:4 (>83%) is found in the diacyl phospholipid subclass because of the preponderence of this subclass in tick salivary glands. Isolated salivary glands incorporated [ 3H]-20:4 primarily (>98%) into the sn-2 position of diacyl PC > PE, with some incorporation into triglycerides. Continued incubation in the absence of labeled 20:4 demonstrated remodeling of [ 3H]-20:4 from PC into PE, and from the diacyl subclass to the alkylacyl subclass in the choline containing phospholipids.

Transmembrane distribution and translocation of spin-labeled plasmalogens in human red blood cells

We have synthesized two new spin-labeled alkenylacyl phospholipids (plasmalogens) in order to investigate the transmembrane distribution and transport of this subclass of glycerophospholipids in human red blood cells. The plasmenylethanolamine analogue diffuses rapidly from the outer to the inner leaflet with a half time at 37°C of 30 min comparable to that of the corresponding diacyl-phosphatidylethanolamine spin-label in an ATP-requiring and N-ethyl maleimide sensitive manner. The plateau corresponds to 79% of the aminophospholipids on the innner leaflet. By contrast, after 4 h incubation less than 20% of the plasmenylcholine spin-labels reach the interior. Thus plasmalogens behave as the corresponding diacyl-lipids. We infer that plasmenylethanolamine is transported from the outer to the inner leaflet of the red cell membrane by the aminophospholipid translocase.

Fatty acid composition of diacyl, alkylacyl, and alkenylacyl phospholipids of control and arachidonate-depleted rat polymorphonuclear leukocytes.

Phospholipid fatty acid composition and phospholipid subclass distribution of control and arachidonate-depleted rat polymorphonuclear leukocytes (PMN) were compared. The 20:4-depleted PMN contained significantly higher amounts of 16:1, 18:1 and 20:3 (delta 5,8,11) and lower amounts of 18:2 and 20:4 than the phospholipids from control cells. Choline-containing glycerophospholipids (CGP) were the major phospholipids of both control and 20:4-depleted cells representing 34% and 37% of the total phospholipids, respectively. Significant amounts of ethanolamine-containing glycerophospholipids (EGP) (29% and 30%) and sphingolipids (20% and 18%) were also present in both cell types. Serine-containing glycerophospholipids (SGP) together with inositol-containing glycerophospholipids (IGP) constituted 16% and 13% of the phospholipids in control and 20:4-depleted cells, respectively. CGP from control cells had significantly higher amounts of 16:0 and 18:2 and lower amounts of 18:0 and 20:4 than EGP, whereas CGP from 20:4-depleted cells has higher amounts of 16:0 and 16:1 and lower amounts of 20:3 than EGP. Analysis of the subclass composition of CGP and EGP revealed that both control and 20:4-depleted cells contained significantly large amounts of alkylacyl-GPC and alkenylacyl-GPE. Small amounts of alkylacyl-GPE and alkenylacyl-GPC were also observed. The predominant fatty acyl residues found in the 1,2-diacyl-GPC, alkylacyl-GPC of control cells were 16:0, 18:0, 18:1, 18:2, and 20:4, while those of 20:4-depleted cells were 16:0, 16:1, 18:0, 18:1, and 20:3. More than 60% of CGP-bound 20:4 of control cells and about 70% of the CGP-bound 20:3 of 20:4-depleted cells were found in their alkylacyl species.(ABSTRACT TRUNCATED AT 250 WORDS)

Preferential synthesis of diacyl and alkenylacyl ethanolamine and choline glycerophospholipids in rabbit platelet membranes.

In rabbit platelet membranes, the contents of alkenylacyl phospholipids (plasmalogen) were 56% of phosphatidylethanolamine and 3% of phosphatidylcholine. This uneven distribution of plasmalogens in each phospholipid class could be attributed to the different substrate specificity of ethanolaminephosphotransferase (EC 2.7.8.1) and cholinephosphotransferase (EC 2.7.8.2). The properties of the enzymes were studied, using endogenous diglycerides and CDP-[3H]ethanolamine or CDP-[14C]choline as substrates. The newly formed phospholipids were mainly diacyl and alkenylacyl and only rarely alkylacyl type. The ratios of the labeled alkenylacyl to diacyl type of phospholipids clearly varied with the concentrations of CDP-ethanolamine or CDP-choline. When 1, 10, and 30 microM CDP-[3H]ethanolamine were used, the labeled phospholipids contained 53, 37, and 27% of the alkenylacyl type, respectively. The apparent Km for CDP-ethanolamine to synthesize alkenylacyl and diacyl types were 2.2 and 8.1 microM. On the other hand, when 1, 10, and 30 microM CDP-[14C]choline were used, the labeled lipids contained 10, 17, and 24% alkenylacyl type, respectively. The apparent Km for CDP-choline to synthesize alkenylacyl and diacyl types were 24 and 4.3 microM. Further, the syntheses of diacyl type of phosphatidylethanolamine and the alkenylacyl type of phosphatidylcholine were markedly inhibited by unlabeled CDP-choline and CDP-ethanolamine, respectively. The two enzymes had opposite substrate specificities, and ethanolaminephosphotransferase showed a high preference to plasmalogen synthesis, especially in the presence of CDP-choline.

Arachidonate mobilization in diacyl, alkylacyl and alkenylacyl phospholipids on stimulation of rat platelets by thrombin and the Ca2+ ionophore A23187.

Platelet stimulation by thrombin or Ca2+ ionophore induces mobilization of arachidonate from lipid stores. We have previously shown that, in [14C]arachidonic acid-prelabelled resting platelets, [14C]arachidonate was transferred from diacyl-sn-glycerophosphocholine to ethanolamine and choline-containing ether phospholipids. This transfer reached an equilibrium after 5 h incubation [Colard, Breton & Bereziat (1984a) Biochem. J. 222, 657-662]. [14C]Arachidonate-prelabelled platelets having reached this transfer equilibrium were used to study the mobilization of arachidonate in etheracyl and diacyl phospholipids. Upon thrombin stimulation, arachidonate decreased in diacyl-sn-glycero-3-phosphoinositol, in alkylacyl- and diacyl-sn-glycero-3-phosphocholine and increased in alkenylacyl- and diacyl-sn-glycero-3-phosphoethanolamine. Upon challenge with Ca2+ ionophore A23187, arachidonate decreased in diacyl-sn-glycero-3-phosphoethanolamine, in diacyl- and alkylacyl-sn-glycero-3-phosphocholine and increased in alkenylacyl-sn-glycero-3-phosphoethanolamine. We also compared arachidonate mobilization in platelets stimulated immediately after [14C]arachidonic acid chase with platelets stimulated after 5 h reincubation. We observed that the arachidonate newly incorporated into diacyl-sn-glycero-3-phosphocholine and triacylglycerols was rapidly released upon stimulation. This suggests the presence in these two lipids of a rapidly-turning-over arachidonate pool.