LC-MS/MS Analysis of Canine Lipoproteins Fractionated Using the Ultracentrifugation-Precipitation Method

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Due to the lack of a gold standard method in canine lipoprotein analysis, it is unclear whether canine high-density lipoprotein (HDL) and low-density lipoprotein (LDL) can be accurately evaluated by the lipoprotein analysis methods used for dogs. This study investigated whether the ultracentrifugation-precipitation (U-P) method was suitable as a gold standard method for analyzing canine lipoprotein. First, the U-P method was compared with a gel permeation high performance liquid chromatography system (GP-HPLC). The concentrations of canine HDL-cholesterol (HDL-C) and LDL-cholesterol (LDL-C) determined by the U-P method correlated closely with those determined by GP-HPLC. However, the canine HDL-C concentration determined by the U-P method was lower than that by GP-HPLC, and the canine LDL-C concentration by the U-P method was higher than that by GP-HPLC. This study showed that some of the canine HDL could be precipitated with heparin manganese chloride solution. Second, the [HDL] and [LDL] fractions separated by the U-P method were analyzed by LC-MS/MS. The [HDL] fraction was found to contain only apolipoprotein A-I, which is an apolipoprotein of HDL, whereas the [LDL] fraction contained both apolipoprotein A-I and apolipoprotein B-100, which is an apolipoprotein of LDL. This data showed that a certain lipoprotein that includes apolipoprotein A-I might precipitate with canine LDL by use of heparin manganese chloride solution. These results indicated that the U-P method is not currently a gold standard method for analyzing canine lipoproteins.