Glycerophosphocholine

Several useful publications describing early direct injection API-electrospray methods appHcable to neurochemical studies include studies on acetylchoHnesterase phosphonylation products (Barak et al., 1997) glycerophosphocholine lipids (Flarrison and Murphy, 1996) receptor antibody characterization (Lewis et al., 1994) neuropeptide Yanalogs (Beck-Sickinger et al., 1994) and large biomolecules (Loo et al.,... 

This enzyme [EC 3.1.1.47], also known as plateletactivating factor acetylhydrolase, 2-acetyl-l-alkyl-glycerophosphocholine esterase, and LDL-associated phospholipase A2, catalyzes the hydrolysis of 2-acetyl-l-alkyl-xn-glycero-3-phosphocholine to yield l-alkyl-xn-glycero-3-phosphocholine and acetate. 

This enzyme [EC 2.3.1.67], also referred to as 1-alkyl-glycerophosphocholine acetyltransferase, catalyzes the reaction of acetyl-CoA with l-alkyl-5 n-glycero-3-phos-phocholine to produce coenzyme A and l-alkyl-2-acetyl-5 n-glycero-3-phosphocholine. 

This enzyme [EC 3.1.1.5] (also referred to as lecithinase B, lysolecithinase, and phospholipase B) catalyzes the hydrolysis of a 2-lysophosphatidylcholine to produce glycerophosphocholine and a fatty acid anion. 

Fig. 4. 500 MHz DOSY spectrum of a D2O solution of a perchloric acid extract of gerbil brain. Assignments of selected signals are indicated as follows ac = acetate ala = alanine cho = choline cr = creatine ere = creatinine etn = ethanolamine GABA = 7-aminobutyric acid glu = glutamate GPC = glycerophosphocholine lac = lactate m-ino = myo-inositol NAA = N-acetylaspartate succ = succinate and tau = taurine. (Raw data...

The location of the double bonds in unsaturated glycerophosphocholine (GPC) lipids is made difficult mainly due to the limited amounts of these compounds isolated from cells. When a thin layer of a GPC lipid is exposed to the action of ozone, partial or total ozonization of the unsaturated fatty acid residues may take place, depending on the particular compound being analyzed. The ozonized hpid is dissolved and analyzed by tandem MS. A typical injection into the instrument may be 5 xL containing 10-20 ng of the lipid. An illustrative example is the ozonized l-stearoyl-2-arachidonoyl-GPC, showing the systematic fragmentation pattern in Scheme 21, typical of the ozonized GPC lipids. Thus, the molecular peak appears increased by one nominal unit (M + H, m/e 1002),... 

GOX (glucose oxidase), 631, 636, 637 GPC (unsaturated glycerophosphocholine), 737 GPO (l-glycerol-3-phosphate oxidase), 633 Graft polymers cellulose, 698 cross-linking initiators, 706 ozonized polymers, 622 Griesbaum co-ozonolysis, 1,2,4-tiioxane antimalarials, 1331... 

Figure 1 In vivo spectra acquired from the human occipital lobe at 4 T (top)and 7 T (bottom) using the same experimental setup and NMR acquisition parameters (TR=3 s and 128 signal averages). PE, phosphoethanolamine PC, phosphocholine Pi, inorganic phosphate GPE, glycerophosphoethanolamine GPC, glycerophosphocholine ...

Hepatic levels of choline, phosphocholine and glycerophosphocholine were reduced as much as 64, 84 and 70%, respectively in male B6C3Fi mice after two weeks administration of diethanolamine (160 mg/kg bw per day) via oral gavage or skin painting. These levels were inversely related to the blood diethanolamine levels (uptake) after the final dose. In contrast, the hepatic levels of sphingomyelin were increased relative to those in control mice, and were directly correlated with blood diethanolamine levels (Stott et al, 2000). 

SCHEME 21. Fragmentation pattern of the ozonide of an unsaturated glycerophosphocholine lipid in electrospray MS... 

Platelet-activating factor (PAF) l-O-alkyl-2-acetyl-sw-glycerophosphocholine... 

Liver-aqueous extract Choline, phosphocholine Glycerophosphocholine Lactate Amino acids Glucose... 

Du L, White RL (2008) Reducing glycerophosphocholine lipid matrix interference effects in biological fluid assays by using high-turbulence liquid chromatography. Rapid Commun Mass Spectrom 22 3362-3370... 

Figure 1. 3,P NMR spectrum of Chinese hamster ovary cells (CHO K1) growing in a hollow-fiber bioreactor (Callies, Jackson, and Brindle, unpublished observations). The assignments are PME, phosphomonoesters inorganic phosphate (predominantly extracellular) PDE, phosphodiesters (predominantly glycerophosphocholine) PCr, phosphocreatine ATP, adenosine-triphosphate (y-, a-, and (1-phosphates) NAD, nicotinamide adenine dinucleotide DPDE, diphosphodiesters.

The observation of phospholipid metabolites in the 3IP NMR spectra of cells and cell extracts (see Figure 1) has led to several studies designed to investigate the control of phospholipid metabolism. These metabolites include phosphocholine (PC), phosphoethanolamine (PE), glycerophosphocholine (GPC), and glycero-phosphoethanolamine (GPE). The phospholipids, themselves, are not observable in vivo, except at low field strengths, due to their relative lack of mobility and consequently broad signals (Murphy et al., 1989). They are observable, however, in nonaqueous cell extracts (Ronen et al., 1990). A primary motivation for these studies has been the observation of altered levels of these intermediates in tumor cells (see below). 

First, anhydrous sodium hydroxide is dissolved in methanol to make a 0.5 M solution. Then the phosphatidylcholine sample, in chloroform-methanol (1 1, v/v), is placed in a Pyrex glass conical centrifuge tube and evaporated to dryness under a stream of nitrogen. To this residue is added a small volume of chloroform (usually 5-10% of the final volume of methanol). Subsequent to completion perhaps the most effective route to structure proof of phosphatidylcholine is one in which the glycerophosphocholine backbone is obtained in an unaltered form. Essentially this involves the cleavage of the ester bonds and can be achieved by any of the four reaction pathways illustrated in Figure 4-3. 

For our purposes, the characterization of the glycerophosphocholine present in the water-rich phase will have center stage, with a more modest discussion of the fatty acid ester fraction. As a footnote to the above experiment, one should measure carefully the distribution of phosphorus in the two fractions the phosphorus should be exclusively in the water-soluble fraction. 

Further insight into the chemical structure of the glycerophosphocholine can be achieved through use of a few analytical procedures outlined as follows. 

In the direct chemical attack for proof of structure of phosphatidylcholine (see the section entitled Glycerophosphochline Characterization ) the decision was made to subject this phospholipid to a base-catalyzed methanolysis. As noted, two products are formed, namely, glycerophosphocholine and the methyl esters of the long-chain fatty acid substituents on the intact phosphatidylcholine. Since the analytical approach to proof of structure of the glycerophosphocholine has been achieved, it is logical now to consider the other product, the methyl esters. 

Among the phospholipases, phospholipase A2 has been of major importance in establishing the positional specificity of fatty acids on the glycerophosphocholine backbone. However, two other phospholipases, namely C... 

This pure glycerophosphocholine can be used as starting material for the synthesis of mixed acid phosphatidylcholine as shown in Figure 4-10. 

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