A high content molecular fragmentation for the analysis of phosphatidylcholines (PC) was achieved utilizing a two-stage [trap (first generation fragmentation) and transfer (second generation fragmentation)] collision-induced dissociation (CID) in combination with travelling-wave ion mobility spectrometry (TWIMS). The novel aspects of this work reside in the fact that a TWIMS arrangement was used to obtain a high level structural information including location of fatty acyl substituents and double bonds for PCs in plasma, and the presence of alkali metal adduct ions such as [M + Li]+ was not required to obtain double bond positions. Elemental compositions for fragment ions were confirmed by accurate mass measurements. A very specific first generation fragment ion m/z 577 (M-phosphoryl choline) from the PC [16:0/18:1 (9Z)] was produced, which by further CID generated acylium ions containing either the fatty acyl 16:0 (C15H31CO+, m/z 239) or 18:1 (9Z) (C17H33CO+, m/z 265) substituent. Subsequent water loss from these acylium ions was key in producing hydrocarbon fragment ions mainly from the α-proximal position of the carbonyl group such as the hydrocarbon ion m/z 67 (+H2C-HC = CH-CH = CH2). Formation of these ions was of important significance for determining double bonds in the fatty acyl chains. In addition to this, and with the aid of 13C labeled lyso-phosphatidylcholine (LPC) 18:1 (9Z) in the ω-position (methyl) TAP fragmentation produced the ion at m/z 57. And was proven to be derived from the α-proximal (carboxylate) or distant ω-position (methyl) in the LPC.Electronic supplementary materialThe online version of this article (doi:10.1007/s13361-011-0172-2) contains supplementary material, which is available to authorized users.

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Despite therapies such as statins, which reduce circulating levels of low density lipoprotein cholesterol (LDL-C), cardiovascular event rates remain high. Numerous epidemiological studies (e.g., the Framingham Heart Study) indicate that high density lipoprotein cholesterol (HDL-C) levels are inversely correlated with cardiovascular risk ( 1-6 ). Therefore, therapies that increase HDL-C have gained recent attention as possible treatments for dyslipidemia and atherosclerosis.Cholesteryl ester transfer protein (CETP) mediates transfer of cholesteryl ester (CE) and triglyceride (TG) between HDL and apoB-containing lipoproteins such as LDL and therefore, represents an attractive target for increasing HDL-C and reducing LDL-C. Indeed, initial clinical trials with torcetrapib established the validity of CETP inhibition as a mechanism for elevation of HDL-C ( 7, 8 ). However, the phase III outcome trial ILLUMINATE demonstrated that torcetrapib treatment was associated with an increase in cardiovascular events and overall mortality, possibly due to off-target effects on blood pressure and circulating adrenal hormones ( 9 ). A series of preclinical studies further corroborated that torcetrapib had compoundspecifi c off-target activity that was unrelated to CETP inhibition ( 10-12 ).Anacetrapib (ANA) is a potent CETP inhibitor that has not demonstrated the off-target activities of torcetrapib in preclinical or clinical studies ( 10,(13)(14)(15). ANA treatment increases HDL-C by over 100% and lowers LDL-C by 30-40% as a monotherapy and when coadministered with statins ( 13-15 ). In a recent 1.5 year safety study in ‫ف‬ 1,600 patients with cardiovascular disease ( 15 ) Cardiovascular disease continues to be a major contributor to morbidity and mortality throughout the world. 29 July 2011. Published, JLR Papers in Press, August 14, 2011 DOI 10.1194 Manuscript received 13 April 2011 and in revised form

high level of cholesterol in the body circulation is strongly associated with atherosclerosis ( 6-10 ). The source of cholesterol comes from different areas: dietary, de novo synthesis, and synthesis in extrahepatic tissues. The liver acts as a cross-junction at which cholesterol is incorporated into HDL/LDL, and then secreted as free cholesterol in the bile or in the form of bile salts/acids.Studies of cholesterol metabolism typically require measurements of static concentrations of cholesterol to identify differences between models or to determine the presence or absence of a disease phenotype. The simultaneous use of stable or radio isotope fl ux analysis can aid in understanding the nature of a metabolic abnormality and yield information regarding the dynamics that contribute to altered or perturbed homeostasis.Questions surrounding cholesterol dynamics have been addressed using isotopic-labeled water for nearly 70 years, beginning with the pioneering studies of Schoenheimer The liver plays a vital role in cholesterol metabolism ( 1-5 ) and homeostasis. In addition to this, production of bile acids from cholesterol also plays an important role in the secretion and degradation of plasma lipoproteins. A 25 September 2010. Published, JLR Papers in Press, September 30, 2010 DOI 10.1194 In vivo D2O labeling to quantify static and dynamic changes in cholesterol and cholesterol esters by high resolution LC/MS Manuscript received 27 July 2010 and in revised form