Cholesterol

Lipoprotein Lipid Apolipoproteins Soluble in organic solvents and nearly insoluble in water Yield fatty acids on hydrolysis Complex alcohols that combine with fatty acids to form esters Lipoprotein An association of a core lipids with coat phospholipid and protein For solubility Apolipoproteins Protein components of lipoprotein.

Classification of Clinically Important lipids

Structure of cholesterol

Cholesterol biosynthesis

Esterification of cholesterol mediated by ACAT & LCA

Structure of a typical lipoprotein particle

apo B-100—containing lipoproteins VLDL, IDL, Lp(a), LDL

Contributors to total cholesterol in normal people LDL Two thirds HDL One third IDL and Lp(a) 2 to 3 mg/dL (each) Total chol = VLDL chol + LDL chol + HDL chol

Classification of LDL.Total. and HDL Cholesterol (mg/dL)

Reference methods To establish the accuracy of lipid and lipoprotein measurements Using the same basis (protocol ) for accuracy that had been used in developing the relationships between lipid and lipoprotein concentration and CHD. From studies, cut points for the risk characterization in patients were derived.

The accuracy of existing or newly developed methods could be assessed.

Methods and procedures commonly used Reference Method (total cholesterol) Chemical method Hydrolyze the cholesteryl esters. alcoholic KOH extracted from the mixture with hexane dried in vacuo acetic acid, acetic anhydride, and sulfuric acid Color development, 620 nm pure cholesterol as the calibrator. Expressed as mg/dL = mmoI/L x 38.7

HDL cholesterol Reference Method Prepare the HDL-containing fraction. a combination of ultracentrifugation and polyanion precipitation Ultracentrifugation Supernate VLDL and any chylomicrons accumulate as a floating layer Infranate IDL, LDL, Lp(a), HDL, and the other serum proteins Precipitation with heparin sulfate and MnCl2 Supernate HDL Precipitate IDL, LDL, Lp(a) The cholesterol in HDL fraction is then quantitated

V-LDL & LDL cholesterol VLDL chol = [Total chol] – [d > 1.006g/mL chol] LDL cholesterol Cholesterol infranate – cholesterol supernate (IDL, LDL, Lp(a), HDL— HDL fraction) The Friedewald Equation [LDL choI] = [Total chol] - [HDL chol]-[Triglyceride]/5 Unacceptable at TG concentrations > 400 mgldL

Methods Reference methods Routine Methods are complex, time consuming, at least partially manual, and require a high level of expertise for reliable operation Routine Methods Enzymatic methods

Enzymatic method

Interference Competition with the oxidation reaction bilirubin, ascorbic acid, and hemoglobin. adding substances such as bilirubin oxidase and dual wavelength readings to minimize the effects of hemolysis β-hydroxy sterols and plant sterols (e.g., β-sitosterol) can also react. are generally at very low concentrations ( so,not significant)

Sources of Variation in Lipid and Lipoprotein Measurements Analytical Variation Physiological Variation Contribute about 70 to 98% of the overall variance Variation for triglyceride is considerably higher

+, minimal to moderate increase ++, moderate to high increase -, minimal to moderate decrease -, moderate to high decrease NC, essentially no change or trend.

Recommendations Cholesterol Triglyceride, HDL and LDL cholesterol Two serial samples obtained at least 1 wk apart be used Fasting or non-fasting Triglyceride, HDL and LDL cholesterol Two to three serial specimens are recommended 12-h fast, or 9-h Specimens Serum or plasma (EDTA plasma)

Sampling Specimen storage in the same position on each occasion Serum or plasma should be removed from cells within 3 h Specimens can be stored for up to 3 d at 4 °C Up to several wk at —20 °C at —70 °C or lower for longer periods